Mitochondrial ND2 Gene Research Articles

Unrecognised (species) diversity in New Guinean passerine birds

ABSTRACT Species represent an important unit for the study of diversity, but may not always be delimited consistently across regions and clades. Many of these taxonomic inconsistencies are due to the variable views of taxonomists. In recent years, however, new methodologies have attempted to circumvent this problem by assigning more objective criteria for the delimitation of species, drawing on a wide range of data such as DNA, morphology, vocalisation and ecology. Here, we apply a genetic screening approach in which we sequence the mitochondrial gene ND2 for all recognised subspecies of 16 species in eight genera (a mix of lowland and montane species) from the geologically complex tropical island of New Guinea. We show that populations within some species are genetically highly divergent despite little morphological differentiation, but we also find an example in which populations from five morphologically distinct species are genetically very similar. Overall, our data show higher levels of genetic differentiation than expected, but also highlight the difficulty of predicting which groups contain unrecognised diversity. These results are interesting in their own right, but also have implications for further analyses that focus on increasing our understanding of how diversity builds up over time.

A new species of crocodile newt Tylototriton (Caudata: Salamandridae) from northern Myanmar (Burma)

ABSTRACTA new species of crocodile newt from populations on opposite sides of the Ayeyarwady Basin, Myanmar, previously referred to as Tylototriton cf. verrucosus, is described on the basis of colour pattern, morphology and phylogenetic data from the mitochondrial gene ND2. Tylototriton panwaensis sp. nov. is most closely related to T. verrucosus + T. shanjing but bears a unique suite of colour pattern and morphological characters separating it from T. verrucosus and T. shanjing as well as the other geographically proximate species T. ngarsuensis and T. shanorum. Molecular and morphological data presented herein (that have been used elsewhere) continue to fuel the issue of separate specific allocation (or not) to T. verrucosus and T. shanjing but go no further to resolve the controversy. The description of this new species adds to a rapidly growing database of newly described species of amphibians and reptiles from the upland, mountainous regions of eastern Myanmar.urn:lsid:zoobank.org:pub:BE851CE6-81BD-488C-A616-39A948A2B9C6

Revision of the taxonomic status of Synthesium elongatum (Ozaki, 1935) (Brachycladiidae), an intestinal digenean of narrow-ridged finless porpoise (Neophocaena asiaeorientalis).

Synthesium elongatum (Brachycladiidae) is an intestinal digenean described from the finless porpoise (Neophocaena asiaeorientalis) in Japan. Few records of this species exist and there is a remarkable morphological similarity between S. elongatum and S. tursionis, such that a synonymy between the species has been suggested previously. However, no morphological and/or molecular analysis has been carried out to clarify the taxonomic status of S. elongatum. In this study, we collected specimens of Synthesium sp. from N. asiaeorientalis in western Japan. The specimens possess lobed testes within the third quarter of the body, a round ovary, and vitellaria extending to level of uterine field, which are diagnostic characters for both S. elongatum and S. tursionis. They were morphologically identified to S. elongatum or S. tursionis due to the fact that the available morphometric data for both species overlap remarkably. A molecular analysis of the mitochondrial ND3 gene showed that the pairwise nucleotide distances between these specimens and S. tursionis were small, and phylogenetic analysis showed that these specimens and S. tursionis are in the same clade. Therefore, it was indicated that S. elongatum and S. tursionis are the same species and, consequently, S. elongatum is a synonym of S. tursionis.

A re-description of Cyrtodactylus chrysopylos Bauer (Squamata: Gekkonidae) with comments on the adaptive significance of orange coloration in hatchlings and descriptions of two new species from eastern Myanmar (Burma).

The karstic foothills and hilly western edge of the Shan Plateau of eastern Myanmar continue to be sources of discovery for new species of the gekkonid genus Cyrtodactylus. Two new karst-associated species within this unique landscape from Mandalay Region-C. aunglini sp. nov. from the Kyauk Nagar Cave and C. myaleiktaung sp. nov. from Mya Leik Taung Mountain-are described on the basis of color pattern, morphology, and genetics. A molecular phylogeny of the C. gansi group (defined herein) based on 1481 base pairs of the mitochondrial gene ND2 and its flanking tRNAs place C. aunglini sp. nov. as the sister species to C. gansi and C. myaleiktaung sp. nov. as the sister species of C. chrysopylos. A new population of C. chrysopylos from a boulder-strewn, karstic ridge near the village of Yane, Shan State at an elevation 951 m is reported here. Genetic and geographic variation as well as differences in natural history between it and the lowland cave population of C. chrysopylos from the type locality in the Panlaung-Pyadalin Cave Wildlife Sanctuary, Shan State at 303 m are discussed. Hatchlings of the new population from Yane are nearly uniformly bright-orange in color. We provide evidence in support of a hypothesis claiming this color functions as a mechanism for crypsis during low levels of illumination and is not involved in intraspecific communication.

