竹叶臭蛙的分类学研究Ⅰ．各地理居群的分类探讨（两栖纳：蛙科）

采用形态特征比较,对7省13个居群的竹叶臭蛙Odorrana versabilis的标本进行了研究,结果表明：它们的形态特征差异明显,可以分为3个形态型;同时经数值性状标准判别分析和DNA指纹分析印证其结果与形态学研究结果相吻合,据此将3个形态型标本确定为3个种,其中包括2新种。本文重新界定了3个种的分布范围。     

竹叶臭蛙的分类学研究Ⅰ.各地理居群的分类探讨(两栖纲:蛙科)

采用形态特征比较 ,对 7省 13个居群的竹叶臭蛙Odorranaversabilis的标本进行了研究 ,结果表明 :它们的形态特征差异明显 ,可以分为 3个形态型 ;同时经数值性状标准判别分析和DNA指纹分析印证其结果与形态学研究结果相吻合。据此将 3个形态型标本确定为 3个种 ,其中包括2新种。本文重新界定了 3个种的分布范围。     

竹叶臭蛙的分类学研究I.各地理居群的分类探讨(两栖纲:蛙科)

采用形态特征比较,对7省13个居群的竹叶臭蛙Odorrana ve,rsabilis的标本进行了研究,结果表明:它们的形态特征差异明显,可以分为3个形态型;同时经数值性状标准判别分析和DNA指纹分析印证其结果与形态学研究结果相吻合.据此将3个形态型标本确定为3个种,其中包括2新种.本文重新界定了3个种的分布范围.     

竹叶臭蛙的分类学研究Ⅱ.两新种记述(两栖纲:蛙科)

据记载竹叶臭蛙Odorrana versabilis分布于广西,贵州,湖南,福建,浙江,安徽,广东,海南,作者对比研究了7个省的13个居群标本的形态特征,可分为3个形态型,这3个形态型标本的特征彼此差异明显,被确定为3个不同的种,其中包括2个新种,即小竹叶臭蛙O.exiliversabilis sp.nov。和鸭嘴臭蛙O.nasuta sp.nov.,为印证形态分类的结果,又采用形态数值性状标准差别分析和DNA指纹实验分析,其结果与形态特征分类研究结果一致。本文对2个新种的形态和生物学资料进行了记述。     

中国臭蛙属两新种记述(无尾目：蛙科)

对云南臭蛙Odorrana andersonii各地居群的形态特征进行了深入研究,将原定名为云南臭蛙的9个地区标本分为3个形态型。这3个形态型被确定为3个不同的种,即云南臭蛙和2个新种：景东臭蛙O.jingdongensis sp.nov.和海南臭蛙O.hainanensis sp.nov.。海南臭蛙Odorrana hainanensis sp.nov.,正模CIB64Ⅲ3916,雄性成体59.5mm;采自海南白沙县鹦歌岭,海拔520;1964年8月25日。景东臭蛙Odorrana jingdongensis sp.nov.,正模CIB581505,雌性成体体长90.4mm;采自云南景东县新民乡,海拔1480m,1958年5月29日。模式标本保存在中国科学院成者生物研究所。     

重庆市发现宜章臭蛙

2014年7~8月在重庆市南川区三泉镇采集到5号臭蛙类标本,经形态特征比较和DNA数据比对及遗传距离分析,鉴定为宜章臭蛙(Odorrana yizhangensis),属重庆市新纪录。重庆南川的宜章臭蛙与模式产地标本相比,指关节下瘤雌蛙较雄蛙明显;有外掌突;趾关节下瘤明显,趾间全蹼;后肢前伸贴体时胫跗关节超过吻端。重庆南川区与湖南宜章经度和纬度相距均约5°,为宜章臭蛙已知分布的最西点。宜章臭蛙的形态和遗传分化及分布格局,是否由于我国滇西高山峡谷地区、粤桂湘赣南岭山地和湘渝鄂边境地区特殊地理效应及第四纪冰期气候的反复变化造成的,值得进一步探讨。     

陕西省发现绿臭蛙

在陕西省宁强县青木川国家级自然保护区采集了6只臭蛙类标本,经形态特征比较,鉴定为绿臭蛙(Odorrana margaretae),为陕西省首次发现。本文对其特征和分布进行了讨论。     

