Symbiotic Protists Research Articles

Transmission dynamics of symbiotic protist communities in the termite gut: association with host adult eclosion and dispersal.

The fidelity of vertical transmission is a critical factor in maintaining mutualistic associations with microorganisms. The obligate mutualism between termites and intestinal protist communities has been maintained for over 130 million years, suggesting the faithful transmission of diverse protist species across host generations. Although a severe bottleneck can occur when alates disperse with gut protists, how protist communities are maintained during this process remains largely unknown. In this study, we examined the dynamics of intestinal protist communities during adult eclosion and alate dispersal in the termite Reticulitermes speratus. We found that the protist community structure in last-instar nymphs differed significantly from that in workers and persisted intact during adult eclosion, whereas all protists disappeared from the gut during moults between worker stages. The number of protists in nymphs and alates was substantially lower than in workers, whereas the proportion of protist species exhibiting low abundance in workers was higher in nymphs and alates. Using a simulation-based approach, we demonstrate that such changes in the protist community composition of nymphs and alates improve the transmission efficiency of whole protist species communities. This study thus provides novel insights into how termites have maintained mutualistic relationships with diverse gut microbiota for generations.

Description and prevalence of gregarines infecting the amphipod Gammarus pulex, in the Water of Leith, Scotland, UK

Gregarines are symbiotic protists that are found in a broad spectrum of invertebrates, including insects, crustaceans, and annelids. Among these the globally distributed amphipod Gammarus pulex is one of the earliest recognized hosts for aquatic gregarines and is prevalent among macroinvertebrates in freshwater environments. In this study, samples of G. pulex were collected in the Water of Leith river, Scotland, UK. Gregarines were identified using light and scanning electron microscopy as well as standard molecular techniques. We identified three septate eugregarine symbionts—Heliospora longissima, Cephaloidophora gammari, and the here newly characterized Cephaloidophora conus n. sp. (formerly Cephaloidophora sp.) associated with Gammarus pulex in the Water of Leith. Prevalences for identified gregarine species were calculated and seasonal dynamics of gregarine infections/colonization were analyzed. Prevalences were highest in autumn and spring reaching almost 50 %. While the two Cephaloidophora species showed similar colonization patterns, the prevalence of Heliospora showed an opposite trend. Identifying gregarine infection/colonization patterns is one step towards better understanding the gregarine–host relationship, as well as possible impacts of the gregarines on their hosts.

Protist symbionts of termites: diversity, distribution, and coevolution.

The symbiosis between termites and their hindgut protists is mutually obligate and vertically inherited. It was established by the late Jurassic in the cockroach ancestors of termites as they transitioned to wood feeding. Since then, protist symbionts have been transmitted from host generation to host generation by proctodeal trophallaxis (anal feeding). The protists belong to multiple lineages within the eukaryotic superphylum Metamonada. Most of these lineages have evolved large cells with complex morphology, unlike the non-termite-associated Metamonada. The species richness and taxonomic composition of symbiotic protist communities varies widely across termite lineages, especially within the deep-branching clade Teletisoptera. In general, closely related termites tend to harbour closely related protists, and deep-branching termites tend to harbour deep-branching protists, reflecting their broad-scale co-diversification. A closer view, however, reveals a complex distribution of protist lineages across hosts. Some protist taxa are common, some are rare, some are widespread, and some are restricted to a single host family or genus. Some protist taxa can be found in only a few, distantly related, host species. Thus, the long history of co-diversification in this symbiosis has been complicated by lineage-specific loss of symbionts, transfer of symbionts from one host lineage to another, and by independent diversification of the symbionts relative to their hosts. This review aims to introduce the biology of this important symbiosis and serve as a gateway to the diversity and systematics literature for both termites and protists. A searchable database with all termite-protist occurrence records and taxonomic references is provided as a supplementary file to encourage and facilitate new research in this field.

Commensal protists in reptiles display flexible host range and adaptation to ectothermic hosts.

