Comparative evaluation of therapeutic effects of silymarin and hydrocortisone on clinical and hematological alterations, and organ injury (liver and heart) in a low-dose canine lipopolysaccharide-induced sepsis model.

The present study aimed to examine the effectiveness of silymarin compared to hydrocortisone on clinical and hematological alterations and organ injury (liver and heart) in a low-dose canine lipopolysaccharide (LPS)-induced sepsis model. Fifteen clinically healthy dogs were randomly categorized into three equal groups: Two dogs in group A, LPS (0.10 µg kg-1, IV) was injected (control, n = 5); Group B was similar to group A, with the difference that silymarin bolus (10.00 mg kg-1, IV, once) was injected 40 min after LPS injection. Group C was similar to group B with the difference that hydrocortisone bolus (2.00 mg kg-1, IV, once) was administrated instead of silymarin. Five mL of blood was collected at baseline, 1, 3, and 6 hr of the study. Septic control dogs experienced a significant reduction in red blood cells count (RBC), hemoglobin (Hb), and hematocrit (HCT) and a significant elevation in serum activities of aspartate aminotransferase (AST), alkaline phosphatase (ALP), lactate dehydrogenase (LDH), creatine kinase isoenzyme MB (CK-MB), and plasma cardiac troponin I (cTnI) concentration. We noticed a significant increase in RBCs, Hb, and HCT, and a significant decrease in AST, ALP, LDH, CK-MB, and cTnI in the silymarin group in comparison with hydrocortisone and control group. Our results suggested that silymarin had a positive influence on sepsis due to protecting RBCs, and decreasing organ (heart and liver) injury. These findings supported the hypothesis that silymarin could be more effective than routine corticosteroid therapy in sepsis.

Evaluation of 5 methods for diagnosing failure of passive transfer in 160 Holstein calves.

BACKGROUND: Inadequate absorption of colostral IgG1 is termed failure of transfer of passive immunity (FTPI). Dairy calves with FTPI have increased mortality and morbidity in their first 6 months of life. OBJECTIVES: This study compared the clinical performance of 5 methods for diagnosing FTPI in Holstein calves. METHODS: An observational study was performed using 160 Holstein heifer calves. Serum was harvested at 48 hours of age, and FTPI was assessed using a digital Brix refractometer for total solids measurements, and digital refractometry and the biuret method to measure serum total protein (STP) concentrations. Serum gamma-glutamyltransferase activity was measured with an automated analyzer, and serum IgG was measured with the zinc sulfate turbidity test and an enzyme-linked immunosorbent assay. Diagnostic test performance was compared with that of the reference method (FTPI defined as a serum total IgG concentration <10 g/L). Test performance was evaluated using the area under the receiver operating characteristic curve, the sensitivity, the specificity, and the positive likelihood ratio at the optimal test cut point, and by calculating the kappa coefficient. RESULTS: A serum digital Brix percentage of <7.8% and an STP concentration of <52 g/L measured using digital refractometry were the best methods to identify calves with FTPI. The STP concentration measured with digital refractometry was 0.1 g/L lower than that measured with the biuret method. CONCLUSIONS: The digital Brix refractometer and the digital refractometer provide accurate and clinically useful methods for identifying dairy calves with FTPI. In this study, the excellent performance of the Brix refractometer was likely due to the use of a fixed sample volume (200 µL) and a uniform sample temperature at the time of measurement.

Comparison of rapid diagnostic tests to detect Mycobacterium avium subsp. paratuberculosis disseminated infection in bovine liver.

