Die wissenschaftliche Begründung für die Entscheidung des portugiesischen Berufungsgerichts zur Aufhebung der Quarantäne für vier Personen auf den Azoren …
Portugiesisches Berufungsgericht hält PCR-Tests für unzuverlässig und hebt Quarantäne auf (tkp.at)
… liefert eine vernichtende Experten-Kritik an Drostens PCR-Testverfahren zur Feststellung von Sars-Cov 2 Infektionen mit den verheerenden Konsequenzen seit dem Frühjahr:
Im erläuternden Video von Gunnar Kaiser hervorragend erklärt:
Portugiesisches Berufungsgericht hält PCR-Tests für unzuverlässig und hebt Quarantäne auf (tkp.at)
Portugiesisches Berufungsgericht hält PCR-Test für unzuverlässig – reitschuster.de
CORONA 42: Corona-Knast trifft hier Juristen, in der Schweiz bereits auch Ärzte – barth-engelbart.de (barth-engelbart.de) wie Dr. Binder, der an dem „Corman-Drosten Review Report“ mitgearbeitet hat – u.a. zusammen mit Dr. Michael Yeadon BSs(Hons) Biochem Tox U Surrey, PhD Pharmacology U Surrey. Managing Director, Yeadon Consulting Ltd, former Pfizer Chief Scientist, United Kingdom ,
Und dieser Dr. Michael Yeadon, der Spitzenwissenschaftler und Ex-Pfizer-Vizepräsident warnt auch öffentlich eindringlich vor der Impfung mit dem schnellentwickelten Stoff aus dem Hause BioNTech/Pfizer.
CORMAN-DROSTEN REVIEW REPORT
KURATIERT VON EINEM INTERNATIONALEN KONSORTIUM VON WISSENSCHAFTLERN IN LIFE SCIENCES (ICSLS)
Bericht Corman-Drosten et al. Eurosurveillance 2020
Dieser umfangreiche Überprüfungsbericht wurde der Eurosurveillance-Redaktion am 27. November 2020 offiziell über ihr Einreichungsportal vorgelegt, das diesem Überprüfungsbericht beigefügt ist, ist ein Widerrufsantragsschreiben, das von allen Haupt- und Mitautoren unterzeichnet wurde. Erste und letzte aufgeführte Namen sind die ersten und zweiten Hauptautoren. Alle Namen dazwischen sind Co-Autoren.
Externe Peer-Review des RTPCR-Tests zum Nachweis von SARS-CoV-2 zeigt 10 wesentliche wissenschaftliche Mängel auf molekularer und methodischer Ebene: Folgen für falsch positive Ergebnisse.
Pieter Borger(1), Bobby Rajesh Malhotra(2) , Michael Yeadon(3) , Clare Craig(4), Kevin McKernan(5) , Klaus Steger(6) , Paul McSheehy(7) , Lidiya Angelova(8), Fabio Franchi(9), Thomas Binder(10), Henrik Ullrich(11) , Makoto Ohashi(12), Stefano Scoglio(13), Marjolein Doesburg-van Kleffens(14), Dorothea Gilbert(15), Rainer Klement(16), Ruth Schruefer(17), Berber W. Pieksma(18), Jan Bonte(19), Bruno H. Dalle Carbonare(20), Kevin P. Corbett(21), Ulrike Kämmerer(22)
Abstrakt
In der Publikation „Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR“ (Eurosurveillance 25(8) 2020) präsentieren die Autoren einen diagnostischen Workflow und rt-qPCR-Protokoll zur Detektion und Diagnose von 2019-nCoV (jetzt bekannt als SARS-CoV-2), die sie für validiert halten, sowie eine robuste diagnostische Methode für den Einsatz in Laborumgebungen im öffentlichen Gesundheitswesen.
Angesichts aller Folgen, die sich aus dieser Veröffentlichung für Gesellschaften weltweit ergeben, führte eine Gruppe unabhängiger Forscher eine Punkt-für-Punkt-Überprüfung der oben genannten Veröffentlichung durch, in der 1) alle Komponenten des vorgestellten Testdesigns überprüft wurden, 2) die RT-qPCR-Protokollempfehlungen w.r.t. good laboratory practice bewertet wurden, und 3) Parameter, die mit einschlägiger wissenschaftlicher Literatur auf diesem Gebiet untersucht wurden.
Das veröffentlichte RT-qPCR-Protokoll zur Detektion und Diagnose von 2019-nCoV und das Manuskript leiden unter zahlreichen technischen und wissenschaftlichen Fehlern, darunter unzureichendes Primerdesign, ein problematisches und unzureichendes RT-qPCR-Protokoll und das Fehlen einer genauen Testvalidierung. Weder der vorgelegte Test noch das Manuskript selbst erfüllen die Voraussetzungen für eine akzeptable wissenschaftliche Veröffentlichung. Darüber hinaus werden schwerwiegende Interessenkonflikte der Autoren nicht erwähnt. Schließlich bedeutet der sehr kurze Zeitraum zwischen Einreichung und Annahme der Veröffentlichung (24 Stunden), dass hier entweder kein systematischer Peer-Review-Prozess durchgeführt wurde oder von problematischer schlechter Qualität. Wir liefern überzeugende Beweise für mehrere wissenschaftliche Unzulänglichkeiten, Fehler und Fehler.
Angesichts der hier vorgestellten wissenschaftlichen und methodischen Fehler sind wir zuversichtlich, dass die Redaktion von Eurosurveillance keine andere Wahl hat, als die Veröffentlichung zurückzuziehen.
CONCISE REVIEW REPORT
Dieses Papier wird zahlreiche schwerwiegende Mängel im Corman-Drosten-Papier aufzeigen, deren Bedeutung zu einer weltweiten Fehldiagnose von Infektionen geführt hat, die SARS-CoV-2 zugeschrieben werden und mit der Krankheit COVID-19 in Verbindung gebracht werden. Wir sind mit strengen Blockaden konfrontiert, die das Leben und die Existenzgrundlage vieler Menschen zerstört haben, der eingeschränkte Zugang zu Bildung und diese von Regierungen auf der ganzen Welt auferlegten Beschränkungen sind ein direkter Angriff auf die Grundrechte und ihre persönlichen Freiheiten, was zu Kollateralschäden für ganze Volkswirtschaften auf globaler Ebene führt. Es gibt zehn fatale Probleme mit dem Corman-Drosten-Papier, die wir in den folgenden Abschnitten näher erläutern und erläutern werden.
Das erste und wichtigste Problem ist, dass der Roman Coronavirus SARS-CoV-2 (in der Publikation 2019-nCoV und im Februar 2020 SARS-CoV-2 von einem internationalen Konsortium von Virenexperten) auf silico (theoretischen) Sequenzen basiert, von einem Labor in China [1] geliefert, da den Autoren zu dieser Zeit weder Kontrollmaterial von infektiöser („live“) oder inaktivierter SARS-CoV-2 noch isolierte genomische RNA des Virus zur Verfügung stand. Bislang wurde von der Autorenschaft keine Validierung auf der Grundlage isolierter SARS-CoV-2-Viren oder ihrer RNA in voller Länge durchgeführt. Nach Corman et al.:
„Wir wollten eine robuste Diagnosemethodik für den Einsatz in Laborumgebungen für die öffentliche Gesundheit entwickeln und einsetzen, ohne dass Virenmaterial zur Verfügung steht.“ [1]
Der Schwerpunkt sollte dabei auf den beiden genannten Zielen liegen: a) Entwicklung und b) Einführung eines diagnostischen Tests fürden Einsatz im Labor für öffentliche Gesundheit . Diese Ziele sind nicht erreichbar, ohne dass tatsächliches Virusmaterial zur Verfügung steht (z. B. zur Bestimmung der infektiösen Viruslast). In jedem Fall kann nur ein Protokoll mit maximaler Genauigkeit das obligatorische und primäre Ziel in jedem Szenario-Ergebnis dieser Größenordnung sein. Kritische Viruslastbestimmung ist eine obligatorische Information, und es liegt in der Gruppenverantwortung von Christian Drosten, diese Experimente durchzuführen und die entscheidenden Daten zur Verfügung zu stellen.
Diese in Silico-Sequenzen wurden jedoch zur Entwicklung einer RT-PCR-Testmethodik zur Identifizierung des vorgenannten Virus verwendet. Dieses Modell beruhte auf der Annahme, dass das neuartige Virus SARS-CoV aus dem Jahr 2003 sehr ähnlich ist, da beide Beta-Coronaviren sind.
Der PCR-Test wurde daher unter Verwendung der genomischen Sequenz von SARS-CoV als Kontrollmaterial für die Sarbeco-Komponente
entwickelt; Das wissen wir aus unserer persönlichen E-Mail-Kommunikation mit [2] einem der Co-Autoren des Corman-Drosten-Papiers. Diese Methode zum Modellieren von SARS-CoV-2 wurde im Corman-Drosten-Papier wie folgt beschrieben:
„die Einrichtung und Validierung eines diagnostischen Workflows für das Screening 2019-nCoV und eine spezifische Bestätigung, die in Ermangelung verfügbarer Virusisolate oder Originalproben von Patienten entwickelt wurde. Design und Validierung wurden durch die enge genetische Verwandtheit mit dem SARS-CoV 2003 ermöglicht und durch den Einsatz synthetischer Nukleinsäuretechnologie unterstützt.“
Die Reverse Transcription-Polymerase Chain Reaction (RT-PCR) ist eine wichtige biomolekulare Technologie, um seltene RNA-Fragmente, die im Voraus bekannt sind, schnell zu erkennen. Im ersten Schritt werden die in der Probe vorhandenen RNA-Moleküle umgekehrt transkribiert, um cDNA zu ergeben. Die cDNA wird dann in der Polymerase-Kettenreaktion mit einem spezifischen Primerpaar und einem thermostabilen DNA-Polymerase-Enzym verstärkt. Die Technologie ist hochempfindlich und ihre Nachweisgrenze ist theoretisch 1 Molekül cDNA. Die Spezifität der PCR wird stark von biomolekularen Konstruktionsfehlern beeinflusst.
What is important when designing an RT-PCR Test and the quantitative RT-qPCR test described in the Corman-Drosten publication?
1. The primers and probes:
a) the concentration of primers and probes must be of optimal range
(100-200 nM)
b) must be specific to the target-gene you want to amplify
c) must have an optimal percentage of GC content relative to the total nitrogenous bases (minimum 40%, maximum 60%)
d) for virus diagnostics at least 3 primer pairs must detect 3 viral genes (preferably as far apart as possible in the viral genome)
2. The temperature at which all reactions take place:
a) DNA melting temperature (>92°)
b) DNA amplification temperature (TaqPol specific)
c) Tm; the annealing temperature (the temperature at which the primers and probes reach the target binding/detachment, not to exceed 2 ̊C per primer pair). Tm heavily depends on GC content of the primers
3. The number of amplification cycles (less than 35; preferably 25-30 cycles);
In case of virus detection, >35 cycles only detects signals which do not correlate with infectious virus as determined by isolation in cell culture [reviewed in 2]; if someone is tested by PCR as positive when a threshold of 35 cycles or higher is used (as is the case in most laboratories in Europe & the US), the probability that said person is actually infected is less than 3%, the probability that said result is a false positive is 97% [reviewed in 3]
4. Molecular biological validations; amplified PCR products must be validated either by running the products in a gel with a DNA ruler, or by direct DNA sequencing
5. Positive and negative controls should be specified to confirm/refute specific virus detection
6. There should be a Standard Operational Procedure (SOP) available
SOP unequivocally specifies the above parameters, so that all laboratories are able to set up the exact same test conditions. To have a validated universal SOP is essential, because it enables the comparison of data within and between countries.
MINOR CONCERNS WITH THE CORMAN-DROSTEN PAPER
1. In Table 1 of the Corman-Drosten paper, different abbreviations are stated – “nM” is specified, “nm” isn’t. Further in regards to correct nomenclature, nm means “nanometer” therefore nm should read nM here.
2. It is the general consensus to write genetic sequences always in the 5’-3’ direction, including the reverse primers. It is highly unusual to do alignment with reverse complementary writing of the primer sequence as the authors did in figure 2 of the Corman-Drosten paper. Here, in addition, a wobble base is marked as “y” without description of the bases the Y stands for.
3. Two misleading pitfalls in the Corman-Drosten paper are that their Table 1 does not include Tm-values (annealing-temperature values), neither does it show GC-values (number of G and C in the sequences as %-value of total bases).
MAJOR CONCERNS WITH THE CORMAN-DROSTEN PAPER
A) BACKGROUND
The authors introduce the background for their scientific work as: “The ongoing outbreak of the recently emerged novel coronavirus (2019-nCoV) poses a challenge for public health laboratories as virus isolates are unavailable while there is growing evidence that the outbreak is more widespread than initially thought, and international spread through travelers does already occur”.
According to BBC News [4] and Google Statistics [5] there were 6 deaths world-wide on January 21st 2020 – the day when the manuscript was submitted. Why did the authors assume a challenge for public health laboratories while there was no substantial evidence at that time to indicate that the outbreak was more widespread than initially thought?
As an aim the authors declared to develop and deploy robust diagnostic methodology for use in public health laboratory settings without having virus material available. Further, they acknowledge that “The present study demonstrates the enormous response capacity achieved through coordination of academic and public laboratories in national and European research networks.”
B) METHODS AND RESULTS
1. Primer & Probe Design
1a) Erroneous primer concentrations
Reliable and accurate PCR-test protocols are normally designed using between 100 nM and 200 nM per primer [7]. In the Corman-Drosten paper, we observe unusually high and varying primer concentrations for several primers (table 1). For the RdRp_SARSr-F and RdRp_SARSr-R primer pairs, 600 nM and 800 nM are described, respectively. Similarly, for the N_Sarbeco_F and N_Sarbeco_R primer set, they advise 600 nM and 800 nM, respectively [1].
It should be clear that these concentrations are far too high to be optimal for specific amplifications of target genes. There exists no specified reason to use these extremely high concentrations of primers in this protocol. Rather, these concentrations lead to increased unspecific binding and PCR product amplification.
Table1: Primers and probes (adapted from Corman-Drosten paper; erroneous primer concentrations are highlighted)
1b) Unspecified (“Wobbly”) primer and probe sequences
To obtain reproducible and comparable results, it is essential to distinctively define the primer pairs. In the Corman-Drosten paper we observed six unspecified positions, indicated by the letters R, W, M and S (Table 2). The letter W means that at this position there can be either an A or a T; R signifies there can be either a G or an A; M indicates that the position may either be an A or a C; the letter S indicates there can be either a G or a C on this position.
This high number of variants not only is unusual, but it also is highly confusing for laboratories. These six unspecified positions could easily result in the design of several different alternative primer sequences which do not relate to SARS-CoV-2 (2 distinct RdRp_SARSr_F primers + 8 distinct RdRp_SARS_P1 probes + 4 distinct RdRp_SARSr_R). The design variations will inevitably lead to results that are not even SARS CoV-2 related. Therefore, the confusing unspecific description in the Corman-Drosten paper is not suitable as a Standard Operational Protocol. These unspecified positions should have been designed unequivocally.
These wobbly sequences have already created a source of concern in the field and resulted in a Letter to the Editor authored by Pillonel et al. [8] regarding blatant errors in the described sequences. These errors are self-evident in the Corman et al. supplement as well.
Table 2: Primers and probes (adapted from Corman-Drosten paper; unspecified (“Wobbly”) nucleotides in the primers are highlighted)
The WHO-protocol (Figure 1), which directly derives from the Corman-Drosten paper, concludes that in order to confirm the presence of SARS-CoV-2, two control genes (the E-and the RdRp-genes) must be identified in the assay. It should be noted, that the RdPd-gene has one uncertain position (“wobbly”) in the forward-primer (R=G/A), two uncertain positions in the reverse-primer (R=G/A; S=G/C) and it has three uncertain positions in the RdRp-probe (W=A/T; R=G/A; M=A/C). So, two different forward primers, four different reverse primers, and eight distinct probes can be synthesized for the RdPd-gene. Together, there are 64 possible combinations of primers and probes!
The Corman-Drosten paper further identifies a third gene which, according to the WHO protocol, was not further validated and deemed unnecessary:
“Of note, the N gene assay also performed well but was not subjected to intensive further validation because it was slightly less sensitive.”
This was an unfortunate omission as it would be best to use all three gene PCRs as confirmatory assays, and this would have resulted in an almost sufficient virus RNA detection diagnostic tool protocol. Three confirmatory assay-steps would at least minimize-out errors & uncertainties at every fold-step in regards to “Wobbly”-spots. (Nonetheless, the protocol would still fall short of any “good laboratory practice”, when factoring in all the other design-errors).
As it stands, the N gene assay is regrettably neither proposed in the WHO-recommendation (Figure 1) as a mandatory and crucial third confirmatory step, nor is it emphasized in the Corman-Drosten paper as important optional reassurance “for a routine workflow” (Table 2).
Consequently, in nearly all test procedures worldwide, merely 2 primer matches were used instead of all three. This oversight renders the entire test-protocol useless with regards to delivering accurate test-results of real significance in an ongoing pandemic.
Figure 1: The N-Gene confirmatory-assay is neither emphasized as necessary third step in the official WHO Drosten-Corman protocol-recommendation below [8] nor is it required as a crucial step for higher test-accuracy in the Eurosurveillance publication.
1c) Erroneous GC-content (discussed in 2c, together with annealing temperature (Tm))
1d) Detection of viral genes
RT-PCR is not recommended for primary diagnostics of infection. This is why the RT-PCR Test used in clinical routine for detection of COVID-19 is not indicated for COVID-19 diagnosis on a regulatory basis.
“Clinicians need to recognize the enhanced accuracy and speed of the molecular diagnostic techniques for the diagnosis of infections, but also to understand their limitations. Laboratory results should always be interpreted in the context of the clinical presentation of the patient, and appropriate site, quality, and timing of specimen collection are required for reliable test results”. [9]
However, it may be used to help the physician’s differential diagnosis when he or she has to discriminate between different infections of the lung (Flu, Covid-19 and SARS have very similar symptoms). For a confirmative diagnosis of a specific virus, at least 3 specific primer pairs must be applied to detect 3 virus-specific genes. Preferably, these target genes should be located with the greatest distance possible in the viral genome (opposite ends included).
Although the Corman-Drosten paper describes 3 primers, these primers only cover roughly half of the virus’ genome. This is another factor that decreases specificity for detection of intact COVID-19 virus RNA and increases the quote of false positive test results.
