Major Metabolic End Products Research Articles

Description of Veillonella absiana sp. nov., isolated from pig faeces.

Two Gram-stain-negative cocci anaerobes were isolated from pig faeces and designated as strains YH-vei2232T and YH-vei2233. Phylogenetic analysis using 16S rRNA gene sequences revealed that the isolates were most closely related to Veillonella rogosae KCTC 5967T, with 97.0% similarity. Analysis of housekeeping gene sequences (rpoB) revealed the strain formed a sub-cluster within the genus Veillonella. The average nucleotide identities between the two isolates and the most closely related strains within genus Veillonella were <75.0%. The major fatty acids were Summed Feature 8 and C16 : 1 ω9c. The cell wall peptidoglycan contained meso-diaminopimelic acid. The major metabolic end products of isolates were propionic and acetic acids. The genomic DNA G+C contents of both strains were 40.1 mol%. The chemotaxonomic, phenotypic and phylogenetic properties of YH-vei2232T (=KCTC 25748T=NBRC 116427T) and YH-vei2233 (=KCTC 25749=NBRC 116428) suggest that they represent a novel species of the genus Veillonella, for which the name Veillonella absiana sp. nov. is proposed.

Segatella asaccharophila sp. nov., an anaerobic pectinophile isolated from a two-phase methane fermentation system.

An obligately anaerobic, Gram-stain-negative, non-spore-forming and non-motile rod (strain LPYR103-PreT) was isolated from a two-phase methane fermentation system. Using 16S rRNA gene sequence-based phylogenetic analysis, strain LPYR103-PreT was classified in the genus Segatella. The 16S rRNA gene sequence similarity, average nucleotide identity and digital DNA-DNA hybridization between strain LPYR103-PreT and its phylogenetically nearest species - Segatella cerevisiae JCM 30867T - were 94.4, 77.9 and 23.4%, respectively. The genome size of strain LPYR103-PreT was 3 326 733 bp, and its genomic DNA G+C content was 44.05%. The most abundant cellular fatty acid was anteiso-C15 : 0. The growth of strain LPYR103-PreT was stimulated by the addition of pectin, d-galacturonate and d-glucuronate; in contrast, the strain exhibited poor growth in the presence of common sugars, such as glucose. Therefore, strain LPYR103-PreT was classified as a pectinophile - a bacterium that shows a preference for pectin and a few related compounds as substrates. Glucose is degraded by type strains of 12 species belonging to the genus Segatella; thus, strain LPYR103-PreT is the first described pectinophile belonging to this genus. Strain LPYR103-PreT produced succinate and acetate as its major metabolic end products. Based on the differences in the phylogenetic, genomic, physiological and chemotaxonomic characteristics of strain LPYR103-PreT and related species, the name Segatella asaccharophila sp. nov. is proposed to accommodate strain LPYR103-PreT (= NRIC 0997T = JCM 37351T=DSM 118531T = KCTC 25923T).

Integrated transcriptomic and metabolomic analysis reveals the potential mechanisms underlying indium-induced inhibition of root elongation in wheat plants

Soil contamination by indium, an emerging contaminant from electronics, has a negative impact on crop growth. Inhibition of root growth serves as a valuable biomarker for predicting indium phytotoxicity. Therefore, elucidating the molecular mechanisms underlying indium-induced root damage is essential for developing strategies to mitigate its harmful effects. Our transcriptomic findings revealed that indium affects the expression of numerous genes related to cell wall composition and metabolism in wheat roots. Morphological and compositional analysis revealed that indium induced a 2.9-fold thickening and a 17.5 % increase in the content of cell walls in wheat roots. Untargeted metabolomics indicated a substantial upregulation of the phenylpropanoid biosynthesis pathway. As the major end product of phenylpropanoid metabolism, lignin significantly accumulated in root cell walls after indium exposure. Together with increased lignin precursors, enhanced activity of lignin biosynthesis-related enzymes was observed. Moreover, analysis of the monomeric content and composition of lignin revealed a significant enrichment of p-hydroxyphenyl (H) and syringyl (S) units in root cell walls under indium stress. The present study contributes to the existing knowledge of indium toxicity. It provides valuable insights for developing sustainable solutions to address the challenges posed by electronic waste and indium contamination on agroecosystems.

