Clonal Dissemination of Antifungal-Resistant Candida haemulonii, China.

Candida haemulonii, a relative of C. auris, frequently shows antifungal resistance and is transmissible. However, molecular tools for genotyping and investigating outbreaks are not yet established. We performed genome-based population analysis on 94 C. haemulonii strains, including 58 isolates from China and 36 other published strains. Phylogenetic analysis revealed that C. haemulonii can be divided into 4 clades. Clade 1 comprised strains from China and other global strains; clades 2-4 contained only isolates from China, were more recently evolved, and showed higher antifungal resistance. Four regional epidemic clusters (A, B, C, and D) were identified in China, each comprising ≥5 cases (largest intracluster pairwise single-nucleotide polymorphism differences <50 bp). Cluster A was identified in 2 hospitals located in the same city, suggesting potential intracity transmissions. Cluster D was resistant to 3 classes of antifungals. The emergence of more resistant phylogenetic clades and regional dissemination of antifungal-resistant C. haemulonii warrants further monitoring.

A fine-scale map of genome-wide recombination in divergent Escherichia coli population.

Recombination is one of the most important molecular mechanisms of prokaryotic genome evolution, but its exact roles are still in debate. Here we try to infer genome-wide recombination within a species, utilizing a dataset of 149 complete genomes of Escherichia coli from diverse animal hosts and geographic origins, including 45 in-house sequenced with the single-molecular real-time platform. Two major clades identified based on physiological, clinical and ecological characteristics form distinct genetic lineages based on scarcity of interclade gene exchanges. By defining gene-based syntenies for genomic segments within and between the two clades, we build a fine-scale recombination map for this representative global E. coli population. The map suggests extensive within-clade recombination that often breaks physical linkages among individual genes but seldom interrupts the structure of genome organizational frameworks as well as primary metabolic portfolios supported by the framework integrity, possibly due to strong natural selection for both physiological compatibility and ecological fitness. In contrast, the between-clade recombination declines drastically when phylogenetic distance increases to the extent where a 10-fold reduction can be observed, establishing a firm genetic barrier between clades. Our empirical data suggest a critical role for such recombination events in the early stage of speciation where recombination rate is associated with phylogenetic distance in addition to sequence and gene variations. The extensive intraclade recombination binds sister strains into a quasisexual group and optimizes genes or alleles to streamline physiological activities, whereas the sharply declined interclade recombination split the population into clades adaptive to divergent ecological niches.

Compositional and functional aberrance of the gut microbiota in treatment-naïve patients with primary Sjögren's syndrome.

OBJECTIVES: To investigate the compositional and functional characteristics of the gut microbiota in primary Sjögren's syndrome (pSS) and compare them with those in systemic lupus erythematosus (SLE). METHODS: Stool samples from 78 treatment-naïve pSS patients and 78 matched healthy controls were detected by shotgun metagenomic sequencing and compared with those from 49 treatment-naïve SLE patients. The virulence loads and mimotopes of the gut microbiota were also assessed by sequence alignment. RESULTS: The gut microbiota of treatment-naïve pSS patients had lower richness and evenness and showed a different community distribution than that of healthy controls. The microbial species enriched in the pSS-associated gut microbiota included Lactobacillus salivarius, Bacteroides fragilis, Ruminococcus gnavus, Clostridium bartlettii, Clostridium bolteae, Veillonella parvula, and Streptococcus parasanguinis. Lactobacillus salivarius was the most discriminating species in the pSS patients, especially in those with interstitial lung disease (ILD). Among the differentiating microbial pathways, the superpathway of l-phenylalanine biosynthesis was also further enriched in pSS complicated with ILD. There were more virulence genes carried by the gut microbiota in pSS patients, most of which encoded peritrichous flagella, fimbriae, or curli fimbriae, three types of bacterial surface organelles involved in bacterial colonization and invasion. Five microbial peptides with the potential to mimic pSS-related autoepitopes were also enriched in the pSS gut. SLE and pSS shared significant gut microbial traits, including community distribution, altered microbial taxonomy and pathways, and enriched virulence genes. However, Ruminococcus torques was depleted in pSS patients but enriched in SLE patients compared to healthy controls. CONCLUSIONS: The gut microbiota in treatment-naïve pSS patients was disturbed and shared significant similarity with that in SLE patients.

