The Neotropical Region is known for its biodiversity and ranks third in number of known termite species. However, biogeographic and phylogeographic information of termites of this region is limited compared to other world geographic regions. Nasutitermes corniger is widely distributed in the region and is of considerable economic importance. The goal of this study was to describe the phylogeography of N. corniger in the Neotropical Region, to better understand its evolutionary processes.

Labial glands are present in all castes and developmental stages of all termite species. In workers, their secretion contains a food-marking pheromone and digestive enzymes, while soldier secretion plays a defensive role. However, these functions were studied only in a limited set of species, and do not allow drawing general conclusions. Hence, we have investigated the chemical composition of the labial gland extracts from soldiers and workers in 15 termite species belonging to 6 families using an integrative approach based on proteomic and small-molecule profiling. We confirmed the presence of hydroquinone and cellulase in the labial glands of workers, and we identified new toxic compounds in soldiers and workers of several species. Our results highlight the dual role of labial gland secretion, i.e. the defensive role in soldiers and workers of several termite species, and the digestive function in workers.

The mitochondrial genome of the Reticulitermes ovatilabrum is 15,913 bp in length and encodes 37 genes including 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNA), 2 ribosomal RNA genes (rRNA), and a non-coding control region (D-loop). The percentage of A/T (65.59%) is much higher than that of C/G (34.41%). The phylogenetic tree revealed that R. ovatilabrum was closest to R. kanmonensi and R. periflaviceps. The mitochondrial genome of the R. ovatilabrum provides a resource for evolutional analysis within termites especially Reticulitermes.

The soldier of Rugitermes aridus sp. nov. is described from a xeric, termite-depauperate region of central Peru. Rugitermes rufus sp. nov. and R. volcanensis sp. nov. are described from soldiers and dealated imagos collected in a mesic forest of Amboró National Park in western Bolivia. The imago of R. rufus is unique among all described Rugitermes species in that the head capsule is reddish orange and the pronotum is brown. The imago head and pronotum are both brown in R. volcanensis. A phylogenetic and GMYC barcode analyses were performed with the COI gene. These analyses confirmed the three new species and revealed a high undescribed diversity of Rugitermes in the New World.

We report the first complete mitochondrial genome of an important pest of timber, the drywood termite Cryptotermes havilandi. The gene content and synteny of the mitochondrial genome of C. havilandi is identical to that of other termite species reported to date. It is composed 13 protein-coding genes, two ribosomal RNA genes, and 22 transfer RNA genes. Our phylogenetic tree, that includes the mitochondrial genomes of 14 species of Kalotermitidae, including C. havilandi, resolves the phylogenetic position of C. havilandi within Kalotermitidae.

The subfamily Syntermitinae comprises a group of Neotropical termites with 18 genera and 101 species described. It has been considered a natural group, but relationships among the genera within the subfamily remain uncertain, and some genera appear to be non-monophyletic. Here, we provide a comprehensive phylogeny including six Neotropical species of Termitinae as outgroup, 42 Syntermitinae species as ingroup, 92 morphological characters (from external and internal anatomy of soldier and worker castes) and 117 molecular sequences (109 obtained for this study and 8 from GenBank) of 4 gene regions (41 and 22 from Cytochrome Oxidase I and II respectively, 19 from Cytochrome b, and 35 from 16S rDNA). Morphological and molecular data were analyzed in combination, with the Bayesian inference method, and the important aspects of termite biology, defense and feeding habits are discussed based on the resulting tree. Although useful for providing diagnostic characters, the morphology of the soldier caste reveals several cases of convergence; whereas the feeding habit shows indications of evolutionary significance.

Rugitermesursulae sp. nov. is described from a sample collected inside a dead branch in a tropical dry forest of Colombia's Caribbean coast using molecular information and external morphological characters of the imago and soldier castes. Rugitermesursulae sp. nov. soldiers and imagoes are the smallest among all described Rugitermes species. The imago's head capsule coloration is dark castaneous, while the pronotum is contrastingly pale yellow. Our description includes soldier characters, such as subflangular elevation and shape of the antennal sockets, that can help in identification of samples lacking imagoes.

