Rod-shaped Cells Research Articles

Raineya orbicola gen. nov., sp. nov. a slightly thermophilic bacterium of the phylum Bacteroidetes and the description of Raineyaceae fam. nov.

An isolate, designated SPSPC-11T, with an optimum growth temperature of about 50 °C and an optimum pH for growth between 7.5 and 8.0, was recovered from a hot spring in central Portugal. Based on phylogenetic analysis of its 16S rRNA sequence, the new organism is most closely related to the species of the genus Thermonema but with a pairwise sequence similarity of <85 %. The isolate was orange-pigmented, formed non-motile long filaments and rod-shaped cells that stain Gram-negative. The organism was strictly aerobic, oxidase-positive and catalase-positive. The major fatty acids were iso-C15:0, iso-C15 : 0 2-OH and iso-C17 : 0 3-OH. The major polar lipids were one aminophospholipid, two aminolipids and three unidentified lipids. Menaquinone 7 was the major respiratory quinone. The DNA G+C content of strain SPSPC-11T was 37.6 mol% (draft genome sequence). The high quality draft genome sequence corroborated many of the phenotypic characteristics of strain SPSPC-11T. Based on genotypic, phylogenetic, physiological and biochemical characterization we describe a new species of a novel genus represented by strain SPSPC-11T (=CECT 9012T=LMG 29233T) for which we propose the name Raineya orbicola gen. nov., sp. nov. We also describe the family Raineyaceae to accommodate this new genus and species.

A Microfluidic Device for Massively Parallel, Whole-lifespan Imaging of Single Fission Yeast Cells.

Whole-lifespan single-cell analysis has greatly increased our understanding of fundamental cellular processes such as cellular aging. To observe individual cells across their entire lifespan, all progeny must be removed from the growth medium, typically via manual microdissection. However, manual microdissection is laborious, low-throughput, and incompatible with fluorescence microscopy. Here, we describe assembly and operation of the multiplexed-Fission Yeast Lifespan Microdissector (multFYLM), a high-throughput microfluidic device for rapidly acquiring single-cell whole-lifespan imaging. multFYLM captures approximately one thousand rod-shaped fission yeast cells from up to six different genetic backgrounds or treatment regimens. The immobilized cells are fluorescently imaged for over a week, while the progeny cells are removed from the device. The resulting datasets yield high-resolution multi-channel images that record each cell's replicative lifespan. We anticipate that the multFYLM will be broadly applicable for single-cell whole-lifespan studies in the fission yeast (Schizosaccharomyces pombe) and other symmetrically-dividing unicellular organisms.

Chromosome Translocation Inflates &lt;i&gt;Bacillus subtilis&lt;/i&gt; Forespores and Impacts Cellular Morphology

The means by which the physicochemical properties of different cellular components together determine bacterial cell shape remain poorly understood. Here, we investigate a programmed cell-shape change during B. subtilis sporulation, when a rod-shaped vegetative cell is transformed to an oval spore. Asymmetric cell division generates a bigger mother cell and a smaller, hemispherical forespore. The septum traps the forespore chromosome, which is translocated to the forespore by SpoIIIE. Simultaneously, forespore size increases as it is reshaped into an ovoid. Using genetics, timelapse microscopy, cryo-electron tomography and mathematical modeling, we demonstrate that forespore growth relies on membrane synthesis and SpoIIIE-mediated chromosome translocation, but not on peptidoglycan or protein synthesis. Our data suggest that the hydrated nucleoid swells and inflates the forespore, displacing ribosomes to the cell periphery, stretching septal peptidoglycan and reshaping the forespore. Our results illustrate how simple biophysical interactions between core cellular components contribute to cellular morphology.

Status Epilepticus Triggers Time-Dependent Alterations in Microglia Abundance and Morphological Phenotypes in the Hippocampus.