Compositional and functional aberrance of the gut microbiota in treatment naïve patients with primary Sjögren's syndrome.

OBJECTIVES: To investigate the compositional and functional characteristics of the gut microbiota in primary Sjögren's syndrome (pSS) and compare them with those in systemic lupus erythematosus (SLE). METHODS: Stool samples from 78 treatment naïve pSS patients and 78 matched healthy controls were detected by shotgun metagenomic sequencing and compared with those from 49 treatment naïve SLE patients. The virulence loads and mimotopes of the gut microbiota were also assessed by sequence alignment. RESULTS: The gut microbiota of treatment naïve pSS patients had lower richness and evenness and showed a different community distribution than that of healthy controls. The microbial species enriched in the pSS-associated gut microbiota included Lactobacillus salivarius, Bacteroides fragilis, Ruminococcus gnavus, Clostridium bartlettii, Clostridium bolteae, Veillonella parvula, and Streptococcus parasanguinis. Lactobacillus salivarius was the most discriminating species in the pSS patients, especially in those with interstitial lung disease (ILD). Among the differentiating microbial pathways, the superpathway of l-phenylalanine biosynthesis was also further enriched in pSS complicated with ILD. There were more virulence genes carried by the gut microbiota in pSS patients, most of which encoded peritrichous flagella, fimbriae, or curli fimbriae, three types of bacterial surface organelles involved in bacterial colonization and invasion. Five microbial peptides with the potential to mimic pSS-related autoepitopes were also enriched in the pSS gut. SLE and pSS shared significant gut microbial traits, including the community distribution, altered microbial taxonomy and pathways, and enriched virulence genes. However, Ruminococcus torques was depleted in pSS patients but enriched in SLE patients compared to that in healthy controls. CONCLUSIONS: The gut microbiota in treatment naïve pSS patients was disturbed and shared significant similarity with that in SLE patients.

Novel autoantibodies identified in ACPA-negative rheumatoid arthritis.

OBJECTIVES: Lack of effective biomarkers in anti-citrullinated protein antibody (ACPA)-negative rheumatoid arthritis (RA) impedes early diagnosis and treatment. This study aimed to identify novel autoantibodies in RA and verify their diagnostic values in ACPA-negative RA based on protein microarray technology. METHODS: A total of 1011 sera from 559 RA (276 ACPA-positive and 283 ACPA-negative), 239 disease controls (DCs) and 213 healthy controls (HCs) were collected and sampled on two sequential microarrays and ELISA and western blot (WB) detection, for novel autoantibodies discovery, validation and verification, respectively. The high-density protein microarray printed with a broad spectrum of recombinant human proteins was first employed to screen candidate autoantibodies, then focused microarrays composed of candidate autoantigens were used for validation, followed by ELISA and WB to verify the presence of the most promising candidates in ACPA-negative RA. RESULTS: Nine novel autoantibodies were identified by two sequential microarrays with positivity 17.93%-27.59% and specificities >90% in ACPA-negative RA. Among these, anti-PTX3 and anti-DUSP11 autoantibodies presented with the highest sensitivity and were consistently verified by ELISA and WB. Pooling samples of all cohorts, the positivities of anti-PTX3 and anti-DUSP11 in ACPA-negative RA were 27.56% and 31.80%, respectively, similar to those in ACPA-positive RA, and significantly higher than in HCs (4.69% and 2.35%) and DCs (10.04% and 8.49%) (p<0.0001). Combination of anti-PTX3 with anti-DUSP11 significantly increased the diagnostic sensitivity (38.00%) with specificity of 88.72%, regardless of ACPA status. CONCLUSION: Anti-PTX3 and anti-DUSP11 autoantibodies are newly identified biomarkers for diagnosis of ACPA-negative RA.