Therefore, even if we obtain three positive signals (i.e. the three primer pairs give 3 different amplification products) in a sample, this does not prove the presence of a virus. A better primer design would have terminal primers on both ends of the viral genome. This is because the whole viral genome would be covered and three positive signals can better discriminate between a complete (and thus potentially infectious) virus and fragmented viral genomes (without infectious potency). In order to infer anything of significance about the infectivity of the virus, the Orf1 gene, which encodes the essential replicase enzyme of SARS-CoV viruses, should have been included as a target (Figure 2). The positioning of the targets in the region of the viral genome that is most heavily and variably transcribed is another weakness of the protocol.
Kim et al. demonstrate a highly variable 3’ expression of subgenomic RNA in Sars-CoV-2 [23]. These RNAs are actively monitored as signatures for asymptomatic and non-infectious patients [10]. It is highly questionable to screen a population of asymptomatic people with qPCR primers that have 6 base pairs primer-dimer on the 3 prime end of a primer (Figure 3).
Apparently the WHO recommends these primers. We tested all the wobble derivatives from the Corman-Drosten paper with Thermofisher’s primer dimer web tool [11]. The RdRp forward primer has 6bp 3prime homology with Sarbeco E Reverse. At high primer concentrations this is enough to create inaccuracies.
Of note: There is a perfect match of one of the N primers to a clinical pathogen (Pantoea), found in immuno-compromised patients. The reverse primer hits Pantoea as well but not in the same region (Figure 3).
These are severe design errors, since the test cannot discriminate between the whole virus and viral fragments. The test cannot be used as a diagnostic for SARS-viruses.
Figure 2: Relative positions of amplicon targets on the SARS coronavirus and the 2019 novel coronavirus genome. ORF: open reading frame; RdRp: RNA-dependent RNA polymerase. Numbers below amplicon are genome positions according to SARS-CoV, NC_004718 [1];
Figure 3: A test with Thermofischer’s primer dimer web tool reveals that the RdRp forward primer has a 6bp 3`prime homology with Sarbeco E Reverse (left box). Another test reveals that there is a perfect match for one of the N-primers to a clinical pathogen (Pantoea) found in immuno-compromised patients (right box).
2. Reaction temperatures
2a) DNA melting temperature (>92°).
Adequately addressed in the Corman-Drosten paper.
2b) DNA amplification temperature.
Adequately addressed in the Corman-Drosten paper.
2c) Erroneous GC-contents and Tm
The annealing-temperature determines at which temperature the primer attaches/detaches from the target sequence. For an efficient and specific amplification, GC content of primers should meet a minimum of 40% and a maximum of 60% amplification. As indicated in table 3, three of the primers described in the Corman-Drosten paper are not within the normal range for GC-content. Two primers (RdRp_SARSr_F and RdRp_SARSr_R) have unusual and very low GC-values of 28%-31% for all possible variants of wobble bases, whereas primer E_Sarbeco_F has a GC-value of 34.6% (Table 3 and second panel of Table 3).
It should be noted that the GC-content largely determines the binding to its specific target due to its three hydrogen bonds in base pairing. Thus, the lower the GC-content of the primer, the lower its binding-capability to its specific target gene sequence (i.e. the gene to be detected). This means for a target-sequence to be recognized we have to choose a temperature which is as close as possible to the actual annealing-temperature (best practise-value) for the primer not to detach again, while at the same time specifically selecting the target sequence.
If the Tm-value is very low, as observed for all wobbly-variants of the RdRp reverse primers, the primers can bind non-specifically to several targets, decreasing specificity and increasing potential false positive results.
The annealing temperature (Tm) is a crucial factor for the determination of the specificity/accuracy of the qPCR procedure and essential for evaluating the accuracy of qPCR-protocols. Best-practice recommendation: Both primers (forward and reverse) should have an almost similar value, preferably the identical value.
We used the freely available primer design software Primer-BLAST [12, 25] to evaluable the best-practise values for all primers used in the Corman-Drosten paper (Table 3). We attempted to find a Tm-value of 60° C, while similarly seeking the highest possible GC%-value for all primers. A maximal Tm difference of 2° C within primer pairs was considered acceptable. Testing the primer pairs specified in the Corman-Drosten paper, we observed a difference of 10° C with respect to the annealing temperature Tm for primer pair1 (RdRp_SARSr_F and RdRp_SARSr_R). This is a very serious error and makes the protocol useless as a specific diagnostic tool.
Additional testing demonstrated that only the primer pair designed to amplify the N-gene (N_Sarbeco_F and N_Sarbeco_R) reached the adequate standard to operate in a diagnostic test, since it has a sufficient GC-content and the Tm difference between the primers (N_Sarbeco_F and N_Sarbeco_R) is 1.85° C (below the crucial maximum of 2° C difference). Importantly, this is the gene which was neither tested in the virus samples (Table 2) nor emphasized as a confirmatory test. In addition to highly variable melting temperatures and degenerate sequences in these primers, there is another factor impacting specificity of the procedure: the dNTPs (0.4uM) are 2x higher than recommended for a highly specific amplification. There is additional magnesium sulphate added to the reaction as well. This procedure combined with a low annealing temperature can create non-specific amplifications. When additional magnesium is required for qPCR, specificity of the assay should be further scrutinized.
The design errors described here are so severe that it is highly unlikely that specific amplification of SARS-CoV-2 genetic material will occur using the protocol of the Corman-Drosten paper.
Table 3: GC-content of the primers and probes (adapted from Corman-Drosten paper; aberrations from optimized GC-contents are highlighted. Second Panel shows a table-listing of all Primer-BLAST best practices values for all primers and probes used in the Corman-Drosten paper by Prof. Dr. Ulrike Kämmerer & her team
3. The number of amplification cycles
It should be noted that there is no mention anywhere in the Corman-Drosten paper of a test being positive or negative, or indeed what defines a positive or negative result. These types of virological diagnostic tests must be based on a SOP, including a validated and fixed number of PCR cycles (Ct value) after which a sample is deemed positive or negative. The maximum reasonably reliable Ct value is 30 cycles. Above a Ct of 35 cycles, rapidly increasing numbers of false positives must be expected .
PCR data evaluated as positive after a Ct value of 35 cycles are completely unreliable.
Citing Jaafar et al. 2020 [3]: “At Ct = 35, the value we used to report a positive result for PCR, <3% of cultures are positive.” In other words, there was no successful virus isolation of SARS-CoV-2 at those high Ct values.
Further, scientific studies show that only non-infectious (dead) viruses are detected with Ct values of 35 [22].
Between 30 and 35 there is a grey area, where a positive test cannot be established with certainty. This area should be excluded. Of course, one could perform 45 PCR cycles, as recommended in the Corman-Drosten WHO-protocol (Figure 4), but then you also have to define a reasonable Ct-value (which should not exceed 30). But an analytical result with a Ct value of 45 is scientifically and diagnostically absolutely meaningless (a reasonable Ct-value should not exceed 30). All this should be communicated very clearly. It is a significant mistake that the Corman-Drosten paper does not mention the maximum Ct value at which a sample can be unambiguously considered as a positive or a negative test-result. This important cycle threshold limit is also not specified in any follow-up submissions to date.
Figure 4: RT-PCR Kit recommendation in the official Corman-Drosten WHO-protocol [8]. Only a “Cycler”-value (cycles) is to be found without corresponding and scientifically reasonable Ct (Cutoff-value). This or any other cycles-value is nowhere to be found in the actual Corman-Drosten paper.
4. Biomolecular validations
To determine whether the amplified products are indeed SARS-CoV-2 genes, biomolecular validation of amplified PCR products is essential. For a diagnostic test, this validation is an absolute must.
Validation of PCR products should be performed by either running the PCR product in a 1% agarose-EtBr gel together with a size indicator (DNA ruler or DNA ladder) so that the size of the product can be estimated. The size must correspond to the calculated size of the amplification product. But it is even better to sequence the amplification product. The latter will give 100% certainty about the identity of the amplification product. Without molecular validation one can not be sure about the identity of the amplified PCR products. Considering the severe design errors described earlier, the amplified PCR products can be anything.
Also not mentioned in the Corman-Drosten paper is the case of small fragments of qPCR (around 100bp): It could be either 1,5% agarose gel or even an acrylamide gel.
The fact that these PCR products have not been validated at molecular level is another striking error of the protocol, making any test based upon it useless as a specific diagnostic tool to identify the SARS-CoV-2 virus.
5. Positive and negative controls to confirm/refute specific virus detection.
The unconfirmed assumption described in the Corman-Drosten paper is that SARS-CoV-2 is the only virus from the SARS-like beta-coronavirus group that currently causes infections in humans. The sequences on which their PCR method is based are in silico sequences, supplied by a laboratory in China [23], because at the time of development of the PCR test no control material of infectious (“live”) or inactivated SARS-CoV-2 was available to the authors. The PCR test was therefore designed using the sequence of the known SARS-CoV as a control material for the Sarbeco component (Dr. Meijer, co-author Corman-Drosten paper in an email exchange with Dr. Peter Borger) [2].
All individuals testing positive with the RT-PCR test, as described in the Corman-Drosten paper, are assumed to be positive for SARS-CoV-2 infections. There are three severe flaws in their assumption. First, a positive test for the RNA molecules described in the Corman-Drosten paper cannot be equated to “infection with a virus”. A positive RT-PCR test merely indicates the presence of viral RNA molecules. As demonstrated under point 1d (above), the Corman-Drosten test was not designed to detect the full-length virus, but only a fragment of the virus. We already concluded that this classifies the test as unsuitable as a diagnostic test
for SARS-virus infections.
Secondly and of major relevance, the functionality of the published RT-PCR Test was not demonstrated with the use of a positive control (isolated SARS-CoV-2 RNA) which is an essential scientific gold standard.
Third, the Corman-Drosten paper states:
“To show that the assays can detect other bat-associated SARS-related viruses, we used the E gene assay to test six bat-derived faecal samples available from Drexler et al. […] und Muth et al. […]. These virus-positive samples stemmed from European rhinolophid bats. Detection of these phylogenetic outliers within the SARS-related CoV clade suggests that all Asian viruses are likely to be detected. This would, theoretically, ensure broad sensitivity even in case of multiple independent acquisitions of variant viruses from an animal reservoir.”
This statement demonstrates that the E gene used in RT-PCR test, as described in the Corman-Drosten paper, is not specific to SARS-CoV-2.
The E gene primers also detect a broad spectrum of other SARS viruses.
The genome of the coronavirus is the largest of all RNA viruses that infect humans and they all have a very similar molecular structure. Still, SARS-CoV1 and SARS-CoV-2 have two highly specific genetic fingerprints, which set them apart from the other coronaviruses. First, a unique fingerprint-sequence (KTFPPTEPKKDKKKK) is present in the N-protein of SARS-CoV and SARS-CoV-2 [13,14,15]. Second, both SARS-CoV1 and SARS-CoV2 do not contain the HE protein, whereas all other coronaviruses possess this gene [13, 14]. So, in order to specifically detect a SARS-CoV1 and SARS-CoV-2 PCR product the above region in the N gene should have been chosen as the amplification target. A reliable diagnostic test should focus on this specific region in the N gene as a confirmatory test. The PCR for this N gene was not further validated nor recommended as a test gene by the Drosten-Corman paper, because of being “not so sensitive” with the SARS-CoV original probe [1].
Furthermore, the absence of the HE gene in both SARS-CoV1 and SARS-CoV-2 makes this gene the ideal negative control to exclude other coronaviruses. The Corman-Drosten paper does not contain this negative control, nor does it contain any other negative controls. The PCR test in the Corman-Drosten paper therefore contains neither a unique positive control nor a negative control to exclude the presence of other coronaviruses. This is another major design flaw which classifies the test as unsuitable for diagnosis.
6. Standard Operational Procedure (SOP) is not available
There should be a Standard Operational Procedure (SOP) available, which unequivocally specifies the above parameters, so that all laboratories are able to set up the identical same test conditions. To have a validated universal SOP is essential, because it facilitates data comparison within and between countries. It is very important to specify all primer parameters unequivocally. We note that this has not been done. Further, the Ct value to indicate when a sample should be considered positive or negative is not specified. It is also not specified when a sample is considered infected with SARS-CoV viruses. As shown above, the test cannot discern between virus and virus fragments, so the Ct value indicating positivity is crucially important. This Ct value should have been specified in the Standard Operational Procedure (SOP) and put on-line so that all laboratories carrying out this test have exactly the same boundary conditions. It points to flawed science that such an SOP does not exist. The laboratories are thus free to conduct the test as they consider appropriate, resulting in an enormous amount of variation. Laboratories all over Europe are left with a multitude of questions; which primers to order? which nucleotides to fill in the undefined places? which Tm value to choose? How many PCR cycles to run? At what Ct value is the sample positive? And when is it negative? And how many genes to test? Should all genes be tested, or just the E and RpRd gene as shown in Table 2 of the Corman-Drosten paper? Should the N gene be tested as well? And what is their negative control? What is their positive control?
The protocol as described is unfortunately very vague and erroneous in its design that one can go in dozens of different directions. There does not appear to be any standardization nor an SOP, so it is not clear how this test can be implemented.
7. Consequences of the errors described under 1-5: false positive results.
The RT-PCR test described in the Corman-Drosten paper contains so many molecular biological design errors (see 1-5) that it is not possible to obtain unambiguous results. It is inevitable that this test will generate a tremendous number of so-called “false positives”. The definition of false positives is a negative sample, which initially scores positive, but which is negative after retesting with the same test. False positives are erroneous positive test-results, i.e. negative samples that test positive. And this is indeed what is found in the Corman-Drosten paper. On page 6 of the manuscript PDF the authors demonstrate, that even under well-controlled laboratory conditions, a considerable percentage of false positives is generated with this test:
“In four individual test reactions, weak initial reactivity was seen however they were negative upon retesting with the same assay. These signals were not associated with any particular virus, and for each virus with which initial positive reactivity occurred, there were other samples that contained the same virus at a higher concentration but did not test positive. Given the results from the extensive technical qualification described above, it was concluded that this initial reactivity was not due to chemical instability of real-time PCR probes and most probably to handling issues caused by the rapid introduction of new diagnostic tests and controls during this evaluation study.” [1]
The first sentence of this excerpt is clear evidence that the PCR test described in the Corman-Drosten paper generates false positives. Even under the well-controlled conditions of the state-of-the-art Charité-laboratory, 4 out of 310 primary-tests are false positives per definition. Four negative samples initially tested positive, then were negative upon retesting. This is the classical example of a false positive. In this case the authors do not identify them as false positives, which is intellectually dishonest.
Another telltale observation in the excerpt above is that the authors explain the false positives away as “handling issues caused by the rapid introduction of new diagnostic tests”. Imagine the laboratories that have to introduce the test without all the necessary information normally described in an SOP.
8. The Corman-Drosten paper was not peer-reviewed
Before formal publication in a scholarly journal, scientific and medical articles are traditionally certified by “peer review.” In this process, the journal’s editors take advice from various experts (“referees”) who have assessed the paper and may identify weaknesses in its assumptions, methods, and conclusions. Typically a journal will only publish an article once the editors are satisfied that the authors have addressed referees’ concerns and that the data presented supports the conclusions drawn in the paper.” This process is as well described for Eurosurveillance [16].
The Corman-Drosten paper was submitted to Eurosurveillance on January 21st 2020 and accepted for publication on January 22nd 2020. On January 23rd 2020 the paper was online. On January 13th 2020 version 1-0 of the protocol was published at the official WHO website [17], updated on January 17th 2020 as document version 2-1 [18], even before the Corman-Drosten paper was published on January 23rd at Eurosurveillance.
Normally, peer review is a time-consuming process since at least two experts from the field have to critically read and comment on the submitted paper. In our opinion, this paper was not peer-reviewed. Twenty-four hours are simply not enough to carry out a thorough peer review. Our conclusion is supported by the fact that a tremendous number of very serious design flaws were found by us, which make the PCR test completely unsuitable as a diagnostic tool to identify the SARS-CoV-2 virus. Any molecular biologist familiar with RT-PCR design would have easily observed the grave errors present in the Corman-Drosten paper before the actual review process. We asked Eurosurveillance on October 26th 2020 to send us a copy of the peer review report. To date, we have not received this report and in a letter dated November 18th 2020, the ECDC as host for Eurosurveillance declined to provide access without providing substantial scientific reasons for their decision. On the contrary, they write that “disclosure would undermine the purpose of scientific investigations.” [24].
9. Authors as the editors
A final point is one of major concern. It turns out that two authors of the Corman-Drosten paper, Christian Drosten and Chantal Reusken, are also members of the editorial board of this journal [19]. Hence there is a severe conflict of interest which strengthens suspicions that the paper was not peer-reviewed. It has the appearance that the rapid publication was possible simply because the authors were also part of the editorial board at Eurosurveillance. This practice is categorized as compromising scientific integrity.
SUMMARY CATALOGUE OF ERRORS FOUND IN THE PAPER
The Corman-Drosten paper contains the following specific errors:
1. There exists no specified reason to use these extremely high concentrations of primers in this protocol. The described concentrations lead to increased nonspecific bindings and PCR product amplifications, making the test unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus.
2. Six unspecified wobbly positions will introduce an enormous variability in the real world laboratory implementations of this test; the confusing nonspecific description in the Corman-Drosten paper is not suitable as a Standard Operational Protocol making the test unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus.
3. The test cannot discriminate between the whole virus and viral fragments. Therefore, the test cannot be used as a diagnostic for intact (infectious) viruses, making the test unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus and make inferences about the presence of an infection.
4. A difference of 10° C with respect to the annealing temperature Tm for primer pair1 (RdRp_SARSr_F and RdRp_SARSr_R) also makes the test unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus.
5. A severe error is the omission of a Ct value at which a sample is considered positive and negative. This Ct value is also not found in follow-up submissions making the test unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus.
6. The PCR products have not been validated at the molecular level. This fact makes the protocol useless as a specific diagnostic tool to identify the SARS-CoV-2 virus.
7. The PCR test contains neither a unique positive control to evaluate its specificity for SARS-CoV-2 nor a negative control to exclude the presence of other coronaviruses, making the test unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus.
8. The test design in the Corman-Drosten paper is so vague and flawed that one can go in dozens of different directions; nothing is standardized and there is no SOP. This highly questions the scientific validity of the test and makes it unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus.
9. Most likely, the Corman-Drosten paper was not peer-reviewed making the test unsuitable as a specific diagnostic tool to identify the SARS-CoV-2 virus.
