Computational Study on the Wealth Conformations of Hepatitis C Virus Envelope Glycoprotein 2 and Role of Its Glycan Coat.

Hepatitis C virus (HCV) is a major cause of chronic liver disease and hepatocellular carcinoma. Antibody development efforts mainly revolve around HCV envelope glycoprotein 2 (E2), which mediates host cell entry by interacting with several cell surface receptors, including CD81. We still have limited knowledge about the structural ensembles and the dynamic behavior of both the CD81 binding sites and the glycans on E2. Here, multiple microsecond-long, all-atom molecular dynamics (MD) simulations, as well as a Markov state model (MSM), were performed to provide an atomistic perspective on the dynamic nature of E2 and its glycans. End-to-end accessibility analyses outline a complete overview of the vulnerabilities of the glycan shield of E2, which may be exploited in therapeutic efforts. Additionally, the Markov state model built from the simulation maps four metastable states for AS412 and three metastable states for the front layer in CD81 binding sites, while binding with HEPC3 would induce a conformation selection for both of them. Overall, this work presents hitherto unseen functional and structural insights into E2 and its glycan coat, providing a new theoretical foundation to control the conformational plasticity of E2 that could be harnessed for vaccine development.

PDLIM2 can inactivate the TGF-ß/Smad pathway to inhibit the malignant behavior of ovarian cancer cells.

PDZ-LIM domain-containing Protein 2 (PDLIM2) has been reported to be downregulated in ovarian cancer. However, its exact function and mechanism in regulating ovarian cancer progression have not been elucidated. This work researched the exert effect and mechanism of PDLIM2 on ovarian cancer progression. Briefly, PDLIM2 expression in clinical tissues of ovarian cancer patients and cells was investigated by qRT-PCR and Western blot. The function of PDLIM2 on the proliferation, colony formation, migration and invasion of ovarian cancer cells was explored via cell counting kit-8, colony formation and Transwell assays. To verify whether PDLIM2 regulates ovarian cancer progression via regulating the transforming growth factor-ß (TGF-ß)/Smad pathway, exogenous TGF-ß (10 ng/mL) treatment was performed on the PDLIM2-overexpressed ovarian cancer cells. PDLIM2 effect on the in vivo growth of ovarian cancer cells was researched by establishing a xenograft tumor model. Immunohistochemistry and Western blot were performed to protein expression in cells and tissues. As a result, PDLIM2 was low-expressed in ovarian cancer tissues/cells. PDLIM2 upregulation attenuated the proliferation, colony formation, migration, invasion and epithelial-mesenchymal transition (EMT) of ovarian cancer cells, and inactivated the TGF-ß/Smad pathway. The opposite results were found in the PDLIM2-silenced ovarian cancer cells. Exogenous TGF-ß treatment abrogated the inhibition of PDLIM2 on the malignant behavior of ovarian cancer cells. PDLIM2 upregulation attenuated the in vivo growth and EMT of ovarian cancer cells. Thus, PDLIM2 attenuates the proliferation, migration, invasion and EMT of ovarian cancer cells via inactivating the TGF-ß/Smad pathway. PDLIM2 may be a usefully target for ovarian cancer treatment.

Fibrocyte: A missing piece in the pathogenesis of fibrous epulis.

OBJECTIVES: To explore the role of fibrocytes in the recurrence and calcification of fibrous epulides. METHODS: Different subtypes of fibrous epulides and normal gingival tissue specimens were first collected for histological and immunofluorescence analyses to see if fibrocytes were present and whether they differentiated into myofibroblasts and osteoblasts upon stimulated by transforming growth factor-ß1 (TGF-ß1). Electron microscopy and elemental analysis were used to characterize the extracellular microenvironment in different subtypes of fibrous epulides. Human peripheral blood mononuclear cells (PBMCs) were subsequently isolated from in vitro models to mimic the microenvironment in fibrous epulides to identify whether TGF-ß1 as well as the calcium and phosphorus ion concentration in the extracellular matrix (ECM) of a fibrous epulis trigger fibrocyte differentiation. RESULTS: Fibrous epulides contain fibrocytes that accumulate in the local inflammatory environment and have the ability to differentiate into myofibroblasts or osteoblasts. TGF-ß1 promotes fibrocytes differentiation into myofibroblasts in a concentration-dependent manner, while TGF-ß1 stimulates the fibrocytes to differentiate into osteoblasts when combined with a high calcium and phosphorus environment. CONCLUSIONS: Our study revealed fibrocytes play an important role in the fibrogenesis and osteogenesis in fibrous epulis, and might serve as a therapeutic target for the inhibition of recurrence of fibrous epulides.

