First insights into past biodiversity of giraffes based on mitochondrial sequences from museum specimens – Corrigendum

The advent of direct sequencing via the polymerase chain reaction (PCR) has opened up the possibility of molecular studies on museum specimens. Here we analyze genetic variation in populations over time by applying PCR to DNA extracted from museum specimens sampled from populations of one species over the last 78 years. Included in this study were 43 museum specimens of the Panamint kangaroo rat Dipodomys panamintinus from localities representing each of three geographically distinct subspecies. These specimens were originally collected and prepared as dried skins in 1911, 1917, or 1937. For each specimen, a 225-bp segment of the mitochondrial genome was sequenced. These mitochondrial DNA sequences were compared to those of 63 specimens collected at the same localities in 1988. The three subspecies were nearly completely distinct. Only 2 of the 106 individuals shared mitochondrial types between subspecies. For all three localities, the diversity levels were maintained between the two temporal samples. The concordance observed between the two temporally separate phylogenies supports the use of museum specimens for phylogenetic inference. This study demonstrates the accuracy and routine nature of the use of museum specimens in the analysis of mitochondrial sequence variation in natural populations and, importantly, that a temporal aspect can now be added to such studies.

Morphological, mitochondrial, and nuclear phylogenomic data were combined to address phylogenetic and species delimitation questions in cave-limited Cicurina spiders from central Texas. Special effort was focused on specimens and cave locations in the San Antonio region (Bexar County), home to four eyeless species listed as US Federally Endangered. Sequence capture experiments resulted in the recovery of ~200–400 homologous ultra-conserved element (UCE) nuclear loci across taxa, and nearly complete COI mitochondrial DNA sequences from the same set of individuals. Some of these nuclear and mitochondrial sequences were recovered from “standard” museum specimens without special preservation of DNA material, including museum specimens preserved in the 1990s. Multiple phylogenetic analyses of the UCE data agree in the recovery of two major lineages of eyeless Cicurina in Texas. These lineages also differ in mitochondrial clade membership, female genitalic morphology, degree of troglomorphy (as measured by relative leg length), and are mostly allopatric across much of Texas. Rare sympatry was confirmed in Bexar County, where members of the two major clades sometimes co-exist in the same karst feature. Both nuclear phylogenomic and mitochondrial data indicate the existence of undescribed species from the San Antonio region, although further sampling and collection of adult specimens is needed to explicitly test these hypotheses. Our data support the two following species synonymies (Cicurina venii Gertsch, 1992 = Cicurina madla Gertsch, 1992; Cicurina loftini Cokendolpher, 2004 = Cicurina vespera Gertsch, 1992), formally proposed here. Overall, our taxonomy-focused research has many important conservation implications, and again highlights the fundamental importance of robust taxonomy in conservation research.

Collecting mitochondrial genome sequences from museum collections has garnered increasing attention in recent years. Currently, several sequencing strategies have been proposed for this purpose, including direct DNA pooling, DNA labelling and mtDNA enrichment. It is important to understand the advantages and limitations of these methods before embarking on large‐scale sequencing efforts. However, there is currently no comprehensive comparison and evaluation of the performance of these sequencing methods. In this study, we evaluated the performance of four different strategies for sequencing mitogenomes by comparing reagent expense, experiment time, bioinformatics time, and the length and the base error rate of the obtained mitochondrial sequences. The four sequencing strategies mainly differ in: (1) whether to label the DNA of each specimen and (2) whether to perform mtDNA enrichment. To reduce experiment difficulty and expenses, we presented an ‘in‐situ’ DNA labelling protocol for cost‐effectively ligating barcode sequences to DNA samples and used PCR‐generated baits for mtDNA enrichment. The widely adopted ‘direct pooling’ sequencing strategy is the fastest but performs the worst among the four strategies. Of the 96 samples analysed, this approach yields averagely only 670 bp of mitochondrial sequences per sample. The most complicated sequencing strategy, which labels the DNA of each specimen and performs mtDNA enrichment, performs the best, producing mitogenome sequences of ~9.3 kb per sample. The sequencing accuracy is highly related to the sequencing depths of mtDNA but not the storage time of samples. We hope that our comparisons will assist researchers in selecting suitable strategies for large‐scale sequencing of mitogenomes from museum specimens, thereby advancing museomics.