Distribution and Origin of Two Forms of the Marsh Frog Pelophylax ridibundus Complex (Anura, Ranidae) from Kamchatka Based on Mitochondrial and Nuclear DNA Data

The formation of the first populations of the marsh frog (the Pelophylax ridibundus complex) near Petropavlovsk-Kamchatsky and in the Paratunka River valley must have been the result of human introduction in the late 1980s. At present, more than 20 localities of this species are recorded in Kamchatka. For a more precise definition of the taxonomic status of P. ridibundus sensu lato, samples from five populations (altogether, 30 individuals) from southeastern and central Kamchatka are analyzed using molecular methods. In all frogs, a mitochondrial DNA type specific for the “eastern” form (=the Anatolian P. cf. bedriagae), but not for the “western” form (=the Central European P. ridibundus), is revealed. However, the results of nuclear DNA analysis of marsh frogs from Kamchatka reveal alleles specific for both of the forms, “eastern” and “western,” with a frequency ratio of about 2 : 1. The results of sequencing the mitochondrial ND2 gene and nuclear SAI-1 gene suggest that the “ancestor” individuals might have been introduced into Kamchatka from the Volga–Don interfluve or Ciscaucasia. The absence of both haplotype and nucleotide diversity in the samples studied suggests a single successful introduction that involved a low number of frogs stemming from a single locality.

Description of a new deep-water dogfish shark from Hawaii, with comments on the Squalusmitsukurii species complex in the West Pacific.

Dogfish sharks of the genus Squalus are small, deep-water sharks with a slow rate of molecular evolution that has led to their designation as a series of species complexes, with low between-species diversity relative to other taxa. The largest of these complexes is named for the Shortspine spurdog (Squalusmitsukurii Jordan & Snyder), a medium-sized dogfish shark common to warm upper slope and seamount habitats, with a putative circumglobal distribution that has come under investigation recently due to geographic variation in morphology and genetic diversity. The Hawaiian population of Squalusmitsukurii was examined using both morphological and molecular analyses, putting this group in an evolutionary context with animals from the type population in Japan and closely-related congeners. External morphology differs significantly between the Hawaiian and Japanese S.mitsukurii, especially in dorsal fin size and relative interdorsal length, and molecular analysis of 1,311 base pairs of the mitochondrial genes ND2 and COI show significant, species-level divergence on par with other taxonomic studies of this genus. The dogfish shark in Hawaii represents a new species in the genus, and the name Squalushawaiiensis, the Hawaiian spurdog, is designated after the type location.

Geographically structured genetic variation in Ptychozoon lionotum (Squamata: Gekkonidae) and a new species from an isolated volcano in Myanmar.

A molecular phylogenetic analysis of Parachute Geckos (Genus Ptychozoon Kuhl van Hasselt, 1822) based on the mitochondrial gene ND2 indicates that a newly discovered population from the Mt. Popa volcano-a habitat island in the northern portion of the Bago Yoma mountains, Myanmar-is a new species, P. popaense sp. nov. This species is part of a clade that contains P. bannanense Wang, Wang, Liu, 2016 and P. lionotum Annandale, 1905 of Indochina. Ptychozoon popaense sp. nov. is morphologically most similar to its sister species P. lionotum which manifests considerable geographic substructuring of genetic variation but differs from the nominate taxon by an uncorrected pairwise sequence divergence of 16.0-17.1% and by discrete differences in morphology and color pattern. This discovery highlights the unique, insular nature of the Bago Yoma mountains of the Ayeyarwady Basin, which support other endemic gekkonids. It also underscores the growing diversity in this highly derived clade of cryptic, parachuting, geckos characterized by highly divergent genetic lineages, which may indicate the presence of additional, unrecognized species.