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne's disease, a chronic enteritis in cattle and other domestic and wild ruminants. The presence of MAP in tissues other than intestines and associated lymph nodes, such as meat and liver, is a potential public health concern. In the present study, the relationship between the results of rapid diagnostic tests of the Johne's disease, such as serum ELISA, rectal scraping PCR, and acid-fast staining, and the presence of MAP in liver was evaluated. Blood, liver, and rectal scraping samples were collected from 200 slaughtered cattle with unknown Johne's disease status. ELISA was performed to determine the MAP antibody activity in the serum. Acid-fast staining was performed on rectal scraping samples, and PCR was performed on rectal scraping and liver samples. PCR-positive liver samples were used for mycobacterial culture. Overall, the results of this study demonstrated that MAP can be detected and cultured from liver of slaughtered cattle and rapid diagnostic tests of Johne's disease have limited value in detecting cattle with MAP infection in liver. These findings show that the presence of MAP in liver tissue may occur in cows with negative results for rapid diagnostic tests and vice versa. Hence, liver might represent another possible risk of human exposure to MAP. Given concerns about a potential zoonotic role for MAP, these results show the necessity to find new methods for detecting cattle with MAP disseminated infection.

Genotypic analysis of virulence genes and antimicrobial profile of diarrheagenic Escherichia coli isolated from diseased lambs in Iran.

The aim of the present study was to determine the analysis of virulence genes and antimicrobial profile of diarrheagenic Escherichia coli isolated from diseased lambs. Two hundred ninety E. coli isolates were recovered from 300 rectal swabs of diarrheic lambs and were confirmed by biochemical tests. The pathotype determination was done according to the presence of genes including f5, f41, LTI, STI, bfp, ipaH, stx 1 , stx 2 , eae, ehlyA, cnf 1 , cnf 2 , cdIII, cdIV, and f17 by PCR method. Sixty-six isolates (23.72%) possessed the STI gene and categorized into entrotoxigenic E. coli (ETEC). Nine isolates (3.1%) and five isolates (1.72%) were positive for the cnf1 and cnf2 genes which categorized into necrotoxic E. coli (NTEC). Hundred and seventeen isolates (40.34%) harbored stx 1 and/or stx 2 and classified as Shiga toxin-producing E. coli (STEC). Thirteen isolates (4.48%) were assigned to atypical entropathogenic E. coli (aEPEC) and possessed eae gene. Two isolates (0.68%) were positive for ipaH gene and were assigned to entroinvasive E. coli (EIEC). Statistical analysis showed a specific association between eae gene and STEC pathotype (P < 0.0001). The most prevalent resistance was observed against lincomycin (96.5%) and the lowest resistance was against kanamycine (56.02%), respectively. The high prevalence of STEC and ETEC indicates that diarrheic lambs represent an important reservoir for humans. ETEC may play an important role for frequent occurrence of diarrhea in lambs observed in this region. Due to high antibiotic resistance, appropriate control should be implemented in veterinary medicine to curb the development of novel resistant isolates.

Simple Indirect Enzyme-Linked Immunosorbent Assay to Detect Antibodies Against Bovine Viral Diarrhea Virus, Based on Prokaryotically Expressed Recombinant MBP-NS3 Protein.

BACKGROUND: Bovine viral diarrhea (BVD) is an economically important disease of cattle distributed worldwide. Diagnosis of BVD relies on laboratory-based detection of its viral causing agent or virus specific antibodies and the most common laboratory method for this purpose is Enzyme-Linked Immunosorbent Assay (ELISA). OBJECTIVES: The current study was aimed to develop a simple indirect ELISA to detect antibodies against Bovine Viral Diarrhea Virus (BVDV) in the sera of infected cattle. MATERIALS AND METHODS: A new simple indirect ELISA method was developed to detect BVDV infection by prokaryotically (Escherichia coli, BL21 strain) expressed recombinant whole nonstructural protein 3 (NS3) of BVDV (NADL strain). Four hundred bovine serum samples were evaluated by the newly developed NS3-ELISA and virus neutralization test (VNT) as the gold standard method to diagnose BVD. Among these samples, 289 sera had been previously tested by a commercial ELISA kit. RESULTS: Statistical analyses showed a very high correlation between the results of the developed NS3-ELISA and VNT (kappa coefficient = 0.935, P < 0.001), with the relative sensitivity and specificity of 94% and 98.8%, respectively. There was also a high correlation between the results of NS3-ELISA and the commercial ELISA kit (kappa coefficient = 0.802, P < 0.001) with the relative sensitivity and specificity of 90.72% and 91.15%, respectively. CONCLUSIONS: The newly developed simple indirect ELISA showed high sensitivity and specificity with respect to VNT. Developing such a simple, sensitive, and specific ELISA which is much less expensive than the available commercial ELISA kits can improve the detection of BVDV infections, help to eliminate the disease from herds, and decrease economic losses caused by this disease.

