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1.  Effects of single nucleotide polymorphism marker density on degree of genetic variance explained and genomic evaluation for carcass traits in Japanese Black beef cattle 
BMC Genetics  2014;15:15.
Japanese Black cattle are a beef breed whose meat is well known to excel in meat quality, especially in marbling, and whose effective population size is relatively low in Japan. Unlike dairy cattle, the accuracy of genomic evaluation (GE) for carcass traits in beef cattle, including this breed, has been poorly studied. For carcass weight and marbling score in the breed, as well as the extent of whole genome linkage disequilibrium (LD), the effects of equally-spaced single nucleotide polymorphisms (SNPs) density on genomic relationship matrix (G matrix), genetic variance explained and GE were investigated using the genotype data of about 40,000 SNPs and two statistical models.
Using all pairs of two adjacent SNPs in the whole SNP set, the means of LD (r 2 ) at ranges 0–0.1, 0.1–0.2, 0.2–0.5 and 0.5–1 Mb were 0.22, 0.13, 0.10 and 0.08, respectively, and 25.7, 13.9, 10.4 and 6.4% of the r 2 values exceeded 0.3, respectively. While about 90% of the genetic variance for carcass weight estimated using all available SNPs was explained using 4,000–6,000 SNPs, the corresponding percentage for marbling score was consistently lower. With the conventional linear model incorporating the G matrix, correlation between the genomic estimated breeding values (GEBVs) obtained using 4,000 SNPs and all available SNPs was 0.99 for carcass weight and 0.98 for marbling score, with an underestimation of the former GEBVs, especially for marbling score.
The Japanese Black is likely to be in a breed group with a relatively high extent of whole genome LD. The results indicated that the degree of marbling is controlled by only QTLs with relatively small effects, compared with carcass weight, and that using at least 4,000 equally-spaced SNPs, there is a possibility of ranking animals genetically for these carcass traits in this breed.
PMCID: PMC3913948  PMID: 24491120
Marbling score; Carcass weight; Japanese black cattle; SNP marker density; Linkage disequilibrium; Genomic evaluation; Genomic relationship matrix; Genetic variance estimation
2.  Origin of Contamination and Genetic Diversity of Escherichia coli in Beef Cattle 
The possible origin of beef contamination and genetic diversity of Escherichia coli populations in beef cattle, on carcasses and ground beef, was examined by using random amplification of polymorphic DNA (RAPD) and PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the fliC gene. E. coli was recovered from the feces of 10 beef cattle during pasture grazing and feedlot finishing and from hides, carcasses, and ground beef after slaughter. The 1,403 E. coli isolates (855 fecal, 320 hide, 153 carcass, and 75 ground beef) were grouped into 121 genetic subtypes by using the RAPD method. Some of the genetic subtypes in cattle feces were also recovered from hides, prechilled carcasses, chilled carcasses, and ground beef. E. coli genetic subtypes were shared among cattle at all sample times, but a number of transient types were unique to individual animals. The genetic diversity of the E. coli population changed over time within individual animals grazing on pasture and in the feedlot. Isolates from one animal (59 fecal, 30 hide, 19 carcass, and 12 ground beef) were characterized by the PCR-RFLP analysis of the fliC gene and were grouped into eight genotypes. There was good agreement between the results obtained with the RAPD and PCR-RFLP techniques. In conclusion, the E. coli contaminating meat can originate from cattle feces, and the E. coli population in beef cattle was highly diverse. Also, genetic subtypes can be shared among animals or can be unique to an animal, and they are constantly changing.
PMCID: PMC154492  PMID: 12732550
3.  Transcriptomic markers meet the real world: finding diagnostic signatures of corticosteroid treatment in commercial beef samples 
The use of growth-promoters in beef cattle, despite the EU ban, remains a frequent practice. The use of transcriptomic markers has already proposed to identify indirect evidence of anabolic hormone treatment. So far, such approach has been tested in experimentally treated animals. Here, for the first time commercial samples were analyzed.
Quantitative determination of Dexamethasone (DEX) residues in the urine collected at the slaughterhouse was performed by Liquid Chromatography-Mass Spectrometry (LC-MS). DNA-microarray technology was used to obtain transcriptomic profiles of skeletal muscle in commercial samples and negative controls. LC-MS confirmed the presence of low level of DEX residues in the urine of the commercial samples suspect for histological classification. Principal Component Analysis (PCA) on microarray data identified two clusters of samples. One cluster included negative controls and a subset of commercial samples, while a second cluster included part of the specimens collected at the slaughterhouse together with positives for corticosteroid treatment based on thymus histology and LC-MS. Functional analysis of the differentially expressed genes (3961) between the two groups provided further evidence that animals clustering with positive samples might have been treated with corticosteroids. These suspect samples could be reliably classified with a specific classification tool (Prediction Analysis of Microarray) using just two genes.
Despite broad variation observed in gene expression profiles, the present study showed that DNA-microarrays can be used to find transcriptomic signatures of putative anabolic treatments and that gene expression markers could represent a useful screening tool.
PMCID: PMC3541986  PMID: 23110699
DNA-microarray; LC-MS; Anabolic treatment; Cattle; Skeletal muscle; Urine
4.  Proteome differences associated with fat accumulation in bovine subcutaneous adipose tissues 
Proteome Science  2010;8:14.
