Follow-Up and Comparative Assessment of SARS-CoV-2 IgA, IgG, Neutralizing, and Total Antibody Responses After BNT162b2 or mRNA-1273 Heterologous Booster Vaccination.

BACKGROUND: Priming with ChAdOx1 followed by heterologous boosting is considered in several countries. Nevertheless, analyses comparing the immunogenicity of heterologous booster to homologous primary vaccination regimens and natural infection are lacking. In this study, we aimed to conduct a comparative assessment of the immunogenicity between homologous primary vaccination regimens and heterologous prime-boost vaccination using BNT162b2 or mRNA-1273. METHODS: We matched vaccinated naïve (VN) individuals (n = 673) with partial vaccination (n = 64), primary vaccination (n = 590), and primary series plus mRNA vaccine heterologous booster (n = 19) with unvaccinated naturally infected (NI) individuals with a documented primary SARS-CoV-2 infection (n = 206). We measured the levels of neutralizing total antibodies (NTAbs), total antibodies (TAbs), anti-S-RBD IgG, and anti-S1 IgA titers. RESULTS: Homologous primary vaccination with ChAdOx1 not only showed less potent NTAb, TAb, anti-S-RBD IgG, and anti-S1 IgA immune responses compared to primary BNT162b2 or mRNA-1273 vaccination regimens (p < 0.05) but also showed ~3-fold less anti-S1 IgA response compared to infection-induced immunity (p < 0.001). Nevertheless, a heterologous booster led to an increase of ~12 times in the immune response when compared to two consecutive homologous ChAdOx1 immunizations. Furthermore, correlation analyses revealed that both anti-S-RBD IgG and anti-S1 IgA significantly contributed to virus neutralization among NI individuals, particularly in symptomatic and pauci-symptomatic individuals, whereas among VN individuals, anti-S-RBD IgG was the main contributor to virus neutralization. CONCLUSION: The results emphasize the potential benefit of using heterologous mRNA boosters to increase antibody levels and neutralizing capacity particularly in patients who received primary vaccination with ChAdOx1.

Serological assessment of SARS-CoV-2 infection during the first wave of the pandemic in Louisville Kentucky.

Serological assays intended for diagnosis, sero-epidemiologic assessment, and measurement of protective antibody titers upon infection or vaccination are essential for managing the SARS-CoV-2 pandemic. Serological assays measuring the antibody responses against SARS-CoV-2 antigens are readily available. However, some lack appropriate characteristics to accurately measure SARS-CoV-2 antibodies titers and neutralization. We developed an Enzyme-linked Immunosorbent Assay (ELISA) methods for measuring IgG, IgA, and IgM responses to SARS-CoV-2, Spike (S), receptor binding domain (RBD), and nucleocapsid (N) proteins. Performance characteristics of sensitivity and specificity have been defined. ELISA results show positive correlation with microneutralization and Plaque Reduction Neutralization assays with infectious SARS-CoV-2. Our ELISA was used to screen healthcare workers in Louisville, KY during the first wave of the local pandemic in the months of May and July 2020. We found a seropositive rate of approximately 1.4% and 2.3%, respectively. Our analyses demonstrate a broad immune response among individuals and suggest some non-RBD specific S IgG and IgA antibodies neutralize SARS-CoV-2.

Serological diagnostic for SARS-CoV-2: an experimental External Quality Assessment Scheme.

OBJECTIVES: Numerous analytical systems, rapidly made available on the market throughout the SARS-CoV-2 pandemic, aim to detect COVID-19, and to continuously update and improve the same systems. Medical laboratory professionals have also developed in-house analytical procedures in order to satisfy the enormous volume of requests for tests. These developments have highlighted the need control the analytical procedures used in order to guarantee patient safety. The External Quality Assessment (EQA) Scheme, an important quality assurance tool, aims to guarantee high standard performance for laboratory and analytical procedures. The aim of the present study was to report on the results collected in an experimental EQA scheme for the serological diagnosis of SARS-CoV-2. METHODS: All qualitative results collected in the different EQA surveys were summarized in order to identify the percentage of laboratory results in relation to typology of antibodies, results and samples. RESULTS: A total of 4,867 data sets were collected. The analysis of EQA data made, demonstrates a better agreement among laboratories results for total Ig than single immunoglobulins (IgG, IgM, IgA) in the case samples positive for SARS-CoV-2, and a wide divergence between IgM results for positive samples (only 34.9% were correct). Results for negative controls and specificity controls demonstrated a better overall agreement than results for positive samples. CONCLUSIONS: Working in collaboration with the IVD manufacturers, laboratory professionals must strive to achieve harmonization of results, and to develop well-defined protocols complying with the ISO 15189 requirements.

