External Quality Assessment (EQA) scheme for serological diagnostic test for SARS-CoV-2 detection in Sicily Region (Italy), in the period 2020-2022.

OBJECTIVES: Since December 2019, worldwide public health has been exposed to a severe acute respiratory syndrome caused by Coronavirus-2. Serological testing is necessary for retrospective assessment of seroprevalence rates, and the determination of vaccine response and duration of immunity. For this reason, it was necessary to introduce a panel of tests able to identify and quantify Covid-19 antibodies. METHODS: As a Regional Reference Centre, the CRQ Laboratory (Regional Laboratory for the Quality Control) developed and conducted an External Quality Assessment (EQA) panel of assays, to evaluate the quality of various methods, that were used by 288 Sicilian laboratories, previously authorized on behalf of the Public Health Service. RESULTS: The performance test was based on pooled samples with different levels of concentration of antibodies. 97 , 98, and 95 % of the participating laboratories tested all samples correctly in 2020, 2021, and 2022 respectively. The best performance was observed in the test of total Ig. The general performance of laboratories improved over the years. CONCLUSIONS: The incorrect diagnosis had and could still have important implications on vaccination cycles. Only through the effort of laboratory professionals, and the extension of the EQA scheme, a better harmonization of methods, protocols, and thus results, to guarantee a better healthcare system, will be possible.

Potential Usefulness of IgA for the Early Detection of SARS-CoV-2 Infection: Comparison With IgM.

Serological testing can be a powerful complementary approach to achieve timely diagnosis of severe acute respiratory coronavirus 2 (SARS-CoV-2) infection, along with nucleic acid detection. Immunoglobulin (Ig) A antibodies are less frequently utilized to detect SARS-CoV-2 infection than IgM and IgG antibodies, even though IgA antibodies play an important role in protective immunity against SARS-CoV-2. This review discusses the differences in kinetics and assay performance between IgA and IgM antibodies and the factors influencing antibody responses. It highlights the potential usefulness of analyzing IgA antibodies for the early detection of SARS-CoV-2 infection. The early appearance of IgA and the high sensitivity of IgA-based immunoassays can aid in diagnosing coronavirus disease 2019. However, because of cross-reactivity, it is important to recognize the only moderate specificity of the early detection of SARS-CoV-2 IgA antibodies against spike antigens. Either the analysis of antibodies targeting the nucleocapsid antigen or a combination of antibodies against the nucleocapsid and spike antigens may strengthen the accuracy of serological evaluation.

[The Identification Idea of Antibodies Against Ku and Other High-Frequency Antigens].

OBJECTIVE: This study was aimed to provide ideas for identifying the antibodies to high-frequency antigens by analyzing a female case of high-frequency antigen antibody (anti-Ku) using serological and sequencing method. METHODS: The methods for identification of blood group, erythrocyte antigen, screening and identification of antibody were used to detect the blood type and antibody in the proband. The proband's serum and reagent screening cells treated with Sulfhydryl reagent were applied to judge the type and characteristics of this antibodies when reacted with the regaent screening cells or proband's serum respectively. Gene sequencing was used to determine the genotype of the proband's blood group. RESULTS: The proband's red blood cells were determined as O type RhD positive, whose serum showed strong positive reaction to antibody-screening cells and antibody identification cells with the same intensity in saline and IAT medium, however, the self-cells showed negative effect. The Direct Antihuman Globulin of proband's red blood cells also showed weak positive reaction, and the other blood types were CcEe, Jk(a+b-), P1-, Le(a-b -), Lu (a-b +), K-, k-, Kp(a-b-). Serum of the proband treated with 2-ME still react with three groups of screening cells in IAT medium. The reaction intensity of proband's serum was also unchanged with the cells modified with papain and bromelain, but showed negative effect when the cells were treated with sulfhydryl agents including DTT and 2-ME. Gene sequencing revealed that the KEL genotype of the patient was KEL*02N.24 . This patient had a rare K0 phenotype. CONCLUSION: The rare Kell-null blood group (also known as K0) were identified by serological and molecular tests in the proband who produced both IgG and IgM type of antibody to high-frequency antigen (anti-Ku). These two methods are of great significance in the identification of this rare blood group as well as the antibody to high frequency antigen.

Corrigendum: The outbreak of seasonal goat's brucellosis in the Swat ecosystem of Khyber Pakhtunkhwa.

No abstract available.

Improved Antibody Detection for Canine Leptospirosis: ELISAs Modified Using Local Leptospiral Serovar Isolates from Asymptomatic Dogs.

