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Examination of 69 strains of Yersinia enterocolitica which represented 20 serotypes and nontypable isolates for HeLa cell infectivity by a roller tube technique that provided a quantitative index of infection showed that infectivity (index, greater than 3.50) was confined to strains of serotypes O:8, O:3, O:5, 27, O:9, O:1, O:1,2,3, O:2,3, O:4,32, and O:21. All strains that were HeLa positive were sucrose positive and negative for salicin, esculin, rhamnose, raffinose, melibiose, alpha-methylglucoside, and citrate. All HeLa-negative strains were either sucrose and salicin positive or were sucrose negative. Twenty-one strains were examined for virulence by the ability to produce guinea pig conjunctivitis and diarrhea in mice. Positive strains were limited to those that were HeLa positive and were autoagglutination positive and calcium dependent at 35 degrees C. There was no association between virulence and the ability to produce enterotoxin measured by the infant mouse assay. Loss of autoagglutinability and calcium dependency was accompanied by loss of virulence, but HeLa cell infectivity was unchanged. The results suggest that at least two properties are necessary for virulence: the presence of the V and W antigens, mediated by the same plasmid as autoagglutination and calcium dependency, and an invasive factor demonstrated in vitro by HeLa cell infectivity. These virulence properties are found in only certain biotypes of Y. enterocolitica.

The toroviruses, Berne virus (BEV) and Breda virus (BRV), are recognized pathogens of horses and cattle, respectively. Torovirus-like particles (TVLPs) that are immunologically related to BRV have been reported as etiological agents of gastroenteritis in humans. Of the toroviruses, only BEV has been shown to replicate in cell culture. Hence, these agents can be routinely detected only by electron microscopy (EM), although serological testing has been used as well. Our studies have provided supporting evidence that the TVLPs detected in the stool specimens of pediatric patients with gastroenteritis are human toroviruses. By EM, these particles are morphologically similar to BEV and BRV. Thin-section electron microscopy revealed that TVLPs contain toroidal-shaped nucleocapsids. Viruses purified from human fecal specimens agglutinate rabbit erythrocytes. BRV antiserum as well as convalescent sera from patients with gastroenteritis whose stools contain TVLPs were shown to contain antibodies that react with purified TVLPs as demonstrated by hemagglutination inhibition, immunoelectron microscopy, and immunoblotting. RNA extracted from partially purified TVLP preparations is amplifiable by RT-PCR using primers bracketing a 219-base region at the 3' end of the Berne virus genome. Sequence analysis of amplicons from five isolates showed a high degree of identity with the corresponding BEV sequence.

A homologous polyclonal antibody was produced in a rabbit to the 104-kilodalton (kDa) protein hemolysin ofActinobacilluspleuropneumoniae serotype 1 strain CM-5. In immunoblots, this antibody recognized a similar 104-kDa protein produced in culture supernatants by A. pleuropneumoniae serotypes 1 to 12 and taxon "Minor group" in addition to Pasteurella haemolytica, ActinobaciUus suis, and alpha-hemolysin-producing Escherichia coli (but only weakly in the latter two organisms). These results were reproduced by using a mouse monoclonal antibody to the CM-5 104-kDa protein hemolysin, except that the monoclonal antibody bound more strongly to the alpha-hemolysin produced by E. coli, only weakly to the 104-kDa protein produced by "Minor group," and not at all to any extracellular antigens produced by A. suis. Pigs experimentally infected with A. pleuropneumoniae serotypes 1 to 10 and A. suis produced an antibody that recognized the 104-kDa hemolysin produced by CM-5. A pig challenged with a "Minor group" strain did not have such antibodies. Rabbit antiserum produced against the leukotoxin of P. haemolytica and alpha-hemolysin-producing E. coli also recognized the CM-5 hemolysin, but the latter only weakly. The hemolytic activity produced by CM-5 in culture supernatant was neutralized strongly by the pig serum to serotypes 1, 2, 5, 6, 9, and 10 and A. suis, only partially by serotype 8 antiserum and the rabbit antiserum to P. haemolytica leukotoxin, and not at all by the antiserum to serotypes 3, 4, and 7 and "Minor group" and the E. coli alpha-hemolysin. These results indicate that a similar but not identical 104-kDa protein is produced in vitro and in vivo by all serotypes of A. pleuropneumoniae and may be related to cytolysins produced by other gram-negative bacteria.

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