Primers for identification of type and other archived specimens of Aphrodes leafhoppers (Hemiptera, Cicadellidae)

Primers were developed for leafhoppers of the genus Aphrodes amplifying 84-244 bp fragments of the mitochondrial cytochrome oxidase subunit I gene. DNA was extracted from legs of over 100-year-old archived museum specimens, amplified and sequenced. The fragments contain sufficient variation to unequivocally identify the different species. The majority of the analysed museum specimens, including three specimens of the syntype series for the UK endemic species A. aestuarina (Edwards 1908), were found to have been assigned to the wrong species. This work clearly underlines the need to validate museum specimens using molecular methods where identity is in doubt, based on reliable standards for species discrimination.     

A case study to quantify the value of voucher specimens for invertebrate conservation: ant records in Lower Austria

Ant records for Lower Austria date back to 1802. Since then, the number of presently recognized central European ant species has increased from 34 to 165. Taxonomic changes and former misidentifications have unfortunately made published historical data unreliable. Therefore, in preparing for the publication of the Red List of Ants of Lower Austria, all voucher specimens from the available major collections were redetermined at the species level. This amounted to a total of 18500 pre-1980 specimens, and 42000 post-1980 specimens. Currently, there are 111 ant species recorded for Lower Austria (including three introduced species). Twelve of these are published here for the first time for Lower Austria, six for all Austria, and one of these is a so far unknown species, discovered among historic voucher specimens. Voucher specimens provide information on the pre-1980 situation of 108 (=97% of the known) species and 32% of the surface area of Lower Austria.     

贵州黏菌初报

通过对作者2005年采自贵州的119份黏菌标本和从湿室培养10份树皮基物获得的19份黏菌标本进行的鉴定分类研究,以及对保存在中国科学院微生物研究所真菌标本馆和中国科学院昆明植物研究所隐花植物标本馆的共21份黏菌标本的鉴定复核,报告了目前已明确了的贵州黏菌51种,分隶属于6目9科21属,其中44种为贵州新记录种,白鳞钙丝菌和红柄发菌为中国新记录种。     

Harvest mites of the family Trombiculidae in the steppe zone of the southeastern Ukraine

Virgin and cultivated biotopes of the Sea of Azov area of Donetsk Province were investigated for parasite coenoses of small mammals. 469 animals and 210 nests were collected, Apodemus sylvaticus, Mus musculus, Microtus arvalis and Cricetulus migratorius being the most abundant species. In all 2940 larvae of trombiculids (18 species) were found on animals and in their nests (2846 specimens on animals and 94 specimens in nests). Two species, Hoffmannina theodori and Doloisia skljari, are new for science, the latter being the first find of the genus Doloisia member in the fauna of the USSR. The majority of parasites are collected from rodents inhabiting virgin biotopes (2328 specimens) while in cultivated biotopes 189 specimens were collected. More rich is the species composition of trombiculids in virgin biotopes (13 species) as compared to that in cultivated ones (5 species), Neotrombicula earis, N. japonica, N. autumnalis and N. vulgaris being mass species. 8 species of larvae are recorded from the nests of rodents. Seasonal dynamics of the abundance of mass species, their occurrence, index of abundance and infection intensity are shown.     

中国荔波世界自然遗产地动物模式标本名录

了解荔波世界自然遗产地动物模式标本分类学研究历史, 可为遗产地履行联合国教科文组织世界遗产中心的要求以及生物多样性保护工作提供参考。本文采用内容分析法, 对采自该遗产地并确立为模式种的物种, 收集相关文献并进行整理和分析, 编制了中国荔波世界自然遗产地动物模式标本名录。结果表明: (1)荔波世界自然遗产地动物模式标本共有204种, 隶属于3门6纲22目60科; (2)动物模式标本物种采集最早可追溯到1955年, 最近为2013年, 两者相隔58年, 而采集高峰期为1995-2004年; (3)以国内学者3人合力发表的物种最多(93种, 占总种数的45.59%); (4)模式物种的橱柜时间严重滞后, 从采集到发表滞后最长的达47年; (5)模式标本涉及66位采集人(或团队), 以国内学者居多(63位, 占总人数的95.45%); (6)遗产地动物模式标本被保存于27家国内外研究机构或私人收藏, 国内收藏1,504号标本, 国外收藏7号; (7)遗产地动物模式标本物种发表于169篇文献, 中文文献有155篇, 外文文献有14篇。随着研究的深入, 荔波世界自然遗产地有更多的新种不断被发现, 说明遗产地物种资源丰富, 生物多样性存在被低估的可能性, 暗示遗产地的物种分类研究工作仍需要投入大量的精力。     