Epitope mapping of bovine viral diarrhea virus nonstructural protein 3.

Six consecutive overlapped coding regions (F1-F6) of whole NS3 molecule of bovine viral diarrhea virus (BVDV) were cloned into pMAL-c2X plasmid vector and expressed in Escherichia coli cells (BL21 strain). The recombinant proteins were then purified by amylose resin to determine the most immunogenic domain(s) of the NS3 molecule. Evaluation of the recombinant proteins was carried out by indirect ELISAs using several bovine sera (previously characterized by virus neutralization test, a commercial ELISA kit, and a newly developed NS3-ELISA) and 6 monoclonal antibodies. The experiments showed that the most immunogenic domain of the NS3 protein was the fourth designed fragment (F4), a 122 amino-acid (AA) region of about 13.5 kDa (nucleotide 1003-1368; residue 335-456). Purified recombinant F4 was also evaluated as single ELISA antigen (F4-ELISA) for the detection of anti-BVDV antibodies in sera of infected cattle. Although this small recombinant fragment of NS3 protein was almost completely soluble and expressed more efficient respect to whole NS3 molecule, it did not show enough sensitivity and specificity to be a proper substitute for NS3 as ELISA antigen to detect specific antibodies against BVDV. However, statistical analyses showed a medium correlation between the results of the developed F4-ELISA and virus neutralization test (kappa coefficient=0.63, P<0.001), with the relative sensitivity and specificity of 78.05% and 84.91%, respectively, suggesting the potential use of this fragment as an ELISA antigen along with other antigens or monoclonal antibody(s) in a competitive ELISA.

Prevalence of Listeria monocytogenes, Vibrio parahaemolyticus, Staphylococcus aureus, and Salmonella spp. in seafood products using multiplex polymerase chain reaction.

Although several etiological agents can be transmitted through seafood consumption, Listeria monocytogenes, Vibrio parahaemolyticus, Staphylococcus aureus, and Salmonella spp. are considered among the most important pathogens in terms of public health and disease. In this study, multiplex polymerase chain reaction (PCR), as a rapid and cost-effective method, was used to determine the prevalence of these pathogens in 245 samples of raw/fresh, frozen, and ready-to-eat (RTE) seafood products marketed in Iran. The prevalence of L. monocytogenes in raw/fresh fish and shrimp samples was 1.4%, whereas 2.9% of the raw/fresh fish and 7.1% of the shrimp samples were contaminated with V. parahaemolyticus. No contamination with L. monocytogenes and V. parahaemolyticus was found in frozen and RTE seafood products. The prevalence of S. aureus was found to be higher than other investigated pathogens. S. aureus was detected in 5% of the raw/fresh samples of fish and shrimp, 17.5% of the frozen, and 12.3% of the RTE samples. Further, our findings indicate that 2.9% of the fish samples, 4.3% of the shrimp samples, and 1.5% of the RTE samples were contaminated with Salmonella spp. Owing to the potential hazard of these pathogenic bacteria, multiplex PCR can provide a rapid and cost-effective method for the surveillance of these pathogens in seafood products.

Seroprevalence of Toxoplasma gondii in water buffaloes (Bubalus bubalis) in South-West of Iran.

