Nanobdella aerobiophila gen. nov., sp. nov., a thermoacidophilic, obligate ectosymbiotic archaeon, and proposal of Nanobdellaceae fam. nov., Nanobdellales ord. nov. and Nanobdellia class. nov.

A co-culture of a novel thermoacidophilic, obligate symbiotic archaeon, designated as strain MJ1T, with its specific host archaeon Metallosphaera sedula strain MJ1HA was obtained from a terrestrial hot spring in Japan. Strain MJ1T grew in the co-culture under aerobic conditions. Coccoid cells of strain MJ1T were 200-500 nm in diameter, and attached to the MJ1HA cells in the co-culture. The ranges and optima of the growth temperature and pH of strain MJ1T in the co-culture were 60-75 °C (optimum, 65-70 °C) and pH 1.0-4.0 (optimum, pH 2.5), respectively. Core lipids of dialkyl glycerol tetraethers (GDGT)-3 and GDGT-4 were highly abundant in MJ1T cells concentrated from the co-culture. Strain MJ1T has a small genome (0.67 Mbp) lacking genes for biosynthesis of essential biomolecules, such as nucleotides, lipids and ATP. The genomic DNA G+C content was 24.9 mol%. The 16S rRNA gene sequence of strain MJ1T was most closely related to that of the cultivated species, 'Nanopusillus acidilobi' strain N7A (85.8 % similarity). Based on phylogenetic and physiological characteristics, we propose the name Nanobdella aerobiophila gen. nov., sp. nov. to accommodate the strain MJ1T (=JCM 33616T=DSM 111728T). In addition, we propose the names Nanobdellaceae fam. nov., Nanobdellales ord. nov., and Nanobdellia class. nov. to accommodate the novel genus.

Naphthoquinone Metabolites Produced by Monacrosporium ambrosium, the Ectosymbiotic Fungus of Tea Shot-Hole Borer, Euwallacea fornicatus, in Stems of Tea, Camellia sinensis.

The tea shot-hole borer beetle (TSHB, Euwallacea fornicatus) causes serious damage in plantations of tea, Camellia sinensis var. assamica, in Sri Lanka and South India. TSHB is found in symbiotic association with the ambrosia fungus, Monacrosporium ambrosium (syn. Fusarium ambrosium), in galleries located within stems of tea bushes. M. ambrosium is known to be the sole food source of TSHB. Six naphthoquinones produced during spore germination in a laboratory culture broth of M. ambrosium were isolated and identified as dihydroanhydrojavanicin, anhydrojavanicin, javanicin, 5,8-dihydroxy-2-methyl-3-(2-oxopropyl)naphthalene-1,4-dione, anhydrofusarubin and solaniol. Chloroform extracts of tea stems with red-colored galleries occupied by TSHB contained UV active compounds similar to the above naphthoquinones. Laboratory assays demonstrated that the combined ethyl acetate extracts of the fungal culture broth and mycelium inhibited the growth of endophytic fungi Pestalotiopsis camelliae and Phoma multirostrata, which were also isolated from tea stems. Thus, pigmented naphthoquinones secreted by M. ambrosium during spore germination may prevent other fungi from invading TSHB galleries in tea stems. The antifungal nature of the naphthoquinone extract suggests that it protects the habitat of TSHB. We propose that the TSHB fungal ectosymbiont M. ambrosium provides not only the food and sterol skeleton necessary for the development of the beetle during its larval stages, but also serves as a producer of fungal inhibitors that help to preserve the purity of the fungal garden of TSHB.

The structured diversity of specialized gut symbionts of the New World army ants.

Symbiotic bacteria play important roles in the biology of their arthropod hosts. Yet the microbiota of many diverse and influential groups remain understudied, resulting in a paucity of information on the fidelities and histories of these associations. Motivated by prior findings from a smaller scale, 16S rRNA-based study, we conducted a broad phylogenetic and geographic survey of microbial communities in the ecologically dominant New World army ants (Formicidae: Dorylinae). Amplicon sequencing of the 16S rRNA gene across 28 species spanning the five New World genera showed that the microbial communities of army ants consist of very few common and abundant bacterial species. The two most abundant microbes, referred to as Unclassified Firmicutes and Unclassified Entomoplasmatales, appear to be specialized army ant associates that dominate microbial communities in the gut lumen of three host genera, Eciton, Labidus and Nomamyrmex. Both are present in other army ant genera, including those from the Old World, suggesting that army ant symbioses date back to the Cretaceous. Extensive sequencing of bacterial protein-coding genes revealed multiple strains of these symbionts coexisting within colonies, but seldom within the same individual ant. Bacterial strains formed multiple host species-specific lineages on phylogenies, which often grouped strains from distant geographic locations. These patterns deviate from those seen in other social insects and raise intriguing questions about the influence of army ant colony swarm-founding and within-colony genetic diversity on strain coexistence, and the effects of hosting a diverse suite of symbiont strains on colony ecology.

