Transcriptional profiling of cecal gene expression in probiotic- and Salmonella-challenged neonatal chicks.

Probiotics are currently used to improve health and reduce enteric pathogens in poultry. However, the mechanisms by which they reduce or prevent disease are not known. Salmonella are intracellular pathogens that cause acute gastroenteritis in humans, and infections by nontyphoid species of Salmonella also can result in diarrhea, dehydration, and depression in poultry. Frequently, however, no clinical signs of infection are apparent in poultry flocks. In this study, day-of-hatch chicks were challenged with Salmonella enterica serovar Enteritidis (SE) and treated 1 h later with a poultry-derived, Lactobacillus-based probiotic culture (FloraMax-B11, Pacific Vet Group USA Inc., Fayetteville, AR). Cecae were collected 12 and 24 h posttreatment for Salmonella detection and RNA isolation for microarray analysis of gene expression. At both 12 and 24 h, SE was significantly reduced in chicks treated with the probiotic as compared with the birds challenged with only SE (P < 0.05). Microarray analysis revealed gene expression differences among all treatment groups. At 12 h, 170 genes were expressed at significantly different levels (P < 0.05), with a minimum difference in expression of 1.2-fold. At 24 h, the number of differentially regulated genes with a minimum 1.2-fold change was 201. Pathway analysis revealed that at both time points, genes associated with the nuclear factor kappa B complex, as well as genes involved in apoptosis, were significantly regulated. Based on this analysis, probiotic-induced differential regulation of the genes growth arrest-specific 2 (GAS2) and cysteine-rich, angiogenic inducer, 61 (CYR61) may result in increased apoptosis in the cecae of chicks. Because Salmonella is an intracellular pathogen, we suggest that increased apoptosis may be a mechanism by which the probiotic culture reduces Salmonella infection.

Effect of lactic acid bacteria probiotic culture treatment timing on Salmonella Enteritidis in neonatal broilers.

In the present study, a series of experiments were conducted to evaluate the ability of a combination of 3 ATCC lactobacilli (LAB3) or a commercially available probiotic culture (PROB) to reduce Salmonella enterica serovar Enteritidis (Salmonella Enteritidis) in broiler chicks. Additionally, we varied the timing of PROB administration in relationship to Salmonella challenge and determined the influence on recovery of enteric Salmonella. In experiments 1 to 3, chicks were randomly assigned to treatment groups and were then challenged via oral gavage with Salmonella Enteritidis. Chicks were treated 1 h after Salmonella Enteritidis challenge with LAB3 or PROB. Twenty-four hours posttreatment, cecal tonsils were collected for recovery of enteric Salmonella. In experiments 4 to 7, day-of-hatch chicks were randomly assigned to treatment groups and were then treated with PROB via oral gavage and placed into pens. Chicks were challenged with Salmonella Enteritidis 24 h after treatment via oral gavage. At 24 h after Salmonella Enteritidis challenge, cecal tonsils were collected and recovery of enteric Salmonella was determined. In experiments 8 to 10, 1-d-old chicks were randomly assigned to treatment groups and were then challenged via oral gavage with Salmonella Enteritidis and placed into pens. Chicks were treated 24 h after challenge with PROB via oral gavage. Twenty-four hours post PROB treatment, cecal tonsils were collected and enriched as described above. It was found that PROB significantly reduced cecal Salmonella Enteritidis recovery 24 h after treatment as compared with controls or LAB3-treated chicks in experiments 1 to 3 (P<0.05). Administration of PROB 24 h before Salmonella Enteritidis challenge significantly reduced recovery of Salmonella Enteritidis in 2 out of 4 experiments and no reduction in cecal Salmonella Enteritidis was observed when chicks were challenged with Salmonella Enteritidis and treated 24 h later with PROB. These data demonstrate that PROB more effectively reduced Salmonella Enteritidis than LAB3, and the timing of PROB treatment affects Salmonella Enteritidis-associated reductions.

Comparison of beak-trimming methods on early broiler breeder performance.

