BACKGROUND: Brucellosis is a zoonotic disease caused by a bacterial pathogen belonging to the genus Brucella. It is one of the most frequent bacterial zoonoses globally but unfortunately, it is still considered as a neglected disease in the developing world. Keeping in view, this study was conducted to determine the prevalence and risk determinants of brucellosis in large ruminants of peri-urban and rural areas of district Multan-Pakistan. For this purpose, blood samples (n = 490) were collected from the cattle (n = 245) and buffalo (n = 245) population of the study area and subjected to preliminary screening of brucellosis using local and imported RBPT reagents. All the samples were further analyzed using commercially available multi-specie indirect ELISA kit followed by their confirmation by PCR using genus and species-specific primers. Data obtained from lab analysis and questionnaires were subjected to statistical analysis for Pearson Chi-square, Odds Ratio and Confidence intervals (95%).
RESULTS: The results showed that the maximum seropositivity was recorded with local RBPT reagent (VRI, Pakistan; 12.45%; 95%CI = 9.72-15.65%) followed by RBPT-IDEXX (12.24%; 95%CI = 9.52-15.45%) and RBPT-ID.vet (11.84%; 95%CI = 9.18-14.95%) however statistical difference was non-significant (P = 0.956). The ELISA results showed an overall seroprevalence rate of 11.22% (95%CI = 8.59-14.33%) with comparatively higher rate in cattle (12.65%; 95%CI = 8.82-17.44%) as compared to buffaloes (9.80%; 95%CI = 6.49-14.15%). The PCR analysis confirmed the presence of genus Brucella in all seropositive samples whereas frequency of B. abortus and B. melitensis in seropositive samples was 80% and 20%, respectively. The co-existence of both species was also observed in 5.45% samples. The statistical analysis showed a significant association of bovine brucellosis with herd size, breed, reproductive disorders, mode of insemination, educational status and farmers\' awareness about brucellosis (P < 0.05). Conversely, locality, age, weight, gender, pregnancy status, parity and puberty status had no associations with brucellosis (P > 0.05).
CONCLUSIONS: In conclusion, brucellosis is prevalent in large ruminants of district Multan, Pakistan. It is suggested to devise and implement stringent policies for the effective control and prevention of brucellosis in the region. Further, the current situation also warrants the need to strengthen interdisciplinary coordination among veterinarians and physicians in one health perspective to ensure and strengthen the human and animal health care systems in the region.

Toxoplasma gondii is a pathogen that poses a serious threat to human health and causes significant economic losses to the global livestock industry. The prevalence of toxoplasmosis infection has been reported to be high in humans and animals around the world, but the occurrence of the disease has not yet been reported in water buffaloes in Guangxi Zhuang Autonomous Region, southern China. To understand the overall seroprevalence of T. gondii infection in Guangxi, a total of 1041 water buffalo and 114 cat serum samples were examined using an indirect enzyme-linked immunosorbent assay (I-ELISA). Of the 1041 water buffaloes analyzed, an overall seroprevalence of 52.9% (551/1041) was obtained, with year, season, and city location being significant factors affecting the rate of T. gondii infection in water buffaloes (P < 0.001). The results also revealed a high seroprevalence of 57% (65/114) in cats. Given that buffalo milk and meat products are vital food sources, these findings suggest that toxoplasmosis in water buffaloes may be a public health threat. This study provides the first T. gondii seroprevalence data in Guangxi, which could contribute to the prevention and control of toxoplasmosis in the region.

Bovine brucellosis is endemic in cattle in India, however not much is known on the prevalence of this disease in stray cattle populations of the country. This study aimed to estimate the prevalence and identify risk factors associated with brucellosis in the stray cattle populations reared in cow shelters (gaushalas) of Punjab, India. Blood samples were collected from 587 cattle reared in 23 cow shelters in 23 districts (one per district) of the Punjab and were tested using Rose Bengal plate test (RBPT), standard tube agglutination test (STAT) and Indirect Enzyme Linked Immunosorbent Assay (i-ELISA). Information on the sex and breed of the animal, total cattle population and presence of a separate shed for parturition were collected. An animal was considered exposed to Brucella infection based on a positive RBPT or STAT test and a positive i-ELISA test. Explanatory variables for the animal level disease status outcome variable were sex and breed of the animal and at the shelter level were shelter cattle population size and presence of a separate shed for parturition. Univariable binomial exact logistic regression analysis was conducted to assess the association of each explanatory variable with the binary outcome variable. Sixty-two animals were seropositive on RBPT, with an apparent seroprevalence of 10.56% (95% confidence interval [CI]: 8.33%, 13.31%) and the estimated true seroprevalence of 11.48% (95% CI: 8.9%, 14.64%). Sixty three animals were seropositive using STAT [apparent seroprevalence of 10.73% (95% CI: 8.48%, 13.50%) and the estimated true seroprevalence of 10.69% (95% CI: 8.27%, 13.67%)], and 68 using i-ELISA [an apparent seroprevalence of 11.58% (95% CI: 9.24%, 14.43%) and the estimated true seroprevalence of 13.28% (95% CI: 10.50%, 16.66%)]. Cross bred cattle had a lower risk of being test positive (odds ratio 0.16, p = 0.04) as compared to indigenous cattle. Due to a ban on cow slaughter in the country, roaming stray cattle infected with brucellosis present a permanent risk of introduction of disease to the dairy farms and other vulnerable populations.