Genotypic differences in CC224, CC363, CC449 and CC446 of Moraxella catarrhalis isolates based on whole genome SNP, MLST and PFGE typing.

Understanding the evolutionary path of M. catarrhalis from macrolide-susceptible to macrolide-resistant organism, is important for hindering macrolide resistance from propagation. Multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE) and whole genome SNP typing (WGST), as useful and practical typing tools, have both advantages and disadvantages. We studied the utility of these 3 typing methods, including the level of agreement, consistency and drawbacks, in characterizing M. catarrhalis clones and clonal complexes. We focused on four clonal complexes [CC224, CC363, CC449 (CCN10) and CC446 (CCN08)] and found that PFGE and WGST had a high level of agreement and a proper consistency of the same clone or very closely related clones, while MLST is less discriminatory for different clones. Furthermore, we also established an evolutionary distance cut-off value for "The same clone". Moreover, we detected macrolide-resistant M. catarrhalis in CC224, which had previously been considered as a macrolide-susceptible clonal complex. A higher number of isolates belonged to ST215 compared to ST446, implying that ST215 is more likely to be the primary founder. Our study also demonstrated that all the four clonal complexes belong to the M. catarrhalis lineage 1, which is considered to be related to increased virulence potential and serum resistance. We also observed that copB II was highly related to CC449 and LOS type B was mainly confined in CC224. In conclusion, these findings provide further insight into the evolutionary characteristics of M. catarrhalis.

Peripheral blood mononuclear cell microRNA profiles in syphilitic patients with serofast status.

Syphilis is a chronic sexually transmitted disease caused by infection with Treponema pallidum, which can invade various system organs, leading to clinical manifestations such as neurosyphilis, ocular syphilis, and cardiovascular syphilis and seriously endangering human health. Serofast status is a common outcome after syphilis treatment that presents an important clinical problem. At present, the etiology of serofast status remains unknown. A systematic investigation of the microRNA (miRNA) expression profiles in peripheral blood mononuclear cells (PBMCs) of patients with serofast status or secondary syphilis and of healthy control subjects was conducted using small RNA-seq. The expression of miRNAs was further confirmed by real-time fluorescence quantitative PCR (qPCR) assays. The data reveal a specific miRNA expression profile that was displayed in cells from patients with serofast status. Known and novel predicted (np)-miRNAs were also identified and verified, such as miR-338-5p, np-miR-163, np-miR-128, np-miR-244, and np-miR-5, which together may be used as indicators for treatment evaluation. The functions of genes targeted by the miRNAs differentially expressed in serofast status patients were further analyzed; these genes were found to be involved in various biological functions, such as T-cell receptor signaling pathways, metabolism, and growth. Our study presents the first systematic landscape of miRNAs in PBMCs from patients with serofast status and proposes specific miRNAs linked with serofast status. Our results provide further evidence that serofast status is closely related to host immune function. Additionally, the miRNA expression profile in PBMCs of patients with serofast status generated by this work offers insight into the complex immune network in humans. We hope our results can provide new insights into the pathogenesis of serofast status.

Cross-sectional Whole-genome Sequencing and Epidemiological Study of Multidrug-resistant Mycobacterium tuberculosis in China.