Status epilepticus (SE) is defined by the occurrence of prolonged “non-stop” seizures that last for at least 5 min. SE provokes inflammatory responses including the activation of microglial cells, the brain’s resident immune cells, which are thought to contribute to the neuropathology and pathophysiology of epilepsy. Microglia are professional phagocytes that resemble peripheral macrophages. Upon sensing immune disturbances, including SE, microglia become reactive, produce inflammatory cytokines, and alter their actin cytoskeleton to transform from ramified to amoeboid shapes. It is widely known that SE triggers time-dependent microglial expression of pro-inflammatory cytokines that include TNFα and IL-1β. However, less is known in regards to the spatiotemporal progression of the morphological changes, which may help define the extent of microglia reactivity after SE and potential function (surveillance, inflammatory, phagocytic). Therefore, in this study, we used the microglia/macrophage IBA1 marker to identify and count these cells in hippocampi from control rats and at 4 h, 3 days, and 2 weeks after a single episode of pilocarpine-induced SE. We identified, categorized, and counted the IBA1-positive cells with the different morphologies observed after SE in the hippocampal areas CA1, CA3, and dentate gyrus. These included ramified, hypertrophic, bushy, amoeboid, and rod. We found that the ramified phenotype was the most abundant in control hippocampi. In contrast, SE provoked time-dependent changes in the microglial morphology that was characterized by significant increases in the abundance of bushy-shaped cells at 4 h and amoeboid-shaped cells at 3 days and 2 weeks. Interestingly, a significant increase in the number of rod-shaped cells was only evident in the CA1 region at 2 weeks after SE. Taken together, these data suggest that SE triggers time-dependent alterations in the morphology of microglial cells. This detailed description of the spatiotemporal profile of SE-induced microglial morphological changes may help provide insight into their contribution to epileptogenesis.

Mechanisms Connecting the Conserved Protein Kinases Ssp1, Kin1, and Pom1 in Fission Yeast Cell Polarity and Division

Connections between the protein kinases that function within complex cell polarity networks are poorly understood. Rod-shaped fission yeast cells grow in ahighly polarized manner, and genetic screens haveidentified many protein kinases, including theCaMKK-like Ssp1 and the MARK/PAR-1 family kinase Kin1, that are required for polarized growth and cell shape, but their functional mechanisms and connections have been unknown [1-5]. We found that Ssp1 promotes cell polarity by phosphorylating the activation loop of Kin1. Kin1 regulates cell polarity and cytokinesis through unknown mechanisms [4-7]. We performed a large-scale phosphoproteomic screen and found that Kin1 phosphorylates itself and Pal1 to promote growth at cell tips, and these proteins are interdependent for localization to growing cell tips. Additional Kin1 substrates for cell polarity and cytokinesis (Tea4, Mod5, Cdc15, and Cyk3) were also phosphorylated by asecond kinase, the DYRK family member Pom1 [8]. Kin1 and Pom1 were enriched at opposite ends of growing cells, and they phosphorylated largely non-overlapping sites on shared substrates. Combined inhibition of both Kin1and Pom1 led to synthetic defects in their shared substrates Cdc15 andCyk3, confirming a non-redundant functional connection through shared substrates. These findings uncover a new Ssp1-Kin1 signaling pathway, and define its functional and mechanistic connection with Pom1 signaling for cellpolarity and cytokinesis. These kinases are conserved in many eukaryotes including humans, suggesting that similar connections and mechanisms might operate in a broad range of cells.

Developmental stage influences chromosome segregation patterns and arrangement in the extremely polyploid, giant bacterium Epulopiscium sp. type B.

Few studies have described chromosomal dynamics in bacterial cells with more than two complete chromosome copies or described changes with respect to development in polyploid cells. We examined the arrangement of chromosomal loci in the very large, highly polyploid, uncultivated intestinal symbiont Epulopiscium sp. type B using fluorescent in situ hybridization. We found that in new offspring, chromosome replication origins (oriCs) are arranged in a three-dimensional array throughout the cytoplasm. As development progresses, most oriCs become peripherally located. Siblings within a mother cell have similar numbers of oriCs. When chromosome orientation was assessed in situ by labeling two chromosomal regions, no specific pattern was detected. The Epulopiscium genome codes for many of the conserved positional guide proteins used for chromosome segregation in bacteria. Based on this study, we present a model that conserved chromosomal maintenance proteins, combined with entropic demixing, provide the forces necessary for distributing oriCs. Without the positional regulation afforded by radial confinement, chromosomes are more randomly oriented in Epulopiscium than in most small rod-shaped cells. Furthermore, we suggest that the random orientation of individual chromosomes in large polyploid cells would not hamper reproductive success as it would in smaller cells with more limited genomic resources.