Butyrate Differentiates Permissiveness to Clostridioides difficile Infection and Influences Growth of Diverse C. difficile Isolates.

A disrupted "dysbiotic" gut microbiome engenders susceptibility to the diarrheal pathogen Clostridioides difficile by impacting the metabolic milieu of the gut. Diet, in particular the microbiota-accessible carbohydrates (MACs) found in dietary fiber, is one of the most powerful ways to affect the composition and metabolic output of the gut microbiome. As such, diet is a powerful tool for understanding the biology of C. difficile and for developing alternative approaches for coping with this pathogen. One prominent class of metabolites produced by the gut microbiome is short-chain fatty acids (SCFAs), the major metabolic end products of MAC metabolism. SCFAs are known to decrease the fitness of C. difficile in vitro, and high intestinal SCFA concentrations are associated with reduced fitness of C. difficile in animal models of C. difficile infection (CDI). Here, we use controlled dietary conditions (8 diets that differ only by MAC composition) to show that C. difficile fitness is most consistently impacted by butyrate, rather than the other two prominent SCFAs (acetate and propionate), during murine model CDI. We similarly show that butyrate concentrations are lower in fecal samples from humans with CDI than in those from healthy controls. Finally, we demonstrate that butyrate impacts growth in diverse C. difficile isolates. These findings provide a foundation for future work which will dissect how butyrate directly impacts C. difficile fitness and will lead to the development of diverse approaches distinct from antibiotics or fecal transplant, such as dietary interventions, for mitigating CDI in at-risk human populations. IMPORTANCE Clostridioides difficile is a leading cause of infectious diarrhea in humans, and it imposes a tremendous burden on the health care system. Current treatments for C. difficile infection (CDI) include antibiotics and fecal microbiota transplant, which contribute to recurrent CDIs and face major regulatory hurdles, respectively. Therefore, there is an ongoing need to develop new ways to cope with CDI. Notably, a disrupted "dysbiotic" gut microbiota is the primary risk factor for CDI, but we incompletely understand how a healthy microbiota resists CDI. Here, we show that a specific molecule produced by the gut microbiota, butyrate, is negatively associated with C. difficile burdens in humans and in a mouse model of CDI and that butyrate impedes the growth of diverse C. difficile strains in pure culture. These findings help to build a foundation for designing alternative, possibly diet-based, strategies for mitigating CDI in humans.

Access to highly specialized growth substrates and production of epithelial immunomodulatory metabolites determine survival of Haemophilus influenzae in human airway epithelial cells.

Haemophilus influenzae (Hi) infections are associated with recurring acute exacerbations of chronic respiratory diseases in children and adults including otitis media, pneumonia, chronic obstructive pulmonary disease and asthma. Here, we show that persistence and recurrence of Hi infections are closely linked to Hi metabolic properties, where preferred growth substrates are aligned to the metabolome of human airway epithelial surfaces and include lactate, pentoses, and nucleosides, but not glucose that is typically used for studies of Hi growth in vitro. Enzymatic and physiological investigations revealed that utilization of lactate, the preferred Hi carbon source, required the LldD L-lactate dehydrogenase (conservation: 98.8% of strains), but not the two redox-balancing D-lactate dehydrogenases Dld and LdhA. Utilization of preferred substrates was directly linked to Hi infection and persistence. When unable to utilize L-lactate or forced to rely on salvaged guanine, Hi showed reduced extra- and intra-cellular persistence in a murine model of lung infection and in primary normal human nasal epithelia, with up to 3000-fold attenuation observed in competitive infections. In contrast, D-lactate dehydrogenase mutants only showed a very slight reduction compared to the wild-type strain. Interestingly, acetate, the major Hi metabolic end-product, had anti-inflammatory effects on cultured human tissue cells in the presence of live but not heat-killed Hi, suggesting that metabolic endproducts also influence HI-host interactions. Our work provides significant new insights into the critical role of metabolism for Hi persistence in contact with host cells and reveals for the first time the immunomodulatory potential of Hi metabolites.

