Two New Species of the Genus Eucorydia (Blattodea: Corydiidae) from the Nansei Islands in Southwest Japan.

Two new species of the cockroach genus Eucorydia Hebard, 1929 from the Nansei Islands in Southwest Japan were compared to two closely related congeners, Eucorydia yasumatsui Asahina, 1971 and Eucorydia dasytoides (Walker, 1868). Eucorydia donanensis Yanagisawa, Sakamaki, and Shimano sp. nov. from Yonaguni-jima Island was characterized by an overall length of 12.5-14.5 mm in males. The dorsal side of the male abdomen was entirely dark purple and there was an obscure orange band running down the middle of the tegmen. Eucorydia tokaraensis Yanagisawa, Sakamaki, and Shimano sp. nov. was characterized by an overall length of 12.0-13.0 mm in males and a distinct orange band running down the middle of the tegmen. Eucorydia yasumatsui, E. donanensis, E. tokaraensis and the zonata population of E. dasytoides were divided into four lineages in a maximum-likelihood tree generated from a dataset concatenated from five (two nuclear, 28S rRNA, histone H3, and three mitochondrial, COII, 12S rRNA, 16S rRNA) genes. We recognized the three Japanese lineages E. yasumatsui, E. donanensis, and E. tokaraensis as distinct species, which were also supported by the pairwise genetic distances (5.4-7.8%, K2P) of the COI sequences. Morphometric analysis was performed on the genitalia. A principal component analysis plot revealed that the sizes of the genitalia in the three Japanese species were similar to each other and smaller than that of the zonata population of E. dasytoides. The analysis also revealed that the three Japanese species were distinguished from each other by combinations of the sizes of L3 and L7 sclerites and the shape of R2 sclerite, with some overlapping exceptions.

Long rDNA amplicon sequencing of insect-infecting nephridiophagids reveals their affiliation to the Chytridiomycota and a potential to switch between hosts.

Nephridiophagids are unicellular eukaryotes that parasitize the Malpighian tubules of numerous insects. Their life cycle comprises multinucleate vegetative plasmodia that divide into oligonucleate and uninucleate cells, and sporogonial plasmodia that form uninucleate spores. Nephridiophagids are poor in morphological characteristics, and although they have been tentatively identified as early-branching fungi based on the SSU rRNA gene sequences of three species, their exact position within the fungal tree of live remained unclear. In this study, we describe two new species of nephridiophagids (Nephridiophaga postici and Nephridiophaga javanicae) from cockroaches. Using long-read sequencing of the nearly complete rDNA operon of numerous further species obtained from cockroaches and earwigs to improve the resolution of the phylogenetic analysis, we found a robust affiliation of nephridiophagids with the Chytridiomycota-a group of zoosporic fungi that comprises parasites of diverse host taxa, such as microphytes, plants, and amphibians. The presence of the same nephridiophagid species in two only distantly related cockroaches indicates that their host specificity is not as strict as generally assumed.

The power of neuropeptide precursor sequences to reveal phylogenetic relationships in insects: A case study on Blattodea.

Recent state-of-the-art analyses in insect phylogeny have exclusively used very large datasets to elucidate higher-level phylogenies. We have tested an alternative and novel approach by evaluating the potential phylogenetic signals of identified and relatively short neuropeptide precursor sequences with highly conserved functional units. For that purpose, we examined available transcriptomes of 40 blattodean species for the translated amino acid sequences of 17 neuropeptide precursors. Recently proposed intra-ordinal relationships of Blattodea, based on the analysis of 2370 protein-coding nuclear single-copy genes (Evangelista et al., 2019), were corroborated with maximum support. The functionally different precursor units were analyzed separately for their phylogenetic information. Although the degree of information was different in the different sequence motifs, all precursor units contained phylogenetic informative data at the ordinal level, and their separate analysis did not reveal contradictory topologies. This study is the first comprehensive exploitation of complete neuropeptide precursor sequences of arthropods in such a context and demonstrates the applicability of these rather short but conserved sequences for an alternative, fast and simple analysis of phylogenetic relationships.

