Colin R. Young

The Absence of Cecal Colonization of Chicks by a Mutant of Campylobacter jejuni not Expressing Bacterial Fibronectin-Binding Protein

Campylobacter jejuni is a common cause of human gastrointestinal illness throughout the world. Infections with C. jejuni and Campylobacter coli are frequently acquired by eating undercooked chicken. The ability of C. jejuni to become established in the gastrointestinal tract of chickens is believed to involve binding of the bacterium to the gastrointestinal surface. A 37-kD outer membrane protein, termed CadF, has been described that facilitates the binding of Campylobacter to fibronectin. This study was conducted to determine whether the CadF protein is required for C. jejuni to colonize the cecum of newly hatched chicks. Day-of-hatch chicks were orally challenged with C. jejuni F38011, a human clinical isolate, or challenged with a mutant in which the cadF gene was disrupted via homologous recombination with a suicide vector. This method of mutagenesis targets a predetermined DNA sequence and does not produce random mutations in unrelated genes. The parental C. jejuni F38011 readily colonized the cecum of newly hatched chicks. In contrast, the cadF mutant was not recovered from any of 60 chicks challenged, indicating that disruption of the cadF gene renders C. jejuni incapable of colonizing the cecum. CadF protein appears to be required for the colonization of newly hatched leghorn chickens.

Detection of salmonella in poultry using a silicon chip-based biosensor.

Salmonella typhimurium was detected to levels as low as 119 CFUs using the Threshold Immunoassay System. This immunoassay system utilizes solution-based binding of the biotin and fluorescein labeled antibodies to salmonella, followed by filtration-capture of the immunocomplex on a biotin-coated nitrocellulose membrane. Lastly, an anti-fluorescein urease conjugate is bound to the immunocomplex. Detection of the bound immunocomplex is made possible via the silicon chip-based light-addressable potentiometric sensor. In the presence of the urea, urease converts the substrate to ammonia and CO2 and this results in a pH change at the silicon surface. The resultant pH change is monitored with time and the signal output is reported in microV s(-1). An experiment whereby chicken carcass washings were fortified with salmonella showed a recovery of 90%, indicating that the technique can be used to test for salmonella under these conditions. Precautions must be used with this instrument as sample debris will affect sample flow through the membrane and hence the signal output.

Dose response and organ invasion of day-of-hatch Leghorn chicks by different isolates of Campylobacter jejuni.

Colonization of the ceca and organ invasion by different isolates of Campylobacter jejuni were investigated in day-of-hatch leghorn chicks. This model of Campylobacter colonization of the ceca demonstrates that 1) day-of-hatch birds do not naturally contain cecal Campylobacter, 2) ceca can be colonized with C. jejuni by oral gavage and not by cloacal inoculation; 3) C. jejuni can be recovered from the ceca up until at least 7 days postinoculation, 4) cecal colonization occurs when as little as 10(2) colony-forming units is orally inoculated into chicks, and 5) different C. jejuni isolates vary both in their ability to colonize the ceca and in their ability to invade the liver. These studies demonstrate that we have a working animal model for Campylobacter colonization for day-of-hatch chicks. This animal model is being used to examine intervention strategies such as vaccines by which Campylobacter can be reduced or removed from the food animal.

Development of a Cross-Reactive Monoclonal Antibody to Sulfonamide Antibiotics: Evidence for Structural Conformation-Selective Hapten Recognition

A unique anti-sulfonamide antibody-secreting hybridoma that cross-reacts with several sulfonamides was isolated. This was possible by using a N-sulfanilyl-4-aminobenzoic acid hapten-protein conjugate as the immunogen. Most of the antibodies that were detected in immunized mice did not recognize the free drug. However, by screening a large number of antibody-secreting hybridomas, cell lines were isolated that produced antibodies which recognized the free drug. The sensitivities of one such monoclonal antibody (MAb), referred to as Sulfa-1, for sulfanitran, sulfapyridine and sulfathiazole (expressed as IC50 values), were 1.41, 22.8, and 322 ng ml−1, respectively. Molecular modeling studies showed that the calculated minimum energy conformation of the hapten used as immunogen was different than those of the cross-reactive drugs. It was postulated that the MAb was derived from a cell line responsive to a form of the immunogen in which the structural conformation of the hapten was different than the molecular ...

Inoculation of Chicks with Viable Non-Colonizing Strains of Campylobacter jejuni: Evaluation of Protection Against a Colonizing Strain

We have treated chicks with viable non-colonizing mutant strains of Campylobacter jejuni to test these as a possible vaccine. We found that intramuscular inoculation with and without adjuvant, and with or without a concomitant oral dose of non-colonizing strains, failed to provoke protective immunity.

Effects of feed withdrawal and transport on cecal environment and Campylobacter concentrations in a swine surgical model

The objective of the present study was to evaluate how feed withdrawal and transportation influenced the cecal environment and cecal populations of Campylobacter in swine. Four miniature Yucatan gilts (8.8 kg), naturally infected with Campylobacter jejuni, were surgically implanted with cecal cannulas. The gilts were fasted for 48 h. Samples of cecal contents were collected for 7 days prior to and for 7 days after the fast, and mean values were determined for pH, volatile fatty acids (VFA), and CFU enumeration of C. jejuni. This was replicated three times. In another trial, gilts (full-fed) were transported in a livestock trailer for 4 h and cecal samples were collected before and after transport and analyzed for pH, VFA, and CFU. Following a 48-h fast, cecal pH increased (P < 0.05) by 1 unit; acetic and propionic acids decreased (P < 0.05) by 61% and 71%, respectively; and there was a twofold log10 increase (P < 0.05) in CFU/g cecal content of C. jejuni. Values of pH, VFA, and CFU of C. jejuni did not change in cecal samples from gilts following transportation. These data are important for food safety considerations because feed withdrawal, commonly associated with shipping and slaughter, can increase Campylobacter concentrations in the pig intestinal tract.

