Experimental infection of newly weaned pigs with human and porcine strains of Serpulina pilosicoli.

Cultures of Serpulina pilosicoli 95/1000, isolated from a pig with porcine intestinal spirochetosis (PIS), and S. pilosicoli WesB, isolated from an Aboriginal child with diarrhea, were used to infect 5-week-old newly weaned pigs. Four of 12 pigs infected with strain 95/1000 and 2 of 12 pigs infected with strain WesB became colonized and developed watery, mucoid diarrhea within 2 to 11 days postinfection. Affected pigs all had moderate subacute mucosal colitis, with gross and histological changes similar to those previously reported in both natural and experimentally induced cases of PIS. Silver-stained histological sections of the colon and cecum from affected pigs demonstrated spirochetes within dilated intestinal crypts, where they were associated with neutrophilic exocytosis and mucus secretion. Sections from one pig infected with strain 95/1000 showed large numbers of spirochetes attached by one end to the colonic epithelium, a feature consistent with PIS. This study confirms the role of S. pilosicoli in the etiology of PIS and provides evidence that S. pilosicoli strains of human origin have pathogenic potential in an animal model.     

Cloning and DNA sequence analysis of an immunogenic glucose-galactose MglB lipoprotein homologue from Brachyspira pilosicoli, the agent of colonic spirochetosis

Colonic spirochetosis (CS) is a newly emerging infectious disease of humans and animals caused by the pathogenic spirochete Brachyspira (formerly Serpulina) pilosicoli. The purpose of this study was to characterize an antigen that was recognized by antibodies present in sera of challenge-exposed pigs. The gene encoding the antigen was identified by screening a plasmid library of human B. pilosicoli strain SP16 (ATCC 49776) genomic DNA with hyperimmune and convalescent swine sera. The predicted amino acid sequence encoded by the cloned B. pilosicoli gene had a high degree of similarity and identity to glucose-galactose MglB lipoprotein. Located 106 bp downstream of the putative mglB gene was a 3'-truncated open reading frame with 73.8% similarity and 66.3% identity to mglA of Escherichia coli, suggesting a gene arrangement within an operon which is similar to those of other bacteria. A single copy of the gene was present in B. pilosicoli, and homologous sequences were widely conserved among porcine intestinal spirochetes Serpulina intermedia, Brachyspira innocens, Brachyspira murdochii, and the avian Brachyspira alvinipulli, but not in porcine Brachyspira hyodysenteriae, human Brachyspira aalborgi, and porcine Treponema succinifaciens. The deduced molecular weight of the mature MglB lipoprotein was consistent with expression by the cloned gene of a polypeptide with an apparent molecular weight of 36,000, as determined by Western blot analysis and [(3)H]palmitate labeling. Because mucin is the principal constituent of the colonic mucus gel and consists of glycoproteins that can serve as the substrate for growth and chemotaxis of B. pilosicoli in vitro, a role for MglB in mucosal localization of the spirochete appears consistent with the pathogenesis of CS. However, the presence of homologous sequences in closely related but nonpathogenic commensal spirochetes suggests that other virulence determinants may be required for pathogenesis.     

Immunoblot reactivity of polyclonal and monoclonal antibodies with periplasmic flagellar proteins FlaA1 and FlaB of porcine Serpulina species.

The periplasmic-flagellum (PF) proteins of Triton X-100-soluble and Triton X-100-insoluble sodium dodecyl sulfate-treated fractions from reference and field strains of Serpulina hyodysenteriae, Serpulina innocens, and Serpulina pilosicoli were characterized by Western blotting with a rabbit polyclonal antibody (PAb) specific for the 44-kDa PF sheath protein of S. hyodysenteriae (Z. Li, F. Dumas, D. Dubreuil, and M. Jacques, J. Bacteriol. 175:8000-8007, 1993) and a murine monoclonal antibody (MAb), designated 7G2, specific for the PF core FlaB proteins of S. hyodysenteriae. The MAb 7G2 reacted with a conserved epitope present in the 37-, 34-, and 32-kDa PF core FlaB proteins of all Serpulina species. This suggested that the core FlaB proteins are conserved among porcine Serpulina species. An immunoreactive band of approximately 44 kDa was present with all S. hyodysenteriae, S. innocens, and S. pilosicoli strains that were reacted with the PAb. The specificities of the PAb and the MAb for the FlaA1 and FlaB proteins of Serpulina species were confirmed by N-terminal amino acid sequencing of 44- and 37-kDa proteins, respectively, of S. hyodysenteriae and S. pilosicoli. Results from this study provide further evidence that the 44-kDa protein FlaA1 and the 37-, 34-, and 32-kDa FlaB proteins are conserved among porcine Serpulina species.     

