Thermococcus argininiproducens sp. nov., an arginine biosynthesis archaeal species isolated from the Central Indian Ocean ridge.

A strictly anaerobic hyperthermophilic archaeon, designated strain IOH2T, was isolated from a deep-sea hydrothermal vent (Onnuri vent field) area on the Central Indian Ocean Ridge. Strain IOH2T showed high 16S rRNA gene sequence similarity to Thermococcus sibiricus MM 739T (99.42 %), Thermococcus alcaliphilus DSM 10322T (99.28 %), Thermococcus aegaeus P5T (99.21 %), Thermococcus litoralis DSM 5473T (99.13 %), 'Thermococcus bergensis' T7324T (99.13 %), Thermococcus aggregans TYT (98.92 %) and Thermococcus prieurii Bio-pl-0405IT2T (98.01 %), with all other strains showing lower than 98 % similarity. The average nucleotide identity and in silico DNA-DNA hybridization values were highest between strain IOH2T and T. sibiricus MM 739T (79.33 and 15.00 %, respectively); these values are much lower than the species delineation cut-offs. Cells of strain IOH2T were coccoid, 1.0-1.2 µm in diameter and had no flagella. Growth ranges were 60-85 °C (optimum at 80 °C), pH 4.5-8.5 (optimum at pH 6.3) and 2.0-6.0 % (optimum at 4.0 %) NaCl. Growth of strain IOH2T was enhanced by starch, glucose, maltodextrin and pyruvate as a carbon source, and elemental sulphur as an electron acceptor. Through genome analysis of strain IOH2T, arginine biosynthesis related genes were predicted, and growth of strain IOH2T without arginine was confirmed. The genome of strain IOH2T was assembled as a circular chromosome of 1 946 249 bp and predicted 2096 genes. The DNA G+C content was 39.44 mol%. Based on the results of physiological and phylogenetic analyses, Thermococcus argininiproducens sp. nov. is proposed with type strain IOH2T (=MCCC 4K00089T=KCTC 25190T).

Proteiniclasticum aestuarii sp. nov., isolated from tidal flat sediment, and emended descriptions of the genus Proteiniclasticum and Proteiniclasticum ruminis.

A novel bacterium, designated SCR006T, was isolated from tidal flat sediment from Suncheon Bay, Republic of Korea. Cells of strain SCR006T were strictly anaerobic, motile cocci, Gram-reaction-negative, and catalase- and oxidase-negative. Growth was observed at 4-41 °C (optimum, 34-37 °C), at pH 6.5-10.0 (optimum, pH 7.0-7.5) and in presence of 0-8 % NaCl (optimum, 0-2 %). Fermentation products of peptone-yeast-glucose medium were acetate and ethanol. Results of phylogenetic analyses based on 16S rRNA gene sequences indicated that strain SCR006T had high sequence similarity to Proteiniclasticum ruminis D3RC-2T (97.9 %), followed by Youngiibacter multivorans DSM 6139T (95.9 %) and Youngiibacter fragilis 232.1T (95.0 %). The average nucleotide identity value between strain SCR006T and P. ruminis DSM 24773T was 72.7 %, which strongly supported that strain SCR006T reresents a novel species within the genus Proteiniclasticum. The major cellular fatty acids are iso-C15 : 0 (27.2 %) and anteiso-C15 : 0 (16.9 %). The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, two unidentified phospholipids, an unidentified aminolipid and five unidentified lipids. The genomic size was 3.2 Mb with genomic DNA G+C content of 45.6 mol%. The results of 16S rRNA-based and genome-based phylogenetic tree analyses indicated that SCR006T should be assigned to the genus Proteiniclasticum. Strain SCR006T could be distinguished from P. ruminis D3RC-2T by its growth conditions, cell morphology and genomic characteristics. Based on the phenotypic, phylogenetic, genomic and chemotaxonomic features, strain SCR006T represents a novel species, for which the name Proteiniclasticum aestuarii sp. nov. is proposed, with the type strain SCR006T (=KCTC 25245T= JCM 34531T).

Alkalibacter rhizosphaerae sp. nov., a CO-utilizing bacterium isolated from tidal flat sediment, and emended description of the genus Alkalibacter.

