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To determine the incidence of Listeria monocytogenes in raw milk, an isolation method was evaluated and used to analyze milk from three areas of the United States. The incidence varied by area from 0% in California to 7% in Massachusetts, with an overall incidence of 4.2%. The highest incidence found in any area during a single sampling period was 12% in Massachusetts in March 1985. During that same sampling, the incidence for all Listeria species was 26%. Of the 27 L. monocytogenes strains isolated during the survey, 25 were pathogenic in adult mice. One of three Listeria ivanovii isolated was pathogenic. No other isolates demonstrated pathogenicity.

Enterotoxigenic Escherichia coli (ETEC) strains that produce multiple enterotoxins are important causes of severe dehydrating diarrhea in human beings and animals, but the relative importance of these enterotoxins in the pathogenesis is poorly understood. Gnotobiotic piglets were used to study the importance of heat-labile enterotoxin (LT) in infection with an ETEC strain that produces multiple enterotoxins. LT ؊ (⌬eltAB) and complemented mutants of an F4؉ LT ؉ STb ؉ EAST1 ؉ ETEC strain were constructed, and the virulence of these strains was compared in gnotobiotic piglets expressing receptors for F4 ؉ fimbria. Sixty percent of the piglets inoculated with the LT ؊ mutant developed severe dehydrating diarrhea and septicemia compared to 100% of those inoculated with the nalidixic acid-resistant (Nal r ) parent and 100% of those inoculated with the complemented mutant strain. Compared to piglets inoculated with the Nal r parent, the mean rate of weight loss (percent per hour) of those inoculated with the LT ؊ mutant was 67% lower (P < 0.05) and that of those inoculated with the complemented strain was 36% higher (P < 0.001). Similarly, piglets inoculated with the LT ؊ mutant had significant reductions in the mean bacterial colony count (CFU per gram) in the ileum; bacterial colonization scores (square millimeters) in the jejunum and ileum; and clinical pathology parameters of dehydration, electrolyte imbalance, and metabolic acidosis (P < 0.05). These results indicate the significance of LT to the development of severe dehydrating diarrhea and postdiarrheal septicemia in ETEC infections of swine and demonstrate that EAST1, LT, and STb may be concurrently expressed by porcine ETEC strains.

Although heat-stable (ST) and heat-labile (LT) enterotoxins produced by enterotoxigenic Escherichia coli (ETEC) have been documented as important factors associated with diarrheal diseases, investigations assessing the contributions of individual enterotoxins to the pathogenesis of E. coli infection have been limited. To address the individual roles of enterotoxins in the diarrheal؉ astA STb ؉ strain did not, although diarrhea developed in several piglets. The changes in the blood packed-cell volume and plasma total protein of gnotobiotic piglets inoculated with the LT-positive strains were significantly greater than those of pigs inoculated with the K88 astA/pBR322 strain (P ‫؍‬ 0.012, P ‫؍‬ 0.002). Immunochemistry image analysis also suggested that LT enhanced bacterial colonization in a gnotobiotic piglet model. This investigation suggested that LT is a major contributor to the virulence of K88؉ ETEC and that isogenic constructs are a useful tool for studying the pathogenesis of ETEC infection.Escherichia coli strains that colonize the small intestines, invade intestinal epithelial cells, and/or produce one or more toxins are important causes of diarrheal disease in both farm animals and humans. The virulence of enterotoxigenic E. coli (ETEC) is believed to be associated with the production of fimbrial adhesins and enterotoxins (1,19,35,36,51,54). Fimbrial adhesins mediate the attachment of bacteria to the surface of host epithelium cells and allow bacterial colonization. Fimbriae produced by different ETEC strains are quite diverse (21). In swine, ETEC strains that produce K88 (F4) or F18 are the most common currently associated with diarrheal diseases (19). These fimbriae apparently bind to glycoconjugates in the porcine enterocyte brush borders, and the absence of the respective glycoconjugate renders the animal resistant to bacterial colonization and consequent diarrheal diseases (14,15,20,48,49).Enterotoxins, including heat-stable enterotoxins (STa and STb) and heat-labile enterotoxin (LT) (23,25,39,45), have been found to disrupt intestinal fluid homeostasis and to cause hypersecretion of fluid and electrolytes through activation of adenylate cyclase (by LT) or guanylate cyclase (by STa) in small intestinal mucosal cells (26, 34). There are two major serogroups of LT found among E. coli strains: LT-I and LT-II. LT-I is associated with diarrheal diseases of both humans and animals, while LT-II is typically associated with diarrheal disease in animals. STs are small and monomeric molecules and may be associated with either human or animal disease (45, 55). STa and STb are the two classes of STs first recognized and differ from each other in both structure and enzyme activity (11,12). STa is produced by ETEC and other bacteria, while STb is found only associated with ETEC. A third ST, enteroaggregative E. coli (EAEC) EAST1, has been more recently identified. It is a plasmid-mediated enterotoxin, of low molecular weight and is frequently but not exclusively associated with EAEC isolated from children with p...

