Associations between biosecurity practices and bovine digital dermatitis in Danish dairy herds

The relationship between biosecurity and digital dermatitis (DD) was evaluated in 8,269 cows from a convenience sample of 39 freestall dairy herds. The hypothesis was that poor implementation of biosecurity was associated with higher within-herd prevalence of DD. All lactating cows were scored as negative or positive for DD at the hind legs during milking in the milking parlor. Information about biosecurity was obtained through questionnaires addressed to farmers, on-farm observations, and information from the Danish Cattle Database (www.seges.dk). These assessment tools covered potential infection sources of DD pathogens to susceptible cows (e.g., via animals, humans, manure, vehicles, equipment, and facilities). External and internal biosecurity measures were explanatory variables in 2 separate logistic regression models, whereas within-herd DD prevalence was the outcome. Overall DD prevalence among cows and herds were 24 and 97%, respectively; the within-herd DD prevalence ranged from 0 to 56%. Poor external biosecurity measures associated with higher prevalence of DD were recent animal purchase, access to pasture, lack of boots available for visitors, farm staff working at other dairy farms as well, hoof trimming without a professional attending, and animal transporters having access to cattle area. For internal biosecurity, higher DD prevalence were associated with infrequent hoof bathing, manure scraping less than 8 times a day, manure removal direction from cows to heifers, animal pens' exit without water hoses, manure-handling vehicle used in other activities, and water troughs contaminated with manure. These findings showed that improvements on biosecurity may be beneficial for controlling DD in dairy herds. The study is relevant for farmers facing problems with DD, as well as hoof trimmers, advisors, and veterinarians, who can use the results for optimized recommendations regarding biosecurity in relation to DD. Furthermore, our results might be considered by future studies investigating DD pathogen reservoirs and transmission routes.     

Simultaneous identification of bovine and equine DNA in milks and dairy products inferred from triplex TaqMan real-time PCR technique

Koumiss is a popular dairy product in many lands, traditionally prepared from mare milk with spontaneous fermentation. Mare milk and its fermented derivates are more expensive than cow milk and its fermented derivates, and the possibility exists for producers and dealers to adulterate equine products with bovine items. In this work, we described the development of a triplex real-time PCR based on species-specific TaqMan probes for identification of bovine and equine DNA in milks and dairy products. In addition, a novel designed endogenous control was simultaneously amplified to eliminate possible false negatives. With this methodology, bovine and equine DNA were specifically identified by employing developed primers and probes. The limits of detection of this method were 0.001 ng for cow milk, yogurt, and mare milk, and 0.005 ng for sour soup and koumiss, respectively. In addition, the triplex real-time PCR assay for authentication of animal-derived products was effectively validated using binary DNA and milk mixtures, exhibiting well in terms of specificity, sensitivity, and reproducibility. In short, the triplex PCR assay was verified to be a time-saving and money-saving technique for the identification of bovine and equine DNA in milks and dairy products.     

Randomized clinical trial of tetracycline hydrochloride bandage and paste treatments for resolution of lesions and pain associated with digital dermatitis in dairy cattle

