Head-butting as an Early Indicator of Reproductive Disinhibition in the Termite Zootermopsis nevadensis

In lower termites, functionally sterile larval helpers are totipotent—capable of becoming reproductively active with the loss of their colony’s king or queen. Full reproductive development may take several weeks, but initiation of this developmental response most likely occurs shortly after colony members detect when a reproductive-specific signal is missing. We investigated the early response of termite helpers to the removal of their king and queen in the basal termite species Zootermopsis nevadensis. Within 6–12 h after reproductives were removed, helpers displayed an increase in head-butting—a behavior associated with dominance in other termite species as well as in closely related roaches. The loss of just one reproductive, either the king or queen, was also sufficient to cause an increase in head-butting. We did not find evidence, however, that this response was sex-specific: males and females were equally likely to increase head-butting independent of the sex of the reproductive that was removed. Finally, we discovered that reproductive-specific compounds present on the cuticle of king and queen termites were also present in their feces, but the presence of the feces did not seem sufficient to inhibit the increased head-butting after the reproductives were removed. Collectively, these results indicate that termite workers readily detect the loss of reproductives in their colony and that they at least initially respond in a non sex-specific manner.     

Passage and Survival of Chlamydospores of Phytophthora cinnamomi Rands, the Causal Agent of Forest Dieback Disease, Through the Gastrointestinal Tracts of Termites and Wild Birds

Chlamydospores of Phytophthora cinnamomi Rands have been shown to survive in the intestinal tracts of termites (Nasutitermes exitiosus) and two species of forest birds indigenous to West Australian jarrah forests. Viable chlamydospores were recovered from bird feces within the normal rate of passage time for food through the gut. The above factors would allow these creatures to function as vectors for the spores.     

培菌白蚁菌圃和粪便微生物多样性分析

【背景】培菌白蚁是属于白蚁科的一类与鸡枞菌属真菌共生的高等白蚁,其与体内肠道微生物和体外菌圃微生物形成三维共生体系。【目的】分析培菌白蚁菌圃和粪便的微生物多样性,并与肠道微生物进行比较。【方法】通过Illumina MiSeq高通量测序方法对培菌白蚁菌圃和粪便样品进行细菌16S rRNA基因和真菌ITS测序分析。【结果】高通量测序获得培菌白蚁菌圃和粪便样品细菌和真菌的有效序列和OTU数目。5个样品细菌OTU数目在90-199之间,而真菌OTU在10-58之间,细菌的种类多样性明显大于真菌。不论是细菌还是真菌,粪便样品的OTU数目多于菌圃样品。经物种分类分析,菌圃样品主要优势细菌是变形菌门(Proteobacteria),其相对含量超过82.4%;其次是拟杆菌门(Bacteroidetes)和厚壁菌门(Firmicutes);粪便样品中优势细菌为拟杆菌门,其次是变形菌门,粪便优势菌属为别样杆菌属和营发酵单胞菌属,这与培菌白蚁肠道菌多样性组成一致。培菌白蚁菌圃和粪便样品共生真菌主要为担子菌门(Basidiomycota)和子囊菌门(Ascomycota)。菌圃优势真菌为鸡枞菌属(Termi...     

Nitrogen fluxes from treefrogs to tank epiphytic bromeliads: an isotopic and physiological approach

Diverse invertebrate and vertebrate species live in association with plants of the large Neotropical family Bromeliaceae. Although previous studies have assumed that debris of associated organisms improves plant nutrition, so far little evidence supports this assumption. In this study we used isotopic (¹⁵N) and physiological methods to investigate if the treefrog Scinax hayii, which uses the tank epiphytic bromeliad Vriesea bituminosa as a diurnal shelter, contributes to host plant nutrition. In the field, bromeliads with frogs had higher stable N isotopic composition (δ¹⁵N) values than those without frogs. Similar results were obtained from a controlled greenhouse experiment. Linear mixing models showed that frog feces and dead termites used to simulate insects that eventually fall inside the bromeliad tank contributed, respectively, 27.7% (±0.07 SE) and 49.6% (±0.50 SE) of the total N of V. bituminosa. Net photosynthetic rate was higher in plants that received feces and termites than in controls; however, this effect was only detected in the rainy, but not in the dry season. These results demonstrate for the first time that vertebrates contribute to bromeliad nutrition, and that this benefit is seasonally restricted. Since amphibian–bromeliad associations occur in diverse habitats in South and Central America, this mechanism for deriving nutrients may be important in bromeliad systems throughout the Neotropics.     

