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PubMed1996

Identification of the swine pathogen Serpulina hyodysenteriae in rheas (Rhea americana).

Recently intestinal spirochetes were isolated from rheas in Ohio and Iowa with a necrotizing typhlocolitis. These intestinal spirochetes, strains R1 and NIV-1, were characterized and compared with other intestinal spirochetes, including strains of S. hyodysenteriae. Both rhea spirochetes were indole positive, strongly beta-hemolytic, grew under a 1% O2:99% N2 atmosphere, and were morphologically similar to spirochetes in the genus Serpulina. Analysis of rRNA gene restriction patterns (ribotypes), and immunoblots of whole cell proteins, indicated both spirochetes were similar to Serpulina hyodysenteriae strains from swine. Comparisons of nearly complete sequences (> 1458 bases) of the 16S rRNA gene of the two rhea spirochetes with S. hyodysenteriae strains confirmed that rhea spirochetes R1 and NIV-1 were strains of S. hyodysenteriae. These results indicate that S. hyodysenteriae has a broader host range than previously recognized.

Animals↗
PubMed1998

Identification of porcine Serpulina strains in routine diagnostic bacteriology.

Serpulina strains from pigs were identified using simple tests. The large size of S. hyodysenteriae in stainings from colonic contents and faeces was found useful for the presumptive differentiation of this major pathogenic species from the other Serpulinae. However, this morphological characteristic gets lost upon cultivation. The 'ring phenomenon' aided to confirm the strong haemolysis typical for S. hyodysenteriae. The weakly haemolytic species S. innocens, S. pilosicoli, S. intermedia and S. hyodysenteriae could be differentiated with the help of the indole spot test and two or four other simple enzymatic tests. Nearly half of the S. hyodysenteriae strains isolated in Belgium were indole-negative. Such strains have only rarely been reported earlier, and were absent among the strains from other European countries examined.

Animals↗
PubMed1999

Differentiation of Serpulina species by NADH oxidase gene (nox) sequence comparisons and nox-based polymerase chain reaction tests.

The NADH oxidase genes (nox) of 18 strains of intestinal spirochaetes were partially sequenced over 1246 bases. Strains examined included 17 representatives from six species of the genus Serpulina, and the type strain 513A(T) of the human intestinal spirochaete Brachyspira aalborgi. Sequences were aligned and used to investigate phylogenetic relationships between the organisms. Nox sequence identities between strains within the genus Serpulina were within the range 86.3-100%, whilst the nox gene of B. aalborgi shared between 78.8-83.0% sequence identity with the nox sequences of the various Serpulina strains. A phenogram produced based on sequence dissimilarities was in good agreement with the current classification of species in the genus Serpulina, although an atypical strongly beta-haemolytic porcine strain (P280/1), previously thought to be S. innocens, appeared distinct from other members of this species. Primer pairs were developed from the nox sequence alignments for use in polymerase chain reaction (PCR) identification of the pathogenic species S. hyodysenteriae (NOX1), S. intermedia (NOX2), and S. pilosicoli (NOX3), and for the combined non-pathogenic species S. innocens and S. murdochii (NOX4). The PCRs were optimised using 80 strains representing all currently described species in the genus Serpulina, as well as the type strain of B. aalborgi. Tests NOX1 and NOX4 specifically amplified DNA from all members of their respective target species, whilst tests NOX2 and NOX3 were less sensitive. NOX2 amplified DNA from all 10 strains of S. intermedia from pigs but from only 4 of 10 strains from chickens, whilst NOX3 amplified DNA from only 18 of 21 S. pilosicoli strains, even at low stringency. Tests NOX1 and NOX4 should prove useful in veterinary diagnostic laboratories, whilst NOX2 and NOX3 require further refinement.

Animals↗
PubMed1999

Sequence characterization of two new members of a multi-gene family in Serpulina hyodysenteriae (B204) with homology to a 39 kDa surface exposed protein: vspC and D.

