DNA methylation analysis on satellite I region in blastocysts obtained from somatic cell cloned cattle

Many observations have been made on cloned embryos and on adult clones by somatic cell nuclear transfer (SCNT), but it is still unclear whether the progeny of cloned animals is presenting normal epigenetic status. Here, in order to accumulate the information for evaluating the normality of cloned cattle, we analyzed the DNA methylation status on satellite I region in blastocysts obtained from cloned cattle. Embryos were produced by artificial insemination (AI) to non‐cloned or cloned dams using semen from non‐cloned or cloned sires. After 7 days of AI, embryos at blastocyst stage were collected by uterine flushing. The DNA methylation levels in embryos obtained by using semen and/or oocytes from cloned cattle were similar to those in in vivo embryos from non‐cloned cattle. In contrast, the DNA methylation levels in SCNT embryos were significantly higher (P < 0.01) than those in in vivo embryos from non‐cloned and cloned cattle, approximately similar to those in somatic cells used as donor cells. Thus, this study provides useful information that epigenetic status may be normal in the progeny of cloned cattle, suggesting the normality of germline cells in cloned cattle.     

Effect of ensiled hop (Humulus lupulus L.) residues on plasma acetate turnover rate in sheep

An isotope dilution method using [1‐¹³C]sodium acetate was applied to determine the effect of feeding ensiled hop (Humulus lupulus L.) residues on plasma acetate turnover rate in six adult crossbred sheep. The sheep were fed 63 g/kg body weight (BW)^0.75/day of either mixed hay of orchardgrass (Dactylis glomerata L.) and reed canarygrass (Phalaris arundinacea L.) and round bale silage at 3:1 ratio (Hay‐diet), or another where round bale silage was replaced by ensiled hop residues (Hop‐diet) with a crossover design each of a 3‐week period. The isotope dilution method was performed on day 21 of each dietary treatment. Dry matter digestibility was similar between diets, and nitrogen (N) digestibility was lower (P = 0.001) for Hop‐diet than Hay‐diet. However, N retention did not differ between diets. Plasma acetate concentration was lower (P = 0.04) for Hop‐diet than Hay‐diet, and the turnover rate of plasma acetate did not differ between diets. Plasma concentration of lactate and non‐esterified fatty acids were similar between diets. Hop‐diet was found almost comparable to Hay‐diet on plasma acetate turnover rate in the present experimental conditions. Therefore, it could be concluded that hop residues partially could be used as an alternative to traditionally used round bale silage for rearing sheep.     

Analysis of methanogenic archaeal communities of rumen fluid and rumen particles from Korean black goats

Molecular diversity of methanogens in the rumen of Korean black goats was investigated with 16S rRNA gene clone libraries using methanogen‐specific primers. The libraries were composed of rumen fluid‐associated methanogens (FAM) and rumen particle‐associated methanogens (PAM) from rumen‐fistulated Korean black goats. Among the 141 clones of the FAM library, the sequences were mostly related to two phyla, the Methanobacteriaceae family (77.3%) and the Thermoplasmatales family (22.7%); and among the 68 clones of the PAM library, sequences were also mainly clustered in the two phyla, the Thermoplasmatales family (63.24%) and the Methanobacteriaceae family (35.29%). Most of the sequenced clones in the two libraries were closely related to uncultured methanogenic archaeon. Quantitative real‐time PCR revealed that PAM (8.97 log 10) had significantly higher (P < 0.01) density of methanogens by the methanogenic 16S rRNA gene copies than FAM (7.57 log 10). The two clone libraries also showed difference in Shannon index (FAM library 1.70 and PAM library 1.59) and Chao 1 estimator (FAM library 18 and PAM library 17 operational taxonomic units). Apparent differences found in the microbial community from the two 16S rRNA gene libraries could be a result of such factors as the chemical and physical nature of the target material surface, types or component of diets, the interaction between the methanogens and other microbes, and age of the experimental goats.     