Morphological adaptation for ectosymbiont maintenance and transmission during metamorphosis in Lagria beetles.

The diversity and success of holometabolous insects is partly driven by metamorphosis, which allows for the exploitation of different niches and decouples growth and tissue differentiation from reproduction. Despite its benefits, metamorphosis comes with the cost of temporal vulnerability during pupation and challenges associated with tissue reorganizations. These rearrangements can also affect the presence, abundance, and localization of beneficial microbes in the host. However, how symbionts are maintained or translocated during metamorphosis and which adaptations are necessary from each partner during this process remains unknown for the vast majority of symbiotic systems. Here, we show that Lagria beetles circumvent the constraints of metamorphosis by maintaining defensive symbionts on the surface in specialized cuticular structures. The symbionts are present in both sexes throughout larval development and during the pupal phase, in line with a protective role during the beetle's immature stages. By comparing symbiont titer and morphology of the cuticular structures between sexes using qPCR, fluorescence in situ hybridization, and micro-computed tomography, we found that the organs likely play an important role as a symbiont reservoir for transmission to female adults, since symbiont titers and structures are reduced in male pupae. Using symbiont-sized fluorescent beads, we demonstrate transfer from the region of the dorsal symbiont-housing organs to the opening of the reproductive tract of adult females, suggesting that symbiont relocation on the outer surface is possible, even without specialized symbiont adaptations or motility. Our results illustrate a strategy for holometabolous insects to cope with the challenge of symbiont maintenance during metamorphosis via an external route, circumventing problems associated with internal tissue reorganization. Thereby, Lagria beetles keep a tight relationship with their beneficial partners during growth and metamorphosis.

Taxonomy and molecular phylogeny of two poorly known ciliate genera, Balantidion and Acropisthium (Protista: Ciliophora: Litostomatea), including a new species of Balantidion.

The class Litostomatea Small & Lynn, 1981 is a morphologically diverse ciliate group including hundreds of free-living and endocommensal species. The genera Acropisthium Perty, 1852 and Balantidion Eberhard, 1862 previously consisted of one free-living freshwater species each. Here, we not only highlight additional morphological features of the two type species, but also investigate a new species, Balantidion foissneri sp. nov., isolated from a river flowing through Lake Weishan, China, based on complementary methods, i.e., living morphology, stained preparations, and 18S rRNA gene sequence data. Balantidion foissneri sp. nov. can be distinguished from the type species, B. pellucidum Eberhard, 1862, by the body size (115-170 × 50-80 µm vs. 70-100 × 25-45 µm in B. pellucidum), oral bulge (distinct vs. indistinct), extrusome shape (filiform vs. rod-shaped), and the number of somatic kineties (46-60 vs. 25-40). In Balantidion species, pre-encystment trophonts show similarly-shaped polymorphic cytoplasmic lepidosomes destined to adorn the outer surface of the resting cyst. Based on the current knowledge, assignment of Balantidion to the family Acropisthiidae Foissner & Foissner, 1988 is proposed. In addition, phylogenetic analyses based on molecular data show that the two Balantidion species form a fully-supported clade to which Acropisthium mutabile has a sister relationship.

Effects on Brood Development in the Carpenter Ant Camponotus vicinus Mayr after Exposure to the Yeast Associate Schwanniomyces polymorphus Kloecker.

The yeast Schwanniomyces polymorphus is associated with the infrabuccal pocket in the carpenter ant Camponotus vicinus (Hymenoptera: Formicidae), but its role in ant development is poorly defined. The potential effects of this yeast on brood development were examined on sets of larval groups and workers over a 12 week period. Worker-larval sets were fed variations of a completely artificial, holidic diet and exposed or not exposed to live S. polymorphus. Worker-larval sets in half of the experiment were defaunated using a two-step heat and chemical process. Brood development and number of adult ants produced were significantly affected by the heat/chemical defaunation process. Compared to worker-larval groups fed a basal, complete diet, all treatments resulted in no or deleterious larval development. Brood weights and number of worker ants produced from the original larval sets at initiation were significantly higher in non-defaunated ant groups fed a diet lacking both B vitamins and cholesterol and exposed to live S. polymorphus. We propose that this yeast may help ants to more efficiently assimilate nutrients when fed nutrient-deficient diets, particularly those deficient in sterols.