A novel anaerobic, rod-shaped, non-motile bacterium, designated strain ES005T, was isolated from tidal flat sediments near the rhizosphere of Phragmites australis at Eulsukdo Island, Republic of Korea. A polyphasic approach revealed that cells of the strain were Gram-stain-positive, catalase- and oxidase-negative, non-spore-forming rods. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strain ES005T belonged to the family Eubacteriaceae, class Clostridia and showed the highest sequence similarity to Alkalibacter mobilis (97.52 %) and followed by Alkalibacter saccharofermentans Z-79820T (96.72%). The OrthoANI value between strain ES005T and A. mobilis was 69.67 %. Strain ES005T grew optimally at 33-37 °C, at pH 6.0-7.0 and in the presence of 1-2 % (w/v) NaCl. Growth in 12.5 % CO atmosphere was observed. Acetate and formate were end products of fructose fermentation and growth on CO. The major cellular fatty acids of strain ES005T were C14 : 0 (39.1 %) and C16 : 0 (26.6 %). The major polar lipids were diphoshatidylgycerol, phosphatidylglycerol and three unidentified phospholipids. The DNA G+C content of strain ES005T was 46.9 mol%. Based on the phenotypic, phylogenetic, genomic and chemotaxonomic features of the isloate, strain ES005T represents a novel species, for which the name Alkalibacter rhizosphaerae sp. nov. is proposed. The type strain is ES005T (=KCTC 25246T=JCM 34530T).

Immune profile by multiplexed immunohistochemistry associated with recurrence after chemoradiation in rectal cancer.

BACKGROUND AND AIM: Evidence has emerged that a pretreatment immune profile in rectal cancer is associated with response to chemoradiotherapy (CRT) and recurrence after CRT. However, few studies have evaluated the immune profile differences after CRT regarding recurrence and nonrecurrence. METHODS: We included patients with advanced rectal cancer treated with CRT and surgery with recurrence within 1 year in a recurrence group. After sex and age matching with the recurrence group, patients with no recurrence for 3 years after CRT were included in a nonrecurrence group. We extracted the immune profile, including CD3 and CD8, from the surgical specimen after CRT using multispectral fluorescence immunohistochemistry and compared the two groups. RESULTS: The immune profiles of 65 patients with rectal cancer were assessed; 30 were included in the recurrence group and 35 were included in the nonrecurrence group. CD3+ and CD8+ T lymphocyte densities were significantly higher in the nonrecurrence group than in the recurrence group (CD3+ ; P < 0.001, CD8+ ; P = 0.003) in the primary tumor. Consistent results were found in epithelial and stromal cells. Compared with the recurrence group, the distinct profiles of co-expressed immune markers in the nonrecurrence group were revealed (CD3+ CD8+ , P = 0.001; CD3+ CD8+ PD-L1- , P = 0.001; CD3+ CD8+ FOXP3- PD-L1- , P = 0.001). CONCLUSIONS: Vigorous CD3+ and CD8+ T cell priming post-CRT was prominent in the nonrecurrence group compared with that of the recurrence group. This finding suggests that differences in immune profiles may have clinical significance even after CRT.

A simple biosynthetic pathway for 2,3-butanediol production in Thermococcus onnurineus NA1.

The biosynthetic pathway of 2,3-butanediol (2,3-BDO) production from pyruvate under anaerobic conditions includes three enzymes: acetolactate synthase (ALS), acetolactate decarboxylase (ALDC), and acetoin reductase (AR). Recently, in anaerobic hyperthermophilic Pyrococcus furiosus, it has been reported that acetoin, a precursor of 2,3-BDO, is produced from pyruvate by ALS through a temperature-dependent metabolic switch. In this study, we first attempted to produce 2,3-BDO from Thermococcus onnurineus NA1 using a simple biosynthetic pathway by two enzymes (ALS and AR) at a high temperature. Two heterologous genes, acetolactate synthase (alsS) from Pyrococcus sp. NA2 and alcohol dehydrogenase (adh) from T. guaymacensis, were introduced and expressed in T. onnurineus NA1. The mutant strain produced approximately 3.3 mM 2,3-BDO at 80 °C. An acetyl-CoA synthetase IIIα (TON_1001) was further deleted to enhance 2,3-BDO production, and the mutant strain showed a 25% increase in the specific production of 2,3-BDO. Furthermore, when carbon monoxide (CO) gas was added as a reductant, specific production of 2,3-BDO increased by 45%. These results suggest a new biosynthetic pathway for 2,3-BDO and demonstrate the possibility of T. onnurineus NA1 as a platform strain for 2,3-BDO production at high temperatures.

