Childhood screening for type 1 diabetes comparing automated multiplex Antibody Detection by Agglutination-PCR (ADAP) with single plex islet autoantibody radiobinding assays.

BACKGROUND: Two or more autoantibodies against either insulin (IAA), glutamic acid decarboxylase (GADA), islet antigen-2 (IA-2A) or zinc transporter 8 (ZnT8A) denote stage 1 (normoglycemia) or stage 2 (dysglycemia) type 1 diabetes prior to stage 3 type 1 diabetes. Automated multiplex Antibody Detection by Agglutination-PCR (ADAP) assays in two laboratories were compared to single plex radiobinding assays (RBA) to define threshold levels for diagnostic specificity and sensitivity. METHODS: IAA, GADA, IA-2A and ZnT8A were analysed in 1504 (54% females) population based controls (PBC), 456 (55% females) doctor's office controls (DOC) and 535 (41% females) blood donor controls (BDC) as well as in 2300 (48% females) patients newly diagnosed (1-10 years of age) with stage 3 type 1 diabetes. The thresholds for autoantibody positivity were computed in 100 10-fold cross-validations to separate patients from controls either by maximizing the χ2-statistics (chisq) or using the 98th percentile of specificity (Spec98). Mean and 95% CI for threshold, sensitivity and specificity are presented. FINDINGS: The ADAP ROC curves of the four autoantibodies showed comparable AUC in the two ADAP laboratories and were higher than RBA. Detection of two or more autoantibodies using chisq showed 0.97 (0.95, 0.99) sensitivity and 0.94 (0.91, 0.97) specificity in ADAP compared to 0.90 (0.88, 0.95) sensitivity and 0.97 (0.94, 0.98) specificity in RBA. Using Spec98, ADAP showed 0.92 (0.89, 0.95) sensitivity and 0.99 (0.98, 1.00) specificity compared to 0.89 (0.77, 0.86) sensitivity and 1.00 (0.99, 1.00) specificity in the RBA. The diagnostic sensitivity and specificity were higher in PBC compared to DOC and BDC. INTERPRETATION: ADAP was comparable in two laboratories, both comparable to or better than RBA, to define threshold levels for two or more autoantibodies to stage type 1 diabetes. FUNDING: Supported by The Leona M. and Harry B. Helmsley Charitable Trust (grant number 2009-04078), the Swedish Foundation for Strategic Research (Dnr IRC15-0067) and the Swedish Research Council, Strategic Research Area (Dnr 2009-1039). AL was supported by the DiaUnion collaborative study, co-financed by EU Interreg ÖKS, Capital Region of Denmark, Region Skåne and the Novo Nordisk Foundation.

Bayesian evaluation of temporal changes in sensitivity and specificity of three serological tests for multiple circulating strains of rabbit haemorrhagic disease virus.

Competition and indirect ELISAs are currently being used to monitor rabbit haemorrhagic disease viruses (RHDV1 and RHDV2) in rabbits worldwide. Temporal changes in the sensitivity (Se) and specificity (Sp) of RHDV1 competition-ELISA (cELISA1), RHDV2 competition-ELISA (cELISA2), and RHDV1 Immunoglobulin G (IgG1) ELISA, were investigated using Bayesian Latent Class models (BCLM) in the Australian wild rabbit population where both viruses circulate simultaneously and a long-term serological dataset exists. When cELISA1 was compared to IgG1 ELISA, the Se of cELISA1 improved while the Sp of IgG1 ELISA declined over the 2011-21. This corresponded with a decline in the true RHDV1 prevalence in 2018-21, suggesting that a large proportion of RHDV1 exposed rabbits survived the introduction and dominance of RHDV2 up to approximately 2017/2018, after which they died and were not replaced. The Se and Sp estimates for 2014-15 for both cELISA1 and IgG1 ELISA, and the true prevalence when analysing all three tests together were similar to those obtained from the analysis of cELISA1/IgG1 ELISA. The same was also true for the Se and Sp of cELISA2 and IgG1 ELISA estimates from 2018 onwards. This suggests that RHDV1 was the dominant infection type in 2014-15, but RHDV2 was the dominant infection type in 2018-21. Further, the increase in Se of cELISA2 and the low Sp of IgG1 ELISA in the cELISA2/IgG1 ELISA analysis, compared to the Se of cELISA2 and Sp of IgG1 ELISA when analysing all three tests together suggests that the underlying infection status was more influenced by RHDV2 and that the higher Se of IgG1 ELISA is due to cross-reaction of RHDV2 antibodies on IgG1 ELISA. The true prevalence data suggest that RHDV2 exposure peaked in 2017. Our findings show that test characteristics changed in response to the changing virus prevalences over time. IgG1 ELISA, currently having a high Se, should be used to monitor both viruses and will perform better than both cELISAs.