Previous cloning and sequencing of clones from a genomic library constructed from Serpulina hyodysenteriae B204 had identified a tandem pair of open reading frames, identified as vspA and vspB (variable surface protein) expected to encode proteins with homology to ( but not identical with) a 39 kDa surface exposed membrane protein from this animal pathogen. Additional screening of the genomic library was performed to retrieve the remainder of the vspB gene using new oligonucleotide probes based upon the cloned gene sequences. Not only was this goal met but we also discovered two more adjacent and related vsp genes (vspC and vspD) and have completely sequenced them. They are all in a parallel orientation and appear to have a set of similar but distinct regulatory elements that may control separate expression of their open reading frames (ORFs). Thus, there are four contiguous vsp genes which are predicted to encode a family of structurally conserved proteins. The four adjacent open reading frames (ORFs) are of similar size (384-389 codons) and share from 83% to 90% identity in their amino acid sequence. Preliminary data suggests there may be yet another homologous gene copy in a distal location of S. hyodysenteriae that faithfully encodes the 39 kDa surface protein. The organization and homologies of these highly conserved multiple gene copies are discussed.

Amino Acid Sequence↗
PubMed1999

Emended descriptions of indole negative and indole positive isolates of Brachyspira (Serpulina) hyodysenteriae.

Two type/reference strains of Brachyspira (B.) hyodysenteriae, 14 Belgian and German indole negative, and 14 Belgian, German and Swedish indole positive field isolates of strongly beta-haemolytic intestinal spirochaetes were compared by pulsed-field gel electrophoresis (PFGE) patterns, biochemical reaction patterns, 16S rDNA sequences and MIC determinations of six antibacterial substances. Three tests for indole production, including a spot indole test, were compared with congruent results. All field isolates were classified as B. hyodysenteriae due to a high genetic and phenotypic similarity with the type strains. The Belgian and German indole negative isolates had identical and unique PFGE patterns for the tested restriction enzymes MluI and SalI, as well as identical 16S rDNA sequences, and they could not be differentiated by any of the methods used. Seven unique PFGE patterns were achieved from the 14 indole positive field isolates. The patterns were identical and unique for epidemiologically related isolates. Type/reference strains and isolates without known relation to other tested isolates showed unique banding patterns. The MICs of tylosin, tiamulin, erythromycin, clindamycin, carbadox and virginiamycin were determined in broth for all isolates. In contrast to Belgian and German isolates, the majority of the Swedish field isolates were susceptible to tylosin, erythromycin and clindamycin. Probable pathways of infection for some of the Swedish isolates were determined. The PFGE patterns of epidemic clones of B. hyodysenteriae remained stable for a period of up to 8 years. In vivo development of resistance to macrolide and lincosamide antibiotics due to use of tylosin was clearly indicated for two epidemic clones.

Animals↗
PubMed2000

Evaluation of a 23S rDNA polymerase chain reaction assay for identification of Serpulina intermedia, and strain typing using pulsed-field gel electrophoresis.

A polymerase chain reaction assay, amplifying a 1027 base pair portion of the 23S rDNA gene, was evaluated for identification of the intestinal spirochaete Serpulina intermedia. A total of 34 strains of S. intermedia isolated from pigs and chickens and 195 strains of other related species were tested. The optimised assay correctly identified all the S. intermedia strains, but generated 11 false positive reactions, giving a test sensitivity of 100% and a test specificity of 94.3%. The false positive reactions were generated from strains of four different species of intestinal spirochaetes, and the product was of the original predicted size. This suggests that the primer sites selected on the 23S rRNA gene were not completely specific for S. intermedia. Pulsed-field gel electrophoresis was then developed to investigate diversity amongst the S. intermedia strains. All strains tested had distinct DNA banding patterns using Mlu1, although three isolates from chickens on the same farm appeared closely related. The collection exhibited considerable genetic diversity, and strains from pigs and chickens were distributed in clusters throughout the dendrogram produced. The most closely related porcine and avian strains shared only 62% similarity.

Animals↗
PubMed2000

Influences of diet and vaccination on colonisation of pigs by the intestinal spirochaete Brachyspira (Serpulina) pilosicoli.

