Veterinary Research Forum

Abstract Prostate cancer is a very common cancer in men, affecting approximately 1.40 million men worldwide in 2020. To improve the quality of life and survival of both animals and humans, effective therapeutic approaches have been developed and evaluated using animal models. The rat model of prostate cancer induced by a multi-step protocol that consists of a sequential administration of flutamide, followed by testosterone propionate, then the administration of N-methyl-N-nitrosourea, and finally subcutaneous implantation of tubes filled with crystalline testosterone, is one of the most frequently used for prostate cancer research. However, the lack of standardization in procedures for prostate cancer induction, sample collection, and analysis represents a challenge for researchers. To address this issue, we aim to provide investigators with a detailed, step-by-step guide to implementing a rat model of prostate cancer, based on our extensive experience in this field. First, we briefly review the prostate cancer-induced protocols found in the literature, then we provide a detailed description of the prostate cancer rat model implemented by our team. After, we explore the rats’ prostate monitoring during the experiment protocol through imaging modalities, such as ultrasonography, computed tomography, and magnetic resonance imaging. We also describe animal welfare monitoring based on a table of humane endpoints, as well as data collection, such as biological variables and prostate samples. In sum, this article will ensure the quality of results and enable their comparison among different researchers using this rat model.

Abstract Pasteurella multocida is a Gram-negative bacterium causing economically significant diseases in cattle. This study aimed to determine P. multocida susceptibility to different antibiotics and antibiotic alternatives. In this study, 246 samples (180 nasal swabs and 66 lung tissue specimens) were collected from cattle showing respiratory manifestations in Egypt. Suspected P. multocida colonies following culture were subjected to polymerase chain reaction (PCR) for molecular confirmation of the isolates. A multiplex PCR was employed to identify P. multocida capsular groups. Susceptibility of the isolated P. multocida to different antibiotics and nanoparticles as antibiotic alternatives including silver (AgNPs), chitosan (CNPs) and curcumin (CurNPs) were tested using broth microdilution method. Thirty-two P. multocida isolates were obtained, kmt1 gene was detected in these isolates, and molecular capsular types classification revealed that all isolates were belonged to the capsular type A. Based on broth microdilution method findings, 20 (62.50%) isolates were considered as multi-drug resistant (MDR); the isolates were most sensitive to danofloxacin and kanamycin, whereas they were most resistant to doxycycline and tilmicosin. Antibiotic alternatives showed high anti-microbial activity against tested isolates with minimum inhibitory concentrations ranging from 1.56 - 6.25 μg mL^-1, 156 - 625 μg mL^-1, and 128 - 512 μg mL^-1 for AgNPs, CNPs and CurNPs, respectively. Our finding demonstrated that MDR P. multocida was evident in cattle in Egypt. Although antibiotic alternatives showed promising in vitro anti-microbial effects against MDR isolates, additional studies are required to be actually applicable in veterinary practices.

Abstract Lecithotrophic larvae utilize extensive yolk reserves for early development. In this study, the effect of egg docosahexaenoic acid (DHA):eicosapentaenoic acid (EPA) ratios (i.e., 5.92, 10.08, 11.66, and 14.53) on the emerging larvae foregut development of rainbow trout was examined. Larvae samples were taken from day 22 to 36 post-fertilization. Thin whole body longitudinal sections were prepared and stained by Hematoxylin and Eosin and Alcian blue procedure. The sections were examined regarding epithelial layer thickness, intestinal fold height and mucosal layer thickness along with number of enterocytes and goblet cells. Results indicated that maximum thickness of the epithelium was observed on day 36 post fertilization in larvae hatched from eggs with DHA:EPA ratios of 14.53 and 10.08. The highest and lowest intestinal folds height were also observed in larvae hatched from eggs with DHA:EPA ratios of 10.08 and 14.53, respectively. The mucosal-submucosa layer thickness was the highest in larvae hatched from eggs with DHA:EPA ratio of 10.08. Enterocyte’s count was the highest in larvae obtained from eggs with DHA:EPA ratio of 10.08 on day 36 post-fertilization. The highest and lowest number of goblet cells were enumerated in larvae obtained from eggs with DHA:EPA ratios of 5.53 and 14.53, respectively. In conclusion, our results revealed that feeding rainbow trout broodstock with diet contained highly unsaturated fatty acids (HUFA):polyunsaturated fatty acids (PUFA) ratio of 0.28 could result in the egg with DHA:EPA ratio of 10.08 which in turn yielded larvae with better foregut development parameters compared to those larvae emerged from the eggs with increased DHA :EPA ratio.