The fat components of red meat products have been of interest to researchers due to the health aspects of excess fat consumption by humans. We hypothesized that differences in protein expression have an impact on adipose tissue formation during beef cattle development and growth. Therefore, in this study we evaluated the differences in the discernable proteome of subcutaneous adipose tissues of 35 beef crossbred steers [Charolais × Red Angus (CHAR) (n = 13) and Hereford × Angus (HEAN) (n = 22)] with different back fat (BF) thicknesses. The goal was to identify specific protein markers that could be associated with adipose tissue formation in beef cows.
Approximately 541-580 protein spots were detected and compared in each crossbred group, and 33 and 36 protein spots showed expression differences between tissues with high and low BF thicknesses from HEAN and CHAR crossbed, respectively. The annexin 1 protein was highly expressed in both crossbred steers that had a higher BF thickness (p < 0.05) and this was further validated by a western blot analysis. In 13 tissues of CHAR animals and 22 tissues of HEAN animals, the relative expression of annexin 1 was significantly different (p < 0.05) between tissues with high and low BF thicknesses.
The increased expression of annexin 1 protein has been found to be associated with higher BF thickness in both crossbred steers. This result lays the foundation for future studies to develop the protein marker for assessing animals with different BF thickness.
PMCID: PMC2853513  PMID: 20298566
5.  Association of selected SNP with carcass and taste panel assessed meat quality traits in a commercial population of Aberdeen Angus-sired beef cattle 
The purpose of this study was to evaluate the effects of eight single nucleotide polymorphisms (SNP), previously associated with meat and milk quality traits in cattle, in a population of 443 commercial Aberdeen Angus-cross beef cattle. The eight SNP, which were located within five genes: μ-calpain (CAPN1), calpastatin (CAST), leptin (LEP), growth hormone receptor (GHR) and acylCoA:diacylglycerol acyltransferase 1 (DGAT1), are included in various commercial tests for tenderness, fatness, carcass composition and milk yield/quality.
A total of 27 traits were examined, 19 relating to carcass quality, such as carcass weight and fatness, one mechanical measure of tenderness, and the remaining seven were sensory traits, such as flavour and tenderness, assessed by a taste panel.
An SNP in the CAPN1 gene, CAPN316, was significantly associated with tenderness measured by both the tenderometer and the taste panel as well as the weight of the hindquarter, where animals inheriting the CC genotype had more tender meat and heavier hindquarters. An SNP in the leptin gene, UASMS2, significantly affected overall liking, where animals with the TT genotype were assigned higher scores by the panellists. The SNP in the GHR gene was significantly associated with odour, where animals inheriting the AA genotype produced steaks with an intense odour when compared with the other genotypes. Finally, the SNP in the DGAT1 gene was associated with sirloin weight after maturation and fat depth surrounding the sirloin, with animals inheriting the AA genotype having heavier sirloins and more fat.
The results of this study confirm some previously documented associations. Furthermore, novel associations have been identified which, following validation in other populations, could be incorporated into breeding programmes to improve meat quality.
PMCID: PMC2714298  PMID: 19555501
6.  Growth hormone-releasing hormone (GHRH) polymorphisms associated with carcass traits of meat in Korean cattle 
BMC Genetics  2006;7:35.
Cold carcass weight (CW) and longissimus muscle area (EMA) are the major quantitative traits in beef cattle. In this study, we found several polymorphisms of growth hormone-releasing hormone (GHRH) gene and examined the association of polymorphisms with carcass traits (CW and EMA) in Korean native cattle (Hanwoo).
By direct DNA sequencing in 24 unrelated Korean cattle, we identified 12 single nucleotide polymorphisms within the 9 kb full gene region, including the 1.5 kb promoter region. Among them, six polymorphic sites were selected for genotyping in our beef cattle (n = 428) and five marker haplotypes (frequency > 0.1) were identified. Statistical analysis revealed that -4241A>T showed significant associations with CW and EMA.
Our findings suggest that polymorphisms in GHRH might be one of the important genetic factors that influence carcass yield in beef cattle. Sequence variation/haplotype information identified in this study would provide valuable information for the production of a commercial line of beef cattle.
PMCID: PMC1524984  PMID: 16749938
7.  Zeranol may increase the risk of leptin-induced neoplasia in human breast 
Oncology Letters  2010;2(1):101-108.
Breast cancer and obesity are serious health problems and their relationship has been studied for many years. Leptin is mainly secreted by adipocytes and plays a key role in breast cancer development. Leptin expression is up-regulated in obese individuals and promotes breast cancer cell growth. On the other hand, exposure to environmental estrogens has been found to be directly related to breast cancer. Zeranol (Z) is a non-steroidal anabolic growth promoter used in the beef industry in the US. This study focused on the evaluation of Z and Z-containing sera (ZS) and its adverse health risk to human consumption of Z-containing meat produced from Z-implanted beef cattle. We hypothesized that Z increases the risk of breast neoplasia in women, particularly in obese women. A cell proliferation assay, ELISA analysis, RT-PCR and Western blot analysis were conducted. Our study demonstrated that Z and ZS collected from Z-implanted heifers stimulated the proliferation of primary cultured human normal breast epithelial cells (HNBECs) by up-regulating cyclin D1 expression. Leptin increased the sensitivity of HNBECs to Z, and Z increased the ability of HNBECs to secrete leptin. These results suggest an interaction between leptin and Z in HNBECs. Furthermore, Z may play a role in leptin-induced breast neoplasia.
