Undertaking behaviour is an essential activity in social insects. Corpses are often recognized by a postmortem change in a chemical signature. Reticulitermes flavipes responded to corpses within minutes of death. This undertaking behaviour did not change with longer postmortem time (24 h); however, R. flavipes exhibited distinctively different behaviours toward dead termites from various origins. Corpses of the congeneric species, Reticulitermes virginicus, were buried onsite by workers with a large group of soldiers guarding the burial site due to the risk of interspecific competition; while dead conspecifics, regardless of colony origin, were pulled back into the holding chamber for nutrient recycling and hygienic purposes. The burial task associated with congeneric corpses was coupled with colony defence and involved ten times more termites than retrieval of conspecific corpses. Our findings suggest elicitation of undertaking behaviour depends on the origin of corpses which is associated with different types of risk.
Termites are highly eusocial insects and show a division of labor whereby
morphologically distinct individuals specialize in distinct tasks. In the
lower termite Reticulitermes flavipes (Rhinotermitidae),
non-reproducing individuals form the worker and soldier castes, which
specialize in helping (e.g., brood care, cleaning, foraging) and
defense behaviors, respectively. Workers are totipotent juveniles that can
either undergo status quo molts or develop into soldiers or neotenic
reproductives. This caste differentiation can be regulated by juvenile
hormone (JH) and primer pheromones contained in soldier head extracts (SHE).
Here we offered worker termites a cellulose diet treated with JH or SHE for
24-hr, or held them with live soldiers (LS) or live neotenic reproductives
(LR). We then determined gene expression profiles of the host termite gut
and protozoan symbionts concurrently using custom cDNA oligo-microarrays
containing 10,990 individual ESTs.
JH was the most influential treatment (501 total ESTs affected), followed by
LS (24 ESTs), LR (12 ESTs) and SHE treatments (6 ESTs). The majority of JH
up- and downregulated ESTs were of host and symbiont origin, respectively;
in contrast, SHE, LR and LS treatments had more uniform impacts on host and
symbiont gene expression. Repeat “follow-up” bioassays
investigating combined JH + SHE impacts in relation to
individual JH and SHE treatments on a subset of array-positive genes
revealed (i) JH and SHE treatments had opposite impacts on gene expression
and (ii) JH + SHE impacts on gene expression were generally
intermediate between JH and SHE.
Our results show that JH impacts hundreds of termite and symbiont genes
within 24-hr, strongly suggesting a role for the termite gut in JH-dependent
caste determination. Additionally, differential impacts of SHE and LS
treatments were observed that are in strong agreement with previous studies
that specifically investigated soldier caste regulation. However, it is
likely that gene expression outside the gut may be of equal or greater
importance than gut gene expression.
Metagenomics; Soldier head extract; Microarray; Caste differentiation; Live soldier; Live reproductive
Previous studies have explored the relationship between termite branch tunnel geometry and foraging efficiency in a model simulation in which foraging efficiency, γ, for two termite species, Coptotermes formosanus Shiraki and Reticulitermes flavipes (Kollar) (Isoptera: Rhinotermitidae), was investigated in response to two variables, the probability of tunnel branching (Pbranch) and the probability of tunnel branch termination (Pterm). It was found that simulated tunnel patterns based on empirical data did not have maximum foraging efficiency. We hypothesized that termites could increase their foraging efficiency in response to landscape heterogeneity. The present study investigated how termites could control the two variables, Pbranch and Pterm, in response to the external environment in terms of tunnel network connectivity. It was found that the best simulated strategy for C. formosanus and R. flavipes termites would occur if both Pbranch and Pterm were increased together. This study provides possible mechanisms for foraging strategies in subterranean termites and a baseline for future empirical work.
Subterranean termite; foraging efficiency; network connectivity; foraging strategy
Undertaking behavior is a significant adaptation to social life in enclosed nests. Workers are known to remove dead colony members from the nest. Such behavior prevents the spread of pathogens that may be detrimental to a colony. To date, little is known about the ethological aspects of how termites deal with carcasses.