Multicenter evaluation of four immunoassays for the performance of early diagnosis of COVID-19 and assessment of antibody responses of patients with pneumonia in Taiwan.

BACKGROUND/PURPOSE: Our study goals were to evaluate the diagnostic performance of four anti-SARS-CoV-2 antibodies tests and the differences in dynamic immune responses between COVID-19 patients with and without pneumonia. METHODS: We collected 184 serum samples from 70 consecutively qRT-PCR-confirmed COVID-19 patients at four participating hospitals from 23 January 2020 to 30 September 2020. COVID-19 pneumonia was defined as the presence of new pulmonary infiltration. Serum samples were grouped by the duration after symptom onset on a weekly basis for antibody testing and analysis. The four immunoassays: Beckman SARS-CoV-2 IgG/IgM (Beckman Test), Siemens (ADVIA Centaur®) SARS-CoV-2 Total (COV2T) (Siemens Test), SBC COVID-19 IgG ELISA (SBC Test) and EliA SARS-CoV-2-Sp1 IgG/IgM/IgA P2 Research (EliA Test) were used for detecting the SARS-CoV-2 specific antibodies. RESULTS: The sensitivity of all tests reached 100% after 42 days of symptom onset. Siemens Test, the only test detecting total anti-SARS-CoV-2 antibodies, had the best performance in the early diagnosis of COVID-19 infection (day 0-7: 77%; day 8-14: 95%) compared to the other 3 serological tests. All tests showed 100% specificity except SBC Test (98%). COVID-19 patients with pneumonia had significantly higher testing signal values than patients without pneumonia (all p values < 0.05, except EliA IgM Test). However, Siemens Test and SBC Test had highest probability in early prediction of the presence of COVID-19 pneumonia. CONCLUSION: Chronological analysis of immune response among COVID-19 patients with different serological tests provides important information in the early diagnosis of SARS-CoV-2 infection and prediction of the risk of pneumonia after infection.

Disease monitoring with quantitative serum IgA levels provides a more reliable response assessment in multiple myeloma patients.

Unlike IgG monoclonal proteins (MCPs), IgA MCP quantification is unreliable due to beta-migration of IgA MCPs on serum protein electrophoresis (SPEP). The utility of nephelometric quantitative IgA (qIgA) to monitor IgA multiple myeloma (MM) is unclear. We retrospectively studied disease response kinetics using qIgA versus MCPs by SPEP, and developed and validated novel qIgA disease assessment criteria in 491 IgA MM patients. The SPEP MCP nadir occurred a median of 41 (IQR 0-102) days before the qIgA. The median time to achieve a partial response (PR) was shorter using standard IMWG versus qIgA response criteria (32 vs 58 days, p < 0.001). Stratification by qIgA criteria, unlike IMWG criteria, led to clear separation of the progression-free survival curves of patients achieving a PR or very good PR. There was a consistent trend toward earlier detection of disease progression using qIgA versus IMWG progression criteria. In conclusion, monitoring IgA MM using MCP-based IMWG criteria may be falsely reassuring, given that MCP levels on SPEP decrease faster than qIgA levels. The qIgA response criteria more accurately stratify patients based on the progression risk and may detect disease progression earlier, which may lead to more consistent measurement of trial endpoints and improved patient outcomes.

Assessment of commercial SARS-CoV-2 antibody assays, Jamaica.

