No consistent effects of humans on animal genetic diversity worldwide

Human impacts on genetic diversity are poorly understood yet critical to biodiversity conservation. We used 175 247 COI sequences collected between 1980 and 2016 to assess the global effects of land use and human density on the intraspecific genetic diversity of 17 082 species of birds, fishes, insects and mammals. Human impacts on mtDNA diversity were taxon and scale‐dependent, and were generally weak or non‐significant. Spatial analyses identified weak latitudinal diversity gradients as well as negative effects of human density on insect diversity, and negative effects of intensive land use on fish diversity. The observed effects were predominantly associated with species turnover. Time series analyses found nearly an equal number of positive and negative temporal trends in diversity, resulting in no net monotonic trend in diversity over this time period. Our analyses reveal critical data and theory gaps and call for increased efforts to monitor global genetic diversity.     

Performance comparison of genetic markers for high‐throughput sequencing‐based biodiversity assessment in complex communities

Metabarcode surveys of DNA extracted from environmental samples are increasingly popular for biodiversity assessment in natural communities. Such surveys rely heavily on robust genetic markers. Therefore, analysis of PCR efficiency and subsequent biodiversity estimation for different types of genetic markers and their corresponding primers is important. Here, we test the PCR efficiency and biodiversity recovery potential of three commonly used genetic markers – nuclear small subunit ribosomal DNA (18S), mitochondrial cytochrome c oxidase subunit I (COI) and 16S ribosomal RNA (mt16S) – using 454 pyrosequencing of a zooplankton community collected from Hamilton Harbour, Ontario. We found that biodiversity detection power and PCR efficiency varied widely among these markers. All tested primers for COI failed to provide high‐quality PCR products for pyrosequencing, but newly designed primers for 18S and 16S passed all tests. Furthermore, multiple analyses based on large‐scale pyrosequencing (i.e. 1/2 PicoTiter plate for each marker) showed that primers for 18S recover more (38 orders) groups than 16S (10 orders) across all taxa, and four vs. two orders and nine vs. six families for Crustacea. Our results showed that 18S, using newly designed primers, is an efficient and powerful tool for profiling biodiversity in largely unexplored communities, especially when amplification difficulties exist for mitochondrial markers such as COI. Universal primers for higher resolution markers such as COI are still needed to address the possible low resolution of 18S for species‐level identification.     

Incomplete estimates of genetic diversity within species: Implications for DNA barcoding

DNA barcoding has greatly accelerated the pace of specimen identification to the species level, as well as species delineation. Whereas the application of DNA barcoding to the matching of unknown specimens to known species is straightforward, its use for species delimitation is more controversial, as species discovery hinges critically on present levels of haplotype diversity, as well as patterning of standing genetic variation that exists within and between species. Typical sample sizes for molecular biodiversity assessment using DNA barcodes range from 5 to 10 individuals per species. However, required levels that are necessary to fully gauge haplotype variation at the species level are presumed to be strongly taxon‐specific. Importantly, little attention has been paid to determining appropriate specimen sample sizes that are necessary to reveal the majority of intraspecific haplotype variation within any one species. In this paper, we present a brief outline of the current literature and methods on intraspecific sample size estimation for the assessment of COI DNA barcode haplotype sampling completeness. The importance of adequate sample sizes for studies of molecular biodiversity is stressed, with application to a variety of metazoan taxa, through reviewing foundational statistical and population genetic models, with specific application to ray‐finned fishes (Chordata: Actinopterygii). Finally, promising avenues for further research in this area are highlighted.     

Recent multiple introductions of the gall‐forming aphid Pemphigus bursarius into Japanese islands

Recently, the number of collection records of Pemphigus galls from Populus nigra has been increasing in Japan. To identify the galls on P. nigra, mitochondrial COI sequences were analyzed from galling aphid samples collected on P. nigra in Tokyo and Hokkaido. From the BLAST search and neighbor‐joining (NJ) analysis, the aphid samples were identified as Pemphigus bursarius, which has not been recorded from Japan. Two samples from Tokyo and Hokkaido showed a genetic difference of 0.30%. This result suggests that different strains of P. bursarius might have been introduced into the Japanese islands at least twice.     

