Quantification of classical swine fever virus load by one-step TaqMan real-time RT-PCR assay

315-322A fluorogenic-probe hydrolysis (TaqMan) real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay based on amplification of a 93 bp fragment from 5’un-translated region (5’UTR) of classical swine fever virus was used for detection and absolute quantification of the virus in clinical and tissue samples. For determining analytical sensitivity, an in vitro transcript RNA containing 5’UTR of classical swine fever virus (CSFV) strain Alfort187 from plasmid pCRXLV324-6 was used as a positive control and a standard for quantification of CSFV genomic RNA copies. The real-time quantitative RT-PCR (qRT-PCR) assay was used to assess the CSFV shedding from naturally infected pigs in whole blood, nasal swab and also in tissue samples. Used qRT-PCR was specific and sensitive as it could detect as low as 16.3 copies of CSFV genomic RNA. The assay was also reproducible as shown by satisfactory low intra-assay (0.80 % to 1.87 %) and inter-assay (1.00% to 3.80%) coefficient of variation with an efficiency of 102.3% and R2 of 0.993. Thus, the real-time qRT-PCR assay described here allows rapid, specific and sensitive laboratory detection and quantification of CSFV genomic RNA copies

Unravelling key genes associated with ovine Brucellosis by differential gene expression analysis: A holistic bioinformatics study

Ovine Brucellosis, caused by Brucella ovis bacteria, is a pathognomonic reproductive infectious disease of sheep that causes epididymitis in rams (male sheep) and placental inflammation in ewes (female sheep) leading to reduced fertility. The specific molecular process that causes alterations in genome of sheep during brucellosis is not yet fully understood. This study aimed to identify key host genes associated with the pathogenesis of ovine brucellosis caused by B. ovis. The GSE35614 dataset containing six healthy and six Brucella ovis infected sample of rams in the chronic phase 2 was obtained from the NCBI GEO database to examine and detect any differences in gene expression (DEGs). Functional and pathway enrichment analyse of the DEGs were performed along with the construction of protein-protein interaction network. Next, functional modules and hub genes were clustered and identified respectively, using the MCODE plugin. As a result, a total of 316 differentially expresses genes were filtered according to the provided cut-off criteria. The enriched DEGs were related to extracellular matrix interaction, cell adhesion mediated by integrin, angiogenesis, and inflammatory response. Furthermore, the hub gene analysis resulted in five hub genes namely, FN1, FBN1, CDH1, CD44, and SPP1, were up-regulated during the infection which could lead to reproductive disorders in sheep. In conclusion, the DEGs, functional and pathways terms, along with hub genes identified in the current study can provide prospective targets for the early diagnosis and treatment of brucellosis and provide insight into the molecular mechanism underlying the alterations that occur during brucellosis in sheep

MOLECULAR DETECTION, ISOLATION, AND PATHOLOGY OF BOVINE TUBERCULOSIS IN AN ORGANIZED FARM IN ASSAM, INDIA

Bovine tuberculosis (bTB) is a well-known zoonotic disease that affects cattle all over the world and results in significant economic loss, particularly in impoverished nations. The present communication describes the pathology, isolation, and molecular detection of Mycobacterium in an organized farm in Assam which has previous records on animals with confirmed M. bovis infection. During the period 2020-2021, a total of 40 animals (4 males and 36 females) of one year and above were included in the present study for screening of bovine tuberculosis by single intradermal comparative tuberculin test (SICCT). The milk, nasal swabs were collected from only tuberculin positive cattle and the tissue samples from necropsied animal and then processed for bacteriology, histopathology, and molecular detection from direct samples. Out of 40, four cows showed positive reactors by SICCT, and out of these four, one animal died. At necropsy, there was the presence of circumscribed yellowish-white lesions of various sizes and numbers. The smear prepared from granulomatous tissue samples showed the presence of acid-fast bacilli by Ziehl–Neelsen stain. Mycobacterium could be isolated from tissue samples. The DNA extracted from the samples could amplify Mycobacteria genus specific hsp65 gene and MTBC specific a 123-bp segment of the insertion sequence IS6110

Uklanjanje enterotoksigenih sojeva bakterije Escherichia coli u pokusno inficirane prasadi pasivnom imunizacijom goveđim kolostrumskim protutijelima.

