Viral ImmunologyVol. 36, No. 1 EditorialFree AccessImmunity Debt, a Gap in Learning, or Immune Dysfunction?Robert F. Needle and Rodney S. RussellRobert F. Needlehttps://orcid.org/0000-0001-8844-4981Division of BioMedical Sciences, Faculty of Medicine, Memorial University, St. John's, Canada.Public Health and Microbiology Laboratory, Eastern Health, St. John's, Canada.Search for more papers by this author and Rodney S. RussellAddress correspondence to: Dr. Rodney Russell, Division of BioMedical Sciences, Faculty of Medicine, Memorial University, 300 Prince Philip Drive, St. John's A1C 5S7, Canada E-mail Address: [email protected]E-mail Address: [email protected]Division of BioMedical Sciences, Faculty of Medicine, Memorial University, St. John's, Canada.Search for more papers by this authorPublished Online:16 Jan 2023https://doi.org/10.1089/vim.2022.0204AboutSectionsPDF/EPUB Permissions & CitationsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail As we bring in the new year of 2023, and say farewell to the year when many of us got acquainted with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) for the first time, the “new normal” is looking much as predicted. Many people have been vaccinated against coronavirus disease 2019 (COVID-19), and many now have hybrid immunity. Anyone watching the pandemic closely would have expected the new normal to include most people living much as they did before the pandemic, but others still having to be extra careful. We expected things to settle down and for most of society to function as it always did, but once everyone accepted that this virus was here to stay, there came the realization that many individuals would not get back to normal as quickly as others would.Unfortunately, the immunologically vulnerable still have to be very careful, especially if they do not yet know how they will fare with COVID-19. Anyone with any kind of immunocompromised state may struggle with clearing a SARS-CoV-2 infection, and more importantly, might not realize that they may not have responded well to the COVID-19 vaccines they received. These immunologically vulnerable individuals were probably already being extra careful about influenza and other infections, but now they have another virus to worry about, one that is still very prevalent and highly contagious within our communities.On the topic of community prevalence, at the beginning of the pandemic, there was much debate over the potential effectiveness of masks and physical distancing with respect to containing SARS-CoV-2, but it became very clear very early that these public health measures indeed helped to reduce transmission rates. One of the more obvious impacts of such measures was evident when the usual respiratory viruses failed to arrive in their normal numbers during 2020 and 2021 when extensive public health guidelines and encouragement of masking were still in place in much of the world. It is not surprising now to see respiratory viruses, such as influenza virus, respiratory syncytial virus, adenoviruses, rhinoviruses, among others, returning with what feels like a vengeance.Collecting data on prevalence and severity also comes with new and interesting challenges. Substantial investment into respiratory testing during the pandemic has led to increased testing of all respiratory pathogens, including testing of different patient populations and different viruses. So while epidemiology confirms that influenza virus and respiratory syncytial virus were prevalent earlier than usual this year, the percent positive remains within historical ranges. This begs the question, would the current higher percentage positive numbers still differ had a similar more targeted patient population been tested prepandemic? Or, are the laboratory data revealing that respiratory viruses are just more common this year, and it is this larger number of cases that is causing the influx seen in many hospitals? In the end, this is another topic for research to increase our understanding around the many impacts of the COVID-19 pandemic.What may not have been so predictable was the severity of these common respiratory infections. When “flu season” began in 2022, signals were emerging that indicated pediatric hospitals were experiencing more traffic than usual. As the data began to emerge, we saw the usual respiratory viruses arriving earlier and in higher numbers than usual. This increase in children needing medical attention and even hospitalization, unfortunately, has led to some dangerous theories. For example, the term “immunity debt” has emerged, and basically proposes that children's immune systems are now weaker because they have not been exposed to as many pathogens as they would have been in the absence of the public health response to the pandemic.However, the term immunity debt is misleading, and we see this situation as more of a slight “gap in education” that will be made up in short time. Of course children's immune systems are not weaker, but if they have not been exposed to certain viruses on the usual schedule, then they are missing the memory response they would have made had they been infected with some of these viruses for the first time over the past couple of years.On one hand, what is being perceived and immunity debt may just be a delay in education of the immune system that will disappear over the next year as children's immune systems make up for lost time by finally seeing the viruses that have been less prevalent than usual over the past 2 years. However, we cannot dismiss this situation so easily because there are some things we still cannot explain. We know that many viruses actively cause immune dysfunction, although this is more commonly observed in the context of chronic viral infections, and often includes documented T cell exhaustion as well as epigenetic and immunological scarring.Long-term immune dysfunction is not typically caused by acute infections that resolve relatively quickly. With most SARS-CoV-2-infected children experiencing mild COVID-19 symptoms, it is unlikely that their subsequent exaggerated responses to other viruses are linked to SARS-CoV-2-induced immune dysfunction, but some scientists are beginning to ask whether this might be the case.Another theory that might explain the increased severity of current respiratory viral infections could be the simple timing of these infections. In many cases now, we are seeing 5- and 6-year-old children getting introduced to certain viruses for the first time, when in the past, they would have encountered these viruses at the age of 2 or 3 years. So are they experiencing heightened inflammatory responses now at later ages because their immune systems are more developed and, therefore, respond in a more vigorous manner that can include a cytokine storm and extensive inflammation?The other very important immune aspect that needs more attention in this discussion is that of maternal antibodies. At least for very young children, they may have also missed the important benefit normally provided by maternal antibodies acquired during breastfeeding. Then, with females of child-bearing age also experiencing comparably less respiratory infections in the past few years, and many pregnant women potentially trying harder than usual not to get infected with viruses, there had to be a gap in maternal antibody coverage compared with nonpandemic years.And then there is long COVID-19, which is still very much a mystery with respect to how we define it, why some get it and others do not, and what impact it has on the immune system. In individuals with long COVID-19, it would not be surprising to see a detrimental effect on how well the immune system can subsequently deal with other viral infections, but it will take time to determine whether this is the case.Many people would now argue that the pandemic is over, and perhaps the urgency of it is, but we are still very much living within the impacts and the fallout of the pandemic. The topic of viral immunology continues to be in the forefront of our minds because, for example, the resurgence of respiratory viruses is affecting the daily lives of anyone with, or interacting with, children. It will be interesting going forward to know the answers to these questions around immunity gaps and immune dysfunction. Despite all the torment and tragedy that came with the pandemic, it has been, and will continue to be, an intense period of learning for the field of viral immunology.FiguresReferencesRelatedDetailsCited byNo signs of “tripledemic” in Finnish children6 March 2023 | Journal of Medical Virology, Vol. 95, No. 3 Volume 36Issue 1Jan 2023 InformationCopyright 2023, Mary Ann Liebert, Inc., publishersTo cite this article:Robert F. Needle and Rodney S. Russell.Immunity Debt, a Gap in Learning, or Immune Dysfunction?.Viral Immunology.Jan 2023.1-2.http://doi.org/10.1089/vim.2022.0204Published in Volume: 36 Issue 1: January 16, 2023PDF download