Methodology and Principal Findings
In this study, we tested the responses to carcasses of four species from different subterranean termite taxa: Coptotermes formosanus Shiraki and Reticulitermes speratus (Kolbe) (lower termites) and Microcerotermes crassus Snyder and Globitermes sulphureus Haviland (higher termites). We also used different types of carcasses (freshly killed, 1-, 3-, and 7-day-old, and oven-killed carcasses) and mutilated nestmates to investigate whether the termites exhibited any behavioral responses that were specific to carcasses in certain conditions. Some behavioral responses were performed specifically on certain types of carcasses or mutilated termites. C. formosanus and R. speratus exhibited the following behaviors: (1) the frequency and time spent in antennating, grooming, and carcass removal of freshly killed, 1-day-old, and oven-killed carcasses were high, but these behaviors decreased as the carcasses aged; (2) the termites repeatedly crawled under the aging carcass piles; and (3) only newly dead termites were consumed as a food source. In contrast, M. crassus and G. sulphureus workers performed relatively few behavioral acts. Our results cast a new light on the previous notion that termites are necrophobic in nature.
We conclude that the behavioral response towards carcasses depends largely on the nature of the carcasses and termite species, and the response is more complex than was previously thought. Such behavioral responses likely are associated with the threat posed to the colony by the carcasses and the feeding habits and nesting ecology of a given species.
Light microscopy and scanning and transmission electron microscopy were used to examine the in situ morphology of the gut microbiota of Reticulitermes flavipes and Caoptotermes formosanus. Laboratory-maintained termites were used and, for R. flavipes, specimens were also prepared immediately after collection from a natural infestation. The latter endeavor enabled a study of different castes and developmental stages of R. flavipes and revealed differences in the microbiota of field versus laboratory specimens. The termite paunch microbiota consisted of an abundance of morphologically diverse bacteria and protozoa. Thirteen bacterial morphotypes in the paunch were described in detail: seven were observed only in R. flavipes, three were observed only in C. formosanus, and three were common to both termite species. The paunch epithelium was densely colonized by bacteria, many of which possessed holdfast elements that secured them tightly to this tissue and to other bacterial cells. Besides bacteria, the protozoan Pyrsonympha vertens adhered to the paunch epithelium of R. flavipes by means of an attachment organelle. Cuplike indentations were present on the paunch epithelial surface and were sites of bacterial aggregation. Ultrastructural features of cups suggested their involvement in ion absorption. In addition to the paunch, the midgut was also colonized by bacteria that were situated between epithelial microvilli. Results suggest that bacteria are an integral part of the gut ecosystem.
The disruption of host-symbiont interactions through the use of antibiotics can help elucidate microbial functions that go beyond short-term nutritional value. Termite gut symbionts have been studied extensively, but little is known about their impact on the termite's reproductive output. Here we describe the effect that the antibiotic rifampin has not only on the gut microbial diversity but also on the longevity, fecundity, and weight of two termite species, Zootermopsis angusticollis and Reticulitermes flavipes. We report three key findings: (i) the antibiotic rifampin, when fed to primary reproductives during the incipient stages of colony foundation, causes a permanent reduction in the diversity of gut bacteria and a transitory effect on the density of the protozoan community; (ii) rifampin treatment reduces oviposition rates of queens, translating into delayed colony growth and ultimately reduced colony fitness; and (iii) the initial dosages of rifampin had severe long-term fitness effects on Z. angusticollis. Taken together, our findings demonstrate that the antibiotic-induced perturbation of the microbial community is associated with prolonged reductions in longevity and fecundity. A causal relationship between these changes in the gut microbial population structures and fitness is suggested by the acquisition of opportunistic pathogens and incompetence of the termites to restore a pretreatment, native microbiota. Our results indicate that antibiotic treatment significantly alters the termite's microbiota, reproduction, colony establishment, and ultimately colony growth and development. We discuss the implications for antimicrobials as a new application to the control of termite pest species.