Cesarean-derived piglets were reared for 5 wk under germfree conditions or monoassociated with a benign Escherichia coli (G58-1) or a enterohemorrhagic strain (933D) derived from O157:H7, and immunized i.p. with the T-dependent (TD) Ags fluorescein-labeled (FL) keyhole limpet hemocyanin or trinitrophenylated (TNP) keyhole limpet hemocyanin and the type 2 T-independent Ags TNP-Ficoll or FL-Ficoll. Only colonized piglets showed an increase in serum IgG, IgA, and IgM and had serum Abs to FL, TNP, and colonizing bacteria. While serum Abs to FL or TNP appeared following colonization alone, secondary responses were restricted to piglets immunized using TD carriers. While animals colonized with 933D had significantly higher total serum IgG and IgM levels and specific IgG Abs than those colonized with G58-1, no differences were seen in serum IgA levels, B cell diversification in the ileal Peyer’s patches, and specific activity (ELISA activity per micrograms of Ig) of pre-boost serum IgG and IgM anti-TNP and anti-FL Abs. Serum IgA Abs to TNP, FL, or bacteria were not detected. Ag-driven responses, as measured by an increase in specific Ab activity, were only observed in secondary responses to TD Ags and to colonizing, pathogenic E. coli. We propose that germline-encoded, isotype-switched B cells in newborn piglets differentiate to Ab-secreting cells 1) after stimulation by bacteria-activated APCs or 2) through direct stimulation by bacterial products. We further propose that Ag-driven systemic responses require both bacterial colonization and TD Ags translocated to the peritoneum.

Abstract. Attaching and effacing Escherichia coli (AEEC) adhere to mucosal epithelium in both small and large intestine and induce a distinctive lesion characterized by an irregular scalloped appearance of the epithelial layer. Infection with attaching and effacing E. coli was detected in 14 calves, 7 pigs, 2 lambs, and 3 dogs. Affected animals were from farms and kennels in South Dakota, Minnesota, Iowa, Nebraska, and Wisconsin. Ages of affected animals were calves, 2 days to 4 months; pigs, l-6 weeks; lambs, 1 week; and dogs, 7-8 weeks. Clinical signs included diarrhea in all animals, but other nonenteric disease problems were present in some animals. Concurrent infection with other enteropathogens was detected in 9 calves and 5 pigs. Infection with AEEC appeared to be the sole cause of illness and death in some animals. There was evidence of intestinal hemorrhage in 5 of the calves and in all 3 dogs. Attaching and effacing lesions varied from small scattered foci to widespread involvement of large areas of intestinal mucosa. Verotoxin was produced by E. coli strains isolated from 9 calves, but not by strains from pigs, lambs, or dogs.

The matrix protein (M1) of influenza A virus is generally viewed as a key orchestrator in the release of influenza virions from the plasma membrane during infection. In contrast to this model, recent studies have indicated that influenza virus requires expression of the envelope proteins for budding of intracellular M1 into virus particles. Here we explored the mechanisms that control M1 budding. Similarly to previous studies, we found that M1 by itself fails to form virus-like-particles (VLPs). We further demonstrated that M1, in the absence of other viral proteins, was preferentially targeted to the nucleus/perinuclear region rather than to the plasma membrane, where influenza virions bud. Remarkably, we showed that a 10-residue membrane targeting peptide from either the Fyn or Lck oncoprotein appended to M1 at the N terminus redirected M1 to the plasma membrane and allowed M1 particle budding without additional viral envelope proteins. To further identify a functional link between plasma membrane targeting and VLP formation, we took advantage of the fact that M1 can interact with M2, unless the cytoplasmic tail is absent. Notably, native M2 but not mutant M2 effectively targeted M1 to the plasma membrane and produced extracellular M1 VLPs. Our results suggest that influenza virus M1 may not possess an inherent membrane targeting signal. Thus, the lack of efficient plasma membrane targeting is responsible for the failure of M1 in budding. This study highlights the fact that interactions of M1 with viral envelope proteins are essential to direct M1 to the plasma membrane for influenza virus particle release.The late phase of the influenza A virus replication cycle is marked by the occurrence of assembly and budding at the plasma membrane of infected cells, which leads to the separation of virion and host cell membranes and ultimately results in the production of infectious virus particles. This critical step is a highly concerted process driven largely by protein-protein, protein-lipid, and protein-nucleic acid interactions (34, 40). It has been established for many years that four viral structural components, namely, the matrix protein (M1), hemagglutinin (HA), neuraminidase (NA), and M2, are actively involved in the assembly and budding process (34,35,40), although the identities of these inter-and intramolecular interactions and regulatory mechanisms for influenza A virus assembly and budding are unclear. It has also been suggested that interactions of M1 with various cytoplasmic tails (CTs) of HA, NA, and M2 are critical to drive the assembly and release of influenza A virions from the surface of infected cells (1,5,10,18,25,29,30,68). To date, these interactions have been largely speculative because direct interactions have been demonstrated only for M1 and M2 (5, 18, 29).Early investigations into the budding machinery of influenza A virus using vaccinia virus-and baculovirus-based expression systems indicated that M1 was the only viral protein absolutely required for the assembly of virus particles (14,15...