BACKGROUND: The performance of the Roche Elecsys® Anti-SARS-CoV-2, Abbott Architect SARS-CoV-2 IgM, Abbott Architect SARS-CoV-2 IgG, Euroimmun SARS-CoV-2 IgA, Euroimmun SARS-CoV-2 IgG ELISA, and Trillium IgG/IgM rapid assays was evaluated in Jamaica. METHODS: Diagnostic sensitivities of the assays were assessed by testing serum samples from SARS-CoV-2 PCR-confirmed persons and diagnostic specificity was assessed by testing serum samples collected during 2018-2019 from healthy persons and from persons with antibodies to a wide range of viral infections. RESULTS: Serum samples collected ≥14 days after onset of symptoms, or an initial SARS-CoV-2 RT-PCR positive test for asymptomatics, showed diagnostic sensitivities ranging from 67.9 to 75.0% when including all possible disease severities and increased to 90.0-95.0% when examining those with moderate to critical disease. Grouping moderate to critical disease showed a significant association with a SARS-CoV-2 antibody positive result for all assays. Diagnostic specificity ranged from 96.7 to 100.0%. For all assays examined, SARS-CoV-2 real-time PCR cycle threshold (Ct) values of the initial nasopharyngeal swab sample testing positive were significantly different for samples testing antibody positive versus negative. CONCLUSIONS: These data from a predominantly African descent Caribbean population show comparable diagnostic sensitivities and specificities for all testing platforms assessed and limited utility of these tests for persons with asymptomatic and mild infections.

Assessment of SARS-CoV-2 serological tests for the diagnosis of COVID-19 through the evaluation of three immunoassays: Two automated immunoassays (Euroimmun and Abbott) and one rapid lateral flow immunoassay (NG Biotech).

BACKGROUND: The emergence of new SARS-CoV-2 has promoted the development of new serological tests that could be complementary to RT-PCR. Nevertheless, the assessment of clinical performances of available tests is urgently required as their use has just been initiated for diagnose. OBJECTIVES: The aim of this study was to assess the performance of three immunoassays for the detection of SARS-CoV-2 antibodies. METHODS: Two automated immunoassays (Abbott SARS-CoV-2 CLIA IgG and Euroimmun Anti-SARS-CoV-2 ELISA IgG/IgA assays) and one lateral flow immunoassay (LFIA NG-Test® IgG-IgM COVID-19) were tested. 293 specimens were analyzed from patients with a positive RT-PCR response, from patients with symptoms consistent with COVID-19 but exhibiting a negative response to the RT-PCR detection test, and from control group specimens. Days since symptoms onset were collected from clinical information sheet associated with respiratory tract samples. RESULTS: Overall sensitivity for IgG was equivalent (around 80 %) for CLIA, ELISA and LFIA. Sensitivity for IgG detection, >14 days after onset of symptoms, was 100.0 % for all assays. Overall specificity for IgG was greater for CLIA and LFIA (more than 98 %) compared to ELISA (95.8 %). Specificity was significantly different between IgA ELISA (78.9 %) and IgM LFIA (95.8 %) (p < 0.05). The best agreement was observed between CLIA and LFIA assays (97 %; k = 0.936). CONCLUSION: Excellent sensitivity for IgG detection was obtained >14 days after onset of symptoms for all immunoassays. Specificity was also excellent for IgG CLIA and IgG LFIA. Our study shows that NG-Test® is reliable and accurate for routine use in clinical laboratories.

Fast and sensitive diagnosis of autoimmune disorders through amperometric biosensing of serum anti-dsDNA autoantibodies.

This work reports the first amperometric biosensor involving the use of neutravidin-functionalized magnetic microbeads (NA-MBs) modified with a biotinylated-anti-dsDNA (b-dsDNA) as efficient magnetic microcarriers to selectively capture anti-dsDNA autoantibodies (IgG, IgA and IgM AAbs) present in the sera of patients with rheumatoid arthritis (RA). Subsequently, the attached anti-dsDNA AAbs are detected with a mixture of conventional HRP-labeled secondary antibodies (HRP-anti-human IgG/IgM/IgA mixture). The biorecognition event is monitored by amperometric transduction using the hydroquinone (HQ)/H2O2 system upon capturing the modified MBs on the surface of screen-printed carbon electrodes (SPCEs). The developed bioplatform exhibits a linear calibration plot ranging from 1 to 200 IU mL-1 with a LOD of 0.3 IU mL-1 for anti-dsDNA AAbs standards. In addition, the biosensor allows performing the determination of the anti-dsDNA AAbs levels directly in 100-times diluted serum samples from patients diagnosed with RA and in just 75 min. The obtained results are in agreement with those provided by an ELISA kit and allow discrimination between positive and negative samples.