Beta-glucosidase (BG) is known as a multifunctional enzyme for social maintenance in terms of both cellulose digestion and social communication in termites. However, the expression profiles of each BG gene and their evolutionary history are not well understood. First, we cloned two types of BG homologs (RsBGI and RsBGII) from the termite Reticulitermes speratus (Kolbe). Gene expression analyses showed that RsBGI expression levels of primary queens and kings from 30 to 100 days after colony foundation were high, but those of reproductives dropped after day 400. Extremely low gene expression levels of RsBGI were observed in eggs, whereas workers had significantly higher expression levels than those of soldiers and other colony members. Consequently, RsBGI gene expression levels changed among each developmental stage, and RsBGI was shown to be involved in cellulose digestion. On the other hand, the RsBGII gene was consistently expressed in all castes and developmental stages examined, and notable expression changes were not observed among them, including in eggs. It was indicated that RsBGII is a main component involved in social communication, for example, the egg-recognition pheromone shown in this species previously. Finally, we obtained partial gene homologs from other termite and cockroach species, including the woodroach (genus Cryptocercus), which is the sister group to termites, and performed molecular phylogenetic analyses. The results showed that the origin of the BG gene homologs preceded the divergence of termites and cockroaches, suggesting that the acquisition of multifunctionality of the BG gene also occurred in cockroach lineages.

We present the description of a new genus and species of soldierless termites from South America. Rustitermes boteroi Constantini, Castro & Scheffrahn, gen. et sp. nov. can be identified by the morphology of the enteric valve, with six slightly asymmetric cushions, each one forming a central pouch made of scales smaller than those between the cushions. The new genus features two characteristic rows of thick bristles on the interior margin of the fore tibia, and is supported by COI molecular sequence data. This species is distributed from Tobago to northern Argentina.

Termites have developed a wide array of defensive mechanisms. One of them is the mandibulate soldier caste that crushes or pierces their enemies. However, in several lineages of Termitinae, soldiers have long and slender mandibles that cannot bite but, instead, snap and deliver powerful strikes to their opponents. Here, we use morphological and molecular evidence to describe Roisinitermesebogoensis Scheffrahn, gen. & sp. n. from near Mbalmayo, Cameroon. Soldiers of R.ebogoensis are unique among all other kalotermitid soldiers in that they possess snapping mandibles. The imago of R.ebogoensis is also easily distinguished from all other Kalotermitidae by the lack of ocelli. Our study reveals a new case of parallel evolution of snapping mandibles in termites, a complex apparatus responsible of one of the fastest biological acceleration rates measured to date.

Termites are major decomposers in terrestrial ecosystems and the second most diverse lineage of social insects. The Kalotermitidae form the second-largest termite family and are distributed across tropical and subtropical ecosystems, where they typically live in small colonies confined to single wood items inhabited by individuals with no foraging abilities. How the Kalotermitidae have acquired their global distribution patterns remains unresolved. Similarly, it is unclear whether foraging is ancestral to Kalotermitidae or was secondarily acquired in a few species. These questions can be addressed in a phylogenetic framework. We inferred time-calibrated phylogenetic trees of Kalotermitidae using mitochondrial genomes of ∼120 species, about 27% of kalotermitid diversity, including representatives of 21 of the 23 kalotermitid genera. Our mitochondrial genome phylogenetic trees were corroborated by phylogenies inferred from nuclear ultraconserved elements derived from a subset of 28 species. We found that extant kalotermitids shared a common ancestor 84 Ma (75-93 Ma 95% highest posterior density), indicating that a few disjunctions among early-diverging kalotermitid lineages may predate Gondwana breakup. However, most of the ∼40 disjunctions among biogeographic realms were dated at <50 Ma, indicating that transoceanic dispersals, and more recently human-mediated dispersals, have been the major drivers of the global distribution of Kalotermitidae. Our phylogeny also revealed that the capacity to forage is often found in early-diverging kalotermitid lineages, implying the ancestors of Kalotermitidae were able to forage among multiple wood pieces. Our phylogenetic estimates provide a platform for critical taxonomic revision and future comparative analyses of Kalotermitidae.

The fauna of termites (Isoptera) preserved in Early Eocene amber from the Cambay Basin (Gujarat, India) are described and figured. Three new genera and four new species are recognized, all of them Neoisoptera - Parastylotermes krishnai Engel & Grimaldi, sp. n. (Stylotermitidae); Prostylotermes kamboja Engel & Grimaldi, gen. et sp. n. (Stylotermitidae?); Zophotermes Engel, gen. n., with Zophotermes ashoki Engel & Singh, sp. n. (Rhinotermitidae: Prorhinotermitinae); and Nanotermes isaacae Engel & Grimaldi, gen. et sp. n. (Termitidae: Termitinae?). Together these species represent the earliest Tertiary records of the Neoisoptera and the oldest definitive record of Termitidae, a family that comprises >75% of the living species of Isoptera. Interestingly, the affinities of the Cambay amber termites are with largely Laurasian lineages, in this regard paralleling relationships seen between the fauna of bees and some flies. Diversity of Neoisoptera in Indian amber may reflect origin of the amber deposit in Dipterocarpaceae forests formed at or near the paleoequator.

