Isolation and genomic characterization of Staphylococcus aureus bacteriophages from Chennai, India.

We isolated and characterized two lytic bacteriophages against Staphylococcus aureus named TANUVAS_MVC-VPHSA1 and TANUVAS_MVC-VPHSA2, with the aim of investigating their genomic and structural features. The bacteriophages belong to the Caudoviricetes, and their genomes have sizes of 50,505 and 50,516 base pairs with a GC content of 41.4%.

Prevalence and Characterization of Staphylococcus aureus Isolated from Retail Raw Milk Samples in Chennai, India.

Milk is an important source of food, and it is also a nutrient-rich medium, which can harbor multiple microorganisms. Staphylococcus aureus is an important foodborne pathogen in food-producing animals, and there have been many reports on its infection and antimicrobial resistance (AMR), which has significant global public health concerns. This study was designed to isolate, characterize, and analyze the AMR pattern of S. aureus from milk samples collected in Chennai, India. A total of 259 raw milk samples from 3 groups: dairy farms, local vendors, and retail outlets were analyzed, and it was found that 34% (89/259) were positive for S. aureus. Positive isolates were further characterized by pulsed-field gel electrophoresis and isolates recovered from different sources, study areas, and locations showed high genetic diversity with no similarity. The presence of AMR has been further assessed by phenotypic methods as per CLSI-M100 performance standards, and all the isolates were susceptible to ampicillin/sulbactam, mupirocin, and tylosin. Additionally, all of the isolates were resistant to ampicillin. There were 28 isolates categorized as multidrug-resistant, which showed resistance to more than 2-3 classes of antimicrobials. This is the first report of inducible clindamycin resistance and mupirocin sensitivity pattern from S. aureus isolates recovered from milk. This study established the occurrence varied with genetic diversity in the isolates prevalent in the study area and divergence pattern of AMR S. aureus. The AMR in these isolates and with methicillin-resistant S. aureus could pose a serious threat to food safety and economic implications.

Effects of Core Antigen Bacterin with an Immunostimulant on Piglet Health and Performance Outcomes When Challenged with Enteric and Respiratory Pathogens.

A total of 90 pigs, approximately one day of age, were used in a 42-day study to evaluate whether Endovac-Porci, a core antigen vaccine with an immunostimulant, provides piglets with broad-spectrum protection against the enteric and respiratory effects of Gram-negative bacteria. This study was a single-site, randomized, prospective, blinded, comparative placebo-controlled design. Individual pigs were randomly allocated to 1 of 2 treatments in a randomized design. An individual pig was considered the experimental unit for the farrowing phase (Study day 0 to 21), and the pen was considered the experimental unit for the nursery phase (Study day 21 to 42). Thus, there were 45 replications per treatment during the farrowing phase and 15 replications per treatment during the nursery phase. Treatments included a control product (saline; CP) and an investigational product (Endovac-Porci; IVP). On Study day 23, all pigs were challenged with enterotoxigenic Escherichia coli strain expressing K88 (F4) fimbriae and Pasteurella multocida. Individual pigs were weighed and feed consumption was measured to determine body weight gain, average daily gain, and feed-to-gain ratio. Clinical and fecal scores and overall health were recorded daily. Overall, administering the IVP to pigs led to an increase (p < 0.01) in body weight gain and average daily gain compared to pigs administered the CP. Pigs administered the IVP had reduced (p < 0.01) mortality compared to pigs administered the CP. There was a Study day × treatment interaction on clinical and fecal scores (p < 0.01). There was also a main effect of Study day where clinical and fecal scores increased (p < 0.01) as the Study day increased. Treatment also had an effect on clinical and fecal scores, where pigs administered the IVP had lower (p < 0.01) clinical and fecal scores compared to pigs administered the CP. In conclusion, administering pigs with the Endovac-Porci vaccination significantly improved the performance (i.e., body weight, body weight gain, and average daily gain) and health (i.e., clinical and fecal scores), while reducing the overall mortality in pigs challenged with E. coli K88 orally and Pasteurella multocida intranasally post-weaning. Results from this study suggest that Endovac-Porci could provide broad-spectrum protection against enteric and respiratory effects of Gram-negative bacteria in piglets.

Comparison of lidocaine alone or in combination with a local nerve block of ethanol, bupivacaine liposome suspension, or oral meloxicam to extend analgesia after scoop dehorning in Holstein calves.

