Veterinary Research Forum

Abstract Pasteurella multocida is a widespread zoonotic pathogen associated with severe hepatic inflammatory responses. Although traditional medicinal plants such as Tribulus terrestris L. (TL) have demonstrated anti-inflammatory properties, their protective effects against Pasteurella multocida-induced liver injury remain insufficiently studied. This study established a Pasteurella multocida-induced liver injury model to evaluate the protective efficacy of TL extract. Thirty-two male Kunming mice were randomly divided into four groups: control, model, penicillin-streptomycin, and TL extract. Therapeutic interventions were conducted by daily intraperitoneal injection of P. multocida (50 CFU/mL) followed by treatment with antibiotics or TL extract (1000 mg/kg/day) for 7 consecutive days. The therapeutic potential of T. terrestris extract was evaluated using histopathological analyses (hematoxylin and eosin (H&E) staining and Masan staining), RT-qPCR, Western blot, immunohistochemistry, and immunofluorescence to assess inflammatory cytokine expression and NF-κB pathway activation. The extract significantly attenuated hepatic fibrosis and inflammatory cell infiltration in infected mice. Following Pasteurella multocida infection, NF-κB pathway activation may indicate by upregulated NF-κB1 and downregulated IκBα expression in mouse liver, alongside increased pro-inflammatory cytokines TNF-α and IL-1β. Treatment with antibiotics or TL reversed these changes. TL likely suppressed NF-κB activation by downregulating receptor TNFRsf1a. Immunohistochemistry and immunofluorescence further demonstrated decreased phosphorylation of IκBα and reduced IL-1β and TNF-α after treatment, indicating effective inhibition of NF-κB signaling and inflammation. These effects were comparable to those observed with antibiotic treatment. Our research reveals that TL extract alleviates Pasteurella multocida-induced hepatic inflammation by inhibiting NF-κB activation, highlighting its potential as a plant-derived therapeutic agent for bacterial liver injury.

Abstract Foot-and-mouth disease virus (FMDV) continues to pose a major threat to the livestock industry worldwide due to its high transmissibility and substantial economic consequences. For diagnostic and vaccinal purposes, necessitate the exploration of novel immunogenic targets is pivotal. Here, antigenic determinant (an epitope) sequence comprising core of 12 amino acids from VP1 protein of foot-and-mouth disease virus (FMDV) was designed by immuno informatics. To improve the physio-chemical and stability characteristics of the final peptide, additional residues were incorporated until it resulted in the synthesis of a 23-amino acid peptide (VARTLPTSFNYGAIKATRVTDEC, 2.5 Kda and PI 8.17). After chemical synthetizing with 95 % purity, the peptide was conjugated to bovine serum albumin (BSA) via EDC/NHS cross linker (Peptide: EDC: Sulfo-NHS: BSA (carrier protein): 10:40:50:1 in MES buffer (50 Mm)) and formulated with an aluminum hydroxide adjuvant. Twelve male New Zealand White rabbits were randomly assigned into three groups and main group received the peptide-BSA conjugate. Sera were direct to Dot blot, double immunodiffusion (DID) and serum neutralization (SN) tests, and also indirect peptide ELISA. The synthetic peptide elicited strong antibody response by primary positive results in dot blot and DID test (positive in 1:4 serum dilution) as well as it showed cross-reactivity with whole antigen. Although only low SN titers (1:8) were observed. Moreover, the peptide ELISA confirmed specific antibody responses against O serotype (OD of 2.4 at day 55 in 1/10 dilutions). Altogether, rationally designed epitope are promising regimes for the future development of FMDV novel diagnostic tools and vaccine cocktails.

Abstract An emerging global threat is the development of resistance to various antibiotic groups among pathogenic bacteria, including P. multocida, The excessive use of antibiotics and presence of antibiotic resistance carriers play a significant role in the development of multiple resistances to antimicrobial agents. Given the role and importance of plasmids in the development of antibiotic resistance, it is important to prevent the transfer, replication, and multiplication of plasmids by eliminating plasmids. In this study, the role of plasmids in the development and transmission of antibiotic resistance in P. multocida was investigated and evaluated. This study was performed on of P. multocida isolates were collected from apparently healthy and ailing animals (February to September 2014). Identification of P. multocida isolates using standard biochemical tests and kmt1 gene amplification. After initial tests and plasmid DNA extraction, five plasmid-containing isolates were selected for further work. To compare antibiotic resistance after plasmid removal, the disk diffusion method was used. Based on the results, it was determined that all plasmid-containing isolates that showed a high level of antibiotic resistance (especially with large plasmids) became sensitive to antimicrobial agents after plasmid removal. Therefore, according to the results obtained, the role and importance of plasmids in the development of antibiotic resistance in P. multocida is determined. It was also determined in this study that the existence and persistence of plasmids by P. multocida bacteria is of great importance in the role of plasmids in the development of antibiotic resistance.