Accuracy of serological tests for bovine brucellosis: A systematic review and meta-analysis.

The direct methods for diagnosis of bovine brucellosis have several limitations, therefore serological tests are the basis for the diagnosis of the disease. However, a meta-analysis estimating the diagnostic sensitivity (DSe) and diagnostic specificity (DSp) on the main tests used in bovine brucellosis control programs worldwide has not been performed. This systematic review and meta-analysis aimed to estimate the DSe, DSp and thereby accuracy of serological tests individually used in the diagnosis of bovine brucellosis. The databases CABI, Cochrane Library, PubMed/MEDLINE, SciELO, Scopus and Web of Science were used to select articles. The search resulted in 5308 studies, of which 71 were selected for systematic review using quality assessment tools and 65 studies were included in the meta-analysis. For the meta-analysis, 178 assays and 11 different serological tests were considered. To estimate DSe and DSp of the tests, studies were divided according to animal selection for the studies: (1) studies that carried out a random or consecutive selection of participants (noncasecontrol studies) and (2) all studies, including casecontrol studies. Considering only the non-case-control studies to estimate the DSe, the tests that exhibited the best and worst performance were the iELISA test (indirect enzyme immunoassay - bacterial suspension as antigen - BS) (96.5%, 95% CI: 94.1-97.9%) and 2ME (2- mercaptoethanol test) (85.0%, 95% CI: 79.6-89.1%), respectively; while for DSp, the FPA (fluorescence polarization assay) (99, 7%, 95% CI: 99.5-99.8%) and PCFIA tests (protein concentration fluorescence immunoassay) (78.5%, 95% CI: 70.0-85.1%) showed better and worse performance, respectively. Overall, our results showed an overestimation in the DSe and DSp of the eleven serological tests assessed when casecontrol studies were included in the meta-analysis, which is a concern considering its impacts on the time and costs associated with populational diagnosis of the diseases, since several of these tests are routinely used in the control and eradication programs of bovine brucellosis worldwide. Furthermore, the tests that exhibited the best DSe and DSp, iELISA (BS) and FPA, respectively, are relatively easy to perform and interpret and the test which showed the best overall accuracy was FPA.

Modelling variation in test sensitivity for monitoring leptospirosis in beef cattle.

When Bayesian latent class analysis is used for diagnostic test data in the absence of a gold standard test, it is common to assume that any unknown test sensitivities and specificities are constant across different populations. Indeed this assumption is often necessary for model identifiability. However there are a number of practical situations, depending on the type of test and the nature of the disease, where this assumption may not be true. We present a case study of using a microscopic agglutination test to diagnose leptospiroris infection in beef cattle, which strongly suggests that sensitivity in particular varies among herds. We develop and fit an alternative model in which sensitivity is related to within-herd prevalence, and discuss the statistical and epidemiological implications.

Diagnostic Sensitivity of Blood Culture, Intraoperative Specimen, and Computed Tomography-Guided Biopsy in Patients with Spondylodiscitis and Isolated Spinal Epidural Empyema Requiring Surgical Treatment.