The American Veterinary Medical Association recommends the use of practices that reduce or eliminate pain and discomfort associated with dehorning. Identification of an effective, long-acting local anesthetic that is practical for producers to implement and reduces pain from dehorning would benefit animal welfare. Thirty-two Holstein bulls and heifers were enrolled. The objective of this study was to compare the efficacy and duration of activity of bupivacaine liposome suspension (BUP; n = 8), ethanol (ETH; n = 8), or meloxicam (LID + MEL; n = 8) co-administered with lidocaine compared with lidocaine only (LID; n = 8), and to quantify their effect on pain biomarkers and behaviors after scoop dehorning with cauterization in approximately 20-wk-old calves. Outcome variables collected included infrared thermography (IRT), mechanical nociceptive threshold (MNT), visual analog scale (VAS) scoring, and blood sampling for serum cortisol and prostaglandin E2 metabolites (PGEM). There was evidence of a sex effect for MNT, with bulls demonstrating a higher threshold (13.74 kgf) compared with heifers (12.12 kgf). There was a treatment by time interaction for cortisol concentrations (ng/mL). At 2 h, the BUP group had higher cortisol values (17.32 ng/mL) than the LID + MEL group (3.10 ng/mL). Heifers also had higher mean cortisol values (13.88 ng/mL) compared with bulls (6.96 ng/mL). There was a treatment by time interaction for PGEM concentration. Calves in the LID + MEL group had lower PGEM values at 4 and 8 h (10.23 and 9.12 pg/mL) than at -24, 0, and 0.5 h (20.38, 27.27, and 22.59 pg/mL, respectively). At 4 h, the LID + MEL group had lower PGEM concentrations (10.23 pg/mL) than the ETH group (27.08 pg/mL). At 8 h, the LID + MEL group had lower PGEM concentrations (9.12 pg/mL) than both the ETH and BUP groups (24.80 and 20.52 pg/mL). Thus, LID + MEL reduced cortisol and prostaglandin metabolite concentrations more effectively than ETH + LID or BUP + LID administered as a local infiltration and cornual block, respectively, before scoop dehorning followed by cauterization. The treatments administered in the present study did not seem to extend the duration of analgesia beyond the currently recommended multimodal approach, including local anesthesia and systemic analgesia such as lidocaine and meloxicam. Evidence from the current study suggests that sex influences pain biomarkers such as nociceptive threshold and cortisol concentration, with males having a higher nociceptive threshold and lower cortisol responses.

Prevalence of brucellosis in livestock of African and Asian continents: A systematic review and meta-analysis.

Brucellosis is a highly contagious bacterial disease that mainly affects ruminants, but it may affect equines, canines, and felines. The disease is of utmost significance from an economic standpoint in countries where there is no national brucellosis prevention and eradication policy in operation. A systematic review was done to estimate disease burden, incidences, prevalence, and geographical distribution critical in planning appropriate intervention strategies for the control and prevention of Brucellosis. Research articles that were published during the period 2000-2020 were considered for this study after reinforced scrutiny by two independent authors. Meta-regression was used to examine heterogeneity, and subgroup and sensitivity analyses were used to calculate residual heterogeneity and the pooled prevalence of Brucellosis in livestock. Confounders such as geography, a diagnostic test, and species had the greatest R 2 values of 17.8, 8.8, and 2.3%, respectively, indicating the presence of heterogeneity and necessitating more research into sensitivity and subgroup analysis. The combined pooled prevalence of brucellosis in both Asia and African countries was 8% when compared to 12% in the Indian livestock population. The findings of our systematic review and meta-analysis indicate that brucellosis continues to be an important animal and public health concern in developing countries of Asia and Africa, as evidenced by the prevalence rate of brucellosis in these regions. Our findings suggested that well-planned epidemiological surveillance studies in different geographic settings are needed to generate reliable data on disease burden including the economic loss in Asian and African countries.

Behavioral and performance response associated with administration of intravenous flunixin meglumine or oral meloxicam immediately prior to surgical castration in bull calves.

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.

Isolation and characterization of lactic acid bacteria with potential probiotic activity and further investigation of their activity by α-amylase and α-glucosidase inhibitions of fermented batters.