BACKGROUND: The increase in multidrug-resistant tuberculosis (MDR-TB) severely hampers tuberculosis prevention and control in China, a country with the second highest MDR-TB burden globally. The first nationwide drug-resistant tuberculosis surveillance program provides an opportunity to comprehensively investigate the epidemiological/drug-resistance characteristics, potential drug-resistance mutations, and effective population changes of Chinese MDR-TB. METHODS: We sequenced 357 MDR strains from 4600 representative tuberculosis-positive sputum samples collected during the survey (70 counties in 31 provinces). Drug-susceptibility testing was performed using 18 anti-tuberculosis drugs, representing the most comprehensive drug-resistance profile to date. We used 3 statistical and 1 machine-learning methods to identify drug-resistance genes/single-nucleotide polymorphisms (SNPs). We used Bayesian skyline analysis to investigate changes in effective population size. RESULTS: Epidemiological/drug-resistance characteristics showed different MDR profiles, co-resistance patterns, preferred drug combination/use, and recommended regimens among 7 Chinese administrative regions. These factors not only reflected the serious multidrug co-resistance and drug misuse but they were also potentially significant in facilitating the development of appropriate regimens for MDR-TB treatment in China. Further investigation identified 86 drug-resistance genes/intergenic regions/SNPs (58 new), providing potential targets for MDR-TB diagnosis and treatment. In addition, the effective population of Chinese MDR-TB displayed a strong expansion during 1993-2000, reflecting socioeconomic transition within the country. The phenomenon of expansion was restrained after 2000, likely attributable to the advances in diagnosis/treatment technologies and government support. CONCLUSIONS: Our findings provide an important reference and improved understanding of MDR-TB in China, which are potentially significant in achieving the goal of precision medicine with respect to MDR-TB prevention and treatment.

Emergence of a Multidrug-Resistant Hypervirulent Klebsiella pneumoniae Sequence Type 23 Strain with a Rare blaCTX-M-24-Harboring Virulence Plasmid.

Here, we report a multidrug-resistant hypervirulent Klebsiella pneumoniae (MDR-HvKP) strain of sequence type 23 (ST23) with a rare hybrid plasmid harboring virulence genes and blaCTX-M-24, and we analyze the genetic basis for relationship between genotypes and MDR-hypervirulence phenotypes. Further analysis indicates that the hybrid plasmid is formed by IS903D-mediated intermolecular transposition of the blaCTX-M-24 gene into the virulence plasmid. The emergence of MDR-HvKP strains, especially those carrying drug-resistant virulent plasmids, poses unprecedented threats/challenges to public health. This is a dangerous trend and should be closely monitored.

PGAP-X: extension on pan-genome analysis pipeline.

BACKGROUND: Since PGAP (pan-genome analysis pipeline) was published in 2012, it has been widely employed in bacterial genomics research. Though PGAP has integrated several modules for pan-genomics analysis, how to properly and effectively interpret and visualize the results data is still a challenge. RESULT: To well present bacterial genomic characteristics, a novel cross-platform software was developed, named PGAP-X. Four kinds of data analysis modules were developed and integrated: whole genome sequences alignment, orthologous genes clustering, pan-genome profile analysis, and genetic variants analysis. The results from these analyses can be directly visualized in PGAP-X. The modules for data visualization in PGAP-X include: comparison of genome structure, gene distribution by conservation, pan-genome profile curve and variation on genic and genomic region. Meanwhile, result data produced by other programs with similar function can be imported to be further analyzed and visualized in PGAP-X. To test the performance of PGAP-X, we comprehensively analyzed 14 Streptococcus pneumonia strains and 14 Chlamydia trachomatis. The results show that, S. pneumonia strains have higher diversity on genome structure and gene contents than C. trachomatis strains. In addition, S. pneumonia strains might have suffered many evolutionary events, such genomic rearrangements, frequent horizontal gene transfer, homologous recombination, and other evolutionary process. CONCLUSION: Briefly, PGAP-X directly presents the characteristics of bacterial genomic diversity with different visualization methods, which could help us to intuitively understand dynamics and evolution in bacterial genomes. The source code and the pre-complied executable programs are freely available from http://pgapx.ybzhao.com .

Bioinformatics Analysis and Characterization of Highly Efficient Polyvinyl Alcohol (PVA)-Degrading Enzymes from the Novel PVA Degrader Stenotrophomonas rhizophila QL-P4.

