Roger B. Harvey

Probiotics, prebiotics and competitive exclusion for prophylaxis against bacterial disease

Abstract The microbial population of the intestinal tract is a complex natural resource that can be utilized in an effort to reduce the impact of pathogenic bacteria that affect animal production and efficiency, as well as the safety of food products. Strategies have been devised to reduce the populations of food-borne pathogenic bacteria in animals at the on-farm stage. Many of these techniques rely on harnessing the natural competitive nature of bacteria to eliminate pathogens that negatively impact animal production or food safety. Thus feed products that are classified as probiotics, prebiotics and competitive exclusion cultures have been utilized as pathogen reduction strategies in food animals with varying degrees of success. The efficacy of these products is often due to specific microbial ecological factors that alter the competitive pressures experienced by the microbial population of the gut. A few products have been shown to be effective under field conditions and many have shown indications of effectiveness under experimental conditions and as a result probiotic products are widely used in all animal species and nearly all production systems. This review explores the ecology behind the efficacy of these products against pathogens found in food animals, including those that enter the food chain and impact human consumers.

Bactericidal effect of sodium chlorate on Escherichia coli O157:H7 and Salmonella Typhimurium DT104 in rumen contents in vitro.

Escherichia coli O157:H7 and Salmonella Typhimurium DT104 are important foodborne pathogens affecting the beef and dairy industries and strategies are sought to rid these organisms from cattle at slaughter. Both pathogens possess respiratory nitrate reductase that also reduces chlorate to the lethal chlorite ion. Because most anaerobes lack respiratory nitrate reductase, we hypothesized that chlorate may selectively kill E. coli O157:H7 and Salmonella Typhimurium DT104 but not potentially beneficial anaerobes. In support of this hypothesis, we found that concentrations of E. coli O157:H7 and Salmonella Typhimurium DT104 were reduced from approximately 1,000,000 colony forming units (CFU) to below our level of detection (< or = 10 CFU) following in vitro incubation (24 h) in buffered ruminal contents (pH 6.8) containing 5 mM added chlorate. In contrast, chlorate had little effect on the most probable number (mean +/- SD) of total culturable anaerobes (ranging from 9.9 +/- 0.72 to 10.7 +/- 0.01 log10 cells/ml). Thus, chlorate was bactericidal to E. coli O157:H7 and Salmonella Typhimurium DT104 but not to potentially beneficial bacteria. The bactericidal effect of chlorate was concentration dependent (less at 1.25 mM) and markedly affected by pH (more bactericidal at pH 6.8 than pH 5.6).

Efficacy of Zeolitic Ore Compounds on the Toxicity of Aflatoxin to Growing Broiler Chickens

Commercially available zeolitic ore compounds, when incorporated into the diets at 0.5%, were evaluated for their ability to reduce the deleterious effects of 3.5 mg aflatoxin/kg feed on growing broiler chickens from 1 day to 3 weeks of age. In a series of four experiments, the compounds used included the following: mordenite (particle size of -20 mesh; Zeomite); clinoptilolite (particle size of -20 mesh; Zeobrite); SC Zeolite (particle size of -20 mesh); and clinoptilolite (particle size of -35 mesh; Clino 1) or clinoptilolite (particle sizes of -20 plus +35 mesh; Clino 2). Results demonstrated that 0.5% Zeobrite, Clino 1, or Clino 2 added to aflatoxin-contaminated diets did not significantly (P < 0.05) diminish the toxicity of high concentrations of aflatoxin to growing broiler chicks. Zeomite mordenite ore reduced the toxicity of aflatoxin to growing chicks by 41%, as indicated by weight gains, liver weight, and serum biochemical measurements, which compares favorably with its in vitro binding capacity to aflatoxin. SC Zeolite reduced weight-gain toxicity of aflatoxin by approximately 29%.

Foodborne Campylobacter: Infections, Metabolism, Pathogenesis and Reservoirs

Campylobacter species are a leading cause of bacterial-derived foodborne illnesses worldwide. The emergence of this bacterial group as a significant causative agent of human disease and their propensity to carry antibiotic resistance elements that allows them to resist antibacterial therapy make them a serious public health threat. Campylobacter jejuni and Campylobacter coli are considered to be the most important enteropathogens of this genus and their ability to colonize and survive in a wide variety of animal species and habitats make them extremely difficult to control. This article reviews the historical and emerging importance of this bacterial group and addresses aspects of the human infections they cause, their metabolism and pathogenesis, and their natural reservoirs in order to address the need for appropriate food safety regulations and interventions.

