Crossing experiments and behavioral observations reveal reproductive incompatibility among three putative species of the whitefly Bemisia tabaci

Abstract The whitefly Bemisia tabaci has a global distribution and extensive genetic diversity. Recent phylogenetic analyses as well as crossing experiments suggest that B. tabaci is a complex composed of > 20 cryptic species, but more crossing studies are required to examine the reproductive compatibility among the putative species and thus further clarify the systematics of this species complex. We conducted crossing experiments and behavioral observations to investigate the reproductive compatibility between the Mediterranean, Asia II 3, and Asia II 1 putative species of B. tabaci collected from Zhejiang, China. Female progeny were never produced in inter-species crosses, demonstrating a lack of egg fertilization; while 55%–75% females were produced in all the intra-species treatments. Continuous behavioral observations showed that frequent courtship events occurred in both intra-species treatments and inter-putative species crosses. However, copulation events occurred only in the three intra-species treatments with one exception: that one copulation event occurred between Asia II 3 and Mediterranean in the crosses where two cohorts of females and males of different putative species were enclosed together in a small arena but were not allowed access to their intra-specific mates for a long period of time. These data demonstrated complete reproductive isolation between the Mediterranean, Asia II 3, and Asia II 1 putative species, and further showed that the isolation is due to lack of copulation. Demonstration of reproductive isolation between the Mediterranean and two indigenous putative species from China provides further evidence for the existence of cryptic species within the B. tabaci complex.     

The presence of six cryptic species of the whitefly Bemisia tabaci complex in China as revealed by crossing experiments

Abstract Recent phylogenetic analysis using mitochondrial cytochrome oxidase I (mtCOI) sequences of Bemisia tabaci worldwide indicates that the whitefly comprises at least 24 morphologically indistinguishable but genetically distinct cryptic species. While evidence of reproductive isolation has been reported for some of the putative species, more extensive crossing experiments are required to clarify the systematics of this species complex. In this study, we established laboratory cultures for six putative species of B. tabaci collected in China. We conducted 22 inter‐species crosses among the six putative species. The data and those reported previously were collated, and the combined dataset covered all the 30 possible inter‐species crosses among the six putative species. Intra‐species controls always produced female and male progeny and the proportions of females in the first generation (F1) ranged from 56% to 70%. However, in inter‐species crosses female progeny were rarely produced, and the few F1 females produced in four of the 30 inter‐species crosses were either sterile or significantly weaker in viability. These results demonstrate a pattern of complete reproductive isolation among the six putative species and show that they are six cryptic species in the B. tabaci complex.     

Evidence of gene flow between sympatric populations of the Middle East‐Asia Minor 1 and Mediterranean putative species of Bemisia tabaci

Bemisia tabaci is a complex of putative species that exhibit a strong geographical pattern. Crossing experiments have revealed various degrees of reproductive isolation between these nascent species, ranging between fertile first‐generation hybrids (F1) and no F1 at all. However, the relevance of these results under natural conditions is generally not known. The worldwide invasion of the putative species Middle East‐Asia Minor 1 (MEAM1) has caused secondary contacts between allopatric species, which in turn provide an opportunity to detect potential hybrids in nature. A total of 346 female B. tabaci were collected in 2003 and 2005 in the North East of Morocco and assigned to MEAM1 (119), Mediterranean (Med) (225) and a new putative species (2) using mitochondrial cytochrome oxidase (mtCOI) gene sequences. MEAM1 and Med individuals were characterized at seven microsatellite loci. MEAM1 and Med were found to be sympatric in 11 of 12 samples (6 fields/year). As previously reported from Spain, MEAM1 frequency decreased over time. The genetic data are consistent with a recent introduction of MEAM1. A Bayesian clustering analysis (Structure ) distinguished two groups, which were 100% consistent with the mtCOI groups. From several lines of evidence, two individuals were identified as hybrids. Assignment profiles using NewHybrids and allele composition indicated that they were not F1 hybrids. The results are discussed in relation to the secondary endosymbiont infection status determined on a sample of individuals, and the contrasting outcomes of the reported crossing experiments between MEAM1 and Med.     

