Antigen Enzyme Immunoassay Research Articles

Diagnosis of Coccidioidomycosis with the Second-Generation Miravista IgG and IgM Enzyme Immunoassay and the Role of Adding Miravista Coccidioides Antigen Detection to Immunodiagnostic Assays.

In the present study, we validate and compare the second-generation Miravista Coccidioides IgG and IgM enzyme immunoassays (EIA) (MiraVista Diagnostics [MVD] Ab EIA) to Meridian Diagnostics Coccidioides IgG and IgM EIA (Meridian Ab EIA), immunodiffusion (ID) and complement fixation (CF). We also evaluated whether the addition of Coccidioides antigen testing to anti-Coccidioides antibody testing increased the sensitivity for the diagnosis of currently active coccidioidomycosis. We retrospectively studied 555 patients evaluated at Valleywise Health Medical Center between January 2013 and May 2017 for whom coccidioidomycosis was suspected and samples were submitted to MVD for testing. Specimens were tested for antigen in the MVD antigen enzyme immunoassay (MVD Ag EIA) and for IgG and IgM antibodies with MVD and Meridian Diagnostics EIAs. ID and CF were obtained from medical records. Sensitivity and specificity were 83.0% and 91.1% or MVD Ab EIA, 69.3% and 99.7% for Meridian Ab EIA, 85.4% and 100% for ID and 65.5% and 100% for CF. Combined MVD antigen and antibody detection by EIA and ID resulted in increased sensitivity in disseminated and pulmonary disease (MVD Ag/MVD Ab: 100%, 88.3%; MVD Ag/Meridian Ab: 98.2%, 78.6%; and MVD Ag/ID: 100%, 91.7%). The detection of antibodies by MVD EIA was more sensitive than Meridian EIA or CF but similar to ID. This study supports the use of antigen testing in immunocompromised patients and those with suspected disseminated disease. Furthermore, the addition of antigen detection by EIA to antibody detection resulted in higher sensitivity of all serological tests.

Detection of Histoplasma capsulatum and Blastomyces dermatitidis antigens in serum using a single quantitative enzyme immunoassay.

Histoplasma and Blastomyces antigen detection assays are commonly used diagnostic tools. However, a high level of cross-reactivity between these antigens prevents definitive pathogen identification by these assays alone. Retrospective analysis of 3,529 patients with Histoplasma and Blastomyces antigen testing performed on the same serum sample yielded an overall percent agreement of 99.3% (3,506 of 3,529; kappa: 0.859) between the two assays, suggesting that use of a single assay to detect both antigens may be an alternative diagnostic approach. We assessed performance of the Gotham BioTech Blastomyces antigen (GBA) enzyme immunoassay (EIA) (Portland, Maine) for detection of Blastomyces and Histoplasma antigens in serum. Comparison to the MiraVista Diagnostics Blastomyces (MVB) EIA showed 100% positive (24 of 24), negative (57 of 57), and overall (81 of 81) percent agreement. Additionally, 171 sera were used to compare the GBA EIA to the MiraVista Diagnostics Histoplasma (MVH) EIA, which showed 91.3% (63 of 69), 98% (100 of 102), and 95.3% (163 of 171) positive, negative, and overall percent agreement, respectively. Among eight patients with discordant GBA/MVH EIA results, seven had additional fungal testing performed, and results suggested that the MVH and GBA results were inaccurate for two and five samples, respectively. Overall, this study suggests that the GBA EIA has a high level of agreement with both of the commonly used, individual Blastomyces and Histoplasma antigen EIAs. By taking advantage of the high level of cross-reactivity between Blastomyces and Histoplasma antigen EIAs, utilization of a single antigen detection assay for these fungi provides an opportunity to optimize test utilization and decrease patient cost while maintaining a high level of diagnostic accuracy.

Evaluation of an enzyme immunoassay and immunodiffusion for detection of anti-Histoplasma antibodies in serum from cats and dogs.

