Billy M. Hargis

Provision of lactose to molting hens enhances resistance to Salmonella enteritidis colonization.

Older leghorn hens, more than 50 weeks of age, were divided into three groups designated 1, unmolted controls; 2, molted; or 3, molted treated with lactose. Forced molt was induced by 14 days of feed removal. Lactose was provided to the hens in group 3 as 2.5% (wt/vol) of the daily drinking water. Each hen in all groups was challenged orally with 10(5) Salmonella enteritidis (SE) cells on day 7 of feed removal. The study was repeated in three replicated trials. The concentrations of acetic, propionic, and total volatile fatty acids (VFA) in the cecal contents of the molted hens in groups 2 and 3 decreased significantly (P < 0.05) on days 6 and 14 of molt compared with the unmolted controls. Forced molt had no apparent effect on pH or on the oxidation-reduction potential of the ceca. Compared to the unmolted controls, SE cecal and spleen and liver colonization was significantly increased (P < 0.05) in the molted hens in group 2. Compared to the molted hens in group 2, SE cecal and spleen and liver colonization was significantly decreased (P < 0.05) in two of three trials in the hens in group 3 provided with lactose. The results suggested that the increased susceptibility of molting hens to SE colonization may be associated with decreased fermentation and production of VFA by cecal bacteria or by a depletion of the number of VFA-producing bacteria present in the ceca. The results further suggest that providing lactose in the drinking water during molting may significantly enhance resistance to SE colonization.

Effect of a Selected Lactobacillus spp.-Based Probiotic on Salmonella enterica Serovar Enteritidis-Infected Broiler Chicks

Abstract The effect of a Lactobacillus spp.–based probiotic (FM-B11TM) on Salmonella enterica serovar Enteritidis (SE) recovery was evaluated in liquid (Expt. 1) and lyophilized (Expt. 2) forms in two separate experiments with two trials each. For each trial, 80 broiler chicks were randomly allocated into two treatments: control and probiotic culture. All chicks were challenged with SE (∼104 colony-forming units [cfu]) upon arrival at our laboratory. In both experiments, probiotic culture was administered in the drinking water for 3 consecutive days at a final concentration of approximately 106 cfu/ml, beginning 1 hr after SE challenge. Cecal tonsils were aseptically removed at 24 and 72 hr postchallenge, followed by enrichment and plating on xylose lactose deoxycholate (XLD) agar for the presence or absence of Salmonella-typical colonies. In Expt. 1, a significant reduction (P < 0.05) in SE-positive samples was observed in both trials at 24 and 72 hr postchallenge. Additionally, in Expt. 2, the lyophilized probiotic decreased (P < 0.05) SE recovery at both 24 and 72 hr postchallenge compared with the control group in trial 1. In trial 2, SE evaluation was performed only at 72 hr after challenge and fewer (P < 0.001) treated samples were positive for SE. Results showed that application of either liquid or lyophilized probiotic culture in the drinking water for 3 consecutive days can help to reduce SE recovery from young birds, although further research is needed to elucidate the mechanism of this response.

Effect of Feed Withdrawal on Campylobacter in the Crops of Market-Age Broiler Chickens

The presence of Campylobacter and Salmonella on poultry meat products remains a significant public health concern. Previous research has indicated that feed withdrawal may significantly increase Salmonella contamination of broiler crops and that crop contents may serve as an important source of Salmonella carcass contamination at commercial processing. The present study evaluated the effect of preslaughter feed withdrawal on the incidence of Campylobacter isolation in crops of market-age commercial broiler chickens prior to capture and transport to the processing plant. The incidence of Campylobacter isolation from the crop was determined immediately before and after feed withdrawal in 40 7-wk-old broiler chickens obtained from each of nine separate broiler houses. Ceca were collected from broilers in six of the same flocks for comparison with the crop samples. Feed withdrawal caused a significant (P < 0.025) increase in Campylobacter-positive crop samples in seven of the nine houses sampled. Furthermore, the total number of Campylobacter-positive crops increased significantly (P < 0.001) from 90/360 (25%) before feed removal to 224/359 (62.4%) after the feed withdrawal period. Alternatively, feed withdrawal did not significantly alter the Campylobacter isolation frequency from ceca. Similar to our previous studies with Salmonella, the present results suggest that preharvest feed withdrawal increases the frequency of Campylobacter crop contamination and, thus, provides a source of Campylobacter contamination of carcasses at commercial processing.

