Optimizing heartworm diagnosis in dogs using multiple test combinations

Abstract Background Various heartworm (HW) diagnostic testing modalities detect products of, or reactions to, different life cycle stages of Dirofilaria immitis. Microfilariae (Mf) can be directly visualized in blood, antigen (Ag) from immature and adult heartworms may be detected on commercial assays, and antibody (Ab) tests detect the host immune response to larval stages. Ag and Mf tests are commonly used in dogs, which frequently carry adult HW infections, but Ab tests have only been validated for use in cats. In some HW-infected dogs, Ag is blocked by immune complexing leading to false-negative results. Heat-treatment (HT) to disrupt these complexes can increase the sensitivity of HW Ag tests. The aim of this study was to compare different methods for diagnosing HW infection in dogs at high risk using individual and paired diagnostic tests, including an exploration of using Ab tests designed for cats to test canine samples. Methods One hundred stray adult (≥ 2-year-old) dogs in Florida shelters were tested using Mf, HW Ag, and HW Ab tests (feline HW Ab tests currently not commercially validated/approved for use in dogs); two versions of each test platform were used. Results Fourteen dogs tested positive using point-of-care (POC) Ag tests; an additional 2 dogs tested positive with microtiter well assay, and an additional 12 dogs tested positive using HT Ag testing. For individual tests, Ag test sensitivity/specificity compared to HT Ag was 50–57%/100%, and Ab tests were 46–64%/82–94%. Sensitivity estimates for individual tests were higher when comparing to non-HT Ag. Pairing POC Ag tests with Mf tests improved sensitivity without loss of specificity, while pairing POC Ag and Ab tests modestly increased sensitivity at the expense of specificity. Conclusions Screening dogs for HW infection using both POC Ag and Mf detection, which is recommended by the American Heartworm Society, improved diagnostic performance in this study compared to single Ag test use, but may have missed more than one in four infected dogs. The need to improve access to highly accurate, rapid, and inexpensive large-scale HW testing for dogs in animal shelters remains largely unmet by current testing availability. The development of practical and validated protocols that incorporate heat or chemical treatment to disrupt Ag-Ab complexes in POC testing or decreasing the cost and time required for such testing in reference laboratories might provide solutions to this unmet need. Similar studies performed in countries where the prevalence of parasites such as D. repens or A. vasorum is different to the USA could potentially yield very different positive predictive values for both HT and non-HT Ag tests. Graphic abstract

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Diagnostic testing for feline panleukopenia in a shelter setting: a prospective, observational study

Journal of Feline Medicine and Surgery ◽

10.1177/1098612x211005301 ◽

2021 ◽

pp. 1098612X2110053

Author(s):

Linda S Jacobson ◽

Kyrsten J Janke ◽

Jolene Giacinti ◽

J Scott Weese

Keyword(s):

Point Of Care ◽

Diagnostic Testing ◽

False Negative ◽

Clinical Signs ◽

Test Results ◽

Feline Panleukopenia Virus ◽

Negative Results ◽

Copy Numbers ◽

Rectal Swabs ◽

Vaccination History

Objectives The aim of this study was to optimize the diagnosis of feline panleukopenia virus (FPV) in a shelter setting by: (1) comparing the results of the canine parvovirus IDEXX SNAP Parvo (SNAP) point-of-care ELISA with a commercial FPV quantitative real-time PCR (qPCR) test; (2) assessing whether vomit and anal/rectal swabs could be used for early diagnosis; and (3) clarifying the interpretation of weak-positive SNAP test results. Methods The study included shelter cats and kittens with incomplete or unknown vaccination history that had clinical signs suspicious for feline panleukopenia and fecal SNAP and PCR tests performed within 24 h of onset. Feces, anal/rectal swabs and vomit were tested using SNAP and PCR, with fecal PCR utilized as the reference standard. Results One hundred and forty-five cats were included. Seventeen were diagnosed with FPV infection and 62 were negative; 66 could not be individually designated because they were co-housed. Sensitivity was as follows: fecal SNAP 55% (n = 102; 95% confidence interval [CI] 32–77); swab SNAP 30% (n = 55; 95% CI 7–65); swab PCR 77% (n = 55; 95% CI 46–95); and vomit PCR 100% (n = 17; 95% CI 16–100). Specificity was high (96–100%) for all sample and test types. For PCR-positive fecal samples, true-positive SNAP tests (including weak positives) had significantly higher DNA viral copy numbers than false-negative SNAP tests ( P = 0.0031). Conclusions and relevance The SNAP ELISA should be viewed as an initial diagnostic test to rule in feline panleukopenia. Positive fecal SNAP test results, including weak positives, are highly likely to be true positives in clinically affected animals. Negative results in clinically affected animals are unreliable and should be followed up with PCR testing.