Evaluation of Phylogenetic Characteristics of Iranian Honeybee (Apis mellifera meda) Populations based on Mitochondrial ND2 Gene

Evaluation of Phylogenetic Characteristics of Iranian Honeybee (Apis mellifera meda) Populations based on Mitochondrial ND2 Gene

The smallest ‘true chameleon’ from Madagascar: a new, distinctly colored species of the Calumma boettgeri complex (Squamata, Chamaeleonidae)

On a recent expedition to eastern Madagascar, we discovered a distinct new species of the genusCalummathat we describe here using an integrative approach combining morphology, coloration, osteology and molecular genetics.Calummaroalokosp. n.has a dermal rostral appendage and occipital lobes, and belongs to theC.boettgericomplex, within the Madagascar-endemic pheneticC.nasutumspecies group. It is readily distinguished from other species of theC.boettgericomplex by a characteristic two-toned body coloration and small body size with a snout-vent length of 45.6 mm in an adult male. The osteology of the skull, with a prominent maxilla and broad parietal, is similar to the closest related species,C.uetzi. Analysis of uncorrected genetic distances within theC.nasutumgroup using the mitochondrial gene ND2 shows a minimum pairwise distance of 11.98% toC.uetzifrom the Sorata massif and Marojejy National Park >500 km north of the type locality ofC.roalokosp. n.. Given an apparently small range (potentially <300 km2), located entirely outside of any nationally-protected areas, we recommend this new species be classified as Endangered under criterion B1ab(iii) of the IUCN Red List. The discovery of clearly distinct species likeC.roalokosp. n.in an area of Madagascar that is comparatively thoroughly surveyed highlights the critical role of continued field surveys for understanding the true extent of Madagascar’s spectacular biodiversity.

Phylogeography and geographic variation in the Red-bellied Woodpecker (Melanerpes carolinus): characterization of mtDNA and plumage hybrid zones

ABSTRACT We used sequences of the mitochondrial ND2 gene, the nuclear TGFB-2 intron-5, and differences in plumage pattern to assay geographic variation in the Red-bellied Woodpecker (Melanerpes carolinus). Ten population samples from throughout its range indicated that ∼23% of the mitochondrial genetic variation was distributed among populations, largely due to the occurrence of a clade of haplotypes restricted to the Florida peninsula with a local frequency of 82%. In a hierarchical analysis, 37% of the mtDNA genetic variance was distributed between the Florida peninsula and elsewhere. A strong signal of population growth was detected in the non-peninsular portion of the range. Otherwise, geographic variation in haplotype frequencies was minor, with no evidence of isolation-by-distance. Nuclear intron data showed little variation among populations (Fst = 0.007). These results suggest recent secondary contact of 2 differentiated taxa following expansion from allopatric Pleistocene refugia. We modeled the ...

Genomic population structure aligns with vocal dialects in Palm Cockatoos (Probosciger aterrimus); evidence for refugial late-Quaternary distribution?

ABSTRACTSpecies persistence and maintenance of genetic diversity are strongly affected by dispersal and historical distribution, especially when species depend on habitat that is non-uniform or fluctuates dramatically with changing climate. Australo-Papuan rainforest has fluctuated dramatically since the last glacial maximum (around 20 kya). To understand how prehistoric climate fluctuation affected population connectivity and genetic diversity in a rainforest edge species, we screened 27 Palm Cockatoo samples from Cape York Peninsula (Australia) and southern Papua New Guinea (PNG) in 1132 single nucleotide polymorphisms in 342 nuclear loci and the mitochondrial ND2 gene. We also modelled the birds’ distribution at present, mid-Holocene (~6 kya) and the last glacial maximum (~21 kya). Population differentiation in nuclear genomic data among Australian populations aligns with distribution contraction to mountainous refugia at the mid-Holocene (~6 kya). Lack of nuclear divergence between PNG and Australia may reflect late-Holocene recolonisation, but different ND2 haplotypes suggest early stages of divergence. Although admixed individuals suggest some gene flow, recent movement restriction to/from Australian refugia is suggested by a unique ND2 haplotype, genomic divergence and a vocal dialect boundary shown previously. Our results show how prehistoric climate fluctuation affects present-day and future species conservation in dynamic rainforest edge ecosystems.

