Non-O157 verotoxigenic Escherichia coli and beef: A Canadian perspective
Alexander Gill, Colin O. Gill

Abstract — Verotoxigenic Escherichia coli (VTEC) are important foodborne pathogens in Canada. VTEC of the O157:H7 serogroup have been the focus of regulatory action and surveillance in both Canada and the USA, due to their role in a number of high profile outbreaks. However, there is increasing evidence that other VTEC serogroups cause a substantial proportion of human illness.

This issue is of particular importance to the cattle industry due to the role of beef as a vehicle for VTEC transmission. In this review, the evidence for non-O157 VTEC as cause of human illness in Canada and the potential for Canadian beef and cattle to serve as a source of VTEC are presented. In addition, the available strategies for the control of VTEC in cattle and beef are discussed.

Clinical signs and their association with herd demographics and porcine reproductive and respiratory syndrome (PRRS) control strategies in PRRS PCR-positive swine herds in Ontario
Beth Young, Cate Dewey, Zvonimir Poljak, Thomas Rosendal, Susy Carman

Abstract —The purposes of this study were to describe the clinical signs observed in PRRS positive herds during a porcine reproductive and respiratory syndrome (PRRS) outbreak in Ontario and to determine associations between these clinical signs and herd demographics and PRRS control strategies. All PRRS polymerase chain reaction- (PCR)-positive submissions to a diagnostic laboratory between September 1, 2004 and August 31, 2007 were identified (n = 1864). After meeting eligibility requirements and agreeing to voluntary study participation, producers from 455 of these submissions were surveyed for information on clinical signs observed in their herds, herd demographics, and PRRS control strategies used in their herds at the time that the PCR-positive samples were taken. Larger herd size was associated with an increased risk of reporting abortion, weakborn piglets, off-feed sows, and sow mortality in sow herds, and with an increased risk of reporting mortality in finishing herds. When disease control strategies were examined, use of a commercial PRRS vaccine in sows and gilts was associated with a decreased risk of reporting weakborn pigs and high pre-weaning mortality, while the use of serum inoculation in breeding animals was associated with an increased risk of reporting off-feed sows and sow mortality. Providing biofeedback of stillborn/mummified piglets, placenta or feces to gilts was associated with an increased risk of reporting respiratory disease and mortality in finishing pigs while all-in/all-out flow in farrowing rooms was associated with an increased risk of reporting sow mortality and weakborn piglets.

Relationship between estimated finishing-pig space allowance and in-transit loss in a retrospective survey of 3 packing plants in Ontario in 2003
Charles Haley, Catherine E. Dewey, Tina Widowski, Robert Friendship

Abstract — The objective of this study was to determine the association between space allowance and in-transit loss of finishing pigs going to select abattoirs in Ontario during summer weather conditions. The study included data from 2- or 3-tiered trailers transporting = 130 pigs in June, July, and August 2003 to 3 packers that processed 76% of Ontario market pigs. Daily in-transit loss data were merged with packer data to determine the number of pigs on each trailer. Space allowance (in square meters per pig) was estimated from the percentage of each trailer’s capacity that was filled by the load size. Actual pig weights were not available. Hourly temperature and relative humidity were obtained from 2 local Ontario weather stations. In-transit loss increased with environmental temperature, by 6.6 times at temperatures between 28°C and 34.2°C compared with < 17°C. At space allowances between 0.44 and 0.43 m2/pig compared with = 0.515 m2/pig, in-transit losses increased 2.12 times when environmental temperatures were < 21°C. Temperature is likely a more important determinant of in-transit loss than space allowance. However, in-transit losses in hot weather are likely to be reduced by increasing space allowance or by adding a cooling device.