Transcriptomic profiling and its implications for the H2 production of a non-methanogen deficient in the frhAGB-encoding hydrogenase.

The F420-reducing hydrogenase of methanogens functions in methanogenesis by providing reduced coenzyme F420 (F420H2) as an electron donor. In non-methanogens, however, their physiological function has not been identified yet. In this study, we constructed an ΔfrhA mutant, whose frhA gene encoding the hydrogenase α subunit was deleted, in the non-methanogenic Thermococcus onnurineus NA1 as a model organism. There was no significant difference in the formate-dependent growth between the mutant and the wild-type strains. Interestingly, the mutation in the frhA gene affected the expression of genes involved in various cellular functions such as H2 oxidation, chemotactic signal transduction, and carbon monoxide (CO) metabolism. Among these genes, the CO oxidation gene cluster, enabling CO-dependent growth and H2 production, showed a 2.8- to 7.0-fold upregulation by microarray-based whole transcriptome expression profiling. The levels of proteins produced by this gene cluster were also significantly increased not only under the formate condition but also under the CO condition. In a controlled bioreactor, where 100% CO was continuously fed, the ΔfrhA mutant exhibited significant increases in cell growth (2.8-fold) and H2 production (3.4-fold). These findings strongly imply that this hydrogenase is functional in non-methanogens and is related to various cellular metabolic processes through an unidentified mechanism. An understanding of the mechanism by which the frhA gene deletion affected the expression of other genes will provide insights that can be applied to the development of strategies for the enhancement of H2 production using CO as a substrate.

Formate-driven growth coupled with H(2) production.

Although a common reaction in anaerobic environments, the conversion of formate and water to bicarbonate and H(2) (with a change in Gibbs free energy of ΔG° = +1.3 kJ mol(-1)) has not been considered energetic enough to support growth of microorganisms. Recently, experimental evidence for growth on formate was reported for syntrophic communities of Moorella sp. strain AMP and a hydrogen-consuming Methanothermobacter species and of Desulfovibrio sp. strain G11 and Methanobrevibacter arboriphilus strain AZ. The basis of the sustainable growth of the formate-users is explained by H(2) consumption by the methanogens, which lowers the H(2) partial pressure, thus making the pathway exergonic. However, it has not been shown that a single strain can grow on formate by catalysing its conversion to bicarbonate and H(2). Here we report that several hyperthermophilic archaea belonging to the Thermococcus genus are capable of formate-oxidizing, H(2)-producing growth. The actual ΔG values for the formate metabolism are calculated to range between -8 and -20 kJ mol(-1) under the physiological conditions where Thermococcus onnurineus strain NA1 are grown. Furthermore, we detected ATP synthesis in the presence of formate as a sole energy source. Gene expression profiling and disruption identified the gene cluster encoding formate hydrogen lyase, cation/proton antiporter and formate transporter, which were responsible for the growth of T. onnurineus NA1 on formate. This work shows formate-driven growth by a single microorganism with protons as the electron acceptor, and reports the biochemical basis of this ability.

Characterization of the frhAGB-encoding hydrogenase from a non-methanogenic hyperthermophilic archaeon.

The F420-reducing hydrogenase has been known as a key enzyme in methanogenesis. Its homologs have been identified in non-methanogenic hyperthermophilic archaea, including Thermococcus onnurineus NA1, but neither physiological function nor biochemical properties have been reported to date. The enzyme of T. onnurineus NA1 was distinguished from those of other methanogens and the members of the family Desulfurobacteriaceae with respect to the phylogenetic distribution of the α and ß subunits, organization of frhAGB genes and conservation of F420-coordinating residues. RT-qPCR and Western blot analyses revealed frhA gene is not silent but is expressed in T. onnurineus NA1 grown in the presence of sulfur, carbon monoxide, or formate. The trimeric enzyme complex was purified to homogeneity via affinity chromatography from T. onnurineus NA1 and exhibited catalytic activity toward the electron acceptors such as viologens and flavins but not the deazaflavin coenzyme F420. This is the first biochemical study on the function of the frhAGB-encoding enzyme from a non-methanogenic archaea.