Cardioprotective Effect of Hydroxysafflor Yellow A via the Cardiac Permeability Transition Pore.

Myocardial ischemia damages cardiac myocytes in part via opening of the mitochondrial permeability transition pore. Preventing this pore's opening is therefore a useful therapeutic goal in treating cardiovascular disease. Hydroxysafflor yellow A has been proposed as a nontoxic alternative to other agents that modulate mitochondrial permeability transition pore opening. In this study, we proposed that hydroxysafflor yellow A prevents mitochondrial permeability transition pore formation in anoxic cardiac myocytes, and thus protects the cell from damage seen during reoxygenation of the cardiac myocytes. Experiments with hydroxysafflor yellow A transport in aerobic myocytes show that roughly 50% of the extracellular dye concentration crosses the cell membrane in a 2-h incubation. In our anoxia/reoxygenation protocol, hydroxysafflor yellow A modulated both the reduction of viability and the loss of rod-shaped cells that attend anoxia and reoxygenation. Hydroxysafflor yellow A's protective effect was similar to that of cyclosporin A, an agent known to inhibit mitochondrial permeability transition pore opening. In additional experiments, plated myocytes were loaded with calcein/MitoTracker Red, then examined for intracellular dye distribution/morphology after anoxia/reoxygenation. Hydroxysafflor yellow A-containing cells showed a cardioprotective pattern similar to that of cyclosporin A (an agent known to close the mitochondrial permeability transition pore). We conclude that hydroxysafflor yellow A can enter the cardiac myocyte and is able to modulate anoxia/reoxygenation-induced damage by interacting with the mitochondrial permeability transition pore.

FIRST REPORT OF PANTOEA STEWARTII subsp. STEWARTII CAUSING JACKFRUIT BRONZING DISEASE IN MEXICO

“Jackfruit bronzing”, an emerging disease affecting jackfruit (Artocarpus heterophyllus) caused by the bacterium Pantoea stewartii, had only been reported in the Philippines and Malaysia. It is characterized by yellow-orange to reddish discoloration of affected pulps and rags (Gapasin et al., 2014; Zulperio et al., 2017). In autumn 2016, the disease was suspected in major growing areas in Nayarit, Mexico, where jackfruits with typical fruit bronzing symptoms were observed and collected. The fruits were excised and the sampled tissues were disinfected with 1% sodium hypochlorite (2 min), then rinsed with sterilized water and placed on Luria-Bertani (LB) agar medium. Ten bacterial isolates with characteristics particular to Pantoea spp. (Gapasin et al., 2014) and resembling the P. stewartii subsp. stewartii (Smith) Mergaert et al. (ATCC8199) type strain, showing round, yellow pigmented, smooth, translucent colonies, Gram-negative and rod-shaped cells and positive for catalase, protease and amylase production, were obtained and selected for further examination. The cpsD and hrpS genes obtained from a typical Mexican bacterial isolate from jackfruit JM-5´ (deposited in the Bacterial Collection of the Autonomus University of Nayarit) (GenBank accession Nos. KY965964 and KY965965) with primers specific to Pantoea stewartii (CPSL1-CPSR2c and HRP1d-HRP3c; Coplin et al., 2002) showed 98-99% similarity to reference strains of P. stewartii subsp. stewartii (EU215384, AB894429-27, AF282857). Furthermore, sequence analysis of a 16S rDNA region (KY965963), amplified with bacterial universal primers FD1-RD1 (Weisburg et al., 1991), showed that our JM-5´ bacterial isolate was 99% identical to Pantoea stewartii subsp. stewartii (KM508073, HG792421). Phylogenetic analyses of sequences were conducted, using MEGA 7 software, by the Maximum Likelihood method. The cspD and 16S rDNA sequences were compared with the available sequences for these genes in genomes of P. stewartii subsp. stewartii, while the hrpS gene sequence was compared against hrpS sequences of phytopatogenic bacteria available in the database, including that of P. stewartii subsp. stewartii. Dinoroseobacter shibae (CP000830) was included as an outgroup. Analysis based on cspD sequence indicated that Mexican bacterial isolates clustered together in the clade of P. stewartii subsp. stewartii. Moreover hrpS and 16S rDNA sequences obtained from our bacterial isolate JM-5´grouped respectively with hrpS and 16S rDNA genes from available genomes of P. stewartii subsp. stewartii. Pathogenicity tests showed characteristic symptoms of fruit bronzing disease 14 days post inoculation in jackfruits inoculated by injection with a bacterial suspension (10 ml, 108 cfu ml-1 ). No symptoms were observed in control fruits inoculated with sterile water. Bacteria re-isolated from inoculated jackfruits were identical to the original isolate showing the morphological, biochemical and microscopic characteristics described above. Furthermore the PCR analyses of the cpsD and hrpS genes revealed that JM-J1, JM-J6, JM-J11 reisolates generated amplicons with similar sizes to those obtained from the P. stewartii subsp. stewartii ATCC8199 type strain. Based on morphological, biochemical, physiological, pathogenicity tests and molecular analyses, we conclude that Mexican bacterial isolates from Jackfruit can be identified as Pantoea stewartii subsp. stewartii. This is the first record of jackfruit bronzing disease in Mexico.