10. We find severe conflicts of interest for at least four authors, in addition to the fact that two of the authors of the Corman-Drosten paper (Christian Drosten and Chantal Reusken) are members of the editorial board of Eurosurveillance. A conflict of interest was added on July 29 2020 (Olfert Landt is CEO of TIB-Molbiol; Marco Kaiser is senior researcher at GenExpress and serves as scientific advisor for TIB-Molbiol), that was not declared in the original version (and still is missing in the PubMed version); TIB-Molbiol is the company which was “the first” to produce PCR kits (Light Mix) based on the protocol published in the Corman-Drosten manuscript, and according to their own words, they distributed these PCR-test kits before the publication was even submitted [20]; further, Victor Corman & Christian Drosten failed to mention their second affiliation: the commercial test laboratory “Labor Berlin”. Both are responsible for the virus diagnostics there [21] and the company operates in the realm of real time PCR-testing.
In light of our re-examination of the test protocol to identify SARS-CoV-2 described in the Corman-Drosten paper we have identified concerning errors and inherent fallacies which render the SARS-CoV-2 PCR test useless.
CONCLUSION
The decision as to which test protocols are published and made widely available lies squarely in the hands of Eurosurveillance. A decision to recognise the errors apparent in the Corman-Drosten paper has the benefit to greatly minimise human cost and suffering going forward.
Is it not in the best interest of Eurosurveillance to retract this paper? Our conclusion is clear. In the face of all the tremendous PCR-protocol design flaws and errors described here, we conclude: There is not much of a choice left in the framework of scientific integrity and responsibility.
REFERENCES
[1] Corman Victor M, Landt Olfert, Kaiser Marco, Molenkamp Richard, Meijer Adam, Chu Daniel KW, Bleicker Tobias, Brünink Sebastian, Schneider Julia, Schmidt Marie Luisa, Mulders Daphne GJC, Haagmans Bart L, van der Veer Bas, van den Brink Sharon, Wijsman Lisa, Goderski Gabriel, Romette Jean-Louis, Ellis Joanna, Zambon Maria, Peiris Malik, Goossens Herman, Reusken Chantal, Koopmans Marion PG, Drosten Christian. Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR. Euro Surveill. 2020;25(3):pii=2000045. https://doi.org/10.2807/1560-7917.ES.2020.25.3.2000045
[2] Email communication between Dr. Peter Borger & Dr. Adam Meijer: Supplementary Material
[3] Jafaar et al., Correlation Between 3790 Quantitative Polymerase Chain Reaction–Positives Samples and Positive Cell Cultures, Including 1941 Severe Acute Respiratory Syndrome Coronavirus 2 Isolates. https://academic.oup.com/cid/advance-article/doi/10.1093/cid/ciaa1491/5912603
[4] BBC, January 21st 2020: https://www.bbc.com/news/world-asia-china-51185836;
Archive: https://archive.is/0qRmZ
[5] Google Analytics – COVID19-deaths worldwide: https://bit.ly/3fndemJ
Archive: https://archive.is/PpqEE
[6] Laboratory testing for COVID-19 Emergency Response Technical Centre, NIVD under
China CDC March 15th, 2020: http://www.chinacdc.cn/en/COVID19/202003/P020200323390321297894.pdf
[7] Real-Time PCR Handbook Life Technologies: https://www.thermofisher.com/content/dam/LifeTech/global/Forms/PDF/real-time-pcr-
handbook.pdf
Nolan T, Huggett J, Sanchez E.Good practice guide for the application of quantitative PCR (qPCR) First Edition 2013
[8] Trestan Pillonel et al, Letter to the editor: SARS-CoV-2 detection by real-time RT-PCR: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7268274/
[9] Kurkela, Satu, and David WG Brown. “Molecular-diagnostic techniques.” Medicine 38.10
(2009): 535-540.
[10] Wolfel et al., Virological assessment of hospitalized patients with COVID-2019
https://www.nature.com/articles/s41586-020-2196-x
[11] Thermofischer Primer Dimer Web Tool: https://www.thermofisher.com/us/en/home/brands/thermo-scientific/molecular-biology/molecular-biology-learning-center/molecular-biology-resource-library/thermo-scientific-web-tools/multiple-primer-analyzer.html
Supplementary Material
[12] Primer-BLAST, NCBI – National Center for Biotechnology Information: https://www.ncbi.nlm.nih.gov/tools/primer-blast/
[13] Marra MA, Steven JMJ, Caroline RA, Robert AH, Angela BW et al. (2003) Science. The
Genome sequence of the SARS-associated coronavirus. Science 300(5624): 1399-1404.
[14] Severe acute respiratory syndrome coronavirus 2 isolate Wuhan-Hu-1, complete
genome: https://www.ncbi.nlm.nih.gov/nuccore/MN908947
[15] Borger P. A SARS-like Coronavirus was expected but nothing was done to be prepared. Am J Biomed Sci Res 2020. https://biomedgrid.com/pdf/AJBSR.MS.ID.001312.pdf
https://www.researchgate.net/publication/341120750_A_SARS-
like_Coronavirus_was_Expected_but_nothing_was_done_to_be_Prepared;
Archive: https://archive.is/i76Hu
[16] Eurosurveillance paper evaluation / review process: https://www.eurosurveillance.org/evaluation
[17] Official recommendation of the Corman-Drosten protocol & manuscript by the WHO,published on January 13th 2020 as version 1.0 of the document:
https://www.who.int/docs/default-source/coronaviruse/wuhan-virus-assay-
v1991527e5122341d99287a1b17c111902.pdf; archive: https://bit.ly/3m3jXVH
[18] Official WHO-recommendation for the Corman / Drosten RT-qPCR-protocol, which
directly derives from the Eurosurveillance-publication, document-version 2-1, published on
17th January 2020: https://www.who.int/docs/default-source/coronaviruse/protocol-v2-
1.pdf?sfvrsn=a9ef618c_2
[19] Eurosurveillance Editorial Board, 2020: https://www.eurosurveillance.org/upload/site-
assets/imgs/2020-09-Editorial%20Board%20PDF.pdf;
Archive: https://bit.ly/2TqXBjX
[20] Instructions For Use LightMix SarbecoV E-gene plus EAV Control, TIB-Molbiol & Roche
Molecular Solutions, January 11th 2020: https://www.roche-as.es/lm_pdf/MDx_40-0776_96_Sarbeco-E-
gene_V200204_09164154001 (1).pdf
Archive, timestamp – January 11th 2020: https://archive.is/Vulo5;
Archive: https://bit.ly/3fm9bXH
[21] Christian Drosten & Victor Corman, responsible for viral diagnostics at Labor Berlin:
https://www.laborberlin.com/fachbereiche/virologie/
Archive: https://archive.is/CDEUG
[22] Tom Jefferson, Elizabeth Spencer, Jon Brassey, Carl Heneghan Viral cultures for COVID-
19 infectivity assessment. Systematic review. Systematic review doi:
https://doi.org/10.1101/2020.08.04.20167932 https://www.medrxiv.org/content/10.1101/2020.08.04.20167932v4
[23] Kim et al.,The Architecture of SARS-CoV-2 Transcriptome:
https://www.sciencedirect.com/science/article/pii/S0092867420304062
[24] ECDC reply to Dr. Peter Borger, 18th November 2020:
Supplementary Material
[25] Prof. Dr. Ulrike Kämmerer & team, survey & Primer-BLAST table:
Supplementary Material
Additional literature:
Description RT-PCR RKI Germany, on page 10 of this link:
https://www.rki.de/DE/Content/Gesundheitsmonitoring/Gesundheitsberichterstattung/GBE
DownloadsJ/JoHM_S5_2020_Studienprotokoll_CORONA_MONITORING_lokal.pdf?__blob=p
ublicationFile
Author’s Affiliations:
1) Dr. Pieter Borger (MSc, PhD), Molecular Genetics, W+W Research Associate, Lörrach, Germany
2) Rajesh Kumar Malhotra (Artist Alias: Bobby Rajesh Malhotra), Former 3D Artist / Scientific Visualizations at CeMM – Center for Molecular Medicine of the Austrian Academy of Sciences (2019-2020), University for Applied Arts – Department for Digital Arts Vienna, Austria
3) Dr. Michael Yeadon BSs(Hons) Biochem Tox U Surrey, PhD Pharmacology U Surrey. Managing Director, Yeadon Consulting Ltd, former Pfizer Chief Scientist, United Kingdom
4) Dr. Clare Craig MA, (Cantab) BM, BCh (Oxon), FRCPath, United Kingdom
5) Kevin McKernan, BS Emory University, Chief Scientific Officer, founder Medical Genomics, engineered the sequencing pipeline at WIBR/MIT for the Human Genome Project, Invented and developed the SOLiD sequencer, awarded patents related to PCR, DNA Isolation and Sequencing, USA
6) Prof. Dr. Klaus Steger, Department of Urology, Pediatric Urology and Andrology, Molecular Andrology, Biomedical Research Center of the Justus Liebig University, Giessen, Germany
7) Dr. Paul McSheehy (BSc, PhD), Biochemist & Industry Pharmacologist, Loerrach, Germany
8) Dr. Lidiya Angelova, MSc in Biology, PhD in Microbiology, Former researcher at the National Institute of Allergy and Infectious Diseases (NIAID), Maryland, USA
9) Dr. Fabio Franchi, Former Dirigente Medico (M.D) in an Infectious Disease Ward, specialized in “Infectious Diseases” and “Hygiene and Preventive Medicine”, Società Scientifica per il Principio di Precauzione (SSPP), Italy
10) Dr. med. Thomas Binder, Internist and Cardiologist (FMH), Switzerland
11) Prof. Dr. med. Henrik Ullrich, specialist Diagnostic Radiology, Chief Medical Doctor at the Center for Radiology of Collm Oschatz-Hospital, Germany
12) Prof. Dr. Makoto Ohashi, Professor emeritus, PhD in Microbiology and Immunology, Tokushima University, Japan
13) Dr. Stefano Scoglio, B.Sc. Ph.D., Microbiologist, Nutritionist, Italy
14) Dr. Marjolein Doesburg-van Kleffens (MSc, PhD), specialist in Laboratory Medicine (clinical chemistry), Maasziekenhuis Pantein, Beugen, The Netherlands
15) Dr. Dorothea Gilbert (MSc, PhD), PhD Environmental Chemistry and Toxicology. DGI Consulting Services, Oslo, Norway
16) Dr. Rainer J. Klement, PhD. Department of Radiation Oncology, Leopoldina Hospital Schweinfurt, Germany
17) Dr. Ruth Schruefer, PhD, human genetics/ immunology, Munich, Germany,
18) Dra. Berber W. Pieksma, General Practitioner, The Netherlands
19) Dr. med. Jan Bonte (GJ), Consultant Neurologist, The Netherlands
20) Dr. Bruno H. Dalle Carbonare (Molecular biologist), IP specialist, BDC Basel, Switzerland
21) Dr. Kevin P. Corbett, MSc Nursing (Kings College London) PhD (London South Bank) Social Sciences (Science & Technology Studies) London, England, United Kingdom
22) Prof. Dr. Ulrike Kämmerer, specialist in Virology / Immunology / Human Biology / Cell Biology, University Hospital Würzburg, Germany
Author’s Contributions:
PB: Planned and conducted the analyses and research, conceptualising the manuscript.
BRM: Planned and conducted the research, conceptualising the figures and manuscript.
MY: Proofreading the analyses and research.
KMcK: Conducted the analyses and research, conceptualized the manuscript.
KS: Conducted the analyses and research.
PMcS: Proofreading the analyses and research.
LA: Proofreading the analyses and research.
FF: Proofreading the analyses and research.
TB: Proofreading the analyses and research.
HU: Proofreading the analyses and research.
MO: Proofreading the analyses and research.
SS: Proofreading the analyses and research.
MDvK: Proofreading the analyses and research.
DG: Proofreading the analyses and research.
RJK: Proofreading the analyses and research.
RS: Proofreading the analyses and research, and the manuscript.
BWK: Proofreading the analyses and research.
RvV: Proofreading the analyses and research.
JB: Proofreading the analyses and research.
KC: Proofreading the analyses and research.
UK: Planned and conducted the analyses and research, conceptualising the manuscript.
Additional Proof-Readers:
Saji N Hameed, Environmental Informatics, University of Aizu, Tsuruga, Ikki-machi, Aizuwakamatsu-shi, Fukushima, Japan
Howard R. Steen, MA Chem. Eng. Cantab, Former Research Manager, Germany
Addendum
Update 2.12.2020:
Author Contribution Dr. Michael Yeadon changed to:
Proofreading the analyses and research.
Author Affiliation Kevin Mckernan changed to:
Medicinal Genomics.next »
Comments
Sebe Vpgel says:Thanks for your excellent work!Reply
robvdz says:Beside all of this, I won’t let somebody that is not medically schooled fidle around in my nose cavities for what ever reason.Reply
Bayaba says:I won’t let someone who is medically trained do that. That includes my own brother, who is an MD and who is the president and CEO of the family practice he runs.Reply
Madaload says:Unless, of course, the government makes testing mandatory. Slovakia already had a few rounds of full scale (all citizens) AG testing. It wasn’t mandatory per se. But if you didn’t get tested, you didn’t get a certificate and had to self isolate for 10 days. If you didn’t get tested in the second round, same thing with 14 days isolation and your employer wouldn’t let you work without the cert. After serious protests, the government now says AG positive results will be validated by PCR tests and 3 rounds of this “non mandatory” testing by Christmas.Reply
Lieveke says:How sad ! Will be with Slovakia in mind…Reply
Dr. med. dent. Klaus Wilhelm Rocholl says:Congratulation – and my deepest and most sincere admiration for your impressive work.
I hope you maybe literally helped to save the world!ReplyWim Sturm says:Thank you for your great work!Facts outweigh fiction and open people’s eyes to this ridiculous fictional reality that has been created in the world based on the Corman Drosten paper.Thank you again for distinguishing fiction from reality with you retraction paper.Reply
J. says:Great job! Heroes of the universe!!!Reply
LIVIANA* says:Thank you
Dankuwel
Obrigado
Merci
Vielen dank
Und viel ErfolgReplyDUC says:Thanks for publishing what I have been saying in essence all along (but sure not in such detail). Lets hope there are consequences which are in balance with the damage done to the world population.M.sc., D.sc., former researcher in molecular genetics, HIV, immunology, among others 6 y at NIH(USA)Reply
Liam King says:Glad to hear you’ve been saying this all along.Where were you shouting about this when you couldn’t be heard?Let me know so I can warn others to stay clear of such ineffectual places (and I will of course point them here instead).Reply
D. Krüger says:Ein wundervoller Hoffnungsschimmer am Horizont der dem ganzen Irrsinn hoffentlich ein baldiges Ende setzt bevor noch mehr unnötiges Leid und Elend verursacht wird.Herzlichen Dank, für Ihre wissenschaftliche Integrität die einigen Ihrer Kollegen offenbar abhanden gekommen ist !Reply
Hoijtink says:Good to see that at least some scientists still use common sense and brains. In my book you are heroes. Sadly it remains to be seen what politicians will do next, they have their own (hidden?) agenda.Thank you all ….Reply
Solveig Warren says:Thank you for telling the truth in such a professional manner and using science appropriately! You are true heroes of our Universe! It is a tough job to to stand up for honesty with the Media having a one track mind!Reply
Marlee Ponich says:Much love and gratitude!Reply
Autoglas says:Thank you for all … I hope the bestReply
Fred K. says:Many thanks for the extremely good and bitterly necessary work! I hope that this work can make a fundamental contribution to finally putting a stop to the madness. If the faulty paper is not voluntarily withdrawn by the authors, I very much hope that there is a way to force this through the courts with the help of dedicated lawyers. Thank you for your extraordinary commitment!Reply
Christoph Schmitz (Univ.Prof. Dr.med.) says:I cannot comment on PCR issues; this is not my field of expertise (I am a neuroanatomist at LMU Munich/Germany with approximately 200 papers listed in PubMed). I would just like to comment on the “points of major concern” #8 and #9 outlined above:#8: it is indeed feasible (and not unusual) to perform a scientifically sound peer-review of a manuscript within a few hours after having obtained the request by a journal, particularly if you are an expert in the field. The fact that the manuscript that is discussed here was accepted for publication one day after submission does not mean that it was not peer-reviewed.#9: every serious academic journal has an internal policy that manuscripts submitted by a member (or members) of the editorial board are handled and reviewed by other members of the editorial board of the journal. The fact that Christian Drosten has served as corresponding author of the manuscript described here does not imply that “scientific integrity was compromised”. In particular, this fact may not support the “suspicions that the paper was not peer-reviewed”.In summary, I warmly recommend to separate the “points of major concern” #8 and #9 outlined above from the other points of major concern in order to place this important discussion on a more objective footing.Reply
Dr. Frank Potthast says:In my opinion, you cannot separate the issues; if the editorial board has commmon sense, that this publication should be accepted for publication within a few hours, the scientific quality must be double-checked if you don´t want to risk accuses of wangle.
The mistakes concerning good laboratory practice are so obvious, that I cannot believe, that it wouldn´t have been noticed by any of the experts.ReplyMaritta Mathis says:With all due respect, have you not actually recognized the dimensions and scope of this scandal, that you only criticize these two points, but are silent overall about the outrageously unscientific approach (where I live this is called fraud)?Reply
PD Dr. Jörg Gerke says:Dear Dr. Schmitz,
I understand your points and find them reasonable. The scientific content should be discriminated from the “social” content.
However, a review within 24 hours is very unusual at least in the field of my experience. Instead the publication of the reviews by the reviewer of the Journal should be recommended.Reply
Evgeniy Gilenko says:Think, the scope of the comment above is restricted to these two subtopics. Even though the practice of approval processes and publishing of scientific articles relies in my opinion mainly just on scientific integrity of reviewers, which I personally would like to trust, and the with regards to the massive impact of the study – direct or not – on the economies, societies etc., I simply would assume, that at the moment of submission and publication there were no evidence of pandemic outbreak of the SARS-COV-2 and therefore no self-evident need for deeper review of the study. Shortly saying, they probably didn’t think the study would have any significant impact on anything. What I am asking myself instead, is why the first critical review appears only on 27th of November and was not conducted or initiated by the journal itself?Reply
Ralph Nimmann says:VERY good question “why the first critical review appears only on 27th of November and was not conducted or initiated by the journal itself?”
