Charles M. Scanlan

Effect of a Defined Continuous-Flow Derived Bacterial Culture and Dietary Lactose on Salmonella typhimurium Colonization in Broiler Chickens

A defined bacterial culture protective against Salmonella typhimurium cecal colonization in broiler chicks was derived utilizing a continuous-flow (CF) culture apparatus. Chicks receiving the CF culture in combination with a diet containing dietary lactose were protected against cecal colonization by S. typhimurium. The culture consisted of a mixture of gram-positive and gram-negative facultative and strictly anaerobic bacteria. The isolates were identified as Enterococcus avium, two strains of Enterococcus faecalis (designated A and B), Lactococcus lactis, Lactobacillus animalis, a Lactobacillus that could not be identified to species level (designated strain CMS), Citrobacter freundii, Escherichia coli, E. fergusonii, Bifidobacterium animals, and Propionibacterium acidipropionici. Results indicated that CF cultures can be used as a tool to identify bacteria which are antagonistic to S. typhimurium in the chick cecum.

Inoculation of broiler chicks with a continuous-flow derived bacterial culture facilitates early cecal bacterial colonization and increases resistance to Salmonella typhimurium

Experiments were conducted to examine the effect of a continuous-flow (CF) derived bacterial culture and of dietary lactose on colonization level by anaerobic bacteria in the ceca of broiler chicks. Cecal facultative and strictly anaerobic bacteria colony forming units (CFU), total volatile fatty acid (TVFA), lactic and propionic acid concentrations in 3-d-old chicks, and cecal log10 Salmonella typhimurium colonization and propionic acid concentrations in 10-d-old chicks were measured. Treatment groups were control diet, 2% lactose diet, CF culture + control diet, and CF culture + 2% lactose diet. Groups inoculated with CF culture had decreased (P < 0.05) Salmonella in cecal contents at 10 d of age. The level of Salmonella protection was significantly (P < 0.05) correlated with increased cecal bacteria CFU, TVFA, and propionic acid concentrations in 3-d-old chicks. Results indicated that inoculating newly hatched chicks with CF culture containing cecal microflora originally obtained from adult chickens, facilitates early cecal colonization by native cecal microflora, and this is associated with protection against Salmonella .

Clostridium difficile in retail meat and processing plants in Texas

The incidence and severity of disease associated with toxigenic Clostridium difficile have increased in hospitals in North America from the emergence of newer, more virulent strains. Toxigenic C. difficile has been isolated from food animals and retail meat with potential implications of transfer to human beings. The objective of the present study was to determine the prevalence of C. difficile in pork from sausage manufacturing plants and retail meat in Texas. Twenty-three C. difficile isolates were detected from 243 meat samples (9.5%) from 3 sausage-manufacturing plants and 5 retail meat outlets from 2004 to 2009. Twenty-two isolates were positive for toxins A, B, and binary toxin, and were characterized as toxinotype V, PFGE type-NAP7, or “NAP7-variant.” Susceptibilities to 11 antimicrobial agents in the current study were similar to those reported previously for toxinotype V isolates, although the results suggested somewhat reduced resistance than reported for other meat, animal, or human clinical toxinotype V isolates.

Competitive Exclusion of Salmonella enteritidis in Leghorn Chicks: Comparison of Treatment by Crop Gavage, Drinking Water, Spray, or Lyophilized Alginate Beads

The protective effect of cecal bacteria cultures on Salmonella enteritidis cecal colonization was evaluated. Competitive-exclusion cultures were administered by crop gavage, in first drinking water, by whole body spray, or encapsulated in alginate beads and provided in feed pans. Leghorn chicks were treated with cultures of cecal bacteria on the day of hatch and challenged orally with 10(4) S. enteritidis 2 days after treatment. Salmonella cecal colonization was evaluated 7 days after challenge. No Salmonella organisms were detected in the ceca of chicks treated with cecal cultures by crop gavage. Chicks treated with cecal cultures in the drinking water or by spray application showed comparable protection and significant decreases (P < 0.05) in the number of Salmonella in the cecal contents compared with untreated controls. The consumption of cecal bacteria encapsulated in alginate beads significantly decreased (P < 0.05) Salmonella cecal colonization compared with control treatment, but it provided less protection than the other treatment methods evaluated.