The odd couple: contrasting phylogeographic patterns in two sympatric sibling species of woodlouse‐hunter spiders in the Canary Islands

Comparative phylogeography seeks for commonalities in the spatial demographic history of sympatric organisms to characterize the mechanisms that shaped such patterns. The unveiling of incongruent phylogeographic patterns in co‐occurring species, on the other hand, may hint to overlooked differences in their life histories or microhabitat preferences. The woodlouse‐hunter spiders of the genus Dysdera have undergone a major diversification on the Canary Islands. The species pair Dysdera alegranzaensis and Dysdera nesiotes are endemic to the island of Lanzarote and nearby islets, where they co‐occur at most of their known localities. The two species stand in sharp contrast to other sympatric endemic Dysdera in showing no evidence of somatic (non‐genitalic) differentiation. Phylogenetic and population genetic analyses of mitochondrial cox1 sequences from an exhaustive sample of D. alegranzaensis and D. nesiotes specimens, and additional mitochondrial (16S, L1, nad1) and nuclear genes (28S, H3) were analysed to reveal their phylogeographic patterns and clarify their phylogenetic relationships. Relaxed molecular clock models using five calibration points were further used to estimate divergence times between species and populations. Striking differences in phylogeography and population structure between the two species were observed. Dysdera nesiotes displayed a metapopulation‐like structure, while D. alegranzaensis was characterized by a weaker geographical structure but greater genetic divergences among its main haplotype lineages, suggesting more complex population dynamics. Our study confirms that co‐distributed sibling species may exhibit contrasting phylogeographic patterns in the absence of somatic differentiation. Further ecological studies, however, will be necessary to clarify whether the contrasting phylogeographies may hint at an overlooked niche partitioning between the two species. In addition, further comparisons with available phylogeographic data of other eastern Canarian Dysdera endemics confirm the key role of lava flows in structuring local populations in oceanic islands and identify localities that acted as refugia during volcanic eruptions.     

Effects of habitat fragmentation on genetic diversity and gene flow among the Homidia socia (Collembola: Entomobryidae) populations in the Thousand Island Lake

In the present study, partial sequences of the mitochondrial cytochrome oxidase subunit I (COI) gene of 22 island populations of the springtail Homidia socia in the Thousand Island Lake were sequenced. Across all sequences, 37 haplotypes were identified for the 510‐bp mitochondrial (mt) DNA COI gene. Haplotype 2 was the most common, and was distributed in the most of the 22 island populations. Haplotype diversity ranged from 0.065 to 0.733, and the total genetic diversity was 0.56216. The genetic characteristics of the 22 island populations were analyzed using the fixation index and gene flow, with values of 0.00043–0.94900 and 0.02703–703.72540, respectively. Comparison between (island area and isolations) with population genetic diversity revealed that there were no significant correlations between them, except for a significant correlation between the number of haplotypes and island area. Mantel tests showed that there was no significant correlation between geographic distance and genetic distance among various groups. All the results indicated that there were no obvious relationships between island characteristics and the genetic diversity of the springtails. We consider that the low dispersal capacity of springtails and the island patches surrounded by water in the Thousand Island Lake are the major factors affecting the genetic diversity of H. socia.     

