The nucleotide sequence of a chromosomally encoded antigen-expressing gene of Borrelia coriaceae was determined and used as a target for the polymerase chain reaction (PCR). Two primer sets were designed specifying the amplification of 269- and 701-bp DNA fragments. Primer set I, producing the short amplicon, was tenfold more sensitive than primer set II. As little as 10 fg of purified B coriaceae DNA could consistently be detected. The PCR assays, containing controlled numbers of whole spirochetes, allowed detectable amplification of 2 to 10 organisms. An internal, nonradioactively labeled gene-specific probe verified specificity of the PCR amplicons. Neither primer set cross-reacted with other related spirochetes. This PCR assay was adapted and found suitable for identification of B. coriaceae in biological samples, such as blood and thymus. Evidence for presence of B. coriaceae in biological samples was not found in tissue samples obtained from experimentally infected cows and their fetuses. These data failed to establish a definite association between B. coriaceae and epizootic bovine abortion.

Genus-, subspecies-, and serotype 1-specific antigens of Chlamydia psittaci were characterized by immunoblot analysis, using monoclonal antibodies that recognize 2 C psittaci strains: AB7 isolated from an ewe that had aborted, and iB1 isolated from feces of a healthy ewe. Genus-specific epitopes were detected on lipopolysaccharide, on a 47-kd protein, and on a 27- to 30-kd doublet. Subspecies-specific epitopes were located on a 30-kd protein, and a 80- to 90-kd protein region was identified, which bore subspecies- and serotype 1-specific epitopes. These 80- to 90-kd proteins were highly reactive with serum from ewes that had aborted and could be a useful antigen for diagnosis of chlamydial induced abortion of ruminants.

The role of porcine parvovirus (PPV) in inducing reproductive failure in swine has been extensively documented. However, information is not available as to the risk of ppv transmission by embryo transfer. Using the polymerase chain reaction (PCR) technique, PPV-specific DNA was detected in association with 4-day-old porcine embryos incubated in vitro in the presence of NADL-8 strain of PPV, despite attempts to rid the embryos of virus by either washing or treatment with pronase or trypsin. The presence of PPV in embryos collected from acutely infected swine was not detected by PCR, although PPV DNA was detected in the proximal portion of the reproductive tract during the early stages of infection. Viral-specific nucleic acid was not detected in embryos transferred from infected donors to seronegative recipients and retrieved and assayed on the 15th and 32nd days of gestation. Results of the use of PCR to detect PPV associated with swine female reproductive tract and embryos ascribe minimal risk to the transmission of PPV to seronegative recipients through embryo transfer.

We used monoclonal antibodies and immunohistologic examination of lymph nodes, to elucidate the pathogenesis of lymphosarcoma induced by, infection with bovine leukemia virus (BLV). The superficial cervical lymph nodes from 3 BLV-infected but apparently healthy sheep and 5 sheep with full-blown lymphosarcoma were examined. We also investigated the integration of bovine leukemia provirus by use of Southern blotting. In lymph nodes from sheep lacking clinical signs of infection, in which the provirus had been integrated at multiple sites in the genome, many large hypertrophic follicles were observed in the cortex. These follicles had germinal centers consisting of CD4+T cells and B cells that expressed surface IgM (sIgM) and major histocompatibility complex (MHC) class-II antigens, but not B cell-specific B2 molecule. The percentage of CD4+T cells in the cortex was significantly (P < 0.05) higher than that of the controls and sheep with lymphosarcoma. In all sheep with lymphosarcoma, the lymph nodes were completely destroyed by proliferating neoplastic cells, and in addition, small atrophic follicles, which consisted of normal B-cell marker-positive cells, were seen near the trabecula and the subcapsule. In these instances, neoplastic cells appeared to be a monoclonal population derived from a single CD5- B-cell lineage and to be classified as 2 types, CD5-CD4-CD8-B2+MHC class-II+sIgM+ and CD5-CD4-CD8-B2+MHC class-II+sIgM-. Moreover, CD8+T cells infiltrated diffusely throughout the tumorous lymph nodes apart from the atrophic follicles, and CD4+T cells were observed around atrophic follicles. Both types of T cells were small-size, normal lymphocytes with round and noncleaved nuclei, and were apparently non-neoplastic cells. In fact, after separation by use of a panning method, it seems that, in blood mononuclear cells from BLV-infected sheep without clinical signs of infection, but in lymphosarcomatous stages, the proviral genome was integrated only in B cells and not in T cells. Thus, we conclude that the host's immune response may be still maintained at a lymphosarcomatous stage.