I added a summary of this peer review on
https://healthtruth.info/weve-got-it-all-wrong-fighting-the-virus/#pcrReply
Els van Veen says:Danke! Dankuwel! Thank you!Ik ben een Nederlandse huisarts en hoop dat nu snel het krankzinnige testen (buiten de kliniek) kan worden gestaakt.
De lockdowns opgeheven.
De mondkapjes weggedaan.
De wetenschap in ere hersteld.
Het recht zal winnen van het onrecht en de leugen.ReplyDipl.Psych. Hans-J. Steiner says:Interesting work, however – after all tthese hoax-reports and “scientific” looking statements of people all over the wolrd, which came finally out, not being cited corectly or just abused for never done citations or just the person was “virtual”, there will be much work to immunize that work from “Faktencheckers” and other discreditions. That will be even the more relevant work to be done urgently to make this paper an evicent part of public, scientific and political discussion!Reply
Udo says:Dear Hans,that is a very valid point. I wonder already since a long time about the “circular reasoning” in the so called “fact check”, as generally speaking they don’t make sense or the find “one specialst” who will explain it (putting a few minutes of effort in it).Reply
Bobby says:https://www.dailywire.com/news/candace-owens-challenges-fact-checker-and-winsReply
Tatjana Z. says:Danke für die Antwort! Dann hoffe ich sehr, dass ihr auch gehört werdet!Reply
Monika says:Thank You All for the great work! God bless you!Reply
Dr .Andreas Gloge says:Man muß das Wahre immer wiederholen, weil auch der Irrtum um uns her immer wieder gepredigt wird, und zwar nicht von einzelnen, sondern von der Masse. In Zeitungen und Enzyklopädien, auf Schulen und Universitäten, überall ist der Irrtum oben auf, und es ist ihm wohl und behaglich, im Gefühl der Majorität, die auf seiner Seite ist.Johann Wolfgang von GoetheReply
Gertrud Adam says:Vielen Dank, kannte ich noch gar nicht. Autorenschaft überraschte mich, als ich sie drunter entdeckte.Reply
Brigid says:Thank you for this excellent piece of work which clarifies in detail and in a factual way what is felt by many. This is the proof. My hope is that this will enlighten those who need to know.Reply
ursula b. says:The more one tries to opress truth the brighter it shines throughout the universe.
Thank you so much for your great work and dedication. With many small steps we will win the race 1nce and foreverReplySTRNTVRVLND says:Let’s hope this effort will put a end to the disproportionate mesures. Please make this understandable for all by also presenting this information on micro-level instead of moleculare.May a revolution be upon usReply
B Anderson says:So, if the test to confirm you have covid-19 is useless…what virus is the vaccine that is soon to be distributed going to prevent you from getting?Reply
Eva says:You are right. And The Expert answers: “Good question, thank you. Next question, please…”Reply
human says:Actually, we are no longer permitted to question anything.Because once questions are allowed, the answers will inevitably indict extremely powerful individuals and organizations of crimes against humanity.Reply
Angela says:Exactly. This is the more pressing question. WHAT IS IN THE VACCINE?
Judging from their behaviour so far, I don’t trust the intentions of authorities worldwide.
Next question: What else is to come?
Believe me, lockdown and other measures will look benign in comparison to what else they will unleash on us.Reply
Maria says:Thank you for doing this work and bringing real science to the table. Hoepfully this will end this epidemic of false positives which is causing so much human suffering. The cost of lockdowns: https://www.aier.org/article/cost-of-us-lockdowns-a-preliminary-report/Reply
Robert Michel says:Thank you,If you have an hammer – every thing looks like a nail….
It becomes dangerious if political leader says “we will be only rescued,
when we would have that vacination” and Media is following narrowminded.We are spending Billions just on test-positiv cases on a RTPCR-test of one Corona-Virus,
instead looking on infektion problems in general.Where are the programs to prevent infected people will not become hostpital patients?Why not use unspecific inflammation marker CRP test to prevent infections in hospitals
and nursing homes? Every worker could be tested on CRP strikings before his shift, just
one drop blood, <5€ test and 5 minutes. There are many infection beside of Corona aviodable,
when health (also of the workes from doctor to the cleaning stuff) and not the duty rosta
and financial proffit would roule (have higest priority).
Influnenca, Streptokken, Pneumokokken, MSRA… all other infectes spread by doctors are
dangerious for their patients. In case of a CRP strike, more diagnostic should follow
before first contact with patients.
Over 200 doctors and medical workes had died in Italy in spring, not because the virus
is so agressive, major reason has been the working condition, to countious to work even
with illness.Such test could help to stop the need to work full time with mask.Why not having just a study about this, that the medical and care busines could do more
than washing hands and wearing masks.A Chinese guidline how to deal with corona cases from March 2020 recomended to do
CRP tests in early stages it was translated and recomended by an German association of
eye clinics: http://www.vsdar.de/corona/
From March to May did I request action by authorities in NRW and Germany nd try to make
this public with small demoes in front of the German health ministery in Bonn, and in
the center of Bonn – see: http://www.corona-demo.deIMHO a CRP could help to distinguish between persons with inhalated Corona-Gen and being
infected, too. But officials in Germany (RKI) count death with several negativ PCR-test
to the Corona-Death-counter: https://heise.de/-4973792.I’m just an civil engineer, but I learnd as helper in 1994 in Goma and Bukavu (Kongo)
during the colera epedime from experienced developing professionls that it is important
to have an open exchance about challanges, ressources, ideas, and experiances.So thank you again for your review, what do you think about CRP tests?
Robert Michel, GermanyReplyRuud van Wees says:Thanks so much, all of you, the real scientists annex freedom fighters.
So many branches of science these days seem corrupted and sent into deadend streets by scientists bowing for the mammon or other kinds of pressure.
I dearly hope this is the beginning of the end of this politicized corona nightmare. If not, then we know for sure there is another dark agenda behind it.ReplyEric Vieira says:The dark agenda seems to be coordinated by the U.N. and WHO. Great Reset? Agenda 21? Agenda 2030? The virus outbreak in Wuhan was even negated by WHO for a while, allowing the virus to spread worldwide. That the more SAR-Cov2 virus-specific primers as negative/positive controls were not recommended tells a tale by itself.Reply
jb says:Remember, this pandemic has started with hiding the truth that dr Li Wenliang told. Please keep in mind his last words in his poem: “…Goodbye, my dear ones.
Farewell, Wuhan, my hometown.
Hopefully, after the disaster,
You’ll remember someone once
Tried to let you know the truth as soon as possible.
Hopefully, after the disaster,
You’ll learn what it means to be righteous….”ReplyVlrdngr says:What if… He is talking about something else entirely, and the whole covid story was just a smokescreen or a cover up?Reply
Rehabilitation says:I was suggested this blog by my cousin. I am not sure whether this post
is written by him as nobody else know such detailed about my
problem. You are amazing! Thanks!Replyfrank says:Why you removed the reply’s from willem engel, who is talking about a fungus.Reply
Mario Wolf says:Excellent. Hopefully this clarification will have the necessary impactReply
Guy Verstraeten says:Eindelijk , en nu hopen dat de onzin ophoudt. Please keep giving updates about the retraction itself ! Thank you so much.Reply
Helga Smilga says:Thanks to the Bravehearts within the world of science (honest and courageous) the hinges of this utter madness will slowly but surely begin to creak.Reply
Andre N. says:Thank you for Work, this is a hope, for all People in the World.
For democracy, the rule of law and freedom.
Especially for the scientists who have made it their mission to work scientifically.
The truth always wins!ReplyJos K. says:Excellent work!Reply
Dorothee O’Sullivan Burchard says:Thank you for this excellent work! It will help to rectify the erroneous claims and measures put in place that curtail the human rights of millions of people! Concerns remain as to whether politicians of governments across the globe take this on board. If not, people need to rise up and continue the peaceful fight for their liberties.Reply
Michiel de Jong says:Thank you for what you have done for society! We are in debt to you all.Reply
Gerlinde Hörr says:Herzlichen Dank für Ihre Mühen! Ein Lichtblick und Hoffnungsschimmer nach acht düsteren Monaten voller Irrsinn! Danke, danke – vielmals!Reply
Jack AVALONE says:You all need to spread the #TRUTH on twitter.#President #TRUMP will surely see it.Reply
Tanya Sutterfield says:I am deeply grateful for the service, we are indebted to all involved. I pray this is received and used to change the devastating course we are currently on and cease the criminal actions being perpetrated on humanity.Reply
Already Provided says:Great work. A small point:
I’m not quite sure about the first part of your definition of a false positive.“The definition of false positives is a negative sample, which initially scores positive, but which is negative after retesting with the same test”.–followed by:“False positives are erroneous positive test-results, i.e. negative samples that test positive.”I would say the second sentence is correct, but not the first sentence. “initially positive but then testing negative negative after retesting” is in my view a case of imprecision (random error) of the test near the limit of detection, not a false positive.Check out Hedderich, M Sachs L, “Angewandte Statistik” 17th edition, section 4.5 p 186 “Der diagnostische Test”, Table 4.6. According to that source, a false positive occurs when:A sample from a patient *who does not have the disease* gives a positive result in the test.[conditional probability would be P(T+|K-)]Of course the critical point is defining what the “disease” is. If it is defined as a infectious state attributable to Sars-Cov-2 then you could argue that *all* results from this test are false positive.ReplyE.M. Oneal says:My profound apologies! I had in fact submitted the comment to the other thread on the “Retraction request letter”, not this one, and you had already accepted it there. Thank you for your prompt response.Reply
E.M. ONeal says:Some remarks by Prof. Ulrike Kämmerer in an interview by the “Corona Ausschuss” meade it clear to me that there is another source of false positives that are not covered by the statistical definition.There are extraneous factors that introduce contamination during the lab test procedure “on the bench” (or before!). Possibilities are the operator sneezing or positive control mnaterial finding its way into the test, or cross- contamination from other samples.That could account for the same sample testing positive on a re-test and would also be a false positive. Although one would expect that such errors are excluded in an accredited lab.There are myriad other potential sources of extraneous false positives, starting with test kit production (contaminated swabs?), through the sample collection from the subject, lab procedures, data analysis and reporting.Reply
https://www.ndr.de/nachrichten/info/16-Coronavirus-Update-Wir-brauchen-Abkuerzungen-bei-der-Impfstoffzulassung,podcastcoronavirus140.htmlNatürlich schadet es nicht auf Kritikpunkte mehrfach hinzuweisen, aber bis jetzt hat dieser Hauptkritikpunkt zumindest nicht dazu geführt, dass die Veröffentlichung zurück gezogen wurde.Schöne Grüße
TatjanaReply
Dave Spars says:Thank you, giving me hope.Reply
Stephen says:I am horrified to read this appalling misunderstanding of molecular biology and how PCRs work. This is as bad as HIV denialism. If you don’t understand how primer concentrations work and how realtime PCR works and how much of the genome was amplified, please stay at home and let the rest of us get on with dealing with COVID.Reply
Martin says:Could you please clarify in detail what has been done wrong by the authors of the review?
Which points in the process of rtPCR did they not understand fully?
Thanks for your answer!ReplyPeter Looman says:Dit is toch geen serieuze reactie. Ik weet zeker dat de schrijvers open staan als er inhoudelijk fouten aangetoond worden door andere deskundigen. De reactie van Stephen heeft op deze manier de waarde van een gemiddeld Twitterbericht (en die is in mijn mening zeer laag).Reply
Chrisje says:Even trolls are targeting scientists. What’s new?Reply
roland brautigam says:It’s clowns like you. You have the arrogance to claim that Peter Borger and Kevin McKernan and Mike Yeates don’t know how primer concentrations work without coming with a counter argument?Reply
John Weir says:Please do not try to initiate a slanging match.No-one will bite.Reply
Hanno says:I Love your reply Stephen
Quackery never died, snakeoil was never short, times don’t change, it all comes back in cycles…Replyura soul says:I am horrified to read this appalling interjection into a hotly contested topic which accuses people of being dangerously reckless with peoples lives without even inserting any verifiable claims or evidence.Reply
Thomas Ellenberg says:I’m more horrified by ad hominem spammers who can’t formulate a valid counter argument.Reply
Jabra says:Thanks for your hard work! Hopefully it will be retracted.Reply
Dr. Jörg Haberstock says:Tolle Arbeit, Danke! Wie ist es zu erklären, dass die ganze Welt nirgendwo aktualisierte PCR-Standards mit SOP zu Covid entwickelt hat, wieso greifen die Ringversuche zum Qualitätsmanagement nicht ? Wie kann das alles weltweit und über mehr als 9 Monate unbemerkt geblieben sein? Das macht mich ratlos. Danke für Euren MutReply
Arnold Achen says:Was Sie verlangen wird von den Behörden nicht gefördert, weil sie mit den Konzerninteressen verflochten sind, die eine Fortführung der inszenierten Pandemie forcieren:https://www.corona-schadensersatzklage.de/corona-status-quo-erste-pcr-klage-ist-anhaengig-gemacht-worden/Und warum sollen die Labors an mehr QC interessiert sein wenn sie aktuell durch die reine Massentestung ein Vermögen verdienen?Reply
Caro oh oh says:So, yes, this qPCR is not the best designed one, but due to the circumstances (designed in January as a broad Sars-corovnavirus detection test), this can be understood. Yet, this does not mean that the test does not work. It has been validated a lot by a lot of labs and instances. Every lab has to do a validation/verification of the used tests. Furthermore internal and external controls are taken into account. You can find more details in the paper (and other papers) and also in the news article: https://www.rd.nl/meer-rd/gezondheid/pcr-test-overleeft-stortvloed-aan-kritiek-1.1718351. Next to this, a whole plethora of real-time PCR tests are available on the market.And yes, having viral RNA in your nose is not the same as being infectious. However, it shows that you have been in contact with the virus. Together with symptoms, this diagnostic test can confirm a diagnosis. Concerning asymptomatic cases, it might be of interest to consider the viral load. It is a valuable point that persons with a low detected viral RNA concentration might not be infectious (yet), or not very well swabbed if no human control gene is taken along.I will summarize the other issues I have with this report, as discussing all of them would lead us too far.1. High primer concentrations: Primer concentrations between100-900 nM are standard (depending on the assay and also the supermix). As the primers contain wobble bases, rather high concentrations make sense. Furthermore, dNTP concentrations are not off from standard conditions.
2. In general wobble bases rather have a negative effect on PCR efficiency (as the correct primer might be exhausted). Here, this seems not the case. On the other hand, it seems that the RdRp assay has a lower sensitivity ((Vogels et al. https://www.nature.com/articles/s41564-020-0761-6), possibly due to a mismatch with some Sars-Cov-2 genomes.
3. Good real-time PCR designs are set-up to detect short fragments (preferably under 150 bp) to obtain a good reaction efficiency. Furthermore, lots of viruses have been sequenced by now (take a look in the NCBI,GISAID,Nextstrain databases for example). The RdRp assay will be transcribed less then the ORF1a transcript, as a frameshift is necessary to transcribe RdRp. Hence, this could theoretically lead to a lower sensitivity of the RdRp assay.
4. The RdRp-assay indeed has not the best design. However, this is a confirmatory assay and it has rather a lower sensitivity (see Voghels et al.). Yes, the E-gene assay might also detect Sars-CoV-1, but this virus is not really going around (and is also causing severe disease).
Off note, concerning melting temperatures, the theoretical Tm calculations should take into account the reaction conditions. Furthermore it is not required to mention Tms, nor GC contents in publications (as you can just copy the primer sequence into an oligo analyzer tool). It is way more useful to validate the annealing temperature in practice (with a gradient PCR for example).
Concerning the primer dimers: as a probe will only detect specific amplicons and not primer dimers, these probably have rather a negative effect on detection.
5. A Cycle Quantification (Cq) threshold is not a unit and is workflow specific. Every lab will have to do its own validation. A Cq value will be dependent on the swab, transport, RNA-extraction, reverse transcription, PCR assay (design, supermix, sample, instrument, plastics) and analysis. You can maybe correlate viral load and time since symptom onset with infectiousness, but not nationwide Cq values (as this will at least be lab dependent, this is not even taking intralab variation into account).
6. I cannot judge about the validation protocol, as probably not every step is described. Melting curves during optimization or sequencing of amplicons is indeed good practice. But again, this assay is a confirmatory assay and has been wet-lab validated. (Gels are IMO for scientists stuck in the nineties and are risks for amplicon contamination.)
7. The test has been validated on negative, positive and other viral controls (read the paper!).
8. The paper and protocol on the WHO website contain a quite well described protocol. Furthermore, each laboratory has to do a proper validation. Btw, it would be unsuitable (in terms of supply chain for example) that every lab is using exactly the same protocol. Reference standards would be useful (but I guess they will come).
