Surveys of microbial populations in environmental niches of interest often utilize sequence variation in the gene encoding the ribosomal small subunit (the 16S rRNA gene). Generally, these surveys target the 16S genes using semi-degenerate primers to amplify portions of a subset of bacterial species, sequence the amplicons in bulk, and assign to putative taxonomic categories by comparison to databases purporting to connect specific sequences in the main variable regions of the gene to specific organisms. Due to sequence length constraints of the most popular bulk sequencing platforms, the primers selected amplify one to three of the nine variable regions, and taxonomic assignment is based on relatively short stretches of sequence (150–500 bases). We demonstrate that taxonomic assignment is improved through reduced unassigned reads by including a survey of near-full-length sequences specific to the target environment, using a niche of interest represented by the upper respiratory tract (URT) of cattle. We created a custom Bovine URT database from these longer sequences for assignment of shorter, less expensive reads in comparisons of the upper respiratory tract among individual animals. This process improves the ability to detect changes in the microbial populations of a given environment, and the accuracy of defining the content of that environment at increasingly higher taxonomic resolution.
The virulence and pathogenicity of bacterial pathogens are related to their adaptability to changing environments. One process enabling adaptation is based on minor changes in genome sequence, as small as a few base pairs, within segments of genome called simple sequence repeats (SSRs) that consist of multiple copies of a short sequence (from one to several nucleotides), repeated in series. SSRs are found in eukaryotes as well as prokaryotes, and length variation in them occurs at frequencies up to a million-fold higher than bacterial point mutations through the process of slipped strand mispairing (SSM) by DNA polymerase during replication. The characterization of SSR length by standard sequencing methods is complicated by the appearance of length variation introduced during the sequencing process that obscures the lower abundance repeat number variants in a population. Here we report a computational approach to correct for sequencing process-induced artifacts, validated for tetranucleotide repeats by use of synthetic constructs of fixed, known length. We apply this method to a laboratory culture of Histophilus somni, prepared from a single colony, and demonstrate that the culture consists of populations of distinct sequence phase and length variants at individual tetranucleotide SSR loci.
Additional file 3. Interpretive criteria for selected antimicrobial drugs. Table containing the following information related to MIC testing: antimicrobial, concentration(s) tested and threshold criteria for susceptible, intermediate or resistant.
Studies were conducted to examine the ability of lung auscultation to predict bovine respiratory disease complex (BRDC) case outcome in feeder cattle. In the first study Lung sounds were obtained and lung auscultation scores (LAS) were assigned by trained personnel. In the first study antemortem physical examination and LAS were correlated with lung pathology and lesion scores when the cattle at harvest. There was a correlation (R 2 =0.89; P <0.0001) between ante-mortem LAS and post-mortem lung lesion score. Next, a field study was conducted to evaluate LAS and case outcome in feeder cattle treated for BRDC(n = 4,341 head). Lung auscultation score (P <0.01) and rectal temperature (P <0.01) were predictive of cattle with increased risk for BRDC retreatment and death loss rates. The model-adjusted probability of a calf with a lung auscultation score of 2 requiring retreatment was 13% whereas the re-treatment probability for cattle assigned a LAS of 9 was 63%. A rise in rectal temperature from 100 to 108 oF correlated with a 266% increase in likelihood to be retreated for BRDC. The risk of death from BRDC increased nearly 2,200%, from 1.7% to 39%, as lung auscultation score severity proceeded from a score of 2 to 9. The risk of death from BRDC increased 196% as rectal temperature proceeded from 100 to 108 oF. These studies demonstrate that lung auscultation score and rectal temperature could be used as tools to better predict BRDC case outcomes in feeder cattle and facilitate targeted management decisions. Introduction Bovine respiratory disease complex (BRDC) is the most common, and costly, disease in feedyard cattle. Significant amounts of money are spent on preventing and treating BRDC in the feedyard in just vaccines and antimicrobials alone. Loneragan estimated initial treatment costs alone in the calendar year of 1999 to be 45.7 million dollars 7 . Total economic losses were recently estimated to be as high as $692 million dollars in a survey