Abstract Pasteurella multocida and Mannheimia haemolytica are significant bacteria of respiratory tract infections in cattle. Identifying whether these bacteria are commensal or pathogenic is crucial for understanding their role in predisposing calves to respiratory tract infections. This study aimed to determine the prevalence and pathogenic characteristics of P. multocida and M. haemolytica in nasal swab samples of clinically healthy calves in Konya, Turkey, and to assess antibiotic resistance profiles. Using conventional culture methods, 5 samples (1.67%) from 4 different farms were positive for P. multocida and 3 samples (1%) were positive for M. haemolytica. PCR of nasal swab samples yielded P. multocida positivity in 17 (5.67%) samples and M. haemolytica positivity in 13 (4.33%), including all culture-positive samples. While all P. multocida isolates were atypical for the capsule, 18% of the PCR-positive samples were L2, 70% L3A, and 12% L8 serotypes. The ompA, omp87, hghA, and tbpA genes were the most prevalent, while the presence of other virulence genes varied. High rates of ampicillin/cloxacillin (100%) and streptomycin (80%) resistance were observed among the isolates. The rates of serotypes A1 and A6 in M. haemolytica PCR-positive samples were 53.85% and 46.15%, respectively. The ssa-1 gene was detected in 33.33% of the isolates. Moderate resistance to trimethoprim/sulfamethoxazole, streptomycin, and neomycin (66.67%) was recorded. The findings revealed the presence of P. multocida and M. haemolytica with various virulence characteristics and antimicrobial resistance profiles in the nasal microbiota of healthy calves, providing novel insights that could inform improved herd management, disease control, and treatment strategies.

Abstract The Accessory Gene Regulator (agr) operon as part of the quorum-sensing system, is responsible for regulating numerous virulence factors in Staphylococci. Furthermore, the ability of the opportunistic pathogen Staphylococcus pseudintermedius to form biofilm is a significant contributor to chronic diseases. Our aim was to evaluate of the association between different agr groups and the expression of the icaD gene related to biofilm formation among S. pseudintermedius isolates from nasal samples. Fifty nasal swabs were collected from broiler chickens and phenotypic and molecular tests were performed to isolate S. pseudintermedius from the samples and investigate their agr genotype. The biofilm production capacity of the isolates was determined using both a microtiter plate assay and amplification of the icaD gene. The S. pseudintermedius was detected in the 86.84% of samples. The assessment of agr typing results revealed that most of the strains (69.70%) were classified in agr group I. The relative frequency of biofilm producing S. pseudintermedius isolates was 75.76% and all of them harbored the icaD gene. The expression level of the icaD determinant was significantly higher among isolates in agr group I than other agr groups. These results suggest that broiler chickens may act as a reservoir for the transmission of S. pseudintermedius to humans which should be considered a potential zoonotic bacterium capable of producing different virulence factors like biofilms.

Abstract Wild birds play a significant role in the dissemination of micro-organisms, including fungal agents, through feathers, skin debris, and feces, both locally and across regions via migration. Identifying fungal carriage in wild birds is crucial for understanding the prevalence of pathogenic fungi and developing effective strategies to prevent and control mycotic infections. This study investigated fungal carriage in wild pigeons, doves, water birds, birds of prey, and small passerines in Balıkesir province of Türkiye, an ecologically important region characterized by key wetlands, two national parks, and major migratory bird routes. A total of 46 wild bird samples were analyzed by culture method, as the “gold standard”, and several clinically important fungal isolates were obtained and subsequently further analyzed by PCR. Fungi were detected across all bird categories, with 27 birds (58.70%) testing positive for at least one fungal species. In total, 10 distinct fungal taxa were isolated from the wild birds. The most frequently identified organisms were Mucor spp. (28.26%) and Aspergillus niger (6.52%), followed by Sporothrix schenckii (4.35%), Candida glabrata (4.30%), Arthroderma terrestre (2.17%), Trichophyton mentagrophytes (2.17%), Nannizzia gypsea (2.17%), Aspergillus flavus (2.17%), Aspergillus fumigatus (2.17%), Lichtheimia spp. (2.17%), and Beauveria spp. (2.17%). Prevalence was the highest in birds of prey and small passerines (77.78%), moderate in pigeons and doves (58.82%), and the lowest in water birds (27.27%). These results emphasize the role of wild birds as potential reservoirs and vectors of fungal agents, highlighting the importance of continued surveillance and biosecurity measures to mitigate the risk of environmental and zoonotic transmissions.