9. Funny, as this report is also not peer reviewed, nor has a DOI, and hence, cannot be pubpeer reviewed.
Regardless of this paper, a diagnostic procedure in a lab does not require publication and peer review. It requires wet lab validation.In conclusion, the design of the RdRp assay is for sure not the best (but there is a good explanation for this). This remains a confirmatory assay after screening with the E-gene assay (or in a multiplex nowadays). The Charite protocol has been extensively validated and remains a valid diagnostic tool.Reply
Next to this, a whole plethora of real-time PCR tests are available on the market.Answer: That’s not the subject of the criticism of the specific publication.Quote: And yes, having viral RNA in your nose is not the same as being infectious. However, it shows that you have been in contact with the virus. Together with symptoms (!!! Yes, but nobody tests symptomatic persons only), this diagnostic test can confirm a diagnosis. Concerning asymptomatic cases, it might be of interest to consider the viral load. It is a valuable point that persons with a low detected viral RNA concentration might not be infectious (yet), or not very well swabbed if no human control gene is taken along.Answer: Yes, but nobody tests symptomatic persons only!Quote: 1. High primer concentrations: Primer concentrations between100-900 nM are standard (depending on the assay and also the supermix). As the primers contain wobble bases, rather high concentrations make sense.Answer: Maybe – but not in the case of the E- and N-Gene PCR without wobble bases.Quote: 2. In general wobble bases rather have a negative effect on PCR efficiency (as the correct primer might be exhausted). Here, this seems not the case. On the other hand, it seems that the RdRp assay has a lower sensitivity ((Vogels et al. https://www.nature.com/articles/s41564-020-0761-6), possibly due to a mismatch with some Sars-Cov-2 genomes.Answer: Yes, but why the mismatches – the genomes were available at the time of submitting the manuscript and the Vogels paper is from Jul 10 2020.Quote: 3. Good real-time PCR designs are set-up to detect short fragments (preferably under 150 bp) to obtain a good reaction efficiency.Answer: Correct.Quote: Furthermore, lots of viruses have been sequenced by now (take a look in the NCBI,GISAID,Nextstrain databases for example). The RdRp assay will be transcribed less then the ORF1a transcript, as a frameshift is necessary to transcribe RdRp. Hence, this could theoretically lead to a lower sensitivity of the RdRp assay.Answer: not of interest – its about the publication from January.Quote: 4. The RdRp-assay indeed has not the best design. However, this is a confirmatory assay and it has rather a lower sensitivity (see Voghels et al.). Yes, the E-gene assay might also detect Sars-CoV-1, but this virus is not really going around (and is also causing severe disease.Answer: Yes – but for an “novel” virus the detection system must be highly specific.Quote: Off note, concerning melting temperatures, the theoretical Tm calculations should take into account the reaction conditions. Furthermore it is not required to mention Tms, nor GC contents in publications (as you can just copy the primer sequence into an oligo analyzer tool). It is way more useful to validate the annealing temperature in practice (with a gradient PCR for example).Answer: 10 degree difference is a no-go and yes, everybody optimizes the PCR primers for GC and melting temperature before ordering them… so a rubbish argument.Quote: Concerning the primer dimers: as a probe will only detect specific amplicons and not primer dimers, these probably have rather a negative effect on detection.Answer: This is correct.Quote: 5. A Cycle Quantification (Cq) threshold is not a unit and is workflow specific. Every lab will have to do it’s own validation. A Cq value will be dependent on the swab, transport, RNA-extraction, reverse transcription, PCR assay (design, supermix, sample, instrument, plastics) and analysis. You can maybe correlate viral load and time since symptom onset with infectiousness, but not nationwide Cq values (as this will at least be lab dependent, this is not even taking intralab variation into account).Answer: Well – they have “validated” their PCR so they should have shown their PCR data and CT values – and indeed every lab had to adapt the Test inhouse – but this point is missing in the publication – so not ok.Quote: 6. I cannot judge about the validation protocol, as probably not every step is described. Melting curves during optimization or sequencing of amplicons is indeed good practice. But again, this assay is a confirmatory assay and has been wet-lab validated. (Gels are IMO for scientists stuck in the nineties and are risks for amplicon contamination.)Answer: no – not wet-lab validated: no clear results for negative and positive controls are shown (including CT).Reply
Randomer says:Thank you for confirming the paper by stating three times that the design might not be the best one. Cheers.Reply
theasdgamer says:“Together with symptoms, this diagnostic test can confirm a diagnosis.”Confirm a diagnosis for what purpose? Adding delays for testing decreases prognosis. Patients are often dilatory about testing and most patients max their viral load on day 3 post symptom onset and maybe contact their primary care physician on day 2 post symptom onset best case. If the doctor won’t treat with an antiviral, who cares about any PCR test? A doctor will treat the symptoms of a URTI. If you do treat with an antiviral, you can’t wait for the return of test results to begin treating. If the antiviral works, what purpose does the PCR test serve?And I’m not a physician.Reply
Lothar Lammfromm says:Yes, you are not a physician and you don’t know anything about Covid-19-Science.This sentence is pure rubbish:“most patients max their viral load on day 3 post symptom onset”No, usually (but this can vary) it is the day before (!) system onset.Reply
ShowMeTheFacts says:“…what purpose does the PCR test serve?”1) To maximize the Casedemic
2) To use as junk science to justify totalitarian policy (businesses restrictions, community activities, sports, etc.)
3) To use as junk science to quarantine non-infectious teachers, students, staff, etc.
4)…
5)…
6)…Reply
Dipl. Ing. (FH), M. Eng. Andreas Macher says:Um dem nächsten Wahnsinn einer Pseudo-Epedemie vorzubeugen müssen unbedingt rechtzeitig vor den unsicheren Test wirklich sichere Tests für die wahrscheinlich virulenten Virusarten entwickelt werden. Damit nimmt man der Impfindustrie den Spielraum, den sie mit den falsch positiven Tests in dieser Epedemie hatten, für die Zukunft. Als nächstes Target zeichnet sich MERS ab.Wenn dieses kompetente Team dafür sorgen würde, dass die relevanten Varianten von MERS wirklich sicher detektiert werden können, wird es nicht noch so eine Panikreaktion in der Bevölkerung geben, wie wir es mit SARS-CoV-2 erleben mussten.Reply
Linda Weingärtner says:Die Panikreaktion der Bevölkerung kam durch die Angstmache der Politik und der Medien zustande.
Die kritischen Stimmen der Experten werden ja bis dato immer noch nicht gehört.ReplyJulian says:Well, you don’t have to worry about MERS.Because dr. Drosten himself said in a 2014 interview that PCR isn’t suitable for detecting MERS.
He admits the PCR technique is too sensitive, which leads to an explosion in case numbers and that the media tends to blow up these case numbers out of proportion.https://www.wiwo.de/technologie/forschung/virologe-drosten-im-gespraech-2014-die-who-kann-nur-empfehlungen-aussprechen/9903228-2.htmlReply
Ordinary Doc says:I do not understand the technicalities of pcr testing. I am however an experienced clinician and I understand what I see in my everyday practice. What you are saying seems absolutely correct. False positives++. Well done and good luck.Reply
Danae says:God bless you all for this incredible work. It is time to cleanse our system from monetarism which is subverting science, medical reserach, press and politics. We cannot have independent, impartial science if it is “financed/bribed” by groups of have clear conflicts of interest. This paper is an important step to stopp the attack on civil liberties and stands between many people being injured by vaccination or even death or infertility. No vaccination is without risks, there are no studies ever being done on how different vaccinations interact with each other. Where there is risk there must be choice.
God bless you and protect you and my he help us all to stay for truth, love and peace.Reply
Michael Wiedom says:Herzlichen Dank für diese hervorragende Arbeit. Bin gespannt, wie es jetzt weitergehen wird.Reply
Petra von Kopp says:Danke… Danke…Reply
OrAnd says:Thanks for your work, I do have a comprehension question regarding 3.: How can there be false positives at all by this PCR-test? In the text (3) and reference [2][3] I can see only points stating why there can be false negatives (not false positives). Am I missing something here?Reply
Kevin McKernan says:We present evidence of promiscuous primers. These can create both False Negatives and False Positives. There are additional sources of FPs as the assay fails to discern infectious from non-infectious people and when used on asymptomatic people, the term ‘case’ must be infectious. This is exacerbated by the selection of a reduced number of assays and assays positioned on the 3′ region of the genome. Wolfel et al describes qPCR amplicons that do a better job discerning infectious from non-infectious people. Drosten is an author on this paper. This would have been a good time to withdraw the assays failing to discern the difference between infectious cases and non-infections false cases.Reply
Ruud Brouwer says:Will this report be peer reviewed? I really doubt it will make it throughReply
RHB says:Well done to all involved. Meticulous scrutiny with massive implications. Not my area of detailed expertise but, if I’m reading broadly right, could even bring down the leadership of the country I’m posting from.Substantiates gut feeling going right back to March, as per blog comments by “Ancient Briton,” “RHB” and “Aweson Walles” on Derek Lowe’s In the Pipeline blog (dominated March onwards by virus postings):https://blogs.sciencemag.org/pipeline/archives/2020/03/12/real-information-a-public-good#comment-31346816 March, 2020 at 9:09 amhttps://blogs.sciencemag.org/pipeline/archives/2020/11/18/vaccine-possibilities19 November, 2020 at 6:49 amhttps://blogs.sciencemag.org/pipeline/archives/2020/11/23/oxford-az-vaccine-efficacy-data#comment-33360129 November, 2020 at 10:21 amReply
She claims peer review was done
She claims they started working on the protocol from 12 January when China released the genetic code
She claims the guidelines for the PCR test were changed by RIVM on 25 September but that these should not change the outcome. I have a few statistics which prove the opposite.
Adam Meijer states that false positives could be 0,5-4%. 4% is catastrophic already.
Article is under “conspiracy theory” header…ReplyJoerg says:Hallo,
Danke fuer die Recherche, das Corman-Drosten et al.Paper war halt ein Schnellschuss.Mich wuerde interessieren, ab wann (im Zeitverlauf, zB ab September2020?) es bessere Tests (weniger Kreuzreaktionen, weniger Falsch-Positive, 3Gen-Prinzip, spezifischer, Beachtung von Ct-Werten) in den Test-Laboratorien gibt? Gibt’s ein Labormediziner, der hier mit liest?Ich kann mir nicht vorstellen, dass eine Vielzahl von Laboren immer noch die gewobelten RdRp-Primer verwendet? Oder?LG JoergReplyM. Hoffmann says:Bessere Tests für was? Für ein Virus das unter Umständen gar nicht existiert–laut Angaben des CDC gibt es kein messbares Isolat von sars-cov-2 weltweit– oder sich funkionell nicht von üblichen Coronaviren unterscheidet und auf jeden Fall denen gegenüber keine höhere Mortalität aufweist?Was soll das klinische Korrelat zum “Test” sein? Welche therapeutischen Massnahmen sollen auf Grund von welchem Testergebnis eingeleitet werden und ergeben dies überhaupt einen Sinn, außer Unterdrückung?Haben Sie in der Vergangenheit je einen Test für Grippe oder schwere Erkältung durchgemacht und was hat das gebracht?Laut der Dokumentation zu Event 201 handelt es sich ohnehin um eine globale Simulation zur Bereicherung von “öffentlich-privaten Partnerschaften”.Es ist die Jagd nach einem Phantom, das benutzt wird, schwere psychologische und wirtschaftliche Schäden anzurichten.Reply
Sonst jemand?Sie antworten dagegen auf Fragen, die ich nicht gestellt habe. Gerne aber meine Meinung dazu:
a) “Virus das unter Umstaenden nicht existiert” “und auf jeden Fall denen gegenüber keine höhere Mortalität aufweist”
Belege dazu? Sind erhoehte Mortalitaeten in anderen Laendern als in D also alle gefaket? Wie erklaeren Sie erhoehte Mortalitaeten (zB auf http://www.euromomo.eu/graphs-and-maps) bei unseren Nachbarn?
b) “klinische Korrelat zum “Test” sein?”
Wie waere es mit dem Versuch, die Verbreitung etwas einzudaemmen, um Gesundheitssysteme/Intensivbetten nicht zu ueberlasten? Also keine klinische Konsequenz sondern eine epidemiologische?!
c) “Haben Sie in der Vergangenheit je einen Test für Grippe oder schwere Erkältung durchgemacht”
Nein, warum auch? Gibt’s denn solche? Bin nicht in der Risikogruppe
d) “Laut der Dokumentation zu Event 201”
Sie meinen, http://www.weforum.org/great-reset/? Da haette ich viel zu tun, alle dubiosen Traeume, Stilblueten und Ideen im Internet zu verfolgen
e) “Es ist die Jagd nach einem Phantom, das benutzt wird, schwere psychologische und wirtschaftliche Schäden anzurichten.”
Wissen Sie, dass manche Kommentare mehr ueber den Kommentator aussagen als ihnen Sinn und Zweck innewohnt?
Meinen Sie wirklich, alles geschieht aus boesartiger Taktik?
Meist reichen Angst, Unwissenheit, Inkompetenz, Unfaehigkeit aus, um fast alles “gut gemeinte” zu erklaeren … meine Erfahrung jedenfalls.
Sie kaempfen gegen Windmuehlen?!
LG JoergReplyKane says:Ähmm.. Quelle [3] dieses Reports spricht eindeutig von Isolaten “1941 SARS-CoV-2 isolates could be obtained”
Also du stimmst diesem Report hier zu, aber dessen Quellen lehnst du ab? Versteh ich das so richtig?Reply
ich weiss nicht, was du meinst? Wolltest du unter einem anderen Kommentar kommentieren?Quelle [3] finde ich interessant und verstehe sie genauso wie du? (also es wurden sehr wohl viele Covid19-Isolate untersucht sowohl per RT-PCR als auch in Subkultur).
Bei RT-PCR gibt’s halt eine gewisse Gefahr von Falschpositiven, auch gibt es einen laborspezifischen Effekt (Labore mit mehr / weniger Expertise?) aber es ist trotzdem die beste aller Nachweismethoden fuer epidemiologische Studien (Schnelligkeit, Durchsatz) und so zu tun, als wuerde da “Irgendwas” rumamplifiziert ist m.E. weiter weg von der Realitaet als 0,5-2%? Fehler.
Alle machen Fehler, nix ist vollkommen in dieser Welt.
Trotzdem ist es ein sinnvolles und berechtigtes Ansinnen fuer mehr Sensitivitaet und Spezifitaet beim Corona-Testen einzustehen! Ein erster Schritt waere viell. bei jedem Test auch den Ct-Wert mit zu liefern? und Getestete mit Ct >30 und ohne Symptome sollten berechtigterweise maulig werden!
LG JoergReply
Francine says:Ik heb een vraag over paragraaf 3 op blz 6. Waarom is daar letterlijk de persoonlijke mening over de PCR test overgenomen van de twee rechters van het Hof van beroep in Lissabon inzake het onrechtmatig in quarantaine plaatsen van vier Duitse toeristen. De zaak waardoor Wybren van Haga meende te moeten melden in de Kamer dat de rechter in Portugal de PCR test naar de prullenbak heeft verwezen.
Deze ‘conclusie’ is te lezen onder punt 17 van het arrest. Zij hebben deze aanname gemaakt na een foutief interpreteren van het onderzoek van Jafaar et al. Dit is dus geenszins de conclusie van de studie van Jafaar et al. Ook onder reviewed 3 maakt u een fout, het gaat hier niet primair om het aantal cycli (35) van de PCR waarboven besmetting < 3% is. Het gaat om het vermogen waarop besmette samples nog in staat zijn cellen in kweek te infecteren, dat was na 3 weken < 3% Dit om zicht te krijgen op duur van isolatie van covid-19 patiënten.
Morgen dient een zaak over de bemoeienis van beide rechters bij de Hoge Justitiële Raad van Portugal.
Volgens het SCM hebben de rechters hier hun bevoegdheden overschreden. Zij hadden geen opmerkingen moeten maken over de mate van betrouwbaarheid van de tests die momenteel worden gebruikt. Daarom zullen ze morgen op 2 december vragen moeten beantwoorden in een tuchtonderzoek door de Justitiële Raad, uitspraak door het Superior Council of Magistrature of de twee rechters juist gehandeld hebben.
Ik ben benieuwd of u hiervan op de hoogte bent en zal de uitspraak gevolgen voor de tekst op blz.6 paragraaf 3 hebben.ReplyRich says:These are theoretical weaknesses you may find in most scientific paper. As many of the reviewers are affiliated with biomedical institutions, why don’t you just show experimentally the predicted consequences for false positives? When the assay is really that bad, this shouldn’t be difficult!?Reply
Adam says:“of course I did not spend too much time for this… “… so why did you leave a comment? Sometimes it is better to su. The authors did spend a lot of time to identirfy the weaknesses of this “study”. It seems we have here a Drosten Fan Boy? For you this is almost religious…Reply
Kevin McKernan says:See my comment on the thread.
We don’t need to perform any experiments as we have provided 4 manuscripts that have already documented this effect.Reply
Carlyn Mildrum says:immaculate articleReply
Bölecke says:Ich bin begeistert. Jetzt weiß ich warum die (unsere ?) Regierung nicht mit schlüssigen Erklärungen die Situation beschrieben hat.Reply
Kim ten Napel says:Following this Corman/ Drosten retraction paper with interest and hope for a better world.
How this ONE PCR test can cause such panic and be used to control the people is remarkable,as this one PCR test has so much power why hasn’t it been tested and retested by every country.
I applaud Peter Borger & Co for this admirable difficult investigation.Reply
infectious disease epidemiologist—you never get herd immunity.
You don’t get it for malaria, you don’t get it for HIV, you don’t get
it for Ebola.”With particular reference to HIV , am I the only person to see this statement as utterly ridiculous ?Expert comment if someone would be so kind.Sorry this is somewhat off-topic.ReplyDr. med. Karla Lehmann says:Herzlichen Dank für Ihre äußerst wertvolle und notwendige kritische Einschätzung dieses CormanDrosten-Papers. Ihre Arbeit hat mir die Zuversicht zurückgegeben, daß es noch Wissenschaftler gibt, die die Dinge hinterfragen und der Sache auf den Grund gehen.Reply
Inhope says:Does any of you, the authors, SPEAK to humans you know?!
If you have any credibility around you, why haven’t you disseminated all this personally?
You know, word of mouth is very potent! The interlocutors would diffuse it further with something like this:
“I know this person. I trust them. I will act as they advise.” And so on.
I guess you are just tinkering with details.
Because if you knew that “viruses” are not real, you would have spoken long ago, and this report would have been unnecessary!
I am close to dying from old age. I have had time to acquire knowledge in more fields than anyone I heard of. Quantum Physics, Artificial Intelligence/Robotics, Electrodynamics, Philosophy of Science.
The last one mentioned provided the space where I, and others, could really take apart all the preposterous statements and outright lies of the “scientists” all over.
They all go to the brainwashing centers called universities and get poisoned by older brainwashed “professors”. They have no chance of waking up later on because they never leave the fetid “educational” environment that raised them. One in a million does, but then they are afraid of losing their grants or professorships! So they continue in the complicity to murder and maim!
Have you heard of Thomas Kuhn?
Stefan Lanka?
One quit physics because of what I criticize, the other has been speaking for decades against his original domain: “virology”.
Get informed.
And get honest.Reply
Peter Barnum says:I have two immediate questions:1) The primer dimers you found are between one primer for the E gene assay and one for the RdRp assay. In real life testing, the two test are conducted in different wells, aren’t they? So these two primers should never be in the same pot and therefore the potential dimers don’t matter!?
