Association of Ovar-DRB1 alleles with innate immune responses in sheep.

BACKGROUND: Major histocompatibility complex (MHC) is the best characterised genetic region associated with adaptive immune responses, including humoral and cell-mediated immunities. OBJECTIVES: In this study, the association of MHC class II alleles with inflammatory cytokines and acute-phase proteins was evaluated in sheep population. METHODS: Allelic diversity of second exon of ovine DRB1 locus (Ovar-DRB1.2) was determined in 100 indigenous Iranian Lori-Bakhtiari fat-tailed sheep using restriction fragment length polymorphism and direct sequencing methods. The association of DRB1.2 alleles with inflammatory cytokines (interleukin-1ß, IL-1ß; IL-6 and tumour necrosis factor-α) and acute-phase proteins (serum amyloid A, alpha-1-acid glycoprotein and haptoglobin) was examined using generalised linear model and multivariate regression analysis. RESULTS: Seven distinct RsaI restriction patterns and fourteen alleles were identified in this population. Allele DRB1*2101 showed a negative influence on the IL-6 response and was associated with lower serum level of IL-6. DRB1.2 heterozygous individuals also showed higher haptoglobin concentration than homozygotes. CONCLUSIONS: These results provide additional support for the association between Ovar-DRB1 alleles and regulation of immune responses in sheep population. Description of MHC polymorphism and its role in the controlling of immune responses will increase our understanding of host-pathogen interactions, and ultimately facilitate the selection of disease-resistant flocks in genetic breeding programs.

The detection of canine anti-sperm antibody following parenteral immunization of bitches against homogenized whole sperm.

BACKGROUNDS: The development of a canine-specific method of immunocontraception is one of the non-invasive controlling strategies for humanely decreasing the dog population. This study was aimed to investigate the potential of whole sperm in stimulating the immune system and producing specific anti-sperm antibodies (ASAs) in female dogs. Mature, mixed-breed bitches were subcutaneously immunized with high (200 × 106 cells/mL) and low (100 × 106 cells/mL) doses of sperm vaccine, emulsified with Freund's adjuvants. Booster immunizations were given at weeks 1, 2, 4, and 6, and serum samples were collected at days 0, 14, 28, 42, 63, and 84 prior to each immunization. Reproductive tract samples, including vaginal and uterine lavages, were also collected by flushing each section with sterile PBS at the end of the experiment. Canine anti-sperm antibody titer and specificity in sera and genital secretions were measured using an enzyme-linked immunosorbent assay technique. RESULTS: Specific anti-sperm antibodies were detected in the serum of both high and low dose groups and were significantly higher than those observed in the controls. A high dose of sperm induced elevated immune responses over the low dose antigen. Immunization with a high dose of sperm increased the level of ASAs in the uterine secretions and vaginal secretions significantly. Higher ASAs were observed to have transduced to the uterine lumen compared to the vagina. CONCLUSIONS: Based on the results obtained in this study, parenteral immunization with whole sperm can induce a high level of specific antibodies in the serum and genital secretions of female dogs and the response would be dose-dependent.

CONTEXTE: Le développement d'une méthode d'immunocontraception spécifique à la race canine constitue l'une des stratégies non invasives pour réduire humainement la population canine. Cette étude a pour objectif d'évaluer le potentiel du sperme entier à stimuler le système immunitaire des femelles et à produire des anticorps anti-sperme spécifiques (ASA) chez ces dernières. Des chiennes matures de race croisée sont immunisées par voir sous-cutanée avec soit de fortes doses (200 millions de cellules/mL) soit de faibles doses (100 millions de cellules/mL) de vaccin constitué de sperme entier émulsifié avec des adjuvants de Freund. Des vaccinations de rappel sont faites aux semaines 1, 2, 4 et 6, et des échantillons sanguins sont prélevés aux jours 0, 14, 28, 42, 63 et 84 avant chaque immunisation. Des échantillons de l'appareil reproducteur, incluant des lavages vaginaux et utérins, sont recueillis par flushing de chaque section avec du PBS stérile à la fin de l'expérimentation. Le titre et la spécificité des anticorps anti-sperme entier canin dans le sérum et les sécrétions génitales ont été mesurés par la technique de dosage ELISA. RÉSULTATS: Des anticorps anti-sperme entier spécifiques ont été détectés dans le sérum des femelles immunisées tant avec de faibles que de fortes doses, et de façon significativement plus élevé que chez le groupe témoin. Une dose forte de sperme entier induit des réponses immunitaires élevées par rapport à l'antigène à faible dose. L'immunisation avec une forte dose de sperme entier augmente de façon significative le niveau d'ASA dans les sécrétions utérines et dans les sécrétions vaginales. On a observé que les ASA ont plus été transduits vers la lumière utérine que vers la lumière vaginale. CONCLUSIONS: Basée sur les résultats obtenus dans la présente étude, l'immunisation parentérale par du sperme entier peut induire un taux élevé d'anticorps spécifiques dans le sérum et le sécrétions génitales de chiennes ; et la réponse serait dose-dépendante.