Abstract Non-typhoidal Salmonella is a predominant etiological agent of foodborne infections globally. Salmonella serovars are present throughout the chicken's digestive tract, particularly in the ceca. This study, conducted from October 2024 to April 2025, employed cultural methods to isolate and identify Salmonella from samples collected at the broiler chicken wet market in Sulaymaniyah province, Iraq. The research additionally examined the prevalence of S. enterica serovar Typhimurium in these samples by PCR. Two-hundred and ten samples were obtained from the cecum, cutting boards, knives, body swabs, workers' hands, and water. Salmonella was isolated from the samples and identified by culturing and molecular techniques. Consequently, 103 individuals (49.04%) tested positive for Salmonella via culture. The PCR results revealed that the contamination rate for S. enterica was 74 (71.84%); 47 (63.51%) were S. Typhimurium, and 33 (70.21%) of S. Typhimurium possessed the hilA gene. The highest prevalence was observed in chopping boards, with 91.67% for S. enterica, whereas the maximum rate for S. Typhimurium was recorded in water, at 100%. The phylogenetic tree indicates that the Iraqi isolate with accession number of PV250092 belongs to the principal group of isolates, exhibiting a bootstrap support value of 100%, signifying a robust genetic association with them. To conclude, this research demonstrated a notable prevalence of Salmonella in the broiler chicken processing environment, particularly a high incidence of S. Typhimurium isolates. The placement of the Iraqi isolate within the global isolate clade suggests a recent common ancestor with other isolates, indicating global transmission pathways.

Abstract This study aimed to identify and characterize nitrifying bacteria and their associated bacteriophages from swiftlet house environments as a preliminary step toward developing biological control strategies for nitrate contamination in edible bird nests,a food safety concern due to the excessive nitrite and nitrate accumulation. A total of 80 environmental samples were collected between October and November 2024 from four swiftlet houses in Central Kalimantan, Indonesia, including 20 samples each from feces, pond water, soil, and edible bird nests (Aerodramus fuciphagus). Phenotypic characterization (culture, Gram staining, and biochemical tests) and genotypic identification (PCR and 16S rRNA gene sequencing) were performed. Bacteriophages were screened using spot tests and plaque assays. The study resulted in the isolation and identification of 15 Nitrobacter isolates. Molecular analysis confirmed all isolates belonged to the Nitrobacter genus. One isolate, Nitrobacter AP6, was selected as a representative host for phage screening due to its consistent growth and plaque clarity. Spot tests revealed four bacteriophages capable of lysing Nitrobacter AP6. These findings demonstrate the feasibility of isolating nitrifying bacteria and phages from swiftlet environments and provide a foundation for future biocontrol development, though further in vitro and in situ validation is required to assess effectiveness and safety.

Abstract Porcine parvoviruses (PPVs) are globally recognized as significant contributors to reproductive failure in swine, primarily due to their association with fetal death. Infection in pregnant sows can lead to severe reproductive disorders including stillbirth, mummification, embryonic death and infertility. A recombinase polymerase amplification assay targeting the variable region of the outer capsid protein gene of the PPV-7 genome was developed and systematically optimized under a range of reaction conditions. The assay showed optimal amplification at a constant temperature of 35.00 ˚C for 25 min, using 0.72 µM of each forward and reverse primer and 14.00 mM magnesium acetate. It demonstrated high sensitivity, reliably detecting as few as 2,050 copies of viral nucleic acids in both the conventional and fluorescent dye-based formats. The assay also showed high specificity, exhibiting no cross-reactivity with other common porcine pathogens such as porcine sapelovirus, porcine circovirus and classical swine fever virus. Of the 167 field samples tested, 23 were positive for PPV-7, corresponding to a positivity rate of 13.77%. Operating at a low and constant temperature, the assay eliminates the need for advanced laboratory equipment, making it highly suitable for pen-side application in field settings. In conclusion, this novel assay demonstrated strong potential for field-based detection of PPV-7 circulating within the swine population of Haryana, India, marking the first report of its kind from this region. Further validation using samples from clinically affected herds will strengthen its diagnostic applicability.