Different phylogeographic patterns in two Japanese Silpha species (Coleoptera: Silphidae) affected by climatic gradients and topography

To reveal differences in phylogeographic patterns of flightless insect species occurring in different regions of Japan, we studied the phylogeography and demographic history of Silpha beetles occurring in cool-temperate habitats of two major islands, Honshu and Hokkaido, using sequences of the mitochondrial cytochrome oxidase subunit I (COI) gene. Honshu has a more mountainous topography, and cool-temperate habitats occur discontinuously, whereas Hokkaido, located to the north of Honshu, has more continuous cool-temperate habitats. A species endemic to Honshu, S. longicornis occurs on Honshu, whereas S. perforata occurs on Hokkaido and the East Asian continent. Our results indicate that the ancestors of S. longicornis colonized Honshu via a south-west route c . 0.7 Mya and the species has highly divergent populations in isolated mountainous areas of Honshu, whereas S. perforata colonized Hokkaido via a northern route less than 90 000 years ago and has less divergent geographic populations. During the last glacial period, S. perforata was probably restricted to refugia in southern Hokkaido and later expanded into northern Hokkaido, whereas S. longicornis populations existed in many isolated refugia, probably because of the complex topography of Honshu. Thus, our study demonstrates that, even between closely related species, interactions among biology, latitudinal climatic gradients and topography can produce different phylogeographic patterns.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 98 , 452–467.     

Mitochondrial DNA analysis of the introduced fall webworm, showing its shift in life cycle in Japan

The fall webworm, Hyphantria cunea (Drury) (Lepidoptera: Arctiidae), was introduced from North America into Japan in 1945. For the first three decades after its introduction, its life cycle was bivoltine. Thereafter, its life cycle shifted to trivoltine in south‐western areas of Japan. Two hypotheses have been proposed for the process that led to the shift in voltinism: one based on a single and the other on multiple independent colonizations. To test these hypotheses, mitochondrial (mt)DNA sequences were analyzed in the black‐headed type of 14 Japanese, one Korean and two North American populations of H. cunea. In addition, the same regions of mtDNA were compared with the red‐headed type of two North American populations. In the black‐headed type, mtDNA sequences were the same in all Japanese populations and in the Korean population, but sequences of the North American populations differed from each other and from those of the other populations. These results suggest that the process of the shift in voltinism occurred originally in Japan, and that the Japanese and the Korean population of H. cunea originated from a relatively small area in North America.     

基于线粒体细胞色素c氧化酶亚基I基因序列的帘蛤科贝类分子系统发育研究

对21种帘蛤科贝类线粒体细胞色素c氧化酶亚基Ⅰ(cytochrome c oxidase subunit I,COI)基因核苷酸序列进行了分析,以探讨这一序列在种质鉴定、分子系统发生研究中的应用价值。测序结果表明,所有物种扩增片段长度均为707 bp(含引物),序列A+T含量(62.4%—67.8%)明显高于G+C含量。物种间共有变异位点379个,其中简约信息位点334个;此区段共编码235个氨基酸,种间共有氨基酸变异位点100个。以COI基因片段序列为标记,用中国蛤蜊(Mactra chinensis)作外群,构建了35种帘蛤科贝类(其中14种贝类COI序列从GenBank下载)的系统发生树,结合拓扑结构分析和序列比对分析,结果表明:支持将短文蛤(Meretrix petechinalis)和丽文蛤(M.lusoria)订为文蛤(M.meretrix)的同物异名的观点,建议将丽文蛤和短文蛤订为文蛤的地理亚种;支持将薄片镜蛤(Dosinia corrugata)和D.angulosa订为2个独立种的观点;认为将波纹巴非蛤(Paphia undulata)和织锦巴非蛤(P.textile)订为2个独立种是合适的。COI基因序列含有丰富的遗传信息,适合作为帘蛤科贝类种群遗传结构和系统发生研究的分子标记。     

PCR gut analysis reveals that Tenuiphantes tenuis (Araneae: Linyphiidae) is a potentially significant predator of Argentine stem weevil,Listronotus bonariensis (Coleoptera: Curculionidae), in New Zealand pastures

Polymerase chain reaction (PCR) gut analysis was conducted on specimens of the introduced spider Tenuiphantes tenuis collected from dairy pasture in Canterbury, New Zealand. PCR primers were specifically designed to amplify a fragment of the mitochondrial gene cytochrome c oxidase subunit 1 (COI) from Listronotus bonariensis and revealed that this major pasture pest species is consumed in the field by T. tenuis. The field predation rate of L. bonariensis by T. tenuis was estimated from our PCR results together with published data on the degradation of DNA and the density of T. tenuis in Canterbury pastures. We found that T. tenuis is a potentially significant predator of L. bonariensis in New Zealand pastures.     