Diversity of sandflies (Psychodidae: Phlebotominae) captured in sandstone caves from Central Amazonia, Brazil

In the present paper we describe the diversity of phlebotomine sandflies collected in three sandstone caves in the municipality of Presidente Figueiredo, state of Amazonas, Brazil. The phlebotomines were captured during 2006 with CDC light traps. Guano samples from inside the Gruta Refúgio do Maruaga were collected to investigate the presence of immature specimens. A total of 2,160 adult phlebotomines representing 15 species were captured. Pintomyia pacae was the dominant species in Gruta dos Animais (1,723 specimens) and Gruta dos Lages (50 specimens) and Deanemyia maruaga new comb (280 specimens) was the dominant species in Gruta Refúgio do Maruaga. A total of 18 guano samples were collected and seven of these samples included immature specimens. A total of 507 immature specimens were captured; 495 of these specimens were larvae and 12 were pupae. The presence of paca (Agouti paca) footprints near Gruta dos Animais and Gruta dos Lages suggests the association of Pi. pacae with this rodent. This finding may explain the abundance of Pi. pacae in these locations, while the species is relatively rare in the forest. Deanemyia maruaga is a cave species that uses guano to breed during its immature stages. Adult specimens of this species are apparently parthenogenetic and autogenous and represent the second record of parthenogenesis for the subfamily Phlebotominae.     

Examination of naturally exposed bottlenose dolphins (Tursiops truncatus) for Microsporidia, Cryptosporidium, and Giardia

Bottlenose dolphins (Tursiops truncatus) captured in the estuarine waters off the coasts of South Carolina and Florida were examined for the presence of Microsporidia, Cryptosporidium sp., and Giardia sp. DNA extracted from feces or rectal swabs was amplified by polymerase chain reaction using parasite-specific small subunit ribosomal RNA gene primers. All positive specimens were subjected to gene sequence analysis. Of 83 dolphins, 17 were positive for Microsporidia. None was positive for Cryptosporidium or Giardia. Gene sequence data for each of the positive specimens were compared with data in GenBank. Fourteen specimens were found similar to, but not identical to, the microsporidian species Kabatana takedai, Tetramicra brevifilum, and Microgemma tinca, reported from fish, and possibly represent parasites of fish eaten by dolphins. Gene sequence data from 3 other specimens had approximately 87% similarity to Enterocytozoon bieneusi, a species known primarily to infect humans and a variety of terrestrial mammals, including livestock, companion animals, and wildlife. It is not clear if these specimens represent a species from a terrestrial source or a closely related species unique to dolphins. There were neither clinical signs nor age- or gender-related patterns apparent with the presence of these organisms.     

Direct identification of slowly growing Mycobacterium species by analysis of the intergenic 16S-23S rDNA spacer region (ISR) using a GelCompar II database containing sequence based optimization for restriction fragment site polymorphisms (RFLPs) for 12 enzymes

To obtain Mycobacterium species identification directly from clinical specimens and cultures, the 16S-23S rDNA spacer (ISR) was amplified using previously published primers that detect all Mycobacterium species. The restriction enzyme that could potentially produce the most restriction fragment length polymorphisms (RFLPs) was determined from all available ISR DNA sequences in GenBank to produce a novel data set of RFLPs for 31 slowly growing Mycobacterium species. Subsequently a GelCompar II database was constructed from RFLPs for 10 enzymes that have been used in the literature to differentiate slowly growing Mycobacterium species. The combination of Sau96I and HaeIII were the best choice of enzymes for differentiating clinically relevant slowly growing Mycobacterium species. A total of 392 specimens were studied by PCR with 195 negative and 197 positive specimens. The ISR-PCR product was digested with HaeIII (previously reported) and Sau96I (new to this study) to obtain a Mycobacterium species identification based on the ISR-RFLPs. The species identification obtained by ISR-RFLP was confirmed by DNA sequencing (isolate numbers are shown in parentheses) for M. avium (3), M. intracellulare (4), M. avium complex (1), M. gordonae (2) and M. tuberculosis (1). The total number of specimens (99) identified were from culture (67), Bactectrade mark 12B culture bottles (11), EDTA blood (3), directly from smear positive specimens (13), tissue (4) and urine (1). Direct species identification was obtained from all 13/13 smear positive specimens. The total number of specimens (99) were identified as M. tuberculosis (41), M. avium (7), M. avium complex (11), M. intracellulare MIN-A (20), M. flavescens (2), M. fortuitum (10), M. gordonae (4), M. shimoidei (1), M. ulcerans (1) and M. chelonae (2). This method reduces the time taken for Mycobacterium species identification from 8-10 weeks for culture and biochemical identification; to 4-6 weeks for culture and ISR-RFLP; to 2 days for smear-positive specimens by ISR-RFLP. The precise 2 day identification obtained may provide significant advantages in clinical management.     