from 2005 conducted by the National Agricultural Statistics Service, a division of the United States Department of Agriculture 12 . Death loss associated with BRDC in feeder cattle has been well documented. Vogel and Parrott published a feedyard mortality survey detailing data from 59 feedyards (38,593,575 head of cattle) in seven Midwestern states from January 1990 – May 1993. Deaths due to BRDC were 44.1% of total deaths in these feedyards. Digestive disorders attributed to 25.9%, and “other” causes accounted for 28.6% of all deaths over the three and a half year period 12 . Another estimate is provided by the United States Department of Agriculture (USDA) National Animal Health Monitoring System (NAHMS), in which data were collected from 21,753,082 head of cattle that were placed on feed from 1994 to 1999 (121 feedyards; 12 states) 8 . An analysis of these data found that the mortality tended to increase (P = 0.09) from 10.3 deaths per 1,000 head of cattle in 1994 to 14.2 deaths per 1,000 head of cattle in 1999 for all etiologies. Respiratory mortality rates increased from 5.4 deaths per 1000 head in 1994 to 8.7 per 1000 head in 1999 8 . Aside from obvious economic losses from death loss of cattle, there are performance issues attributed to this disease. Gardner et al. found that steers without lung lesions attributed to BRDC at slaughter had improved average daily gain (ADG) relative to cattle that had BRDC lung lesions (3.48 vs. 3.09 lb/d, P <0.01) 5 . Similarly, Bryant et al. showed that BRDC lung lesions present at slaughter was associated with a decrease in ADG (0.057 lb/d) in single source calves and up to a 0.65 lb/d decrease in ADG in commingled calves compared to cattle in their respective groups that did not have lung lesions at slaughter 2 . More recently researchers in South Africa found that the negative effect on ADG in feeder cattle due to the presence of lung lesions at slaughter was 0.051 lb/d decrease compared to ADG in cattle without lung lesions at slaughter 11 . Additionally, the researchers reported that the presence of lesions at slaughter was associated with an additional 5.5 d to finish cattle. In the U.S. feedyard system, disease detection starts with the pen riders. Pen riders are in the pens looking at cattle at least once per day for signs of any illnesses. Clinical signs that are used to identify cattle possibly afflicted with BRDC include respiratory rate, respiratory character, rumen fill, observed anorexia, nasal discharge, ocular discharge, and depression 1 . Chute-side feedyard diagnostics consist of examining clinical signs and taking rectal temperatures. Decisions on therapeutic regimens are often outlined in a treatment protocol and are generally based on a rectal temperature greater than some arbitrary cutoff, for example, ≥103.5 oF 3 . Rectal temperatures can vary from the influence of such factors as environmental temperature, relative humidity, heat of fermentation, exercise, excitement, and anxiety. These changes in rectal temperature are the result of physiologic change rather than a pathologic one 4 . Vogel et al found that for each unit (F) increase in maximum ambient temperature, rectal temperature in all cattle pulled increased 0.07 oF (P <0.01) 13 . In the same study, they found that the rectal temperatures in cattle pulled and treated for BRDC had no association with the risk of re-treatment or mortality 13 . Using elevated rectal temperature as the sole diagnostic tool beyond clinical signs is essentially treating on the basis of depression with undifferentiated fever. Treatments based solely on this may lead to unnecessary antimicrobial use 1 . Additional diagnostic tools, such as a stethoscope, may aid in decision making; however, the literature is largely devoid of reports associating the use of a stethoscope as a diagnostic aid to better identify and manage BRDC or predict BRDC-case outcome in feeder cattle. Therefore, the objectives of this paper are to validate a thoracic auscultation scoring system by correlating ante-mortem lung sounds with post-mortem lung lesions and then evaluate chute-side diagnostic tools to predict case outcome in the feeder cattle treated for BRDC
Moraxella bovoculi is a recently described bacterium that is associated with infectious bovine keratoconjunctivitis (IBK) or "pinkeye" in cattle. In this study, closed circularized genomes were generated for seven M. bovoculi isolates: three that originated from the eyes of clinical IBK bovine cases and four from the deep nasopharynx of asymptomatic cattle. Isolates that originated from the eyes of IBK cases profoundly differed from those that originated from the nasopharynx of asymptomatic cattle in genome structure, gene content and polymorphism diversity and consequently placed into two distinct phylogenetic groups. These results suggest that there are genetically distinct strains of M. bovoculi that may not associate with IBK.
