Gut microbe Lactiplantibacillus plantarum undergoes different evolutionary trajectories between insects and mammals.

BACKGROUND: Animals form complex symbiotic associations with their gut microbes, whose evolution is determined by an intricate network of host and environmental factors. In many insects, such as Drosophila melanogaster, the microbiome is flexible, environmentally determined, and less diverse than in mammals. In contrast, mammals maintain complex multispecies consortia that are able to colonize and persist in the gastrointestinal tract. Understanding the evolutionary and ecological dynamics of gut microbes in different hosts is challenging. This requires disentangling the ecological factors of selection, determining the timescales over which evolution occurs, and elucidating the architecture of such evolutionary patterns. RESULTS: We employ experimental evolution to track the pace of the evolution of a common gut commensal, Lactiplantibacillus plantarum, within invertebrate (Drosophila melanogaster) and vertebrate (Mus musculus) hosts and their respective diets. We show that in Drosophila, the nutritional environment dictates microbial evolution, while the host benefits L. plantarum growth only over short ecological timescales. By contrast, in a mammalian animal model, L. plantarum evolution results to be divergent between the host intestine and its diet, both phenotypically (i.e., host-evolved populations show higher adaptation to the host intestinal environment) and genomically. Here, both the emergence of hypermutators and the high persistence of mutated genes within the host's environment strongly differed from the low variation observed in the host's nutritional environment alone. CONCLUSIONS: Our results demonstrate that L. plantarum evolution diverges between insects and mammals. While the symbiosis between Drosophila and L. plantarum is mainly determined by the host diet, in mammals, the host and its intrinsic factors play a critical role in selection and influence both the phenotypic and genomic evolution of its gut microbes, as well as the outcome of their symbiosis.

Cytoplasmic polyadenylation by TENT5A is required for proper bone formation.

Osteoblasts orchestrate bone formation through the secretion of type I collagen and other constituents of the matrix on which hydroxyapatite crystals mineralize. Here, we show that TENT5A, whose mutations were found in congenital bone disease osteogenesis imperfecta patients, is a cytoplasmic poly(A) polymerase playing a crucial role in regulating bone mineralization. Direct RNA sequencing revealed that TENT5A is induced during osteoblast differentiation and polyadenylates mRNAs encoding Col1α1, Col1α2, and other secreted proteins involved in osteogenesis, increasing their expression. We postulate that TENT5A, possibly together with its paralog TENT5C, is responsible for the wave of cytoplasmic polyadenylation of mRNAs encoding secreted proteins occurring during bone mineralization. Importantly, the Tent5a knockout (KO) mouse line displays bone fragility and skeletal hypomineralization phenotype resulting from quantitative and qualitative collagen defects. Thus, we report a biologically relevant posttranscriptional regulator of collagen production and, more generally, bone formation.

In Vitro Interactions of TiO2 Nanoparticles with Earthworm Coelomocytes: Immunotoxicity Assessment.

Titanium dioxide nanoparticles (TiO2 NPs) are manufactured worldwide. Once they arrive in the soil environment, they can endanger living organisms. Hence, monitoring and assessing the effects of these nanoparticles is required. We focus on the Eisenia andrei earthworm immune cells exposed to sublethal concentrations of TiO2 NPs (1, 10, and 100 µg/mL) for 2, 6, and 24 h. TiO2 NPs at all concentrations did not affect cell viability. Further, TiO2 NPs did not cause changes in reactive oxygen species (ROS) production, malondialdehyde (MDA) production, and phagocytic activity. Similarly, they did not elicit DNA damage. Overall, we did not detect any toxic effects of TiO2 NPs at the cellular level. At the gene expression level, slight changes were detected. Metallothionein, fetidin/lysenin, lumbricin and MEK kinase I were upregulated in coelomocytes after exposure to 10 µg/mL TiO2 NPs for 6 h. Antioxidant enzyme expression was similar in exposed and control cells. TiO2 NPs were detected on coelomocyte membranes. However, our results do not show any strong effects of these nanoparticles on coelomocytes at both the cellular and molecular levels.