Cryptic diversity and species delimitation in the Xiphinema americanum‐group complex (Nematoda: Longidoridae) as inferred from morphometrics and molecular markers

The Xiphinema americanum‐group constitutes a complex of about 55 species of polyphagous plant‐ectoparasitic nematodes with a worldwide distribution. This group of plant‐parasitic nematodes is one of the most difficult dagger nematode species complexes for diagnosis because the morphology is very conservative and morphometric characters often overlap. We conducted nematode surveys in cultivated and wild olives in southern Spain from 2012 to 2014, from which we identified 16 nematode populations of the X. americanum‐group, five of which were tentatively identified as belonging to three new species and are described herein as X iphinema plesiopachtaicum sp. nov., X iphinema vallense sp. nov. , and X iphinema astaregiense sp. nov. , and 11 populations belonging to nine known species: Xiphinema brevisicum, Xiphinema duriense, Xiphinema incertum, Xiphinema luci, Xiphinema madeirense, Xiphinema opisthohysterum, Xiphinema pachtaicum, Xiphinema parapachydermum, and Xiphinema rivesi. A phenetic study based on multivariate factor analyses was developed to compare some of these related species by using morphometric features. In the factor analysis the first four factors accounted for 73.1% of the total variance of the selected characters, identifying body length, body length/maximum body width (a), body length/pharyngeal length (b), body length/tail length (c), and tail length/body width at anus (c′) ratios, distance from anterior end to vulva as percentage of body length (V), stylet length, oral aperture‐guiding ring distance, and lip region width as key morphometric characters to differentiate a restricted set of species within the X. pachtaicum‐subgroup that includes X. plesiopachtaicum sp. nov. and X. vallense sp. nov. Multivariate analysis of variance using these specific characters allowed to differentiate species in the X. pachtaicum complex or groups of them using morphometric characters (body length, a, b, c, c′, V, stylet length, lip region width, oral aperture‐guiding ring distance, female tail length, and hyaline region length). Phylogenetic analyses based on nuclear ribosomal DNA genes [D2‐D3 expansion segments of large ribosomal subunit 28S, and internal transcribed spacer 1 (ITS1)] and the protein‐coding mitochondrial gene, cytochrome c oxidase subunit 1 (coxI) were congruent, showing two main clades separating most of the species of X. americanum‐subgroup ‘sensu stricto’ from the X. pachtaicum‐subgroup. Agreement between phylogenetic trees and some morphological characters (viz. total stylet length, vulva position, and a ratio) were tested by reconstruction of their histories on rRNA‐based trees using parsimony and Bayesian approaches. Thus, integrative taxonomy, based on a combination of multivariate morphological and molecular analyses constitutes a new insight into the identification of X. americanum‐group species. © 2015 The Linnean Society of London     

Identification of forensically important blow fly species (Diptera: Calliphoridae) in China by mitochondrial cytochrome oxidase I gene differentiation

Abstract Unambiguous and rapid sarcosaphagous insect species identification is an essential requirement for forensic investigations. Although some insect species are difficult to classify morphologically, they can be effectively identified using molecular methods based on similarity with abundant authenticated reference DNA sequences in local databases. However, local databases are still relatively incomplete in China because of the large land area with distinct regional conditions. In this study, 75 forensically important blow flies were collected from 23 locations in 16 Chinese provinces, and a 278‐bp segment of the cytochrome oxidase subunit I gene of all specimens was successfully sequenced. Phylogenetic analysis of the sequenced segments showed that all Calliphorid specimens were properly assigned into nine species with relatively strong supporting values, thus indicating that the 278‐bp cytochrome oxidase subunit one region is suitable for identification of Calliphorid species. The clear difference between intraspecific threshold and interspecific divergence confirmed the potential of this region for Calliphorid species identification, especially for distinguishing between morphologically similar species. Intraspecific geographic variations were observed in Lucilia sericata (Meigen, 1826) and Lucilia caesar (Linnaeus, 1758).     