Assessment of IgA anti-PT and IgG anti-ACT reflex testing to improve Bordetella pertussis serodiagnosis in recently vaccinated subjects.

OBJECTIVES: Quantifying IgG antibodies to pertussis toxin (PT) is the most specific and sensitive method for the serodiagnosis of a Bordetella pertussis infection. Since PT is a component of acellular pertussis vaccines, anti-PT IgG is also induced by vaccination, precluding pertussis serodiagnosis based exclusively on anti-PT IgG in recently vaccinated subjects. Here, we aim to identify additional B. pertussis-specific serological markers that can discriminate between infection and recent vaccination. METHODS: The clinical usefulness of measuring IgA directed to the vaccine antigen PT and IgG directed to non-vaccine antigens (Fim2/3, LPS, ACT, CatACT) was evaluated in nine well characterized subject groups, aged 10-89 years (n = 390). Serum anti-PT IgG levels (>125 IU/mL) served as an indicator for a recent B. pertussis infection. Comparing symptomatic pertussis-infected subjects (n = 140) with recently vaccinated, non-infected subjects (n = 100) revealed the optimal cut-off, accuracy, sensitivity and specificity for each single parameter. RESULTS: For pertussis diagnosis in recently vaccinated subjects, the measurement of anti-PT IgA (cut-off 15 IU/mL) and anti-ACT IgG (cut-off 15 U/mL) resulted in accuracies of 95% (91.5-97.1) and 87.5% (82.7-91.1), sensitivities of 92.9% (87.4-96.0) and 83.6% (76.5-88.8) and specificities of 98% (93.0-99.4) and 93% (86.3-96.6), respectively. Comparing anti-PT IgA levels between the youngest (10-19 years, n = 38) and oldest (70-89 years, n = 17) age groups revealed an age-dependent increase in antibody levels in pertussis-infected subjects (p < 0.0001). CONCLUSIONS: Reflex testing of anti-PT IgA and anti-ACT IgG improves pertussis serodiagnosis in recently vaccinated symptomatic subjects with elevated anti-PT IgG levels. Furthermore, both markers can discriminate between vaccination and recent infection in pertussis serosurveillance studies.

Improvements in diagnosis and risk assessment of primary and secondary antiphospholipid syndrome.

Classification criteria for antiphospholipid syndrome have not been updated since the revised Sapporo classification criteria were published in 2006. These criteria have limitations in that they omit nonclassical manifestations (hematologic and neurologic), include anticardiolipin and anti-ß2-glycoprotein I immunoglobulin (Ig)M isotypes, and do not separately consider primary (no autoimmune disease) or secondary (usually systemic lupus erythematosus) disease. Recent findings in antiphospholipid antibody include fluctuation of antiphospholipid antibodies, recognition that IgA isotypes do confer risk, identification of the role of complementopathy in catastrophic antiphospholipid syndrome, and elucidation of the role of thrombosis risk equations.

Multilaboratory Assessment of Epstein-Barr Virus Serologic Assays: the Case for Standardization.