Gold nanoparticles coated with polyvinylpyrrolidone and sea urchin extracellular molecules induce transient immune activation.

We report that the immunogenicity of colloidal gold nanoparticles coated with polyvinylpyrrolidone (PVP-AuNPs) in a model organism, the sea urchin Paracentrotus lividus, can function as a proxy for humans for in vitro immunological studies. To profile the immune recognition and interaction from exposure to PVP-AuNPs (1 and 10 µg mL-1), we applied an extensive nano-scale approach, including particle physicochemical characterisation involving immunology, cellular biology, and metabolomics. The interaction between PVP-AuNPs and soluble proteins of the sea urchin physiological coelomic fluid (blood equivalent) results in the formation of a protein "corona" surrounding the NPs from three major proteins that influence the hydrodynamic size and colloidal stability of the particle. At the lower concentration of PVP-AuNPs, the P. lividus phagocytes show a broad metabolic plasticity based on the biosynthesis of metabolites mediating inflammation and phagocytosis. At the higher concentration of PVP-AuNPs, phagocytes activate an immunological response involving Toll-like receptor 4 (TLR4) signalling pathway at 24 hours of exposure. These results emphasise that exposure to PVP-AuNPs drives inflammatory signalling by the phagocytes and the resolution at both the low and high concentrations of the PVP-AuNPs and provides more details regarding the immunogenicity of these NPs.

Bringing SEM and MSI Closer Than Ever Before: Visualizing Aspergillus and Pseudomonas Infection in the Rat Lungs.

A procedure for processing frozen rat lung tissue sections for scanning electron microscopy (SEM) from deeply frozen samples initially collected and stored for matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) was developed. The procedure employed slow thawing of the frozen sections while floating on the surface and melting in a fixative solution. After the float-washing step, the sections were dehydrated in a graded ethanol series and dried in a critical point dryer. The SEM generated images with well-preserved structures, allowing for monitoring of bacterial cells and fungal hyphae in the infected tissue. Importantly, the consecutive nonfixed frozen sections were fully compatible with MALDI-MSI, providing molecular biomarker maps of Pseudomonas aeruginosa. The protocol enables bimodal image fusion in the in-house software CycloBranch, as demonstrated by SEM and MALDI-MSI.

Author Correction: Bacterial nanotubes as a manifestation of cell death.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Bacterial nanotubes as a manifestation of cell death.

Bacterial nanotubes are membranous structures that have been reported to function as conduits between cells to exchange DNA, proteins, and nutrients. Here, we investigate the morphology and formation of bacterial nanotubes using Bacillus subtilis. We show that nanotube formation is associated with stress conditions, and is highly sensitive to the cells' genetic background, growth phase, and sample preparation methods. Remarkably, nanotubes appear to be extruded exclusively from dying cells, likely as a result of biophysical forces. Their emergence is extremely fast, occurring within seconds by cannibalizing the cell membrane. Subsequent experiments reveal that cell-to-cell transfer of non-conjugative plasmids depends strictly on the competence system of the cell, and not on nanotube formation. Our study thus supports the notion that bacterial nanotubes are a post mortem phenomenon involved in cell disintegration, and are unlikely to be involved in cytoplasmic content exchange between live cells.

The torpedo effect in Bacillus subtilis: RNase J1 resolves stalled transcription complexes.

RNase J1 is the major 5'-to-3' bacterial exoribonuclease. We demonstrate that in its absence, RNA polymerases (RNAPs) are redistributed on DNA, with increased RNAP occupancy on some genes without a parallel increase in transcriptional output. This suggests that some of these RNAPs represent stalled, non-transcribing complexes. We show that RNase J1 is able to resolve these stalled RNAP complexes by a "torpedo" mechanism, whereby RNase J1 degrades the nascent RNA and causes the transcription complex to disassemble upon collision with RNAP. A heterologous enzyme, yeast Xrn1 (5'-to-3' exonuclease), is less efficient than RNase J1 in resolving stalled Bacillus subtilis RNAP, suggesting that the effect is RNase-specific. Our results thus reveal a novel general principle, whereby an RNase can participate in genome-wide surveillance of stalled RNAP complexes, preventing potentially deleterious transcription-replication collisions.