Prediction of Early Liver Failure in Pediatric Patients Admitted to Intensive Care Unit.

BACKGROUND Hepatic dysfunction has been associated with poor prognosis in critically ill patients. We aimed to investigate the incidence of early liver dysfunction and its association with probable predictive variables in a group of Iranian patients. METHODS The study was conducted on 149 pediatric patients referred to the pediatric intensive care unit (PICU), Shiraz University of Medical Sciences, Shiraz, Iran between April and October 2016. Serum levels of liver aminotransferase, alkaline phosphatase, total bilirubin, direct bilirubin, and international normalized ratio (INR) were recorded in 24, 48, and 96 hours after admission. RESULTS On the first day of admission, direct bilirubin was the least (9.1%) and abnormal alkaline phosphatase level was the most (66.9%) common abnormalities. Abnormal levels of all tests except alkaline phosphatase were predictive of increased rate of mortality. In univariable logistic regression, abnormal aminotransferases (ALT and AST), INR, total bilirubin, and direct bilirubin had significant relationship with patients' mortality after 24, 48, and 96 hours. In multivariable logistic regression only ALT and INR in the first 24 hours had significant relationship with mortality in final model. Although univariate logistic regression revealed a significant relationship between AST and ALT levels with PICU length of stay, no significant relationship was observed between these variables and PICU length of stay (except AST in the first 24 hours) in multivariable analysis. CONCLUSION Increase in liver enzymes may predict mortality and increased PICU length of stay in critically ill children.

Association between DLA-DRB1.2 allelic diversity and development of mammary gland tumors in dogs.

BACKGROUND: The major histocompatibility complex (MHC) is the best-characterized genetic region related to resistance/susceptibility to a wide range of infectious and immune-mediated diseases. Evidences suggest that MHC class II genes may play an important role in developing different types of tumors including breast cancer. Canine mammary gland tumors (CMTs) are the most common neoplasms in female dogs. In the current study, the association of canine MHC class II DLA-DRB1.2 genotypes with development of mammary gland tumor profiles in dogs was investigated. DLA-DRB1.2 allelic diversity was determined in 40 dogs (18 CMT cases and 22 controls) using HRM technique and DNA sequencing. Association of the DLA-DRB1.2 genotypes with CMT profiles was expressed as odds ratio (OR). RESULTS: Based on the histopathological typing of tumors, CMT cases were categorized into 4 groups: simple carcinoma, complex carcinoma, carcinoma arising in a benign tumor and special types of carcinoma. A total of eight HRM profiles (A to H) were identified in dogs sampled. The association study revealed a significant correlation between DLA-DRB1.2 genotypes with different CMT profiles. The E genotype was significantly associated with increased risk of carcinoma arising in a benign tumor, and the B genotype represented a positive correlation with complex carcinoma. Significant association was also observed between the heterozygosity of DLA-DRB1.2 genotypes and decreased risk of developing tumor in dogs. CONCLUSIONS: These results provide additional support for the association between DLA-DRB1 genes and development of mammary gland tumors in dogs and could potentially be used for early diagnosis of neoplasia and identifying susceptible dogs.

Evaluation of humoral immune responses to enterotropic lentogenic VG/GA vaccine of Newcastle disease in commercial turkey poults (Meleagris gallopavo).