Phocaeicola faecicola sp. nov., isolated from porcine faeces.

An obligately anaerobic, non-motile, Gram-negative and rod-shaped strain (AGMB03916T) was isolated from faeces of a 2-week-old piglet raised at the National Institute of Animal Science in Wanju, Republic of Korea. Growth of strain AGMB03916T occurred at 30-45 °C (optimum, 37 °C), at pH 6-9 (optimum, pH 8) and in the presence of 0.5-1.0 % (w/v) NaCl. Based on the results of 16S rRNA gene sequence analyses, strain AGMB03916T was closely related to two validly published species of the genus Phocaeicola, Phocaeicola plebeius and Phocaeicola coprocola. The 16S rRNA gene sequence similarity of strain AGMB03916T compared to P. plebeius M12T (=KCTC 5793T) and P. coprocola M16T (=KCTC 5443T) were 96.3 and 95.0 %, respectively. The genomic DNA G+C content of strain AGMB03916T was 46.4 mol%. The average nucleotide identity values between strain AGMB03916T and the reference strains were 74.9-78.5 %. Cells were able to utilize d-glucose, lactose, sucrose, maltose, salicin, aesculin hydrolysis, cellobiose and raffinose. The major end product of metabolism was acetate. The major cellular fatty acids (>10 %) of the isolate were iso-C15 : 0, anteiso-C15 : 0, C16 : 0, C16 : 0 3-OH and summed feature 11 (iso-C17 : 0 3-OH and/or C18 : 2 DMA). On the basis of the genotypic, biochemical, chemotaxonomic, phenotypic and phylogenetic data, strain AGMB03916T represents a novel species of the genus Phocaeicola, for which the name Phocaeicola faecicola sp. nov. is proposed. The type strain is AGMB03916T (=KCTC 25014T=GDMCC 1.2574T).

Description of Anaerostipes faecalis sp. nov., a new segmented filamentous bacterium isolated from swine faeces.

A novel, strictly anaerobic, gram-negative, segmented filamentous bacterium strain AGMB03513T, was isolated from the faeces of a 5-month-old pig. Phylogenetic analysis based on the 16S rRNA gene indicated that the isolate was a member of the family Lachnospiraceae, and the closest strain was Anaerostipes butyraticus. Strain AGMB03513T formed a lineage within the genus Anaerostipes and was closely related to A. butyraticus DSM 22094T (= KCTC 15125T, 95.8%), Anaerostipes hadrus DSM 3319T (= KCTC 15606T, 95.5%), Anaerostipes caccae DSM 14662T (= KCTC 15019T, 94.0%), and Anaerostipes rhamnosivorans DSM 26241T (= KCTC 15316T, 93.4%). Strain AGMB03513T grew at temperatures between 30 and 45°C, within a pH range of 7.0-9.0, and in medium containing up to 1.5% NaCl. Cells were found to utilise D-glucose, D-mannitol, D-lactose, D-saccharose, D-maltose, D-xylose, L-arabinose, D-mannose, and D-sorbitol, and acetate was identified as the major end product of metabolism. The major components of the cellular fatty acids were C12:0, C16:0, and C18:0. In addition, the bacterium contained meso-diaminopimelic acid in the cell wall. According to the comparative analysis of the whole genome sequence, the DNA G + C content of strain AGMB03513 was 37.0mol%. In addition, Average nucleotide identity (ANI), average amino acid identity (AAI), and digital DNA-DNA hybridisation (dDDH) values were obtained in comparisons of strain AGMB03513T with reference strains of species in the genus Anaerostipes. ANI values were found to be between 71.0 and 75.7%, AAI values between 66.6 and 73.2%, and dDDH values between 19.5 and 21.4%. All the data were below the threshold range for species determination. Based on phenotypic, phylogenetic, biochemical, chemotaxonomic, and genomic characteristics, we considered it reasonable to assign a novel species status to strain AGMB03513T, for which we propose the name Anaerostipes faecalis sp. nov. The type strain is AGMB03513T (= KCTC 25020T = NBRC 114896T).