Understanding past and present genetic diversity, in particular in endangered species such as the rhinoceroses, is of paramount importance for a series of aspects in natural history, evolutionary systematics and conservation. As it turned out from several recent studies even in eminent museum specimens the historical context including its provenance often remains unresolved. At the same time modern molecular genetic techniques make this material more and more available also for integrative studies. With probably less than fifty extant specimens, among the Asian rhinoceroses the Javan rhinoceros, Rhinoceros sondaicus, is one of the most critically endangered mammal species, rendering also each of its rare museum specimens of great significance. We here apply available DNA isolation and sequencing techniques to a horn of a specimen housed at the Zoological Museum in Hamburg with indication as to derive from the extinct conspecific Sumatra population. In comparison with already existing mitochondrial gene fragment sequence data of Asian rhino populations, we were able to verify the identification of this particular museum specimen as of the nearly equally rare Sumatran rhinoceros, Dicerorhinus sumatrensis, instead as of the extremely rare R. sondaicus.

BackgroundThe Indonesian island of Sulawesi has a complex geological history. It is composed of several landmasses that have arrived at a near modern configuration only in the past few million years. It is the largest island in the biodiversity hotspot of Wallacea—an area demarcated by the biogeographic breaks between Wallace’s and Lydekker’s lines. The mammal fauna of Sulawesi is transitional between Asian and Australian faunas. Sulawesi’s three genera of squirrels, all endemic (subfamily Nannosciurinae: Hyosciurus, Rubrisciurus and Prosciurillus), are of Asian origin and have evolved a variety of phenotypes that allow a range of ecological niche specializations. Here we present a molecular phylogeny of this radiation using data from museum specimens. High throughput sequencing technology was used to generate whole mitochondrial genomes and a panel of nuclear ultraconserved elements providing a large genome-wide dataset for inferring phylogenetic relationships.ResultsOur analysis confirmed monophyly of the Sulawesi taxa with deep divergences between the three endemic genera, which predate the amalgamation of the current island of Sulawesi. This suggests lineages may have evolved in allopatry after crossing Wallace’s line. Nuclear and mitochondrial analyses were largely congruent and well supported, except for the placement of Prosciurillus murinus. Mitochondrial analysis revealed paraphyly for Prosciurillus, with P. murinus between or outside of Hyosciurus and Rubrisciurus, separate from other species of Prosciurillus. A deep but monophyletic history for the four included species of Prosciurillus was recovered with the nuclear data.ConclusionsThe divergence of the Sulawesi squirrels from their closest relatives dated to ~9.7–12.5 million years ago (MYA), pushing back the age estimate of this ancient adaptive radiation prior to the formation of the current conformation of Sulawesi. Generic level diversification took place around 9.7 MYA, opening the possibility that the genera represent allopatric lineages that evolved in isolation in an ancient proto-Sulawesian archipelago. We propose that incongruence between phylogenies based on nuclear and mitochondrial sequences may have resulted from biogeographic discordance, when two allopatric lineages come into secondary contact, with complete replacement of the mitochondria in one species.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-016-0650-z) contains supplementary material, which is available to authorized users.

AimThe Atlantic sturgeon, Acipenser oxyrinchus, was long considered an American species, but recent discoveries based on analyses of archaeological remains proved its presence in the Baltic Sea and in France. This came as a surprise, as the European sturgeon, Acipenser sturio, was thought to be the only sturgeon species present on the European Atlantic coast. It was hypothesized that migrants from North America founded the Baltic population of Atlantic sturgeons around 1200 years ago.LocationThe sampling is composed of five sturgeon archaeological remains (5000–1800 yr bp) and 21 naturalized or ethanol‐preserved museum specimens from the 19th and 20th centuries, originating from the French Atlantic coast.MethodsAnalyses of ancient DNA were performed following the strict criteria of the field to avoid contamination and to ensure sequence authentication. A 210‐bp fragment of the control region (CR) was sequenced and three microsatellite loci were sized and sequenced.ResultsOut of a total of 26 samples, three archaeological remains and six museum specimens produced A. oxyrinchus mitochondrial or nuclear sequences; two of these museum samples showed signs of hybridization between A. sturio and A. oxyrinchus. All the other samples yielded only A. sturio DNA sequences, and a new CR haplotype was described in this species.Main conclusionsMolecular evidence of sympatry and natural hybridization between A. sturio and A. oxyrinchus on the French Atlantic coast – specifically the hybrid evidence of two museum specimens collected from the Seine River in 1823 and 1858 – challenge our understanding of the species' past relationships. In light of these findings, new hypotheses are presented to explain the history and geographical range of A. oxyrinchus in Europe.

