This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.Page 1 The aims of this study were to use multilocus sequence typing (MLST) to i) investigate 26 the population structure, diversity and molecular epidemiology of the weakly 27 haemolytic anaerobic intestinal spirochaete Brachyspira intermedia, and ii) determine 28 the relationship of the species to the other two indole-positive but strongly haemolytic 29Brachyspira species -B. hyodysenteriae and "B. suanatina". Seventy-seven B. 30 intermedia isolates from pigs and chickens were analysed, with the nucleotide 31 sequences of seven conserved genomic loci examined for each. B. intermedia was 32 genetically diverse, with the 77 isolates being divided into 71 sequence types (STs) and 33 64 amino acid types (AATs). Many distinct groups of B. intermedia isolates were 34 identified, with some isolates being separated from others by large genetic distances. 35Although both pig and chicken isolates were found in most groups, suggesting that 36 cross-species transmission of such isolates may occur, some isolates from pigs were 37 located in small groups that did not include chicken isolates, and vica versa.

Avian intestinal spirochaetosis causes problems including delayed onset of lay and wet litter in adult chickens, and results from colonization of the caecae/rectum with pathogenic intestinal spirochaetes (genus Brachyspira). Because avian intestinal spirochaetosis has not previously been studied in South East Asia, this investigation was undertaken in Malaysia. Faecal samples were collected from 25 farms and a questionnaire was administered. Brachyspira species were detected by polymerase chain reaction in 198 of 500 (39%) faecal samples from 20 (80%) farms, including 16 (94%) layer and four (50%) breeder farms. Pathogenic Brachyspira pilosicoli was identified in five (29%) layer and two (25%) breeder farms whilst pathogenic Brachyspira intermedia was detected in nine (53%) layer and one (12.5%) of the breeder farms. Twelve (80%) layer farms had egg production problems and 11 (92%) were positive for Brachyspira: three (25%) for B. pilosicoli and six (50%) for B. intermedia. Of three breeder farms with egg production problems, one was colonized with B. pilosicoli. Three of ten layer farms with wet litter were positive for B. pilosicoli and six for B. intermedia. Of four breeder farms with wet litter, one was colonized with B. pilosicoli and one with B. intermedia. No significant associations were found between colonization and reduced egg production or wet litter, perhaps because so many flocks were colonized. A significant association (P = 0.041) occurred between a high prevalence of colonization and faecal staining of eggs. There were significant positive associations between open-sided housing (P = 0.006), and flocks aged >40 weeks (P < 0.001) and colonization by pathogenic species.

Background:Melioidosis is a fatal emerging infectious disease of both man and animal caused by bacteria Burkholderia pseudomallei. Variations were suggested to have existed among the different B. pseudomallei clinical strains/genotypes which may implicate bacterial susceptibility and resistance toward antibiotics.Aim:This study was designed to determine whether the phenotypic antibiotic resistance pattern of B. pseudomallei is associated with the source of isolates and the genotype.Materials and Methods:A collection of 111 B. pseudomallei isolates from veterinary cases of melioidosis and the environments (soil and water) were obtained from stock cultures of previous studies and were phylogenetically characterized by multilocus sequence typing (ST). The susceptibility to five antibiotics, namely meropenem (MEM), imipenem, ceftazidime (CAZ), cotrimoxazole (SXT), and co-amoxiclav (AMC), recommended in both acute and eradication phases of melioidosis treatment were tested using minimum inhibitory concentration antibiotics susceptibility test.Results:Majority of isolates were susceptible to all antibiotics tested while few resistant strains to MEM, SXT, CAZ, and AMC were observed. Statistically significant association was found between resistance to MEM and the veterinary clinical isolates (p<0.05). The likelihood of resistance to MEM was significantly higher among the novel ST 1130 isolates found in veterinary cases as compared to others.Conclusion:The resistance to MEM and SXT appeared to be higher among veterinary isolates, and the novel ST 1130 was more likely to be resistant to MEM as compared to others.

This study was designed to determine the genotype and the phylogeny of Burkholderia pseudomallei isolated from veterinary cases and from the animal environments in Peninsular Malaysia. The Malaysian B. pseudomallei population were then compared to those found elsewhere. A total of 113 isolates from veterinary cases (35) and the environment (56 from soil and 22 from water) were characterized using multilocus sequence typing (MLST). Two novel alleles, allele 97 and 69 of the gene locus ace and lepA respectively were recovered. Isolates were resolved into 12 distinct sequence types (STs) out of which five were novel, namely ST1130, ST1131, ST1338, ST1339 and ST1367. The isolates from veterinary cases co-clustered with those from the environment. B. pseudomallei isolates in this study were highly clonal and have descended from a common ancestor clonal complex (CC) 48 found in Southeast Asia. This study shows that veterinary case isolates are often caused by similar STs, with similar populations found in the direct animal environment and those previously reported to cause human infections in Malaysia and elsewhere. Isolates of B. pseudomallei from human infections have been given more attention, with a comparatively lower focus on isolates from animals and the farm environment. This study highlighted the genotype and phylogeny of B. pseudomallei isolated from animals and the environment and their relations to the isolates from human cases reported in Malaysia and elsewhere. Most STs reported in this study, from veterinary cases and animal environment are similar to those previously reported as causing human infections in Malaysia and elsewhere. Therefore, even though direct zoonosis is uncommon, monitoring melioidosis occurrences in animals can provide insights on the bacterial strains infecting humans.

The antimicrobial capability of oil containing medium-chain fatty acids and their corresponding monoglycerides, known as Enhanced Virgin Coconut Oil (EVCO) against microorganisms isolated from bovine mastitis milk was studied. EVCO contains an effective amount of medium-chain free fatty acids (caprylic, capric and lauric acid) and their corresponding derivatives (monocaprylin, monocaprin and monolaurin) can act as antimicrobial agent with broad spectrum of antimicrobial properties. An in vitro assessment of EVCO against bovine mastitis microorganisms isolated from locally collected mastitis milk samples was conducted to determine its minimum bactericidal concentration (MBC>99). In general, EVCO exhibited its great antimicrobial capability to kill all mastitis microorganisms isolated from the mastitis milk sample at different concentrations of MBC>99 value. In fact, EVCO was found more effective to kill Mycoplasma sp. and gram positive microbes rather than gram negative microbes, especially in Mycoplasma bovis, Acholeplasma oculi, Enterococcus sp., Streptococcus sp. and Staphylococcus aureus with lower MBC>99 values in comparison with the synthetic antibiotic, Mastivet. A further investigation of the antimicrobial activities of EVCO against Staphylococcus aureus and Mycoplasma bovis against time revealed that EVCO was more efficient in killing these two microorganisms in a shorter time at a lower concentration as opposed to Mastivet. These findings indicated that EVCO has great potential application in postmilking teat germicide, which can be used as a preventive measure to control mastitis cases, especially against environmental mastitis pathogens.