The purpose of this study was to determine whether methods used to control swine dysentery (SD), caused by the intestinal spirochaete Brachyspira (Serpulina) hyodysenteriae, would also be effective in controlling porcine intestinal spirochaetosis (PIS) caused by the related spirochaete Brachyspira (Serpulina) pilosicoli. Weaner pigs in Groups I (n=8) and II (n=6) received a standard weaner pig diet based on wheat and lupins, whilst Group III (n=6) received an experimental diet based on cooked white rice and animal protein. Pigs in Group II were vaccinated intramuscularly twice at a 3-week-interval with a formalinised bacterin made from B. pilosicoli porcine strain 95/1000 resuspended in Freund's incomplete adjuvant. Eleven days later pigs in all groups were infected orally with 10(10) cells of strain 95/1000 on three successive days. One control pig in Group I developed acute diarrhoea, and at post-mortem had a severe erosive colitis with end-on attachment of spirochaetes to the colonic epithelium. All other pigs developed transient mild diarrhoea and had moderate patchy colitis at post-mortem 3 weeks later. B. pilosicoli was isolated from the faeces of all pigs, except for one fed rice, and was isolated from the mesenteric nodes of three pigs from Group I and from one vaccinated pig in Group II. Consumption of the rice-based diet, but not vaccination, delayed and significantly (p<0.001) reduced the onset of faecal excretion of B. pilosicoli after experimental challenge. Vaccination induced a primary and secondary serological response to B. pilosicoli, as measured using sonicated whole cells of strain 95/1000 as an ELISA plate coating antigen. Antibody titres in the vaccinated pigs then declined, despite intestinal colonisation by B. pilosicoli. Both groups of unvaccinated animals also failed to develop a post-infection increase in circulating antibody titres.

Animal Feed↗
PubMed1997

Fusospirochetal superinfection of pre-existing oral lesion in patients with acquired immunodeficiency syndrome.

Three patients with AIDS presented with nonbleeding, painful, fetid, oral ulcers overlaid with a grayish-black semiadherent membrane at the sites of a pre-existing lesion. These lesions persisted despite treatment directed toward the primary etiology (cytomegalovirus, Kaposi's sarcoma). Gram- and Giemsa-stained smears of teased membrane fragments revealed an impressive bacterial flora with fusiforms and Borrelia-type spirochetes. Prompt treatment with penicillin brought amelioration of symptoms and sloughing of the overlaying membrane.

AIDS-Related Opportunistic Infections↗
PubMed1999

Spirochetal infection of the nervous system.

Neurologic infection is a characteristic feature of spirochetes. The neurologic manifestations of spirochetal infection are a source of continuing public concern: Lyme neuroborreliosis in endemic areas, neurosyphilis in HIV infected patients, and neuroborreliosis during outbreaks of relapsing fever. These are reviewed in this article. The techniques for diagnosis and recommendations in the management of these infections are also discussed.

Anti-Bacterial Agents↗
PubMed2000

Diarrhea in growing-finishing swine.

Regardless of the etiology of an enteric disease in nursery age to finisher swine, making a prompt and accurate diagnosis is crucial. Eliciting a complete history, assessing clinical signs and pathology, and selecting and interpreting laboratory tests are essential components in achieving this. Early detection and diagnosis of enteric disease is particularly critical in the nursery through finisher phase because of economic impacts. Recurrent topics when discussing control and prevention of enteric diseases are reducing stress and improving pig comfort and reducing or eliminating exposure through sanitation and biosecurity. These are not new concepts; in fact, prior to the advent of antimicrobials, they were the mainstay of treatment of enteric diseases. With concern over the use of antimicrobials in food animal production increasing, exploiting disease ecology to control enteric diseases is increasing in importance. New vaccines and bacterins for postweaning swine enteric diseases are needed tools to exploit the pig's immune system. Recent advances in diagnostic capabilities allow an increase in understanding and exploitation of disease ecology.

Animals↗
PubMed1995

DNA probe and polymerase chain reaction procedure for the specific detection of Serpulina hyodysenteriae.