Table of nucleotide polymorphism allele locations and genotypes used to initially type all 1133 M. haemolytica isolates (first table). Due to the size of the dataset this is the first of two tables that contain the genotypes (Additional files 2 and 3). The table contains the locations and allele scores for 11,306 of 16,447 nucleotide polymorphisms that were used to generate the tree of Fig. 1. Each polymorphism was unambiguously scored in 95% or more of the isolates. (ZIP 4344 kb)
OBJECTIVE To compare predictive values, extent of agreement, and gamithromycin susceptibility between bacterial culture results of nasopharyngeal swab (NPS) and bronchoalveolar lavage fluid (BALF) samples obtained from calves with bovine respiratory disease (BRD). ANIMALS 28 beef calves with clinical BRD. PROCEDURES Pooled bilateral NPS samples and BALF samples were obtained for bacterial culture from calves immediately before and at various times during the 5 days after gamithromycin (6 mg/kg, SC, once) administration. For each culture-positive sample, up to 12 Mannheimia haemolytica, 6 Pasteurella multocida, and 6 Histophilus somni colonies underwent gamithromycin susceptibility testing. Whole-genome sequencing was performed on all M haemolytica isolates. For paired NPS and BALF samples collected 5 days after gamithromycin administration, the positive and negative predictive values for culture results of NPS samples relative to those of BALF samples and the extent of agreement between the sampling methods were determined. RESULTS Positive and negative predictive values of NPS samples were 67% and 100% for M haemolytica, 75% and 100% for P multocida, and 100% and 96% for H somni. Extent of agreement between results for NPS and BALF samples was substantial for M haemolytica (κ, 0.71) and H somni (κ, 0.78) and almost perfect for P multocida (κ, 0.81). Gamithromycin susceptibility varied within the same sample and between paired NPS and BALF samples. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated culture results of NPS and BALF samples from calves with BRD should be interpreted cautiously considering disease prevalence within the population, sample collection relative to antimicrobial administration, and limitations of diagnostic testing methods.
Abstract The objective of this study was to determine the effects of flunixin meglumine or meloxicam on behavioral response and performance characteristics associated with surgical castration in crossbred bulls. Intact male Bos taurus calves (n = 252; averaging 176 kg) were randomly allocated into one of three treatment groups within pen: control (CON), flunixin meglumine (FLU; 2.2 mg/kg intravenous injection), or meloxicam (MEL; 2.0 mg/kg per os). The individual animal was the experimental unit. Calves were individually weighed on days 0 and 14 of the trial to evaluate performance outcomes. On study day 0, treatments were administered, according to their random allocation, immediately prior to surgical castration using the Henderson tool method. Visual analog scale (VAS) assessment and categorical attitude score (CAS) were collected on days −1, 0 (6 h post-castration), 1, 2, 3, and 4 in the study. The VAS was assigned using a 100 mm horizontal line with “normal” labeled at one end of the line and “moribund” at the other end of the horizontal line. The masked observer assigned a mark on the horizontal line based upon the observed severity of pain exhibited by that individual animal. The CAS was assigned by the same observer using five different categories with a score of 0 being “normal”. Average daily gain tended (P = 0.09) to be associated with the treatment group, and MEL had a greater (P = 0.04) average daily gain through day 14 compared with CON. A significant (P < 0.01) treatment by day interaction was indicated for VAS score, and MEL had lower VAS scores on days 0, 1, 2, and 3 post-castration compared with CON; FLU had lower VAS scores on days 0 and 1 compared with CON. A significant treatment by day interaction was not present (P = 0.25) for CAS. The FLU had lesser percent CAS ≥1 (17.5%; P = 0.05) compared with CON (29.4%); MEL has lesser percent CAS ≥1 observations (14.9%; P = 0.01) compared with CON. The median VAS increased as CAS was more severe. Results indicated MEL and FLU calves temporally improved behavioral responses following surgical castration with positive numerical trends for a 14 d average daily gain (ADG). The VAS system appeared to be an effective method of subjective evaluation of pain in beef calves in this study. Route of administration, duration of therapy, and low relative cost make oral meloxicam a reasonable analgesic treatment in calves when administered at the time of surgical castration.
