The emergence of novel macrolide resistance island in Macrococcus caseolyticus and Staphylococcus aureus of food origin.

Food-derived Staphylococcaceae species with severe antimicrobial resistance, especially Staphylococcus aureus, is a major threat to public health. Macrococcus caseolyticus (M. caseolyticus) is a member of the Staphylococcaceae family which plays a vital role in fermented products and disease causation in animals. In our previous study, several Staphylococcus aureus antibiotic-resistant island msr (SaRImsr) were found in multidrug-resistant S. aureus. In this study, novel SaRImsr, SaRImsr-III emerged from S. aureus. Another novel SaRImsr-like further emerged in M. caseolyticus from food. These isolates' prevalence and genetic environment were investigated and characterized to understand the distribution and transmission of these novel SaRImsr strains. All SaRImsr-positive S. aureus isolates exhibited a multidrug resistance (MDR) phenotype, within which a series of antimicrobial resistance genes (ARGs) and virulence factor genes (VFs) were identified. In addition, three SaRImsr types, SaRImsr-I (15.1 kb), SaRImsr-II (16-17 kb), and SaRImsr-III (18 kb) carrying mef(D)-msr(F), were identified in these isolates' chromosomes. SaRImsr-(I-III) contains a site-specific integrase gene int and operon mef(D)-msr(F). SaRImsr-III has an additional orf3-orf4-IS30 arrangement downstream of mef(D) and msr(F). Moreover, the SaRImsr-like and macrolide-resistant transposon Tn6776 forming a novel mosaic structure coexisted in one M. caseolyticus isolate. Within this mosaic structure, the macrolide-resistant genes mef(D)-msr(F) were absent in SaRImsr-like, whereas an operon, mef(F)-msr(G), was identified in Tn6776. The SaRImsr-(I-III) and SaRImsr-like structure were inserted into the rpsI gene encoding the 30S ribosomal protein S9 in the chromosome. Excision and cyclisation of SaRImsr-III, SaRImsr-like, operon mef(D)-msr(F), and orf3-orf4-IS30 arrangements were confirmed using two-step PCR. This study is the first to report MDR S. aureus harbouring novel SaRImsr-III and M. caseolyticus containing novel mosaic structures isolated from retail foods. Similar SaRImsr-type resistant islands' occurrence and propagation in Staphylococcaceae species require continuous monitoring and investigation.

Identification and characterization of a novel GNAT superfamily Nα -acetyltransferase from Salinicoccus halodurans H3B36.

Nα -acetyl-α-lysine was found as a new type of compatible solutes that acted as an organic cytoprotectant in the strain of Salinicoccus halodurans H3B36. A novel lysine Nα -acetyltransferase gene (shkat), encoding an enzyme that catalysed the acetylation of lysine exclusively at α position, was identified from this moderate halophilic strain and expressed in Escherichia coli. Sequence analysis indicated ShKAT contained a highly conserved pyrophosphate-binding loop (Arg-Gly-Asn-Gly-Asn-Gly), which was a signature of the GNAT superfamily. ShKAT exclusively recognized free amino acids as substrate, including lysine and other basic amino acids. The enzyme showed a wide range of optimal pH value and was tolerant to high-alkali and high-salinity conditions. As a new member of the GNAT superfamily, the ShKAT was the first enzyme recognized free lysine as substrate. We believe this work gives an expanded perspective of the GNAT superfamily, and reveals great potential of the shkat gene to be applied in genetic engineering for resisting extreme conditions.

Investigating the microbial-influenced corrosion of UNS S32750 stainless-steel base alloy and weld seams by biofilm-forming marine bacterium Macrococcus equipercicus.

This study investigates the microbial-influenced corrosion of UNS S32750 super-duplex stainless-steel joints fabricated using different welding methods. Herein, the samples were introduced into a medium inoculated with Macrococcus equipercicus isolated from a marine environment. Confocal laser scanning microscopy and atomic force microscopy were used to characterise the topography and formation of pits in the corroded samples, respectively. Potentiodynamic polarisation studies were conducted on both the base alloy and weld seams exposed for 30 and 60 days in the experimental system inoculated with M. equipercicus and un-inoculated system. Results indicate that the thickness of the biofilm formed due to this bacterium increased and became heterogeneous with an increase in the exposure time, thereby resulting in micro-pits. Bacterial colonisation was observed in all the coupons after exposure to the inoculated medium. Although micro-pits were observed in all the coupons, the base metal and flux-cored arc weld seams showed highest sensitivity to bacterial attack.