Serpulina (Treponema) hyodysenteriae, a Gram-negative anaerobic spirochete, is the causative agent of swine dysentery, a mucohaemorrhagic diarrheal disease in which lesions are confined to the large intestine of pigs. A DNA probe and polymerase chain reaction (PCR) amplification procedures which are specific, rapid , and sensitive for the detection of S.hyodysenteriae have been developed. Clone pF12 from a plasmid library of S.hyodysenteriae B204 genomic DNA was identified as a clone specific for S.hyodysenteriae but not for S.innocens by differential hybridization screening with S.hyodysenteriae and S.innocens genomic DNA probes. A DNA probe consisting of a 1.3 kb restriction fragment from pF12 was found to be highly specific for S. hyodysenteriae and detected 10(5) bacterial cells. A PCR procedure using primers derived from this fragment yielded a single product which was specifically generated for S.hyodysenteriae template DNA and not for other control cells DNA. PCR provided increased sensitivity with the direct detection of as few as 10 S.hyrodysenteriae cells. The PCR procedure could detect S.hyodysenteriae cells in seeded faecal matter. Moreover the PCR assay was able to detect most S. hyodysenteriae field isolates of serotypes 8 and 9. These tools have diagnostic application in veterinary microbiology.

Animals↗
PubMed1999

Oral flora and pathogenic organisms.

Oral microbial flora consist of numerous bacterial taxa, ranging from aerobes through fastidious anaerobes, and fungi, viruses, and protozoa. Many of these bacteria are unique to the oral cavity. The organisms exist in a complex interrelationship that is regulated and maintained by physical and metabolic microbial interactions, and by environmental factors, such as saliva and diet. Many of these organisms are relatively harmless, although others are significant pathogens, producing local and systemic diseases in healthy and compromised individuals.

Actinomycetaceae↗
PubMed1993

The role of hemolysin(s) in the pathogenesis of Serpulina hyodysenteriae.

Serpulina (Treponema) hyodysenteriae, an anaerobic beta hemolytic spirochaete, is the etiologic agent of swine dysentery. Not much is known at present about the virulence factors of S. hyodysenteriae. However, the hemolysin production of this bacterium is generally accepted to be a virulence factor. To study the exact role of hemolysin production in the pathogenesis of swine dysentery, the gene encoding a hemolysin, tly, was cloned and its nucleotide sequence determined. After inactivation of this gene, the virulence of a tly-minus mutant in mice was tested. The mutant had reduced hemolysis indicating that the tly-encoded hemolysin was not the only hemolysin produced by S. hyodysenteriae. Mice infected with the tly-minus mutant had fewer cecal lesions than mice infected with the wild-type S. hyodysenteriae. It was concluded that the tly-encoded hemolysin might be an important virulence factor, but not the only one. Since it was demonstrated that spirochaetes can be transformed through electroporation, this has made a genetic approach to elucidate the pathogenesis of spirochaetal infections possible.

Animals↗
PubMed1998

Cooperative haemolysis between weakly-beta haemolytic human intestinal spirochaetes and Clostridium perfringens.

Interactions between human intestinal spirochaetes (HIS) related to intestinal spirochaetosis and intestinal pathogenic anaerobic bacteria were investigated by searching for the presence of cooperative haemolysis among 39 strains of weakly beta-haemolytic human intestinal spirochaetes and Clostridium perfringens alpha-toxin producers on plates carrying six different sheep blood agar media. An area of intense cooperative haemolysis (about 3-10 mm) was observed between all tested spirochaetal strains and C. perfringens where the clostridial alpha-toxin diffused toward the colonies of the spirochaetes overlapping part of their growth zone. The cooperative haemolysis was a potentiation of the haemolysis due to the single cultivation of human intestinal spirochaetes and C. perfringens and was observed after anaerobic incubation for 24-48 hours when both bacteria at a concentration range of 10(8)-10(3) CFU/ml were streaked at a distance of 3-10 mm to each other. A cooperative haemolysis was also observed between C. perfringens and weakly beta-haemolytic spirochaetes related to porcine and avian intestinal spirochaetosis and the spirochaete causing swine dysentery. The present study indicated that the damage produced in vitro by the clostridial alpha-toxin was enhanced only on the red blood cells which were in proximity to the HIS colonies causing the complete lysis of the erythrocytes. It is hence possible that the potentiation of the damage to red blood cells observed in vitro mimics an in vivo damage on the membranes of enterocytes to which HIS are attached when intestinal spirochaetosis occurs and when cytolysins similar to the alpha-toxin are available in the intestine of the host.