Peptoniphilus nemausensis sp. nov. A new Gram-positive anaerobic coccus isolated from human clinical samples, an emendated description of the genus Peptoniphilus and an evaluation of the taxonomic status of Peptoniphilus species with not validly published names

A Gram-positive, anaerobic coccus isolated from a human surgical site infection was previously shown to belong to an unknown species of the genus Peptoniphilus initially proposed as ‘Peptoniphilus nemausus’ sp. nov., based on both 16S rRNA gene sequence identity of 97.9% with the most closely related species Peptoniphilus coxii and an individualized phylogenetic branching within the genus Peptoniphilus. A polyphasic characterization of the novel species is proposed herein. Whole genome sequence analysis showed an average nucleotide identity value of 84.75% and digital DNA-DNA hybridization value of 28.9% against P. coxii type strain. The strain displayed unique features among members of the genus Peptoniphilus, as it was able to hydrolyze aesculin, and produced acetate as the major metabolic end-product without associated production of butyrate. Growth was observed under microaerophilic conditions. From all these data, the isolate is confirmed as belonging to a new Peptoniphilus species, for which the name Peptoniphilus nemausensis sp. nov. is proposed. The type strain is 1804121828T (=LMG 31466T = CECT 9935T). A database survey using a highly polymorphic partial sequence of the 16S rRNA gene of P. nemausensis revealed P. nemausensis to be a particularly rare skin-associated species in humans. An emendated description of the Peptoniphilus genus is proposed based on a review of the characteristics of the 12 new species with validly published names since the genus description in 2001 and of P. nemausensis. Finally, the relationships between members of the genus Peptoniphilus were explored based on whole genome sequence analysis in order to clarify the taxonomic status of not yet validly published species showing that three pairs of species should be considered as synonyms: Peptoniphilus timonensis and ‘Peptoniphilus phoceensis’, Peptoniphilus lacydonensis and ‘Peptoniphilus rhinitidis’, Peptoniphilus tyrrelliae and Peptoniphilus senegalensis.

Peptoniphilus faecalis sp. nov., isolated from swine faeces.

An obligately anaerobic, Gram-positive, non-motile, coccus-shaped bacterial strain designated AGMB00490T was isolated from swine faeces. 16S rRNA gene sequence-based phylogenetic analysis indicated that the isolate belongs to the genus Peptoniphilus and that the most closely related species is Peptoniphilus gorbachii WAL 10418T (=KCTC 5947T, 97.22 % 16S rRNA gene sequence similarity). Whole genome sequence analysis determined that the DNA G+C content of strain AGMB00490T was 31.2 mol% and moreover that the genome size and numbers of tRNA and rRNA genes were 2 129 517 bp, 34 and 10, respectively. Strain AGMB00490T was negative for oxidase and urease; positive for catalase, indole production, arginine arylamidase, leucine arylamidase, tyrosine arylamidase and histidine arylamidase; and weakly positive for phenylalanine arylamidase and glycine arylamidase. The major cellular fatty acids (>10 %) of the isolate were determined to be C16 : 0 and C18 : 1 ω9c. Strain AGMB00490T produced acetic acid as a major end product of metabolism. Accordingly, phylogenetic, physiologic and chemotaxonomic analyses revealed that strain AGMB00490T represents a novel species for which the name Peptoniphilus faecalis sp. nov. is proposed. The type strain is AGMB00490T (=KCTC 15944T=NBRC 114159T).

Prebiotic Efficacy of Oligosaccharides Derived from the Immunomodulatory Glycoprotein (Agp) Purified from Ajowan (Trachyspermum ammiL.)

