Phylogeny and evolution of hemipteran insects based on expanded genomic and transcriptomic data.

BACKGROUND: Hemiptera is the fifth species-rich order of insects and the most species-rich order of hemimetabolous insects, including numerous insect species that are of agricultural or medical significance. Despite much effort and recent advance in inferring the Hemiptera phylogeny, some high-level relationships among superfamilies remain controversial. RESULTS: We sequenced the genomes of 64 hemipteran species from 15 superfamilies and the transcriptomes of two additional scale insect species, integrating them with existing genomic and transcriptomic data to conduct a comprehensive phylogenetic analysis of Hemiptera. Our datasets comprise an average of 1625 nuclear loci of 315 species across 27 superfamilies of Hemiptera. Our analyses supported Cicadoidea and Cercopoidea as sister groups, with Membracoidea typically positioned as the sister to Cicadoidea + Cercopoidea. In most analyses, Aleyrodoidea was recovered as the sister group of all other Sternorrhyncha. A sister-group relationship was supported between Coccoidea and Aphidoidea + Phylloxeroidea. These relationships were further supported by four-cluster likelihood mapping analyses across diverse datasets. Our ancestral state reconstruction indicates phytophagy as the primary feeding strategy for Hemiptera as a whole. However, predation likely represents an ancestral state for Heteroptera, with several phytophagous lineages having evolved from predatory ancestors. Certain lineages, like Lygaeoidea, have undergone a reversal transition from phytophagy to predation. Our divergence time estimation placed the diversification of hemipterans to be between 60 and 150 million years ago. CONCLUSIONS: By expanding phylogenomic taxon sampling, we clarified the superfamily relationships within the infraorder Cicadomorpha. Our phylogenetic analyses supported the sister-group relationship between the superfamilies Cicadoidea and Cercopoidea, and the superfamily Membracoidea as the sister to Cicadoidea + Cercopoidea. Our divergence time estimation supported the close association of hemipteran diversification with the evolutionary success and adaptive radiation of angiosperms during the Cretaceous period.

Evolution of satDNAs on holocentric chromosomes: insights from hemipteran insects of the genus Mahanarva.

Satellite DNAs (satDNAs) constitute one of the main components of eukaryote genomes and are involved in chromosomal organization and diversification. Although largely studied, little information was gathered about their evolution on holocentric species, i.e., diffuse centromeres, which, due to differences in repeat organization, could result in different evolutionary patterns. Here, we combined bioinformatics and cytogenetic approaches to evaluate the evolution of the satellitomes in Mahanarva holocentric insects. In two species, de novo identification revealed a high number of satDNAs, 110 and 113, with an extreme monomer length range of 18-4228 bp. The overall abundance of satDNAs was observed to be 6.67% in M. quadripunctata and 1.98% in M. spectabilis, with different abundances for the shared satDNAs. Chromosomal mapping of the most abundant repeats of M. quadripunctata and M. spectabilis on other Mahanarva reinforced the dynamic nature of satDNAs. Variable patterns of chromosomal distribution for the satDNAs were noticed, with the occurrence of clusters on distinct numbers of chromosomes and at different positions and the occurrence of scattered signals or nonclustered satDNAs. Altogether, our data demonstrated the high dynamism of satDNAs in Mahanarva with the involvement of this genomic fraction in chromosome diversification of the genus. The general characteristics and patterns of evolution of satDNAs are similar to those observed on monocentric chromosomes, suggesting that the differential organization of genome compartments observed on holocentric chromosomes compared with monocentric chromosomes does not have a large impact on the evolution of satDNAs. Analysis of the satellitomes of other holocentric species in a comparative manner will shed light on this issue.

Two new species of Erythroneurini from China (Hemiptera, Cicadellidae, Typhlocybinae).