Histoplasma antigen and anti-Histoplasma antibody detection are used to support the diagnosis of histoplasmosis. There is a paucity of published data on antibody assays. Our primary hypothesis was that anti-Histoplasma immunoglobulin G (IgG) antibody detection using enzyme immunoassay (EIA) will be more sensitive as compared to immunodiffusion (ID). Thirty-seven cats and 22 dogs with proven or probable histoplasmosis; 157 negative control animals. Residual stored sera were tested for anti-Histoplasma antibodies using EIA and ID. Results of urine antigen EIA were reviewed retrospectively. Diagnostic sensitivity was calculated for all three assays and compared between immunoglobulin G (IgG) EIA and ID. The diagnostic sensitivity of urine antigen EIA and IgG EIA, interpreted in parallel, was reported. Sensitivity of IgG EIA was 30/37 (81.1%; 95% confidence interval [CI], 68.5%-93.4%) in cats and 17/22 (77.3%; 95% CI, 59.8%-94.8%) in dogs. Diagnostic sensitivity of ID was 0/37 (0%; 95% CI, 0%-9.5%) in cats and 3/22 (13.6%; 95% CI, 0%-28.0%) in dogs. Immunoglobulin G EIA was positive in all animals (2 cats and 2 dogs) with histoplasmosis but without detectable antigen in urine. Diagnostic specificity of IgG EIA was 18/19 (94.7%; 95% CI, 74.0%-99.9%) in cats and 128/138 (92.8%; 95% CI, 87.1%-96.5%) in dogs. Antibody detection by EIA can be used to support the diagnosis of histoplasmosis in cats and dogs. Immunodiffusion has an unacceptably low diagnostic sensitivity and is not recommended.

Diversity of rotavirus strains circulating in Haiti before and after introduction of monovalent vaccine

BackgroundHaiti introduced a monovalent human group A rotavirus (RVA) vaccine (Rotarix) into its routine infant immunization program in April 2014. The goal of the surveillance program was to characterize RVA strains circulating in Haiti before and after RVA vaccine introduction. MethodsStool samples were collected from children <5 years old presenting with acute gastroenteritis at 16 hospitals in Haiti. RVA antigen enzyme immunoassay (EIA) testing was performed, and G and P genotypes were determined for positive specimens. In this study, genotype data for samples collected from May 2012 through April 2014 (the pre-vaccine introduction era) and May 2014 through July 2019 (post-vaccine introduction era) were analyzed. ResultsA total of 809 specimens were tested by the Centers for Disease Control and Prevention. During the pre-vaccine introduction era (May 2012 through April 2014), G12P[8] was the predominant genotype, detected in 88–94% of specimens. There was a high prevalence of the equine-like G3P[8] genotype among Haitian children with RVA after vaccine introduction. ConclusionsThe predominance of equine-like G3P[8] in three of five RVA seasons post-vaccine introduction suggests possible vaccine-specific selection pressure in Haiti. These temporal variations in RVA genotype predominance will require continued monitoring in Haiti as the vaccination program continues.

The utility of Coccidioides antigen and antibody detection in cerebrospinal fluid in the diagnosis of canine central nervous system coccidioidomycosis.

To evaluate the utility of enzyme immunoassays (EIAs) for the detection of Coccidioides antigen and antibody in CSF in the diagnosis of CNS coccidioidomycosis in dogs. 51 dogs evaluated for CNS disease in a single specialty center in Tucson in 2016. Excess CSF after routine analysis was banked after collection from dogs presented to the neurology service. Samples were tested by EIA for presence of Coccidioides antigen and antibody. Clinical data were collected from medical records retrospectively. 22 dogs were diagnosed with CNS coccidioidomycosis (CCM) or another neurologic disease (non-CCM). These groups of dogs overlapped in the presenting complaints, MRI results, and routine CSF analysis results. Four dogs, all with CCM, had positive antigen EIA results. With clinical diagnosis used as the reference standard, CSF antigen testing had low sensitivity (20%) but high specificity (100%) for diagnosis of CCM. Ten dogs with CCM and 4 dogs with other diagnoses had antibody detected in CSF by EIA. Sensitivity of CSF antibody testing was 46%, specificity was 86%, and positive and negative predictive values for the study population were 71% and 68%, respectively. Diagnosis of CNS coccidioidomycosis in dogs in an endemic region was hampered by overlap of clinical signs with other neurologic disorders and the low sensitivity of confirmatory diagnostics. The evaluated Coccidioides-specific EIAs performed on CSF can aid in the diagnosis. A prospective study is warranted to corroborate and refine these preliminary findings.