Lipo-oligosaccharide profiles of Serpulina pilosicoli strains and their serological cross-reactivities.

The purpose of this study was to determine the presence of lipopolysaccharide-like material in the intestinal spirochaete Serpulina pilosicoli and the extent of antigenic cross-reactivity of this material in different strains of the species. Hot water-phenol, aqueous-phase extracts from five porcine and three human strains of S. pilosicoli, and from seven strains of four other Serpulina spp., were separated by SDS-PAGE and silver-stained profiles were obtained. All S. pilosicoli strains had a predominant band at c. 16 kDa. Some also had a partial ladder-like profile, consistent with the presence of semi-rough lipo-oligosaccharide (LOS); this was more obvious in Western immunoblot analysis. LOS from each S. pilosicoli strain was serologically distinct in immunoblot analysis and there was no cross-reactivity with other Serpulina spp. The serological diversity found amongst the LOS of S. pilosicoli strains may help to explain why individual people and animals can suffer repeated infections with different strains of the organism.     

PCR Amplification from Fixed Tissue Indicates Frequent Involvement of Brachyspira aalborgi in Human Intestinal Spirochetosis

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.     

Comparative prevalences of Brachyspira aalborgi and Brachyspira (Serpulina) pilosicoli as etiologic agents of histologically identified intestinal spirochetosis in Australia

DNA from gastrointestinal biopsy specimens from 28 Australian patients with histologic evidence of intestinal spirochetosis (IS) was subjected to PCRs to amplify segments of the 16S rRNA and NADH oxidase genes of Brachyspira aalborgi and Brachyspira (Serpulina) pilosicoli. B. aalborgi was identified in specimens from 24 (85.7%) patients and B. pilosicoli in those from 4 (14.3%) patients (2 of whom were also positive for B. aalborgi). For two patients, no product was amplified. This study demonstrates that B. aalborgi is much more commonly involved in histologically identified IS in Australian patients than is B. pilosicoli. This is the first report of amplification of B. pilosicoli DNA from humans with IS.     

Identification and characterization of Serpulina pilosicoli isolates recovered from the blood of critically ill patients.

The phenotypic and genetic characteristics of spirochetes isolated from the blood of one U.S. and six French patients with severe clinical disease or impaired immunity were examined. All spirochetes were anaerobic, weakly beta-hemolytic, positive for hippurate hydrolysis, and negative for beta-glucosidase activity. Cell lengths ranged from 4 to 8 microm, and each isolate had between 8 and 12 periplasmic flagella per cell. These features were consistent with the spirochetes' being Serpulina pilosicoli, the agent of intestinal spirochetosis. All isolates were positive in a PCR assay amplifying a portion of the S. pilosicoli 16S rRNA gene, and they all grouped with fecal isolates of S. pilosicoli in multilocus enzyme electrophoresis (MLEE). The blood isolates could be differentiated from each other by MLEE, although the U.S. and two French isolates were closely related. Apparently S. pilosicoli may translocate from the large intestine to establish spirochetemia. The clinical significance of this finding remains uncertain and requires further investigation.     

Diagnostic Examination of Human Intestinal Spirochetosis by Fluorescent In Situ Hybridization for Brachyspira aalborgi, Brachyspira pilosicoli, and Other Species of the Genus Brachyspira (Serpulina)

Human intestinal spirochetosis, characterized by end-on attachment of densely packed spirochetes to the epithelial surface of the large intestines as a fringe has been associated with the weakly beta-hemolytic spirochetes Brachyspira aalborgi and Brachyspira (Serpulina) pilosicoli. In this study, fluorescent in situ hybridization with oligonucleotide probes targeting 16S or 23S rRNA of B. aalborgi, B. pilosicoli, and the genus Brachyspira was applied to 40 sections of formalin-fixed, paraffin-embedded intestinal biopsy specimens from 23 Danish and 15 Norwegian patients with histologic evidence of intestinal spirochetosis. Five biopsy specimens from patients without intestinal spirochetosis and three samples from pigs with experimental B. pilosicoli colitis were examined as well. In addition, the 16S ribosomal DNAs of two clinical isolates of B. aalborgi were sequenced, and a PCR procedure was developed for the identification of B. aalborgi in cultures. The genotypic characteristics of the two clinical isolates showed very high (99.5%) similarity with two existing isolates, the type strain of B. aalborgi and a Swedish isolate. Hybridization with the Brachyspira genus-specific probe revealed a brightly fluorescing fringe of spirochetes on the epithelia of 39 biopsy specimens, whereas 1 biopsy specimen was hybridization negative. The spirochetes in biopsy specimens from 13 Danish and 8 Norwegian patients (55.3%) were identified as B. aalborgi. The spirochetes in the biopsy specimens from the other 17 patients hybridized only with the Brachyspira probe, possibly demonstrating the involvement of as-yet-uncharacterized Brachyspira spirochetes in human intestinal spirochetosis.     