PMCID: PMC3412469  PMID: 22870137
zeranol; leptin; breast cancer; primary cultured human normal breast epithelial cells
8.  Canadian beef quality audit 1998-99. 
The Canadian Veterinary Journal  2001;42(2):121-126.
The second beef quality audit was conducted in Canada in 1998-99 to determine the prevalence of quality defects in slaughtered cattle and to monitor changes since the first audit in 1995. Approximately 0.6% of the number of cattle processed annually in Canada were evaluated. Brands were observed on 49% and tag was observed on 43% of the hides. Both brands and tag had increased from 1995. Seventy percent of the cattle were polled and 5% had full horns; thus, the number of horned cattle had decreased from 1995. Bruises were found on 54% of the carcasses, which was a decrease from 78% in 1995. Sixty-eight percent of the bruises were minor, 28% major, and 4% critical in severity. The distribution of bruises on the carcass was 17% on the chuck, 36% on the rib, 30% on the loin, and 16% on the round. Grubs were observed on 0.008% of the carcasses, and surface injection site lesions were observed on 0.2% of the whole carcasses, a decrease from the 1.3% seen in 1995. Seventy-two percent of the livers were passed for human food and 14% for pet food; 14% were condemned. Approximately 64% of the liver losses were due to abscesses. Five percent of the heads and tongues and 0.3% of the whole carcasses were condemned. The hot carcass weight was highly variable in all cattle, averaging 353 kg (s = 43). The average ribeye area was 90 cm2 (s = 13). Both hot carcass weight and ribeye area had increased from 1995. The average grade fat was 9 mm (s = 5), ranging from 0 mm to 48 mm. Lean meat yield averaged 58.8% (s = 4.6). One percent of the carcasses were devoid of marbling, 17% were Canada A, 49% were Canada AA, 32% were Canada AAA, and 1% were Canada Prime, which was an increase in marbling from 1995. Dark cutters were found in 1% of all carcasses; 1% of steers, 0.5% of heifers, 3% of cows, and 14% of bulls. Three percent of the carcasses were underfinished and 13% were overfinished. The number of overfinished carcasses had increased from 1995. Stages, steers with bullish traits, were infrequently observed in 0.5% of the steers, and 0.2% of the steers and 0.3% of the heifers had poor conformation. Yellow fat was not observed in any steers or heifers, but it was found on 65% of the cow carcasses. Only 0.6% of the heifers had an aged carcass, based on skeletal maturity. Based on August 1998 to July 1999 prices, it was estimated that the Canadian beef industry lost $82.62 per head processed, or $274 million annually, from quality nonconformities, which was an increase from 1995. Additional improvements in management, feeding, handling, genetics, marketing, and grading are needed in the beef industry to reduce quality defects.
PMCID: PMC1476488  PMID: 11272455
9.  Clostridium perfringens strains from bovine enterotoxemia cases are not superior in in vitro production of alpha toxin, perfringolysin O and proteolytic enzymes 
Bovine enterotoxemia is a major cause of mortality in veal calves. Predominantly veal calves of beef cattle breeds are affected and losses due to enterotoxemia may account for up to 20% of total mortality. Clostridium perfringens type A is considered to be the causative agent. Recently, alpha toxin and perfringolysin O have been proposed to play an essential role in the development of disease. However, other potential virulence factors also may play a role in the pathogenesis of bovine enterotoxemia. The aim of this study was to evaluate whether strains originating from bovine enterotoxemia cases were superior in in vitro production of virulence factors (alpha toxin, perfringolysin O, mucinase, collagenase) that are potentially involved in enterotoxemia. To approach this, a collection of strains originating from enterotoxemia cases was compared to bovine strains isolated from healthy animals and to strains isolated from other animal species.
Strains originating from bovine enterotoxemia cases produced variable levels of alpha toxin and perfringolysin O that were not significantly different from levels produced by strains isolated from healthy calves and other animal species. All tested strains exhibited similar mucinolytic activity independent of the isolation source. A high variability in collagenase activity between strains could be observed, and no higher collagenase levels were produced in vitro by strains isolated from enterotoxemia cases.
Bovine enterotoxemia strains do not produce higher levels of alpha toxin, perfringolysin O, mucinase and collagenase, as compared to strains derived from healthy calves and other animal species in vitro.
PMCID: PMC3913962  PMID: 24479821
10.  Use of Antibody Responses against Locus of Enterocyte Effacement (LEE)-Encoded Antigens To Monitor Enterohemorrhagic Escherichia coli Infections on Cattle Farms 
Applied and Environmental Microbiology  2013;79(12):3677-3683.