The CodY-dependent clhAB2 operon is involved in cell shape, chaining and autolysis in Bacillus cereus ATCC 14579.

The Gram-positive pathogen Bacillus cereus is able to grow in chains of rod-shaped cells, but the regulation of chaining remains largely unknown. Here, we observe that glucose-grown cells of B. cereus ATCC 14579 form longer chains than those grown in the absence of glucose during the late exponential and transition growth phases, and identify that the clhAB2 operon is required for this chain lengthening phenotype. The clhAB2 operon is specific to the B. cereus group (i.e., B. thuringiensis, B. anthracis and B. cereus) and encodes two membrane proteins of unknown function, which are homologous to the Staphylococcus aureus CidA and CidB proteins involved in cell death control within glucose-grown cells. A deletion mutant (ΔclhAB2) was constructed and our quantitative image analyses show that ΔclhAB2 cells formed abnormal short chains regardless of the presence of glucose. We also found that glucose-grown cells of ΔclhAB2 were significantly wider than wild-type cells (1.47 μm ±CI95% 0.04 vs 1.19 μm ±CI95% 0.03, respectively), suggesting an alteration of the bacterial cell wall. Remarkably, ΔclhAB2 cells showed accelerated autolysis under autolysis-inducing conditions, compared to wild-type cells. Overall, our data suggest that the B. cereus clhAB2 operon modulates peptidoglycan hydrolase activity, which is required for proper cell shape and chain length during cell growth, and down-regulates autolysin activity. Lastly, we studied the transcription of clhAB2 using a lacZ transcriptional reporter in wild-type, ccpA and codY deletion-mutant strains. We found that the global transcriptional regulatory protein CodY is required for the basal level of clhAB2 expression under all conditions tested, including the transition growth phase while CcpA, the major global carbon regulator, is needed for the high-level expression of clhAB2 in glucose-grown cells.

Constant Flux of Spatial Niche Partitioning through High-Resolution Sampling of Magnetotactic Bacteria.

