Acceptability and Preferences of Dry HR HPV Self-Sampling Mailed Kits Among Canadian Women: A Cross-Sectional Study.

Human papillomavirus (HPV), a sexually transmitted disease, is identified as the source of 99.7% of cervical cancers. Screening for cervical cancer using oncogenic HPV (high-risk [HR] HPV) detection is more sensitive than traditional cytology. However, few Canadian data exist on HR HPV self-sampling. OBJECTIVE: To evaluate the acceptability of HR HPV self-sampling by patients, the percentage of correctly collected samples, the return rate of mailed kits, and the HPV positivity rate in a population sample based on different cervical cancer risk factors. METHODS: We conducted an observational cross-sectional study on HPV primary cervical cancer screening with self-collected cervicovaginal samples through mail service. RESULTS: A total of 400 kits were mailed and 310 kits were returned, making a return rate of 77.5%. Of these, 84.2% of patients were very satisfied with this method and 95.8% (297/310) of patients would choose self-sampling over cytology as their primary screening method. All patients would recommend this screening method to their friends or family members. Of the samples, 93.8% could be analyzed correctly and the HPV positivity rate was 11.7%. CONCLUSION: In this large and random sample, there was a strong interest in self-testing. Offering HR HPV self-sampling could increase access to cervical cancer screening. The self-screening method could also be a part of the solution to reaching under-screened populations, in particular, those who do not have a family doctor or avoid gynaecologic exams because of pain or anxiety.

Natural spring water gargle samples as an alternative to nasopharyngeal swabs for SARS-CoV-2 detection using a laboratory-developed test.

The objective of this study was to validate the use of spring water gargle (SWG) as an alternative to oral and nasopharyngeal swab (ONPS) for SARS-CoV-2 detection with a laboratory-developed test. Healthcare workers and adults from the general population, presenting to one of two COVID-19 screening clinics in Montréal and Québec City, were prospectively recruited to provide a gargle sample in addition to the standard ONPS. The paired specimens were analyzed using thermal lysis followed by a laboratory-developed nucleic acid amplification test (LD-NAAT) to detect SARS-CoV-2, and comparative performance analysis was performed. An individual was considered infected if a positive result was obtained on either sample. A total of 1297 adult participants were recruited. Invalid results (n = 18) were excluded from the analysis. SARS-CoV-2 was detected in 144/1279 (11.3%) participants: 126 from both samples, 15 only from ONPS, and 3 only from SWG. Overall, the sensitivity was 97.9% (95% CI: 93.7-99.3) for ONPS and 89.6% (95% CI: 83.4-93.6; p = 0.005) for SWG. The mean ONPS cycle threshold (Ct ) value was significantly lower for the concordant paired samples as compared to discordant ones (22.9 vs. 32.1; p < 0.001). In conclusion, using an LD-NAAT with thermal lysis, SWG is a less sensitive sampling method than the ONPS. However, the higher acceptability of SWG might enable a higher rate of detection from a population-based perspective. Nonetheless, in patients with a high clinical suspicion of COVID-19, a repeated analysis with ONPS should be considered. The sensitivity of SWG using NAAT preceded by chemical extraction should be evaluated.

Validation of a Gradient Diffusion Method (Etest) for Testing of Antimicrobial Susceptibility of Aerococcus urinae to Fluoroquinolones.

Aerococcus urinae is a urinary pathogen with well-described resistance to fluoroquinolones. This study aimed to validate the gradient diffusion (GD) method (Etest) on cation-adjusted Mueller-Hinton agar with 5% sheep blood for testing the susceptibilities of Aerococcus urinae to the antimicrobial agents ciprofloxacin and levofloxacin and to compare the Etest to the broth microdilution (BMD) method from CLSI document M45-A3. Agar dilution (AD), as recommended by EUCAST, was used as an alternative reference method to arbitrate discrepancies or address technical issues. Aerococcus urinae isolates from urinary specimens were prospectively collected between June 2016 and December 2017 from six hospitals in Quebec, Canada, and identifications were confirmed using Vitek MS with the IVD 3.0 database. Of the 207 isolates tested using BMD, 37 (17.9%) showed trailing and 19 (9.2%) showed insufficient growth; these were tested using AD. Also, 38 isolates (18.4%) for ciprofloxacin and 13 isolates (6.3%) for levofloxacin showed a lack of essential or categorical agreement between the Etest and BMD and were also tested by AD. By use of a combined reference method (BMD or AD), the susceptibility rates of Aerococcus urinae were 82.6% and 81.6% for ciprofloxacin and levofloxacin, respectively. Categorical agreement between GD and the combined reference methods was 95.2% for ciprofloxacin and 97.1% for levofloxacin, with no very major error identified. Major and minor error rates were 0.6% and 4.3% for ciprofloxacin and 1.2% and 1.9% for levofloxacin. Overall, antimicrobial susceptibility testing (AST) using the Etest on sheep blood agar showed good agreement with the reference methods and can be considered by clinical laboratories wishing to perform AST on Aerococcus urinae isolates.