Intensified exploration of sub-Saharan Africa during the 18th and 19th centuries led to many newly described giraffe subspecies. Several populations described at that time are now extinct, which is problematic for a full understanding of giraffe taxonomy. In this study, we provide mitochondrial sequences for 41 giraffes, including 19 museum specimens of high importance to resolve giraffe taxonomy, such as Zarafa from Sennar and two giraffes from Abyssinia (subspecies camelopardalis), three of the first southern individuals collected by Levaillant and Delalande (subspecies capensis), topotypes of the former subspecies congoensis and cottoni, and giraffes from an extinct population in Senegal. Our phylogeographic analysis shows that no representative of the nominate subspecies camelopardalis was included in previous molecular studies, as Zarafa and two other specimens assigned to this taxon are characterized by a divergent haplogroup, that the former subspecies congoensis and cottoni should be treated as synonyms of antiquorum, and that the subspecies angolensis and capensis should be synonymized with giraffa, whereas the subspecies wardi should be rehabilitated. In addition, we found evidence for the existence of a previously unknown subspecies from Senegal (newly described in this study), which is now extinct. Based on these results, we propose a new classification of giraffes recognizing three species and 10 subspecies. According to our molecular dating estimates, the divergence among these taxa has been promoted by Pleistocene climatic changes resulting in either savannah expansion or the development of hydrographical networks (Zambezi, Nile, Lake Chad, Lake Victoria).

Recent or incomplete speciation events can often lead to conflicting patterns of genetic and morphological variation, due to a combination of hybridization, incomplete lineage sorting and limited morphological divergence among genetically distinct species. This situation complicates the reconciliation of molecular systematic results with morphology-based taxonomy. For example, six-rayed sea stars of the genus Leptasterias have been the subject of considerable taxonomic controversy. Prior molecular studies established that Leptasterias subgenus Hexasterias consists of several genetically discrete clades, but extensive morphological variability has prevented the application of formal taxonomic nomenclature to these groups. In the present study, molecular and morphological approaches were combined to reconcile, where possible, the traditional taxonomy of Fisher (1930: US Nat Mus Bull 76:1–356) with recent molecular systematic results. Partial mitochondrial DNA control region sequences were collected from 29 Fisher-identified museum specimens representing seven nominal species of Leptasterias subgenus Hexasterias, to establish the relationship between traditional taxa and molecular lineages. Additional DNA sequences were obtained from 52 recently collected individuals representing the L.aleutica (Fisher, 1930)/L.camtschatica (Brandt, 1835) complex; these sea stars were collected on Kanaga Island, Alaska, in August 1997. Morphological data were collected for 227 museum specimens collected before 1930, and 309 specimens collected between 1988 and 1998 from locations in the eastern North Pacific. The analyses indicate that three nominal species within the Camtschatica section of subgenus Hexasterias, L.alaskensis (Verrill, 1914), L.hexactis (Stimpson, 1862), and L.leptodoma (Fisher, 1930), correspond to separate major clades of a molecular phylogeny, permitting the re-description of these taxa using molecular characters. Extensive sharing of mitochondrial DNA sequences by L.aleutica and L.camtschatica suggests recent genetic divergence of these two species and possible incomplete lineage sorting or frequent hybridization. The lack of diagnostic molecular differences between these two nominal species means that they are currently identifiable only by morphological criteria. Determining the status of three other nominal species, L.aequalis (Stimpson, 1862), L.asteira (Fisher, 1930) and L.pusilla (Fisher, 1930) will require additional molecular and morphological data.