BACKGROUND: the successful treatment of spondylodiscitis (SD) and isolated spinal epidural empyema (ISEE) depends on early detection of causative pathogens, which is commonly performed either via blood cultures, intraoperative specimens, and/or image-guided biopsies. We evaluated the diagnostic sensitivity of these three procedures and assessed how it is influenced by antibiotics. METHODS: we retrospectively analyzed data from patients with SD and ISEE treated surgically at a neurosurgery university center in Germany between 2002 and 2021. RESULTS: we included 208 patients (68 [23-90] years, 34.6% females, 68% SD). Pathogens were identified in 192 cases (92.3%), including 187 (97.4%) pyogenic and five (2.6%) non-pyogenic infections, with Gram-positive bacteria accounting for 86.6% (162 cases) and Gram-negative for 13.4% (25 cases) of the pyogenic infections. The diagnostic sensitivity was highest for intraoperative specimens at 77.9% (162/208, p = 0.012) and lowest for blood cultures at 57.2% (119/208) and computed tomography (CT)-guided biopsies at 55.7% (39/70). Blood cultures displayed the highest sensitivity in SD patients (SD: 91/142, 64.1% vs. ISEE: 28/66, 42.4%, p = 0.004), while intraoperative specimens were the most sensitive procedure in ISEE (SD: 102/142, 71.8% vs. ISEE: 59/66, 89.4%, p = 0.007). The diagnostic sensitivity was lower in SD patients with ongoing empiric antibiotic therapy (EAT) than in patients treated postoperatively with targeted antibiotic therapy (TAT) (EAT: 77/89, 86.5% vs. TAT: 53/53, 100%, p = 0.004), whereas no effect was observed in patients with ISEE (EAT: 47/51, 92.2% vs. TAT: 15/15, 100%, p = 0.567). CONCLUSIONS: in our cohort, intraoperative specimens displayed the highest diagnostic sensitivity especially for ISEE, whereas blood cultures appear to be the most sensitive for SD. The sensitivity of these tests seems modifiable by preoperative EAT in patients with SD, but not in those with ISEE, underscoring the distinct differences between both pathologies.

Direct comparison of clinical diagnostic sensitivity of saliva from buccal swabs versus combined oro-/nasopharyngeal swabs in the detection of SARS-CoV-2 B.1.1.529 Omicron.

BACKGROUND/PURPOSE: While current guidelines recommend the use of respiratory tract specimens for the direct detection of SARS-CoV-2 infection, saliva has recently been suggested as preferred sample type for the sensitive detection of SARS-CoV-2 B.1.1.529 (Omicron). By comparing saliva collected using buccal swabs and oro-/nasopharyngeal swabs from patients hospitalized due to COVID-19, we aimed at identifying potential differences in virus detection sensitivity between these sample types. METHODS: We compare the clinical diagnostic sensitivity of paired buccal swabs and combined oro-/nasopharyngeal swabs from hospitalized, symptomatic COVID-19 patients collected at median six days after symptom onset by real-time polymerase chain reaction (PCR) and antigen test. RESULTS: Of the tested SARS-CoV-2 positive sample pairs, 55.8% were identified as SARS-CoV-2 Omicron BA.1 and 44.2% as Omicron BA.2. Real-time PCR from buccal swabs generated significantly higher quantification cycle (Cq) values compared to those from matched combined oro-/nasopharyngeal swabs and resulted in an increased number of false-negative PCR results. Reduced diagnostic sensitivity of buccal swabs by real-time PCR was observed already at day one after symptom onset. Similarly, antigen test detection rates were reduced in buccal swabs compared to combined oro-/nasopharyngeal swabs. CONCLUSION: Our results suggest reduced clinical diagnostic sensitivity of saliva collected using buccal swabs when compared to combined oro-/nasopharyngeal swabs in the detection of SARS-CoV-2 Omicron in symptomatic individuals.

Optimising urinary catecholamine metabolite diagnostics for neuroblastoma.