中国四川省蛙科一新种——南江臭蛙(两栖纲:无尾目)

对分布于四川、重庆市、贵州、湖北、湖南、安徽和福建的花臭蛙[Odorrana(Odorrana)schmackeri(Boettger,1892)]标本进行了形态比较,结果发现四川省南江和万源县的花臭蛙标本与其他产地的花臭蛙标本在形态特征上存在明显区别,主要是:1)成体雄雌体长之比值较小,约为1∶1.34;2)犁骨齿列短,两内侧间距宽,距内鼻孔远;3)背面深色斑点周围无浅色边缘;4)股后部深色斑大而稀疏;5)趾间蹼较弱,第四趾蹼达远端关节下瘤;6)腹内成熟卵的动植物极均为乳白色。故将四川省南江和万源的标本订为新种--南江臭蛙[Odorrana(Odorrana)nanjiangensis,sp.nov.]。     

中国蛙科一新种(两栖纲,无尾目)

对分布于广西龙胜和湖南宜章的龙胜臭蛙Odorrana(Odorrana)lungshengensis作了进一步比较研究,发现两者的形态特征有明显区别,并将湖南宜章的标本命名为新种--宜章臭蛙Odorrana(Odorrana)yizhangensis sp.nov..     

光雾臭蛙的分布新纪录及地理变异

2009年8月在湖北省保康县五道峡自然保护区采到1种臭蛙类标本,与湖北省已有记录的绿臭蛙Odorrana margaretae明显不同,经与四川南江光雾山标本进行形态特征比较和DNA序列比对,鉴定为光雾臭蛙O.kuangwuensis。湖北省保康县光雾臭蛙体长大于模式产地标本;指序3、4、2、1,第2指与第1指几等长;关节下瘤显著,第2～4指具指基下瘤;4肢背面绿色与黑酱色横纹相间排列,横纹间无云状斑,股、胫部横纹3～4条,跗部和前臂2～3条。湖北省保康县五道峡与四川省南江县光雾山相距近5个经度,是否因地理隔离造成种群间的差异,有待进一步研究。     

云南三种稻蝗基因组DNA的RAPD多态性分析

应用RAPD技术对采自云南龙陵两交水和腾冲曲石马鹿冲的24头稻蝗进行了基因组DNA多态性分析,10条随机引物共产生129条清晰、稳定的谱带.基于Jaccard相似系数,用Between-groups Linkage法得出的聚类图与基于Nei's遗传距离分别用UPGMA和NJ法构建的分子系统树基本一致:供试的所有稻蝗个体分为3个聚类簇:龙陵两交水的3、7～8号个体与腾冲曲石马鹿冲(海拔1450m)的8个个体聚为一支,龙陵两交水的1、2、4～6号个体聚为另一支,两支相聚后与腾冲曲石马鹿冲(海拔1 550 m)的8个个体聚为一起.形态学鉴定表明:龙陵两交水的3、7～8号个体为日本稻蝗,1、2、4～6号个体为小稻蝗,腾冲曲石马鹿冲(海拔1450 m)的8个个体均为日本稻蝗,腾冲曲石马鹿冲(海拔1550 m)的个体为有待进一步鉴定的未知种.形态学鉴定结果与聚类结果完全吻合,充分展示了RAPD技术在区分近缘物种方面的独到优势.比较不同采集地的日本稻蝗、小稻蝗的RAPD分析结果,发现云南的这两种稻蝗在同物种不同种群中的遗传多样性最为丰富,从一个侧面显示出我国云南昆虫的物种多样性及物种内部丰富的遗传多样性.     

Comparative analysis of Borrelia isolates from southeastern USA based on randomly amplified polymorphic DNA fingerprint and 16S ribosomal gene sequence analyses

Fifty-three southern USA Borrelia isolates were characterized using randomly amplified polymorphic DNA fingerprinting analysis (RAPD). Twenty-nine types were recognized among 37 B. andersonii strains, seven types among eight B. bissettii strains, and seven types among seven B. burgdorferi sensu stricto strains. Strain TXW-1 formed a separate RAPD type. Nearly complete sequences of the rrs genes from 17 representative southern Borrelia were determined. The similarity values were found to be 96-100% within the B. burgdorferi sensu lato (s.l.) complex, 94-99% among the relapsing fever borreliae, and 93-99% between the two complexes. Phylogenetic analysis indicated that all the Borrelia strains we analyzed could be divided into two parts: the B. burgdorferi s.l. complex and the relapsing fever borreliae complex. TXW-1 segregated with the North American relapsing fever borreliae and formed a separate subbranch.     