Association between dental visit behavior and mortality: a nationwide longitudinal cohort study from NHANES.

OBJECTIVES: The benefits of professional dental treatment for oral diseases have been widely investigated. However, it is unclear whether professional dental treatment provides additional benefits for improving general health. MATERIALS AND METHODS: Data were obtained from the US National Health and Nutrition Examination Survey (NHANES) 1999 to 2004 and 2011 to 2018 cycles. A total of 36,174 participants were included and followed-up for mortality until December 31, 2019. Dental visit behavior was defined as the time interval of last dental visit (TIDV, < 0.5 year, 0.5-1 year, 1-2 years, 2-5 years, and > 5 years) and the main reasons of the last dental visit (treatment, examination, and other reasons). The Cox proportional risk model was used to estimate the hazard ratio (HR) and 95% confidence interval (CI). RESULTS: Compared with participants with time interval of less than 0.5 year, the multivariate-adjusted HRs and 95%CI for participants with time interval of more than 5 years were 1.45 (1.31, 1.61) for all-cause mortality (P trend < 0.0001), 1.49 (1.23, 1.80) for cardiovascular diseases mortality (P trend = 0.0009) and 1.53 (1.29, 1.81) for cancer mortality (P trend = 0.013). Compared with dental visit for examination, participants who had their dental visit for treatment had higher risk for mortality. For participants with dental visit for examination, TIDV of less than 1 year showed lower risk for mortality, whereas TIDV of less than 0.5 year is recommend for population with dental visit for treatment. CONCLUSIONS: Poor dental visit behavior is associated with an increased risk of mortality. Further well-designed studies are needed to confirm the association between professional dental visit and mortality. CLINICAL RELEVANCE: This study highlights the potential benefits of regular dental visits in maintaining general health.

Digital workflow for periodontal splinting with a guided device.

OBJECTIVE: To detail a technique for bonding periodontal splint precisely in a digital workflow. CLINICAL CONSIDERATIONS: Periodontal splinting can be considered to stabilize the mobile teeth, especially for mandibular anterior teeth. Reliable bonding of periodontal splints is a prerequisite for successful clinical performance. However, when bonding the indirect splint to place or making direct splint intraorally, there is a significant risk of mobile teeth drifting away from the splint. To guide accurate insertion of periodontal splint with no risk of displacement of mobile teeth, a guide device fabricated by digital workflow is introduced in this article. CONCLUSIONS: Periodontal compromised teeth can be provisionally fixed during splinting, with the help of the guided device, and precise bonding of the splint is readily accomplished by using such digital workflow. This technique is not only applicable to the lingual splints, but also suitable for the labial ones. CLINICAL SIGNIFICANCE: The use of a guided device, after being digitally designed and fabricated, enables to stabilize the mobile teeth, in case of any displacement during splinting. It is straightforward, and beneficial to reduce the risk of complications, such as debonding of the splint, and secondary occlusal trauma.

A method to improve positioning of denture teeth on denture bases for CAD-CAM complete dentures: A dental technique.

In the digital workflow of complete denture fabrication, one solution for producing computer-aided design and computer-aided manufacturing dentures has been to mill the denture teeth and base separately and then bond them together. The correct bonding of the denture teeth and base is important to reproduce the designed occlusion in the definitive prosthesis. A novel technique is described to assist in the accurate positioning of denture teeth on the denture base by constructing auxiliary positioning slots on the denture base and auxiliary positioning posts on the denture teeth. The technique can assist in accurately assembling CAD-CAM milled complete dentures and may shorten chairside time by reducing clinical occlusal adjustment.

[Analysis on identification and traceability of one non-toxigenic Corynebacterium diphtheriae from a patient with diabetic foot in Hainan province].