Genetic identification of members of the Bemisia tabaci cryptic species complex from South Africa reveals native and introduced haplotypes

The whitefly Bemisia tabaci cryptic species complex contains some important agricultural pest and virus vectors. Members of the complex have become serious pests in South Africa (SA) because of their feeding habit and their ability to transmit begomovirus species. Despite their economic importance, studies on the biology and distribution of B. tabaci in SA are limited. To this end, a survey was made to investigate the diversity and distribution of B. tabaci cryptic species in eight geographical locations (provinces) in SA, between 2002 and 2009, using the mitochondrial cytochrome oxidase I (mtCOI) sequences. Phylogenetic analysis revealed the presence of members from two endemic sub‐Saharan Africa (SSAF) subclades coexisting with two introduced putative species. The SSAF‐1 subclade includes cassava host‐adapted B. tabaci populations, whereas the whiteflies collected from cassava and non‐cassava hosts formed a distinct subclade, referred to as SSAF‐5, and represent a new subclade among previously recognized southern Africa clades. Two introduced cryptic species, belonging to the Mediterranean and Middle East–Asia minor 1 clades, were identified and include the B and Q types. The B type showed the widest distribution, being present in five of the eight provinces explored in SA, infesting several host plants and predominating over the indigenous haplotypes. This is the first report of the occurrence of the exotic Q type in SA alongside the more widely distributed B type. Furthermore, mtCOI PCR‐RFLP was developed for the SA context to allow rapid discrimination between the B, Q and SSAF putative species. The capacity to manage pests and disease effectively relies on knowledge of the identity of the agents causing the damage. Therefore, this study contributes to the understanding of South African B. tabaci species diversity, information needed for the development of knowledge‐based disease management practices.     

Genetic diversity and geographic distribution of Bemisia tabaci and Trialeurodes vaporariorum in Costa Rica

The tobacco whitefly Bemisia tabaci (Gennadius) cryptic species complex and of the greenhouse whitefly Trialeurodes vaporariorum (Westwood) are extensively reported as destructive pests in vegetable crops worldwide. A survey was conducted in 2011 and 2012 to determine the occurrence and genetic diversity present in the populations of these whiteflies in the major vegetable production areas of Costa Rica. Insect samples were collected from sweet pepper (Capsicum annuum L.), tomato (Solanum lycopersicum L.), common bean (Phaseolus vulgaris L.) and weeds present in commercial crops either in open field or greenhouse conditions. PCR‐RFLP analysis of mitochondrial cytochrome c oxidase subunit 1 gene (mtCOI) sequences of 621 whitefly individuals confirmed the presence of the Mediterranean (MED) type of the B. tabaci and of T. vaporariorum in most sampled regions. Also, individuals of the Middle East‐Asia Minor 1 (MEAM1) type of the B. tabaci were observed in low numbers. Contingency analyses based on type of crop, geographical region, whitefly species, year of collection and production system confirmed that T. vaporariorum was the most frequent species in vegetable production areas in Costa Rica, both in greenhouses and in open fields. B. tabaci MED is likely spreading to new areas of the country, whereas B. tabaci MEAM1 was mostly absent or rarely found. Comparisons of mtCOI sequences from B. tabaci individuals revealed the presence of four B. tabaci sequence haplotypes (named MED‐i, MED‐ii, MEAM1‐i, MEAM1‐xviii) in Costa Rica, three of them identical to B. tabaci haplotypes previously reported in the Western Hemisphere and other parts of the world. Analysis of sequences of T. vaporariorum individuals revealed a more complex population with the presence of 11 haplotypes, two of which were identical to T. vaporariorum sequences reported from other countries.     