Diminution of Campylobacter colonization in neonatal pigs reared off-sow.

Pigs may be a natural reservoir of Campylobacter and can be colonized as early as 24 h after birth. The purpose of the present study was to evaluate what effect early removal of piglets from Campylobacter-positive sows has on Campylobacter prevalence in neonates. In two trials, piglets were removed from sows within 24 h of birth and were reared in nurseries isolated from sows for 21 days. From the neonates rectal swabs were cultured for Campylobacter, and Campylobacter status of the isolated piglets was compared to that of littermates reared on sows. The nurseries consisted of wire-floored farrowing crates that were equipped with heaters and self-feeders. In trial I, the Campylobacter prevalence in nursery-reared piglets was 13 of 14 on day 2 and 0 of 14 on day 20. Campylobacter prevalence in the sow-reared piglets was 8 of 9 from days 2 to 20. In trial II, 12 of 29 on day 2, and 5 of 26 on day 20, of the nursery-reared piglets were culture positive for Campylobacter. For the sow-reared piglets, Campylobacter prevalence was 7 of 15 on day 1 and 15 of 15 (100%) on day 20. These data suggest that successful permanent colonization of the gut by Campylobacter is probably related to constant exposure of piglets to Campylobacter-positive feces. Campylobacter prevalence may be diminished in neonates that are reared off-sow in isolated nurseries.

Cecal Colonization of Chicks by Porcine Strains of Campylobacter coli

SUMMARY. Ten genotypically distinct strains of Campylobacter coli were isolated from a swine production facility. These porcine isolates were then orally inoculated into day-of-hatch leghorn chicks and were excellent colonizers of the chick cecum. Campylobacter coli recovered from inoculated chickens were genotypically identical to the challenge strain. The absence of host specificity suggests a possible movement of strains among swine, field animals and birds, and poultry houses.

Anti‐Inflammatory Effects of Ergotamine in Steers

The objective of this experiment was to investigate whether the ergot alkaloid, ergotamine (ET), an alkaloid used to model fescue toxicosis in cattle, modifies the response of cattle to endotoxin (LPS) challenge. Steers (n = 16) were divided into the following treatment groups: control (C), ergotamine (ET), endotoxin (LPS), and ET + LPS. ET and ET + LPS groups received a single bolus intravenous injection of ET (40 microg. kg. body wt(-1)), whereas C and LPS steers received a single bolus injection of sterile vehicle. Thirty minutes after ET/vehicle administration, a single bolus intravenous injection of LPS (0.2 microg. kg. body wt(-1)) was given. Blood was collected at various time points for 48 hr post. Endotoxin increased rectal temperature (RT) and the circulating levels of tumor necrosis factor-alpha (TNF-alpha), cortisol, haptoglobin (Hp), thromboxane B(2) (TXB(2)). The circulating Hp, TNF-alpha, and TXB(2) increases were blunted by pretreatment with ET compared with ET + LPS. Ergotamine by itself increased circulating cortisol and RT, whereas it decreased serum prolactin (PRL). Therefore, whereas administration of LPS at 0.2 microg/kg to steers resulted in an expected response, the combination of ET + LPS attenuated major effects of LPS alone. Thus, acute administration of ET appeared to be anti-inflammatory as it decreased the inflammatory response to LPS, an effect likely driven at least in part by the ET-caused cortisol increase.

Anti-Inflammatory Effects of Ergotamine in Steers (44562)

The objective of this experiment was to investigate whether the ergot alkaloid, ergotamine (ET), an alkaloid used to model fescue toxicosis in cattle, modifies the response of cattle to endotoxin (LPS) challenge. Steers (n = 16) were divided into the following treatment groups: control (C), ergotamine (ET), endotoxin (LPS), and ET + LPS. ET and ET + LPS groups received a single bolus intravenous injection of ET (40 μg · kg · body wt-1), whereas C and LPS steers received a single bolus injection of sterile vehicle. Thirty minutes after ET/vehicle administration, a single bolus intravenous injection of LPS (0.2 μg · kg · body wt-1) was given. Blood was collected at various time points for 48 hr post. Endotoxin increased rectal temperature (RT) and the circulating levels of tumor necrosis factor-α (TNF-α), cortisol, haptoglobin (Hp), thromboxane B2 (TXB2). The circulating Hp, TNF-α, and TXB2 increases were blunted by pretreatment with ET compared with ET + LPS. Ergotamine by itself increased circulating cortisol and RT, whereas it decreased serum prolactin (PRL). Therefore, whereas administration of LPS at 0.2 μg/kg to steers resulted in an expected response, the combination of ET + LPS attenuated major effects of LPS alone. Thus, acute administration of ET appeared to be anti-inflammatory as it decreased the Inflammatory response to LPS, an effect likely driven at least in part by the ET-caused cortisol increase.