Mitochondrial Clades Research Articles

A Pleistocene Biogeography in Miniature: The Small‐Scale Evolutionary History of Podarcis lusitanicus (Squamata, Lacertidae)

ABSTRACTAimRecent literature is unveiling more complex responses of biodiversity to Pleistocenic climatic oscillations than those derived from initial models based solely on southern glacial macrorefugia. Here, we zoom in to delve into the genetic diversity and population structure of Podarcis lusitanicus, a lizard species native to the north‐western Iberian Peninsula.LocationNorth‐western Iberian Peninsula.TaxonPodarcis lusitanicus.MethodsUtilising a comprehensive multilocus approach, we explored the species' phylogeography and demographic history at a very fine spatial scale and timeframe. Concurrently, we evaluated both current and historical habitat suitability for the species, linking it to the observed demographic trends. We examined DNA sequences from two mitochondrial (cytb and ND4) and three nuclear genes (ACM4, MC1R and PDC) in 230 specimens collected from 21 populations spanning most of the species' distribution range.ResultsOur findings corroborate cytonuclear discordances in Podarcis, with nuclear genes failing to identify the mitochondrial clades in P. lusitanicus. Despite its separation from other Podarcis species approximately 4 million years ago, its diversification occurred much more recently, around 0.4 million years ago, resulting in three monophyletic lineages that are geographically coherent. This pattern suggests that some lineages of P. lusitanicus might have gone extinct after the split from the common ancestor. Pleistocene climatic oscillations profoundly influenced the demographic history of the species. During warmer periods, such as the Last Interglacial, populations expanded and came into secondary contact, whereas during the Last Glacial Maximum, populations contracted and became isolated in multiple micro‐refugia. The subsequent Mid‐Holocene warming facilitated population recovery, range expansion and lineage admixture.Main ConclusionsThis study underscores the intricate interplay between isolation and dispersal at a microscale, driven by Pleistocene climatic fluctuations, which have profoundly shaped the genetic structure and diversity of Iberian terrestrial biodiversity and may remain overlooked by biogeographic studies at coarser scales.

Different life strategies of closely related louse species in sympatry: specialist and “generalist“ lineages of Polyplax serrata

The origin and significance of host specificity are intriguing questions in parasitology. In the case of single-host versus multiple-host parasites, this topic integrates with the concept of the specialist/generalist trade-off. We use the model of sucking lice Polyplax serrata and rodent hosts Apodemus, to address these concepts. Polyplax serrata was shown to form a complex genetic structure, with a strictly specific S lineage living on Apodemus flavicollis, and a less specific N lineage on A. flavicollis and Apodemus sylvaticus. Moreover, the S lineage formed two mitochondrial clades with geographically exclusive distributions and a narrow hybrid zone, providing an opportunity to test the hypothesis that hybrids suffer a decrease in fitness. We sampled 451 individual lice from two host species at 103 localities. We used prevalences and intensities as proxies of fitness, which the parasites realize on their host. The S lineage, strictly specific to Apodemus flavicollis, reached significantly higher prevalences and intensities on its host compared with the N lineage. Conversely, the N lineage occurred with high prevalence and intensity on A. sylvaticus but tended to use also A. flavicollis when the louse populations became too dense. We discuss possible mechanisms behind this difference (particularly interspecific competition as a typical phenomenon in the specialist/generalist systems). We conclude that a parasite’s “choice”, not accessibility of the host or interspecific competition, is the main factor affecting the louse prevalences. We suggest that historical differences in geographic distribution of both lice and mice may provide a possible explanation for the observed life strategy differences. In contrast to the convincing picture in S and N lineage prevalences, we did not detect an expected drop in fitness in hybrids. We consider instability of the hybrid zone, or decline in abundance of the respective hosts, as possible explanations for this result.