There is a rare case of an elderly diabetic with diabetic foot infection at Hainan General Hospital in September 2021, which was diagnosed as Corynebacterium diphtheriae infection incidentally on routine culture with conventional methods and molecular biological approaches, to aid in diagnosis in clinical practice. Owing to smear staining, Albert staining and VITEK 2 system, automated identification systems viz matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) confirmed combing with 16S ribosomal RNA (16S rRNA) gene has been used for the taxonomic classification of bacteria. Otherwise, toxin gene tox was done for diphtheria toxin synthesis. The isolate was Gram-stain-positive, rod-like arrangement with irregular thickness, with characteristic metachromatic granules, ferment most sugars and homology of 16S rRNA analyses with C. diphtheriae NCTC11397T (MW682323.1) was greater than a 100% possibility, toxin gene tox was negative. The findings lay the foundation to clinical identify and trace of non-toxigenic C. diphtheriae. Moreover, this work provides insights into the non-toxigenic C.diphtheriae that contribute to recognized risk of non-toxigenic C.diphtheriae infections.

Recurrence characteristics and clinicopathological results of borderline ovarian tumors.

BACKGROUND: This study aimed to investigate the clinical and pathological characteristics, and the recurrence and prognostic factors of borderline ovarian tumors (BOTs). METHODS: The data of 286 patients admitted to hospital and followed up for more than ten months were analyzed retrospectively to study the clinicopathological characteristics and related factors of recurrence. RESULTS: The median age of the patients was 42.06 ± 14.97 years, and the duration of the follow-up ranged from 10-109 months. During the follow-up period, 40 patients had a recurrence. Of these patients, 36 were ≤ 40 years, and patients with premenopausal recurrence accounted for 20.5% (36/176). In patients undergoing conservative treatment or radical operations, the recurrence rates were 21.3% and 1.8%, respectively, and they were 13.4% (36/268) in patients at Federation International of Gynecology and Obstetrics (FIGO) stage I, and 22.2% (4/18) in patients at an advanced stage. Postoperative pathology revealed that 40 patients had micropapillary tumors, among whom ten patients (25%) had a recurrence, and 19 patients had complications with interstitial infiltration. Of these 19 patients, six had a recurrence (31.5%). Another 22 patients had complications with calcified sand bodies; among these, eight patients (36.4%) had a recurrence. All the differences were statistically significant (P < 0.05). There were four cancer-related deaths during the follow-up period. Late FIGO stage, conservative operation, and a high level of carbohydrate antigen 125 (CA125) were independent risk factors for the recurrence of BOTs. CONCLUSION: BOTs usually occur in women under 40 years, have an occult onset, and half of the patients have no obvious clinical manifestations. Serum CA125 level can be used as a tumor marker to detect BOTs and the risk of its recurrence. Operation mode and FIGO stage are important independent factors for the recurrence of BOTs.

Genomic analysis of Helicobacter himalayensis sp. nov. isolated from Marmota himalayana.

BACKGROUND: Helicobacter himalayensis was isolated from Marmota himalayana in the Qinghai-Tibet Plateau, China, and is a new non-H. pylori species, with unclear taxonomy, phylogeny, and pathogenicity. RESULTS: A comparative genomic analysis was performed between the H. himalayensis type strain 80(YS1)T and other the genomes of Helicobacter species present in the National Center for Biotechnology Information (NCBI) database to explore the molecular evolution and potential pathogenicity of H. himalayensis. H. himalayensis 80(YS1)T formed a clade with H. cinaedi and H. hepaticus that was phylogenetically distant from H. pylori. The H. himalayensis genome showed extensive collinearity with H. hepaticus and H. cinaedi. However, it also revealed a low degree of genome collinearity with H. pylori. The genome of 80(YS1)T comprised 1,829,936 bp, with a 39.89% GC content, a predicted genomic island, and 1769 genes. Comparatively, H. himalayensis has more genes for functions in "cell wall/membrane/envelope biogenesis" and "coenzyme transport and metabolism" sub-branches than the other compared helicobacters, and its genome contained 42 virulence factors genes, including that encoding cytolethal distending toxin (CDT). CONCLUSIONS: We characterized the H. himalayensis 80(YS1)T genome, its phylogenetic position, and its potential pathogenicity. However, further understanding of the pathogenesis of this potentially pathogenic bacterium is required, which might help to manage H. himalayensis-induced diseases.