Genetic and phenotypic characterization of intestinal spirochetes colonizing chickens and allocation of known pathogenic isolates to three distinct genetic groups.

Infection with intestinal spirochetes has recently been recognized as a cause of lost production in the poultry industry. Little is known about these organisms, so a collection of 56 isolates originating from chickens in commercial flocks in Australia, the United States, The Netherlands, and the United Kingdom was examined. Strength of beta-hemolysis on blood agar, indole production, API ZYM enzyme profiles, and cellular morphology were determined, and multilocus enzyme electrophoresis was used to analyze the extent of genetic diversity among the isolates. The results were compared with those previously obtained for well-characterized porcine intestinal spirochetes. The chicken isolates were genetically heterogeneous. They were divided into 40 electrophoretic types distributed among six diverse genetic groups (groups b to g), with a mean genetic diversity of 0.587. Strains in two groups (groups d and e) may represent new species of Serpulina, and the groups contained only strains isolated from chickens. Three genetic groups contained isolates previously shown to be pathogenic for chickens. These corresponded to the proposed species "Serpulina intermedius," to an unnamed group (group e), and to Serpulina pilosicoli. Two of the chicken isolates (one "S. intermedius" and one S. pilosicoli isolate) were strongly beta-hemolytic, two (both "S. intermedius") had an intermediate level of beta-hemolysis, and the rest were weakly beta-hemolytic. Fourteen isolates of "S. intermedius" produced indole, as did one isolate from group d. Isolates identified as S. pilosicoli resembled porcine isolates of this species, having four to six periplasmic flagella inserted subterminally in a single row at each end of the cell, and had tapered cell ends. All other spirochetes were morphologically similar, having seven or more periplasmic flagella and blunt cell ends. The identification of three genetic groups containing pathogenic isolates provides an opportunity for more detailed epidemiologic studies with these pathogens and for the development of improved diagnostic tests.     

Eosinophilic venulitis in the small intestines in a mouse model of late asthma

The allergen-unchallenged enteric lesions in late allergic asthma are largely unknown. To clarify this point, BALB/c mice were sensitized by ovalbumin (OVA)/aluminum adjuvant intraperitoneally two times (on days 0 and 10) and then challenged with OVA intranasally on day 14 (asthma group). Four days after the challenge, small intestinal lesions were examined. By this treatment, diarrhea was not observed in the asthma group. Compared to the controls with or without OVA sensitization and/or OVA challenge, the asthma group developed eosinophilic venulitis without an increase in mucosal mast cells in small intestines, whereas intestinal epithelial cells were relatively intact. A few numbers of interleukin (IL)-4⁺ and IL-5⁺ lymphoid cells were recognized in intestines in the asthma group, but not in the controls. Expression of vascular cell adhesion molecule-1 on venular endothelium and eotaxin-2⁺ eosinophils, but not epithelial cells, in intestines were detected in the asthma group, but not in the controls. Total IgE, OVA-specific IgE and eotaxin, and IL-5, but not interferon-γ, were produced systemically in the asthma group compared to the controls. The present study suggests that eosinophilic venulitis without mast cells in the intestine may be induced by the systemic, but not by local, helper T 2-type responses. In addition, eosinophilic venulitis in small intestines may be subclinical enteric lesions.     

Reduced virulence of Serpulina hyodysenteriae hemolysin-negative mutants in pigs and their potential to protect pigs against challenge with a virulent strain.