Environmental factors like climate change and captive breeding can impact the gut microbiota and host health. Therefore, conservation efforts for threatened species may benefit from understanding how these factors influence animal microbiomes. Parabasalid protists are members of the mammalian microbiota that can modulate the immune system and impact susceptibility to infections. However, little is known about parabasalids in reptiles. Here, we profile reptile-associated parabasalids in wild and captive reptiles and find that captivity has minimal impact on parabasalid prevalence or diversity. However, because reptiles are cold-blooded (ectothermic), their microbiotas experience wider temperature fluctuation than microbes in warm-blooded animals. To investigate whether extreme weather patterns affect parabasalid-host interactions, we analyzed the gene expression in reptile-associated parabasalids and found that temperature differences significantly alter genes associated with host health. These results expand our understanding of parabasalids in this vulnerable vertebrate group and highlight important factors to be taken into consideration for conservation efforts.

Identification of a New Endo-β-1,4-xylanase Prospected from the Microbiota of the Termite Heterotermes tenuis.

Xylanases are hemicellulases that break down xylan to soluble pentoses. They are used for industrial purposes, such as paper whitening, beverage clarification, and biofuel production. The second-generation bioethanol production is hindered by the enzymatic hydrolysis step of the lignocellulosic biomass, due to the complex arrangement established among its constituents. Xylanases can potentially increase the production yield by improving the action of the cellulolytic enzyme complex. We prospected endo-β-1,4-xylanases from meta-transcriptomes of the termite Heterotermes tenuis. In silico structural characterization and functional analysis of an endo-β-1,4-xylanase from a symbiotic protist of H. tenuis indicate two active sites and a substrate-binding groove needed for the catalytic activity. No N-glycosylation sites were found. This endo-β-1,4-xylanase was recombinantly expressed in Pichia pastoris and Escherichia coli cells, presenting a molecular mass of approximately 20 kDa. Enzymatic activity assay using recombinant endo-β-1,4-xylanase was also performed on 1% xylan agar stained with Congo red at 30 °C and 40 °C. The enzyme expressed in both systems was able to hydrolyze the substrate xylan, becoming a promising candidate for further analysis aiming to determine its potential for application in industrial xylan degradation processes.

Phylogenetic identification of symbiotic protists of five Chinese Reticulitermes species indicates a cospeciation of gut microfauna with host termites.

Symbiotic protists play important roles in the wood digestion of lower termites. Previous studies showed that termites generally possess host-specific flagellate communities. The genus Reticulitermes is particularly interesting because its unique assemblage of gut flagellates bears evidence for transfaunation. The gut fauna of Reticulitermes species in Japan, Europe, and North America had been investigated, but data on species in China are scarce. For the first time, we analyzed the phylogeny of protists in the hindgut of five Reticulitermes species in China. A total of 22 protist phylotypes were affiliated with the family Trichonymphidae, Teranymphidae, Trichomonadidae, and Holomastigotoididae (Phylum Parabasalia), and 45 protist phylotypes were affiliated with the family Pyrsonymphidae (Phylum Preaxostyla). The protist fauna of these five Reticulitermes species is similar to those of Reticulitermes species in other geographical regions. The topology of Trichonymphidae subtree was similar to that of Reticulitermes tree. All Preaxostyla clones were affiliated with the genera Pyrsonympha and Dinenympha (Order Oxymonadida) as in the other Reticulitermes species. The results of this study not only add to the existing information on the flagellates present in other Reticulitermes species but also offer the opportunity to test the hypotheses for the coevolution of symbiotic protists with their host termites.

Influence of Zwitterionic Buffer Effects with Thermal Modification Treatments of Wood on Symbiotic Protists in Reticulitermes grassei Clément.