Beak trimming is necessary in commercial broiler breeders to prevent or decrease trauma as they mature. Two common beak-trimming methods were evaluated by early performance comparison with nontrimmed chicks (NBT). The robotic electrocautery device (ECD) trims and cauterizes the beak tip. The robotic infrared beak-trimming device (IBT) applies an infrared light beam to destroy the live basal tissue while leaving the hard corneum intact for the first approximately 10 d. In 2 experiments, day-of-hatch Ross 708 by-product chicks were obtained from a local hatchery, where 1/3 of the chicks were trimmed using IBT. All chicks were then transported to another hatchery where 1/3 were trimmed using ECD and 1/3 were NBT. Personnel at each hatchery were highly experienced and skilled with their respective technique. All chicks were then transported to University of Arkansas facilities. Before placement in each experiment, chicks were individually neck-tagged and weighed, and in experiment 1, beaks were measured using a digital caliper. A small but significant transient reduction in BW gain was observed at 14 d due to ECD as compared with NBT controls, although ECD was not different than IBT in experiment 1. In experiment 2, IBT birds were significantly heavier at 11 d by 7.8 and 8.7 g than the NBT or ECD, respectively. However, at d 21 and 42, no significant differences in BW or BW gain were observed. When beak trimming was performed on day of hatch by skilled and experienced personnel, little measurable effect on early performance was observed during the first 6 wk of life. Decreased broiler performance is generally considered a sensitive indication of physical or psychogenic stress. Given the marked reduction in beak-inflicted trauma with beak trimming birds as they reach sexual maturity, these results suggest that when properly performed, neither of these beak-trimming methods causes sufficient physical or psychogenic stress to markedly affect early growth rate.

Evaluation of Salmonella-lytic properties of bacteriophages isolated from commercial broiler houses.

Because of recent interest in bacteriophage therapy in poultry, information regarding the interaction of bacteriophages and potential host bacteria in the environment should be collected. The present studies were initiated with a rather typical commercial broiler integrator within the south-central United States to examine environmental Salmonella levels in two broiler complexes, attempt to isolate Salmonella-lytic bacteriophages, and elucidate a possible reason for differing apparent Salmonella prevalence. Significantly (P < 0.05) less Salmonella was isolated from houses in complex 1 (15/44 [34%] Salmonella-positive drag swabs) as compared to houses in complex 2 (22/24 [92%]). A total of seven Salmonella-lytic bacteriophages were isolated from Salmonella-positive environments, and two bacteriophages were isolated from a single Salmonella-negative house. During the initial bacteriophage isolation, individual bacteriophages did not replicate in the Salmonella host isolated from the same environment, and lysis of additional Salmonella hosts relied on high numbers of bacteriophage to be present. This suggests that the presence of these bacteriophages in the environment of a commercial broiler house had little to no effect on the presence of Salmonella. This study highlights the need to find additional bacteriophage sources, more effective isolation methods, and more innovative approaches to using bacteriophages to treat enteric disease.

Evaluation of a Lactobacillus-based probiotic culture for the reduction of Salmonella enteritidis in neonatal broiler chicks.

We evaluated the effect of a Lactobacillus-based probiotic culture (FM-B11) for reduction of Salmonella Enteritidis in neonatal broiler chicks. In all experiments, chicks were challenged with approximately 10(4) cfu of Salmonella Enteritidis upon arrival at our laboratory, and the treatments were administered 1 h postchallenge. Cecal tonsil samples were obtained 24 h posttreatment and enriched for Salmonella Enteritidis recovery. The first experiment compared the effects of oral administration of doses of 10(4), 10(6), and 10(8) cfu/chick. In this experiment, doses of 10(6) and 10(8) both significantly reduced Salmonella Enteritidis recovery compared with controls (15 vs. 85% Salmonella Enteritidis positive), but 10(4) cfu did not significantly reduce Salmonella Enteritidis recovery. The second experiment compared the efficacy of oral administration of the live probiotic culture, with or without supernatant removed, to inactivated cultures or supernatant alone. Live probiotic organisms, with or without supernatant, significantly reduced Salmonella Enteritidis in this experiment, but inactivated or cell-free treatments did not reduce Salmonella Enteritidis. In the final 2 experiments, differing doses of probiotic culture were administered on the vent lips, where the treatment was taken into the lower gastrointestinal tract via cloacal drinking. Concentrations of probiotic culture from 10(2) to 10(7) cfu/chick significantly reduced Salmonella Enteritidis, and there was no difference in Salmonella Enteritidis recovery between treatment concentrations. These data suggest that this Lactobacillus-based probiotic culture may be efficacious for reduction of Salmonella Enteritidis in neonatal chicks.