Ajowan (Trachyspermum ammi L.) spice has proven medicinal and nutraceutical properties. A purified glycoprotein (Agp; 30.7 kDa) with established immunomodulatory properties from ajowan was hydrolyzed with trifluoroacetic acid. The resultant glycan was analyzed for prebiotic activity. The probiotic microbes, L. delbrueckii (NCDC 10), L. acidophilus (NCDC 11) and L. casei (NCDC 17) showed potential utilization of the Agp hydrolysates. α-Arabinofuranosidase and β-galactopyranosidase are the major enzymes involved in the metabolism of oligosaccharides, and the microbes produced acetate as the major metabolic end product. Two major oligosaccharides from Agp hydrolysate were purified for structural characterization (ESI-MS, GLC, and NMR spectroscopy). Agp-oligosaccharide I is composed of three units of galactose and one unit of galacturonic acid, whereas Agp-oligosaccharide II is composed of two arabinose and two galactose residues of which one of the arabinoses is acetylated. This study may have future implications for the design of novel immune boosters and prebiotic food products.

A Sulfoglycolytic Entner-Doudoroff Pathway in Rhizobium leguminosarum bv. trifolii SRDI565.

Rhizobia are nitrogen-fixing bacteria that engage in symbiotic relationships with plant hosts but can also persist as free-living bacteria in the soil and rhizosphere. Here, we show that free-living Rhizobium leguminosarum SRDI565 can grow on the sulfosugar sulfoquinovose (SQ) or the related glycoside SQ-glycerol using a sulfoglycolytic Entner-Doudoroff (sulfo-ED) pathway, resulting in production of sulfolactate (SL) as the major metabolic end product. Comparative proteomics supports the involvement of a sulfo-ED operon encoding an ABC transporter, sulfo-ED enzymes, and an SL exporter. Consistent with an oligotrophic lifestyle, proteomics data revealed little change in expression of the sulfo-ED proteins during growth on SQ versus mannitol, a result confirmed through biochemical assay of sulfoquinovosidase activity in cell lysates. Metabolomics analysis showed that growth on SQ involves gluconeogenesis to satisfy metabolic requirements for glucose-6-phosphate and fructose-6-phosphate. Metabolomics analysis also revealed the unexpected production of small amounts of sulfofructose and 2,3-dihydroxypropanesulfonate, which are proposed to arise from promiscuous activities of the glycolytic enzyme phosphoglucose isomerase and a nonspecific aldehyde reductase, respectively. The discovery of a rhizobium isolate with the ability to degrade SQ builds our knowledge of how these important symbiotic bacteria persist within soil.IMPORTANCE Sulfonate sulfur is a major form of organic sulfur in soils but requires biomineralization before it can be utilized by plants. Very little is known about the biochemical processes used to mobilize sulfonate sulfur. We show that a rhizobial isolate from soil, Rhizobium leguminosarum SRDI565, possesses the ability to degrade the abundant phototroph-derived carbohydrate sulfonate SQ through a sulfoglycolytic Entner-Doudoroff pathway. Proteomics and metabolomics demonstrated the utilization of this pathway during growth on SQ and provided evidence for gluconeogenesis. Unexpectedly, off-cycle sulfoglycolytic species were also detected, pointing to the complexity of metabolic processes within cells under conditions of sulfoglycolysis. Thus, rhizobial metabolism of the abundant sulfosugar SQ may contribute to persistence of the bacteria in the soil and to mobilization of sulfur in the pedosphere.

Monitoring of cell culture conditions and early prediction of the quality of an osteogenic cell-based medicinal product

