Margaret L. Khaitsa

Epidemiological evidence for Mycobacterium avium subspecies paratuberculosis as a cause of Crohn's disease

Mycobacterium avium subspecies paratuberculosis is the causative agent of Johnes disease, a chronic enteritis in ruminants including cattle, sheep, goats, and farmed deer. Recently, this bacterium has received an increasingly wide interest because of a rapidly growing body of scientific evidence which suggests that human infection with this microorganism may be causing some, and possibly all, cases of Crohns disease. Recent studies have shown that a high percentage of people with Crohns disease are infected with M. avium subsp. paratuberculosis; whether the association of this bacterium and Crohns disease is causal or coincidental is not known. Crohns disease is a gastrointestinal disease in humans with similar histopathological findings to those observed in the paucibacillary form of Johnes disease in cattle. The search for risk factors in Crohns disease has been frustrating. However, epidemiologists have gathered enough information that points to an association between M. avium subsp. paratuberculosis and Crohns disease. This paper reviews epidemiological models of disease causation, the major philosophical doctrines about causation, the established epidemiological criteria for causation, and the currently known epidemiological evidence of M. avium subsp. paratuberculosis as a possible cause of Crohns disease.

Comparative effect of direct-fed microbials on fecal shedding of Escherichia coli O157:H7 and Salmonella in naturally infected feedlot cattle.

The effect of direct-fed microbials (DFM) on fecal shedding of Escherichia coli O157:H7 and Salmonella in naturally infected feedlot cattle was evaluated in a clinical trial involving 138 feedlot steers. Following standard laboratory methods, fecal samples collected from steers were evaluated for change in the detectable levels of E. coli O157:H7 and Salmonella shed in feces after DFM treatment. Sampling of steers was carried out every 3 weeks for 84 days. A significant reduction (32%) in fecal shedding of E. coli O157:H7 (P < 0.001), but not Salmonella (P = 0.24), was observed among the treatment steers compared with the control group during finishing. The probability of recovery of E. coli O157:H7 from the feces of treated and control steers was 34.0 and 66.0%, respectively. Steers placed on DFM supplement were almost three times less likely to shed E. coli O157:H7 (odds ratio, 0.36; 95% confidence interval, 0.25 to 0.53; P < 0.001) in their feces as opposed to their control counterparts. The probability of recovery of Salmonella from the feces of the control (14.0%) and the treated (11.3%) steers was similar. However, the DFM significantly reduced probability of new infections with Salmonella among DFM-treated cattle compared with controls (nontreated ones). It appears that DFM as applied in our study are capable of significantly reducing fecal shedding of E. coli O157:H7 in naturally infected cattle but not Salmonella. The factors responsible for the observed difference in the effects of DFM on E. coli O157:H7 and Salmonella warrants further investigation.

Antimicrobial Drug Resistance and Molecular Characterization of Salmonella Isolated from Domestic Animals, Humans and Meat Products

OBJECTIVES 1) To characterize and determine genotypic relatedness of Salmonella serovars commonly isolated from domestic animals and humans in North Dakota, and 2) to assess their role in transferring antimicrobial resistance (AMR) to humans. MATERIALS AND METHODS A total of 434 Salmonella isolates obtained from 1) feces of apparently healthy feedlot, range, and dairy cattle in North Dakota; 2) clinical samples from sick or dead animals submitted to North Dakota State University-Veterinary Diagnostic Laboratory (2000-2005); 3) previous meat product surveillance studies in North Dakota; and 4) 179 samples from human patients in North Dakota (2000-2005) by the North Dakota Department of Health were studied. The isolates were initially serotyped and later genotyped by pulsed-field gel electrophoresis (PFGE) to investigate their relatedness. The National Antimicrobial Resistance Monitoring Systems panel was used to compare AMR profiles of animal and human isolates to assess a possible role of domestic animals in transfer of AMR to humans. RESULTS Salmonella Typhimurium was the predominant serotype in both humans (13.4%) and domestic animals (34.3%), followed by Newport in animals (2.6%) and human (3.9%). Salmonella Arizona (0.7%), Salmonella Give (0.9%), and Salmonella Muenster (3.5%) were isolated from sick or dead animals. PFGE results confirmed occurrence of similar Salmonella genotypes in both domestic animals and humans. AMR profiles showed that most animal strains were multidrug resistant. A single human isolate had PFGE and multidrug resistance profiles similar to a major cattle genotype, suggesting a possible AMR transmission from cattle to humans. CONCLUSION AND APPLICATION: Similar Salmonella genotypes were infecting domestic animals and humans in North Dakota. The AMR levels were higher in domestic animal isolates than in humans, implying that the occurrence of AMR in animal isolates may not translate directly into AMR in human isolates in North Dakota. This is helpful in determining future policies regarding antimicrobial drug use in domestic animals and humans.