Abstract Bovine tuberculosis is a chronic bacterial disease primarily caused by Mycobacterium bovis, a member of the Mycobacterium tuberculosis complex (MTBC), with significant zoonotic implications. This study aimed to detect MTBC in wildlife species, specifically nilgai (Boselaphus tragocamelus) and sambar deer (Rusa unicolor), using gross pathology, histopathology, acid-fast staining, and molecular confirmation. Necropsied tissue samples were collected during post-mortem examination of a nilgai and a sambar deer from the Assam State Zoo, Guwahati, India. Macroscopically, multiple granulomatous tubercles of varying sizes were observed in the lungs and liver, with creamy white caseous material marked upon sectioning. Ziehl-Neelsen staining of the tissue smears from granulomatous lesions confirmed the presence of acid-fast bacilli. Microscopic examination of tuberculosis granulomas revealed a central necrotic mass surrounded by inflammatory cell infiltration, including Langerhans-type giant cells. Molecular confirmation of MTBC infection was achieved by amplifying hsp65 and IS1081 in tissue samples, further validated by Basic Local Alignment Search Tool for nucleotide analysis following Sanger dideoxy sequencing. In conclusion, this study confirmed the presence of tuberculosis in these wildlife species through an integrated approach combining pathology, microbiology, and molecular diagnostics, highlighting the need to understand pathogen entry into the herd and prevent potential spillover.

Abstract Mycobacterium avium subsp. paratuberculosis (MAP), the causative agent of Johne’s disease and a potential contributor to Crohn’s disease, presents a significant challenge due to its resistance to conventional antibiotics. This necessitates the development of innovative strategies for prevention and treatment. This study aimed to evaluate the anti-bacterial activity of pathogen-specific antibodies derived from chicken egg yolks (immunoglobulin Y [IgY]) and the postbiotics from lactic acid bacteria against MAP. Immunoglobulin Y antibodies were produced by immunizing hens with formalin-killed MAP strain antigens. The IgY was extracted and purified, and the anti-MAP titers were quantified by indirect enzyme-linked immunosorbent assay. The minimum inhibitory concentration of different concentrations of specific anti-MAP IgY and the mixture of postbiotics (from four different probiotic strains, including Lactobacillus reuteri, Lactobacillus rhamnosus, Lactobacillus acidophilus, and Pediococcus acidilactici) individually and in combination against MAP was determined at various time intervals. Anti-MAP IgY titers in egg yolks increased within 2 weeks of immunization, reaching peak levels at 6 weeks. Growth inhibition assays revealed that postbiotics concentration as low as 6.25 mg mL^-1 effectively inhibited MAP growth. Anti-MAP IgY demonstrated anti-bacterial activity with a minimum inhibitory concentration of 50.00 mg mL^-1, while the combined IgY-posbiotics treatment achieved MAP growth inhibition at a minimum inhibitory concentration of 3.125 mg mL^-1. The findings of the study suggest that combination therapy with specific IgY and postbiotics may be a promising preventive strategy for controlling MAP infections. Further in vivo studies are needed to elucidate the underlying mechanisms and optimize the application of this approach for broader use in veterinary and human medicine.

Abstract This study aimed to detect Coxiella burnetii, Chlamydia abortus, and Brucella species in the abomasal contents of aborted ruminant fetuses from the Central Anatolia region of Türkiye using PCR between 2020 and 2023. The abomasal contents of a total of 97 aborted fetuses from cattle, sheep, and goats with a history of abortion, collected between the years 2020 and 2023, were tested in this study. As a result of PCR analysis of 97 abomasal contents, four (4.10%; 95.00% confidence interval [CI]: 1.33 - 10.82) of them were C. abortus, including three sheep and one goat. Two (2.10%; 95.00% CI: 0.36 - 7.96) of them were C. burnetii, including one sheep and one cow. A total of 60 (61.90%; 95.00% CI: 51.40 - 71.37) samples from 47 cattle, nine sheep, and four goats were determined by Brucella genus-specific PCR. Following multiplex PCR analysis of the positive Brucella spp. samples, 39 (65.00%; 95.00% CI: 51.52 - 76.55) samples were identified as B. abortus, including two sheep, one goat, and 36 cattle. Additionally, 19 (31.70%; 95.00% CI: 20.60 - 45.09) isolates were identified as Brucella melitensis, including five sheep, two goats, and 12 cattle. In two sheep samples, both B. melitensis and C. abortus were identified from the same animals. In conclusion, Brucella spp. were the predominant abortion-causing pathogens, with C. abortus also contributing significantly. Effective control strategies under the One Health approach are essential to prevent the uncontrolled spread and inter-species transmission of these zoonotic agents in the region and country.