At least two indigenous species of the Bemisia tabaci complex are present in Brazil

Bemisia tabaci is one of the most important global agricultural insect pests, being a vector of emerging plant viruses such as begomoviruses and criniviruses that cause serious problems in many countries. Although knowledge of the genetic diversity of B. tabaci populations is important for controlling this pest and understanding viral epidemics, limited information is available on this pest in Brazil. A survey was conducted in different locations of São Paulo and Mato Grosso states, and the phylogenetic relationships of B. tabaci individuals from 43 populations sampled from different hosts were analysed based on partial mitochondrial cytochrome oxidase 1 gene (mtCOI) sequences. According to the recently proposed classification of the B. tabaci complex, which employs the 3.5% mtCOI sequence divergence threshold for species demarcation, most of the specimens collected were found to belong to the Middle East‐Asia Minor 1 species, which includes the invasive populations of the commonly known B biotype, within the Africa/Middle East/Asia Minor high‐level group. Three specimens collected from Solanun gilo and Ipomoea sp. were grouped together and could be classified in the New World species that includes the commonly known A biotype. However, six specimens collected from Euphorbia heterophylla, Xanthium cavanillesii and Glycine maxima could not be classified into any of the 28 previously proposed species, although according to the 11% mtCOI sequence divergence threshold, they belong to the New World high‐level group. These specimens were classified into a new recently proposed species named New World 2 that includes populations from Argentina. Middle East‐Asia Minor 1, New World and New World 2 were differentiated by RFLP analysis of the mtCOI gene using TaqI enzyme. Taq I analysis in silico also differentiates these from Mediterranean species, thus making this method a convenient tool to determine population dynamics, especially critical for monitoring the presence of this exotic pest in Brazil.     

Performance on different host plants of an alien and an indigenous Bemisia tabaci from China

The capacity of the Middle East‐Asia Minor 1 putative species of the whitefly Bemisia tabaci (Gennadius) species complex, commonly referred to as the ‘B biotype’, to invade has often been linked to its presumed wider host range than the indigenous competitors. To determine whether this alien putative species and the indigenous Asia II 1 whitefly putative species, commonly referred to as the ‘ZHJ2 biotype’, differ in their ability to use different host plants, we compared their development, survival and reproduction on eight crop species/cultivars that are commonly cultivated in Zhejiang, China. Of the eight host plants tested, B performed substantially better than ZHJ2 on squash, tomato and tobacco, B and ZHJ2 preformed equally well on cotton and sweet potato, while ZHJ2 performed better than B on kidney bean and pepper. These results indicate that while B generally has a wider host range than many indigenous B. tabaci, an indigenous B. tabaci can perform as well as or better on some host plants. These results combined with the cropping patterns in Zhejiang suggested that the differential capacity to use various host plants between whitefly species is important in mediating the process of invasion by an alien whitefly species.     

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.     

Genetic structure of the whitefly Bemisia tabaci populations in Colombia following a recent invasion

The whitefly Bemisia tabaci (Gennadius) is one of the most important pests causing economic losses in a variety of cropping systems around the world. This species was recently found in a coastal region of Colombia and has now spread inland. To investigate this invasive process, the genetic structure of B. tabaci was examined in 8 sampling locations from 2 infested regions (coastal, inland) using 9 microsatellite markers and the mitochondrial COI gene. The mitochondrial analysis indicated that only the invasive species of the B. tabaci complex Middle East–Asia Minor 1 (MEAM 1 known previously as biotype B) was present. The microsatellite data pointed to genetic differences among the regions and no isolation by distance within regions. The coastal region in the Caribbean appears to have been the initial point of invasion, while the inland region in the Southwest showed genetic variation among populations most likely reflecting founder events and ongoing changes associated with climatic and topographical heterogeneity. These findings have implications for tracking and managing B. tabaci.     