Enterohemorrhagic Escherichia coli (EHEC) is a significant zoonotic pathogen causing severe disease associated with watery and bloody diarrhea, hemorrhagic colitis, and the hemolytic-uremic syndrome (HUS) in humans. Infections are frequently associated with contact with EHEC-contaminated ruminant feces. Both natural and experimental infection of cattle induces serum antibodies against the LEE-encoded proteins intimin, EspA, EspB, and Tir and the Shiga toxins Stx1 and Stx2, although the latter are poorly immunogenic in cattle. We determined whether antibodies and/or the kinetics of antibody responses against intimin, Tir, EspA, and/or EspB can be used for monitoring EHEC infections in beef cattle herds in order to reduce carcass contamination at slaughter. We examined the presence of serum antibodies against recombinant O157:H7 E. coli intimin EspA, EspB, and Tir during a cross-sectional study on 12 cattle farms and during a longitudinal time course study on two EHEC-positive cattle farms. We searched for a possible correlation between intimin, Tir, EspA, and/or EspB antibodies and fecal excretion of EHEC O157, O145, O111, O103, or O26 seropathotypes. The results indicated that serum antibody responses to EspB and EspA might be useful for first-line screening at the herd level for EHEC O157, O26, and most likely also for EHEC O103 infections. However, antibody responses against EspB are of less use for monitoring individual animals, since some EHEC-shedding animals did not show antibody responses and since serum antibody responses against EspB could persist for several months even when shedding had ceased.
PMCID: PMC3675955  PMID: 23563950
11.  Effect of administration of oat beta-glucan on immune parameters of healthy and immunosuppressed beef steers. 
In order to assess the effect of oat beta-glucan (ObetaG) administration on immune parameters of beef steers, 3 experiments were carried out. In experiment 1, the in vitro effect of ObetaG on the proliferation of blood lymphocytes, with or without the presence of dexamethasone (DXM), was evaluated. In experiment 2, groups of 12 healthy steers were administered ObetaG or saline solution and immunized with ovalbumin (OVA). Immune parameters studied included IgG antibody levels to OVA, proliferation responses of blood lymphocytes to OVA, and blood leukocyte differential cell counts. For experiment 3, groups of 10 steers were treated with ObetaG and DXM, DXM only, or saline solution, and immunized with OVA and keyhole limpet hemocyanin (KLH). Serum antibody responses to OVA and KLH, serum IgG concentration levels, blastogenic responses of blood lymphocytes to OVA and KLH, differential blood leukocyte numbers, and iron and zinc concentration in serum were tested to evaluate the effect of ObetaG to overcome immunosuppression. The in vitro treatment of naive blood lymphocytes with ObetaG did not increase their ability to proliferate; however, when ObetaG was added to cultures of DXM-treated lymphocytes, a significant (P < 0.05 to P < 0.001) reversion of the immunosuppressive effect of DXM occurred. Administration of ObetaG to clinically healthy steers did not induce significant changes on any of the immune parameters studied. The administration of ObetaG to DXM-treated steers provoked, on Day 25, a significant increase in IgG anti-OVA (P < 0.01) and anti-KLH (P < 0.05) responses vs the DXM only group. On Day 25, the specific proliferation responses of lymphocytes, to both OVA and KLH, were significantly increased (P < 0.05) in ObetaG+DXM group compared to DXM group. On Day 4, a significant increase in the number of leukocytes (P < 0.01) and neutrophils (P < 0.001), and a significant decrease in the number of monocytes (P < 0.05) were observed in the group treated with DXM only compared to ObetaG+DXM group. No significant differences were observed in iron and zinc concentration between ObetaG+DXM and DXM groups. These results indicated that ObetaG did not influence immune responses of naive cells in vitro or of healthy steers in vivo; however, when cells or animals were treated with DXM, ObetaG significantly restored some of the specific and non-specific immune parameters studied.
PMCID: PMC1189562  PMID: 10534005
12.  Beef in an Optimal Lean Diet study: effects on lipids, lipoproteins, and apolipoproteins123 
Background: A Step I diet with lean beef compared with lean white meat both decrease LDL cholesterol. To our knowledge, no studies have evaluated a low–saturated fatty acid (SFA) (<7% calories) diet that contains lean beef.
Objective: We studied the effect on LDL cholesterol of cholesterol-lowering diets with varying amounts of lean beef [ie, Dietary Approaches to Stop Hypertension (DASH): 28 g beef/d; Beef in an Optimal Lean Diet (BOLD): 113 g beef/d; and Beef in an Optimal Lean Diet plus additional protein (BOLD+): 153 g beef/d] compared with that of a healthy American diet (HAD).
Design: Thirty-six hypercholesterolemic participants (with LDL-cholesterol concentrations >2.8 mmol/L) were randomly assigned to consume each of the 4 diets (HAD: 33% total fat, 12% SFA, 17% protein, and 20 g beef/d), DASH (27% total fat, 6% SFA, 18% protein, and 28 g beef/d), BOLD (28% total fat, 6% SFA, 19% protein, and 113 g beef/d), and BOLD+ (28% total fat, 6% SFA, 27% protein, and 153 g beef/d) for 5 wk.
