Termite Neotermes Koshunensis Research Articles

Damage and ecological characteristics of termite Neotermes koshunensis on Taiwan cherry tree Cerasus campanulata in Okinawa Island

Abstract Termites are typical house pests that can also be harmful pests for living trees, although this topic has not received much attention. To clarify the damage to trees caused by dry-wood termites (Neotermes koshunensis), and the ecological characteristics of such damage, we conducted a study on Taiwan cherry trees (Cerasus campanulata) grown in parks, historical sites, and streets in the central-southern area of Okinawa Island, a subtropical region. Damage by N. koshunensis was confirmed in 21 of the 36 sites surveyed (58.3%) and in 76 of the 1076 trees surveyed (7.1%). However, damage by Formosan subterranean termites (Coptotermes formosanus), a house pest, was only observed at six sites (16.7%), and 10 trees (0.9%), indicating that most damage was caused by N. koshunensis. Furthermore, the probability of the presence of damaged trees close to other damaged trees was significantly higher than the probability of the presence of damaged trees close to undamaged trees, implying that destructive colonies spread from damaged trees to other trees nearby. Therefore, care for trees in the initial stages of termite invasion seems to be the best method for preventing the spread N. koshunensis damage to other trees in the same area. To our knowledge, this study is the first to report that dry-wood termite N. koshunensis are harmful pests for Taiwan cherry trees in Okinawa Island. Our results could be used to establish dry-wood termite countermeasures for garden tree species.

Characteristics of dispersal flight and disperser production in an Asian dry-wood termite, Neotermes koshunensis (Isoptera, Kalotermitidae)

Differences in the dispersal flight patterns among termite families are correlated with the difference between the two life history characteristics exhibited by this group: “separate-piece nesters” versus “single-piece nesters.” However, information remains limited on the phenology and the life history characteristics of single-piece nesters, impeding our understanding of this topic. We report the flight phenology of an Asian single-piece nester termite Neotermes koshunensis on Okinawa Island, Japan. In 1983–1984, a light-trap survey showed that N. koshunensis exhibited an extended dispersal flight period from late April to early November, peaking in June, with a female-biased sex ratio. Between 1983 and 2012, the collection of 134 whole colonies of N. koshunensis from the surrounding area confirmed the presence of alates and pre-alate nymphs within the colonies over 7 months, reflecting the extended flight season of this termite species, probably in association with the extended dispersal flight season. However, in some cases, alates and pre-alate nymphs were also retained in the colonies after the dispersal flight season (i.e., winter, from December to February). The daily number of trapped alates in 1983 was positively correlated with temperature and relative humidity; however, alate production inside the colony was also positively correlated with temperature, relative humidity, and precipitation. Thus, these environmental factors might promote the flight activity of this termite by enhancing alate production inside the colony. Furthermore, temperature also had a significantly positive effect in the model incorporating the density of alates in the colony, along with environmental factors; thus, temperature might facilitate the release of alate from colonies. The accumulation of information on the phenology and life history characteristics of alate advances our understanding of the different dispersal strategies used by termites, providing insights into how the different families have evolved.

Facultative parthenogenesis in the Ryukyu drywood termite Neotermes koshunensis.

Parthenogenesis is a relatively rare reproductive mode in nature compared to sex. In social insects, the evolution of parthenogenesis has a notable impact on their life histories. Some termites with parthenogenetic ability produce numerous non-dispersing supplementary queens asexually, whereas other castes are produced via sexual reproduction. This asexual queen succession (AQS) system is adaptive because hundreds of the asexual queens improve the reproductive potential of the colony and maintain the genetic diversity within the colony. However, the evolutionary process of the AQS system remains unclear because parthenogenetic species without this system are unknown. Here, we report facultative parthenogenesis in the drywood termite Neotermes koshunensis. Although the eggs produced by females isolated from males hatched, the hatching rate of those eggs was lower than that of the eggs produced by females kept with males. These parthenogenetic offspring inherited only the maternal alleles and showed high homozygosity, which indicates that the mechanism of ploidy restoration is terminal fusion. A previous study showed that most colonies of this species have a single queen or orphan; thus, the AQS system has not evolved despite their parthenogenetic ability. Further investigations of N. koshunensis will reveal how parthenogenesis evolved and its role in the insect societies.