Risk Factors Associated with Anthrax Outbreak in Animals in North Dakota, 2005 : A Retrospective Case-Control Study

Objective. We identified the risk factors associated with the anthrax outbreak of 2005 in animals in North Dakota. Methods. Medical records of the 2005 anthrax outbreak were obtained from the Veterinary Diagnostic Laboratory at North Dakota State University. Additional data were obtained from the North Dakota state veterinarians office, and supplemental questionnaires were administered to producers. The data obtained included ecological and environmental factors, animal health factors, and management factors. Results. Anthrax occurred from July 1 to October 12, 2005. The cases were located in eastern North Dakota around the Red River Basin. Ransom, LaMoure, and Barnes counties reported most cases (71%). Species affected included cattle, bison, horses, sheep, elk, deer, pigs, and llamas. The predominant symptom was sudden death (38%) followed by bleeding from orifices (17%). Chi-square analysis indicated significant differences between case and control premises on the following variables: death reported on neighboring pasture, vaccination period, dry conditions, wet conditions, antibiotic use, multiple vaccination, and type of predator (coyote). Factors that significantly (p<0.05) predicted anthrax occurrences on the final logistic regression model were vaccination, use of antibiotics during an outbreak, and period of vaccine administration (before or during the outbreak). Conclusions. The characteristics of the anthrax outbreak regarding time and place of occurrence, animals affected, clinical signs reported, and mortality rate were consistent with previous reports of natural anthrax outbreaks in animals. A number of factors that significantly predicted anthrax occurrence in animals in the 2005 outbreak in North Dakota were identified. This information is important in planning appropriate control and prevention measures for anthrax, including recommending the right vaccination and treatment regimens in managing future anthrax outbreaks.

Fecal shedding of Escherichia coli O157:H7 in North Dakota feedlot cattle in the fall and spring.

Cattle are an important reservoir of Escherichia coli O157:H7, which can lead to contamination of food and water, and subsequent human disease. E. coli O157:H7 shedding in cattle has been reported as seasonal, with more animals shedding during summer and early fall than during winter. North Dakota has relatively cold weather, especially in winter and early spring, compared with many other regions of the United States. The objective was to assess fecal shedding of E. coli O157:H7 in North Dakota feedlot cattle over the fall, winter, and early spring. One hundred forty-four steers were assigned randomly to 24 pens on arrival at the feedlot. Samples of rectal feces were obtained from each steer four times (October and November 2003, and March and April 2004) during finishing. On arrival (October 2003), 2 (1.4%) of 144 cattle were shedding E. coli O157:H7. The shedding increased significantly to 10 (6.9%) of 144 after 28 days (November 2003), to 76 (53%) of 143 at the third sampling (March 2004), and dropped significantly to 30 (21%) of 143 at the fourth (last) sampling (March 2004) before slaughter. Unfortunately, we were unable to sample the cattle during winter because of the extreme weather conditions. Sampling time significantly (P < 0.0001) influenced variability in E. coli O157:H7 shedding, whereas herd (P = 0.08) did not. The prevalence of E. coli O157:H7 shedding in North Dakota steers in fall and early spring was comparable to what has been reported in other parts of the United States with relatively warmer weather. Further research into E. coli O157:H7 shedding patterns during extreme weather such as North Dakota winters is warranted in order to fully assess the seasonal effect on the risk level of this organism.