Magnetotactic bacteria (MTB) swim along magnetic field lines in water. They are found in aquatic habitats throughout the world, yet knowledge of their spatial and temporal distribution remains limited. To help remedy this, we took MTB-bearing sediment from a natural pond, mixed the thoroughly homogenized sediment into two replicate aquaria, and then counted three dominant MTB morphotypes (coccus, spirillum, and rod-shaped MTB cells) at a high spatiotemporal sampling resolution: 36 discrete points in replicate aquaria were sampled every ∼30 days over 198 days. Population centers of the MTB coccus and MTB spirillum morphotypes moved in continual flux, yet they consistently inhabited separate locations, displaying significant anticorrelation. Rod-shaped MTB were initially concentrated toward the northern end of the aquaria, but at the end of the experiment, they were most densely populated toward the south. The finding that the total number of MTB cells increased over time during the experiment argues that population reorganization arose from relative changes in cell division and death and not from migration. The maximum net growth rates were 10, 3, and 1 doublings day-1 and average net growth rates were 0.24, 0.11, and 0.02 doublings day-1 for MTB cocci, MTB spirilla, and rod-shaped MTB, respectively; minimum growth rates for all three morphotypes were -0.03 doublings day-1 Our results suggest that MTB cocci and MTB spirilla occupy distinctly different niches: their horizontal positioning in sediment is anticorrelated and under constant flux.IMPORTANCE Little is known about the horizontal distribution of magnetotactic bacteria in sediment or how the distribution changes over time. We therefore measured three dominant magnetotactic bacterium morphotypes at 36 places in two replicate aquaria each month for 7 months. We found that the spatial positioning of population centers changed over time and that the two most abundant morphotypes (MTB cocci and MTB spirilla) occupied distinctly different niches in the aquaria. Maximum and average growth and death rates were quantified for each of the three morphotypes based on 72 sites that were measured six times. The findings provided novel insight into the differential behavior of noncultured magnetotactic bacteria.

Marivibrio halodurans gen. nov., sp. nov., a marine bacterium in the family Rhodospirillaceae isolated from underground rock salt.

Gram-negative, spiral or curved rod-shaped cells of a bacterial strain, designated ZC80T, were isolated from a rock salt sample collected at Yunnan salt mine, China. Analysis of the strain's 16S rRNA gene sequence revealed a clear affiliation of this novel strain within the family Rhodospirillaceae. Strain ZC80T formed a robust cluster with Pelagibius litoralis CL-UU02T at a 16S rRNA gene sequence similarity level of 88.1 %. Strain ZC80T shared no more than 91.0 % 16S rRNA gene sequence similarity with the type strains of other species in the family Rhodospirillaceae. Strain ZC80T was able to grow in the presence of 2-15 % (w/v) NaCl, and grew at 10-50 °C and pH 6.0-10.0. The major fatty acids were C19 : 0 cyclo ω8c (41.3 %). The major isoprenoid quinone was ubiquinone 10 (Q-10). The major polar lipids were phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol and an unidentified aminolipid. The DNA G+C content of strain ZC80T was 60.8 mol%. On the basis of phylogenetic analyses and chemotaxonomic and physiological data, strain ZC80T is considered to represent a novel species of a new genus in the family Rhodospirillaceae, for which the name Marivibrio halodurans gen. nov., sp. nov. is proposed. The type strain of Marivibrio halodurans is ZC80T (=CGMCC 1.15697T=NBRC 112461T).

Nocardioides taihuensis sp. nov., isolated from fresh water lake sediment.

A novel actinobacterial strain, designated X17T, was isolated from the sediment of Taihu Lake in China and its taxonomic position was investigated by using a polyphasic approach. The isolate formed milky-white colonies comprising aerobic, Gram-stain-positive, rod-shaped cells. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the organism belonged to the genus Nocardioides and consistently formed a distinct cluster with Nocardioides agariphilus JCM 16020T and Nocardioides islandensis MSL 26T, sharing 95.5 and 94.8 % 16S rRNA gene sequence similarities, respectively. The genomic DNA G+C content was 69.9 mol%. Chemotaxonomically, the isolate contained ll-diaminopimelic acid in the cell-wall peptidoglycan, MK-8 (H4) as the predominant menaquinone, and diphosphatidylglycerol, phosphatidylglycerol and phosphatidylinositol as the polar lipids found in the cell wall. The major fatty acids were 18 : 1 ω9c (38.6 %), 16 : 0 iso (20.3 %), 15 : 0 iso (6.8 %) and 18 : 0 (5.8 %). Based on its physiological, biochemical and chemotaxonomic characteristics, the strain represents a novel species of the genus Nocardioides, for which the name Nocardioides taihuensis sp. nov. (type strain X17T=CGMCC 4.7318T=NBRC 112321T) is proposed.

Rhodopseudomonas telluris sp. nov., a phototrophic alphaproteobacterium isolated from paddy soil.