Digesta retention time and recovery rates of ants and termites in Chinese pangolins (Manis pentadactyla)

Pangolins are myrmecophagous mammals whose biology and ecology remain poorly studied. Termite mandibles and ant head capsules are the two primary remains found in pangolin feces. Determining the retention time of insect cuticles is important for understanding the digestive physiology of pangolins, while determining the recovery rate of termites and ants in feces is required to estimate the number of these prey items that are consumed by pangolins. In this study, the authors conducted feeding trials with captive Chinese pangolins (Manis pentadactyla). Sixty grams of the fungus-growing termite Odontotermes formosanus (18,816 individuals) and 15–20 g of the yellow crazy ant Anoplolepis gracilipes (14,400–19,200 individuals) were fed to each pangolin. After feeding, pangolin feces were collected daily for 1 week. The authors also assessed the accuracy of using chromium (III) oxide powder (Cr₂O₃) as a proxy for determining gut passage time, as has been done in previous studies. The results showed that remaining termite mandibles and ant head capsules in feces peaked at 66 and 90 hr after feeding and their recovery rates were 0.35 ± 0.10 and 0.65 ± 0.04, respectively. In both feeding trials, the retention time of Cr₂O₃ was much shorter than that of the termite mandibles and ant head capsules, indicating that Cr₂O₃ is not an appropriate indicator for estimating food retention time of myrmecophagous animals. Our results revealed that the ant head capsules were preserved better in feces compared with the termite mandibles, suggesting that termites may be considerably underestimated in the feces of wild pangolins.     

Why is the number of primary tunnels of the formosan subterranean termite,Coptotermes formosanus Shiraki (Isoptera: Rhinotermidae), restricted during foraging?

Subterranean termites forage by digging a network of tunnels to come into contact with food sources. When 1000 termites (Coptotermes formosanus Shiraki) were placed in a laboratory arena, 6.7 primary tunnels were constructed. The aim of this study was to explain the empirical observation in which termites restrict the number of primary tunnels. To this end, we constructed a model to simulate termite tunnel patterns based on empirical data and to calculate food transportation efficiency, γ, for the tunnel patterns. The efficiency was defined as the ratio of the number of encountered food particles to the sum of the shortest length from the location of encountered food particles to the initial position of growth of the tunnel. The γ was maximized when the number of primary tunnels was 5 or 6, which was fairly consistent with the empirical number of primary tunnels. This result indicated that termites may restrict the number of their primary tunnels to improve the transportation efficiency, which is directly related to their survival.     

“Methanoplasmatales,” Thermoplasmatales-Related Archaea in Termite Guts and Other Environments,Are the Seventh Order of Methanogens

The Euryarchaeota comprise both methanogenic and nonmethanogenic orders and many lineages of uncultivated archaea with unknown properties. One of these deep-branching lineages, distantly related to the Thermoplasmatales, has been discovered in various environments, including marine habitats, soil, and also the intestinal tracts of termites and mammals. By comparative phylogenetic analysis, we connected this lineage of 16S rRNA genes to a large clade of unknown mcrA gene sequences, a functional marker for methanogenesis, obtained from the same habitats. The identical topologies of 16S rRNA and mcrA gene trees and the perfect congruence of all branches, including several novel groups that we obtained from the guts of termites and cockroaches, strongly suggested that they stem from the same microorganisms. This was further corroborated by two highly enriched cultures of closely related methanogens from the guts of a higher termite (Cubitermes ugandensis) and a millipede (Anadenobolus sp.), which represented one of the arthropod-specific clusters in the respective trees. Numerous other pairs of habitat-specific sequence clusters were obtained from the guts of other termites and cockroaches but were also found in previously published data sets from the intestinal tracts of mammals (e.g., rumen cluster C) and other environments. Together with the recently described Methanomassiliicoccus luminyensis isolated from human feces, which falls into rice cluster III, the results of our study strongly support the idea that the entire clade of “uncultured Thermoplasmatales” in fact represents the seventh order of methanogenic archaea, for which the provisional name “Methanoplasmatales” is proposed.     