The limits of the metapopulation: Lineage fragmentation in a widespread terrestrial salamander (Plethodon cinereus)

Abstract In vicariant species formation, divergence results primarily from periods of allopatry and restricted gene flow. Widespread species harboring differentiated, geographically distinct sublineages offer a window into what may be a common mode of species formation, whereby a species originates, spreads across the landscape, then fragments into multiple units. However, incipient lineages usually lack reproductive barriers that prevent their fusion upon secondary contact, blurring the boundaries between a single, large metapopulation-level lineage and multiple independent species. Here we explore this model of species formation in the Eastern Red-backed Salamander (Plethodon cinereus), a widespread terrestrial vertebrate with at least six divergent mitochondrial clades throughout its range. Using anchored hybrid enrichment data, we applied phylogenomic and population genomic approaches to investigate patterns of divergence, gene flow, and secondary contact. Genomic data broadly match most mitochondrial groups but reveal mitochondrial introgression and extensive admixture at several contact zones. While species delimitation analyses in BPP supported five lineages of P. cinereus, genealogical divergence indices (gdi) were highly sensitive to the inclusion of admixed samples and the geographic representation of candidate species, with increasing support for multiple species when removing admixed samples or limiting sampling to a single locality per group. An analysis of morphometric data revealed differences in body size and limb proportions among groups, with a reduction of forelimb length among warmer and drier localities consistent with increased fossoriality. We conclude that P. cinereus is a single species, but one with highly structured component lineages of various degrees of independence.

Mitochondrial DNA evidence reflects high genetic divergence of Amynthas aspergillum (Oligochaeta: Megascolecidae) in southern China.

Amynthas aspergillum (Perrier, 1872), a natural resource used in traditional Chinese medicine (Guang-dilong) with high economic value, is widely distributed in forests and farmland habitats in the hilly areas of southern China. To investigate the extent of genetic differentiation and diversity in A. aspergillum, a population genetic structure study was performed on 157 samples from 75 locations in southern China using the mitochondrial genes COI, COII, 12S rRNA, 16S rRNA, and NDI. The results indicated that A. aspergillum had a high level of genetic diversity, and variation within populations was the main source of the total variation. Six deeply divergent mitochondrial clades (I-VI) were detected using both phylogenetic tree and haplotype network analyses. This finding was supported by the high Kimura two-parameter genetic distance and the pairwise fixation index value obtained based on the COI gene. No significant phylogeographic structures were observed. The widespread geographic distribution of clades II, IV, and VI suggested a recent demographic expansion based on multiple analysis results. These results include a high level of Hd and low π, star-shaped haplotype network structures with a high number of less frequent haplotypes, significantly negative neutrality test values, and a unimodal mismatch distribution pattern. The divergence time estimates and reconstruction of the ancestral area revealed that A. aspergillum originated in Guangxi Province and underwent initial intraspecific diversification in the early Pliocene to generate clade I. Then, it gradually dispersed eastward and rapidly differentiated into clades II-V during the Pleistocene. The Yunnan-Guizhou Plateau and Nanling and Wuyi Mountains might act as geographical barriers for the spread of A. aspergillum to the west and north.

Systematic revision of the eastern Queensland land snail genus Figuladra Köhler and Bouchet, 2020 based on mitochondrial phylogenetics and comparative morpho-anatomy (Eupulmonata, Camaenidae)

ABSTRACT The current study revises the systematic taxonomy of Figuladra, a genus of camaenid land snails endemic to eastern Queensland, based on the investigation of many recently collected ethanol-preserved samples as well as historic museum material, including types. We employed comparative analyses to investigate the variation in morphological features (shell, penial anatomy) and performed phylogenetic analyses of mitochondrial 16S rDNA sequences to test species hypotheses. Phylogenetic analyses recovered three principal mitochondrial clades, one occurring in mid-east Queensland, one in south-east Queensland, and another in the upland areas of south-east Queensland. We recognise altogether 15 accepted species, including four that are newly described herein (i.e., Figuladra finlaysoni sp. nov., F. robertirwini sp. nov., F. bromileyorum sp. nov. and F. vidulus sp. nov.). The status of 11 previously described species has been assessed and revised diagnoses are provided for each of these. Species of the south-east Queensland clade revealed comparatively low levels of genetic and morphological differentiation and may represent examples of more recent, conceivably ongoing, speciation. Figuladra species have allopatric distributions with their ranges being separated by altitudinal or riverine barriers. Particularly the St Lawrence Gap dry corridor has been identified as a major biogeographic barrier separating two principal clades of Figuladra.

