PCR扩增ZFY,ZFX序列鉴定奶牛性别的灵敏度研究

从克隆、测序所得奶牛ＺＦＹ、ＺＦＸ序列中设计１对高特异性引物（或探针），即分别特异于牛ＺＦＹ、ＺＦＸ的ＺＦ３、ＺＦ４，与克隆时使用的引物ＺＦ２搭配成ＺＦ２、ＺＦ３和ＺＦ２、ＺＦ４。利用这２对引物和ＺＦ１、ＺＦ２对奶牛基因组ＤＮＡ进行ＮｅｓｔＰＣＲ来判断性别，结果达到用超微量（８ｐｇ）ＤＮＡ，即可检出ＺＦＹ、ＺＦＸ序列而判定雌雄的灵敏度；用非同位素标记试剂盒（ＤＩＧ－Ｓｙｓｔｅｍ）标记ＺＦ３、ＺＦ４作为探针，对ＺＦ１、ＺＦ２扩增产物点杂交来判断性别，也达到同样的灵敏度     

Timing of Puberty and Its Relationship with Body Growth and Season in Male Raccoons (Procyon lotor) in Hokkaido

The raccoon (Procyon lotor), indigenous to North America, has naturalized in Japan as an invasive alien species, having been introduced into the country in the 1970s. In Hokkaido, the northernmost island of Japan, feral raccoons have been increasing in number and spreading throughout the island. The age at the onset of puberty for raccoons is important for estimating individual lifetime reproductive success and population growth. The present study investigated the timing of and potential factors affecting the onset of puberty in male raccoons in Hokkaido. External characteristics and histology of testes were studied in 151 male feral raccoons and in 1 captive juvenile. For the majority of feral yearling raccoons, prepubertal development began in May, and spermatozoa production began in October prior to their second mating season. However, some larger juveniles attained puberty during the juvenile period. The captive juvenile, which was fed throughout the winter, attained puberty only 11 months after birth. These results suggest that if male raccoons can achieve enough body growth before the first mating season, puberty can be attained early. In both juveniles and yearlings, spermatozoa production was only observed after autumn. This timing coincided with the recrudescence of seasonally active spermatogenesis in adult males. Therefore, attaining puberty in male raccoons appears to require both adequate body nutrient development and several environmental factors that control seasonal testicular changes.     

PCR法鉴定雏鸽性别的初步研究

雏鸽的雌雄外貌形态差别甚微,极难区别。为有效准确地鉴别雏鸽的性别,淘汰非目的性别,从而降低饲养成本,提高经济效益。本研究通过采用PCR方法,选定特殊的性别CHD1基因及鉴定引物,克隆出雏鸽特定性别基因序列鉴定雏鸽的雌雄。结果表明:在雏鸽雌性样本中扩增出1条带(777 bp),而雄性中未见扩增带,显示出CHD1基因的性别特异性,PCR扩增性别鉴定结果与样本解剖后对性腺观察的形态学鉴定结果也完全一致。在87份样品中,阳性克隆结果为25,阴性克隆结果为62,故87只雏鸽中雌鸽25只,雄鸽62只,其准确率达100%。     

牛胚胎性别鉴定荧光定量PCR方法的优化与应用

旨在建立一种可检测牛早期胚胎性别的复合探针体系,减少常规PCR方法电泳所带来的污染,提高鉴定准确率,降低鉴定成本.本研究以SRY为牛胚胎性别鉴定的目标基因,以进口YCD-PCR性别鉴定试剂盒为对照组(n=9543),荧光定量PCR的单引物分别扩增体系(荧光单扩,FSPSA,n=6570)和双引物混合扩增体系(荧光双扩,...     

A Real-Time PCR for Detection and Quantification of Mycoplasma ovipneumoniae

A real-time PCR for detection and quantification of M. ovipneumoniae was developed using 9 recently sequenced M. ovipneumoniae genomes and primers targeting a putative adhesin gene p113. The assay proved to be specific and sensitive (with a detection limit of 22 genomic DNA) and could quantify M. ovipneumoniae DNA over a wide linear range, from 2.2 × 10² to 2.2 × 10⁷ genomes.     