BACKGROUND: The leafhopper genus Empoascanara Distant, 1918 encompasses 81 species, most of which are distributed in Afrotropical, Oriental and Australian Realm. The leafhopper genus Kapsa Dworakowska, 1972 encompasses 23 species, mainly known from the Oriental and Australian Realms. NEW INFORMATION: Two new species of the leafhopper tribe Erythroneurini from Guizhou Province, China, Empoascanaradichotomus sp. nov. and Kapsasinuose sp. nov. are described and illustrated. Identification keys to the males of the genera Empoascanara and Kapsa in China are proposed.

Revision of the New World leafhopper tribe Faltalini (Hemiptera: Cicadellidae: Deltocephalinae) and the evolution of brachyptery.

The taxonomy of the deltocephaline leafhopper tribe Faltalini (13 genera, 63 species) is revised. A key to all genera and keys to species for each genus are provided. All genera are described and at least one species of each genus is illustrated including images of the habitus, male and female genitalia, and scanning electron microscope (SEM) images of the head, face, and other characters. The new genera Dietrichana n.gen. [type species D. pampas n. sp.] and Paraclorindaia n.gen. [type species P. pinguis n. sp.] are described and the following 36 new species are described: Ackbaria jujuyensis n. sp. [Argentina], A. ojosverdes n. sp. [Argentina], Aequcephalus laplata n. sp. [Chile], Bonamus jussaral n. sp. [Brazil], Clorindaia obrienorum n. sp. [Uruguay], C. otamendi n. sp. [Argentina], Dietrichana pampas n. sp. [Argentina], Faltala catalanoae n. sp. [Argentina, Uruguay], F. paradellae n. sp. [Argentina], F. viscacha n. sp. [Argentina, Uruguay], Hecullus mexicanus n. sp. [Mexico], Kramerana saltensis n. sp. [Argentina], Paraclorindaia pinguis n. sp. [Argentina, Uruguay], P. vesperis n. sp. [Argentina], Tenucephalus admirabilis n. sp. [Argentina], T. bartletti n. sp. [Peru], T. brevilamina n. sp. [Argentina], T. cerrado n. sp. [Brazil], T. ecuadorensis n. sp. [Ecuador], T. flagellifer n. sp. [Peru], T. heppneri n. sp. [Peru], T. iguazu n. sp. [Argentina], T. interstinctus n. sp. [Peru], T. johnsoni n. sp. [Brazil], T. lingulatus n. sp. [Bolivia, Peru], T. manuensis n. sp. [Peru], T. misiones n. sp. [Argentina], T. musettiae n. sp. [Brazil], T. nielsoni n. sp. [Brazil], T. novafriburgo n. sp. [Brazil], T. parana n. sp. [Argentina], T. spectabilis n. sp. [Peru], T. takiyae n. sp. [Brazil, Peru], T. umbrinus n. sp. [Brazil], T. viperinus n. sp. [Argentina], and Virganana danasensis n. sp. [Ecuador]. The new combinations Paraclorindaia brasileira (Zahniser) n.comb. [transferred from Clorindaia] and Tenucephalus amabilis (Linnavuori Heller) n.comb. [transferred from Bonamus] are recognized. A phylogenetic analysis based on 51 morphological characters scored for 42 taxa including two outgroup taxa was performed. The resulting phylogeny resolved Faltalini as monophyletic, but the tribe remains difficult to diagnose because it encompasses a large amount of morphological diversity in characters traditionally used to define tribes of Deltocephalinae including head morphology, wing length and venation, and male genitalia. Despite this diversity, members of the tribe share the following characters: 1) phragma of male genital capsule partly developed into sclerotized plates or setose bulbous lobes (exception: Acrolithus); 2) first valvula shape relatively straight; 3) first valvula dorsal sculpturing separated from dorsal margin by unsculptured band; 4) first valvula dorsal sculpturing pattern maculose, with maculae separate or only slightly overlapping, or granulose; 5) first valvula with distinctly delimited ventral sculptured area; and 6) second valvula without dorsal teeth. Two major clades within the tribe were resolved and strongly supported, consisting of (Bonamus, Tenucephalus) and (Hecalocorica, (Hecullus, (Acrolithus, (Ackbaria, (Aequcephalus, Dietrichana, Virganana, Clorindaia, Faltala (Kramerana, Paraclorindaia)))))). A stepwise evolution of brachyptery in the latter clade is suggested by the phylogeny, beginning with evolution of subbrachypterous females in Hecullus and Acrolithus and culminating in completely brachypterous males and females in the monophyletic "Faltala group" of eight genera. It is hypothesized that a transition from an ancestral forested habitat to an open or grassland habitat facilitated the evolution of brachyptery. The hindwings and associated thoracic structures of eight species of Faltalini and three other Deltocephalinae were imaged with SEM. Brachyptery in Cicadellidae and other insects are briefly reviewed and discussed considering the hypothesis proposed here.