Results: There was a decrease in total cholesterol (TC) and LDL-cholesterol concentrations (P < 0.05) after consumption of the DASH (−0.49 ± 0.11 and −0.37 ± 0.09 mmol/L, respectively), BOLD (−0.48 ± 0.10 and −0.35 ± 0.9 mmol/L, respectively), and BOLD+ (−0.50 ± 0.10 and −0.345 ± 0.09 mmol/L, respectively) diets compared with after consumption of the HAD (−0.22 ± 0.10 and −0.14 ± 0.10 mmol/L, respectively). Apolipoprotein A-I, C-III, and C-III bound to apolipoprotein A1 particles decreased after BOLD and BOLD+ diets compared with after the HAD, and there was a greater decrease in apolipoprotein B after consumption of the BOLD+ diet than after consumption of the HAD (P < 0.05 for both). LDL cholesterol and TC decreased after consumption of the DASH, BOLD, and BOLD+ diets when the baseline C-reactive protein (CRP) concentration was <1 mg/L; LDL cholesterol and TC decreased when baseline CRP concentration was >1 mg/L with the BOLD and BOLD+ diets.
Conclusions: Low-SFA, heart-healthy dietary patterns that contain lean beef elicit favorable effects on cardiovascular disease (CVD) lipid and lipoprotein risk factors that are comparable to those elicited by a DASH dietary pattern. These results, in conjunction with the beneficial effects on apolipoprotein CVD risk factors after consumption of the BOLD and BOLD+ diets, which were greater with the BOLD+ diet, provide support for including lean beef in a heart-healthy dietary pattern. This trial was registered at as NCT00937898.
PMCID: PMC3238465  PMID: 22170364
13.  Exploring the Sources of Bacterial Spoilers in Beefsteaks by Culture-Independent High-Throughput Sequencing 
PLoS ONE  2013;8(7):e70222.
Microbial growth on meat to unacceptable levels contributes significantly to change meat structure, color and flavor and to cause meat spoilage. The types of microorganisms initially present in meat depend on several factors and multiple sources of contamination can be identified. The aims of this study were to evaluate the microbial diversity in beefsteaks before and after aerobic storage at 4°C and to investigate the sources of microbial contamination by examining the microbiota of carcasses wherefrom the steaks originated and of the processing environment where the beef was handled. Carcass, environmental (processing plant) and meat samples were analyzed by culture-independent high-throughput sequencing of 16S rRNA gene amplicons. The microbiota of carcass swabs was very complex, including more than 600 operational taxonomic units (OTUs) belonging to 15 different phyla. A significant association was found between beef microbiota and specific beef cuts (P<0.01) indicating that different cuts of the same carcass can influence the microbial contamination of beef. Despite the initially high complexity of the carcass microbiota, the steaks after aerobic storage at 4°C showed a dramatic decrease in microbial complexity. Pseudomonas sp. and Brochothrix thermosphacta were the main contaminants, and Acinetobacter, Psychrobacter and Enterobacteriaceae were also found. Comparing the relative abundance of OTUs in the different samples it was shown that abundant OTUs in beefsteaks after storage occurred in the corresponding carcass. However, the abundance of these same OTUs clearly increased in environmental samples taken in the processing plant suggesting that spoilage-associated microbial species originate from carcasses, they are carried to the processing environment where the meat is handled and there they become a resident microbiota. Such microbiota is then further spread on meat when it is handled and it represents the starting microbial association wherefrom the most efficiently growing microbial species take over during storage and can cause spoilage.
PMCID: PMC3723795  PMID: 23936168
14.  Recombinant Moraxella bovoculi cytotoxin-ISCOM matrix adjuvanted vaccine to prevent naturally occurring infectious bovine keratoconjunctivitis 
A randomized, blinded, controlled field trial was conducted during summer 2006 in a northern California, USA, herd of beef cattle to evaluate the efficacy of a recombinant Moraxella bovoculi cytotoxin subunit vaccine to prevent naturally occurring infectious bovine keratoconjunctivitis (IBK; pinkeye). A convenience sample comprised of 127 steers were administered a subcutaneous dose of either adjuvant alone (ISCOM matrices; control group) or recombinant M. bovoculi cytotoxin carboxy terminus adjuvanted with ISCOM matrices (MbvA group) and were boostered 21 days later. The steers were examined once weekly for 15 weeks for evidence of IBK. No significant difference in the cumulative proportion of corneal ulcerations was detected between groups. Compared to the control calves, the MbvA vaccinates had significantly higher increases in serum neutralizing titers to M. bovoculi hemolysin between week 0 and week 6. The prevalence of M. bovis isolations was higher from ulcerated eyes of calves vaccinated with MbvA as compared to control calves. Vaccination of calves against the carboxy terminus of M. bovoculi RTX toxin resulted in significant increases in serum hemolysin neutralizing titers and may modulate organism type cultured from ulcerated eyes of calves in herds where both M. bovis and M. bovoculi exist. Use of M. bovoculi antigens alone in vaccines to prevent IBK may not be beneficial in herds where IBK is associated with both M. bovoculi and M. bovis.