Abstract Curcumin has been employed in traditional medicine for over a millennium to treat various ailments, and its global use is now widespread. Chinese medicine relies heavily on curcumin as a primary element and uses it to cure infectious diseases, skin disorders, depression, and stress. It has cardioprotective, neuroprotective, and anti-diabetic properties, as well as pharmacological effects on disorders like type II diabetes, atherosclerosis, and human immunodeficiency virus replication. The anti-cancer activity of curcumin has been studied extensively with notable improvements in gastrointestinal, melanoma, urogenital, breast, and lung malignancies. We investigated the anti-inflammatory effects of curcumin on expression of tumor necrosis factor (TNF)-α, c-Fos, and interleukin (IL)-6 genes in brain and liver tissue owing to the effects of ketamine anesthesia on postnatal rats. The thalamic and hepatic tissues were collected without anesthesia, immediately after anesthesia, and 4 and 12 hr after anesthesia in control and curcumin treated postnatal rats. The results showed that glucose, triglyceride, high- and low-density lipoprotein levels were lowered with curcumin treatment. We also found that ketamine increased c-Fos and inflammatory cytokines like TNF-α and IL-6, all of which contribute to inflammation. Brain and liver immunohistochemistry studies confirmed the real-time polymerase chain reaction findings. Curcumin injections alone may be effective in decreasing ketamine-induced inflammation in both brain and liver tissues.

Abstract The most serious problem in public health is salmonellosis, a common disease in horse. The aim of this study was to investigate the shedding of Salmonella serotypes in healthy Caspian pony. We examined 143 pony's fecal samples collected from the north of Iran belonging to different ages and sexes. Samples were cultured, then identification of isolates were performed by common bacteriological methods and polymerase chain reaction (PCR). The PCR was also used to explore the presence of fimA and salmonella secreted effector L (SseL) genes as virulence factors in the isolates and all were assigned to antibiotic susceptibility test via disc diffusion method. Results showed two fecal samples (1.39%) contaminated with Salmonella and further examination demonstrated the isolates belonging to S. enterica serotype typhimurium. Both serotypes were isolated from female and ˂6 years of age group of ponies and we detected fimA and SseL genes in the isolates. Observing multiple drug resistance and virulence genes in isolates is of utmost importance from both clinical and public health perspectives. It is highly likely that we face instances of salmonellosis in animals or humans that lead to severe infections and fail to respond to treatment in future. This study revealed that the occurrence of Salmonella was low in ponies, however, regarding the presence of virulence factors with multidrug resistant trend in this zoonotic bacterium, establishment of good hygienic measurement to prevent the transmission of bacteria between animal and human is necessary. 