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Diagnostic performance and clinical implications of rapid SARS-CoV-2 antigen testing in Mexico using real-world nationwide COVID-19 registry data

10.1101/2021.01.02.21249141 ◽

2021 ◽

Author(s):

Omar Yaxmehen Bello-Chavolla ◽

Neftali Eduardo Antonio-Villa ◽

Luisa Fernández-Chirino ◽

Enrique C. Guerra ◽

Carlos A. Fermín-Martínez ◽

...

Keyword(s):

Diagnostic Performance ◽

Large Scale ◽

False Negative ◽

High Specificity ◽

Adverse Outcomes ◽

Clinical Implications ◽

Rt Pcr ◽

Predictive Values ◽

Epidemic Dynamics ◽

Negative Results

ABSTRACTBACKGROUNDSARS-CoV-2 testing capacity is important to monitor epidemic dynamics. Given difficulties of large-scale RT-PCR implementation, rapid antigen tests (Rapid Ag-T) have been proposed as alternatives in settings such as Mexico.OBJECTIVESTo evaluate diagnostic performance of Rapid Ag-T for SARS-CoV-2 infection and its associated clinical implications compared to RT-PCR testing in Mexico.METHODSWe analyzed data from the COVID-19 registry of the Mexican General Directorate of Epidemiology up to December 31st, 2020 (n=3,374,165) and cases with both RT-PCR and Rapid Ag-T (n=18,446). We evaluated diagnostic performance using accuracy measures and assessed time-dependent changes in AUROC. We also explored test discordances as predictors of hospitalization, intubation, severe COVID-19 and mortality.RESULTSRapid Ag-T is primarily used in Mexico City. Rapid Ag-T have low sensitivity 37.6% (95%CI 36.6-38.7), high specificity 95.4% (95%CI 95.1-95.8) and acceptable positive 86.1% (95%CI 85.0-86.6) and negative predictive values 67.2% (95%CI 66.2-69.2). Rapid Ag-T has optimal diagnostic performance up to days 7-10 after symptom, and its performance is modified by testing location, comorbidity, and age. RT-PCR(-) / Rapid Ag-T(+) cases had higher risk of adverse COVID-19 outcomes and were older, RT-PCR(+)/ Rapid Ag-T(-) cases had slightly higher risk or adverse outcomes and ≥7 days from symptom onset. Cases detected with rapid Ag-T were younger, without comorbidities, and milder COVID-19 course.CONCLUSIONSRapid Ag-T could be used as an alternative to RT-PCR for large scale SARS-CoV-2 testing in Mexico. Interpretation of Rapid Ag-T results should be done with caution to minimize the risk associated with false negative results.

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Evaluation of SARS-CoV-2 antigen-based rapid diagnostic kits in Pakistan: formulation of COVID-19 national testing strategy

Virology Journal ◽

10.1186/s12985-021-01505-3 ◽

2021 ◽

Vol 18 (1) ◽

Author(s):

Umar Saeed ◽

Sara Rizwan Uppal ◽

Zahra Zahid Piracha ◽

Azhar Rasheed ◽

Zubair Aftab ◽

...