Digital dermatitis is an infectious disease that causes lameness in dairy cattle, a primary welfare concern of the dairy industry. One of the common treatments for this painful hoof disease is through the application of an antibiotic bandage that must be removed following treatment. The objectives of this randomized clinical trial were to determine if topical application of tetracycline hydrochloride in a paste would be as therapeutically effective for the treatment of digital dermatitis as a powdered form of tetracycline hydrochloride held in place by a bandage, and to quantify pain associated with digital dermatitis lesions. Two hundred and fourteen Holstein cow hooves with digital dermatitis lesions were randomly assigned to 1 of 3 treatments: a tetracycline hydrochloride paste, tetracycline hydrochloride powder held in place with a bandage for 2 d, or a negative (untreated) control. Lesions were examined at 2 time periods: 3 to 7 d posttreatment and 8 to 12 d posttreatment to determine healing rates. Nociceptive thresholds were measured using a pressure algometer to quantify the pain at the lesion site. The tetracycline hydrochloride paste was as effective as the powdered bandage treatment in terms of healing rates, with 47.4 and 57.1% hooves healed at 8 to 12 d posttreatment, respectively. Both treatments were more effective than the control, in which no lesions healed 8 to 12 d following initial examination. Mean (±SE) nociceptive thresholds for active, healing, and healed lesions differed, with limb-withdrawal response occurring at 7.45 (±0.67) kg, 12.84 (±1.85) kg, and censored to 25 kg (maximum value of algometer) of force applied, respectively. However, active lesions were not consistently associated with pain, as maximum force was tolerated when applied to 19% of active lesions, perhaps due to variability in stoicism between individual cattle or due to changes in pain during the progression of infection. In conclusion, tetracycline hydrochloride paste was as effective as tetracycline hydrochloride bandage, eliminating the need for bandage removal following treatment application. Digital lesions can be painful during both active and healing stages, suggesting the need for treatment and husbandry interventions for pain mitigation.     

Conventional identification of Streptococcus uberis isolated from bovine mastitis in Argentinean dairy herds

The objective of this study was to evaluate a conventional scheme for identifying Streptococcus uberis strains isolated from bovine mastitis. Seventy-five gram-positive, catalase-negative cocci were collected from cows with mastitis from 19 dairy herds located in the east-central region of Argentina. Five American Type Culture Collection strains and bovine isolates were identified by the API 20 Strep system and by restriction fragment length polymorphism analysis of 16S rDNA. A conventional scheme based on 11 biochemical tests was selected for identification of Strep. uberis strains: the Christie-Atkins-Munch-Petersen reaction; hydrolysis of Arg, esculin, and sodium hippurate; growth in inulin, mannitol, raffinose, salicin, and sorbitol; and growth at 45°C and in 6.5% NaCl. Reference strains and 25 bovine isolates were classified accurately to the species level by the conventional scheme in a blind assay. Each reference strain and each bovine isolate were identified as belonging to the same species following these 3 methods. The remaining 50 isolates identified as Strep. uberis by the API 20 Strep system and 16S rDNA RFLP were assayed by the conventional scheme. This scheme correctly identified 47 (94%) of 50 isolates as Strep. uberis by comparing their biochemical profile with that of the reference strain. Three (6%) of the 50 isolates were classified as Strep. uberis by the API 20 Strep system and by 16S rDNA RFLP and were identified as Enterococcus faecalis by the conventional scheme. Thirty percent of the Strep. uberis strains showed biochemical profiles identical to the Strep. uberis American Type Culture Collection 27958 strain. Seventy percent of the Strep. uberis strains demonstrated variability compared with the reference strain, resulting in 19 different biochemical profiles. The conventional scheme proposed in this study resulted in a relatively low number of misidentifications and could biochemically identify not only typical, but also atypical Strep. uberis strains. This conventional scheme can be considered an adequate method for identifying Strep. uberis strains isolated from bovine mastitis because of its affordable cost in developing countries, and it may contribute to determining the frequency of isolation of Strep. uberis strains in Argentinean dairy herds.     

Short communication: Screening stable flies and house flies as potential vectors of digital dermatitis in dairy cattle

Digital dermatitis is a bacterial infection that causes lesions above the heel bulbs on cattle hooves, and several bacterial species from the genus Treponema are suspected to be causative agents of this polymicrobial condition. Transmission of the bacteria to healthy cows is understudied, particularly with regard to potential insect vectors. Therefore, the objective of this research was to determine if flies captured from a dairy farm known to have digital dermatitis are contaminated with Treponema bacteria. The DNA-based assays did not detect any Treponema phagedenis from stable flies and house flies collected at a dairy experiencing an outbreak of digital dermatitis. Other potential means of bacterial transmission are discussed.     

