Squash bees host high diversity and prevalence of parasites in the northeastern United States.

The squash bee Eucera (Peponapis) pruinosa is emerging as a model species to study how stressors impact solitary wild bees in North America. Here, we describe the prevalence of trypanosomes, microsporidians and mollicute bacteria in E. pruinosa and two other species, Bombus impatiens and Apis mellifera, that together comprise over 97% of the pollinator visitors of Cucurbita agroecosystems in Pennsylvania (United States). Our results indicate that all three parasite groups are commonly detected in these bee species, but E. pruinosa often exhibit higher prevalences. We further describe novel trypanosome parasites detected in E. pruinosa, however it is unknown how these parasites impact these bees. We suggest future work investigates parasite replication and infection outcomes.

Prevalence and antibiotic resistance of rectal Mollicutes in HIV-infected men who have sex with men at the University Hospital of Dresden, Germany.

BACKGROUND: Rectal sexually transmitted infections (STI) are common in men having sex with men (MSM). Mycoplasma genitalium is increasingly being reported in this localization, but due to frequent lack of symptoms at this site, clinical significance is still unclear. Rectal prevalence of Mycoplasma hominis and Ureaplasma species is not well studied so far. We aimed to investigate the prevalence and antibiotic sensitivity of rectal Mollicutes in our HIV-cohort. METHODS: In 227 MSM presenting for annual STI-screening, 317 anorectal swabs were collected from January 2017 to December 2018. PCR was performed for detection of Chlamydia trachomatis, Neisseria gonorrhoeae, M. genitalium and also culture for M. hominis and Ureaplasma spec. RESULTS: Prevalence for M. genitalium, M. hominis, Ureaplasma spec., C. trachomatis and N. gonorrhoeae was 8.2%, 7.3%, 12.0%, 5.1% and 1.9%, respectively. Patients were asymptomatic with few exceptions. Seroprevalence of syphilis in 227 MSM was 41.9%. In 20 strains of M. genitalium, resistance-associated mutations to macrolides and quinolones were found in 60% and 30%, respectively; in five strains (25%) to both. M. hominis and Ureaplasma spec. frequently occurred combined, mostly in significant quantity consistent with infection. M. hominis and Ureaplasma spec. regularly showed sensitivity to tetracycline. CONCLUSION: At screening, rectal colonization with Mollicutes was common in our patients, but rarely caused symptoms. Due to rising antibiotic resistance of M. genitalium against quinolones, therapeutic options are increasingly limited. Treatment should be guided by antibiotic resistance testing including quinolones. In persisting anorectal symptoms, M. hominis and Ureaplasma spec. should also be taken into account.

The evolution of abdominal microbiomes in fungus-growing ants.

The attine ants are a monophyletic lineage that switched to fungus farming ca. 55-60 MYA. They have become a model for the study of complex symbioses after additional fungal and bacterial symbionts were discovered, but their abdominal endosymbiotic bacteria remain largely unknown. Here, we present a comparative microbiome analysis of endosymbiotic bacteria spanning the entire phylogenetic tree. We show that, across 17 representative sympatric species from eight genera sampled in Panama, abdominal microbiomes are dominated by Mollicutes, α- and γ-Proteobacteria, and Actinobacteria. Bacterial abundances increase from basal to crown branches in the phylogeny reflecting a shift towards putative specialized and abundant abdominal microbiota after the ants domesticated gongylidia-bearing cultivars, but before the origin of industrial-scale farming based on leaf-cutting herbivory. This transition coincided with the ancestral single colonization event of Central/North America ca. 20 MYA, documented in a recent phylogenomic study showing that almost the entire crown group of the higher attine ants, including the leaf-cutting ants, evolved there and not in South America. Several bacterial species are located in gut tissues or abdominal organs of the evolutionarily derived, but not the basal attine ants. The composition of abdominal microbiomes appears to be affected by the presence/absence of defensive antibiotic-producing actinobacterial biofilms on the worker ants' cuticle, but the significance of this association remains unclear. The patterns of diversity, abundance and sensitivity of the abdominal microbiomes that we obtained explore novel territory in the comparative analysis of attine fungus farming symbioses and raise new questions for further in-depth research.

Genome and evolution of the arbuscular mycorrhizal fungus Diversispora epigaea (formerly Glomus versiforme) and its bacterial endosymbionts.

