Indistinguishable Species Research Articles

Genetic differentiation between sandfly populations of Phlebotomus chinensis and Phlebotomus sichuanensis (Diptera: Psychodidae) in China inferred by microsatellites

BackgroundPhlebotomus chinensis is a primary vector of visceral leishmaniasis; it occurs in various biotopes with a large geographical distribution, ranging from Yangtze River to northeast China. Phlebotomus sichuanensis, a species closely related to P. chinensis in high altitude regions, has a long term disputation on its taxonomic status. Both species occur in the current epidemic regions and are responsible for the transmission of leishmaniasis. Population genetic analysis will help to understand the population structure and infer the relationship for morphologically indistinguishable cryptic species. In this study, microsatellite markers were used for studying the genetic differentiation between P. chinensis and P. sichuanensis.MethodsSandflies were collected in 6 representative localities in China in 2005-2009. Ten microsatellite loci were used to estimate population genetic diversity. The intra-population genetic diversity, genetic differentiation and effective population size were estimated.ResultsAll 10 microsatellite loci were highly polymorphic across populations, with high allelic richness and heterozygosity. Hardy-Weinberg disequilibrium was found in 23 out of 60 (38.33%) comparisons associated with heterozygote deficits, which was likely caused by the presence of null allele and the Wahlund effect. Bayesian clustering analysis revealed three clusters. The cluster I included almost all specimens in the sample SCD collected at high altitude habitats in Sichuan. The other two clusters were shared by the remaining 5 populations, SCJ in Sichuan, GSZ in Gansu, SXL and SXX in Shaanxi and HNS in Henan. The diversity among these 5 populations was low (FST = -0.003-0.090) and no isolation by distance was detected. AMOVA analysis suggested that the variations were largely derived from individuals within populations and among individuals. Consistently, the analysis of ribosomal DNA second internal transcribed spacer (ITS2) sequence uncovered three types of variants, which corresponded with the three gene pools revealed by microsatellites.ConclusionsThe data suggested that the SCD population carried a distinct gene pool, which was differentiated from the other populations. The high altitude ecological habitats, distinctive ITS2 and herein divergence inferred by microsatellite loci support the species status of P. sichuanensis. The P. chinensis populations did not have a significant divergence from each another.

Evidence for horizontal transmission of secondary endosymbionts in the Bemisia tabaci cryptic species complex.

Bemisia tabaci (Hemiptera: Aleyrodidae) is a globally distributed pest composed of at least 34 morphologically indistinguishable cryptic species. At least seven species of endosymbiont have been found infecting some or all members of the complex. The origin(s) of the associations between specific endosymbionts and their whitefly hosts is unknown. Infection is normally vertical, but horizontal transmission does occur and is one way for new infections to be introduced into individuals. The relationships between the different members of the cryptic species complex and the endosymbionts have not been well explored. In this study, the phylogenies of different cryptic species of the host with those of their endosymbionts were compared. Of particular interest was whether there was evidence for both coevolution and horizontal transmission. Congruence was observed for the primary endosymbiont, Portiera aleyrodidarum, and partial incongruence in the case of two secondary endosymbionts, Arsenophonus and Cardinium and incongruence for a third, Wolbachia. The patterns observed for the primary endosymbiont supported cospeciation with the host while the patterns for the secondary endosymbionts, and especially Wolbachia showed evidence of host shifts and extinctions through horizontal transmission rather than cospeciation. Of particular note is the observation of several very recent host shift events in China between exotic invader and indigenous members of the complex. These shifts were from indigenous members of the complex to the invader as well as from the invader to indigenous relatives.