Sulfur-Oxidizing Symbionts without Canonical Genes for Autotrophic CO2 Fixation.

Since the discovery of symbioses between sulfur-oxidizing (thiotrophic) bacteria and invertebrates at hydrothermal vents over 40 years ago, it has been assumed that autotrophic fixation of CO2 by the symbionts drives these nutritional associations. In this study, we investigated "Candidatus Kentron," the clade of symbionts hosted by Kentrophoros, a diverse genus of ciliates which are found in marine coastal sediments around the world. Despite being the main food source for their hosts, Kentron bacteria lack the key canonical genes for any of the known pathways for autotrophic carbon fixation and have a carbon stable isotope fingerprint that is unlike other thiotrophic symbionts from similar habitats. Our genomic and transcriptomic analyses instead found metabolic features consistent with growth on organic carbon, especially organic and amino acids, for which they have abundant uptake transporters. All known thiotrophic symbionts have converged on using reduced sulfur to gain energy lithotrophically, but they are diverse in their carbon sources. Some clades are obligate autotrophs, while many are mixotrophs that can supplement autotrophic carbon fixation with heterotrophic capabilities similar to those in Kentron. Here we show that Kentron bacteria are the only thiotrophic symbionts that appear to be entirely heterotrophic, unlike all other thiotrophic symbionts studied to date, which possess either the Calvin-Benson-Bassham or the reverse tricarboxylic acid cycle for autotrophy.IMPORTANCE Many animals and protists depend on symbiotic sulfur-oxidizing bacteria as their main food source. These bacteria use energy from oxidizing inorganic sulfur compounds to make biomass autotrophically from CO2, serving as primary producers for their hosts. Here we describe a clade of nonautotrophic sulfur-oxidizing symbionts, "Candidatus Kentron," associated with marine ciliates. They lack genes for known autotrophic pathways and have a carbon stable isotope fingerprint heavier than other symbionts from similar habitats. Instead, they have the potential to oxidize sulfur to fuel the uptake of organic compounds for heterotrophic growth, a metabolic mode called chemolithoheterotrophy that is not found in other symbioses. Although several symbionts have heterotrophic features to supplement primary production, in Kentron they appear to supplant it entirely.

Two new Geosmithia species in G. pallida species complex from bark beetles in eastern USA.

Species of Geosmithia are cosmopolitan but understudied fungi, and most are associated with phloem-feeding bark beetles on various woody hosts. We surveyed 207 bark and ambrosia beetles from 37 species in the eastern USA for associated fungi. The community is dominated by species in the G. pallida species complex (GPSC) and included several Geosmithia isolates that appear to be new to science. The new Geosmithia isolates exhibited the characteristic brownish-colored colonies typical for the G. pallida species complex and were phylogenetically resolved as two genealogically exclusive lineages based on a concatenated multilocus data set based on the internal transcribed spacers (ITS) of the nuc rDNA (ITS1-5.8S-ITS2 = ITS), and the translation elongation factor 1-α (TEF1-α), ß-tubulin (TUB2), and RNA polymerase II second largest subunit (RPB2) genes. Two new Geosmithia species, G. brunnea and G. proliferans, are proposed, and their morphological traits and phylogenetic placements are presented.

Inside Out: Archaeal Ectosymbionts Suggest a Second Model of Reduced-Genome Evolution.

Reduced-genome symbionts and their organelle counterparts, which have even smaller genomes, are essential to the lives of many organisms. But how and why have these genomes become so small? Endosymbiotic genome reduction is a product of isolation within the host, followed by massive pseudogenization and gene loss often including DNA repair mechanisms. This phenomenon can be observed in insect endosymbionts such as the bacteria Carsonella ruddii and Buchnera aphidicola. Yet endosymbionts are not the only organisms with reduced genomes. Thermophilic microorganisms experience selective pressures that cause their genomes to become more compact and efficient. Nanoarchaea are thermophilic archaeal ectosymbionts that live on the surface of archaeal hosts. Their genomes, a full order of magnitude smaller than the Escherichia coli genome, are very small and efficient. How have the genomes of nanoarchaea and late-stage insect endosymbionts, which live in drastically different environments, come to mirror each other in both genome size and efficiency? Because of their growth at extreme temperatures and their exterior association with their host, nanoarchaea appear to have experienced genome reduction differently than mesophilic insect endosymbionts. We suggest that habitat-specific mechanisms of genome reduction result in fundamentally different pathways for these two groups of organisms. With this assertion, we propose two pathways of symbiosis-driven genome reduction; isolation-symbiosis experienced by insect endosymbionts and thermal-symbiosis experienced by nanoarchaea.