The role of the Serpulina hyodysenteriae hemolysin encoded by the tlyA gene in the pathogenesis of swine dysentery (SD) was studied. tlyA mutants of two S. hyodysenteriae strains (B204 and C5) were tested for virulence in pigs. None of the animals developed SD. However, after infection with wild-type strain B204 or C5, the incidence of SD was 100 or 60%, respectively. Thus, the tlyA-encoded hemolysin of S. hyodysenteriae is an important virulence factor in SD. The potential of these mutants to protect pigs against challenge with a virulent S. hyodysenteriae strain was also studied. After challenge with wild-type strain B204, 50% of pigs previously inoculated with the B204 tlyA mutant were protected, whereas all control pigs contracted SD. None of the pigs previously inoculated with the C5 tlyA mutant developed SD upon challenge with wild-type strain B204, whereas 40% of the control pigs developed SD in this experiment. Thus, previous colonization with S. hyodysenteriae tlyA mutants in pigs provides partial protection against challenge with a virulent S. hyodysenteriae strain.     

Characterization of two DNA probes specific for Serpulina hyodysenteriae.

Two DNA probes, one 1.1- and one 0.75-kb probe, specific for Serpulina hyodysenteriae were isolated from a genomic library generated from virulent S. hyodysenteriae 5380. These probes are highly specific and react with all S. hyodysenteriae strains tested. Under stringent conditions, the DNA probes did not react with the nonpathogenic species Serpulina innocens or with other species of enteric bacteria, including Escherichia coli. Both probes are able to detect S. hyodysenteriae in colony blot hybridizations, and when applied to fecal specimens, they can detect 10(4) S. hyodysenteriae cells in 0.1 g of seeded fecal matter. Both probes can detect S. hyodysenteriae in fecal specimens from swine with clinical signs of swine dysentery after experimental challenge and from swine from a herd with an acute outbreak of swine dysentery. These probes have application as a diagnostic tool in veterinary microbiology.     

Evaluation of blood culture systems for detection of the intestinal spirochaete Brachyspira (Serpulina) pilosicoli in human blood

The anaerobic intestinal spirochaete Brachyspira (Serpulina) pilosicoli has been isolated from the bloodstream of French patients by manual blood culture systems. The purpose of this study was to determine whether the automated and manual blood culture systems used in Australia are suitable for growth and detection of this organism. Strains of B. pilosicoli were added to human blood to give concentrations ranging from 1 x 10(4) to 1 x 10(1) spirochaetes/ml and 10-ml volumes were inoculated into the media. Three strains of B. pilosicoli grew slowly in all manual Hemoline and BBL Septi-Chek formulations tested. Subcultures taken between 2 and 10 days after inoculation yielded growth only after incubation for a further 5-8 days. Growth and automated detection were achieved in the BACTEC system with Anaerobic/F medium with or without Fastidious Organism Supplement. Minimum time to signal for nine strains varied between 5.6 and 14.9 days, with a minimum concentration of 10(1) spirochaetes/ml of blood being detected. None of nine strains gave a positive signal in the BacT/Alert system when FAN Anaerobic culture bottles were used; however, four strains were detected by subculture taken at 7 or 14 days after inoculation. When Anaerobic medium was used in the BacT/Alert system, two of three strains gave a signal and the other strain grew and was detected by subculture. Spirochaetaemias caused by B. pilosicoli may be unrecognised because detection time by the signal or subculture exceeds 5 days.     

Attaching and effacing adherence of Vero cytotoxin-producing Escherichia coli to rabbit intestinal epithelium in vivo.

Vero cytotoxin-producing Escherichia coli (VTEC) strains have been associated with sporadic cases and outbreaks of hemorrhagic colitis and with the hemolytic uremic syndrome in humans. Since adherence of enteric pathogens to epithelial surfaces is often a prerequisite for the subsequent delivery of bacterial enterotoxins and mucosal invasion, we evaluated intestinal adherence by 18 VTEC strains, which were of human origin and belonged to 10 distinct serotypes, 7 days after enteral challenge of rabbits. A total of 23 postweaning rabbits (body weight, 1 kg) were each fed 2 X 10(8) VTEC, and 5 rabbits were challenged with an equal number of fecal commensal E. coli strains as controls. Each rabbit was monitored daily for the development of diarrhea. At 7 days after infection the proximal jejunum, distal ileum, cecum, and proximal colon were removed from each rabbit and examined for the presence of adherent organisms under light microscopy, after Giemsa staining of Formalin-fixed secretions, and by transmission electron microscopy. Nonbloody diarrhea developed in 16 of 23 VTEC-infected rabbits in contrast to 0 of 5 infected with commensal E. coli strains (P less than 0.02). Organisms were adherent to surface epithelial cells in the ceca (20 of 23 rabbits), proximal colons (9 of 23), and distal ilea (6 of 23) of VTEC-infected rabbits. Intimate attaching and effacing binding of bacteria to intestinal epithelial cells, in regions where the normal microvillous membrane architecture had been disrupted, was observed under electron microscopy for VTEC of multiple serotypes. In contrast, no organisms were adherent to the jejuni. Adherence of organisms was not seen in any portion of the intestines of rabbits that were challenged with commensals. These findings indicate that multiple serotypes of VTEC demonstrate intimate attaching and effacing binding to rabbit enterocytes and colonocytes in vivo. In addition to bacterial binding in the ceca and colons, VTEC adhere to enterocytes in the distal small intestines of per orally infected postweaning rabbits.     