Immune response following vaccination against Salmonella Enteritidis using 2 commercial bacterins in laying hens
Thi Q.L. Tran, Sylvain Quessy, Ann Letellier, Annie Desrosiers, Martine Boulianne

Abstract — The humoral and cell-mediated immune (CMI) response to 2 commercial killed Salmonella Enteritidis (SE) vaccines (Layermune and MBL SE4C) was evaluated in laying hens. Layers were distributed in 2 experimental groups. The first received a single immunization at 16 wk of age, while the second experimental group was immunized at 12 wk of age and again at 18 wk of age. Serum immunoglobulin (Ig)G antibodies were measured using a commercial SE ELISA kit and showed persistent levels from 3 to 32 and 34 wk post-vaccination. The vaccination protocol using 2 immunizations showed a higher seroconversion level than the single vaccination. However, our results for bacterial intracellular survival indicated that IgG titers were not linked with bacterial killing. Local IgA production was measured in the intestines and oviducts with an in-house SE whole cell antigen ELISA. Only the MBL SE4C vaccine elicited IgA antibody production when tested on intestine and oviduct mucosal secretions, 3-weeks post-vaccination in both immunization protocol groups. To evaluate the CMI response, the splenic T-cells and B-cells populations were analyzed using flow cytometry. The CD3/B-cell ratio decreased 3 wk after the second immunization in the twice vaccinated Layermune group due to an increase in B-cells.

Expression of vascular endothelial growth factor and endothelial nitric oxide synthase is increased in the placenta of sheep at high altitude in the Andes
Víctor H. Parraguez, Miljenko A. Atlagich, Bessie Urquieta, Marco Galleguillos, Mónica De los Reyes, David L. Kooyman, Silvia Araneda, Luis A. Raggi

Abstract — Fetal weight and the placenta of sheep at high altitude (HA) are affected by hypoxia. Placental changes (an increase in placental size and vascularization) are greater in ewes from populations that have lived for several generations at HA than in those exposed during just 1 gestation. This study investigated placental expression of vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS), 2 molecules involved in placental angiogenesis that could be upregulated by hypoxia. Two groups of ewes were maintained at HA (3589 m) during pregnancy: HA-native ewes (group HH) and ewes native to lowlands but moved to HA immediately after the diagnosis of pregnancy (group LH). A control group (LL) was kept at sea level. Near term, placentomes were removed, weighed, and processed for immunohistochemical detection of VEGF and eNOS, as well as for vascular area measurement. Placental weight was significantly higher in the HH group than in the LH and LL groups; between the latter 2 groups there was no significant difference. The placental area occupied by vasculature was significantly greater in both the HA groups than in the LH group; the number of placentomes was greatest in the LL group. The density of VEGF and eNOS in the placentome tissue was significantly greater in both HA groups than in the LL group. Although the density of VEGF was significantly lower in the HH group than in the LH group, no differences were observed in eNOS density between the HH and LH animals. These results demonstrate that chronic hypoxia upregulates the expression of placental VEGF and eNOS, suggesting an important role of these molecules in the placental response to HA hypoxia. In addition, an attenuated response to hypoxia in VEGF synthesis may be part of the long-term process of adaptation to HA.

Effects of ketamine-xylazine intravenous bolus injection on cardiovascular function in rabbits
Christine Baumgartner, Melanie Bollerhey, Johanna Ebner, Lién Laacke-Singer, Tibor Schuster, Wolf Erhardt

Abstract —The direct effects of ketamine-xylazine (KET-XYL) on vascular function have not been investigated in rabbits. The short-term cardiovascular effects of intravenous (IV) KET-XYL bolus injection, therefore, should be investigated using vascular ultrasonography.

In this prospective experimental study, KET-XYL anesthesia was induced IV in 9 female New Zealand White rabbits before 3 defined test bolus injections of KET-XYL were given IV. Before and for 10 min after each KET-XYL injection vascular and hemodynamic variables were recorded at the left common carotid artery (ACC) after the 1st injection, and at the abdominal aorta (AA) after the 2nd injection. Echocardiography was performed after the 3rd injection to investigate changes in cardiac parameters.

Ketamine-xylazine IV caused a significant increase in vessel diameter at the ACC and AA. Average volumetric flow significantly decreased at the ACC and pulsatility index significantly decreased at the AA. Fractional shortening (FS) and heart rate significantly decreased, while mean arterial blood pressure initially increased.