Cyclospora cayetanensis is an emerging foodborne parasite that causes cyclosporiasis, an enteric disease of humans. Domestically acquired outbreaks have been reported in Canada every spring or summer since 2013. To date, investigations into the potential sources of infection have relied solely on epidemiological data. To supplement the epidemiological data with genetic information, we genotyped 169 Canadian cyclosporiasis cases from stool specimens collected from 2010 to 2021 using an existing eight-marker targeted amplicon deep (TADS) scheme specific to C. cayetanensis as previously described by the US Centers for Disease Control and Prevention (CDC). This is the first study to genotype Canadian Cyclospora cayetanensis isolates, and it focuses on evaluating the genotyping performance and genetic clustering. Genotyping information was successfully collected with at least part of one of the markers in the TADS assay for 97.9% of specimens, and 81.1% of cyclosporiasis cases met the minimum requirements to genetically cluster into 20 groups. The performance of the scheme suggests that examining cyclosporiasis cases genetically will be a valuable tool for supplementing epidemiological outbreak investigations and to minimize further infections. Further research is required to expand the number of discriminatory markers to improve genetic clustering.

Tuberculosis is a significant cause of morbidity worldwide and is a priority at the provincial and federal levels in Canada. It is known that tuberculosis transmission networks are complex and span many years as well as different jurisdictions and countries. MIRU-VNTR is a universal tuberculosis genotyping method that utilizes a 24-loci pattern and it has shown promise in identifying inter and intrajurisdictional clusters within Canada. MIRU-VNTR data collected over 10 years from the National Reference Centre for Mycobacteriology (NRCM) were analyzed in this study. Some clusters were unique to a single province/territory, while others spanned multiple provinces and/or territories in Canada. The use of a universal laboratory test can enhance contact tracing, provide geographical information on circulating genotypes, and hence, aid in tuberculosis investigation by public health. The housing of all data on one platform, technical ease of the method, easy exchange of data between jurisdictions, and strong collaboration with laboratories and surveillance units at the provincial and federal levels have the potential to identify possible outbreaks in real time.