Evaluation of tetracycline in milk following extra-label administration of topical tetracycline for digital dermatitis in dairy cattle

Digital dermatitis (DD) is a painful infectious foot lesion commonly treated topically with extra-label tetracycline. Our objectives were to determine the concentrations of tetracycline in milk and plasma and to calculate a withdrawal interval following topical application at various doses. Another objective was to evaluate agreement between tests for measuring tetracycline in milk. A randomized block trial was conducted on 2 farms, where 50 cows with active DD lesions on 2 feet were allocated to 1 of 5 treatment groups (n = 10 cows per group). Treatment groups consisted of topical applications of tetracycline hydrochloride, in a paste or as a powdered form under a bandage, at 3 different dosing levels (2, 5, and 25 g) on each of the 2 affected feet. Following enrollment and treatment, samples were collected from milk, teat skin, and blood every 8 to 24 h for up to 7 d postdosing. Concentrations of tetracycline were measured by liquid chromatography-mass spectrometry and milk samples were further tested using the Charm ROSA TET test (Charm Sciences Inc., Lawrence, MA). Tetracycline was present in milk, plasma, and teat skin from all treatment groups. Tetracycline concentrations varied depending on time of sampling, method of application, and dosing level. At 8 h post-treatment, 11% of cows had tetracycline present in milk higher than 100 ng/mL (ppb) but none higher than 300 ng/mL. The 25-g treatment group had the longest estimated withdrawal interval, the highest observed concentrations (210–244 ng/mL) of tetracycline present in milk, and the longest observed consecutive period of tetracycline presence (from 8 to 72 h) among all treatment groups. Compared with liquid chromatography-mass spectrometry, the Charm test had a sensitivity of 77 and 100% for measuring tetracycline in milk at ≥30 and ≥100 ng/mL, respectively. Post-treatment samples of the teat skin were taken from 15 cows on 6 occasions, and every cow had tetracycline present in at least 1 of those 6 samples. This confirms an association between topical DD treatment with tetracycline and contamination of the teat. A total of 22% of blood samples had detectable tetracycline, and the majority (63%) occurred at 8 h post-treatment. At 100 ng/mL, the estimated cow-level milk withdrawal interval ranged from 0 to 70 h. At 300 ng/mL, the estimated cow-level withdrawal interval ranged from 0 to 34 h, and was 0 h at the bulk tank level. We recommend that conservative measures be adopted after extra-label use of topical tetracycline for DD treatment, including using a low dose and strategic post-treatment testing for tetracycline-class drugs in milk.     

Prevalence and risk factors for udder cleft dermatitis in dairy cattle

Udder cleft dermatitis (UCD) is a skin lesion in dairy cattle mostly located at the anterior junction between the udder and the abdominal wall or between the front quarters. Relatively little is known about causative factors for UCD, and few studies have investigated prevalence and risk factors of UCD. Therefore, the aim of this study was to investigate the prevalence of UCD in a random sample of dairy herds with freestalls and milking parlors in a county of Sweden. Thirty dairy herds participated in the study. Each herd was visited once at milking, when every third cow was investigated for presence of UCD. Associations between UCD and milk production, breed, parity, days in milk, claw health, and udder health on the herd and cow levels were also investigated. In addition, a case-control study was performed in 6 herds with a high prevalence of UCD to investigate associations between udder conformation or mange and UCD. Udder cleft dermatitis was found in 18.4% of the 1,084 cows included in the study. The within-herd cow prevalence varied between zero and 39%, with an average of 18.5%. Risk factors for UCD at the herd level were a high proportion of Swedish Red cows and a high production level. At the cow level, breed, parity, and production level were identified as risk factors. The highest risk of having UCD was found in high-producing Swedish Red cows that had calved at least 3 times. Veterinary-treated clinical mastitis was associated with UCD, but cow composite somatic cell count was not. A strong anterior udder attachment was a protective factor, but signs of mange had no association with UCD. The primary cause of UCD is still unclear, and more research is needed to identify the best ways to prevent the development of this animal welfare problem.     