Immunoprophylaxis of Salmonella enteritidis Infection by Lymphokines in Leghorn Chicks

Chickens were treated prophylactically with the soluble products from Con A-stimulated T-lymphocytes from Salmonella enteritidis-infected chickens in order to investigate the effect of such prophylactic treatment on organ invasion by S. enteritidis. At 18 days of age, chicks were injected intraperitoneally with one of the following: A) the Amicon YM 10 ultrafiltrate from immunized chickens, B) the Amicon YM 10 ultraretentate material from immunized chickens, or C) the Amicon YM 10 ultraretentate material from control nonimmune chickens. Thirty minutes after lymphokine injection, all birds were challenged per os with 10(8) colony-forming units of S. enteritidis. At both 1 and 6 days post-challenge, prophylactic treatment of chickens with the immune retentate resulted in a 51-60% reduction in S. enteritidis organ invasion. The reduction in S. enteritidis organ invasion was associated with a significant increase in lamina propria thickness based on morphometric analysis (P < 0.05). These results demonstrate that the prophylactic administration of S. enteritidis-immune lymphokines induces protection against S. enteritidis organ invasion, which is associated with a measurable microanatomical change in the cecal mucosa.

Survival of Salmonella in the Crop Contents of Market-Age Broilers During Feed Withdrawal

Recent studies have indicated that crop contamination increases during preslaughter feed withdrawal and that contaminated crop contents may serve as an important source of Salmonella entry into poultry processing plants. During the present study, we evaluated the effect of preslaughter feed withdrawal on crop pH and Salmonella crop contamination in broilers from three commercial broiler flocks. The effect of experimental feed withdrawal on crop pH, lactic acid concentration, and Salmonella crop contamination was also evaluated in market-age broilers challenged experimentally with Salmonella typhimurium. Crop pH increased significantly (P < 0.05) from 3.64 +/- 0.25 before feed removal to 5.14 +/- 0.72 after 8 hr of feed withdrawal in broilers from commercial flocks. The incidence of Salmonella crop contamination in the commercial broilers increased (P < 0.05) from 3.3% before feed removal to 12.6% after 8 hr of feed withdrawal. Similarly, crop pH increased (P < 0.05) by a magnitude of approximately 1 unit in broilers after 8 hr of experimental feed withdrawal. The population of S. typhimurium in the crops of the experimentally challenged broilers increased (P < 0.05) by approximately 1 log unit during the 8-hr experimental feed withdrawal. The concentration of lactic acid in the crop of the broilers during experimental feed withdrawal decreased (P < 0.01) from a range of 119-135 mumol/ml before feed removal to a range of 22-32 mumol/ml after 8 hr of feed withdrawal. The results indicated that feed withdrawal resulted in a decrease in lactic acid in the crop, accompanied by an increase in crop pH, and an increase in Salmonella crop contamination.

Dosage titration of a characterized competitive exclusion culture to inhibit Salmonella colonization in broiler chickens during growout.

Broiler chicks were spray treated on the day of hatch with titrated dosages (10(6), 10(7), or 10(8) anaerobic CFU) of a characterized competitive exclusion culture (CF3) and challenged orally on day 3 with 10(4) CFU of Salmonella typhimurium. On day 10, cecal contents from control and CF3-treated chicks were cultured for S. typhimurium to determine the minimal efficacious dosage of the CF3 culture. The experiment was repeated in three replicated trials. Resistance to Salmonella cecal colonization was dosage related and progressively enhanced at the 10(7)- and 10(8)-CFU dosages compared with the 10(6)-CFU dosage. The 10(7)-CFU dosage was selected as the minimal effective dosage and evaluated for efficacy during a 43-day broiler growout study. Six hundred broilers were spray treated on the day of hatch and compared with 600 controls. One-half of the control and CF3-treated birds were challenged orally on day 3 with 10(4) CFU of S. typhimurium and designated seeders. The remaining unchallenged birds were designated contacts. Compared with the controls, the recovery of Salmonella cells from the ceca of the CF3-treated broilers was significantly decreased (P < 0.01) in the challenged seeders on days 21 and 43 of growout. Salmonella contamination of floor pen litter was significantly reduced (P < 0.05) in pens of CF3-treated birds compared with controls. The transmission of Salmonella cells from seeder to contact birds in the same pens was decreased significantly (P < 0.01). The results indicated that treatment of broiler chicks on the day of hatch with the 10(7)-CFU dosage of CF3 culture effectively increased resistance to S. typhimurium challenge during growout to market age.