Abstract Biofilm formation is a key virulence factor in Staphylococcus aureus, contributing to bacterial persistence, antimicrobial resistance, and chronic infections. This study aimed to investigate the presence of biofilm-associated genes (fib, fnbA, fnbB, clfA, and clfB) in S. aureus isolates from dogs in Ilam, Iran. From December 2022 to September 2023, 250 swab samples were collected from nasal, oral, and rectal sites of dogs, yielding 81 S. aureus isolates confirmed by PCR amplification of the nuc gene. The prevalence of biofilm-associated genes varied, with clfA, clfB, and fnbA detected in 98.80% of isolates, fib in 63.00%, and fnbB in 16.00%. Notably, fnbA, clfA, and clfB were present in all rectal isolates, while fnbB was absent in this group. The findings highlighted the widespread presence of biofilm-related genes in S. aureus from dogs, suggesting their potential role in colonization and zoonotic transmission. The high prevalence of adhesion-associated genes underscored the need for monitoring biofilm-forming S. aureus in companion animals to mitigate antimicrobial resistance and public health risks.

Abstract Rabies is a viral disease affecting both land-dwelling and flying mammals. In India, dogs have been the primary source of rabies. This study aimed to investigate the role of wild animals in the transmission and persistence of the rabies virus and analyse the viral genome to understand its characteristics. In the present study, a brain sample from a fox (Vulpes vulpes) found dead in the free-ranging area of Mannamangalam forest station, Thrissur, Kerala state, India, was collected for rabies screening by fluorescent antibody staining and molecular techniques. Viral proteins and nucleic acids were detected and the samples were subsequently analyzed using next-generation sequencing for characterization. The genome analysis revealed that the virus belonged to the type I genotype of the arctic-like lineage. The phylogenetic analysis indicated that the fox virus strain shared close homology with all Indian isolates irrespective of the host species and was clustered in the same arctic-like lineage, denoting the distribution of a similar lineage and genotype across all regions of India. The deduced amino acid variations of nucleoprotein and glycoprotein genes also revealed a pattern of similarity amongst isolates of Indian origin and differed from isolates of other geographical locations and lineages. This study based on genome-wide surveillance could offer novel insights into the genetic makeup of currently circulating strains in the human-wildlife conflict and their continuing spread and persistence of rabies.

Abstract Staphylococcus aureus is one of food poisoning bacteria. This study assessed the prevalence, virulence factors, and antimicrobial susceptibility profiles of S. aureus isolated from frozen fish and shrimp in Egypt. Ninety samples from frozen fish fillets and shrimps (45 for each) were examined for S. aureus prevalence using VITEK 2 compact system, followed by molecular confirmation by nuc gene, virulence characterization, and its resistance genes. The overall prevalence rate of S. aureus was 14.44% (13/90). Fish fillet had the highest mean total S. aureus count (9.50 ± 3.50 × 10⁵ colony forming unit g^-1), followed by shrimp (7.50 ± 3.30 × 10⁵ colony-forming unit g^-1), with a non-significant difference among fish fillet and shrimp. All confirmed S. aureus isolates were lecithinase producers and showed β-hemolysis, and coagulase positive, and confirmed molecularly coa gene positive. All isolates were resistant to ampicillin (100%), both erythromycin and clindamycin (69.23%), and tetracycline (61.53%), followed by vancomycin (46.15%). However, all isolates were sensitive to linezolid, tigecycline (84.70%), and trimethoprim/sulfamethoxazole (61.53%). Twelve (92.30%) phenotypically cefoxitin and oxacillin-resistant and molecularly mecA recovered S. aureus isolates were confirmed as methicillin-resistant S. aureus, while based on vancomycin-resistant pheno-typically, and molecularly vanA recovered S. aureus isolates were confirmed as vancomycin-resistant S. aureus. The emergence of multidrug-resistant methicillin-resistant S. aureus and vancomycin-resistant S. aureus in frozen fish fillets and shrimp indicates public health hazards, so there is a need for food safety measures alongside reliable detection methods of resistant bacteria along the food chain.