When piglets fail to receive colostrum from the sow, as an alternative means passive immunization can be carried out with antibodies derived from non-maternal sources. In this study, pregnant cows were vaccinated intramuscularly and intramammarily with killed ETEC vaccine and developed a high level of colostral antibodies. These antibodies were specifically of IgG and IgA. Day-old piglets fed with colostrum containing a high titre of IgG survived the challenge infection. However, 75% piglets fed with colostrum from unvaccinated cows, and 100% piglets receiving no colostrum, died from challenge infection.Ako prasad nije posisala kolostrum od krmače može se pasivno imunizirati kolostrumskim protutijelima podrijetlom od krava. Steone krave bile su vakcinirane intramuskularno i intramamarno inaktiviranim enterotoksigenim sojem bakterije E. coli. U njihovu kolostrumu dokazana je visoka razina specifičnih protutijela razreda IgG i IgA. Jednodnevna prasad koja je dobila kolostrum s visokim titrom protutijela IgG preživjela je izazivačku infekciju, dok je 75% prasadi koja je dobila kolostrum od nevakciniranih krava uginulo od izazivačke infekcije. Uginula je i sva prasad koja nije dobila kolostrum

Bronchoalveolar lavage is an ideal tool in evaluation of local immune response of pigs vaccinated with Pasteurella multocida bacterin vaccine

Aim: The aim was to study the bronchoalveolar lavage (BAL) technique in evaluating the local immune response of pig immunized with Pasteurella multocida bacterin vaccine. Materials and Methods: Weaned piglets were immunized with formalin-inactivated P52 strain of P. multocida bacterin and evaluated for pulmonary immune response in BAL fluid. BAL was performed before vaccination and at different post vaccination days. The BAL fluid was assayed using enzyme-linked immunosorbent assay to study the development of P. multocida specific antibody isotypes and also evaluated for different cell populations using standard protocol. Results: The average recovery percentage of BAL fluid varies from 58.33 to 61.33 in vaccinated and control group of piglets. The BAL fluid of vaccinated pigs showed increase in antibody titer up to 60th days post vaccination (8.98±0.33), IgG being the predominant isotype reached maximum titer of 6.12±0.20 on 45th days post vaccination, followed by IgM and a meager concentration of IgA could be detected. An increased concentration of the lymphocyte population and induction of plasma cells was detected in the BAL fluid of vaccinated pigs. Conclusion: Though intranasal vaccination with P. multocida plain bacterin vaccine could not provoke a strong immune response, but is promising as lymphocyte population was increased and plasma cells were detected. BAL can be performed repeatedly up to 3/4 months of age in pigs to study pulmonary immune response without affecting their health

Seroprevalence of contagious ecthyma in goats of Assam: An analysis by indirect enzyme-linked immunosorbent assay

Aim: The objective of this study was to screen the prevalence of contagious ecthyma (CE) among the goat population of Assam owing to its high prevalence rate. Materials and Methods: In this study, a total of 231 serum samples were collected from 12 districts of Assam during September 2013 to July 2014. The serum samples were tested for the presence of antibodies against Orf virus (ORFV) by indirect enzyme-linked immunosorbent assay (ELISA). Indirect ELISA was standardized using purified Orf reference virus produced in bulk in primary lamb testes cells. Results: Studies on seroprevalence showed 76.62% of goats were seropositive. The total number of animals were divided into different age groups starting from 0-2 months, 2-4 months, 4-6 months, and above 8 months and accordingly highest prevalence of antibodies against ORFV was recorded in the age-group above 8 months of age. Significantly, lower rates of infection were observed in goats of age group 2-4 months. This study recorded that seropositivity from naturally infected animals and in contact apparently healthy animals to be 53.67% and 46.32%, respectively. Conclusion: The results indicated that CE is a prevalent infection in goats of Assam, and the healthy population is at increased risk of infection

Detection of foot-and-mouth disease virus type O in recovered as well as healthy cattle to study carrier status in Assam

Foot and mouth disease (FMD), one of the most contagious diseases of animals, affects different host species including wild animals. Asymptomatic FMD recovered animals may remain as carrier, which may be threat to other healthy animals. Hence, it is necessary to monitor the carrier status of the FMD recovered animals to effectively prevent further spread of the disease. Out of all the seven serotypes of FMD, O serotype is most commonly found in livestock. Therefore, in the present study, we chose to detect serotype ‘O’ in oropharyngeal fluid (OP) and to quantify cytokines, viz. IL-1α, IL-1β and IL-2. A total of 30 OP fluids and 30 blood samples were collected from 10 animals (1 in-contact healthy animal) for 3 months post infection. FMD O serotype could be detected in all the animals (100%). The RQ values were found to be 0.014 to 63.118 and 0.162 to 46.889 for IL-1α and IL-1β genes respectively, while insignificant RQ values were obtained for IL-2. In the second and third months, two animals showed down regulation for IL-1α gene, while IL-1β and IL-2 genes were down regulated in 7 animals and in all 10 animals, respectively for all the three months