Enhancing the processivity of a family B-type DNA polymerase of Thermococcus onnurineus and application to long PCR.

Mechanisms that allow replicative DNA polymerases to attain high processivity are often specific to a given polymerase and cannot be generalised to others. Amplification efficiency is lower in family B-type DNA polymerases than in family A-type (Taq) polymerases because of their strong 3'-5' exonuclease-activity. Here, we have red the exonuclease domain of the Thermococcus onnurineus NA1 (TNA1) DNA polymerase, especially Asn210 to Asp215 residues in Exo II motif (NXXXFD), to improve the processivity. N213D mutant protein had higher processivity and extension rate than the wild-type TNA1 DNA polymerase, retaining a lower mutation frequency than recombinant Taq DNA polymerase. Consequently, the N213D mutant could amplify target DNA up to 13.5 kb in length from human genomic DNA and 16.2 kb in length from human mitochondrial DNA while wild-type TNA1 amplified target DNA of 2.7 kb in length from human genomic DNA.

Hearing impairment increases the risk of hip fracture-related mortality and recurrent hip fractures: A propensity score matching analysis.

PURPOSE: Despite the ongoing rise in hip fractures and the adverse effects of hearing impairment (HI) on increased mortality and morbidity, research addressing the influence of HI on mortality risk or complications in patients with hip fractures remains absent. This study aimed to analyze the effects of HI on mortality and treatment outcomes among patients with hip fracture. METHODS: We retrospectively collected data from consecutive patients diagnosed with hip fractures between January 2007 and March 2022 who had auditory examination records. From the initially enrolled 265 patients, data for 58 with HI and 58 without HI (control group) were extracted using a 1:1 propensity score matching. The primary outcome included comparison of mortality rates, and the secondary outcome encompassed the comparison of postoperative medical and surgical complications. RESULTS: The 1-year cumulative mortality rate was not significantly different between the HI and control groups, but the overall cumulative mortality rate was significantly higher in the HI than in the control group (63.0 % and 48.6, respectively; P = 0.046) in a follow-up period of up to 16 years. The HI group had a significantly higher incidence of "second hip fractures due to falls" than the control group (P = 0.016), although no differences in other medical and surgical complications were revealed. CONCLUSIONS: Awareness of the long-term risk of higher mortality when managing patients with hip fracture and HI is important. To reduce the risk of second hip fractures, paying more attention to fall prevention education and taking a more proactive approach, especially for those with HI.

CO-dependent H2 production by genetically engineered Thermococcus onnurineus NA1.

Hydrogenogenic CO oxidation (CO + H(2)O → CO(2) + H(2)) has the potential for H(2) production as a clean renewable fuel. Thermococcus onnurineus NA1, which grows on CO and produces H(2), has a unique gene cluster encoding the carbon monoxide dehydrogenase (CODH) and the hydrogenase. The gene cluster was identified as essential for carboxydotrophic hydrogenogenic metabolism by gene disruption and transcriptional analysis. To develop a strain producing high levels of H(2), the gene cluster was placed under the control of a strong promoter. The resulting mutant, MC01, showed 30-fold-higher transcription of the mRNA encoding CODH, hydrogenase, and Na(+)/H(+) antiporter and a 1.8-fold-higher specific activity for CO-dependent H(2) production than did the wild-type strain. The H(2) production potential of the MC01 mutant in a bioreactor culture was 3.8-fold higher than that of the wild-type strain. The H(2) production rate of the engineered strain was severalfold higher than those of any other CO-dependent H(2)-producing prokaryotes studied to date. The engineered strain also possessed high activity for the bioconversion of industrial waste gases created as a by-product during steel production. This work represents the first demonstration of H(2) production from steel mill waste gas using a carboxydotrophic hydrogenogenic microbe.

Genomic potential and physiological characteristics of C1 metabolism in novel acetogenic bacteria.

