Objective—To evaluate the role of the nuclear factor-κB (NF-κB) in the response of bovine monocytes to exposure to Mycobacterium avium subsp paratuberculosis (MAP).
Sample Population—Monocytes from healthy adult Holstein cows that were known to be negative for MAP infection.
Procedures—Monocytes were incubated with MAP organisms with or without a specific inhibitor of the NF-κB pathway (pyrrolidine dithiocarbamate), and activation of the NF-κB pathway was detected by use of an electrophorectic mobility shift assay. The capacities of monocytes to express tumor necrosis factor (TNF)-α, interleukin (IL)-10, and IL-12; to acidify phagosomes; to phagocytize and kill MAP organisms; and to undergo apoptosis were evaluated.
Results—Addition of MAP organisms to monocytes activated the NF-κB pathway as indicated by increased NF-κB–DNA binding. Addition of pyrrolidine dithiocarbamate prevented nuclear translocation of NF-κB, decreased expression of TNF-α and IL-10, and increased IL-12 expression. Treatment of MAP-exposed monocytes with pyrrolidine dithiocarbamate increased the rate of apoptosis but failed to alter phagosome acidification, organism uptake, or organism killing by those cells.
Conclusions and Clinical Relevance—Results indicated that NF-κB rapidly translocated to the nucleus after exposure of bovine monocytes to MAP organisms. These data suggest that NF-κB is involved in initiation of inflammatory cytokine transcription and inhibition of apoptosis but that it is not directly involved in phagosome acidification or organism killing.
Objective—To assess effects of disease severity, sampling instrument, and processing technique on extracted DNA yield and detection rate for feline herpesvirus-1 (FHV-1) via PCR assay.
Sample Population—Crandell-Rees feline kidney (CRFK) cells grown in vitro and conjunctival samples from 40 eyes of 20 cats.
Procedures—Samples of CRFK cells (collected by use of a swab or cytology brush, with or without suspension in PBS solution) underwent DNA extraction; DNA yield was quantified spectrophotometrically. In affected cats, signs of herpetic disease were subjectively assessed. Conjunctival swab and brush samples were collected bilaterally for measurement of DNA concentration; a defined mass (DM) of DNA and defined volume (DV) of sample were assessed for FHV-1 via PCR assays.
Results—For CRFK cells, DNA yields from unsuspended swabs and brushes were greater than for suspended swabs and brushes; suspended swab samples yielded less DNA than suspended brush samples. For conjunctival samples, DNA yields from swabs were greater than for brushes. Clinical score was not correlated with double-stranded DNA yield collected via either sampling instrument; however, cats with FHV-1–positive assay results had higher clinical scores than cats with FHV-1–negative results. Detection of FHV-1 in swab and brush samples was similar. Double-stranded DNA yield and FHV-1 detection were inversely related via DM-PCR assay. The DV-PCR assay had a significantly higher FHV-1 detection rate than the DM-PCR assay.
Conclusions and Clinical Relevance—The DV-PCR assay of DNA extracted from an unsuspended swab sample was the preferred method for assessment of conjunctival shedding of FHV-1 in cats.
Objective—To determine whether soil concentrations of total or virulent Rhodococcus equi differed among breeding farms with and without foals with pneumonia caused by R equi.
Sample Population—37 farms in central Kentucky.
Procedures—During January, March, and July 2006, the total concentration of R equi and concentration of virulent R equi were determined by use of quantitative bacteriologic culture and a colony immunoblot technique, respectively, in soil specimens obtained from farms. Differences in concentrations and proportion of virulent isolates within and among time points were compared among farms.
Results—Soil concentrations of total or virulent R equi did not vary among farms at any time point. Virulent R equi were identified in soil samples from all farms. Greater density of mares and foals was significantly associated with farms having foals with pneumonia attributable to R equi. Among farms with affected foals, there was a significant association of increased incidence of pneumonia attributable to R equi with an increase in the proportion of virulent bacteria between samples collected in March and July.
Conclusions and Clinical Relevance—Results indicated that virulent R equi were commonly recovered from soil of horse breeding farms in central Kentucky, regardless of the status of foals with pneumonia attributable to R equi on each farm. The incidence of foals with pneumonia attributable to R equi can be expected to be higher at farms with a greater density of mares and foals.
Objective—To determine the adsorptive capability of di-tri-octahedral smectite (DTOS) on Clostridium perfringens alpha, beta, and beta-2 exotoxins and equine colostral antibodies.
Sample Population—3 C perfringens exotoxins and 9 colostral samples.
Procedures—Alpha, beta, and beta-2 exotoxins were individually co-incubated with serial dilutions of DTOS or bismuth subsalicylate, and the amount of toxin remaining after incubation was determined via toxin-specific ELISAs. Colostral samples from healthy mares were individually co-incubated with serial dilutions of DTOS, and colostral IgG concentrations were determined via single radial immunodiffusion assay.
