Experimental infectious respiratory disease in groups of calves: lobar distribution, variance, and sample-size requirements for vaccine evaluation.

The distribution and variance of respiratory disease produced with aerosols of bovine herpesvirus 1 (BHV-1) and Mannheimia haemolytica in control (183 calves in 44 experiments) and vaccinated calves were studied in experiments conducted at the Animal Diseases Research Institute, Lethbridge, Alberta, from 1975 to 1989. All calves had been born and raised at this institute and exposed similarly for 5 min by means of a face mask to viral and bacterial aerosols produced by a Collison atomizer (particles < 3 microm in diameter). We summarized the macroscopic pathological responses of pneumonia (main end point), tonsillitis, tracheitis, and other microbiologic and experimental variables. We also summarized the lobar distribution of pneumonia in 202 control and 192 vaccinated calves with this disease model and in calves similarly exposed to parainfluenza 3 virus/M. haemolytica or BHV-1/Pasteurella multocida. Pneumonia in control calves began in ventral tissues of all lobes, with lobar preferences, and progressed dorsally, the dorsal parts of both large caudal lobes being least affected. A high variance of pneumonia was evident within and among experiments. From the magnitude of variance observed in the control groups, the number of calves per group required in vaccine-challenge studies using this BHV-1/M. haemolytica disease model was estimated. Such estimates are required for any disease model used in vaccine-challenge studies.

Aerobiology of a high-line speed cattle abattoir.

The operation of the high-line speed cattle abattoir studied follows a plant-created hazard analysis and critical control point (HACCP) plan that is recognized by the Canadian Food Inspection Agency. Measurement of bioaerosols is not a part of this plan. In this study CFUs in air of selected abattoir processes were enumerated after impinging air onto tryptic soy agar plates with a slit air sampler for 10 to 20 min. The total viable count (TVC) per liter of air was calculated for each sample following incubation at 30 degrees C for 24 h. Monthly samples were collected on the hide removal floor and the carcass dressing floor from March 1998 to April 1999. Mud tag, dirt, and wetness of incoming hides were scored subjectively on the hide removal floor. The other processes were sampled in 3 separate months. The TVC at two locations on the hide removal floor (center of hide removal floor [CHF] and top of hide puller [THP]) had a strong association to each other (r = 0.84; P < 0.001). The mean TVC at the CHF and THP was 10.0 and 11.5, respectively, and the TVC for individual samples ranged from 2 to 42 at these locations. The TVC means for all the other processes ranged from 0.01 to 0.7. Tag and TVC on the hide removal floor had a different seasonal distribution with TVC being highest in the warm months (April to October 1998) and lowest for November to April 1999. No significant relations between TVC and the dirt and wetness variables were evident for the CHF and THP locations on the hide removal floor. It was concluded that the control of aerosols in the hide removal floor should be treated as a critical control point in the HACCP plan.

Microbiological testing of raw, boxed beef in the context of hazard analysis critical control point at a high-line-speed abattoir.

The efficacy of cold storage of raw, bagged, boxed beef was assessed microbiologically at a high-line-speed abattoir (270 carcasses per h). At the time of this study, plant management was in the process of creating a hazard analysis critical control point plan for all processes. Aerobic bacteria, coliforms, and type 1 Escherichia coli were enumerated (5 by 5-cm excision samples, hydrophobic grid membrane filter technology) before and after cold storage of this final product produced at six fabrication tables. In addition, the temperature-function integration technique (TFIT) was used to calculate the potential number of generations of E. coli during the first 24 or 48 h of storage of the boxed beef. Based on the temperature histories (total of 60 boxes, resulting from 12 product cuts, five boxes from each of two fabrication tables on each of 6 sampling days, and six types of fabrication tables), TFIT did not predict any growth of E. coli (with or without lag) for the test period. This was verified by E. coli mean log10 values of 0.65 to 0.42 cm2 (P > 0.05) determined by culture before and after the cooling process, respectively. Counts of aerobic bacteria and coliforms were significantly reduced (P < 0.001 and P < 0.05, respectively) during the initial period of the cooling process. There were significant microbiological differences (P < 0.05) between table-cut units.