Mitochondrial DNA variability and development of a PCR diagnostic test for populations of the whitefly Bemisia afer (priesner and hosny)

The whitefly, Bemisia afer (Hemiptera; Aleyrodidae), is emerging as a major agricultural pest. The current identification methods based on adult and pupal morphology are laborious and unreliable. A diagnostic polymerase chain reaction (PCR) protocol was developed for the first time in this study to discriminate B. afer from other whitefly species. Primers specific to mitochondrial cytochrome oxidase 1 gene (mtCOI) were designed to amplify a band of approx 650 bp. The PCR products were sequenced from B. afer samples collected from Malawi, Tanzania, Uganda, Zanzibar, and the United Kingdom. Phylogenetic analyses of mtCOI sequences and those of reference B. afer sequences clustered the African B. afer separately from the UK and Chinese populations and from other whitefly species. The African cluster was divided into two clades by parsimony and neighbor-joining methods. This indicates the existence of at least two genotypic clusters of B. afer, which are diverged by 0.8 to 3.2% nucleotide (nt) identities. Analysis of molecular variance indicated that these differences were the result of within population variation but were insufficient to identify discrete populations. Among the whitefly species used in the analysis, B. afer was equally dissimilar to Bemisia tabaci and Bemisia tuberculata (21.3–26.2% nt identities). As is the case for B. tabaci, these data show that mtCOI sequences are informative also for identifying B. afer variants, which lack distinguishing morphological features.     

Whole-genome single nucleotide polymorphism and mating compatibility studies reveal the presence of distinct species in sub-Saharan Africa Bemisia tabaci whiteflies

In sub-Saharan Africa cassava growing areas, two members of the Bemisia tabaci species complex termed sub-Saharan Africa 1 (SSA1) and SSA2 have been reported as the prevalent whiteflies associated with the spread of viruses that cause cassava mosaic disease (CMD) and cassava brown streak disease (CBSD) pandemics. At the peak of CMD pandemic in the late 1990s, SSA2 was the prevalent whitefly, although its numbers have diminished over the last two decades with the resurgence of SSA1 whiteflies. Three SSA1 subgroups (SG1 to SG3) are the predominant whiteflies in East Africa and vary in distribution and biological properties. Mating compatibility between SSA1 subgroups and SSA2 whiteflies was reported as the possible driver for the resurgence of SSA1 whiteflies. In this study, a combination of both phylogenomic methods and reciprocal crossing experiments were applied to determine species status of SSA1 subgroups and SSA2 whitefly populations. Phylogenomic analyses conducted with 26 548 205 bp whole genome single nucleotide polymorphisms (SNPs) and the full mitogenomes clustered SSA1 subgroups together and separate from SSA2 species. Mating incompatibility between SSA1 subgroups and SSA2 further demonstrated their distinctiveness from each other. Phylogenomic analyses conducted with SNPs and mitogenomes also revealed different genetic relationships among SSA1 subgroups. The former clustered SSA1-SG1 and SSA1-SG2 together but separate from SSA1-SG3, while the latter clustered SSA1-SG2 and SSA1-SG3 together but separate from SSA1-SG1. Mating compatibility was observed between SSA1-SG1 and SSA1-SG2, while incompatibility occurred between SSA1-SG1 and SSA1-SG3, and SSA1-SG2 and SSA1-SG3. Mating results among SSA1 subgroups were coherent with phylogenomics results based on SNPs but not the full mitogenomes. Furthermore, this study revealed that the secondary endosymbiont—Wolbachia—did not mediate reproductive success in the crossing assays carried out. Overall, using genome wide SNPs together with reciprocal crossings assays, this study established accurate genetic relationships among cassava-colonizing populations, illustrating that SSA1 and SSA2 are distinct species while at least two species occur within SSA1 species.     