Results—Di-tri-octahedral smectite decreased the amount of each C perfringens exotoxin in co-incubated samples in a dose-dependent manner and was more effective than bismuth subsalicylate at reducing exotoxins in vitro. Decreases in the concentration of IgG were detected in samples of colostrum that were combined with DTOS at 1:4 through 1:16 dilutions, whereas no significant decrease was evident with DTOS at the 1:32 dilution.
Conclusions and Clinical Relevance—Di-tri-octahedral smectite effectively adsorbed C perfringens exotoxins in vitro and had a dose-dependent effect on the availability of equine colostral antibodies. Results suggested that DTOS may be an appropriate adjunctive treatment in the management of neonatal clostridiosis in horses. In vivo studies are necessary to fully assess the clinical efficacy of DTOS treatment.
Objective—To estimate seroprevalence of bluetongue virus (BTV) and the geographic distribution of seropositive cattle herds in Illinois and western Indiana.
Sample Population—10,585 serum samples obtained from cattle in 60 herds during 3 transmission seasons (2000 through 2002).
Procedures—In a longitudinal study, serum samples were tested for BTV antibodies by use of a competitive ELISA. Four geographic zones were created by use of mean minimum January temperature. A multivariable mixed-effects logistic regression model with a random effect for herd was used to estimate seropositive risk for zone, age of cattle, herd type, and transmission season.
Results—Overall, BTV antibodies were detected in 156 (1.5%) samples. Estimated seroprevalence in 2000, 2001, and 2002 was 1.49%, 0.97%, and 2.18%, respectively. Risk of being seropositive for BTV was associated with geographic zone and age. Seroprevalence increased progressively from northern to southern zones, with no evidence of BTV infection in the northernmost zone. In the southernmost zone, annual seroprevalence ranged from 8.65% to 11.00%. Adult cattle were 2.35 times as likely as juvenile cattle to be seropositive.
Conclusions and Clinical Relevance—Overall seroprevalence was lower than has been reported for Illinois cattle. Bluetongue virus antibodies were distributed heterogeneously in this region. Only in the southernmost zone was seroprevalence consistently > 2%. Regionalization of BTV risk based on state borders does not account for such variability. Serologic data could be combined with landscape, climate, and vector data to develop predictive models of BTV risk within transitional regions of the United States.
Objective—To determine the efficacy of tinidazole for treatment of cats with experimentally induced Tritrichomonas foetus infection.
Animals—8 specific-pathogen-free kittens.
Procedures—Tinidazole was tested for activity against a feline isolate of T foetus in vitro. Kittens were infected orogastrically with the same isolate and treated or not with tinidazole (30 mg/kg, PO, q 24 h for 14 days). Amoxicillin was administered 28 weeks after completion of tinidazole administration to induce diarrhea. Feces were repeatedly tested for T foetus by use of PCR assay and microbial culture for 33 weeks.
Results—Tinidazole killed T foetus at concentrations ≥ 10 μg/mL in vitro. In experimentally induced infection, tinidazole administered at 30 mg/kg decreased T foetus below the limit of molecular detection in 2 of 4 cats. Recrudescent shedding of T foetus, as elicited by amoxicillin-induced diarrhea, was diminished in cats that received prior treatment with tinidazole.
Conclusions and Clinical Relevance—Although tinidazole decreased the detection of T foetus and treated cats were resistant to later efforts to incite the infection, inability of tinidazole to eradicate infection in many cats poses a serious impediment to the drug’s effectiveness in practice.
Objective—To compare sensitivity of a complement fixation (CF) test and competitive ELISA (cELISA) for detection of Anaplasma marginale in experimentally infected steers.
Animals—40 crossbred (Angus-Simmental) steers.
Procedures—Steers were inoculated with 2.6 × 109 A marginale–infected erythrocytes (day 0). Blood samples were collected on days 9, 13, 20, 28, 34, 41, 61, 96, 126, and 156 days after inoculation. The percentage of parasitized erythrocytes (PPE) was determined by microscopic examination of stained blood films, and sera were evaluated with the CF test and cELISA by use of USDA-approved methods. Sensitivity and agreement (κ statistic) between the 2 methods were determined. Persistent infections were confirmed by inoculation of blood obtained from infected steers into susceptible, splenectomized calves.
Results—9 days after inoculation, sensitivity of the cELISA was 47.5%, whereas the CF test failed to identify seropositive steers. After day 13, sensitivity of the cELISA and CF test was 100% and 20%, respectively. During peak parasitemia (day 20), sensitivity of the cELISA and CF test was 100%. Thereafter, sensitivity of the CF test fluctuated between 7.5% and 37.5%, whereas sensitivity of the cELISA remained at 100%. Overall sensitivity of the cELISA and CF test was 94.8% and 26.5%, respectively (κ statistic, 0.039).