Arbuscular mycorrhizal (AM) fungi form endosymbioses with most plants, and they themselves are hosts for Mollicutes/Mycoplasma-related endobacteria (MRE). Despite their significance, genomic information for AM fungi and their MRE are relatively sparse, which hinders our understanding of their biology and evolution. We assembled the genomes of the AM fungus Diversispora epigaea (formerly Glomus versiforme) and its MRE and performed comparative genomics and evolutionary analyses. The D. epigaea genome showed a pattern of substantial gene duplication and differential evolution of gene families, including glycosyltransferase family 25, whose activities are exclusively lipopolysaccharide biosynthesis. Genes acquired by horizontal transfer from bacteria possibly function in defense against foreign DNA or viruses. The MRE population was diverse, with multiple genomes displaying characteristics of differential evolution and encoding many MRE-specific genes as well as genes of AM fungal origin. Gene family expansion in D. epigaea may enhance adaptation to both external and internal environments, such as expansion of kinases for signal transduction upon external stimuli and expansion of nucleoside salvage pathway genes potentially for competition with MRE, whose genomes lack purine and pyrimidine biosynthetic pathways. Collectively, this metagenome provides high-quality references and begins to reveal the diversity within AM fungi and their MRE.

Reconstructing the functions of endosymbiotic Mollicutes in fungus-growing ants.

Mollicutes, a widespread class of bacteria associated with animals and plants, were recently identified as abundant abdominal endosymbionts in healthy workers of attine fungus-farming leaf-cutting ants. We obtained draft genomes of the two most common strains harbored by Panamanian fungus-growing ants. Reconstructions of their functional significance showed that they are independently acquired symbionts, most likely to decompose excess arginine consistent with the farmed fungal cultivars providing this nitrogen-rich amino-acid in variable quantities. Across the attine lineages, the relative abundances of the two Mollicutes strains are associated with the substrate types that foraging workers offer to fungus gardens. One of the symbionts is specific to the leaf-cutting ants and has special genomic machinery to catabolize citrate/glucose into acetate, which appears to deliver direct metabolic energy to the ant workers. Unlike other Mollicutes associated with insect hosts, both attine ant strains have complete phage-defense systems, underlining that they are actively maintained as mutualistic symbionts.

Highly divergent Mollicutes symbionts coexist in the scorpion Androctonus australis.

Androctonus australis is one of the most ubiquitous and common scorpion species in desert and arid lands from North Africa to India and it has an important ecological role and social impact. The bacterial community associated to this arachnid is unknown and we aimed to dissect its species composition in the gut, gonads, and venom gland. A 16S rRNA gene culture-independent diversity analysis revealed, among six other taxonomic groups (Firmicutes, Betaproteobacteria, Gammaproteobacteria, Flavobacteria, Actinobacteria, and Cyanobacteria), a dominance of Mollicutes phylotypes recorded both in the digestive tract and the gonads. These related Mollicutes include two Spiroplasma phylotypes (12.5% of DGGE bands and 15% of clones), and a new Mycoplasma cluster (80% of clones) showing 16S rRNA sequence identities of 95 and 93% with Mollicutes detected in the Mexican scorpions Centruroides limpidus and Vaejovis smithi, respectively. Such scorpion-associated Mollicutes form a new lineage that share a distant ancestor with Mycoplasma hominis. The observed host specificity with the apparent phylogenetic divergence suggests a relatively long co-evolution of these symbionts with the scorpion hosts. From the ecological point of view, such association may play a beneficial role for the host fitness, especially during dormancy or molt periods.

Horizontal Gene Transfers in Mycoplasmas (Mollicutes).

The class Mollicutes (trivial name "mycoplasma") is composed of wall-less bacteria with reduced genomes whose evolution was long thought to be only driven by gene losses. Recent evidences of massive horizontal gene transfer (HGT) within and across species provided a new frame to understand the successful adaptation of these minimal bacteria to a broad range of hosts. Mobile genetic elements are being identified in a growing number of mycoplasma species, but integrative and conjugative elements (ICEs) are emerging as pivotal in HGT. While sharing common traits with other bacterial ICEs, such as their chromosomal integration and the use of a type IV secretion system to mediate horizontal dissemination, mycoplasma ICEs (MICEs) revealed unique features: their chromosomal integration is totally random and driven by a DDE recombinase related to the Mutator-like superfamily. Mycoplasma conjugation is not restricted to ICE transmission, but also involves the transfer of large chromosomal fragments that generates progenies with mosaic genomes, nearly every position of chromosome being mobile. Mycoplasmas have thus developed efficient ways to gain access to a considerable reservoir of genetic resources distributed among a vast number of species expanding the concept of minimal cell to the broader context of flowing information.