Verification of the hygienic adequacy of beef carcass cooling processes by microbiological culture and the temperature-function integration technique.

To enhance food safety and keeping quality, beef carcasses are cooled immediately after leaving the slaughter floor. Within hazard analysis and critical control point (HACCP) systems, this cooling process needs to be monitored by the industry and verified by regulatory agencies. This study assessed the usefulness of the temperature-function integration technique (TFIT) for the verification of the hygienic adequacy of two cooling processes for beef carcasses at one abattoir. The cooling process passes carcasses through a spray cooler for at least 17 h and a holding cooler for at least 7 h. The TFIT is faster and cheaper than culture methods. For spray cooler 1, the Escherichia coli generations predicted by TFIT for carcass surfaces (pelvic and shank sites) were compared to estimated E. coli counts from 120 surface excision samples (rump, brisket, and sacrum; 5 by 5 by 0.2 cm) before and after cooling. Counts of aerobic bacteria, coliforms, and E. coli were decreased after spray cooler 1 (P < or = 0.001). The number of E. coli generations (with lag) at the pelvic site calculated by TFIT averaged 0.85 +/- 0.19 and 0.15 +/- 0.04 after emerging from spray coolers 1 and 2, respectively. The TFIT (with lag) was considered convenient and appropriate for the inspection service to verify HACCP systems for carcass cooling processes.

The incidence of caseous lymphadenitis in Alberta sheep and assessment of impact by vaccination with commercial and experimental vaccines.

In Alberta, caseous lymphadenitis (CLA) is one of the leading causes of lamb and mutton carcass condemnation. In this study, serologic results confirmed a high (50-94%) incidence of exposure to Corynebacterium pseudotuberculosis, the causative agent of CLA, in mature, unvaccinated sheep in southern Alberta. To assess the efficacy and impact of vaccination with 2 commercial (Glanvac-6 and Case-Vac) and 1 experimental (WC+ MDP-GDP) CLA vaccines, a series of 3 field trials in 3249 ewes and lambs was conducted in affected flocks from 1992-1996. Efficacy was assessed from the serological response to vaccination, prevalence and size of injection site reactions by treatment, and the incidence of CLA abscesses. Overall, agglutinating antibody titres to C. pseudotuberculosis in lambs vaccinated with WC+MDP-GDP and Case-Vac remained significantly elevated above nonvaccinated control lambs for the 12 mo period after the initial vaccination. Lambs vaccinated with the WC/MDP-GDP maintained higher titres (P < 0.06) than those vaccinated with Case-Vac for the period from 6 to 12 mo after vaccination. Agglutinating antibody titres for lambs vaccinated with Glanvac did not differ from those of controls at any point during the 12 mo period after vaccination. The number of injection site reactions was elevated in lambs vaccinated with Glanvac as compared to those vaccinated with WC+MDP-GDP but the size of injection site reactions did not significantly differ. Sheep vaccinated with WC+ MDP-GDP also had a reduced incidence of putative CLA abscesses, although confirmation of the presence of C. pseudotuberculosis was only successful in a small number of instances.

Genetic diversity of potato determined by random amplified polymorphic DNA analysis.

The RAPD procedure was used to establish genetic diversity of 28 potato genotypes including siblings and genotypes with no immediate relationship. In addition amplified DNA from three parents and Solanum chacoense were compared with that from six progeny to determine the genetic relationships. Amplification of genomic DNA from the 28 genotypes using PCR and 12 decamer primers yielded 158 amplified DNA fragments, ranging in size from 490 to 3200 bp. A total of 128 unique RAPD fragments were observed among the 28 potato genotypes. Similarity measures and principal coordinate analysis generally reflected the expected trends in relationships of the full and half-sib potato genotypes. However there were important exceptions to this general trend and it appears that related varieties can be as genetically different as varieties with no immediate relationship. The data suggest that RAPD analysis used in conjunction with pedigree information can provide a superior measure of genetic divergence than analysis based solely on pedigree information.