Paraliobacillus zengyii sp. nov., a slightly halophilic and extremely halotolerant bacterium isolated from Tibetan antelope faeces.

Two rod-shaped, slightly halophilic and extremely halotolerant bacterial strains (X-1125T and X-1174), which were Gram-stain-positive, facultatively anaerobic and motile with peritrichous flagella, were isolated from the faeces of Tibetan antelopes. Their optimal temperature, NaCl concentration and pH for growth were 28 °C, 3 % (w/v) NaCl and pH 7.5, respectively. Based on the results of 16S rRNA gene sequences, and phylogenetic and phylogenomic analyses, their nearest phylogenetic neighbours were Paraliobacillussediminis KCTC 33762T (98.4 % similarity), Paraliobacillusquinghaiensis CGMCC 1.6333T (96.9 %) and Paraliobacillusryukyuensis NBRC 100001T (95.9 %) while the 16S rRNA genes of strains X-1125T and X-1174 were highly similar (99.7 %) to each other. The polar lipids comprised diphosphatidylglycerol, two unidentified phospholipids and four unidentified lipids. MK-7 was the sole menaquinone (100 %). The cell wall contained alanine, glycine, glutamic acid and meso-diaminopimelic acid. The major fatty acids (>9 %) were anteiso-C15 : 0, anteiso-C17 : 0 and C16 : 1ω11c. The in silico DNA-DNA hybridization value between strains X-1125T and X-1174 was 97.8 % (well above the species threshold), but their values were lower than the 70 % threshold with the three closely related type strains. Strains X-1125T and X-1174 had DNA G+C contents (mol%) of 35.2 and 35.1 %, respectively. Based on the presented data, strains X-1125T and X-1174 hereby represent a novel species of the genus Paraliobacillus, for which the name Paraliobacillus zengyii sp. nov. is proposed. The type strain is X-1125T (=DSM 107811T=CGMCC 1.16464T).

Pelistega ratti sp. nov. from Rattus norvegicus of Hainan island.

Two strains (NLN63T and NLN82) of Gram-stain-negative, oxidase- and catalase-positive, bacilli-shaped organisms were isolated from the faecal samples of two separate Rattus norvegicus in Baisha county of Hainan Province, Southern PR China. Phylogenetic analysis based on the near full-length 16S rRNA sequences revealed that strain NLN63T belongs to the genus Pelistega, having maximum similarity to Pelistega suis CCUG 64465T (97.1 %), Pelistega europaea CCUG 39967T (96.2 %) and Pelistega indica DSM 27484T (96.2 %), respectively. The phylogenomic tree built on 553 core genes from genomes of 20 species in the genus Pelistega and other adjacent genera further confirmed that strains NLN63T and NLN82 form a distinct subline and exhibit specific phylogenetic affinity with P. europaea CCUG 39967T. In digital DNA-DNA hybridization analyses, strain NLN63T showed low estimated DNA reassociation values (21.4-22.6 %) with the type strains of the species in the genus Pelistega. The DNA G+C contents of strains NLN63T and NLN82 were 37.3 and 37.1 mol%, respectively. Strain NLN63T had a unique MALDI-TOF MS profile, contained Q-8 as the major quinone and C16 : 0, summed feature 8 (C18 : 1 ω7c/C18 : 1 ω6c or both) and summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c or both) as the dominant fatty acids. Based upon these polyphasic characterization data obtained from the present study, a novel species of the genus Pelistega, Pelistega ratti sp. nov., is proposed with NLN63T (=GDMCC 1.1697T=JCM 33788T) as the type strain.

Associations among gastroesophageal reflux disease, mental disorders, sleep and chronic temporomandibular disorder: a case-control study.