An integrative phylogenomic approach illuminates the evolutionary history of cockroaches and termites (Blattodea).

Phylogenetic relationships among subgroups of cockroaches and termites are still matters of debate. Their divergence times and major phenotypic transitions during evolution are also not yet settled. We addressed these points by combining the first nuclear phylogenomic study of termites and cockroaches with a thorough approach to divergence time analysis, identification of endosymbionts, and reconstruction of ancestral morphological traits and behaviour. Analyses of the phylogenetic relationships within Blattodea robustly confirm previously uncertain hypotheses such as the sister-group relationship between Blaberoidea and remaining Blattodea, and Lamproblatta being the closest relative to the social and wood-feeding Cryptocercus and termites. Consequently, we propose new names for various clades in Blattodea: Cryptocercus + termites = Tutricablattae; Lamproblattidae + Tutricablattae = Kittrickea; and Blattoidea + Corydioidea = Solumblattodea. Our inferred divergence times contradict previous studies by showing that most subgroups of Blattodea evolved in the Cretaceous, reducing the gap between molecular estimates of divergence times and the fossil record. On a phenotypic level, the blattodean ground-plan is for egg packages to be laid directly in a hole while other forms of oviposition, including ovovivipary and vivipary, arose later. Finally, other changes in egg care strategy may have allowed for the adaptation of nest building and other novelties.

New Cockroaches (Dictyoptera: Blattodea) from French Guiana and a Revised Checklist for the Region.

Although French Guiana is one of the greatest hotspots of cockroach biodiversity on Earth, there are still undocumented species. From both newly collected and museum specimens, we provide species descriptions for Buboblatta vlasaki sp. nov., Lamproblatta antoni sp. nov., and Euhypnorna bifuscina sp. nov. and report new geographic records for species in the genera Epilampra Burmeister, Euphyllodromia Shelford, Ischnoptera Burmeister, and Euhypnorna Hebard. Finally, we update the checklist of species known from the region to 163 total species records from French Guiana, making it the second greatest hotspot of known cockroach biodiversity on Earth.

Ribosomal DNA internal transcribed spacer 2 sequence analysis and phylogenetic comparison of seven cockroach species in northwestern Iran.

OBJECTIVES: The current study was conducted to identify cockroach species (Blattodea) of northwestern Iran in public places using morphological characteristics and ribosomal DNA internal transcribed spacer 2 (rDNA-ITS2). Sequences were analyzed with Basic Local Alignment Search Tool (BLAST) searches, Neighbor-Joining methods based on and Tamura-Nei phylogenetic analyses. In addition, eight cockroach rDNA-ITS2 sequences from China, India, Iran and the United States obtained from GenBank were compared to those obtained in this study. RESULTS: Specimens collected in Iran were identified as Periplaneta americana (L.), Shelfordella lateralis (Walker), Blatta orientalis (L.) (Blattodea: Blattidae), Blattella germanica (L.), Supella longipalpa (F.) (Blattodea: Ectobiidae), Polyphaga aegyptiaca (L.), and Polyphaga saussurei (Dohrn) (Blattodea: Corydiidae). rDNA-ITS2 nucleotide sequence analysis showed 100% similarity between P. aegyptiaca and P. saussurei species collected from Iran despite morphological differences. However, ITS2 sequence of P. americana submitted from China showed 30.49-31.71% difference to P. americana sequences from Iran and the United States. The results highlight the importance of morphological identification of cockroach species before conducting molecular techniques.

Historical biogeography of the termite clade Rhinotermitinae (Blattodea: Isoptera).

