Specific host metabolite and gut microbiome alterations are associated with bone loss during spaceflight.

Understanding the axis of the human microbiome and physiological homeostasis is an essential task in managing deep-space-travel-associated health risks. The NASA-led Rodent Research 5 mission enabled an ancillary investigation of the gut microbiome, varying exposure to microgravity (flight) relative to ground controls in the context of previously shown bone mineral density (BMD) loss that was observed in these flight groups. We demonstrate elevated abundance of Lactobacillus murinus and Dorea sp. during microgravity exposure relative to ground control through whole-genome sequencing and 16S rRNA analyses. Specific functionally assigned gene clusters of L. murinus and Dorea sp. capable of producing metabolites, lactic acid, leucine/isoleucine, and glutathione are enriched. These metabolites are elevated in the microgravity-exposed host serum as shown by liquid chromatography-tandem mass spectrometry (LC-MS/MS) metabolomic analysis. Along with BMD loss, ELISA reveals increases in osteocalcin and reductions in tartrate-resistant acid phosphatase 5b signifying additional loss of bone homeostasis in flight.

Transcriptome of Epibiont Saccharibacteria Nanosynbacter lyticus Strain TM7x During the Establishment of Symbiosis.

Saccharibacteria Nanosynbacter lyticus strain TM7x is a member of the broadly distributed candidate phylum radiation. These bacteria have ultrasmall cell sizes, have reduced genomes, and live as epibionts on the surfaces of other bacteria. The mechanisms by which they establish and maintain this relationship are not yet fully understood. The transcriptomes of the epibiont TM7x and its host bacteria Schaalia odontolytica strain XH001 were captured across the establishment of symbiosis during both the initial interaction and stable symbiosis. The results showed a dynamic interaction with large shifts in gene expression for both species between the initial encounter and stable symbiosis, notably in transporter genes. During stable symbiosis, the host XH001 showed higher gene expression for peptidoglycan biosynthesis, mannosylation, cell cycle and stress-related genes, whereas it showed lower expression of chromosomal partitioning genes. This was consistent with the elongated cell shape seen in XH001 infected with TM7x and our discovery that infection resulted in thickened cell walls. Within TM7x, increased pili, type IV effector genes, and arginine catabolism/biosynthesis gene expression during stable symbiosis implied a key role for these functions in the interaction. Consistent with its survival and persistence in the human microbiome as an obligate epibiont with reduced de novo biosynthetic capacities, TM7x also showed higher levels of energy production and peptidoglycan biosynthesis, but lower expression of stress-related genes, during stable symbiosis. These results imply that TM7x and its host bacteria keep a delicate balance in order to sustain an episymbiotic lifestyle. IMPORTANCE Nanosynbacter lyticus type strain TM7x is the first cultivated member of the Saccharibacteria and the candidate phyla radiation (CPR). It was discovered to be ultrasmall in cell size with a highly reduced genome that establishes an obligate epibiotic relationship with its host bacterium. The CPR is a large, monophyletic radiation of bacteria with reduced genomes that includes Saccharibacteria. The vast majority of the CPR have yet to be cultivated, and our insights into these unique organisms to date have been derived from only a few Saccharibacteria species. Being obligate parasites, it is unknown how these ultrasmall Saccharibacteria, which are missing many de novo biosynthetic pathways, are maintained at a high prevalence within the human microbiome as well as in the environment.

Draft Genome Sequences of Nine "Candidatus Nanosynbacter sp. HMT-352" Strains Cultured from the Human Oral Cavity.

Here, we report draft genome sequences for nine strains of "Candidatus Nanosynbacter sp. HMT-352." These strains and their sequences were used to interrogate strain-level variations in host range, gene content, and growth dynamics among the phylum "Candidatus Saccharibacteria."

Strain-Level Variation and Diverse Host Bacterial Responses in Episymbiotic Saccharibacteria.

