Bacterial flora of liver abscesses in crossbred beef cattle and Holstein steers fed finishing diets with or without tylosin.

Holstein steers raised for beef production consistently have a higher prevalence and more severe form of liver abscesses than cattle of beef breeds. A study was conducted to compare bacterial flora of liver abscesses collected from multiple abattoirs from 4 groups of cattle, arranged in a 2 × 2 factorial design, consisting of crossbred cattle and Holstein steers, and each group fed a finishing diet supplemented with or without tylosin. A total of 383 liver abscess samples, consisting of 94 and 81 from crossbred cattle and 89 and 119 from Holstein steers fed finishing diets with or without tylosin, respectively, were subjected for anaerobic and aerobic bacterial isolations. The minimum inhibitory concentrations (MIC) of tylosin to the predominant bacterial species were determined. The likelihood chi-square test was performed to assess unadjusted differences in bacterial prevalence proportions between the 2 types of cattle (crossbred and Holstein steers) and feed type (tylosin or no tylosin). There was no interaction between cattle type and tylosin inclusion on the prevalence of any of the bacterial species isolated. Liver abscesses from Holstein steers yielded a higher total number of isolates compared to liver abscesses from crossbred cattle (1060 vs. 788). subsp. was isolated from all abscesses. The prevalence of subsp. was 19.1% and was not affected by the cattle type or tylosin. The prevalence of was higher ( < 0.01) in crossbred cattle (73.7%) compared to Holstein steers (29.8%). Also, the prevalence of was higher in abscesses from tylosin-fed (66.1%) cattle than no tylosin-fed cattle (35%). The overall prevalence of was 25.3% and was similar ( = 0.58) between cattle type, but the prevalence was lower ( < 0.01) in tylosin-fed (16.9%) compared to no tylosin-fed group (33%). Mean MIC of tylosin for and were similar across both cattle types and tylosin inclusion. Although bacterial flora of liver abscesses from Holstein steers appeared to be more diverse than that of crossbred cattle, there was no difference in the prevalence of the and and in fact, prevalence of was higher in crossbred than Holstein steers. Therefore, the difference in bacterial flora is not the likely reason for higher prevalence and severity of liver abscesses in Holstein steers than crossbred beef cattle.

Pharmacokinetics and pharmacodynamics of gamithromycin in pulmonary epithelial lining fluid in naturally occurring bovine respiratory disease in multisource commingled feedlot cattle.

The objectives of this study were to determine (i) whether an association exists between individual pharmacokinetic parameters and treatment outcome when feeder cattle were diagnosed with bovine respiratory disease (BRD) and treated with gamithromycin (Zactran(®) ) at the label dose and (ii) whether there was a stronger association between treatment outcome and gamithromycin concentration in plasma or in the pulmonary epithelial lining fluid (PELF) effect compartment. The study design was a prospective, blinded, randomized clinical trial utilizing three groups of 60 (362-592 lb) steers/bulls randomly allocated within origin to sham injection or gamithromycin mass medication. Cattle were evaluated daily for signs of BRD by a veterinarian blinded to treatment. Animals meeting the BRD case definition were enrolled and allocated to a sample collection scheme consisting of samples for bacterial isolation (bronchoalveolar lavage fluid and nasopharyngeal swabs) and gamithromycin concentration determination (PELF and plasma). Gamithromycin susceptibility of M. haemolytica (n = 287) and P. multocida (n = 257) were determined using broth microdilution with frozen panels containing gamithromycin at concentrations from 0.03 to 16 µg/mL. A two-compartment plasma pharmacokinetic model with an additional compartment for gamithromycin in PELF was developed using rich data sets from published and unpublished studies. The sparse data from our study were then fit to this model using nonlinear mixed effects modeling to estimate individual parameter values. The resulting parameter estimates were used to simulate full time-concentration profiles for each animal in this study. These profiles were analyzed using noncompartmental methods so that PK/PD indices (AUC24 /MIC, AUC∞ /MIC, CMAX /MIC) could be calculated for plasma and PELF (also T>MIC) for each individual. The calculated PK/PD indices were indicative that for both M. haemolytica and P. multocida a higher drug exposure in terms of concentration, and duration of exposure relative to the MIC of the target pathogen, was favorable to a successful case outcome. A significant association was found between treatment success and PELF AUC0-24 /MIC for P. multocida. The calves in this study demonstrated an increased clearance and volume of distribution in plasma as compared to the healthy calves in two previously published reports. Ultimately, the findings from this study indicate that higher PK/PD indices were predictive of positive treatment outcomes.