Genetic groups and endosymbiotic microbiota of the Bemisia tabaci species complex in Japanese agricultural sites

The whitefly Bemisia tabaci is a cryptic species complex of at least 24 genetically distinct species. Thus far, one obligate and seven facultative symbiotic bacteria have been reported from the B. tabaci species complex. Both genetic groups and infected symbionts are extremely important to estimate the pest status of B. tabaci. In this study, we collected 340 whiteflies from 39 agricultural sites, covering an entire region of the B. tabaci habitat in Japan, and examined the genotypes and symbiont community composition at subspecies level. Use of the cleaved amplified polymorphic sequence technique and mitochondrial cytochrome oxidase subunit I gene sequencing detected five genetic groups: indigenous species JpL and Asia II 6, invasive species Middle East‐Asia Minor 1 (MEAM1) and Mediterranean Subclade Q1 (MED Q1), and a genetic group previously undetected in Asia, Mediterranean Subclade Q2 (MED Q2). The genetic groups exhibited characteristic infection statuses with regard to their facultative symbionts, as observed in other countries. The endosymbiotic microbiota of the Japanese MED Q1 was different from that in neighbouring countries, but similar to that in the French or Uruguayan MED Q1. These results may indicate that Japanese MED Q1 species have not invaded from neighbouring countries, but from distant countries by international transportation. All Japanese MED Q2 species were infected with Rickettsia, some of which are regarded as conferring a female‐biased sex ratio and fitness benefit on B. tabaci. The results suggest that MED Q2 may be prevalent in Japan and neighbouring countries.     

Occurrence of three genotypic clusters of Bemisia tabaci and the rapid spread of the B biotype in south India

The whitefly, Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae), is generally considered to have originated from the Indian subcontinent, although little information has so far been collected on the molecular diversity of populations present in this region. The genetic diversity of B. tabaci populations from Karnataka State, south India was analysed using the random amplified polymorphic DNA‐polymerase chain reaction (RAPD‐PCR) technique and partial mitochondrial cytochrome oxidase I (mtCOI) gene sequences (689 bases) of 22 selected samples. A total of 108 whitefly samples analysed by RAPD‐PCR produced 89 polymorphic bands, and cluster analyses grouped them according to their geographic origin into ‘north’ and ‘south’ Karnataka. Phylogenetic analysis of mtCOI gene sequences with reference B. tabaci sequences from other Asian countries divided them into three genotypic clusters. Each cluster was supported with high bootstrap values (82–100%) and the individuals belonging to each cluster shared high nucleotide identities (up to 100%). This indicated at least three distinct genotypes, apparently indigenous to India, which are also present in China, Malaysia, Nepal, Pakistan, and Thailand. These coexist with the B biotype, which was first reported in India in 1999, and has since spread rapidly to other states in south India. The B biotype was more common than the indigenous B. tabaci, in locations where it had been present for more than 2 years. This is reminiscent of the situation in the Americas during the early 1990s, where the B biotype replaced existing biotypes and caused unprecedented losses to agriculture.     

DNA Barcoding of Bemisia tabaci Complex (Hemiptera: Aleyrodidae) Reveals Southerly Expansion of the Dominant Whitefly Species on Cotton in Pakistan

Background

Although whiteflies (Bemisia tabaci complex) are an important pest of cotton in Pakistan, its taxonomic diversity is poorly understood. As DNA barcoding is an effective tool for resolving species complexes and analyzing species distributions, we used this approach to analyze genetic diversity in the B. tabaci complex and map the distribution of B. tabaci lineages in cotton growing areas of Pakistan.