Abstract Goose parvovirus causes major economic losses in the waterfowl industry due to the high mortality. Therefore, it is essential to establish protection/control objectives in the fight against the disease. The genome of goose parvovirus consists of three structural proteins, including VP1, VP2, and VP3. The VP2 is a candidate antigen in developing vaccines and diagnostic kits. This study aimed to produce the VP2 protein from a local goose parvovirus strain that causes serious infections in geese in Türkiye using the baculovirus expression vector system. To achieve this, the VP2 gene was first amplified by polymerase chain reaction, followed by purification and insertion into the pENTR™/TEV/D-TOPO™ entry vector. Then, the target gene in the pENTR™/TEV/D-TOPO™ vector was transferred to linear N-Term BaculoDirect™ DNA through LR recombination (site-specific recombination between attL and attR sites). The construct was transfected into Spodoptera frugiperda cells. To verify the production of baculoviral virions, a band of approximately 600 bp in length was obtained as a result of polymerase chain reaction amplification using external primer sets for both the VP2 gene and expression vector. The obtained band was purified and sequenced for confirmation. In addition, to confirm the production of the recombinant protein, western blot analysis was conducted utilizing the V5 epitope located at the N-terminus of the expressed protein, resulting in the detection of a ~65.00 kDa band corresponding to the VP2 gene. To detect protein expression in S. frugiperda cells infected with the recombinant baculovirus, immunofluorescence analysis was performed using the same epitope. 

Abstract Rotavirus is one of the major causes of diarrhea in different animal species and has a bad economic impact due to the losses in neonates and productivity. To investigate the occurrence of this infection in bovine calves, three localities in Khartoum State, Sudan, were selected. A total of 200 fecal samples were collected from diarrheic calves; 100 from Khartoum and 50 from each of Khartoum Bahari and Omdurman provinces. Collected samples were screened for a group A rotavirus antigen using enzyme-linked immunosorbent assay (ELISA). Positive results were seen in 40.00% of samples; the highest prevalence of 42.00% was found in samples from Khartoum province. Five ELISA-positive samples were examined under electron microscope, and characteristic wheel-like appearance of rotavirus was visualized. Polyacrylamide gel electrophoresis was also applied on 15 of the positive samples; eight samples showed different polyacrylamide gel electrophoretic group A rotavirus long profile with different patterns. The results showed that the occurrence of rotavirus infection in cattle in Khartoum State is increasing.

Abstract Infectious bronchitis virus (IBV) is an important pathogen in the poultry industry causing avian infectious bronchitis that is, an acute but highly contagious disease affecting the upper respiratory tract, kidneys and reproductive tract. The 3rd passage of a polymerase chain reaction confirmed nephropathogenic IBV isolate was used for this study. Heat stability for 5, 10, 15, 20, and 30 min at 56.00 ˚C, pH sensitivity at pH 3.00, 7.00, 9.00, and 11.00 ultraviolet (UV) irradiation for 10, 15, 20, and 30 min, and chloroform sensitivity were studied. The IBV isolate was found to be susceptible to a temperature of 56.00 ˚C for 5 min and above, UV irradiation within 10 min, chloroform treatment and to pH 11.00 while being resistant to pH 3.00 and 9.00. The second part of the study investigated in vitro effectiveness of the disinfection potential of several commercially used disinfectants in Pakistan against the IBV isolate. For this purpose, Virkon S, Bromosept, and Beloran were employed for the virus inactivation test. Following the IBV challenge for contact time of 1, 5, 10, and 30 min, we counted the number of embryos that died after incubation. Results showed that suitable dilution of disinfectant for the recommended contact period could kill the virus. The maximum susceptibility was seen in the case of Virkon S which killed the virus in just 1 min. Thus, IBV could be killed using commercially available Virkon S, Beloran, and Bromosept after being used in recommended concentrations for recommended contact time.

Abstract Foot-and-mouth disease virus (FMDV) is a highly transmissible pathogen causing severe economic losses in the global livestock sector. Frequent outbreaks of FMDV type-O in Pakistan highlight the need for continuous genomic and antigenic surveillance to track its evolution. This study aimed to isolate and molecularly characterize FMDV type-O from an outbreak in the Punjab province of Pakistan using in vitro cell culture techniques. Samples were processed for viral isolation on a susceptible cell line, followed by RNA extraction. The VP1 gene, pivotal for antigenicity and immunogenicity, was amplified using a one-step polymerase chain reaction protocol. Purified amplicons underwent sequencing, and the nucleotide sequences were translated into amino acid sequences for further analysis. Protein three-dimensional modeling and in silico comparison were performed against the vaccinal seed strain PanAsia-2. The VP1 sequence analysis revealed notable genetic variability among the isolates, indicating adaptive evolution. Structural and antigenic modeling uncovered key differences between the field isolates and vaccinal strain, suggesting potential antigenic drift, which could undermine vaccine performance. The study underscores the dynamic evolution of FMDV type-O in Pakistan and the critical importance of ongoing genomic monitoring to refine vaccine strategies and enhance outbreak control.