New data on the distribution of the Vipera ammodytes (Linnaeus, 1758) mitochondrial lineages place their contact zone in western Bulgaria

Molecular studies have outlined several mitochondrial lineages of Vipera ammodytes, but the limits of their distribution ranges remain unclear due to limited sampling throughout the vast range of the species. One particularly understudied area is the Eastern Balkan Peninsula where at least three lineages occur, showing ranges that may be partly overlapping. We used two mitochondrial markers (cytb and ND2) to unveil mtDNA distribution patterns based on vipers from 31 localities across Bulgaria. Our results confirmed the presence of the north-eastern (NE) and the south-eastern (SE) mitochondrial clade in Bulgaria, the latter being represented by its southern (S) and eastern (E) subclades. Among the sampled localities, two were suspected to be potential contact zones between these mtDNA lineages based on old morphology-derived distributional records. The NE clade was absent from both potential contact zones. However, our data showed that in western Bulgaria, populations of this clade establish contact with populations of the E subclade in at least one area, and also come close to contact with populations of the S subclade. These results indicate the need for more detailed research in the potential areas of contact in western Bulgaria, integrating morphological data with extensive mitochondrial and nuclear DNA-sampling to better understand the phylogeographic patterns of intraspecific differentiation in V. ammodytes.

Diversity and evolution of the MHC class II DRB gene in the Capra sibirica experienced a demographic fluctuation in China

The major histocompatibility complex (MHC) genes are the most polymorphic genes in vertebrates, and their proteins play a critical role in adaptive immunity for defense against a variety of pathogens. MHC diversity was lost in many species after experiencing a decline in size. To understand the variation and evolution of MHC genes in the Siberian ibex, Capra sibirica, which has undergone a population decline, we analyzed the variation of the second exon of MHC class II DRB genes in samples collected from five geographic localities in Xinjiang, China, that belong to three diverged mitochondrial clades. Consequently, we identified a total of 26 putative functional alleles (PFAs) with 260 bp in length from 43 individuals, and found one (for 27 individuals) to three (for 5 individuals) PFAs per individual, indicating the presence of one or two DRB loci per haploid genome. The Casi-DRB1*16 was the most frequently occurring PFA, Casi-DRB1*22 was found in only seven individuals, 14 PFAs occurred once, 7 PFAs twice, implying high frequency of rare PFAs. Interestingly, more than half (15) of the PFAs were specific to clade I, only two and three PFAs were specific to clades II and III, respectively. So, we assume that the polygamy and sexual segregation nature of this species likely contributed to the allelic diversity of DRB genes. Genetic diversity indices showed that PFAs of clade II were lower in nucleotide, amino acid, and supertype diversity compared to those of the other two clades. The pattern of allele sharing and FST values between the three clades was to some extent in agreement with the pattern observed in mitochondrial DNA divergence. In addition, recombination analyses revealed no evidence for significant signatures of recombination events. Alleles shared by clades III and the other two clades diverged 6 million years ago, and systematic neighbor grids showed Trans-species polymorphism. Together with the PAML and MEME analyses, the results indicated that the DRB gene in C. sibirica evolved under balancing and positive selection. However, by comparison, it can be clearly seen that different populations were under different selective pressures. Our results are valuable in understanding the diversity and evolution of the DRB gene in a mountain living C. sibirica and in making decisions on future long-term protection strategies.

Genetic diversity and complex structure of the European Roe Deer population at a continental scale

Abstract Although the European Roe Deer (Capreolus capreolus) is one of the most common and widespread ungulate species in Europe and inhabiting a variety of habitats, few studies have addressed its population structure at a large spatial scale using nuclear genetic data. The aims of our study were to: (i) investigate genetic diversity, level of admixture, and genetic structure across European Roe Deer populations; (ii) identify barriers to gene flow; and (iii) reveal factors that have impacted the observed pattern of population genetic structure. Using 12 microsatellite loci, we analyzed 920 European Roe Deer samples from 16 study sites from northern, southern, central, and eastern Europe. The highest genetic diversity was found in central and eastern sites, and lowest in the northern and southern sites. There were 2 main groups of genetically related populations in the study area—one inhabiting mainly Fennoscandia, and the second in the continental part of Europe. This second population was further divided into 3 to 5 spatially distributed genetic clusters. European Roe Deer belonging to the Siberian mitochondrial DNA clade, inhabiting large parts of eastern Europe, were not identified as a separate population in the analysis of microsatellite loci. No isolation by distance (IBD) was detected between roe deer from the fennoscandian and the continental study sites, but the Baltic Sea was inferred to be the main barrier to gene flow. Only weak IBD was revealed within the continental population. Three lower-level genetic barriers were detected in the western, southern, and eastern parts of the study area. The main factors inferred as shaping the observed genetic diversity and population structure of European Roe Deer were postglacial recolonization, admixture of different populations of the species originating from several Last Glacial Maximum refugial areas, and isolation of several study sites.