Prompted by our limited understanding of the degradation of lignin and lignin-derived aromatic metabolites in termites, we studied the metabolism of monoaromatic model compounds by termites and their gut microflora. Feeding trials performed with [ring-U-(sup14)C]benzoic acid and [ring-U-(sup14)C]cinnamic acid revealed the general ability of termites of the major feeding guilds (wood and soil feeders and fungus cultivators) to mineralize the aromatic nucleus. Up to 70% of the radioactive label was released as (sup14)CO(inf2); the remainder was more or less equally distributed among termite bodies, gut contents, and feces. Gut homogenates of the wood-feeding termites Nasutitermes lujae (Wasmann) and Reticulitermes flavipes (Kollar) mineralized ring-labeled benzoic or cinnamic acid only if oxygen was present. In the absence of oxygen, benzoate was not attacked, and cinnamate was only reduced to phenylpropionate. Similar results were obtained with other, nonlabeled lignin-related phenylpropanoids (ferulic, 3,4-dihydroxycinnamic, and 4-hydroxycinnamic acids), whose ring moieties underwent degradation only if oxygen was present. Under anoxic conditions, the substrates were merely modified (by side chain reduction and demethylation), and this modification occurred at the same time as a net accumulation of phenylpropanoids formed endogenously in the gut homogenate, a phenomenon not observed under oxic conditions. Enumeration by the most-probable-number technique revealed that each N. lujae gut contained about 10(sup5) bacteria that were capable of completely mineralizing aromatic substrates in the presence of oxygen (about 10(sup8) bacteria per ml). In the absence of oxygen, small numbers of ring-modifying microorganisms were found (<50 bacteria per gut), but none of these microorganisms were capable of ring cleavage. Similar results were obtained with gut homogenates of R. flavipes, except that a larger number of anaerobic ring-modifying microorganisms was present (>5 x 10(sup3) bacteria per gut). Neither inclusion of potential cosubstrates (H(inf2), pyruvate, lactate) nor inclusion of hydrogenotrophic partner organisms resulted in anoxic ring cleavage in most-probable-number tubes prepared with gut homogenates of either termite. The oxygen dependence of aromatic ring cleavage by the termite gut microbiota is consistent with the presence, and uptake by microbes, of O(inf2) in the peripheral region of otherwise anoxic gut lumina (as reported in the accompanying paper [A. Brune, D. Emerson, and J. A. Breznak, Appl. Environ. Microbiol. 61:2681-2687, 1995]). Taken together, our results indicate that microbial degradation of plant aromatic compounds can occur in termite guts and may contribute to the carbon and energy requirement of the host.
Social insects such as termites express dramatic polyphenism (the occurrence of multiple forms in a species on the basis of differential gene expression) both in association with caste differentiation and between castes after differentiation. We have used cDNA macroarrays to compare gene expression between polyphenic castes and intermediary developmental stages of the termite Reticulitermes flavipes.
Social insects such as termites express dramatic polyphenism (the occurrence of multiple forms in a species on the basis of differential gene expression) both in association with caste differentiation and between castes after differentiation. We have used cDNA macroarrays to compare gene expression between polyphenic castes and intermediary developmental stages of the termite Reticulitermes flavipes.
We identified differentially expressed genes from nine ontogenic categories. Quantitative PCR was used to quantify precise differences in gene expression between castes and between intermediary developmental stages. We found worker and nymph-biased expression of transcripts encoding termite and endosymbiont cellulases; presoldier-biased expression of transcripts encoding the storage/hormone-binding protein vitellogenin; and soldier-biased expression of gene transcripts encoding two transcription/translation factors, two signal transduction factors and four cytoskeletal/muscle proteins. The two transcription/translation factors showed significant homology to the bicaudal and bric-a-brac developmental genes of Drosophila.
Our results show differential expression of regulatory, structural and enzyme-coding genes in association with termite castes and their developmental precursor stages. They also provide the first glimpse into how insect endosymbiont cellulase gene expression can vary in association with the caste of a host. These findings shed light on molecular processes associated with termite biology, polyphenism, caste differentiation and development and highlight potentially interesting variations in developmental themes between termites, other insects, and higher animals.
We investigated the bacterial gut microbiota from 32 colonies of wood-feeding termites, comprising four Microcerotermes species (Termitidae) and four Reticulitermes species (Rhinotermitidae), using terminal restriction fragment length polymorphism analysis and clonal analysis of 16S rRNA. The obtained molecular community profiles were compared statistically between individuals, colonies, locations, and species of termites. Both analyses revealed that the bacterial community structure was remarkably similar within each termite genus, with small but significant differences between sampling sites and/or termite species. In contrast, considerable differences were found between the two termite genera. Only one bacterial phylotype (defined with 97% sequence identity) was shared between the two termite genera, while 18% and 50% of the phylotypes were shared between two congeneric species in the genera Microcerotermes and Reticulitermes, respectively. Nevertheless, a phylogenetic analysis of 228 phylotypes from Microcerotermes spp. and 367 phylotypes from Reticulitermes spp. with other termite gut clones available in public databases demonstrated the monophyly of many phylotypes from distantly related termites. The monophyletic “termite clusters” comprised of phylotypes from more than one termite species were distributed among 15 bacterial phyla, including the novel candidate phyla TG2 and TG3. These termite clusters accounted for 95% of the 960 clones analyzed in this study. Moreover, the clusters in 12 phyla comprised phylotypes from more than one termite (sub)family, accounting for 75% of the analyzed clones. Our results suggest that the majority of gut bacteria are not allochthonous but are specific symbionts that have coevolved with termites and that their community structure is basically consistent within a genus of termites.