Inhibition of Salmonella enteritidis cecal and organ colonization in leghorn chicks by a defined culture of cecal bacteria and dietary lactose

The effect of oral inoculation with a defined culture of cecal bacteria and provision of dietary lactose on colonization resistance to invasive Salmonella enteritidis was evaluated in leghorn chicks. A defined mixed culture composed of 11 indigenous cecal bacteria capable of utilizing lactose or lactose fermentation products as a primary carbon source was isolated and maintained in continuous-flow culture. Combined treatment with the defined culture and dietary lactose significantly decreased (P < 0.01) the number of Salmonella in the cecal contents and the number of Salmonella cecal-culture-positive chicks in four replicated trials. Additionally, Salmonella colonization of the spleen, liver, and cecal tonsils was significantly decreased (P < 0.05) in each of the trials. The results indicate that the resistance of leghorn chicks to S. enteritidis cecal and organ colonization is effectively increased by inoculation with a defined culture of cecal bacteria and provision of lactose in the diet.

In Vitro Inhibition of the Growth of Salmonella typhimurium and Escherichia coli 0157:H7 by Bacteria Isolated from the Cecal Contents of Adult Chickens

A Veillonella species and Enterococcus durans were isolated from the cecal contents of adult broilers. Mixed cultures of Veillonella and E. durans inhibited the growth of Salmonella typhimurium and Escherichia coli 0157:H7 on media containing 2.5% lactose (w/v). The growth of S. typhimurium or E. coli 0157:H7 was not inhibited by mixed cultures containing Veillonella and E. durans on media containing only 0.25% lactose or by pure cultures of Veillonella or E. durans on media containing either 0.25% or 2.5% lactose. The mixed cultures of Veillonella and E. durans produced significantly (P<0.05) more acetic, propionic, and lactic acids in media containing 2.5% lactose than in media containing 0.25% lactose. The inhibition of the enteropathogens was related to the production of lactic acid from lactose by the E. durans and the production of acetic and propionic acids from lactic acid by the Veillonella .

A Bacteriologic Study of Scabby-Hip Lesions from Broiler Chickens in Texas

Broilers from commercial flocks experiencing a 10–60% incidence of scabby-hip lesions at processing were examined, and selected skin lesions were cultured. Over 70% of the lesions were associated with traumatic excoriations, particularly on the caudal dorsal convexity of the birds. Most lesions were observed on birds that were 5 weeks of age or older. From the 27 specimens cultured, Clostridium perfringens was isolated in pure culture from 4 lesions and Staphylococcus species from 10 lesions. Pure cultures of staphylococci were recovered from 4 lesions, and 2–5 different staphylococci were isolated from 6 lesions. Eight staphylococci were identified as S. sciuri, 8 as S. simulans, 2 as S. epidermidis, 2 as S. lentus, 2 as S. warneri, 1 as S. cohnii, and 1 as S. intermedius. Fifty cutaneous specimens from 10 5-week-old normal broilers were cultured. A total of 197 isolates were identified including 65 S. sciuri, 52 S. lentus, 24 S. simulans, 12 S. hyicus, 11 S. warneri, 9 S. cohnii, 9 S. gallinarium, 8 S. xylosus, and 7 S. epidermidis.

Use of differential rumen fluid-based carbohydrate agar media for culturing lactose-selected cecal bacteria from chickens

A rumen fluid-based differential carbohydrate agar medium for enumerating chicken cecal carbohydrate-utilizing bacteria was tested. Eleven bacteria isolated from a continuous-flow culture that had been seeded with chicken cecal contents were inoculated on fifteen different media in an anaerobic environment. These media included various levels of rumen fluid (0%, 5%, 10%, 16%) and carbohydrate (glucose [GLU], galactose [GAL], lactose[LAC]). Viable cell recovery (colony-forming units [CFU]/direct cell counts × 100%) was higher for 16% rumen fluid-supplemented carbohydrate media than the lower concentrations of rumen fluid-supplemented carbohydrate media. In addition, average viable cell recovery of all 11 bacteria on differential carbohydrate media were significantly (P < 0.05) higher in galactose and lactose media than on the glucose media. Data indicated that 16% rumen-fluid based media can be used to enumerate anaerobic, lactose-fermenting subgroups of chicken cecal bacteria.