Acetogenic bacteria can utilize C1 compounds, such as carbon monoxide (CO), formate, and methanol, via the Wood-Ljungdahl pathway (WLP) to produce biofuels and biochemicals. Two novel acetogenic bacteria of the family Eubacteriaceae ES2 and ES3 were isolated from Eulsukdo, a delta island in South Korea. We conducted whole genome sequencing of the ES strains and comparative genome analysis on the core clusters of WLP with Acetobacterium woodii DSM1030T and Eubacterium limosum ATCC8486T. The methyl-branch cluster included a formate transporter and duplicates or triplicates copies of the fhs gene, which encodes formyl-tetrahydrofolate synthetase. The formate dehydrogenase cluster did not include the hydrogenase gene, which might be replaced by a functional complex with a separate electron bifurcating hydrogenase (HytABCDE). Additionally, duplicated copies of the acsB gene, encoding acetyl-CoA synthase, are located within or close to the carbonyl-branch cluster. The serum bottle culture showed that ES strains can utilize a diverse range of C1 compounds, including CO, formate, and methanol, as well as CO2. Notably, ES2 exhibited remarkable resistance to high concentrations of C1 substrates, such as 100% CO (200 kPa), 700 mM formate, and 500 mM methanol. Moreover, ES2 demonstrated remarkable growth rates under 50% CO (0.45 h-1) and 200 mM formate (0.34 h-1). These growth rates are comparable to or surpassing those previously reported in other acetogenic bacteria. Our study introduces novel acetogenic ES strains and describes their genetic and physiological characteristics, which can be utilized in C1-based biomanufacturing.

H2 production from CO, formate or starch using the hyperthermophilic archaeon, Thermococcus onnurineus.

The hyperthermophilic archaeon, Thermococcus onnurineus, was grown in media supplemented with either CO, formate, or starch. H(2) was produced with each substrate with respective maximum rates of 1.55, 3.83 and 2.66 mmol H(2)/l h. The yields (mol H(2)/mol substrate) were 0.98, 1 and 3.13, respectively. This microbe is the first example where a single microorganism can grow and produce H(2) using CO, formate or starch as substrate.

Blood KL-6 predicts prognosis in primary Sjögren's syndrome-associated interstitial lung disease.

Interstitial lung disease associated with primary Sjögren's syndrome (SJS-ILD) has a variable clinical course. We aimed to investigate the role of blood biomarkers in predicting prognosis for SJS-ILD. Clinical data of 46 SJS-ILD patients were retrospectively reviewed. Plasma biomarker levels, including Krebs von den Lungen-6 (KL-6), CC chemokine ligand 18 (CCL18), chitinase-3-like-1 (YKL-40), interleukin-4 receptor alpha (IL-4Ra), and matrix metalloproteinase-7 (MMP-7) were measured using the multiplex Luminex assays (R&D Systems, Minneapolis, USA). The median follow-up period was 69.0 months. The mean age of the patients was 59.4 years; 17.4% were men. The KL-6 level was significantly higher in non-survivors (n = 12; 119.6 vs. 59.5 pg/mL, P = 0.037) than survivors (n = 34), while the levels of the other biomarkers did not differ. Receiver operating characteristic analysis indicated that KL-6 shows the best performance for predicting survival (area under the curve = 0.705, P = 0.037; best cut-off value = 53.5 pg/mL). Multivariable Cox analysis that was adjusted by age and diffusing capacity for carbon monoxide suggested a high KL-6 level (> 53.5 pg/mL) as an independent prognostic factor for survival (hazard ratio = 5.939, 95% confidence interval 1.312-26.881, P = 0.021). Our results suggest that blood KL-6 might be a useful in predicting the prognosis for patients with SJS-ILD.

Bioconversion of CO to formate by artificially designed carbon monoxide:formate oxidoreductase in hyperthermophilic archaea.

Ferredoxin-dependent metabolic engineering of electron transfer circuits has been developed to enhance redox efficiency in the field of synthetic biology, e.g., for hydrogen production and for reduction of flavoproteins or NAD(P)+. Here, we present the bioconversion of carbon monoxide (CO) gas to formate via a synthetic CO:formate oxidoreductase (CFOR), designed as an enzyme complex for direct electron transfer between non-interacting CO dehydrogenase and formate dehydrogenase using an electron-transferring Fe-S fusion protein. The CFOR-introduced Thermococcus onnurineus mutant strains showed CO-dependent formate production in vivo and in vitro. The maximum formate production rate from purified CFOR complex and specific formate productivity from the bioreactor were 2.2 ± 0.2 µmol/mg/min and 73.1 ± 29.0 mmol/g-cells/h, respectively. The CO-dependent CO2 reduction/formate production activity of synthetic CFOR was confirmed, indicating that direct electron transfer between two unrelated dehydrogenases was feasible via mediation of the FeS-FeS fusion protein.