INTRODUCTION: The analysis of urinary catecholamine metabolites is a cornerstone of neuroblastoma diagnostics. Currently, there is no consensus regarding the sampling method, and variable combinations of catecholamine metabolites are being used. We investigated if spot urine samples can be reliably used for analysis of a panel of catecholamine metabolites for the diagnosis of neuroblastoma. METHODS: Twenty-four-hour urine or spot urine samples were collected from patients with and without neuroblastoma at diagnosis. Homovanillic acid (HVA), vanillylmandelic acid (VMA), dopamine, 3-methoxytyramine, norepinephrine, normetanephrine, epinephrine and metanephrine were measured by high-performance liquid chromatography coupled with fluorescence detection (HPLC-FD) and/or ultra-performance liquid chromatography coupled with electrospray tandem mass spectrometry (UPLC-MS/MS). RESULTS: Catecholamine metabolite levels were measured in urine samples of 400 neuroblastoma patients (24-hour urine, n = 234; spot urine, n = 166) and 571 controls (all spot urine). Excretion levels of catecholamine metabolites and the diagnostic sensitivity for each metabolite were similar in 24-hour urine and spot urine samples (p > .08 and >.27 for all metabolites). The area under the receiver-operating-characteristic curve (AUC) of the panel containing all eight catecholamine metabolites was significantly higher compared to that of only HVA and VMA (AUC = 0.952 vs. 0.920, p = .02). No differences were observed in metabolite levels between the two analysis methods. CONCLUSION: Catecholamine metabolites in spot urine and 24-hour urine resulted in similar diagnostic sensitivities. The Catecholamine Working Group recommends the implementation of spot urine as standard of care. The panel of eight catecholamine metabolites has superior diagnostic accuracy over VMA and HVA.

Profile likelihood confidence interval for the prevalence assessed by an imperfect diagnostic test.

We present a new confidence interval for the prevalence of a disease for a situation when sensitivity and specificity of the diagnostic test are estimated from validation samples independent of the study sample. The new interval is based on profile likelihood and incorporates an adjustment improving the coverage probability. Its coverage probability and expected length were assessed by simulation and compared to two other methods for this problem, namely those by Lang and Reiczigel (2014) and Flor et al. (2020). Expected length of the new interval is less than that of the Lang and Reiczigel interval while its coverage is about the same. Comparison to the Flor interval resulted in similar expected length but higher coverage probabilities for the new interval. All in all, the new interval proved to be better than both its competitors.

Diagnostic sensitivity and specificity of immunohistochemistry for the detection of rabies virus in domestic and wild animals in South Africa.

We estimated the diagnostic sensitivity (DSe) and specificity (DSp) of an immunohistochemistry (IHC) protocol compared to the direct fluorescent antibody test (DFAT), which is the gold standard test for rabies diagnosis. We obtained brain samples from 199 domestic and wild animal cases (100 DFAT-negative, 99 DFAT-positive), by convenience sampling from 2 government-accredited rabies virus (RABV) testing laboratories in South Africa, between February 2015 and August 2017. Tissues that had been stored at 4-8°C for several days to weeks at the 2 accredited laboratories were formalin-fixed and paraffin-embedded. Nighty-eight cases tested IHC-positive using a polyclonal anti-RABV nucleoprotein antibody and a polymer detection system. The overall DSe and DSp for the RABV IHC test were 98% (95% CI: 93-100%) and 99% (95% CI: 95-100%), respectively. Domestic dogs accounted for 41 of 98 RABV IHC-positive cases, with the remainder in 4 domestic cats, 25 livestock, and 28 wildlife. Herpestidae species, including 7 meerkats and 9 other mongoose species, were the most frequently infected wild carnivores, followed by 11 jackals. Three cases in domestic dogs had discordant test results; 2 cases were IHC-/DFAT+ and 1 case was IHC+/DFAT-. Considering the implications of a false-negative rabies diagnosis, participating in regular inter-laboratory comparisons is vital, and a secondary or confirmatory method, such as IHC, should be performed on all submitted specimens, particularly negative cases with human contact history.

Evaluation and comparison of the sensitivity of three commercial RT-qPCR kits used for the detection of SARS-CoV-2 in Santiago, Chile.