721. Diagnosis of Histoplasmosis Using the MVista Histoplasma Galactomannan Antigen Qualitative Lateral Flow-Based Immunoassay; A Multicenter Study

Abstract Background Accurate and timely methods for the diagnosis of histoplasmosis in endemic resource-limited settings are largely lacking. Histoplasma galactomannan antigen detection by enzyme immunoassay (EIA) is the most widely used method for the diagnosis of acute pulmonary and disseminated histoplasmosis in the United States (USA). EIA methods have constraints in resource-limited settings including cost, turnaround time, and the need for large reference laboratories, leading to missed or delayed diagnoses and poor outcomes. Lateral flow assays (LFA) are practical methods that can be used in this setting for Histoplasma antigen detection. Methods Frozen urine specimens were submitted to MiraVista (MVista) for Histoplasma antigen EIA testing from three academic medical centers in highly endemic areas of the USA. They were also blinded and tested for the MVista Histoplasma LFA by skilled MVista technologists. Medical records were reviewed for clinical information. Patients were classified as controls or cases of histoplasmosis. Cases were divided into proven or probable, pulmonary, or disseminated, immune competent or immune suppressed, and mild, moderate, or severe. Results 352 subjects were enrolled, including 66 cases of histoplasmosis (44 proven, 22 probable) and 286 controls. Most of the cases were immunocompromised (68%). 76% had disseminated histoplasmosis. 6% were mild, 66% moderate, and 28% severe. A high degree of concordance was found between LFA and EIA results (kappa 0.837, OR 372.7, LR 204, p&amp;lt; 0.001). Overall, the sensitivity and specificity of the LFA were 78.8% and 99.3% respectively (kappa 0.84, p&amp;lt; 0.001). The sensitivity was higher in proven cases (93.2%), in patient with disseminated (94.7%), moderate (80%) and severe disease (94%), and those with galactomannan levels ≥ 2 ng/mL (97.7%). Specificity was 99.3% in proven cases, 99.3% in patient with moderate and severe disease, and 96.4% in those with galactomannan levels ≥ 2 ng/mL. Table 1. Statistical characteristics of the LFA test for histoplasmosis in different categories. PPV: Positive Predictive Value. NPV: Negative Predictive Value. EIA: Enzyme Immunoassay. The LFA test for histoplasmosis is more accurate in patients with high burden of infection. Conclusion The MVista Histoplasma galactomannan LFA may meet the need for accurate rapid diagnosis of histoplasmosis in resource-limited settings, especially in patients with relatively high disease burden, potentially reducing morbidity and mortality. Disclosures Melissa Minderman, Bachelor's Degree, Molecular Biology, MiraVista Diagnostics (Employee) Suphansa Gunn, Bachelor's Degree, psychology, MiraVista Diagnostics (Employee) Lawrence J. Wheat, MD, MiraVista Diagnostics (Employee)

Disseminated Rasamsonia argillacea species complex infections in 8 dogs.

BackgroundClinical features, treatment, and outcome of opportunistic infections with Rasamsonia spp., a nonpigmented filamentous mold, are not well documented in dogs.ObjectivesDescribe clinical, radiographic, pathologic features, and outcome of dogs with disseminated Rasamsonia species complex infections.AnimalsEight client‐owned dogs.MethodsRetrospective case series. Medical records were reviewed to describe signalment, history, clinicopathologic and imaging findings, microbiologic and immunologic results, cyto‐ and histopathologic diagnoses, treatment, and outcome.ResultsPresenting complaints were nonspecific with anorexia (n = 5) and back pain (n = 4) most common. Five dogs were German Shepherd dogs. Six dogs had multifocal discospondylitis and 2 had pleural effusion. Six dogs had Rasamsonia piperina and 2 had Rasamsonia argillacea infections with isolates identified using DNA sequencing. Rasamsonia spp. were isolated by urine culture in 5 of 7 dogs. Five of 6 dogs had positive serum Aspergillus galactomannan antigen enzyme immunoassay (EIA) results. Median survival time was 82 days, and 317 days for dogs that survived to discharge. Four died during initial hospitalization (median survival, 6 days). All isolates had low minimum effective concentrations (MECs) to echinocandins with variable minimum inhibitory concentrations (MICs) for azole antifungal drugs.Conclusions and Clinical ImportanceRasamsonia spp. infections in dogs are associated with multisystemic disease involving the vertebral column, central nervous system, kidneys, spleen, lymph nodes, lungs, and heart. The infection shares clinical features with other systemic mold infections and can be misidentified when using phenotypical microbiologic methods. Molecular techniques are required to identify the organism and guide appropriate antifungal treatment.