BACKGROUND: Villegas-Glisson/University of Georgia (VG/GA) strain of Newcastle disease virus (NDV) is recommended for the initial vaccination of commercially reared turkey poults. However, the vaccine-induced antibody responses have not been studied in this species. The level of systemic humoral immune responses against the NDV was investigated in commercial turkey poults vaccinated with the VG/GA vaccine. One hundred eighty-two hybrid strain of turkey poults (Meleagris gallopavo) were divided randomly into vaccinated and unvaccinated groups. The vaccinated group was given the VG/GA vaccine at 10 and 20 days of age. To investigate the vaccine immunity, the level of specific IgY and IgA in serum samples were determined using ELISA and haemagglutination inhibition assays (HI). The biological half-life of maternal antibodies was also determined before the immunization. RESULTS: VG/GA-specific antibodies were detected in the vaccinated turkey poults and were significantly higher in the vaccinated group compared to the unvaccinated group. IgY and IgA antibodies showed a significant increase in titers 14 days after the second vaccination and reached a peak on day 35 of age. The correlation coefficient and intra-rater reliability showed a significant correlation between the HI titers and IgY/IgA ELISA values. Maternal IgY and IgA levels were found to decline in the serum with half-lifes of 7.68 ± 2.35 and 2.18 ± 0.82 days, respectively. CONCLUSIONS: Enterotropic lentogenic VG/GA vaccine induced a marked humoral immune response against the NDV in turkey poults. The positive correlation between IgY and IgA highlights the role of these two antibody classes in controlling the Newcastle disease in turkey poults.

Phylogenetic analysis of goat warble fly (Przhevalskiana silenus) based on mitochondrial COI gene.

The larvae of the genus Przhevalskiana (Diptera: Oestridae) are the causative agents of subcutaneous myiasis in goats. Several species have been grouped under this genus based on the morphology of different larval stages, albeit with a lot of uncertainties. Thus, application of genetic tools seems to be helpful for taxonomy. During this study, the cytochrome oxidase I (COI) gene was targeted for the characterization of larval stages of goat warble fly. Fragments of 606 bp were amplified for all the specimens. Based on the COI gene analysis, all the recovered specimens were identified as larvae of Przhevalskiana silenus. Molecular data on the genus is relatively rare but present isolates revealed about 87-89% identity with previous isolates of P. silenus. According to the phylogenetic data, the present isolates branched (as a sister group) with a number of Hypoderma spp. including H. bovis, H. diana, H. lineatum and H. sinense. The present findings confirmed that the COI gene could be a suitable marker for genetic characterization and identification of larvae up to the species level.

Association between Helicobacter pylori Infection and Iron Deficiency Anemia in School-aged Iranian Children.

OBJECTIVES: To find the relationship between Helicobacter pylori infection and iron deficiency anemia in school-aged children. METHODS: 71 children with dyspepsia, epigastric and vague abdominal pain attending a tertiary medical center in Iran underwent upper gastrointestinal endoscopy and were investigated for H. Pylori infection. Hemoglobin, mean corpuscular volume (MCV), serum ferritin, total iron binding capacity (TIBC) and serum iron levels were compared between children with or without H. pylori infection. RESULTS: H. pylori infection was detected in 42 (59.1%) patients. Proportion of children with iron deficiency anemia was not statistically different between two groups (26.2% vs. 14.3%; P=0.48). While hemoglobin was significantly lower in children with H. pylori infection (P=0.01), there were no significant differences in serum level of ferritin, iron, mean corpuscular volume and total iron binding capacity. CONCLUSIONS: Presence of H. pylori does not seem to play an important role in the pathophysiology and development of iron deficiency anemia in school-aged Iranian population.

Production, purification, and evaluation of quail immunoglobulin Y against Salmonella typhimurium and Salmonella enteritidis.

Salmonella species have been the major foodborne problems in food production systems, with Salmonella enterica serovars typhimurium (S. typhimurium) and enteritidis (S. enteritidis) being among the more common isolates. The oral administration of chicken egg yolk specific antibodies (IgYs) has been established as an efficient alternative for treatment and prevention of gastrointestinal pathogens including Salmonella. The present study was aimed to investigate the possible production of specific IgYs against Salmonella typhimurium and Salmonella enteritidis in quail egg yolks. Salmonella spp.-free female Japanese quails (Coturnix coturnix japonica) were intramuscularly immunized with formalin or heat-inactivated Salmonella immunogens (1.0 × 109 CFU/mL) emulsified with Freund adjuvants. Egg yolk IgYs were purified using ammonium sulfate precipitation method. Anti-Salmonella IgYs titer and specificity were determined using enzyme-linked immunosorbent assay (ELISA) and western blot analysis. Salmonella specific IgYs detected in the immunized quails were significantly higher than those of the control group, which confirmed the immunization procedure. Specific IgYs against S. typhimurium and S. enteritidis were identified in both groups immunized with heat or formalin-inactivated immunogens. However, formalin-inactivated immunogens induced relatively higher immune responses over the heat-inactivated ones. Quail anti-Salmonella IgYs showed a high specificity to their corresponding immunogens, with moderate cross-reactivity to other members of Enterobacteriaceae family. Quail can be regarded as a valuable and inexpensive source for producing large-scale of specific antibodies that can be used for immunodiagnostic and immunotherapeutic purposes.