Introduction: The COVID-19 pandemic is still in force, causing global public health challenges and threats. Although vaccination and herd immunity have proven to be the most efficient way to control the pandemic, massive and early testing of patients using the RT-qPCR technique is crucial for constant genomic surveillance. The appearance of variants of SARS-CoV-2 with new mutations can reduce the efficiency of diagnostic detection. In this sense, several commercial RT-qPCR kits have been the target of extensive analysis because low assay performance could lead to false-negative diagnoses. Methods: In this study, we evaluated the performance of three commercial RT-qPCR kits; Thermo Fisher (TaqMan 2019-nCoV Assay Kit v1), BGI and Roche (LightCycler® Multiplex RNA Virus Master) used for the diagnosis of COVID-19 throughout the pandemic in Santiago de Chile. Results: Under our best assay conditions, we found significant differences in Cq amplification values for control and viral probes, against the same nasopharyngeal swab samples (NPSs). In addition, in some cases, the sensitivity of the RT-qPCR kits decreased against viral variants. Conclusion: Our study suggests evaluating the RT-qPCR kits used to detect SARS-CoV-2 because variants such as Omicron, which has several mutations, can compromise their detection and underestimate viral circulation.

Next-Generation Sequencing Gene Panels in Inheritable Cardiomyopathies and Channelopathies: Prevalence of Pathogenic Variants and Variants of Unknown Significance in Uncommon Genes.

The diffusion of next-generation sequencing (NGS)-based approaches allows for the identification of pathogenic mutations of cardiomyopathies and channelopathies in more than 200 different genes. Since genes considered uncommon for a clinical phenotype are also now included in molecular testing, the detection rate of disease-causing variants has increased. Here, we report the prevalence of genetic variants detected by using a NGS custom panel in a cohort of 133 patients with inherited cardiomyopathies (n = 77) or channelopathies (n = 56). We identified 82 variants, of which 50 (61%) were identified in genes without a strong or definitive evidence of disease association according to the NIH-funded Clinical Genome Resource (ClinGen; "uncommon genes"). Among these, 35 (70%) were variants of unknown significance (VUSs), 13 (26%) were pathogenic (P) or likely pathogenic (LP) mutations, and 2 (4%) benign (B) or likely benign (LB) variants according to American College of Medical Genetics (ACMG) classifications. These data reinforce the need for the screening of uncommon genes in order to increase the diagnostic sensitivity of the genetic testing of inherited cardiomyopathies and channelopathies by allowing for the identification of mutations in genes that are not usually explored due to a currently poor association with the clinical phenotype.

Evaluation of Exudative Pleural Effusions: A Multicenter, Prospective, Observational Study.

PURPOSE: The aim of this study is to determine the diagnostic performances of pleural procedures in undiagnosed exudative pleural effusions and to evaluate factors suggestive of benign or malignant pleural effusions in tertiary care centers. METHODS: This was a multicenter prospective observational study conducted between January 1 and December 31, 2018. A total of 777 patients with undiagnosed exudative pleural effusion after the initial work-up were evaluated. The results of diagnostic procedures and the patients' diagnoses were prospectively recorded. Sensitivity, specificity, and accuracy estimates with 95% confidence intervals were used to examine the performance of pleural procedures to detect malignancy. RESULTS: The mean age ± SD of the 777 patients was 62.0 ± 16.0 years, and 68.3% of them were male. The most common cause was malignancy (38.3%). Lung cancer was the leading cause of malignant pleural effusions (20.2%). The diagnostic sensitivity and accuracy of cytology were 59.5% and 84.3%, respectively. The diagnostic sensitivity of image-guided pleural biopsy was 86.4%. The addition of image-guided pleural biopsy to cytology increased diagnostic sensitivity to more than 90%. Thoracoscopic biopsy provided the highest diagnostic sensitivity (94.3%). The highest diagnostic sensitivity of cytology was determined in metastatic pleural effusion from breast cancer (86.7%). CONCLUSION: The diagnostic performance increases considerably when cytology is combined with image-guided pleural biopsy in malignant pleural effusions. However, to avoid unnecessary interventions and complications, the development of criteria to distinguish patients with benign pleural effusions is as important as the identification of patients with malignant pleural effusions.