The prevalence of antibodies to Toxoplasma gondii was conducted in 300 buffaloes from Ahvaz, Kouzestan province, southwest of Iran. Blood sera were screened using a Modified agglutination test (MAT) incorporating 2-mercaptoethanol. Positive reactions in sera dilutions above 1:25 were considered as indicative for the presence of T. gondii antibodies. The overall prevalence of infection in the animals was 14.33% with titers of 1:25 in 21, 1:50 in 12, 1:100 in 6, 1:200 in 2 and 1:400 in 2. The prevalence was different in relation to the sex with buffaloes with 19.7% and 7% in females and males respectively. These results indicate that T. gondii infection in water buffaloes of Khouzestan is relatively high and consumption of buffalo meat may be a risk factor for humans in Ahvaz, southwest of Iran.

Metallothionein biosynthesis as a detoxification mechanism in mercury exposure in fish, spotted scat (Scatophagus argus).

It is of crucial importance to study on the biomarkers types to assess the specification of the pollutants and health status of marine ecosystems in environmental evaluation projects. In this respect, total metallothionein biosynthesis and mercury bioaccumulation in the liver and gills under acute mercury exposure were investigated in fish, Scat (Scatophagus argus). Spotted scat was exposed to different mercury concentrations (0, 10, 20, 30) for 24, 48, 72 h. Total MT levels were determined by enzyme-linked immunosorbent assay (ELISA) method. Mercury contents were determined through cold vapor atomic absorption spectrometry (CVAAS). Induction of MT during exposure was tissue specific, displaying different response pattern in gills and liver. Mercury accumulated in liver much higher than in gills and the latter also showed lower MT level (P<0.05). MT biosynthesis in liver showed a significant (P<0.05) increase after exposure to different mercury concentration with increase in exposure time, whereas total MT content did not significantly (P>0.05) change in gills except for 72 h exposure at 30 µg l(-1). Nonetheless, the relationship between MT biosynthesis and Mercury bioaccumulation in both tissues was significant (P<0.05). The results suggest that this form of MT in S. argus was Hg inducible and could be extended as a biomarker of mercury pollution in marine ecosystems.

Occurrence of anti-Toxoplasma gondii antibodies in female cattle in south-west of Iran.

Toxoplasmosis, caused by Toxoplasma gondii, is a significant disease in livestock and humans. In Iran, studies shows that T. gondii infection in humans is relatively high and prevalence is associated mainly with consumption of undercooked meat or meat products. We have examined 450 serum samples from female cattle distributed over all Ahvaz, the center of Khouzestan province, south-west of Iran. IgG antibodies to T. gondii were assayed by the modified agglutination test using whole tachyzoites of T. gondii, and found in 71 (15.77%) of 450 cattle with titers of 1:25 in 38, 1:50 in 18, 1:100 in 11, 1:200 in three and 1:400 in one. Titers of antibodies were decreased in cattle over 2 years old. These results indicate that T. gondii infection in cattle of Khouzestan is relatively considerable, but not very high and consumption of beef may be a source of infection for humans in south-west of Iran.

Role of Clostridium perfringens in causing abomasal ulcers in buffalo.

In this study, the correlation between abomasal ulcers and presence of Clostridium perfringens (C. perfringens) was evaluated in 80 (50 affected and 30 non affected) randomly slaughtered buffaloes in Ahvaz slaughterhouse. Immediately after the slaughter, the abomasums was isolated and an incision was made on the wall of it. Then the abomasums were emptied and its interior was washed with water. The inner surface was examined for presence of abnormal lesion. Ulcers from affected and piece of abomasa from non affected buffaloes were cultured. Cultures were also made from contents of all samples and smears were also prepared from affected and non affected tissues. Cultures from content samples (12%) of 50 ulcerated abomasa were positive for C. perfringens while the agents were isolated from 1 content (3.3%) of non ulcerated abomasa. There was no statistical difference between presence of C. perfringens in contents and abomasal ulcers. Totally C. perfringens were isolated from ulcers of 6 (12%) ulcerated and tissues of 3 (10%) non ulcerated cases. Statistical analysis showed no correlation between presences of C. perfringens and abomasal ulcers. There was no statistical difference between sex and age of the affected animals. In conclusion C. perfringens seems not to be solely, a cause ofabomasal ulcers in buffaloes.