Social insects rely heavily on pheromone communication to maintain their sociality. Egg protection is one of the most fundamental social behaviours in social insects. The recent discovery of the termite-egg mimicking fungus ‘termite-ball’ and subsequent studies on termite egg protection behaviour have shown that termites can be manipulated by using the termite egg recognition pheromone (TERP), which strongly evokes the egg-carrying and -grooming behaviours of workers. Despite the great scientific and economic importance, TERP has not been identified because of practical difficulties. Herein we identified the antibacterial protein lysozyme as the TERP. We isolated the target protein using ion-exchange and hydrophobic interaction chromatography, and the MALDI-TOF MS analysis showed a molecular size of 14.5 kDa. We found that the TERP provided antibacterial activity against a gram-positive bacterium. Among the currently known antimicrobial proteins, the molecular size of 14.5 kDa limits the target to lysozyme. Termite lysozymes obtained from eggs and salivary glands, and even hen egg lysozyme, showed a strong termite egg recognition activity. Besides eggs themselves, workers also supply lysozyme to eggs through frequent egg-grooming, by which egg surfaces are coated with saliva containing lysozyme. Reverse transcript PCR analysis showed that mRNA of termite lysozyme was expressed in both salivary glands and eggs. Western blot analysis confirmed that lysozyme production begins in immature eggs in queen ovaries. This is the first identification of proteinaceous pheromone in social insects. Researchers have focused almost exclusively on hydrocarbons when searching for recognition pheromones in social insects. The present finding of a proteinaceous pheromone represents a major step forward in, and result in the broadening of, the search for recognition pheromones. This novel function of lysozyme as a termite pheromone illuminates the profound influence of pathogenic microbes on the evolution of social behaviour in termites.
Termites have the unique ability to exploit lignocellulose as a primary nutrition source. Traditionally, termite lignocellulose digestion has been considered as a gut-symbiont-mediated process; however, in recent years the importance of host digestive capabilities have become apparent. Despite this growing understanding, how digestive enzymes from different origins specifically collaborate (i.e., additively or synergistically) has remained largely unknown. In a recent study, we undertook translational-genomic studies to address these questions in the lower termite Reticulitermes flavipes (Isoptera: Rhinotermitidae) and its symbiotic gut fauna. We used a combination of native gut tissue preparations and recombinant enzymes derived from the host gut transcriptome to identify synergistic collaborations between host and symbiont, and also among enzymes produced exclusively by the host termite. These findings provided important new evidence of synergistic collaboration among enzymes in the release of fermentable monosaccharides from wood lignocellulose, and laid a foundation for future integrative studies into termite digestion, symbiosis and eusociality.
termite; lignocellulose; cellulose; laccase; symbiosis; microbiome; end-product inhibition; functional genomics
Social insects nesting in soil environments are in constant contact with entomopathogens but have evolved a range of defence mechanisms, resulting in both individual and social immunity that reduce the chance for epizootics in the colony, as in the case of subterranean termites. Coptotermes formosanus uses its faeces as building material for its nest structure that result into a ‘carton material’, and here, we report that the faecal nest supports the growth of Actinobacteria which provide another level of protection to the social group against entomopathogens. A Streptomyces species with in vivo antimicrobial activity against fungal entomopathogens was isolated from the nest material of multiple termite colonies. Termite groups were exposed to Metarhizium anisopliae, a fungal entomopathogen, during their foraging activity and the presence of Streptomyces within the nest structure provided a significant survival benefit to the termites. Therefore, this report describes a non-nutritional exosymbiosis in a termite, in the form of a defensive mutualism which has emerged from the use of faecal material in the nesting structure of Coptotermes. The association with an Actinobacteria community in the termite faecal material provides an extended disease resistance to the termite group as another level of defence, in addition to their individual and social immunity.
