Goose Parvovirus (GPV) also known as Derzy’s disease is aninfectious viral disease of waterfowl which causes serious economic loss in industrial production of geese and Muscovy ducks. In year 2014, Derzy’s disease was detected in Pekin ducks from Sarawak. The affected farm recorded up to 50% mortality, affecting only young ducklings (starting at the age of 3 weeks). Polymerase chainreaction (PCR) from liver samples were performed based on partial region of VP3 gene of GPV, generated amplicon of 801 bp. Sequence analysis showed that the isolate shared 99% sequence similaritywith goose parvovirus strain YBLJ and YZYZ20130304 from China. Phylogeny based on VP3 showed that this isolate is grouped under Asian strains. This is the first report of GPV in Malaysia focusingon the molecular analysis. Notably, this study revealed that GPV not only can be detected from goose and Muscovy but also from Pekin duck.

OBJECTIVE To evaluate a hypervariable octameric oligonucleotide fingerprints (HOOF-Prints) assay for identification of and discrimination between wild-type and vaccine strains of Brucella melitensis. SAMPLE Brucella melitensis vaccine strain M5 and wild-type strain M43. PROCEDURES 8 pairs of primers (alterable, octameric nucleotides) were designed on the basis of a biological analysis of 8 flanking sequences in the DNA of B melitensis. The HOOF-Prints technique was used to identify wild-type and vaccine strains of B melitensis. Phylogenetic analysis of short, polymorphic fragments of DNA from B melitensis strains M5 and M43 was performed. RESULTS Variable-number tandem repeat DNA segments of B melitensis vaccine strain M5 and wild-type strain M43 were successfully amplified by means of PCR assay. All target gene fragments ranged in size from 100 to 300 bp. Separate phylogenetic analysis of each Brucella strain revealed considerable differences between the vaccine and wild-type strains. CONCLUSIONS AND CLINICAL RELEVANCE The results of this study suggested the HOOF-Prints assay may be useful for discriminating vaccine strains of B melitensis from wild-type strains. This ability could allow discrimination between animals that are seropositive because of vaccination against B melitensis and those that are seropositive because of B melitensis infection and could decrease the likelihood of importing Brucella-infected animals.

The primary objectives of this study were to estimate the prevalence of Campylobacter fetus subsp. venerealis (Cfv) and Tritrichomonas foetus in breeding bulls from a sentinel cohort of cow-calf herds in western Canada and to estimate the association between positive test status and non-pregnancy. The final objective was to evaluate the application of these tests when: i) screening bulls in the absence of a recognized problem with reproductive performance, and ii) testing for diagnosis of poor pregnancy rates. The crude apparent bull prevalence for Cfv was 1.1% [95% confidence interval (CI): 0.5% to 2.1%; 8/735] and herd prevalence was 2.6% (95% CI: 0.3% to 9.0%; 2/78). The crude apparent bull prevalence for T. foetus was < 0.001% (95% CI: 0.0% to 0.5%; 0/735) and herd prevalence was < 0.001% (95% CI: 0.0% to 4.6%; 0/78). Cows from herds where at least 1 bull was test positive for Cfv were 2.35 times more likely (95% CI: 1.01% to 5.48%; P = 0.047) to not be pregnant than those with no positive bulls. Polymerase chain reaction (PCR) testing of preputial material collected into phosphate-buffered saline (PBS) was recommended for screening for T. foetus when the pre-test probability of infection was > 1%. The same test for Cfv was not recommended for screening moderate- and low-risk herds due to the high risk of false positives. Tests for both T. foetus and Cfv can be used to investigate herds with reproductive problems when also ruling out other risk factors. Regardless of the type of test used, however, 3 negative tests are required to rule out infection in high-risk situations.