Leptospirosis is a zoonotic disease of significant concern for human and animal health, with domestic animals, including dogs, acting as reservoirs for human infection. Serology is widely used for leptospirosis diagnosis, even though the standard microscopic agglutination test (MAT) using a panel of serovars lacks specificity and can lead to detection limitations in certain regions. In this study, we aimed to develop an antibody detection tool for dogs using an indirect enzyme-linked immunosorbent assay (ELISA) with a set of local serovar isolates, including Paidjan, Dadas, and Mini, to enhance the accuracy of leptospirosis surveillance in our region. The specificity and sensitivity of various antigen preparations, namely leptospiral whole-cell protein (WCP), total membrane protein (TMP), and outer membrane protein (OMP), were assessed using sera from infected and non-infected dogs, as well as negative puppy sera. Leptospirosis diagnosis was supported using a genus-specific nested polymerase chain reaction test on all collected sera. Protein preparations were validated using SDS-PAGE and Western blotting analysis. In the results, the standard MAT failed to detect antibodies in any of the dogs confirmed as being infected using PCR and isolation, highlighting its limitations. In contrast, the OMP-based ELISAs using local isolates of Leptospira serovars gave positive results with sera from all infected dogs, and negative results with sera from all dogs from non-endemic areas. IgG titres of infected and unvaccinated dogs from endemically affected areas were significantly higher than those in non-endemic regions. Using the OMP-based IgG/ELISAs with the local serovar Dadas resulted in higher specificity and lower sensitivity than when using the WCP- and TMP-based IgG/ELISAs. Agreement analysis revealed fair and moderate concordance between OMP-based IgG/ELISAs and PCR results, whereas slight and fair agreement was observed between OMP-based ELISAs and the MAT. Overall, the modified OMP-based IgG/ELISAs, utilising relevant local serovar isolates from dogs, demonstrated improved accuracy in detecting leptospirosis in the study area, overcoming the limitations of the MAT. This study highlights the importance of identifying and incorporating these local circulating serovar isolates into serological techniques for leptospirosis diagnosis and surveillance.

Post-therapeutic cure criterion in chronic Chagas disease using Trypanosoma cruzi chimeric proteins.

Chagas disease (CD) is a neglected disease caused by Trypanosoma cruzi Chagas, 1909. Causative treatment can be achieved with two drugs: benznidazole or Nifurtimox. There are some gaps that hinder progress in eradicating the disease. There is no test that can efficiently assess cure control after treatment. Currently, the decline in anti-T. cruzi antibody titres is assessed with conventional serological tests, which can take years. However, the search for new markers of cure must continue to fill this gap. The present study aimed to evaluate the decline in serological titres using chimeric proteins after treatment with benznidazole in chronic patients diagnosed with CD. It was a prospective cross-sectional cohort study between 2000 and 2004 of T. cruzi-positive participants from the Añatuya region (Argentina) treated with benznidazole. Serum samples from ten patients were collected before treatment (day zero) and after the end of treatment (2, 3, 6, 12, 24 and 36 months). For the detection of anti-T. cruzi antibodies, an indirect ELISA was performed using two chimeric recombinant proteins (IBMP-8.1 and IBMP-8.4) as antigens. The changes in reactivity index within the groups before and after treatment were evaluated using the Friedman test. All participants experienced a decrease in serological titres after treatment with benznidazole, especially IBMP-8.1. However, due to the small number of samples and the short follow-up period, it is premature to conclude that this molecule serves as a criterion for sustained cure. Further studies are needed to validate tests based on these or other biomarkers to demonstrate parasitological cure.

Development and evaluation of a test strip for the rapid detection of antibody against equine infectious anemia virus.

Equine infectious anemia (EIA) is a contagious disease of horses caused by the equine infectious anemia virus (EIAV). The clinical signs at the acute phase include intermittent high fever, thrombocytopenia, hemorrhage, edema, and anemia. The clinical signs at chronic and relapsing subclinical levels include emaciation and progressive weakness. Surviving horses become lifelong carriers because of the integration of the viral genome into that of the host, and these horses can produce and transmit the virus to other animals. This increases the difficulty of imposing practical control measures to prevent epidemics of this disease. Serological tests measuring the antibodies in equine sera are considered to be a reliable tool for the long-term monitoring of EIA. However, the standard serological tests for EIV either have low sensitivity (e.g., agar gel immunodiffusion test, AGID) or are time consuming to perform (e.g., ELISA and western blotting). The development of a rapid and simple method for detecting the disease is therefore critical to control the spread of EIA. In this study, we designed and developed a colloidal gold immunochromatographic (GICG) test strip to detect antibodies against EIAV based on the double-antigen sandwich. Both the p26 and gp45 proteins were used as the capture antigens, which may help to improve the positive detection rate of the strip. We found that the sensitivity of the test strip was 8 to 16 times higher than those of two commercially available ELISA tests and 128 to 256 times higher than AGID, but 8 to 16 times lower than that of western blotting. The strip has good specificity and stability. When serum samples from experimental horses immunized with the attenuated EIAV vaccine (n = 31) were tested, the results of the test strip showed 100% coincidence with those from NECVB-cELISA and 70.97% with AGID. When testing clinical serum samples (n = 1014), the test strip surprisingly provided greater sensitivity and a higher number of "true positive" results than other techniques. Therefore, we believe that the GICG test strip has demonstrated great potential in the field trials as a simple and effective tool for the detection of antibodies against EIAV. KEY POINTS: • A colloidal gold immunochromatographic (GICG) fast test strip was developed with good specificity, sensitivity, stability, and repeatability • The test strip can be used in point-of-care testing for the primary screening of EIAV antibodies • Both the p26 and gp45 proteins were used as the capture antigens, giving a high positive detection rate in the testing of experimentally infected animal and field samples.

