نشریه تحقیقات دامپزشکی و فرآورده های بیولوژیک
سال سی و سوم شماره 4 (پیاپی 129، زمستان 1399)

Adenoviruses are common infectious agents in poultry around the world and show a wide range of clinical signs and symptoms. The importance of the presence of adenovirus infections, in addition to economic losses, is associated with other viral diseases.The purpose of this study was the molecular detection of Fowl adenoviruses and histopathological  examination of liver lesions in broiler chickens suspected of inclusion body hepatitis in Markazi and Ghom provinces, Iran. In this regard, the liver samples with macroscopic lesions were collected from dead birds of ten broiler chicken flocks of these provinces in the winter of 97 and spring 98. The chickens of these flocks were at the age of 6 to 41 days. For molecular analysis, PCR was performed on L1 (Loop1) region of the hexon gene. For histopathology, a portion of the liver tissues were placed in 10% formalin. After routine procedures in the pathology laboratory, microscopically examined. Eventually, 9 flocks from the 20 flocks were positive in the PCR test. These positive products were sequenced. These adenoviruses belonged to the D and E genotypes and D11 and 8b serotypes of avian adenoviruses. Also, in the microscopic study of the affected liver, necrosis with several heterophilia infiltration around these regions and within the liver sinusoid was observed. In the nucleus of healthy hepatocytes around the necrosis regions, the inclusion of intra-basophilic nucleus was observed. Confirmation of the presence of the IBH agent in these provinces, the need for prevention programs in mother and broiler farms should be considered.

Clostridium novyi is endospore- forming, obligate anaerobic bacteria of the class clostridia. Cultivation of this bacterium in common culture is one of the problems with this bacterium. It is pathogenic, causing a wide variety of diseases in man and animals the spore of this bacterium has also received much attention in the treatment of cancer due to its ability to grow in anaerobic conditions inside solid tumors. In this study, the optimum growth conditions for vegetative and spore form was investigated. Bacterial culture was investigated in different solid media including BHI, Blood Agar, F medium and NP medium in vegetative form. Also, a suitable spore media was introduced, the spore purification method, an easy method for bacterial count and spore aspiration were investigated. The best rich media for C.novyi in vegetative form are F and NP media. That is, by adding horse blood to these media culture, the growth be lux but is not a sufficient factor for growth. There was an almost linear relationship between bacterial absorption and bacterial count for Clostridium novyi. Also, the difference in the number of spores germination which heating at 80 ° C for 15 minutes was significant.

Clostridium perfringens is the causative agent of enterotoxaemia, which is related to severe economic losses. The evaluation of immunity of beta antitoxin, that prevented animals against the disease, is measured using potency (serum neutralization) technique that it has disadvantages. So Enzyme-linked Immunosorbent Assay (ELISA) has been proposed as an alternative to SN. In this research, before and after purification of beta toxin, MLD and amount of protein was measured using Lowry method. Then, positive and negative control antiserum and samples antiserum were prepared. Then, we were measured beta antitoxin by ELISA and serum neutralization simultaneously and the results were analyzed by SPSS software.The negative control cut-off was calculated 0.485 and cross-examination was shown that beta toxin used in this study has no cross reaction with other toxins of Clostridium perfringens and its repeatability was assessed using standard deviation (for Medium specimen: SD=0.033 CV=7.25) The results of this study showed that there is a significant agreement between in vivo and in vitro tests for serum samples of vaccinated rabbits by enterotoxaemia vaccine. Linear regression analysis gave correlation coefficients of 0.84 for the indirect ELISA, with a significance level of p < 0.01. Sensitivity, positive and negative predictive value of ELISA in these samples was 100%, 84.21% and 100%, respectively. Finally, ELISA systems are suitable candidates to replace the MNT used for detection antibody in laboratory animals. However, further research in this field is needed for target animals.