Diagnostic Performance of Loop-Mediated Isothermal Amplification and Ultrasensitive Rapid Diagnostic Tests for Malaria Screening Among Pregnant Women in Kenya.

BACKGROUND: Screen-and-treat strategies with sensitive diagnostic tests may reduce malaria-associated adverse pregnancy outcomes. We conducted a diagnostic accuracy study to evaluate new point-of-care tests to screen pregnant women for malaria at their first antenatal visit in western Kenya. METHODS: Consecutively women were tested for Plasmodium infection by expert microscopy, conventional rapid diagnostic test (cRDT), ultra sensitive RDT (usRDT), and loop-mediated isothermal amplification (LAMP). Photoinduced electron-transfer polymerase chain reaction (PET-PCR) served as the reference standard. Diagnostic performance was calculated and modelled at low parasite densities. RESULTS: Between May and September 2018, 172 of 482 screened participants (35.7%) were PET-PCR positive. Relative to PET-PCR, expert microscopy was least sensitive (40.1%; 95% confidence interval [CI], 32.7%-47.9%), followed by cRDT (49.4%; 95% CI, 41.7%-57.1), usRDT (54.7%; 95% CI, 46.9%-62.2%), and LAMP (68.6%; 95% CI, 61.1%-75.5%). Test sensitivities were comparable in febrile women (n = 90). Among afebrile women (n = 392), the geometric-mean parasite density was 29 parasites/µL and LAMP (sensitivity = 61.9%) and usRDT (43.2%) detected 1.74 (95% CI, 1.31-2.30) and 1.21 (95% CI, 88-2.21) more infections than cRDT (35.6%). Per our model, tests performed similarly at densities >200 parasites/µL. At 50 parasites/µL, the sensitivities were 45%, 56%, 62%, and 74% with expert microscopy, cRDT, usRDT, and LAMP, respectively. CONCLUSIONS: This first-generation usRDT provided moderate improvement in detecting low-density infections in afebrile pregnant women compared to cRDTs.

Evaluation of the artus® Prep&Amp UM RT-PCR for detection of SARS-CoV-2 from nasopharyngeal swabs without prior nucleic acid eluate extraction.

Here we describe a retrospective clinical evaluation of the QIAGEN artus® SARS-CoV-2 Prep&Amp UM RT-PCR assay that detects SARS-CoV-2 RNA without the need for a nucleic acid eluate extraction procedure. Using Roche SARS-CoV-2 RT-PCR on the cobas® 8800 platform as a reference standard, a total of 225 confirmed SARS-CoV-2 positive and 320 negative nasopharyngeal swabs in viral transport media, were used to evaluate the artus® assay. Using the RT-PCR cycle threshold as a semi-quantitative marker of viral load, an assessment of over 370,000 SARS-CoV-2 RT-PCR positive results was used in the design of the reference positive specimen cohort. The viral load of all reference positive specimens used in the evaluation was a unique and accurate representation of the range and levels of SARS-CoV-2 positivity observed over a 13-month period of the COVID-19 pandemic. The artus® RT-PCR detects the presence of SARS-CoV-2 RNA, an internal control, and the human RNase P gene to ensure specimen quality. The diagnostic sensitivity of artus® was 92.89% with a specificity of 100%. To assess the analytical sensitivity, a limit of detection was performed using the 1st WHO NIBSC SARS-CoV-2 international standard, recording a 95% LOD of 1.1 × 103 IU/ml. The total invalid rate of specimens was 7.34% due to a lack of detectable RNase P (Ct >35). The artus® SARS-CoV-2 Prep&Amp UM RT-PCR assay is a new rapid RT-PCR assay, which may be considered to produce acceptable levels of diagnostic sensitivity and specificity whilst potentially halving the laboratory processing time.