Population variability of Bemisia tabaci (Genn.) in different hosts

The silverleaf whitefly, Bemisia tabaci (Genn.) (Hemiptera: Aleyrodidae), is a cryptic species complex that contains some of the most damaging pests in tropical and subtropical regions. Recent studies have indicated that this complex is composed of at least 24 distinct and morphologically indistinguishable species that mainly differ in their ability to transmit phytoviruses, adapt to hosts, and induce physiological changes in certain hosts. The importance of this species has been increasing worldwide, because it serves as a phytovirus vector, particularly for geminiviruses, in economically important crops. Here, we aimed to examine the population variability of B. tabaci populations inhabiting 6 agricultural crops grown in 5 regions of Brazil and 1 region of the USA; BRrep [Brasília (DF, Brazil) - cabbage], ILsoj [Urbana (IL/USA) - soybean], BJabo [Bom Jesus da Lapa (BA, Brazil) - pumpkin], CPsoj [Campinas (SP, Brazil) - soybean], UBman [Ubatuba (SP, Brazil) - cassava], and PEmel [Petrolina (PE, Brazil) - melon]. Thirteen polymorphic loci with 50 alleles were observed, with an average of 2.37 (range: 2.00-2.91) alleles per population. The UBman and PEmel B. tabaci populations were the most differentiated, which was probably caused by insect adaptation to the host plant and the use of insecticides. A 33.87% inter-population variation was observed, indicating that microsatellites may be used to measure differentiation among these B. tabaci populations. Based on the comparison of microsatellites in the current study, only the Middle East-Asia Minor 1 population of B. tabaci was found in the six populations.

Will the Real Bemisia tabaci Please Stand Up?

Since Panayiotis Gennadius first identified the whitefly, Aleyrodes tabaci in 1889, there have been numerous revisions of the taxonomy of what has since become one of the world's most damaging insect pests. Most of the taxonomic revisions have been based on synonymising different species under the name Bemisia tabaci. It is now considered that there is sufficient biological, behavioural and molecular genetic data to support its being a cryptic species complex composed of at least 34 morphologically indistinguishable species. The first step in revising the taxonomy of this complex involves matching the A. tabaci collected in 1889 to one of the members of the species complex using molecular genetic data. To do this we extracted and then amplified a 496 bp fragment from the 3′ end of the mitochondrial DNA cytochrome oxidase one (mtCOI) gene belonging to a single whitefly taken from Gennadius' original 1889 collection. The sequence identity of this 123 year-old specimen enabled unambiguous assignment to a single haplotype known from 13 Mediterranean locations across Greece and Tunisia. This enabled us to unambiguously assign the Gennadius A. tabaci to the member of the B. tabaci cryptic species complex known as Mediterranean or as it is commonly, but erroneously referred to, as the ‘Q-biotype’. Mediterranean is therefore the real B. tabaci. This study demonstrates the importance of matching museum syntypes with known species to assist in the delimitation of cryptic species based on the organism's biology and molecular genetic data. This study is the first step towards the reclassification of B. tabaci which is central to an improved understanding how best to manage this globally important agricultural and horticultural insect pest complex.

Spatial and host‐plant partitioning between coexisting Bemisia tabaci cryptic species in Tunisia

AbstractThe whitefly Bemisia tabaci is a species complex including at least 24 morphologically indistinguishable species among which the Mediterranean (Med) and Middle East‐Asia Minor I (MEAMI) species containing the biotypes commonly known as Q and B, respectively. These B and Q biotypes (hereafter referred to as MEAMI and Med species) are the most invasive agricultural pests of the B. tabaci complex worldwide. The spread of MEAMI and more recently of Med species into regions already invaded by other B. tabaci populations has been frequently seen to lead to their displacement by Med species. In Tunisia, in contrast to usual observations in the Mediterranean basin, Med and MEAMI species have been seen to co‐occur in the main crop producing regions. Based on fine population genetics and field spatial distribution analyses, we found that the co‐existence of these two interacting species was based on habitat partitioning including spatial and host‐plant partitioning. Although they co‐occurred at larger spatial scales, they excluded one another at sample scale. We observed neither spatial overlapping nor hybridization between MEAMI and Med B. tabaci. Vegetable crops were the main hosts for MEAMI specimens while 99.1% of the B. tabaci collected on the ornamental, Lantana camara, were Med specimens. Different patterns of genetic diversity were observed between the two species, as well as among Med specimens sampled on the ornamental versus vegetables, with the highest genetic diversity found in Med B. tabaci sampled on L. camara. These findings lead us to focus our discussion on the role played by lantana, human pressure, and competition, in the spatial and genetic patterns observed in the whitefly B. tabaci.

Cryptic fungal species unmasked

The word cryptic is derived from the Greek adjective krupt�s which means hidden. Morphologically indistinguishable species that have been revealed by molecular phylogenetic methods, and 37ultimately only recognised by their DNA sequences, are referred to as cryptic species. The importance of cryptic species for plant pathologists is that they may have significant differences in the severity of diseases they cause, host range and geographic distribution. It is these differences that are of concern to many biosecurity agencies, particularly in Australia.