BACKGROUND: Temporomandibular disorders (TMDs) are a family of pain-related disorders associated with impaired function in the jaw, temporomandibular joint and muscles of mastication. Our objectives were to evaluate the association between chronic TMD and gastresophageal reflux disease (GERD) and to determine whether mental disorders or undermined sleep mediates this association. METHODS: We conducted a case-control study involving 1522 consecutive adult patients with chronic TMD and 1522 matched controls from 2 hospitals in China. All participants were aged between 18 and 70 years and were recruited from July 2017 to April 2018 Chronic TMD was diagnosed by trained dentists using the criteria in the Orofacial Pain Prospective Evaluation and Risk Assessment Study. Trained gastroenterologists made blinded diagnoses of GERD according to the Montreal definition and classification (at least 2 d of mild symptoms, or 1 d of moderate or severe symptoms per week). We used validated questionnaires to evaluate psychological status and sleep quality. RESULTS: Of the study participants, we identified 132 patients and 61 controls with GERD. Using conditional logistic regression analysis, we identified GERD as a risk factor for TMD (odds ratio 2.74, 95% confidence interval 1.88 to 3.98). Mediation analyses identified that somatization, anxiety and undermined sleep moderately mediated the relation between TMD and GERD. INTERPRETATION: Our study suggests that symptomatic GERD is associated with chronic, painful TMD, and somatization, anxiety and undermined sleep mediate this association to a certain extent. Due consideration should be given to the evaluation and management of gastrointestinal symptoms and mental disorders in the combined therapy for painful TMD.

Collagen intrafibrillar mineralization as a result of the balance between osmotic equilibrium and electroneutrality.

Mineralization of fibrillar collagen with biomimetic process-directing agents has enabled scientists to gain insight into the potential mechanisms involved in intrafibrillar mineralization. Here, by using polycation- and polyanion-directed intrafibrillar mineralization, we challenge the popular paradigm that electrostatic attraction is solely responsible for polyelectrolyte-directed intrafibrillar mineralization. As there is no difference when a polycationic or a polyanionic electrolyte is used to direct collagen mineralization, we argue that additional types of long-range non-electrostatic interaction are responsible for intrafibrillar mineralization. Molecular dynamics simulations of collagen structures in the presence of extrafibrillar polyelectrolytes show that the outward movement of ions and intrafibrillar water through the collagen surface occurs irrespective of the charges of polyelectrolytes, resulting in the experimentally verifiable contraction of the collagen structures. The need to balance electroneutrality and osmotic equilibrium simultaneously to establish Gibbs-Donnan equilibrium in a polyelectrolyte-directed mineralization system establishes a new model for collagen intrafibrillar mineralization that supplements existing collagen mineralization mechanisms.

Co-delivery of paclitaxel and gemcitabine by methoxy poly(ethylene glycol)-poly(lactide-coglycolide)-polypeptide nanoparticles for effective breast cancer therapy.

Traditional chemotherapeutic drugs have shown limited clinical curative effects in antitumor therapy. The application of multidrug combination and adjuvant-drug carriers is a feasible strategy to overcome the limitations while minimizing the dosage of single drug and acquiring the synergistic effects in tumor therapy. However, the systemic toxicity, drug resistance, and tumor recurrence are still unavoidable. Here we develop core-shell nanoparticles (NPs) to encapsulate paclitaxel (PTX) and gemcitabine (GEM) for breast cancer therapy. We find that the NPs could encapsulate PTX and GEM, with an encapsulation efficiency of 96.3 and 95.13%, respectively. Moreover, the drug loading of these NPs is 2.71% (PTX) and 2.64% (GEM). Notably, the co-delivery of GEM and PTX performs enhanced anticancer effect compared with the PTX alone or GEM alone therapy at the same concentration, which indicates a synergistic effect. Moreover, encapsulation of PTX and GEM by methoxy poly(ethylene glycol)-poly(lactide-coglycolide) also shows enhanced anticancer effects (81.5% tumor inhibition) and reduced systemic toxicity in vivo compared with free drugs (65% tumor inhibition). Together with those results, co-delivery of PTX and GEM by methoxy poly(ethylene glycol)-poly(lactide-coglycolide) might have important potencies in clinical applications for breast cancer therapy.

Isolation and characterization of Streptococcus respiraculi sp. nov. from Marmota himalayana (Himalayan marmot) respiratory tract.