Metabolomics-based profiling of three terminal alkene-producing Jeotgalicoccus spp. during different growth phase.

Production of terminal alkenes by microbes has gained importance due to its role as a chemical feedstock in commercial industries. Jeotgalicoccus species has been widely unexplored despite being well-known as a natural producer of terminal alkene, catalyzing the one-step fatty acid decarboxylation reaction by OleTJE cytochrome P450. In this study, widely targeted ion-pair LC-MS/MS was used to monitor central carbon metabolism of Jeotgalicoccus halotolerans JCM 5429, Jeotgalicoccus huakuii JCM 8176, and Jeotgalicoccus psychrophilus JCM 5429 at logarithmic and stationary phases. Growth and production profile of terminal alkene, alcohols and organic acids were also measured. Among the three strains used in this study, J. halotolerans and J. psychrophilus showed higher terminal alkene production compared to J. huakuii. All strains achieved maximum terminal alkene production at logarithmic phase and therefore, detailed analysis of the metabolite profiles of the three strains were performed in logarithmic phase. PCA analysis showed that the strains were discriminated based on their ability to produce terminal alkene along PC1 and some of the important metabolites corresponding to this separation is the acetyl-CoA and 2-oxoglutarate. This study is the first report on metabolite profiling of three Jeotgalicoccus spp. in different growth phases. The results from this study can provide a better understanding of the changes that occur in the metabolome level during growth and production of terminal alkene in Jeotgalicoccus species.

Development of the first gene expression system for Salinicoccus strains with potential application in bioremediation of hypersaline wastewaters.

Salinicoccus salsiraiae IM408 (=CGMCC13032) is a novel halophilic bacterium that we isolated from the saline soil of Da Gang Oilfield. It tolerates 60 g/l sodium chloride and up to 123 g/l (1.5 M) sodium acetate and has shown a potential application in bioremediation of wastewater with high salt and high chemical oxygen demand (COD). Two plasmids, pS408-1 and pS408-2, were identified in S. salsiraiae IM408, and the sequences and copy numbers of the plasmids were determined. Based on these plasmids, two shuttle vectors containing a replicon for Escherichia coli, ampicillin, and chloramphenicol resistance genes, as well as the replicon from pS408-1 or pS408-2, were constructed and named as pTCS101 and pTCS201, respectively. A suitable host strain, named S. salsiraiae PE01, was also developed from the wild-type by plasmid elimination. Using the plasmid pTCS101 as an expression vector, L-lactate dehydrogenase from Staphylococcus aureus was expressed successfully in S. salsiraiae PE01. This is the first gene expression system for the Salinicoccus genus. It has provided the potential for expression of desired proteins or for establishment of desired pathways in Salinicoccus strains, which would make these halophiles more advantageous in future biotechnological applications.

Mutagenesis and redox partners analysis of the P450 fatty acid decarboxylase OleTJE.

The cytochrome P450 enzyme OleTJE from Jeotgalicoccus sp. ATCC 8456 is capable of converting free long-chain fatty acids into α-alkenes via one-step oxidative decarboxylation in presence of H2O2 as cofactor or using redox partner systems. This enzyme has attracted much attention due to its intriguing but unclear catalytic mechanism and potential application in biofuel production. Here, we investigated the functionality of a select group of residues (Arg245, Cys365, His85, and Ile170) in the active site of OleTJE through extensive mutagenesis analysis. The key roles of these residues for catalytic activity and reaction type selectivity were identified. In addition, a range of heterologous redox partners were found to be able to efficiently support the decarboxylation activity of OleTJE. The best combination turned out to be SeFdx-6 (ferredoxin) from Synechococcus elongatus PCC 7942 and CgFdR-2 (ferredoxin reductase) from Corynebacterium glutamicum ATCC 13032, which gave the highest myristic acid conversion rate of 94.4%. Moreover, Michaelis-Menton kinetic parameters of OleTJE towards myristic acid were determined.

Catalytic Determinants of Alkene Production by the Cytochrome P450 Peroxygenase OleTJE.