Candida “Psilosis” – electromigration techniques and MALDI-TOF mass spectrometry for phenotypical discrimination

Biofilm-positive strains of Candida parapsilosis are the second most common yeasts responsible for bloodstream infections. This pathogen is difficult to identify by standard methods from other phenotypically indistinguishable species, biofilm-negative Candida orthopsilosis and biofilm-positive Candida metapsilosis. From a medical point of view, important information is especially whether the strains form biofilm. The biofilm formation enables yeast to colonize artificial surfaces thereby protecting the yeast cell against antifungal agents. The commonly used genotypic methods including different modifications of the polymerase chain reaction have some disadvantages. Therefore, a rapid and reliable method able to identify phenotypically indistinguishable C. "psilosis" species is still of great interest. In this study, the four well-established analytical techniques: gel isoelectric focusing, sodium dodecyl sulfate polyacrylamide gel electrophoresis, two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, were applied in order to discriminate C. "psilosis" species. The ability of these techniques to differentiate between biofilm-positive and biofilm-negative strains was further investigated. Our results have revealed that the proposed methods, especially matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of intact yeast cells, can be used as the efficient tools for discrimination and identification of the phenotypically indistinguishable microorganisms.

Evolution of Anopheles gambiae in Relation to Humans and Malaria

The closely related and morphologically indistinguishable mosquito species in the Afrotropical Anopheles gambiae complex differ dramatically in their contribution to malaria transmission, ranging from major vectors through minor or locally important vectors and nonvectors. Radiation of the A. gambiae complex and ongoing diversification within its nominal species appears to be a product of recent and rapid adaptation to environmental heterogeneities, notably those of anthropogenic origin. Polytene chromosome and genomic analyses suggest that paracentric chromosomal inversions and possibly other low-recombination regions have played instrumental roles in this process by facilitating ecotypic differentiation both within and across semipermeable species boundaries. Forthcoming complete genome sequences from several members of the A. gambiae complex will provide powerful tools to accelerate ongoing investigation of how genetic diversification of populations and species has shaped behavioral and physiological traits, such as vector competence, that bear on vectorial importance.

Use of consensus sequences and genetic networks to identify members of the Bemisia tabaci cryptic species complex in Egypt and Syria

AbstractThe sweet potato whitefly, Bemisia tabaci (Gennadius), is a cryptic species complex composed of at least 24 different morphologically indistinguishable species. The considerable differences in the pest status across the complex, and the ability of some to develop resistance to, insecticides make awareness of their identity critical in terms of developing effective control measures. Previously, phylogenetic reconstructions have been used to identify different B. tabaci, but this approach is no longer necessary because of the existence of mitochondrial cytochrome oxidase one consensus sequences for each of the known species. We therefore use these consensus sequences to determine the identities of the members of the complex in Syria and Egypt and then used genetic networks to reveal the pattern of their genetic relatedness to other haplotypes in the species to which they were assigned. The results showed the presence of three species in Syria, AsiaII 1, Middle East–Asia Minor 1 (this equates to the global invader known commonly as the B biotype) and Mediterranean (this equates to the global invader known commonly as the Q biotype). Egypt was shown to have two cryptic species, Middle East–Asia Minor 1 and Mediterranean. In Syria, Middle East–Asia Minor 1 was found around Damascus only (south‐west Syria), while Mediterranean was found throughout Aleppo (northern Syria) and Hama (north central Syria). AsiaII 1 was found around Hims (south central Syria) and Damascus (south‐western Syria). In Egypt, Mediterranean was found in Cairo and Ismailia (central Upper Egypt), while Middle East–Asia Minor 1 was found in the remaining all parts of Upper Egypt, Suez, North Sinai, Port Said, Dakahila, Behera and Alexandria which cover the main agricultural zone of Egypt. Genetic relatedness of Syrian and Egyptian populations with each other and with rest of world is also discussed.

First Report of Cercospora apii Leaf Spot on Capsicum chinense in Brazil.