Termites are the principal decomposers in tropical and subtropical ecosystems around the world. Time-calibrated molecular phylogenies show that some lineages of Neoisoptera diversified during the Oligocene and Miocene, and acquired their pantropical distribution through transoceanic dispersal events, probably by rafting in wood. In this paper, we intend to resolve the historical biogeography of one of the earliest branching lineages of Neoisoptera, the Rhinotermitinae. We used the mitochondrial genomes of 27 species of Rhinotermitinae to build two robust time-calibrated phylogenetic trees that we used to reconstruct the ancestral distribution of the group. Our analyses support the monophyly of Rhinotermitinae and all genera of Rhinotermitinae. Our molecular clock trees provided time estimations that diverged by up to 15.6 million years depending on whether or not 3rd codon positions were included. Rhinotermitinae arose 50.4-64.6 Ma (41.7-74.5 Ma 95% HPD). We detected four disjunctions among biogeographic realms, the earliest of which occurred 41.0-56.6 Ma (33.0-65.8 Ma 95% HPD), and the latest of which occurred 20.3-34.2 Ma (15.9-40.4 Ma 95% HPD). These results show that the Rhinotermitinae acquired their distribution through a combination of transoceanic dispersals and dispersals across land bridges.

The evolution of sociality in termites from cockroaches: A taxonomic and phylogenetic perspective.

Despite multiple studies and advances, sociality still puzzles evolutionary biologists in numerous ways, which might be partly addressed with the advent of sociogenomics. In insects, the majority of sociogenomic studies deal with Hymenoptera, one of the two groups that evolved eusociality with termites. But, to fully grasp the evolution of sociality, studies must obviously not restrict to eusocial lineages. Multiple kinds of social system transitions have been recorded and they all bring complementary insights. For instance, cockroaches, the closest relatives to termites, display a wide range of social interactions and evolved convergently subsocial behaviors (i.e., brood care). In this context, we emphasize the need for natural history, taxonomic, and phylogenetic studies. Natural history studies provide the foundations on which building hypotheses, whereas taxonomy provides the taxa to sample to test these hypotheses, and phylogenetics brings the historical framework necessary to test evolutionary scenarios of sociality evolution.

Topological support and data quality can only be assessed through multiple tests in reviewing Blattodea phylogeny.

Assessing support for molecular phylogenies is difficult because the data is heterogeneous in quality and overwhelming in quantity. Traditionally, node support values (bootstrap frequency, Bayesian posterior probability) are used to assess confidence in tree topologies. Other analyses to assess the quality of phylogenetic data (e.g. Lento plots, saturation plots, trait consistency) and the resulting phylogenetic trees (e.g. internode certainty, parameter permutation tests, topological tests) exist but are rarely applied. Here we argue that a single qualitative analysis is insufficient to assess support of a phylogenetic hypothesis and relate data quality to tree quality. We use six molecular markers to infer the phylogeny of Blattodea and apply various tests to assess relationship support, locus quality, and the relationship between the two. We use internode-certainty calculations in conjunction with bootstrap scores, alignment permutations, and an approximately unbiased (AU) test to assess if the molecular data unambiguously support the phylogenetic relationships found. Our results show higher support for the position of Lamproblattidae, high support for the termite phylogeny, and low support for the position of Anaplectidae, Corydioidea and phylogeny of Blaberoidea. We use Lento plots in conjunction with mutation-saturation plots, calculations of locus homoplasy to assess locus quality, identify long branch attraction, and decide if the tree's relationships are the result of data biases. We conclude that multiple tests and metrics need to be taken into account to assess tree support and data robustness.

The Complete Mitogenome of the Wood-Feeding Cockroach Cryptocercus meridianus (Blattodea: Cryptocercidae) and Its Phylogenetic Relationship among Cockroach Families.

In this study, the complete mitochondrial genome of Cryptocercus meridianus was sequenced. The circular mitochondrial genome is 15,322 bp in size and contains 13 protein-coding genes, two ribosomal RNA genes (12S rRNA and 16S rRNA), 22 transfer RNA genes, and one D-loop region. We compare the mitogenome of C. meridianus with that of C. relictus and C. kyebangensis. The base composition of the whole genome was 45.20%, 9.74%, 16.06%, and 29.00% for A, G, C, and T, respectively; it shows a high AT content (74.2%), similar to the mitogenomes of C. relictus and C. kyebangensis. The protein-coding genes are initiated with typical mitochondrial start codons except for cox1 with TTG. The gene order of the C. meridianus mitogenome differs from the typical insect pattern for the translocation of tRNA-SerAGN, while the mitogenomes of the other two Cryptocercus species, C. relictus and C. kyebangensis, are consistent with the typical insect pattern. There are two very long non-coding intergenic regions lying on both sides of the rearranged gene tRNA-SerAGN. The phylogenetic relationships were constructed based on the nucleotide sequence of 13 protein-coding genes and two ribosomal RNA genes. The mitogenome of C. meridianus is the first representative of the order Blattodea that demonstrates rearrangement, and it will contribute to the further study of the phylogeny and evolution of the genus Cryptocercus and related taxa.