The Jeotgalicoccus sp. peroxygenase cytochrome P450 OleTJE (CYP152L1) is a hydrogen peroxide-driven oxidase that catalyzes oxidative decarboxylation of fatty acids, producing terminal alkenes with applications as fine chemicals and biofuels. Understanding mechanisms that favor decarboxylation over fatty acid hydroxylation in OleTJE could enable protein engineering to improve catalysis or to introduce decarboxylation activity into P450s with different substrate preferences. In this manuscript, we have focused on OleTJE active site residues Phe79, His85, and Arg245 to interrogate their roles in substrate binding and catalytic activity. His85 is a potential proton donor to reactive iron-oxo species during substrate decarboxylation. The H85Q mutant substitutes a glutamine found in several peroxygenases that favor fatty acid hydroxylation. H85Q OleTJE still favors alkene production, suggesting alternative protonation mechanisms. However, the mutant undergoes only minor substrate binding-induced heme iron spin state shift toward high spin by comparison with WT OleTJE, indicating the key role of His85 in this process. Phe79 interacts with His85, and Phe79 mutants showed diminished affinity for shorter chain (C10-C16) fatty acids and weak substrate-induced high spin conversion. F79A OleTJE is least affected in substrate oxidation, whereas the F79W/Y mutants exhibit lower stability and cysteine thiolate protonation on reduction. Finally, Arg245 is crucial for binding the substrate carboxylate, and R245E/L mutations severely compromise activity and heme content, although alkene products are formed from some substrates, including stearic acid (C18:0). The results identify crucial roles for the active site amino acid trio in determining OleTJE catalytic efficiency in alkene production and in regulating protein stability, heme iron coordination, and spin state.

Capture of micrococcin biosynthetic intermediates reveals C-terminal processing as an obligatory step for in vivo maturation.

Thiopeptides, including micrococcins, are a growing family of bioactive natural products that are ribosomally synthesized and heavily modified. Here we use a refactored, modular in vivo system containing the micrococcin P1 (MP1) biosynthetic genes (TclIJKLMNPS) from Macrococcus caseolyticus str 115 in a genetically tractable Bacillus subtilis strain to parse the processing steps of this pathway. By fusing the micrococcin precursor peptide to an affinity tag and coupling it with catalytically defective enzymes, biosynthetic intermediates were easily captured for analysis. We found that two major phases of molecular maturation are separated by a key C-terminal processing step. Phase-I conversion of six Cys residues to thiazoles (TclIJN) is followed by C-terminal oxidative decarboxylation (TclP). This TclP-mediated oxidative decarboxylation is a required step for the peptide to progress to phase II. In phase II, Ser/Thr dehydration (TclKL) and peptide macrocycle formation (TclM) occurs. A C-terminal reductase, TclS, can optionally act on the substrate peptide, yielding MP1, and is shown to act late in the pathway. This comprehensive characterization of the MP1 pathway prepares the way for future engineering efforts.

Abyssicoccus albus gen. nov., sp. nov., a novel member of the family Staphylococcaceae isolated from marine sediment of the Indian Ocean.

A Gram-stain positive, aerobic, non-motile, asporogenous, coccoid shaped bacterium, designated YIM M12140(T), was isolated from a marine sediment sample collected from the Indian Ocean. Phylogenetic analysis showed that strain YIM M12140(T) forms a separate clade within the family Staphylococcaceae. Strain YIM M12140(T) shares high 16S rRNA gene sequence similarity with Macrococcus brunensis DSM 19358(T) (92.9 %). The isolate was found to grow at 0-10 % (w/v) NaCl (optimum, 2-3 %), pH 6.0-10.0 (optimum, pH 8.0) and temperature 5-40 °C (optimum, 28 °C). The polar lipids were identified as diphosphatidylglycerol, phosphatidylglycerol, one unidentified aminophospholipid and two unidentified polar lipids. The major cellular fatty acids of the strain were identified as anteiso-C15:0, -C17:0, iso-C16:0, anteiso-C19:0 and C20:0. The respiratory menaquinones were found to be MK-6 (94 %) and MK-7 (6 %). The cell wall amino acids were found to contain Lys, Ala, Glu, Gly, Asp, Ser and Thr. Whole cell sugars were identified as mannose, ribose, rhamnose, glucose, galactose and xylose. The G+C content of the genomic DNA of strain YIM M12140(T) was determined to be 42.4 mol %. Based on phenotypic, chemotaxonomic data and phylogenetic analysis, it is proposed that strain YIM M12140(T) represents a novel species of a new genus in the family Staphylococcaceae, for which the name Abyssicoccus albus gen. nov., sp. nov. is proposed. The type strain is YIM M12140(T) (= DSM 29158(T) = CCTCC AB 2014213(T)).