Two new species of Aufidus (Hemiptera, Cercopidae) from New Guinea.

Two new species of the cercopid genus Aufidus Stål, 1863, A. alacaligus sp. nov. and A. albonigrus sp. nov., are described and illustrated, Photos of the type specimens with the illustrations of the male terminalia are provided. A key to the species found in New Guinea is presented.

Comparative analysis of mitochondrial genomes of Nirvanini and Evacanthini (Hemiptera: Cicadellidae) reveals an explicit evolutionary relationship.

Mitogenomes of five leafhopper species, Chudania hellerina and Concaveplana rufolineata in Nirvanini, Carinata rufipenna, Evacanthus danmainus and E. heimianus representing Evacanthini, were sequenced. The lengths of these five mitogenomes range from 15,044 (C. hellerina) to 15,680 bp (E. heimianus). All five mitogenomes exhibit similar base composition, gene size and codon usage of protein-coding genes. All 22 tRNA genes have typical cloverleaf secondary structures, except for trnS1 (AGN) which appears to lack the dihydrouridine arm. The two included Nirvanini species employ the anticodon TCT instead of the commonly used GCT in trnS1 (AGN). Genes nad2, atp8 and nad6 were highly variable while cox1 and cob showed the lowest nucleotide diversity. Phylogenetic analyses of two concatenated nucleotide datasets, incorporating the newly sequenced taxa and other available membracoid mitogenomes, recovered each included leafhopper subfamily as monophyletic with evacanthine tribes Nirvanini and Evacanthini forming monophyletic sister clades. A relationship among Evacanthinae, Cicadellinae and Typhlocybinae received moderate branch support.

First Description of Adult Males of Four Species of the Treehopper genus Tricentrus Stål from China (Hemiptera, Membracidae, Centrotinae, Tricentrini).

Adult males are described and illustrated for the first time for four Chinese species of the treehopper genus Tricentrus previously known only from females: T. albipennis Kato, 1930, T. floripinnae Yuan Cui, 1987, T. foliocornatus, Yuan Fan, 2002 and T. oedothorectoidis Yuan Fan, 2002.

New genus and species of Leptopsaltriini (Hemiptera: Cicadidae: Cicadinae) from China and Vietnam, with colour-changing behaviour reported for the first time in Cicadoidea.

One new cicada genus Versicolora gen. nov. and two new species, V. ziyongi sp. nov. from China and V. bellula sp. nov. from China and Vietnam, are described. The new genus is placed in the tribe Leptopsaltriini of the subfamily Cicadinae. The relationship of this new genus to other related taxa is discussed. Versicolora ziyongi sp. nov. camouflages itself on the bark of the host-plants and gradually changes its body colour when captured. This colour-changing behaviour is recorded for the first time in Cicadoidea, which provides innovative information for ecomorphological study of this remarkable species and other cicadas that potentially exhibit this behaviour.

Structural features and phylogenetic implications of four new mitogenomes of Centrotinae (Hemiptera: Membracidae).