Bolus injections of KET-XYL IV produced a transient vasodilatation at the ACC and AA. Despite central vasodilatation, bradycardia, and decrease of FS and average volumetric flow (VFave), mean arterial blood pressure did not significantly decrease indicating well-preserved cardiovascular compensatory mechanism after the ratio and doses of KET-XYL IV bolus injections used in this study.

Prediction of serum ionized calcium concentration by serum total calcium measurement in cats
Patricia A. Schenck, Dennis J. Chew

Abstract — Feline serum samples (n = 434) were classified as hypercalcemic, normocalcemic, or hypocalcemic based on both total calcium (tCa) and ionized calcium (iCa) concentrations. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), positive diagnostic likelihood ratio (PDLR), and negative diagnostic likelihood ratio (NDLR) were calculated for prediction of hypercalcemia and hypocalcemia in all samples, in hypoalbuminemic cats, and in those with chronic renal failure (CRF) as compared with cats that had other conditions. Diagnostic discordance in prediction of iCa using tCa was 40%. Sensitivity of tCa in prediction of ionized hypercalcemia was low and specificity was high. The PDLR for prediction of ionized hypercalcemia or hypocalcemia was low in all cats, especially in those with CRF. Due to the high level of diagnostic discordance, tCa should not be used to predict iCa concentration. Concentration of iCa should be measured directly when accurate assessment of calcium status is needed.

A comparison of the immune responses of dogs exposed to canine distemper virus (CDV) — Differences between vaccinated and wild-type virus exposed dogs
Danielle Perrone, Scott Bender, Stefan Niewiesk

Abstract — Canine distemper virus (CDV)-specific immune response was measured in different dog populations. Three groups of vaccinated or wild-type virus exposed dogs were tested: dogs with a known vaccination history, dogs without a known vaccination history (shelter dogs), and dogs with potential exposure to wild-type CDV. The use of a T-cell proliferation assay demonstrated a detectable CDV-specific T-cell response from both spleen and blood lymphocytes of dogs. Qualitatively, antibody assays [enzyme-linked immunosorbent assay (ELISA) and neutralization assay] predicted the presence of a T-cell response well, although quantitatively neither antibody assays nor the T-cell assay correlated well with each other. An interesting finding from our study was that half of the dogs in shelters were not vaccinated (potentially posing a public veterinary health problem) and that antibody levels in dogs living in an environment with endemic CDV were lower than in vaccinated animals.

Molecular characterization of the VP1 gene of a Mexican isolate of infectious pancreatic necrosis virus
Magda Barrera-Mejía, José Simón-Martínez, Raúl Ulloa-Arvizu, Celene Salgado-Miranda, Edgardo Soriano-Vargas

Abstract — The presence of infectious pancreatic necrosis virus (IPNV) in salmonids predominantly produces a high mortality rate in first-feeding fry. Genomic analysis of the vp2 gene sequence is most commonly used to determine the genetic diversity of IPNV isolates. Recently, information obtained from the vp1 gene allowed for efficient analysis of the genetic diversity of IPNV. In this study, the vp1 gene from a Mexican IPNV isolate was characterized and compared with IPNV isolates from Europe, North America, and Asia. The results indicate that the Mexican isolate is most closely related genetically to the 2310 strain from Spain.

Serodiagnosis of porcine reproductive and respiratory syndrome virus infection with the use of glycoprotein 5 antigens
Hyoju Pyo, Jayoung Seo, Gukhyun Suh, Kwanghyun Kim, Jaeil Lee, Taejung Kim