Correlation of the lung microbiota with metabolic profiles in bronchoalveolar lavage fluid in HIV infection.

BACKGROUND: While 16S ribosomal RNA (rRNA) sequencing has been used to characterize the lung's bacterial microbiota in human immunodeficiency virus (HIV)-infected individuals, taxonomic studies provide limited information on bacterial function and impact on the host. Metabolic profiles can provide functional information on host-microbe interactions in the lungs. We investigated the relationship between the respiratory microbiota and metabolic profiles in the bronchoalveolar lavage fluid of HIV-infected and HIV-uninfected outpatients. RESULTS: Targeted sequencing of the 16S rRNA gene was used to analyze the bacterial community structure and liquid chromatography-high-resolution mass spectrometry was used to detect features in bronchoalveolar lavage fluid. Global integration of all metabolic features with microbial species was done using sparse partial least squares regression. Thirty-nine HIV-infected subjects and 20 HIV-uninfected controls without acute respiratory symptoms were enrolled. Twelve mass-to-charge ratio (m/z) features from C18 analysis were significantly different between HIV-infected individuals and controls (false discovery rate (FDR) = 0.2); another 79 features were identified by network analysis. Further metabolite analysis demonstrated that four features were significantly overrepresented in the bronchoalveolar lavage (BAL) fluid of HIV-infected individuals compared to HIV-uninfected, including cystine, two complex carbohydrates, and 3,5-dibromo-L-tyrosine. There were 231 m/z features significantly associated with peripheral blood CD4 cell counts identified using sparse partial least squares regression (sPLS) at a variable importance on projection (VIP) threshold of 2. Twenty-five percent of these 91 m/z features were associated with various microbial species. Bacteria from families Caulobacteraceae, Staphylococcaceae, Nocardioidaceae, and genus Streptococcus were associated with the greatest number of features. Glycerophospholipid and lineolate pathways correlated with these bacteria. CONCLUSIONS: In bronchoalveolar lavage fluid, specific metabolic profiles correlated with bacterial organisms known to play a role in the pathogenesis of pneumonia in HIV-infected individuals. These findings suggest that microbial communities and their interactions with the host may have functional metabolic impact in the lung.

Identification of N(α)-acetyl-α-lysine as a probable thermolyte and its accumulation mechanism in Salinicoccus halodurans H3B36.

Salinicoccus halodurans H3B36 is a moderate halophile that was isolated from a 3.2-m-deep sediment sample in Qaidam Basin, China. Our results suggest that N(α)-acetyl-α-lysine can accumulate and act as a probable thermolyte in this strain. The accumulation mechanism and biosynthetic pathway for this rare compatible solute were also elucidated. We confirmed that the de novo synthesis pathway of N(α)-acetyl-α-lysine in this strain starts from aspartate and passes through lysine. Through RNA sequencing, we also found an 8-gene cluster (orf_1582-1589) and another gene (orf_2472) that might encode the biosynthesis of N(α)-acetyl-α-lysine in S. halodurans H3B36. Orf_192, orf_193, and orf_1259 might participate in the transportation of precursors for generating N(α)-acetyl-α-lysine under the heat stress. The transcriptome reported here also generated a global view of heat-induced changes and yielded clues for studying the regulation of N(α)-acetyl-α-lysine accumulation. Heat stress triggered a global transcriptional disturbance and generated a series of actions to adapt the strain to heat stress. Furthermore, the transcriptomic results showed that the regulon of RpoN (orf_2534) may be critical to conferring heat stress tolerance and survival to S. halodurans.

Comparative Proteomic Insights into the Lactate Responses of Halophilic Salinicoccus roseus W12.