Phylogeography of the American crocodile, Crocodylus acutus (Crocodylia: Crocodylidae) in Colombia: a conservation perspective

Abstract Crocodylus acutus is the most widely distributed crocodile in the Neotropics, playing a key role in the ecosystems it inhabits. However, unsustainable use and habitat degradation have depleted its populations across its range. In Colombia, it is classified as Endangered by local authorities, and a legislative framework for its protection has been created, implementing hunting bans and breeding programmes. Successful implementation of these programmes depends on the existence of baseline genetic information, which is currently insufficient for the species. We assess the genetic diversity, phylogeography, and demographic patterns of the American crocodile in Colombia based on three mitochondrial markers (cox1, cytb, and trnP/trnF/D-Loop) and propose actions for its management and conservation. We processed 33 American crocodile samples from both wild and captive populations and performed genetic variability, phylogenetic, phylogeographical, and neutrality tests, including previously reported sequences. We found evidence of high genetic variability in Colombia, the existence of clearly differentiated mitochondrial phylogenetic clades, and possible human-influenced or stochastic demographic variation events. We propose guidelines for the translocation of American crocodiles based on our findings and present evidence of possible dispersal events in the Caribbean and Magdalena regions. Finally, we suggest the implementation of management units and identify research priorities for future work.

Phylogeography of the Cow Knob Salamander (Plethodon punctatus Highton): populations on isolated Appalachian mountaintops are disjunct but not divergent

Although many studies have examined how taxa responded to Pleistocene climate fluctuations in the Appalachian Mountains, impacts on high-elevation endemics of Central Appalachia are not yet understood. We use mitochondrial (ND4 & Cytb) and nuclear (GAPD) DNA sequences to investigate the phylogeography of the Cow Knob Salamander (Plethodon punctatus), a woodland species from Central Appalachian highlands thought to have origins in the Pleistocene. Data from 72 tail tips representing 25 sites revealed that the species comprises two geographically cohesive mitochondrial clades with a narrow, putative contact zone on Shenandoah Mountain. Molecular clock estimates indicate the clades diverged in the Middle Pleistocene. The population size of the Southern clade appears to have remained stable for at least 50,000 years. Despite spanning several isolated mountain systems, the Northern clade has exceptionally low genetic diversity, probably due to recent demographic expansion. Palaeodemographic hypothesis testing supported a scenario in which a founder effect characterized the Northern clade as it diverged from the Southern clade. Species distribution models predicted no suitable habitat for the species during the Last Glacial Maximum. Ultimately, Pleistocene glacial climates may have driven the species from the northern half of its current range, with recolonization events by members of the Northern clade as climates warmed. Density dependent processes may now maintain a narrow contact zone between the two clades.

Every branch in its niche: intraspecific variation in habitat suitability of a widely distributed small mammal, the harvest mouse Micromys minutus

Climate and land use changes re-shape the distribution of species around the world, so understanding biogeographical patterns is key to protect and manage wildlife, particularly in the case of threatened or declining species. Species with wide ranges are usually considered as common, yet the occurrence of intraspecific variability and adaptation to local conditions may pose serious challenges to their conservation. In this paper, we selected the harvest mouse Micromys minutus as a model species to assess the role of intraspecific genetic variability in shaping the environmental preferences and distribution of a widely distributed mammal across Europe. By applying an integrative approach combining species distribution modelling and phylogenetic reconstruction, we assessed the clade-specific habitat suitability of all mitochondrial clades of the species occurring in Europe, unveiling distinct bioclimatic niches. Most of Central Europe results as suitable to M. minutus, whereas different clades showed distinct distributions of suitable areas across Europe, with limited overlap. We then combined mitochondrial analyses with clade-specific suitability maps to determine the geographic delimitations of Italian and central European clades. We showed that individuals from the northernmost Italian regions actually belong to the central European clade, consistent with the biogeographical patterns of other small mammals, and indicating potential past dispersal between the Italian Peninsula and Central Europe, or human-assisted translocations. Our results highlight the importance of intraspecific variability in shaping biogeographical patterns of widely distributed species, also representing a reproducible exercise to understand their environmental preferences and eventually fostering their conservation.