In this study, the ITS-1 region of ribosomal RNA genes of six bovine Eimeria species: E. alabamensis, E. auburnensis, E. bovis, E. cylindrica, E. ellipsoidalis and E. zuernii analysed by PCR and the results of molecular methods analysed to find any relationship between presence of parasites and diarretic status. Studies were conducted in calves aged less than 3 months for a period of 2 years. During the study period, 160 dung samples were collected from neonatal calves and examined first microscopically and then by molecular techniques. Stools were analyzed for the presence of Eimeria Spp. oocysts by Sheather's Sugar Flotation Solution method. DNA was extracted and PCR were done to identify six cattle-adapted specie (E.alabamensis, E.auburnensis, E.bovis, E.cylindrica, E.ellipsoidalis and E.zuernii). Eimeria spp. infection was detected in 15 (9.37%) of 160 calves without a history of diarrhea infection and pathogenic Eimeria spp. was asymptomatic in all animals that was not positively related to the diarrheic status of the calves (P>0.05). According to microscopic analsis 6 species (E.alabamensis, E.auburnensis, E.bovis, E.canadensis, E.ellipsoidalis, and E. zuernii) identified and with molecular technique 3 species (E.auburnensis, E.bovis, E. zuernii) were detected. Different risk factors were considered to affect the rate of infection of calves with this protozoan parasite. For controlling disease, Immune status of the calves should be improved by providing sufficient amount of colostrum within the first 24 hours after birth and good hygiene as well as reducing and monitoring stress levels caused by weaning, a change in feed and overcrowding.

Anaplasma is an intracellular microorganism that lives in blood cells of host, causing illness and impose care costs. This study was aimed to investigate molecular and microscopic presence of genus and Anaplasma species in camels of Golestan province. Blood samples were collected from adult slaughtered camels in Inche Bourun industrial slaughterhouse for two years. Microscopic examination was performed by preparing Giemsa-stained blood smear. Molecular test was performed by primary amplification of DNA fragments and then they confirmed by performing the nested-PCR. The specific primers of 16S rRNA and msp4 genes were used to detect different species of Anaplasma. The proliferation of the primary and the nested PCR products showed that genus Anaplasma in 33 out of 90 samples (36.37 percent) were positive. Among samples including Anaplasma 2.22, 22,22 and 13.33 percentage were detected by specific PCR for A. marginale, A. phagocytophilum and A. bovis, respectively. None of the samples showed the presence of Anaplasma centrale. Microscopic observation of anaplasmic bodies in red blood cells (RBC) indicated the presence of Anaplasma genus in 33.33 percent of blood smears. By comparing molecular and microscopic tests, the sensitivity, specificity and accuracy to identify the genus Anaplasma were 21.21, 66.67 and 50 percent, respectively. The results of this study showed that Anaplasma was present among camels in Golestan province. According to the results, molecular study of Anaplasma yields more accurate results than microscopic examination.

The objective of this experiment was to investigate the effects of different levels of milk thistle powder and vitamin E on liver healthy, serum parameters and production indices in laying hens. The 2×3 factorial experiment was managed in a randomized complete design with 72 Hy- Line (W36) laying hens in six treatments and four replicates of three birds each. The experimental period lasted for a total of six weeks. Treatments included different levels of milk thistle powder (0, 100 and 200 mg/kg) and vitamin E (recommended by NRC; 5 mg/kg and more than recommended by NRC; 80 mg/kg). The effect of different levels of milk thistle powder and vitamin E on production indices were not significant in the throughout the experimental period. The best feed conversion ratio was observed in hens fed with rations included 80 mg/kg of vitamin E in the sixth week of experiment (p < 0.05). Laying hens were fed with milk thistle powder (200 mg/kg) had the best feed conversion ratio compared with the control group in the fifth week of experiment (p < 0.05). The effect of different levels of milk thistle powder and vitamin E on egg quality and serum parameters were not significant. Dietary milk thistle powder (200 mg/kg), as compared with the control, decreased serum activity of alanine aminotransferase (p < 0.05). In conclusion, diet supplementation with milk thistle powder (100 and 200 mg/kg) and vitamin E (more than recommended by NRC) had no positive effects on production indices and egg quality in laying hens.