Evaluation of a novel monoclonal antibody-based enzyme immunoassay for detection of Histoplasma antigen in urine of dogs.

BackgroundCommercially available, noninvasive testing options for histoplasmosis are limited outside of the United States.ObjectivesTo describe the diagnostic performance of a novel Histoplasma antigen enzyme immunoassay (IM EIA) for the diagnosis of histoplasmosis in dogs.AnimalsTwenty dogs with histoplasmosis, 79 dogs without histoplasmosis, and 11 unclassified dogs providing 202 urine samples.MethodsThis a prospective study using stored urine samples. Samples were analyzed with the IM EIA and with the commercially available Histoplasma antigen EIA (MV EIA). Dogs were classified based on final proven diagnosis and performance of the IM EIA was described and compared with the MiraVista enzyme immunoassay (MV EIA).ResultsThe diagnostic sensitivity (DSe), specificity (DSp), and accuracy (DAc) of the IM EIA were 70% (51%‐89%), 99% (97%‐100%), and 93% (81%‐100%), respectively. The DSe, DSp, and DAc for the MV EIA were 95% (85%‐100%), 99% (97%‐100%), and 98% (95%‐100%), respectively. The area under the receiver operator characteristic curve was significantly smaller for IM EIA (0.87) as compared with MV EIA (0.97, P = .03). This was primarily due to 6 false negative IM EIA results, 4 from dogs with disease localized to the gastrointestinal tract. The MV EIA was positive in 5/6 of these dogs.Conclusions and Clinical ImportanceThe IM EIA might be useful for the diagnosis of disseminated histoplasmosis in dogs, but clinical usefulness will be limited in dogs with histoplasmosis localized to the GI tract.

Screening for invasive fungal disease using non-culture-based assays among inpatients with advanced HIV disease at a large academic hospital in South Africa.

Despite widespread access to antiretroviral therapy (ART), the burden of advanced HIV disease in South Africa is high. This translates into an increased risk of AIDS-related opportunistic infections, including invasive mycoses. Using a limited number of non-culture-based diagnostic assays, we aimed to determine the prevalence of invasive mycoses and tuberculosis among hospitalised adults with very advanced HIV (CD4 counts<100cells/µL) at a large academic hospital. We conducted interviews and prospective medical chart reviews. We performed point-of-care finger stick and serum cryptococcal antigen lateral flow assays; serum (1 → 3) ß-D-glucan assays; urine Histoplasma galactomannan antigen enzyme immunoassays and TB lipoarabinomannan assays. We enrolled 189 participants from 5280 screened inpatients. Fifty-eight per cent were female, with median age 37years (IQR: 30-43) and median CD4 count 32cells/µL (IQR: 13-63). At enrolment, 60% (109/181) were receiving ART. Twenty-one participants (11%) had a diagnosis of an invasive mycosis, of whom 53% (11/21) had cryptococcal disease. Thirteen participants (7%) had tuberculosis and a concurrent invasive mycosis. ART-experienced participants were 60% less likely to have an invasive mycosis than those ART-naïve (adjusted OR: 0.4; 95% CI 0.15-1.0; P=.03). Overall in-hospital mortality was 13% (invasive mycosis: 10% [95% CI 1.2-30.7] versus other diagnoses: 13% (95% CI 8.4-19.3)). One in ten participants had evidence of an invasive mycosis. Diagnosis of proven invasive fungal disease and differentiation from other opportunistic infections was challenging. More fungal-specific screening and diagnostic tests should be applied to inpatients with advanced HIV disease.

HIV-Associated Histoplasmosis: Current Perspectives.