Comparison of saliva with oral and nasopharyngeal swabs for SARS-CoV-2 detection on various commercial and laboratory-developed assays.

The accurate laboratory detection of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a crucial element in the fight against coronavirus disease 2019 (COVID-19). Reverse transcription-polymerase chain reaction testing on combined oral and nasopharyngeal swab (ONPS) suffers from several limitations, including the need for qualified personnel, the discomfort caused by invasive nasopharyngeal sample collection, and the possibility of swab and transport media shortage. Testing on saliva would represent an advancement. The aim of this study was to compare the concordance between saliva samples and ONPS for the detection of SARS-CoV-2 on various commercial and laboratory-developed tests (LDT). Individuals were recruited from eight institutions in Quebec, Canada, if they had SARS-CoV-2 RNA detected on a recently collected ONPS, and accepted to provide another ONPS, paired with saliva. Assays available in the different laboratories (Abbott RealTime SARS-CoV-2, Cobas® SARS-CoV-2, Simplexa™ COVID-19 Direct, Allplex™ 2019-nCoV, RIDA®GENE SARS-CoV-2, and an LDT preceded by three different extraction methods) were used to determine the concordance between saliva and ONPS results. Overall, 320 tests were run from a total of 125 saliva and ONPS sample pairs. All assays yielded similar sensitivity when saliva was compared to ONPS, with the exception of one LDT (67% vs. 93%). The mean difference in cycle threshold (∆C t ) was generally (but not significantly) in favor of the ONPS for all nucleic acid amplification tests. The maximum mean ∆​​​​​C t was 2.0, while individual ∆C t varied importantly from -17.5 to 12.4. Saliva seems to be associated with sensitivity similar to ONPS for the detection of SARS-CoV-2 by various assays.

SARS-CoV-2 seroprevalence in health care workers from 10 hospitals in Quebec, Canada: a cross-sectional study.

BACKGROUND: The COVID-19 pandemic has disproportionately affected health care workers. We sought to estimate SARS-CoV-2 seroprevalence among hospital health care workers in Quebec, Canada, after the first wave of the pandemic and to explore factors associated with SARS-CoV-2 seropositivity. METHODS: Between July 6 and Sept. 24, 2020, we enrolled health care workers from 10 hospitals, including 8 from a region with a high incidence of COVID-19 (the Montréal area) and 2 from low-incidence regions of Quebec. Eligible health care workers were physicians, nurses, orderlies and cleaning staff working in 4 types of care units (emergency department, intensive care unit, COVID-19 inpatient unit and non-COVID-19 inpatient unit). Participants completed a questionnaire and underwent SARS-CoV-2 serology testing. We identified factors independently associated with higher seroprevalence. RESULTS: Among 2056 enrolled health care workers, 241 (11.7%) had positive SARS-CoV-2 serology. Of these, 171 (71.0%) had been previously diagnosed with COVID-19. Seroprevalence varied among hospitals, from 2.4% to 3.7% in low-incidence regions to 17.9% to 32.0% in hospitals with outbreaks involving 5 or more health care workers. Higher seroprevalence was associated with working in a hospital where outbreaks occurred (adjusted prevalence ratio 4.16, 95% confidence interval [CI] 2.63-6.57), being a nurse or nursing assistant (adjusted prevalence ratio 1.34, 95% CI 1.03-1.74) or an orderly (adjusted prevalence ratio 1.49, 95% CI 1.12-1.97), and Black or Hispanic ethnicity (adjusted prevalence ratio 1.41, 95% CI 1.13-1.76). Lower seroprevalence was associated with working in the intensive care unit (adjusted prevalence ratio 0.47, 95% CI 0.30-0.71) or the emergency department (adjusted prevalence ratio 0.61, 95% CI 0.39-0.98). INTERPRETATION: Health care workers in Quebec hospitals were at high risk of SARS-CoV-2 infection, particularly in outbreak settings. More work is needed to better understand SARS-CoV-2 transmission dynamics in health care settings.

Fast and Accurate Bacterial Species Identification in Urine Specimens Using LC-MS/MS Mass Spectrometry and Machine Learning.