Abstract Bovine respiratory disease (BRD) is a significant disease in the cattle industry worldwide. The interaction between environmental factors, hosts, livestock management, and viral and bacterial pathogens causes this disease. Viruses are crucial in the initiation and progression of BRD. This study was the first to investigate the prevalence of BRD viruses using the reverse transcription polymerase chain reaction method in nasal and eye conjunctival swabs and blood samples of 115 BRD calves in the central desert of Iran. At least one investigated virus was detected in 44 animals (38.26%). The detection rates of bovine viral diarrhea virus, bovine coronavirus, bovine adenovirus, bovine respiratory syncytial virus, bovine herpes virus-1, and bovine para influenza virus-3 were 20.00, 14.78, 5.21, 0.86, 0.00, and 0.00%, respectively. Three animals (2.60%) had a simultaneous infection with two viruses. Detection of bovine viral diarrhea virus, bovine coronavirus, and bovine adenovirus was correlated. The virus infection rates were 31.81 and 44.66% in five sampled cities. The virus detection rate in infected animals was related to the nose (26 animals; 50.09%), nose and eyes (seven animals; 15.90%), eyes (seven animals; 15.90%), nose, eyes, and blood (three animals; 6.81%), and blood (one animal; 2.27%) samples. The virus detection rate in different samples was in separate clusters. Monitoring and controlling the circulation of bovine viral diarrhea virus and bovine coronavirus in the central desert of Iran is vital due to the high detection rate. Our results highlight the necessity of investigating other viruses and bacterial agents related to the BRD in the study area.

Abstract Respiratory infections are considered within the major constraints of animal production; viruses are the major causing pathogens. This study aimed to elucidate the prevalence of parainfluenza virus-3 (PIV-3), bovine viral diarrhea virus, and respiratory syncytial virus (RSV) in sheep and goats and the existence of co-infections. A total of 270 sheep and 220 goat pneumonic lung tissues were collected from slaughterhouses in four different areas. Enzyme-linked immunosorbent assay was used to detect the antigen of the three viruses, fluorescent antibody technique and polymerase chain reaction confirmed enzyme-linked immuno-sorbent assay positive results. Prevalence detected for PIV-3 was 11.10% in sheep and 9.50% in goats, pestivirus was 10.40% in sheep and 7.70% in goats, and RSV was 17.80% in sheep and 5.00% in goats. Detected co-infections were 5.60% for PIV-3 and pestivirus in sheep and 4.00% in goats and pestivirus and RSV was observed only in goats (1.40%). Co-infection of the three viruses was detected in only one goat sample (1.00%). The existence of the three viruses in sheep and goats was confirmed. To our knowledge, this is the first report of the co-infections of PIV-3, pestivirus, and RSV in sheep and goats in the studied areas.

Abstract Bovine ephemeral fever (BEF) is a debilitating disease of cattle and water buffaloes. Bovine ephemeral fever viruses (BEFVs) form four phylogenetic lineages including the Middle East, East of Asia, Australia and Africa, while the exotic viral strains have also been detected in different geographic areas. We characterized eight BEFVs from different regions of Iran during a period of seven years from 2015 to 2022. Sequencing the entire length of the G gene, the BEFVs were classified in the Middle Eastern lineage with the maximum of 99.73% and minimum of 97.30% nucleotide identity. The all Iranian and Turkish BEFVs detected during the large epizootic in 2020 were clustered phylogenetically together. However, no amino acid variation was observed between the Iranian viruses detected in 2020 and those identified before 2020 in the Middle Eastern lineage suggesting that host, environmental and other genetic factor (s) might have involved in occurrence of the epizootic in 2020. Two BEFVs detected during 2022 outbreak from Kermanshah and Narmashir in the west and east of Iran, respectively, were clustered in two distinct groups as a novel amino acid substitution H51Y in the epitope G3 was also identified in Kermanshah 2022 sequence. These results imply that the Middle Eastern lineage replaced the previously circulated East Asian BEFVs in Iran during 2012 to 2013 and also signify the emergence of new BEFVs due to the intra-lineage evolution. Continuous monitoring of the circulating viruses and identifying the potential vector (s) and its biology help better understand epizootiology of BEFV in the high-risk region.