Histoplasmosis is an endemic mycosis caused by Histoplasma capsulatum. Infection develops by inhalation of microconidia from environmental sites inhabited by birds and bats. Disseminated disease is the usual presentation due to impaired cellular immunity. Common clinical manifestations include fever, fatigue, malaise, anorexia, weight loss, and respiratory symptoms. Histoplasma antigen detection is the most sensitive method for diagnosis. The sensitivity of the MVista® Quantitative Histoplasma antigen enzyme immunoassay is 95–100% in urine, over 90% in serum and bronchoalveolar lavage (BAL) antigen and 78% in cerebral spinal fluid (CSF). A proven diagnosis can be established by culture or pathology with sensitivities between 70% and 80%. The sensitivity of antibody detection by immunodiffusion or complement fixation was between 60% and 70%. Diagnosis using molecular methods has not been adequately validated for implementation and FDA cleared assays are unavailable. Liposomal amphotericin B should be used for 1–2 weeks followed by itraconazole for at least one year until CD4 counts are above 150 cells/mm3, HIV viral load is below 400 copies/mL and Histoplasma urine antigen is negative. Serum itraconazole level should be monitored to avoid drug toxicity. Antigen should be measured periodically to establish that treatment is effective and to assist in identifying relapse. The incidence of immune reconstitution inflammatory syndrome is low but it must be considered in patients who are thought to be failing antifungal treatment as it does not respond to changing antifungal agents but rather to initiation of corticosteroid therapy. In this review, we discuss pathogenesis, clinical manifestations, diagnosis and treatment based on personal experience and relevant publications.

Advances in Diagnosis of Progressive Pulmonary and Disseminated Coccidioidomycosis.

BackgroundAntibody detection is the main method for diagnosis of coccidioidomycosis, but it has limitations. The Coccidioides antigen enzyme immunoassay is recommended for testing cerebrospinal fluid in suspected meningitis. Reports on urine and serum antigen detection evaluated small numbers of patients who were mostly immunocompromised. The purpose of this study was to assess the accuracy of combined antibody and antigen detection for diagnosis.MethodsA retrospective study, including all patients in whom Coccidioides antigen detection in serum was performed between January 2013 and May 2017, was conducted at Valleywise Health Medical Center (formerly Maricopa Integrated Health System). Sensitivity and specificity of antigen and antibody were evaluated in 158 cases and 487 controls.ResultsThe sensitivity of antibody detection by immunodiffusion (ID) was 84.2%. The sensitivity of antigen detection was 57.0% if both urine and serum were tested and 36.7% if urine alone was tested. The sensitivity of combining antigen and ID antibody detection was 93.0%. The sensitivity of urine and serum antigen detection was 55.4% in proven and 58.7% in probable cases, 79.1% in disseminated and 41.6% in pulmonary cases, and 74.7% in immunocompromised and 40.0% in immunocompetent patients. Specificity was 99.4% for antigen detection and 96.5% for ID antibody detection. Diagnostic accuracy was 95.4% for ID antibody and antigen detection, 93.6% for ID antibody alone, and 89.1% for pathology or culture.ConclusionsThese findings support combined antibody and antigen detection for diagnosis of progressive coccidioidomycosis. The diagnosis may have been missed if antigen detection was not performed.

Fever in the returned traveller: the utility of the Platelia Dengue NS1 antigen enzyme immunoassay for the diagnosis of dengue in a non-endemic setting

Investigating fever in the returned traveller can be difficult and costly. Dengue is one of the most frequently reported aetiologies. NS1 is a non-structural dengue virus protein detectable during acute infection. The aim of this report is to describe the utility of the Platelia Dengue NS1 antigen enzyme immunoassay (EIA) for detection of dengue in a non-endemic region compared to a composite gold standard of contemporaneous molecular testing and seroconversion. We performed a retrospective analysis of all dengue serology tests from 6 February 2012 to 5 December 2018. Dengue serology and in-house flavivirus molecular results were identified using the laboratory information management system. Dengue serology was performed using the Bio-Rad Platelia Dengue NS1 antigen EIA, and Abbott Panbio Dengue IgG and IgM EIA. True positive NS1 result was defined as positive molecular test within one week of the positive NS1 result or seroconversion within 120 days. NS1 negative samples that remained negative to all dengue markers on repeat more than 10 and up to 120 days after were labelled as true negatives. More than 75% of cases had a serology pattern consistent with primary dengue. Sensitivity and specificity of NS1 Ag EIA was 96.4% (95% CI 92.3-98.7%) and 98.4% (95% CI 94.5-99.8%), respectively. Performance was poorer in serotype 4 infections (sensitivity 50%). Platelia Dengue NS1 antigen EIA test performance in the returned traveller cohort fulfils the remit as a single diagnostic test for acute dengue infection.