Fast identification of microbial species in clinical samples is essential to provide an appropriate antibiotherapy to the patient and reduce the prescription of broad-spectrum antimicrobials leading to antibioresistances. MALDI-TOF-MS technology has become a tool of choice for microbial identification but has several drawbacks: it requires a long step of bacterial culture before analysis (≥24 h), has a low specificity and is not quantitative. We developed a new strategy for identifying bacterial species in urine using specific LC-MS/MS peptidic signatures. In the first training step, libraries of peptides are obtained on pure bacterial colonies in DDA mode, their detection in urine is then verified in DIA mode, followed by the use of machine learning classifiers (NaiveBayes, BayesNet and Hoeffding tree) to define a peptidic signature to distinguish each bacterial species from the others. Then, in the second step, this signature is monitored in unknown urine samples using targeted proteomics. This method, allowing bacterial identification in less than 4 h, has been applied to fifteen species representing 84% of all Urinary Tract Infections. More than 31,000 peptides in 190 samples were quantified by DIA and classified by machine learning to determine an 82 peptides signature and build a prediction model. This signature was validated for its use in routine using Parallel Reaction Monitoring on two different instruments. Linearity and reproducibility of the method were demonstrated as well as its accuracy on donor specimens. Within 4h and without bacterial culture, our method was able to predict the predominant bacteria infecting a sample in 97% of cases and 100% above the standard threshold. This work demonstrates the efficiency of our method for the rapid and specific identification of the bacterial species causing UTI and could be extended in the future to other biological specimens and to bacteria having specific virulence or resistance factors.

A Cluster of Three Cases of Hantavirus Pulmonary Syndrome among Canadian Military Personnel.

Hantavirus pulmonary syndrome (HPS) is a rare illness in eastern Canada. We present three cases of HPS among military personnel in Quebec. The three cases shared a common exposure to mouse excreta while engaged in military training in Alberta, a western province of Canada.

Comparison of rectal swabs and fecal samples for the detection of Clostridioides difficile infections with a new in-house PCR assay.

The detection of Clostridioides difficile infections (CDI) relies on testing the stool of patients by toxin antigen detection or PCR methods. Although PCR and antigenic methods have significantly reduced the time to results, delays in stool collection can significantly add to the turnaround time. The use of rectal swabs to detect C. difficile could considerably reduce the time to diagnosis of CDI. We developed a new rapid PCR assay for the detection of C. difficile and evaluated this PCR assay on both stool and rectal swab specimens. We recruited a total of 623 patients suspected of C. difficile infection. Stool samples and rectal swabs were collected from each patient and tested by our PCR assay. Stool samples were also tested by the cell cytotoxicity neutralization assay (CCNA) as a reference. The PCR assay detected C. difficile in 60 stool specimens and 61 rectal swabs for the 64 patients whose stool samples were positive for C. difficile by CCNA. The PCR assay detected an additional 35 and 36 stool and rectal swab specimens positive for C. difficile, respectively, for sensitivity with stools and rectal swabs of 93.8% and 95.3%, specificity of 93.7% and 93.6%, positive predictive values of 63.2% and 62.9%, and negative predictive values of 99.2% and 99.4%. Detection of C. difficile using PCR on stools or rectal swabs yielded reliable and similar results. The use of PCR tests on rectal swabs could reduce turnaround time for CDI detection, thus improving CDI management and control of C. difficile transmission. IMPORTANCE: Clostridioides difficile infection (CDI) is the leading cause of healthcare-associated diarrhea, resulting in high morbidity, mortality, and economic burden. In clinical laboratories, CDI testing is currently performed on stool samples collected from patients with diarrhea. However, the diagnosis of CDI can be delayed by the time required to collect stool samples. Barriers to sample collection could be overcome by using a rectal swab instead of a stool sample. Our study showed that CDI can be identified rapidly and reliably by a new PCR assay developed in our laboratory on both stool and rectal swab specimens. The use of PCR tests on rectal swabs could reduce the time for the detection of CDI and improve the management of this infection. It should also provide a useful alternative for infection-control practitioners to better control the spread of C. difficile.

Duration of isolation and contagiousness in coronavirus disease 2019 (COVID-19) patients receiving tocilizumab and dexamethasone: A case series.

We describe 10 patients with severe coronavirus disease 2019 (COVID-19) who received tocilizumab and dexamethasone. We correlated isolation duration with cycle thresholds (Ct) values of nucleic acid amplification tests, clinical state and viral cultures. Isolation duration exceeded 21 days for 7 patients due to positive viral cultures or Ct values <30.