Keyword(s):

Gold Standard ◽

Injection Drug Users ◽

Drug Users ◽

Diagnostic Testing ◽

False Negative ◽

Cross Sectional Study ◽

Nasopharyngeal Swab ◽

Rt Pcr ◽

Cross Sectional ◽

Negative Results

AbstractRapid diagnosis of SARS-CoV-2 during pandemic enables timely treatment and prevention of COVID-19. Evaluating the accuracy and reliability of rapid diagnostic testing kits is crucial for surveillance and diagnosis of SARS-CoV-2 infections in general population, injection drug users, multi-transfused populations, healthcare workers, prisoners, barbers and other high risk populations. The aim of this study was to evaluate performance and effectiveness of nasopharyngeal swab (NSP) and saliva based rapid antigen detection testing kits in comparison with USFDA approved triple target gold standard real-time polymerase chain reaction. A cross-sectional study was conducted on 33,000 COVID-19 suspected patients. From RT-PCR positive patients, nasopharyngeal swab (NSP) and saliva samples were obtained for evaluation of rapid COVID-19 testing kits (RDT). 100/33,000 (0.3%) of specimens were RT-PCR positive for SARS-CoV-2. Among RT-PCR positive, 62% were males, 34% were females, and 4% were children. The NSP-RDT (Lepu Medical China) analysis revealed 53% reactivity among males, 58% reactivity among females, and 25% reactivity among children. However saliva based RDT (Lepu Medical China) analysis showed 21% reactivity among males and 23% among females, and no reactivity in children. False negative results were significantly more pronounced in saliva based RDT as compared to NSP-RDT. The sensitivity of these NSP-RDT and saliva based RDT were 52% and 21% respectively. The RDTs evaluated in this study showed limited sensitivities in comparison to gold standard RT-PCR, indicating that there is a dire need in Pakistan for development of suitable testing to improve accurate COVID-19 diagnosis in line with national demands.

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Average Weighted Accuracy: Pragmatic Analysis for a Rapid Diagnostics in Categorizing Acute Lung Infections (RADICAL) Study

Clinical Infectious Diseases ◽

10.1093/cid/ciz437 ◽

2019 ◽

Vol 70 (12) ◽

pp. 2736-2742 ◽

Cited By ~ 1

Author(s):

Ying Liu ◽

Ephraim L Tsalik ◽

Yunyun Jiang ◽

Emily R Ko ◽

Christopher W Woods ◽

...

Keyword(s):

Diagnostic Test ◽

Diagnostic Yield ◽

False Negative ◽

Rapid Diagnostics ◽

Likelihood Ratios ◽

Clinical Value ◽

Predictive Values ◽

Negative Results ◽

Treatment Standard ◽

Lung Infections

Abstract Patient management relies on diagnostic information to identify appropriate treatment. Standard evaluations of diagnostic tests consist of estimating sensitivity, specificity, positive/negative predictive values, likelihood ratios, and accuracy. Although useful, these metrics do not convey the tests’ clinical value, which is critical to informing decision-making. Full appreciation of the clinical impact of a diagnostic test requires analyses that integrate sensitivity and specificity, account for the disease prevalence within the population of test application, and account for the relative importance of specificity vs sensitivity by considering the clinical implications of false-positive and false-negative results. We developed average weighted accuracy (AWA), representing a pragmatic metric of diagnostic yield or global utility of a diagnostic test. AWA can be used to compare test alternatives, even across different studies. We apply the AWA methodology to evaluate a new diagnostic test developed in the Rapid Diagnostics in Categorizing Acute Lung Infections (RADICAL) study.

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The role of ultrasonography in the diagnosis of talocalcaneal coalitions

Acta Radiologica ◽

10.1177/0284185120944911 ◽

2020 ◽

pp. 028418512094491

Author(s):

Tiezheng Wang ◽

Hengtao Qi ◽

Kai Rong ◽

Shuqian Zhang ◽

Shougang Bao ◽

...

