Retrospective use of whole genome sequencing to better understand an outbreak of Salmonella enterica serovar Mbandaka in New South Wales, Australia

Introduction@#Salmonella enterica serovar Mbandaka is an infrequent cause of salmonellosis in New South Wales (NSW) with an average of 17 cases reported annually. This study examined the added value of whole genome sequencing (WGS) for investigating a non-point source outbreak of Salmonella ser. Mbandaka with limited geographical spread.@*Methods@#In February 2016, an increase in Salmonella ser. Mbandaka was noted in New South Wales, and an investigation was initiated. A WGS study was conducted three months after the initial investigation, analysing the outbreak Salmonella ser. Mbandaka isolates along with 17 human and non-human reference strains from 2010 to 2015.@*Results@#WGS analysis distinguished the original outbreak cases (n = 29) into two main clusters: Cluster A (n = 11) and Cluster B (n = 6); there were also 12 sporadic cases. Reanalysis of food consumption histories of cases by WGS cluster provided additional specificity when assessing associations. Discussion: WGS has been widely acknowledged as a promising high-resolution typing tool for enteric pathogens. This study was one of the first to apply WGS to a geographically limited cluster of salmonellosis in Australia. WGS clearly distinguished the outbreak cases into distinct clusters, demonstrating its potential value for use in real time to support non-point source foodborne disease outbreaks of limited geographical spread.

Establishment of digital polymerase chain reaction-ribotyping and database for Clostridium difficile genotyping / 中华传染病杂志

Objective To develop a digital polymerase chain reaction (PCR) ribotyping method and database for Clostridium difficile genotyping.Methods Sequencer based fluorescence capillary gel electrophoresis was used,instead of agarose gel electrophoresis,to establish the digital PCR-ribotyping of Clostridium difficile.Forty Clostridium difficile reference strains,consisting of 10 PCR-ribotypes (RT),were genotyped by the new digital PCR-ribotyping method to set-up the database.Results The sequencer based fluorescence capillary gel electrophoresis correctly detected PCR-ribotyping products of the 40 reference strains,and showed as digital figure;significant differences of these digital figures were found between the 10 RT.High similar digital figures were shown in twenty-one RT027 strains,three RT002 strains and two RT014 strains.However,seven RT001 strains were typed as four subtypes,and two RT014 strains as two subtypes,respectively.Conclusion A digital PCR-ribotyping,and a reference database consisting of 10 RT are successfully established.

Detection of genotype and toxin associated gene of hypervirulent Clostridium difficile clinical isolates / 国际检验医学杂志

Objective To investigate the genotype and variance of toxin associated genes of moxifloxacin‐resistant Clostridium difficile clinical isolates in Sydney .Methods Twenty‐two moxifloxacin‐resistant Clostridium difficile clinical isolates were collected from Sydney ,which were genotyped by using sequencer capillary gel electrophoresis based PCR‐ribotyping ,and toxin A and B cod‐ing gene tcdA and tcdB ,and binary toxin coding gene cdtA and cdtB were detected by using PCR method .Toxin regulator gene tc‐dC was analyzed by using PCR‐sequencing ,and was aligned with reference sequence of VPI 10463 (Genbank accession number :X92982) ,and the tcdC sequence types of all 22 isolates were identified by using blast tool in NCBI .Results Twenty‐one isolates were genotyped as hypervirulent PCR‐ribotypes 027 (RT027) ,and one isolate as RT078 ;all 22 isolates contained tcdA and tcdB for toxin A and B and cdtA and cdtB for binary toxin (tcdA+ tcdB+ cdtA+ cdtB+ ) .The tcdC sequence types of the 21 RT027 i‐solates belong to sc1 ,and that of the one RT078 isolate belongs to WA39 .Compared with tcdC reference sequence of VPI 10463 ,a consecutive 18 bp deletion (nt341 to 379) and one nucleotide deletion at position 117 were found in the 21 RT027 isolates ,and a consecutive 39 bp deletion (nt330 to 368) and one nucleotide mutation at position 184(C> T) were found in the one RT078 isolate . Conclusion Clostridium difficile hypervirulent RT027 was the common moxifloxacin resistant genotype ;Clostridium difficile hy‐pervirulent RT027 and RT078 clinical isolates contained genes for toxin A and B and binary toxin ,and contained gene sequence mu‐tation in toxin regulator gene tcdC .

PCR-ribotyping for genotyping Clostridium dif ficile clinical isolates / 国际检验医学杂志

Objective To investigate the genotype and production of toxin A and B of C .difficile clinical isolates collected from Sydney ,Australia .Methods Sixty‐eight C .difficile clinical isolates were collected from Westmead Hospital ,the University of Sydney ,which were genotyped by using PCR‐ribotyping ,and toxin A ,B coding gene tcdA ,tcdB were detected by using PCR meth‐od .Results Thirty‐one PCR‐ribotypes (RTs) were confirmed in the 68 C .difficile clinical isolates ,RT014 (19 .1% ) and RT002 (11 .8% ) were the common genotypes .Sixty‐four of 68 (94 .1% ) isolates contained tcdA and tcdB for toxin A and B .Conclusion The common prevalent PCR‐ribotypes of C .difficile were RT014 and RT002 in Sydney ,most of the C .difficile clinical isolates contained toxin A and B .