To explore the variation and phylogenetic utility of mitogenomes among lineages of the diverse hemipteran superfamily Membracoidea, we sequenced four new mitogenomes of four treehopper species of the subfamily Centrotinae (Membracidae): Hypsauchenia hardwichii, Leptocentrus albolineatus, Maurya qinlingensis, and Tricentrus brunneus. The mitogenomes are 15,508 to 16,467 bp in size, and comprise the typical set of 37 mitochondrial genes and a large non-coding region (AT-rich region). Gene organization, nucleotide composition and codon usage of protein-coding genes (PCGs) are similar to those of most other sequenced Membracidae mitogenomes. All PCGs start with a typical ATN or TTG and end with TAA/G or the incomplete stop codon (a single T). All transfer RNA genes can be folded into typical clover-leaf secondary structures, except for trnS1. The location, length and AT content of the rrnS and rrnL genes are highly conserved in the Membracidae mitogenomes. In contrast, the AT-rich control region is highly variable in length and in numbers of tandem repeats present. Phylogenetic analyses based on the nucleotide and amino acid sequence data of 13 PCGs from 59 species of Membracoidea and four outgroups (Cercopoidea and Cicadoidea species) recovered Membracoidea as monophyletic with strong support, and Cicadellidae as paraphyletic with respect to Aetalionidae + Membracidae, in agreement with previous analyses. Relationships among membracoid subfamilies were also in general agreement with results from prior studies. The monophyly of Centrotinae is strongly supported, with relationships among tribes recovered as ((Centrotini+ (Tricentrini + Antialcidini)) + ((Leptobelini + Hypsauchenini) + Leptocentrini).

A new species in the grass feeding leafhopper genus Hecalus Stål from Pakistan (Cicadellidae: Deltocephalinae: Hecalini).

A new leafhopper species Hecalus rawalakotensis sp. nov. is described and illustrated from Rawalakot, Azad Jammu and Kashmir, Pakistan. A key is provided to distinguish all species in this genus from Pakistan.

Revision of the leafhopper genus Soractellus Evans (Cicadellidae: Deltocephalinae) with description of a new species from Pakistan.

The leafhopper genus Soractellus Evans, 1966 is reviewed and a new short-winged species, Soractellus lalianensis sp. nov. from Pakistan, is described and illustrated. A key to distinguish all species in this genus is given.

Nomenclatural changes in the grass-feeding Mexican leafhopper genus Cocrassana Blocker & Larsen (Hemiptera: Cicadellidae: Deltocephalinae: Athysanini)

ABSTRACT Two nomenclatural changes are proposed in the tribe Athysanini (Cicadellidae: Deltocephalinae). Chlorotettix sexvarus DeLong, 1959, is newly placed in the genus Cocrassana Blocker & Larsen and is a senior synonym of the type species of Cocrassana, C. riepmai Blocker & Larsen, 1991, syn. nov. A revised diagnosis and illustration of the species are provided.

Ultrastructure of spermatozoa in three cicada species from China (Hemiptera, Cicadomorpha, Cicadidae).

The ultrastructure of mature spermatozoa of three cicada species, Subpsaltria yangi, Karenia caelatata, and Platypleura kaempferi, was investigated using epifluorescence and transmission electron microscopies. This is the first investigation of the sperm ultrastructure of species in the subfamily Tibicininae and the tribe Sinosenini, represented by S. yangi and K. caelatata, respectively. The three species all produce two or three types of spermatozoa with various lengths, viz., polymegaly. The centriolar adjunct of spermatozoa in S. yangi shows a granular substructure, which is different from that of other cicada species, suggesting that spermatozoa in Tibicininae may have their own characteristics in comparison with other cicadas. The centriolar adjunct of spermatozoa of K. caelatata displays characteristics similar to that of the Cicadinae. Combined with other morphological characters, it is reasonable to remove K. caelatata and its allies (i.e., Sinosenini) from Cicadettinae to Cicadinae. The study of sperm ultrastructure, particularly in the species of Tibicininae and Sinosenini, expands the spermatological research of Cicadidae and provides more information for phylogenetic analysis of Cicadidae.