Polyvinyl alcohol (PVA) is used widely in industry, and associated environmental pollution is a serious problem. Herein, we report a novel, efficient PVA degrader, Stenotrophomonas rhizophila QL-P4, isolated from fallen leaves from a virgin forest in the Qinling Mountains. The complete genome was obtained using single-molecule real-time (SMRT) technology and corrected using Illumina sequencing. Bioinformatics analysis revealed eight PVA/vinyl alcohol oligomer (OVA)-degrading genes. Of these, seven genes were predicted to be involved in the classic intracellular PVA/OVA degradation pathway, and one (BAY15_3292) was identified as a novel PVA oxidase. Five PVA/OVA-degrading enzymes were purified and characterized. One of these, BAY15_1712, a PVA dehydrogenase (PVADH), displayed high catalytic efficiency toward PVA and OVA substrate. All reported PVADHs only have PVA-degrading ability. Most importantly, we discovered a novel PVA oxidase (BAY15_3292) that exhibited higher PVA-degrading efficiency than the reported PVADHs. Further investigation indicated that BAY15_3292 plays a crucial role in PVA degradation in S. rhizophila QL-P4. Knocking out BAY15_3292 resulted in a significant decline in PVA-degrading activity in S. rhizophila QL-P4. Interestingly, we found that BAY15_3292 possesses exocrine activity, which distinguishes it from classic PVADHs. Transparent circle experiments further proved that BAY15_3292 greatly affects extracellular PVA degradation in S. rhizophila QL-P4. The exocrine characteristics of BAY15_3292 facilitate its potential application to PVA bioremediation. In addition, we report three new efficient secondary alcohol dehydrogenases (SADHs) with OVA-degrading ability in S. rhizophila QL-P4; in contrast, only one OVA-degrading SADH was reported previously.IMPORTANCE With the widespread application of PVA in industry, PVA-related environmental pollution is an increasingly serious issue. Because PVA is difficult to degrade, it accumulates in aquatic environments and causes chronic toxicity to aquatic organisms. Biodegradation of PVA, as an economical and environment-friendly method, has attracted much interest. To date, effective and applicable PVA-degrading bacteria/enzymes have not been reported. Herein, we report a new efficient PVA degrader (S. rhizophila QL-P4) that has five PVA/OVA-degrading enzymes with high catalytic efficiency, among which BAY15_1712 is the only reported PVADH with both PVA- and OVA-degrading abilities. Importantly, we discovered a novel PVA oxidase (BAY15_3292) that is not only more efficient than other reported PVA-degrading PVADHs but also has exocrine activity. Overall, our findings provide new insight into PVA-degrading pathways in microorganisms and suggest S. rhizophila QL-P4 and its enzymes have the potential for application to PVA bioremediation to reduce or eliminate PVA-related environmental pollution.

Precision methylome characterization of Mycobacterium tuberculosis complex (MTBC) using PacBio single-molecule real-time (SMRT) technology.

Tuberculosis (TB) remains one of the most common infectious diseases caused by Mycobacterium tuberculosis complex (MTBC). To panoramically analyze MTBC's genomic methylation, we completed the genomes of 12 MTBC strains (Mycobacterium bovis; M. bovis BCG; M. microti; M. africanum; M. tuberculosis H37Rv; H37Ra; and 6 M. tuberculosis clinical isolates) belonging to different lineages and characterized their methylomes using single-molecule real-time (SMRT) technology. We identified three (m6)A sequence motifs and their corresponding methyltransferase (MTase) genes, including the reported mamA, hsdM and a newly discovered mamB. We also experimentally verified the methylated motifs and functions of HsdM and MamB. Our analysis indicated the MTase activities varied between 12 strains due to mutations/deletions. Furthermore, through measuring 'the methylated-motif-site ratio' and 'the methylated-read ratio', we explored the methylation status of each modified site and sequence-read to obtain the 'precision methylome' of the MTBC strains, which enabled intricate analysis of MTase activity at whole-genome scale. Most unmodified sites overlapped with transcription-factor binding-regions, which might protect these sites from methylation. Overall, our findings show enormous potential for the SMRT platform to investigate the precise character of methylome, and significantly enhance our understanding of the function of DNA MTase.