Effect of probiotic treatment in broiler chicks on intestinal macrophage numbers and phagocytosis of Salmonella enteritidis by abdominal exudate cells.

Previous data have indicated that a Lactobacillus-based probiotic culture (FM-B11) is efficacious in reducing Salmonella Enteritidis colonization within 24 h when administered within 1 h of challenge. We hypothesized that the innate immune system, specifically macrophages, may play a role in the observed reduction of Salmonella Enteritidis colonization with probiotic treatment. Day-of-hatch chicks were challenged with Salmonella Enteritidis and then treated with the probiotic culture 1 h later. Three other treatment groups were not treated (negative control), challenged only, or treated with probiotic only. In all experiments, probiotic treatment on the day of hatch reduced (P < 0.05) cecal Salmonella Enteritidis recovery as compared with the control treatment. In experiments 1 and 2, immunohistochemistry was used to evaluate the presence of macrophages (KUL01+) in the ileum and cecum of 7 to 10 chicks per group at 24 h posttreatment. In experiment 1, the number of macrophages observed per 10,000 microm(2) in the ileum of Salmonella Enteritidis-challenged chicks was higher (P < 0.05) than that of nonchallenged chicks (4.87 +/- 0.31 vs. 3.05 +/- 0.19). In the cecum, there were more (P < 0.05) macrophages per 10,000 microm(2) in chicks receiving probiotic treatment without challenge than in negative control chicks (5.32 +/- 0.41 vs. 3.66 +/- 0.35). However, in experiment 2 we found no differences among treatments in the numbers of macrophages for both the ileum and cecum. Experiments 3 and 4 were performed to evaluate the ability of Sephadex-elicited abdominal exudate cells (AEC) from chicks to phagocytose Salmonella Enteritidis in vitro. Abdominal exudate cells were isolated from the abdominal cavity, maintained in tissue culture plates overnight, and then assayed for phagocytic activity by coincubating with Salmonella Enteritidis. In experiment 3, more (P < 0.05) Salmonella Enteritidis was recovered from AEC derived from probiotic-treated chicks than in any other treatment. However, in experiment 4, all treatments resulted in similar levels of elicited AEC, and phagocytosis of Salmonella Enteritidis was at low levels in all groups. Although not conclusive, the modest differences detected in experiments 1 and 3, and the fact that those differences were not repeatedly detectable, suggest that these macrophage-related changes were not solely responsible for the reductions of Salmonella Enteritidis following probiotic treatment.

Ability of bacteriophages isolated from different sources to reduce Salmonella enterica serovar enteritidis in vitro and in vivo.

Salmonella enterica serovar Enteritidis-lysing bacteriophages isolated from poultry or human sewage sources were used to reduce Salmonella Enteritidis in vitro and in experimentally infected chicks. Cocktails of 4 different bacteriophages obtained from commercial broiler houses (CB4Ø) and 45 bacteriophages from a municipal wastewater treatment plant (WT45Ø) were evaluated. In experiment 1, an in vitro crop assay was conducted with selected bacteriophage concentrations (10(5) to 10(9) pfu/mL) to determine ability to reduce Salmonella Enteritidis in the simulated crop environment. Following 2 h at 37 degrees C, CB4Ø or WT45Ø reduced Salmonella Enteritidis recovery by 1.5 or 5 log, respectively, as compared with control. However, CB4Ø did not affect total SE recovery after 6 h, whereas WT45Ø resulted in up to a 6-log reduction of Salmonella Enteritidis. In experiment 2, day-of-hatch chicks were challenged orally with 3 x 10(3) cfu/chick Salmonella Enteritidis and treated cloacally with 1 x 10(9) WT45Ø pfu/chick 1 h postchallenge. One hour later, chicks were treated or not with a commercially available probiotic (Floramax-B11). Both treatments significantly reduced Salmonella Enteritidis recovery from cecal tonsils at 24 h following vent lip application as compared with controls, but no additive effect was observed with the combination of bacteriophages and probiotic. In experiment 3, day-of-hatch chicks were challenged orally with 9 x 10(3) cfu/chick Salmonella Enteritidis and treated via oral gavage with 1 x 10(8) CB4Ø pfu/chick, 1.2 x 10(8) WT45Ø pfu/chick, or a combination of both, 1 h postchallenge. All treatments significantly reduced Salmonella Enteritidis recovered from cecal tonsils at 24 h as compared with untreated controls, but no significant differences were observed at 48 h following treatment. These data suggest that some bacteriophages can be efficacious in reducing SE colonization in poultry during a short period, but with the bacteriophages and methods presently tested, persistent reductions were not observed.