2) Even if primer dimers form and are elongated by the polymerase, this would not lead to increased fluorescence, since a probe-based assay is used in contrast to intercalating dyes which would indeed just indicate any dsDNA being present.Regards
PeterReply
Drosten is an author on this paper that declares the need to replace the primers that were reviewed in 24 hours.You cant have it both ways. If they are promiscuous primers and known to be leading to false positives or false negatives in the literature, then the Author of such disinformation has an obligation to correct record when peer review was clearly compromised.2)Etievant et al“The E Charité and N2 US CDC assays were positive for all specimens, including negative samples and negative controls (water). These false-positive results were explored (details below), but the
sensitivity of these assays was not further assessed.”“No false-positive results were obtained on clinical samples that tested negative for SARS-CoV-2 and/or positive for other viruses than SARS-CoV-2, except for E Charité and N2 US CDC, which were positive for all specimens.Sensitivity was first assessed using SARS-CoV-2 cell culture supernatants. Using both specific SARS-CoV-2 (S) and non-specific (NS; detecting SARS-CoV-2, SARS-CoV, and bat-SARS-related CoVs) RdRp Charité assays”Notice the term “non-specific” Charite’ primers.“Thus, the false-positive amplification obtained using E Charité
might be derived from a contamination (amplicon size at 121 bp) but could also be associated with an aspecific amplification (amplicon size at 84 bp)”https://www.mdpi.com/2077-0383/9/6/1871This paper stops evaluating the Charite’ primer half way through the study due to performance issues.3)Jung et al“On the other hand, the RdRp_SARSr (Charité) set shows less effective amplification than the other two primer sets at all reaction temperatures including their recommended extension temperature (58 °C). Unexpected amplifications from NTC samples were observed with the RdRp_SARSr (Charité) set. The electrophoresis and melting curve analysis showed nonspecific amplification at lower positions (Lane 5, Figure S5b) and temperatures (Figure S5a) than the result of specific amplification with the RdRP_SARSr (Charité) set.”Evidence of Non Template controls amplifying in multiple peer reviewed journals will directly result in quarantine of non-infectious people and create a chain reaction of legal liability.4)Gand et al.
“Interestingly, for Assay_2_RdRp-P2, similar false-positive results as obtained in our in silico study were obtained in the wet lab by Chan and colleagues, who detected SARS-CoV when using the probe P2 targeting the RdRp gene that is considered strictly specific to SARS-CoV-2 [30]”“The sensitivity of Assay_2_RdRp-P2 (Charité) was already demonstrated in the wet lab to be lower than that of other assays investigated in this study”These four manuscripts all point to flaws in these primers.Christian Drosten is an author on one of these manuscripts that concludes they must be replaced!The error prone primers still exist on the WHO website with a link to Eurosurveillance.The liability of these erroneous methods leads back to Eurosurveillance and once they have been informed of the issue, we are confident they will do the right thing and retract the paper. A failure to retract the paper absorbs the liability as opposed to placing that liability onto the source of the deception; the authors themselves.Citations are contained in this thread.
https://t.co/2St1wmWLAH?amp=1Reply
https://www.eurosurveillance.org/content/10.2807/1560-7917.ES.2020.25.3.2000045#html_fulltextUsing the E and RdRp gene assays, we tested a total of 297 clinical samples from patients with respiratory disease from the biobanks of five laboratories that provide diagnostic services (one in Germany, … (Table 2). In total, this testing yielded no false positive outcomes.In four individual test reactions, weak initial reactivity was seen but they were negative upon retesting with the same assay. These signals were not associated with any particular virus, and for each virus with which initial positive reactivity occurred, there were other samples that contained the same virus at a higher concentration but did not test positive. Given the results from the extensive technical qualification described above, it was concluded that this initial reactivity was not due to chemical instability of real-time PCR probes but most probably to handling issues caused by the rapid introduction of new diagnostic tests and controls during this evaluation study.Notice here:The one sentence states “no false positive outcome”. The next sentence states: “In four individual test reactions, weak initial reactivity was seen but they were negative upon retesting with the same assay”.The first sentence becomes irrelevant and wrong, the second sentence states 4 (!) false positives out of 310.In my opinion this is the kind of “cautionary manner” that led to the current situation worldwide, contradictions and the all-of-a-sudden-scientific acceptance of those, embedded in “cautionary manners”.The truth here is:Four in ‘n=310’ primary-tests => false positives = 1.3 %.1) This is a direct alegre to the many reported cases of “false positives”, then “negatives”.Further:2) Christian Drosten publicly states that this is his “validation concerning cross-reactivity”, we tend to see this as a pseudo-validation:https://twitter.com/c_drosten/status/1309755692232904704This is no validation, there are 4 false positives, it’s clearly stated in the text accompanied to table 2 of the CD-paper. Further, table 2 does not state a) which “validated clinical sample with known virus” caused the false positive, b) which gene assay.2) The editors of Eurosurveillance further state in their communications: https://www.eurosurveillance.org/content/10.2807/1560-7917.ES.2020.25.21.2001035Quote:Our strategy during establishment was to use a synthetic target for the SARS-CoV-2 E gene assay, while validating amplification of a full virus genome RNA using the RdRp assay that is specific for both, SARS-CoV and SARS-CoV-2, with the latter not being available to us in the form of an isolate or clinical sample at the time. Based on experimental validation, it later turned out that the mismatched base pairs do not reduce RT-PCR sensitivity and are not to be seen as the reason for somewhat higher Ct values with the RdRp assay as compared to the E gene assay [3].Notice here:“Based on experimental validation, it later turned out that the mismatched base pairs do not reduce RT-PCR sensitivity …”Again we have this “cautionary manner”: “The basic copies do not correspond to each other BUT that does NOT effect the sensitivity.”… Trust us.Nonsense! The supplementary data & material of this correspondance does not confirm that, which is provided with this editor’s / author’s communication response.
RdRp needs to be pulled.Drosten is an author. Same journal.Can’t have it both ways.The WHO still points to old and faulty primers and the first review clearly missed the error. We expect this to be withdrawn soon!Everything else is irrelevant concerning your supposedly-not-answered-question in regards to our review report on the CD-protocol, and also the referenced nature article (chinese study): We know that they are not using the CD-protocol, so it is irrelevant for our review report, which focuses on the Corman et al. publication and nothing else. This is a fallacity in itself.5) We know that most of the western hemisphere uses 2 confirmatory assays. We know that China uses much better protocols (see point 3) than the CD-protocol. We know that Taiwan even uses 3 confirmatory assays – as everyone should (also compare the specific sections on this in our review report). AND we know that Thailand uses two genes when arriving there, and when initially tested positive, then another 5 genes are used for confirmation to decide if the person is indeed positive or not, thus nearly no cases in Thailand.So in summary I’d say, your question is heavily irrelevant in regards to our review report because it sets the scope outside the Corman-Drosten Protocol review (it’s not so hard to stay on topic, our review report sets the framework).
control samples with the RNA of other Coronaviruses were shown.So its a publication on the detection of RNA from one Patient (and unusual: stool sample – why not from respiratory samples?) in different dilutions….And not supporting anything about specificity but only sensitivity of the dilution series of this one specific sample.
NN says:This website here is nonsense. See here for the reasons why: https://twitter.com/BMauschen/status/1333468806203793411Reply
Tony says:I would like to share some comments or questions:
1… nm means “nanometer” ….
This is a typo, no more and no less, and any lab technician will recognise it immediately.
2.”First, a positive test for the RNA molecules described in the Corman-Drosten paper cannot be equated to “infection with a virus”
Where else would the RNA come from, if not through replication of the virus in the host. Could high RNA concentrations be caused by “contamination”?
3.”…the Corman-Drosten test was not designed to detect the full-length virus, but only a fragment of the virus. We already concluded that this classifies the test as unsuitable as a diagnostic test…”
Detection of the full-length construct would be too costly, time demanding and therefore counterproductive. If you consider the PCR assay unsuitable for diagnosis, what about PCR-based liquid biopsies?
4.”Secondly and of major relevance, the functionality of the published RT-PCR Test was not demonstrated with the use of a positive control (isolated SARS-CoV-2 RNA) …”
The test does not work, because it was not originally validated with the isolated virus? Sounds weird to me.
5.”The PCR test in the Corman-Drosten paper therefore contains neither a unique positive control nor a negative control to exclude the presence of other coronaviruses.”
So far I remember, there were a lot of positive and negative controls included.(?)6. “The first sentence of this excerpt is clear evidence that the PCR test described in the Corman-Drosten paper generates false positives”
„however they were negative upon retesting with the same assay“. Positives will be retested anyway, even by amplification of another gene, isn’t is?ReplyStringer says:Scenario:1. SARS-Cov2 effectively does not exist except as a theoretical construction in a computer
2. It is part of the pandemic simulation called “Event 201” which aims at traumatizing and controlling the population and enabling public-private partnerships to plunder cash and resources on a massive scale
3. There is no preparation of the virus available in any lab worldwide in amounts sufficient to perform independent determinations of its identity, defined as a complete elucidation of the viral structure and characterization of its biological activity
4. There is no preparation of the virus available in any lab worldwide in amounts sufficient for use as a laboratory standard
5. No such virus has been isolated from any human dead or alive in sufficient quantities to perform a complete determination of identity (structure and biological activity) in order to compare it with a standard
6. Therefore the current PCR test is “detecting” RNA fragments from common corona viruses that have been around for years
7. Even if the test is substantially improved, it will be chasing a phantom however many sequences may be simultanously targeted
8. There is no indication of the clinical utility of the PCR test except to tyrannize the population by enforcing quarantine and other punishment
9. The clinical utility of the PCR test for the tested persons is zero, otherwise such tests would have been in use for at least a decade to support diagnosis and therapy of influenza or heavy cold cases.Factual and logical rebuttals are cordially invited.Reply
1) (non existence) Ask ill people or read cases, look to excess mortality charts (besides Germany)
2) (conspiracy) Honestly, do you feel traumatized & controlled? Actually, we are far through the tunnel already. Fear, ignorance, incompetence, incapacity are enough to explain the most.
3) (no big amounts of virus preps) Can’t comment on this one (no expert).
4) (no lab standard available) Could be synthesized/PCR-amplified of infectious material?
5) similar as 3) see 3)
6) (common corona virus detected) Why then is there an excess mortality of positive tested patients?
7) (there is no improved test) That’s what I’m asking. Why not? There are specific genes and discriminating sequences (stable enough/without mutation pressure) compared to common corona viruses?
8) (no clinical advantage) Yes. The tests are established to monitor the spreading, to shelter the healthcare system. Epidemiological reasons! (NOT clinical)
9) similar as 8) see 8)
Kind regards JoergReplyDr Solomon says:There is no excess mortality in UK where I practice as a doctor.
The numbers presented ( 59K) also include those thousands who were literally murdered in their care homes by Boris Johnson Govt as they were denied access to hospital.It also includes those who died months later from other causes and had tested positive (false positive) earlier.It also includes thousands of people who died in their homes due to heart attacks, strokes, infections and cancers because they were frightened to death by BBC propaganda and did not come to hospitals! So are you still surprised by slightly increased numbers of death this year ??
Doctors think and Robots don’t !
Kind RegardsReply
http://www.euromomo.eu/graphs-and-maps
Please, scroll down to the UK graph, there is a tremendous peak in excess mortality in the calendar week 15 2020 (z-score 36).
Imagine, if you say “all these data (for other countries too) are a concerted fraud”, you better bring up proofs beside selective, specific, personal observations?
Why is the „fraud“ so naïve, leaving out countries as Germany (there is indeed no excess mortality) making a “conspiracy theory” more difficult?
Who should have ordered this fraud? How could it be concerted among different countries which are fighting each other on the level of EU policies?
The answer is: VERY, VERY PROBABLY THE GRAPHS PRESENTED ARE NOT WITHOUT MISTAKES BUT IN TOTAL THEY ARE THE BEST ESTIMATE OF THE TRUTH WE HAVE?!Off course, one could and should earnestly discuss all the governmental actions according the “pandemic” (besides it is more or less severe).
But it is to easy to borrow “simple” answers for complex questions! This is typical human, but behold and think.Thanks for your service as a doctor, much power and strength for your duty.
JoergReply
https://www.youtube.com/watch?v=0JcVglSdQ-c
That does not come temporally there!Better check the facts and don’t blindly believe what you pick up on the Internet.
https://www.nature.com/articles/s41419-020-2721-8It was not until March 28 that hydroxychloroquine (HCQ) was briefly introduced by the U.S. Food and Drug Administration (FDA).
There were already many patients in the hospitals, do you really think that all of them had administered HCQ from 29th March (week 14) and in week 15 (peak) so many were dead? What about all the people who died in nursing homes or at home? Do you think they all received HCQ from their GPs as of 29 March?
If you have verifiable sources of it, give it to me.Of course, it is true that many suboptimal treatments took place in March/Apr, but thanks to God, the mortality rate has decreased over time. A merit of the research, the doctors, the nursing staff.
As I said, it is easy to discuss whether many restrictions on the part of politics are exaggerated.
Nevertheless, don’t get involved in every witch-hunt! Please consider the scientifically verifiable facts, not just opinions on YouTube.Kind regards Joerg
https://www.rubikon.news/artikel/die-medikamenten-tragodie
Dass im Fruehjahr viele Behandlungsfehler vorkamen ist richtig. Dazu gab es ja Gott-Sei-Dank eine schnelle Lernkurve.
Die Sterbestatistiken sind sehr heterogen, es gab keine einheitlichen Zaehlweisen in den Laendern, uswTrotzdem bitte ich Sie die Charts bei Euromomo zu studieren:
– Es ist mE genauso ueber das Ziel hinausschiessend, die gesamte Datenlage zu verharmlosen.
– Wie erklaeren Sie laenderspezifische Uebersterblichkeitspeaks im Herbst (in A, B, F, I, SLO, CH)? Sind diese immer noch “unfaehig Patienten besser zu behandeln”? Sind da immernoch “Medikamenten-Tester” am Werk?Glauben Sie mir, viele Massnahmen in D finde ich uebertrieben, mir waere selbstverantwortliches, freiheitliches Handeln lieber als Ge- und Verbote (zB wie in S, CH). Aber die dtsch. Mehrheit will es, die Politik setzt es um.Thesen, weshalb es in manchen Laendern weniger schwere Covid19-Verlaeufe gibt, koennten sein: weniger Vorbelastungen? gutes Gesundheitssystem? bessere indiv. Immunsysteme? genet. Disposition? vorherige Kontakte mit milden Corona-Viren? usw. Aber das hilft uns jetzt nicht.
Bald haben wir es geschafft und dann entspannen sich alle hoffentlich wieder.LG Joerg
A. Berglund says:1)You cannot prove that the “ill people” and “cases” are infected with “SARS-COv2”. The “diagnostic test” for RNA fragments is useless. It is a random number generator on the set (0,1).2)”through the tunnel”–I’m sure the proprietors of hotels, restaurants, fitness and yoga studios and the travel industry who have lost up to 80% of their revenue compared to 2019, and the workers who have lost their jobs, will derive great comfort from those words. You must have a well-paid government job.(6)and (8)the diagnostic test is useless for any purposeReply
tim says:it’s revolution time…Reply
Pierre Lutgen IFBV-Belgerb says:The same questionable PCR testing is used to hide the inadequacies and failures of ACT (Coartem, Coarsucam, Artequick) malaria therapyReply
Rik Breuer says:I would go one step further and question the validity of the “finding” of the virus itself.I read the paper regarding the Chinese scientists in Wuhan taking a sample from “one” sick patient and “identifying” the new virus SARS-CoV-2. They didn’t isolate the virus (that never happened anyway), they didn’t fulfill Koch’s postulates and without a Gold Standard they couldn’t prove that this “virus” is causing an flu like illness which can’t be clinical diagnosed.I would say, this whole Covid19-pandemic is a scam from beginning to end.I have looked up excess mortality in US and Germany. There is none.Reply
Kevin Corbett says:I agree with you.Reply
Terrence Joseph Bennett says:Thank you for doing science!
Shame on those who prefer Crapitalism to truthReplyheusenbeck says:Why is “A) BACKGROUND” a major concern with the corman drosten paper?“According to BBC News [4] and Google Statistics [5] there were 6 deaths world-wide on January 21st 2020 – the day when the manuscript was submitted. Why did the authors assume a challenge for public health laboratories while there was no substantial evidence at that time to indicate that the outbreak was more widespread than initially thought?”To quote Pieter Burger from another paper he released in april 2020:“It was common knowledge that some strain of coronavirus-sooner or later-was going to cause a pandemic. It was known since the SARS-CoV-outbreak in 2003. In 2013 and 2015, the world was informed that a variant of SARS-CoV in bats was emerging as a threat for humans.”
[https://www.researchgate.net/publication/341120750_A_SARS-like_Coronavirus_was_Expected_but_nothing_was_done_to_be_Prepared]Like any virologist Drosten warned about the threat of emerging infectious diseases and as we know, he co-discovered SARS-CoV1, developed the first diagnostic test and contributed to research on MERS early on. So i dont know what the implification of your “major concern A)” is. Do you expect him or any scientist working in this field, with the necessary resources, to sit back and check google statistics till death rate is alarming?Reply
Before we are all robotized in near future, it is time to wake up. A global Consortium of Doctors and Nurses backed by the people is the solution.Together, We Can Make A DIFFERENCE!
IT IS NOW OR NEVER…………ReplyBusu says:Thank you for this important job!
What are your next steps? In Germany there are no reports about this study, I don’t know are there to prevent this study.ReplyFranz says:Thanks for your huge effort giving a review on this paper. Would be great to get some more information about practical work of authors done on detection of SARS-CoV-2.Reply
– One should rather honor that within days and largely based on alignments of SARS-like viruses such well-working PCR assays have been designed, one of which (E gene) is still the reliable backbone of diagnosticsTechnically:
– Why discuss annealing temperatures when you can show experimentally that these PCRs work?
– Why discuss wobble bases when the RdRP PCR is demonstrably specific and sensitive (statement from practice: we had more than once a false base in the primer, which made a specific PCR-assay better instead of worse)
– Why discuss the ‘right’ amount of primers? If the reverse RdRP primer is too weak (too short) and contains wobble bases, then it is only logical to use more of them [higher concentration]
-> PCR IS MUCH MORE EMPIRICAL THAN DESIGN THEORY!
Auf deutsch: “Entscheidend ist, was hinten rauskommt!”Sensitivity & Specificity:
– In the January publication sensitivity (5 copies) and specificity (other cold pathogens do not make a signal) were determined experimentally.
– What is a diagnostic PCR? To find negative samples negative and find positive samples. These three PCR assays do this reliably.Improvements since Jan:
– For RdRP an interaction with human DNA had been overlooked. The RdRP assay has different primers since March.What has been researched since Jan?
– Well over 100,000 viruses have been sequenced since January and they are virtually all identical.