Soil Respiration of the Dahurian Larch (Larix gmelinii) Forest and the Response to Fire Disturbance in Da Xing'an Mountains, China.

Despite the high frequency of wildfire disturbances in boreal forests in China, the effects of wildfires on soil respiration are not yet well understood. We examined the effects of fire severity on the soil respiration rate (Rs) and its component change in a Dahurian Larch (Larix gmelinii) in Northeast China. The results showed that Rs decreased with fire burning severity. Compared with the control plots, Rs in the low burning severity plots decreased by 19%, while it decreased by 28% in the high burning severity plots. The Rs decrease was mainly due to a decreased autotrophic respiration rate (Ra). The temperature sensitivity (Q 10) of Rs increased after the low severity fire disturbances, but it decreased after the high severity fire disturbance. The Rs were triggered by the soil temperature, which may explain most of the Rs variability in this area. Our study, for the first time, provides the data-based foundation to demonstrate the importance of assessing CO2 fluxes considering both fire severity and environmental factors post-fire in boreal forests of China.

Who lives in a fungus? The diversity, origins and functions of fungal endobacteria living in Mucoromycota.

Bacterial interactions with plants and animals have been examined for many years; differently, only with the new millennium the study of bacterial-fungal interactions blossomed, becoming a new field of microbiology with relevance to microbial ecology, human health and biotechnology. Bacteria and fungi interact at different levels and bacterial endosymbionts, which dwell inside fungal cells, provide the most intimate example. Bacterial endosymbionts mostly occur in fungi of the phylum Mucoromycota and include Betaproteobacteria (Burkhoderia-related) and Mollicutes (Mycoplasma-related). Based on phylogenomics and estimations of divergence time, we hypothesized two different scenarios for the origin of these interactions (early vs late bacterial invasion). Sequencing of the genomes of fungal endobacteria revealed a significant reduction in genome size, particularly in endosymbionts of Glomeromycotina, as expected by their uncultivability and host dependency. Similar to endobacteria of insects, the endobacteria of fungi show a range of behaviours from mutualism to antagonism. Emerging results suggest that some benefits given by the endobacteria to their plant-associated fungal host may propagate to the interacting plant, giving rise to a three-level inter-domain interaction.

Generous hosts: What makes Madagascar periwinkle (Catharanthus roseus) the perfect experimental host plant for fastidious bacteria?

Although much attention has been paid to the metabolism and biosynthesis of monoterpene alkaloids in Catharanthus roseus, its value as an experimental host for a variety of agriculturally and economically important phytopathogenic bacteria warrants further study. In the present study, we evaluated the chemical composition of the phloem and xylem saps of C. roseus to infer the nutritional requirements of phloem- and xylem-limited phytopathogens. Periwinkle phloem sap consisted of a rich mixture of sugars, organic acids, amino acids, amines, fatty acids, sugar acids and sugar alcohols while xylem contained similar compounds in lesser concentrations. Plant sap analysis may lead to a better understanding of the biology of fastidious Mollicutes and their complex nutritional requirements, and to successful culture of phytoplasmas and other uncultured phloem-restricted bacteria such as Candidatus Liberibacter asiaticus, the causal agent of huanglongbing in citrus.

Endogone, one of the oldest plant-associated fungi, host unique Mollicutes-related endobacteria.

Glomeromycota have been considered the most ancient group of fungi capable of positively interacting with plants for many years. Recently, other basal fungi, the Endogone Mucoromycotina fungi, have been identified as novel plant symbionts, challenging the paradigm of Glomeromycota as the unique ancestral symbionts of land plants. Glomeromycota are known to host endobacteria and recent evidences show that also some Mucoromycotina contain endobacteria. In order to examine similarities between basal groups of plant-associated fungi, we tested whether Endogone contained endobacteria. Twenty-nine Endogone were investigated in order to identify Mollicutes-related endobacteria (Mre). Fruiting bodies were processed for transmission electron microscopy and molecularly investigated using fungal and Mre-specific primers. We demonstrate that Mre are present inside 13 out of 29 Endogone: endobacteria are directly embedded in the fungal cytoplasm and their 16S rDNA sequences cluster together with the ones retrieved from Glomeromycota, forming, however, a separate new clade. Our findings provide new insights on the evolutionary relations between Glomeromycota, Mucoromycotina and endobacteria, raising new questions on the role of these still enigmatic microbes in the ecology, evolution and diversification of their fungal hosts during the history of plant-fungal symbiosis.