Cell-mediated immunity was evaluated in intestinal, respiratory, and systemic lymphoid tissues of pigs exposed when 11 days old to virulent transmissible gastroenteritis virus (TGEV), attenuated TGEV, or porcine respiratory coronavirus (PRCV), 3 antigenically related porcine coronaviruses with distinct enteric and respiratory tissue tropisms. Mononuclear cells were prepared from mesenteric lymph nodes (MLN), bronchial lymph nodes (BLN), and spleens of pigs and tested for virus-specific responses by use of lymphocyte proliferation assays. Vigorous MLN and BLN proliferation responses to virulent TGEV and PRCV, respectively, at postinoculation days 8 to 24 were strongly associated with prior detection of TGEV in rectal swab samples and PRCV in nasal swab samples. Gastrointestinal disease and intestinal virus replication, assessed on the basis of rectal virus shedding, were almost exclusively found in the virulent TGEV-inoculated pigs, even though virulent TGEV and a high dose of attenuated TGEV elicited the highest proliferation responses in MLN. Pigs exposed to PRCV or attenuated TGEV did not have clinical signs of disease, and only 1 pig given a high dose of attenuated TGEV shed virus in feces. Porcine respiratory coronavirus replicated in the respiratory tract after either oronasal or aerosol inoculation of virus and induced strong BLN, but not MLN, proliferation responses. A high dose of attenuated TGEV (4 X 10(8) plaque-forming units) was more effective than a lower dose of attenuated TGEV (7 X 10(6) plaque-forming units) in eliciting significant lymphocyte proliferation in MLN and BLN. Cellular immune function, assessed on the basis of mitogen-induced proliferation of lymphocytes, was comparable for all 3 sources of lymphocytes and was not adversely affected by exposure to any of the pigs. The tissue tropism of TGEV and PRCV was associated with induction of virus-specific cell-mediated immune responses, as evidenced by substantial lymphocyte proliferation responses in MLN and BLN, mucosa-associated lymph nodes adjacent to the primary sites of virus replication. The failure of PRCV strain ISU-1 to replicate in the intestinal tract correlated with poor virus-specific cellular immune responses in MLN.

Sixteen IgA-deficient German Shepherd Dogs with small intestinal bacterial overgrowth were randomized into 2 groups. One group was fed a chicken-based kibble diet; the other was fed the same diet, but with 1% fructo-oligosaccharides supplemented at the expense of cornstarch. After being exposed to the diets for 46 to 51 days, the group that ate the supplemented diet had significantly (P = 0.04) fewer aerobic/facultative anaerobic bacterial colony-forming units in fluid from the duodenum/proximal part of the jejunum, as well as in the duodenal mucosa. We could not detect significant differences in the species of bacteria found in the intestine of these 2 groups of dogs. We conclude that at least some dietary carbohydrates can affect small intestinal bacterial populations in dogs with small intestinal bacterial overgrowth.