– The diagnostic regions were rarely affected by mutations (once Roche cobas and once CDC).Conclusion:
– Without ‘new’ results (sequences) there are few needs for new assays [because the current ones work]Kind regards JoergReplyMax says:same her – much fuzz about something that doesn’t matter in practice. To the “reviewers”:.) you criticise that many of the steps of validation have been done with virtual data, but you never tried the assay on the bench. You have just gone through it “virtually”. You had months of time to proof that the assay does not work (in the lab, on the bench!), but you did not even try to do it..) why is it important for you to discriminate between a virus fragment and whole virus? how should it get there if not via infection? There are no virus-pcr-detection systems, that detect the “whole virus”, it’s always a specific fragment you are searching for..) There are no Wobbly-positions in the 2019-nCoV specific primer..) there is no SOP and this could be improved, yes. but drosten gives information about sample preparation, primer-sequences, limits of detection and setup for PCR. Any routine lab can handle this information and setup their tests..) The cT value already has been corrected by drosten in an interview to a recommendation to cT ~30. But as this differs between labs, it should not be taken too precise, but validated by the labs. much more important are control samples, which are taken by serious labs anyway – positive virus sample, positive human DNA-sample, negative load,..stay safe!Reply
.) you criticise that many of the steps of validation have been done with virtual data, but you never tried the assay on the bench. You have just gone through it “virtually”. You had months of time to proof that the assay does not work (in the lab, on the bench!), but you did not even try to do it.ANSWER:
We have now several labs worldwide, which report that the C-D test does not work in their labs and generates false positive results in different PCR machines. So this test cannot be sold as the golden standard.QUOTE:
.) why is it important for you to discriminate between a virus fragment and whole virus? how should it get there if not via infection? There are no virus-pcr-detection systems, that detect the “whole virus”, it’s always a specific fragment you are searching for.ANSWER:
Bad news for virus diagnostics. PCR virus diagnostics should only be to support differential diagnosis to exclude other virus diseases. It should never be used as a screening device, since virus-parts are inhaled all day long and trapped in the mucous of nose and lungs, exactly there were the samples are taken from.QUOTE:
.) There are no Wobbly-positions in the 2019-nCoV specific primer.
.) there is no SOP and this could be improved, yes. but drosten gives information about sample preparation, primer-sequences, limits of detection and setup for PCR. Any routine lab can handle this information and setup their tests.ANSWER:
Bad science and even worse for diagnotics. There must be an online SOP, which must be used in all labs, germany-wide, world wide. And the SOP should be online update when required.QUOTE:
.) The cT value already has been corrected by drosten in an interview to a recommendation to cT ~30. But as this differs between labs, it should not be taken too precise, but validated by the labs. much more important are control samples, which are taken by serious labs anyway – positive virus sample, positive human DNA-sample, negative load.ANSWERS:
When Ct is different Germanwide, worldwide, nothing can be compared. This is very bad science.QUOTE:
stay safe!ANSWER:
Stay free!Reply- 3) this can be enhanced, yes, but does not make the test itself bad. every trained lab-worker handling with pcr knows what to do, when getting drostens publication!4.) well, this is fact with every PCR, and also every other (diagnostic) test. unless you use the exact same reagents (including LOTs) and the same equipment (and you can never do this in two different labs) you will have differences. Thats what trained lab-workers are here and you must not let beginners/student or simply untrained personnel do the diagnostics! even in the same lab you have to re-evaluate and re-validate your SOPs from time to time, also if you do not change anything. thats science. thats good science. relying on SOPs someone else made up with different reagents (included buffer solutions from different companies etc) is bad science.Reply
Lorenzo says:Max, how can a lab practically validate its results? You talk about control samples, I assume a _certain_ positive and a _certain_ negative, how can a lab acquire these? Regarding the samples I’m thinking about actual swabs material with a mix of human and multiple bacterial and viral genomes. I’ve read papers where water with just individual viruses was used as a negative, it seems quite different from the real world situation.Reply
Overall:
– One should rather honor that within days and largely based on alignments of SARS-like viruses such well-working PCR assays have been designed, one of which (E gene) is still the reliable backbone of diagnosticsANSWER:
Indeed, but wouldn’t it be nice to know what it is detecting? It was not molecularly validated. It may detect any coronavirus and probably other viruses as well.QUOTE:
Technically:
– Why discuss annealing temperatures when you can show experimentally that these PCRs work?
– Why discuss wobble bases when the RdRP PCR is demonstrably specific and sensitive (statement from practice: we had more than once a false base in the primer, which made a specific PCR-assay better instead of worse)
– Why discuss the ‘right’ amount of primers? If the reverse RdRP primer is too weak (too short) and contains wobble bases, then it is only logical to use more of them [higher concentration]
-> PCR IS MUCH MORE EMPIRICAL THAN DESIGN THEORY!
Auf deutsch: “Entscheidend ist, was hinten rauskommt!”ANSWER:
We agree, but the Test is designed so sloppy, we simply don’t know what positive results mean.QUOTE:
Sensitivity & Specificity:
– In the January publication sensitivity (5 copies) and specificity (other cold pathogens do not make a signal) were determined experimentally.
– What is a diagnostic PCR? To find negative samples negative and find positive samples. These three PCR assays do this reliably.ANSWER:
No, there is nothing in the test to exclude other viruses.QUOTE:
Improvements since Jan:…ANSWER:
Irrelevant for our external review report.QUOTE:
What has been researched since Jan?
– Well over 100,000 viruses have been sequenced since January and they are virtually all identical.ANSWER:
He means “puzzled together from 200 bp pieces” using a prespecified SARS-CoV-2 template.QUOTE:
– The diagnostic regions were rarely affected by mutations (once Roche cobas and once CDC).ANSWER:
There are now over 30’000 difference Sequences collected, including indels, in glue: http://cov-glue.cvr.gla.ac.uk/#/home So, if 100,000 sequences have been collected, 1 in 3 is different.QUOTE:
Conclusion:
– Without ‘new’ results (sequences) there are few needs for new assays [because the current ones work]ANSWER:
We have now reports from all over the world that the PCR test does not work as described in the Corma-Drosten paper. Positive often even depends on the instruments used. This test should not be propagated as the golden standard, because it is not golden.Reply
– One should rather honor that within days and largely based on alignments of SARS-like viruses such well-working PCR assays have been
designed, one of which (E gene) is still the reliable backbone of diagnosticsANSWER (ADDITIONALLY TO DR.BORGER’S ANSWER):
There was NOR REAL nCoV2019 positive control in the test validation.QUOTE:
Technically:
– Why discuss annealing temperatures when you can show experimentally that these PCRs work?
– Why discuss wobble bases when the RdRP PCR is demonstrably specific and sensitive (statement from practice: we had more than once a false base in the primer, which made a specific PCR-assay better instead of worse)ANSWER:
This is not a good laboratory practice – normally you can create an PCR that is optimized for the gen target to be detected, especially in a case where sequences are available in the Genbank (or viruses GISAID). The Chinese CDC managed to fulfill ths aspect for both the primer pairs and probes for their very early PCR, so why did the authors of the cormandrosten PCR publication such a weak PCR compared to the Chinese scientists?QUOTE:
Why discuss the ‘right’ amount of primers? If the reverse RdRP primer is too weak (too short) and contains wobble bases, then it is only logical to use more of them [higher concentration]ANSWER:
There is no need to include wobble bases in a PCR when the correct gene sequence is known (se figure 2 in the Cormandrosten paper). One can easily select a primer pair that fits and can be used at low concentrations with the optimal melting temperature around 60°C by one of the many good database “Primer design” tools . In daily laboratory practice you create 2-3 different primer pairs for the same target, order them and then then you test them in the lab with correct positive and negative controls to figure out which pair works best. After further validation with “real” samples (tissue, swabs, cell culture, etc. you can publish it. Well, maybe this it common practice for “normal” labs
>>>> only, maybe not for TOP-virology labs….<<<<<Reply
wilhelm Lehberger says:Es sollte selbstverständlich sein, dass im wissenschaftlichen Bereich und gemäß dem Anspruch einer demokratischen Gesellschaft Positionen ausgetauscht, geprüft und insbesondere im Falle derart gravierender Auswirkungen öffentlich erörtert werden können. Dies findet nicht statt und findet sich auch hier nur ansatzweise wieder.
Besorgte Nachfragen werden ignoriert und zu oft treten die übelsten menschlichen Eigenschaften zutage – denunzieren und diffamieren. Wenn also ein Ehepaar vor einer Reise Antigentests und in zeitlichem Abstand einen PCR-Schnelltest beim Hausarzt vornimmt, vorsorglich 3 Wochen private Kontakte ausschließt und der PCR-Test dann 1x negativ + 1x positiv ausfällt, dann ist das angesichts der hohen Infektionsgefahr nicht zu erklären. Das ist kein Einzelfall und nach dem Informationsstand nicht zu erklären. Es traten auch in der folgenden Quarantäne (die das Gesundheitsamt erst nach Ablauf und 4 Wochen nach dem Test anordnete) keine spezifischen Symptome auf. Was heißt das nun? Wurde man nun infiziert oder nicht? Wenn ja, ist man dann “geheilt” oder “immun”? Bringt dazu ein erneuter Antigentest Erkenntnisse? War hingegen der Test falsch, bedeutet das ca.320 € für die Tests und 3.500 € Stornokosten (die Versicherungsbedingungen wurden zum 1.9. angepasst). Faktisch ist damit die Reisefreiheit auf unbestimmte Zeit aufgehoben, weil sie keinen “richtigen” PCR-Test innerhalb von 24 Std. nicht bekommen. Die “offizielle” Antwort wäre sicher, man solle ja nicht verreisen (gilt nicht für Pendler u. Geschäftsleute). Da muss man sich über Proteste nicht wundern und schon sind wir wieder (auch hier) bei Rechtsradikalen und Verschwörungstheoretikern. Da empfehle ich doch den Blick über den linken Zaun, da gibt es erhebliche Zweifel bis zum Protestaufruf. Und wenn Klaus Schwab und das Weltwirtschaftforum Verschwörungstheorien befördern (wie das in einem Kommentar anklang),zu dem u.a. Minister und Staatschefs anreisen, dann sollte das die Politik klarstellen. Dazu die alte JUSO-Frage: Wem nützt es? (Und wer zahlt es?)ReplyV. Scholz says:Dieser Test ist ein Zufallszahlgenerator.RA Fuellmich redet nicht umsonst vom “vorsätzlichen sittenwidrigen Betrug”.Reply
Das finden sie in allen Charts.
Erklaerung: Der qPCR-Test mag eine gewisse Fehlerrate (je nach Labor 0,x – 1,y%?) haben, aber er zeigt NICHT Zufall an und es werden auch mehr in’s Krankenhaus eingeliefert, wenn die Rate Positiv-Getestet/Anzahl-Tests hoeher ist. Das koennen sie auch bei ordentlichen Datenaufarbeitungen deutlich erkennen: https://www.querschuesse.de/corona-faktencheck/Wut und Enttaeuschung ueber Einschraenkungen, WIE Entscheidungen getroffen wurden, ist verstaendlich, aber ein schlechter Ratgeber!Positiv: in D stagnieren oder sinken die Infektionsraten OHNE, dass das Gesundheitssystem nur in die Naehe der Ueberlastung kam. Wir sind ziemlich sicher aus dem Groebsten raus!Immer locker bleiben, oder verklagen? JoergReply
Oscar Drinnon says:The use of bald-faced lies about an epidemic to conceal a political agenda was exemplified as early as 1968 in Stanely Kubrick’s famous film “2001, A space odyssey”.The android-like bureaucrat “Dr. Heywood Floyd” informs moon base scientists that an epidemic is being used as a cover story for the news blackout after the discovery of the black monolith. He further informs them that they will have to sign a written oath of secrecy:‘– https://www.youtube.com/watch?v=PdbhZBpqNPM –‘Reply
Michael says:Eurosurveillance has reacted:
https://www.eurosurveillance.org/content/10.2807/1560-7917.ES.2020.25.48.2012031Let’s hope science wins!ReplyGyarmati says:No way. They will reject the above paper and the retraction demand. The narrative cannot be threatened.Reply
Frank Visser says:This contradicts your story that the C/D PCR is defective, is completely aspecific and can’t even work:Comparative Performance of SARS-CoV-2 Detection Assays Using Seven Different Primer-Probe Sets and One Assay Kit“We found that the most sensitive assays were those that used the E-gene primer-probe set described by Corman et al. (V. M. Corman, O. Landt, M. Kaiser, R. Molenkamp, et al., Euro Surveill 25:2000045, 2020, https://doi.org/10.2807/1560-7917.ES.2020.25.3.2000045) and the N2 set developed by the CDC (Division of Viral Diseases, Centers for Disease Control and Prevention, 2020, https://www.cdc.gov/coronavirus/2019-ncov/downloads/rt-pcr-panel-primer-probes.pdf). All assays tested were found to be highly specific for SARS-CoV-2, with no cross-reactivity with other respiratory viruses observed in our analyses regardless of the primer-probe set or kit used.”Reply
https://www.eurosurveillance.org/content/10.2807/1560-7917.ES.2020.25.21.2001035Quote:
Our strategy during establishment was to use a synthetic target for the SARS-CoV-2 E gene assay, while validating amplification of a full virus genome RNA using the RdRp assay that is specific for both, SARS-CoV and SARS-CoV-2, with the latter not being available to us in the form of an isolate or clinical sample at the time. Based on experimental validation, it later turned out that the mismatched base pairs do not reduce RT-PCR sensitivity and are not to be seen as the reason for somewhat higher Ct values with the RdRp assay as compared to the E gene assay [3]“More mismatches? No problem, test works, we assure you”.Reply
Ale vd sluis says:nu maar weer hopen of niet, dat u uw inteletuaeel vermogen ten positive weet te aan te wenden. ik begrijp de frustratie maar denk echt dat er betere manieren zijn om uw gelijk aan t tonen als dat er al uberhaupt al toe doet.Kom op peter, waar blijft die toegevooegde waardeReply
Robert says:Heeft u bij Peter in het dorp gewoond? Aan uw Nederlands te lezen heeft u in ieder geval niet op dezelfde school gezeten. Mag ik u vragen wat u zelf vindt van de toegevoegde waarde van uw opmerkingen in een verder behoorlijk volwassen discussie?Reply
A. van der Sluis says:Op uw eerste vraag: ja. Uit mijn nederlands trekt u de verkeerde conclusie. Het antwoord op uw tweede vraag moet u misschien zoeken in het artikel in die Zeit; Shitstorms Steht den Wissenschaftlern bei! (Heeft u ongetwijfeld gelezen.) Het is welliswaar een epinierend artikel en ik vermoed dat u het als niet relevant zult bestempelen?Reply
Emalsen says:No mention that the protocol in question was updated or of any further development in testing. Instead the authors, in comments above, made clear that their criticism aims solely at the original paper from nearly a year ago.
In wich scale this specific protocol was or might still being used, the report unfortunately does not investigate. So any impact of misdiagnosis, even if they prove this old protocol eventually somehow guilty, stays unfounded. I wonder what they try to gain here?Reply
https://www.eurosurveillance.org/content/10.2807/1560-7917.ES.2020.25.48.2012031#html_fulltextP.S.: Announced changes via Christian Drosten’s podcast don’t count. We want it in written official form.ReplyRob says:That’s an issue that could be resolved by one short email from every test kit’s manufacturer, if it’s not already included in their literature. We already know the cyclic thresholds are way too high for all the tests, I suspect they probably contain many if not most or all of the issues raised here – easy enough to find out though.Reply
Willy Schmid says:Please try to stop testing worldwide contacting the right persons/organisations who are ableto do this. By now, almost everybody knows that the PCR-test is not good at all. But in all countries, they continue to test, making more lockdowns. Now testing is even made with a quick test, which is less precise than the PCR-test according to the manufacturer Roche. Thanks.Reply
Y. Gielens says:I have not seen an analysis of the effects of variability in the sampling procedure, sample storage and sample extraction on the test results. (These are all steps prior to the test proper).Examples of variables:
–Design and material of swabs. Are they fit for purpose? Have they been tested for contaminants that can cause false-positives? Is the production process controlled?–Nature and amount of sample removed onto the swab from the oral or nasal cavity: (a) what matrix is actually sampled? Mucus? Water? Cell layers? (b) How much sample is removed, measured in microliters (fluid) or micrograms (solid)?–What is the extraction variability from the matrix and from swab to swab?–Effect of sampling site (oral or nasal). Is one targeted or both? Why?–Stability of the molecular species to be anayzed (oligonicelotides?) in the sample after collection and during transport: temperature, mechanical stress?–Efficiency of extraction from the sample matrix/swab of the molecular entities(oligonucleotides) to be analyzed ?Have all of these factors been tested, for example by spike/recovery experiments?If two subjects “A” and “B” have the same amounts of RNA at the nasal/oral sampling sites, but subject “A” tests positive and subject “B” does not, the discrepancy could be due to variability in any of the above factors. Just one example: removing double the amount of sample material onto the swab from the nose and throat of A compared to B would suffice.These are just technical points apart from the main issue of whether the molecules being analyzed have amy clinical relevance. But as long as (incredibly)such tests are still being performed, they have to be considered.ReplyEllis Mulder says:Thank you so much! Anyone should know this! I’m so proud of you all!Reply
Christl Meyer says:Thank you. It´s the same with testing for “HIV”.
I am fighting since more than 20 years for truth.
https://www.academia.edu/11649973/GENE_OR_VIRUS_IN_HEALTH_AND_DISEASE_ITS_ALL_ABOUT_SELF_AND_NON-SELFReplyMichael says:“The E gene primers also detect a broad spectrum of other SARS viruses.”Sollte da nicht “corona viruses” stehen anstatt “SARS viruses”?Passt auch besser zum Rest des Paragraphs“Still, SARS-CoV1 and SARS-CoV-2 have two highly specific genetic fingerprints, which set them apart from the other coronaviruses. First, a unique fingerprint-sequence (KTFPPTEPKKDKKKK) is present in the N-protein of SARS-CoV and SARS-CoV-2 [13,14,15]. Second, both SARS-CoV1 and SARS-CoV2 do not contain the HE protein, whereas all other coronaviruses possess this gene [13, 14].”Reply
Jack Leenders says:Deep respect for your profound work, fingers crossed for the impact
ReplyGeam Aston Martin V8 Convertible 1978 says:I constantly emailed this blog post page to all my contacts, because if like to
read it after that my contacts will too. https://anunturi-parbrize.ro/index.php?cauta=geam+aston+martinReplyHans Kleber says:debunked again
https://twitter.com/MarionKoopmans/status/1333002771738611712this time from a Virologist.Reply
2) it seems like Marion Koopmans either didn’t read our review report or has any other kind of Attention Deficit Problem – non of our concern, here is an extensive response by Kevin Mckernan: https://threadreaderapp.com/thread/1333846936332464129.html
3) I personally can’t take anyone serious who references the low-tier “virology down under” blog by Ian M. Mackay, the analogy therefor would be: To reference Alex Jones & Infowars and declare it as a valid resource for any scientific discussion.Reply
Contra el Encierro says:Dear authors,Thank you so much for you work and for your very good idea in making this live web site (with comments): this is what is needed and this is what they do not want: an open in-depth debate. You are at the highest point of your life. Congratulations.We have just published a full translation of your report in Spanish:
https://contraelencierro.blogspot.com/2020/12/review-report-corman-drosten-et-al.htmlWe will follow comments and news in your site very closely. Thanks again:https://contraelencierro.blogspot.comCourage!Reply
Meikel says:Liebes Team!Vielen Dank für Ihre Arbeit!