Abstract — Glycoprotein 5 (GP5) of porcine reproductive and respiratory syndrome virus (PRRSV) has been studied extensively as a target for vaccine development. This study evaluated the serodiagnostic application of PRRSV GP5 by enzyme-linked immunosorbent assay (ELISA). Two immunodominant peptides (VR #1 and VR #2) and two neutralizing ectodomain-containing peptides (Ecto #1 and Ecto #2), as well as recombinant GP5 (rGP5) as a control, were prepared. Serum from unvaccinated pigs was screened for the antibodies that bind to these peptide and protein antigens. The results were compared with those from a commercially available diagnostic ELISA kit (HerdChek), which uses the nucleocapsid (N) protein as an antigen. Only VR #1+#2 showed a result statistically similar to that of N protein. Ecto #1 and Ecto #2 had a lower sensitivity than VR #1+#2 and rGP5. The peptides and rGP5 showed significant associations with the N protein (P < 0.05 or 0.01), which suggests that GP5 may also be a candidate serodiagnostic antigen. Since antibodies against GP5 persist much longer than those against the N protein, GP5 itself and some of its fragments are thought to be good targets for serodiagnosis. In addition, the presence of antibodies against the PRRSV structural antigens showed significant antigen-dependent differences.

Prion genotypes of scrapie-infected Canadian sheep 1998–2008
Noel P. Harrington, Katherine I. O’Rourke, Yuqin Feng, Jasmine Rendulich, Cathleen DiFruscio, Aru Balachandran

Abstract — This report describes the genetics of the prion protein gene (PRNP) at codons 136, 154, and 171 for sheep diagnosed with naturally acquired classical scrapie in Canada between 1998 and 2008. Genotyping analysis was performed on 249 sheep with confirmed classical scrapie infection representing 98 flocks from 6 provinces. A further case-control analysis of 3 of these flocks compared the genotypes between infected sheep (n = 72) and those of their healthy flockmates (n = 1990). The incidence of classical scrapie in the Canadian sheep population was highly associated with the ARQ haplotype (91.8%) and the ARQ/ARQ genotype (91.6%). In addition, the ARQ haplotype was found at significantly higher frequency in scrapie-infected sheep when compared with their healthy flockmates. Comparison with other published data suggests that the scrapie risk of PRNP genotypes differs between Canada and countries where the VRQ allele is associated with the highest susceptibility to infection.

Characteristics of the molecular diversity of the outer membrane protein A gene of Haemophilus parasuis
Cheng Tang, Bin Zhang, Hua Yue, Falong Yang, Guoqing Shao, Quan Hai, Xiaofei Chen, Dingqian Guo

Abstract — The molecular diversity of the gene encoding the outer membrane protein A (OmpA) of Haemophilus parasuis has been unclear. In this study, the structural characteristics, sequence types, and genetic diversity of ompA were investigated in 15 H. parasuis reference strains of different serovars and 20 field isolates. Three nucleotide lengths of the complete open reading frame (ORF) of ompA were found: 1098 base pairs (bp), 1104 bp, and 1110 bp. The OmpA contained 4 hypervariable domains, mainly encoding the 4 putative surface-exposed loops, which makes it a potential molecular marker for genotyping. Western blot analysis showed that the recombinant OmpAs of serovars 4 and 5 could cross-react with antiserum to all 15 serovars. Hence, although ompA of H. parasuis exhibited high variation among serovars, this variation did not seem to affect the strong antigenic characteristics of OmpA.

In situ detection of urease-positive Helicobacter pylori-like organisms on swine gastric mucosa
Kathleen A. McIntosh, Steven Krakowka, Susan S. Ringler, John A. Ellis

Abstract — The objective of this study was to improve the visual localization of urease activity of Helicobacter pylori-like organisms (HPLO) on swine gastric mucosa by in vitro optimization of the urea concentration and pH indicator of a urease test reagent. Five 21-day-old conventional pigs were infected orally with HPLO (3 pigs) or Brucella broth alone (2 pigs). At 17 d after infection the pigs were euthanized and their stomachs excised and tested for HPLO by a modified urease test formulation sprayed onto the gastric mucosa, as well as confirmatory culture and isolation of HPLO from urease-positive sites. This study showed improved detection of HPLO in porcine gastric mucosa with the use of a modified urease test formulation containing 5% urea and the pH indicator bromocresol purple compared with the use of a conventional formulation of 2% urea and phenol red. This test can readily be applied to achieve a presumptive diagnosis of HPLO in cases of gastritis or gastric esophageal ulceration in pigs.