The origin of the Tibetan Mastiff and species identification of Canis based on mitochondrial cytochrome c oxidase subunit I (COI) gene and COI barcoding

DNA barcoding is an effective technique to identify species and analyze phylogenesis and evolution. However, research on and application of DNA barcoding in Canis have not been carried out. In this study, we analyzed two species of Canis, Canis lupus (n = 115) and Canis latrans (n = 4), using the cytochrome c oxidase subunit I (COI) gene (1545 bp) and COI barcoding (648 bp DNA sequence of the COI gene). The results showed that the COI gene, as the moderate variant sequence, applied to the analysis of the phylogenesis of Canis members, and COI barcoding applied to species identification of Canis members. Phylogenetic trees and networks showed that domestic dogs had four maternal origins (A to D) and that the Tibetan Mastiff originated from Clade A; this result supports the theory of an East Asian origin of domestic dogs. Clustering analysis and networking revealed the presence of a closer relative between the Tibetan Mastiff and the Old English sheepdog, Newfoundland, Rottweiler and Saint Bernard, which confirms that many well-known large breed dogs in the world, such as the Old English sheepdog, may have the same blood lineage as that of the Tibetan Mastiff.     

Incongruence of morphology and genetic markers in Hyles tithymali (Lepidoptera: Sphingidae) from the Canary Islands

Hyles t. tithymali on the Canary Islands has been observed to occur in two larval morphotypes, connected by intermediate forms along a geographical cline from east to west. In this study, it was tested whether this distribution of phenotypes reflects a genealogical division of the population. mtDNA sequence data (COI + II, tRNA-leu) and genomic fingerprints from intersimple sequence repeat (ISSR)-PCR data were used. The sequence data had low variation (max. 0.4%), and phylogenetic analyses did not reveal groups that correlated with the morphotype. The samples did not group according to their island of origin and the most common haplotype was shared among all islands. Although nine haplotypes occurred only on the westernmost islands, the data showed little phylogeographical structure. The population of H. t. tithymali appears to reflect a comparatively rapid and recent colonization event of the Canary Islands. The ISSR-PCR data were very variable and did not reveal patterns corresponding to morphological variation or geographical distribution. Although the two morphs observed may represent the first stage of differentiation between two lineages, the recent origin of H. t. tithymali provided insufficient time for complete lineage sorting of ancestral polymorphism. Hence, the population of Hyles t. tithymali on the Canary Islands appears genetically more homogeneous than that was expected from the phenotypic distribution of the two morphotypes in the population.     

DNA barcoding reveals extraordinary cryptic diversity in an amphipod genus: implications for desert spring conservation

DNA barcoding has revealed unrecognized species in several animal groups. In this study we have employed DNA barcoding to examine Hyalella, a taxonomically difficult genus of amphipod crustaceans, from sites in the southern Great Basin of California and Nevada, USA. We assessed the extent of species diversity using a species screening threshold (SST) set at 10 times the average intrapopulation cytochrome c oxidase subunit I (COI) haplotype divergence. Despite the fact that this threshold approach is more conservative in delineating provisional species than the phylogenetic species concept, our analyses revealed extraordinary levels of cryptic diversity and endemism. The SST discriminated two provisional species within Hyalella sandra, and 33 provisional species within Hyalella azteca. COI nucleotide divergences among these provisional species ranged from 4.4% to 29.9%. These results have important implications for the conservation of life in desert springs - habitats that are threatened as a result of groundwater over-exploitation.     