Keyword(s):

False Negative ◽

Fibrous Tissue ◽

Joint Space ◽

Clinical Suspicion ◽

Youden Index ◽

Imaging Method ◽

Predictive Values ◽

Negative Results ◽

Reduced Space ◽

Sensitivity Specificity

Background Patients with talocalcaneal coalitions (TCC) often undergo computed tomography (CT). However, ultrasonography diagnosis of TCC has been seldom done according to the literature. Purpose To investigate the accuracy of ultrasonography in diagnosing TCC compared to CT. Material and Methods Ninety-seven consecutive patients with a clinical suspicion of TCC were included. Ultrasonography was used to assess the classification and complication of TCC. The main sonographic criteria for a positive diagnosis in cases of osseous coalition were the joint space between the medial surface of talar head and the underlying sustentaculum tali of calcaneus disappearing and being replaced by a continuous hyperechoic bony structure. In cases of fibrous coalition, ultrasonography revealed a reduced space of the joint associated with an irregular, angular appearance of its outline and hypoechoic fibrous tissue inside. These data were compared with CT findings. κ statistic was applied to determine the level of agreement. The sensitivity, specificity, positive and negative predictive values, accuracy, and Youden index of ultrasonography as a diagnostic method were assessed. Results Ultrasonography findings were positive in 20 of 97 patients with a clinical suspicion of TCC. The diagnosis was confirmed by CT in 21 patients. There were one false-positive result and two false-negative results by ultrasonography. The κ value was 0.907. The sensitivity, specificity, positive and negative predictive values, accuracy, and Youden index of ultrasonography were 90.5%, 98.7%, 95.0%, 97.4%, 96.9%, and 0.892, respectively. Conclusion Ultrasonography could be a reliable, accurate, and non-radioactive diagnostic imaging method in diagnosis of patients with suspected TCC.

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False-Negative Results in Point-of-Care Qualitative Human Chorionic Gonadotropin (hCG) Devices Due to Excess hCGβ Core Fragment

Clinical Chemistry ◽

10.1373/clinchem.2008.121210 ◽

2009 ◽

Vol 55 (7) ◽

pp. 1389-1394 ◽

Cited By ~ 46

Author(s):

Ann M Gronowski ◽

Mark Cervinski ◽

Ulf-Håkan Stenman ◽

Alison Woodworth ◽

Lori Ashby ◽

...

Keyword(s):

Chorionic Gonadotropin ◽

Human Chorionic Gonadotropin ◽

Point Of Care ◽

False Negative ◽

The Core ◽

Core Fragment ◽

Negative Results ◽

False Negative Results ◽

Positive Results ◽

Urine Specimens

Abstract Background: During pregnancy, human chorionic gonadotropin (hCG) immunoreactivity in urine consists of intact hCG as well as a number of hCG variants including the core fragment of hCGβ (hCGβcf). We identified 3 urine specimens with apparent false-negative results using the OSOM® hCG Combo Test (Genzyme Diagnostics) qualitative hCG device and sought to determine whether an excess of 1 of the fragments or variants might be the cause of the interference. Methods: We measured concentrations of hCG variants in the urine from 3 patients with apparent false-negative hCG results. Purified hCG variants were added to urines positive for hCG and tested using the OSOM, ICON® 25 hCG (Beckman Coulter), and hCG Combo SP® Brand (Cardinal Health) devices. Results: Dilution of these 3 urine samples resulted in positive results on the OSOM device. Quantification of hCG variants in each of the 3 patient urine specimens demonstrated that hCGβcf occurred in molar excess of intact hCG. Addition of purified hCGβcf to hCG-positive urines caused false-negative hCG results using the OSOM and ICON qualitative urine hCG devices. Conclusions: Increased concentrations of hCGβcf can cause false-negative results on the OSOM and ICON qualitative urine hCG devices. .