PMCID: PMC2855018  PMID: 20217228
Moraxella bovoculi; Moraxella bovis; Cytotoxin; Subunit; Infectious bovine keratoconjunctivitis
15.  A rapid method for authentication of Buffalo (Bubalus bubalis) meat by Alkaline Lysis method of DNA extraction and species specific polymerase chain reaction 
Buffalo (Bubalus bubalis) meat is a major item of export from India but export of beef i.e. meat from cattle (Bos indicus) is prohibited. Also, adulteration of buffalo meat with that of beef (meat from cattle) is a common fraudulent practice because of prohibition on cow slaughter in most states of India. Food analysts require precise identification techniques to implement such regulations. In the present study, a method of DNA extraction by Alkaline lysis from meat samples and speciation of buffalo meat using species specific Polymerase Chain Reaction (PCR) has been reported. Alkaline lysis technique is a rapid method which involves triturating meat with four volumes of 0.2N NaOH, dilution of resultant liquid extract with eight volumes of 0.2N NaOH, heating the mix 75 °C for 20 min followed by neutralization with eight volumes of 0.04N Tris HCl. Entire procedure of DNA extraction takes less than 30 min and it is economical as it involves less expensive chemicals. Method was successfully applied in animal byproducts also viz., liver, heart and kidney. For authentication of buffalo meat, pair of primers was designed based on mitochondrial D loop gene nucleotide sequence. PCR amplification using the designed primers gave amplicon of size 482 bp in buffalo and no amplification was detected in closely related species viz., cattle, sheep and goat meat samples. Results of the assay were highly repetitive and reliable. An export sample referred by export regulation authorities was also analyzed by using the Alkaline lysis method of DNA extraction and species specific PCR which enabled authentication of meat within 5 h.
PMCID: PMC3550954  PMID: 24425899
Bubalus bubalis; PCR; DNA; Alkaline lysis; Mitochondrial D loop gene; Speciation
16.  Clostridium perfringens in Meat and Meat Products 
Applied Microbiology  1965;13(3):352-357.
A total of 262 specimens of meat and meat dishes were examined for the presence of Clostridium perfringens. Of this total, 161 were raw, unprocessed beef, veal, lamb, pork, or chicken; 101 were processed meats and meat dishes. C. perfringens was isolated from 113 (43.1%) of these specimens. The highest percentage of contamination (82%) was found in veal cuts, and the lowest (4.7%) in sliced sandwich meats and spreads. Only 2 of the 113 isolates were shown to produce heat-resistant spores, which indicates a very low incidence (0.8%) of contamination. These findings indicate that outbreaks of C. perfringens food-borne disease in the Cincinnati area are caused principally by the contamination of the food with vegetative cells or spores of the organism after cooking. Studies of the effects of various holding temperatures on the growth of C. perfringens indicated that, in the range of 5 to 15 C, no multiplication would occur, but that viable cells would still be present at the end of a 5-day holding period. Extremely rapid growth occurred at temperatures around 45 C, and complete inhibition of growth was accomplished between 49 and 52 C.
PMCID: PMC1058257  PMID: 14325274
17.  Effect of Organic Acids on Escherichia coli O157:H7 and Staphylococcus aureus Contaminated Meat 
Appropriate and safe antibacterial agents able to decontaminate meat surfaces have long been big concern of meat industry. In an attempt to manage beef carcass contamination, spray wash treatments utilizing three concentrations (1, 1.5 and 2%) of acetic, lactic, propionic and formic acids were performed to evaluate their efficacy in reducing numbers of Escherichia coli O157:H7 and Staphylococcus aureus on meat tissues. The procured beef pieces of freshly slaughtered animals were decontaminated with hot water and then inoculated with E. coli O157:H7 and S. aureus individually which then were spray washed with organic acids separately. The total plate count of the treated samples showed that the populations of bacteria decreased after being exposed to organic acids. Spray wash of formic acid resulted in the highest reduction of both bacterial species on meat surface. Significantly, higher log reductions were obtained for S. aureus than E. coli O157:H7. It was concluded that organic acids are highly effective in decontaminating meat surfaces and organic acids are shown to be safe, simple, efficient, and cheap modality of meat decontamination which can be highly recommended for industrial scales.
PMCID: PMC2729390  PMID: 19696918
Meat; beef; Escherichia coli; O157:H7; Staphylococcus aureus; acetic acid; lactic acid; propionic acid; formic acid; food safety.
18.  Variants in the 3' UTR of General Transcription Factor IIF, polypeptide 2 affect female calving efficiency in Japanese Black cattle 
BMC Genetics  2013;14:41.
Calving efficiency can be described as the measure of a cow’s ability to produce viable offspring within a specific period of time. This trait is crucial in beef cattle because calves are necessary both for the production of beef and for heifer replacements. Recently, the number of calves produced at 4 years of age (NCP4) has been used to evaluate the calving efficiency of Japanese Black cattle. To identify variants associated with calving efficiency in Japanese Black cattle, we conducted a genome-wide association study (GWAS) using 688 animals with extreme NCP4 values selected from 15,225 animals.