Cultural and Biochemical Characterization of Actinobacillus and Actinobacillus-Like Species from Ram Lambs with Epididymitis

Cellular, colonial, cultural, and biochemical characteristics of 25 field strains of gram-negative pleomorphic bacilli from rams with epididymitis were compared with Actinobacillus actinomycetemcomitans American Type Culture Collection (ATCC) strain 29522 and Actinobacillus seminis ATCC strain 15768. Three field strains were identified as A. actinomycetemcomitans, 15 as A. seminis, and 2 as Haemophilus agni; however, 5 strains (3 in group A and 2 in group B) were not identified as species in the genera Actinobacillus, Haemophilus, or Pasteurella based on the taxonomic criteria in Bergeys manual of systematic bacteriology. The 5 Actinobacillus-like organisms in groups A and B were predominantly gram-negative coccobacilli and exhibited less pleomorphism than the 2 Actinobacillus species. The colonial morphologies of groups A and B were similar to the 2 Actinobacillus species but were smaller in diameter and had a pale yellow color. Groups A and B, like the actinobacilli, were facultative anaerobic and capnophilic, did not grow on MacConkey agar, and were catalase-positive and oxidase-positive. Group A reduced nitrate but group B did not. The A. seminis strains utilized ornithine, and group A utilized arginine; but group B did not utilize either omithine or arginine. All strains failed to utilize lysine or tryptophane. All strains produced acid but no gas from glucose, and the utilization of other carbohydrates varied markedly both between and within the 5 groups of bacteria. Based on carbohydrate utilization profiles, the 15 field strains of A. seminis represented 13 biotypes, but with a commercial semi-quantitative enzyme system only 1 biotype was identified. All the A. actinomycetemcomitans, A. seminis, and group B strains produced moderate to high levels of alkaline phosphatase and acid phosphatase, but the group A strains were negative. Groups A and B were negative for beta-glucuronidase, but all the A. actinomycetemcomitans and A. seminis strains produced this enzyme.

Current Concepts of Competitive Exclusion Cultures for the Control of Salmonellae in Domestic Poultry

Two defined competitive exclusion (CE) cultures (CF-I and CF-II) and a characterized CE culture (CF-III), which are composed of mixtures of nonpathogenic bacteria, were developed from anaerobic continuous-flow (CF) cultures that had been inoculated with cecal contents from adult chickens. After the primary CF cultures attained homeostasis, 13 bacteria (11 facultative anaerobes and 2 obligate anaerobes) representing 6 genera were isolated from CF-I; II bacteria (9 facultative anaerobes and 2 obligate anaerobes) representing 7 genera were isolated from CF-II; and 29 bacteria (15 facultative anaerobes and 14 obligate anaerobes) representing 14 genera were isolated from CF-III. Newly hatched chicks were treated orally with each primary CF culture; challenged on day 3 with Salmonella typhimurium; and cultured on day 10. Each culture significantly (p < 0.05) reduced salmonellae intestinal colonization and organ invasion. From the reconstituted CF-I and CF-II cultures, all organisms were isolated and their fermentation parameters and efficacy against salmonellae challenge were similar, if not identical, to the primary cultures. The CF-I and CF-II cultures satisfied the 5 requirements for defined CE cultures: 1) the primary CE culture must be efficacious; 2) all bacteria must be isolated and identified; 3) the fermentation parameters of the reconstituted CE culture must be similar to those of the primary culture; 4) all bacteria from the reconstituted culture must be isolated and identified; and 5) the efficacy of the reconstituted culture must be very similar to the primary culture. From these integrated studies, 3 mechanisms were demonstrated for preventing the enteric colonization of salmonellae in newly hatched chicks that were pretreated with CE cultures. First, the component organisms in the CE culture establish a normal enteric flora prior to salmonellae exposure. Second, the CE organisms compete with salmonellae for essential nutrients. Third, the CE organisms produce concentrations of volatile fatty acids at low pH levels that are bacteriostatic for salmonellae.