Odor Aversion and Pathogen-Removal Efficiency in Grooming Behavior of the Termite Coptotermes formosanus

The results of biocontrol with entomopathogens in termites have been discouraging because of the strong social hygiene behavior for removing pathogens from termite colonies. However, the mechanism of pathogen detection is still unclear. For the successful application of biopesticides to termites in nature, it would be beneficial to identify substances that could disrupt the termite’s ability to perceive pathogens. We hypothesized that termites can perceive pathogens and this ability plays an important role in effective hygiene behavior. In this study, pathogen-detection in the subterranean termite Coptotermes formosanus was investigated. We performed quantitative assays on conidia removal by grooming behavior using epifluoresence microscopy and Y-maze tests to examine the perception of fungal odor by termites. Three species each of high- and low-virulence entomopathogenic fungi were used in each test. The results demonstrated that termites removed conidia more effectively from a nestmate’s cuticle if its odor elicited stronger aversion. Highly virulent pathogens showed higher attachment rates to termite surfaces and their odors were more strongly avoided than those of low-virulence isolates in the same species. Moreover, termites appeared to groom each other more persistently when they had more conidia on their bodies. In brief, insect perception of pathogen-related odor seems to play a role in the mechanism of their hygiene behavior.     

Occurrence of fungi in combs of fungus-growing termites (Isoptera: Termitidae, Macrotermitinae)

Fungus-growing termites cultivate their mutualistic basidiomycete Termitomyces species on a substrate called a fungal comb. Here, the Suicide Polymerase Endonuclease Restriction (SuPER) method was adapted for the first time to a fungal study to determine the entire fungal community of fungal combs and to test whether fungi other than the symbiotic cultivar interact with termite hosts. Our molecular analyses show that although active combs are dominated by Termitomyces fungi isolated with direct Polymerase Endonuclease Restriction - Denaturing Gradient Gel Electrophoresis (PCR-DGGE), they can also harbor some filamentous fungi and yeasts only revealed by SuPER PCR-DGGE. This is the first molecular evidence of the presence of non-Termitomyces species in active combs. However, because there is no evidence for a species-specific relationship between these fungi and termites, they are mere transient guests with no specialization in the symbiosis. It is however surprising to notice that termite-associated Xylaria strains were not isolated from active combs even though they are frequently retrieved when nests are abandoned by termites. This finding highlights the implication of fungus-growing termites in the regulation of fungi occurring within the combs and also suggests that they might not have any particular evolutionary-based association with Xylaria species.     

Deterministic Assembly of Complex Bacterial Communities in Guts of Germ-Free Cockroaches

The gut microbiota of termites plays important roles in the symbiotic digestion of lignocellulose. However, the factors shaping the microbial community structure remain poorly understood. Because termites cannot be raised under axenic conditions, we established the closely related cockroach Shelfordella lateralis as a germ-free model to study microbial community assembly and host-microbe interactions. In this study, we determined the composition of the bacterial assemblages in cockroaches inoculated with the gut microbiota of termites and mice using pyrosequencing analysis of their 16S rRNA genes. Although the composition of the xenobiotic communities was influenced by the lineages present in the foreign inocula, their structure resembled that of conventional cockroaches. Bacterial taxa abundant in conventional cockroaches but rare in the foreign inocula, such as Dysgonomonas and Parabacteroides spp., were selectively enriched in the xenobiotic communities. Donor-specific taxa, such as endomicrobia or spirochete lineages restricted to the gut microbiota of termites, however, either were unable to colonize germ-free cockroaches or formed only small populations. The exposure of xenobiotic cockroaches to conventional adults restored their normal microbiota, which indicated that autochthonous lineages outcompete foreign ones. Our results provide experimental proof that the assembly of a complex gut microbiota in insects is deterministic.     

Vertical transmission as the key to the colonization of Madagascar by fungus-growing termites?

