Unraveling swine hepatitis E in the central region of Argentina through ELISA development and epidemiological insights.

Hepatitis E virus (HEV) is a public health concern globally, causing acute viral hepatitis in humans. Genotype-3 HEV (HEV-3), the most frequently genotype detected in South America, is zoonotic and the main reservoirs are the domestic pig and wild boar. Circulation of HEV-3 in Argentina has been confirmed in humans as well as in pig herds, wild boar and environmental waters. However, data are scarce mainly due to the inaccessibility of serological assays in this country. In order to provide insights in the epidemiology of HEV in swine in Argentina, we developed an indirect ELISA based on the native recombinant protein ORF2 and conducted a serological survey to determine the prevalence of seropositive swine in small-scale pig farms in the central region of Argentina. The method was evaluated in a panel of 157 serum samples, resulting in relative sensitivity of 98.6 % (95 % CI 95 %-100 %) and relative specificity of 97.7 % (95 % CI 94 %-100 %) compared to a commercial test. An almost perfect agreement was obtained between the two tests (Kappa index of 0.961). A survey on 294 samples from 49 small-scale farms resulted in a seropositivity rate of 54 %. Seropositive animals were found in 34 out of 49 (69.4 %) farms. Most of the farms (70.6 %) had over 50 % of seropositive animals. The wide spreading of HEV in the swine population of Tandil, Argentina, underscore the need to better understand the epidemiology of HEV in the region, enabling the implementation of targeted interventions to mitigate the impact of this virus on public health.

Exploring hepatitis E virus seroprevalence and associated risk factors among the human population in Tandil, Buenos Aires, Argentina.

Background: Hepatitis E virus (HEV) infection is a common cause of acute clinical hepatitis worldwide and is emerging as a disease in Argentina. It is primarily transmitted through contaminated water and food, following the fecal-oral route. Furthermore, is a zoonotic disease with swine as the primary reservoir. Prevalence of HEV infection in humans in several regions of Argentina remains unknown. Objectives: (i) Determine the seroprevalence of HEV among the human population in Tandil, Buenos Aires, Argentina; (ii) Evaluate its association with demographic, socioeconomic and other risk exposures variables, and (iii) Describe and analyze spatial patterns related to HEV infection. Methods: From August 2020 to July 2021, serum samples were collected from 969 individuals aged 1-80 years. Seroprevalence and 95% Confidence Interval was determined. To assess the factors associated with the presence of anti-HEV antibodies, associations between the variables and seropositivity were evaluated through bivariate and multivariate analysis. Spatial scanning for clusters of positivity was carried out. Factors associated with these clusters were also assessed. Results: Anti-HEV antibodies were detected in 4.64% (IC 95% 3.27-6.02) of samples. Dark urine was associated with seropositivity (p = 0.02). Seropositivity was linked with the presence of natural water courses near their households (p = 0.02); the age (p = 0.04); and previous travel to Europe (p = 0.04). A spatial cluster of low rates of HEV seropositivity was detected, with greater distance of the households to water courses associated to the cluster, and male sex inversely associated to it. Discussion and conclusion: This study is the first study to investigate the prevalence of HEV in the population from Tandil, Buenos Aires, Argentina. Considering HEV infection in the differential diagnosis in individuals presenting acute hepatitis is highlighted. The incorporation of HEV testing into blood screening policies should be mandatory. Factors related to the infection and spatial patterns of high and low risk were determined, and should be considered when implementing specific preventive measures.

A multispecies competitive nanobody-based ELISA for the detection of antibodies against hepatitis E virus.

The hepatitis E virus (HEV) is an emergent zoonotic virus causing viral hepatitis worldwide. Clinically, hepatitis E is not easily distinguished from other types of acute viral hepatitis. There is a need for HEV diagnostic assays to detect and prevent interspecies transmission among susceptible populations. Nanobodies (Nbs) are expressed recombinantly in different systems, produced with high yields, and have superior physicochemical properties compared with conventional antibodies (Ab). Several Nbs against ORF2, the capsid protein and main antigen, were selected and produced in E. coli. Nb39 and Nb74 specifically recognized HEV ORF2 (genotypes 3 and 4). A competitive ELISA (cELISA) was developed and validated using a reference panel of human (n = 86) and swine sera (n = 116) tested in comparison with a commercial kit. The optimal cutoff values determined by ROC analysis were 69.16% (human) and 58.76% (swine); the sensitivity and specificity were high: 97.4% (95% CI 86.5-99.5%) and 95.8% (95% CI 86.0-98.8%) for human vs. 100% (95% CI 93.5-100%) and 98.3% (95% CI 91.0-99.7%) for swine. Further, the cELISA detected total anti-HEV antibodies in wild boar, deer, and mice. To our knowledge, this is the first report of production of Nbs against HEV-3 ORF2 for diagnostic purposes.

