Hang Dai

Genomic Portrait of the Evolution and Epidemic Spread of a Recently Emerged Multidrug-Resistant Shigella flexneri Clone in China

ABSTRACT Shigella flexneri is the major cause of shigellosis in developing countries. A new S. flexneri serotype, Xv, appeared in 2000 and replaced serotype 2a as the most prevalent serotype in China. Serotype Xv is a variant of serotype X, with phosphoethanolamine modification of its O antigen mediated by a plasmid that contained the opt gene. Serotype Xv isolates belong to sequence type 91 (ST91). In this study, whole-genome sequencing of 59 S. flexneri isolates of 14 serotypes (serotypes 1 to 4, Y, Yv, X, and Xv) indicated that ST91 arose around 1993 by acquiring multidrug resistance (MDR) and spread across China within a decade. A comparative analysis of the chromosome and opt-carrying plasmid pSFXv_2 revealed independent origins of 3 serotype Xv clusters in China, with different divergence times. Using 18 cluster-dividing single-nucleotide polymorphisms (SNPs), SNP typing divided 380 isolates from 3 provinces (Henan, Gansu, and Anhui) into 5 SNP genotypes (SGs). One SG predominated in each province, but substantial interregional spread of SGs was also evident. These findings suggest that MDR is the key selective pressure for the emergence of the S. flexneri epidemic clone and that Shigella epidemics in China were caused by a combination of local expansion and interregional spread of serotype Xv.

Rapid and Sensitive Detection of Listeria ivanovii by Loop-Mediated Isothermal Amplification of the smcL Gene

A loop-mediated isothermal amplification (LAMP) assay for rapid and sensitive detection of the L. ivanovii strains had been developed and evaluated in this study. Oligonucleotide primers specific for L. ivanovii species were designed corresponding to smcL gene sequences. The primers set comprise six primers targeting eight regions on the species-specific gene smcL. The LAMP assay could be completed within 1 h at 64°C in a water bath. Amplification products were directly observed by the Loopamp Fluorescent Detection Reagent (FD) or detected by agarose gel electrophoresis. Moreover, the LAMP reactions were also detected by real-time measurement of turbidity. The exclusivity of 77 non-L. ivanovii and the inclusivity of 17 L. ivanovii were both 100% in the assay. Sensitivity of the LAMP assay was 250 fg DNA and 16 CFU per reaction for detection of L. ivanovii in pure cultures and simulated human stool. The LAMP assay was 10 and 100-fold more sensitive than quantitative PCR (qPCR) and conventional PCR assays,respectively. When applied to human stool samples spiked with low level (8 CFU/0.5 g) of L. ivanovii strains, the new LAMP assay described here achieved positive detection after 6 hours enrichment. In conclusion, the new LAMP assay in this study can be used as a valuable, rapid and sensitive detection tool for the detection of L. ivanovii in field, medical and veterinary laboratories.

SXT/R391 integrative and conjugative elements in Proteus species reveal abundant genetic diversity and multidrug resistance.

SXT/R391 integrative and conjugative elements (ICEs) are self-transmissible mobile genetic elements that are found in most members of Enterobacteriaceae. Here, we determined fifteen SXT/R391 ICEs carried by Proteus isolates from food (4.2%) and diarrhoea patients (17.3%). BLASTn searches against GenBank showed that the fifteen SXT/R391 ICEs were closely related to that from different Enterobacteriaceae species, including Proteus mirabilis. Using core gene phylogenetic analysis, the fifteen SXT/R391 ICEs were grouped into six distinct clusters, including a dominant cluster and three clusters that have not been previously reported in Proteus isolates. The SXT/R391 ICEs shared a common structure with a set of conserved genes, five hotspots and two variable regions, which contained more foreign genes, including drug-resistance genes. Notably, a class A β-lactamase gene was identified in nine SXT/R391 ICEs. Collectively, the ICE-carrying isolates carried resistance genes for 20 tested drugs. Six isolates were resistant to chloramphenicol, kanamycin, streptomycin, trimethoprim-sulfamethoxazole, sulfisoxazole and tetracycline, which are drug resistances commonly encoded by ICEs. Our results demonstrate abundant genetic diversity and multidrug resistance of the SXT/R391 ICEs carried by Proteus isolates, which may have significance for public health. It is therefore necessary to continuously monitor the antimicrobial resistance and related mobile elements among Proteus isolates.

Characterization of Listeria monocytogenes isolated from human Listeriosis cases in China

Characterization of Listeria monocytogenes isolated from human Listeriosis cases in China

Escherichia marmotae sp. nov., isolated from faeces of Marmota himalayana.