Detection of a novel intracellular microbiome hosted in arbuscular mycorrhizal fungi.

Arbuscular mycorrhizal fungi (AMF) are important members of the plant microbiome. They are obligate biotrophs that colonize the roots of most land plants and enhance host nutrient acquisition. Many AMF themselves harbor endobacteria in their hyphae and spores. Two types of endobacteria are known in Glomeromycota: rod-shaped Gram-negative Candidatus Glomeribacter gigasporarum, CaGg, limited in distribution to members of the Gigasporaceae family, and coccoid Mollicutes-related endobacteria, Mre, widely distributed across different lineages of AMF. The goal of the present study is to investigate the patterns of distribution and coexistence of the two endosymbionts, CaGg and Mre, in spore samples of several strains of Gigaspora margarita. Based on previous observations, we hypothesized that some AMF could host populations of both endobacteria. To test this hypothesis, we performed an extensive investigation of both endosymbionts in G. margarita spores sampled from Cameroonian soils as well as in the Japanese G. margarita MAFF520054 isolate using different approaches (molecular phylotyping, electron microscopy, fluorescence in situ hybridization and quantitative real-time PCR). We found that a single AMF host can harbour both types of endobacteria, with Mre population being more abundant, variable and prone to recombination than the CaGg one. Both endosymbionts seem to retain their genetic and lifestyle peculiarities regardless of whether they colonize the host alone or together. These findings show for the first time that fungi support an intracellular bacterial microbiome, in which distinct types of endobacteria coexist in a single cell.

Mollicutes-related endobacteria thrive inside liverwort-associated arbuscular mycorrhizal fungi.

Arbuscular mycorrhizal fungi (AMF) can host Gram-positive endobacteria (BLOs) in their cytoplasm. These have been identified as Mollicutes-related microbes based on an inventory of AMF spores from fungal collections. Bacteria-like organisms (BLOs) of unknown identity have also been reported in the cytoplasm of AMF associated with liverworts, the earliest-diverged extant lineage of land plants. A combination of morphological, molecular and phylogenetic analyses revealed that three samples of two liverwort species (Conocephalum conicum and Lunularia cruciata) growing spontaneously in a botanical garden harboured AMF belonging to Glomerales, and these, in turn, hosted coccoid BLOs. 16S rDNA sequences from these BLOs clustered with the Mollicutes sequences identified from the spore collections but revealed the presence of novel phylotypes. Electron microscopy and fluorescence in situ hybridization (FISH) confirmed the presence of BLOs inside the cytoplasm of AMF hyphae colonizing the liverwort thalli. The high genetic variability of BLOs in liverwort-AMF associations thriving in the same ecological niche raises questions about the mechanisms underlying such diversity.

[Physiological-biochemical and taxonomic studies in mollicutology].

From the beginning of researches in the field of molliculotology and for all the time of existence of special Mycoplasmology Department in the Institute of Microbiology and Virology numerous original results were obtained which are of priority for this science. The collection of strains of various representatives of the class of Mollicutes was formed. Phytopathogenic mollicute strains were first cultived in the elaborated artificial medium and their nature and specific pathogen factors were investigated. Fundamental principles of realization of pathogenic potencies by Mollicutes-agents of plant "yellows" were researched. A number of enzymes of nucleic metabolism and the proteinases, their part as the aggression factors to host organisms was distinguished for the first time, the enzymes localization was studied by cytochemical methods. The composition of carbohydrate part from glycocalix of the mollicutes and microorganisms related to them was studied, and the model of their interaction with the cells of affected organisms was elaborated. The theory and the basis for the practical use of antisignature olygodeoxyribonucleotides as the universal antimicrobial means was formulated. Properties of DNA from mollicutes genome was researched. The system position of these microorganisms and the phylogenetic relations with the representatives of affined taxones was specified. The artificial model system for studying the interaction of phytopathogenic mollicutes with plant cells was created which use helps investigate the particuliarities of the signal and metabolic relations ofmollicutes and cells of host macroorganism. In the course of done researches the changes in the fatty acids composition of the common lipids, in the activity oflectins and enzymes and in the amount of some proteins by the action of mollicute infection were stated.