PanGP: a tool for quickly analyzing bacterial pan-genome profile.

Pan-genome analyses have shed light on the dynamics and evolution of bacterial genome from the point of population. The explosive growth of bacterial genome sequence also brought an extremely big challenge to pan-genome profile analysis. We developed a tool, named PanGP, to complete pan-genome profile analysis for large-scale strains efficiently. PanGP has integrated two sampling algorithms, totally random (TR) and distance guide (DG). The DG algorithm drew sample strain combinations on the basis of genome diversity of bacterial population. The performance of these two algorithms have been evaluated on four bacteria populations with strain numbers varying from 30 to 200, and the DG algorithm exhibited overwhelming advantage on accuracy and stability than the TR algorithm.

Emergence and clonal expansion of Aeromonas hydrophila ST1172 that simultaneously produces MOX-13 and OXA-724.

BACKGROUND: Aeromonas hydrophila infections can cause gastrointestinal symptoms such as diarrhea; however, deep infections are rarely reported. Outbreaks of A. hydrophila are reported more frequently in fish, poultry, and snakes than in humans. This study aimed to track clonal relatedness of deep infections caused by A. hydrophila using whole genome sequencing (WGS). METHODS: We collected three isolates of A. hydrophila in July 19 to August 29, 2019, from patients that underwent spine surgery. Accurate species identification was performed using whole-genome average nucleotide identity (ANI). Antimicrobial susceptibility testing was performed using a VITEK 2 automated AST-N334 Gram-negative susceptibility card system. Antimicrobial resistance and virulence genes were identified using the Comprehensive Antibiotic Resistance Database and Virulence Factor Database VFanalyzer. RESULTS: All three isolates were identified as A. hydrophila based on ANI and multilocus sequence typing analysis revealed that A. hydrophila belonged to a novel sequence type (ST1172). All three isolates were susceptible to amikacin and levofloxacin; however, they were resistant to piperacillin/tazobactam, ceftriaxone, cefuroxime, cefoxitin, and imipenem. Isolate 19W05620 (patient 3) showed increased ceftazidime resistance (minimum inhibitory concentration ≥ 64 µg/mL). All three isolates possessed the same chromosomally encoded ß-lactamases, including blaOXA-724 (ß-lactamase), imiH (metallo-ß-lactamase), and blaMOX-13 (AmpC) in plasmids. CONCLUSIONS: Our study validated the transmission of a novel carbapenem-resistant A. hydrophila sequence type (ST1172) in patients that underwent spine surgery. Control measures should be developed to prevent dissemination of A. hydrophila in the hospital setting.

Tracing the possible evolutionary trends of Morganella morganii: insights from molecular epidemiology and phylogenetic analysis.