Simple SummaryOver the past thirty years, the thermal modification of wood has become a universally recognised and commercialised wood modification process. Thermal modifications may affect wood properties, either positively (dimensional stability and decay resistance) or negatively (mechanical properties). The combination of the impregnation of specific reagents with thermal modification may help to overcome the negative effects on wood properties. In this study, we evaluate the effect of a combination of two zwitterionic buffers, bicine and tricine, and thermal modification of two wood species (beech and spruce) against subterranean termites and their symbiotic fauna. Bicine and tricine treatments alone had a clear influence on wood mass loss and termite survival. The flagellate protist symbiotic community was affected by the treatments and responded differently to them, as a highly adaptable community. However, the combination of bicine with the thermal modification showed a negative effect on termites and their symbionts on both wood species. The combination of these different factors should be further investigated, as these results seem to be promising with regard to the enhancement of the termite resistance of wood.The majority of thermal modification processes are at temperatures greater than 180 °C, resulting in a product with some properties enhanced and some diminished (e.g., mechanical properties). However, the durability of thermally modified wood to termite attack is recognised as low. Recent attempts at combining thermal modification with chemical modification, either prior to or directly after the thermal process, are promising. Buffers, although not influencing the reaction systems, may interact on exposure to certain conditions, potentially acting as promoters of biological changes. In this study, two zwitterionic buffers, bicine and tricine, chosen for their potential to form Maillard-type products with fragmented hemicelluloses/volatiles, were assessed with and without thermal modification for two wood species (spruce and beech), with subsequent evaluation of their effect against subterranean termites (Reticulitermes grassei Clément) and their symbiotic protists. The effect of the wood treatments on termites and their symbionts was visible after four weeks, especially for spruce treated with tricine and bicine and heat treatment (bicine HT), and for beech treated with bicine and bicine and heat treatment (bicine HT). The chemical behaviour of these substances should be further investigated when in contact with wood and also after heat treatment. This is the first study evaluating the effect of potential Maillard reactions with zwitterionic buffers on subterranean termite symbiotic fauna.

Spirotrichonymphea (Parabasalia) symbionts of the termite Paraneotermes simplicicornis

The desert dampwood termite Paraneotermes simplicicornis harbors several species of obligately symbiotic protists that support its nutrition by fermenting lignocellulose. Among them are three morphotypes with the dexiotropic spiraling flagellar bands characteristic of Spirotrichonymphea (Parabasalia). The largest morphotype, characterized by an elongated cell apex with axial columella and internally positioned spiraling flagellar bands, was previously described as Spirotrichonympha polygyra. A smaller morphotype, with similarly internalized flagellar bands but a more rounded posterior without a protruding axostyle, was previously reported but not named. The smallest morphotype has surface flagellar bands and can attach to other protist cells by its apex. In this study, we combine light microscopy of live specimens and 18S rRNA gene sequencing of individually isolated cells to better understand the diversity of symbionts in P. simplicicornis. We found that S. polygyra branches distantly from true Spirotrichonympha, which are associated with Reticulitermes termites. Thus, we propose the new genus Cuppa to accommodate C. polygyra n. comb. (type species) and the similar but smaller morphotype Cuppa taenia n. sp. The undescribed smallest morphotype can be excluded from all previously described Spirotrichonymphea genera by molecular and behavioral evidence, so we propose Fraterculus simplicicornis n. gen., n. sp., to accommodate this organism.

Distribution and relative abundance of three protist genera within the Zootermopsis nevadensis nuttingi hindgut

The termite Zootermopsis nevadensis nuttingi, which inhabits coastal forests of the western United States, plays an important ecological role in the breakdown and digestion of wood. Vital to this role are symbiotic protists residing in the termite’s hindgut. Five protist genera of varying size and morphology make up this gut community, but very little is known about their spatial organization within the hindgut or the number of protist cells per host. To resolve this issue, we used light microscopy and a hemocytometer to determine the distribution of the larger protist genera across hindgut segments. We found that Streblomastix were the most abundant cells in all three segments, but especially in the anterior hindgut. Trichomitopsis cells were significantly more abundant in the posterior hindgut. Trichonympha were the least abundant overall, often reaching their highest abundance in the middle segment but with a more variable distribution. Understanding the distribution of different protists within the hindgut may improve our understanding of the ecological relationships among protists, as well as their individual roles in lignocellulose digestion, contributing to a better understanding of the symbiotic system as a whole.