Conclusions and Clinical Relevance—The cELISA had superior sensitivity for serologic detection of A marginale.The CF test and cELISA each had a high percentage of false-negative results during the prepatent period. These findings are relevant for export certification and anaplasmosis prevention or eradication programs.
Objective—To evaluate a combination of 2 nonantibiotic microbicide compounds, sodium hypochlorite (NaOCl) and polyhexamethylene biguanide (PHMB), as a treatment to suppress or eliminate Salmonella spp from red-eared slider (RES) turtle (Trachemys scripta elegans) eggs and hatchlings.
Sample Population—2,738 eggs from 8 turtle farms in Louisiana.
Procedures—Eggs were randomly sorted into 3 or, when sufficient eggs were available, 4 treatment groups as follows: control, pressure-differential egg treatment with NaOCl and gentamicin, NaOCl and PHMB bath treatment, and pressure-differential egg treatment with NaOCl and PHMB. Bacterial cultures were performed from specimens of eggs and hatchlings and evaluated for Salmonella spp.
Results—RES turtle eggs treated with NaOCl and PHMB as a bath (odds ratio [OR], 0.2 [95% confidence interval (CI), 0.1 to 0.3]) or as a pressure-differential dip (OR, 0.01 [95% CI, 0.001 to 0.07]) or with gentamicin as a pressure-differential dip (OR, 0.1 [95% CI, 0.06 to 0.2]) were significantly less likely to have Salmonella-positive culture results than control-group eggs.
Conclusions and Clinical Relevance—Concern over reptile-associated salmonellosis in children in the United States is so great that federal regulations prohibit the sale of turtles that are < 10.2 cm in length. Currently, turtle farms treat eggs with gentamicin solution. Although this has reduced Salmonella shedding, it has also resulted in antimicrobial resistance. Results of our study indicate that a combination of NaOCl and PHMB may be used to suppress or eliminate Salmonella spp on RES turtle eggs and in hatchlings.
Objective—To determine whether mares are a clinically important source of Rhodococcus equi for their foals.
Sample Population—171 mares and 171 foals from a farm in Kentucky (evaluated during 2004 and 2005).
Procedures—At 4 time points (2 before and 2 after parturition), the total concentration of R equi and concentration of virulent R equi were determined in fecal specimens from mares by use of quantitative bacteriologic culture and a colony immunoblot technique, respectively. These concentrations for mares of foals that developed R equi–associated pneumonia and for mares with unaffected foals were compared. Data for each year were analyzed separately.
Results—R equi–associated pneumonia developed in 53 of 171 (31%) foals. Fecal shedding of virulent R equi was detected in at least 1 time point for every mare; bacteriologic culture results were positive for 62 of 171 (36%) mares at all time points. However, compared with dams of unaffected foals, fecal concentrations of total or virulent R equi in dams of foals with R equi–associated pneumonia were not significantly different.
Conclusions and Clinical Relevance—Results indicate that dams of foals with R equi–associated pneumonia did not shed more R equi in feces than dams of unaffected foals; therefore, R equi infection in foals was not associated with comparatively greater fecal shedding by their dams. However, detection of virulent R equi in the feces of all mares during at least 1 time point suggests that mares can be an important source of R equi for the surrounding environment.
Objective—To determine the interval to provirus and serum antibody detection (via PCR assay and ELISA, respectively) in calves after experimental inoculation with bovine leukemia virus (BLV).
Animals—8 colostrum-deprived, BLV-negative Holstein bull calves (≥ 6 weeks old).
Procedures—Via IM injection, each calf received a fresh whole-blood inoculum (day 0) calculated to contain 2 × 106 lymphocytes. Blood samples for the ELISA and PCR assay were collected from calves immediately prior to inoculation and weekly thereafter for 7 weeks. Mean and median number of weeks to PCR-detected conversion of BLV status and seroconversion were calculated. Point sensitivity and cumulative sensitivity of the 2 assays were calculated at each sample collection. At each sampling time, the proportion of calves identified as infected by the cumulative weekly ELISA and PCR assay results was compared by use of a Fisher exact test.
Results—In 5 calves, conversion of BLV status was detected via PCR assay before seroconversion was identified. However, seroconversion preceded PCR-detected conversion in 2 calves. In 1 calf, both assays yielded positive results at the same test date. These differences were not significant.
Conclusions and Clinical Relevance—In experimentally inoculated BLV-negative calves, conversion of BLV status was detected via PCR assay more quickly than via ELISA; this difference was not significant and probably not clinically important. The PCR assay may be useful as a confirmatory test in animals of exceptional value; tests based on viral identification may become critically important if vaccines against BLV infection are developed and marketed.