Brucellosis is an economically devastating animal disease and has public health concern. Serological methods such as Rose Bengal Plate Test (RBPT), Complement Fixation Test (CFT), and Indirect-Enzyme-Linked Immunosorbent Assay (I-ELISA) have been used to detect brucellosis. However, there is limited comparative evaluation studies and lack of molecular confirmation of the causative agents in the study areas. The study was aimed to compare RBPT, I-ELISA, CFT, and confirmation using Polymerase Chain Reaction (PCR). A total of 2317 sera samples were collected from brucellosis-affected areas of Ethiopia with no vaccination history. All sera were subjected to comparative serological assays. Post-cross tabulation, sensitivity, and specificity were determined using Receiver Operating Characteristics (ROC) curve analysis software. PCR was performed on 54 seropositive samples using genus- and species-specific primers.
Among the 2317 sera tested for comparative serological assays, 189 (8.16%) were positive for RBPT, 191 (8.24%) for I-ELISA, and 48 (2.07%) for CFT. Sensitivity to RBPT was 100% (95%) in shoats and 74% (95%) in cattle. Specificity on RBPT was 98.69% (95%), 99.28% (95%), 100% (95%) in sheep, goats, and cattle, respectively. CFT sensitivity was 4 (95%) in sheep, 9.65 (95%) goats, and 72 (95%) cattle. Specificity on CFT was 100% (95%) for sheep, goats, and cattle. A 223bp Brucella genus-specific and 156bp B. abortus species-specific detected. However, B. melitensis not detected.
In this study, I-ELISA was the most sensitive and specific test. RBPT detected all Brucellosis-infected sheep and goats; nevertheless, it showed false positive in sheep and goats and false negative in cattle. The presence of B. abortus in small and large ruminants was confirmed by PCR. This is the first report of B. abortus detection in small ruminant in Ethiopia. B.abortus detected in non-preferred hosts. The findings suggest further study on molecular epidemiology of Brucella species.

Brucellosis is known as one of the most common zoonotic diseases worldwide affecting both livestock and humans. It causes abortions, reduces milk production, and infertility in infected animals. The disease is routinely diagnosed through three serological techniques, such as rose bengal plate test (RBPT), standard agglutination test (SAT), and indirect enzyme-linked immunosorbent assay (I-ELISA). The aim of this study was to identify and compare the brucellosis seroprevalence among dairy cattle farms through these different serological tests. From 2112 sampled dairy cattle in different parts of Iran, RBPT, SAT, and I-ELISA led to 296 (14.02%), 215 (10.18%), and 297 (14.06%) positive results, respectively. Brucella abortus biovar 3 (62 cases) was identified as the most common cause of brucellosis in tested animals. Our results showed that the specificity and sensitivity of I-ELISA were higher than those obtained by RBPT and SAT. In this study, the overall agreement of RBPT and SAT with I-ELISA reached 95.21% and 94.12% in dairy cattle farms, respectively. Furthermore, Cohen\'s kappa statistical analysis revealed that the best degree of agreement was seen between RBPT and I-ELISA (0.80), followed by RBPT and SAT (0.78) and finally SAT and I-ELISA (0.72), thereby indicating a strong agreement between RBPT and I-ELISA methods and good agreement between SAT and I-ELISA methods. The McNemar analysis also showed that a significant difference exists between positive and negative results determined by SAT and I-ELISA methods (p < 0.0001). However, the positive and negative results determined by I-ELISA and RBPT did not show a significant difference (p = 0.9207). Therefore, I-ELISA was a more specific and sensitive serological test when compared to RBPT and SAT and could remarkably decrease non-specific reaction by improving the serological screening specificity for an accurate brucellosis diagnosis in endemic areas.