Transcriptome analysis of the digestive system of a wood-feeding termite (Coptotermes formosanus) revealed a unique mechanism for effective biomass degradation

BackgroundWood-feeding termite, Coptotermes formosanus Shiraki, represents a highly efficient system for biomass deconstruction and utilization. However, the detailed mechanisms of lignin modification and carbohydrate degradation in this system are still largely elusive.ResultsIn order to reveal the inherent mechanisms for efficient biomass degradation, four different organs (salivary glands, foregut, midgut, and hindgut) within a complete digestive system of a lower termite, C. formosanus, were dissected and collected. Comparative transcriptomics was carried out to analyze these organs using high-throughput RNA sequencing. A total of 71,117 unigenes were successfully assembled, and the comparative transcriptome analyses revealed significant differential distributions of GH (glycosyl hydrolase) genes and auxiliary redox enzyme genes in different digestive organs. Among the GH genes in the salivary glands, the most abundant were GH9, GH22, and GH1 genes. The corresponding enzymes may have secreted into the foregut and midgut to initiate the hydrolysis of biomass and to achieve a lignin-carbohydrate co-deconstruction system. As the most diverse GH families, GH7 and GH5 were primarily identified from the symbiotic protists in the hindgut. These enzymes could play a synergistic role with the endogenous enzymes from the host termite for biomass degradation. Moreover, twelve out of fourteen genes coding auxiliary redox enzymes from the host termite origin were induced by the feeding of lignin-rich diets. This indicated that these genes may be involved in lignin component deconstruction with its redox network during biomass pretreatment.ConclusionThese findings demonstrate that the termite digestive system synergized the hydrolysis and redox reactions in a programmatic process, through different parts of its gut system, to achieve a maximized utilization of carbohydrates. The detailed unique mechanisms identified from the termite digestive system may provide new insights for advanced design of future biorefinery.

Incomplete Co-cladogenesis Between Zootermopsis Termites and Their Associated Protists.

Coevolution is a major driver of speciation in many host-associated symbionts. In the termite-protist digestive symbiosis, the protists are vertically inherited by anal feeding among nest mates. Lower termites (all termite families except Termitidae) and their symbionts have broadly co-diversified over ~170 million yr. However, this inference is based mainly on the restricted distribution of certain protist genera to certain termite families. With the exception of one study, which demonstrated congruent phylogenies for the protist Pseudotrichonympha and its Rhinotermitidae hosts, coevolution in this symbiosis has not been investigated with molecular methods. Here we have characterized the hindgut symbiotic protists (Phylum Parabasalia) across the genus Zootermopsis (Archotermopsidae) using single cell isolation, molecular phylogenetics, and high-throughput amplicon sequencing. We report that the deepest divergence in the Zootermopsis phylogeny (Zootermopsis laticeps [Banks; Isoptera: Termopsidae]) corresponds with a divergence in three of the hindgut protist species. However, the crown Zootermopsis taxa (Zootermopsis angusticollis [Hagen; Isoptera: Termopsidae], Z. nevadensis nevadensis [Hagen; Isoptera: Termopsidae], and Z. nevadensis nuttingi [Haverty & Thorne; Isoptera: Termopsidae]) share the same protist species, with no evidence of co-speciation under our methods. We interpret this pattern as incomplete co-cladogenesis, though the possibility of symbiont exchange cannot be entirely ruled out. This is the first molecular evidence that identical communities of termite-associated protist species can inhabit multiple distinct host species.

Trp residue at subsite - 5 plays a critical role in the substrate binding of two protistan GH26 β-mannanases from a termite hindgut.