Bacteriological and Molecular Detection of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus in Equines of Northern India

Present study was undertaken to study the prevalence of β-haemolytic streptococci in equine of northern temperate region of Jammu and Kashmir, India. One hundred and forty one samples were collected in duplicate from nasopharyngeal tract of diseased (53) and apparently healthy equine (88) for isolation and direct PCR. A total of 77 isolates of streptococci were recovered from 141 samples with an overall prevalence rate of 54.60%. Out of these 77 isolates, 52 were from diseased and 25 from apparently healthy animals. Of the 77 isolates, 4 were identified as Streptococcus equi subsp. equi, 56 as S. equi subsp. zooepidemicus and 17 as S. dysgalactiae subsp. equisimilis. Thus the overall prevalence of S. equi subsp. equi, S. equi subsp. zooepidemicus and S. dysgalactiae subsp. equisimilis was 2.83, 39.71 and 12.05% respectively. The sensitivity of the PCR for the detection of S. equi species was found higher when attempted from direct swab samples.     

牙釉质基因鉴定山羊性别的研究

根据牙釉质(amelogenin,AMEL)同源基因在山羊X、Y染色体上存在不同程度碱基缺失的特点,设计1对引物,对山羊血液基因组DNA混合样进行PCR扩增,将PCR产物纯化、克隆并测序;然后对46个山羊血液DNA样本(♀:22个,♂:24个)进行性别鉴定。结果显示,雌性山羊样品经扩增只产生一条非特异性条带,雄性山羊样品产生一条非特异性条带和一条特异性条带;非特异性片段长度为349 bp,特异性片段为289 bp;46个山羊血液DNA样本鉴定结果与实际性别对比,雌性准确率为100%(22/22),雄性准确率为100%(24/24)。     

Spontaneous Rhabdomyosarcoma in a Common Marmoset ( Callithrix jacchus )

The common marmoset (Callithrix jacchus) is now widely used in various research fields, including toxicology. However, information about the background pathology of this species is scarce. Here, we report a case of rhabdomyosarcoma that spontaneously occurred in a common marmoset. A 44-month-old male common marmoset was euthanized due to bilateral hind limb paralysis. At necropsy, a 2×2×5-cm intramuscular mass was observed in the lower right back. Histologically, the mass was mainly composed of interlacing bundles of spindle-shaped tumor cells. Immunohistochemically, the tumor cells were positive for myogenin, desmin, vimentin and alpha-smooth muscle actin. Ultrastructurally, the tumor cells contained bundles of myofilaments with Z-band-like structures. Thus, the tumor was diagnosed as a rhabdomyosarcoma. To our knowledge, this is the first report of spontaneous rhabdomyosarcoma that was definitely diagnosed in the common marmoset.     

Syphacia (Syphacia) maxomyos sp. n. (Nematoda: Oxyuridae) from Maxomys spp. (Rodentia: Muridae) from Sulawesi and Sumatra,Indonesia

The present report describes Syphacia (Syphacia) maxomyos sp. n. (Nematoda: Oxyuridae) from two species of spiny rats, Maxomys musschenbroekii from Sulawesi and M. whiteheadi from Sumatra. It is characterized by a cephalic plate extending laterally with dorsoventral constriction and stumpy eggs with an operculum rim reaching pole. It is readily distinguishable by the former feature from all of hitherto known representatives of this genus in Indonesia, but it resembles parasites in Murini and Hydromyni rodents in continental Asia and Sahul. This is the first Syphacia species distributed in both the Sunda Shelf and Sulawesi with the exception of Syphacia muris, a cosmopolitan pinworm found in rodents of the of genus Rattus. It is surmised that S. maxomyos is specific to Maxomys and that it was introduced to Sulawesi by dispersal of some Maxomys from the Sunda Shelf.     