IgA antibodies targeting Epstein-Barr virus (EBV) have been proposed for screening for nasopharyngeal carcinoma (NPC). However, methods differ, and the antigens used in these assays differ considerably between laboratories. To enable formal comparisons across a range of established EBV serology assays, we created a panel of 66 pooled serum samples and 66 pooled plasma samples generated from individuals with a broad range of IgA antibody levels. Aliquots from these panels were distributed to six laboratories and were tested by 26 assays measuring antibodies against VCA, EBNA1, EA-EBNA1, Zta, or EAd antigens. We estimated the correlation between assay pairs using Spearman coefficients (continuous measures) and percentages of agreement (positive versus negative, using predefined positivity cutoffs by each assay developer/manufacturer). While strong correlations were observed between some assays, considerable differences were also noted, even for assays that targeted the same protein. For VCA-IgA assays in serum, two distinct clusters were identified, with a median Spearman coefficient of 0.41 (range, 0.20 to 0.66) across these two clusters. EBNA1-IgA assays in serum grouped into a single cluster with a median Spearman coefficient of 0.79 (range, 0.71 to 0.89). Percentages of agreement differed broadly for both VCA-IgA (12% to 98%) and EBNA1-IgA (29% to 95%) assays in serum. Moderate-to-strong correlations were observed across assays in serum that targeted other proteins (correlations ranged from 0.44 to 0.76). Similar results were noted for plasma. We conclude that standardization of EBV serology assays is needed to allow for comparability of results obtained in different translational research studies across laboratories and populations.

Seroprevalence of IgA and IgM antibodies to Bordetella pertussis in healthy Japanese donors: Assessment for the serological diagnosis of pertussis.

Pertussis is a human respiratory infection caused by the gram-negative bacterium, Bordetella pertussis. To evaluate the pertussis burden and vaccine efficacy, diagnosis and epidemiological surveillance should be based on accurate and valid diagnostic methods. Recently, the serodiagnostic tests Novagnost Bordetella pertussis IgA and IgM were approved in Japan for pertussis diagnostics. Although the anti-pertussis toxin (PT) IgG assay has been used for pertussis diagnosis worldwide, little is known about the anti-B. pertussis IgA and IgM assays. In this study, serum samples from 460 healthy donors were examined to determine the seroprevalence of anti-B. pertussis IgA and IgM in a Japanese population, and its correlation with donor age. Our data demonstrated that anti-B. pertussis IgA and IgM are positively and negatively correlated with age (r = 0.27, r = -0.37; P < 0.001, respectively). Age-specific analysis revealed high titers of anti-B. pertussis IgA in adults (46-50 years), while anti-B. pertussis IgM titers were high in schoolchildren (6-10, 11-15 years). When applying the arbitrary cut-off values for these ages, 17.6% and 39.5% of healthy donors were interpreted as pertussis-positive or indeterminate with anti-B. pertussis IgA (46-50 years) and IgM (11-15 years) titers, respectively. Overall, our findings indicated that the Novagnost Bordetella pertussis IgA and IgM testing could be greatly affected by subject age, limiting its value for pertussis diagnosis.

Assessment of Serological Markers of Genital Chlamydia trachomatis Infection Among the Gynaecology Patients attending Babcock University Teaching Hospital, Ilishan-Remo, Ogun State, Nigeria.

Genital Chlamydia trachomatis infection causes significant morbidity and mortality in women. A number of epidemiologic studies have suggested that Polymerase Chain Reaction (PCR) is more accurate as a diagnostic tool for Chlamydia trachomatis. However, the use of serological markers may be cost effective and practical in diagnosing and estimating the burden of the disease in resource limited countries.This study was aimed at determining the serological markers (IgG, IgM and IgA) of Chlamydia trachomatis, evaluate the association between Chlamydia trachomatis infection and the sociodemographic characteristics and clinical diagnosis of the participants. This was a cross sectional hospital-based study in which blood samples from 145 consenting participants were tested for IgG, IgM and IgA antibodies against Chlamydia trachomatis using enzyme linked immunosorbent assay and their clinical diagnosis, retrieved from their case notes. The cumulative prevalence of seropositivity for Chlamydia trachomatis (IgG, IgM, IgA) was 112 (77.2%) while 33 (22.8%) were seronegative. The overall predominant seromarker was IgG 91(62.8%) while IgM and IgA accounted for 85(58.6%) and 54(37.2%) respectively. A statistically significant association was found between Chlamydia trachomatis infection and PID (p value = 0.031), primary infertility (p value 0.011) and level of income (p value= (0,045).

Impact Exerted by Nutritional Risk Screening on Clinical Outcome of Patients with Esophageal Cancer.