Immunoglobulin A response to SARS-CoV-2 infection and immunity.

The novel coronavirus disease (COVID-19) and its infamous "Variants" of the etiological agent termed Severe Acute Respiratory Syndrome Corona Virus 2 (SARS-CoV-2) has proven to be a global health concern. The three antibodies, IgA, IgM, and IgG, perform their dedicated role as main workhorses of the host adaptive immune system in virus neutralization. Immunoglobulin-A (IgA), also known as "Mucosal Immunoglobulin", has been under keen interest throughout the viral infection cycle. Its importance lies because IgA is predominant mucosal antibody and SARS family viruses primarily infect the mucosal surfaces of human respiratory tract. Therefore, IgA can be considered a diagnostic and prognostic marker and an active infection biomarker for SARS CoV-2 infection. Along with molecular analyses, serological tests, including IgA detection tests, are gaining ground in application as an early detectable marker and as a minimally invasive detection strategy. In the current review, it was emphasized the role of IgA response in diagnosis, host defense strategies, treatment, and prevention of SARS-CoV-2 infection. The data analysis was performed through almost 100 published peer-reviewed research reports and comprehended the importance of IgA in antiviral immunity against SARS-CoV-2 and other related respiratory viruses. Taken together, it is concluded that secretory IgA- Abs can serve as a promising detection tool for respiratory viral diagnosis and treatment parallel to IgG-based therapeutics and diagnostics. Vaccine candidates that target and trigger mucosal immune response may also be employed in future dimensions of research against other respiratory viruses.

[COVID-19 vaccination at the University of Alicante (Spain): detection of neutralizing antibodies against SARS-CoV-2 by rapid serological test, a cross-sectional study]. / Vacunación contra la COVID-19 en la Universidad de Alicante (España): detección de anticuerpos neutralizantes frente al sars-CoV-2 mediante test serológico rápido, un estudio transversal.

OBJECTIVE: Neutralizing antibodies against SARS-CoV-2 have shown to be an effective tool for the analysis of the immunity generated against COVID-19. Numerous seroprevalence studies carried out in different groups have made it possible to draw a global map of vaccination coverage through the use of rapid lateral flow immunochromatography serological tests for clinical and epidemiological purposes. The objective of our study was to determine the degree of immunity against SARS-CoV-2 associated with the presence of neutralizing antibodies in administrative staff, teachers and students at the University of Alicante by means of a rapid serological test and to learn about their experience with vaccination against COVID-19. METHODS: A cross-sectional epidemiological study was designed, based on the prevalence of antibodies against the S protein (spike) of SARS-CoV-2. A total of 888 people participated. The study was carried out with a single test (July 6 to July 22, 2021). Using logistic regression, adjusted Odds Ratios were calculated according to sex, age, type of vaccine, number of vaccine doses received, complete vaccination schedule, and having had COVID-19. RESULTS: The vaccines received mostly were Vaxzevria® and Comirnaty®, with 73.3% between both, although 67.2% presented a complete regimen. The results of the OJABIO rapid neutralizing antibody test gave a positive result in 61.4% of the sample. There was a high association between the variables COVID-19 infection, two doses of vaccine, Spikevax® or Comirnaty® vaccine, and eighteen/twenty-nine years old group with a positive result on the OJABIO test. A total of 712 subjects answered the parallel survey (80%) on adverse effects and preferences between the different vaccines against COVID-19. CONCLUSIONS: The vaccination status against COVID-19 in the university community after six months of the start of national immunization strategies reflects low coverage despite the excellent willingness to get vaccinated. Neutralizing antibodies (NAb) rapid tests can be useful to guide immunization strategies and decide when to administer new booster doses.