Reinfection with SARS-CoV-2 in a patient undergoing chemotherapy for lymphoma: Case report.

BACKGROUND: COVID-19 is usually a time-limited disease. However, prolonged infections and reinfections can occur among immunocompromised patients. It can be difficult to distinguish a prolonged infection from a new one, especially when reinfection occurs early. METHODS: We report the case of a 57-year-old man infected with SARS-CoV-2 while undergoing chemotherapy for follicular lymphoma. He experienced prolonged symptomatic infection for 3 months despite a 5-day course of remdesivir and eventually deteriorated and died. RESULTS: Viral genome sequencing showed that his final deterioration was most likely due to reinfection. Serologic studies confirmed that the patient did not seroconvert. CONCLUSIONS: This case report highlights that reinfection can occur rapidly (62-67 d) among immunocompromised patients after a prolonged disease. We provide substantial proof of prolonged infection through repeated nucleic acid amplification tests and positive viral culture at day 56 of the disease course, and we put forward evidence of reinfection with viral genome sequencing.

HISTORIQUE: La COVID-19 est généralement une maladie limitée dans le temps. Toutefois, des infections et réinfections prolongées peuvent survenir chez des patients immunodéprimés. Il peut être difficile de distinguer une infection prolongée d'une nouvelle infection, particulièrement lorsque la réinfection se produit rapidement. MÉTHODOLOGIE: Les auteurs rendent compte du cas d'un homme de 57 ans infecté par le SRAS-CoV-2 alors qu'il était sous chimiothérapie pour soigner un lymphome folliculaire. Il a souffert d'une infection symptomatique prolongée de trois mois, malgré un traitement de cinq jours au remdésivir. Son état s'est finalement détérioré et il est décédé. RÉSULTATS: Le séquençage du génome viral a démontré que la détérioration finale de son état a probablement été causée par une réinfection. Les études sérologiques ont confirmé qu'il n'avait pas présenté de séroconversion. CONCLUSIONS: Le présent rapport de cas établit la possibilité d'une réinfection rapide (au bout de 62 à 67 jours) chez les patients immunodéprimés après une longue maladie. Les auteurs fournissent des preuves substantielles d'une infection prolongée par des tests répétés d'amplification des acides nucléiques et par des cultures virales positives au 56e jour de l'évolution de la maladie, et ils présentent des preuves de réinfection grâce au séquençage du génome viral.

Natural spring water gargle and direct RT-PCR for the diagnosis of COVID-19 (COVID-SPRING study).

BACKGROUND: Nasopharyngeal swab has long been considered the specimen of choice for the diagnosis of respiratory viral infections, including SARS-CoV-2 infection, but it suffers from several drawbacks: its discomfort limits screening acceptability, and it is vulnerable to shortages in both specialized materials and trained healthcare workers in the context of a pandemic. METHODS: We prospectively compared natural spring water gargle to combined oro-nasopharyngeal swab (ONPS) for the diagnosis of coronavirus disease 2019 (COVID-19) in paired clinical specimens (1005 ONPS and 1005 gargles) collected from 987 unique early symptomatic as well as asymptomatic individuals from the community. RESULTS: Using a direct RT-PCR method with the Allplex™ 2019-nCoV Assay (Seegene), the clinical sensitivity of the gargle was 95.3% (95% confidence interval [CI], 90.2 - 98.3%), similar to the sensitivity of the ONPS (93.8%; 95% CI, 88.2 - 97.3%), despite significantly lower viral RNA concentration in gargles, as reflected by higher cycle threshold values. No single specimen type detected all COVID-19 cases. SARS-CoV-2 RNA was stable in gargles at room temperature for at least 7 days. CONCLUSION: The simplicity of this sampling method coupled with the accessibility of spring water are clear advantages in a pandemic situation where testing frequency, turnaround time and shortage of consumables and trained staff are critical elements.

The PRONTO study: Clinical performance of ID NOW in individuals with compatible SARS-CoV-2 symptoms in walk-in centres-accelerated turnaround time for contact tracing.