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Exploring 'best practice' for nucleic acid detection of Neisseria gonorrhoeae

Sexual Health ◽

10.1071/sh07050 ◽

2008 ◽

Vol 5 (1) ◽

pp. 17 ◽

Cited By ~ 40

Author(s):

David M. Whiley ◽

Suzanne M. Garland ◽

Geoffrey Harnett ◽

Gary Lum ◽

David W. Smith ◽

...

Keyword(s):

Nucleic Acid ◽

Neisseria Gonorrhoeae ◽

Best Practice ◽

Current Knowledge ◽

False Negative ◽

High Sensitivity ◽

Nucleic Acid Detection ◽

Predictive Values ◽

Neisseria Species ◽

Negative Results

Nucleic acid detection tests (NADT) have considerable benefits for the detection of Neisseria gonorrhoeae (GC), including high sensitivity across a range of specimen types and use under widely differing settings and conditions. However, sexual health practitioners and others who use data generated by NADT for GC should be aware of some important limitations of these tests. False-positive results caused by cross reaction with commensal Neisseria species have been observed in many assays, and have lead to unacceptably low positive-predictive values in some patient populations. Further, false-negative results can be caused by GC sequence variation, with some gonococci lacking certain NADT target sequences. This review examines the issues associated with gonococcal NADT and considers best practice for use of these assays based on current knowledge. We emphasise the need for supplementary testing and extensive assay validation, and suggest appropriate strategies for these requirements irrespective of the setting in which they are used. Further, we highlight the need to maintain culture-based testing for certain specimen sites as well as for antimicrobial resistance surveillance.

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Performance of SARS-CoV-2 Serology tests: Are they good enough?

10.1101/2020.11.13.20229625 ◽

2020 ◽

Author(s):

Isabelle Piec ◽

Emma English ◽

M Annette Thomas ◽

Samir Dervisevic ◽

William D Fraser ◽

...

Keyword(s):

Sensitivity And Specificity ◽

Respiratory Infections ◽

Point Of Care ◽

False Negative ◽

Rapid Test ◽

Diagnostic Sensitivity ◽

Medical Community ◽

Antibody Detection ◽

Serological Tests ◽

Negative Results

AbstractBackgroundIn the emergency of the SARS-CoV-2 pandemic, great efforts were made to quickly provide serology testing to the medical community however, these methods have been introduced into clinical practice without the complete validation usually required by the regulatory organizations.MethodsSARS-CoV-2 patient samples (n=43) were analysed alongside pre-pandemic control specimen (n=50), confirmed respiratory infections (n=50), inflammatory polyarthritis (n=22) and positive for thyroid stimulating immunoglobulin (n=30). Imprecision, diagnostic sensitivity and specificity and concordance were evaluated on IgG serologic assays from EuroImmun, Epitope Diagnostics (EDI), Abbott Diagnostics and DiaSorin and a rapid IgG/IgM test from Healgen.ResultsEDI and EuroImmun imprecision was 0.02-14.0% CV. Abbott and DiaSorin imprecision (CV) ranged from 5.2% - 8.1% and 8.2% - 9.6% respectively. Diagnostic sensitivity of the assays were 100% (CI: 80-100%) for Abbott, EDI and EuroImmun and 95% (CI: 73-100%) for DiaSorin at ≥14 days post PCR. Only the Abbott assay had a diagnostic specificity of 100% (CI: 91-100%). EuroImmun cross-reacted in 3 non-SARS-CoV-2 respiratory infections and 2 controls. The DiaSorin displayed more false negative results and cross-reacted in six cases across all conditions tested. EDI had one cross-reactive sample. The Healgen rapid test showed excellent sensitivity and specificity. Overall, concordance of the assays ranged from 76.1% to 97.9%.ConclusionsSerological tests for SARS-CoV-2 showed good analytical performance. The head-to-head analysis of samples revealed differences in results that may be linked to the use of nucleocapsid or spike proteins. The point of care device tested demonstrated adequate performance for antibody detection.

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Rapid, sensitive and specific SARS coronavirus-2 detection: a multi-center comparison between standard qRT-PCR and CRISPR based DETECTR.