Reticulitermes chinensis Snyder is an important pest species in China. Serratia marcescens Bizio (SM1) is a potent biological bacterium. In our lab, we found that SM1 can kill R. chinensis. To date, the interaction between R. chinensis and SM1 has not been studied. Here, we explored immune responses of R. chinensis against SM1 using transcriptome sequencing. To elucidate immune-related genes, we identified 126,153 unigenes from R. chinensis. In total, 178 immune-related differentially expressed genes (DEGs) were identified. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that many cellular responses were enriched in the top 20 terms. Then, we systematically analyzed several cellular immune pathways involved in the response of R. chinensis to SM1, including phagocytosis, autophagy, and endocytosis pathways. Furthermore, the expression profiles of the cellular immune-related genes were assessed using quantitative reverse-transcription PCR, and the expression levels of the selected genes were upregulated. Further results revealed SM1-mediated activation of humoral immune responses genes, including Toll, IMD, and melanization pathways, which suggested the involvement of humoral immune responses in the defense against SM1. This research elucidated the mechanisms underlying the immune defense of R. chinensis against SM1, providing a solid theoretical basis for exploiting new immune suppressive agents to control R. chinensis. Moreover, this study will facilitate the better control of R. chinensis using SM1.

The subsocial life style and wood-feeding capability of Cryptocercus gives us an evolutionary key to unlock some outstanding questions in biology. With the advent of the Genomics Era, there is an unprecedented opportunity to address the evolution of eusociality and the acquisition of lignocellulases at the genetic level. However, to quantify gene expression, an appropriate normalization strategy is warranted to control for the non-specific variations among samples across different experimental conditions. To search for the internal references, 10 housekeeping genes from a gut transcriptome of a wood-feeding cockroach, Cryptocercus punctulatus, were selected as the candidates for the RT-qPCR analysis. The expression profiles of these candidates, including ACT, EF1α, GAPDH, HSP60, HSP70, αTUB, UBC, RPS18, ATPase and GST, were analyzed using a panel of analytical tools, including geNorm, NormFinder, BestKeeper, and comparative ΔCT method. RefFinder, a comprehensive ranking system integrating all four above-mentioned algorithms, rated ACT as the most stable reference gene for different developmental stages and tissues. Expression analysis of the target genes, Hex-1 and Cell-1, using the most or the least appropriate reference genes and a single or multiple normalizers signified this research. Our finding is the first step toward establishing a standardized RT-qPCR analysis in Cryptocercus.

Coptotermes suzhouensis (Isoptera: Rhinotermitidae) is a significant subterranean termite pest of wooden structures and is widely distributed in southeastern China. The complete mitochondrial DNA sequence of C. suzhouensis was analyzed in this study. The mitogenome was a circular molecule of 15,764 bp in length, which contained 13 protein-coding genes (PCGs), 22 transfer RNA genes, two ribosomal RNA genes, and an A+T-rich region with a gene arrangement typical of Isoptera mitogenomes. All PCGs were initiated by ATN codons and terminated by complete termination codons (TAA), except COX2, ND5, and Cytb, which ended with an incomplete termination codon T. All tRNAs displayed a typical clover-leaf structure, except for tRNASer(AGN), which did not contain the stem-loop structure in the DHU arm. The A+T content (69.23%) of the A+T-rich region (949 bp) was higher than that of the entire mitogenome (65.60%), and two different sets of repeat units (A+B) were distributed in this region. Comparison of complete mitogenome sequences with those of Coptotermes formosanus indicated that the two taxa have very high genetic similarity. Forty-one representative termite species were used to construct phylogenetic trees by maximum likelihood, maximum parsimony, and Bayesian inference methods. The phylogenetic analyses also strongly supported (BPP, MLBP, and MPBP = 100%) that all C. suzhouensis and C. formosanus samples gathered into one clade with genetic distances between 0.000 and 0.002. This study provides molecular evidence for a more robust phylogenetic position of C. suzhouensis and inferrs that C. suzhouensis was the synonymy of C. formosanus.