In Brazil, Capsicum chinense Jacq. is the predominant species of commercial hot peppers because of its popular citrus-like aroma and adaptability to different soils and climates (4). In June 2010, 30 samples of C. chinense with severe leaf spot were collected from a field in the city of Viçosa, state of Minas Gerais, Brazil. Symptoms were observed on leaves, calyxes, fruits, and stems on most of the plants found in the area. On leaves, symptoms included amphigenous lesions that were initially circular to ellipsoid, 1 to 5 mm in diameter, whitish to tan in the center, and surrounded by a dark brown or reddish purple border. Lesions coalesce and turned necrotic with age. A fungus isolated from the lesions matched well with the description of Cercospora apii Fresen. It formed erumpent stromata that were dark brown and spherical to irregular; fascicule conidiophores were clear brown or pale, straight or curved, unbranched, geniculate, 22.5 to 80 × 5 to 7.5 μm, 0 to 3 septate, subtruncate apex; and conidia were solitary, hyaline to subhyaline, filiform, base truncate, tip acute, straight to curved, 12.5 to 140 × 3.5 to 5 μm, and 0 to 11 septate (1,2). A sample was deposited in the herbarium of the Universidade Federal de Viçosa, Minas Gerais, Brazil (VIC 31415). Identity was confirmed by amplifying part of the calmodulin gene with species-specific primers CercoCal-apii and CercoCal-R (3) of fungal DNA from a single-spore culture. In amplification reaction, initial denaturation step was done at 94°C for 5 min, followed by 40 cycles of denaturation at 94°C (30 s), annealing at 56°C (30 s), and elongation at 72°C (30 s). Primers CercoCal-apii and CercoCal-R amplified a single DNA product of 176 bp, and coupled with the morphological characteristics, confirmed the identity of the fungus as Cercospora apii. To check pathogenicity, a 6-mm-diameter plug of the isolate was removed from the expanding edge of a 21-day-old culture grown on potato dextrose agar (PDA) and placed in contact with the adaxial face of the leaves of 8-week-old C. chinense grown in 2-liter plastic pots with soil substrate. Six plants, one per pot, were inoculated with the isolate and six plants were inoculated with the fungus-free PDA plug. Inoculated plants were maintained in a moist chamber for 24 h and then subsequently kept in a greenhouse at 26°C. Leaf spot was observed in all inoculated plants 15 days after inoculation and symptoms were similar to those expressed in the field. The fungus was reisolated from the inoculated plants and matched well with the description of Cercospora apii. All fungus-free PDA inoculated plants remained healthy. Cercospora apii comprises a complex of 281 morphologically indistinguishable species that can infect an extremely wide host range (2). To our knowledge, this pathogen has the potential to cause significant damage to the hot pepper industry of Brazil.

High‐throughput molecular identification of fish eggs using multiplex suspension bead arrays

The location and abundance of fish eggs provide information concerning the timing and location of spawning activities and can provide fishery-independent estimates of spawning biomass. However, the full value of egg and larval surveys is severely restricted because many species' eggs and larvae are morphologically similar, making species-level identification difficult. Recent efforts have shown that nearly all species of fish may be identified by mitochondrial DNA (mtDNA) sequences (e.g. via 'DNA barcoding'). By taking advantage of a DNA barcode database, we have developed oligonucleotide probes for 23 marine fish species that produce pelagic eggs commonly found in California waters. Probes were coupled to fluorescent microspheres to create a suspension bead array. Biotin-labelled primers were used to amplify the mitochondrial cytochrome oxidase subunit I (COI) and 16S ribosomal rRNA genes from individual fish eggs. The amplicons were then hybridized to the bead array, and after the addition of a reporter fluorophore, samples were analysed by flow cytometry with Luminex 100 instrumentation. Probes specifically targeted eggs that are abundant and/or from morphologically indistinguishable species pairs. Results showed that the 33 different probes designed for this study accurately identified all samples when PCR was successful. Suspension bead arrays have a number of benefits over other methods of molecular identification; these arrays permit high multiplexing, simple addition of new probes, high throughput and lower cost than DNA sequencing. The increasing availability of DNA barcode data for numerous fish faunas worldwide suggests that bead arrays could be developed and widely used for fish egg, larval and tissue identifications.