Establishment of Genome Based Criteria for Classification of the Family Desulfovibrionaceae and Proposal of Two Novel Genera, Alkalidesulfovibrio gen. nov. and Salidesulfovibrio gen. nov.

Bacteria in the Desulfovibrionaceae family, which contribute to S element turnover as sulfate-reducing bacteria (SRB) and disproportionation of partially oxidized sulfoxy anions, have been extensively investigated since the importance of the sulfur cycle emerged. Novel species belonging to this taxon are frequently reported, because they exist in various environments and are easy to culture using established methods. Due to the rapid expansion of the taxon, correction and reclassification have been conducted. The development of high-throughput sequencing facilitated rapid expansion of genome sequence database. Genome-based criteria, based on these databases, proved to be potential classification standard by overcoming the limitations of 16S rRNA-based phylogeny. Although standards methods for taxogenomics are being established, the addition of a novel genus requires extensive calculations with taxa, including many species, such as Desulfovibrionaceae. Thus, the genome-based criteria for classification of Desulfovibrionaceae were established and validated in this study. The average amino-acid identity (AAI) cut-off value, 63.43 ± 0.01, was calculated to be an appropriate criterion for genus delineation of the family Desulfovibrionaceae. By applying the AAI cut-off value, 88 genomes of the Desulfovibrionaceae were divided into 27 genera, which follows the core gene phylogeny results. In this process, two novel genera (Alkalidesulfovibrio and Salidesulfovibrio) and one former invalid genus ("Psychrodesulfovibrio") were officially proposed. Further, by applying the 95-96% average nucleotide identity (ANI) standard and the 70% digital DNA-DNA hybridization standard values for species delineation of strains that were classified as the same species, five strains have the potential to be newly classified. After verifying that the classification was appropriately performed through relative synonymous codon usage analysis, common characteristics were listed by group. In addition, by detecting metal resistance related genes via in silico analysis, it was confirmed that most strains display metal tolerance.

Multiplex Immunofluorescence Assay of Infiltrating Mononu-Clear Cell Subsets in Acute T-Cell-Mediated Rejection and BK Virus-Associated Nephropathy in the Allograft Kidney.

Renal allograft biopsy is the gold standard procedure for diagnosis of kidney rejection via specific pathological changes. To provide a better assessment of immunologic events in acute T-cell-mediated rejection (acute TCMR) and BK virus-associated nephropathy (BKVAN) cases, we used multiplex immunofluorescence staining to identify infiltrating mononuclear cell subsets in the cortex area of transplanted kidneys. Antibodies to CD4, CD8, CD20, CD68, Foxp3, and cytokeratin were used. In cortical interstitium, CD8+ cells were significantly more prevalent in acute TCMR than BKVAN cases (34% vs. 22.8%, p = 0.034). In medulla, CD20+ cells were significantly more prevalent in BKVAN than acute TCMR cases (51.9% vs. 11.3%, p = 0.028).

Enrichment and proteome analysis of a hyperthermostable protein set of archaeon Thermococcus onnurineus NA1.

Thermococcus onnurineus NA1 is a hyperthermophilic archaeon that can be used for the screening of thermophilic enzymes. Previously, we characterized the metabolic enzymes of the cytosolic proteome by two-dimensional electrophoresis/tandem mass spectrometry (2-DE/MS-MS). In this study, we identified a subset of hyperthermostable proteins in the cytosolic proteome using enrichment by in vitro heat treatment and protein identification. After heat treatment at 100°C for 2 h, 13 and 149 proteins were identified from the soluble proteome subset by 2-DE/MS-MS and 1-DE/MS-MS analysis, respectively. Representative proteins included intracellular protease I, thioredoxin reductase, triosephosphate isomerase, putative hydroperoxide reductase, proteasome, and translation initiation factors. Intracellular protease, deblocking aminopeptidases, and fructose-1,6-bisphosphatase were overexpressed in Escherichia coli and biological activity above 85°C was confirmed. The folding transition temperature (Tm) of identified proteins was analyzed using the in silico prediction program TargetStar. The proteins enriched with the heat treatment have higher Tm than the homologous proteins from mesophilic strains. These results suggested that the heat-stable protein set of hyperthermophilic T. onnurineus NA1 can be effectively fractionated and enriched by in vitro heat treatment.

