The present survey was undertaken to determine the mycoflora of broiler houses. Attempts were made to isolate and identify fungi in the dust, feed, litter and water from 21 broiler houses. A total of 166 isolates of fungi was identified as yeast spp. (44%), Aspergillus spp. (30.7%), Verticillium spp. (7.2%), Penicillium spp. (3.6%), Paecilomyces spp. (3.6%), Scopulariopsis spp. (3.0%), Cephalosporium spp. (3.0%), Chrysosporium spp. (2.4%), Cladosporium spp. (1.8%) and Absidia spp. (0.6%).

Up to 60% of cases of equine colitis have no known cause. To improve understanding of the causes of acute colitis in horses, we hypothesized that Clostridium perfringens producing enterotoxin (CPE) and/or beta2 toxin (CPB2) are common and important causes of severe colitis in horses and/or that C. perfringens producing an as-yet-undescribed cytotoxin may also cause colitis in horses. Fecal samples from 55 horses (43 adults, 12 foals) with clinical evidence of colitis were evaluated by culture for the presence of Clostridium difficile, C. perfringens, and Salmonella. Feces were also examined by enzyme-linked immunosorbent assay (ELISA) for C. difficile A/B toxins and C. perfringens alpha toxin (CPA), beta2 toxin (CPB2), and enterotoxin (CPE). Five C. perfringens isolates per sample were genotyped for the following genes: cpa, cpb, cpb2 consensus, cpb2 atypical, cpe (enterotoxin), etx (epsilon toxin), itx (iota toxin), netB (necrotic enteritis toxin B), and tpeL (large C. perfringens cytotoxin). The supernatants of these isolates were also evaluated for toxicity for an equine cell line. All fecal samples were negative for Salmonella. Clostridium perfringens and C. difficile were isolated from 40% and 5.4% of samples, respectively. All fecal samples were negative for CPE. Clostridium perfringens CPA and CPB2 toxins were detected in 14.5% and 7.2% of fecal samples, respectively, all of which were culture-positive for C. perfringens. No isolates were cpe, etx, netB, or tpeL gene-positive. Atypical cpb2 and consensus cpb2 genes were identified in 15 (13.6%) and 4 (3.6%) of 110 isolates, respectively. All equine C. perfringens isolates showed far milder cytotoxicity effects than a CPB-producing positive control, although cpb2-positive isolates were slightly but significantly more cytotoxic than negative isolates. Based on this studied population, we were unable to confirm our hypothesis that CPE and CPB2-producing C. perfringens are common in horses with colitis in Ontario and we failed to identify cytotoxic activity in vitro in the type A isolates recovered.

Pathogens causing significant respiratory disease in growing pigs include Porcine reproductive and respiratory syndrome virus, Porcine circovirus 2, swine influenza virus, porcine respiratory coronavirus, Mycoplasma hyopneumoniae, and Bordetella bronchiseptica. The objective of this research was to characterize the respiratory excretion of these pathogens by acutely infected pigs. Pigs were inoculated under experimental conditions with 1 pathogen. Samples were collected from the upper respiratory tract and exhaled air. All pathogens were detected in swabs of the upper respiratory tract, but only M. hyopneumoniae and B. bronchiseptica were detected in expired air from individually sampled, acutely infected pigs. These findings suggest either that the acutely infected pigs did not aerosolize the viruses or that the quantity of virus excreted was below the detection threshold of current sampling or assay systems, or both, at the individual-pig level.

Objective-To determine the susceptibility of strains of Pasteurella multocida subsp multocida isolated from lung specimens of pigs with pneumonia to 20 antimicrobials and to evaluate the emergence of resistance to those antimicrobials in Spain during the past 2 decades. Sample Population-63 isolates recovered from 1987 to 1988 and 132 isolates recovered from 2003 to 2004. Procedure-A broth microdilution method was used to determine minimal inhibitory concentration (MIC) range and values for MIC50 and MIC90. Resistance of a strain to an antimicrobial agent was determined by use of the breakpoint value when available. Results-Isolates were generally susceptible to penicillin, ampicillin, ceftiofur, gentamicin, apramycin, neomycin, spectinomycin, chlortetracycline, erythromycin, tilmicosin, enrofloxacin, and florfenicol, and most isolates were resistant to clindamycin, tylosin tartrate, and tiamulin regardless of the time period. A substantial increase in resistance to sulfachlorpiridazine, sulfadimethoxine, sulfathiazole, and trimethoprim-sulfamethoxazole was observed, and a minor increase in resistance to oxytetracycline was also detected. Several multiresistance patterns were observed, most frequently among isolates recovered in the 2003 to 2004 interval. Conclusions and Clinical Relevance-Ceftiofur, florfenicol, and enrofloxacin are recommended for treatment of infections caused by P multocida subsp multocida in Spain. Increased frequency of resistance to oxytetracycline and sulfonamide drugs may be a contraindication for their use.

