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.

Responses of the skin over the dorsum to solar UV irradiation (2 hours/d for 6 consecutive days) were investigated in hairless descendants of Mexican hairless dogs. Assessment of skin color changes, using a spectrophotometer, indicated that luminance values began to decrease from the third, day of UV irradiation, reached the minimal value at 3 weeks, and almost recovered 12 weeks after completion of UV irradiation. The number of the dihydroxyphenylalanine-positive melanocytes increased significantly (P < 0.01) from the third day of UV irradiation, reached its maximal value at 2 weeks, and recovered to normal at 12 weeks after completion of UV irradiation. On the second day of UV irradiation, the epidermis became focally thick, with disarrangement of component cells that had degenerative changes. In addition, a few so-called sunburn cells with pyknotic nuclei were seen in the epidermis. On the third day of UV irradiation, apparent suntan reaction developed, and a large number of epithelial cell in the epidermis were heavily pigmented with melanin granules. At 12 weeks after completion of UV irradiation, the epidermis appeared almost normal. On the other hand, significant changes were not detected in the dermis throughout the study.

To differentiate Mycoplasma hyopneumoniae, the cause of mycoplasmal pneumonia in pigs, from M flocculare and M hyorbinis, an assay, using the polymerase chain reaction to amplify a segment of the 16S rRNA gene sequence, was developed. The assay was found to be useful for identification of field isolates, as well as for identification of laboratory-adapted strains. Amplification of DNA from M hyopneumoniae and M flocculare resulted in products of 200 and 400 base pairs, respectively. The DNA from M hyorbinis was not amplified. The assay was sensitive enough to detect as little as 1,000 genome equivalents of M hyopneumoniae and M flocculare DNA. Sensitivity was increased 100-fold by increasing the concentration of magnesium ion in the reaction buffer from 2 to 4 mM; however, DNA from M hyorbinis was also amplified under these conditions. The DNA from several walled bacteria and from other mycoplasmas was also tested, but none of these DNA samples was amplified, suggesting that the assay was specific for porcine mycoplasmas.

Alterations in the size and functions of porcine alveolar macrophages exposed to sublytic amounts of heat-labile, hemolytic cytotoxin produced by Actinobacillus pleuropneumoniae (App) serotype 1, strain HS54 into the culture medium were studied in vitro. Alveolar macrophages were sensitive to the cytotoxin; treatment of the macrophages with low concentrations of cytotoxin (0.016 hemolytic unit) resulted in severe, irreversible cell swelling. However, high doses of cytotoxin (2.0 hemolytic units) were required to cause substantial cell death, as indicated by the influx of propidium iodide into and release of lactate dehydrogenase from cells. Macrophages exposed to low, sublytic doses of cytotoxin failed to migrate toward chemoattractant, were unable to attach to glass, and failed to phagocytize optimally opsonized erythrocytes. Macrophages already attached to glass surfaces detached when exposed to sublytic doses of cytotoxin. The swelling and impairment of functions of alveolar macrophages observed in this study could not be attributed to endotoxic effects, because heat treatment of the cytotoxin preparation for 60 minutes at 60 C resulted in complete loss of cytotoxicity. We conclude that sublytic doses of heat-labile, hemolytic cytotoxic substances produced by App depress alveolar macrophage function at concentrations likely to develop in association with acute pulmonary infection with App. The Apx (A pleuropneumoniae Rtx toxins) exotoxins secreted by the bacteria into culture medium were considered responsible for the toxic activity of the cytotoxin preparation. The Apx of the App field strain used in this study were likely to be similar to those of serotype-1 reference strain (S4707). Analysis by use of DNA-DNA hybridization indicated that genomic DNA of the field strain contained sequences similar to those encoding structural protein of ApxI (apxIA) and ApxII (apxIIA) of the serotype-1 reference strain. Therefore, Apx produced by the field strain of App used in this study are likely to be of similar pathogenic importance worldwide.

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.