Comparison of Mannheimia haemolytica isolates from an outbreak of bovine respiratory disease.

The objective of this study was to determine the clonal relatedness of Mannheimia haemolytica isolates responsible for an outbreak of bovine respiratory disease in a commercial feedlot. The isolates were obtained from the lungs of 21 calves with fatal pneumonia that were part of a group of 206 total calves. All isolates were serotyped and analyzed by pulsed-field gel electrophoresis (PFGE) and for antibiotic sensitivity patterns. ELISA and immunoblotting assays were performed to compare serum antibody levels to M. haemolytica antigens in calves with fatal pneumonia to those calves that survived the outbreak. Isolates were categorized into 14 different PFGE groups based on 90% similarity. Two Group D isolates (1 and 6), and 3 Group H isolates (14, 15, and 16) were characterized as 100% similar. Antimicrobial susceptibility profiles defined 8 groups based on differences in patterns of resistance between isolates. The two 100% similar isolates from PFGE Group D were both in susceptibility Group 1. All but isolate 14 from PFGE Group H (3, 15, 16, and 19) were in susceptibility Group 4a. Serum antibody levels to M. haemolytica antigens in the dead calves were not different than the antibody levels in the 185 calves that survived the outbreak. Immunoblots of selected isolates from each of the PFGE groups demonstrated only minimal differences in antigenic profiles between strains when reacted with serum from calves that either died from or survived the outbreak. Based on the characteristics of these isolates, multiple strains of M. haemolytica were responsible for fatal pneumonia during this outbreak.

The pharmacokinetics and effects of meloxicam, gabapentin, and flunixin in postweaning dairy calves following dehorning with local anesthesia.

Approved analgesic compounds in cattle are not currently available in the United States due to the lack of validated pain assessment methods and marker residue depletion studies. In this study, we compared the pharmacokinetic parameters and effect of preemptive analgesics administered to calves subjected to dehorning with local anesthesia. Holstein steers were randomly assigned to receive one of the following treatments per os (PO) or intravenously (IV) (n = 8/group): meloxicam (1 mg/kg PO), gabapentin (15 mg/kg PO), meloxicam (1 mg/kg), and gabapentin (15 mg/kg) PO, flunixin (2.2 mg/kg IV), or a placebo. Plasma drug, haptoglobin, substance P (SP) concentrations, serum cortisol concentrations, ocular thermography, mechanical nociceptive threshold (MNT), and average daily gain (ADG) were evaluated. Data were analyzed using mixed-effects models and noncompartmental pharmacokinetic analysis. Meloxicam, gabapentin, and meloxicam with gabapentin at the present doses did not reduce cortisol concentrations. Analgesic-treated calves had significantly lower plasma SP concentrations and improved ADG compared with controls. Flunixin calves had reduced circulating cortisol compared with controls. Meloxicam-treated calves showed an increase in MNT at two horn bud sites compared with the other treatments. Analgesics improved ADG and reduced biomarkers of pain, but effects differed by compound and route of administration.

Attenuation of acute plasma cortisol response in calves following intravenous sodium salicylate administration prior to castration.