Fiberoptic bronchoscopy was performed in pigs to assess bacterial contamination of bronchoalveolar lavage fluids (BALF) obtained by use of the method and to determine the aerobic bacterial species in bronchoalveolar airways of healthy pigs. Bacterial contamination of BALF caused by insertion of the bronchoscope was evaluated, using a chromogenic bacterial tracer strain, and was found to be 0.22% of total colony-forming units (CFU), with range between 0 and 1.6%. A total of 164 pulmonary-healthy pigs from 6 closed herds were selected. The BALF obtained from these pigs were examined bacteriologically. Bacteria could not be isolated from 10.4% of all BALF; 5.5% of the BALF samples yielded pure cultures; and 84.1% yielded mixed aerobic bacterial growth. In BALF from 29.2% of the pigs, less than or equal to 5 X 10(2) CFU of bacteria/ml were isolated. The total number of bacteria in BALF from 50% of the pigs varied between 5 X 10(2) and 10(3) CFU/ml; 10.4% of BALF samples contained between 10(3) CFU/ml and 5 X 103 CFU/ml. More than 1 bacterial species were isolated from a single lung lavage of 84.1% of the pigs. Up to 6 species were isolated from a single BALF sample. A total of 443 bacterial isolates were differentiated into 25 bacterial genera and species. Samples of BALF yielded staphylococci (67.6%: Staphylococcus hyicus from 13.4% of the samples and S aureus from 2.4%), alpha-hemolytic streptococci (49.4%), Escherichia coli (42.1%), non-hemolytic streptococci (26.2%), Klebsiella spp (18.3%), micrococci (12.8%), and Coryneformes (11.0%). Other bacterial species were found, but less frequently. In our study, BALF from all pigs yielded < 5 X 103 CFU/ ml. Thus, low numbers of bacteria known to be facultative pathogens were isolated from BALF without causing detectable pneumonia.

Twenty-five quarters of 12 dairy cows, 3 to 8 years old, with a bacteriologic history of freedom from infection with Streptococcus uberis were inoculated via the teat canal with S uberis (23 quarters) or sterile medium (2 quarters). The cows were sent to slaughter 1, 3, or 6 days later. Acute inflammatory response involving accumulation of large numbers of polymorphonuclear, neutrophilic leukocytes (neutrophils) in the secretory acini was recognized after 24 hours in infected cows. After 6 days, the neutrophil response was still evident, but infiltration of septa by lymphocytes, septal edema, extensive vacuolation of secretory cells, focal necrosis of alveoli, small outgrowths of the secretory and ductular epithelium, and widespread hypertrophy of the ductular epithelium also were recognized. Early stages of involution and fibrosis also were evident at that stage. Streptococci were identified by immunoperoxidase labeling, free or phagocytosed, in macrophages; in the alveolar lumina, adherent to damaged secretory or ductular epithelium; in the subepithelium and septal tissue; and in lymphatic vessels and lymph nodes. The importance of the macrophage as the primary phagocytic cell is highlighted, and doubt is cast on the value of the exuberant neutrophil response by the host in defense of the gland.

Conjunctival swab specimens from healthy pigs were cultured to determine normal microbial population. Four commercial swine operations were selected for study. Pigs of 4 age groups were tested: nursing pigs, nursery pigs, feeder pigs, and sows. Swab specimens were taken from the conjunctival sac of each pig. Bacterial, fungal, and mycoplasmal growth was determined separately. Chlamydia sp was detected by use of an ELISA. Bacteria were recovered from 98% of specimens evaluated. alpha-Streptococcus sp (89%) was the most commonly recovered organism, followed by Staphylococcus epidermidis (39%) and Staphylococcus sp (39%). Mycoplasma sp was not detected in any of the specimens. Chlamydia sp was identified in 28% of all specimens evaluated. These results are similar to reports of normal conjunctival flora in other domestic animals.

Members of the genus Salmonella were identified in feces from horses, using the polymerase chain reaction (PCR) and genus-specific olignucleotide primers. Feces from healthy horses were determined to be culture-negative for Salmonella spp. Fecal samples were inoculated with known numbers of colony-forming units (CFU) of S anatum, S derby, S enteritidis, S heidelberg, S newport, and S typhimurium. The DNA was extracted from fecal samples and amplified by PCP, using genus-specific primers. Sensitivity of the assay extended to 10(3) CFU of Salmonella sp/g of feces, sensitivity of microbiologic culture with enrichment extended to 10(2) CFU of Salmonella sp/g of feces. Feces that were not inoculated with Salmonella spp were negative by the PCR. Detection of salmonellae in feces was possible, using the PCR, within 10 to 12 hours from the time of submission of samples.