New records and updated checklist of Cicadomorpha (Hemiptera: Auchenorrhyncha) species from Greece.

Seventeen auchenorrhynchan species of the family Cicadellidae are recorded from Greece for the first time: Euscelidius variegatus (Kirschbaum 1858), Euscelis ohausi (Wagner 1939), Psammotettix notatus (Melichar 1896), Psammotettix striatus (Linnaeus 1758), Psammotettix sabulicola (Curtis 1837), Anzygina honiloa (Kirkaldy 1906), Arboridia versuta (Melichar 1897), Edwardsiana ishidae (Matsumura 1932), Edwardsiana tersa (Edwards 1914), Eupteryx decemnotata (Rey 1891), Eupteryx curtisii (Flor 1861), Eupteryx rostrata (Ribaut 1936), Frutioidia sanguinosa (Rey 1891), Zygina rosea (Flor 1861), Zygina suavis (Rey 1891), Zygina tiliae (Fallen 1806), Anaceratagallia frisia (Wagner 1939). Faunistic and distributional data notes are given for each species and a complete, updated species checklist for Greece is provided.

Re-description of the sperm morphology of Dalbulus maidis (Hemiptera: Cicadellidae: Deltocephalinae).

The corn leafhopper, Dalbulus maidis, which is one of the most important vectors of maize diseases, had the morphology of its spermatozoa described more than 40 years ago. However, the description is imprecise, requiring certain rectifications. Hence, we re-describe the sperm morphology of this species using light microscopy and transmission electron microscopy. Their spermatozoa measured from 118.1 to 128.5 µm in length, this value being about three times smaller than what was previously described. The head region is formed by a small acrosome and the nucleus. The acrosome is paracrystalline and has the bifurcated base that is docked in two cavities on one side of the anterior region of the nucleus. The nucleus, which is linear and thin, measured from 19.3 to 22.9 µm in length, and exhibited homogeneously compact chromatin. In cross-section, it has a thumbtack shape, except in the anterior and posterior regions, where it is oval. The flagellum consists of an axoneme with 9 + 9 + 2 microtubules, two similar mitochondrial derivatives, two accessory bodies each associated with a small sub-ellipsoidal structure, and a 'center-flagellar material'. In all the spermatozoa observed, the final portion of the flagellum was branched into four filaments. Some of these morphological features are now being re-described, while others have been observed for the first time in the Cicadellidae family, as the center-flagellar material.

A new species of Cicadatra Kolenati, 1857 from China, with primary phylogenetic analyses of the tribe Cicadatrini (Hemiptera: Cicadidae).

A new species of the genus Cicadatra Kolenati, 1857, C. hei sp. nov., from Sichuan Province, China is described. A key to the species of Cicadatra from China is provided. Morphological characters for 9 taxa of the tribe Cicadatrini and for 3 outgroup taxa are examined to present the first phylogenetic reconstruction within the Cicadatrini. The monophyly of Cicadatrini is supported, and Cicadatrini is more allied to the outgroup Cicadetta of the subfamily Cicadettinae (= Tibicininae auct.) from TNT and WinClada analyses. The systematic status of the tribe Cicadatrini is discussed. The genus Cicadalna Boulard, 2006 is recognized to be a junior synonym of Emathia Stål, 1866, and Cicadalna takensis Boulard, 2006 is transferred from Cicadalna to Emathia to become Emathia takensis (Boulard, 2006) comb. nov.

First finding of a dual-meaning X wave for phloem and xylem fluid ingestion: Characterization of Scaphoideus titanus (Hemiptera: Cicadellidae) EPG waveforms.