The mutualism between fungus-growing termites (Macrotermitinae) and their mutualistic fungi (Termitomyces) began in Africa. The fungus-growing termites have secondarily colonized Madagascar and only a subset of the genera found in Africa is found on this isolated island. Successful long-distance colonization may have been severely constrained by the obligate interaction of the termites with fungal symbionts and the need to acquire these symbionts secondarily from the environment for most species (horizontal symbiont transmission). Consistent with this hypothesis, we show that all extant species of fungus-growing termites of Madagascar are the result of a single colonization event of termites belonging to one of the only two groups with vertical symbiont transmission, and we date this event at approximately 13 Mya (Middle/Upper Miocene). Vertical symbiont transmission may therefore have facilitated long-distance dispersal since both partners disperse together. In contrast to their termite hosts, the fungal symbionts have colonized Madagascar multiple times, suggesting that the presence of fungus-growing termites may have facilitated secondary colonizations of the symbiont. Our findings indicate that the absence of the right symbionts in a new environment can prevent long-distance dispersal of symbioses relying on horizontal symbiont acquisition.     

Role of bacteria in maintaining the redox potential in the hindgut of termites and preventing entry of foreign bacteria

The hindgut of the lower termites, Mastotermes darwiniensis and Coptotermes lacteus and the higher termite Nasutitermes exitiosus were made aerobic by exposure of the termites to pure oxygen, a procedure which killed their spirochaetes and their protozoa (lower termites only). The time taken for the hindgut to become anaerobic after the termites were restored to normal atmospheric conditions ranged from 2 to 4.5 hr. After oxygen treatment the number of gut bacteria increased some six- to ten-fold in all termite species, indicating that the bacteria are poised to use oxygen entering the gut. Removal of all the hindgut microbiota by feeding tetracycline caused the hindgut to become aerobic in M. darwiniensis and N. exitiosus. The transferring of M. darwiniensis to fresh wood, free of antibiotic, resulted in the return of the normal flora and the eventual establishment of anaerobic conditions in the hindgut. Thus the bacteria appear to be important in maintaining anaerobic conditions in the gut. Attempts to determine whether the protozoa (in the lower termites) played any part in maintaining the Eh of the hindgut were unsuccessful. Serratia marcescens failed to colonise the gut of normal C. lacteus and transiently colonized (for 5 days) the gut of normal N. exitiosus. Transient colonization by S. marcescens (from 6 to 10 days) occurred in N. exitiosus when its hindgut spirochaetes were killed and in C. lacteus when its spirochaetes and protozoa were killed, indicating a possible role for the spirochaetes and/or protozoa in influencing the bacteria allowed to reside in the hindgut. Exposure of normal termites to Serratia provoked an increase in the numbers of the normal gut bacteria.     

Microplate-based high throughput screening procedure for the isolation of lipid-rich marine microalgae

Background

Termites are highly effective at degrading lignocelluloses, and thus can be used as a model for studying plant cell-wall degradation in biological systems. However, the process of lignin deconstruction and/or degradation in termites is still not well understood.

Methods

We investigated the associated structural modification caused by termites in the lignin biomolecular assembly in softwood tissues crucial for cell-wall degradation. We conducted comparative studies on the termite-digested (i.e. termite feces) and native (control) softwood tissues with the aid of advanced analytical techniques: ¹³C crosspolarization magic angle spinning and nuclear magnetic resonance (CP-MAS-NMR) spectroscopy, flash pyrolysis with gas chromatography mass spectrometry (Py-GC/MS), and Py-GC-MS in the presence of tetramethylammonium hydroxide (Py-TMAH)-GC/MS.

Results

The ¹³C CP/MAS NMR spectroscopic analysis revealed an increased level of guaiacyl-derived (G unit) polymeric framework in the termite-digested softwood (feces), while providing specific evidence of cellulose degradation. The Py-GC/MS data were in agreement with the ¹³C CP/MAS NMR spectroscopic studies, thus indicating dehydroxylation and modification of selective intermonomer side-chain linkages in the lignin in the termite feces. Moreover, Py-TMAH-GC/MS analysis showed significant differences in the product distribution between control and termite feces. This strongly suggests that the structural modification in lignin could be associated with the formation of additional condensed interunit linkages.

Conclusion

Collectively, these data further establish: 1) that the major β-O-4' (β-aryl ether) was conserved, albeit with substructure degeneracy, and 2) that the nature of the resulting polymer in termite feces retained most of its original aromatic moieties (G unit-derived). Overall, these results provide insight into lignin-unlocking mechanisms for understanding plant cell-wall deconstruction, which could be useful in development of new enzymatic pretreatment processes mimicking the termite system for biochemical conversion of lignocellulosic biomass to fuels and chemicals.     