Symbiotic protists in the hindgut of termites provide a novel enzymatic resource for efficient lignocellulytic degradation of plant biomass. In this study, two β-mannanases, RsMan26A and RsMan26B, from a symbiotic protist community of the lower termite, Reticulitermes speratus, were successfully expressed in the methylotrophic yeast, Pichia pastoris. Biochemical characterization experiments demonstrated that both RsMan26A and RsMan26B are endo-acting enzymes and have a very similar substrate specificity, displaying a higher catalytic efficiency to galactomannan from locust bean gum (LBG) and glucomannan than to β-1,4-mannan and highly substituted galactomannan from guar gum. Homology modeling of RsMan26A and RsMan26B revealed that each enzyme displays a long open cleft harboring a unique hydrophobic platform (Trp79) that stacks against the sugar ring at subsite - 5. The Km values of W79A mutants of RsMan26A and RsMan26B to LBG increased by 4.8-fold and 3.6-fold, respectively, compared with those for the native enzymes, while the kcat remained unchanged or about 40% of that of the native enzyme, resulting in the decrease in the catalytic efficiency by 4.8-fold and 9.1-fold, respectively. The kinetic values for glucomannan also showed a similar result. These results demonstrate that the Trp residue present near the subsite - 5 has an important role in the recognition of the sugar ring in the substrate.

Effects of lignins as diet components on the physiological activities of a lower termite, Coptotermes formosanus Shiraki

We investigated the effects of lignins as diet components on the physiological activities of a lower termite, Coptotermes formosanus Shiraki. Artificial diets composed of polysaccharides with and without purified lignins (milled-wood lignins) from Japanese cedar (softwood), Japanese beech (hardwood), and rice (grass), were fed to C. formosanus workers. The survival and body mass of the workers as well as the presence of three symbiotic protists in the hindguts of the workers were then periodically examined. The survival rates of workers fed on diets containing lignins were, regardless of the lignocellulose diet sources, significantly higher than those of workers fed on only polysaccharides. In addition, it was clearly observed that all the tested lignins have positive effects on the maintenance of two major protists in the hindguts of C. formosanus workers, i.e., Pseudotrichonympha grassii and Holomastigotoides hartmanni. Overall, our data suggest that the presence of lignin is crucial to maintaining the physiological activities of C. formosanus workers during their lignocellulose decomposition. Our data also suggested that some components, possibly minerals and/or non-structural carbohydrates, in grass lignocellulose negatively affect the survival of C. formosanus workers as well as the present rate of the symbiotic protists in their hindguts.

The effects of various lignocelluloses and lignins on physiological responses of a lower termite, Coptotermes formosanus

We investigated the physiological responses of a lower termite, Coptotermes formosanus, fed on various lignocelluloses and purified lignins (milled-wood lignins, MWLs) from Japanese cedar (softwood), Japanese beech (hardwood), and rice (grass). Termite survival, body mass, and the changes of the symbiotic protists in the hindgut of workers were observed for 4 weeks. The survival, body mass and presence of both Pseudotrichonympha grassii and Holomastigotoides hartmanni in the hindgut of workers fed on rice lignocellulose at the 4th week of observation were significantly lower than those of the workers fed on Japanese cedar and Japanese beech lignocellulose samples, whereas there was no significant difference in Spirotrichonympha leidyi among all the diets. The three purified MWLs, regardless of their structural differences, did not show any significant differences for the termites’ survival or body mass or the survival of all the three protists. The three MWL diets resulted in significantly lower termite survival compared to starvation, although these diets showed no significant effects on body mass or the protist profiles. Overall, lignins are hardly utilized as a nutrient source by C. formosanus workers and are even rather detrimental to termites when fed on solely.