Low Prevalence of Insulin Resistance Among Iranian Patients with Chronic Hepatitis C Virus Infection: A Case-Control Study.

BACKGROUND: Association between Chronic Hepatitis C virus (CHC) infection and type 2 diabetes mellitus has been challenging in recent decades. Despite of extensive research in this area, there is no general agreement on the direct effect of HCV infection on insulin resistance. PATIENTS AND METHODS: The study was performed in 52 CHC patients (mean age = 39.48) and 52 sex-matched healthy Iranian controls, referred to the Hepatitis Clinic, Department of Gastroenterohepatology, Shiraz University of medical sciences, Shiraz, Iran, from 2012 to 2015. Fasting blood glucose level, fasting insulin level and insulin resistance defined as a Homeostasis Model Assessment of Insulin Resistance (HOMA-IR) index were determined and compared between two groups. RESULTS: Insulin resistance was present in 26.9% of CHC patients and 34.62% of healthy controls. Mean HOMA index was 1.93 in patients and 2.18 in controls. There were no statistically significant differences between patient and control groups with regard to fasting insulin level, fasting blood glucose, HOMA index and insulin resistance. HOMA index and fasting insulin level were significantly higher in IR CHC patients relative to IR controls. Fasting blood glucose was also significantly higher in controls younger than 40 years. CONCLUSION: Results obtained in this study showed that chronic hepatitis C cannot be considered as a risk factor for insulin resistance and diabetes in Iranian population. However, regular screening for insulin resistance is recommended in CHC patients with age ≥ 40 years and fasting blood glucose ≥ 100 mg/dl.

Association Study of Glutathione S-transferases Gene Polymorphisms (GSTM1 and GSTT1) with Ulcerative Colitis and Crohn's Disease in the South of Iran.

BACKGROUND: Inflammatory bowel diseases (IBDs), including ulcerative colitis (UC) and Crohn's disease (CD), are chronic inflammatory disorders of the gastrointestinal tract. A combination of environmental factors and interactions with a genetic predisposition are suggested to play an important role in the etiology and pathogenesis of the IBD. Glutathione S-transferases (GSTs) are multifunctional enzymes involved in the cellular oxidative stress handling. Possible associations between GSTs gene polymorphisms and susceptibility to UC and CD have been reported in different population. The relationship between GSTM1 and GSTT1 deletion polymorphisms and susceptibility to UC and CD were investigated in the Iranian population. MATERIALS AND METHODS: The study was performed in 106 IBD patients and 243 age- and sex-matched healthy Iranian controls consulting the IBD registry center of the Motahari Clinic, Shiraz University of Medical Sciences, Shiraz, Iran, between 2011 and 2013. GSTM1 and GSTT1 genotyping were performed using multiplex polymerase chain reaction and differences in the distribution of gene polymorphisms were analyzed statistically between the studied groups. RESULTS: Statistically significant higher frequency of GSTM1 null genotype was observed in IBD patients (P = 0.01) and in the subgroup of patients with UC (P = 0.04) compared to healthy controls, whereas this was not true for CD patients. No significant association was found between GSTT1 gene polymorphism and UC or CD. CONCLUSIONS: Absence of GSTT1 functional gene does not play an important role in the pathophysiology and development of IBD, UC, and CD in Iranian population whereas GSTM1 null genotype could be considered as a possible genetic predisposing factor for more susceptibility to IBD and UC.

LEI0258 microsatellite variability and its association with humoral and cell mediated immune responses in broiler chickens.

Major histocompatibility complex (MHC) has a profound influence on disease resistance or susceptibility, productivity and important economic traits in chicken. Association of the MHC with a wide range of immune responses makes it a valuable predictive factor for the disease pathogenesis and outcome. The tandem repeat LEI0258 is a genetic marker which is located within the B locus of chicken MHC and strongly associated with serologically defined haplotypes. LEI0258 microsatellite marker was applied to investigate the MHC polymorphism in Ross 308 broiler chicken (N=104). Association of LEI0258 alleles with humoral and cell mediated immune responses to Newcastle disease (ND), Infectious bursal disease (IBD) and Avian influenza (AI) vaccines were also examined. LEI0258 polymorphism was determined by PCR-based fragment analysis, and association of LEI0258 alleles with immune responses were evaluated using multivariate regression analysis and GLM procedures. A total of seven alleles ranging from 195 to 448bp were found, including two novel alleles (263 and 362bp) that were unique in Ross 308 broiler population. Association study revealed a significant influence of MHC alleles on humoral and cellular immune responses in Ross population (P<0.05). Alleles 385 and 448bp were associated with increased peripheral blood lymphocyte proliferation response. Alleles 300, 362 and 448bp had a positive effect on immune responses to Infectious bursal disease vaccine, and allele 263bp was significantly correlated with elevated antibody titer against Newcastle disease vaccine. Results obtained from this study confirmed the important role of MHC as a candidate gene marker for immune responses that could be used in genetic improvement of disease-resistant traits and resource conservation in broiler population.