termites; symbiosis; Actinobacteria; nesting behaviour; entomopathogens
Termites live in nests that can differ in microbial load and thus vary in degree of disease risk. It was hypothesized that termite investment in immune response would differ in species living in nest environments that vary in the richness and abundance of microbes. Using the drywood termite, Incisitermes schwarzi Banks (Isoptera: Kalotermitidae), as a model for species having low nest and cuticular microbial loads, the susceptibility of individuals and groups to conidia of the entomopathogenic fungus, Metarhizium anisopliae Sorokin (Hypocreales: Clavicipitaceae), was examined. The survivorship of I. schwarzi was compared to that of the dampwood termite, Zootermopsis angusticollis Hagen (Termopsidae), a species with comparatively high microbial loads. The results indicated that I. schwarzi derives similar benefits from group living as Z. angusticollis: isolated termites had 5.5 times the hazard ratio of death relative to termites nesting in groups of 25 while termites in groups of 10 did not differ significantly from the groups of 25. The results also indicated, after controlling for the influence of group size and conidia exposure on survivorship, that Z. angusticollis was significantly more susceptible to fungal infection than I. schwarzi, the former having 1.6 times the hazard ratio of death relative to drywood termites. Thus, disease susceptibility and individual investment in immunocompetence may not be dependent on interspecific variation in microbial pressures. The data validate prior studies indicating that sociality has benefits in infection control and suggest that social mechanisms of disease resistance, rather than individual physiological and immunological adaptations, may have been the principle target of selection related to variation in infection risk from microbes in the nest environment of different termite species.
ecological immunology; entomopathogenic fungus; herd immunity; infection control; microbial load
Termite colonies are founded by a pair of primary reproductives. In many species, including subterranean termites (family Rhinotermitidae), the primary king and queen can be succeeded by neotenic reproductives that are produced from workers or nymphs within the colony. It is generally believed that these neotenics inbreed within the colony, sometimes for many generations. Here, we show that primary queens of the North American subterranean termite, Reticulitermes virginicus, are replaced by numerous parthenogenetically produced female neotenics. We collected functional female neotenics from five colonies of R. virginicus in North Carolina and Texas, USA. Genetic analysis at eight microsatellite loci showed that 91–100% of the neotenics present within a colony were homozygous at all loci, indicating that they were produced through automictic parthenogenesis with terminal fusion. In contrast, workers, soldiers and alates were almost exclusively sexually produced by mating between the female neotenics and a single king. This is the second termite species shown to undergo asexual queen succession, a system first described in the Japanese species, Reticulitermes speratus. Thus, the conditional use of sexual and asexual reproduction to produce members of different castes may be widespread within Reticulitermes and possibly other subterranean termites.
parthenogenesis; inbreeding; caste differentiation; microsatellite; breeding system
Subterranean termites are major global pests of wood structures and wood products. Among the most economically important subterranean termite species in the US are Heterotermes aureus, Reticulitermes flavipes, and Coptotermes formosanus. In prior studies, the entomopathogenic nematode, Steinernema riobrave strain 355, exhibited a high level of virulence to H. aureus compared with other nematode species. However, S. riobrave 355 was reported to be poorly or only moderately virulent to R. flavipes and C. formosanus, respectively. We hypothesized that other strains of S. riobrave may possess a high level of virulence to all three termite species. Under laboratory conditions we compared three novel strains of S. riobrave (3-8b, 7-12, and TP) with the 355 strain for virulence to H. aureus, R. flavipes, and C. formosanus workers. H. aureus was very susceptible to all the S. riobrave strains, and termites in all nematode treatments were dead after 4 d. The TP strain of S. riobrave caused greater mortality in R. flavipes and C. formosanus compared to the other nematode strains. Specifically, the TP strain caused 75% and 91% mortality in R. flavipes and C. formosanus, respectively, which was more than 300% and 70% higher than the mortality caused by other strains. Additional studies are warranted to determine the ability of S. riobrave (TP) to control the targeted termite species under field conditions.