BACKGROUND:Serological assays designed to detect SARS-CoV-2 antibodies are being used in serological surveys and other specialized applications. As a result, and to ensure that the outcomes of serological testing meet high quality standards, evaluations are required to assess the performance of these assays and the proficiency of laboratories performing them.METHODS:A panel of 60 plasma/serum samples from blood donors who had reverse transcriptase-polymerase chain reaction (RT-PCR) confirmed SARS-CoV-2 infections and 21 SARS-CoV-2 negative samples were secured and distributed to interested laboratories within Canada (n = 30) and the United States (n = 1). Participating laboratories were asked to provide details on the diagnostic assays used, the platforms the assays were performed on, and the results obtained for each panel sample. Laboratories were blinded with respect to the expected outcomes.RESULTS:The performance of the different assays evaluated was excellent, with the high-throughput platforms of Roche, Ortho, and Siemens demonstrating 100% sensitivity. Most other high-throughput platforms had sensitivities of >93%, with the exception of the IgG assay using the Abbott ARCHITECT which had an average sensitivity of only 87%. The majority of the high-throughput platforms also demonstrated very good specificities (>97%).CONCLUSION:This proficiency study demonstrates that most of the SARS-CoV-2 serological assays utilized by provincial public health or hospital laboratories in Canada have acceptable sensitivity and excellent specificity.

The ability to detect antibodies to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently under investigation with various performance characteristics and indications for use. In this article, we analyzed the ability of the Abbott SARS-CoV-2 immunoglobulin class G (IgG), EuroImmun SARS-CoV-2 enzyme-linked immunosorbent assay (ELISA) IgG, and EuroImmun SARS-CoV-2 ELISA immunoglobulin class A (IgA) kits to detect evidence of previous infection with SARS-CoV-2. We tested 49 known coronavirus disease-19 (COVID-19) patients and 111 prepandemic stored serology specimens. This resulted in a sensitivity of 95.9%, 100.0%, and 91.3% and a specificity of 98.2%, 98.2%, and 90.8% respectively, using manufacturer recommended cutoffs after inconclusive results (one for EuroImmun IgG and five for EuroImmun IgA) being excluded in the final statistical analyses. Cross-reactivity of hepatitis C virus seropositive specimens was observed resulting in false positives (p < 0.05). If a two-tiered algorithmic approach was applied, that is, testing with Abbott SARS-CoV-2 assay followed by EuroImmun SARS-CoV-2 IgG, 100% specificity and sensitivity could be obtained after six inconclusive results were excluded from data set before statistical analyses. Performance characteristics presented demonstrate the superior performance of IgG class antibodies for investigating previous infections. In addition, utilizing a second antibody test for supplementary testing may significantly enhance performance, particularly in lower prevalence settings.

Objective: To assess the concordance of high-risk HPV (HR-HPV) testing with the Alinity assay on cervical samples collected with diverse collection/storage protocols (ThinPrep, SurePath, Cervicollect) and to assess inter-assay concordance of HR-HPV testing of cervical cell specimens with Alinity m HR HPV assay (Alinity) vs cobas (R) 4800 HPV assay (cobas). Methods: Specimens were obtained from 560 women attending a Women's Health clinic. Two specimens were obtained from each woman with combinations of two of the three collection devices and aliquots were tested by the two assays. Results: Alinity showed an agreement of 93.9%, Kappa = 0.89 (263/280) between ThinPrep and SurePath specimens; 97.5%, Kappa = 0.95 (347/356) and 92.9%, Kappa = 0.85 (104/112) between ThinPrep and SurePath aliquots taken before or after cytology processing, respectively. Cervi-Collect specimens showed an agreement of 94.6%, Kappa = 0.89 (265/280) with ThinPrep specimens. Compared to cobas, Alinity showed agreements of 94.3%, Kappa = 0.88 (395/419) and 91.8%, Kappa = 0.82 (257/280) between ThinPrep and SurePath specimens, respectively. Alinity and cobas detected genotypes 16/18 and other high-risk HPV types at similar rates and showed similar correlations with cytology grades. Conclusions: Compared to cobas, Alinity performed equally well for detecting HPV in cervical specimens obtained with ThinPrep and SurePath. The Cervi-Collect device compared well to the other collection methods. Alinity is a reliable assay for simultaneous detection of HPV-16/18 and other high-risk genotypes in cervical specimens.