OBJECTIVE: Los anticuerpos neutralizantes frente al SARS-CoV-2 han resultado una herramienta eficaz para el análisis de la inmunidad generada frente a la COVID-19. Numerosos estudios de seroprevalencia realizados en diferentes colectivos han permitido trazar un mapa global sobre la cobertura vacunal mediante el uso de pruebas serológicas rápidas de inmunocromatografía de flujo lateral con fines clínicos y epidemiológicos. El objetivo de nuestro estudio fue determinar el grado de inmunidad frente al SARS-CoV-2 asociado a la presencia de anticuerpos neutralizantes en personal administrativo, docentes y estudiantes de la Universidad de Alicante, mediante un test serológico rápido, así como conocer su experiencia sobre la vacunación frente a la COVID-19. METHODS: Se diseñó un estudio epidemiológico, transversal, basado en la prevalencia de anticuerpos frente a la proteína S (espícula o Spike) del SARS-CoV-2. Participaron un total de 888 personas. El estudio se llevó a cabo con un único test (6 de julio a 22 de julio de 2021). Mediante regresión logística se calcularon Odds Ratios ajustadas según sexo, edad, tipo de vacuna, número de dosis de vacuna recibidas, pauta completa de vacunación y haber padecido la COVID-19. RESULTS: Las vacunas recibidas mayoritariamente fueron Vaxzevria® y Comirnaty®, con un 73,3% entre ambas; el 67,2% presentó pauta completa. Los resultados del test rápido de anticuerpos neutralizantes OJABIO dieron un resultado positivo en el 61,4% de la muestra. La posibilidad de un resultado positivo en el test OJABIO estuvo fuertemente asociada a haber padecido la COVID-19, haber recibido dos dosis, estar vacunado con Spikevax® o Comirnaty® o pertenecer al grupo de dieciocho a veintinueve años. Un total de 712 sujetos respondieron a un cuestionario (80%) paralelo sobre los efectos adversos y las preferencias entre las distintas vacunas contra la COVID-19. CONCLUSIONS: El estado de vacunación frente a la COVID-19 en la comunidad universitaria a los seis meses de la puesta en marcha de las estrategias nacionales de inmunización refleja una baja cobertura asociada, a pesar de la excelente predisposición a vacunarse. Los test rápidos de anticuerpos neutralizantes (AcN) pueden ser de utilidad para orientar las estrategias de inmunización y para decidir el momento de administrar nuevas dosis de refuerzo.

Vacunación contra la COVID-19 en la Universidad de Alicante (España): detección de anticuerpos neutralizantes frente al SARS-CoV-2 mediante test serológico rápido, un estudio transversal / COVID-19 vaccination at the University of Alicante (Spain): detection of neutralizing antibodies against SARS-CoV-2 by rapid serological test, a cross-sectional study

Fundamentos: Los anticuerpos neutralizantes frente al SARS-CoV-2 han resultado una herramienta eficaz para el análisis de la inmunidad generada frente a la COVID-19. Numerosos estudios de seroprevalencia realizados en diferentes colectivos han permitido trazar un mapa global sobre la cobertura vacunal mediante el uso de pruebas serológicas rápidas de inmunocromatografía de flujo lateral con fines clínicos y epidemiológicos. El objetivo de nuestro estudio fue determinar el grado de inmunidad frente al SARS-CoV-2 asociado a la presencia de anticuerpos neutralizantes en personal administrativo, docentes y estudiantes de la Universidad de Alicante, mediante un test serológico rápido, así como conocer su experiencia sobre la vacunación frente a la COVID-19. Métodos: Se diseñó un estudio epidemiológico, transversal, basado en la prevalencia de anticuerpos frente a la proteína S (espícula o Spike) del SARS-CoV-2. Participaron un total de 888 personas. El estudio se llevó a cabo con un único test (6 de julio a 22 de julio de 2021). Mediante regresión logística se calcularon Odds Ratios ajustadas según sexo, edad, tipo de vacuna, número de dosis de vacuna recibidas, pauta completa de vacunación y haber padecido la COVID-19. Resultados: Las vacunas recibidas mayoritariamente fueron Vaxzevria® y Comirnaty ® , con un 73,3% entre ambas; el 67,2% presentó pauta completa. Los resultados del test rápido de anticuerpos neutralizantes OJABIO dieron un resultado positivo en el 61,4% de la muestra. La posibilidad de un resultado positivo en el test OJABIO estuvo fuertemente asociada a haber padecido la COVID-19, haber recibido dos dosis, estar vacunado con Spikevax® o Comirnaty® o pertenecer al grupo de dieciocho a veintinueve años. Un total de 712 sujetos respondieron a un cuestionario (80%) paralelo sobre los efectos adversos y las preferencias entre las distintas vacunas contra la COVID-19...(AU)

Background: Neutralizing antibodies against SARS-CoV-2 have shown to be an effective tool for the analysis of the immunity generated against COVID-19. Numerous seroprevalence studies carried out in different groups have made it possible to draw a global map of vaccination coverage through the use of rapid lateral flow immunochromatography serological tests for clinical and epidemiological purposes. The objective of our study was to determine the degree of immunity against SARS-CoV-2 associated with the presence of neutralizing antibodies in administrative staff, teachers and students at the University of Alicante by means of a rapid serological test and to learn about their experience with vaccination against COVID-19. Methods: A cross-sectional epidemiological study was designed, based on the prevalence of antibodies against the S protein (spike) of SARS-CoV-2. A totalof 888 people participated. The study was carried out with a single test (July 6 to July 22, 2021). Using logistic regression, adjusted Odds Ratios were calculated according to sex, age, type of vaccine, number of vaccine doses received, complete vaccination schedule, and having had COVID-19. Results: The vaccines received mostly were Vaxzevria® and Comirnaty® , with 73.3% between both, although 67.2% presented a complete regimen. The results of the OJABIO rapid neutralizing antibody test gave a positive result in 61.4% of the sample. There was a high association between the variables COVID-19 infection, two doses of vaccine, Spikevax® or Comirnaty® vaccine, and eighteen/twenty-nine years old group with a positive result on the OJABIO test. A total of 712 subjects answered the parallel survey (80%) on adverse effects and preferences between the different vaccines against COVID-19...(AU)

Serological identification of MERS-CoV in camels of Wasit province, Iraq.