Ancient DNA of the Don-Hares Assumes the Existence of Two Distinct Mitochondrial Clades in Northeast Asia

Paleoclimatic changes during the Pleistocene-Holocene transition is suggested as a main factor that led to species extinction, including the woolly mammoth (Mammuthus primigenius), Steller's sea cow (Hydrodamalis gigas) and the Don-hare (Lepus tanaiticus). These species inhabited the territory of Eurasia during the Holocene, but eventually went extinct. The Don-hare is an extinct species of the genus Lepus (Leporidae, Lagomorpha), which lived in the Late Pleistocene-Early Holocene in Eastern Europe and Northern Asia. For a long time, the Don-hare was considered a separate species, but at the same time, its species status was disputed, taking into account both morphological data and mitochondrial DNA. In this study, mitochondrial genomes of five Don-hares, whose remains were found on the territory of Northeastern Eurasia were reconstructed. Firstly, we confirm the phylogenetic proximity of the "young" specimens of Don-hare and mountain or white hare, and secondly, that samples older than 39 Kya form a completely distinct mitochondrial clade.

Genetic diversity of the mantellid frog Blommersia blommersae, and description of a new anuran species from south-eastern Madagascar

ABSTRACT A range-wide assessment of genetic variation in one mitochondrial (16S rRNA) and one nuclear-encoded (Rag-1) gene fragment of the widespread Madagascar frog Blommersia blommersae revealed the presence of up to 12 deep genetic lineages. Many of these differed by genetic distances >3% in the 16S gene. In the Ranomafana area in the southern central east of Madagascar, two mitochondrial lineages differing by 1.2–1.8% in 16S occurred in close syntopy across multiple sites. A phylogeny of representative samples based on multiple mitochondrial genes supported three main mitochondrial clades within B. blommersae, with lineages from the (i) north, (ii) north and northern central east, and (iii) southern central east, respectively. In addition, one lineage from Sainte Luce in the south-east was sister to all other species of Blommersia, and thus clearly not belonging to B. blommersae, to which it has been tentatively assigned previously. Except for the Sainte Luce population, most of the mitochondrial lineages did not show a concordant and consistent differentiation in Rag-1 and no obvious morphological differences were detected among the lineages. We therefore refrain from taxonomic conclusions for them at this time, although indications exist that the populations from the northern central east and southern central east may show certain differences in relative note duration of advertisement calls, which require further study. However, the Sainte Luce individuals were highly divergent in 16S (>10%) and Rag-1, phylogenetically isolated, and characterised by differences in colour pattern. This lineage represents the most evolutionarily divergent species of Blommersia known to date and is herein formally named and described as Blommersia dupreezi sp. nov.

Multiple concordant cytonuclear divergences and potential hybrid speciation within a species complex in Asia

Various environmental factors impact the distribution, population structure, demography and evolutionary trajectory of a bird species, leading to genetic and morphological divergences between populations across its distribution. The Paradoxornis webbianus species complex is found throughout much of East Asia, where its geographically distinct populations exhibit dramatic morphological variation. This has resulted in a hotly debated taxonomy. This study intended to identify genetic divergence patterns and their underlying contributing factors for this species complex. We collected 243 birds, whose data was combined with those available in GenBank to perform phylogeographic analyses using one mitochondrial and six nuclear loci. Six mitochondrial clades were observed in the species complex, while individual-based Bayesian clustering using nuclear markers showed multiple congruent breaks. Overall, the six molecular lineages could be recognized as independent species under the lineage species concept in view of genetic divergence, clade-specific morphological changes and distribution: P. webbianus, P. w. bulomachus, P. alphonsianus, P. a. ganluoensis, P. brunneus brunneus and P. b. ricketti. The estimated divergence times range from 0.46 to 3.36 million years ago, suggesting it was likely impacted by paleoclimatic changes. Interestingly, P. alphonsianus carries two divergent mitochondrial lineages shared with P. webbianus and P. a. ganluoensis, respectively, and analyses based on nuclear loci found a similar pattern. We discussed the various hypotheses for this pattern and argued that P. alphonsianus was likely the result of hybridization between P. webbianus and P. a. ganluoensis. Further data on genome, transcriptome and breeding ecology are needed to address the hypothesis of hybrid speciation and its underlying mechanisms.