Coptotermes formosanus; entomopathogenic nematode; Heterotermes aureus; Reticulitermes flavipes; Steinernema riobrave; termite; biological control
Among the best-known examples of mutualistic symbioses is that between lower termites and the cellulolytic flagellate protists in their hindguts. Although the symbiosis in worker termites has attracted much attention, there have been only a few studies of protists in other castes. We have performed the first examination of protist population dynamics in queens and kings during termite colony foundation. Protist numbers, as well as measurements of hindgut and reproductive tissue sizes, were undertaken at five time points over 400 days in incipient colonies of Reticulitermes speratus, as well as in other castes of mature colonies of this species. We found that protist numbers increased dramatically in both queens and kings during the first 50 days of colony foundation but began to decrease by day 100, eventually disappearing by day 400. Hindgut width followed a pattern similar to that of protist numbers, while ovary and testis widths increased significantly only at day 400. Kings were found to contain higher numbers of protists than queens in incipient colonies, which may be linked to higher levels of nutrient transfer from kings to queens than vice versa, as is known in some other termite species. Protists were found to be abundant in soldiers from mature colonies but absent in neotenics. This probably reflects feeding of soldiers by workers via proctodeal trophallaxis and of reproductives via stomodeal trophallaxis. The results reveal the dynamic nature of protist numbers during colony foundation and highlight the trade-offs that exist between reproduction and parental care during this critical phase of the termite life cycle.
Phylogenetic relationships, diversity, and in situ identification of spirochetes in the gut of the termite Neotermes koshunensis were examined without cultivation, with an emphasis on ectosymbionts attached to flagellated protists. Spirochetes in the gut microbial community investigated so far are related to the genus Treponema and divided into two phylogenetic clusters. In situ hybridizations with a 16S rRNA-targeting consensus oligonucleotide probe for one cluster (known as termite Treponema cluster I) detected both the ectosymbiotic spirochetes on gut protists and the free-swimming spirochetes in the gut fluid of N. koshunensis. The probe for the other cluster (cluster II), which has been identified as ectosymbionts on gut protists of two other termite species, Reticulitermes speratus and Hodotermopsis sjoestedti, failed to detect any spirochete population. The absence of cluster II spirochetes in N. koshunensis was confirmed by intensive 16S ribosomal DNA (rDNA) clone analysis, in which remarkably diverse spirochetes of 45 phylotypes were identified, almost all belonging to cluster I. Ectosymbiotic spirochetes of the three gut protist species Devescovina sp., Stephanonympha sp., and Oxymonas sp. in N. koshunensis were identified by their 16S rDNA and by in situ hybridizations using specific probes. The probes specific for these ectosymbionts did not receive a signal from the free-swimming spirochetes. The ectosymbionts were dispersed in cluster I of the phylogeny, and they formed distinct phylogenetic lineages, suggesting multiple origins of the spirochete attachment. Each single protist cell harbored multiple spirochete species, and some of the spirochetes were common among protist species. The results indicate complex relationships of the ectosymbiotic spirochetes with the gut protists.
A number of social insect species have recently been shown to have genetically influenced caste determination (GCD), challenging the conventional view that caste determination should be strictly environmental. To date, GCD has been found in phylogenetically isolated species; examples of GCD being present in multiple species of a genus are lacking. Through crossing experiments of neotenic (juvenile) reproductives, we have recently provided the first evidence for a royal versus worker GCD in the termite Reticulitermes speratus. To elucidate whether this system is more widespread, we performed crossing experiments using three additional Reticulitermes species. Offspring caste and sex ratios were found to be highly similar to those found previously in R. speratus, raising the possibility that GCD was present in an ancestral lineage of Reticulitermes, and subsequently maintained throughout several episodes of speciation.
worker; neotenic; genetically influenced caste determination
Recent research has shown that low genetic variation in individuals can increase susceptibility to infection and group living may exacerbate pathogen transmission. In the eusocial diploid termites, cycles of outbreeding and inbreeding characterizing basal species can reduce genetic variation within nestmates during the life of a colony, but the relationship of genetic heterogeneity to disease resistance is poorly understood. Here we show that, one generation of inbreeding differentially affects the survivorship of isolated and grouped termites (Zootermopsis angusticollis) depending on the nature of immune challenge and treatment. Inbred and outbred isolated and grouped termites inoculated with a bacterial pathogen, exposed to a low dose of fungal pathogen or challenged with an implanted nylon monofilament had similar levels of immune defence. However, inbred grouped termites exposed to a relatively high concentration of fungal conidia had significantly greater mortality than outbred grouped termites. Inbred termites also had significantly higher cuticular microbial loads, presumably due to less effective grooming by nestmates. Genetic analyses showed that inbreeding significantly reduced heterozygosity and allelic diversity. Decreased heterozygosity thus appeared to increase disease susceptibility by affecting social behaviour or some other group-level process influencing infection control rather than affecting individual immune physiology.