The diverse prey spectrum of the Tanganyikan scale-eater Perissodus microlepis (Boulenger, 1898)

Feeding upon the scales of other fish—lepidophagy—is a highly specialized foraging strategy in fish. Scale-eating is rare in teleosts, yet has evolved several times in East African cichlids, the most famous case being the Perissodini clade in Lake Tanganyika. Here, we examined the prey spectrum of the scale-eater Perissodus microlepis (Boulenger, 1898) via morphological assessment and targeted sequencing (barcoding) of ingested scales. We found that the size of the ingested scales, but not their number, correlates with the body size of scale-eaters. Sequencing of a segment of the mitochondrial ND2 gene in more than 300 scales revealed that P. microlepis feed upon a broad spectrum of prey species. In total, we detected 39 different prey species, reflecting the cichlid community in the rocky littoral zone of Lake Tanganyika. The most common prey were the algae-eaters Petrochromis polyodon, Pe. ephippium, Eretmodus cyanostictus, Tropheus moorii, and Simochromis diagramma, which make up more than half of the diet. The diversity of scales found within scale-eaters and the overall broad prey spectrum suggest that P. microlepis is an opportunistic feeder. Mouth-handedness and body color hue of the scale-eaters do not seem to have an influence on prey choice.

The development of two novel species-specific primers for identifying genus Crassostrea, with focus on C. sikamea and C. ariakensis

Oyster reefs have important ecological functions and environmental service values, such as filtering water, providing fish habitats, maintaining biodiversity, and carbon sequestration. Oyster reef recovery has become one of the most important means for fishery resource conservation and ecosystems restoration of coastal regions. C. sikamea and C. ariakensis are two important reef oysters along the coast of China. However, it is very difficult to identify the larvae of these two oysters. To solve this problem, we developed two pairs of novel species-specific PCR primers for identifying these two oysters. The forward C. sikamea-specific primer is Cs-F: 5′-CGAAGAGGGGCATGATAAATGAGG-3′, and the reverse primer is Cs-R: 5′-ATATGAACTTCTCCAACCTCCCC-3′, both of them within the scope of mitochondrial ND5 gene. The forward C. ariakensis-specific primer is Ca-F: 5′-GGGCAAATAAAAGGCAAAACCC-3′, located between tRNAser and tRNApro, and the reverse primer is Ca-R: 5′-CATAAACTTCTGCAAGACTCCC-3′, lies between tRNApro and small subunit ribosomal RNA (ssrRNA). These two primers can identify the larvae of C. ariakensis and C. sikamea from other oysters rapidly and effectively, with extremely high accurate rate (=100%). This molecular identification method does not require prior sorting of larvae. These two specific primers are good contribution to molecular identification of C. sikamea, C. ariakensis and other oysters, and could be a useful tool for oyster larvae identification and ecological restoration.

Phylogeny of the egg-loving green alga Oophila amblystomatis (Chlamydomonadales) and its response to the herbicides atrazine and 2,4-D

The spotted salamander (Ambystoma maculatum) shares a unique endosymbiotic relationship with the unicellular green alga, Oophila amblystomatis. Despite studies isolating and identifying O. amblystomatis in salamander eggs, the taxonomic identity of the alga remains a point of ongoing debate. In this study, the nuclear SSU rRNA gene was used to characterize two well-supported Oophila clades that include lineages identified from past studies in addition to new isolates from the current study. These two clades do not form a monophyletic group and, furthermore, O. amblystomatis appears to be paraphyletic with numerous other chlamydomonad algae. To gain further insight into the biogeographic variation of the host A. maculatum, the mitochondrial ND4 and control gene regions were examined and the phylogeography was observed to be similar to that noted in the literature. Additionally, the response of O. amblystomatis to atrazine and 2,4-dichlorophenoxyacetic acid following 96 h exposure and a 96 h recovery phase was characterized, as this is a plausible mechanism by which the development of the host salamander could be impaired by herbicides. At the 96 h growth rate, no-observed-effect concentrations were 64 μg/L and 30 mg/L for atrazine and 2,4-D, respectively. We observed full recovery ofO. amblystomatis at these concentrations within 96 h. These data suggest that atrazine and 2,4-D do not pose a significant risk to the symbiotic algae, or, indirectly, to the host salamander. In conclusion, we recommend a revision of the current taxonomy of O. amblystomatis, and demonstrate the need for species identification and thorough phylogenetic reconstruction in toxicity testing to accurately inform risk assessment.