Pain associated with castration in cattle is an animal welfare concern in beef production. This study examined the effect of oral aspirin and intravenous (i.v.) sodium salicylate on acute plasma cortisol response following surgical castration. Twenty bulls, randomly assigned to the following groups, (i) uncastrated, untreated controls, (ii) castrated, untreated controls, (iii) 50 mg/kg sodium salicylate i.v. precastration and (iv) 50 mg/kg aspirin (acetylsalicylic acid) per os precastration, were blood sampled at 3, 10, 20, 30, 40, 50 min and 1, 1.5, 2, 4, 6, 8, 10 and 12 h postcastration. Samples were analyzed by competitive chemiluminescent immunoassay and fluorescence polarization immunoassay for cortisol and salicylate, respectively. Data were analyzed using noncompartmental analysis, a simple cosine model, anova and t-tests. Intravenous salicylate V(d(ss)) was 0.18 L/kg, Cl(B) was 3.36 mL/min/kg and t(1/2 lambda) was 0.63 h. Plasma salicylate concentrations above 25 microg/mL coincided with significant attenuation in peak cortisol concentrations (P = 0.029). Peak salicylate concentrations following oral aspirin administration was <10 microg/mL and failed to attenuate cortisol response. Once salicylate concentrations decreased below 5 microg/mL, cortisol response in the castrated groups was significantly higher than uncastrated controls (P = 0.018). These findings have implications for designing drug regimens to provide analgesia during routine animal husbandry procedures.

DNA extraction from low-biomass carbonate rock: an improved method with reduced contamination and the low-biomass contaminant database.

Caves represent a unique environment in which to study subsurface geomicrobial interactions and processes. One of the primary techniques used to study such geologic samples is molecular phylogenetic analysis, but this technique is hampered by low microbial biomass and calcium in the host rock, often leading to poor and irreproducible DNA extraction. We describe an improved protocol to recover extremely low amounts of DNA from calcium-rich geologic samples. This protocol relies on the use of the synthetic DNA molecule poly-dIdC, to act both as blocking agent and carrier molecule to increase the yield of DNA, and dialysis to remove calcium inhibitors of PCR amplification. Further, we demonstrate that many traditionally used laboratory substrates contain microbial DNA that can be amplified through the polymerase chain reaction (PCR) and contaminate molecular phylogenetic profiles. While the number of potential contaminants can be minimized, it cannot be eliminated from extraction techniques. We have therefore established the low-biomass contaminant (LBC) database, which contains the 16S rRNA gene sequences of species that have been identified as common laboratory contaminants. These identified contaminants provide a reference database to allow investigators to critically evaluate certain species identified within their phylogenetic profile when examining such low-biomass environments.

Rapid quantification of C3a and C5a using a combination of chromatographic and immunoassay procedures.

Monoclonal antibodies were isolated which reacted specifically with the complement cleavage products C3a, C3adR, C5a, and C5adR but not with the parent molecules C3 or C5. In both cases the mAbs showed a higher affinity towards the desArg forms. These mAbs were used as capture antibodies in immunoassays for C3a/C3adR and C5a/C5adR. The immunoassays are based on the ABICAP technology which ensures for a rapid measurement. Due to the large binding capacity and the very short diffusion pathways in the gel-matrix the binding equilibrium between capture antibodies and the antigen is reached whilst the sample is flowing through the column. Therefore this test represents an endpoint assay offering the possibility of using a single calibration curve for a large number of measurements. With the C3adR assay concentrations down to 16 ng/ml C3adR can be detected. The lower detection limit of the C5adR assay is 1 ng/ml C5adR. The tests for C3a/C3adR, and C5a/C5adR can be performed in 20 to 25 min and this rapid processing of plasma samples should permit the application of these parameters for diagnostic purposes and patient management.

Evaluation of the C-terminal C5a effector site with short synthetic C5a analog peptides.

Biological activities have been determined for a series of 18 peptides based on the C-terminal sequence of human or rat C5a. Lysosomal enzyme release was tested in two cell types, the promyelotic leukemia cell line U937 and polymorphonuclear leukocytes. In addition, an ATP-release assay with guinea pig platelets was performed. It was demonstrated that the C-terminal octapeptide 67-74 of human C5a represents the minimal sequence required to induce a measurable biological signal in all assays. Extending this peptide to a length of 21 amino acids produced at best only a slight enhancement of potency. Amino acid replacements with either tryptophanyl or phenylalanyl residues in positions between 65-69 either increased potency (at position 67), or abrogated potency (at position 66) in the two lysosomal enzyme assays. N-terminal acylation with the fluorenylmethoxy-carbonyl-aminohexanoyl group slightly enhanced C5a potency. In desensitization experiments with guinea pig platelets all peptides with a C5a activity were able to desensitize not only the C5a but also the C3a responses.