Two bacterial strains were individually isolated from Marmota himalayana respiratory tracts; the animals were from the Tibet-Qinghai Plateau, PR China. The isolates were Gram-stain-positive, catalase-negative, coccus-shaped, chain-forming organisms. Analysis of 16S rRNA gene sequences indicated that the type strain HTS25T shared 98.0, 97.4, 97.2 and 97.1 % similarity with Streptococcus cuniculi, Streptococcus acidominimus, Streptococcus marmotae and Streptococcus himalayensis respectively. Sequence analysis of the sodA and rpoB genes indicated that HTS25T was closely related to S. marmotae (similarities of 94.7 and 91.4 % respectively). Analysis of groEL sequences showed interspecies similarity of 84.8 % between HTS25T and S. himalayensis. A whole-genome phylogenetic tree reconstructed from 81 core genes from the genomes of 17 members of the genus Streptococcus was used to validate that HTS25T forms a distinct subline from other recognized species of the genus Streptococcus. DNA-DNA hybridization of HTS25T showed a maximum estimated DNA reassociation value of 32.1 % to Streptococcus cuniculi CCUG 65085T. On the basis of the results of phenotypic and phylogenetic analyses, we propose that the two isolates be classified as representing a novel species of the genus Streptococcus, named Streptococcus respiraculi sp. nov. The type strain is HTS25T (=DSM 101998T=CGMCC 1.15531T). The genome of Streptococcus respiraculi sp. nov. strain HTS25T (2 067 971 bp) contains 2001 genes with an average DNA G+C content of 42.7 mol%.

Air Entrapment in Demineralized Dentin Adversely Affects Bonding.

PURPOSE: The present study evaluated the influence of air entrapment within demineralized dentin on bond strength, nanoleakage, and degree of conversion. MATERIALS AND METHODS: A vacuum pump with adjustable pressure was used to control the gaseous pressure of a sealed container during the application of a two-step etch-and-rinse adhesive to demineralized dentin. Atmospheric pressure was used as control, and reduced pressures (0.08, 0.06, 0.04 MPa) as experimental variables. Simulated pulpal pressure was adopted during the bonding procedures. After making composite buildups and 24-h water storage, 48 specimens were occlusogingivally sectioned into beams for microtensile bond strength testing, interfacial morphology observation, and nanoleakage evaluation immediately (6 for each group) or after artificial aging (6 for each group); 20 specimens (5 for each group) were occlusogingivally sectioned into slices for degree of conversion testing. Failure modes were compared using the chi-squared test. The other data were analyzed using ANOVA. RESULTS: When air within composite-dentin interface was thinned with reduced pressures, more thorough adhesive infiltration was achieved, and less distinct nanoleakage as well as higher bond strength were observed compared with control groups, regardless of artificial aging. Mixed failure was predominantly identified, and its percentage was higher in the reduced pressure groups than in the controls. Adhesive application at reduced pressure improved the degree of conversion. CONCLUSION: Air entrapment in the demineralized dentin adversely affects composite-dentin bonds. Adhesive application at reduced pressure is helpful for reducing entrapped air, thereby improving the durability of composite-dentin bonds.

Quaternary ammonium-based biomedical materials: State-of-the-art, toxicological aspects and antimicrobial resistance.

Microbial infections affect humans worldwide. Many quaternary ammonium compounds have been synthesized that are not only antibacterial, but also possess antifungal, antiviral and anti-matrix metalloproteinase capabilities. Incorporation of quaternary ammonium moieties into polymers represents one of the most promising strategies for preparation of antimicrobial biomaterials. Various polymerization techniques have been employed to prepare antimicrobial surfaces with quaternary ammonium functionalities; in particular, syntheses involving controlled radical polymerization techniques enable precise control over macromolecular structure, order and functionality. Although recent publications report exciting advances in the biomedical field, some of these technological developments have also been accompanied by potential toxicological and antimicrobial resistance challenges. Recent evidenced-based data on the biomedical applications of antimicrobial quaternary ammonium-containing biomaterials that are based on randomized human clinical trials, the golden standard in contemporary medicinal science, are included in the present review. This should help increase visibility, stimulate debates and spur conversations within a wider scientific community on the implications and plausibility for future developments of quaternary ammonium-based antimicrobial biomaterials.

Streptococcus himalayensis sp. nov., isolated from the respiratory tract of Marmota himalayana.