Varied prevalence of Clostridium difficile in an integrated swine operation.

The objectives of this study were to compare the prevalence of Clostridium difficile (Cd) among different age and production groups of swine in a vertically integrated swine operation in Texas in 2006 and to compare our isolates to other animal and human isolates. Results are based on 131 Cd isolates from 1008 swine fecal samples and pork trim samples (overall prevalence of 13%). The prevalence (number positive/number tested in production type) of Cd was different between the groups (P<or=0.001), and was highest among suckling piglets at 50.0% (61/122), followed by 23.8% (34/143) for lactating sows and effluent from the farrowing barn, 8.4% (10/119) for nursery, 6.5% (4/62) for pork products, 3.9% (15/382) for grower-finisher, and 3.9% (7/180) for breeding boars and sows. Of the 131 isolates, 122 were positive by PCR for both toxins A (tcdA) and B (tcdB) genes, 129 isolates harbored a 39 base pair deletion in the tcdC gene, 120 isolates were toxinotype V, and all 131 of the isolates were positive for the binary toxin gene cdtB. All isolates were resistant to cefoxitin, ciprofloxacin, and imipenem, whereas all were sensitive to metronidazole, piperacillin/tazobactam, amoxicillin/clavulanic acid, and vancomycin. The majority of isolates were resistant to clindamycin; resistant or intermediate to ampicillin; and sensitive to tetracycline and chloramphenicol. There was an increased (P</=0.001) number of isolates for the timeframe of September to February compared to March to August.

Effect of sodium chlorate on Salmonella Typhimurium concentrations in the weaned pig gut

Salmonella cause economic losses to the swine industry due to disease and compromised food safety. Since the gut is a major reservoir for Salmonella, strategies are sought to reduce their concentration in pigs immediately before processing. Respiratory nitrate reductase activity possessed by Salmonella also catalyzes the intracellular reduction of chlorate (an analog of nitrate) to chlorite, which is lethal to the microbe. Since most gastrointestinal anaerobes lack respiratory nitrate reductase, we conducted a study to determine if chlorate may selectively kill Salmonella within the pig gut. Weaned pigs orally infected with 8 x 10(7) CFU of a novobiocin- and nalidixic acid-resistant strain of Salmonella Typhimurium were treated 8 and 16 h later via oral gavage (10 ml) with 0 or 100 mM sodium chlorate. Pigs were euthanized at 8-h intervals after receiving the last treatment. Samples collected by necropsy were cultured qualitatively and quantitatively for Salmonella and for most probable numbers of total culturable anaerobes. A significant (P < 0.05) chlorate treatment effect was observed on cecal concentrations of Salmonella, with the largest reductions occurring 16 h after receiving the last chlorate treatment. An observed treatment by time after treatment interaction suggests the chlorate effect was concentration dependent. Chlorate treatment may provide a means to reduce foodborne pathogens immediately before harvest.

Genotypic variation among arcobacter isolates from a farrow-to-finish swine facility.

Arcobacter spp. were isolated from nursing sows and developing pigs on three farms of a farrow-to-finish swine operation and market-age pigs at slaughter. Isolates were identified by polymerase chain reaction and genotypic fragment patterns were examined by pulsed-field gel electrophoresis (PFGE). Incidences of Arcobacter-positive samples increased progressively as the pigs aged, resulting in all of the pens at the end of the growth cycle in the finishing barn containing Arcobacter-positive feces. However, only 10 of 350 cecal samples from slaughtered pigs were positive. There was little similarity between genotypic patterns for Arcobacter collected from the three farms. The level of genotypic variation revealed by PFGE suggested that pigs in this farrow-to-finish operation were colonized by multiple Arcobacter parent genotypes that may have undergone genomic rearrangement, common to members of Campylobacteraceae, during successive passages through the animals. Additionally, the level of genotypic diversity seen among Arcobacter isolates from farms of a single farrow-to-finish swine operation suggests an important role for genotypic phenotyping as a source identification and monitoring tool during outbreaks.