Extremophiles use adaptive mechanisms to survive in extreme environments, which is of great importance for several biotechnological applications. A halophilic strain, Salinicoccus roseus W12, was isolated from salt lake in Inner Mongolia, China in this study. The ability of the strain to survive under high sodium conditions (including 20% sodium lactate or 25% sodium chloride, [w/v]) made it an ideal host to screen for key factors related to sodium lactate resistance. The proteomic responses to lactate were studied using W12 cells cultivated with or without lactate stress. A total of 1,656 protein spots in sodium lactate-treated culture and 1,843 spots in NaCl-treated culture were detected by 2-dimensional gel electrophoresis, and 32 of 120 significantly altered protein spots (fold change > 2, p < 0.05) were identified by matrix-assisted laser-desorption ionization time-of-flight mass spectrometry. Among 21 successfully identified spots, 19 proteins were upregulated and 2 were downregulated. The identified proteins are mainly involved in metabolism, cellular processes and signaling, and information storage and processing. Transcription studies confirmed that most of the encoding genes were upregulated after the cells were exposed to lactate in 10 min. Cross-protecting and energy metabolism-related proteins played an important role in lactate tolerance for S. roseus W12.

Green Synthesis of Silver Nano-particles by Macrococcus bovicus and Its Immobilization onto Montmorillonite Clay for Antimicrobial Functionality.

Macrococcus bovicus was locally isolated from soil and used in the green synthesis of nano-scaling silver (NSAg). It was immobilized on a sodic-montmorillonite clay (MMT1) and cetyltrimethylammonium bromide-modified montmorillonite (MMT2) which was also calcined at 300 °C (MMT3). The NSAg clays were characterized by X-ray fluorescence, Fourier transform infrared spectra, X-ray diffractometry, surface area measurement, UV-Vis spectrometry, scanning electron microscope, transmission electron microscope and thermogravimetric analysis. NSAg was confirmed to be included in the interparticular cavities of the clay sheets and its mechanical stability was evidenced. The antimicrobial activity of the NSAg-modified clays was investigated against Staphylococcus aureus, Escherichia coli and Candida albicans using the cup plate and the plate count techniques. The antimicrobial activity of the NSAg clays was confirmed and attributed to the caging of NSAg in MMT cavities. MMT3 was found to inhibit the microbial growth to as high as 65 % as observed from the plate count method. Graphical Abstract Scheme of the biosynthesis of nano-scaling Ag and its immobilization and antimicrobial application.

Radiation-resistant Macrococcus caseolyticus (A) isolated from radiation-processed semidried prawns.

A radiation-resistant bacterial isolate from gamma-radiation-processed (5 kGy) semidried prawns was identified as a new strain of Macrococcus caseolyticus and was designated as M. caseolyticus (A) on the basis of morphological and biochemical characterization and 16S rRNA sequencing. DNA-DNA hybridization studies with M. caseolyticus DSM 20597(T) further confirmed the isolate as M. caseolyticus. Major fatty acids present in M. caseolyticus (A) were C14:0, C16:1ω11c, and C18:1ω9c, whereas C15:0anteiso, C16:0iso, and C18:0iso were absent. The closest match for the isolate, as per fatty acid methyl ester analysis, was M. caseolyticus DSM 20597(T). However, the similarity index was significantly low (0.112), which indicates that the isolate could be a new strain of M. caseolyticus. The decimal reduction dose (D10) for M. caseolyticus (A), M. caseolyticus JCSC5402, and Staphylococcus aureus MTCC96 was 1.18, 0.607, and 0.19 kGy, respectively. This is the first report on radiation resistance of M. caseolyticus. Macrococcus caseolyticus (A) is more resistant to gamma and UV radiation stress than are M. caseolyticus JCSC5402 and S. aureus MTCC96; however, it is sensitive to heat as well as desiccation stress.

Salinicoccus halitifaciens sp. nov., a novel bacterium participating in halite formation.