A longitudinal study of Salmonella shedding and antimicrobial resistance patterns in North Dakota feedlot cattle.

Salmonella is one of the most frequent causes of foodborne illness worldwide, and transmission involves foods of animal origin such as beef. The objective of this study was to monitor the prevalence of Salmonella fecal shedding in feedlot cattle during the finishing period and to assess the antimicrobial resistance patterns of the isolated salmonellae. On arrival at the feedlot, 1 (0.7%) of the 144 steers was shedding Salmonella in its feces. After 28 days on feed, shedding was detected in 8 (5.6%) of the 144 steers. At the third sampling, 19 (13%) of 143 steers were shedding, and the number of shedders continued to increase to 89 (62%) of 143 at the last sampling. Salmonella shedding was significantly influenced (P < 0.0001) by sampling time but not by herd of origin. All Salmonella isolates identified belonged to serotype Typhimurium serovar Copenhagen, a type commonly isolated from Salmonella infections in humans. Antimicrobial resistance testing of the isolates revealed five multidrug resistance patterns, two of which accounted for 104 (95.4%) of 109 of the isolates. All the isolates were susceptible to ceftiofur, and all were resistant to spectinomycin, sulfathiazole, tiamulin, florfenicol, ampicillin, penicillin, chlortetracycline, oxytetracycline, and clindamycin. Data from this study indicate that a high prevalence of antimicrobial-resistant Salmonella strains can sometimes be found in feedlot cattle in North Dakota. These data will contribute to risk assessment of Salmonella shedding by cattle in feedlots and highlight the need to continue preharvest monitoring for this organism.

Environmental and ecological determinants of West Nile virus occurrence in horses in North Dakota, 2002

West Nile virus (WNV) outbreak in North Dakota in 2002 included over 569 horse cases, clustered mainly in the eastern and northeastern parts of the state. The pattern of occurrence observed suggested existence of specific environmental and ecological factors that increased the risk for infection and illness in those locations. We developed a predictive model with factors that explained the pattern of WNV occurrence observed. Results indicated that surface elevation, temperature, precipitation, reported WNV-positive birds, reported WNV-positive humans, and reported WNV-positive mosquitoes were important predictors of occurrence in horses. However, case distance from water bodies was not significant in the model. Future predictive models of WNV occurrence in horses should take into account these factors in order to improve accuracy and reliability. Research into other potential determinants such as horse management factors are required to determine more differential risk factors associated with WNV occurrence in horses.

Comparison of two sampling methods for Escherichia coli O157:H7 detection in feedlot cattle.

Two sampling methods (rectoanal swabs and rectal fecal grabs) were compared for their recovery of Escherichia coli O157:H7 from feedlot cattle. Samples were collected from 144 steers four times during the finishing period by swabbing the rectoanal mucosa with cotton swabs and immediately obtaining feces from the rectum of each individual steer. The number of steers with detectable E. coli O157:H7 increased from 2 of 144 (1.4%) cattle on arrival at the feedlot to 10 of 144 (6.9%) after 1 month, 76 of 143 (52.8%) after 7 months, and 30 of 143 (20.8%) at the last sampling time before slaughter. Wilcoxon signed-rank tests indicated that the two sampling methods gave different results for sampling times 3 and 4 (P < 0.05) but not for sampling time 2 (P = 0.16). Agreement between the two sampling methods was poor (kappa < 0.2) for three of the four sampling times and moderate (kappa = 0.6) for one sampling time, an indication that in this study rectoanal swabs usually were less sensitive than rectal fecal grabs for detection of E. coli O157:H7 in cattle. Overall, the herd of origin was not significantly associated with E. coli O157:H7 results, but the weight of the steers was. Further investigation is needed to determine the effects of potential confounding factors (e.g., size and type of swab, consistency of feces, site sampled, and swabbing technique) that might influence the sensitivity of swabs in recovering E. coli O157:H7 from the rectoanal mucosa of cattle.