Microbiologic evaluation of carcasses before and after washing in a beef slaughter plant.

The effect of washing on the bacterial contamination of beef carcasses in a modern abattoir was evaluated. Twenty-six carcasses were evaluated at the end of the slaughter process before and after washing, and 13 other carcasses were evaluated only after being washed. An excision sample (5 x 5 x 0.5 cm) was collected from 10 sites on each carcass immediately before washing and at an adjacent site immediately after washing. Aerobic bacterial colonies were enumerated, using hydrophobic grid membrane filter technology. After washing, the log10 of the most probable number of growth units/cm2 decreased (P < 0.01) at the lateral rump site, increased (P < 0.01) at the thorax and neck sites, but was unchanged at the other 7 sites, compared with before washing. The sample size required to estimate, within 0.5 log10 units, the mean log10 most probable number of growth units/cm2 at a site for use in future group-carcass evaluations was determined and compared with a previously used sample size definition. It was concluded that the washing process described did not result in a major change in the bacterial contamination of carcasses.

Bacteriological evaluation of groups of beef carcasses before the wash at six Alberta abattoirs.

A method has been developed for the bacteriological evaluation of groups of beef carcasses which can be used to measure the degree of control over hygiene during hide removal and carcass dressing in abattoirs. This method, which enumerates aerobic mesophilic bacteria automatically using a hydrophobic grid membrane filter, was applied at six abattoirs. Two hundred excision samples (5 x 5 x 0.5 cm) were taken at 10 sites on the external surface of a group of 20 carcasses (five carcasses were sampled on each of four consecutive daily visits) for group-carcass evaluation at each abattoir. For each abattoir, the mean log10 Most Probable Number of Growth Units (MPNGU) and between-carcass variance component were obtained for each site and the average over sites. Using the average within-abattoir variance of this study and previously published studies involving 76 additional carcasses (Jericho et al. 1993), it was determined that 20 carcasses are more than adequate to estimate the mean log10 MPNGU per cm2 within 0.5 units at a site. The distribution of the log10 MPNGU per cm2 over the 10 sites was compared for the abattoirs, and sites were found to cluster into 2-4 homogenous groups. The means over sites of log10 MPNGU per cm2 for the abattoirs ranged from 1.52 to 2.64 and were unrelated to line speed.

Microbiological evaluation of groups of beef carcasses: heifers and steers.

Numbers of mesophilic bacteria were estimated on carcasses of 25 heifers and 25 steers of beef breeds in a modern, high-line-speed abattoir. One side of each carcass from each sex was sampled at the end of the kill-floor, before the carcass wash, on each of 25 visits. Two adjacent excision samples (5 x 5 x 0.5 cm) were taken from each of ten sites and processed for automatic enumeration of aerobic bacteria on hydrophobic grid membrane filters. The effects of sex and carcass weight on bacterial counts were examined. Groups of carcasses were examined to determine the sample size required for future assessments of kill-floor hygiene. The log10 of the most probable number of growth units (MPNGU)/cm2 did not differ significantly between heifers and steers (average over the ten sites of 2.2) and there was no effect of carcass weight on bacterial counts for nine of the ten sites. There were, however, highly significant (p < 0.001) differences in the counts between sites and the counts from the ten sites clustered into five homogenous groups. The between-carcass component of variation at a site was generally larger than the within-carcass component. We conclude that, to estimate the mean log10 MPNGU/cm2 at a site to within +/- 0.5 units, future group-carcass evaluations require about 200 samples from 10 (two adjacent samples/site) or 20 carcasses (one sample/site).

A comparative evaluation of two sensitive serum neutralization tests for bovine herpesvirus-1 antibodies.