Strain JC90(T) was isolated from a soda lake in Lonar, India. Strain JC90(T) maintains its external pH to 8.5 and participates in halite formation. Based on 16S rRNA gene sequence similarity studies, strain JC90(T) was found to belong to the genus Salinicoccus and is most closely related to "Salinicoccus kekensis" K164(T) (99.3 %), Salinicoccus alkaliphilus T8(T) (98.4 %) and other members of the genus Salinicoccus (<96.5 %). However Strain JC90(T) is <36 % related (based on DNA-DNA hybridization) with the type strains of "S. kekensis" K164(T) and S. alkaliphilus T8(T). The DNA G+C content of strain JC90(T) was determined to be 46 mol %. The cell-wall amino acids were identified as lysine and glycine. Polar lipids were found to include diphosphatidylglycerol, phosphatidylglycerol, phosphatidyl ethanolamine, an unidentified glycolipid and unidentified lipids (L1,2). Major hopanoids of strain JC90(T) were determined to be bacterial hopane derivatives (BHD1,2), diplopterol, diploptene and two unidentified hopanoids (UH1,2). The predominant isoprenoid quinone was identified as menaquinone (MK-6). Anteiso-C15:0 was determined to be the predominant fatty acid and significant proportions of iso-C14:0, C14:0, iso-C15:0, C16:0, iso-C16:0, iso-C17:0, anteiso-C17:0 and C18:02OH were also detected. The results of physiological and biochemical tests support the molecular evidence and allowed a clear phenotypic differentiation of strain JC90(T) from all other members of the genus Salinicoccus. Strain JC90(T) is therefore considered to represent a novel species, for which the name Salinicoccus halitifaciens sp. nov. is proposed. The type strain is JC90(T) (=KCTC 13894(T) =DSM 25286(T)).

Technological and safety characteristics of Staphylococcaceae isolated from Spanish traditional dry-cured sausages.

The aim of this study was to determine the technological properties (nitrate reductase, proteolytic and lipolytic activities; and the ability to grow at the temperature and pH values of fermenting sausage, and at high NaCl concentrations) and safety characteristics (amino acid decarboxylase and enterotoxigenic activities) of 38 strains of Staphylococcaceae (11 of Staphylococcus epidermidis, 15 of Staphylococcus equorum, 5 of Staphylococcus pasteuri and 7 of Staphylococcus saprophyticus) isolated from Androlla and Botillo, two Spanish traditional sausages, in order to evaluate their suitability as potential starter cultures in the manufacture of these sausages. Most strains were able to grow at 10 °C, in the presence of 10% and 15% NaCl and at pH values of 5.5 and 5.0, except for S. equorum strains, growth of which was reduced at these pH values. The proteolytic activity assessed by the agar plate method showed that 89.5% and 52.6% of the strains were able to hydrolyze sarcoplasmic and myofibrillar proteins, respectively. These results were not confirmed by electrophoretic assays as only 47.2% of the strains changed the SDS-PAGE profile of actin, myosin and/or sarcoplasmic protein extracts. The assessment of the lipolytic activity by titration showed that only 21.0% of the strains can hydrolyze pork fat to any extent; whereas the profiles of the freed fatty acids were different in the different strains. Most of the strains showed decarboxylase activity against histidine, lysine, ornithine and tyrosine, but the quantities of biogenic amines produced were in most cases <25 ppm and <5 ppm for putrescine and cadaverine, respectively. Only four strains (10.5%), of S. epidermidis, produced enterotoxin C.

Inference of the transcriptional regulatory network in Staphylococcus aureus by integration of experimental and genomics-based evidence.

Transcriptional regulatory networks are fine-tuned systems that help microorganisms respond to changes in the environment and cell physiological state. We applied the comparative genomics approach implemented in the RegPredict Web server combined with SEED subsystem analysis and available information on known regulatory interactions for regulatory network reconstruction for the human pathogen Staphylococcus aureus and six related species from the family Staphylococcaceae. The resulting reference set of 46 transcription factor regulons contains more than 1,900 binding sites and 2,800 target genes involved in the central metabolism of carbohydrates, amino acids, and fatty acids; respiration; the stress response; metal homeostasis; drug and metal resistance; and virulence. The inferred regulatory network in S. aureus includes ∼320 regulatory interactions between 46 transcription factors and ∼550 candidate target genes comprising 20% of its genome. We predicted ∼170 novel interactions and 24 novel regulons for the control of the central metabolic pathways in S. aureus. The reconstructed regulons are largely variable in the Staphylococcaceae: only 20% of S. aureus regulatory interactions are conserved across all studied genomes. We used a large-scale gene expression data set for S. aureus to assess relationships between the inferred regulons and gene expression patterns. The predicted reference set of regulons is captured within the Staphylococcus collection in the RegPrecise database (http://regprecise.lbl.gov).

