BackgroundTo complement next-generation sequencing technologies, there is a pressing need for efficient pre-sequencing capture methods with reduced costs and DNA requirement. The Alu family of short interspersed nucleotide elements is the most abundant type of transposable elements in the human genome and a recognized source of genome instability. With over one million Alu elements distributed throughout the genome, they are well positioned to facilitate genome-wide sequence amplification and capture of regions likely to harbor genetic variation hotspots of biological relevance.ResultsHere we report on the use of inter-Alu PCR with an enhanced range of amplicons in conjunction with next-generation sequencing to generate an Alu-anchored scan, or 'AluScan', of DNA sequences between Alu transposons, where Alu consensus sequence-based 'H-type' PCR primers that elongate outward from the head of an Alu element are combined with 'T-type' primers elongating from the poly-A containing tail to achieve huge amplicon range. To illustrate the method, glioma DNA was compared with white blood cell control DNA of the same patient by means of AluScan. The over 10 Mb sequences obtained, derived from more than 8,000 genes spread over all the chromosomes, revealed a highly reproducible capture of genomic sequences enriched in genic sequences and cancer candidate gene regions. Requiring only sub-micrograms of sample DNA, the power of AluScan as a discovery tool for genetic variations was demonstrated by the identification of 357 instances of loss of heterozygosity, 341 somatic indels, 274 somatic SNVs, and seven potential somatic SNV hotspots between control and glioma DNA.ConclusionsAluScan, implemented with just a small number of H-type and T-type inter-Alu PCR primers, provides an effective capture of a diversity of genome-wide sequences for analysis. The method, by enabling an examination of gene-enriched regions containing exons, introns, and intergenic sequences with modest capture and sequencing costs, computation workload and DNA sample requirement is particularly well suited for accelerating the discovery of somatic mutations, as well as analysis of disease-predisposing germline polymorphisms, by making possible the comparative genome-wide scanning of DNA sequences from large human cohorts.

BackgroundCholera is still a significant public health issue in developing countries. The aetiological agent is Vibrio cholerae and only two serogroups, O1 and O139, are known to cause pandemic or epidemic cholera. In contrast, non-O1/non-O139 V. cholerae has only been reported to cause sporadic cholera-like illness and localised outbreaks. The aim of this study was to determine the genetic diversity of non-O1/non-O139 V. cholerae isolates from hospitalised diarrhoeal patients in Zhejiang Province, China.ResultsIn an active surveillance of enteric pathogens in hospitalised diarrhoeal patients, nine non-O1/non-O139 V. cholerae isolates were identified from 746 diarrhoeal stool samples at a rate of 1.2%. These isolates and an additional 31 isolates from sporadic cases and three outbreaks were analysed using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). PFGE divided the isolates into 25 PFGE types while MLST divided them into 15 sequence types (STs). A single ST, ST80, was predominant which persisted over several years in different cities and caused two outbreaks in recent years. Antibiotic resistance varied with the majority of the isolates resistant to sulphamethoxazole/trimethoprim and nearly all isolates either resistant or intermediate to erythromycin and rifampicin. None of the isolates carried the cholera toxin genes or toxin co-regulated pilus genes but the majority carried a type III secretion system as the key virulence factor.ConclusionsNon-O1/non-O139 V. cholerae is an important contributor to diarrhoeal infections in China. Resistance to commonly used antibiotics limits treatment options. Continuous surveillance of non-O1/non-O139 V. cholerae is important for control and prevention of diarrhoeal infections.

Listeriosis is a severe disease with high mortality rate, especially in immunosuppressed individuals. The causative organism Listeria monocytogenes is primarily transmitted to humans through contaminated foods. To gain an understanding of the prevalence of Listeria in Chinese food products, we reviewed relevant papers from journals published in China from 2000 to 2007. The average recovery rate of Listeria spp. was 3.7% (0.1-7.7%) in all food categories in 13 provinces, with raw meat being the leading source. L. innocua (28.9%, 271/937) and L. monocytogenes (25.3%, 237/937) were more commonly isolated, both at higher proportion in all food types. Subtyping schemes in three laboratories in different provinces revealed that the majority of the L. monocytogenes isolates belonged to lineage II (67.1%), followed by lineage I at 31.6%, including the pathogenic serovars 1/2a, 1/2b, and 4b isolates. Lineage III isolates comprising the low-pathogenic serovar 4a were rare. Knowledge of the prevalence of Listeria in various food products in different regions of China may be useful for developing intervention strategies for control of contaminations along the production chains.

Campylobacter is a zoonotic pathogen that causes foodborne diarrheal illness globally. To better understand health risks in Southeastern China, Campylobacter spp. were surveyed in humans and representative poultry products over 3 years. One hundred and ninety-five representative isolates (n = 148, Campylobacter jejuni; n = 45, Campylobacter coli; n = 2 Campylobacter hyointestinalis) were examined for genetic relatedness and antimicrobial susceptibility. Nearly all Campylobacter isolates (99.0%, 193/195) were resistant to at least one class of antimicrobials, and 45.6% (89/195) of the isolates exhibited multidrug resistance. Genotypic analysis revealed high diversity among tested strains. Multilocus sequence typing (MLST) displayed 120 sequence types (STs) including 42 novel STs being added to the PubMLST international database. Sixty-two STs belonged to 16 previously characterized clonal complexes (CCs), of which CC-21, CC-45, CC-464, CC-574, CC-353, and CC-828 were most frequently identified. In addition, pulsed-field gel electrophoresis (PFGE) fingerprinting resulted in 66 PFGE SmaI patterns among the 125 isolates, with eight patterns shared between human and poultry sources. Subtyping data did not correlate with antimicrobial resistance phenotypes. Taken together, this large-scale surveillance study highlights high antimicrobial resistance and molecular features of Campylobacter isolates in Southeastern China.