Morganella morganii, encompassing two subspecies, subsp. morganii and subsp. sibonii, is a common opportunistic pathogen, notable for intrinsic resistance to multiple antimicrobial agents. Despite its clinical significance, research into the potential evolutionary dynamics of M. morganii remains limited. This study involved the analysis of genome sequences from 431 M. morganii isolates, comprising 206 isolates that cause host infections, obtained from this study and 225 from the NCBI genome data sets. A diverse array of antimicrobial resistance genes (ARGs) was identified in M. morganii isolates, including mcr-1, tet(X4), tmexCD-toprJ, and various carbapenemase genes. In addition, a novel blaKPC-2-bearing plasmid with demonstrated conjugative capability was discovered in M. morganii. The majority of virulence-related genes (VRGs), except for the hlyCABD gene cluster, were found in almost all M. morganii. Three novel genospecies of M. morganii were identified, designated as M. chanii, M. variant1, and M. variant2. Compared to M. sibonii, M. chanii genospecies possessed a greater number of flagellar-related genes, typically located within mobile genetic elements (MGEs), suggesting potential for better environmental adaptability. Phylogenetic analysis further disclosed that M. morganii was divided into 12 sequence clusters (SCs). Particularly, SC9 harbored an elevated abundance of ARGs and VRGs, mainly toxin-related genes, and was associated with a higher presence of MGEs compared to non-SC9 strains. The collective findings suggest that M. morganii undergoes evolution driven by the influence of MGEs, thereby significantly enhancing its adaptability to selective pressures of environmental changes and clinical antimicrobial agents.IMPORTANCEThe growing clinical significance of Morganella morganii arises from its abundant virulence factors and antimicrobial resistance genes, resulting in elevated infection rates and increased clinical scrutiny. However, research on the molecular epidemiology and evolutionary trends of M. morganii has been scarce. Our study established a list of virulence-related genes (VRGs) for M. morganii and conducted a large-scale epidemiological investigation into these VRGs. Based on genomic classification, three novel genotypes of M. morganii were identified, representing evolutionary adaptations and responses to environmental challenges. Furthermore, we discovered the emergence of a sequence cluster enriched with antimicrobial resistance genes, VRGs, and mobile genetic elements, attributed to the selective pressure of antimicrobial agents. In addition, we identified a novel conjugative plasmid harboring the blaKPC-2 gene. These findings hold significance in monitoring and comprehending the epidemiology of M. morganii.

Multicenter comparative genomic study of Klebsiella oxytoca complex reveals a highly antibiotic-resistant subspecies of Klebsiellamichiganensis.

BACKGROUND: The Klebsiella oxytoca complex is an opportunistic pathogen that has been recently identified as an actual complex. However, the characteristics of each species remain largely unknown. We aimed to study the clinical prevalence, antimicrobial profiles, genetic differences, and interaction with the host of each species of this complex. METHODS: One hundred and three clinical isolates of the K. oxytoca complex were collected from 33 hospitals belonging to 19 areas in China from 2020 to 2021. Species were identified using whole genome sequencing based on average nucleotide identity. Clinical infection characteristics of the species were analyzed. Comparative genomics and pan-genome analyses were performed on these isolates and an augmented dataset, including 622 assemblies from the National Center for Biotechnology Information. In vitro assays evaluating the adhesion ability of human respiratory epithelial cells and survivability against macrophages were performed on randomly selected isolates. RESULTS: Klebsiella michiganensis (46.6%, 48/103) and K. oxytoca (35.92%, 37/103) were the major species of the complex causing human infections. K. michiganensis had a higher genomic diversity and larger pan-genome size than did K. oxytoca. K. michiganensis isolates with blaoxy-5 had a higher resistance rate to various antibiotics, antimicrobial gene carriage rate, adhesion ability to human respiratory epithelial cells, and survival rate against macrophages than isolates of other species. CONCLUSION: Our study revealed the genetic diversity of K. michiganensis and firstly identified the highly antimicrobial-resistant profile of K. michiganensis carrying blaoxy-5.

A-to-I RNA Editing in Klebsiella pneumoniae Regulates Quorum Sensing and Affects Cell Growth and Virulence.

Millions of adenosine (A) to inosine (I) RNA editing events are reported and well-studied in eukaryotes; however, many features and functions remain unclear in prokaryotes. By combining PacBio Sequel, Illumina whole-genome sequencing, and RNA Sequencing data of two Klebsiella pneumoniae strains with different virulence, a total of 13 RNA editing events are identified. The RNA editing event of badR is focused, which shows a significant difference in editing levels in the two K. pneumoniae strains and is predicted to be a transcription factor. A hard-coded Cys is mutated on DNA to simulate the effect of complete editing of badR. Transcriptome analysis identifies the cellular quorum sensing (QS) pathway as the most dramatic change, demonstrating the dynamic regulation of RNA editing on badR related to coordinated collective behavior. Indeed, a significant difference in autoinducer 2 activity and cell growth is detected when the cells reach the stationary phase. Additionally, the mutant strain shows significantly lower virulence than the WT strain in the Galleria mellonella infection model. Furthermore, RNA editing regulation of badR is highly conserved across K. pneumoniae strains. Overall, this work provides new insights into posttranscriptional regulation in bacteria.