Jeotgalicoccus huakuii sp. nov., a halotolerant bacterium isolated from seaside soil.

A Gram-stain-positive, oxidase- and catalase-positive, non-motile, non-spore-forming, halotolerant, coccoid bacterium, designated strain NY-2(T), was isolated from a seaside soil sample from Shandong Province, China. Strain NY-2(T) was able to grow in the presence of 0-23 % (w/v) NaCl and at pH 4.5-10.0 and 5-42 degrees C; optimum growth was observed with 3-8 % (w/v) total salts and at pH 6.5-8.0 and 28-37 degrees C. Chemotaxonomic analyses, including fatty acid profiles, menaquinones and polar lipids, supported the affiliation of strain NY-2(T) to the genus Jeotgalicoccus. The predominant menaquinone of strain NY-2(T) was menaquinone 7 (MK-7) (100 %) and the major cellular fatty acids were iso-C(15 : 0) (49.0 %) and anteiso-C(15 : 0) (19.6 %). Cellular polar lipids were phosphatidylglycerol, diphosphatidylglycerol and several unidentified phospholipids. The DNA G+C content of strain NY-2(T) was 36.8 mol%. Based on 16S rRNA gene sequence analysis, strain NY-2(T) formed a coherent cluster with Jeotgalicoccus marinus JSM 076033(T), Jeotgalicoccus halotolerans YKJ-101(T) and Jeotgalicoccus psychrophilus YKJ-115(T). Phylogenetic analysis, DNA-DNA relatedness data, phenotypic characteristics and chemotaxonomic data indicated that strain NY-2(T) (=CCTCC AB 208288(T) =JCM 15687(T)) should be classified as the type strain of a novel species of the genus Jeotgalicoccus, for which the name Jeotgalicoccus huakuii sp. nov. is proposed.

Salinicoccus carnicancri sp. nov., a halophilic bacterium isolated from a Korean fermented seafood.

A novel, moderately halophilic bacterium belonging to the genus Salinicoccus was isolated from crabs preserved in soy sauce: a traditional Korean fermented seafood. Colonies of strain Crm(T) were ivory and the cells were non-motile, Gram-positive cocci. The organism was non-sporulating, catalase-positive and oxidase-negative. The major fatty acids of strain Crm(T) were iso-C(15 : 0) (22.0 %), anteiso-C(15 : 0) (40.6 %) and anteiso-C(17 : 0) (12.1 %). The cell wall peptidoglycan contained lysine and glycine, and the major isoprenoid quinone was MK-6. The polar lipids were phosphatidylglycerol, diphosphatidylglycerol and an unidentified glycolipid. The genomic DNA G+C content was 47.8 mol%. Strain Crm(T) was closely related to the type strain of Salinicoccus halodurans, with which it shared 96.9 % 16S rRNA gene sequence similarity. The DNA-DNA hybridization value between strains Crm(T) and S. halodurans DSM 19336(T) was 7.6 %. Based on phenotypic, genetic and phylogenetic data, strain Crm(T) should be classified as a novel species within the genus Salinicoccus , for which the name Salinicoccus carnicancri sp. nov. is proposed. The type strain is Crm(T) (=KCTC 13301(T) =JCM 15796(T)).

Antibiotic resistance in lactic acid bacteria and Micrococcaceae/Staphylococcaceae isolates from artisanal raw milk cheeses, and potential implications on cheese making.

Antibiotic susceptibility against 19 antimicrobial agents was evaluated in isolates of the genera Lactococcus (46 isolates), Leuconostoc (22), Lactobacillus (19), Staphylococcus (8), Enterococcus (7), and Microccoccus/Kocuria (5) obtained from the predominant microflora of nonrecent and recent types of artisanal raw cow's milk cheeses. Beta-lactams showed broad activity against all genera, although leuconostocs and lactobacilli were highly resistant to oxacillin (80% to 95.5%). Resistance to aminoglycosides was frequent for lactococci and enterococci (particularly for streptomycin), whereas lower rates of resistance were detected for lactobacilli and leuconostocs. Technologically interesting traits for the food industry were distributed among isolates that showed different degrees of resistance to common antibiotics. However, isolates showing resistance to less than 2 antibiotics were mainly those with properties of greatest technological interest (acidifying activity, proteolytic/lipolytic activities, or diacetyl production).