Termite pathogens: Transfer of the entomopathogen Metarhizium anisopliae between Reticulitermes sp. termites

Subterranean termites (Reticulitermes sp.) exposed to whole cultures of Metarhizium anisopliae for 4, 8, 12, or 48 hr transfer disease to previously healthy termites. Healthy termites concentrate grooming activity on diseased individuals and thereby become infected. Termites which have been killed by the fungus are avoided by healthy individuals and are less effective in spreading disease than are exposed living termites.     

Acromyrmex Leaf-Cutting Ants Have Simple Gut Microbiota with Nitrogen-Fixing Potential

Ants and termites have independently evolved obligate fungus-farming mutualisms, but their gardening procedures are fundamentally different, as the termites predigest their plant substrate whereas the ants deposit it directly on the fungus garden. Fungus-growing termites retained diverse gut microbiota, but bacterial gut communities in fungus-growing leaf-cutting ants have not been investigated, so it is unknown whether and how they are specialized on an exclusively fungal diet. Here we characterized the gut bacterial community of Panamanian Acromyrmex species, which are dominated by only four bacterial taxa: Wolbachia, Rhizobiales, and two Entomoplasmatales taxa. We show that the Entomoplasmatales can be both intracellular and extracellular across different gut tissues, Wolbachia is mainly but not exclusively intracellular, and the Rhizobiales species is strictly extracellular and confined to the gut lumen, where it forms biofilms along the hindgut cuticle supported by an adhesive matrix of polysaccharides. Tetracycline diets eliminated the Entomoplasmatales symbionts but hardly affected Wolbachia and only moderately reduced the Rhizobiales, suggesting that the latter are protected by the biofilm matrix. We show that the Rhizobiales symbiont produces bacterial NifH proteins that have been associated with the fixation of nitrogen, suggesting that these compartmentalized hindgut symbionts alleviate nutritional constraints emanating from an exclusive fungus garden diet reared on a substrate of leaves.     

Low temperature enhances the ability of the termite-egg-mimicking fungus Athelia termitophila to compete against wood-decaying fungi

A unique symbiosis exists between subterranean termites and the sclerotium-forming fungus Athelia termitophila, which forms termite-egg-mimicking sclerotia called ‘termite balls’. While the sclerotia gain a competitor-free habitat by being harboured by termite eggs, A. termitophila mycelia have to compete with wood-decay fungi in the life stage without termites. To understand its relationship with termites, the factors that affect the ability of A. termitophila to compete with other wood-decay fungi must be clarified. Here, we show that A. termitophila is competitive against other wood-decay fungi at low temperatures. In Petri dish experiments to evaluate the effects of the physicochemical conditions, that A. termitophila experiences in termite nests, on its competitive ability, A. termitophila overcame surrounding fungi in the winter, when termites are less active. Further studies quantifying the effects of A. termitophila on termites in winter will help us to understand this relationship.     

Phylogenetic and Functional Analysis of Gut Microbiota of a Fungus-Growing Higher Termite: Bacteroidetes from Higher Termites Are a Rich Source of β-Glucosidase Genes

Fungus-growing termites, their symbiotic fungi, and microbiota inhibiting their intestinal tract comprise a highly efficient cellulose-hydrolyzing system; however, little is known about the role of gut microbiota in this system. Twelve fosmid clones with β-glucosidase activity were previously obtained by functionally screening a metagenomic library of a fungus-growing termite, Macrotermes annandalei. Ten contigs containing putative β-glucosidase genes (bgl1–10) were assembled by sequencing data of these fosmid clones. All these contigs were binned to Bacteroidetes, and all these β-glucosidase genes were phylogenetically closed to those from Bacteroides or Dysgonomonas. Six out of 10 β-glucosidase genes had predicted signal peptides, indicating a transmembrane capability of these enzymes to mediate cellulose hydrolysis within the gut of the termites. To confirm the activities of these β-glucosidase genes, three genes (bgl5, bgl7, and bgl9) were successfully expressed and purified. The optimal temperature and pH of these enzymes largely resembled the environment of the host’s gut. The gut microbiota composition of the fungus-growing termite was also determined by 454 pyrosequencing, showing that Bacteroidetes was the most dominant phylum. The diversity and the enzyme properties of β-glucosidases revealed in this study suggested that Bacteroidetes as the major member in fungus-growing termites contributed to cello-oligomer degradation in cellulose-hydrolyzing process and represented a rich source for β-glucosidase genes.     