Aim. An antimicrobial and antioxidant propolis and thyme (Thymus vulgaris L.) oil was added to chitosan (chitosan), an edible coating agent. The effects of this coating material on the shelf life and quality of the coated chicken breast samples were evaluated. Material and Method. In this study, chicken breast meat samples were examined in six groups including: control, chitosan, chitosan-alcohol, chitosan - 1% thyme oil, chitosan - 1% propolisexract and chitosan - 1% thyme oil - 1% propolis extract. These samples were stored at 4 ° C for 16 days. Total mesophilic aerobic bacteria (TMAB), total psychophilic aerobic bacteria (TPAB), Lactobacillus spp. and Pseudomonas spp. counts and chemically; pH, aw, Thiobarbutyric acid Reactive Substans (TBARS) and color (L *, a * and b *) values were analyzed on 1st, 4th, 8th, 12th and 16th days of storage. Result. In reviewing the microbiological results; TMAB, TPAB, Lactobacillus spp. and Pseudomonas spp. counts were observed to increase more slowly in the coated groups (P<0.05) during storage. Concerning the results of the physicochemical analysis; TBARS values in the coated groups were lower and observed to increase slower during the storage period (P<0.05). Compared to the ph values of other treated groups, ph value variations in coated groups were observed to be lower (P<0.05). However, it was determined that the coating applications (chitosan, propolis extract, thyme oil) had no effect on aw and color (L *, a * and b *) (P> 0.05).  Conclusion. In this study, the addition of thyme essential oil and propolis extract to the chitosan coating material has been shown to reduce bacterial growth, lipid oxidation and pH change in chicken breast meat samples and to have a positive effect on shelf life. The most effective application on product quality was the treatment gruop of the combination of thyme essential oil, propolis extract and chitosan. As a result, the addition of thyme oil and propolis extract to chitosan coating material could be used for preserving foods. However, more scientific studies are needed on the use of such applications in food preservation.

The trace elements are a serious threat to aquatics as well as human health, as they resistant to degradation and high potential for bioaccumulation in aquatic organisms so present study aimed to evaluate the order and concentration of trace elements accumulated in edible muscle tissue of Common perch (Perca fluviatilis) sampled from Siah Darvishan River considered one of the species of fish as high economic and ecological value and finally, determined its health status for human consumption purposes. In the present experiment, concentration of eleven elements (Se, Fe, Ni, Zn, Pb, Cd, Hg, Cu, Cr, Mn and As) in muscle tissue of 25 Common perch captured by cast net from Siah Darvishan River (Guilan province, Iran) were monitored and evaluated using by Varian® atomic absorption spectroscopy. The mean elements in muscle tissue have been observed, as follows: Zinc (33/418), Iron (20/127), Copper (5/729), Manganese (1/849), Lead (0/864), Selenium (0/691), Arsenic (0/594), Cadmium (0/273), Nickel (0/253), Chromium (0/176) and Mercury (0/089) microgram per gram dry weight, respectively. The results of this study showed that the mean bio- accumulation of all the elements investigated in muscle tissue of Common perch, except the four trace elements; Cadmium, Manganese, Arsenic and Lead were at lower level than the Organization (FAO/WHO) acceptable standard. These results indicate requisiteness for more attention to enhancing factors of these elements in environment of the mentioned species.

The aim of this study was improvement of ram frozen semen efficiency using addition of cysteamine antioxidant to freezing medium. In this experiment, semen samples were collected from five Zandi rams and add to the extender containing 0, 1, 2, 4 and 8 mM cysteamine. In sperm quality evaluation, sperm samples were evaluated and motion parameters, membrane integrity, morphology, acrosome integrity, mitochondria activity, viability and lipid peroxidation were assessed. The results showed using 2 mM cysteamine resulted in improving of sperm total and progressive motility, membrane integrity, mitochondria activity, acrosome integrity, and viability as well as reduction of apoptosis rate and lipid peroxidation. Using the treatment of 8 mM cysteamine caused to reduction of sperm quality after thawing and the antioxidant did not have any effect on sperm morphology. In conclusion, using 2 mM cysteamine in freezing extender could be a helpful strategy for improvement of ram sperm quality after cryopreservation.