Background & Aim Novadip Biosciences developed a technology platform for tissue engineering and manufacturing of osteogenic cell-based pharmaceuticals. During the production process, a flexible medium refreshment strategy is essential to ensure the quality of the final product and the batch-to-batch consistency. The cellular metabolic activity of several dozens of batches was studied. A panel of metabolite concentrations was measured in cell culture supernatant from AMSCs isolation to formulation. Normalization of the data was carried out by calculating absolute and specific production/consumption rates. Methods, Results & Conclusion We defined optimal culture condition for AMSCs. In our setting, the level of metabolites stays in standard production/consumption rates and never reaches critically high values. A continuous consumption of glucose and glutamine is observed, and medium refreshment strategy allows to keep those nutrients at adequate level. A significant production of metabolites is observed throughout the process. The lactate level stays relatively low during the proliferation phase then raises after induction of osteogenic differentiation. It is showed that lactate that is a major end-product of glucose metabolism, reflects the transition between proliferating cells and osteogenic-oriented cells. Ammonia level tends to increase after osteo-induction but does not reach inhibitory concentration. Similarly, LDH level increases after osteo-induction. Glutamate consumption increases while pyruvate level decreases progressively during proliferation phase then go down to a low and stable level after osteoinduction. Phosphate concentration raises drastically during differentiation while the calcium level drops. Thus, the osteogenic differentiation leads to a metabolic shift in glucose, lactate, glutamate, LDH and pyruvate concentrations. Production and consumptions rates could be used to implement a routine-based analysis of waste media. Monitoring the rate of nutrients consumption and metabolites production represents a non-invasive and cost-effective strategy to monitor the process performance and predict quality issues with a high level of confidence.

Heart failure developed after myocardial infarction does not affect gut microbiota composition in the rat.

There is a body of evidence that supports the notion that gut dysbiosis plays a role in the pathogenesis of cardiovascular diseases. Decreased cardiac function can reduce intestinal perfusion, resulting in morphological alterations, which may contribute to changes in the gut microbiota composition in patients with heart failure (HF). In this regard, a germane question is whether changes in gut microbiota composition are a cause or consequence of the cardiovascular disturbance. We tested the hypothesis that the development of HF, after myocardial infarction, would cause gut dysbiosis. Fecal samples were collected before and 6 wk after myocardial infarction or sham surgery. Gut microbiota were characterized by sequencing the bacterial 16S ribosomal DNA. The composition of bacterial communities in the fecal samples was evaluated by calculating three major ecological parameters: 1) the Chao 1 richness, 2) the Pielou evenness, and 3) the Shannon index. None of these indices was changed in either sham or HF rats. The Firmicutes/Bacteroidetes ratio was not altered in HF rats. The number of species in each phylum was also not different between sham and HF rats. β-Diversity analysis showed that the composition of gut microbiota was not changed with the development of HF. Bacterial genera were grouped according to their major metabolic end-products (acetate, butyrate, and lactate), but no differences were observed in HF rats. Therefore, we conclude that HF induced by myocardial infarction does not affect gut microbiota composition, at least in rats, indicating that the dysbiosis observed in patients with HF may precede cardiovascular disturbance.NEW & NOTEWORTHY Our study demonstrated that, following myocardial infarction in rats, heart failure (HF) development did not affect the intestinal microbiota despite distinct differences reported in the gut microbiota of humans with HF. Our finding is consistent with the notion that dysbiosis observed in patients with HF may precede cardiovascular dysfunction and therefore offers potential for early diagnosis and treatment.

Prevotella cerevisiae sp. nov., beer-spoilage obligate anaerobic bacteria isolated from brewery wastewater.

Two obligate anaerobic, Gram-stain-negative, non-spore-forming bacilli (strains SBC 8034T and SBC 8065) were isolated from brewery wastewater. Cells of the two strains were rod-shaped and 0.8×2-5 µm in size. Strains SBC 8034T and SBC 8065 did not grow on Columbia agar or tryptic soy agar II with 5 % sheep blood, brain-heart infusion agar or chocolate agar, but did grow on peptone-yeast-glucose agar and de Man, Rogosa and Sharpe agar using beer instead of water. The organisms produced acetic acid and succinic acid as the major metabolic end-products. Phylogenetic analysis based on 16S rRNA gene sequences revealed that both strains are clearly distinct from all recognized species within the genus Prevotella, but belong to the same species (DDH=85 %). Based on 16S rRNA and hsp60 gene sequencing, along with phenotypic, chemical and biochemical properties, strains SBC 8034T and SBC 8065 were considered to represent a novel species within the genus Prevotella, for which the name Prevotellacerevisiae sp. nov. is proposed. Strain SBC 8034T (=DSM 100619T=JCM 30867T) is the type strain of the proposed novel species.