Epigenome-wide DNA methylation study of whole blood in patients with sporadic amyotrophic lateral sclerosis.

BACKGROUND: Epigenetics, and especially DNA methylation, contributes to the pathogenesis of sporadic amyotrophic lateral sclerosis (SALS). This study aimed to investigate the role of DNA methylation in SALS using whole blood of SALS patients. METHODS: In total, 32 SALS patients and 32 healthy controls were enrolled in this study. DNA was isolated from whole blood collected from the participants. DNA methylation profiles were generated using Infinium MethylationEPIC BeadChip. RESULTS: We identified 34 significant differentially methylated positions (DMPs) in whole blood from SALS patients, compared with the healthy controls. Of these DMPs, five were hypermethylated and 29 were hypomethylated; they corresponded to 13 genes. For the DMPs, ATAD3B and BLK were hypermethylated, whereas DDO, IQCE, ABCB1, DNAH9, FIGN, NRP1, TMEM87B, CCSAP, ST6GALNAC5, MYOM2, and RUSC1-AS1 were hypomethylated. We also identified 12 differentially methylated regions (DMRs), related to 12 genes (NWD1, LDHD, CIS, IQCE, TNF, PDE1C, LGALS1, CSNK1E, LRRC23, ENO2, ELOVL2, and ELOVL2-AS1 ). According to data from the Kyoto Encyclopedia of Genes and Genomes database, DNAH9 and TNF are involved in the amyotrophic lateral sclerosis (ALS) pathway. Correlation analysis between clinical features and DNA methylation profiling indicated that the methylation level of ELOVL2 and ARID1B was positively associated with the age of onset ( r  = 0.86, adjust P   =  0.001) and disease duration ( r  = 0.83, adjust P   =  0.01), respectively. CONCLUSIONS: We found aberrant methylation in DMP- and DMR-related genes, implying that many epigenetic alterations, such as the hypomethylation of DNAH9 and TNF, play important roles in ALS etiology. These findings can be helpful for developing new therapeutic interventions.

A high-fat diet disrupts the hepatic and adipose circadian rhythms and modulates the diurnal rhythm of gut microbiota-derived short-chain fatty acids in gestational mice.

The prevalence of gestational obesity has reached epidemic proportions. Evidence supported that the interactions between the gut microbiota and circadian clocks far reached, affecting host metabolism. Our study aimed to investigate the effect of a high-fat diet (HF) on the hepatic and adipose circadian rhythms in gestational mice and to explore the role of gut microbiota-derived short-chain fatty acids (SCFAs) in mediating the effects. C57BL/6 female mice were randomly fed a standard chow diet (Ctr) or HF prior to and during pregnancy. Samples were collected every 4 h over 24 h (six time points), and 16S rRNA and metabonomics were carried out. Rhythmic patterns were identified and compared using CircaCompare. The results showed that the HF before and during pregnancy significantly induced obesity and worsen glucose tolerance, insulin sensitivity, and lipid metabolism in the gestational mice. Furthermore, the HF significantly disrupted the rhythmic pattern of hepatic and adipose circadian clock genes and downstream metabolic genes. Importantly, our results revealed that the HF altered the diurnal rhythm of the gut microbiota in a diverse manner, which was assessed across three categories: phase shift, loss rhythmicity, and gained rhythmicity. We report here, for the first time, a parallel alteration of the rhythmic phase of butyric acid and butyrate-producing Clostridiaceae_1, which was confirmed by a positive correlation between them. Overall, our research emphasized the importance of the rhythmicity of gut microbiota-derived SCFAs in mediating circadian disruption in response to the HF in gestational mice, which may provide novel insights into the prevention and treatment of gestational obesity.