The taxonomic position of a group of seven closely related lactose-negative enterobacterial strains, which were isolated from fresh faecal samples of Marmota himalayana collected from the Qinghai-Tibetan plateau, China, was determined by using a polyphasic approach. Cells were Gram-reaction-negative, non-sporulating, non-motile, short rods (0.5-1 × 1-2.5 μm). By 16S rRNA gene sequences, the representative strain, HT073016(T), showed highest similarity values with Escherichia fergusonii ATCC 35469(T) at 99.3%, Escherichia coli ATCC 11775(T) at 99.2%, Escherichia albertii LMG 20976(T) at 98.9%, Escherichia hermannii CIP 103176(T) at 98.4%, and Escherichia vulneris ATCC 33821(T) at 97.7%. Phylogenetic analysis based on the 16S rRNA gene sequences showed that the seven strains formed a monophyletic group with five other species of the genus Escherichia. Digital DNA-DNA hybridization studies between strain HT073016(T) and five other species of the genus Escherichia showed that it shared less than 70% DNA-DNA relatedness with all known species of the genus Escherichia, supporting the novel species status of the strain. The DNA G+C content of strain HT073016(T) was 53.8 mol%. On the basis of phenotypic and phylogenetic characteristics, strain HT073016(T) and the six other HT073016(T)-like strains were clearly distinct from the type strains of other recognized species of the genus Escherichia and represent a novel species of the genus Escherichia, for which the name Escherichia marmotae sp. nov. is proposed, with HT073016(T) ( = CGMCC 1.12862(T) = DSM 28771(T)) as the type strain.

Vibrio fujianensis sp. nov., isolated from aquaculture water

A Gram-stain-negative, facultatively anaerobic strain, designated FJ201301T, was isolated from aquaculture water collected from Fujian province, China. Phylogenetic analysis of 16S rRNA gene sequences indicated that strain FJ201301T belonged to the genus Vibrio, formed a distinct cluster with Vibriocincinnatiensis ATCC 35912T and shared the highest similarity with Vibriosalilacus CGMCC 1.12427T. A 15 bp insertion found in the 16S rRNA gene was a significant marker that distinguished strain FJ201301T from several phylogenetic neighbours (e.g. V. cincinnatiensis). Multilocus sequence analysis of eight genes (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA and topA; concatenated 4135 bp sequence) showed that, forming a long and independent phylogenetic branch, strain FJ201301T clustered with V. cincinnatiensis ATCC 35912T, Vibrioinjenensis KCTC 32233T and Vibriometschnikovii CIP 69.14T clearly separated from V. salilacus CGMCC 1.12427T. Furthermore, the highest in silico DNA-DNA hybridization and average nucleotide identity values between strain FJ201301T and the closest related species were 26.3 and 83.1 % with V. cincinnatiensis ATCC 35912T, less than the proposed cutoff levels for species delineation, i.e. 70 and 95 %, respectively. Biochemical, sequence and genomic analysis suggested the designation of strain FJ201301T representing a novel species of the genus Vibrio, for which the name Vibrio fujianensis sp. nov. is proposed. The type strain is FJ201301T (=DSM 104687T=CGMCC 1.16099T).

Shewanella carassii sp. nov., isolated from surface swabs of crucian carp and faeces of a diarrhoea patient

Two strains, 08MAS2251T and LZ2016-166, were isolated from diverse samples in China collected from the surface of crucian carp and the faeces of a diarrhoea patient, respectively. Both strains were pink-orange coloured, Gram-negative, oxidase- and catalase-positive, facultative anaerobic and motile bacteria, produced H2S and reduced nitrates to nitrites. Growth occurred in the presence of 0-9 % (w/v) NaCl and at 10-42 °C. The optimum conditions were with 1 % (w/v) NaCl and at 35 °C. The phylogenetic tree of 16S rRNA gene demonstrated that strains 08MAS2251T and LZ2016-166 clustered in a distinctive clade next to the species Shewanella chilikensis JC5T within the genus Shewanella. Meanwhile, gyrB gene sequence analysis indicated that the two strains formed an independent branch that was clearly separate from all the other Shewanella species with sequence similarities from 68.49 to 95.74 %. The DNA G+C content of strain 08MAS2251T was 52.68 mol%. Genomic relatedness of in silico DNA-DNA hybridization between strain 08MAS2251T and phylogenetic neighbours ranged from 50.5-51.8 %, below the cutoff of 70 %. In addition, corresponding average nucleotide identity values were between 93.01 to 93.49%, which were lower than 95 % threshold. The major fatty acids of strain 08MAS2251T were C17 : 1ω8c (27.2 %), iso-C15 : 0 (22.5 %), summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c; 8.7 %), C16 : 0 (6.2 %), iso-C13 : 0 (5.6 %) and C17 : 0 (4.5 %). Based on phenotypic and genetic analysis, strains 08MAS2251T and LZ2016-166 are identified as a novel species of the genus Shewanella, for which the name Shewanellacarassii sp. nov. is proposed. The type strain is 08MAS2251T (=DSM 104682T=CGMCC 1.16033T).