Revised minimal standards for description of new species of the class Mollicutes (division Tenericutes).

Minimal standards for novel species of the class Mollicutes (trivial term, mollicutes), last published in 1995, require revision. The International Committee on Systematics of Prokaryotes Subcommittee on the Taxonomy of Mollicutes proposes herein revised standards that reflect recent advances in molecular systematics and the species concept for prokaryotes. The mandatory requirements are: (i) deposition of the type strain into two recognized culture collections, preferably located in different countries; (ii) deposition of the 16S rRNA gene sequence into a public database, and a phylogenetic analysis of the relationships among the 16S rRNA gene sequences of the novel species and its neighbours; (iii) deposition of antiserum against the type strain into a recognized collection; (iv) demonstration, by using the combination of 16S rRNA gene sequence analyses, serological analyses and supplementary phenotypic data, that the type strain differs significantly from all previously named species; and (v) assignment to an order, a family and a genus in the class, with an appropriate specific epithet. The 16S rRNA gene sequence provides the primary basis for assignment to hierarchical rank, and may also constitute evidence of species novelty, but serological and supplementary phenotypic data must be presented to substantiate this. Serological methods have been documented to be congruent with DNA-DNA hybridization data and with 16S rRNA gene placements. The novel species must be tested serologically to the greatest extent that the investigators deem feasible against all neighbouring species whose 16S rRNA gene sequences show >0.94 similarity. The investigator is responsible for justifying which characters are most meaningful for assignment to the part of the mollicute phylogenetic tree in which a novel species is located, and for providing the means by which novel species can be identified by other investigators. The publication of the description should appear in a journal having wide circulation. If the journal is not the International Journal of Systematic and Evolutionary Microbiology, copies of the publication must be submitted to that journal so that the name may be considered for inclusion in a Validation List as required by the International Code of Bacteriological Nomenclature (the Bacteriological Code). Updated informal descriptions of the class Mollicutes and some of its constituent higher taxa are available as supplementary material in IJSEM Online.

Comparative RNomics and modomics in Mollicutes: prediction of gene function and evolutionary implications.

Stable RNAs are central to protein synthesis. Ribosomal RNAs make the core of the ribosome and provide the scaffold for accurate translation of mRNAs by a set of tRNA molecules each carrying an activated amino acid. To fulfill these important cellular functions, both rRNA and tRNA molecules require more than the four canonical bases and have recruited enzymes that introduce numerous modifications on nucleosides. Mollicutes are parasitic unicellular bacteria that originated from gram-positive bacteria by considerably reducing their genome, reaching a minimal size of 480 kb in Mycoplasma genitalium. By analyzing the complete set of tRNA isoacceptors (tRNomics) and predicting the tRNA/rRNA modification enzymes (Modomics) among all sequenced Mollicutes (15 in all), our goal is to predict the minimal set of RNA modifications needed to sustain accurate translation of the cell's genetic information. Building on the known phylogenetic relationship of the 15 Mollicutes analyzed, we demonstrate that the solutions to reducing the RNA component of the translation apparatus vary from one Mollicute to the other and often rely on co-evolution of specific tRNA isoacceptors and RNA modification enzymes. This analysis also reveals that only a few modification enzymes acting on nucleotides of the anticodon loop in tRNA (the wobble position 34 as well as in position 37, 3'-adjacent to anticodon) and of the peptidyltransferase center of 23S rRNA appear to be absolutely essential and resistant to gene loss during the evolutionary process of genome reduction.

Evolution of beetle bioluminescence: the origin of beetle luciferin.

Bioluminescence, the conversion of chemical energy into light in living organisms, is dependent on two principal components, an enzyme luciferase and the substrate luciferin. In beetles, the enzyme luciferase has been extensively studied, with significant enzymological, sequence and structural data now available. Furthermore, the enzyme has been employed in a remarkable number of important applications, from microbial detection and medical imaging to GM gene expression studies. However, there is little information regarding the biosynthesis of beetle luciferin, and here we review the literature and speculate as to its evolutionary origins. Luciferin consists of a benzothiazole moiety attached to a thiazole carboxylic acid moiety, the former being rarely observed in nature but the latter being observed in a broad range of biologically derived molecules. Benzothiazoles are, however, observed in melanogenesis and we speculate as to whether this may be relevant to the understanding of luciferin biosynthesis in beetles. This review examines recent novel insights into beetle luciferin recycling and we assess a range of possible biosynthetic mechanisms.