Background: This PRONTO study investigated the clinical performance of the Abbott ID NOWTM (IDN) COVID-19 diagnostic assay used at point of care and its impact on turnaround time for divulgation of test results. Methods: Prospective study conducted from December 2020 to February 2021 in acute symptomatic participants presenting in three walk-in centres in the province of Québec. Results: Valid paired samples were obtained from 2,372 participants. A positive result on either the IDN or the standard-of-care nucleic acid amplification test (SOC-NAAT) was obtained in 423 participants (prevalence of 17.8%). Overall sensitivity of IDN and SOC-NAAT were 96.4% (95% CI: 94.2-98.0%) and 99.1% (95% CI: 97.6-99.8), respectively; negative predictive values were 99.2% (95% CI: 98.7-99.6%) and 99.8% (95% CI: 99.5-100%), respectively. Turnaround time for positive results was significantly faster on IDN. Conclusion: In our experience, IDN use in symptomatic individuals in walk-in centres is a reliable sensitive alternative to SOC-NAAT without the need for subsequent confirmation of negative results. Such deployment can accelerate contact tracing, reduce the burden on laboratories and increase access to testing.

Drug resistance patterns of recombinant herpes simplex virus DNA polymerase mutants generated with a set of overlapping cosmids and plasmids.

Herpes simplex virus (HSV) DNA polymerase (Pol) mutations can confer resistance to all currently available antiherpetic drugs. However, discrimination between mutations responsible for drug resistance and those that are part of viral polymorphism can be difficult with current methodologies. A new system is reported for rapid generation of recombinant HSV type 1 (HSV-1) DNA Pol mutants based on transfection of a set of overlapping viral cosmids and plasmids. With this approach, twenty HSV-1 recombinants with single or dual mutations within the DNA pol gene were successfully generated and subsequently evaluated for their susceptibilities to acyclovir (ACV), foscarnet (FOS), cidofovir (CDV), and adefovir (ADV). Mutations within DNA Pol conserved regions II (A719T and S724N), VI (L778M, D780N, and L782I), and I (F891C) were shown to induce cross-resistance to ACV, FOS, and ADV, with two of these mutations (S724N and L778M) also conferring significant reduction in CDV susceptibility. Mutant F891C was associated with the highest levels of resistance towards ACV and FOS and was strongly impaired in its replication capacity. One mutation (D907V) lying outside of the conserved regions was also associated with this ACV-, FOS-, and ADV-resistant phenotype. Some mutations (K522E and Y577H) within the delta-region C were lethal, whereas others (P561S and V573M) induced no resistance to any of the drugs tested. Recombinants harboring mutations within conserved regions V (N961K) and VII (Y941H) were resistant to ACV but susceptible to FOS. Finally, mutations within conserved region III were associated with various susceptibility profiles. This new system allows a rapid and accurate evaluation of the functional role of various DNA Pol mutations, which should translate into improved management of drug-resistant HSV infections.

Herpes simplex virus isolates with reduced adefovir susceptibility selected in vivo by foscarnet therapy.

Sequential herpes simplex virus (HSV) isolates from AIDS patients receiving foscarnet therapy were evaluated for susceptibility to adefovir. Foscarnet-resistant isolates with DNA polymerase mutations in regions II, VI, and between I and VII were also associated with an important decrease in susceptibility to adefovir (mean IC50 increase: 4.6-fold compared to pre-foscarnet or wild-type isolates) suggesting that adefovir-resistant HSV could be selected in vivo by foscarnet therapy.

Resistance of herpesviruses to antiviral drugs: clinical impacts and molecular mechanisms.

Nucleoside analogues such as acyclovir and ganciclovir have been the mainstay of therapy for alphaherpesviruses (herpes simplex virus (HSV) and varicella-zoster virus (VZV)) and cytomegalovirus (CMV) infections, respectively. Drug-resistant herpesviruses are found relatively frequently in the clinic, almost exclusively among severely immunocompromised patients receiving prolonged antiviral therapy. For instance, close to 10% of patients with AIDS receiving intravenous ganciclovir for 3 months excrete a drug-resistant CMV isolate in their blood or urine and this percentage increases with cumulative drug exposure. Many studies have reported that at least some of the drug-resistant herpesviruses retain their pathogenicity and can be associated with progressive or relapsing disease. Viral mutations conferring resistance to nucleoside analogues have been found in either the drug activating/phosphorylating genes (HSV or VZV thymidine kinase, CMV UL97 kinase) and/or in conserved regions of the viral DNA polymerase. Currently available second line agents for the treatment of herpesvirus infections--the pyrophosphate analogue foscarnet and the acyclic nucleoside phosphonate derivative cidofovir--also inhibit the viral DNA polymerase but are not dependent on prior viral-specific activation. Hence, viral DNA polymerase mutations may lead to a variety of drug resistance patterns which are not totally predictable at the moment due to insufficient information on specific drug binding sites on the polymerase. Although some CMV and HSV DNA polymerase mutants have been found to replicate less efficiently in cell cultures, further research is needed to correlate viral fitness and clinical outcome.