3-甲基-4-硝基酚对雄性小鼠生殖毒性的研究

目的探讨3-甲基-4-硝基酚(PNMC)对雄性小鼠的生殖毒性作用。方法 20只小鼠随机分成阴性对照组和PNMC组(分0.9、9和90mg/kg·bw组),经口灌胃染毒30天,观察睾丸和精子的形态变化。结果染毒组小鼠各剂量睾丸生精细胞发生坏死,曲细精管腔内出现细胞团,精子细胞分化异常,形成均质红染颗粒,睾丸曲细精管生精上皮/直径比值在90mg/kg组明显高于对照组;染毒组小鼠精子畸形率明显高于对照组(P<0.05),并且随着染毒浓度的增加,畸形率有增加的趋势。结论 3-甲基-4-硝基酚不仅可以引起生精细胞坏死和抑制精子形成,还可以诱导精子畸变,从而对生殖系统造成损伤。     

COMPARISON OF SONOGRAPHIC FEATURES OF BENIGN AND NEOPLASTIC DEEP LYMPH NODES IN DOGS

The differentiation of benign vs. neoplastic lymph nodes impacts patient management. Specific sonographic features are typically considered when assessing lymph nodes in dogs. However, the usefulness of these criteria in distinguishing benign vs. malignant lymph nodes remains largely unknown, especially for deep lymph nodes. Our aim was to compare sonographic features in benign and neoplastic deep lymph nodes with the hope of identifying predictive criteria. Thirty‐one deep lymph nodes (16 mesenteric, 10 medial iliac, three hepatic, one sternal, and one cranial mediastinal) in 31 dogs were examined prospectively with B‐mode and Color flow Doppler. Lymph nodes were aspirated using ultrasound‐guidance and final diagnosis were established based on cytologic and/or histopathologic interpretation. Prevalence of each sonographic feature and combinations of two features was calculated for each group and compared using a χ²‐test or Student's t‐test for unequal variances. Ten lymph nodes were benign (hyperplastic and/or inflammatory) and 21 were neoplastic. All were hypoechoic, except for one neoplastic lymph node. Maximal short‐axis diameter (P=0.0006) and long‐axis diameter (P=0.01), and SA/LA ratio (P=0.008) were increased significantly for neoplastic (2.8, 5.5 cm, and 0.50, respectively) vs. benign (1.2, 3.8 cm, and 0.34, respectively) lymph nodes. The prevalence of other features was similar between groups. Doppler evaluation was possible in 77% of lymph nodes, but there was no significant difference between groups. When any two ultrasound features were combined, the only difference between benign and neoplastic lymph nodes was for the combination of contour regularity and appearance of the perinodal fat (P=0.03).     

植物乳杆菌α-半乳糖苷酶基因（melA）的克隆与序列分析

试验以L.plantarum1.557的基因组DNA为模板,通过PCR技术扩增α-半乳糖苷酶基因（melA）基因,PCR产物经纯化回收后克隆至pMD18-T载体中,转化E.coli DH5α感受态细胞,筛选阳性克隆,提取质粒进行SacⅠ和Sph Ⅰ酶切及PCR扩增鉴定,并对melA基因片段进行序列测定。结果表明,测得的melA基因序列全长2240 bp,含有酶切位点、启动子、SD序列及编码738个氨基酸的开放阅读框,理论分子质量为84 ku;4种核苷酸中GC含量为47.45%、AT含量为52.55%;序列中有3个碱基发生了变化,均为无义突变,未导致其推导的氨基酸的改变;序列测定结果与GenBank中登录的序列进行比较,核苷酸和氨基酸序列的同源性除与AY873840相比分别为96.6%和92.2%外,其余均大于99%。试验成功克隆了melA基因,为进一步构建重组表达载体奠定了基础。     

Cuticular hydrocarbons in two parapatric species of ants and their hybrid

Discrimination between nestmates and non‐nestmates in social insects is thought to rely on the pattern of cuticular hydrocarbons. We investigated the cuticular hydrocarbon profiles of 2 parapatric sibling ant species, Temnothorax nylanderi (Förster, 1850) and Temnothorax crassispinus (Karavaiev, 1926), and their hybrid. We found that although the profiles show considerable similarities, a discriminant analysis based on the relative peak areas of cuticular hydrocarbons separates the 3 taxa. The profiles of hybrids were not consistently intermediate between those of the paternal species, suggesting either non‐additive interactions among the parental biosynthetic pathways or systematic differences in environment‐derived odor cues.     