Long-term clinical course and outcome in patients with primary Sjögren syndrome-associated interstitial lung disease.

Interstitial lung disease (ILD) is the most common lung manifestation in patients with Sjögren syndrome (SJS) and is associated with poor outcomes. This study aimed to investigate the long-term clinical course and prognostic factors in patients with SJS-ILD. Clinical data and high-resolution computed tomography (HRCT) images of 62 patients with primary SJS-ILD were retrospectively analyzed (biopsy-proven cases, n = 16). The mean patient age was 59.8 years; 83.9% of the patients were females, and 38.7% showed a usual interstitial pneumonia (UIP) pattern on HRCT. The median follow-up period was 61.5 months. During follow-up, 15 patients (24.2%) died, 7 (11.3%) experienced acute exacerbation (AE), and 27 (43.5%) progressed. The 1-, 3- and 5-year survival rates were 93.5%, 85.8%, and 81.1%, respectively. Age (hazard ratio [HR]: 1.158, P = 0.003), C-reactive protein (CRP) level (HR: 1.212, P = 0.045), FVC (HR: 0.902, P = 0.005), and a UIP pattern on HRCT (HR: 4.580, P = 0.029) were significant prognostic factors in multivariable Cox analysis. In conclusion, death, AE, and ILD progression occurred in 25%, 10%, and 50% of the patients with SJS-ILD, respectively. Older age, higher CRP level, lower FVC, and a UIP pattern on HRCT indicated poor prognosis.

Paradesulfovibrio onnuriensis gen. nov., sp. nov., a chemolithoautotrophic sulfate-reducing bacterium isolated from the Onnuri vent field of the Indian Ocean and reclassification of Desulfovibrio senegalensis as Paradesulfovibrio senegalensis comb. nov.

An anaerobic, rod-shaped, mesophilic, chemolithoautotrophic, sulfate-reducing bacterial strain IOR2T was isolated from a newly found deep-sea hydrothermal vent (OVF, Onnuri Vent Field) area in the central Indian Ocean ridge (11°24'88″ S 66°25'42″ E, 2021 m water depth). The 16S rRNA gene sequence analysis revealed that the strain IOR2T was most closely related to Desulfovibrio senegalensis BLaC1T (96.7%). However, it showed low similarity with the members of the family Desulfovibrionaceae, such as Desulfovibrio tunisiensis RB22T (94.0%), D. brasiliensis LVform1T (93.9%), D. halophilus DSM 5663T (93.7%), and Pseudodesulfovibrio aespoeensis Aspo-2T (93.2%). The strain IOR2T could grow at 23-42°C (optimum 37°C), pH 5.0-8.0 (optimum pH 7.0) and with 0.5-6.5% (optimum 3.0%) NaCl. The strain could use lactate, pyruvate, H2, and glycerol as electron donors and sulfate, thiosulfate, and sulfite as electron acceptors. The major fatty acids of the strain IOR2T were iso-C15:0, iso-C17:0, ante-iso-C15:0, and summed feature 9 (C16:0 methyl/iso-C17:1ω9c). Both the strains IOR2T and BLaC1T could grow with CO2 and H2 as the sole sources of carbon and energy, respectively. Genomic evidence for the Wood-Ljungdahl pathway in both the strains reflects chemolithoautotrophic growth. The DNA G + C content of the strain IOR2T and BLaC1T was 58.1-60.5 mol%. Based on the results of the phylogenetic and physiologic studies, Paradesulfovibrio onnuriensis gen. nov., sp. nov. with the type strain IOR2T (= KCTC 15845T = MCCC 1K04559T) was proposed to be a member of the family Desulfovibrionaceae. We have also proposed the reclassification of D. senegalensis as Paradesulfovibrio senegalensis comb. nov.