γ-Tubulin interacts with E2F transcription factors to regulate proliferation and endocycling in Arabidopsis.

γ-Tubulin is associated with microtubule nucleation, but evidence is accumulating in eukaryotes that it also functions in nuclear processes and in cell division control independently of its canonical role. We found that in Arabidopsis thaliana, γ-tubulin interacts specifically with E2FA, E2FB, and E2FC transcription factors both in vitro and in vivo. The interaction of γ-tubulin with the E2Fs is not reduced in the presence of their dimerization partners (DPs) and, in agreement, we found that γ-tubulin interaction with E2Fs does not require the dimerization domain. γ-Tubulin associates with the promoters of E2F-regulated cell cycle genes in an E2F-dependent manner, probably in complex with the E2F-DP heterodimer. The up-regulation of E2F target genes PCNA, ORC2, CDKB1;1, and CCS52A under γ-tubulin silencing suggests a repressive function for γ-tubulin at G1/S and G2/M transitions, and the endocycle, which is consistent with an excess of cell division in some cells and enhanced endoreduplication in others in the shoot and young leaves of γ-tubulin RNAi plants. Altogether, our data show ternary interaction of γ-tubulin with the E2F-DP heterodimer and suggest a repressive role for γ-tubulin with E2Fs in controlling mitotic activity and endoreduplication during plant development.

Titanium dioxide nanoparticles temporarily influence the sea urchin immunological state suppressing inflammatory-relate gene transcription and boosting antioxidant metabolic activity.

Titanium dioxide nanoparticles (TiO2NPs) are revolutionizing biomedicine due to their potential application as diagnostic and therapeutic agents. However, the TiO2NP immune-compatibility remains an open issue, even for ethical reasons. In this work, we investigated the immunomodulatory effects of TiO2NPs in an emergent proxy to human non-mammalian model for in vitro basic and translational immunology: the sea urchin Paracentrotus lividus. To highlight on the new insights into the evolutionarily conserved intracellular signaling and metabolism pathways involved in immune-TiO2NP recognition/interaction we applied a wide-ranging approach, including electron microscopy, biochemistry, transcriptomics and metabolomics. Findings highlight that TiO2NPs interact with immune cells suppressing the expression of genes encoding for proteins involved in immune response and apoptosis (e.g. NF-κB, FGFR2, JUN, MAPK14, FAS, VEGFR, Casp8), and boosting the immune cell antioxidant metabolic activity (e.g. pentose phosphate, cysteine-methionine, glycine-serine metabolism pathways). TiO2NP uptake was circumscribed to phagosomes/phagolysosomes, depicting harmless vesicular internalization. Our findings underlined that under TiO2NP-exposure sea urchin innate immune system is able to control inflammatory signaling, excite antioxidant metabolic activity and acquire immunological tolerance, providing a new level of understanding of the TiO2NP immune-compatibility that could be useful for the development in Nano medicines.

Francisella tularensis subsp. holarctica Releases Differentially Loaded Outer Membrane Vesicles Under Various Stress Conditions.