Evidence for Clostridium Septicum as a Primary Cause of Cellulitis in Commercial Turkeys

Etiology and methods of immunoprophylaxis against common field cellulitis in commercial turkeys were evaluated. It was determined that intravenous administration (∼10 8 cells/ml) of 1 of 4 isolates of Clostridium septicum from cellulitis lesions rapidly caused the classic lesions of cellulitis followed by death within 36 hr at high doses. When the supernatant alone was injected into turkey poults, signs of depression and ataxia were temporarily observed for up to 20 hr after injection, but no cellulitis lesions were detected. An enzyme-linked immunosorbent assay was developed to measure antibody levels against the isolated C. septicum. This assay was used to predict susceptibility to infection. An experimental formalin-killed bacterin/toxoid was produced from the challenge strain of C. septicum with inactivation timed to allow toxin accumulation and ∼108 cells/ml. This bacterin/toxoid given at day of hatch generated a rapid and persistent antibody response against the homologous C. septicum in the vaccinated birds at 9 weeks (P < 0.001). The ability of this experimental vaccine to protect birds in the field as well as its ability to evaluate unvaccinated flocks to establish the time of seroconversion and the relationship to clinical disease are currently under evaluation.

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.

Comparison of Prophylactic and Therapeutic Efficacy of Salmonella enteritidis-immune Lymphokines against Salmonella enteritidis Organ Invasion in Neonatal Leghorn Chicks

Investigations in our laboratories have indicated that when Salmonella enteritidis (SE)-immune lymphokines--supernatants from concanavalin-A-stimulated T cells derived from SE-immune adult chickens--were administered intraperitoneally to 1-day-old chicks before SE challenge, they conferred protection against SE organ invasion within 24 hr. This resistance mediated by SE-immune lymphokines was associated with a concomitant increase in peripheral blood polymorphonuclear leukocytes that peaked 4 hr after SE challenge. In the present study, we evaluated efficacy of SE-immune lymphokines in protecting chicks against SE organ invasion and alterations in peripheral blood polymorphonuclear leukocyte counts. Administration of SE-immune lymphokines to chicks either 30 min or 6 days before SE challenge caused a significant reduction in SE organ invasion. However, when SE-immune lymphokines were administered 2 days after SE challenge, there was no reduction in SE organ invasion. Both prophylactic (before SE challenge) and therapeutic (after SE challenge) administration of SE-immune lymphokines caused a significant increase in numbers of peripheral blood polymorphonuclear leukocytes. Results from these studies suggest that SE-immune lymphokines have potential value as an effective prophylactic but not as a therapeutic modulator of early resistance to SE organ invasion in neonatal leghorn chicks.

Detection and partial characterization of an anti-steroidogenic peptide from the humoral immune system of the chicken

Phenomenological association of alterations of immune system function at the time of puberty (e.g. involution of the chicken bursa of Fabricius) has led to postulation that the humoral immune system may negatively affect the hypothalamo-adenohypophyseal-gonadal axis of the neonate. Presently, we examined the effect of an acidic aqueous bursa of Fabricius extract, derived from prepubescent chickens, on in vitro basal and LH-stimulated progesterone biosynthesis by isolated ovarian granulosa cells of the largest preovulatory chicken follicles (F1 and F2). Crude extracts of < 5kDa and > 3kDa inhibited LH-stimulated progesterone secretion (P < 0.05). The bioactive component was observed to be heat labile and is sensitive to the endopeptidases chymotrypsin, trypsin and papain. The peptide is not sensitive to the exopeptidase, aminopeptidase M. Partial purification by reversed phase HPLC resulted in a fraction capable of inhibiting in vitro steroidogenesis. This fraction suppressed LH-stimulated progesterone biosynthesis to approximately basal levels (79% suppression). Following removal of the peptide, granulosa cells were capable of LH-stimulated progesterone biosynthesis similar to control cells. Bursal extract significantly inhibited cAMP analog-stimulated progesterone biosynthesis. These data indicate that the anti-steroidogenic peptide derived from the chicken bursa of Fabricius is a single heat labile, amino terminally blocked peptide with bioactivity independent of the gonadotropin receptor of the granulosa cell.