OBJECTIVE: Preoperative nutritional status of patients is closely associated with their recovery after the surgery. This study aims to ascertain the impact exerted by the nutritional risk screening on clinical outcome of patients with esophageal cancer. METHODS: 160 patients with esophageal cancer aged over 60, having got therapy at the First Hospital of Jilin University from Jun 2016 to Feb 2017 were evaluated by adopting the NRS2002. 80 cases of patients got active therapy of nutritional support, and the other patients not supported nutritionally were selected as the control group. The comparison was drawn between two groups in serum albumin, serum immunoglobulin, postoperative complications, hospitalization, and hospitalization expenses. RESULTS: For all the patients, in 3 and 7 days after the surgery, the serum albumin in the nutritionally supported group outstripped that in group without nutritional support (P < 0.05) regardless of the nutritional risk. For the patients in the risk of nutrition, the IgA in the nutritionally supported group outstripped that of group without nutritional support (P < 0.05) in 3 and 7 days before the surgery, and the serum IgG outstripped that of the group without nutritional support in 1 and 3 days before the surgery (P < 0.05). In terms of the patients in the risk of nutrition, the average hospitalization of nutritionally supported group was shorter (P < 0.05), and the average hospitalization expenses were lower compared with those of the group without nutritional support. And for the patients in no risk, the hospitalization expenses of supported group surmounted those of group without nutritional support (P < 0.05), whereas the average hospitalization took on no statistic difference (P > 0.05). CONCLUSION: For the patients in the risk of nutrition, preoperative nutritional support can facilitate the nutritional status and immunization-relative result after surgery, which shall also decrease the average hospitalization and hospitalization cost.

Strategies to improve the efficiency of celiac disease diagnosis in the laboratory.

The demand for testing to detect celiac disease (CD) autoantibodies has increased, together with the cost per case diagnosed, resulting in the adoption of measures to restrict laboratory testing. We designed this study to determine whether opportunistic screening to detect CD-associated autoantibodies had advantages compared to efforts to restrict testing, and to identify the most cost-effective diagnostic strategy. We compared a group of 1678 patients in which autoantibody testing was restricted to cases in which the test referral was considered appropriate (G1) to a group of 2140 patients in which test referrals were not reviewed or restricted (G2). Two algorithms A (quantifying IgA and Tissue transglutaminase IgA [TG-IgA] in all patients), and B (quantifying only TG-IgA in all patients) were used in each group, and the cost-effectiveness of each strategy was calculated. TG-IgA autoantibodies were positive in 62 G1 patients and 69 G2 patients. Among those positive for tissue transglutaminase IgA and endomysial IgA autoantibodies, the proportion of patients with de novo autoantibodies was lower (p=0.028) in G1 (11/62) than in G2 (24/69). Algorithm B required fewer determinations than algorithm A in both G1 (2310 vs 3493; p<0.001) and G2 (2196 vs 4435; p<0.001). With algorithm B the proportion of patients in whom IgA was tested was lower (p<0.001) in G2 (29/2140) than in G1 (617/1678). The lowest cost per case diagnosed (4.63 euros/patient) was found with algorithm B in G2. We conclude that opportunistic screening has advantages compared to efforts in the laboratory to restrict CD diagnostic testing. The most cost-effective strategy was based on the use of an appropriate algorithm.

Implementation of National Institute for Health and Care Excellence (NICE) guidance to measure immunoglobulin A with all coeliac screens: can an affordable solution be devised?

There has been a dramatic increase in requests for coeliac disease (CD) serological screening using immunoglobulin (Ig)A tissue transglutaminase antibodies (IgA-tTG). Recently, the UK National Institute for Health and Care Excellence has revised its guidance, recommending that total IgA should also be measured in all samples. This is justified, as false-negative results may occur with IgA deficiency. However, implementation of this guidance will incur considerable expense. Tests that measure IgA-tTG antibodies can detect IgA deficiency, indicated by low background signal. This provides an opportunity to identify samples containing IgA ≤ 0·2g/l, obviating the need for unselected IgA measurement. We investigated the feasibility of this approach in two centres that use the EliA™ Celikey™ assay or QUANTA Lite® enzyme-linked immunosorbent assay to quantify IgA-tTG antibodies. In both cases, total IgA correlated strongly with background IgA-tTG assay signal. Using the Celikey™ assay, a threshold of < 17·5 response units achieved 100% sensitivity (95% confidence intervals 79·4-100%) for detection of IgA ≤ 0·2g/l, circumventing the need for IgA testing in > 99% of sera. A similar principle was demonstrated for the QUANTA Lite® assay, whereby a threshold optical density of < 0·0265 also achieved 100% sensitivity (95% confidence intervals 78·2-100%) for IgA ≤ 0·2 g/l, avoiding unnecessary IgA testing in 67% of cases. These data suggest that CD screening tests can identify samples reliably containing low IgA in a real-life setting, obviating the need for blanket testing. However, this approach requires careful individualized validation, given the divergent efficiency with which assays identify samples containing low IgA.