兔支气管败血波氏杆菌的分离鉴定及耐药基因检测

本试验旨在了解近年来浙江省兔支气管败血波氏杆菌的感染情况及耐药状况,指导合理用药,检测细菌的耐药基因,探究其耐药机理.根据细菌形态、培养特性、生化试验,结合PCR法对分离菌株进行鉴定;K-B纸片扩散法检测细菌对18种常用抗生素的耐药率;设计耐药基因特异性引物,PCR法扩增耐药基因,并进行测序分析.结果显示,分离鉴定出2012-2014年22株兔支气管败血波氏杆菌;药敏结果显示,分离菌对青霉素G、头孢拉定、链霉素、林可霉素、克林霉素、甲氧嘧啶耐药严重,对多黏菌素B、左氟沙星、强力霉素、四环素等药物敏感,耐β-内酰胺类药物的细菌中检测到耐药基因bla_TEM.结果表明,兔支气管败血波氏杆菌是引起兔呼吸道疾病的主要病原菌,分离菌多重耐药,耐药率较高,检测到的耐药基因与耐药表型相符.     

Genetic and antigenic analysis of Chlamydia pecorum strains isolated from calves with diarrhea

Chlamydia pecorum (designated 22–58) was isolated in 2010 in HmLu-1 cells from the jejunum of a calf which died of necrotizing enterocolitis in Yamaguchi Prefecture, Japan. Immunohistochemical staining identified C. pecorum positive reactions in the jejunal villi. C. pecorum, designated 24–100, was isolated from the feces of a calf with diarrhea in another farm in Yamaguchi Prefecture in 2012. A significant increase in neutralizing antibody titers against C. pecorum was confirmed in paired sera. Nucleotide sequence identities of omp1 genes of the 2 isolates were 100%. The isolates were genetically and antigenically more closely related to C. pecorum Bo/Yokohama strain isolated from cattle with enteritis in Japan than to the other prototype strains, Bo/Maeda isolated from cattle with pneumonia and Ov/IPA isolated from sheep with polyarthritis. These results indicate that C. pecorum strains similar to 22–58 and 24–100 might be endemic in Yamaguchi Prefecture and cause enteric disease in cattle.     

High Prevalence of BlaCTX-M Group Genes in Aeromonas dhakensis Isolated from Aquaculture Fish Species in South Korea

The prevalence of resistant genes against β-lactams in 119 Aeromonas strains was determined. A large number (99.2%) of the present fish strains were resistant to one or more β- lactams including ceftiofur, amoxicillin-clavulanic acid, ampicillin, piperacillin and cefpodoxime. Among antibiotic resistance phenotypes, the simultaneous resistance to all β-lactams occurred in 25.2% (n=30) of all strains, which consisted of 18 strains of A. dhakensis, 8 strains of A. caviae, 2 strains of A. hydrophila and only one strain of A. veronii. For exploring genetic background of the antibiotic resistances, multiple PCR assays were subjected to detect β-lactamase-encoding genes, bla_TEM, bla_OXA-B and bla_CTX-M. In the results, the bla_TEM-1 gene was harbored in all strains, whereas only 3 strains harbored bla_OXA gene. In the case of bla_CTX-M gene, the gene was detected in 21.0% (25 out of 119) of all strains, which countered with 80% (20 out of 25) of A. dhakensis, 8% (2 out of 25) of A. caviae and 12% (3 out of 25) of A. hydrophila. In addition, most of the bla_CTX-M positive strains showed simultaneous resistance to all β-lactams (18 out of 30 strains). In sequence analysis for bla_CTX-M genes detected, they were CTX-M group 1-encoding genes including bla_CTX-M-33 from 3 eel strains of A. dhakensis. Therefore, A. dhakensis obtained from cultured fish could represent a reservoir for spreading genes encoding CTX-M group 1 enzymes and hence should be carefully monitored, especially for its potential risk to public health.     

A case of nontraumatic gas gangrene in a common marmoset (Callithrix jacchus)

The common marmoset is widely used in neuroscience and regenerative medicine research. However, information concerning common marmoset disorders, particularly infectious diseases, is scarce. Here, we report a case of a female common marmoset that died suddenly due to gas gangrene. The animal presented with gaseous abdominal distention at postmortem, and Clostridium perfringens type A was isolated from several tissues. Vacuoles, a Gram-positive bacteremia and intravascular hemolysis were observed microscopically in the muscles, liver and lungs. On the basis of these findings, we diagnosed nontraumatic gas gangrene caused by Clostridium perfringens type A in this common marmoset.