Dimethylated sulfur compounds in symbiotic protists: A potentially significant source for marine DMS(P)

AbstractSymbiosis with micro‐algae (photosymbiosis) is a common feature among marine planktonic protists, but very little is known about the physiology and ecological significance of these associations. High concentrations of dimethylsulfoniopropionate (DMSP), a metabolite produced by marine microalgae, are commonly observed in coral‐microalgae symbiosis, where DMS(P) is involved in multiple physiological functions. Knowledge on concentration and role of DMSP in analogous photosymbiosis in plankton is lacking. Here, we assess the total DMSP (DMSPt = DMSP + DMS) concentration and natural stable isotopes of sulfur across ecologically relevant symbiotic plankton groups, the Radiolaria and Foraminifera. We found that intracellular DMSPt concentrations in microalgal symbionts were among the highest recorded (range = 170–702 mmol L−1), while lower concentrations (range = 0.1–23 mmol L−1) were characteristic of the holobiont (i.e., host‐microalgae). The contribution of symbiotic Radiolaria to the water column particulate DMSPt concentration ranged 0.1–8%. Sulfur isotopic composition (34S) of DMSPt in the Collodaria holobionts was significantly higher than their symbiotic microalgae isolated in culture. Despite their high intracellular DMSPt content, uptake in these holobionts throughout 3‐d incubations was not detected. We observed a systematic 34S depletion (∼ 1.5‰) of DMS relative to DMSP in experimental incubations containing filtered seawater, which we hypothesize is related to the bacterial preference for the uptake of 34S‐depleted DMS. Overall, the results indicate that plankton symbiosis can, at times, represent a potentially important source of DMS(P). Specific differences in 34S provided new insights into sulfur isotopic fractionation associated with DMS(P) biotransformation processes, with potential implications for current interpretations of isotopically tracked biogenic sources of marine aerosols.

Expanding the Knowledge on Lignocellulolytic and Redox Enzymes of Worker and Soldier Castes from the Lower Termite Coptotermes gestroi.

Termites are considered one of the most efficient decomposers of lignocelluloses on Earth due to their ability to produce, along with its microbial symbionts, a repertoire of carbohydrate-active enzymes (CAZymes). Recently, a set of Pro-oxidant, Antioxidant, and Detoxification enzymes (PAD) were also correlated with the metabolism of carbohydrates and lignin in termites. The lower termite Coptotermes gestroi is considered the main urban pest in Brazil, causing damage to wood constructions. Recently, analysis of the enzymatic repertoire of C. gestroi unveiled the presence of different CAZymes. Because the gene profile of CAZy/PAD enzymes endogenously synthesized by C. gestroi and also by their symbiotic protists remains unclear, the aim of this study was to explore the eukaryotic repertoire of these enzymes in worker and soldier castes of C. gestroi. Our findings showed that worker and soldier castes present similar repertoires of CAZy/PAD enzymes, and also confirmed that endo-glucanases (GH9) and beta-glucosidases (GH1) were the most important glycoside hydrolase families related to lignocellulose degradation in both castes. Classical cellulases such as exo-glucanases (GH7) and endo-glucanases (GH5 and GH45), as well as classical xylanases (GH10 and GH11), were found in both castes only taxonomically related to protists, highlighting the importance of symbiosis in C. gestroi. Moreover, our analysis revealed the presence of Auxiliary Activity enzyme families (AAs), which could be related to lignin modifications in termite digestomes. In conclusion, this report expanded the knowledge on genes and proteins related to CAZy/PAD enzymes from worker and soldier castes of lower termites, revealing new potential enzyme candidates for second-generation biofuel processes.