A strain of anoxygenic phototrophic bacteria isolated from paddy soil (designated strain TUT3615T) was studied taxonomically in comparison with Rhodopseudomonasstrain ATCC 17005 as its nearest phylogenetic relative. Strains TUT3615T and ATCC 17005 had budding rod-shaped cells and showed in vivo absorption maxima at 804 and 860 nm in the near infrared region, indicating the presence of bacteriochlorophyll a. The intracytoplasmic membrane system was of the lamellar type parallel to the cytoplasmic membrane. 16S rRNA gene sequence comparisons showed that strains TUT3615T and ATCC 17005 had a 99.7 % level of similarity to one another and were closest to Rhodopseudomonas palustris ATCC 17001T (98.6 % similarity) among the established species of the genus Rhodopseudomonas. Genomic DNA-DNA hybridization studies revealed that strains TUT3615T and ATCC 17005 had an average similarity level of 65 % to one another and of less than 40 % to the available type strains of Rhodopseudomonas species. Results of phenotypic studies showed that strains TUT3615T and ATCC 17005 could be differentiated from one another and from any previously described species of Rhodopseudomonas. The G+C contents of the genomic DNA of strain TUT3615T and ATCC 17005 were 66.3 and 66.5 mol%, respectively. Based on these data, we propose the name Rhodopseudomonas telluris sp. nov. for strain TUT3615T. The type strain is TUT3615T (=KCTC 23279T=NBRC 107609T). We suspend a proposal to reclassify strain ATCC 17005 as a novel species or subspecies until a genome-wide analysis provides more definite information on its taxonomic position.

Dyella caseinilytica sp. nov., Dyella flava sp. nov. and Dyella mobilis sp. nov., isolated from forest soil.

Three strains, DHOB09T, DHOC52T and DHON07T, were isolated from the forest soil of Dinghushan Biosphere Reserve, Guangdong Province, PR China. They were all Gram-stain-negative, aerobic, rod-shaped cells. The ranges (optimum) for the temperature, pH and NaCl concentration for growth of DHOB09T, DHOC52T and DHON07T were 10-42 (25-28) °C, pH 5.5-9.0 (7.0-7.5) and 0-4.0 (0-0.5) % (w/v); 10-42 (28) °C, pH 4.0-7.0 (4.5-6.5) and 0-2.0 (0) % (w/v) and 10-37 (25-28) °C, pH 4.0-7.5 (5.5-6.0) and 0-2.5 (0) % (w/v), respectively. Phylogenetic analysis based on 16S rRNA gene sequences showed that DHOB09T, DHOC52T and DHON07T formed a phyletic cluster with seven species of the genus Dyella within the major clade of Dyella with sequence similarities ranged from 96.9 to 98.6 %. This indicated that the three strains may represent three novel species of the genus Dyella. This result was also strongly supported by the concatenated analysis of partial gyrB, lepA and recA gene sequences. DNA-DNA hybridization between strains DHON07T and DHOB09T, as well as DHON07T and Dyella koreensis BB4T was much lower than 70 %. The G+C content of strains DHOB09T, DHOC52T and DHON07T were 59.4, 60.7 and 59.5 %, respectively. The major fatty acids of the three strains were iso-C15 : 0, iso-C16 : 0 and iso-C17 : 0 and the predominant respiratory lipoquinone was ubiquinone-8. All of the physiological, phylogenetic and chemotaxonomic data showed that strains DHOB09T, DHOC52T and DHON07T are distinctive from each other and from all species of the genus Dyellawith validly published names. Therefore, we suggest that they represent three novel species of the genus, for which the names Dyella caseinilytica sp. nov. (type strain DHOB09T=CGMCC 1.15434T=LMG 29202T), Dyella flava sp. nov. (type strain DHOC52T=NBRC 111979T=KCTC 52128T) and Dyella mobilis sp. nov. (type strain DHON07T=CGMCC 1.15400T=NBRC 111475T) are proposed.