Probiotic microbiota plays a vital role in gastrointestinal health and possesses other beneficial attributes such as antimicrobial and antibiotic agents along with a significant role in the management of diabetes. The present study identifies the probiotic potential of Lactobacillus spp. isolated from three traditionally fermented foods namely, jalebi, medhu vada, and kallappam batters at biochemical, physiological, and molecular levels. By 16S rRNA gene amplification and sequencing, the isolates were identified. A similarity of >98% to Lacticaseibacillus rhamnosus RAMULAB13, Lactiplantibacillus plantarum RAMULAB14, Lactiplantibacillus pentosus RAMULAB15, Lacticaseibacillus paracasei RAMULAB16, Lacticaseibacillus casei RAMULAB17, Lacticaseibacillus casei RAMULAB20, and Lacticaseibacillus paracasei RAMULAB21 was suggested when searched for homology using NCBI database. Utilizing the cell-free supernatant (CS), intact cells (IC), and cell-free extract (CE) of the isolates, inhibitory potential activity against the carbohydrate hydrolyzing enzymes α-glucosidase and α-amylase was assessed. CS, CE, and IC of the isolates had a varying capability of inhibition against α-glucosidase (15.08 to 59.55%) and α-amylase (18.79 to 63.42%) enzymes. To assess the probiotic potential of seven isolates, various preliminary characteristics were examined. All the isolates exhibited substantial tolerance toward gastrointestinal conditions and also demonstrated the highest survival rate (> 99%), hydrophobicity (> 65%), aggregation (> 76%), adherence to HT-29 cells (> 84%), and chicken crop epithelial cells suggesting that the isolates had a high probiotic attribute. Additionally, the strains showed remarkable results in safety assessment assays (DNase and hemolytic), and antibacterial and antibiotic evaluations. The study concludes that the lactic acid bacteria (LAB) characterized possesses outstanding probiotic properties and has antidiabetic effects. In order to obtain various health advantages, LAB can be utilized as probiotic supplements.

Assessment of bovine respiratory disease progression in calves challenged with bovine herpesvirus 1 and Mannheimia haemolytica using point-of-care and laboratory-based blood leukocyte differential assays.

Blood leukocyte differentials can be useful for understanding changes associated with bovine respiratory disease (BRD) progression. By improving turnaround time, point-of-care leukocyte differential assays (PCLD) may provide logistical advantages to laboratory-based assays. Our objective was to assess BRD progression in steers challenged with bovine herpesvirus 1 and Mannheimia haemolytica using point-of-care and laboratory-based blood leukocyte differentials. Thirty Holstein steers (average body weight of 211 kg + 2.4 kg) were inoculated intranasally on day 0 with bovine herpesvirus 1 and intrabronchially on day 6 with Mannheimia haemolytica. Blood leukocytes differentials were measured using both assays from study days 0 to 13. Linear mixed models were fitted to evaluate the associations between: (1) the type of assay (laboratory-based or PCLD) with respect to leukocyte, lymphocyte, and neutrophil concentrations; (2) study day with cell concentrations; and (3) cell concentrations with lung consolidation measured at necropsy. Point-of-care leukocyte, lymphocyte, and neutrophil concentrations were significantly associated (P < 0.05) with the respective cell concentrations obtained from the laboratory-based leukocyte differential. Cell concentrations reported by both assays differed significantly (P < 0.05) over time, indicating shifts from healthy to viral and bacterial disease states. Lymphocyte concentrations, lymphocyte/neutrophil ratios obtained from both assays, and band neutrophil concentrations from the laboratory-based assay were significantly associated (P < 0.05) with lung consolidation, enhancing assessments of disease severity. The PCLD may be a useful alternative to assess BRD progression when laboratory-based leukocyte differentials are impractical.

Genistein: A Potent Anti-Breast Cancer Agent.

Genistein is an isoflavonoid present in high quantities in soybeans. Possessing a wide range of bioactives, it is being studied extensively for its tumoricidal effects. Investigations into mechanisms of the anti-cancer activity have revealed many pathways including induction of cell proliferation, suppression of tyrosine kinases, regulation of Hedgehog-Gli1 signaling, modulation of epigenetic activities, seizing of cell cycle and Akt and MEK signaling pathways, among others via which the cancer cell proliferation can be controlled. Notwithstanding, the observed activities have been time- and dose-dependent. In addition, genistein has also shown varying results in women depending on the physiological parameters, such as the early or post-menopausal states.