Two sensitive serum neutralization (SN) tests for the detection of antibodies to bovine herpesvirus-1 (BHV-1) in bovine sera were evaluated. Both SN tests used a 24 h incubation of test sera with 100 CCID50 of BHV-1 before the addition of susceptible cells. The tests differed in the presence (C test) or absence (D test) of complement and were compared with a standard 1 h incubation SN test and the enzyme-linked immunosorbent assay (ELISA). Although the mean titer of the C test was twofold higher than the mean titer of the D test for 310 sera, the number of samples which were negative was not significantly different between tests. For 100 sera from herds with known reactors, which were negative in a 1 h incubation SN test, 32% tested positive in the C and D tests. Other investigations, including Western immunoblotting and radioimmune precipitation, suggest that the 24 h incubation tests produce some false positive results. In contrast, the 1 h incubation SN test and, to a much lesser extent, the ELISA appear to produce some false negative results. The C test was more sensitive than the D test for detecting an early immune response after experimental infection.

A short incubation serum neutralization test for bovine viral diarrhea virus.

A three day serum neutralization (SN) test for the detection of antibodies to bovine viral diarrhea virus (BVDV), which is an improvement on the existing five day test, is described. The improved test results in a more rapid viral cytopathic effect and utilizes Madin Darby kidney (MDBK) cells, and horse serum as a medium supplement. A comparison of tests utilizing the NADL and the Singer strains of BVDV and the use of either secondary bovine kidney cells with calf serum (BKCS) or continuous MDBK cells with horse serum (MDHS) was performed. Analysis of the SN results of 685 serum samples from 445 Quebec and Ontario cattle showed that there was no difference, as expected, in the means of the SN antibody titers when the NADL strain was used in either the BKCS or MDHS system but SN antibody titers were elevated (p less than 0.01) when the Singer strain was used in the MDHS system. The SN test with the Singer strain also yielded significantly higher titers for sera from 200 Alberta cattle.

Protective effect of inactivated bovine herpesvirus-1 in calves experimentally infected with bovine herpesvirus-1 and Pasteurella haemolytica.

The protective effect of an inactivated whole-virion bovine herpesvirus-1 (BHV-1) immunising inoculum, without adjuvant, against viral-bacterial respiratory disease was studied in three experimental treatment groups of five calves each. One group was boosted 14 days after the first vaccination and at this time the second group received their initial inoculation. Seven days later, calves were challenged with BHV-1 in aerosol and four days after this challenge all calves were exposed to Pasteurella haemolytica A1 in aerosol. Among the three groups, differences in rectal temperature responses four days after viral challenge (P less than 0.01) did not relate to protection. However the main response variable, viral-bacterial pneumonia, was reduced in boosted calves (P less than 0.05).

Protective effect of inactivated Pasteurella haemolytica bacterin challenged in bovine herpesvirus-1 experimentally infected calves.

Formalin-inactivated and sonicated Pasteurella haemolytica bacterins were prepared from 1 h cultures of bacterial pellets in RPMI-1640 medium containing 7% fetal calf serum. The bacterial pellets were obtained from logarithmic phase growth of the organism by centrifugation. The protective effect of the vaccine was evaluated in 43 specific-pathogen-free Hereford crossbred calves and yearlings in three experiments. Cattle were either single vaccinated or boosted via three routes; intratracheally (i.t.), intranasally (i.n.) or intramuscularly (i.m.), using low or high doses. The two low-dose groups were also given supernatant by the same routes and volume as the bacterin. Cattle were challenged by P. haemolytica in aerosol at 24 or 39 days after last vaccination. To enhance the susceptibility of the cattle to this challenge, the cattle were exposed to bovine herpesvirus-1 aerosol 4 days before the bacterial challenge. The extent of pneumonia was significantly less in three groups of cattle (i.n.-i.n.,i.m.-i.n.,i.m.-i.m.) boosted with high dose of the bacterin than in the controls. Protection was observed when challenge isolates were heterologous or homologous to the isolates used to prepare the bacterins. It was also observed that the level of complement fixing antibody or anticytotoxin activity to P. haemolytica did not correlate with the degree of protection.

Electroantennographic responses differentiate sibling species of dingy cutworm complex,Feltia jaculifera (GN.) (Lepidoptera: Noctuidae).