Brucellosis is a zoonotic disease with significant economic and public health impacts. The disease has been found in ruminants, including camels, but clinical diagnosis of camel brucellosis is difficult due to the lack of clinical signs. Thus, this study aimed to estimate the sensitivity (Se) and specificity (Sp) of the Buffered Plate Antigen Test (BPAT), Rose Bengal Test (RBT), and indirect ELISA (i-ELISA) for the diagnosis of Brucella infection in dromedary camels imported from Sudan to Egypt. The secondary objective of the study was to calculate the animal-level true prevalence of Brucella infection in imported camels. A cross-sectional study was carried out on 921 apparently healthy camels randomly selected from those imported from Sudan and kept in the quarantine stations in the Shalateen area of the Red Sea Governorate, Egypt, between June 2018 and January 2019. Serum samples were collected and analyzed using BPAT, RBT, and i-ELISA. The posterior estimates [medians and 95% Bayesian probability intervals (95% BPI)] for Se and Sp of the three serological tests were obtained using Bayesian latent class models (BLCMs). The BLCM was fitted with the assumption that the BPAT and RBT tests were conditionally dependent on the true brucellosis status of camels. All tests had comparable and high Se (>86%) and Sp (>98%). The animal-level true prevalence of Brucella infection in imported camels was 8.6% (95% BPI: 6.8 - 10.7). Based on these findings, the three assays could be used for the initial screening of Brucella infection in camels. However, the BPAT and RBT are more suitable for use in camel brucellosis control and eradication program in Egypt because of their low unit cost and fast turnaround time compared to the i-ELISA. In addition, BPAT and RBT could be performed in the field where in-vivo tests are rarely used due to logistic and management constraints.

Neospora caninum is an important obligate intracellular apicomplexan parasite that causes spontaneous abortions in cattle and leads to huge economic losses to the farming industry. Although a high prevalence of N. caninum infection has been reported in Asia, data on the prevalence of water buffaloes in China remain unclear. To understand the seroprevalence of N. caninum infection in water buffaloes and its definitive host dogs in China, a total of 987 water buffalo sera from Guangxi Province were tested using an indirect enzyme-linked immunosorbent assay. We obtained an overall seroprevalence of 50.9% (502/987) for water buffalo samples. And the positive rate was higher in border cities (56.8%, 425/748) than in central cities (32.3%, 77/239). We further tested 240 serum samples from dogs in Guangxi and found an overall prevalence of 57.9% (139/240). The high prevalence of N. caninum infection in both dogs and water buffaloes was first reported in southern China, and these data will surely contribute to the prevention and control of the disease.

Porcine parvovirus (PPV) is widely prevalent in pig farms. PPV is closely related to porcine respiratory disease complex (PRDC) and porcine circovirus disease (PCVD), which seriously threatens the healthy development of the pig industry. Although commercial antibody detection kits are available, they are expensive and unsuitable for large-scale clinical practice. Here, a soluble VP2 protein of PPV is efficiently expressed in the E. coli expression system. The VP2 protein can be self-assembled into virus-like particles (VLPs) in vitro. After multiple steps of chromatography purification, PPV-VLPs with a purity of about 95% were obtained. An indirect, enzyme-linked immunosorbent assay (I-ELISA), comparable to a commercial PPV kit, was developed based on the purified PPV-VLPs and was used to detect 487 clinical pig serum samples. The results showed that the I-ELISA is a simple, cost-effective, and efficient method for the diagnosis of clinical pig serum and plasma samples. In summary, high-purity, tag-free PPV-VLPs were prepared, and the established VLP-based I-ELISA is of great significance for the sero-monitoring of antibodies against PPV.

Toxin-contaminated foods and beverages are a major source of illness, may cause death, and have a significant negative economic impact worldwide. Aflatoxin B1 (AFB1) is a potent toxin that may induce cancer after chronic low-level exposure. This study developed a quantitative recombinant AflR gene antiserum ELISA technique for aflatoxin B1 detection in contaminated food products. Aflatoxin B1 residuals from 36 food samples were analyzed with HPLC and VICAM. DNA was extracted from aflatoxin-contaminated samples and the AflR gene amplified using PCR. PCR products were purified and ligated into the pGEM-T vector. Recombinant plasmids were sequenced and transformed into competent E. coli (BL21). Molecular size and B-cell epitope prediction for the recombinant protein were assessed. The purified protein was used to induce the production of IgG antibodies in rabbits. Serum IgG was purified and labeled with alkaline phosphatase. Finally, indirect-ELISA was used to test the effectiveness of polyclonal antibodies for detection of aflatoxin B1 in food samples.

Foot-and-mouth disease (FMD) is a devastating animal disease. A previously developed multi-epitope protein B4 vaccine of the FMD virus (FMDV) serotype O provides safety advantages over inactivated vaccines and could be used to prevent and control FMD in pigs. Commercial enzyme-linked immunosorbent assay (ELISA) kits for assessing vaccine efficacy are available for the inactivated vaccines but not for the multi-epitope protein vaccine. In this study, multi-epitope protein B4 was expressed in Escherichia coli, and an indirect ELISA (I-ELISA) was developed to detect antibodies against FMDV serotype O in pigs. The specificity and sensitivity were 96.7% and 95.9%, respectively. B4-vaccinated pigs yielded B4 I-ELISA serum values that were positively correlated with clinical protection against challenge with FMDV serotype O. The I-ELISA\'s ability to detect antibodies from animals vaccinated with the inactivated vaccine was also evaluated, and the B4 I-ELISA values were significantly positively correlated with liquid-phase blocking ELISA (LPBE) titers (r = 0.6708, p < 0.0001); thus, the I-ELISA was also suitable for detection of antibodies from swine vaccinated with the inactivated vaccine.