Survey of Histoplasma capsulatum in bat guano and status of histoplasmosis in Slovenia, Central Europe

There have been increasing reports on the presence of Histoplasma capsulatum in some European countries. The study investigated the presence of Histoplasma in bat guanos, speleologists with records of visiting Histoplasma-endemic regions and patients with histoplasmosis. A commercial ALPHA Histoplasma Antigen enzyme immunoassay was tested as an alternative methodology to detect Histoplasma in environment and compared with polymerase chain reaction (PCR) assays. The presence of Histoplasma antigen in bat guanos was not confirmed by PCR. Among 14 healthy speleologists, two were indicated as having the Histoplasma antigen in urine, but expressed negative PCR-specific results for the Histoplasma antigen. Five unequivocal cases of imported acute pulmonary histoplasmosis in Slovenia between years 2005 and 2016 were confirmed in patients returning from North and South America after visiting hazardous localities e.g., caves with guano, and places with dust. Currently there is no evidence of autochthonous histoplasmosis in Slovenia, or that bat guano is a source of H. capsulatum. Involvement of histoplasmosis in travellers’ and cavers’ morbidity might be underestimated in non-endemic areas. It is crucial to ensure the use of appropriate protective equipment in Histoplasma hazardous localities, to spread information about this hazardous microbe to vulnerable populations and to monitor the health of the environment. A differential diagnosis for a febrile respiratory disease outbreak in patients returning from endemic regions should trigger routine consideration of possible histoplasmosis.

Helicobacter pylori eradication therapy in patients with immune thrombocytopenic purpura: a single-center experience

Objective: The aim was to determine the frequency of Helicobacter pylori infection in patients with immune thrombocytopenic purpura (ITP) and to detect the effect of its eradication therapy on platelet count response in H. pylori-positive patients. Patients and methods: H. pylori stool antigen enzyme immunoassay test was done for 54 patients with ITP. H. pylori-positive patients received standard triple therapy for 2 weeks. Platelet count response to H. pylori eradication therapy was evaluated 2 and 3 months after the treatment. Results: H. pylori infection was positive in 38 (70.4%) patients with ITP. Eradication of H. pylori was achieved in 30 (78.9%) patients (responder group). There were statistically significant increases in platelet count (among the responder group) after H. pylori eradication therapy at 2 and 3 months (P Conclusion: Eradication of H. pylori infection in patients with ITP is associated with significant increase in platelet counts.

The Effect of Red Rose Extract on Helicobacter Pylori Eradication: A Randomized Controlled Clinical Trial

Background Helicobacter pylori (H. pylori) is believed to be the most common bacterial infection worldwide. Herein, we aimed to investigate the effect of red rose extract on H. pylori eradication in 9 to 15 year-old children. Materials and Methods In this randomized controlled clinical trial,by the convenience sampling method 332 children were screened for H. pylori infection. Their stool samples were studied by the H. pylori Antigen Enzyme Immunoassay (EIA) Test Kit. Positive cases were divided into two groups and received either red rose extract or placebo for 14 days. The stool exam was repeated once again at treatment termination. Data were analyzed using SPSS software (version 21.0). Results In total 17.8% (n=59) of the 332 cases were positive for Helicobacter pylori stool antigen (HpSA), and 56 completed the study. The mean HpSA titer remained stable in the red rose extract treated group but it increased in the placebo group following intervention; however, the difference was statistically insignificant (p=0.57). Moreover, the intervention resulted in no difference in the eradication rate between the two groups (p=0.57). Conclusion Red rose extract did not reduce HpSA titer after intervention, so it cannot be solely considered as an alternative for H. pylori eradication. Further studies with a different dose and duration or with combined regimens are recommended.

A survey of primary-care pediatricians regarding the management of Helicobacter pylori infection and celiac disease