Methods/Principal Findings

Sequence diversity in the DNA barcode region (mtCOI-5′) was examined in 593 whiteflies from Pakistan to determine the number of whitefly species and their distributions in the cotton-growing areas of Punjab and Sindh provinces. These new records were integrated with another 173 barcode sequences for B. tabaci, most from India, to better understand regional whitefly diversity. The Barcode Index Number (BIN) System assigned the 766 sequences to 15 BINs, including nine from Pakistan. Representative specimens of each Pakistan BIN were analyzed for mtCOI-3′ to allow their assignment to one of the putative species in the B. tabaci complex recognized on the basis of sequence variation in this gene region. This analysis revealed the presence of Asia II 1, Middle East-Asia Minor 1, Asia 1, Asia II 5, Asia II 7, and a new lineage “Pakistan”. The first two taxa were found in both Punjab and Sindh, but Asia 1 was only detected in Sindh, while Asia II 5, Asia II 7 and “Pakistan” were only present in Punjab. The haplotype networks showed that most haplotypes of Asia II 1, a species implicated in transmission of the cotton leaf curl virus, occurred in both India and Pakistan.

Conclusions

DNA barcodes successfully discriminated cryptic species in B. tabaci complex. The dominant haplotypes in the B. tabaci complex were shared by India and Pakistan. Asia II 1 was previously restricted to Punjab, but is now the dominant lineage in southern Sindh; its southward spread may have serious implications for cotton plantations in this region.     

Diversity of secondary endosymbionts among different putative species of the whitefly Bemisia tabaci

Abstract Endosymbionts are important components of arthropod biology. The whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a cryptic species complex composed of ≥ 28 putative species. In addition to the primary endosymbiont Portiera aleyrodidarum, six secondary endosymbionts (S‐endosymbionts), Hamiltonella, Rickettsia, Wolbachia, Cardinium, Arsenophonus and Fritschea, have been identified in B. tabaci thus far. Here, we tested five of the six S‐endosymbiont lineages (excluding Fritschea) from 340 whitely individuals representing six putative species from China. Hamiltonella was detected only in the two exotic invaders, Middle East‐Asia Minor 1 (MEAM1) and Mediterranean (MED). Rickettsia was absent in Asia II 1 and MED, scarce in Asia II 3 (13%), but abundant in Asia II 7 (63.2%), China 1 (84.7%) and MEAM1 (100%). Wolbachia, Cardinium and Arsenophonus were absent in the invasive MEAM1 and MED but mostly abundant in the native putative species. Furthermore, phylogenetic analyses revealed that some S‐endosymbionts have several clades and different B. tabaci putative species can harbor different clades of a given S‐endosymbiont, demonstrating further the complexity of S‐endosymbionts in B. tabaci. All together, our results demonstrate the variation and diversity of S‐endosymbionts in different putative species of B. tabaci, especially between invasive and native whiteflies.     

A virus and its vector,pepper yellow leaf curl virus and <Emphasis Type="Italic">Bemisia tabaci</Emphasis>, two new invaders of Indonesia

Bemisia tabaci is a species of sap-sucking insect belonging to the Aleyrodidae and are commonly known as whiteflies. The species is made up of a complex of distinct genetic groups which have a strong geographic pattern to their genetic structure. Two members of this complex known as the B and Q biotypes have proven to be particularly invasive, spreading with the aid of trade in ornamental plants, well beyond their home ranges across the Mediterranean Basin, Middle East and Asia Minor. This study uses DNA microsatellites to identify another biological invasion this time involving a B. tabaci from south east Asia. We provide evidence which supports an invasion sometime between 1994 and 1999 of B. tabaci from central Thailand into the Indonesian islands of Sumatra then Java and Bali. The invasion is also associated with the invasion of pepper yellow leaf curl virus, a begomovirus transmitted by B. tabaci, which is also shown to have a probable origin in the same geographic region as the invading whitefly. The consequences of the invasion of a plant-infecting virus and its vector has been a massive increase in the scale and impact of begomoviruses in tomato and chilli production which has seen regional bans imposed on the planting of chilli, an important cash crop for many village farmers in Sumatra and Java.     