Comparative Analysis of the Bonding Modes between Two Antidiabetic Drugs with beta-Glucosidases from Different Species

β-glucosidase is one of the critical enzymes to the type 2 diabetes, which belongs to a large family of glycoside hydrolases. In this article, we attempted to explore the binding modes between two drugs and β-glucosidases by comparing their bonding modes with β-glucosidases from different species. Results showed that the binding orientations and geometrical conformation of synthetic drug (miglitol) and natural product (quercetin) were all different in all active sites, which may be related to the flexibility of the molecules. Compared with the conformations obtained by density functional theory calculations at the B3LYP/6-31G* level, the docking conformations indicated that the -CH2CH2OH group of miglitol and the B ring of quercetin were the most critical groups to the stabilities in the active sites. Finally, protein sequence alignment was performed under default parameters. Although the sequence similarity is not very high between these β-glucosidases, the residues related to the active sites were conservative; especially the one involved in H-bond interactions between three species, namely soil metagenome, Micrococcus antarcticus and termite Neotermes koshunensis.

Refinement of Methods for Sexing Instars and Caste Members in Neotermes koshunensis (Isoptera, Kalotermitidae)

Exact sexing of individuals is important to an understanding of termite societies. We found that the female-specific sexual character of the elongated seventh sternite, known in other termites, is useful for distinguishing the sex of individuals of the dry-wood termite Neotermes koshunensis (Shiraki) from the fifth instar onward. The method described is highly efficient at sexing instars, nymphs, soldiers and alates.

The Unusual Neotenic System of the Asian Dry Wood Termite, Neotermes koshunensis (Isoptera: Kalotermitidae)

In most lower termites, colonies are headed by neotenic reproductives of both sexes after the primary reproductives (i.e., the queen and king) are lost. The production of a neotenic sexual is inhibited by the presence of a primary reproductive of the same sex. We found an exception in the caste system of the dry wood termite Neotermes koshunensis (Kalotermitidae). The neotenic caste is exclusively male. Moreover, production of male neotenics is completely inhibited not only by the presence of a king but also by the presence of a queen. Therefore, it is likely to be difficult for N. koshunensis colonies to recover their reproductive pairs.

Antennal cropping in the Asian dry-wood termite, Neotermes koshunensis

Antennal cropping, a behavior inferred to exist because queens and kings have shorter antennae than fresh alates, is widespread in termites. However, the proximate and ultimate mechanisms underlying this phenomenon remain unclear. We studied the occurrence of antennal cropping in queens and kings of the dry-wood termite Neotermes koshunensis (Kalotermitidae). Observation of the antennal tip structure with scanning electron microscopy and the occurrence of antennal cropping in new kings and queens reared in isolation indicated that self-cropping is an important proximate mechanism. Previous studies inferred that antennal cropping may play a key role in the life-history of alates at the colony-founding stage. However, we also found antennal cropping in adultoid reproductives (secondary reproductives) that had not experienced a colony founding. We propose a new hypothesis that antennal cropping is important for individuals in regulating their physiology when they change from the non-reproducing to the reproducing phase.

Heterologous expression and characterization of a glucose-stimulated β-glucosidase from the termite Neotermes koshunensis in Aspergillus oryzae

Neotermes koshunensis is a lower termite that secretes endogenous β-glucosidase in the salivary glands. This β-glucosidase (G1NkBG) was successfully expressed in Aspergillus oryzae. G1NkBG was purified to homogeneity from the culture supernatant through ammonium sulfate precipitation and anion exchange, hydrophobic, and gel filtration chromatographies with a 48-fold increase in purity. The molecular mass of the native enzyme appeared as a single band at 60 kDa after gel filtration analysis, indicating that G1NkBG is a monomeric protein. Maximum activity was observed at 50 °C with an optimum pH at 5.0. G1NkBG retained 80% of its maximum activity at temperatures up to 45 °C and lost its activity at temperatures above 55 °C. The enzyme was stable from pH 5.0 to 9.0. G1NkBG was most active towards laminaribiose and p-nitrophenyl-β-D-fucopyranoside. Cellobiose, as well as cello-oligosaccharides, was also well hydrolyzed. The enzyme activity was slightly stimulated by Mn(2+) and glycerol. The K(m) and V(max) values were 0.77 mM and 16 U/mg, respectively, against p-nitrophenyl-β-D-glucopyranoside. An unusual finding was that G1NkBG was stimulated by 1.3-fold when glucose was present in the reaction mixture at a concentration of 200 mM. These characteristics, particularly the stimulation of enzyme activity by glucose, make G1NkBG of great interest for biotechnological applications, especially for bioethanol production.