Separation of phenotypically indistinguishable Candida species, C. orthopsilosis, C. metapsilosis and C. parapsilosis, by capillary electromigration techniques

At the current state of laboratory diagnostics, methods for fast identification of phenotypically indistinguishable species are difficult or inaccurate. An example is represented by Candida parapsilosis, which is the second most common yeast species isolated from bloodstream infections. C. parapsilosis comprises a complex of three genetically distinct groups. Genotypes II and III have been designated as the separate species Candida orthopsilosis and Candida metapsilosis, phenotypically indistinguishable. The considerable genetic variability of these newly described yeasts species has caused difficulties in the development of molecular techniques for their precise identification. Similarly, the detection of biofilm formation, which is considered as an important yeast virulence factor, is accompanied by difficulties. In this study we optimize the first precise and reproducible method for the separation and possible identification of C. orthopsilosis, C. metapsilosis and C. parapsilosis as well as the detection of their ability to form biofilm. The method is based on capillary isoelectric focusing and capillary electrophoresis with UV detection. In capillary isoelectric focusing, very narrow pH gradients were established. With such gradients, differences in isoelectric points of biofilm-negative and biofilm-positive species calculated from the migration times of the selected p I markers were below 0.03 p I units. In the capillary zone electrophoresis narrow zones of the cells of Candida species were detected with sufficient resolution. The values of the isoelectric point and the migration velocities of the examined species were independent on the origin of the tested strains. Capillary isoelectric focusing was examined also for the separation and detection of the cultivated biofilm-negative C. parapsilosis in the blood serum.

Analysis of the evolutionary forces shaping mitochondrial genomes of a Neotropical malaria vector complex

Many vectors of human malaria belong to complexes of morphologically indistinguishable cryptic species. Here we report the analysis of the newly sequenced complete mitochondrial DNA molecules from six recognized or putative species of one such group, the Neotropical Anopheles albitarsis complex. The molecular evolution of these genomes had been driven by purifying selection, particularly strongly acting on the RNA genes. Directional mutation pressure associated with the strand-asynchronous asymmetric mtDNA replication mechanism may have shaped a pronounced DNA strand asymmetry in the nucleotide composition in these and other Anopheles species. The distribution of sequence polymorphism, coupled with the conflicting phylogenetic trees inferred from the mitochondrial DNA and from the published white gene fragment sequences, indicates that the evolution of the complex may have involved ancient mtDNA introgression. Six protein coding genes (nad5, nad4, cox3, atp6, cox1 and nad2) have high levels of sequence divergence and are likely informative for population genetics studies. Finally, the extent of the mitochondrial DNA variation within the complex supports the notion that the complex consists of a larger number of species than until recently believed.

Bemisia tabaci: A Statement of Species Status

Bemisia tabaci has long been considered a complex species. It rose to global prominence in the 1980s owing to the global invasion by the commonly named B biotype. Since then, the concomitant eruption of a group of plant viruses known as begomoviruses has created considerable management problems in many countries. However, an enduring set of questions remains: Is B. tabaci a complex species or a species complex, what are Bemisia biotypes, and how did all the genetic variability arise? This review considers these issues and concludes that there is now sufficient evidence to state that B. tabaci is not made up of biotypes and that the use of biotype in this context is erroneous and misleading. Instead, B. tabaci is a complex of 11 well-defined high-level groups containing at least 24 morphologically indistinguishable species.

Incomplete concerted evolution and reproductive isolation at the rDNA locus uncovers nine cryptic species within Anopheles longirostrisfrom Papua New Guinea

BackgroundNuclear ribosomal DNA (rDNA) genes and transcribed spacers are highly utilized as taxonomic markers in metazoans despite the lack of a cohesive understanding of their evolution. Here we follow the evolution of the rDNA second internal transcribed spacer (ITS2) and the mitochondrial DNA cytochrome oxidase I subunit in the malaria mosquito Anopheles longirostris from Papua New Guinea (PNG). This morphospecies inhabits a variety of ecological environments indicating that it may comprise a complex of morphologically indistinguishable species. Using collections from over 70 sites in PNG, the mtDNA was assessed via direct DNA sequencing while the ITS2 was assessed at three levels - crude sequence variation through restriction digest, intragenomic copy variant organisation (homogenisation) through heteroduplex analysis and DNA sequencing via cloning.ResultsGenetic evaluation of over 300 individuals revealed that A. longirostris comprises eight ITS2 PCR-RFLP genotypes and nine ITS2 heteroduplex genotypes showing distinct copy variant organization profiles after PCR amplification. Seven of these nine genotypes were found to be sympatric with other genotypes. Phylogenetic analysis of cloned ITS2 PCR products and mtDNA COI confirmed all nine clades with evidence of reproductive isolation at the rDNA locus. Compensatory base changes in the ITS2 secondary structure or in pseudoknots were absent when closely related species were assessed. Individuals from each ITS2 genotype showed the same copy variant heteroduplex profile suggesting that the rDNA array is fixed within each genotype.ConclusionThe centromere-proximal position of the rDNA array in Anopheles mosquitoes has probably reduced interchromosomal recombination leaving intrachromosomal events responsible for the observed pattern of concerted evolution we see in these mosquitoes. The stability of these intragenomic ITS2 copy variants within individuals and interbreeding populations suggests that rDNA is moving as a single evolutionary unit through natural populations to fixation and has provided a complementary diagnostic tool to the restriction digest for studying genetic discontinuities and species boundaries. In this, the utility of the ITS2 as a universal taxonomic marker is probably contingent on several factors pertaining to spacer dimensions and the genomic location of the rDNA array with respect to recombination and proximity to regions potentially under selection.