Coordination of Chromosome Segregation and Cell Division in Staphylococcus aureus

Productive bacterial cell division and survival of progeny requires tight coordination between chromosome segregation and cell division to ensure equal partitioning of DNA. Unlike rod-shaped bacteria that undergo division in one plane, the coccoid human pathogen Staphylococcus aureus divides in three successive orthogonal planes, which requires a different spatial control compared to rod-shaped cells. To gain a better understanding of how this coordination between chromosome segregation and cell division is regulated in S. aureus, we investigated proteins that associate with FtsZ and the divisome. We found that DnaK, a well-known chaperone, interacts with FtsZ, EzrA and DivIVA, and is required for DivIVA stability. Unlike in several rod shaped organisms, DivIVA in S. aureus associates with several components of the divisome, as well as the chromosome segregation protein, SMC. This data, combined with phenotypic analysis of mutants, suggests a novel role for S. aureus DivIVA in ensuring cell division and chromosome segregation are coordinated.

Pedobacter psychrophilus sp. nov., isolated from fragmentary rock.

Strain P4487AT was isolated during investigation of cultivable bacterial populations of environmental materials sampled at James Ross Island, Antarctica. It revealed Gram-stain-negative short rod-shaped cells producing a pink pigment. Phylogenetic analysis based on 16S rRNA gene sequences allocated strain P4487AT to the genus Pedobacter but showed that the strain represents a distinct intrageneric phylogenetic lineage clearly separated from remaining Pedobacter species. Phylogenetically, strain P4487AT formed a common branch with the Pedobacter arcticus and Pedobacter lignilitoris cluster while the highest value of 94.4 % 16S rRNA gene sequence similarity suggested that Pedobacter lentus is the most closely related species. Biochemical and physiological test results enabled the differentiation of strain P4487AT from all phylogenetically closely related species. Chemotaxonomic analyses of strain P4487AT showed MK-7 as the respiratory menaquinone, sym-homospermidine as the major polyamine, phosphatidylethanolamine and two unidentified lipids as the major polar lipids, presence of sphingolipids, and C16 : 1ω7c/C16 : 1ω6c (summed feature 3), iso-C15 : 0 and iso-C17 : 0 3-OH as the major fatty acids, all of which corresponded with characteristics of the genus Pedobacter. The results showed that strain P4487AT represents a novel species within the genus Pedobacter, for which the name Pedobacter psychrophilus sp. nov. is proposed. The type strain is P4487AT (=CCM 8644T=LMG 29436T).

Paenibacillus polysaccharolyticus sp. nov., a xylanolytic and cellulolytic bacteria isolated from leaves of Bamboo Phyllostachys aureosulcata.

A novel xylanolytic and cellulolytic strain, BL9T, was isolated from leaves of the Bamboo plants maintained at the Tamil Nadu Agricultural University Campus, Coimbatore, India. On the basis of the results of 16S rRNA gene analysis, it was determined to be phylogenetically close to the type strains of Paenibacillus amylolyticus NRRL NRS-290T (98.3 %), Paenibacillus barcinonensis BP-23T (98.1 %), Paenibacillus tundrae A10bT (98.0 %) and Paenibacillus xylanexedens B22aT (97.6 %). The strain stained Gram variable and was aerobic, motile and catalase- and oxidase-positive, with rod-shaped cells. Based upon the genome sequence, the average nucleotide identity with the related species ranged from 66 % to 72 %, and the digital DNA-DNA hybridization value ranged from 13 % to 27 %. The DNA G+C content was 45.6 mol%, meso-diaminopimelic acid was identified as the predominant component of the cell wall, and MK-7 was the only menaquinone in cell membranes. The whole-cell fatty acid profile included C16 : 0, iso-C14 : 0, anteiso-C15 : 0 and iso-C16 : 0. The polar lipids identified were phosphatidylethanolamine, phosphatidylglycerol, phosphatidylserine and diphosphtidylglycerol. On the basis of these polyphasic taxonomic traits, BL9T represents a novel species of the genus Paenibacillus, for which the name Paenibacillus polysaccharolyticus sp. nov. is proposed. The type strain is BL9T (=NBRC 105191T=ICMP 17623T).

Merdimonas faecis gen. nov., sp. nov., isolated from human faeces.