A simple strategy for recovering ultraconserved elements, exons, and introns from low coverage shotgun sequencing of museum specimens: Placement of the partridge genus Tropicoperdix within the galliformes

Natural history museum collections represent a vast source of ancient and historical DNA samples from extinct taxa that can be utilized by high-throughput sequencing tools to reveal novel genetic and phylogenetic information about them. Here, we report on the successful sequencing of complete mitochondrial genome sequences (mitogenomes) from eleven extinct bird species, using de novo assembly of short sequences derived from toepad samples of degraded DNA from museum specimens. For two species (the Passenger Pigeon Ectopistes migratorius and the South Island Piopio Turnagra capensis), whole mitogenomes were already available from recent studies, whereas for five others (the Great Auk Pinguinis impennis, the Imperial Woodpecker Campehilus imperialis, the Huia Heteralocha acutirostris, the Kauai Oo Moho braccathus and the South Island Kokako Callaeas cinereus), there were partial mitochondrial sequences available for comparison. For all seven species, we found sequence similarities of >98%. For the remaining four species (the Kamao Myadestes myadestinus, the Paradise Parrot Psephotellus pulcherrimus, the Ou Psittirostra psittacea and the Lesser Akialoa Akialoa obscura), there was no sequence information available for comparison, so we conducted blast searches and phylogenetic analyses to determine their phylogenetic positions and identify their closest extant relatives. These mitogenomes will be valuable for future analyses of avian phylogenetics and illustrate the importance of museum collections as repositories for genomics resources.

Quintero, E., Ribas, C. C. & Cracraft, J. (2012). The Andean Hapalopsittaca parrots (Psittacidae, Aves): an example of montane‐tropical lowland vicariance. —Zoologica Scripta, 42, 28–43.We describe a phylogenetic and biogeographical pattern connecting high montane biotas to those of the lowlands, as exemplified by the exclusively montane parrot genus Hapalopsittaca and its lowland sister genus Pyrilia, both nested within Tavares et al.’s “amazons and allies” clade. As Hapalopsittaca is the only genus within the “amazons and allies” clade that is exclusively distributed in the Andes, the optimization leads to the inference that the ancestral distribution of the branch leading to Pyrilia + Hapalopsittaca was lowland. Museum specimens were examined to determine basal diagnosably distinct taxonomic units. Based on this analysis, mitochondrial sequences (cyt b and ND2 genes) from 17 individuals, mostly from toe pads, and representing all basal taxa within Hapalopsittaca, were obtained. A divergence‐dating analysis using both nuclear (RAG‐1) and mitochondrial (cyt b) genes was conducted to explore whether the uplift of the Andes coincides temporarily with the origin of this montane group, and thus might be causally linked to its origin. Molecular dating estimates the split between Hapalopsittaca and Pyrilia to have occurred between 6.6 and 8.0 Myr; thus, the timing of this highland/lowland disjunction is consistent with that of the final uplift of the Central Andes, supporting a hypothesis of vicariance due to Andean uplift. These results suggest that the taxonomic assembly of montane biotas may be, at least in part, explained by events of Earth history rather than by long‐distance dispersal and colonization. Diversification within Hapalopsittaca and the origin of current species are more recent in time and probably are related to Pleistocene climatic oscillations, as has been shown in other montane groups.

The black-veined white, Aporia crataegi (Lepidoptera: Pieridae), is distributed from northwest Africa throughout most of Europe and temperate Asia to far eastern Russia and Japan and is presumed to be extinct in South Korea, with only a few dried specimens remaining. A common conservation practice for such near-extinct species is the launch of an introduction program, but prior genetic analysis of donor and donee populations may be essential for the success of such a program. In the present study, we investigated the genetic diversity and the relationships of A. crataegi populations from South Korea and nearby Asian countries, using 11 newly developed A. crataegi-specific microsatellite markers and mitochondrial DNA (mtDNA) sequences from two genes (a total of 1317 bp from COI and CytB). mtDNA-based analyses, using a total of 115 individuals collected from China, Russia, Mongolia, and Japan and museum specimens from South Korea consistently showed long-held, close genetic relationships between A. crataegi populations from Mongolia, China, and Russia and those from South Korea, with the highest diversity detected in the Mongolian population. Principal coordinate analyses and STRUCTURE analyses, based on microsatellite markers, also suggested that A. crataegi from South Korea had a similar gene pool to populations from China, Mongolia, and Russia, with a similar degree of genetic diversity among these populations. Taken together, the mtDNA and microsatellite data suggest that the northern populations, particularly those from Mongolia, can be considered the most suitable population as an immediate donor for the near-extinct South Korean population of A. crataegi.