古田山国家级自然保护区木腐真菌物种多样性及分布

木腐真菌是微生物的一个重要类群, 主要以倒木为生长基质, 通过产生各种水解酶将倒木的纤维素、木质素和半纤维素分解为小分子物质, 对促进森林生态系统中的营养物质循环发挥着重要的生态功能。于2016年8月在浙江古田山国家级自然保护区开展的木腐真菌野外调查, 利用形态学和DNA序列分析对采集的标本进行了物种鉴定, 并分析了木腐真菌的物种组成和地理成分。在采集的158份标本中鉴定木腐真菌45属92种, 其中白腐真菌78种, 褐腐真菌14种。古田山的木腐真菌物种区系组成中, 热带-亚热带成分比例最高。在158份木腐真菌标本中, 97份标本采自直径大于10 cm的倒木或树桩上, 分属于76个种, 是木腐真菌生长的主要基质大小类型; 48份标本采自直径为2-10 cm的枝干上, 分属38个种; 13份标本采自直径小于2 cm的枝干上, 分属12种。不同腐烂等级倒木上生长的真菌数量和种类差异明显, 其中一级腐烂倒木上采集到9份标本(7种), 二级腐烂倒木上采集到86份标本(45种), 三级腐烂倒木上49份标本(29种), 四级腐烂倒木上14份标本(14种)。结果表明, 林分中倒木直径大小和腐烂程度是影响木腐真菌生长与分布的重要因子。     

广东鼎湖山自然保护区生物主模式标本内容分析

为了解鼎湖山物种分类学研究的历史,采用内容分析法对采自广东鼎湖山自然保护区的生物主模式标本信息进行了分析。结果表明,鼎湖山自然保护区的生物主模式标本共含202种生物(亚种或变种),分属3界10门20纲54目104科146属,其中植物界63种、动物界68种、菌物界71种。有194份标本(另8份无采集人信息)涉及77位采集人(或其团队),有67家机构的147人参与新种命名, 67种期刊的140条文献参与新种发表, 38个国内外机构保存了200种的标本,物种在30%的科、目、采集人、第1命名人、期刊和存放地各分配了约70%的数量,呈现类似"二八定律"所述的不平衡现象。1978至1997年是标本采集并命名的高峰期,标本采集率为55.45%、标本命名率为58.91%;植物标本的采集和命名早于动物和真菌。鼎湖山物种的橱柜时间(标本采集到命名间隔的时间)平均为7.56 a,且植物的长于动物和菌物。鼎湖山主模式标本生物的物种密度比邻近保护区的要高。     

Which specimens from a museum collection will yield DNA barcodes? A time series study of spiders in alcohol

We report initial results from an ongoing effort to build a library of DNA barcode sequences for Dutch spiders and investigate the utility of museum collections as a source of specimens for barcoding spiders. Source material for the library comes from a combination of specimens freshly collected in the field specifically for this project and museum specimens collected in the past. For the museum specimens, we focus on 31 species that have been frequently collected over the past several decades. A series of progressively older specimens representing these 31 species were selected for DNA barcoding. Based on the pattern of sequencing successes and failures, we find that smaller-bodied species expire before larger-bodied species as tissue sources for single-PCR standard DNA barcoding. Body size and age of oldest successful DNA barcode are significantly correlated after factoring out phylogenetic effects using independent contrasts analysis. We found some evidence that extracted DNA concentration is correlated with body size and inversely correlated with time since collection, but these relationships are neither strong nor consistent. DNA was extracted from all specimens using standard destructive techniques involving the removal and grinding of tissue. A subset of specimens was selected to evaluate nondestructive extraction. Nondestructive extractions significantly extended the DNA barcoding shelf life of museum specimens, especially small-bodied species, and yielded higher DNA concentrations compared to destructive extractions. All primary data are publically available through a Dryad archive and the Barcode of Life database.     