Genetic variability of Aedes aegypti in the department of Sucre, Colombia, by analysis of the nucleotide sequence of the mitochondrial ND4 gene

Introduction. Aedes aegypti is the most important mosquito species in America for the transmission of viruses of dengue, Zika, Chikungunya and yellow fever. Ecological factors as well as chemical controls can affect the genetic composition of Ae. aegypti populations, which is why its genetic characterization is necessary. Objective. To determine the genetic variability of Ae. aegypti populations in four municipalities of Sucre department, Colombia. Materials and methods. Larvae of Ae. aegypti, collected in the municipalities of Sincelejo, Sampués, Corozal and Guaranda, Sucre department, were reared under laboratory conditions to adult stage. A segment of the mitochondrial ND4 gene which codes for the subunit 4 of the enzyme NADH-dehydrogenase was used as genetic marker. The genetic analysis included the estimation of parameters of nucleotide and haplotype diversity, genetic structure and gene flow. Results. One hundred and eight partial sequences of 357 nucleotides and four nucleotide haplotypes of the ND4 gene of Ae. aegypti were obtained. A significantly high genetic differentiation was found between the Sampués and Guaranda populations (FST=0.59467), Sincelejo and Sampués (FST=0.25637), and Corozal and Guaranda (FST=0.22237). A high gene flow (Nm=infinite) was observed among the populations of Sincelejo and Corozal. Conclusion. There are genetic differences between the Ae. aegypti populations from the municipalities of Sucre department. The presence of a new haplotype of the mitochondrial ND4 gene of Ae. aegypti in Colombia was recorded, detected in the municipality of Sincelejo.

Association between sperm mitochondrial ND2 gene variants and total fertilization failure

ABSTRACTThe objective of this study was to explore the association of sperm mitochondrial ND2 (MT-ND2) gene variants with total fertilization failure (TFF). A retrospective comparative study of 246 cases of fresh in vitro fertilization (IVF) cycles or half-intracytoplasmic sperm injection cycles in the Han Chinese population was performed from July 2011 to May 2017. A total of 59 cases undergoing TFF, and 187 control cases with normal fertilization (fertilization rates >50%) were included. The sperm mitochondrial genovariation was determined using nested sequencing. A total of 32 homoplasmic variants and 47 heteroplasmic variants of MT-ND2 gene were observed in this study. There were no significant differences in the frequencies of the 32 homoplasmic variants of MT-ND2 gene between the TFF and control groups. A total of 53 pair-wise comparisons were performed, and the general characteristics of the IVF failure and control subjects were adjusted in logistic models. Data suggested that there were no significant differences in the frequencies of point 4914, 5320, and 5426 heteroplasmic variants of MT-ND2 gene between the TFF and control groups. In addition, no significant difference was observed in the frequency of mtDNA haplogroup D or haplogroup G between the IVF failure group and the normal fertilization group. This study suggests that the MT-ND2 gene variants might not be associated with TFF.Abbreviations: ATP: adenosine triphosphate; dNTP: deoxy-ribonucleoside triphosphate; FADH2: flavin adenine dinucleotide; FDR: false discovery rate; FSH: follicle-stimulating hormone; IVF: in vitro fertilization; LH: luteinizing hormone; MTATP6: mitochondrially encoded ATP synthase 6; MTCYB: mitochondrially encoded cytochrome b; mtDNA: mitochondrial DNA; MT-ND2: mitochondrial ND2; NADH: nicotinamide adenine dinucleotide; ND2: NADH dehydrogenase subunit 2; OXPHOS: oxidative phosphorylation; PCR: single nucleotide polymorphisms; SNPs: single nucleotide polymorphisms; TFF: total fertilization failure

A new species of Hemiphyllodactylus (Squamata: Gekkonidae) from Hong Kong

A new species of gekkonid, Hemiphyllodactylus hongkongensis sp. nov., is described based on a series of specimens collected from Hong Kong SAR, China. The new species can be distinguished from other known congeners by molecular divergence in the mitochondrial ND2 gene and morphological characters including 5-6 chin scales; a manual lamellar formula of 3-3(4)-4-4; a pedal lamellar formula of 3(4)-4(5)-4(5)-4; 24-25 continuous femoral and precloacal pores; 12-15 dorsal scales and 9-10 ventral scales contained in diameter of eye. At present, the genus Hemiphyllodactylus comprises 22 described species.

How Do Patients Respond to Genetic Testing for Age-related Macular Degeneration?