We investigated the scolytine fauna of the West Himalayan region of India. The present study combines field samples, museum specimens, and published data to catalog the scolytine diversity. We recognize 57 scolytine species from 15 tribes and 25 genera. Five new species Cryphalus himalayensis sp. nov., Dryocoetes asperatus sp. nov., Ernoporus squamosus sp. nov., Scolytoplatypus denticauda sp. nov., and Hypothenemus ficivorus sp. nov. are described from the region. Previously reported and inadequately characterized species are imaged and redescribed. Two new Indian records, seven new regional records, and fifteen new host records are reported. Keys to genera and photographs of each species are provided to facilitate identification. Mitochondrial COX-1 sequences were generated from some Himalayan species and sequences from GenBank were utilized for comparison with other related species. The current monographic study includes an overview of each species' taxonomic diagnosis, range, and host plants. Furthermore, an updated list of 165 scolytine species, distributed in the Himalaya but excluding its eastern part of Arunachal Pradesh, is also provided.

The Lesser Antilles is a biodiversity hot spot but unfortunately human disturbance has taken its toll, causing dramatic population declines and even extinction of numerous endemic species. Nevertheless, today the rediscovery of previously thought extinct species is not uncommon. Often, old museum specimens and their original descriptions are the only information available for such species. The application of molecular phylogenetic relationships to extant species can help to elucidate pivotal information on their ecology and conservation. Erythrolamprus cursor is possibly an extinct colubrid racer from Martinique, currently classified as critically endangered on the International Union for Conservation of Nature Red List. Mitochondrial deoxyribonucleic acid sequences were obtained from four E. cursor specimens from the Muséum national d'Histoire naturelle (Paris) collections. All sequences recovered the same haplotype and the level of divergence between E. cursor and E. juliae, from the nearby island of Dominica, was lower than between other intraspecific distances within other Erythrolamprus. Furthermore, phylogenetic analyses confirm that these two species are sister taxa and share most recent common ancestry. We discuss that published ecological data available for the sister species (E. juliae) may help to elucidate information on this species' natural history, ultimately having important implications for a future conservation management program if E. cursor is to be found. We emphasize the urgent need to conduct an exhaustive survey on the supposed last population of E. cursor at Diamond Rock to establish the survival of this species there, to understand how it may have adapted to such an ecosystem, especially in sympatry of several introduced rodent species.

The Madrean Archipelago in southern Arizona and southwestern New Mexico consists of 22 mountain ranges south of the Mogollon Rim. Herpetofaunal lists for these ranges and a segment of the rim were constructed based on museum specimens from Madrean evergreen woodland, petran montane conifer forest, and interior chaparral. Few or no species have been recorded from these communities in the Sierrita, Little Dragoon, and Big Hatchet mountains, emphasizing the need for additional sampling in the archipelago. A total of 83 species are found in the remaining 20 ranges, with lizards (28) and snakes (35) predominating. No two mountain ranges had the same herpetofauna. Species richness varies from 15 to 44 (mean 28.1). Phenetic analyses of herpetofaunal similarity among the ranges identify three groups: a northern group with eight ranges from the Rincon Mountains to the Pinaleño Mountains to the Sierra Ancha; a southwestern group consisting of the Baboquivari, Santa Rita, Pajarito, and Patagonia mountains; and a southeastern group with seven ranges from the Huachuca and Whetstone mountains to the Animas Mountains; the Mogollon Rim segment is placed as the first “branch” of the phenogram. The analyses place the Patagonia Mountains in the SW group and the Huachuca Mountains in the SE group, although the two are connected by woodland. The Madrean line separating the northern group from the two southern groups approximates the southern limit of interior chaparral. The ranges of the southwestern group are in contact with semitropical Sonoran desert scrub at low elevations, whereas Chihuahuan desert scrub and semidesert grassland surround the southeastern ranges. With few exceptions, published studies of phylogeography within species suggest that divergence among montane populations in the archipelago does not predate the Pleistocene. Phylogeographic analyses using nuclear and mitochondrial gene sequences and including all ranges of the Madrean Archipelago inhabited by a species are needed to determine the extent to which shared historic biogeographic events may underlie the groups of mountain ranges identified on the basis of herpetofaunal similarities in this study.