Background: Since the first human case of Middle East Respiratory Syndrome (MERS) caused by Coronavirus (MERS-CoV) in 2012, several evidence bases have shown one-humped camels as the main reservoir host, from which infection is transmitted to humans. Aim: Serological investigation of MERS in dromedary camels in Wasit province (Iraq), detection severity of infection, and association to some risk factors. Methods: A total of 455 dromedary camels were selected randomly from two main districts in Wasit province, Iraq, during January and April (2023). Sera of all study camels were examined by enzyme-linked immunosorbent assay (ELISA), and titers of positive study animals were categorized according to their severity. Results: Serological testing yielded 37.58% positive animals for MERS infection. According to the severity of positive ODs (titer), a total of 53.22%, 30.99%, 12.28%, and 3.51% showed mild, moderate, strong, and very strong infections, respectively. Regarding risk factors, significant elevation in seropositivity was seen in camels of >3-6 and >6 years old and reduced in camels of £3 years old with an elevated risk of MERS with increased age. Regionally, seropositivity and relative risk were increased in the camels of Shaykh Sa'd when compared with Al-Numaniyah. Regarding sex, no significant variation was detected between seropositive females and males; however, male camels appeared at higher risk than females. Association between the severity of MERS infection and risk factors revealed that there was a significant increase in mild and moderate infections in female camels of >6 years old; whereas strong and very strong infections were seen in male camels of 33-6 years old. Mild and very strong infections were recorded in Shaykh Sa'd; while moderate and strong infections in Al-Numaniyah. Conclusion: The study indicated a longstanding existence of MERS-CoV in camels of Wasit province; therefore, recent infections or active viral excretion are required for confirmation by molecular approaches.

Efficacy of inactivated and RNA particle vaccines against a North American Clade 2.3.4.4b H5 highly pathogenic avian influenza virus in chickens.

Highly pathogenic avian influenza virus (HPAIV) has caused widespread outbreaks in poultry in the Americas. Because of the duration and extent of these outbreaks, vaccine use may be an additional tool to limit virus spread. Three vaccines were evaluated for efficacy in chickens against a current North American clade 2.3.4.4b H5 HPAIV isolate, A/turkey/Indiana/3703-003/2022 H5N1. The vaccines included: 1) a commercial inactivated reverse genetics (rg) generated H5N1 product with a clade 2.3.4.4c H5 hemagglutinin (HA) (rgH5N1); 2) a commercial alphavirus RNA particle (RP) vaccine with the TK/IN/22 HA; and 3) an in-house inactivated rg produced vaccine with the TK/IN/22 HA and a North American lineage N9 neuraminidase (NA) (SEP-22-N9). Both inactivated vaccines were produced with HA genes that were modified to be low pathogenic and with the remaining genes from the PR8 influenza strain. All vaccines provided 100% protection against mortality and morbidity and all vaccines reduced virus shed by the oropharyngeal and cloacal routes significantly compared to sham vaccinates. However, differences were observed among the vaccines in quantities of virus shed at two- and four-days post challenge (DPC). To determine if infected birds could be identified after vaccination to aid surveillance programs, serum was collected from the RP and SEP-22-N9 vaccine groups at 7, 10, and 14 DPC to detect antibody to the NA and nucleoprotein (NP) of the challenge virus by enzyme linked lectin assay (ELLA) and ELISA. As early as 7DPC ELLA detected antibody in sera from 100% of the chickens in the RP vaccinated group and 70% of the chickens in the SEP-22-N9 vaccinated group. Antibody to the NP was detected by commercial ELISA in more than 50% of the birds in the RP vaccinated group at each time point.

The outbreak of migratory goat's brucellosis in the Swat ecosystem of Khyber Pakhtunkhwa.