Unmitigated Surgical Castration in Calves of Different Ages: Cortisol Concentrations, Heart Rate Variability, and Infrared Thermography Findings.

The objective was to characterize physiological responses to unmitigated surgical castration in calves of varying ages. Thirty male Holstein calves of three ages [<6 w (6W); 3 m (3M); 6 m (6M); n = 10] underwent a simulated castration treatment (SHAM) followed 24 h later by castration (CAST). For both treatments, heart rate variability, eye temperature, and cortisol were measured over time from treatment to specified end points to capture the acute response period. Interactions between treatment and age (p = 0.035) and time and age (p < 0.001) were noted for cortisol. The 6W calves had lower cortisol compared to 6M calves at SHAM and CAST. Cortisol of 6W calves decreased from peak to pre-treatment levels faster than 6M calves. An interaction between time and age was reported in squared differences of inter-beat-intervals (RMSSD; p = 0.02) and high-frequency power (HFP; p = 0.05), whereby both responses decreased in 6W calves during the sampling period which was not seen in 3M and 6M calves. Average eye temperature (AET) differed by age (p = 0.0018) whereby 6W calves had lower AET than 6M calves (p = 0.0013) regardless of treatment and time. The findings suggest that responses to unmitigated surgical castration seem to be mediated by the autonomic nervous system in an age-related manner.

Unmitigated Surgical Castration in Calves of Different Ages: Electroencephalographic and Neurohormonal Findings.

Castration is a common management procedure employed in North American cattle production and is known to cause a pain response. The present study was designed to investigate the effect of unmitigated surgical castration on the electroencephalography (EEG) responses and plasma substance P (SP) concentrations in calves of different ages under the same experimental conditions. Thirty male Holstein calves in three age categories [<6 weeks (6W); 3 months (3M); 6 months (6M); 10 calves per age group] were used in the study. Calves were subjected to a simulated castration session (SHAM) followed 24 h later by surgical castration (CAST) without analgesia. An EEG analysis was performed before the procedure (i.e., baseline), at treatment, and 0-5, 5-10, and 10-20 min post-treatment for both SHAM and CAST, respectively. Blood samples were collected immediately prior to both treatments (time 0) and again at 1, 2, 4, 8, and 12 h after both treatments. The EEG results showed a three-way interaction between treatment, age, and time for delta and beta absolute power, beta relative power, total power, and median frequency (p = 0.004, p = 0.04, p = 0.04, p = 0.03, and p = 0.008, respectively). Following CAST, EEG total power decreased, and median frequency increased relative to SHAM in 6W and 3M calves only following treatment. For 6W and 3M calves, delta and beta absolute power increased at CAST and at later time points relative to SHAM. Marginal evidence for two-way interactions was noted between time and treatment and between age and treatment on the concentration of SP (p = 0.068 and p = 0.066, respectively). Substance P concentrations decreased in CAST treatment compared to SHAM at the later times (8 h: p = 0.007; 12 h: p = 0.048); 6W calves showed lower SP concentration at CAST relative to SHAM (p = 0.017). These findings indicate variation in EEG responses and in SP concentrations following unmitigated surgical castration in calves and that these responses may be age specific. These EEG findings have implications for supporting the perception of the pain associated with surgical castration in young calves and emphasize the urgency of pain mitigation strategies during routine husbandry practices such as castration, as typically implemented in North American cattle management.

Evaluating the utility of a CO2 surgical laser for piglet castration to reduce pain and improve wound healing: a pilot study.