A comprehensive DNA barcode database for Central European beetles with a focus on Germany: adding more than 3500 identified species to BOLD

Beetles are the most diverse group of animals and are crucial for ecosystem functioning. In many countries, they are well established for environmental impact assessment, but even in the well‐studied Central European fauna, species identification can be very difficult. A comprehensive and taxonomically well‐curated DNA barcode library could remedy this deficit and could also link hundreds of years of traditional knowledge with next generation sequencing technology. However, such a beetle library is missing to date. This study provides the globally largest DNA barcode reference library for Coleoptera for 15 948 individuals belonging to 3514 well‐identified species (53% of the German fauna) with representatives from 97 of 103 families (94%). This study is the first comprehensive regional test of the efficiency of DNA barcoding for beetles with a focus on Germany. Sequences ≥500 bp were recovered from 63% of the specimens analysed (15 948 of 25 294) with short sequences from another 997 specimens. Whereas most specimens (92.2%) could be unambiguously assigned to a single known species by sequence diversity at CO1, 1089 specimens (6.8%) were assigned to more than one Barcode Index Number (BIN), creating 395 BINs which need further study to ascertain if they represent cryptic species, mitochondrial introgression, or simply regional variation in widespread species. We found 409 specimens (2.6%) that shared a BIN assignment with another species, most involving a pair of closely allied species as 43 BINs were involved. Most of these taxa were separated by barcodes although sequence divergences were low. Only 155 specimens (0.97%) show identical or overlapping clusters.     

中国科学院华南植物园标本馆维管植物标本数据统计分析

为充分利用植物标本信息,从植物标本与物种信息系统数据库中对中国科学院华南植物园标本馆维管植物标本数据进行了统计和分析。结果表明,该馆馆藏标本统计时已录入696298号、716469份腊叶标本,分别隶属于354科5813属48707种(包含亚种、变种及变型),其中包括蕨类植物59科253属2105种(包含亚种、变种及变型)、裸子植物11科64属 557种(包含亚种、变种及变型)、被子植物284科5496属46045种(包含亚种、变种及变型)。标本分别采自中国33个省、自治区、直辖市、特别行政区以及其它107个国家或地区(98个国家,9个地区或岛屿);国内标本采自广东省的最多,国外标本采自美国的最多。其中模式标本6351号,隶属于194科938属4373种。还对标本馆今后的采集工作提出了建议。这些数据为科学研究提供了宝贵的数据资源。     

Lessons from genome skimming of arthropod‐preserving ethanol

Field‐collected specimens of invertebrates are regularly killed and preserved in ethanol, prior to DNA extraction from the specimens, while the ethanol fraction is usually discarded. However, DNA may be released from the specimens into the ethanol, which can potentially be exploited to study species diversity in the sample without the need for DNA extraction from tissue. We used shallow shotgun sequencing of the total DNA to characterize the preservative ethanol from two pools of insects (from a freshwater habitat and terrestrial habitat) to evaluate the efficiency of DNA transfer from the specimens to the ethanol. In parallel, the specimens themselves were subjected to bulk DNA extraction and shotgun sequencing, followed by assembly of mitochondrial genomes for 39 of 40 species in the two pools. Shotgun sequencing from the ethanol fraction and read‐matching to the mitogenomes detected ~40% of the arthropod species in the ethanol, confirming the transfer of DNA whose quantity was correlated to the biomass of specimens. The comparison of diversity profiles of microbiota in specimen and ethanol samples showed that ‘closed association’ (internal tissue) bacterial species tend to be more abundant in DNA extracted from the specimens, while ‘open association’ symbionts were enriched in the preservative fluid. The vomiting reflex of many insects also ensures that gut content is released into the ethanol, which provides easy access to DNA from prey items. Shotgun sequencing of DNA from preservative ethanol provides novel opportunities for characterizing the functional or ecological components of an ecosystem and their trophic interactions.     