Francisella tularensis is a Gram-negative, facultative intracellular bacterium, causing a severe disease called tularemia. It secretes unusually shaped nanotubular outer membrane vesicles (OMV) loaded with a number of virulence factors and immunoreactive proteins. In the present study, the vesicles were purified from a clinical isolate of subsp. holarctica strain FSC200. We here provide a comprehensive proteomic characterization of OMV using a novel approach in which a comparison of OMV and membrane fraction is performed in order to find proteins selectively enriched in OMV vs. membrane. Only these proteins were further considered to be really involved in the OMV function and/or their exceptional structure. OMV were also isolated from bacteria cultured under various cultivation conditions simulating the diverse environments of F. tularensis life cycle. These included conditions mimicking the milieu inside the mammalian host during inflammation: oxidative stress, low pH, and high temperature (42°C); and in contrast, low temperature (25°C). We observed several-fold increase in vesiculation rate and significant protein cargo changes for high temperature and low pH. Further proteomic characterization of stress-derived OMV gave us an insight how the bacterium responds to the hostile environment of a mammalian host through the release of differentially loaded OMV. Among the proteins preferentially and selectively packed into OMV during stressful cultivations, the previously described virulence factors connected to the unique intracellular trafficking of Francisella were detected. Considerable changes were also observed in a number of proteins involved in the biosynthesis and metabolism of the bacterial envelope components like O-antigen, lipid A, phospholipids, and fatty acids. Data are available via ProteomeXchange with identifier PXD013074.

Sea Urchin Extracellular Proteins Design a Complex Protein Corona on Titanium Dioxide Nanoparticle Surface Influencing Immune Cell Behavior.

Extensive exploitation of titanium dioxide nanoparticles (TiO2NPs) augments rapid release into the marine environment. When in contact with the body fluids of marine invertebrates, TiO2NPs undergo a transformation and adhere various organic molecules that shape a complex protein corona prior to contacting cells and tissues. To elucidate the potential extracellular signals that may be involved in the particle recognition by immune cells of the sea urchin Paracentrotus lividus, we investigated the behavior of TiO2NPs in contact with extracellular proteins in vitro. Our findings indicate that TiO2NPs are able to interact with sea urchin proteins in both cell-free and cell-conditioned media. The two-dimensional proteome analysis of the protein corona bound to TiO2NP revealed that negatively charged proteins bound preferentially to the particles. The main constituents shaping the sea urchin cell-conditioned TiO2NP protein corona were proteins involved in cellular adhesion (Pl-toposome, Pl-galectin-8, Pl-nectin) and cytoskeletal organization (actin and tubulin). Immune cells (phagocytes) aggregated TiO2NPs on the outer cell surface and within well-organized vesicles without eliciting harmful effects on the biological activities of the cells. Cells showed an active metabolism, no oxidative stress or caspase activation. These results provide a new level of understanding of the extracellular proteins involved in the immune-TiO2NP recognition and interaction in vitro, confirming that primary immune cell cultures from P. lividus can be an optional model for swift and efficient immune-toxicological investigations.

Bordetella Adenylate Cyclase Toxin Inhibits Monocyte-to-Macrophage Transition and Dedifferentiates Human Alveolar Macrophages into Monocyte-like Cells.