The role of multiple partners in a digestive mutualism with a protocarnivorous plant

Background and aims

The protocarnivorous plant Paepalanthus bromelioides (Eriocaulaceae) is similar to bromeliads in that this plant has a rosette-like structure that allows rainwater to accumulate in leaf axils (i.e. phytotelmata). Although the rosettes of P. bromelioides are commonly inhabited by predators (e.g. spiders), their roots are wrapped by a cylindrical termite mound that grows beneath the rosette. In this study it is predicted that these plants can derive nutrients from recycling processes carried out by termites and from predation events that take place inside the rosette. It is also predicted that bacteria living in phytotelmata can accelerate nutrient cycling derived from predators.

Methods

The predictions were tested by surveying plants and animals, and also by performing field experiments in rocky fields from Serra do Cipó, Brazil, using natural abundance and enriched isotopes of ¹⁵N. Laboratory bioassays were also conducted to test proteolytic activities of bacteria from P. bromelioides rosettes.

Key Results

Analyses of ¹⁵N in natural nitrogen abundances showed that the isotopic signature of P. bromelioides is similar to that of carnivorous plants and higher than that of non-carnivorous plants in the study area. Linear mixing models showed that predatory activities on the rosettes (i.e. spider faeces and prey carcass) resulted in overall nitrogen contributions of 26·5 % (a top-down flux). Although nitrogen flux was not detected from termites to plants via decomposition of labelled cardboard, the data on ¹⁵N in natural nitrogen abundance indicated that 67 % of nitrogen from P. bromelioides is derived from termites (a bottom-up flux). Bacteria did not affect nutrient cycling or nitrogen uptake from prey carcasses and spider faeces.

Conclusions

The results suggest that P. bromelioides derive nitrogen from associated predators and termites, despite differences in nitrogen cycling velocities, which seem to have been higher in nitrogen derived from predators (leaves) than from termites (roots). This is the first study that demonstrates partitioning effects from multiple partners in a digestion-based mutualism. Despite most of the nitrogen being absorbed through their roots (via termites), P. bromelioides has all the attributes necessary to be considered as a carnivorous plant in the context of digestive mutualism.     

Farmers’ perception of termites in agriculture production and their indigenous utilization in Northwest Benin

Background

Although termites are considered as agricultural pests, they play an important role in maintaining the ecosystem. Therefore, it matters to investigate the farmers’ perception of the impacts of the termites on the agriculture and their indigenous utilization.

Methods

A semi-structured questionnaire was used to interview 94 farmers through 10 villages of Atacora department, in the northwestern region of Benin, to obtain information for the development of successful strategies of termite management and conservation. Their perceptions on the importance and management of termites along with the indigenous nomenclature and utilization of termite mounds were assessed. Termite species identified by farmers were collected and preserved in 80% alcohol for identification.

Results

Eight crops were identified by farmers as susceptible to termites with maize, sorghum, and yam as being the most susceptible. According to farmers, the susceptibility to termites of these crops is due to their high-water content and sweet taste. A total of 27 vernacular names of termites were recorded corresponding to 10 species, Amitermes evuncifer, Macrotermes subhyalinus, and Trinervitermes oeconomus being the most damaging termite species. All the names given to termite species had a meaning. The drought was identified by farmers as the main factor favouring termite attacks. Demolition of termite mounds in the fields was the most commonly reported control method. Salt and other pesticides were commonly used by farmers to protect stored farm products. The lack of effective control methods is the main constraint for termite management. In northwestern Benin, farmers reported different purpose utilizations of termite mounds and termites.

Conclusions

The study has shown that farmers perceived termites as pests of several agricultural crops and apply various indigenous control practices whose efficiency need to be verified. Utilization of termites and termite mound soil as food and medicinal resources underlines the need for a more focused approach to termite control for the conservation of non-pest termite species. The sensitization of farmers on the importance of termites as well as the development of an integrated control method to combat termite pests proved necessary.