Animals↗
PubMed2003

The basis of persistent bacterial infections.

Selected bacterial pathogens, such as Helicobacter pylori and Mycobacterium tuberculosis, establish persistent infections in mammalian hosts despite activating inflammatory and antimicrobial responses. The strategies used to overcome host defense responses vary with the anatomical location of the infection but often rely on deliberate manipulations of the host cell responses. Phylogenetically unrelated bacteria can share similar strategies for the establishment of persistence and, in selected examples, one even can define homologous "persistence" genes. Such observations suggest that persistent infection is a specific phase in infection pathogenesis rather than a fortuitous imbalance in the host-pathogen interaction.

Animals↗
PubMed1996

Pericoronitis and tonsillitis: clinical and darkfield microscopy findings.

OBJECTIVE: Upper respiratory tract infections and tonsillitis and pharyngitis in particular increase the risk of lower third molar pericoronitis. The study was made to investigate clinical and microbiologic associations between pericoronitis and tonsillitis. STUDY DESIGN: The subjects were 20-year-old Finnish male conscripts. Thirty-eight patients had tonsillitis without and 33 patients had tonsillitis with a symptom-free erupting lower third molar; 27 patients had pericoronitis but healthy tonsils. All subjects were examined clinically. Bacterial samples were taken from healthy and diseased tonsils and from healthy and diseased third molar pericoronal pockets. The microbes were classified morphologically with the use of a darkfield microscope. RESULTS: Eighty-two percent of infected tonsils had deep crypts and 52% of them exudated pus compared with 36% (p < 0.001) and 0% (p < 0.001) in healthy tonsils, respectively. Infected lower third molars were less than half erupted and had on average 8.8 mm deep pericoronal pockets compared with pockets 5.5 mm deep recorded in symptom-free cases (p < 0.001). Spirochetes were more common in infected pockets than in symptom-free cases comprising 8.2% versus 3.0% of the total bacterial count (p = 0.044). Rods were more common in infected tonsils than healthy ones (8.9% versus 4.8%, p = 0.041). CONCLUSION: There were similarities in morphologic microbiota of pericoronitis and tonsillitis. However, clinical findings of tonsils did not appear to link with the findings of third molars.

Adult↗
PubMed2004

Fusospirochetosis causing necrotic oral ulcers in patients with HIV infection.

Under certain permissive circumstances, normally occurring fusiform bacteria and Borrelia spirochetes can result in a symbiotic overgrowth that leads to necrotic oral ulcers (stomatitis), gingivitis, and periodontitis. These lesions are collectively known as oral fusospirochetosis and may be under-appreciated in patients with HIV infection and AIDS. Fusospirochetal oral ulcers in patients with HIV are often large, necrotic, and malodorous; they respond completely to penicillin. We report 3 patients with HIV infection and fusospirochetal ulcerative stomatitis and review the clinical presentation, microbiologic diagnosis, potential pathogenesis, and treatment of these lesions.

AIDS-Related Opportunistic Infections↗
PubMed2003

Brachyspira hyodysenteriae contains eight linked gene copies related to an expressed 39-kDa surface protein.

A tandemly linked set of four open reading frames (ORFs), identified as vspA-D (variable surface protein) had been identified from previous cloning and sequencing of clones from a genomic library constructed from Brachyspira hyodysenteriae strain B204. The predicted translation products of these closely related genes were homologous to (but not identical with) a characterized 39-kDa surface-exposed membrane protein from this animal pathogen. Additional screening of the genomic library has been performed to retrieve what are believed to be additional vsp genes including the one expected to encode this 39-kDa protein. Four new vsp genes have been identified and found to be associated in a second set of four tandemly linked alleles. This new gene cluster of 7481 nucleotides is not adjacent to the original vspA-D gene cluster described but does appear to have arisen from a gene (region) duplication event. The new vsp genes (identified as vspE-H) are oriented parallel to one another and appear to have a set of similar but distinct regulatory elements that may control separate expression of their ORFs. The four adjacent ORFs are of similar size (361-390 codons) and share from 83% to 90% identity in their amino acid sequence. The organization and homologies of these highly conserved multiple gene copies are discussed.

Alleles↗