Infections with weakly haemolytic Brachyspira species in pigs with miscellaneous chronic diseases

Laser capture microdissection in combination with fluorescent in situ hybridization was used to identify an unknown species of spirochetes from the pig colonic mucosa. The 16S rRNA gene was PCR amplified, and the closest related type strain was Treponema bryantiiT (90.1%). The spirochete, here named “Candidatus Treponema suis,” was associated with colitis, including invasion of the surface epithelium as well as superficial parts of the mucosa.

Identification of a Novel, Invasive, Not-Yet-Cultivated Treponema sp. in the Large Intestine of Pigs by PCR Amplification of the 16S rRNA Gene

Laser microdissection: A promising tool for exploring microorganisms and their interactions with hosts.

Avian intestinal spirochetosis (AIS), an important but neglected disease in laying hens, is caused by Brachyspira pilosicoli, B. intermedia, and B. alvinipulli. Poultry are also frequently colonized by putatively nonpathogenic species such as B. murdochii and B. innocens. We evaluated the differentiation of Brachyspira species by 3 methods: sequencing of the reduced nicotinamide adenine dinucleotide (NADH) oxidase gene (nox), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and a new multiplex (m)PCR targeting genes such as the tryptophanase A gene (tnaA) and the p-aminobenzoyl-glutamate hydrolase subunit B gene (abgB). Sequencing of 414 bp of the nox PCR amplification products generated from 41 pure cultures of avian Brachyspira isolates allowed presumptive species identification in 33 isolates with at least 99% identity in basic local alignment search tool analysis, including B. pilosicoli, B. intermedia, B. murdochii, B. innocens, and “B. pulli”. MALDI-TOF MS...

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Differentiation of Brachyspira spp. isolated from laying hens using PCR-based methods and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry:

The work described in this thesis was concerned with identifying the prevalence and risk factors associated with colonisation by the intestinal spirochaete Brachyspira pilosicoli in:
 
humans: long term residents of Perth, Western Australia (WA) and Indonesians either living temporarily in Perth or as long term residents in urban and rural areas of Bali, Indonesia,
 
animals: domestic animals including alpacas, birds, cattle, cats, chickens, dogs, doves, ducks, goats, horses, pigs, and sheep (housed at a wide variety of places around Perth), and a range of wild animals housed in various Zoos and wildlife centres in WA
 
This study shows that for humans:
 
• Brachyspira pilosicoli was significantly more prevalent in Indonesians of all sub groups, be they temporary residents of Perth (94% - 216 faecal samples from 180 individuals), or long term residents of Indonesia (126% - 992 faecal samples from 617 individuals) compared with long term residents of Australia living mainly in Perth (02% of 766 sampled), even in those with gastrointestinal complaints This suggests a relationship between a high prevalence of B pilosicoli and living in Indonesia;
• In Bali, B pilosicoli was significantly more prevalent in the impoverished urban area of Sesetan (203-234%) where the husbandry of pigs is poor and effluent treatment is non-existent compared to four traditional farming villages (Badung, Karang Suwung, Melinggih, Payangan Desa) (33-226%) In the latter villages effluent and drainage is better and there is less likely to be contamination of drinking water
• There was no significant association between the presence of B pilosicoli and the presence of clinical symptoms including headaches, abdominal pains, diarrhoea, joint/muscular pain and constipation
• Amongst Indonesians living in Indonesia, there was no significant difference in the prevalence of B pilosicoli between people with and without contact with animals and between farmers and other occupational groups
• Indonesians visiting Perth who were positive for B pilosicoli originated from nine cities and five main islands in Indonesia This suggests that B pilosicoli is endemic throughout Indonesia
• Strain typing of isolates of B pilosicoli showed that they were genetically heterogenous and did not show any consistent pattern with respect to geographical location, family of origin or disease status Isolates from the same individual were sometimes unrelated, suggesting the probability of re-infection with another strain between the samplings
• Some households (~7%) had more than one member positive for B pilosicoli Strain analysis suggested transmission between family members, and this could be due to either faecal-oral transmission, or from a common external source, such as contaminated water
• B pilosicoli was cultured from only 02% of Australians This low prevalence may be a result of little or no exposure to B pilosicoli due to good personal hygiene and environmental sanitation
• B pilosicoli strain H1b and H171 that were isolated from healthy Indonesians were able to colonise mice and day-old chickens, and induced clinical signs of pasty faeces in the latter Histological sections showed mild typhlitis and typical end-on attachment of B pilosicoli to the caecal epithelial mucosa of the chickens This finding suggests that the human isolates had pathogenic potential
 
This study showed that for animals investigated:
 
• Intestinal spirochaetes were cultured from 464% (13/28) of bilbies with 143% (4/28) positive for B pilosicoli Spirochaetes were also cultured from the faeces of two Western Barred bandicoots and one (12%) kangaroo
• Intestinal spirochaetes were not isolated from any alpacas, cattle, goats, horses, pigs, and sheep but were detected in 405% of ducks, 143% of chickens, 149% of ostriches and 15% of cats
• Few pets that are commonly kept in households (dogs, cats and aviary birds) were colonised, suggesting that they are not an important focus of B pilosicoli infection in Australia

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Prevalence, risk factors and molecular epidemiology of Brachyspira pilosicoli in humans and animals

A hippurate-negative biovariant of Brachyspira pilosicoli (B. pilosicolihipp-) is occasionally isolated in diarrhoeic pigs in Finland, often concomitantly with hippurate-positive B. pilosicoli or Lawsonia intracellularis. We studied pathogenicity of B. pilosicolihipp- with special attention paid to avoiding co-infection with other enteric pathogens. Pigs were weaned and moved to barrier facilities at the age of 11 days. At 46 days, 8 pigs were inoculated with B. pilosicolihipp- strain Br1622, 8 pigs were inoculated with B. pilosicoli type strain P43/6/78 and 7 pigs were sham-inoculated. No signs of spirochaetal diarrhoea were detected; only one pig, inoculated with P43/6/78, had soft faeces from day 9 to 10 post inoculation. The pigs were necropsied between days 7 and 23 after inoculation. Live pigs were culture-negative for Brachyspira spp., but B. pilosicolihipp- was reisolated from necropsy samples of two pigs. The lesions on large colons were minor and did not significantly differ between the three trial groups. In silver-stained sections, invasive spirochaetes were detected in colonic mucosae of several pigs in all groups. Fluorescent in situ hybridisation for genus Brachyspira, B. pilosicoli and strain Br1622 was negative. However, in situ detection for members of the genus Leptospira was positive for spirochaete-like bacteria in the colonic epithelium of several pigs in both infected groups as well as in the control group. L. intracellularis, Salmonella spp., Yersinia spp. and intestinal parasites were not detected. The failure of B. pilosicoli strains to cause diarrhoea is discussed with respect to infectivity of the challenge strains, absence of certain intestinal pathogens and feed and management factors.

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Neither Hippurate-negative Brachyspira pilosicoli nor Brachyspira pilosicoli Type Strain Caused Diarrhoea in Early-weaned Pigs by Experimental Infection

K. Ahlsten

Papers

Neither Hippurate-negative Brachyspira pilosicoli nor Brachyspira pilosicoli Type Strain Caused Diarrhoea in Early-weaned Pigs by Experimental Infection