Saccharibacteria (TM7), which are obligate episymbionts growing on the surface of host bacteria, may play an important role in oral disease, such as periodontitis (1, 2). As TM7 is a newly cultured lineage of bacteria, its research is limited by the small number of isolated representatives relative to the number of TM7 genomes assembled from culture-independent studies (3-5). A comprehensive view of both TM7 taxa and TM7 strain-level variations remains opaque. In this study, we expanded our previously developed TM7 baiting method into using many host bacteria in parallel, which allowed us to obtain 37 TM7 strains from the human oral cavity. These strains were further classified into low-enrichment (LE, n = 24) and high-enrichment (HE, n = 13) groups based on their proficiency at propagating on host bacteria. Of the 13 HE strains, 10 belong to "Candidatus Nanosynbacter sp." strain HMT-352 (human microbial taxon) (6), enabling us to explore both the phenotypic and genomic strain variations within a single TM7 species. We show that TM7 HMT-352 strains exhibit a diverse host range and varied growth dynamics during the establishment of their episymbiotic relationship with host bacteria. Furthermore, despite HMT-352 strains sharing a majority of their genes, we identified several gene clusters that may play a pivotal role in host affinity. More importantly, our comparative analyses also provide TM7 gene candidates associated with strain-level phenotypic variation that may be important for episymbiotic interactions with host bacteria. IMPORTANCE Candidate phylum radiation (CPR) bacteria comprise a poorly understood phylum that is estimated to encompass ∼26% of all diversity of domain bacteria. Among CPR bacteria, the Saccharibacteria lineage (TM7) is of particular interest, as it is found in high abundance in the mammal microbiome and has been associated with oral disease. While many CPR genomes, TM7 included, have been acquired through culture-independent methods, only a small number of representatives have been isolated. Such isolated representatives, however, shed light on the physiology, pathogenesis, and episymbiotic interactions of TM7. Combined with genomic analyses, experiments involving isolated representatives can distinguish phylogenetic to phenotypic discrepancies and better identify genes of importance. In this study, we utilized multiple host bacteria in parallel to isolate TM7 bacteria and examined strain-level variation in TM7 to reveal key genes that may drive TM7-host interactions. Our findings accentuate that broad phylogenetic characterization of CPR is the next step in understanding these bacteria.

Mocs1 (Molybdenum cofactor synthesis 1) may contribute to lifespan extension in Drosophila.

While evaluating the effect on lifespan of decreased ribosomal protein (Rp) expression in Drosophila, we discovered a potential function in the same process for the Molybdenum cofactor synthesis 1 (Mocs1) gene. We utilized the UAS-GAL4 inducible system, by crossing tissue-specific GAL4 drivers to the Harvard Drosophila Transgenic RNAi Project (TrIP) responder lines for Rp gene knockdown. We also employed a negative control that knocked down a gene unrelated to Drosophila (GAL4). Relative to the genetic background in which no driven transgenes were present, lifespan was significantly lengthened in females, both for Rp knockdown and the negative GAL4 control. We reasoned that the Mocs1 gene, located immediately downstream of the integration site on the third chromosome where all the TrIP responders are targeted might be responsible for the lifespan effects observed, due to the potential for upregulation using the UAS-GAL4 system. We repeated the lifespan experiment using an enhancer trap in the same location as the TrIP transgenes, and found that lifespan was significantly lengthened in females that possessed both the driver and responder, relative to controls, implicating Mocs1 in the biology of aging.

Swimming Kinematics of Cyprids of the Barnacle Balanus glandula.

Larvae of barnacles typically pass through naupliar and cyprid planktonic stages before settlement and metamorphosis. As the final larval stage, cyprids swim much faster than nauplii and in turbulent fluid environments with high shears as they seek habitat. Cyprids swim with six pairs of reciprocating thoracic appendages and use two anterior antennules during settlement. Our understanding of how thoracic appendages generate movement is limited due to short stroke intervals (∼5 ms) that impede observations of the shape and trajectory of appendages. Here, we used high-speed videography to observe both free-swimming and tethered cyprids of the intertidal acorn barnacle Balanus glandula to produce a comprehensive description of thoracic appendage swimming kinematics. Cyprids used a drag-based method of swimming: their six pairs of thoracic appendages moved through metachronal power strokes and synchronous recovery strokes similar to the thoracopod motions in calanoid copepods during escape swimming. During the power stroke, plumose setae on each appendage pair spread laterally into a high surface area and high drag paddle composed of a meshwork of fused setules. This interconnected setal array collapsed into a low surface area and low drag shape during the recovery stroke. These effective swimming appendages allowed cyprids to move upward at an average speed of 1.4 cm/s (∼25 body lengths/s) with an average beat frequency of 16 beats/s, and reach an instantaneous velocity of up to 6 cm/s. Beat frequency of the thoracic appendages was significantly associated with speed, with higher beat frequencies indicating faster swimming speed. At their average speed, cyprids moved at the intermediate Reynolds number of ∼10, in which both viscous and inertial forces affected movement. Cyprids could alter swimming direction by sweeping the posterior-most appendage pair to one side and beating the remaining thoracic appendages synchronously through the power stroke with greater motion on the outside of their turn. These results greatly enhance our understanding both of cyprid motility and how small planktonic organisms can use swimming appendages with fused setule arrays to reach high swimming speeds and affect directional changes.