Evaluation of Pet‐Related Management Factors and the Risk of Salmonella spp. Carriage in Pet Dogs from Volunteer Households in Ontario (2005–2006)

The purpose of this study was to determine pet‐related management factors that may be associated with the presence of Salmonella spp. in feces of pet dogs from volunteer households. From October 2005 until May 2006, 138 dogs from 84 households in Ontario were recruited to participate in a cross‐sectional study. Five consecutive daily fecal samples were collected from each dog and enrichment culture for Salmonella spp. was performed. A higher than expected number of the dogs (23.2%; 32/138) had at least one fecal sample positive for Salmonella, and 25% (21/84) of the households had at least one dog shedding Salmonella. Twelve serotypes of Salmonella enterica subsp. enterica were identified, with the predominant serotypes being Typhimurium (33.3%; 13/39), Kentucky (15.4%; 6/39), Brandenburg (15.4%; 6/39) and Heidelberg (12.8%; 5/39). Univariable logistic regression models were created with a random effect for household to account for clustering. Statistically significant risk factors for a dog testing positive included having contact with livestock, receiving a probiotic in the previous 30 days, feeding a commercial or homemade raw food diet, feeding raw meat and eggs, feeding a homemade cooked diet, and having more than one dog in the household. In two‐variable models that controlled for feeding raw food, the non‐dietary variables were no longer statistically significant. These results highlight the potential public health risk of including raw animal products in canine diets.     

Temporal distribution and pathogenicity of the predominant tomato‐infecting begomoviruses in Taiwan

Between 1998 and 2009, the four tomato‐infecting begomovirus species detected in Taiwan were Ageratum yellow vein Hualien virus (AYVHuV), Tomato leaf curl Taiwan virus (ToLCTWV), Tomato yellow leaf curl Thailand virus (TYLCTHV) and a newly defined species Tomato leaf curl Hsinchu virus (ToLCHsV). AYVHuV was detected occasionally in 2003 and ToLCHsV only in 2000–2001, whilst ToLCTWV was detected throughout the period. TYLCTHV was first detected in 2005. Between 1998 and 2005, >99% of the begomovirus‐positive samples were infected with ToLCTWV. In 2007 in western Taiwan, 16% of the positive samples were infected with ToLCTWV, 35% with TYLCTHV and 49% with mixed infection (ToLCTWV/TYLCTHV). In contrast, in eastern Taiwan the proportions were 84% ToLCTWV, 2% TYLCTHV and 14% mixed infection. However, throughout Taiwan in 2008–2009, most positive samples were either identified as TYLCTHV (51%) or mixed infection (ToLCTWV/TYLCTHV; 41%), and only 8% were ToLCTWV. This shows a clear trend of shifting from ToLCTWV to TYLCTHV and mixed infection over a short time period in Taiwan. Sequence analyses indicated that tomato‐infecting AYVHuV, an apparent recombinant between ToLCTWV and AYVHuV from Ageratum, represents a new strain Hsinchu. TYLCTHV Taiwan isolates were highly similar to each other, whereas ToLCTWV isolates had greater diversity and were classified into three strains which had one country‐wide and two local distributions. ToLCTWV and TYLCTHV were confirmed as monopartite and bipartite begomoviruses, respectively, by agroinfection followed by transmission with Bemisia tabaci biotype B. In addition, TYLCTHV was found to be mechanically transmissible together with viral DNA‐B.     

Tracking Phytophthora infestans with SSR markers within and between seasons – a field study in Sweden

The dynamics of a late blight epidemic and sexual reproduction in Phytophthora infestans were studied in an experimental field in mid‐Sweden. The field was inoculated with six isolates of P. infestans taken from another potato field where sexual reproduction of the pathogen was suspected. Three weeks after inoculation single‐lesion leaflets were sampled and the resulting isolates characterized using microsatellites (SSRs) and mating type as markers. Among the 151 isolates analysed, the inoculum genotypes constituted more than 80% of the genotypes found, with three other genotypes making up the remainder. The following year, P. infestans obtained from soil samples taken from this field were analysed, and six novel genotypes were identified. Genotypes from the previous summer’s population were not detected. Analysis of the genotypes recovered was consistent with them being recombinants, with the previous summer’s population acting as parents. These findings are consistent with the hypothesis that oospores produced during a summer epidemic in Sweden can overwinter and cause infection the next year.