Characterisation of Lactococcus lactis isolates from herbs, fruits and vegetables for use as biopreservatives against Listeria monocytogenes in cheese

The control of Listeria monocytogenes in soft cheese manufacture remains a challenge due to its widespread occurrence, ability to grow at refrigerated temperatures and salt resistance. The application of antimicrobial compounds produced by lactic acid bacteria (LAB) as natural preservatives for improving cheese quality and safety has attracted significant research interest. The aim of this study was to select non-dairy origin LAB strains with anti-Listeria activity and to characterise their industrially relevant properties for potential roles as biopreservative adjunct cultures for use in cheese production. A total of 897 LAB isolated from herbs, fruits and vegetables were screened for their anti-Listeria activity using an MRS agar overlay assay. Fourteen isolates with strong activity were identified by 16S rRNA gene sequencing as Lactococcus lactis and all contained the nisin gene cluster. Of these fourteen isolates, four strains also showed detectable anti-Listeria activity in a milk medium. These wild Lc. lactis strains possessed slow lactose utilisation and acidification capacity in milk despite exhibiting high growth rates. This was likely due to the adoption of the mixed acid fermentation pathway since ethanol was a major metabolic end product. Analysis of volatiles in milk and laboratory scale cheese fermented with wild and industrial Lc. lactis strain combination using headspace solid phase microextraction (SPME) GC-MS identified up to a 300–400 fold increase in the production of 3-methylbutanal, an important cheese odorant with a malty flavour. Whole genome sequencing of four wild strains provided some genetic insights into their phenotypes. The wild Lc. lactis lack the lactose utilisation operon (lacABCDFEG) commonly found in industrial strains, but contain genes encoding a putative β-galactosidase and a previously characterised alternative lactose assimilation pathway with slow lactose uptake. Wild strains also contain genes encoding α-keto-acid decarboxylase (kdcA) required for 3-methylbutanal production. These results provide insight into technological properties of wild non-dairy Lc. lactis strains which could be potentially used as biopreservative adjunct cultures to control the growth of L. monocytogenes in cheese.

Caecibacterium sporoformans gen. nov., sp. nov., an anaerobic, butyrate-producing, spore-forming bacterium isolated from chicken caecum.

Strains of a Gram-stain-negative, rod-shaped and immotile bacterium were isolated from broiler chicken caecal content. The isolates required strict anaerobic conditions for growth, formed spores, were catalase-positive and oxidase-negative. They produced butyrate as the major metabolic end product in reinforced clostridial medium broth. The genomic DNA G+C content of the isolated strains was 32.5-34.6 mol%. The major cellular fatty acids were C16 : 0 FAME, C14 : 0 FAME, C19 : 0CYC 9,10DMA and C16 : 0DMA. The fatty acid composition of the cell wall showed no similarity to any strain in the midi database. 16S rRNA gene sequence analysis showed that the nearest phylogenetic neighbours were Anaerostipes hadrus and Clostridium populeti (92 % sequence similarity) within Clostridium cluster XIVa of the phylum Firmicutes. Therefore, a novel genus is proposed, with the name Caecibacterium sporoformans gen. nov., sp. nov. The type strain of Caecibacterium sporoformans is LMG 27730T=DSM 26959T.

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).

Rarimicrobium hominis gen. nov., sp. nov., representing the fifth genus in the phylum Synergistetes that includes human clinical isolates.