Variation and diversity of symbiotic protist composition in the damp-wood termite Hodotermopsis sjoestedti

SUMMARY It is well recognized that composition of termite symbiotic protist community generally show hostspecies specificity at species-to-species level. On the other hand, studies focused on population or colony level variations of the protist composition are few, although such studies are necessary for understanding the termite-protist mutualism. In this study we investigated symbiotic protist fauna of 34 Hodotermopsis sjoestedti nests collected from five islands in the Ryukyu Archipelago, Taiwan Is., and southern part of Chinese continent. Thirteen parabasalid species and six oxymonad species, which were reported from the host termite collected in Yakushima Is., were recognized in this study. Four parabasalids were identified as recently described species from H. sjoestedti collected in Vietnam. The fauna was the most speciesrich among the termite hosts studied to date, possibly reflecting large host body size. The prevalence rates of the symbiotic protist species are generally high, 100% in all populations for all protist species, except Gen. A sp. and Hoplonympha sp. All host nests had identical protist species composition except for the presence or absence of Gen. A sp. Two host populations lacked this species. In Yakushima Is., both types of termite nests were found, which exhibited mutually exclusive distribution. The distribution pattern suggests that the host termite is a multiple-site nester and nest-budding occurs in the field.

Metatranscriptome of the protistan community in Reticulitermes flaviceps.

The hindgut of lower termites harbors various symbiotic protists, which perform varied functions in lignocellulose decomposition. As termites are social insects, the species and numbers of these flagellated protists in the termite gut vary among the different castes. Juvenile hormones (JHs) can regulate caste differentiation in termites. In this study, we used the juvenile hormone analog fenoxycarb to induce termite workers (Reticulitermes flaviceps) to differentiate into pre-soldiers. A metatranscriptomic investigation of the protistan community was then performed by 454 pyrosequencing. From a thorough analysis based on 597312 generated reads, we found that the starch and sucrose metabolism pathway was the most abundant pathway across the metatranscriptome. The current study demonstrates that the metatranscriptome of the protistan community in termites contains an abundance of lignocellulase, which plays a vital role in termite nutrition.

Temporal variations in symbiotic hindgut protist community of the subterranean termite Reticulitermes lucifugus Rossi in Sicily

The dynamics of symbiotic protist communities of Reticulitermes lucifugus “Sicily” were investigated to assess the effects of seasonal environmental conditions and the termite’s feeding activity. In worker and soldier castes, the total protist population (total abundance), individual species abundances, and species proportions in their hindguts were examined monthly from December 2012 to February 2014. The main protist species taken into account were Dinenympha fimbriata Kirby, D. gracilis Leidy, Pyrsonympha flagellata Gras., Spirotrichonympha flagellata Grassi u. Foa, Holomastigotes elongatum Grassi u. Foa, and Trichonympha agilis Leidy, while others were not specifically identified, but were counted together. Seasonal comparisons between worker and soldier castes were made. Further, monthly data of workers were examined. We always found the presence of symbiontic protists in worker and soldier hindguts, therefore we hypothesize that termites fed without periods of interruption. In both castes the total protist population and species abundance showed variable trends with the highest values in summer. Workers were found to have significantly larger populations of protists and abundances of individual species than soldiers. Similar protist species were observed in both castes and in all periods, except for H. elongatum which was sometimes absent in soldiers. In hindguts of workers and soldiers the proportions of the protist species often showed a significant variation across seasons except for S. flagellata and T. agilis. In both castes we always detected the highest percentage of Dinenympha spp. with respect to the other species. Elements that might influence these aspects are discussed.

The GH26 β-mannanase RsMan26H from a symbiotic protist of the termite Reticulitermes speratus is an endo-processive mannobiohydrolase: Heterologous expression and characterization

Symbiotic protists in the gut of termites are prominent natural resources for enzymes involved in lignocellulose degradation. Here we report expression, purification, and biochemical characterization of a glycoside hydrolase family 26 mannanase RsMan26H from the symbiotic protist of the lower termite, Reticulitermes speratus. Biochemical analysis of RsMan26H demonstrates that this enzyme is an endo-processive mannobiohydrolase producing mannobiose from oligo- and polysaccharides, followed by a minor accumulation of oligosaccharides larger than mannobiose. To our knowledge, this is the first report describing the unique mannobiohydrolase enzyme from the eukaryotic origin.