DNA barcoding provides distinction between Radix Astragali and its adulterants

Based on variable nuclear and/or organellar DNA sequences among vastly divergent species as well as morphologically indistinguishable species, DNA barcoding is widely applicable in species identification, biodiversity studies, forensic analyses, and authentication of medicinal plants. The roots of Astragalus membranaceus and A. membranaceus var. mongholica are commonly used as Radix Astragali in several Asian countries, including China, Japan, and Korea. However, in addition to the two species recorded in the Chinese Pharmacopoeia, there are twenty-three species from different genera including Astragalus, Oxytropis, Hedysarum, and Glycyrrhiza, which have been used as adulterants not only in trading markets but also by the herbal medicine industry. Therefore, a simple, reliable, and accurate classification method is important for distinguishing authentic Radix Astragali from its adulterants. In this study, we acquired data for 37 samples from four related genera within the family Fabaceae. Then we compared four candidate DNA barcoding markers using ITS, matK, rbcL, and coxI sequences from nuclear, chloroplast, and mitochondrial genomes, all commonly used for plants to identify genetic variations among genera, intraspecies, and interspecies. We observed higher divergences among genera and interspecies for ITS, which have the average Kimura 2-parameter distances of 4.5% and 14.1%, respectively, whereas matK was found to have sufficient divergence at the intraspecific level. Moreover, two indels detected in the matK sequence are useful for PCR studies in distinguishing Radix Astragali from its adulterants. This study suggests that the combined barcoding regions of ITS and matK are superior barcodes for Radix Astragali and further studies should focus on evaluating the applicability and accuracy of such combined markers for a wide range of traditional Chinese herbs.

Good species behaving badly: Non-monophyly of black fly sibling species in the Simulium arcticum complex (Diptera: Simuliidae)

Mitochondrial based phylogenetic reconstructions often show deviations from species-level monophyly. We used the Simulium arcticum species complex (Diptera: Simuliidae) as a model system for interpreting non-monophyly in light of chromosomal data supporting species status of siblings. For cytogenetic identification of morphologically indistinguishable black fly sibling species, larvae must be preserved in Carnoy’s solution, a fixative known to degrade DNA. Consequently, we reconstructed phylogenetic relationships based on 12S, COII, cyt b, and ITS-1 gene sequences obtained from larvae sampled from presumed taxon-pure localities. As species composition at ‘taxon-pure’ sites may have changed at the time of sampling, we performed a second study that aimed to: (1) assess phylogenetic relationships among cytologically verified members of the S. arcticum species complex using COI and COII gene sequences; (2) determine whether useable genetic information could be gleaned from Carnoy’s fixed specimens; and (3) determine the extent to which Carnoy’s fixative degrades DNA over time. We consistently obtained genetic data from material stored in Carnoy’s solution for two to threemonths. Genetic analysis of samples fixed in Carnoy’s solution for up to sixyears indicates that larvae preserved for a maximum of five years can provide useable information for molecular analysis. Our preliminary and cytologically confirmed phylogenetic analyses demonstrate that mitochondrial DNA fails to resolve species-level monophyly of chromosomally distinct S. arcticum taxa. As results of analyses based on cytologically verified larvae mirror those of our preliminary study, we rule out imperfect taxonomy as the reason for species-level non-monophyly. Although we cannot confidently reject either inadequate phylogenetic information or incomplete lineage sorting as the cause of non-monophyly, the sharing of alleles between sympatric siblings suggests introgressive hybridization between taxa. We conclude that the patterns present in the S. arcticum phylogeny likely represent the initial stages of chromosome based sibling speciation.