We identified genetic variants on bovine chromosome 12 (BTA12) that were associated with NCP4. The General Transcription Factor IIF, polypeptide 2 (GTF2F2), located in the 132 kbp-associated region, proved to be in strong linkage disequilibrium. We found 15 associated variants in the promoter and the 3' UTR regions. Consistent with this finding, transcripts of GTF2F2 derived from the haplotype (Q) with the increased number of calves were 1.33-fold more abundant than q-derived transcripts. Furthermore, luciferase assays revealed that the activity of the 3' UTR, a region that includes nine SNPs, was higher in constructs with the Q haplotype than in those with the q haplotype by approximately 1.35-fold. In contrast, the activity of the promoter region did not differ between haplotypes. The association was replicated in an independent sample of 827 animals that were randomly selected from the remainder of the cohort from the same farms used in the GWAS. In the replicated population, the frequency of the Q haplotype is 0.313, and this haplotype accounts for 2.69% of the total phenotypic variance. The effect of the Q to q haplotype substitution on NCP4 was 0.054 calves. These findings suggest that variants in the 3' UTR of GTF2F2 affect the level of GTF2F2 mRNA, which is associated with calving efficiency.
This GWAS has identified variants in the 3’ UTR of GTF2F2 that were associated with the NCP4 of Japanese Black cattle, and this association was validated in an independent sample. The Q haplotype will be immediately useful in improving the calving efficiency of Japanese Black cattle.
PMCID: PMC3656791  PMID: 23663537
Calving efficiency; Number of calves produced at 4 years of age (NCP4); Genome-wide association study; General Transcription Factor IIF, polypeptide 2 (GTF2F2); Beef cattle
19.  Isolation of Leptospira hardjo from kidneys of Ontario cattle at slaughter. 
Kidneys from 117 cattle from 110 Ontario farms were examined at slaughter for leptospires. Leptospira hardjo (hardjo-bovis A) was isolated from 11 kidneys and L. kennewicki from one. The isolations were all made (12/89, 13.5%) from beef cattle from feedlots, no isolates being obtained from dairy or beef cattle from extensive farms (0/28). Isolations were only made from cattle with antibody titers (greater than or equal to 20) against the serovars recovered. Isolation was more sensitive than immunofluorescence in identifying leptospira, particularly in animals with low antibody titers against L. hardjo. Leptospira were isolated from two kidneys with multiple gross lesions of focal nephritis, but there was no correlation between the presence of scanty kidney lesions and isolations of leptospira. Leptospira hardjo infection appears to be common in Ontario feedlot cattle.
PMCID: PMC1255308  PMID: 3300922
20.  Characterization of two Pro-opiomelanocortin gene variants and their effects on carcass traits in beef cattle 
BMC Genetics  2011;12:2.
Carcass quantity (lean meat yield) and quality (degree of marbling) in beef cattle determines much of their economic value. Consequently, it is important to study genes that are part of the appetite pathway and that may ultimately affect carcass composition. Pro-opiomelanocortin is a prohormone that codes for many different peptides, several of which are involved in the appetite pathway. A single nucleotide polymorphism (SNP) c.288C>T in pro-opiomelanocortin (POMC) has previously been associated with hot carcass weight (HCW) and shipping weight (Ship wt) in beef cattle.
While developing a commercial real time PCR test for POMC c.288C>T a 12 bp deletion (POMC c.293_304delTTGGGGGCGCGG) was identified. The deletion results in the removal of four amino acids (a valine, two glycines, and an alanine). Both the POMC c.288C>T and the deletion were genotyped in 386 crossbred steers and evaluated for associations with carcass traits. The animals with one copy of the deletion had a significantly smaller carcass rib-eye area (7.91 cm2; P = 0.02) in comparison to homozygous normal animals. Significant associations were observed between POMC c.288C>T with start-of-finishing weight (SOF WT; P = 0.04), hot carcass weight (HCW; P = 0.02), average fat and grade fat (both P = 0.05), carcass rib-eye area (REA; P = 0.03) and marbling (P = 0.02).
These results suggest that it could be beneficial for beef producers to know both the deletion and POMC c.288C>T genotypes when making marketing and culling decisions.
PMCID: PMC3022757  PMID: 21205304
21.  Anthrax outbreak in a Swedish beef cattle herd - 1st case in 27 years: Case report 
After 27 years with no detected cases, an outbreak of anthrax occurred in a beef cattle herd in the south of Sweden. The outbreak was unusual as it occurred in winter, in animals not exposed to meat-and-bone meal, in a non-endemic country.
The affected herd consisted of 90 animals, including calves and young stock. The animals were kept in a barn on deep straw bedding and fed only roughage. Seven animals died during 10 days, with no typical previous clinical signs except fever. The carcasses were reportedly normal in appearance, particularly as regards rigor mortis, bleeding and coagulation of the blood. Subsequently, three more animals died and anthrax was suspected at necropsy and confirmed by culture and PCR on blood samples.
The isolated strain was susceptible to tetracycline, ciprofloxacin and ampicillin. Subtyping by MLVA showed the strain to cluster with isolates in the A lineage of Bacillus anthracis.
Environmental samples from the holding were all negative except for two soil samples taken from a spot where infected carcasses had been kept until they were picked up for transport.
The most likely source of the infection was concluded to be contaminated roughage, although this could not be substantiated by laboratory analysis. The suspected feed was mixed with soil and dust and originated from fields where flooding occurred the previous year, followed by a dry summer with a very low water level in the river allowing for the harvesting on soil usually not exposed. In the early 1900s, animal carcasses are said to have been dumped in this river during anthrax outbreaks and it is most likely that some anthrax spores could remain in the area.