CO2 surgical lasers are widely used for procedures in veterinary and human medicine. There is evidence to suggest surgery using a CO2 laser reduces pain and swelling and improves healing time compared with surgery with a scalpel. Millions of piglets in North America are surgically castrated each year using a scalpel. Therefore, piglet welfare may be improved by making refinements to the surgical procedure. The objectives of this preliminary study were to determine the ability of a CO2 surgical laser to (1) reduce pain and (2) improve wound healing of piglets undergoing surgical castration. Two-day-old male Yorkshire × Landrace piglets were used and randomly assigned to 1 of 3 treatments (n = 10 piglets/treatment group): surgical castration with the CO2 laser, surgical castration with a scalpel, or sham (uncastrated control). Piglets were video recorded in their pens for 1 hr preprocedure and from 0 to 2, 6 to 8, and at 24 hr postprocedure for behavior scoring. Surgical site images were collected at baseline, 0, 8, 24, 48, 72, 96, 120, 144, and 168 hr postcastration for wound healing assessment. Infrared thermography images of the surgical site were also taken at baseline, 0, 0.5, 8, and 24 hr postprocedure to assess inflammation. Finally, blood was collected from each piglet at baseline and 0.5 hr postcastration to assess cortisol levels, prostaglandin E metabolite and pig-major acute phase protein concentration. Laser-castrated piglets displayed more pain behaviors across the observation period than scalpel-castrated piglets (P = 0.05). Laser-castrated piglets also displayed significantly more agonistic behavior than both scalpel-castrated piglets (P = 0.005) and sham piglets (P = 0.036); yet, laser-castrated piglets had significantly lower temperatures at the site of incision compared with scalpel-castrated piglets (P = 0.0211). There was no significant difference in wound healing or any of the blood parameters assessed between laser-castrated and scalpel-castrated piglets. There was evidence of thermal tissue damage on the scrotum of piglets that were castrated using the CO2 laser. This may have resulted in the unremarkable healing time and the increased pain behavior observed in this study. The surgical laser technique should be refined before conclusions can be made regarding the utility of a CO2 laser for piglet castration.

Longitudinal Characterization of Prevalence and Concentration of Shiga Toxin-Producing Escherichia coli Serogroups in Feces of Individual Feedlot Cattle.

The objective of this study was to quantify the frequency, distribution, and variability of fecal shedding and super-shedding of Shiga toxin-producing Escherichia coli (STEC) serogroups O26, O45, O103, O111, O121, O145, and O157 in feedlot cattle over time. A total of 750 fecal grab samples were collected over a 5-week period (June-July 2017) from 150 cattle housed in 10 pens at a commercial feedlot operation. Samples were subjected to culture-based methods and real-time quantitative polymerase chain reaction for STEC detection and quantification. Cumulative animal-level prevalence estimates were 9.5%, 5.2%, and 15.8% for STEC O157, non-O157 STEC serogroups only (STEC-6), and for all STEC serogroups tested (STEC-7), respectively, with the prevalence of STEC O157 and STEC-7 significantly differing between weeks (p < 0.01). Most of the variability in fecal shedding for STEC O157, STEC-6, and STEC-7 was between pens, rather than between cattle. Over the 5-week period, 10 animals (6.7%) persistently shed STEC non-O157 over 3 or more consecutive weeks, whereas 2 animals (1.3%) intermittently shed STEC non-O157 on nonconsecutive weeks. Fifteen animals (10.0%) shed multiple STEC serogroups within the same fecal sample and five animals (3.3%) shed multiple serogroups at super-shedding levels, higher than 104 CFU (colony-forming units)/g, in the same sample. The presence of a super-shedder in a pen was significantly associated with a greater within pen-level prevalence of STEC-6 (p = 0.01). This study gives further insights into intermittent and persistent shedding and super-shedding patterns of STEC serogroups in individual feedlot cattle, which can enable the development and effective application of preharvest and periharvest interventions, as well as surveillance strategies, for these pathogens.

Quantification of Bacteria Indicative of Fecal and Environmental Contamination from Hides to Carcasses.

Fecal bacteria, which reside in the gastrointestinal tract of cattle, can contaminate beef carcasses during processing. In beef cattle slaughter plants, the presence and concentrations of generic Escherichia coli, coliforms, Enterobacteriaceae (EB), and total aerobic bacteria are monitored as indicator organisms of fecal and environmental contamination. The objectives of this study were as follows: (1) to determine the concentrations of generic E. coli, coliforms, EB, and aerobic bacteria on beef carcasses at different processing points in Midwestern commercial beef slaughter plants during the summer, spring, and fall seasons; and (2) to estimate bacterial transfer on carcasses during the hide removal and evisceration processes. Hide and carcass surface sample swabs were collected from slaughtered cattle at four large commercial processing plants. At each plant visit (3 visits to each of the 4 plants) and during 3 seasons, 20 samples were collected at 5 points: hide-on (hide of animal near exsanguination pit), hide-off carcass, pre-evisceration carcass, postevisceration carcass, and postintervention carcass, for a total of 3600 samples. Bacterial concentrations were determined using 3M™ Petrifilm™ plates. Associations between season and processing plant with concentrations of E. coli, coliforms, EB, and total aerobic bacteria, overall, between hide-on and hide-off, and between pre- and post-evisceration, were evaluated using multilevel mixed-effects linear regression models. Bacterial concentrations on beef carcasses significantly decreased throughout processing. Moreover, hide removal was an important source of carcass contamination, given bacterial concentrations detected on hide-off carcass samples were the highest, and bearing in mind that carcass muscle surfaces should be sterile. Results from this study indicate that the interventions applied by the processing plants were effective, as they probably contributed to the significant reduction of bacterial concentrations of carcasses.