Seroprevalencia y factores de riesgo de brucelosis y leptospirosis en cerdos en comunidades rurales de Argentina

RESUMEN Objetivo. Determinar la seroprevalencia de brucelosis y leptospirosis en cerdos de granjas de pequeña escala en áreas rurales del partido de Tandil, Argentina; analizar los factores de riesgo asociados; describir los serogrupos de Leptospira spp. prevalentes y determinar la distribución espacial de ambas enfermedades. Materiales y métodos. Se obtuvieron 340 muestras de suero. Se utilizaron la prueba de aglutinación en placa, la prueba de Rosa de Bengala y el Test de Polarización de la Fluorescencia para detectar anticuerpos contra Brucella spp. Se aplicó la Prueba de Aglutinación Microscópica para el diagnóstico de leptospirosis. Se utilizaron métodos estadísticos para evaluar los factores asociados a las infecciones. Resultados. Se observaron seroprevalencias de 0 y 22,6% para brucelosis y leptospirosis, respectivamente. Los serogrupos de Leptospira más prevalentes fueron: Canicola, Ballum, Icterohaemorrhagiae y Pomona. Los principales factores de riesgo asociados a la infección por leptospirosis fueron el mayor número de cerdos, la presencia de equinos y la presencia de jabalíes cerca o dentro de las granjas. Se asoció el uso de subproductos de cereal como alimento para los animales y el uso de cajón para el destete. Se detectó un agrupamiento espacial significativo de seropositividad a Leptospira spp. en un área de baja altitud. Conclusiones. La brucelosis probablemente esté controlada, pero Leptospira spp. se encuentra presente en estas producciones. El conocimiento de la seroprevalencia de la brucelosis y la leptospirosis, su distribución espacial y los factores de riesgo asociados puede ser útil para la prevención y el control de enfermedades zoonóticas endémicas en la región.

ABSTRACT Objectives. To determine the seroprevalence of brucellosis and leptospirosis in pigs reared on small-scale farms in rural areas of the district of Tandil, Buenos Aires Province, Argentina; to analyse the associated risk factors; to describe the prevalent Leptospira spp. serogroups; and to determine the spatial distribution of both diseases. Materials and methods. A total of 340 serum samples were obtained. Buffer Plate Agglutination Test, Rose Bengal Test, and Fluorescence Polarisation Assay were used to detect antibodies against Brucella spp. Microscopic Agglutination Test was applied for serological diagnosis of leptospirosis. Statistical methods were used to evaluate the factors associated with the infections. Results. Seroprevalence of 0% and 22.6% was observed for brucellosis and leptospirosis, respectively. The most prevalent Leptospira serogroups identified were Canicola, Ballum, Icterohaemorrhagiae and Pomona. The main risk factors associated with leptospirosis infection were farms with a higher number of pigs and the presence of horses and wild boars near or within the farms. The use of milling by-products of cereal grains as animal feed and the presence of weaning cages were also associated. A relevant spatial cluster of seropositivity to Leptospira spp. was identified in a low-altitude area. Conclusions. Our results suggest that brucellosis is probably controlled, but Leptospira spp. are present in the farming systems under study. Knowledge of brucellosis and leptospirosis seroprevalence, spatial distribution and associated risk factors can be useful for the prevention and control of endemic zoonotic diseases in the region.

Early events following bovine leukaemia virus infection in calves with different alleles of the major histocompatibility complex DRB3 gene.

Cattle maintaining a low proviral load (LPL) status after bovine leukaemia virus (BLV) infection have been recognized as BLV controllers and non-transmitters to uninfected cattle in experimental and natural conditions. LPL has been associated with host genetics, mainly with the BoLA class II DRB3 gene. The aim of this work was to study the kinetics of BLV and the host response in Holstein calves carrying different BoLA-DRB3 alleles. Twenty BLV-free calves were inoculated with infected lymphocytes. Two calves were maintained uninfected as controls. Proviral load, total leukocyte and lymphocyte counts, anti-BLVgp51 titres and BLVp24 expression levels were determined in blood samples at various times post-inoculation. The viral load peaked at 30 days post-inoculation (dpi) in all animals. The viral load decreased steadily from seroconversion (38 dpi) to the end of the study (178 dpi) in calves carrying a resistance-associated allele (*0902), while it was maintained at elevated levels in calves with *1501 or neutral alleles after seroconversion. Leukocyte and lymphocyte counts and BLVp24 expression did not significantly differ between genetic groups. Animals with < 20 proviral copies/30 ng of DNA at 178 dpi or < 200 proviral copies at 88 dpi were classified as LPL, while calves with levels above these limits were considered to have high proviral load (HPL) profiles. All six calves with the *1501 allele progressed to HPL, while LPL was attained by 6/7 (86%) and 2/6 (33%) of the calves with the *0902 and neutral alleles, respectively. One calf with both *0902 and *1501 developed LPL. This is the first report of experimental induction of the LPL profile in cattle.

Association of TNF-α gene promoter region polymorphisms in bovine leukemia virus (BLV)-infected cattle with different proviral loads.

Tumor necrosis factor alpha (TNF-α) is a pleiotropic cytokine involved in the immune response against viral and other infections. Its expression levels are affected by a polymorphism in the promoter region of the gene. Bovine leukemia virus is a retrovirus that infects cattle and develops two different infection profiles in the host. One profile is characterized by a high number of proviral copies integrated into the host genome and a strong immune response against the virus, while the most relevant property of the other profile is that the number of copies integrated into the host genome is almost undetectable and the immune response is very weak. We selected a population of cattle sufficiently large for statistical analysis and classified them according to whether they had a high or low proviral load (HPL or LPL). Polymorphisms in the promoter region were identified by PCR-RFLP. The results indicated that, in the HPL group, the three possible genotypes were normally distributed and that, in the LPL group, there was a significant association between the proviral load and a low frequency of the G/G genotype at position -824.