Recent pre-harvest supplementation strategies to reduce carriage and shedding of zoonotic enteric bacterial pathogens in food animals

Abstract Food-borne bacterial illnesses strike more than 76 million North Americans each year. Many of these illnesses are caused by animal-derived foodstuffs. Slaughter and processing plants do an outstanding job in reducing bacterial contamination after slaughter and during further processing, yet food-borne illnesses still occur at an unacceptable frequency. Thus, it is imperative to widen the window of action against pathogenic bacteria. Attacking pathogens on the farm or in the feedlot will improve food safety all the way to the consumer’s fork. Because of the potential improvement in overall food safety that pre-harvest intervention strategies can provide, a broad range of preslaughter intervention strategies are currently under investigation. Potential interventions include direct anti-pathogen strategies, competitive enhancement strategies and animal management strategies. Included in these strategies are competitive exclusion, probiotics, prebiotics, antibiotics, antibacterial proteins, vaccination, bacteriophage, diet, and water trough interventions. The parallel and simultaneous application of one or more preslaughter strategies has the potential to synergistically reduce the incidence of human food-borne illnesses by erecting multiple hurdles, thus preventing entry of pathogens into the food chain. This review emphasizes work with Escherichia coli O157:H7 to illustrate the various strategies.

Comparison of Two Hydrated Sodium Calcium Aluminosilicate Compounds to Experimentally Protect Growing Barrows from Aflatoxicosis

Two formulations of hydrated sodium calcium aluminosilicate (HSCAS-1 and HSCAS-3), anti-caking agents for mixed feed, were added to the diets of growing barrows and were evaluated for their potential to diminish the clinical signs of aflatoxicosis. The experimental design consisted of 8 barrows (2 replicates of 4 each/treatment) assigned to 1 of the following 6 treatment diets (total of 48): 1) 0 g of HSCAS-1 or HSCAS-3 and 0 mg of aflatoxin (AF)/kg of feed (control); 2) 5 g HSCAS-1/kg of feed; 3) 5 g HSCAS-3/kg of feed; 4) 3 mg AF/kg of feed; 5) 3 mg AF plus 5 g HSCAS-1/kg of feed; or 6) 3 mg AF plus 5 g HSCAS-3/kg of feed. Barrows were maintained in indoor concrete-floored pens, with feed and water -available for 28 days (from 8 to 12 weeks of age). Barrows were observed twice daily and weighed weekly, and blood samples were collected at day 28 for hematologic, immunologic, and serum biochemical measurements. At the termination of the study, barrows were euthanized and necropsied. Barrow body weight gains were diminished, compared to those of controls, by consumption of AF alone and both of the AF plus HSCAS diets; however, the AF plus HSCAS-1 and AF plus HSCAS-3 barrow body weight gains were significantly greater (P < 0.05) than those of the AF-alone barrows, No toxic responses or performance differences were noticed for barrows consuming either of the HSCAS-alone diets. Serum concentrations of alkaline phosphatase, gamma glutamyltransferase, calcium, cholesterol, albumin, triglycerides, and urea nitrogen were altered in barrows of the AF-alone treatment. The use of HSCAS prevented most but not all of the AF-induced changes in biochemical values. Immunologic measurements that were adversely affected by AF included mitogen-induced lymphoblastogenesis and peritoneal macrophage activity and function. The addition of HSCAS to AF-contaminated diets protected barrows from some of these toxic changes. Although immunologic measurements in the AF plus HSCAS groups were significantly different than those of the AF-alone group, values were still not equivalent to those of controls. These findings suggest that HSCAS-1 and HSCAS-3 are equal in their ability to protect against the toxicity of AF. Although these compounds may offer a novel approach to the preventive management of aflatoxicosis in livestock and poultry, HSCAS is not approved by the US Food and Drug Administration for treatment of animal diets for prevention of mycotoxicosis.

Effects of phyllosilicate clay on the metabolic profile of aflatoxin B1 in Fischer-344 rats

The phyllosilicate clay, hydrated sodium calcium aluminosilicate (HSCAS), has been shown to prevent aflatoxicosis in farm animals by reducing the bioavailability of aflatoxin. The present study was designed to determine the effects of HSCAS on the metabolism of aflatoxin B1 (AFB1) in an aflatoxin-sensitive species. Male Fischer-344 rats were orally dosed with 1.0, 0.5, 0.25 and 0.125 mg AFB1/kg body weight alone and in combination with 0.5% HSCAS. Urine samples were collected after 6, 24, 36, and 48 h. Aflatoxin M1 (AFM1) and aflatoxin P1 (AFP1) were detected in most urine samples, with or without HSCAS. AFM1 was found to be the major metabolite. Metabolite concentrations were significantly decreased in the presence of HSCAS, and more importantly, no additional metabolites were detected. Our results suggest that the AFB1-HSCAS complex was not significantly dissociated in vivo, and support earlier findings that HSCAS tightly binds aflatoxin.