Performance of multiple diagnostic methods in assessing the progression of bovine respiratory disease in calves challenged with infectious bovine rhinotracheitis virus and Mannheimia haemolytica1.

The objective of this study was to evaluate the diagnostic performance of chute-side diagnostic methods for detecting physiological and pathological changes as indicators of early bovine respiratory disease (BRD) in calves experimentally inoculated with infectious bovine rhinotracheitis virus (IBR) and Mannheimia haemolytica (Mh). A challenge study was performed over 14 d in 30 Holstein steers [average weight (±SEM) = 211 kilograms (kg) ± 2.4 kg] inoculated on day 0 with IBR and on day 6 with Mh. Diagnostic methods included clinical illness scores (CIS), lung auscultation using a computer-aided stethoscope (CAS), rectal temperature, facial thermography, pulse oximetry, and bilateral thoracic ultrasonography. Animals were randomized into 1 of 5 necropsy days (days 6, 7, 9, 11, and 13) when the percentage of lung consolidation was estimated. The effect of study day on the results of the diagnostic methods and associations between each diagnostic method's values with lung consolidation measured at necropsy were determined with mixed models. Values for all diagnostic methods differed significantly (P < 0.01) by day. During the IBR phase (days 0 to 6) calves had "normal" to "moderate" CIS, whereas during the Mh phase (days 6.5 to 13) scores were predominantly "severe" to "moribund." Similarly, CAS scores were "normal" and "mild acute" during the IBR phase and "mild acute" to "moderate acute" after the Mh challenge. Oxygen saturation did not differ significantly between days 0, 1, 2, 4, and 6; however, significantly decreased 12 h after inoculation with Mh (P < 0.05). Mean lung consolidation between animal's right and left side recorded by ultrasound was 0.13% (±0.07) before the inoculation with Mh. However, during the Mh phase, mean consolidation increased significantly over time (P < 0.05). The percentage of lung consolidation at necropsy ranged from 1.7% (±0.82) on day 6 to 55.4% (±7.49) on day 10. Clinical illness scores, rectal temperature, facial thermography, oxygen saturation, and ultrasonography were significantly associated (P < 0.05) with lung consolidation at necropsy. In addition, there was a significant trend (P = 0.07) between CAS and lung consolidation scores at necropsy. These chute-side diagnostic methods are useful for detecting disease progression on animals with early stages of BRD.

Spiral Plating Method To Quantify the Six Major Non-O157 Escherichia coli Serogroups in Cattle Feces.

Cattle are a major reservoir of the six major Shiga toxin-producing non-O157 Escherichia coli (STEC) serogroups (O26, O45, O103, O111, O121, and O145) responsible for foodborne illnesses in humans. Besides prevalence in feces, the concentrations of STEC in cattle feces play a major role in their transmission dynamics. A subset of cattle, referred to as super shedders, shed E. coli O157 at high concentrations (≥4 log CFU/g of feces). It is not known whether a similar pattern of fecal shedding exists for non-O157. Our objectives were to initially validate the spiral plating method to quantify the six non-O157 E. coli serogroups with pure cultures and culture-spiked fecal samples and then determine the applicability of the method and compare it with multiplex quantitative PCR (mqPCR) assays for the quantification of the six non-O157 E. coli serogroups in cattle fecal samples collected from commercial feedlots. Quantification limits of the spiral plating method were 3 log, 3 to 4 log, and 3 to 5 log CFU/mL or CFU/g for individual cultures, pooled pure cultures, and cattle fecal samples spiked with pooled pure cultures, respectively. Of the 1,152 cattle fecal samples tested from eight commercial feedlots, 122 (10.6%) and 320 (27.8%) harbored concentrations ≥4 log CFU/g of one or more of the six serogroups of non-O157 by spiral plating and mqPCR methods, respectively. A majority of quantifiable samples, detected by either spiral plating (135 of 137, 98.5%) or mqPCR (239 of 320, 74.7%), were shedding only one serogroup. Only one of the quantifiable samples was positive for a serogroup carrying Shiga toxin (stx1) and intimin (eae) genes; 38 samples were positive for serogroups carrying the intimin gene. In conclusion, the spiral plating method can be used to quantify non-O157 serogroups in cattle feces, and our study identified a subset of cattle that was super shedders of non-O157 E. coli. The method has the advantage of quantifying non-O157 STEC, unlike mqPCR that quantifies serogroups only.