Multi-event capture-recapture models estimate the diagnostic performance of serological tests for myxoma and rabbit haemorrhagic disease viruses in the absence of reference samples.

Estimation of the diagnostic performance of serological tests often relies on another test assumed as a reference or on samples of known infection status, yet both are seldom available for emerging pathogens in wildlife. Longitudinal disease serological data can be analysed through multi-event capture-mark-recapture (MECMR) models accounting for the uncertainty in state assignment, allowing us to estimate epidemiological parameters such as incidence and mortality. We hypothesized that by estimating the uncertainty in state assignment, MECMR models estimate the diagnostic performance of serological tests for rabbit haemorrhagic disease virus (RHDV) and myxoma virus (MYXV). We evaluated this hypothesis on longitudinal serological data of three tests of RHDV and one test of MYXV in two populations of the European rabbit (Oryctolagus cuniculus algirus). First, we selected the optimal cut-off threshold for each test using finite mixture models, a reference method not relying on reference tests or samples. Second, we used MECMR models to compare the diagnostic sensitivity (Se) and specificity (Sp) of the three tests for RHDV. Third, we compared the estimates of diagnostic performance by MECMR and finite mixture models across a range of cut-off values. The MECMR models showed that the RHDV test employing GI.2 antigens (Se: 100%) outperformed two tests employing GI.1 antigens (Se: 21.7% ± 8.6% and 8.7% ± 5.9%). At their selected cut-offs (2.0 for RHDV GI.2 and 2.4 for MYXV), the estimates of Se and Sp were concordant between the MECMR and finite mixture models. Over the duration of the study (May 2018 to September 2020), the monthly survival of European rabbits seropositive for MYXV was significantly higher than that of seronegative rabbits (82.7% ± 4.9% versus 61.5% ± 12.7%) at the non-fenced site. We conclude that MECMR models can reliably estimate the diagnostic performance of serological tests for RHDV and MYXV in European rabbits. This conclusion could extend to other diagnostic tests and host-pathogen systems. Longitudinal disease surveillance data analysed through MECMR models allow the validation of diagnostic tests for emerging pathogens in novel host species while simultaneously estimating epidemiological parameters.

Development of indirect ELISA and its evaluation in comparison with KOH hydrolysis and fungal culture for the immuno diagnosis of Trichophyton rubrum and Trichophyton mentagrophytes infection in humans.

Trichophyton is a keratinophilic fungus that can invade keratinized tissues of humans and cause superficial mycoses called dermatophytosis. The objective of this study was to develop and evaluate indirect ELISA in comparison with gold standard methods such as direct microscopic examination of KOH mounts and fungal culture for the diagnosis of Trichophyton infection in humans. The present investigation reports the production and partial purification of T. rubrum mycelial antigens and production of specific polyclonal antibodies. It also reports the development and optimization of indirect ELISA and evaluation of its potential in comparison with gold standard methods for the diagnosis of Trichophyton infection in humans. The diagnostic sensitivity and specificity of Trichophyton indirect ELISA was 93.75% and 93.33% respectively. The positive and negative predictive values were high as well, found to be 93.75% and 90.00% respectively indicating usefulness of the assay. In all comparisons, the correlation coefficient (r) value was >0.5 indicating strong correlation between KOH hydrolysis test, fungal culture method and indirect ELISA. A significant correlation coefficient of 0.856 (P < 0.0001) was obtained between indirect ELISA and fungal culture method. This shows a good agreement between fungal culture method and indirect ELISA. The present study clearly shows diagnostic performance of Trichophyton indirect ELISA developed in this study is efficient as fungal culture method for the diagnosis of Trichophyton infection in humans.

Comment on Skrebinska et al. Who Could Be Blamed in the Case of Discrepant Histology and Serology Results for Helicobacter pylori Detection? Diagnostics 2022, 12, 133.