Structural and functional analysis of three β-glucosidases from bacterium Clostridium cellulovorans, fungus Trichoderma reesei and termite Neotermes koshunensis

β-glucosidases (EC 3.2.1.21) cleave β-glucosidic linkages in disaccharide or glucose-substituted molecules and play important roles in fundamental biological processes. β-Glucosidases have been widely used in agricultural, biotechnological, industrial and medical applications. In this study, a high yield expression (70-250 mg/l) in Escherichia coli of the three functional β-glucosidase genes was obtained from the bacterium Clostridium cellulovorans (CcBglA), the fungus Trichoderma reesei (TrBgl2), and the termite Neotermes koshunensis (NkBgl) with the crystal structures of CcBglA, TrBgl2 and NkBgl, determined at 1.9Å, 1.63Å and 1.34Å resolution, respectively. The overall structures of these enzymes are similar to those belonging to the β-retaining glycosyl hydrolase family 1, which have a classical (α/β)(8)-TIM barrel fold. Each contains a slot-like active site cleft and a more variable outer opening, related to its function in processing different lengths of β-1,4-linked glucose derivatives. The two essential glutamate residues for hydrolysis are spatially conserved in the active site. In both TrBgl2 and NkBgl structures, a Tris molecule was found to bind at the active site, explaining the slight inhibition of hydrolase activity observed in Tris buffer. Manganese ions at 10mM exerted an approximate 2-fold enzyme activity enhancement of all three β-glucosidases, with CcBglA catalyzing the most efficiently in hydrolysis reaction and tolerating Tris as well as some metal inhibition. In summary, our results for the structural and functional properties of these three β-glucosidases from various biological sources open important avenues of exploration for further practical applications.

Molecular Phylogenetic Position of the Genera Stephanonympha and Caduceia (Parabasalia) Inferred from Nuclear Small Subunit rRNA Gene Sequences

Nuclear small subunit (SSU) rRNA gene sequences were obtained by polymerase chain reaction from trichomonad symbionts of termites that belong to the Devescovinidae (Caduceia versatilis) and polymastigont Calonymphidae (Stephanonympha nelumbium). The unidentified SSU rRNA sequence Nk3, previously obtained from the termite Neotermes koshunensis, has also been shown to derive from a Stephanonympha sp. by in situ hybridization. These sequences were analysed in a broad phylogeny including nearly all identified parabasalid sequences available in the databases, and some as yet unidentified sequences likely deriving from the new order Cristamonadida (Devescovinidae, Calonymphidae, and hypermastigids Lophomonadida). A global phylogeny of parabasalids reveals a partial agreement between the clades identified in this work and the last classification of this phylum into four orders. However, this classification is still incongruent with our data and new taxonomic considerations are proposed. The analysis confirms the monophyly of the Cristamonadida and separates this order into two groups: the first unites nearly all the Devescovinidae including Caduceia and the Calonymphidae Coronympha and Metacoronympha, whereas the second group is composed of a few Devescovinidae, Lophomonadida, and Calonymphidae such as Stephanonympha. Caduceia is closely related to Devescovina, corroborating the marked morphological similarity between these two genera whereas Stephanonympha groups together with the Calonymphidae Snyderella and Calonympha. These data also confirm the polyphyly of the families Devescovinidae and Calonymphidae and support the arrangement of the axostyle-pelta complexes as a valuable character for taxonomic considerations within the Calonymphidae.

Alate differentiation and compound-eye development in the dry-wood termite Neotermes koshunensis (Isoptera, Kalotermitidae)

In social insects, caste-specific characters develop in the postembryonic differentiation processes. However, the mechanisms of caste-specific organ development have yet to be elucidated. In order to obtain insights into the relationship between caste differentiation and the regulation of organ development, we determined the caste-developmental pathway and observed compound-eye development accompanying alate differentiation in the dry-wood termite, Neotermes koshunensis. As previously reported in other Neotermes, this species has a linear caste-developmental pathway, comprising six larval- and two nymphal-instar stages. Although the apparent eye formation occurs during the last nymphal stages, just prior to the imaginal molt, individuals possess eye primordia from the first larval-instar stage. The outer morphological structure of the eye was observed from the third larval-instar stage. The detailed differentiation of cells constituting ommatidia appeared to occur in relatively young larval instars (fourth stage), although the pigmentation of pigment cells and detailed structural formation of ommatidia occurred during the final stage of alate development, i.e., during the late second nymphal-instar stage. This suggests that eye development is arrested in the larval stages, and then resumed during the late nymphal stage to complete functional eye formation, which is required for nuptial flight. In comparison to major hemimetabolous insects, which possess functional compound eyes even at the first instar larva, this termite species shows the heterochronic shift in terms of compound-eye development.