Monocytes arriving at the site of infection differentiate into functional effector macrophages to replenish the resident sentinel cells. Bordetella pertussis, the pertussis agent, secretes an adenylate cyclase toxin-hemolysin (CyaA) that binds myeloid phagocytes through complement receptor 3 (CD11b/CD18) and swiftly delivers its adenylyl cyclase enzyme domain into phagocytes. This ablates the bactericidal capacities of phagocytes through massive and unregulated conversion of cytosolic ATP into the key signaling molecule cAMP. We show that exposure of primary human monocytes to as low a concentration as 22.5 pM CyaA, or a low (2:1) multiplicity of infection by CyaA-producing B. pertussis bacteria, blocks macrophage colony-stimulating factor (M-CSF)-driven differentiation of monocytes. CyaA-induced cAMP signaling mediated through the activity of protein kinase A (PKA) efficiently blocked expression of macrophage markers, and the monocytes exposed to 22.5 pM CyaA failed to acquire the characteristic intracellular complexity of mature macrophage cells. Neither M-CSF-induced endoplasmic reticulum (ER) expansion nor accumulation of Golgi bodies, mitochondria, or lysosomes was observed in toxin-exposed monocytes, which remained small and poorly phagocytic and lacked pseudopodia. Exposure to 22.5 pM CyaA toxin provoked loss of macrophage marker expression on in vitro differentiated macrophages, as well as on primary human alveolar macrophages, which appeared to dedifferentiate into monocyte-like cells with upregulated CD14 levels. This is the first report that terminally differentiated tissue-resident macrophage cells can be dedifferentiated in vitro The results suggest that blocking of monocyte-to-macrophage transition and/or dedifferentiation of the sentinel cells of innate immunity through cAMP-elevating toxin action may represent a novel immune evasion strategy of bacterial pathogens.IMPORTANCE Macrophages are key sentinel cells of the immune system, and, as such, they are targeted by the toxins produced by the pertussis agent Bordetella pertussis The adenylate cyclase toxin (CyaA) mediates immune evasion of B. pertussis by suspending the bactericidal activities of myeloid phagocytes. We reveal a novel mechanism of potential subversion of host immunity, where CyaA at very low (22 pM) concentrations could inhibit maturation of human monocyte precursors into the more phagocytic macrophage cells. Furthermore, exposure to low CyaA amounts has been shown to trigger dedifferentiation of mature primary human alveolar macrophages back into monocyte-like cells. This unprecedented capacity is likely to promote survival of the pathogen in the airways, both by preventing maturation of monocytes attracted to the site of infection into phagocytic macrophages and by dedifferentiation of the already airway-resident sentinel cells.

Morphology of the vasa vasorum in coronary arteries of the porcine heart: A new insight.

INTRODUCTION: The vasa vasorum interna were described during the last decade as a special kind of vessels originating directly from the lumen of the paternal artery and participating in the nourishment of its wall, especially of the aorta and coronary arteries. At the same time, their existence was repeatedly denied/negated by many other authors. AIM: The purpose of the actual study was the anatomical verification of the existence of the vasa vasorum interna in porcine coronary arteries. MATERIALS AND METHODS: The vascular supply was studied on the wall of the anterior interventricular branch of the left coronary artery on 36 hearts taken from healthy pigs. Light microscopy, vascular injections and scanning electron microscopy were used for the analysis of 141 samples. RESULTS: In only two cases small arteries resembling vasa vasorum interna and originating directly from the lumen of the coronary artery were found. But, in both cases these vessels ran without branching, passed over the whole thickness of adventitia and branched in the wider periarterial space. In contrast to this all feeding arteries of the vasa vasorum arose from the larger branches of the paternal artery, branched entirely in its adventitia and did not enter the media. CONCLUSION: Due to the very low incidence of these small arteries originating from the lumen of the paternal artery and the absence of their participation on the nourishment of the arterial wall we came to the conclusion that it is not suitable to use the term "vasa vasorum interna" for their designation.

Differential Effects of the Flavonolignans Silybin, Silychristin and 2,3-Dehydrosilybin on Mesocestoides vogae Larvae (Cestoda) under Hypoxic and Aerobic In Vitro Conditions.

Mesocestoides vogae larvae represent a suitable model for evaluating the larvicidal potential of various compounds. In this study we investigated the in vitro effects of three natural flavonolignans-silybin (SB), 2,3-dehydrosilybin (DHSB) and silychristin (SCH)-on M. vogae larvae at concentrations of 5 and 50 µM under aerobic and hypoxic conditions for 72 h. With both kinds of treatment, the viability and motility of larvae remained unchanged, metabolic activity, neutral red uptake and concentrations of neutral lipids were reduced, in contrast with a significantly elevated glucose content. Incubation conditions modified the effects of individual FLs depending on their concentration. Under both sets of conditions, SB and SCH suppressed metabolic activity, the concentration of glucose, lipids and partially motility more at 50 µM, but neutral red uptake was elevated. DHSB exerted larvicidal activity and affected motility and neutral lipid concentrations differently depending on the cultivation conditions, whereas it decreased glucose concentration. DHSB at the 50 µM concentration caused irreversible morphological alterations along with damage to the microvillus surface of larvae, which was accompanied by unregulated neutral red uptake. In conclusion, SB and SCH suppressed mitochondrial functions and energy stores, inducing a physiological misbalance, whereas DHSB exhibited a direct larvicidal effect due to damage to the tegument and complete disruption of larval physiology and metabolism.