SIGNIFICANCEThe American Academy of Ophthalmology currently recommends against routine genetic testing for complex diseases such as age-related macular degeneration (AMD). The results of this study demonstrate that patients are very interested in predictive genetic testing for AMD, find the information useful, and make behavioral changes as a result of the information.PURPOSEThe goal of this project was to conduct a pilot AMD genomic medicine study.METHODSEligible patients were aged 50 to 65 years with no personal history of AMD. DNA samples were genotyped for five single-nucleotide polymorphisms (SNPs) in the CFH gene, one SNP in the ARMS-2 gene, one SNP in the C3 gene, and one SNP in the mitochondrial ND2 gene. A risk score was calculated utilizing a model based on odds ratios, lifetime risk of advanced AMD and known population prevalence of genotype, haplotype, and smoking risk. The study optometrist provided the patient's risk score and counseling for personal protective behaviors. Telephone interviews were conducted 1 to 3 months after the counseling visit.RESULTSOne hundred one subjects (85%) participated in the genetic testing; 78 (77.2%) were female. Follow-up interviews were conducted with 94 participants (93.1%). More than half (n = 48) of the participants said that they were motivated to participate in the study because they had a family member with AMD or another eye or genetic disorder. Despite low risk levels, many participants reported making changes as a result of the genetic testing. Twenty-seven people reported making specific changes, including wearing sunglasses and brimmed hat and taking vitamin supplements. Another 16 people said that they were already doing the recommended activities, including wearing glasses, quitting smoking, and/or taking vitamins.CONCLUSIONSInterest in genetic testing for future risk of AMD was high in this population and resulted in support to continue current health behaviors or incentive to improve behaviors related to eye health.

Climate-driven mitochondrial selection: A test in Australian songbirds.

Diversifying selection between populations that inhabit different environments can promote lineage divergence within species and ultimately drive speciation. The mitochondrial genome (mitogenome) encodes essential proteins of the oxidative phosphorylation (OXPHOS) system and can be a strong target for climate-driven selection (i.e., associated with inhabiting different climates). We investigated whether Pleistocene climate changes drove mitochondrial selection and evolution within Australian birds. First, using phylogeographic analyses of the mitochondrial ND2 gene for 17 songbird species, we identified mitochondrial clades (mitolineages). Second, using distance-based redundancy analyses, we tested whether climate predicts variation in intraspecific genetic divergence beyond that explained by geographic distances and geographic position. Third, we analysed 41 complete mitogenome sequences representing each mitolineage of 17 species using codon models in a phylogenetic framework and a biochemical approach to identify signals of selection on OXPHOS protein-coding genes and test for parallel selection in mitolineages of different species existing in similar climates. Of 17 species examined, 13 had multiple mitolineages (range: 2-6). Climate was a significant predictor of mitochondrial variation in eight species. At least two amino acid replacements in OXPHOS complex I could have evolved under positive selection in specific mitolineages of two species. Protein homology modelling showed one of these to be in the loop region of the ND6 protein channel and the other in the functionally critical helix HL region of ND5. These findings call for direct tests of the functional and evolutionary significance of mitochondrial protein candidates for climate-associated selection.

Molecular systematics of the North American chub genus Macrhybopsis(Teleostei: Cyprinidae).

The North American fish genus Macrhybopsis (Teleostei: Cyprinidae) as presently conceived comprises 12 species and occurs in much of interior eastern North America. Variation in the mitochondrial ND2 gene and the nuclear S7 intron 1 reveal conflicting gene-tree relationships for deeper nodes, which are assumed to represent past introgression and heterospecific mitochondrial fixation. The results support monophyly for the wide-ranging M. aestivalis complex with successive sister relationships to M. gelida, M. meeki, and M. storeriana. The current species-level taxonomy of Macrhybopsis is generally supported. Species status is supported for the morphologically distinct M. australis and M. tetranema, both of which are genetically introgressed by M. hyostoma. The results agree with previous suggestions that the wide-ranging M. hyostoma harbors cryptic species. Similar crypticity is indicated for the poorly sampled M. storeriana; a sample from the Pearl River shows 8% ND2 divergence from two Mississippi River populations. Within the M. aestivalis complex, there are only two examples of geographic overlap among mtDNA phylogroups. One involves co-occurrence of the highly divergent M. marconis and M. cf. hyostoma, and the other is the detection of the apparently anthropogenic occurrence of mitochondrial DNA from a Red River form, either M. cf. hyostoma or M. australis, in the Cimarron River of the Arkansas River basin.