Modified SHI medium supports growth of a disease-state subgingival polymicrobial community in vitro.

Developing a laboratory model of oral polymicrobial communities is essential for in vitro studies of the transition from healthy to diseased oral plaque. SHI medium is an enriched growth medium capable of supporting in vitro biofilms with similar diversity to healthy supragingival inocula; however, this medium does not maintain the diversity of gram-negative bacteria more associated with subgingival plaque. Here, we systematically modified SHI medium components to investigate the impacts of varying nutrients and develop a medium capable of supporting a specific disease-state subgingival community. A diseased subgingival plaque sample was inoculated in SHI medium with increasing concentrations of sucrose (0%, 0.1%, 0.5%), fetal bovine serum (FBS) (0%, 10%, 20%, 30%, 50%), and mucin (0.1, 2.5, 8.0 g/L) and grown for 48 hrs, then the 16S rRNA profiles of the resulting biofilms were examined. In total, these conditions were able to capture 89 of the 119 species and 43 of the 51 genera found in the subgingival inoculum. Interestingly, biofilms grown in high sucrose media, although dominated by acidogenic Firmicutes with a low final pH, contained several uncultured taxa from the genus Treponema, information that may aid culturing these periodontitis-associated fastidious organisms. Biofilms grown in a modified medium (here named subSHI-v1 medium) with 0.1% sucrose and 10% FBS had a high diversity closest to the inoculum and maintained greater proportions of many gram-negative species of interest from the subgingival periodontal pocket (including members of the genera Prevotella and Treponema, and the Candidate Phyla Radiation phylum Saccharibacteria), and therefore best represented the disease community.

Complete Genome Sequence of Strain BB001, a Novel Epibiont Bacterium from the Candidate Phylum Saccharibacteria (TM7).

Strain BB001 is cultivated from the human oral cavity on its basibiont bacterial host Actinomyces sp. It is an ultrasmall bacterium with a reduced genome that grows obligately on its bacterial host. BB001 is the first member of human microbiome taxon 957.

Permanently Fused Setules Create Unusual Folding Fans Used for Swimming in Cyprid Larvae of Barnacles.

Many crustacean swimming appendages carry arrays of plumose setae-exoskeletal, feather-like structures of long bristles (setae) with short branches (setules) distributed along two sides. Although closely spaced, setae are not physically interconnected. Setal arrays function during swimming as drag-based leaky paddles that push the organism through water. Barnacle cyprids, the final, non-feeding larval stage, swim with six pairs of legs (thoracopods) that open and close setal arrays in alternating high-drag power strokes and low-drag recovery strokes. While studying cyprid swimming, we found that their thoracopods contained setae permanently cross-linked by fused setules. These cuticular connections would seem highly unlikely because setae are individually produced exoskeletal secretions, and the connections imply unknown processes for the production or modification of crustacean setae. We describe the morphology and function of plumose setae on cyprids of Balanus glandula and other species across the clade Cirripedia. Setules from adjacent plumose setae are seamlessly joined at their tips and occur in three distinct linkage patterns. Thoracopods lack muscles to open and close the array; interconnected setae are instead pulled apart, producing a paddle-like fan with high drag when appendages spread laterally during power strokes. Setules are spring-like, passively closing setae into tight bundles with low drag during recovery strokes. The linked setules occur in the three main clades of the Cirripedia. This cuticular arrangement is effective in swimming, may eliminate the need for muscles to close the setal array, and may represent a unique swimming structure within the Crustacea.