新疆传统乳制品中乳酸菌的分离鉴定

以5种新疆当地传统乳制品为原料进行乳酸菌的分离和鉴定，共分离出36株凝乳状态较好的乳酸菌，对其中16株产酸速率较快的菌株利用部分生理生化和糖发酵试验进行了鉴定。结果表明，9株为乳酸乳球菌乳酸亚种，1株为干酪乳杆菌假植物亚种，2株未知，需进二步鉴定，其余4株杆菌为乳酸乳杆菌。     

Risk factors for increased rates of sole ulcers, white line disease, and digital dermatitis in dairy cattle from twenty-seven farms in England and Wales

Claw lesion treatment records were recorded by farmers on 27 dairy farms (3,074 cows, 36,432 records) in England and Wales between February 2003 and February 2004. These were combined with farm environment and management data collected using a combination of direct observations, interviews with farmers, and milk recording data. Multilevel models were constructed for the 3 most frequently reported lesions related to lameness, namely, sole ulcers, white line disease, and digital dermatitis. Risks associated with an increased incidence of sole ulcers were parity 4 or greater, the use of roads or concrete cow tracks between the parlor and grazing, the use of lime on free stalls, and housing in free stalls with sparse bedding for 4 mo or more. The risks for white line disease were increasing parity and increasing herd size, cows at pasture by day and housed at night, and solid grooved concrete floors in yards or alleys. Solid grooved flooring was also associated with an increased risk of digital dermatitis, and cows 6 or more months after calving had a decreased risk of a first case of digital dermatitis. These results improve our understanding of the specific risks for 3 important lesions associated with bovine lameness and could be used as interventions in future clinical studies targeted at the reduction of specific lesions.     

Short communication: Prevalence of digital dermatitis in Canadian dairy cattle classified as high,average, or low antibody- and cell-mediated immune responders

Lameness is a major animal welfare issue affecting Canadian dairy producers, and it can lead to production, reproduction, and health problems in dairy cattle herds. Although several different lesions affect dairy cattle hooves, studies show that digital dermatitis is the most common lesion identified in Canadian dairy herds. It has also been shown that dairy cattle classified as having high immune response (IR) have lower incidence of disease compared with those animals with average and low IR; therefore, it has been hypothesized that IR plays a role in preventing infectious hoof lesions. The objective of this study was to compare the prevalence of digital dermatitis in Canadian dairy cattle that were classified for antibody-mediated (AMIR) and cell-mediated (CMIR) immune response. Cattle (n = 329) from 5 commercial dairy farms in Ontario were evaluated for IR using a patented test protocol that captures both AMIR and CMIR. Individuals were classified as high, average, or low responders based on standardized residuals for AMIR and CMIR. Residuals were calculated using a general linear model that included the effects of herd, parity, stage of lactation, and stage of pregnancy. Hoof health data were collected from 2011 to 2013 by the farm's hoof trimmer using Hoof Supervisor software (KS Dairy Consulting Inc., Dresser, WI). All trim events were included for each animal, and lesions were assessed as a binary trait at each trim event. Hoof health data were analyzed using a mixed model that included the effects of herd, stage of lactation (at trim date), parity (at trim date), IR category (high, average, and low), and the random effect of animal. All data were presented as prevalence within IR category. Results showed that cows with high AMIR had significantly lower prevalence of digital dermatitis than cattle with average and low AMIR. No significant difference in prevalence of digital dermatitis was observed between high, average, and low CMIR cows. These results indicate that having more robust AMIR is associated with lower prevalence of digital dermatitis hoof lesions.     

Prevalence of digital dermatitis using mirror scoring in Finnish freestall dairy herds