Five strains of Gram-positive-staining, catalase-negative, coccus-shaped, chain-forming organisms isolated separately from the respiratory tracts of five Marmota himalayana animals in the Qinghai-Tibet Plateau of China were subjected to phenotypic and molecular taxonomic analyses. Comparative analysis of the 16S rRNA gene indicated that these singular organisms represent a new member of the genus Streptococcus, being phylogenetically closest to Streptococcus marmotae DSM 101995T (98.4 % similarity). The groEL, sodA and rpoB sequence analysis showed interspecies similarity values between HTS2T and Streptococcus. marmotae DSM 101995T, its closest phylogenetic relative based on 16S rRNA gene sequences, of 98.2, 78.8 and 93.7 %, respectively. A whole-genome phylogenetic tree built from 82 core genes of genomes from 16 species of the genus Streptococcus validated that HTS2T forms a distinct subline and exhibits specific phylogenetic affinity with S. marmotae. In silico DNA-DNA hybridization of HTS2T showed an estimated DNA reassociation value of 40.5 % with Streptococcus. marmotae DSM 101995T. On the basis of their phenotypic characteristics and phylogenetic findings, it is proposed that the five isolates be classified as representatives of a novel species of the genus Streptococcus, Streptococcus himalayensis sp. nov. The type strain is HTS2T (=DSM 101997T=CGMCC 1.15533T). The genome of Streptococcus himalayensis sp. nov. strain HTS2T contains 2195 genes with a size of 2 275 471 bp and a mean DNA G+C content of 41.3 mol%.

Streptococcuspantholopis sp. nov., isolated from faeces of the Tibetan antelope (Pantholops hodgsonii).

Two bacterial strains were isolated from faecal samples of Tibetan antelopes. The isolates were Gram-stain-positive, catalase-negative, coccus-shaped organisms that were tentatively identified as representing a novel streptococcal species based on their morphological features, biochemical test results and phylogenomic findings. Comparative 16S rRNA gene sequencing studies confirmed that the organisms were members of the genus Streptococcus, but they did not correspond to any recognized species of the genus. The nearest phylogenetic relative of the unknown coccus was Streptococcus ursoris NUM 1615T (93.4 % 16S rRNA gene sequence similarity). Analysis of groEL and rpoB gene sequences of the novel isolates showed interspecies divergence of 27.0 and 22.2 %, respectively, from the type strain of its closest 16S rRNA gene phylogenetic relative, S. ursoris. The complete genome of strain TA 26T has been sequenced. Digital DNA-DNA hybridization studies between strain TA 26T and other species of the genus Streptococcus deposited in the GenBank database showed less than 70 % DNA-DNA relatedness, supporting a novel species status of the strain. On the basis of their genotypic and phenotypic differences from recognized Streptococcus species, the two isolates represent a novel species of the genus Streptococcus, for which the nameStreptococcus pantholopis sp. nov. (type strain TA 26T=CGMCC 1.15667T=DSM 102135T) is proposed.

Streptococcusmarmotae sp. nov., isolated from the respiratory tract of Marmota himalayana.

Five strains of a Gram-stain-positive, catalase-negative, α-haemolytic, coccus-shaped chain-forming organism were isolated separately from the lower respiratory tracts of five animals of Marmota himalayana in the endemic area of plague, the Qinghai-Tibet Plateau, China. Based on their morphological characteristics, biochemical features and molecular phylogenetic studies, the strains were placed as representing a new member of the genus Streptococcus. Comparative 16S rRNA gene sequence studies indicated that strain HTS5T shared 96.5, 96.2 and 96.0 % similarity with Streptococcus gallinaceus CCUG 42692T, Streptococcus parasanguinis ATCC 15912T and Streptococcus suis ATCC 43765T, respectively. Sequence analysis of its rpoB and sodA genes showed that strain HTS5T was most closely related to Streptococcus cuniculi CCUG 65085T with 9.2 and 10.9 % interspecies divergence, respectively. The whole genome phylogenetic tree based on 339 core genes of 65 Streptococcus genomes confirmed that HTS5T belongs to a distinct lineage that is well separated from recognized species of the genus Streptococcus. In silico DNA-DNA hybridization using 65 available genomes from GenBank showed that HTS5T displayed less than 70 % DNA-DNA relatedness with the other 65 species of the genus Streptococcus deposited in the GenBank database. The genome of strain HTS5T (2 322 791 bp) contained 2377 genes and had a G+C content of 41.6 mol%. Therefore, the five strains are considered to represent a novel species of the genus Streptococcus for which the name Streptococcusmarmotae sp. nov. is proposed. The type strain is HTS5T (=DSM 101995T=CGMCC 1.15534T).