Brucellosis is a major threat to public health especially in developing countries including Pakistan. This study reveals the characterisation of Brucella species affecting humans and goats in the Swat region of Khyber Pakhtunkhwa, Pakistan. Blood samples were collected from shepherds and goats and analysed by Rose Bengal precipitation test (RBPT), standard plate agglutination test (SPAT), polymerase chain reaction (PCR) and Sanger sequencing of 16S rRNA gene. The findings of the study indicated 24% (36/150) and 11.3% (17/150) positivity for Brucella abortus and Brucella melitensis, respectively, in human samples. In samples of goats, 26.66% (40/150) were positive for B. abortus and 16.66% (25/150) samples were positive B. melitensis by SPAT. The species-specific PCR confirmed B. abortus in 24% (36/150) of human samples and 26.66% (17/150) of goat samples by targeting the IS711 locus. The remaining seropositive samples were confirmed as B. melitensis using IS711 M species-specific primer. The sequences of the amplified fragments of the 16S rRNA gene were blasted, and phylogenetic analysis revealed that Brucella species circulating in the Swat district were closely related to B. melitensis and B. abortus reported from India, China, Philippines, and the United States (US) showing the existence of the possible epidemiological linkage among the Brucella species. This study concluded that there was a higher prevalence of B. abortus (26.6%) in humans and goats compared to B. melitensis (16.6%). These results revealed that the Brucella species were circulating in both humans and goats in the study areas. The findings of the study concluded that B. abortus and B. melitensis were circulating in goats and shepherds with a higher prevalence of B. abortus than B. melitensis. Furthermore, the Brucella species identified in Swat were phylogenetically related to the Brucella species reported from India, China, Philippines and the US.Contribution: The proposed study covers the scope of the journal. The species of the genus Brucella affect both animals and shepherds. This study investigates the seroprevalence of brucellosis in shepherds and goats in different geographical areas in the Swat district. The phylogenetic analysis of the Brucella spp. identified in Swat showed close relationships to the Brucella species reported in India, China, Philippines and the US, which shows the possible epidemiological linkages between the Brucella spp.

The Antibodies' Response to SARS-CoV-2 Vaccination: 1-Year Follow Up.

The use of vaccines has allowed the containment of coronavirus disease 2019 (COVID-19) at a global level. The present work aims to add data on vaccination by evaluating the level of neutralizing antibodies in individuals who have received a three-vaccination series. For this purpose, we ran a surveillance program directed at measuring the level of IgG Abs against the Receptor Binding Domain (RBD) and surrogate virus neutralizing Ab (sVNT) anti-SARS-CoV-2 in the serum of individuals undergoing vaccination. This study was performed on employees from the University of Rome Tor Vergata and healthcare workers from the University Hospital who received the Vaxzevria vaccine (n = 56) and Comirnaty vaccine (n = 113), respectively. After the second dose, an increase in both RBD and sVNT Ab values was registered. In individuals who received the Comirnaty vaccine, the antibody titer was about one order of magnitude higher after 6 months from the first dose. All participants in this study received the Comirnaty vaccine as the third dose, which boosted the antibody response. Five months after the third dose, nearly one year from the first injection, the antibody level was >1000 BAU/mL (binding antibody units/mL). According to the values reported in the literature conferring protection against SARS-CoV-2 infection, our data indicate that individuals undergoing three vaccine doses present a low risk of infection.

T cell immune response predicts survival in severely ill COVID-19 patients requiring venovenous extracorporeal membrane oxygenation support.

Introduction: There is a critical gap in understanding which SARS-CoV-2 patients would benefit most from venovenous extracorporeal membrane oxygenation (VV-ECMO) support. The potential role of a dysregulated immune response is still unclear in this patient population. Objectives: To assess the potential predictive value of SARS-CoV-2 specific cellular and humoral immune responses for survival in critically ill COVID-19 patients requiring VV-ECMO. Methods: We conducted a prospective single-center observational study of unvaccinated patients requiring VV-ECMO support treated at the intensive care unit of Semmelweis University Heart and Vascular Center between March and December 2021. Peripheral blood samples were collected to measure the humoral and cellular immune statuses of the patients at the VV-ECMO cannulation. Patients were followed until hospital discharge. Results: Overall, 35 COVID-19 patients (63% men, median age 37 years) on VV-ECMO support were included in our study. The time from COVID-19 verification to ECMO support was a median (IQR) of 10 (7-14) days. Of the patients, 9 (26%) were discharged alive and 26 (74%) died during their hospital stay. Immune tests confirmed ongoing SARS-CoV-2 infection in all the patients, showing an increased humoral immune response. SARS-CoV-2-specific cellular immune response was significantly higher among survivors compared to the deceased patients. A higher probability of survival was observed in patients with markers indicating a higher T cell response detected by both QuantiFeron (QF) and flow cytometry (Flow) assays. (Flow S1 CD8+ ≥ 0.15%, Flow S1 CD4+ ≥ 0.02%, QF CD4 ≥ 0.07, QF whole genome ≥ 0.59). In univariate Cox proportional hazard regression analysis BMI, right ventricular (RV) failure, QF whole genome T cell level, and Flow S1 CD8+ T cell level were associated with mortality, and we found that an increased T cell response showed a significant negative association with mortality, independent of BMI and RV failure. Conclusion: Evaluation of SARS-CoV-2 specific T cell response before the cannulation can aid the risk stratification and evaluation of seriously ill COVID-19 patients undergoing VV-ECMO support by predicting survival, potentially changing our clinical practice in the future.