A strictly anaerobic Gram-stain-positive, non-motile and non-spore-forming bacterial strain, BR31T, was isolated from a faecal sample of a healthy human. Bacterial colonies were ivory-coloured on GAM agar and composed of rod-shaped cells with rounded ends approximately 1.4-2.1×0.5-0.6 µm in size. According to comparative analysis based on 16S rRNA gene sequences, strain BR31T formed a distinct phylogenetic lineage that reflects a new genus within Clostridium cluster XIVa in the family Lachnospiraceae, with highest similarity to Eubacterium contortum DSM 3982T (94.6 %). The DNA G+C content was calculated to be 47.0 mol% from whole genome sequencing. Predicted genes associated with synthesis of teichuronic acid, polyamines, polar lipids and diaminopimelic acid were detected. The peptidoglycan contained meso-diaminopimelic acid and the main polar lipid detected was phosphatidylglycerol. The major metabolic end product of glucose was acetic acid, which was in agreement with those of most members of the family. However, the profile of major cellular fatty acids (C16 : 0, C14 : 0, summed feature 4 and C13 : 0) and overall enzyme activity demonstrated phenotypic differentiation of strain BR31T from other closely related genera. Thus, based on distinct phenotypic, phylogenetic, genotypic and chemotaxonomic characteristics, strain BR31T is considered to represent a novel species of a new genus, for which the name Merdimonas faecis gen. nov., sp. nov. is proposed. The type strain of Merdimonas faecis is BR31T (=KCTC 15482T=JCM 30748T).

An evolutionary optimization of a rhodopsin-based phototrophic metabolism in Escherichia coli

BackgroundThe expression of the Gloeobacter rhodopsin (GR) in a chemotrophic Escherichia coli enables the light-driven phototrophic energy generation. Adaptive laboratory evolution has been used for acquiring desired phenotype of microbial cells and for the elucidation of basic mechanism of molecular evolution. To develop an optimized strain for the artificially acquired phototrophic metabolism, an ancestral E. coli expressing GR was adaptively evolved in a chemostat reactor with constant illumination and limited glucose conditions. This study was emphasized at an unexpected genomic mutation contributed to the improvement of microbial performance.ResultsDuring the chemostat culture, increase of cell size was observed, which were distinguished from that of the typical rod-shaped ancestral cells. A descendant ET5 strain was randomly isolated from the chemostat culture at 88-days. The phototrophic growth and the light-induced proton pumping of the ET5 strain were twofold and eightfold greater, respectively, than those of the ancestral E. coli strain. Single point mutation of C1082A at dgcQ gene (encoding diguanylate cyclase, also known as the yedQ gene) in the chromosome of ET5 strain was identified from whole genome sequencing analysis. An ancestral E. coli complemented with the same dgcQ mutation from the ET5 was repeated the subsequently enhancements of light-driven phototrophic growth and proton pumping. Intracellular c-di-GMP, the product of the diguanylate cyclase (dgcQ), of the descendant ET5 strain was suddenly increased while that of the ancestral strain was negligible.ConclusionsNewly acquired phototrophic metabolism of E. coli was further improved via adaptive laboratory evolution by the rise of a point mutation on a transmembrane cell signaling protein followed by increase of signal molecule that eventually led an increase proton pumping and phototrophic growth.

Cell fiber-based three-dimensional culture system for highly efficient expansion of human induced pluripotent stem cells

Human pluripotent stem cells are a potentially powerful cellular resource for application in regenerative medicine. Because such applications require large numbers of human pluripotent stem cell-derived cells, a scalable culture system of human pluripotent stem cell needs to be developed. Several suspension culture systems for human pluripotent stem cell expansion exist; however, it is difficult to control the thickness of cell aggregations in these systems, leading to increased cell death likely caused by limited diffusion of gases and nutrients into the aggregations. Here, we describe a scalable culture system using the cell fiber technology for the expansion of human induced pluripotent stem (iPS) cells. The cells were encapsulated and cultured within the core region of core-shell hydrogel microfibers, resulting in the formation of rod-shaped or fiber-shaped cell aggregations with sustained thickness and high viability. By encapsulating the cells with type I collagen, we demonstrated a long-term culture of the cells by serial passaging at a high expansion rate (14-fold in four days) while retaining its pluripotency. Therefore, our culture system could be used for large-scale expansion of human pluripotent stem cells for use in regenerative medicine.