Abstract The Q fever is a zoonotic bacterial infection caused by an obligate intra-cellular bacterium, Coxiella burnetii. Members of the Canidae family (Mammalia), including dogs and foxes, are potential reservoirs of C. burnetii, which has a wide host range from mammals and birds to arthropods (primarily ticks). Infected dogs can transmit the disease to other animals and humans. This study aimed to investigate the presence of C. burnetii in dogs and ticks collected from infested dogs in the Kars, Ardahan, and Iğdir provinces of Türkiye by serological and molecular methods. Three hundred canine serum samples were analyzed for phase I and phase II C. burnetii antibodies using indirect enzyme-linked immunosorbent assay. Whole blood samples (n = 300) from the dogs sampled for sera and 184 ticks randomly collected from these dogs were also analyzed for C. burnetii with touch-down polymerase chain reaction. The ticks were classified according to the taxonomic characteristics. In result, 107 tick DNA samples collected from individual females and pooled males were evaluated. The C. burnetii was detected in 3.73% (of the tick samples. However, C. burnetii was not detected in any of the canine blood samples by polymerase chain reaction. Out of the 300 dogs, 18.33% presented antibodies against C. burnetii in their blood serum. When assessed for location, C. burnetii seropositivity was found to be significantly high especially in the Northeastern Anatolia region (18.33%). Study data highlighted the zoonotic risk of ticks, demonstrating that ticks on dogs can carry C. burnetii.

Abstract Bovine subclinical mastitis represents a major cause of severe economic losses in dairy farms. This research aimed to detect the antimicrobial resistance trends of Staphylococcus aureus and to determine the presence of mecA, mphC, lnuA, tetK and tetL antimicrobial resistance genes in raw bulk milk in the period between December 2023 and February 2024. One hundred raw bulk cow milk samples were gathered from different dairy farms in Egypt. The prevalence of subclinical bovine mastitis was 65.00% using California mastitis test. The prevalence of isolated S. aureus was 46.15% via bacterial culturing and all isolates (n = 30) were confirmed via hemolytic activity, catalase and coagulase test, and gram staining followed by polymerase chain reaction targeting nuc1 gene. Antimicrobial sensitivity test was applied on all confirmed S. aureus isolates utilizing the disk diffusion method on Mueller-Hinton agar. The highest resistance was verified for tetracycline at 100% followed by erythromycin and clindamycin at 56.66 and 16.66%, respectively. The highest sensitivity at 100% was verified for amikacin, ampicillin, amoxicillin plus clavulanic acid, ampicillin plus sulbactam, ciprofloxacin, colistin, gentamicin, imipenem, tobramycin, doxycycline and vancomycin. Multidrug resistance was found in 20.00% of the total isolates. Methicillin resistant S. aureus represented by mecA gene was identified in 83.33% of isolates. Macrolides resis­tant S. aureus represented by mphC gene was identified in 16.66% of isolates. Lincosamide resistant S. aureus represented by inuA gene was identified in 66.66% of isolates. Tetracycline resistant S. aureus represented by tetK and tetL genes was detected in 23.33 and 53.33% of isolates, respectively. This study provided antibiotic-resistant S. aureus profiles to dairy farms to avoid treatment failure, adverse effects on animal health and economic impact for the owner of the animal.

Abstract The transmission of Escherichia coli (E. coli) containing virulent genes from animals to humans and the environment poses significant public health challenges. This study aimed to detect the virulence factor of the E. coli heme-utilization gene A (chuA) in E. coli isolated from the feces of apparently healthy horses in the island of Sumbawa, Indonesia. The study utilized 52 fecal samples from a total horse population of 283, calculated using the disease detection formula. Fresh feces were collected immediately after excretion and placed in buffered peptone water for subsequent analysis. The samples were then isolated on eosin methylene blue media and identified using biochemical tests. Identified E. coli strains were further examined for detecting the chuA gene using polymerase chain reaction techniques. The E. coli was successfully isolated and identified in 11 (21.15%) of the 52 collected fecal samples. Polymerase chain reaction analysis detected the chuA gene in 8 (15.38%) E. coli isolates at 279 bp on gel electrophoresis. The close interaction between horses and humans in the island of Sumbawa,  Indonesia, may facilitate the spread of E. coli. Thus, surveillance is needed to employ a One Health approach to monitor E. coli strains encoding the chuA gene and other virulence factors to control their dissemination.