The COVID-19 pandemic has led to the influx of immunoassays for the detection of antibodies towards severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into the global market. The Canadian Public Health Laboratory Network Serology Task Force undertook a nationwide evaluation of twelve laboratory and 6 point-of-care based commercial serological assays for the detection of SARS-CoV-2 antibodies. We determined that there was considerable variability in the performance of individual tests and that an orthogonal testing algorithm should be prioritized to maximize the accuracy and comparability of results across the country. The manual enzyme immunoassays and point-of-care tests evaluated had lower specificity and increased coefficients of variation compared to automated enzyme immunoassays platforms putting into question their utility for large-scale sero-surveillance. Overall, the data presented here provide a comprehensive approach for applying accurate serological assays for longitudinal sero-surveillance and vaccine trials while informing Canadian public health policy.

The discovery of curative antiviral drugs for a chronic disease such as HCV infection has encouraged drug discovery in the context of other viruses for which no curative drugs currently exist. Since we currently face a novel virus that has caused a pandemic, the need for new antiviral agents is more apparent than ever.

With emergence of pandemic COVID-19, rapid and accurate diagnostic testing is essential. This study compared laboratory-developed tests (LDTs) used for the detection of SARS-CoV-2 in Canadian hospital and public health laboratories, and some commercially available real-time RT-PCR assays. Overall, analytical sensitivities were equivalent between LDTs and most commercially available methods.

Background: Group B Streptococcus (GBS) testing during pregnancy and application of intrapartum prophylaxis to carriers prevents early-onset disease due to GBS in neonates. New testing methods may be more accurate and cost-effective than conventional cultures. Objective: To compare performance and cost-effectiveness of three chromogenic agars and two direct latex agglutinations after carrot-broth enrichment, using in-house polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) as reference standards. Methods: A total of 285 consecutive vaginal–rectal swabs were enriched with carrot broth and then tested using conventional Streptococcus selective agar (SSA), ChromAGAR Strep B (Colourex; Alere ULC, Ontario), ChromID Strepto B (bioMérieux Canada, Quebec), Brilliance GBS (Oxoid Company Inc, Ontario) and two latex agglutination kits: PathoDxtra Strep Grouping Reagent Kit (Thermofisher Scientific, Oxoid Company, Ontario) and MEDStrep (Alere ULC, Ontario). In-house PCR and LAMP reference methods were performed on frozen carrot broth. Results: Of the 285 samples received, 244 were analyzed by in-house PCR and 195 by LAMP. The conventional method (SSA) is less sensitive than PCR, but equally specific. Chromogenic agars were as sensitive as or more sensitive than SSA, and cost as much as or more than SSA. Direct latex antigens were less sensitive and specific than SSA, and cost less than SSA. In-house PCR was more sensitive than LAMP. Conclusion: Chromogenic media perform approximately as well as the conventional method (SSA) but are costlier. Direct latex agglutination methods do not achieve adequate performance. The ideal limit of detection for PCR is unknown, since the risk to the newborn of maternal carriage of a low inoculum of GBS is unknown.

BackgroundPreviously we studied the antibiotic susceptibility of invasive Haemophilus influenzae collected in Canada from 1990 to 2006 and characterized isolates by serotype, MLST and ftsI gene sequencing for significant PBP3 mutations.ObjectivesTo provide an update based on isolates collected from 2007 to 2014.MethodsA total of 882 case isolates were characterized by serotype using slide agglutination and PCR. MLST was carried out to determine ST. Isolates were tested for β-lactamase production, presence of significant PBP3 mutations and antibiotic susceptibility by disc diffusion against 14 antibiotics. MIC values of three antibiotics were determined for 316 isolates using microbroth dilution.ResultsNon-typeable H. influenzae accounted for 54.6% of the isolates and 45.4% were serotypeable, predominantly type a (23.1%), type b (8.3%) and type f (10.8%). The overall rate of ampicillin resistance due to β-lactamase production was 16.4% and increased from 13.5% in 2007-10 to 19% in 2011-14. Significant PBP3 mutations were identified in 129 isolates (14.6%) with 23 (2.6%) also producing β-lactamase. MLST identified related STs (ST-136, ST-14 and ST-367) associated exclusively with genetically β-lactamase-negative, ampicillin-resistant isolates and confirmed previously reported associations between significant PBP3 mutations and ST.ConclusionsA significant increase in β-lactamase-producing isolates was observed from 2007 to 2014; the rate of significant PBP3 mutations has increased since previously reported and 52.5% of non-typeable H. influenzae now show resistance markers. Resistance to trimethoprim/sulfamethoxazole was common and no resistance to fluoroquinolones or third-generation cephalosporins was found.