Complete mitochondrial genomes of two blattid cockroaches, Periplaneta australasiae and Neostylopyga rhombifolia, and phylogenetic relationships within the Blattaria.

Complete mitochondrial genomes (mitogenomes) of two cockroach species, Periplaneta australasiae and Neostylopyga rhombifolia, 15,605 bp and 15,711 bp in length, respectively, were determined. As reported for other cockroach mitogenomes, the two mitogenomes possessed typical ancestral insect mitogenome gene composition and arrangement. Only several small intergenic spacers were found: one, which was common in all sequenced cockroach mitogenomes except for the genus Cryptocercus, was between tRNA-Ser (UCN) and ND1 and contained a 7bp highly conserved motif (WACTTAA). Three different types of short tandem repeats in the N. rhombifolia control region (CR) were observed. The homologous alignments of these tandem repeats with other six cockroach mitogenome CRs revealed a low similarity. Three conserved sequence blocks (CSB) were detected in both cockroach mitochondrial CRs. CSB1 was specific for blattinine mitogenomes and was highly conserved with 95% similarity, speculating that this block was a possible molecular synapomorphy for this subfamily. CSB3 located nearby downstream of CSB1 and has more variations within blattinine mitogenomes compared with CSB1. The CSB3 was capable of forming stable stem-loop structure with a small T-stretch in the loop portion. We assessed the influence of four datasets and two inference methods on topology within Orthopteroidea. All genes excluding the third codon positions of PCGs could generate more stable topology, and higher posterior probabilities than bootstrap values were presented at some branch nodes. The phylogenetic analysis with different datasets and analytical methods supported the monophyly of Dictyoptera and supported strongly the proposal that Isoptera should be classified as a family (Termitidae) of the Blattaria. Specifically, Shelfordella lateralis was inserted in the clade Periplaneta. Considering the K2P genetic distance, morphological characters, and the phylogenetic trees, we suggested that S. lateralis should be placed in the genus Periplaneta.

Distribution of Karyotypes of the Cryptocercus punctulatus Species Complex (Blattodea: Cryptocercidae) in Great Smoky Mountains National Park.

During the period between 1999 and 2006, wood-feeding cockroaches in the Cryptocercus punctulatus Scudder species complex were collected throughout Great Smoky Mountains National Park, USA. The chromosome numbers of insects from 59 sites were determined, and phylogenetic analyses were performed based on mitochondrial COII and nuclear ITS2 DNA. The distribution of the three male karyotypes found in the park (2n = 37, 39, and 45) is mapped and discussed in relation to recent disturbances and glacial history. Clades of the three karyotype groups meet near the ridgeline separating North Carolina from Tennessee in the center of the park, suggesting that these may have originated from separate lower elevation refugia after the last glacial maximum. The timing of divergence and a significant correlation between elevation difference and genetic distance in two of the clades supports this hypothesis. The ecological role of the cockroaches in the park is discussed.

Origin of origami cockroach reveals long-lasting (11 Ma) phenotype instability following viviparity.