To consider the effect of supplementation bentonite and wheat starch on performance, carcass characteristics, blood parameters and value pH of gastrointestinal tract and litter and ilium microbial population intestinal morphology characteristics of broilers, a completely randomized design test was done with four treatments and five replicate for each treatment. In this experimental Two hundred one day-old mixed sex broilers (Ross breed) were divided into four treatments with 10 chicks per each replicate (pen). Treatments were contain: 1- Basal diet without additive (control) 2- 2- Basal diet plus bentonite 3- Basal diet plus wheat starch 4- Basal diet plus bentonite and wheat starch. The diets were fed to broilers at 42 days and water and feed were fed ad-libitum. Range of feed intake and gain of each replicate were weighted weekly and took blood to determine blood parameters from wing vein of birds. The results showed no significant difference between the experimental groups on gain body and feed conversion ratio but in starter period significantly affect feed intake. Addition of bentonite and wheat starch to diet had not significant effect carcass characteristics. The result of this experiment showed during the whole period of experiment, using bentonite and wheat starch no had significantly effect on cholesterol, triglyceride and VLDL of serum but effect on glucose serum. Treatments significantly effect on PDI in starter and grower period. Effect Experimental treatments no had significant on microbial population. Result indicated that between treatments no had significant on villi length and villus/crypt ratio but were statistically significant (p < 0.05) villi thickness and level.

The aim of this study was to investigate the effect of different levels of Myrtle leaf powder on quantitative and qualitative parameters of sperm, antioxidant capacity of semen plasma, and blood superoxide dismutase activity (SOD) in Arabi ram in non-breeding season. 12 adult Arabic rams were used in 3 experimental treatments with 4 replicates per treatment. The treatments consisted of 3 levels of Myrtle leaf powder (0, 12.5 and 25 g/kg of DM). After 42 days feeding of experimental treatments, semen samples were collected and blood samples were obtained once a week for 4 consecutive weeks and the quantitative and qualitative parameters of sperm and total antioxidant capacity of semen were measured. Also, blood samples were subjected to measure testosterone level and SOD activity. 25 g/kg of myrtle powder had the best effect on improvement the sperm and semen characteristics include the motility (73.88%), viability (74.19%), morphological defects (10%), plasma membrane integrity (43.38%), semen volume (1.91 ml) and spermatozoa concentration (1.39 × 109/ml) as well as increased the seminal plasma total antioxidant capacity (TAC) (109.38 μmol/litr) and blood plasma SOD (500.19 U/ml) (p < 0.05). The blood sera testosterone concentration was not affected by treatments. A positive and significant correlation was found between the TAC and SOD with quantitative and qualitative parameters of sperm in Arabi ram (p < 0.05). In conclusion, the use of Myrtle leaf powder in diet improved reproductive performance, seminal plasma TAC and blood SOD activity in Arabi ram. The best result was obtained from 25 g/kg of this plant.

Scorpion sting is one of the serious problems for public health especially in Khuzestan province. The genus Hemiscorpius scorpions are a major factor to scorpion sting and its related death particularly in children. Eleven specimens of Hemiscorpius lepturus scorpion were collected from 6 regions of Khuzestan province and after identification, genomic DNA extraction was performed by manual method of phenol / chloroform. The Internal transcribed spacers (ITS2) genes with about 350 nucleotides was amplified by PCR and then sequenced using forward and reverse primers. The nucleotide sequence was compared using the nBLAST program available at NCBI Web site GenBank database. The results of the phylogenetic showed that the samples can be identified in two clusters A and B, with 85% correlation. Cluster A is subdivided into three subclusters A1 (including: H8, H9, H11M, H5, H44, H2 and H10), A2 (H22 and H4), and A3 (H10M). The specimens in these subclusters, showed a correlation between 54-85%. The studied sequences showed similarity (81.7-98.6%) with the only Hemiscorpius lepturus scorpion sequence (KU341981) recorded at the GenBank database beside a number of scorpions of the Hemiscorpionidae family. The only H9M scorpion specimen in cluster B with a relatively high genetic distance (11.7–19.1%) was discriminated from the rest of the specimens, indicating that this specimen probably belongs to a subspecies different from the Hemiscorpius lepturus scorpion of Khuzestan province.