Diamond nanoparticles suppress lateral growth of bacterial colonies.

Diamond nanoparticles (DNPs) of various types have been recently reported to possess antibacterial properties. Studies have shown a decrease of the colony forming ability on agar plates of the bacteria that had been previously co-incubated with DNPs in the suspension. Before plating, bacteria with DNPs were adequately diluted in order to obtain a suitable number of colony forming units. However, residual DNPs were still present on an agar plate, concentrated on the surface during the plating process; this introduces a potential artifact which might affect colony growth. The effect of DNPs remaining on the surface, alongside growing bacteria, has not been previously investigated. In this work, we present the experiments designed to investigate the effect of DNPs on bacterial survival and on the growth of the bacterial colony on a solid media. We employed Escherichia coli and Bacillus subtilis as models of Gram-negative and Gram-positive bacteria, respectively, and Proteus mirabilis as a model of bacterium exhibiting swarming motility on the surfaces. We analyzed the number, area, and weight of bacterial colonies grown on the agar surface covered with DNPs. We did not observe any bactericidal effect of such applied DNPs. However, in all bacterial species used in this work, we observed the appreciable reduction of colony area, which suggests that DNPs obstruct either bacterial growth or motility. The most obvious effect on colony growth was observed in the case of motile P. mirabilis. We show that DNPs act as the mechanical barrier blocking the lateral colony growth.

σI from Bacillus subtilis: Impact on Gene Expression and Characterization of σI-Dependent Transcription That Requires New Types of Promoters with Extended -35 and -10 Elements.

The σI sigma factor from Bacillus subtilis is a σ factor associated with RNA polymerase (RNAP) that was previously implicated in adaptation of the cell to elevated temperature. Here, we provide a comprehensive characterization of this transcriptional regulator. By transcriptome sequencing (RNA-seq) of wild-type (wt) and σI-null strains at 37°C and 52°C, we identified ∼130 genes affected by the absence of σI Further analysis revealed that the majority of these genes were affected indirectly by σI The σI regulon, i.e., the genes directly regulated by σI, consists of 16 genes, of which eight (the dhb and yku operons) are involved in iron metabolism. The involvement of σI in iron metabolism was confirmed phenotypically. Next, we set up an in vitro transcription system and defined and experimentally validated the promoter sequence logo that, in addition to -35 and -10 regions, also contains extended -35 and -10 motifs. Thus, σI-dependent promoters are relatively information rich in comparison with most other promoters. In summary, this study supplies information about the least-explored σ factor from the industrially important model organism B. subtilisIMPORTANCE In bacteria, σ factors are essential for transcription initiation. Knowledge about their regulons (i.e., genes transcribed from promoters dependent on these σ factors) is the key for understanding how bacteria cope with the changing environment and could be instrumental for biotechnologically motivated rewiring of gene expression. Here, we characterize the σI regulon from the industrially important model Gram-positive bacterium Bacillus subtilis We reveal that σI affects expression of ∼130 genes, of which 16 are directly regulated by σI, including genes encoding proteins involved in iron homeostasis. Detailed analysis of promoter elements then identifies unique sequences important for σI-dependent transcription. This study thus provides a comprehensive view on this underexplored component of the B. subtilis transcription machinery.

The enzymatic de-epithelialization technique determines denuded amniotic membrane integrity and viability of harvested epithelial cells.