The case indicates that untypical cases in non-endemic areas may be missed to a larger extent than previously thought. Field tests allowing a preliminary risk assessment of animal carcasses would be helpful for increased sensitivity of detection and prevention of further exposure to the causative agent.
PMCID: PMC2826306  PMID: 20122147
22.  In Vitro Colonization of the Muscle Extracellular Matrix Components by Escherichia coli O157:H7: The Influence of Growth Medium, Temperature and pH on Initial Adhesion and Induction of Biofilm Formation by Collagens I and III 
PLoS ONE  2013;8(3):e59386.
Enterohemorrhagic Escherichia coli (EHEC) O157:H7 are responsible for repeated food-poisoning cases often caused by contaminated burgers. EHEC infection is predominantly a pediatric illness, which can lead to life-threatening diseases. Ruminants are the main natural reservoir for EHEC and food contamination almost always originates from faecal contamination. In beef meat products, primary bacterial contamination occurs at the dehiding stage of slaughtering. The extracellular matrix (ECM) is the most exposed part of the skeletal muscles in beef carcasses. Investigating the adhesion to the main muscle fibrous ECM proteins, insoluble fibronectin, collagen I, III and IV, laminin-α2 and elastin, results demonstrated that the preceding growth conditions had a great influence on subsequent bacterial attachment. In the tested experimental conditions, maximal adhesion to fibril-forming collagens I or III occurred at 25°C and pH 7. Once initially adhered, exposure to lower temperatures, as applied to meat during cutting and storage, or acidification, as in the course of post-mortem physiological modifications of muscle, had no effect on detachment, except at pHu. In addition, dense biofilm formation occurred on immobilized collagen I or III and was induced in growth medium supplemented with collagen I in solution. From this first comprehensive investigation of EHEC adhesion to ECM proteins with respect to muscle biology and meat processing, new research directions for the development of innovative practices to minimize the risk of meat contamination are further discussed.
PMCID: PMC3596346  PMID: 23516631
23.  Apparent Prevalence of Beef Carcasses Contaminated with Mycobacterium avium subsp. paratuberculosis Sampled from Danish Slaughter Cattle 
Presence of Mycobacterium avium subsp. paratuberculosis (MAP) in beef has been reported as a public health concern because asymptomatically infected cattle may contain MAP in tissues that are used for human consumption. Associations between MAP carcasses contamination and animal characteristics such as age, breed, production type, and carcass classification were assessed. Cheek muscles from 501 carcasses were sampled cross-sectionally at a Danish abattoir and tested for presence of viable MAP and MAP DNA by bacterial culture and IS900 realtime PCR, respectively. Cheek muscle tissues from carcasses of two dairy cows were positive by culture whereas 4% of the animals were estimated with ≥10 CFU/gram muscle based on realtime PCR. Age was found to be associated with carcass contamination with MAP. The observed viable MAP prevalence in beef carcasses was low. However, detection of MAP and MAP DNA in muscle tissues suggested that bacteremia occurred in slaughtered cattle.
PMCID: PMC3087313  PMID: 21547261
24.  Genome-wide association study for backfat thickness in Canchim beef cattle using Random Forest approach 
BMC Genetics  2013;14:47.
Meat quality involves many traits, such as marbling, tenderness, juiciness, and backfat thickness, all of which require attention from livestock producers. Backfat thickness improvement by means of traditional selection techniques in Canchim beef cattle has been challenging due to its low heritability, and it is measured late in an animal’s life. Therefore, the implementation of new methodologies for identification of single nucleotide polymorphisms (SNPs) linked to backfat thickness are an important strategy for genetic improvement of carcass and meat quality.
The set of SNPs identified by the random forest approach explained as much as 50% of the deregressed estimated breeding value (dEBV) variance associated with backfat thickness, and a small set of 5 SNPs were able to explain 34% of the dEBV for backfat thickness. Several quantitative trait loci (QTL) for fat-related traits were found in the surrounding areas of the SNPs, as well as many genes with roles in lipid metabolism.
These results provided a better understanding of the backfat deposition and regulation pathways, and can be considered a starting point for future implementation of a genomic selection program for backfat thickness in Canchim beef cattle.
PMCID: PMC3680339  PMID: 23738659
Bovine; Lipid metabolism; Machine learning; Single nucleotide polymorphism (SNP); Subcutaneous fat; Tropical composite cattle
25.  Antibody titers to Pasteurella haemolytica A1 in Ontario beef cattle. 
Indirect bacterial agglutination titers to Pasteurella haemolytica A1 were determined in serum, thoracic, pericardial, or peritoneal fluid from cattle necropsied as part of the Bruce County Beef Project in 1979-80 and 1980-81. Antibody titers were also assayed in serum from 84 calves on entry to feedlots in the fall of 1979. Titers on entry were low compared to antibody levels at necropsy. Cattle which died with pneumonia, in particular those dying of fibrinous pneumonia (shipping fever), had lower levels of antibody to P. haemolytica than did those dying of other causes.
PMCID: PMC1320292  PMID: 6756619

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