Age and growth in Peale's dolphin (Lagenorhynchus australis) in subantarctic waters off southern South America

The Peale's dolphin, Lagenorhynchus australis, is a mainly coastal and one of the most often seen species off southernmost South America from 33°S off Chile and 38°S off Argentina to south of Cape Horn. Although a common species, its IUCN status is “Data Defficient”, so any information on this species is relevant. Age, growth and physical maturity were examined in skeletons of 57 specimens of this species, mostly from the coasts of Tierra del Fuego, Argentina; this represents most of the known specimens of this species available in museum and private collections. Most specimens were by-catch in shore-set gillnets or had been harpooned for crab bait. Age was determined for 23 specimens, all from Tierra del Fuego. The maximum age was 13 years for a 199 cm female. The largest female in this survey was 210 cm, the largest male 205 cm in total length (12 years), but this probably does not represent the maximum length for this species. Only two animals, of 12 and 13 years, were physically mature. Animals reach asymptotic length (188 cm) with less than the 30% of the vertebral epiphyses fused. The animals in this study were mainly subadults, as has been found for other southern by-caught dolphins. Growth was studied for total length using 18 osteological characters with the Gompertz model. We propose that zygomatic width can be used to estimate total length for incomplete beach-cast specimens.     

重庆发现云南小狭口蛙及其系统发育关系

2021年5月在重庆武隆仙女山国家森林公园采集到3号小狭口蛙标本,经形态学及分子生物学鉴定为云南小狭口蛙（Glyphoglossus yunnanensis）,属重庆市两栖动物分布新记录种。本文给出了这3号标本的17项外形特征测量值,并描述了其生境特征。对10个不同地理种群的云南小狭口蛙种群相关16S rRNA基因片段构建最大似然树表明,重庆武隆标本与云南小狭口蛙产地标本序列聚在一起,且其间的遗传距离为0.6%,远远小于小狭口蛙属物种之间的遗传距离。重庆武隆的这一分布点,使得对云南小狭口蛙在我国分布的认知往东北延伸了超过350 km。云南小狭口蛙在重庆武隆的分布新记录能帮助理解该物种潜在分布区以及谱系地理。     

印尼新几内亚巴布亚省豕蛙属4新种(两栖纲:姬蛙科)(英文)

姬蛙科豕蛙属(Choerophryne)已知5种,几乎都分布于新几内亚岛东部地区,已有物种的描述多基于较少数量的标本,由于标本贫乏,迄今无 后续研究.本文针对采集于1998-2003年的50号标本,对该属进行了再研究,并描述了分布 于新几内亚岛的巴布亚西北地区鲜为人知的4新种.与同属已知物种比较,新种的有效性得 到来自形态、声谱以及分子数据的综合分析结果的支持.在新几内亚岛西部地区豕蛙属4新 种中,至少有3种的种群密度很高,从而极大增加了对该属的了解     

Identification of dry-pinned museum specimens of the sibling species Anopheles gambiae and An.arabiensis (Diptera: Culicidae) using non-radioactively labelled DNA probes

Non-radioactively labelled DNA probes were tested for their ability to identify dried museum specimens of Anopheles gambiae and its sibling species An.arabiensis . The specimens were the progeny of wild-caught females collected in 1991 from villages in western Kenya. Three years later, specimens whose identity was known to the second author were provided 'blind' to the first author for identification with oligonucleotide probes (SH 5 and SH 4 derived from pAngss and pAngsl, respectively) using a simplified squash-blotting protocol for non-radioactive probes. All specimens were successfully identified with whole-body squashes, and the results agreed with previous identifications of parents or siblings based on rDNA-polymerase chain reaction . The amounts of DNA released by squash-blotting were just sufficient for identification by those experienced in the technique, but not for squashes of heads or thoraces alone. An aim of the study was to determine whether squash-blot methods of identification might be useful for establishing the genetic identity of name-bearing type specimens of sibling species held in museum collections.     

中国几种剧毒鹅膏菌的ITS序列分析

测定了来自中国不同地区的9个剧毒鹅膏菌的核糖体内转录间隔区(ITS)核苷酸序列.以湖南鹅膏为外类群,用ITS序列构建了系统进化树.结果表明:两个不同产地的欧氏鹅膏存在一定的差异,但仍可聚为同一组;欧氏鹅膏与其它几种剧毒鹅膏的亲缘关系较远;两个根据形态特征鉴定为灰花纹鹅膏的标本可能是两个不同的种;欧氏鹅膏、致命鹅膏和黄盖鹅膏白色变种这3个产生白色子实体的种系统演化上不是同源的.