10.1101/2020.07.27.20147249 ◽

2020 ◽

Cited By ~ 2

Author(s):

Eelke Brandsma ◽

Han JMP Verhagen ◽

Thijs J.W. van de Laar ◽

Eric C.J. Claas ◽

Marion Cornelissen ◽

...

Keyword(s):

Point Of Care ◽

Diagnostic Testing ◽

Added Value ◽

N Gene ◽

Analytical Sensitivity ◽

Qrt Pcr ◽

Negative Results ◽

Point Of Care Test ◽

Ribonucleoprotein Complexes ◽

Large Patient

Recent advances in CRISPR-based diagnostics suggest that DETECTR, a combination of isothermal reverse transcriptase loop mediated amplification (RT-LAMP) and subsequent Cas12 bystander nuclease activation by amplicon targeting ribonucleoprotein complexes, could be a faster and cheaper alternative to qRT-PCR without sacrificing sensitivity/specificity. Here we compare qRT-PCR with DETECTR to diagnose COVID-19 on 378 patient samples and report a 95% reproducibility. Patient sample dilution assays suggest a higher analytical sensitivity of DETECTR compared to qRT-PCR, however, this was not confirmed in a large patient cohort. The data showed that both techniques are equally sensitive in detecting SARS-CoV-2 providing an added value of DETECTR to the currently used qRT-PCR platforms. For DETECTR, different gRNAs can be used simultaneously to obviate negative results due to mutations in N-gene. Lateral flow strips, suitable as a point of care test (POCT), showed a 100% correlation to the high-throughput DETECTR assay. Importantly, DETECTR was 100% specific for SARS-CoV-2 and did not detect other human coronaviruses. As there is no need for specialized equipment, DETECTR could be rapidly implemented as a complementary technically independent approach to qRT-PCR thereby increasing the testing capacity of medical microbiological laboratories and relieving the existent PCR-platforms for routine non- SARS-CoV-2 diagnostic testing.

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Multiplex Target-Redundant RT-LAMP for Robust Detection of SARS-CoV-2 Using Fluorescent Universal Displacement Probes

10.1101/2021.08.13.21261995 ◽

2021 ◽

Author(s):

Enos C Kline ◽

Nuttada Panpradist ◽

Ian T Hull ◽

Qin Wang ◽

Amy K Oreskovic ◽

...

Keyword(s):

Molecular Diagnostics ◽

Detection System ◽

Point Of Care ◽

False Negative ◽

Cost Effective ◽

Internal Amplification Control ◽

Robust Detection ◽

Negative Results ◽

False Negative Results ◽

Target Redundancy

AbstractThe increasing prevalence of variant lineages during the COVID-19 pandemic has the potential to disrupt molecular diagnostics due to mismatches between primers and variant templates. Point-of-care molecular diagnostics, which often lack the complete functionality of their high throughput laboratory counterparts, are particularly susceptible to this type of disruption, which can result in false negative results. To address this challenge, we have developed a robust Loop Mediated Isothermal Amplification assay with single tube multiplexed multi-target redundancy and an internal amplification control. A convenient and cost-effective target specific fluorescence detection system allows amplifications to be grouped by signal using adaptable probes for pooled reporting of SARS-COV-2 target amplifications or differentiation of the Internal Amplification Control. Over the course of the pandemic, primer coverage of viral lineages by the three redundant sub-assays has varied from assay to assay as they have diverged from the Wuhan-Hu-1 isolate sequence, but aggregate coverage has remained high for all variant sequences analyzed, with a minimum of 97.4% (Variant of Interest: Eta). In three instances (Delta, Gamma, Eta), a high frequency mismatch with one of the three sub-assays was observed, but overall coverage remained high due to multi-target redundancy. When challenged with extracted human samples the multiplexed assay showed 100% sensitivity for samples containing greater than 30 copies of viral RNA per reaction, and 100% specificity. These results are further evidence that conventional laboratory methodologies can be leveraged at the point-of-care for robust performance and diagnostic stability over time.

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