环境DNA技术在淡水底栖大型无脊椎动物多样性监测中的应用

环境DNA (eDNA)是指生物有机体在环境中(例如土壤、沉积物或水体)遗留下的DNA片段。eDNA技术是指从环境中提取DNA片段进行测序以及数据分析来反映环境中的物种或群落信息。与传统方法相比, eDNA技术具有高灵敏度、省时省力、无损伤等优点。目前, eDNA技术已成为一种新的水生生物监测方法, 主要应用于水生生物的多样性研究、濒危和稀有动物的物种状态及外来入侵动物扩散动态的监测等。本文从eDNA技术在水生生物多样性监测应用领域的发展历程、eDNA技术的操作流程以及其在监测淡水底栖大型无脊椎动物方面的应用进展、技术优势和局限性五个方面进行了综述。最后, 本文对eDNA技术在淡水底栖大型无脊椎动物多样性监测应用的发展趋势和前景作出展望。     

High diversity and local endemism in Aotearoa New Zealand's groundwater crustacean fauna

We used DNA barcoding to assess the diversity and distribution of New Zealand''s groundwater amphipods and isopods (Crustacea) and to determine whether biodiversity and endemism within tectonically active New Zealand are similar to those of more tectonically stable continents. Sixty‐five wells were sampled in seven aquifers across four regions within the North and South islands of New Zealand, and resident invertebrates were morphologically identified and then assessed using sequencing of the mitochondrial DNA cytochrome c oxidase subunit one (COI) gene. Invertebrates were found in 54 wells. Of the 228 individual amphipods and isopods found in 36 of the wells, 154 individuals were successfully sequenced for COI (68% success rate) from 25 wells, with at least one well in each aquifer containing sequenced individuals. Of the 45 putative species identified using Barcode Index Numbers (BINs), 30 BINs (78% of all taxa and 83% of amphipods) were previously unrecorded. Substantial morphologically cryptic, species‐level diversity was revealed, particularly within the amphipod Family Paraleptamphopidae. Similarly, one isopod taxon morphologically identified as Cruregens fontanus was assigned to five well‐separated BINs based on COI sequences. Endemism appeared high, with all taxa regionally endemic; 87% of species were restricted to one aquifer and more than 50% restricted to one well. Non‐saturated species accumulation curves indicated that, while additional sampling may increase the range of some currently identified taxa, additional range‐restricted taxa are also likely to be discovered. Patterns of diversity and short‐range endemism were similar to those found elsewhere, including locations which are more tectonically stable. The predominance of local endemism within New Zealand''s groundwater fauna suggests that land‐use activities and groundwater extraction require careful evaluation to minimize threats to groundwater biodiversity.     

Polymorphism observed in mitochondrial genes of red flour beetle,Tribolium castaneum (Coleoptera: Tenebrionidae) of different origin in laboratory cultures

Tribolium castaneum, a pest of stored grain, has a uniform morphology, preventing the visual identification of strains from different areas. Polymorphisms in the nucleotide sequences of the mitochondrial genes of this species were examined, and combined into seven haplotypes among the test insect specimens originating from Japan, Thailand, and Canada. These results suggested the potential for geographical differentiation.     

Amplified fragment length polymorphism confirms reciprocal monophyly in Chrysomya putoria and Chrysomya chloropyga: a correction of reported shared mtDNA haplotypes

Reinvestigation of mitochondrial haplotypes previously reported to be shared between the Afrotropical blowflies Chrysomya putoria Weidemann and Chrysomya chloropyga Weidemann (Diptera: Calliphoridae) revealed an error resulting from the misidentification of specimens. Preliminary amplified fragment length polymorphism (AFLP) analysis of the original and additional individuals again failed to find reciprocal monophyly, leading to a re-examination of the specimens for diagnostic male genitalic characters that were first described following the earlier study. Four of the original study specimens were found to have been misidentified, and definitive analysis of both mtDNA and AFLP genotypes using phylogenetic analysis and genetic assignment showed that each species was indeed reciprocally monophyletic. In addition to correcting the earlier error, this study illustrates how AFLP analysis can be used for efficient and effective specimen identification through both phylogenetic analysis and genetic assignment, and suggests that the latter method has special advantages for identification when no conspecific specimens are represented in the reference database.