Digital dermatitis (DD) is a severe bacterial hoof disease found worldwide. The disease can be classified into 5 different stages, denoted as M1 to M4 and M4.1, by clinical examination. The main objective of this study was to estimate prevalence of DD lesions in Finnish freestall dairy cattle population through hind feet inspection of standing cows with a mirror. Another aim was to estimate the sensitivity and specificity of mirror scoring on standing cows in a pen or in a milking parlor without washing the feet. Three veterinarians visited 81 randomly selected herds across the country. During the herd visits, hind feet of standing cows (n = 7,010) were scored with a mirror without washing the feet, either when the cows were standing in a pen (n = 4,992) or in the milking parlor (n = 2018). In total, 128 cows (111 from pen and 17 from milking parlor) including 256 feet were chosen with cross-sectional sampling and scored in a trimming chute. Animal-level sensitivity for scoring M2 lesions with a mirror was 55% and specificity was 97%; for all active DD lesions (M1, M2, or M4.1), sensitivity was 36% and specificity was 96%. Sensitivity for scoring any DD lesions was 90% and specificity was 82%. The bias-corrected sensitivity and specificity for scoring any DD lesions were 79% and 92%, respectively. The bias-corrected sensitivity and specificity for scoring M2 DD lesions were 10% and 100%. We found M2 lesions in 12.1% of the study herds, and true herd-level prevalence was the same. Altogether, 33.3% (true prevalence 28.4%) of the herds had either M1, M2, or M4.1 DD lesions. However, only 0.7% (true prevalence 5.4%) of cows in total had active M2 lesions. The within-herd prevalence of M2 lesions (in herds where at least 1 cow had a M2 lesion) was 5.7% and varied between 0.4% and 18.8%. Herds with active DD lesions also had more any DD lesions than herds without active DD lesions. The herd-level prevalence was higher than previously thought, with only 1 herd without any DD lesions. However, the animal-level prevalence of active DD lesions was relatively low. Farmers and veterinarians need to be informed of the disease and possible control measures. Because of the low within-herd prevalence, the control of the disease might be easier than in countries where DD is widespread. Further studies are needed to identify factors associated with DD prevalence in Finnish dairy herds.     

辣白菜中分离含内源性质粒的魏斯氏菌及其鉴定

为了探讨魏斯氏菌在辣白菜中起的作用和获得与明串珠菌相关乳酸细菌的质粒,从辣白菜中分离了魏斯氏菌。辣白菜是朝鲜民族的传统食品,在低温条件下大白菜经过乳酸细菌发酵而成的。将辣白菜汁液均匀涂布在含2%(m/v)CaCO3的MRS平板上,30℃恒温培养箱中培养24h～36h,在中国内地首次分离获得了魏斯氏菌。通过对该菌进行镜检、生理生化特性检测以及16S rDNA序列的系统发育树分析确定其分类地位。结果表明其属于Weissella cibaria,命名为kimshi006,在NCBI上的登录号为HM369807。这一魏斯氏菌菌株含有2条内源性质粒,其大小分别为大约3000bp和8000bp。     

A randomized trial to evaluate the effect of a trace mineral premix on the incidence of active digital dermatitis lesions in cattle

A balanced, parallel-group, single-blinded randomized efficacy study divided into 2 periods was conducted to evaluate the effect of a premix containing higher than typically recommended levels of organic trace minerals and iodine (HOTMI) in reducing the incidence of active digital dermatitis (DD) lesions acquired naturally and induced by an experimental infection challenge model. For the natural exposure phase of the study, 120 healthy Holstein steers 5 to 7 mo of age without signs of hoof disease were randomized into 2 groups of 60 animals. The control group was fed a standard trace mineral supplement and the treatment group was fed the HOTMI premix, both for a period of 60 d. On d 60, 15 steers free of macroscopic DD lesions were randomly selected from each group for the challenge phase and transported to an experimental facility, where they were acclimated and then challenged within a DD infection model. The same diet group allocation was maintained during the 60 d of the challenge phase. The primary outcome measured was the development of an active DD lesion greater than 20 mm in diameter across its largest dimension. No lesions were identified during the natural exposure phase. During the challenge phase, 55% (11/20) and 30% (6/20) of feet were diagnosed with an active DD lesion in the control and treatment groups, respectively. Diagnosis of DD was confirmed by histopathologic demonstration of invasive Treponema spp. within eroded and hyperplastic epidermis and ulcerated papillary dermis. All DD confirmed lesions had dark-field microscopic features compatible with DD and were positive for Treponema spp. by PCR. As a secondary outcome, the average DD lesion size observed in all feet was also evaluated. Overall mean (standard deviation) lesion size was 17.1 (2.36) mm and 11.1 (3.33) mm for the control and treatment groups, respectively, with this difference being driven by acute DD lesions >20 mm. A trend existed for the HOTMI premix to reduce the total DD infection rate and the average size of the experimentally induced lesions. Further research is needed to validate the effect of this intervention strategy in the field and to generate prevention and control measures aimed at optimizing claw health based on nutritional programs.     