Feedlot- and Pen-Level Prevalence of Enterohemorrhagic Escherichia coli in Feces of Commercial Feedlot Cattle in Two Major U.S. Cattle Feeding Areas.

The objective of this study was to determine feedlot- and pen-level fecal prevalence of seven enterohemorrhagic Escherichia coli (EHEC) belonging to serogroups (O26, O45, O103, O111, O121, O145, and O157, or EHEC-7) in feces of feedlot cattle in two feeding areas in the United States. Cattle pens from four commercial feedlots in each of the two major U.S. beef cattle areas were sampled. Up to 16 pen-floor fecal samples were collected from each of 4-6 pens per feedlot, monthly, for a total of three visits per feedlot, from June to August, 2014. Culture procedures including fecal enrichment in E. coli broth, immunomagnetic separation, and plating on selective media, followed by confirmation through polymerase chain reaction (PCR) testing, were conducted. Generalized linear mixed models were fitted to estimate feedlot-, pen-, and sample-level fecal prevalence of EHEC-7 and to evaluate associations between potential demographic and management risk factors with feedlot and within-pen prevalence of EHEC-7. All study feedlots and 31.0% of the study pens had at least one non-O157 EHEC-positive fecal sample, whereas 62.4% of pens tested positive for EHEC O157; sample-level prevalence estimates ranged from 0.0% for EHEC O121 to 18.7% for EHEC O157. Within-pen prevalence of EHEC O157 varied significantly by sampling month; similarly within-pen prevalence of non-O157 EHEC varied significantly by month and by the sex composition of the pen (heifer, steer, or mixed). Feedlot management factors, however, were not significantly associated with fecal prevalence of EHEC-7. Intraclass correlation coefficients for EHEC-7 models indicated that most of the variation occurred between pens, rather than within pens, or between feedlots. Hence, the potential combination of preharvest interventions and pen-level management strategies may have positive food safety impacts downstream along the beef chain.

Pooling of Immunomagnetic Separation Beads Does Not Affect Detection Sensitivity of Six Major Serogroups of Shiga Toxin-Producing Escherichia coli in Cattle Feces.

Shiga toxin-producing Escherichia coli (STEC) of the serogroups O26, O45, O103, O111, O121, and O145, often called non-O157 STEC, are foodborne pathogens. Cattle are asymptomatic reservoirs for STEC; the organisms reside in the hindgut and are shed in the feces, which serve as the source of food product contaminations. Culture-based detection of non-O157 STEC involves an immunomagnetic separation (IMS) step to capture the specific serogroups in complex matrices, such as feces. The IMS procedure is time consuming and labor intensive because of the need to subject each fecal sample to six individual beads. Therefore, our objective was to evaluate whether pooling of IMS beads affects sensitivity of non-O157 STEC detection compared with using individual IMS beads. The evaluation was done by comparing detection of serogroups in feces spiked with pure cultures (experiments 1 and 2) and from feces (n = 384) of naturally shedding cattle (experiment 3). In spiked fecal samples, detection with pools of three, four, six, or seven beads was similar to, or at times higher than, detection with individual IMS beads. In experiment 3, the proportions of fecal samples that tested positive for the six serogroups as detected by individual or pooled beads were similar. Based on noninferiority tests, detection with pooled beads was not substantially inferior to detection with individual beads (P > 0.05). In conclusion, the pooling of IMS beads is a better option for detection of STEC serogroups in fecal samples compared with individual beads because the procedure saves time and labor and has the prospect of a higher throughput.