Serologic evolution and follow-up to IgG antibodies of infants born to mothers with gestational COVID.

BACKGROUND: It is known that SARS-CoV-2 antibodies from pregnant women with SARS-CoV-2 infection during pregnancy cross the placenta but the duration and the protective effect of these antibodies in infants is scarce. METHODS: This prospective study included mothers with SARS-COV-2 infection during pregnancy and their infants from April 2020 to March 2021. IgG antibodies to SARS-CoV-2 spike protein were performed on women and infants at birth and at two and six months during follow-up. Anthropometrical measures and physical and neurological examinations and a clinical history of symptoms and COVID-19 diagnosis were collected. Simple linear regression was performed to compare categorical and continuous variables. To compare the mother's and infant's antibody titers evolution, a mixed linear regression model was used. A predictive model of newborn antibody titers at birth has been established by means of simple stepwise linear regression. RESULTS: 51 mother-infant couples were included. 45 (90%) of the mothers and 44 (86.3%) of the newborns had a positive serology al birth. These antibodies were progressively decreasing and were positive in 34 (66.7%) and 7 (13.7%) of infants at 2 and 6 months, respectively. IgG titers of newborns at birth were related to mothers' titers, with a positive moderate correlation (Pearson's correlation coefficient: 0.82, p < 0,001). Fetal/maternal antibodies placental transference rate was 1.3 (IQR: 0.7-2.2). The maternal IgG titers at delivery and the type of maternal infection (acute, recent, or past infection) was significantly related with infants' antibody titers at birth. No other epidemiological or clinical factors were related to antibodies titers. Neurodevelopment, psychomotor development, and growth were normal in 94.2% of infants in the third follow-up visit. No infants had a COVID-19 diagnosis during the follow-up period. CONCLUSIONS: Transplacental transfer of maternal antibodies is high in newborns from mothers with recent or past infection at delivery, but these antibodies decrease after the first months of life. Infant's IgG titers were related to maternal IgG titers at delivery. Further studies are needed to learn about the protective role of maternal antibodies in infants.

Multiplex assay for the simultaneous detection of antibodies against small ruminant lentivirus, Mycobacterium avium subsp. paratuberculosis, and Brucella melitensis in goats.

Background and Aim: Brucellosis, paratuberculosis (PTb), and infections caused by small ruminant lentivirus (SRLV), formerly known as caprine arthritis encephalitis virus (CAEV), adversely affect goat production systems. Nonetheless, commonly used diagnostic tests can only determine one analyte at a time, increasing disease surveillance costs, and limiting their routine use. This study aimed to design and validate a multiplex assay for antibody detection against these three diseases simultaneously. Materials and Methods: Two recombinant proteins from the SRLV (p16 and gp38), the native hapten of Brucella melitensis, and the paratuberculosis-protoplasmic antigen 3 from Mycobacterium avium subsp. paratuberculosis (MAP) were used to devise and assess a multiplex assay. Conditions for the Luminex® multiplex test were established and validated by sensitivity, specificity, repeatability, and reproducibility parameters. Cut-off points for each antigen were also established. Results: The 3-plex assay had high sensitivity (84%) and specificity (95%). The maximum coefficients of variation were 23.8% and 20.5% for negative and positive control samples, respectively. The p16 and gp38 SRLV antigens are 97% and 95%, similar to the CAEV sequence found in GenBank, respectively. Conclusion: The multiplex test can be effectively used for the simultaneous detection of antibodies against SRLV, MAP and B. melitensis in goats.

Temporal pattern of humoral immune response in mild cases of COVID-19.

BACKGROUND: Understanding the humoral response pattern of coronavirus disease 2019 (COVID-19) is one of the essential factors to better characterize the immune memory of patients, which allows understanding the temporality of reinfection, provides answers about the efficacy and durability of protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and consequently helps in global public health and vaccination strategy. Among the patients who became infected with SARS-CoV-2, the majority who did not progress to death were those who developed the mild COVID-19, so understanding the pattern and temporality of the antibody response of these patients is certainly relevant. AIM: To investigate the temporal pattern of humoral response of specific immunoglobulin G (IgG) in mild cases of COVID-19. METHODS: Blood samples from 191 COVID-19 real-time reverse transcriptase-polymerase chain reaction (RT-qPCR)-positive volunteers from the municipality of Toledo/ Paraná/Brazil, underwent two distinct serological tests, enzyme-linked immunosorbent assay, and detection of anti-nucleocapsid IgG. Blood samples and clinicoepidemiological data of the volunteers were collected between November 2020 and February 2021. All assays were performed in duplicate and the manufacturers' recommendations were strictly followed. The data were statistically analyzed using multiple logistic regression; the variables were selected by applying the P < 0.05 criterion. RESULTS: Serological tests to detect specific IgG were performed on serum samples from volunteers who were diagnosed as being positive by RT-qPCR for COVID-19 or had disease onset in the time interval from less than 1 mo to 7 mo. The time periods when the highest number of participants with detectable IgG was observed were 1, 2 and 3 mo. It was observed that 9.42% of participants no longer had detectable IgG antibodies 1 mo only after being infected with SARS-CoV-2 and 1.57% were also IgG negative at less than 1 mo. At 5 mo, 3.14% of volunteers were IgG negative, and at 6 or 7 mo, 1 volunteer (0.52%) had no detectable IgG. During the period between diagnosis by RT-qPCR/symptoms onset and the date of collection for the study, no statistical significance was observed for any association analyzed. Moreover, considering the age category between 31 and 59 years as the exposed group, the P value was 0.11 for the category 31 to 59 years and 0.32 for the category 60 years or older, showing that in both age categories there was no association between the pair of variables analyzed. Regarding chronic disease, the exposure group consisted of the participants without any comorbidity, so the P value of 0.07 for the category of those with at least one chronic disease showed no association between the two variables. CONCLUSION: A temporal pattern of IgG response was not observed, but it is suggested that immunological memory is weak and there is no association between IgG production and age or chronic disease in mild COVID-19.