The leafhopper Scaphoideus titanus (Hemiptera: Cicadomorpha: Cicadellidae), an invasive deltocephaline species introduced into Europe from North America, is the vector of the most important phytoplasma disease in European viticulture, flavescence dorée. In this first electropenetrography (EPG) study of S. titanus, we characterized its feeding waveforms and defined their biological meanings. Four typical waveform phases (pathway, X wave, sustained ingestion, and interruption) and four families within those phases (A, B, C, and N) were characterized using DC EPG technology. We proposed biological meanings for these waveforms based on excreta pH-ingestion correlations, presence of X waves, and comparison with previous AC, DC, and AC-DC EPG waveforms conducted on Cicadomorpha. We observed that sustained (i.e., >10min) ingestion by a deltocephaline leafhopper can occur from both xylem and phloem vascular cells. Waveform C2x represented ingestion of xylem fluid, and two waveforms represented behaviors when stylets were inserted into phloem sieve elements: C2p variant 1 (C2p-1), which may represent salivation (perhaps simultaneous with ingestion), and C2p variant 2 (C2p-2), which represented active ingestion. Furthermore, we found that the EPG-recorded X wave has a dual meaning by occurring prior to sustained ingestion from either phloem or xylem. This X wave was very similar in appearance to the model X wave of sharpshooters, an entirely different leafhopper subfamily, Cicadellinae. All cicadellines are obligate xylem-ingesters. Such a "dual-meaning X wave" will provide insights into how the feeding tactics of S. titanus relate to other sheath-feeding hemipterans, and will provide support for future research to clarify the role of this leafhopper as a vector of plant pathogens.

Mixed xylem and phloem sap ingestion in sheath-feeders as normal dietary behavior: Evidence from the leafhopper Scaphoideus titanus.

In phytophagous piercing-sucking insects, salivary sheath-feeding species are often described as xylem- or phloem-sap feeding specialists. Because these two food sources have very different characteristics, two feeding tactics are often associated with this supposed specialization. Studying the feeding behavior of insects provides substantial information on their biology, ecology, and evolution. Furthermore, study of feeding behavior is of primary importance to elucidate the transmission ability of insects that act as vectors of plant pathogens. In this study, we compared the durations of ingestion performed in xylem versus phloem by a leafhopper species, Scaphoideus titanus Ball, 1932. This was done by characterizing and statistically analyzing electrical signals recorded using the electropenetrography technique, derived from the feeding behaviors of males and females. We identified three groups of S. titanus based on their feeding behavior: 1) a group that reached the phloem quickly and probed for a longer time in phloem tissue than the other groups, 2) a group that reached the xylem quickly and probed for a longer time in xylem tissue than the other groups, and 3) a group where individuals did not ingest much sap. In addition, the numbers and durations of waveforms representing ingestion of xylem and phloem saps differed significantly depending on the sex of the leafhopper, indicating that the two sexes exhibit different feeding behaviors. Males had longer phloem ingestion events than did females, which indicates that males are greater phloem feeders than females. These differences are discussed, specifically in relation to hypotheses about evolution of sap feeding and phytoplasma transmission from plant to plant.

Duocrassana longula, new genus and species of leafhopper (Hemiptera: Cicadellidae: Deltocephalinae) from southern Mexico and its relationship to other genera in Athysanini.

The new leafhopper genus, Duocrassana Pinedo-Escatel, Zahniser & Dietrich, gen. nov., and its type species, Duocrassana longula Pinedo-Escatel, Zahniser & Dietrich sp. nov., are described and illustrated based on material from Oaxaca, Mexico. The relationship with other genera in the tribe Athysanini is discussed, highlighting the importance of male genitalia for its identification.

Bambusimukaria, a new bamboo-feeding leafhopper genus from China, with description of one new species (Hemiptera, Cicadellidae, Deltocephalinae, Mukariini).

A new genus and species, Bambusimukaria quinquepunctata gen. & sp. n., feeding on bamboo in Guizhou and Fujian, China, are described and illustrated. The characters of crown, frontoclypeus, forewing venations and male genitalia place the new genus in the tribe Mukariini.