After more than one hundred years, a new drywood termite of the genus Proneotermes is described from the tropical dry forest in the Caribbean coast of Colombia. Morphological and genetic analyses are given for Proneotermes macondianussp. n. This termite occurs in tropical dry forests in small colonies inside thin branches of dry wood. The soldier of Proneotermes macondianus is smaller and the genal horns are angled outward compared to the other two described Proneotermes species. The imago wings are unusually short and wide. Genetic analyses for COII, 12S, and 16S genes show less than three percent difference between sample localities of Proneotermes macondianus. Intergeneric comparison with selected kalotermitid genera indicates that Bifiditermes is the most closely related genus of those sequenced. New morphological descriptions and morphometric measurements of Proneotermes latifrons based on the soldier caste are also included. Neotropical locality records for Proneotermes latifrons and Proneotermes perezi are provided.

Disjunctitermes insularisgen. n. & sp. n. is described from workers collected on Guadeloupe and in Peru and is the first soldierless termite found on a deep-water island. As with many soldierless and soil-feeding termite species, the enteric valve morphology is an essential diagnostic character of D. insularis. The D. insularis sequence cluster, derived from a barcode analysis with twelve other described genera of New World Apicotermitinae, is well resolved. Results of a stochastic dynamic spread model suggest that the occurrence of D. insularis on Guadeloupe may be the result of a pre-Colombian overwater dispersal event from mainland South America.

Carbon dioxide (CO2) is a pervasive chemical stimulus that plays a critical role in insect life, eliciting behavioral and physiological responses across different species. High CO2 concentration is a major feature of termite nests, which may be used as a cue for locating their nests. Termites also survive under an elevated CO2 concentration. However, the mechanism by which elevated CO2 concentration influences gene expression in termites is poorly understood.

The insect olfactory system plays pivotal roles in insect survival and reproduction through odor detection. Morphological and physiological adaptations are caste-specific and evolved independently in workers, soldiers, and reproductives in termites. However, it is unclear whether the olfactory system is involved in the division of labor in termite colonies. In the present study, the antennal sensilla of alates, workers, soldiers, nymphs, and larvae of the termite Reticulitermes aculabialis Tsai et Hwang ( Isoptera: Rhinotermitidae) were investigated. Transcriptomes were used to detect olfactory genes, and differential expression levels of olfactory genes were confirmed in various castes by qRT-PCR analysis. Nine types of sensilla were identified on the antennae of R. aculabialis, and soldiers possessed all 9 types. In 89,475 assembled unigenes, we found 16 olfactory genes, including 6 chemosensory protein (CSP) and 10 odorant-binding protein (OBP) genes. These OBP genes included 8 general odorant-binding protein genes (GOBPs) and 2 pheromone-binding protein-related protein (PBP) genes. Five CSP genes were more highly expressed in alates than in workers, soldiers, larvae, and nymphs, and the expression levels of CSP6 were significantly higher in nymphs. Seven GOBP and two PBP genes exhibited significantly higher expression levels in alates, and there were no significant differences in the expression levels of GOBP2 among workers, soldiers, alates, and larvae. These results suggest that alates, as primary reproductives, have unique expression patterns of olfactory genes, which play key roles in nuptial flight, mate seeking, and new colony foundation.

The recombinant genotypes that can be produced when closely related species mate improve the genetic diversity of the population. Among closely related species, the link between interspecific reproduction behaviors and genetic diversity has barely been studied. Reticulitermes chinensis and R. flaviceps, which live close to each other, were used as research subjects in our study to find out how preferring conspecifics affects reproductive behavior between species. We discovered that neither R. chinensis nor R. flaviceps displayed preference behavior for conspecifics. Males of R. chinensis and R. flaviceps chased and groomed not only intraspecific females but also interspecific females. In a brief period of time, 2 mating behaviors, intra- and interspecific mating, were also observed. There were no significant differences in the duration of each behavior (tandem, grooming, and mating) between interspecies and intraspecies partners. Moreover, genetic analysis showed both interspecific mating and intraspecific mating can produce living offspring when the 2 types of mating occur in a colony. Our findings showed that there was no obvious intraspecific preference between the 2 species of termite Reticulitermes when it came to tandem, grooming, and mating, which not only makes it easier for interspecific hybridization to occur but also sheds light on the genetic diversity.