Listeriosis in a peri-urban area: Cultural and molecular characterization of Listeria monocytogenes isolated from encephalitic goats

Background and Aim: Listeriosis in food animals bears a significant threat to human health. Detailed investigations into the cause facilitate proper management of the disease. This study reports the cultural, pathological, and molecular characterization of Listeria monocytogenes isolated from encephalitic goats from peri-urban Guwahati, Assam. Materials and Methods: Out of nine suspected samples, five positive isolates of L. monocytogenes were subjected to bacteriological, biochemical, and molecular tests. The genus and species-specific L. monocytogenes 16S rRNA and prs genes were amplified by polymerase chain reaction (PCR) to yield 1200 and 370 bp sized products, respectively. The encephalitic form of the disease was characterized by circling movement, high fever, and terminal recumbence. Results: All the five isolates were confirmed to be L. monocytogenes based on PCR amplification of genus and species-specific 16S rRNA and prs gene products. The isolates were sensitive to ciprofloxacin, oxytetracycline (OTC), and norfloxacin, but resistant to doxycycline and erythromycin. A high dose of OTC was used in a goat at the early stage of clinical symptom and the animal recovered clinically. Conclusion: Listeriosis in goats could pose a significant public health threat as the meat (occasionally milk) or meat products from goats are widely consumed by the people of Assam. Understanding the molecular epidemiological aspects of L. monocytogenes infections of food animal species should, therefore, be the priority in this part of the country

Decoding bovine coronavirus immune targets: an epitope informatics approach

Abstract Bovine coronavirus (BCoV) poses a significant threat to the global cattle industry, causing both respiratory and gastrointestinal infections in cattle populations. This necessitates the development of efficacious vaccines. While several inactivated and live BCoV vaccines exist, they are predominantly limited to calves. The immunization of adult cattle is imperative for BCoV infection control, as it curtails viral transmission to calves and ameliorates the impact of enteric and respiratory ailments across all age groups within the herd. This study presents an in silico methodology for devising a multiepitope vaccine targeting BCoV. The spike glycoprotein (S) and nucleocapsid (N) proteins, which are integral elements of the BCoV structure, play pivotal roles in the viral infection cycle and immune response. We constructed a remarkably effective multiepitope vaccine candidate specifically designed to combat the BCoV population. Using immunoinformatics technology, B-cell and T-cell epitopes were predicted and linked together using linkers and adjuvants to efficiently trigger both cellular and humoral immune responses in cattle. The in silico construct was characterized, and assessment of its physicochemical properties revealed the formation of a stable vaccine construct. After 3D modeling of the vaccine construct, molecular docking revealed a stable interaction with the bovine receptor bTLR4. Moreover, the viability of the vaccine’s high expression and simple purification was demonstrated by codon optimization and in silico cloning expression into the pET28a (+) vector. By applying immunoinformatics approaches, researchers aim to better understand the immune response to bovine coronavirus, discover potential targets for intervention, and facilitate the development of diagnostic tools and vaccines to mitigate the impact of this virus on cattle health and the livestock industry. We anticipate that the design will be useful as a preventive treatment for BCoV sickness in cattle, opening the door for further laboratory studies

Not Available

Not AvailableBackground and aim: Torque teno viruses (TTVs) are circular, single-stranded DNA viruses, which infect a wide range of animals including livestock and companion animals. Swine TTVs (torque teno sus viruses [TTSuVs]) are thought to act as a primary or coinfecting pathogen in pathological conditions such as porcine dermatitis and nephropathy syndrome and post-weaning multisystemic wasting syndrome. So far, the presence of the virus has not been reported in India. Considering that TTSuVs have the potential to cross the species barrier into humans and that pork consumption is common in North-Eastern states of India, the current study aims to investigate the presence of TTSuV in the Indian pig population. Materials and methods: A total of 416 samples were collected during 2014-2018, from both apparently healthy pigs and also from pigs suspected of having died from classical swine fever and/or porcine reproductive and respiratory syndrome. These samples were screened for TTSuV infection by polymerase chain reaction (PCR) and DNA sequencing techniques. Results: The presence of the virus was confirmed in 110 samples from 12 different states of India. Phylogenetic analysis of the nucleotide sequences obtained from the PCR products indicated the presence of viruses of both Iotatorquevirus and Kappatorquevirus genera in India. Conclusion: The study is the first report on the presence of TTSuVs in India and highlights the circulation of both genera of the virus in the country.Not Availabl