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PCR amplification from fixed tissue indicates frequent involvement of Brachyspira aalborgi in human intestinal spirochetosis.

ABSTRACT: PCR procedures amplifying portions of the 16S rRNA and NADH oxidase genes of Brachyspira aalborgi and Serpulina pilosicoli were applied to DNA extracted from paraffin-embedded human colonic or rectal tissues from 30 Norwegian, Australian, and U.S. patients, 16 of whom had histologic evidence of intestinal spirochetosis (IS). B. aalborgi-specific sequences were identified by PCR in 10 of the IS patients (62.5%) but none of the others, while S. pilosicoli sequences were not detected in tissues from any patient. Direct sequencing of products from three of the positive samples provided further confirmation of the presence of B. aalborgi. B. aalborgi may be a more common cause of intestinal spirochetosis than has been previously thought.

SUBMITTER: Mikosza AS

PROVIDER: S-EPMC85046 | biostudies-literature | 1999 Jun

REPOSITORIES: biostudies-literature

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PCR amplification from fixed tissue indicates frequent involvement of Brachyspira aalborgi in human intestinal spirochetosis.

Mikosza A S AS La T T Brooke C J CJ Lindboe C F CF Ward P B PB Heine R G RG Guccion J G JG de Boer W B WB Hampson D J DJ

Journal of clinical microbiology 19990601 6

PCR procedures amplifying portions of the 16S rRNA and NADH oxidase genes of Brachyspira aalborgi and Serpulina pilosicoli were applied to DNA extracted from paraffin-embedded human colonic or rectal tissues from 30 Norwegian, Australian, and U.S. patients, 16 of whom had histologic evidence of intestinal spirochetosis (IS). B. aalborgi-specific sequences were identified by PCR in 10 of the IS patients (62.5%) but none of the others, while S. pilosicoli sequences were not detected in tissues fro ...[more]

PMID: 10325391

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Project description:Primary ovarian insufficiency (POI) is one of the major causes of female infertility associated with the premature loss of ovarian function in about 3.7% of women before the age of 40. This disorder is highly heterogeneous and can manifest with a wide range of clinical phenotypes, ranging from ovarian dysgenesis and primary amenorrhea to post-pubertal secondary amenorrhea, with elevated serum gonadotropins and hypoestrogenism. The ovarian defect still remains idiopathic in some cases; however, a strong genetic component has been demonstrated by the next-generation sequencing (NGS) approach of familiar and sporadic POI cases. As recent evidence suggested an oligogenic architecture for POI, we developed a target NGS panel with 295 genes including known candidates and novel genetic determinants potentially involved in POI pathogenesis. Sixty-four patients with early onset POI (range: 10-25 years) of our cohort have been screened with 90% of target coverage at 50×. Here, we report 48 analyzed patients with at least one genetic variant (75%) in the selected candidate genes. In particular, we found the following: 11/64 patients (17%) with two variants, 9/64 (14%) with three variants, 9/64 (14%) with four variants, 3/64 (5%) with five variants, and 2/64 (3%) with six variants. The most severe phenotypes were associated with either the major number of variations or a worse prediction in pathogenicity of variants. Bioinformatic gene ontology analysis identified the following major pathways likely affected by gene variants: 1) cell cycle, meiosis, and DNA repair; 2) extracellular matrix remodeling; 3) reproduction; 4) cell metabolism; 5) cell proliferation; 6) calcium homeostasis; 7) NOTCH signaling; 8) signal transduction; 9) WNT signaling; 10) cell death; and 11) ubiquitin modifications. Consistently, the identified pathways have been described in other studies dissecting the mechanisms of folliculogenesis in animal models of altered fertility. In conclusion, our results contribute to define POI as an oligogenic disease and suggest novel candidates to be investigated in patients with POI.

Project description:BackgroundThe anaerobic spirochete Brachyspira pilosicoli colonizes the large intestine of various species of birds and mammals, including humans. It causes "intestinal spirochetosis", a condition characterized by mild colitis, diarrhea and reduced growth. This study aimed to sequence and analyse the bacterial genome to investigate the genetic basis of its specialized ecology and virulence.Methodology/principal findingsThe genome of B. pilosicoli 95/1000 was sequenced, assembled and compared with that of the pathogenic Brachyspira hyodysenteriae and a near-complete sequence of Brachyspira murdochii. The B. pilosicoli genome was circular, composed of 2,586,443 bp with a 27.9 mol% G+C content, and encoded 2,338 genes. The three Brachyspira species shared 1,087 genes and showed evidence of extensive genome rearrangements. Despite minor differences in predicted protein functional groups, the species had many similar features including core metabolic pathways. Genes distinguishing B. pilosicoli from B. hyodysenteriae included those for a previously undescribed bacteriophage that may be useful for genetic manipulation, for a glycine reductase complex allowing use of glycine whilst protecting from oxidative stress, and for aconitase and related enzymes in the incomplete TCA cycle, allowing glutamate synthesis and function of the cycle during oxidative stress. B. pilosicoli had substantially fewer methyl-accepting chemotaxis genes than B. hyodysenteriae and hence these species are likely to have different chemotactic responses that may help to explain their different host range and colonization sites. B. pilosicoli lacked the gene for a new putative hemolysin identified in B. hyodysenteriae WA1. Both B. pilosicoli and B. murdochii lacked the rfbBADC gene cluster found on the B. hyodysenteriae plasmid, and hence were predicted to have different lipooligosaccharide structures. Overall, B. pilosicoli 95/1000 had a variety of genes potentially contributing to virulence.Conclusions/significanceThe availability of the complete genome sequence of B. pilosicoli 95/1000 will facilitate functional genomics studies aimed at elucidating host-pathogen interactions and virulence.

Dataset Information

PCR amplification from fixed tissue indicates frequent involvement of Brachyspira aalborgi in human intestinal spirochetosis.

Publications

PCR amplification from fixed tissue indicates frequent involvement of Brachyspira aalborgi in human intestinal spirochetosis.

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