Serology remains the mainstay for diagnosis of Epstein-Barr virus (EBV) infection. This study compared two automated platforms (BioPlex 2200 and Architect i2000SR) to test three EBV serological markers: viral capsid antigen (VCA) immunoglobulins of class M (IgM), VCA immunoglobulins of class G (IgG) and EBV nuclear antigen-1 (EBNA-1) IgG. Using sera from 65 patients at various stages of EBV disease, BioPlex demonstrated near-perfect agreement for all EBV markers compared to a consensus reference. The agreement for Architect was near-perfect for VCA IgG and EBNA-1 IgG, and substantial for VCA IgM despite five equivocal results. Since the majority of testing in our hospital was from adults with EBNA-1 IgG positive results, post-implementation analysis of an EBNA-based algorithm showed advantages over parallel testing of the three serologic markers. This small verification demonstrated that both automated systems for EBV serology had good performance for all EBV markers, and an EBNA-based testing algorithm is ideal for an adult hospital.

Background: Previously we studied the antibiotic susceptibility of invasive Haemophilus influenzae collected in Canada from 1990 to 2006 and characterized isolates by serotype, MLST and ftsI gene sequencing for significant PBP3 mutations. Objectives: To provide an update based on isolates collected from 2007 to 2014. Methods: A total of 882 case isolates were characterized by serotype using slide agglutination and PCR. MLST was carried out to determine ST. Isolates were tested for β-lactamase production, presence of significant PBP3 mutations and antibiotic susceptibility by disc diffusion against 14 antibiotics. MIC values of three antibiotics were determined for 316 isolates using microbroth dilution. Results: Non-typeable H. influenzae accounted for 54.6% of the isolates and 45.4% were serotypeable, predominantly type a (23.1%), type b (8.3%) and type f (10.8%). The overall rate of ampicillin resistance due to β-lactamase production was 16.4% and increased from 13.5% in 2007–10 to 19% in 2011–14. Significant PBP3 mutations were identified in 129 isolates (14.6%) with 23 (2.6%) also producing β-lactamase. MLST identified related STs (ST-136, ST-14 and ST-367) associated exclusively with genetically β-lactamase-negative, ampicillin-resistant isolates and confirmed previously reported associations between significant PBP3 mutations and ST. Conclusions: A significant increase in β-lactamase-producing isolates was observed from 2007 to 2014; the rate of significant PBP3 mutations has increased since previously reported and 52.5% of non-typeable H. influenzae now show resistance markers. Resistance to trimethoprim/sulfamethoxazole was common and no resistance to fluoroquinolones or third-generation cephalosporins was found.

The recent emergence of a severe respiratory disease caused by enterovirus D68 prompted investigation into whether Canadian hospital and provincial laboratories can detect this virus using commercial and laboratory-developed assays. This study demonstrated analytical sensitivity differences between commercial and laboratory-developed assays for the detection of enterovirus D68.

BACKGROUND:Serogroup B Neisseria meningitidis (MenB) has always been a major cause of invasive meningococcal disease (IMD) in Canada. With the successful implementation of a meningitis C conjugate vaccine, the majority of IMD in Canada is now caused by MenB.OBJECTIVE:To investigate IMD case isolates in Atlantic Canada from 2009 to 2013. Data were analyzed to determine the potential coverage of the newly licensed MenB vaccine.METHODS:Serogroup, serotype and serosubtype antigens were determined from IMD case isolates. Clonal analysis was performed using multilocus sequence typing. The protein-based vaccine antigen genes were sequenced and the predicted peptides were investigated.RESULTS:The majority of the IMD isolates were MenB (82.5%, 33 of 40) and, in particular, sequence type (ST)-154 B:4:P1.4 was responsible for 47.5% (19 of 40) of all IMD case isolates in Atlantic Canada. Isolates of this clone expressed the PorA antigen P1.4 and possessed the nhba genes encoding for Neisseria heparin-binding antigen peptide 2, which together matched exactly with two of the four components of the new four-component meningococcal B vaccine. Nineteen MenB isolates had two antigenic matches, another five MenB and one meningitis Y isolate had one antigenic match. This provided 75.8% (25 of 33) potential coverage for MenB, or a 62.5% (25 of 40) overall potential coverage for IMD.CONCLUSION:From 2009 to 2013, IMD in Atlantic Canada was mainly caused by MenB and, in particular, the B:4:P1.4 ST-154 clone, which accounted for 47.5% of all IMD case isolates. The new four-component meningococcal B vaccine appeared to offer adequate coverage against MenB in Atlantic Canada.