Escherichia coli O26 in feedlot cattle: fecal prevalence, isolation, characterization, and effects of an E. coli O157 vaccine and a direct-fed microbial.

Escherichia coli O26 is second only to O157 in causing foodborne, Shiga toxin-producing E. coli (STEC) infections. Our objectives were to determine fecal prevalence and characteristics of E. coli O26 in commercial feedlot cattle (17,148) that were enrolled in a study to evaluate an E. coli O157:H7 siderophore receptor and porin (SRP(®)) vaccine (VAC) and a direct-fed microbial (DFM; 10(6) colony-forming units [CFU]/animal/day of Lactobacillus acidophilus and 10(9) CFU/animal/day of Propionibacterium freudenreichii). Cattle were randomly allocated to 40 pens within 10 complete blocks; pens were randomly assigned to control, VAC, DFM, or VAC+DFM treatments. Vaccine was administered on days 0 and 21, and DFM was fed throughout the study. Pen-floor fecal samples (30/pen) were collected weekly for the last 4 study weeks. Samples were enriched in E. coli broth and subjected to a multiplex polymerase chain reaction (PCR) designed to detect O26-specific wzx gene and four major virulence genes (stx1, stx2, eae, and ehxA) and to a culture-based procedure that involved immunomagnetic separation and plating on MacConkey agar. Ten presumptive E. coli colonies were randomly picked, pooled, and tested by the multiplex PCR. Pooled colonies positive for O26 serogroup were streaked on sorbose MacConkey agar, and 10 randomly picked colonies per sample were tested individually by the multiplex PCR. The overall prevalence of E. coli O26 was higher (p<0.001) by the culture-based method compared to the PCR assay (22.7 versus 10.5%). The interventions (VAC and or DFM) had no impact on fecal shedding of O26. Serogroup O26 was recovered in pure culture from 23.9% (260 of 1089) of O26 PCR-positive pooled colonies. Only 7 of the 260 isolates were positive for the stx gene and 90.1% of the isolates possessed an eaeß gene that codes for intimin subtype ß, but not the bfpA gene, which codes for bundle-forming pilus. Therefore, the majority of the O26 recovered from feedlot cattle feces was atypical enteropathogenic E. coli, and not STEC.

Prevalence of Shiga toxin-producing Escherichia coli and associated virulence genes in feces of commercial feedlot cattle.

The objective of this study was to determine the prevalence of Shiga toxin-producing Escherichia coli (STEC) serogroups and associated virulence genes in feces of commercial feedlot cattle. During March to May 2011, fecal samples were collected from individual cattle (n=960) in 10 cohorts (cattle subpopulations within a feedlot) comprising 17,148 total steers that originated from 48 backgrounding operations in six U.S. states. Fecal samples were enriched in E. coli broth and subjected to two detection protocols: (1) an 11-gene multiplex polymerase chain reaction (PCR) that identifies seven O serogroups (O26, O45, O103, O111, O121, O145, and O157) and four virulence genes (stx1, stx2, eae, and ehxA) applied to extracted total DNA ("direct PCR"); and (2) cultural procedures that involve immunomagnetic separation (IMS) with O26, O103, and O111 beads, plating on a nondifferential MacConkey agar, followed by the multiplex PCR of pooled colonies ("culture-based method"). Generalized linear mixed models were used to adjust prevalence estimates for clustering. Based on direct PCR detection, O157 (49.9%) was the most prevalent O serogroup followed by O26 (20.3%), O103 (11.8%), O121 (10.7%), O45 (10.4%), O145 (2.8%), and O111 (0.8%). Cumulative adjusted prevalence estimates were 22.3, 24.6, and 0.01% for O26, O103, and O111 serogroups, respectively, based on culture-based methods. However, prevalence varied significantly by cohort (p-values<0.05) for O26, O121, and O157 based on direct PCR, and for O26, O103, and O111 serogroups based on culture-based methods. Results of this study indicate that all seven STEC serogroups were identified in feedlot cattle feces, with O157, O26, and O103 being the most prevalent serogroups. A substantial proportion of serogroup-positive samples did not harbor Shiga toxin genes; thus, additional elucidation of the potential human health risk is required. Further evaluation of diagnostic methods for non-O157 STEC is needed given their impact on prevalence estimation.