Refined Global Analysis of Bemisia tabaci (Hemiptera: Sternorrhyncha: Aleyrodoidea: Aleyrodidae) Mitochondrial Cytochrome Oxidase 1 to Identify Species Level Genetic Boundaries

Abstract Identifying species boundaries within morphologically indistinguishable cryptic species complexes is often contentious. For the whitefly Bemisia tabaci (Gennadius) (Hemiptera: Sternorrhyncha: Aleyrodoidea: Aleyrodidae), the lack of a clear understanding about the genetic limits of the numerous genetic groups and biotypes so far identified has resulted in a lack of consistency in the application of the terms, the approaches used to apply them and in our understanding of what genetic structure within B. tabaci means. Our response has been to use mitochondrial gene cytochrome oxidase one to consider how to clearly and consistently define genetic separation. Using Bayesian phylogenetic analysis and analysis of sequence pairwise divergence we found a considerably higher number of genetic groups than had been previously determined with two breaks in the distribution, one at 11% and another at 3.5%. At >11% divergence, 11 distinct groups were resolved, whereas at >3.5% divergence 24 groups were identified. Consensus sequences for each of these groups were determined and were shown to be useful in the correct assignment of sequences of unknown origin. The 3.5% divergence bound is consistent with species level separations in other insect taxa and suggests that B. tabaci is a cryptic species composed of at least 24 distinct species. We further show that the placement of Bemesia atriplex (Froggatt) within the B. tabaci in group adds further weight to the argument for species level separation within B. tabaci. This new analysis, which constructs consensus sequences and uses these as a standard against which unknown sequences can be compared, provides for the first time a consistent means of identifying the genetic bounds of each species with a high degree of certainty.

Reproductive incompatibility among genetic groups of Bemisia tabaci supports the proposition that the whitefly is a cryptic species complex

The worldwide distribution and extensive genetic diversity of the whitefly Bemisia tabaci has long been recognized. However, whether B. tabaci is a complex species or a species complex has been a subject of debate. Recent phylogenetic analyses suggest that B. tabaci is a cryptic species complex composed of at least 24 morphologically indistinguishable species. Here, we conducted crossing experiments and demonstrated reproductive incompatibility among three of the 24 putative species. Our data and those of previously reported crossing experiments among various putative species of B. tabaci were collated to reveal the pattern of reproductive isolation. The combined results provide strong support to the proposition that B. tabaci is a cryptic species complex.

Using the molecular toolbox to compare harmful algal blooms in upwelling systems

Harmful algal blooms (HABs) are now generally recognized as occurring over a wide range of habitats from oligotrophic to hypernutrified, and appear to be expanding globally. Unlike many other ecosystems impacted by HABs, upwelling systems worldwide share a common set of physical parameters and are likely to respond similarly, regardless of locale. The Core Research Project on HABs in Upwelling Systems, a component of the scientific programme on the Global Ecology and Oceanography of Harmful Algal Blooms (GEOHAB), promotes a comparative approach to identify the similarities and differences in the manifestation of HAB events in these systems. As applied to the goals of this programme, molecular techniques are a powerful suite of tools for HAB species identification, for determining genetic similarity within morphologically indistinguishable species, and ultimately, for assessing spatial and temporal patterns in ecophysiological responses in these upwelling systems. Knowledge of HAB organisms will be enhanced by comparing and contrasting the responses of these organisms in similar upwelling regions. Here, we provide an update on the availability of molecular and genetic tools for comparative HAB programmes in upwelling systems, focusing on four broad applications: cell enumeration and identification, molecular phylogenetics, functional/comparative genomics, and applications of high throughput sequencing methods. We highlight the rapid evolution, the promise, and the potential pitfalls, of the molecular toolbox, focusing on specific examples of how scientists and resource managers currently apply these methods. Specific examples are developed using relevant case studies from the California, Benguela and Iberian systems. We summarise by providing a synthesis of future research directions and goals that would be particularly relevant to advancing the comparative method for HAB genetics with an emphasis on upwelling systems.