isoptera; termite; life history; immunity; social behaviour
Clark-type oxygen microelectrodes and glass pH microelectrodes, each with a tip diameter of <=10 (mu)m, were used to obtain high-resolution profiles of oxygen concentrations and pH values in isolated termite guts. Radial oxygen profiles showed that oxygen penetrated into the peripheral hindgut contents up to about 150 to 200 (mu)m below the epithelial surface in both the lower termite Reticulitermes flavipes (Kollar) and the higher termite Nasutitermes lujae (Wasmann). Only the central portions (comprising less than 40% of the total volume) of the microbe-packed, enlarged hindgut compartments ("paunches") were completely anoxic, indicating that some members of the hindgut microbiota constitute a significant oxygen sink. From the slopes of the oxygen gradients, we estimated that the entire paunches (gut tissue plus resident microbiota) of R. flavipes and N. lujae accounted for 21 and 13%, respectively, of the respiratory activity of the intact animals. Axial oxygen profiles also confirmed that in general, only the paunches were anoxic in their centers, whereas midguts and posterior hindgut regions contained significant amounts of oxygen (up to about 50 and 30% air saturation, respectively). A remarkable exception to this was the posterior portion of an anterior segment (the P1 segment) of the hindgut of N. lujae, which was completely anoxic despite its small diameter ((apprx=)250 (mu)m). Axial pH profiles of the guts of Nasutitermes nigriceps (Haldeman) and Microcerotermes parvus (Haviland) revealed that there were extreme shifts as we moved posteriorly from the midgut proper (pH (apprx=)7) to the P1 segment of the hindgut (pH >10) and then to the P3 segment (paunch; pH (apprx=)7). The latter transition occurred at the short enteric valve (P2 segment) and within a distance of less than 500 (mu)m. In contrast, R. flavipes, which lacks a readily distinguishable P1 segment, did not possess a markedly alkaline region, and the pH around the midgut-hindgut junction was circumneutral. The oxic status of the peripheral hindgut lumen and its substantial oxygen consumption, together with previous reports of large numbers of aerobic and facultatively anaerobic bacteria in the hindgut microflora, challenge the notion that termite hindguts are a purely anoxic environment and, together with the steep axial pH gradients in higher termites, refine our concept of this tiny microbial habitat.
Termites may have high exposure to both pathogenic and competitive fungal species. Previous studies have shown anti-fungal properties of the primary components (α-pinene and limonene) of Nasutitermes frontal glands that are present on soldiers. In this study, the termite Nasutitermes acajutlae (Isoptera: Termitidae) was used to examine if the growth of naturally occurring fungi was inhibited by soldiers, as compared to workers that do not have frontal glands. Soldiers and workers were killed, surface sterilized, and placed on nutrient agar either singly or in combinations with other termites. Time until appearance of fungus and growth of visible fungal colonies was determined. Fungus appeared significantly earlier in cultures with single workers than with single soldiers (P < 0.001). Once fungus appeared, there were no significant differences in growth rate. When worker and/or soldier fluids were combined in one culture, fungal growth appeared later in cultures containing soldiers; growth was significantly slower in colonies with 5 workers and 5 soldiers than in cultures with 5 workers alone; P < 0.001. Finally, growth appeared later in cultures with 5 soldiers than in cultures with one soldier, suggesting a dose-response. Fungi that grew from termites were mostly non-pathogenic, suggesting the anti-fungal properties of soldiers may inhibit both pathogens and competitors.
Termite; parasitism; competition
The parabasalian symbionts of lower termite hindgut communities are well-known for their large size and structural complexity. The most complex forms evolved multiple times independently from smaller and simpler flagellates, but we know little of the diversity of these small flagellates or their phylogenetic relationships to more complex lineages. To understand the true diversity of Parabasalia and how their unique cellular complexity arose, more data from smaller and simpler flagellates are needed. Here, we describe two new genera of small-to-intermediate size and complexity, represented by the type species Cthulhu macrofasciculumque and Cthylla microfasciculumque from Prorhinotermes simplex and Reticulitermes virginicus, respectively (both hosts confirmed by DNA barcoding). Both genera have a single anterior nucleus embeded in a robust protruding axostyle, and an anterior bundle flagella (and likely a single posterior flagellum) that emerge slightly subanteriorly and have a distinctive beat pattern. Cthulhu is relatively large and has a distinctive bundle of over 20 flagella whereas Cthylla is smaller, has only 5 anterior flagella and closely resembles several other parababsalian genera. Molecular phylogenies based on small subunit ribosomal RNA (SSU rRNA) show both genera are related to previously unidentified environmental sequences from other termites (possibly from members of the Tricercomitidae), which all branch as sisters to the Hexamastigitae. Altogether, Cthulhu likely represents another independent origin of relatively high cellular complexity within parabasalia, and points to the need for molecular characterization of other key taxa, such as Tricercomitus.