豆腐酸浆中一株解淀粉乳杆菌的分离鉴定

利用MRS选择性培养基从自然发酵的豆腐酸浆中分离得到一株产乳酸细菌(编号E36)。初步研究结果表明:该菌株革兰氏染色阳性,菌体呈短杆状,单生或呈短链,菌落呈圆形,乳白色,边缘整齐,表面湿润,不透明;菌株生长过程中,加有溴甲酚紫指示剂的培养基由紫变黄,经生理生化和16S rRNA基因序列分析,初步鉴定该菌为产酸解淀粉乳杆菌。     

酿酒葡萄中内生酵母菌的分离与初步鉴定

采用组织块培养法,从优质酿酒葡萄品种"赤霞珠"中分离出12株内生酵母菌,其中春季4株,夏季3株,秋季5株;从酿酒葡萄的不同组织部位均分离到了内生酵母菌,其中根部6株、茎部2株、叶部1株、果实3株。经形态、培养特征、子囊孢子和假菌丝的观察及生理生化实验,初步鉴定出这些菌株分别为瓶形酵母和克勒克酵母,其中瓶形酵母6株,克勒克酵母6株。     

哈萨克族传统发酵乳酪中发酵菌的分离鉴定

探讨新疆哈萨克族(哈族)传统手工发酵工艺制作的乳酪中所含发酵菌的种类。采用选择性培养基对乳酪中发酵菌分离纯化,然后结合形态学、生理生化特性和16SrRNA、26SrDNAD1/D2区基因序列分析方法进行鉴定。结果表明,从乳酪乳清中分离到两种有价值的微生物一类为酵母菌,另一类为乳酸菌;分别归属为:单孢酿酒酵母(Saccharomyces unisporus)、东方伊萨酵母(Issatchenkia orientalis)和乳酸乳球菌(Lactococcus lactic)。哈族传统发酵乳酪中所含发酵菌的种类与其他地区相比并不完全相同,有其独特的酵母菌资源。     

金沙窖酒酒醅中产香酵母分离与鉴定

从金沙窖酒的酒醅、大曲及环境中筛选得到一株产香酵母FBKL2.0011,将其用于实验室模拟酒厂发酵蒸酒,通过感官评定及气质联用（GC-MS）对酒样挥发性物质进行分析,共检测出43种香气组分.并通过对其个体形态特征、生理生化试验及分子生物学分析,初步鉴定为酵母属（Saccharomyces）.     

贵州黑糯米酒曲优良酵母菌分离筛选与鉴定

从贵州黑糯米酒曲中分离出酵母菌312株。通过菌落形态菌体形态观察及生理生化实验将其初步鉴定至5个属。酵母属57株、假丝酵母属14株、复膜孢酵母属28株、毕赤酵母属212株、德巴利氏酵母属1株。经TTC法、杜氏管法、发酵法及玉米糖化醪法逐级筛选,最终筛选出1株产酒量高产香好的菌株,编号为Q3～J8。并提取菌株Q3-M8ITS序列进行分子生物学鉴定,Q3-J8为布拉酵母（Saccharomyces boulardii）。     

山西老陈醋大曲红曲霉分离鉴定

本试验对山西老陈醋使用的红心大曲中红曲霉进行分离鉴定,从山西老陈醋专用红心大曲中共分离到8株红曲霉,经固体平板培养特征、显微镜镜检个体特征、以及生物学特性和生化试验等,确定其为红曲霉属（Monascus）中的两个种即红色红曲霉（Monascus ruber）和紫色红曲霉（Monascus purpureus）.