Genetic variability,community structure,and horizontal transfer of endosymbionts among three Asia II‐Bemisia tabaci mitotypes in Pakistan

Endosymbionts associated with the whitefly Bemisia tabaci cryptic species are known to contribute to host fitness and environmental adaptation. The genetic diversity and population complexity were investigated for endosymbiont communities of B. tabaci occupying different micro‐environments in Pakistan. Mitotypes of B. tabaci were identified by comparative sequence analysis of the mitochondria cytochrome oxidase I (mtCOI) gene sequence. Whitefly mitotypes belonged to the Asia II‐1, ‐5, and ‐7 mitotypes of the Asia II major clade. The whitefly–endosymbiont communities were characterized based on 16S ribosomal RNA operational taxonomic unit (OTU) assignments, resulting in 43 OTUs. Most of the OTUs occurred in the Asia II‐1 and II‐7 mitotypes (r² = .9, p < .005), while the Asia II‐5 microbiome was less complex. The microbiome OTU groups were mitotype‐specific, clustering with a basis in phylogeographical distribution and the corresponding ecological niche of their whitefly host, suggesting mitotype‐microbiome co‐adaptation. The primary endosymbiont Portiera was represented by a single, highly homologous OTU (0%–0.67% divergence). Two of six Arsenophonus OTUs were uniquely associated with Asia II‐5 and ‐7, and one occurred exclusively in Asia II‐1, two only in Asia II‐5, and one in both Asia II‐1 and ‐7. Four other secondary endosymbionts, Cardinium, Hemipteriphilus, Rickettsia, and Wolbachia OTUs, were found at ≤29% frequencies. The most prevalent Arsenophonus OTU was found in all three Asia II mitotypes (55% frequency), whereas the same strain of Cardinium and Wolbachia was found in both Asia II‐1 and ‐5, and a single Hemipteriphilus OTU occurred in Asia II‐1 and ‐7. This pattern is indicative of horizontal transfer, suggestive of a proximity between mitotypes sufficient for gene flow at overlapping mitotype ecological niches.     

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

The 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.     

Microbiome diversity and reproductive incompatibility induced by the prevalent endosymbiont Arsenophonus in two species of African cassava Bemisia tabaci whiteflies

A minimum of 13 diverse whitefly species belonging to the Bemisia tabaci (B. tabaci) species complex are known to infest cassava crops in sub‐Saharan Africa (SSA), designated as SSA1‐13. Of these, the SSA1 and SSA2 are the predominant species colonizing cassava crops in East Africa. The SSA species of B. tabaci harbor diverse bacterial endosymbionts, many of which are known to manipulate insect reproduction. One such symbiont, Arsenophonus, is known to drive its spread by inducing reproductive incompatibility in its insect host and are abundant in SSA species of B. tabaci. However, whether Arsenophonus affects the reproduction of SSA species is unknown. In this study, we investigated both the reproductive compatibility between Arsenophonus infected and uninfected whiteflies by inter‐/intraspecific crossing experiments involving the sub‐group three haplotypes of the SSA1 (SSA1‐SG3), SSA2 species, and their microbial diversity. The number of eggs, nymphs, progenies produced, hatching rate, and survival rate were recorded for each cross. In intra‐specific crossing trials, both male and female progenies were produced and thus demonstrated no reproductive incompatibility. However, the total number of eggs laid, nymphs hatched, and the emerged females were low in the intra‐species crosses of SSA1‐SG3A+, indicating the negative effect of Arsenophonus on whitefly fitness. In contrast, the inter‐species crosses between the SSA1‐SG3 and SSA2 produced no female progeny and thus demonstrated reproductive incompatibility. The relative frequency of other bacteria colonizing the whiteflies was also investigated using Illumina sequencing of 16S rDNA and diversity indices were recorded. Overall, SSA1‐SG3 and SSA2 harbored high microbial diversity with more than 137 bacteria discovered. These results described for the first time the microbiome diversity and the reproductive behaviors of intra‐/inter‐species of Arsenophonus in whitefly reproduction, which is crucial for understanding the invasion abilities of cassava whiteflies.