Association of BoLA-DRB3.2 Alleles with BLV Infection Profiles (Persistent Lymphocytosis/Lymphosarcoma) and Lymphocyte Subsets in Iranian Holstein Cattle.

Major histocompatibility complex (MHC) is the best-characterized genetic region associated with resistance and susceptibility to a wide range of diseases. In cattle, the most important example of the relationship between the MHC and infectious diseases has been established by the resistance to Bovine leukemia virus (BLV) infection. The association of the bovine MHC class II BoLA-DRB3.2 alleles with BLV infection profiles was examined. BoLA-DRB3.2 allelic diversity was determined in 190 Iranian Holstein cattle using direct sequencing method. Association of the DRB3.2 alleles with BLV infection profiles was found as the odds ratio. Effects of the alleles on lymphocyte subsets were also evaluated by multivariate regression analysis and GLM procedures. The studied cattle were categorized into three groups: BLV seronegative, BLV seropositive with persistent lymphocytosis (PL), and BLV seropositive with lymphosarcoma (LS). The PL profile was significantly associated with the BoLA-DRB3.2*0101, *1101 and *4201 alleles, although the *3202 allele mediating resistance to PL was observed. Significant association was found between the BoLA-DRB3.2*1802, *3202, and *0901 alleles and susceptibility to LS, while the *0101 and *1101 alleles were associated with resistance to LS. BoLA-DRB3.2 alleles also showed a significant correlation with CD4, CD8, CD21 cells and CD4/CD8 ratio. Allelic differences influence the immune response to BLV infection and developing the disease profile. These differences also have important consequences for tumor resistance.

Chicken major histocompatibility complex polymorphism and its association with production traits.

Major histocompatibility complex (MHC) is the best characterized genetic region controlling disease resistance and immune responses in chicken. MHC genes are also involved in various non-immune functions such as productive traits and reproductive success. The genetic diversity of MHC in an Iranian indigenous chicken (Khorasan) was studied, and association of the MHC alleles with production traits was determined. The MHC polymorphism was ascertained by genotyping the LEI0258 microsatellite locus by PCR-based fragment analysis. LEI0258 microsatellite marker is a genetic indicator for MHC, which is located on microchromosome 16 and strongly associated with serologically defined MHC haplotypes. A total of 25 different LEI0258 alleles (185-493 bp) and 76 genotypes were identified in 313 chickens. An allele of 361 bp had the highest frequency (26.44%), and alleles of 207 and 262 bp had the lowest (0.16%). High level of heterozygosity (87%) and good genotype frequency fit to the Hardy-Weinberg equilibrium was observed in this population (P = 0.238). The association study also revealed a significant influence of MHC alleles on body weight, egg weight, egg laying intensity, and weight of sexual maturity in Khorasan population (P < 0.05). The information obtained from this study indicates a high MHC genetic diversity and the association of MHC alleles with important production traits in Khorasan chicken. These data would be applicable in designing breeding and genetic resource conservation for indigenous chicken populations.

LEI0258 microsatellite variability in Khorasan, Marandi, and Arian chickens.

Microsatellite LEI0258 is a genetic marker for chicken MHC haplotypes and can be used as an indicator of the influence of population genetics on immune responses. LEI0258 microsatellite variability in three Iranian indigenous chicken populations (Khorasan, Marandi, and Arian) was investigated. In total, 142 Khorasan, 42 Marandi, and 58 Arian chickens were examined. Collectively, 25 different alleles and 79 genotypes could be found. The observed levels of heterozygosity were 81% in Khorasan and Marandi and 34% in Arian chickens. Our results indicate that LEI0258 diversity in Marandi chickens is higher than in the other populations. Allelic diversity in Iranian chickens is relatively higher than in the local chicken breeds reported for Brazil and Vietnam.