An ELISA to Detect Antibodies to Bovine Alphaherpesviruses 1 and 5 and Bubaline Alphaherpesvirus 1 in Cattle Sera.

Bovine alphaherpesvirus 1 (subtypes 1.1, 1.2a, and 1.2b), type 5 (subtypes 5a, 5b, and 5c), and bubaline herpesvirus 1 (BuHV-1) induce highly, though not fully cross-reactive serological responses. Most types and subtypes of these viruses circulate particularly in countries of the southern hemisphere, notably Brazil and Argentina. Therefore, the detection of infected animals is important in defining prevention and control strategies, particularly when flocks are destined for international trade. Identification of infected herds is most often achieved by assays that detect antibodies, such as enzyme immunoassays (ELISAs). However, to date, no ELISA has been evaluated in its capacity to detect antibodies to these alphaherpesviruses. Here, an ELISA was developed to detect antibodies to all currently recognized BoAHV-1, BoAHV-5, and BuAHV-1 types/subtypes, and its sensitivity and specificity were determined. Six hundred bovine sera were screened in serum neutralization tests (SN) against the seven viruses. ELISAs prepared with each of the viruses were compared to SN. Subsequently, a combined assay with multiple antigens LISA was prepared by mixing five viral antigens, chosen for their highest sensitivity in the preparative assays. In comparison to SN, the mAgELISA sensitivity was 96.5% with 96.1% specificity (κ = 0.93; PPV = 95.0%; NPV = 97.3%). The findings reveal that the mAgELISA developed here is highly suitable for the detection of antibodies, comparable in sensitivity and specificity to that of SN when performed with all known types and subtypes of bovine and bubaline alphaherpesviruses.

Anticuerpos anti-SARS-CoV-2 en personal de salud de un hospital pediátrico / SARS-CoV-2 antibodies among health care workers from a children's hospital

Introducción. En Argentina, el personal de salud ha sido el primero en vacunarse contra COVID-19, pero todavía existen pocos datos sobre la producción de anticuerpos IgG anti-S. Objetivos. Evaluar IgG específica contra glicoproteína spike del SARS-CoV-2 (IgG anti-S) posvacunación en personal de un hospital pediátrico. Explorar la asociación entre presencia de dichos anticuerpos, edad y antecedente de infección previa. Población y métodos. Estudio transversal que incluyó 193 trabajadores vacunados con los dos componentes de la vacuna Sputnik V. Se pesquisó el título de IgG anti-S y se registraron edad, antecedente de infección previa por SARS-CoV-2 y fecha de la vacunación. Resultados. El 98,6 % de los sujetos generó IgG anti-S. El título fue mayor en quienes habían cursado infección previamente (p <0,001), pero no hubo relación con la edad de los sujetos. Conclusión. Aportamos datos de generación de anticuerpos IgG anti-S posvacunación en personal de salud de un hospital pediátrico y exploramos algunos predictores.

Introduction. In Argentina, health care workers have been the first ones to receive the COVID-19 vaccine, but there are still few data on the production of anti-S IgG antibodies. Objectives. To assess specific IgG against the SARS-CoV-2 spike protein (anti-S IgG) after the vaccination of health care workers from a children's hospital. To explore the association between the presence of these antibodies, age, and history of prior infection. Population and methods. Cross-sectional study in 193 workers who received both doses of the two component Sputnik V vaccine. The anti-S IgG antibody titer was measured and age, history of prior SARS-CoV-2 infection, and date of vaccination were recorded. Results. Anti-S IgG antibodies were produced in 98.6% of the subjects. The titer was higher in those with prior infection (p < 0.001), but no relationship was established with subjects' age. Conclusion. We provide data on post-vaccination production of IgG anti-S antibodies among health care workers from a children's hospital and explore some predictors.