Temporal effects of lactic acid bacteria probiotic culture on Salmonella in neonatal broilers.

We evaluated the ability of a commercially available lactic acid bacteria-based probiotic culture (LAB) to reduce Salmonella Enteritidis or Salmonella Typhimurium in day-of-hatch broiler chicks. In these experiments, chicks were challenged with Salmonella Enteritidis or Salmonella Typhimurium and treated with LAB 1-h postchallenge. Following treatment, cecal tonsils and ceca were aseptically collected for Salmonella Enteritidis or Salmonella Typhimurium enrichment or Salmonella Enteritidis enumeration, respectively. In experiments 1 to 3, LAB significantly reduced the incidence of Salmonella Enteritidis (60 to 70% reduction) or Salmonella Typhimurium (89 to 95% reduction) recovered from the cecal tonsils of day-old broiler chicks 24 h following treatment as compared with controls (P < 0.05). Additionally, administration of LAB caused a >2.9 log(10) reduction of total cecal Salmonella Enteritidis recovered 24 h following treatment as compared with controls (P < 0.05). In experiments 4 to 7, upon sample enrichment LAB significantly reduced the recovery of Salmonella Enteritidis from the cecal tonsils at 24 h, but not 6 or 12 h posttreatment (P < 0.05). However, in experiments 6 and 7, when total cecal Salmonella Enteritidis recovery was enumerated, a significant treatment-associated reduction was observed 12 h posttreatment, although in cecal tonsil samples there was no difference in Salmonella Enteritidis incidence at 12 h (P < 0.05). In these studies, LAB treatment significantly reduced recovery of Salmonella in day-of-hatch broilers.

Use of a specific bacteriophage treatment to reduce Salmonella in poultry products.

Bacteriophages represent a group of viruses that specifically infect and replicate in bacteria and could potentially be used to reduce recovery of Salmonella from poultry carcasses. Bacteriophages were isolated from municipal wastewater in the presence of Salmonella enteritidis phage type 13A (SE). In the first 2 experiments, commercially processed broiler carcass rinse water was pooled and divided. The addition of 10(10) pfu/mL of a single bacteriophage (PHL 4) with selected concentrations of SE reduced (P < 0.05) frequency of SE recovered as compared with the control rinse water sample. In experiments 3 and 4, broiler carcasses were intentionally inoculated with SE, sprayed with selected concentrations of PHL 4, and rinsed for SE enrichment and isolation. Application of 5.5 mL of 10(8) or 10(10) pfu/mL of PHL 4 reduced (P < 0.05) the frequency of SE recovery as compared with controls. In experiments 5 and 6, commercially processed turkeys were rinsed with water containing 72 wild-type bacteriophages isolated against SE, which were amplified in SE, or the Salmonella isolated antemortem from drag swabs from the flock selected for in-plant treatment, or a combination of bacteriophages amplified by each bacterial host. All bacteriophage treatments reduced (P < 0.05) frequency of Salmonella recovery as compared with controls. Sufficient concentrations of an appropriate bacteriophage, or a bacteriophage mixture, can significantly reduce recoverable Salmonella from carcass rinses.

In situ detection and quantification of bursa of fabricius cellular proliferation or apoptosis in normal or steroid-treated neonatal chicks.

Apoptosis, or programmed cell death, is believed to be the mechanism for depletion of lymphocytes recognizing self-antigens following clonal expansion in the bursa of Fabricius. Although bursal apoptosis has previously been shown to increase following in vivo exposure to glucocorticoids, the microanatomical site of induced or normal apoptosis has not been unequivocally established. Presently, we adapted the existing terminal deoxynucleotidal transferase-mediated dUTP nick-end labeling (TUNEL) assay for use with neonatal bursae. Similar to previous reports, TUNEL revealed that normal apoptosis is preferentially, but not exclusively, ongoing in bursal follicular cortical cells. Administration of a single dose of a synthetic glucocorticoid (dexamethasone) or androgen (19-nortestosterone) did not significantly (P < 0.05) alter follicular lymphocyte numbers or apoptosis per unit of area at the time points evaluated post-administration (6 or 24 h). However, administration of 19-Nortestosterone increased the interfollicular epithelial thickness, a change usually associated with edema, within 6 h following treatment. Additionally, administration of the androgen 19-nortestosterone significantly decreased the number of proliferating cells as detected using mouse anti-proliferating cell nuclear antigen (PCNA) as a primary immunohistochemical antibody. In normal (control) bursal sections, occasional follicles consisting of predominantly apoptotic cells were observed (0.26% of follicles). Such follicles were consistently one-tenth the area of normal follicles. This incidental finding may suggest occasional occurrence of a common signal for deletion, such as a common integral or clonal mistake, viral infection, or an aberrant paracrine signal.