Viviparity evolved in bacteria, plants, ˃141 vertebrate lineages (ichthyosaurs, lizards, fishes, mammals, and others), and in 11 of 44 insect orders. Live-birth cockroaches preserved with brood sac (3D recovered two times optically) included Diploptera vladimir, Diploptera savba, Diploptera gemini spp.n., D. sp.1-2, and Stegoblatta irmgardgroehni from Green River, Colorado; Quilchena, Republic; McAbee, Canada; and Baltic amber, Russia (49, 54, and 45 Ma). They evolved from rare and newly evolved Blaberidae; they radiated circumtropically, later expanded into SE Asia, and have now spread to Hawaii and the SE USA. Association of autapomorphic characters that allow for passive and active protections from parasitic insects (unique wing origami pleating identical with its egg case-attacking wasp) suggest a response to high parasitic loads. Synchronized with global reorganization of the biota, morphotype destabilization in roaches lasted approximately 11-22 Ma, including both the adaptation of novel characters and the reduction of others. Thus, while viviparity can be disadvantageous, in association with new Bauplans and/or behaviors, it can contribute to the evolution of taxa with viviparous representatives that are slightly selectively preferred.

Selective sweeps in Cryptocercus woodroach antifungal proteins.

We identified the antifungal gene termicin in three species of Cryptocercus woodroaches. Cryptocercus represents the closest living cockroach lineage of termites, which suggests that the antifungal role of termicin evolved prior to the divergence of termites from other cockroaches. An analysis of Cryptocercus termicin and two ß-1,3-glucanase genes (GNBP1 and GNBP2), which appear to work synergistically with termicin in termites, revealed evidence of selection in these proteins. We identified the signature of past selective sweeps within GNBP2 from Cryptocercus punctulatus and Cryptocercus wrighti. The signature of past selective sweeps was also found within termicin from Cryptocercus punctulatus and Cryptocercus darwini. Our analysis further suggests a phenotypically identical variant of GNBP2 was maintained within Cryptocercus punctulatus, Cryptocercus wrighti, and Cryptocercus darwini while synonymous sites diverged. Cryptocercus termicin and GNBP2 appear to have experienced similar selective pressure to that of their termite orthologues in Reticulitermes. This selective pressure may be a result of ubiquitous entomopathogenic fungal pathogens such as Metarhizium. This study further reveals the similarities between Cryptocercus woodroaches and termites.

New cockroach species of the genus Panchlora Burmeister (Blaberidae, Panchlorinae) from Ecuador.

The genus Panchlora includes 49 species, but only 45 are widely distributed in Central and South America. Most of them are green. The new species herein described presents an ornamental coloration markedly different of all until now described species. Panchlora kozaneki sp. n. is similar to Panchlora pulchella Burmeister, 1838. The number of species known from Ecuador is increased to eight.

Gut Bacterial Community of the Xylophagous Cockroaches Cryptocercus punctulatus and Parasphaeria boleiriana.

Cryptocercus punctulatus and Parasphaeria boleiriana are two distantly related xylophagous and subsocial cockroaches. Cryptocercus is related to termites. Xylophagous cockroaches and termites are excellent model organisms for studying the symbiotic relationship between the insect and their microbiota. In this study, high-throughput 454 pyrosequencing of 16S rRNA was used to investigate the diversity of metagenomic gut communities of C. punctulatus and P. boleiriana, and thereby to identify possible shifts in symbiont allegiances during cockroaches evolution. Our results revealed that the hindgut prokaryotic communities of both xylophagous cockroaches are dominated by members of four Bacteria phyla: Bacteroidetes, Firmicutes, Proteobacteria, and Actinobacteria. Other identified phyla were Spirochaetes, Planctomycetes, candidatus Saccharibacteria (formerly TM7), and Acidobacteria, each of which represented 1-2% of the total population detected. Community similarity based on phylogenetic relatedness by unweighted UniFrac analyses indicated that the composition of the bacterial community in the two species was significantly different (P < 0.05). Phylogenetic analysis based on the characterized clusters of Bacteroidetes, Spirochaetes, and Deltaproteobacteria showed that many OTUs present in both cockroach species clustered with sequences previously described in termites and other cockroaches, but not with those from other animals or environments. These results suggest that, during their evolution, those cockroaches conserved several bacterial communities from the microbiota of a common ancestor. The ecological stability of those microbial communities may imply the important functional role for the survival of the host of providing nutrients in appropriate quantities and balance.

The complete mitochondrial genomes of four cockroaches (Insecta: Blattodea) and phylogenetic analyses within cockroaches.