The dingy cutworm is one of the more common and widely distributed of North American noctuids. Determination of the antennal responses of field-collected male moths to a standard test blend of pheromone components shows that two pheromonal phenotypes of the dingy cutworm occur throughout the prairie region of western Canada and at least two others occur in eastern Canada. The pheromone types are morphologically indistinguishable and, so far as is known, have similar life histories and biology. The two prairie types are broadly sympatric with partially overlapping seasonal flight periods. Although hybrids can be easily produced in the laboratory, there is no evidence of hybridization in the field. The pheromone system of the dingy cutworm is apparently rather plastic, resulting in a mosaic of pheromone types (sibling species) that appear to be reproductively isolated but have not differentiated morphologically.

Efficacy of methods used to test fertility of beef bulls used for multiple-sire breeding under range conditions.

A study was conducted during the 1982, 1983 and 1984 breeding seasons with 277 crossbred bulls, 1 to 3 yr of age, that were evaluated for physical soundness, testicular development, seminal quality, and both sexual and social behavior immediately before exposure to crossbred cow herds ranging in size from 89 to 329 cows. Crossbred cow herds were exposed to 4 to 24 bulls per breeding group (mean of 14) at a mean female: male ratio of 21.2 +/- .6:1 under extensive range conditions for 31 to 62 d (mean 46.6 d). All resulting calves were blood-typed to determine the number of calves sired by each bull as an estimate of his fertility. The mean number of calves sired by 1- (n = 116), 2- (n = 126) and 3-yr-old (n = 35) bulls was 4.7 +/- .1, 8.2 +/- .1 and 10.5 +/- .1, respectively. A regression model for predicting bull fertility under multiple-sire, range breeding conditions was selected that accounted for 29% of the total variance in fertility. Similar models accounted for a greater proportion of variance in fertility of 1-yr-old (37%) than of 2-yr-old bulls (22%). Due to the large amount of unexplained variation, the model could not predict individual bull fertility precisely. However, this study demonstrated that selection of herd sires with large scrotal circumference, low backfat thickness, low levels of primary sperm defects, and a low number of mounts in combination with a moderate number of services during libido testing would be expected to improve fertility of beef bulls used under extensive range conditions.

ELISA for bovine herpesvirus 1 (BHV1) antibody with HIRT supernatant.

Detailed studies were made of the ability of HIRT supernatant (HS) from bovine herpesvirus 1 (BHV1) infected cultures to sensitize plates for enzyme-linked immunosorbent assay (ELISA). The ultracentrifuged pellet of HS had less sensitizing activity than the supernatant, but the antigen was removed completely by 0.22 micron filters and to some extent by 0.45 micron filters; it was minimally affected by sonication; it was destroyed by the action of Pronase but not by DNAase I when a kallikrein inactivator was added and the mixture incubated; incubation with DNAase II had no effect. Thus the presence of DNA was not required for the sensitizing activity of HS and the antigens recognized by antibodies in HS in ELISA were directed to its protein component. Strong reactions were given in immunoblotting of HS from BHV1 infected tissue cultures with anti-BHV1 glycoprotein monoclonal IgG, but HS from uninfected tissue cultures did not react with the same monoclonal IgG.

Scrotal circumference of two-year-old bulls of several beef breeds.

Scrotal circumference (SC) was measured on 7,918 2-yr-old Angus, Charolais, horned and polled Herefords, Limousin, Shorthorn, and Simmental bulls presented to culling committees at six show/sales between 1977 and 1983. Only SC data from bulls within the age range of 24 +/- 4 mo were used. Scrotal circumference data were corrected across breeds for the effects of location-year and sire and were adjusted to a common bull age of 730 d. The adjusted mean SC (+/- SE) for 2-yr-old beef bulls was Simmental, 38.8 +/- 0.10 cm (n = 540); Aberdeen Angus, 37.2 +/- 0.09 cm (n = 629); Charolais, 36.3 +/- 0.09 cm (n = 499); horned Hereford, 36.1 +/- 0.03 cm (n = 3,769); polled Hereford, 35.6 +/- 0.04 cm (n = 2,170); Shorthorn, 34.9 +/- 0.11 cm (n = 231); and Limousin, 32.2 +/- 0.18 cm (n = 80). The authors' recommendations of minimum acceptable SC for 2-yr-old beef bulls are Simmental, 36.0 cm; Angus and Charolais, 35.0 cm; horned and polled Herefords and Shorthorn, 34.0 cm; and Limousin, 33.0 cm.