A multiple-level analysis of hospital team effectiveness.

The purpose of this study was to identify factors at organizational, team, and individual levels that are most predictive of quality improvement team effectiveness. Numerous studies have been conducted on the use of teams in health care. The majority of these studies have focused on organizational-level issues. A few others have focused on team-level issues or individual-level issues. The authors believe that successful use of teams requires a more integrated approach. This study addresses this need by providing a multiple-level analysis of teams in three hospitals. Structured interviews were conducted with hospital administrators as well as quality improvement representatives and team leaders, and written questionnaires were administered to team members. Eight factors were related significantly to the effectiveness of team effort: frequency of team meetings, hours per week conducting team activities, willingness of members to serve on team, selection method of team members, communication of team plans, team member position/composition, team leader performance, and facilitator performance.

Best-practice guidelines for utilizing facilitators.

It appears that the majority of hospitals and other health care organizations use facilitators in team development. While some organizations use facilitators more extensively than others, facilitators appear to serve pivotal roles for teamwork in most health care organizations. A review of the literature, however, suggests that there is no clear consensus regarding the best way to use facilitators. This lack of consensus is, in part, due to the lack of clear best-practice guidelines for utilizing facilitators in health care. The purpose of this article is to provide such guidelines.

The effective team member: avoiding team burnout.

This article outlines specific suggestions for team members designed to help ensure that team membership is a satisfying experience. The suggestions offered provide clear guidelines for the responsibilities individual health care providers must assume when working on teams. Your proactive engagement in addressing the suggestions provided is part of an integrated, holistic approach to teams.

Reproducibility and variation of corneal thickness in different locations in the cornea as measured by an ultrasonic pachymeter.

Corneal thickness was measured in nine positions of nineteen normal eyes with an ultrasonic pachymeter. A standardized protocol was used to examine for intraobserver and interobserver variation, day to day variation, and time of day variation in the measurement of corneal thickness. No statistically significant difference was noted for most points in intraobserver, interobserver, day to day, or different time of day variation measurements. However, it was noted that central measurements of corneal thickness tended to be less variable (more reproducible) than paracentral (p = .0366) and peripheral (.0032) measurements. Paracentral measurements were less variable than peripheral measurements (p = .0323). While this was a small study, 95% confidence intervals were also calculated for each of the nine corneal positions where measurements were performed.

Management of contact lens associated Acanthamoeba keratitis.

Four patients with contact lens associated Acanthamoeba keratitis were treated at Wills Eye Hospital between 1987 and 1989. Two patients had used daily wear soft contact lenses and two had worn hard contact lenses. All four patients were treated with topical antibiotics (propamidine and neomycin), oral ketoconazole, and varying amounts of topical steroids. Two of these patients received topical clotrimazole. All four patients eventually required surgery for either control of their infection (three cases) or control of secondary disease (glaucoma and cataract). Medical therapy for Acanthamoeba keratitis remains problematic, and therefore prevention of this disease by avoidance of risk factors remains paramount.

Direct polyclonal activation of human B lymphocytes by the acquired immune deficiency syndrome virus.

When B lymphocytes from normal human peripheral blood were incubated for 1 hour with the retrovirus that causes the acquired immune deficiency syndrome (AIDS), the B cells showed marked proliferation and differentiation. Proliferative responses to the virus peaked on day 4 and appeared to be independent of accessory cells. This finding was repeated with three separate viral isolates, one of which was from a patient from Zaire. The magnitude of the observed responses was comparable to that seen with standard polyclonal B-cell activators. This phenomenon may be at least partially responsible for the polyclonal B-cell activation seen in patients with AIDS.