OBJECTIVE To develop 2 rapid loop-mediated isothermal amplification (LAMP) assays for detection of Pasteurella multocida DNA (Pm-LAMP assay) and P multocida DNA from strains associated with hemorrhagic septicemia (HS) in cattle and buffalo (HS-LAMP assay). SAMPLE Solutions containing 16 P multocida strains and 9 other bacterial species at various concentrations. PROCEDURES Optimal conditions were determined for running the Pm-LAMP and HS-LAMP assays. The assays were then used to detect DNA of the test organisms. Results of LAMP assays were validated against conventional PCR assays designed for specific detection of P multocida and the B:2 serotype of HS-associated strains. RESULTS Following incubation of sample reaction mixtures for 27 minutes, specificity and sensitivity of the HS-LAMP assay at template DNA amounts as low as 5 pg were 93% and 97%, respectively. When duplicates of each sample were incubated for 28 minutes (a positive result defined as positive results for both reactions of a given sample), specificity and sensitivity of the HS-LAMP assay in the same conditions increased to 100%. The best specificity and sensitivity of Pm-LAMP single (93% and 91%) and duplicate (97% and 98%) reactions at template DNA amounts as low as 10 pg were achieved at 33 and 34 minutes, respectively. CONCLUSIONS AND CLINICAL RELEVANCE These preliminary findings suggested the developed HS-LAMP assay had high sensitivity and specificity for detection of HS-associated P multocida. Additional research is needed to determine the accuracy of the assay for use on clinical specimens obtained in HS-endemic countries such as Pakistan and Thailand.

OBJECTIVE To evaluate the correlation between the density of native gastric Helicobacter spp and the presence of gastric lesions in dogs. ANIMALS 80 dogs of various breeds, sexes, and ages. PROCEDURES Gastroscopic and histologic examinations were performed for all dogs. Helicobacter spp were detected by combining evaluation of urease activity and results of bacteriologic culture, microscopic observation, and a 16S rRNA PCR assay. The density of Helicobacter-like organisms was evaluated with light microscopy by use of Warthin-Starry modified stain. Correlations were evaluated by use of the Spearman correlation analysis. RESULTS Gastritis was found in 55 of 80 dogs and classified as mild (n = 31), moderate (16), or severe (8). Of these 55 dogs, only 8 had clinical signs. Histologic examination revealed some degree of lymphocytic-plasmacytic infiltrate, mild eosinophilia, and neutrophilic inflammation in the lamina propria. Seventy-six dogs had positive results for Helicobacter spp. Helicobacter pylori DNA was not detected. Low density and homogeneous distribution of Helicobacter spp were observed in all gastric zones. CONCLUSIONS AND CLINICAL RELEVANCE A significant correlation between density of Helicobacter spp and gastroscopic or histologic lesions was not detected. These findings supported the contention that there is no correlation between general Helicobacter spp density or numbers and gastritis in dogs.

A comprehensive study was conducted on the clinical observationsincluding clinical history, physical examination along with haematobiochemical alteration on 41 naturally occurring cases of canine babesiosis from Punjab state, India. Examination of 964 dogs revealed 4.25 percent (41/964) prevalence of the disease including 3.84 percent (37) B. gibsoni and 0.41 percent (4) B. canis infected cases. Clinical and parasitological diagnosis was finally confirmed by polymerase chain reaction. A large variation of clinical anifestations including rare findings of paraplegia, blindness, ocular bleeding, immune mediated haemolytic anaemia (IMHA), ascites and skin lesions were observed among the affected animals. Bloodfilms showed anisocytosis and nucleated erythrocytes indicating regenerative anaemia. Blood parameters of the affected dogs revealed significant decrease in Hb, TEC, PCV and thrombocytes. Significantdecrease in lymphocytes was found in B. gibsoni affected animals. The affected dogs showed significant increase in serum bilirubin, ALT, AKP, BUN and creatinine. Haemato-biochemical observations wereindicative of severity of babesiosis in dogs.