Ich hoffe sehr, dass sie zu einem kritischen Blick auf die in meinen Augen strategielose Testung bzgl. SARS-Cov2 führt.
Ich kann nicht beurteilen, ob der Corman-Drosten-Test gut oder schlecht ist. Vielleicht musste es ja wirklich schnell gehen, vielleicht auch nicht (es scheint eher Letzteres).
Prof. Kämmerer erwähnte in der Sitzung 28 (?) des Corona-Ausschuss, dass sinngemäß „ein Praktikant den Test so eben bestanden hätte“.
Natürlich ist es fatal, einen Test mit der Schulnote 4- (so nenne ich da jetzt mal) einzusetzen, um solch gravierende und in die Grundrechte eingreifende Massnahmen zu begründen.
Soweit ich mich über PCRs informieren konnte, weiß ich, dass „sie quasi jede noch so geringe Kleinigkeit finden könnten“, bei geringer Prävalenz mehr false-positiv generieren und zur DIAGNOSTIK nicht gedacht waren.
Trotzdem sind PCRs sicherlich sehr gut, wenn man weiß wann und wie man sie einsetzt und ebenso versteht, dass Ergebnis zu interpretieren.
Letzteres scheint mir Letzteres das derzeit das größte Problem zu sein.
Wie sollte es denn normalerweise in medizinischer Diagnostik laufen?
Der Patient geht zum Arzt, weil er „Probleme“/Symptome hat!
Der Arzt macht seine Anamnese und mögliche körperliche Untersuchungen.
Daraus entsteht eine Hypothese bzgl. der Diagnose, die womöglich noch durch technische Diagnoseverfahren (hier die PCR) unterstützt oder verworfen werden kann.
Kommt nun z.B. ein Pat. in eine Praxis und klagt über die typischen SARS-CoV2 Symptome, wäre der Arzt natürlich geneigt oder verpflichtet einen Test zur Abklärung einzusetzen.
Wählt er/sie nun die PCR, erhält er folgende mögliche Information …
a) Test negativ, es folgt wohl die logische Erklärung „KEIN CORONA“, der Pat hat eine andere Erkältungserkrankung.
b) Test Ergebnis POSITIV
b.1) Der Arzt erhält zum Ergebnis POSITIV auch den CT Wert mit z.B. 40 …
Das Problem beginnt … das Gesundheitsamt verzeichnet den Patienten als CORONA-infiziert, der CT von 40 sagt aber eigentlich … nicht mehr krank oder „nur mal was abbekommen“ … die Infektiösität ist laut dieser Studie (https://www.nature.com/articles/s41467-020-19802-w) quasi 0. Es folgt trotzdem QUARANTÄNE! K1 Personen desgleichen!
b.2) CT ist bei pos. Ergebnis <30 (ich bin nett und nehme den vom RKI empfohlenen Wert) … ja, hier darf man annehmen, das der Pat. wohl infiziert ist bzw. die Viruslast Massnahmen wie eine Quarantäne rechtfertigen würden.Jeder Test ist nur so gut, wie er im Gesamtkontext eingesetzt wird. Auch ein wesentlich besser „gebauter“ PCR Test kann immer noch falsch eingesetzt werden, wenn die Fragen „Wann und wen teste ich?“ und „Wie interpretiere ich das Ergebnis inkl. CT?“ in Bezug auf die Anamnese des behandelnden Arztes nicht bedacht werden! Hier scheinen mir noch viel größere Fallstricke zu lauern.
Problematisch an dem Corman-Drosten-Test wäre für mich auch, sollte er wirklich so „schlecht“ sein, dass er als GOLD-Standard definiert ist.
Jeder eigentlich „bessere“ Test wird durch seine Abweichung zum GOLD-Standard somit als schlecht bewertet.
So hätte wohl auch der AntiGen-Test eine deutlich bessere Reputation, wenn er nicht mit den Positivraten des Drosten-PCR verglichen werden würde. Dazu z.B. auch jenen Diskussion (https://infekt.ch/2020/10/covid-19-antigen-test-schlechter-als-pcr-wirklich/ )oder auch die Aussagen von Frau Prof. Kämmerer im Corona-Ausschuss.
Nochmals danke ich Ihnen allen für die Zeit und Mühen die Sie in dieses Paper gesteckt haben! Diese Arbeit ist einer der letzten Hoffnungsschimmer, die ich noch habe, dass sich „die Dinge“ noch ändern könnten.Replysteve gangloff says:Can you provide a PDF of this document – please forward to my email below. thanksReply
Wolfgang E. says:Thank you for doing science!Reply
Emanuel E. Garcia, MD says:An exemplary scientific critique of the subject matter that deserves worldwide attention. It is abundantly clear to me that the so-called pandemic was driven by non-scientific agendas. Your analysis of the Corman-Drosten report, unlike most of what we get about COVID, represents real science. Thank you.Reply
五十嵐敬冶 says:私も、このレポートが発表される以前に、ドロステンのRT-PCR法には「重大な欠陥」があると気付きました。
彼のプロトコルは2003年のSARSウィルス株のゲノムを参照して策定しています。
中国チームが採取したという検体のゲノムではありません。
「推測による検出方法」なのです。
それ故に、(一人でも多く「陽性者」を捕まえられるように。自説プロトコルが結果を示せるように。)必要以上にCt値(増幅サイクル数)が高く(45に)設定されているのです。ModEdit: Translation:Before this report was published, I also realized that Drosten’s RT-PCR method had “serious flaws.”
His protocol is based on the genome of the 2003 SARS virus strain.
It is not the genome of the sample collected by the Chinese team.
It is a “guessing detection method”.
Therefore, the Ct value (amplification cycle number) is set higher (to 45) than necessary (so that as many “positive people” as possible can be caught. So that the self-explanatory protocol can show the result.).ReplyCharles says:Hi,Thank you for the brave work you are doing, hopefully you can answer these questions i have ?At Amphia Hospital and Bravis Hospital, total
nucleic acids were extracted for RT-PCR after an external
lysis step (1:1 with lysis binding buffer; Roche Diagnostics,
Almere, Netherlands), using MagnaPure96 (Roche) with
an input volume of 500 μL and output volume of 100 μL.The extraction was internally controlled by addition of a
known concentration of phocine distemper virus (PDV) ??.Is this a normal procedure that Koopmans et.al. use ?And what are they selling at https://www.european-virus-archive.com/virus/human-2019-ncov-isolate if there is no Sars-Cov-2 isolatehttps://www.european-virus-archive.com/evag-portal/evag-partners/rivm ?Thank you and we keep monitoring themCharles..Reply
“Culture Medium: DMEM (Dulbecco’s Modified Eagles Medium) (PAA, Cölbe, Germany) with 4.5 g/L Glucose (PAA) supplemented with 10% Foetal Bovine Serum (PAA), 1% Penicillin/Streptomycin 100 x concentrate (Penicillin 10000 U/mL, Streptomycin 10 mg/ mL) (PAA), 1% L-Glutamine 200 mM, 1% Sodium Pyruvate 100 mM (PAA), 1% MEM nonessential amino acids (NEAA) 100 x concentrate (PAA) – Subculture routine: Detachment with an EDTA/trypsin mixture (PAA)”ReplyJan says:Can you explain this for simple humans if possible ?
Reply
Amplification was done in a 7500SDS (Thermofisher) with a
cycling profile of 5 min at 50°C, 20 s at 95°C, 45 cycles of
3 s at 95°C, and 30 s at 58°C.At Elisabeth-TweeSteden Hospital:Amplification with Rotorgene
(QIAgen) consisted of 5 min at 50°C and 15 min at 95°C
followed by 45 cycles of 15 s at 95°C, 30 s at 60°C, and 15 s
at 72°C.Partly based on these data, SARS-CoV-2 was
concluded to have already spread in the population in the
province of North Brabant, which led to a change of
policy, in which containment measures were complemented by targeted physical distance measures, starting in the south of the Netherlands initially and later
comprising the whole country.ReplyRJ says:Fix your zip file. The dcox and pdf files will not extract!Reply
оксана says:огромная благодарность за проделанную работу!теперь все пазлы и сложились.ModeEdit: Translation:huge gratitude for the work done! now all the puzzles have come together.Reply
Su Obreen says:Finally! What ‘VIRUSWAARHEID.NL’ has already established from the start of the COVID19 gate is now being brought to light by other celebrities / prominent figures.Reply
jac says:Deep respect for your work.Reply
E says:FYI in this article the “friend” of P Borger claim that his (Marc Bonte UmcU) rapid test has 100% specificity.Please stop this ongoining noncense!https://www.medrxiv.org/content/10.1101/2020.10.16.20214189v1
https://www.medrxiv.org/content/10.1101/2020.10.16.20214189v1Reply
“In medical diagnosis, test sensitivity is the ability of a test to correctly identify those with the disease (true positive rate),
whereas test SPECIFICITY is the ability of the test to correctly identify those without the disease (true negative rate)”Second: So in your cited paper the authors state [as I understand] that in all cases a negative antibody Covid19 Rapid assay was also in the RT-PCR test negative [95%CI: 99.7-100%].What should be wrong with that? Or what exactly is “ongoing nonsense” in your opinion? I don’t get it?Kind regards JoergReply
1) PCR + and AB + (ca. 90%) SENSITIVITY, true positive rate
2) PCR + and AB – (ca. 10%) false negatives related to AB-test
3) PCR – and AB – (100%) SPECIFICITY, true negative rate
4) PCR – and AB + (0%) false positives related to AB-testReply
And I guess you are expecting a second and about the same letter from ECDC in the very, very far future?.Reply
https://cormandrostenreview.com/false-positives-consequences/I have forwarded your remarks to Dr. Peter Borger, but I think I have answered it for him – he would give you most probably the same answer.Another fact: Eurosurveillance has set-up a comments section now too at their journal portal, it’s new. Is it in reaction to our approach to ensure more discussion and transparency?Reply
U had een bezwaar mogelijkheid tegen het besluit om niet te openbaren. Heeeft u deze gebruikt, zo, nee, waarom niet?Kunt u mij zeggen waarom mijn reply onder nr84 op Robert niet werd geplaatst? Dubbel naam/mail gebruik misschien?
Als het geplaatst kan worden hoef ik daarover geen uitleg maar naar een inhoudelijk reactie ben ik erg benieuwd.
Freonlike groet oan dr.Peter Borger.Reply
Arturas Minajevas says:Thanks alot from Lithuania!Reply
meine zwei Kommentare vom 05.12. wurden geloescht? Warum?
Falls es kein Versehen war, bitte auch alle anderen Kommentare von mir loeschen!
Vielen Dank im Voraus und alles Gute
JoergReply- Tipps fuer die naechsten Beitraege/Blogs:
1) Eine Funktion fuer “neueste 10-20 Beitraege” in einer Spalte rechts oben oder ganz unten.2) Da es ein moderierter Blog ist (Du und andere?), ist es legitim zwischen ARGUMENTEN und MEINUNGEN zu sortieren. Bei kontroversen Themen mischt sich das ja meistens stark und fuehrt zur Unuebersichtlichkeit bzw lenkt von urspruenglicher Intention Argumente zu einem Fachthema zu diskutieren ab (gibt’s auch als Troll-Angriffs-Strategie).
Es waere also moeglich ganz krude Aussagen (aber auch die vielen “prima, weiterso” Bekundungen auf eine “Plauderei-Version” zu diesem Blog-Post zu schieben (“Review report Corman-Drosten et al. Eurosurveillance 2020 Chitchat” und die Fachargumente hier zu lassen …
Aber macht sicher auch viel Arbeit … Hoffentlich lohnt es sich fuer Euch (die viele Arbeit).LG JoergReply
Steffen Jurisch says:Dezember, 7., 2020 Danke für Ihre Arbeit. Leider interessiert diese die Psychoparhen von Politiker keinen Deut – sie machen weiter und drehen die faschistoide Diktaturschraube noch fester an und die Masse bejudelt es noch, weil sie dumm und zu faul ist, sich zu informieren.
Ich hoffe wirklich das der Spuk bald ein Ende hat und die Gerichte sämtliche Politiker und Mitläufer zur Verantwortung zieht…ReplyJo-Anne van Westing says:Goedemorgen, ik ben zooooo blij met alle artsen/wetenschappers die wat van zich laten horen. Ik merk in mijn eigen omgeving dat mensen moe van me worden als ik weer met iets nieuws kom dat aangeeft dat het allemaal niet klopt wat er nu gaande is. Mijn vreemde onderbuik gevoel startte met het uit de handel halen van het boek betreft vaccinaties.Ik stuur dit beoordelingsrapport naar een paar huisartsen in mijn omgeving. Misschien kunnen meer mensen dat doen?Ik vind het zo erg voor jongeren…
Ik hoop, ik hoop, ik hoop…ReplyDaniel says:I hope, I pray, I stand and I fight in heart and spirit for our children and for life!I pray that all doctors who are not compromised and scared step out and forward so together we can make an end to this horrid nightmare imposed to mankind by a select club of psychopaths. Together with the Notice of Liability I believe we can take back our freedom and lives!Reply
Marrie says:It’s easier to fool the people than to convince them that they have been fooled.Reply
RALPH JACKSON says:An excellent report. Our British Government should be taking to the War Tribunal for crimes against its citizensReply
Dave says:Thank you for your excellent work.Here’s an interesting post highlighting some of the problems with the CDC primers earlier this year:https://tomeraltman.net/2020/03/03/technical-problems-COVID-primers.htmlReply
Pjotr says:This is a review of the paper and the therein described method, fine. But, for example in Germany, about 190 test laboratories are delivering PCR results. Do they all use the Corman test as it is described in the Corman paper? I don`t think so. I assume they took it as starting point and modified it. Therefore, it would be intersting to get the test methods from a number of laboratories and compare them,i.e., look at the primers, the cycle numbers and CT etc., they are using. The authors of the CormanDRostenReview should report the outcome of such a comparison of test methods. In addition, was cross laboratory validation done between certified laboratories? Does anyone have a report on this? A cross lab validation could be, e.g., lab A analyzed a large number of samples and sends them to further laboratories B, C, D etc. for blinded analysis (not knowing the results of lab A). Labs B, C, D etc. send their results to lab A. Lab A compares the results, i.e., what is the %agreement on positive and negative PCR results between the labs. Any data available on such a comparison?Reply
Naar nu blijkt zijn de grote hoeveelheid coronatesten uiterst lucratief voor de artsen die vrijgevestigd zijn.
Deze geldstroom is tot op heden verborgen gebleven.Bij welke ziekenhuizen en artsen de honderden miljoenen precies beland zijn, is een zorgvuldig bewaard geheim. De NVMM, het RIVM, het LCDK en de GGD’en weten precies hoeveel testen elk lab heeft verricht.De labs zijn namelijk wettelijk verplicht dat te melden aan de GGD’en. Ook is er sinds 1 juni een IT-systeem (Coron-IT) waarbinnen de labs zulke data doorgeven aan de GGD en het LCDK. Bovendien houden de labs het RIVM op de hoogte met ‘virologische dagstaten’.Daardoor is eenvoudig te berekenen hoeveel omzet elk lab heeft gedraaid, en zou je schattingen kunnen maken van de extra inkomsten van de betrokken artsen. Maar al deze partijen weigeren transparant te zijn over de besteding van honderden miljoenen euro belastinggeld.Zie ook https://www.ftm.nl/artikelen/kluytmans-belang-coronatestenLet op:de volgende lucratieve scam gaan ze draaien met deze nieuwe sneltesten:
In Nederland heeft het OMT [..] beoordeeld dat de Panbio COVID-19-Ag rapid test (Abbott), de SARS-CoV-2 Rapid Antigen Test (Roche), BD Veritor COVID test (Becton Dickinson), Sofia SARS Antigen FIA (Quidel) en Standard F-Covid-19 Ag (SD Biosensor) gebruikt kunnen worden bij mensen met klachten in teststraten.KASSA!ReplyAli says:Your comment displays the very need for a Standard Operating Procedure (SOP) for all labs performing this diagnostic. Just one of the flaws found.Reply
Thomas says:Here a link to a study which strongly indicates that there was no (adequate) peer review:http://www.aukema.org/2020/12/meta-data-analysis-at.htmlIt says:
To assess commonality in the review and acceptance process at eurosurveillance.org, the author collected and analysed meta-data for all 1,595 publications since 01-Jan-2015…
…Except for this one Research article (on 22-jan-2020)(the CormanDrosten Paper, Author’s note), no other article has ever been reviewed and accepted within a single day since 2015.”ReplyHarold says:Drosten on the PCR Test in 2014:“Ja, aber die Methode ist so empfindlich, dass sie ein einzelnes Erbmolekül dieses Virus nachweisen kann. Wenn ein solcher Erreger zum Beispiel bei einer Krankenschwester mal eben einen Tag lang über die Nasenschleimhaut huscht, ohne dass sie erkrankt oder sonst irgend etwas davon bemerkt, dann ist sie plötzlich ein Mers-Fall. Wo zuvor Todkranke gemeldet wurden, sind nun plötzlich milde Fälle und Menschen, die eigentlich kerngesund sind, in der Meldestatistik enthalten. Auch so ließe sich die Explosion der Fallzahlen in Saudi-Arabien erklären. Dazu kommt, dass die Medien vor Ort die Sache unglaublich hoch gekocht haben.”https://amp2.wiwo.de/technologie/forschung/virologe-drosten-im-gespraech-2014-der-koerper-wirdstaendig-von-viren-angegriffen/9903228.htmlReply
Sam Laurey says:Nun, da Herr Drostens PCR-Test als Unsinn prooved, was sind nun die Folgen?
Wie kann man diese Weisheit in die Praxis umsetzen? Bald?
Freundlich grüßeund dank
viel Oder besser spät als nie/Sam LaureyAntwort
Chantal says:Hinterlasse eine Antwort
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