In their article, Skebrinska and colleagues analysed the potential pitfalls of detecting Helicobacter pylori (H. pylori) by serology, histological (Giemsa) and immunohistochemical (IHC) staining. However, in the Introduction, the authors state: "…IHC is recommended only in individuals with active gastritis without H. pylori identification by histochemistry". Although this is a widely-held view, it does not seem to hold up in view of the results of the study by Kocsmár et al., which showed that the diagnostic sensitivity of Giemsa in the absence of activity is only 33.6%, but it is 92.6% in the presence of active gastritis, which is close to the 99.4% sensitivity of IHC. Considering that chronic active gastritis with the features of H. pylori gastritis is also common in other entities, if active inflammation is present in the sample, there is a very small chance that a Giemsa-negative case will be confirmed as H. pylori-positive by IHC. Based on this, the use of IHC is more reasonable in Giemsa-negative cases with no activity in which the etiological role of H. pylori is suggested by clinical, anamnestic or other data. However, it may also be reasonable to routinely use IHC as the primary staining method instead of Giemsa.

Bayesian latent class analysis to estimate the optimal cut-off for the MilA ELISA for the detection of Mycoplasma bovis antibodies in sera, accounting for repeated measures.

The MilA ELISA has been identified as a highly effective diagnostic tool for the detection of Mycoplasma bovis specific antibodies and has been validated for serological use in previous studies. This study aimed to estimate the optimal cut-off and corresponding estimates of diagnostic sensitivity (DSe) and diagnostic specificity (DSp) of the MilA ELISA for testing bovine serum. Serum samples from 298 feedlot cattle from 14 feedlots across four Australian states were tested on entry into the feedlot and approximately 42 days later. The paired serum samples were tested with the MilA ELISA, BIO K302 (Bio-X Diagnostics, Belgium) and BIO K260 (Bio-X Diagnostics, Belgium). A cut-off of 135 AU was estimated to be optimal using Bayesian latent class analysis with three tests in multiple populations, accounting for conditional dependence between tests. At this cut-off, the DSe and DSp of the MilA ELISA were estimated to be 92.1 % (95 % highest probability density [HPD] interval: 87.4, 95.8) and 95.5 % (95 % HPD: 92.4, 97.8), respectively. The DSes of the BIO K260 and BIO K302 ELISAs were estimated to be 60.5 % (95 % HPD: 54.0, 66.9) and 44.6 % (95 % HPD: 38.7, 50.7), respectively. DSps were 95.6 % (95 % HPD: 92.9, 97.7) and 97.8 % (95 % HPD: 95.9, 99.0), respectively. Mycoplasma bovis seroprevalence was remarkably high at follow-up after 42 days on the feedlots. Overall, this study estimated a cut-off, DSe and DSp for the MilA ELISA with less dependence on prior information than previous analyses and demonstrated that the MilA ELISA has higher DSe than the BIO K260 and BIO K302 assays.

Evaluation of canine detection of COVID-19 infected individuals under controlled settings.

Reverse transcription polymerase chain reaction (RT-PCR) is currently the standard diagnostic method to detect symptomatic and asymptomatic individuals infected with Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, RT-PCR results are not immediate and may falsely be negative before an infected individual sheds viral particles in the upper airways where swabs are collected. Infected individuals emit volatile organic compounds in their breath and sweat that are detectable by trained dogs. Here, we evaluate the diagnostic accuracy of dog detection against SARS-CoV-2 infection. Fifteen dogs previously trained at two centres in Australia were presented to axillary sweat specimens collected from known SARS-CoV-2 human cases (n = 100) and non-cases (n = 414). The true infection status of the cases and non-cases were confirmed based on RT-PCR results as well as clinical presentation. Across dogs, the overall diagnostic sensitivity (DSe) was 95.3% (95%CI: 93.1-97.6%) and diagnostic specificity (DSp) was 97.1% (95%CI: 90.7-100.0%). The DSp decreased significantly when non-case specimens were collected over 1 min rather than 20 min (p value = .004). The location of evaluation did not impact the detection performances. The accuracy of detection varied across dogs and experienced dogs revealed a marginally better DSp (p value = .016). The potential and limitations of this alternative detection tool are discussed.