Five human clinical isolates of an unknown, strictly anaerobic, slow-growing, Gram-stain-negative, rod-shaped micro-organism were subjected to a polyphasic taxonomic study. Comparative 16S rRNA gene sequence-based phylogeny showed that the isolates grouped in a clade that included members of the genera Pyramidobacter, Jonquetella, and Dethiosulfovibrio; the type strain of Pyramidobacter piscolens was the closest relative with 91.5-91.7 % 16S rRNA gene sequence similarity. The novel strains were mainly asaccharolytic and unreactive in most conventional biochemical tests. Major metabolic end products in trypticase/glucose/yeast extract broth were acetic acid and propionic acid and the major cellular fatty acids were C13 : 0 and C16 : 0, each of which could be used to differentiate the strains from P. piscolens. The DNA G+C content based on whole genome sequencing for the reference strain 22-5-S 12D6FAA was 57 mol%. Based on these data, a new genus, Rarimicrobium gen. nov., is proposed with one novel species, Rarimicrobium hominis sp. nov., named after the exclusive and rare finding of the taxon in human samples. Rarimicrobium is the fifth genus of the 14 currently characterized in the phylum Synergistetes and the third one in subdivision B that includes human isolates. The type strain of Rarimicrobium hominis is ADV70T ( = LMG 28163T = CCUG 65426T).

Faecalibaculum rodentium gen. nov., sp. nov., isolated from the faeces of a laboratory mouse.

A novel strictly anaerobic strain, ALO17(T), was isolated from mouse faeces and found to produce lactic acid as a major metabolic end product. The isolate was observed to be Gram-stain positive, non-motile, non-spore forming small rods, oxidase and catalase negative, and to form cream-coloured colonies on DSM 104 agar plates. The NaCl range for growth was determined to be 0-2% (w/v). The isolate was found to grow optimally at 37°C, with 0.5% (w/v) NaCl and at pH 7. The cell wall hydrolysates were found to contain ribose as a major sugar. The genomic DNA G+C content was determined to be 52.3mol%. A phylogenetic analysis of the 16S rRNA gene sequence revealed that Holdemanella biformis DSM 3989(T), Faecalicoccus pleomorphus ATCC 29734(T), Faecalitalea cylindroides ATCC 27803(T), and Allobaculum stercoricanis DSM 13633(T) are closely related to the isolate (87.4, 87.3, 86.9 and 86.9% sequence similarity), respectively. The major cellular fatty acids (>10%) of the isolate were identified as C18:1 cis 9 FAME (36.9%), C16:0 FAME (33.7%) and C18:0 FAME (13.2%). In contrast to the tested reference strains, C20:0 FAME (4.0%) was detected only in strain ALO17(T) whilst C16:0 DMA was absent. The isolate also differed in its substrate oxidation profiles from the reference strains by being positive for D-melibiose and stachyose but negative for N-acetyl-D-galactosamine and 3-methyl-D-glucose. On the basis of polyphasic taxonomic evidence from this study, the isolate is concluded to belong to a novel genus within the family Erysipelothricaceae. We propose the name Faecalibaculum rodentium gen. nov., sp. nov. to accommodate strain ALO17(T) (=KCTC 15484(T)=JCM 30274(T)) as the type strain.

Engineering of a Synthetic Metabolic Pathway for the Assimilation of (d)-Xylose into Value-Added Chemicals.

A synthetic pathway for (d)-xylose assimilation was stoichiometrically evaluated and implemented in Escherichia coli strains. The pathway proceeds via isomerization of (d)-xylose to (d)-xylulose, phosphorylation of (d)-xylulose to obtain (d)-xylulose-1-phosphate (X1P), and aldolytic cleavage of the latter to yield glycolaldehyde and DHAP. Stoichiometric analyses showed that this pathway provides access to ethylene glycol with a theoretical molar yield of 1. Alternatively, both glycolaldehyde and DHAP can be converted to glycolic acid with a theoretical yield that is 20% higher than for the exclusive production of this acid via the glyoxylate shunt. Simultaneous expression of xylulose-1 kinase and X1P aldolase activities, provided by human ketohexokinase-C and human aldolase-B, respectively, restored growth of a (d)-xylulose-5-kinase mutant on xylose. This strain produced ethylene glycol as the major metabolic endproduct. Metabolic engineering provided strains that assimilated the entire C2 fraction into the central metabolism or that produced 4.3 g/L glycolic acid at a molar yield of 0.9 in shake flasks.