Efficacy of antimicrobial treatments and vaccination regimens for control of porcine reproductive and respiratory syndrome virus and Streptococcus suis coinfection of nursery pigs

Seventy-six, crossbred, porcine reproductive and respiratory syndrome virus (PRRSV)-free pigs were weaned at 12 days of age and randomly assigned to seven groups of 10 to 11 pigs each. Pigs in group 1 served as unchallenged controls. Pigs in groups 2 to 7 were challenged intranasally with 2 ml of high-virulence PRRSV isolate VR-2385 (10(4.47) 50% tissue culture infective doses per 2 ml) on day 0 of the study (30 days of age). Seven days after PRRSV challenge, pigs in groups 2 to 7 were challenged intranasally with 2 ml of Streptococcus suis serotype 2 (10(8.30) CFU/2 ml). Group 2 pigs served as untreated positive controls. Antimicrobial treatments included daily intramuscular injection with 66,000 IU of procaine penicillin G per kg of body weight on days 8 to 10 (group 3), drinking water medication with 23.1 mg of tiamulin per kg during days 8 to 10 (group 4), and daily intramuscular injection of 5.0 mg of ceftiofur hydrochloride per kg on days 8 to 10 (group 5). Vaccination regimens included two intramuscular doses of an autogenous killed S. suis vaccine (group 6) prior to S. suis challenge or a single 2-ml intramuscular dose of an attenuated live PRRSV vaccine (group 7) 2 weeks prior to PRRSV challenge. Mortality was 0, 63, 45, 54, 9, 40, and 81% in groups 1 to 7, respectively. Ceftiofur treatment was the only regimen that significantly (P < 0. 05) reduced mortality associated with PRRSV and S. suis coinfection. The other treatments and vaccinations were less effective. We conclude that ceftiofur administered by injection for three consecutive days following S. suis challenge was the most effective regimen for minimizing disease associated with PRRSV and S. suis coinfection.     

Experimental infection of broiler breeder hens with the intestinal spirochaete Brachyspira ( Serpulina ) pilosicoli causes reduced egg production

The pathogenic potential of the anaerobic intestinal spirochaetes Brachyspira ( Serpulina ) pilosicoli and Brachyspira innocens was evaluated in adult chickens. Thirty 17-week-old Cobb broiler breeder hens were individually caged in three groups of 10 birds. Control birds (group A) were sham inoculated with sterile broth medium. Birds in the other two groups (groups B and C) were inoculated, respectively, with an isolate of B. innocens or of B. pilosicoli . Birds were monitored daily, and killed at 41 weeks of age. Infection had no consistent effect on body weight gain, but inoculation with B. pilosicoli resulted in a transient increase in faecal water content. B. innocens infection had no effect on egg production, but B. pilosicoli infection caused a delayed onset of laying, and a highly significant reduction in egg production over the first 11 weeks of lay. This study confirms that B. pilosicoli can cause serious egg production losses in adult chickens, while B. innocens is not obviously pathogenic.     

Pathogenicity of porcine intestinal spirochetes in gnotobiotic pigs.