Abstract This study aimed to investigate the effects of zoledronic acid (ZA) and antibacterial CM11 peptide on the osteoinduction and antibacterial properties of bioactive glass (BG). The bioactive glass/gelatin (BG/Gel) composite was synthesized using the sol-gel method. The 2-x minimum inhibitory concentration of the peptide and 4.00 mg mL^-1 of ZA were added to the BG/Gel during fabrication. The BG/Gel composite morphological and structural characteristics and anti-bacterial activities were analyzed using Fourier transform infra-red spectroscopy, scanning electron microscopy and disk diffusion test, respectively. The release of the peptide and ZA from BG/Gel was measured using ultra-violet spectroscopy. After 14 days, the effects of the peptide/ ZA-containing BG/Gel (PZ-BG/Gel) on the growth and differentiation of mesenchymal stem cells were evaluated using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide, calcium and alkaline phosphatase assays, immunocytochemical staining for osteocalcin (OCN) and real-time polymerase chain reaction for OCN, type I collagen, bone morphogenetic protein 2 and Runt-related transcription factor-2 genes. The disk diffusion test showed the anti-microbial activity of the scaffold against multi-drug-resistant isolates of Pseudomonas aeruginosa and Staphylococcus aureus. Analyses showed a significantly higher level of stem cells differentiation into the osteo-genic cells in PZ-BG/Gel scaffold compared to BG/Gel scaffold alone. Accordingly, osteoblast markers were significantly increased in comparison with the control. In conclusion, the osteo-induction and antibacterial properties of BG-based scaffold can be improved using ZA and CM11.

Abstract The present study evaluated the Salmonella isolates obtained from various origins in Iran. Salmonella strains previously recovered and stored in the veterinary microbiology laboratory were serotyped and subjected to antibiotic susceptibility test, detection of the virulence and resistance genes by polymerase chain reaction (PCR), and genotyping by enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR). All Salmonella isolates showed resistance to erythromycin and the most resistance rates were detected for trimethoprim (86.66%), ampicillin (75.00%), and sulfamethoxazole-trimethoprim (63.33%), respectively. In total, 86.33% of the isolates were known as multi-drug resistant and none of the isolates showed resistance to cefepime, nalidixic acid, imipenem, ceftriaxone, and polymyxin B. The virulence genes, invA, sdiA, and hilA besides the tetA resistance gene were identified in all 60 Salmonella strains. The most prevalent resistance genes were respectively tetC (70.00%), sul2 (58.33%), and ereA (55.00%). Statistical analysis revealed a significant difference between Salmonella serotypes associated with the sul1 resistance gene. In ERIC-PCR analysis, 14 distinct clusters were obtained. Statistically, there were significant relationships between the source and ERIC’s genomic pattern and between the serotype of Salmonella isolates and genotypic pattern of ERIC. According to the results, Salmonella serotypes from non-human sources had considerable resistance to different antibiotics and carried significant virulence determinants and resistance genes. In addition, ERIC-PCR showed relevant results in discriminating Salmonella serotypes from other sources.

Abstract Bovine clinical mastitis is an economically important disease in dairy industry worldwide resulting in reduction of milk yield and quality. Among mycotic mastitis, Candida spp. are commonly occurring opportunistic mycosis in immunocompromised animals. The micro-organism’s causing mastitis has high zoonotic potential and has been linked with rapid growth and introduction of antimicrobial resistance between animals and humans. The present study was conducted to isolate and identify the common pathogenic Candida spp. from bovine mastitis cases in India. The isolates were phenotypically characterized by culturing on Sabouraud’s dextrose agar, Hichrome Candida differential agar and germ tube production test. Antibiogram was also performed to determine their antifungal activities. The phenotypically positive isolates were confirmed by polymerase chain reaction (PCR) and genetically analyzed by targeting 18S-ITS1-5.8S-ITS2-28S region specific for Candida spp. and identified the yeast at the species level. The antibiogram showed the isolates were highly sensitive with ketoconazole, clotrimazole and miconazole. The PCR assay identified C. lusitaniae and C. tropicalis based on the two distinctive amplicon sizes (592bp and 737bp) respectively. Also, the sequence analysis and phylogeny confirmed C. lusitaniae in six sequences and C. tropicalis in one sequence. It is worth noting that in this study, the species identification was consistent among PCR and genetic analysis. Therefore, the PCR based identification system of the fungal species performed in this study could be an efficient and time saving tool for early diagnosis of clinical mastitis in milch animal, which allows prompt control and application of speedy effective treatment.