OBJECTIVE To develop a high-resolution melting (HRM) assay to detect the g.66493737C>T polymorphism in the myostatin gene (MSTN) and determine the frequency of 3 previously defined g.66493737 genotypes (T/T, T/C, and C/C) in warmblood horses. SAMPLES Blood samples from 23 horses. PROCEDURES From each blood sample, DNA was extracted and analyzed by standard PCR methods and an HRM assay to determine the MSTN genotype. Three protocols (standard protocol, protocol in which a high-salt solution was added to the reaction mixture before the first melting cycle, and protocol in which an unlabeled probe was added to the reaction mixture before analysis) for the HRM assay were designed and compared. Genotype results determined by the HRM protocol that generated the most consistent melting curves were compared with those determined by sequencing. RESULTS The HRM protocol in which an unlabeled probe was added to the reaction mixture generated the most consistent melting curves. The genotypes of the g.66493737C>T polymorphism were determined for 22 horses (16 by HRM analysis and 20 by sequencing); 14, 7, and 1 had the T/T, T/C, and C/C genotypes, respectively. The genotype determined by HRM analysis agreed with that determined by sequencing for 14 of 16 horses. The frequency of alleles T and C was 79.5% and 20.5%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that HRM analysis may be a faster and more economical alternative than PCR methods for genotyping. Genotyping results might be useful as predictors of athletic performance for horses.

Escherichia coli isolates from chickens with colibacillosis were assigned to phylogenetic groups based on multiplex polymerase chain reaction (PCR) and antibacterial resistance of E. coli belonging to these groups was examined. Furthermore, the gyrA gene of isolates was sequenced and a phylogenetic tree was generated. A total of 84 E. coli isolates were grouped using multiplex PCR of TSPE4.C2, chuA, yjaA, and gadA molecular markers. Four phylogenetic groups were identified with strains divided as follows: 16 in group A (19.05%), 17 in group B1 (20.24%), 23 in group B2 (27.38%), and 28 in group D (33.33%). Escherichia coli isolates belonging to phylogenetic groups B2 and D were resistant to Soltrim and Flumequine unlike the majority of E. coli isolates that belonged to groups A and B1, and which were susceptible to these antibiotics. The phylogenetic results based on gyrA gene sequences from multiplex PCR revealed that E. coli phylogenetic grouping was in accordance with the clusters obtained in the phylogenetic tree. In conclusion, the comparative sequence analysis of gyrA sequences provides a firm framework for an accurate classification of E. coli and related taxa and may constitute a pertinent phylogenetic marker for E. coli.

Colibacillosis is a disease caused by avian pathogenic E. coli (APEC) and is one of the principle cause of morbidity and mortality in poultry worldwide which is represented by a complex syndrome characterized by multiple organ lesions. This study was carried out to determine phylogenetic grouping and virulenceassociated genes contained by E. coli isolates which is related in causing disease in chicken. E. coli isolates obtained from clinical cases of Veterinary ResearchInstitute were re-identified by conventional methods. Phylogenetic grouping of the isolates was determined by triplex polymerase chain reaction (PCR), and the presence of eight virulence genes were identified by multiplex PCR. A total of 125 E. coli isolates were subjected toanalysis of phylogenetic background and virulence associated genes profiling. Phylogenetic analysis demonstrated that most of the E. coli isolated from chicken in this study belonged to group B1 (36.0%),group D (28.0%), group A (27.2%) and group B2 (8.8%). Multiplex PCR analysis demonstrated that 96 (78.6%) of the E. coli isolates harbored at least one virulencegene, while 29 (23.3%) did not contain any virulence genes tested. The most prevalent virulence genes identified were iss (51.2%), followed by iucD (36.0%),tsh (32.8%), vat (16.0%), astA (13.6%), irp2 (11.2%), papC (9.6%) and the least is cva/cvi gene (0%). None of the isolates harbored more than four virulence genes.Each of phylogenetic groups presented with different combinations of virulence genes, with no specific combinations of virulence genes found to correlate withE. coli phylogroups. None of the E. coli isolates harbored more than four virulence genes, suggesting that E. coli isolates from chicken in this study appear to bederived from commensal strains and may relate to environmental predispose factors especially stress factors in the host to establish infection.