Three complete mitochondrial genomes of Blaberidae (Insecta: Blattodea) (Gromphadorhina portentosa, Panchlora nivea, Blaptica dubia) and one complete mt genome of Blattidae (Insecta: Blattodea) (Shelfordella lateralis) were sequenced to further understand the characteristics of cockroach mitogenomes and reconstruct the phylogenetic relationship of Blattodea. The gene order and orientation of these four cockroach genomes were similar to known cockroach mt genomes, and contained 13 protein-coding genes (PCGs), 2 ribosomal RNA (rRNA) genes, 22 transfer RNA (tRNA) genes and one control region. The mt genomes of Blattodea exhibited a characteristics of a high A+T composition (70.7%-74.3%) and dominant usage of the TAA stop codon. The AT content of the whole mt genome, PCGs and total tRNAs in G. portentosa was the lowest in known cockroaches. The presence of a 71-bp intergenic spacer region between trnQ and trnM was a unique feature in B. dubia, but absent in other cockroaches, which can be explained by the duplication/random loss model. Based on the nucleotide and amino acid datasets of the 13 PCGs genes, neighbor-joining (NJ), maximum parsimony (MP), maximum likelihood (ML) and bayesian inference (BI) analyses were used to rebuild the phylogenetic relationship of cockroaches. All phylogenetic analyses consistently placed Isoptera as the sister cluster to Cryptocercidae of Blattodea. Ectobiidae and Blaberidae (Blaberoidea) formed a sister clade to Blattidae. Corydiidae is a sister clade of all the remaining cockroach species with a high value in NJ and MP analyses of nucleotide and amino acid datasets, and ML and BI analyses of the amino acid dataset.

A global molecular phylogeny and timescale of evolution for Cryptocercus woodroaches.

Cryptocercus is a genus of sub-social wood-feeding cockroaches that represents the sister group to the eusocial termites. We generated mitochondrial (12S+16S rRNA, COII), nuclear (28S rRNA) and Blattbacterium endosymbiont (16S+23S rRNA) sequence data for 8 new Chinese species, and combined these with previously available data to undertake the most extensive analysis of phylogenetic relationships within the genus to date. As expected, phylogenetic relationships among Blattabacterium strains were found to be congruent with those of their hosts. Three major clades were found to exist in Asian populations, one representing taxa from the Hengduan mountains in Southwestern China, a second including taxa from Russia, Korea, Northeastern China, and Yunnan in the Hengduan Mountains, and a third including taxa from the Qinling Mountains and Daba Mountains in Central China. A molecular dating analysis using 7 termite fossils to calibrate the molecular clock indicated that the divergence of American and Asian Cryptocercus occurred 55.09Ma (41.55-72.28Ma 95% CI), and that the radiations of American and Asian taxa occurred 28.48Ma (20.83-37.95Ma 95% CI) and 20.97Ma (15.78-27.21Ma 95% CI) respectively. Reconstruction of ancestral geographic distributions using S-DIVA suggested Cryptocercus was originally distributed across both continents, as opposed to ancestral migration of Cryptocercus from one continent to the other. The last common ancestor of Asian Cryptocercus was inferred to have existed in Central China. An examination of male chromosome numbers in Asian Cryptocercus showed that diploid numbers vary from 2n=15 to 2n=41, and indicates the presence of eight new species. Our study represents the most comprehensive phylogenetic and biogeographic study yet performed for this important group of cockroaches.

A taxonomic study of the beetle cockroaches (Diploptera Saussure) from China, with notes on the genus and species worldwide (Blattodea: Blaberidae: Diplopterinae).

Four taxa of beetle cockroaches (Diploptera Saussure, 1864) from South China are described and illustrated, viz., two new species D. elliptica sp. n. and D. naevus sp. n., one new subspecies D. nigrescens guani subsp. n. and one widespread known species D. punctata (Eschscholtz, 1822). The genus and known species from around the world are discussed based on types and other specimens. D. pulchra Anisyutkin, 2007 is now regarded as a junior synonym of D. bicolor Hanitsch, 1925. Whether the populations of D. punctata represent or not different species needs to be studied in the future. A key, a distribution map, and photos of species are provided.