Museomics Provides Insights into Conservation and Education: The Instance of an African Lion Specimen from the Museum of Zoology “Pietro Doderlein”

Innovative technological approaches are crucial to enhance naturalistic museum collections and develop information repositories of relevant interest to science, such as threatened animal taxa. In this context, museomics is an emerging discipline that provides a novel approach to the enhancement and exploitation of these collections. In the present study, the discovery of a neglected lion skeleton in the Museum of Zoology “Pietro Doderlein” of the University of Palermo (Italy) offered the opportunity to undertake a multidisciplinary project. The aims of the study consisted of the following: (i) adding useful information for museographic strategies, (ii) obtaining a new genetic data repository from a vulnerable species, (iii) strengthening public awareness of wildlife conservation, and (iv) sharing new learning material. The remains of the lion were examined with a preliminary osteological survey, then they were restored by means of 3D printing of missing skeletal fragments. Phylogenetic analyses based on cytochrome b sequence clearly indicate that the specimen belongs to the Central Africa mitochondrial clade. At the end of the study, the complete and restored skeleton was exhibited, along with all of the information and data available from this project. This study shows a useful approach for the restoration and enhancement of a museum specimen, with important opportunities for preserving biodiversity and driving specific conservation policies, but also for providing Life Science learning material.

Mitochondrial genomes reveal mid-Pleistocene population divergence, and post-glacial expansion, in Australasian snapper (Chrysophrys auratus)

Glacial cycles play important roles in determining the phylogeographic structure of terrestrial species, however, relatively little is known about their impacts on the distribution of marine biota. This study utilised modern (n = 350) and ancient (n = 26) mitochondrial genomes from Australasian snapper (Chrysophrys auratus) sampled in New Zealand to assess their demographic and phylogeographic history. We also tested for changes in genetic diversity using the up to 750-year-old mitochondrial genomes from pre-European archaeological sites to assess the potential impacts of human exploitation. Nucleotide diversity and haplotype diversity was high (π = 0.005, h = 0.972). There was no significant change in nucleotide diversity over the last 750 years (p = 0.343), with no detectable loss of diversity as a result of indigenous and industrial-scale fishing activity. While there was no evidence for contemporary population structure (AMOVA, p = 0.764), phylogeographic analyses identified two distinct mitochondrial clades that diverged approximately 650,000 years ago during the mid-Pleistocene, suggesting the species experienced barriers to gene flow when sea levels dropped over 120 m during previous glacial maxima. An exponential population increase was also observed around 8000 years ago consistent with a post-glacial expansion, which was likely facilitated by increased ocean temperatures and rising sea levels. This study demonstrates that glacial cycles likely played an important role in the demographic history of C. auratus and adds to our growing understanding of how dynamic climatic changes have influenced the evolution of coastal marine species.

Ancient DNA Reveals Maternal Philopatry of the Northeast Eurasian Brown Bear (Ursus arctos) Population during the Holocene.

Significant palaeoecological and paleoclimatic changes that took place during Late Pleistocene-Early Holocene transition are considered important factors that led to megafauna extinctions. Unlike many other species, the brown bear (Ursus arctos) has survived this geological time. Despite the fact that several mitochondrial DNA clades of brown bears became extinct at the end of the Pleistocene, this species is still widely distributed in Northeast Eurasia. Here, using the ancient DNA analysis of a brown bear individual that inhabited Northeast Asia in the Middle Holocene (3460 ± 40 years BP) and comparative phylogenetic analysis, we show a significant mitochondrial DNA similarity of the studied specimen with modern brown bears inhabiting Yakutia and Chukotka. In this study, we clearly demonstrate the maternal philopatry of the Northeastern Eurasian U. arctos population during the several thousand years of the Holocene.

Forensic Genetic Analyses of Melanistic Iguanas Highlight the Need to Monitor the Iguanid Trade.