The purpose of this study was to identify genes that influence bovine growth process using gene expression profile. Growth can be expressed as the change of live weight based on time unit or graph description of weight against animal age. Growth is the result of different biological processes and genetics determines the highest level that these processes can occur. In this study raw data at birth, 3, 7, 12, 20, 25, and 30 months in TXT format were obtained from ArrayExpress database with E-GEOD-2554 accession number. In the next step, the quality control assessment of the existing raw data was performed using Limma package in R environment by PCA method and then the intra array normalization was done by LOESS method and between samples by quantile method. The criteria for identifying genes with differential expression were the absolute value of fold change larger than 2 and adj P-Value < 0.05. DAVID database was used for investigation of biological pathways and reconstruction of the gene expression network were done by Cytoscape software. The results showed that there were significant differences in expression of 1030 growth-related genes at different ages. 518 genes were associated with increased expression and 512 genes with reduced expression. The Wnt, insulin and IGF-I were main signaling pathways that influences growth process. The most important genes identified in this study were FN1, SOD2, HMOX1, SERPINE1, PCK1, STAT3 and HSPA1A. FN1 gene at 20-25 and 25-30 months of age, SOD2 gene at 0-3 months of age, HMOX1 gene at 0-3 and 12-20 months of age, SERPINE1 gene at 3-7 months of age, PCK1 gene at 20-25 months of age, STAT3 gene at 20-25 months of age and HSPA1A gene at 25-30 months of age have a significant increase in expression. Therefore, the results of this study could provide additional information to better understand the relationship between genes and their biological pathways in the growth process that can be used in animal breeding programs.

Heat stresses have an adverse effect on the oxidative status of the body. In order to investigate the effect of cumin and fennel powder in comparison with monensin on oxidative stress of lambs in heat stress conditions, 15 male lambs were used in a completely randomized design in three groups with five replications. Experimental groups included group one: basis diet, group two: basis diet + 8 g of cumin and fennel powder mixture and group three: basis diet + 30 mg monensin. Blood samples were collected from lambs' vein on day 56 and blood serum antioxidant enzymes and blood parameters were measured. Data were analyzed using SAS statistical software. The supplementation of base diet with cumin and fennel powder resulted in a significant increase in the level of glutathione reduction and antioxidant enzymes activity (superoxide dismutase, catalase, glutathione peroxidase and paraoxonase), but the amount of nitric oxide (inflammation index) and malondialdehyde (lipid peroxidation index) in the blood serum of lambs decreased significantly. The addition of cumin and fennel powder, similar to monensin, did not significantly affect thyroxine hormone concentration, but significantly reduced the amount of aspartate aminotransferase enzyme in lamb's blood serum. Based on the findings of this study, adding cumin powder and fennel to 8 g/kg DM in the diet of lambs can reduce the damage caused by oxidative stress and improve the antioxidant status of lambs in heat stress conditions.

Since preserve ram sperm in cold condition has not sufficient efficiency, the objective of the present study was to assess the effect of different levels (0.5, 1, 1.5, 2, 2.5 and 3%) of soy lecithin nano particles and egg yolk in extender on ram sperm quality during storage cool conditions. Semen samples were collected from three Lori Bakhtiari rams using artificial vagina. Afterward, ejaculates were pooled and then divided into seven aliquots. Aliquots were diluted with extenders and stored at 5°C for 72 hours. Total and progressive motility, sperm viability, plasma membrane functionality (HOS), morphology and DNA fragmentation were evaluated at 0, 3, 6, 24, 48 and 72 hours after starting the experiment. The results showed that the extender containing 1.5% nano-lecithin had the highest percentage of total and progressive motility of sperm after 72 hours of storage at cold conditions. The extender containing 3% nano-lecithin had the lowest viable sperm compared to other treatments (p ≤ 0.05). There was no significant difference between all of the extenders for plasma membrane functionality and morphology parameters. DNA fragmentation rate was bigger in treatments containing 0.5 and 3% lecithin nano particles than other treatments. In general, it seems that extender containing 1.5% nano-lecithin can be an appropriate substitute for semen extenders containing egg yolk to store ram sperm in cool conditions.