Structure and origin of defective genomes contained in serially passaged herpes simplex virus type 1 (Justin).

Restriction enzyme and hybridization analyses have revealed that high-density DNA prepared from passage 15 of serially passaged herpes simplex virus type 1 (Justin) contains three major classes of modified viral DNA molecules, each composed of distinct but closely related types of repeate units. The DNA sequences within the three types of repeat units are colinear with the DNA sequences located at the right end (between coordinates 0.94 and 1.0) of the parental herpes simplex virus type 1 genome. Thus, the three types of repeat units each contain the entire repeat sequence (ac) (which brackets the unique sequences of the small [S] component of herpes simplex virus type 1 DNA) and differ only with respect to the amount of unique S sequences which they contain. The three classes of high-density DNA molecules were found to be stably propagated between passages 6 and 15 of this series.     

Detection and identification of human papillomavirus types isolated from Korean patients with flat warts

Flat warts, also called verruca planna (VP) or juvenile warts, are benign epithelial proliferations of the skin caused by infection with human papillomaviruses (HPV). Several HPV types are known to be associated with flat warts, and particularly HPV type 3 and 10 have been most frequently reported in other countries. In this study, for the detection and typing of human papillomavirus isolated from Korean patients with flat warts, polymerase chain reaction (PCR) and restriction endonuclease digestion were carried out with a set of restriction endonucleases, using the cloned HPV DNA and DNA from clinical specimens. A unique digestion pattern for HPV type 3 and 10, a form of miniature fingerprinting, enabled us to identify HPV type from the amplified fragments. A total of thirty clinical samples, as either frozen tissue or paraffin-embedded tissue, were investigated to verify the type. All the clinical samples except one were con-firmed to be type 3, one of the most frequently observed types in flat warts, and one sample was neither type 3 nor type 10. Further investigation of the unidentified sample by DNA sequencing and sequence alignment with other known HPV types revealed that the sample was a variant of HPV type 94, one of the EV-related HPVs, with the closest evolutionary distance to the HPV type 10 among the known flat wart-associated HPV types.     

Mitochondrial DNA differentiation between two sympatric morphs of striped jack near Japan

Two sympatric morphs (type A with a vertebral number of 25 and type B with a vertebral number of 24) of striped jack Pseudocaranx dentex (Bloch & Schneider) were analysed genetically. A part of the 16S–rRNA region of mtDNA was amplified with polymerase chain reaction for 24 specimens, and a restriction enzyme fragment polymorphism showed significant differences between the two types. While all specimens sampled in Ogasawara were identified as type B, about 90% of striped jack in Oita were type A and 10% were type B. Although the spawning areas of these two types are still unknown, significant genetic differences between the two sympatric morphs show that recruitment and migration patterns might differ from each other. The current system suggests the possibility that the juveniles of type B in Oita may migrate from the Ogasawara Islands.     

Contig assembly of bacterial artificial chromosome clones through multiplexed fluorescence-labeled fingerprinting

A rapid multiplexed fingerprinting method has been developed for bacterial artificial chromosome (BAC) contig assembly. Defined subsets of BAC DNA fragments that result from digestion by three paired restriction endonucleases are labeled with unique fluorescent F-ddATP for each subset. Lists of the labeled fragment size are generated by an ABI 377 DNA sequencer and the GeneScan analysis software and then processed by an assembly program, FPC (Fingerprinted Contigs), to produce contig maps. Data obtained from the multiplexed labeling permit detection of smaller overlaps than is observed when data from a single double-digest are analyzed. The method has been tested on 98 BACs from chromosome 22 regions where large-scale sequencing is under way and also through simulation, using randomly generated BAC clones derived from existing DNA sequence data. In each case, contig assembly results demonstrated the advantages of multiplexed fingerprinting.     