Rotavirus specific maternal antibodies and immune response to RV3-BB neonatal rotavirus vaccine in New Zealand.

BACKGROUND: Maternal antibodies, acquired passively via placenta and/or breast milk, may contribute to the reduced efficacy of oral rotavirus vaccines observed in children in developing countries. This study aimed to investigate the effect of rotavirus specific maternal antibodies on the serum IgA response or stool excretion of vaccine virus after any dose of an oral rotavirus vaccine, RV3-BB, in parallel to a Phase IIa clinical trial conducted at Dunedin Hospital, New Zealand. At the time of the study rotavirus vaccines had not been introduced in New Zealand and the burden of rotavirus disease was evident. METHODS: Rotavirus specific IgG and serum neutralizing antibody (SNA) levels in cord blood and IgA levels in colostrum and breast milk samples collected ∼4 weeks, ∼20 weeks and ∼28 weeks after birth were measured. Infants were randomized to receive the first dose of vaccine at 0-5 d (neonatal schedule) or 8 weeks (infant schedule). Breast feeding was with-held for 30 minutes before and after vaccine administration. The relationship between rotavirus specific IgG and SNA levels in cord blood and IgA in colostrum and breast milk at the time of first active dose of RV3-BB vaccine and level of IgA response and stool excretion after 3 doses of vaccine was assessed using linear and logistic regression. RESULTS: Forty infants received 3 doses of RV3-BB rotavirus vaccine and were included in the analysis of the neonatal and infant groups. Rotavirus specific IgA in colostrum (neonatal schedule group) and breast milk at 4 weeks (infant schedule group) was identified in 14/21 (67%) and 14/17 (82%) of infants respectively. There was little evidence of an association between IgA in colostrum or breast milk IgA at 4 weeks, or between cord IgG or SNA level, and IgA response or stool excretion after 3 doses of RV3-BB, or after one dose (neonatal schedule) (all p>0.05). CONCLUSIONS: The level of IgA in colostrum or breast milk and level of placental IgG and SNA did not impact on the serum IgA response or stool excretion following 3 doses of RV3-BB Rotavirus Vaccine administered using either a neonatal or infant schedule in New Zealand infants.

Development of recombinant vaccine candidate molecule against Shigella infection.

Shigellosis is an acute bacillary diarrheal disease caused by the gram negative bacillus Shigella. The existence of multiple Shigella serotypes and their growing resistance to antibiotics stress the urgent need for the development of vaccine that is protective across all serotypes. Shigella's IpaB antigen is involved in translocon pore formation, promotes bacterial invasion and induces apoptosis in macrophages. S. Typhi GroEL (Hsp 60) is the immunodominant antigen inducing both arms of immunity and has been explored as adjuvant in this study. The present study evaluates the immunogenicity and protective efficacy of recombinant IpaB domain-GroEL fusion protein in mice against lethal Shigella infection. The IpaB domain and GroEL genes were fused using overlap extension PCR and cloned in pRSETA expression vector. Fused gene was expressed in Escherichia coli BL-21 cells and the resulting 90 KDa fusion protein was purified by affinity chromatography. Intranasal (i.n.) immunization of mice with fusion protein increased the IgG and IgA antibody titers as compared to the group immunized with IpaB and GroEL and control PBS immunized group. Also IgG1 and IgG2a antibodies induced in fusion protein immunized mice were higher than co-immunized group. Significant increase in lymphocyte proliferation and cytokine levels (IFN-γ, IL-4 and IL-10), indicates induction of both Th1 and Th2 immune responses in both immunized groups. Immunization with fusion protein protected 90-95% of mice whereas 80-85% survivability was observed in co-immunized group against lethal challenge with S. flexneri, S. boydii and S. sonnei. Passive immunization conferred 60-70% protection in mice against all these Shigella species. Organ burden and histopathology studies also revealed significant decrease in lung infection as compared to the co-immunized group. Since IpaB is the conserved dominant molecule in all Shigella species, this study will lead to an ideal platform for the development of safe, efficacious and cost-effective recombinant vaccine against Shigella serotypes.