Today, the use of antibiotics for the prevention, control and treatment of poultry industry is unavoidable and on the other hand requiring a modern approach that providing high sensitivity and specificity, convenient and rapid method to detection antibiotics. Therefore, in this study, for the first time, a wide range of antibiotics used in poultry industry were studied and monitored based on liquid chromatography techniques. Therefore, two liquid chromatography-mass spectrometers were used to prepare the samples with acetic acid and acetonitrile and z-sep+ compounds and with internal standards of antibiotic groups of sulfonamides, quinolones and tetracycline and comprised the results with conventional common method. Because of using of polymeric interface, absorption a large number of antibiotics was possible, so a wide range of antibiotics could be measured without any setting back in static phase efficiency. At the preparation, the amount of sample used is lower than that of classical methods and the preparation steps get shorter. Steady isotope of internal standard caused controls in the extraction, purification, injection and calculations of possible errors such as ion subsidence, injection error, percent recovery of drug and retention time correction and also the possibility to inject high volumes and increase the sensitivity in this method was improved. Therefore, exploitation two liquid chromatography devices, one for sample preparation provides big advantages, which alone lacks this feature. Meanwhile, simultaneously automate method reduces analysis time, test costs and increases accuracy compared to classical method and the sample preparation gets very convenient and allows the ability to search for antibiotics in the produced green chicken.

The main sources of protein, fat and energy are marine foods, most of which are fish, although severely affected by corruption. The shelf life of fish has been affected by a variety of issues, including microbial, chemical, and physical corruption, which is mainly accompanied by an unpleasant smell of leachate. Of the cases that strongly prevent microbial and chemical corruption, packaging is modified or vacuumed and the use of edible coatings. The aim of this study was to evaluate the antioxidant and antimicrobial properties of chitosan nanocomposite and fennel extract (Foeniculum vulgare) in elephant fillet fillets. In order to increase the shelf life of the fish at the refrigerator temperature, MAP, VAP, chitosan and fennel essential oil were used. Treatments in this study were four groups consisting of a control sample coated with chitosan, coated with fennel and chitosan containing essential oil. Samples were packed in two types of MAP and VAP packaging. In total, nine sample groups were sampled with a control sample in normal packaging. The evaluation of the nanocytosine stability properties, the evaluation of microbial indices and chemical samples showed that using chitosan coating containing fennel essential oil in MAP packaging, it is possible to keep elephant fish for optimal quality at refrigerator temperature for 18 days, while the sample the control lasted only 6 days. The treated sample with chitosan coating and fennel essential oil, along with the MAP package, provided good microbial and chemical qualities. But in the sensory evaluation it was found that on the 27th day this sample earned a modest score and was not evaluated desirable. Counts of microorganisms on day 27, samples containing chitosan and fennel essential oils are in the virtual range. But the chemical and sensory characteristics of the elephant fish showed a shelf life of at least 18 days.

In this research, the effect of Lactobacillus plantarum and Lactobacillus rhamnosus bacteria isolated from the gut of the domestic fish (Tor grypus) on the immune response of common carp (Cyprinus carpio) were evaluated. For this purpose, 300 juvenile carps with average weight of 70 ± 11.5 gr were randomly divided into five groups and were fed with feed containing Lactobacillus plantarum, 5×107 CFU/ml Lactobacillus rhamnosus, bacteria combination, 250 mg of commercial product of Multi-Behsil probiotic and non-probiotic for 60 days, respectively. Blood samples were taken at days 0, 30 and 60 from each group and immunological parameters were analyzed and compared. All groups were challenged with Aeromonas hydrophila bacteria on day 60 of the experiment. The lysozyme activity in treated groups with rhamnosus, combination and commercial probiotic significantly increased compared with control group. The activity of myeloproxidase in commercial probiotic group had a significant increase on day 30. There was also a significant rise in NBT reduction in the combination and commercial probiotics group (p <0.05). The bactericidal activity in the plantarum and combination group on day 30 was significantly higher than control group. Bactericidal activity increased significantly after 60 days in all treated groups. Total immunoglobulin in combination and plantarumgroup rose significantly on days 30 and 60, respectively. After the challenge, the combination diet showed a better survival rate than other groups. The results showed that the two isolated bacteria had probiotic effects and appropriate immune stimulation, especially asmixture.