The human amniotic membrane (HAM) is widely used for its wound healing effect in clinical practice, as a feeder for the cell cultivation, or a source of cells to be used in cell therapy. The aim of this study was to find effective and safe enzymatic HAM de-epithelialization method leading to harvesting of both denuded undamaged HAM and viable human amniotic epithelial cells (hAECs). The efficiency of de-epithelialization using TrypLE Express, trypsin/ ethylenediaminetetraacetic (EDTA), and thermolysin was monitored by hematoxylin and eosin staining and by the measurement of DNA concentration. The cell viability was determined by trypan blue staining. Scanning electron microscopy and immunodetection of collagen type IV and laminin α5 chain were used to check the basement membrane integrity. De-epithelialized hAECs were cultured and their stemness properties and proliferation potential was assessed after each passage. The HAM was successfully de-epithelialized using all three types of reagents, but morphological changes in basement membrane and stroma were observed after the thermolysin application. About 60% of cells remained viable using trypsin/EDTA, approximately 6% using TrypLE Express, and all cells were lethally damaged after thermolysin application. The hAECs isolated using trypsin/EDTA were successfully cultured up to the 5th passage with increasing proliferation potential and decreased stem cell markers expression (NANOG, SOX2) in prolonged cell culture. Trypsin/EDTA technique was the most efficient for obtaining both undamaged denuded HAM and viable hAECs for consequent culture.

Teicoplanin resistance in Staphylococcus haemolyticus is associated with mutations in histidine kinases VraS and WalK.

We investigated the genetic basis of glycopeptide resistance in laboratory-derived strains of S. haemolyticus with emphasis on differences between vancomycin and teicoplanin. The genomes of two stable teicoplanin-resistant laboratory mutants selected on vancomycin or teicoplanin were sequenced and compared to parental S. haemolyticus strain W2/124. Only the two non-synonymous mutations, VraS Q289K and WalK V550L were identified. No other mutations or genome rearrangements were detected. Increased cell wall thickness, resistance to lysostaphin-induced lysis and adaptation of cell growth rates specifically to teicoplanin were phenotypes observed in a sequenced strain with the VraS Q289K mutation. Neither of the VraS Q289K and WalK V550L mutations was present in the genomes of 121S. haemolyticus clinical isolates. However, all but two of the teicoplanin resistant strains carried non-synonymous SNPs in vraSRTU and walKR-YycHIJ operons pointing to their importance for the glycopeptide resistance.

Alloscardovia venturai sp. nov., a fructose 6-phosphate phosphoketolase-positive species isolated from the oral cavity of a guinea-pig (Cavia aperea f. porcellus).

A slightly irregular, short rod-shaped bacterial strain, MOZIV/2T, showing activity of fructose 6-phosphate phosphoketolase was isolated from the oral cavity of a home-bred guinea-pig. Based on comparative 16S rRNA gene sequence analyses, its closest relatives were Alloscardovia omnicolens DSM 21503T and Alloscardovia criceti DSM 17774T with 96.0 and 95.6 % pairwise similarities, respectively. Completeness of the compared sequences was 97.3 and 96.9 %, respectively. Growth was found only under anaerobic conditions. Activities of α- and ß-gluco(galacto)sidases were detected in strain MOZIV/2T, which is characteristic for almost all members of the family Bifidobacteriaceae. Sequencing of other molecular markers (fusA, gyrB and xfp) revealed low gene sequence similarities to A. omnicolens DSM 21503T ranging from 72.7 to 87.5 %. Strain MOZIV/2T differed from other species within the genus Alloscardovia by the presence of C18 : 1ω9t. In addition, much higher proportions of C8 : 0, C11 : 0, C12 : 0, C14 : 1, C16 : 1 and C17 : 0 fatty acids were found in cells of strain MOZIV/2T. The peptidoglycan structure was of type A4α [l-Lys(l-Orn)-d-Asp], which is consistent with its classification within the genus Alloscardovia. The DNA G+C content (45.8 mol%) was lower than those found in other alloscardovia. Phylogenetic studies and evaluation of phenotypic characteristics including the results of biochemical, physiological and chemotaxonomic analyses confirmed the novel species status for strain MOZIV/2T, for which the name Alloscardovia venturai sp. nov. is proposed. The type strain is MOZIV/2T (=DSM 100237T=CCM 8604T=LMG 28781T).