To evaluate site-to-site variation within fecal pats from cattle with regard to detection of Escherichia coli O157 and determine the effect on the accuracy of prevalence estimates of assay of multiple samples collected from the same fecal pat. 120 freshly voided fecal pats collected from 2 beef feedlots. Procedures-5 samples were systematically collected from each fecal pat and analyzed for E coli O157 via selective preenrichment techniques, immunomagnetic separation, and biochemical tests. Presumptive isolates were definitively identified via agglutination assays and polymerase chain reaction techniques. Best estimators of prevalence were calculated from the distribution of E coli O157-positive samples per pat. Of the 120 fecal pats, 96, 13, 4, 2, 3, and 2 fecal pats had 0, 1, 2, 3, 4, and 5 E coli O157-positive samples, respectively. The greatest estimate of E coli O157 prevalence (20%) was achieved when all 5 samples were assessed; this estimate represented a 2.4- fold increase in prevalence, compared with that provided via analysis of 1 sample/pat (8.2%). Compared with assessment of 5 sites/pat, the relative sensitivity of detecting an E coli O157-positive fecal pat via analysis of 1 site/pat was 40.1%. Results suggest that estimates of E coli O157 prevalence derived from sampling of 1 location/pat are likely underestimates of the true prevalence of this pathogen in fecal pats (and by extension, cattle). Additional research is warranted to confirm these results in situations of high and low prevalence and across different feedlots.

Eight Holstein cows, 4 inoculated intracisternally in 1 quarter of the mammary gland with Escherichia coli and 4 noninfected controls, were administered ceftiofur sodium (3 mg/kg of body weight, IV, q 12 hours) for 24 hours, beginning at 14 hours after inoculation of infected cows. All challenge-exposed cows became infected, with mean +/- SEM peak log10 bacterial concentration in milk of 5.03 +/- 0.69 colony-forming units/ml. The infection resulted in systemic signs (mean peak rectal temperature, 41.5 +/- 0.3 C; anorexia; signs of depression) and local inflammation (mean peak albumin concentration in milk, 7.89 +/- 1.71 mg/ml). Ceftiofur was detectable in milk from all challenge-exposed cows, compared with only 1 of 4 noninfected cows, and the mean period after inoculation that ceftiofur was detectable in milk was longer (P < 0.05) in infected (147.7 +/- 27.5 hours) than noninfected cows (1.3 +/- 1.3 hours). However, maximal ceftiofur concentration attained in milk for all cows was 0.28 micrograms/ml, and was 0.20 micrograms/ml or less for all but 2 milk samples collected for 10 days after challenge exposure. Mean serum concentration of ceftiofur peaked at 1.0 +/- 0.3 micrograms/ml and 0.7 +/- 0.1 micrograms/ml for infected and noninfected COWS, respectively. After each ceftiofur dose, mean peak and trough concentrations of ceftiofur in serum did not differ between groups; however, concentration of ceftiofur in serum was higher at 7 hours after each dose in noninfected cows, suggesting more rapid clearance of the drug in infected cows. Ceftiofur was not detected in serum (< 0.05 micrograms/ml) of any cow at or after 120 hours following inoculation of infected cows. Storage of serum samples at -20 C for 3 weeks resulted in a 98.8% decrease in ceftiofur activity, compared with that in fresh serum samples. Eighty-seven percent of this loss occurred 30 minutes after mixing serum and ceftiofur; thus, about 13% of the original activity was lost in storage. Storage of milk samples under similar conditions did not result in loss of ceftiofur activity. Despite acute inflammation, the dosage of ceftiofur used in this trial would not result in drug concentrations in milk above FDA safe concentrations, or above the reported minimum inhibitory concentration for coliform bacteria.