BackgroundAdherence of primary-care pediatricians to guidelines in pediatric gastroenterology is essential to achieve optimal clinical outcomes. The study aim was to examine adherence of primary-care pediatricians to the European and North American Societies for Pediatric Gastroenterology, Hepatology and Nutrition guidelines on the management of Helicobacter pylori (H. pylori) infection and celiac disease.MethodsWe conducted a cross-sectional study during March–July 2017 using the survey platform of Maccabi Healthcare Services, the second largest state-mandated health organization in Israel. We sent the study questionnaire to a random sample of 300 pediatricians via electronic mails and to increase the response rate, we performed a telephone interview. Overall, 108 (36%) pediatricians provided completed questionnaires.ResultsUsing professional guidelines for the management of H. pylori infection and celiac disease was reported by 34 and 37% of pediatricians, respectively. Referral to H. pylori testing was reported by 78 and 52% of pediatricians in children with suspected duodenal ulcer and unexplained iron deficiency anemia, respectively, with the stool antigen enzyme immunoassay being mostly (51%) used as the first choice diagnostic test. Most pediatricians reported prescription of triple therapy; proton pump inhibitors/clarithromycin/amoxicillin (59%) or metronidazole (21%). For celiac disease, overall adherence to all guidelines was high both for initial evaluation and for confirmation of diagnosis.ConclusionsAdherence to the guidelines on management of H. pylori infection was low, while adherence to the guidelines on celiac disease management was high among primary-care pediatricians. Educational interventions are needed to improve H. pylori infection management among primary-care pediatricians.

263. Advances in Diagnosis of Progressive Coccidioidomycos: Experience in 164 Cases and 508 Controls

BackgroundAntibody detection is the main method for diagnosis of coccidioidomycosis but has limitations including sensitivity and turnaround time. The MVISTA Coccidioides antigen enzyme immunoassay (EIA) is recommended for testing CSF in suspected Coccidioides meningitis. The early reports on urine and serum antigen testing evaluated small numbers of patients who were mostly immunocompromised with advanced disease.MethodsA retrospective study, including all patients in whom Coccidioides antigen testing was performed between January 2013 and May 2017, was conducted at Maricopa Integrated Health System (MIHS). Sensitivity and specificity of antigen testing at MiraVista Diagnostics and antibody testing at MIHS or commercial laboratories were evaluated in 164 cases and 508 controls.ResultsThe sensitivity of antigen testing was 51% and specificity was 99%. The sensitivity of antigen detection was highest if both urine and serum were tested (57%) than if only urine was tested (38%). The sensitivity of antibody testing was 84% and the specificity was 94% by immunodiffusion (ID). The sensitivity and specificity of antigen or ID antibody testing both were 94%. Sensitivity of antigen testing was 57% in proven and 58% in probable cases, ID antibody in 85% of proven and 75% of probable and antigen or ID antibody in 93% of proven and 95% of probable cases. Antigen was detected more often in disseminated (79%) than pulmonary cases (42%) as was ID antibody, 91% and 79%, respectively. Antigen testing was more sensitive in immunocompromised (76%) than non-immunocompromised patients (41%) while ID antibody was less sensitive in immunocompromised (74%) than in non-immunocompromised patients (93%). Combined antigen and ID antibody testing provided the highest sensitivity, 94% in all cases, 94% in immunocompromised and 95% in non-immunocompromised patients.ConclusionThese findings support testing urine and serum for Coccidioides antigen and serum for ID antibodies for diagnosis of progressive pulmonary or disseminated coccidioidomycosis. Disclosures All authors: No reported disclosures.

The Feasibility of Metagenomic Next-Generation Sequencing to Identify Pathogens Causing Tuberculous Meningitis in Cerebrospinal Fluid.

PurposeThe application of metagenomic next-generation sequencing (mNGS) in the diagnosis of tuberculous meningitis (TBM) remains poorly characterized. Here, we retrospectively analyzed data from patients with TBM who had taken both mNGS and conventional tests including culture of Mycobacterium tuberculosis (MTB), polymerase chain reaction (PCR) and acid-fast bacillus (AFB) stain, and the sensitivity and specificity of these methods were compared.MethodsWe retrospectively recruited TBM patients admitted to the hospital between December 2015 and October 2018. The first collection of cerebrospinal fluid (CSF) samples underwent both mNGS and conventional tests. In addition, patients with bacterial/cryptococcal meningitis or viral meningoencephalitis were mNGS positive controls, and a patient with auto-immune encephalitis was an mNGS negative control.ResultsTwenty three TBM patients were classified as 12 definite and 11 clinical diagnoses, which were based on clinical manifestations, pathogen evidence, CSF parameters, brain imaging, and treatment response. The mNGS method identified sequences of Mycobacterium tuberculosis complex (MBTC) from 18 samples (18/23, 78.26%). In patients with definite TBM, the sensitivity of mNGS, AFB, PCR, and culture to detect MTB in the first CSF samples were 66.67, 33.33, 25, and 8.33%, respectively. The specificity of each method was 100%. Among the four negative mNGS cases (4/23, 17.39%), three turned out positive by repeated AFB stain. The agreement of mNGS with the total of conventional methods was 44.44% (8/18). Combination of mNGS and conventional methods increased the detection rate to 95.65%. One patient was diagnosed as complex of TBM and cryptococcal meningitis, in which AFB stain and cryptococcal antigen enzyme immunoassay were positive and the DNA of Cryptococcus neoformans was detected by mNGS.ConclusionOur study indicates that mNGS is an alternative method to detect the presence of mycobacterial DNA in CSF samples from patients with TBM and deserves to be applied as a front-line CSF test.