Isolation and characterization of shiga toxin-producing escherichia coli serogroups O26, O45, O103, O111, O113, O121, O145, and O157 shed from range and feedlot cattle from postweaning to slaughter.

Cattle are the main reservoirs for Shiga toxin-producing Escherichia coli (STEC) strains. E. coli O26, O45, O103, O111, O121, O145, and O157 are among the STEC serogroups that cause severe foodborne illness and have been declared as adulterants by the U.S. Department of Agriculture, Food Safety and Inspection Service. The objectives of this study were (i) to estimate the prevalence of non-O157 STEC and E. coli O157 in naturally infected beef cows and in steer calves at postweaning, during finishing, and at slaughter and (ii) to test non-O157 STEC isolates for the presence of virulence genes stx1, stx2, eaeA, and ehlyA. Samples were collected from study animals during multiple sampling periods and included fecal grabs, rectal swabs, and midline sponge samples. Laboratory culture, PCR, and multiplex PCR were performed to recover and identify E. coli and the virulence genes. The prevalence of non-O157 STEC (serogroups O26, O45, O103, O111, O121, O113, and O145) fecal shedding ranged from 8% (4 of 48 samples) to 39% (15 of 38 samples) in cows and 2% (1 of 47 samples) to 38% (9 of 24 samples) in steer calves. The prevalence of E. coli O157 fecal shedding ranged from 0% (0 of 38 samples) to 52% (25 of 48 samples) in cows and 2% (1 of 47 samples) to 31% (15 of 48 samples) in steer calves. In steer calves, the prevalence of non-O157 STEC and E. coli O157 was highest at postweaning, at 16% (15 of 96 samples) and 23% (22 of 96 samples), respectively. Among the 208 non-O157 STEC isolates, 79% (164 isolates) had stx1, 79% (165 isolates) had stx2, and 58% (121 isolates) had both stx1 and stx2 genes. The percentage of non-O157 STEC isolates encoding the eaeA gene was low; of the 165 isolates tested, 8 (5%) were positive for eaeA and 135 (82%) were positive for ehlyA. Findings from this study provide further evidence of non-O157 STEC shedding in beef cows and steer calves particularly at the stage of postweaning and before entry into the feedlot.

Genetic and Phenotypic Characterization of the blaCMY Gene from Escherichia coli and Salmonella enterica Isolated from Food-Producing Animals, Humans, the Environment, and Retail Meat

The bla(CMY-2) family of the ampC beta-lactamase genes confer broad-spectrum resistance to beta-lactam antimicrobials, including ceftriaxone and ceftiofur, as well as to beta-lactamase inhibitors, such as clavulanic acid. Organisms with the bla(CMY-2) phenotype have been recovered from the environment and from retail meat products, posing a potential public health risk. The objectives of this study were to sequence the bla(CMY-2) gene from Escherichia coli and Salmonella enterica from multiple sources that had a reduced susceptibility to ceftriaxone and to determine the effect of observed mutations in the bla(CMY-2) gene on the antimicrobial resistance phenotype (spectrum and minimum inhibitory concentration/susceptibility patterns) of the isolates. The bla(CMY-2) genes from 52 bacterial isolates were sequenced for this study. Sixty-two percent (32/52) were E. coli and 38% (20/52) were S. enterica. Of the 32 E. coli isolates, 30 were found to carry a beta-lactamase gene that was 100% homologous to bla(CMY-2). One of the E. coli isolates was found to contain a gene that was 90% homologous to bla(CMY-2). This isolate also had lower minimum inhibitory concentrations to tetracyclines, streptomycin, and the sulfonamide antimicrobials than are commonly expected for isolates containing the bla(CMY-2). Of the 20 genes obtained from Salmonella isolates, 8 (40%) were found to be homologous to bla(CMY-2), with no altered susceptibility phenotypes observed.