We defined methods for use of luminol-dependent chemiluminescence (LDCL) and superoxide anion (O2-) production as parameters of the oxidative metabolism of neutrophils isolated from 1.5- to 5-week-old neonatal calves. We determined how variations in blood sample handling, agonist preparation, individual variability, and age of calves influenced the LDCL and O2- responses to certain agonists, and defined concentrations of soluble and particulate agonists that maximally stimulated the oxidative metabolism of bovine neutrophils. Oxidative responses, particularly LDCL, were characterized by marked dayto-day variability, differed greatly within and between calves, were partially age-dependent, and were partially dependent on the individual agonist. Superoxide anion production had substantially less variability. We compared the in vitro oxidative (LDCL and O2-) responses of neutrophils isolated from neonatal calves stimulated by defined concentrations of the agonists-latex, phorbol myristate acetate, calcium ionophore, and opsonized zymosan-with responses to formylated oligopeptides and zymosan-activated serum, and to live, dead, live opsonized, and dead opsonized Pasteurella haemolytica organisms. Opsonization of particulates, pathogenic or nonpathogenic, enhanced the LDCL and O2- responses of stimulated neutrophils although P haemolytica was a less potent stimulant of oxidative functions than were nonbiological agonists. We conclude that the generation of reactive oxygen species by bovine neutrophils in response to P haemolytica is highly dependent on the presence of opsonins and is greatly enhanced in live vs killed bacteria. Futhermore, the in vitro generation of reactive oxygen species, including O2- by stimulated neutrophils, may be of biologic importance if similar events occur in vivo, and could have a major role in the pathogenesis of the acute lung injury associated with pneumonic pasteurellosis.

The effect that topical administration of cyclosporine would have on the number and type of microorganisms isolated from the corneal surface of dogs with keratoconjunctivitis sicca was studied. Schirmer tear tests were performed on and corneal swab specimens were collected from 61 eyes of 31 dogs with keratoconjunctivitis sicca prior to and after 3, 6, and 12 months of treatment with cyclosporine. In eyes that responded to cyclosporine treatment (Schirmer tear test value increased by greater than or equal to 5 mm/min, compared with pretreatment value), the percentage of eyes from which bacteria were isolated after 3, 6, and 12 months of treatment was significantly (P < 0.001) less than the percentage from which bacteria were isolated prior to treatment. However, among eyes that did not respond to treatment, we did not detect a significant change over time in prevalence of bacteria or type of bacteria isolated. The percentage of eyes from which fungi were isolated decreased during treatment; however, the small number of eyes in which fungal culture results were initially positive precluded demonstration of a significant change. For all eyes, we did not detect any significant differences over time in the frequency with which specific bacterial genera were isolated, with the exception of beta-hemolytic Streptococcus spp. Opportunistic corneal infections were not detected even though none of the dogs received antibiotics. An increase in production of tears, which contain anti-infection proteins, was believed to be the primary factor responsible for the decrease in the percentage of eyes from which microorganisms could be isolated.

Mycobacterial culture was performed on colostrum, milk, and feces from 126 clinically normal cows of a single herd with high prevalence of Mycobacterium paratuberculosis infection. Thirty-six (28.6 degrees h) cows were determined to be shedding the organism in the feces. Of the 36 fecal culture-positive cows, M paratuberculosis was isolated from the colostrum of 8 (22.2%) and from the milk of 3 (8.3%). Cows that were heavy fecal shedders were more likely to shed the organism in the colostrum than were light fecal shedders.

A rifampicin-resistant Pasteurella haemolytica serotype 1 with 2 added plasmids was used as a colonization-challenge strain in calves to test the resistance to colonization elicited by vaccination. Nine calves were vaccinated with a tissue culture-derived P haemolytica serotype-1 vaccine which, in a prior study, had elicited a serotype-specific inhibition of nasal and tonsillar colonization by the homologous serotype under field conditions. The vaccinates and 9 nonvaccinated control calves were exposed by tonsillar instillation with the challenge strain. The P haemolytica were enumerated in nasal secretion and tonsil wash specimens collected biweekly for 3 weeks. Rifampicin-supplemented agar medium inhibited growth of other bacterial species in the specimens and, thus, increased the sensitivity of detection of the challenge P haemolytica by 100-fold. The challenge strain retained its plasmids during the period of colonization. Inhibition of colonization was evidenced by lower frequency of isolations and fewer isolations of the challenge strain from nasal secretion and tonsil wash specimens of the vaccinates than from those of the nonvaccinates.