Testicular development, daily sperm production and epididymal sperm reserves in 15-mo-old Angus and Hereford bulls: effects of bull strain plus dietary energy.

Bull calves (n = 143) were obtained from two strains of Angus and two strains of Hereford cattle for which replacements were selected on the basis of superior feedlot growth performance on either high- or medium-energy diets. From weaning to slaughter at 15 mo of age, bulls were fed either the high-energy (80% grain + 20% forage) or medium-energy diet (100% forage) corresponding to their strain. Bulls in high-energy diet groups had a greater (P less than .05) scrotal circumference at 12 mo, but not 15 mo of age, than bulls in medium-energy diet groups. Compared with Hereford bulls, Angus had greater (P less than .01) scrotal circumference (36.1 vs 33.9 cm) and greater (P less than .05) paired testes weight (570 vs 464 g) at 15 mo of age. Daily sperm production per gram testicular parenchyma (DSP/g) was affected by strain-diet (P less than .01) but not by breed. Bulls in medium-energy diet groups had 12% greater DSP/g than did high-energy diet bulls (17.4 X 10(6) vs 15.5 X 10(6)). Daily sperm production (DSP) was 9% and 30% greater (P less than .01) for medium-energy diet bulls in 1980 (8.2 X 10(9) vs 7.5 X 10(9)) and 1981 (8.0 X 10(9) vs 6.2 X 10(9)), respectively, compared with high-energy diet bulls. The effect (P less than .01) of breed on DSP was attributed to breed differences in paired testes weight.(ABSTRACT TRUNCATED AT 250 WORDS)

Serum complement-fixing antibody to Pasteurella haemolytica A1 and its effect on experimentally induced pneumonic pasteurellosis in calves.

Ninety-three calves comprising 16 experimental groups were exposed to viral (bovine herpesvirus-1 or parainfluenza-3 virus) and Pasteurella haemolytica aerosols. Serum samples from these calves were tested before and after exposure for antibodies to P haemolytica by a modified direct complement-fixation test. At slaughter of the calves, the extent of pneumonia produced was estimated for each calf and compared with the results of the modified direct complement-fixation tests. The extent of pneumonia was not related (P greater than 0.05) to the amount of anti-P haemolytica antibody produced by either naturally occurring or experimentally induced infection.

Testicular development, epididymal sperm reserves and seminal quality in two-year-old Hereford and Angus bulls: effects of two levels of dietary energy.

The effect of high (HED) and medium energy diets (MED), fed to Hereford (H) and Angus (A) bulls from 6 through 24 mo of age, on scrotal circumference (SC), paired testes weight (PTW), epididymal sperm reserves (ESR) and seminal traits were examined. Over 3 yr, 120 bulls were involved. Angus exceeded H for both SC and PTW. Hereford bulls in yr 2 had smaller SC than in yr 1 or 3 but the response for A was consistent. Year affected PTW. In yr 2 Hereford bulls fed HED had 75% fewer ESR than MED-H bulls (9.3 vs 37.2 X 10(9]. Comparably treated A bulls had similar ESR numbers (29.2 vs 33.4 X 10(9]. In yr 3, epididymal sperm reserves of HED-H were depressed by 35% compared with MED-H (23.1 vs 35.7 X 10(9], whereas HED-A had 14% fewer ESR than did MED-A bulls (28.6 vs 33.1 X 10(9]. It was not obvious why H bulls were more susceptible to the effects of HED. Seminal quality of HED bulls was inferior to that of MED bulls, particularly with respect to progressive motility and the incidence of sperm in which a crater defect of the head was present at 2 yr of age. In yr 2 all seminal traits were severely depressed in 2-yr-old HED-H. Feeding HED to young H and A bulls reduced their reproductive potential.