Competition exclusion, when a single species dominates resources due to superior competitiveness, is seldom observed in nature. Termites compete for resources with deadly consequences, yet more than one species can be found feeding in the same wooden resource. This is especially surprising when drywood species, with colonies of a few hundred, are found cohabiting with subterranean species, with colonies of millions. Termites communicate vibro-acoustically and, as these signals can travel over long distances, they are vulnerable to eavesdropping. We investigated whether drywood termites could eavesdrop on vibration cues from subterranean species. We show, using choice experiments and recordings, that the drywood termite Cryptotermes secundus can distinguish its own species from the dominant competitor in the environment, the subterranean termite Coptotermes acinaciformis. The drywood termite was attracted to its own vibration cues, but was repelled by those of the subterranean species. This response increased with decreasing wood size, corresponding with both increased risk and strength of the cue. The drywood termites appear to avoid confrontation by eavesdropping on the subterranean termites; these results provide further evidence that vibro-acoustic cues are important for termite sensory perception and communication.
aggression; competition; communication; eavesdropping; Isoptera; social information
The subterranean termite, Reticulitermes speratus kyushuensis (Isoptera: Rhinotermitidae), excavate complex tunnel networks below the ground for foraging. The tunnels are either curved or meandering. In our previous study, results showed that termites passed smooth—rounded corners faster than they did around sharp corners. Smooth—rounded corners can be mathematically quantified by the curvature, representing the amount by which a geometric object deviates from a straight line. The present study explored how the time spent inside a tunnel changes in accordance with the degree of tunnel curvature. To do so, artificial tunnels with different curvatures were constructed in acryl substrates. Tunnels were 5 cm in length with widths of W — 2, 3, or 4 mm, and the distance between the two ends of the tunnel was D = 2, 3, 4, or 5 cm. A higher value of D signified a lower curvature. The time (τ) taken by a termite to pass through the tunnel was measured. In the case of W = 2 mm, the values of τ were statistically equal for D = 2, 3, or 4 cm, while τ for D = 5 cm was significantly lesser. In the case of W = 3, τ was statistically more for D = 2 and 3 cm than it was for D = 4 and 5 cm. For W = 4, τ was statistically equal for D = 2 and 3 cm, while τ for D = 4 cm was relatively shorter. Interestingly, the value of τ when D = 5 cm was statistically the same as D = 3 or 4 cm. These resulted from two types of termite behavior: biased walking and zigzag walking.
foraging efficiency; termite tunnel curvature; termite tunnel network; traffic efficiency
The phylogenetic diversity of the intestinal microflora of a lower termite, Reticulitermes speratus, was examined by a strategy which does not rely on cultivation of the resident microorganisms. Small-subunit rRNA genes (16S rDNAs) were directly amplified from the mixed-population DNA of the termite gut by the PCR and were clonally isolated. Analysis of partial 16S rDNA sequences showed the existence of well-characterized genera as well as the presence of bacterial species for which no 16S rDNA sequence data are available. Of 55 clones sequenced, 45 were phylogenetically affiliated with four of the major groups of the domain Bacteria: the Proteobacteria, the spirochete group, the Bacteroides group, and the low-G+C-content gram-positive bacteria. Within the Proteobacteria, the 16S rDNA clones showed a close relationship to those of cultivated species of enteric bacteria and sulfate-reducing bacteria, while the 16S rDNA clones in the remaining three groups showed only distant relationships to those of known organisms in these groups. Of the remaining 10 clones, among which 8 clones formed a cluster, there was only very low sequence similarity to known 16S rRNA sequences. None of these clones were affiliated with any of the major groups within the domain Bacteria. The 16S rDNA gene sequence data show that the majority of the intestinal microflora of R. speratus consists of new, uncultured species previously unknown to microbiologists.