Evaluation of a novel method based on amplification of DNA fragments surrounding rare restriction sites (ADSRRS fingerprinting) for typing strains of vancomycin-resistant Enterococcus faecium

In the search for an effective DNA-typing technique for use in hospital epidemiology, the performance and convenience of a novel assay based on the fingerprinting of bacterial genomes by amplification of DNA fragments surrounding rare restriction sites (ADSRRS fingerprinting) was tested. A large number of vancomycin-resistant Enterococcus faecium (VREM) isolates from haematological ward patients of the Clinical Hospital in Gdańsk were examined. We found that ADSRRS fingerprinting analysis is a rapid method that offers good discriminatory power. The method demonstrated also excellent reproducibility. The usefulness of the ADSRRS fingerprinting method for molecular typing was compared with pulsed field gel electrophoresis (PFGE) method, which is currently considered the gold standard for molecular typing of isolates recovered from patients and the environment in the course of investigation and control of nosocomial outbreaks. Clustering of ADSRRS fingerprinting data matched pulsed field gel electrophoresis data.The features of ADSRRS fingerprinting technique is discussed in comparison with conventional methods. Data presented here demonstrate the complexity of the epidemiological situation concerning VREM that may occur in a single medical ward.     

DNA fingerprints of Bombyx mori L. Testing of genotypic variability of parthenogenetic strains.

A comparative analysis of the total cellular DNA in certain parthenogenetic specimens of the silkworm (Bombyx mori), produced from the females of the parthenogenetic strains by two types of parthenogenesis, has been performed through the application of the DNA fingerprinting method based on M13 phage DNA as a hybridization probe. It has been shown that parental specimens and their genetically identical off-springs produced through ameiotic parthenogenesis have identical patterns of hybridization with the hypervariable DNA fragments. The off-springs produced through the meiotic type of parthenogenesis have individual-specific patterns of hybridization, revealing a high level of polymorphism of individual genotypes. The results obtained testify to the effectiveness and reliability of this promising method for identification of genotypic variability, marking and genomic characterization of parthenogenetic clones in the silkworm.     

HPV genotypes in high grade cervical lesions and invasive cervical carcinoma as detected by two commercial DNA assays, North Carolina, 2001-2006

Background

HPV typing using formalin fixed paraffin embedded (FFPE) cervical tissue is used to evaluate HPV vaccine impact, but DNA yield and quality in FFPE specimens can negatively affect test results. This study aimed to evaluate 2 commercial assays for HPV detection and typing using FFPE cervical specimens.

Methods

Four large North Carolina pathology laboratories provided FFPE specimens from 299 women ages18 and older diagnosed with cervical disease from 2001 to 2006. For each woman, one diagnostic block was selected and unstained serial sections were prepared for DNA typing. Extracts from samples with residual lesion were used to detect and type HPV using parallel and serial testing algorithms with the Linear Array and LiPA HPV genotyping assays.

Findings

LA and LiPA concordance was 0.61 for detecting any high-risk (HR) and 0.20 for detecting any low-risk (LR) types, with significant differences in marginal proportions for HPV16, 51, 52, and any HR types. Discordant results were most often LiPA-positive, LA-negative. The parallel algorithm yielded the highest prevalence of any HPV type (95.7%). HR type prevalence was similar using parallel (93.1%) and serial (92.1%) approaches. HPV16, 33, and 52 prevalence was slightly lower using the serial algorithm, but the median number of HR types per woman (1) did not differ by algorithm. Using the serial algorithm, HPV DNA was detected in >85% of invasive and >95% of pre-invasive lesions. The most common type was HPV16, followed by 52, 18, 31, 33, and 35; HPV16/18 was detected in 56.5% of specimens. Multiple HPV types were more common in lower grade lesions.

Conclusions

We developed an efficient algorithm for testing and reporting results of two commercial assays for HPV detection and typing in FFPE specimens, and describe HPV type distribution in pre-invasive and invasive cervical lesions in a state-based sample prior to HPV vaccine introduction.     

AFLP在分子生物学研究中的应用

扩增片段长度多态性（AFLP）可靠性强,多态检出率高,因而被认为是最有效的DNA指纹分析技术。AFLP已广泛应用于分类学、病理学、种群遗传学、DNA指纹分析的研究和建立数量性状基因图谱,成为最主要的遗传标记。介绍了AFLP的原理、影响因素及其在分子生物学研究中的应用。