Identification and in situ Detection of Two Lineages of Bacteroidales Ectosymbionts Associated with a Termite Gut Protist, Oxymonas sp.

Bacterial attachments often cover the entire surface of flagellated protists in the guts of termites. Based on PCR-amplified 16S rRNA gene sequences, we investigated the phylogenetic positions of the rod-shaped bacteria (ectosymbionts) attached to the protist Oxymonas sp. in the termite Neotermes koshunensis. Two distinct and unique lineages of the ectosymbionts within the order Bacteroidales were identified, each belonging to a cluster exclusively comprised of the sequences from termite gut. We designed two oligonucleotide probes specific for the two lineages, and successfully detected the ectosymbionts, each of which distributed over the entire surface of Oxymonas sp. However, few cells of Oxymonas sp. simultaneously harbored both lineages of the ectosymbionts.

コウシュンシロアリＮｅｏｔｅｒｍｅｓ ｋｏｓｈｕｎｅｎｓｉｓ（Ｓｈｉｒａｋｉ）におけるカースト間セルラーゼ活性の差異と共生原生動物除去の影響

Distributions of two types of cellulase activity (endo-β-1,4-glucanase [EC 3.2.1.4] and β-glucosidase [EC 3.2.1.21]) were measured in the salivary glands and the gut of the xylophagous termite Neotermes koshunensis, and activity was compared among pseudoworker-caste insects, soldier-caste insects and defaunated pseudoworkers in which the symbiotic flagellates were removed by exposing the termites to sunlight for 120 min. Strong cellulase activity was detected in the salivary glands and the hindgut of the pseudoworkers, while the defaunated termites showed a marked reduction in cellulase activity of the hindgut. The salivary glands of soldier termites showed significantly lower cellulase activity than those of pseudoworkers, whereas the hindgut of the soldiers possessed a similar level of activity to that of the pseudoworkers. The present results suggest that the hindgut is the primary site of cellulose digestion in pseudoworkers and soldiers of N. koshuensis, and that the salivary cellulases also play an indispensable role for cellulose digestion in pseudoworkers.

Molecular phylogeny of parabasalids inferred from small subunit rRNA sequences, with emphasis on the Devescovinidae and Calonymphidae (Trichomonadea)

Small subunit rRNA sequences were obtained by polymerase chain reaction from trichomonad symbionts of termites that belong to the polymastigont Calonymphidae, including Snyderella tabogae, Calonympha grassii, and Metacoronympha senta. The yet-unidentified sequence Nk9 previously obtained from the termite Neotermes koshunensis, has also been shown to derive from the Devescovinidae Devescovina sp. by in situ hybridization. These new sequences were analyzed by distance, parsimony, and likelihood methods in a broad phylogeny including all identified parabasalid sequences available in databases. All analyses revealed the emergence of a very well supported Devescovinidae/Calonymphidae group but showed an unexpected dichotomy of the Calonymphidae represented by the “ Coronympha” and “ Calonympha” groups. It strongly suggests that the polymastigont state observed in the Calonymphidae might be explained by at least two independent evolutionary events. In a second phylogenetic analysis, some yet-unidentified parabasalid sequences likely deriving from the Devescovinidae/Calonymphidae taxa, were added to our data set. This analysis confirmed the polyphyly of the Calonymphidae. A tentative identification is proposed for each of these sequences, and hypotheses on the origin of the Devescovinidae and Calonymphidae are discussed. Tritrichomonas foetus or a close relative might be the best candidate for the ancestor of the Devescovinidae, fairly consistent with morphology-based hypotheses. Regarding the Calonymphidae, the origin of the “ Coronympha” group might be found within the Devescovinidae, related to Foaina, whereas the “ Calonympha” group may directly descend from Tritrichomonas or related species.