Simple SummaryThe illegal pet trade remains an ongoing, substantial threat to wild populations, especially small insular populations, and can even lead to extinction. Fraudulent activity within the global reptile trade is known to occur, but its identification through forensic applications depends on knowledge of diversity within wild populations. In this study, we assessed the geographic origin of melanistic iguanas (Iguana iguana), which are only found in nations that have never authorized legal export of live animals. Analysis of genetic data from two pet iguanas in the USA flag these as originating from Saba or Montserrat, from which no export permits have ever been issued, confirming their illegal origin. Despite the international trade in I. iguana, in which tens- if not hundreds of thousands of specimens are traded each year, only a handful of individuals have been genetically assessed. Our work highlights the utility of applying forensic genetic techniques to this trade in order to track and discourage illegal activity.Lizards within the Iguana iguana species complex are among the most common reptilian pets, with the widest natural geographic range among iguanids. Deep phylogenetic divergence distinguishes multiple mitochondrial clades, and several taxonomic changes have recently been proposed. These small populations, typically island endemics, are threatened by numerous factors, including the international pet trade. Recent investigations reveal the absence of required CITES permits for lawful export of animals, providing evidence of ongoing illegal trade. Additional monitoring of trade in iguanas can be achieved through the application of forensic molecular techniques. In this study, two captive melanistic iguanas were genotyped for molecular markers for which geographic distributions of alleles have been established. Mitochondrial sequencing indicates that both animals carry a haplotype known to originate from the islands of Saba and Montserrat, populations taxonomically proposed to be Iguana melanoderma. Genotypes at 15 microsatellite loci are equally consistent with this origin, given the results of a principal component analysis. This first forensic genetic assessment within the extensive I. iguana pet trade highlights the presence of illegal activity. The need for additional forensic assessments of pet-trade iguanas is evident, especially given that their value is driven by variety and rarity, which is further intensified by recent taxonomic changes.

Recent invasion and eradication of two members of the Euwallacea fornicatus species complex (Coleoptera: Curculionidae: Scolytinae) from tropical greenhouses in Europe

Ambrosia beetles of the Euwallacea fornicatus species complex are emerging tree pests with a broad host range including important agricultural crops. Native to Southeast Asia, these species were introduced into various countries, where they cause considerable damage to many tree species. Here we report several outbreaks of E. fornicatus s.l. in Europe. The first individuals were found in 2017 in a palm house of a botanical garden in Poznan (Poland) whereas in 2020 an outbreak was detected in a tropical greenhouse in Merano (Italy). In 2021, two additional outbreaks were detected in two greenhouses in Germany, in Erfurt and Berlin. For both cases in Germany it was possible to trace back the invasion to a distributor of exotic plants in the Netherlands where several infested plants were detected. Molecular analyses show that individuals from Poland and Italy are genetically identical but belong to a different mitochondrial clade than individuals in Germany which are identical to most individuals of two greenhouses in the Netherlands. Moreover, in the two greenhouses in the Netherlands we found beetles that belong to another haplotype of E. fornicatus and two haplotypes of E. perbrevis, a species in the E. fornicatus complex, which has not been previously intercepted in Europe. Our study provides novel insights into the invasion history of E. fornicatus and the eradication measures in Europe. Considering the potential of introduction and establishment of Euwallacea ambrosia beetles, particular attention should be paid to monitor the presence of these pests in tropical greenhouses across Europe.

Shedding light on taxonomic chaos: Diversity and distribution of South Asian skipper frogs (Anura, Dicroglossidae, Euphlyctis)

A known haven of amphibian diversity, South Asia is also a hotspot of taxonomic confusions. Vastly distributed from Saudi Arabia to Myanmar, the dicroglossid genus Euphlyctis (“skittering” or “skipper” frogs) is a representative example. Combining phylogenetic analyses with 16S barcoding and genome size variation of 403 frogs from 136 localities, we examined genetic diversity and distributions across the whole range of Euphlyctis, with a particular focus on taxonomic and nomenclatural issues. We recovered two deeply divergent mitochondrial clades totalling ten lineages that we considered as species, and eight could be attributed valid taxonomic names and junior synonyms. The first clade (subgenus Phrynoderma) is confirmed in South India, Bangladesh and Sri Lanka, and comprises six species: E. karaavali, E. hexadactyla, E. aloysii, E. kerala and two undescribed taxa. Five are endemic to the Western Ghats biodiversity hotspot and four of them form the E. aloysii species complex. The second clade (subgenus Euphlyctis) extends across South Asia and neighbouring regions, and comprises four species: E. ehrenbergii, E. jaladhara, and two widespread lineages erroneously called “E. mudigere” and “E. kalasgramensis” in recent literature, while their oldest valid names appear to be E. cyanophlyctis and E. adolfi, respectively. Additional analyses on this pair of taxa highlighted strong phenotypic resemblance, notable intraspecific phylogeographic structure, and an extensive contact zone along the southern slopes of the Himalaya, with putative signs of genetic introgression. Through an independent investigation of the historical literature, we identified overlooked issues and misconceptions regarding the status of many old and recent taxa, and proposed solutions, such as transferring “E. ghoshi” to the genus Limnonectes. Our study illustrates how range-wide genetic barcoding can clarify taxonomic confusions, and we call to solve remaining issues prior to the description of new taxa.