Novel canine anti-Coccidioides immunoglobulin G enzyme immunoassay aids in diagnosis of coccidioidomycosis in dogs.

The diagnosis of coccidioidomycosis (CM) in dogs is typically based on clinical presentation, serology, and (less frequently) spherule identification. Agar gel immunodiffusion (AGID) is the most commonly employed serological method, but AGID is slow (requiring up to a week for titer). A Coccidioides antigen enzyme immunoassay (EIA) is also available; however, sensitivity is low in CM dogs. An antibody EIA was developed to detect canine immunoglobulin G (IgG) reacting to Coccidioides antigens. Serum was evaluated from dogs with pathology proven CM and/or AGID positive CM, as well as dogs with histoplasmosis, blastomycosis, non-fungal infections, or healthy dogs. A standard curve was used to convert optical density (OD) values into EIA units (EU). Serum and urine samples from CM dogs were also tested in the antigen EIA. Sensitivity and specificity for IgG were 89.2% and 97.2%, respectively, upon evaluation of dogs with proven or probable CM and control dogs. Cross-reactivity was observed in 7.7% and in 6.4% of dogs with histoplasmosis or blastomycosis, respectively. The antigen EIA alone was insensitive (33.8%). Combined IgG and antigen testing increased sensitivity to 93.2%, as three dogs were IgG-negative but had detectable serum or urine antigen. In 22 dogs with proven CM, sensitivity was statistically similar for antibody EIA and AGID (86% and 73%; P = .487). The MiraVista® canine Coccidioides antibody IgG EIA may aid in the diagnosis of CM by improving turnaround time with comparable sensitivity to AGID. Serial or concurrent testing by antibody and antigen EIAs may be beneficial when screening dogs for CM.

Molecular Characterization of Rotavirus Strains Circulating in Enugu Nigeria: 2011 to 2016

Rotavirus gastroenteritis is a major public health concern globally, estimated to cause 215,000 deaths among children < 5 years of age in 2013; with majority of mortality occurring in developing countries. In 2013, it was estimated that Nigeria was the second country with the highest number of rotavirus deaths. Monitoring of circulating rotavirus strains in Enugu, Nigeria is part of on-going rotavirus surveillance before the introduction of rotavirus vaccination. A total of 2694 stool samples were collected from enrolled under 5 years old children with diarrhoea between January 2011 and December 2016 and tested the virus using an antigen enzyme immunoassay. Randomly selected rotavirus positive samples were further characterized by rotavirus genotype methods to identify the G and P types circulating during the study period. Rotavirus was detected in 1242 (46%) of the 2694 samples collected over the six years period. Of these, 867 were randomly selected for genotyping. G and P types could be assigned for 832 samples (96%), while 31 (3.6%) could only be assigned either genotype G or P (partially typed) and 4 (0.4%) could not be assigned genotype G and P (untypeable). The most common G-genotypes detected during the entire study period were G12, G1 and G3 accounting for 27.6%, 21.0% and 16.3% respectively. Mixed G and P-genotypes were commonly detected. Ninety-one of the samples, representing 10.8% (91/839) had mixed G-genotype whilst 130 of the samples representing 15.2% (130/852) had mixed P-genotype. The most common P-genotypes detected were P[8], P[6] and P[4] representing 38.3%, 35.4% and 9.1% respectively. The predominant strain detected was G12P[8] (22.3%) followed by G3P[6] (14.5%), G1P[8] (9.2%) and G1P[6] (8.0%). These data are useful for making an informed decision about the introduction of rotavirus vaccine into the national routine immunization program and to monitor the impact of the vaccine post licensure.