Twelve intestinal spirochete strains of porcine origin were characterized on the basis of their phenotypic properties, by multilocus enzyme electrophoresis, and by pathogenicity testing in gnotobiotic pigs. The spirochetes used included two strains of Serpulina hyodysenteriae (B204 and P18A), two strains of Serpulina innocens (B256 and 4/71), one strain from the proposed new genus and species "Anguillina coli" (P43/6/78), and seven non-S. hyodysenteriae strains recently isolated from United Kingdom pig herds with a history of nonspecific diarrhea and typhlocolitis. By multilocus enzyme electrophoresis, five of these were identified as S. innocens, one was identified as an unspecified Serpulina sp., and one was identified as "A. coli." S. hyodysenteriae B204 and P18A, "A. coli" P43/6/78 and 2/7, and three (22/7, P280/1, and 14/5) of the five S. innocens field isolates induced mucoid feces and typhlocolitis in gnotobiotic pigs. None of the other spirochetes produced clinical signs or large intestinal pathology in this model. The "A. coli" strains induced a more watery diarrhea, with lesions present more proximally in the large intestine, than did the other pathogenic spirochetes. S. innocens 22/7 was also tested for pathogenicity in hysterotomy-derived pigs that had previously been artificially colonized with a spirochete-free intestinal flora and shown to be susceptible to swine dysentery. Despite effective colonization, strain 22/7 did not produce any disease, nor was there any exacerbation of large intestinal pathology or clinical signs when pigs with an experimentally induced existing colitis caused by Yersinia pseudotuberculosis were superinfected with strain 22/7. Certain non-S. hyodysenteriae spirochetes are therefore capable of inducing disease in gnotobiotic pigs, but their role as primary or opportunistic pathogens in conventional pigs remains equivocal.     

Immediate hypersensitivity reaction to beta-lactoglobulin in the epithelium lining the colon of guinea pigs fed cows' milk

Isolated epithelia from the colons of guinea pigs fed cows' milk have been studied in vitro under short circuit current conditions. When challenged with beta-lactoglobulin (beta LG) from the basolateral side, a large, transient inward flowing current was recorded. Pharmacological and ion flux measurements indicated this was due to inappropriate chloride secretion. There was no effect of beta LG on normal epithelia. The effects of challenge with beta LG were abolished by pretreatment with indomethacin, a prostaglandin synthetase inhibitor. By exposing epithelia from water-drinking animals in vitro to serum from milk-drinking guinea pigs sensitivity to beta LG could be conferred.     

Porcine colonic lymphoglandular complex: Distribution,structure, and epithelium

Lymphoepithelium and cells specialized for uptake and transport of foreign matter are characteristic of antigen sampling organs, Including lymphoglandular complexes (LGCs). Distribution, his-tologic structure, and epithelial ultrastructure of co-Ionic lymphoglandular complexes were determined in 5- to 13-week-old pigs. LGCs averaged 1,231 in number per colon, displayed a characteristic distribution pattern, and were most evenly distributed in colons of older pigs. LGCs consisted of well-defined submucosal masses composed of lymphatic nodules and internodu-lar lymphoid tissue penetrated by radially branching extensions of mucosal glands. Epithelial diverticula of each LGC entered the submucosa as a group through a circular collar derived from the muscularis mucosae. LGC epithelium contained goblet cells, cuboidal and columnar enterocytes, enteroendocrine cells, individual and clustered intraepithelial leukocytes, and cells morphologically compatible with follicle-associated epithelial cells/M cells. We regard the colonic LGC as a distinct mucosal lymphoid organ and suggest a significant role for it in local and systemic immune responses. The porcine colonic LGC may serve as a model for the human LGC.     

Isolation of enterotoxigenic Bacteroides fragilis from humans with diarrhea.

Enterotoxigenic Bacteroides fragilis was isolated from stool specimens of 8 of 44 diarrheic individuals (ages, 4 months to 69 years). The individuals had watery diarrhea and intestinal cramping; and infants had hyperthermia, vomiting, and blood in the stools. No recognized enteric pathogens were detected in seven of the eight diarrheic individuals positive for enterotoxigenic B. fragilis. The bacterium produced an enterotoxin detectable in concentrated broth that supported bacterial growth. Fifteen adult rabbits with ligated ceca developed fatal enteric disease following intraileal injection with 5 x 10(9) CFU of enterotoxigenic B. fragilis. Conversely, eight control rabbits injected with nonenterotoxigenic B. fragilis remained clinically normal. As few as 5 x 10(3) CFU of enterotoxigenic B. fragilis caused fatal enteric disease in the rabbit model. Disease in rabbits was characterized by mucoid, often hemorrhagic, diarrhea. The bacterium colonized the caudal small intestine and the colon of the rabbits and caused moderate to severe necrotizing colitis. Enterotoxigenic B. fragilis is widespread in the intestinal tract of diarrheic humans and is enteropathogenic in adult rabbits with ligated ceca. Its possible role in the enteric disease complex merits further study.