New species of Temnocephala (Platyhelminthes, Temnocephalida) ectosymbiont on vulnerable species of aeglids (Crustacea, Anomura) from the Neotropical Region / Nova espécie de Temnocephala (Platyhelminthes, Temnocephalida) ectosimbionte sobre espécies vulneráveis de eglídeos (Crustacea, Anomura) da Região Neotropical

Abstract: A new species of the genus Temnocephala Blanchard, 1849 from southern Brazil was found on two species of anomuran crustaceans, Aegla spinipalma Bond-Buckup & Buckup, 1994 and Aegla grisella Bond-Buckup & Buckup, 1994, the latter classified as a vulnerable species by the "Lista de Referência da Fauna Ameaçada de Extinção no Rio Grande do Sul. Decreto no 41.672, de 11 junho de 2002". The crustaceans were collected from a tributary creek of the Forqueta river, Perau de Janeiro, Arvorezinha and a tributary creek of the Fão river, Pouso Novo, Rio Grande do Sul, Brazil; both localities belong to the Sub-Basin of Forqueta River. The new species differs from seven other temnocephalans epibionts on Aegla Leach, 1820, by having the following characters: 1. a long and slightly curved cirrus, 2. two vaginal sphincters, one proximal, big and asymmetric, and one distal, smaller and symmetric, and; 3. longer than wide, elongated epidermal 'excretory' syncytial plates (EPs), with a almost horizontally central excretory pore, displaced to the anterior portion of the plate. The new species' EP is the largest in total length among epibionts temnocephalans in crustaceans already registered. Regarding the similarities with the male reproductive system of Temnocephala axenosMonticelli, 1898, the new species has important differences in the female reproductive system. It has a larger proximal vaginal sphincter, located in the middle of the vagina, while the smaller distal one is at the extreme end of the organ. Besides that, the vaginal portion between the proximal and distal sphincters is conspicuous, with a strong muscular wall. This is the first record of a species of Temnocephala in the Taquari Valley, as well in the 'Perau de Janeiro', which is an area with a rich endemic fauna.

Resumo: Uma nova espécie do gênero Temnocephala Blanchard, 1849 da região sul do Brasil foi encontrada sobre duas espécies de crustáceos anomuros, Aegla spinipalma Bond-Buckup & Buckup, 1994 e Aegla grisella Bond-Buckup & Buckup, 1994, a última classificada como uma espécie vulnerável pela Lista de Referência da Fauna Ameaçada de Extinção no Rio Grande do Sul. Decreto no 41.672, de 11 junho de 2002. Os crustáceos foram coletados em um arroio tributário do Rio Forqueta, Perau de Janeiro, Arvorezinha e em um arroio tributário do Rio Fão, Pouso Novo, Rio Grande do Sul, Brazil; ambas localidades pertencem a Sub-Bacia do Rio Forqueta. A nova espécie se diferencia dos outros sete temnocefalídeos epibiontes sobre Aegla Leach, 1820 pelos caracteres a seguir: 1. cirro longo e levemente curvo, 2. dois esfíncteres vaginais, um proximal, grande e assimétrico e um distal, menor e simétrico, e, 3. placas sinciciais epidérmicas 'excretoras' (PEs) alongadas, mais longas do que largas, com poro excretor quase central horizontalmente e deslocado para a porção anterior da placa. A PE da nova espécie é a maior em comprimento total entre os temnocefalídeos epibiontes sobre crustáceos registrados até o momento. Embora haja similaridades com o sistema reprodutor masculino de Temnocephala axenosMonticelli, 1898, a nova espécie apresenta diferenças importantes no sistema reprodutor feminino. O esfíncter vaginal proximal é maior, localizado no meio da vagina, enquanto o distal é menor e se localiza no final do órgão. Além disso, a porção da vagina entre os esfíncters proximal e distal é conspícua, com uma forte parede muscular. Esta é a primeira espécie de Temnocephala registrada para o Vale do Taquari, assim como para o Perau de Janeiro, área com uma fauna endêmica rica.