'Pre-endoscopy point of care test (Simtomax- IgA/IgG-Deamidated Gliadin Peptide) for coeliac disease in iron deficiency anaemia: diagnostic accuracy and a cost saving economic model'.

BACKGROUND: International guidelines recommend coeliac serology in iron deficiency anaemia, and duodenal biopsy for those tested positive to detect coeliac disease. However, pre-endoscopy serology is often unavailable, thus committing endoscopists to take routine duodenal biopsies. Some endoscopists consider duodenal biopsy mandatory in anaemia to exclude other pathologies. We hypothesise that using a point of care test at endoscopy could fill this gap, by providing rapid results to target anaemic patients who require biopsies, and save costs by biopsy avoidance. We therefore assessed three key aspects to this hypothesis: 1) the availability of pre-endoscopy serology in anaemia; 2) the sensitivities and cost effectiveness of pre-endoscopy coeliac screening with Simtomax in anaemia; 3) whether other anaemia-related pathologies could be missed by this targeted-biopsy approach. METHODS: Group 1: pre-endoscopy serology availability was retrospectively analysed in a multicentre cohort of 934 anaemic patients at 4 UK hospitals. Group 2: the sensitivities of Simtomax, endomysial and tissue-transglutaminase antibodies were compared in 133 prospectively recruited patients with iron deficiency anaemia attending for a gastroscopy. The sensitivities were measured against duodenal histology as the reference standard in all patients. The cost effectiveness of Simtomax was calculated based on the number of biopsies that could have been avoided compared to an all-biopsy approach. Group 3: the duodenal histology of 153 patients presenting to a separate iron deficiency anaemia clinic were retrospectively reviewed. RESULTS: In group 1, serology was available in 361 (33.8 %) patients. In group 2, the sensitivity and negative predictive value (NPV) were 100 % and 100 % for Simtomax, 96.2 % and 98.9 % for IgA-TTG, and 84.6 % and 96.4 % for EMA respectively. In group 3, the duodenal histology found no causes for anaemia other than coeliac disease. CONCLUSION: Simtomax had excellent diagnostic accuracy in iron deficiency anaemia and was comparable to conventional serology. Duodenal biopsy did not identify any causes other than coeliac disease for iron deficiency anaemia, suggesting that biopsy avoidance in Simtomax negative anaemic patients is unlikely to miss other anaemia-related pathologies. Due to its 100 % NPV, Simtomax could reduce unnecessary biopsies by 66 % if only those with a positive Simtomax were biopsied, potentially saving £3690/100 gastroscopies. TRIAL REGISTRATION: The group 2 study was retrospectively registered with clinicaltrials.gov. Trial registration date: 13(th) July 2016; TRIAL REGISTRATION NUMBER: NCT02834429 .

Towards Universal Screening for Toxoplasmosis: Rapid, Cost-Effective, and Simultaneous Detection of Anti-Toxoplasma IgG, IgM, and IgA Antibodies by Use of Very Small Serum Volumes.

Rapid, cost-effective, and early determination of the serological status of potentially infected individuals, particularly pregnant women, can be critical in preventing life-threatening infections and subsequent fetal congenital abnormalities. An article in this issue of the Journal of Clinical Microbiology (X. Li, C. Pomares, G. Gonfrier, B. Koh, S. Zhu, M. Gong, J. G. Montoya, and H. Dai, J Clin Microbiol 54:1726-1733, 2016, http://dx.doi.org/10.1128/JCM.03371-15) describes an innovative multiplexed immunoassay that offers a path toward universal screening.