Immunomagnetic separation combined with colony immunoblotting for selective enrichment and detection of piliated Lactobacillus rhamnosus strains.

AIMS: Piliated Lactobacillus rhamnosus (pLR) strains have attracted much attention owing to their excellent mucus adhering capacity and immunomodulatory effects. Here, we aimed to develop a rapid, sensitive method for isolating pLR strains in complex ecosystems using immunomagnetic separation (IMS) with colony immunoblotting (CIB). METHODS AND RESULTS: Magnetic nanobeads (diameter: 180 nm) conjugated with anti-pLR SpaA pilin antibodies (anti-SpaA) were prepared and used to preconcentrate pLR strains in samples, followed by confirmation with anti-SpaA-based CIB analysis. Under optimized experimental conditions, IMS-CIB selectively recovered pLR strains from 107  CFU ml-1 of faecal microbiota samples spiked with 2·9 × 101 to 2·4 × 106  CFU ml-1 of pLR strains. No positive colonies were detected in samples without addition of pLR strains. The detection limit of IMS-CIB was 29 CFU pLR ml-1 of faecal microbiota, which is much lower than that of CIB without IMS preconcentration (2·0 × 104  CFU ml-1 ). CONCLUSIONS: IMS-CIB allowed selective preconcentration of pLR strains in highly heterogeneous bacterial suspensions and direct detection of pLR colonies, which remained readily available for subsequent isolation. SIGNIFICANCE AND IMPACT OF THE STUDY: Our findings established an effective method for selective enrichment and detection of pLR strains.

Epidemiological surveillance of Campylobacter jejuni in chicken, dairy cattle and diarrhoea patients.

Campylobacter jejuni of different subtypes were identified in chicken, dairy cattle and diarrhoea patients in China from 2005 to 2006, using multiplex PCR and RFLP. The results indicated that, of the three types of samples, C. jejuni was most frequently detected in poultry of the three types of samples, with an average isolation rate of up to 18.61% and a flock contamination rate of 86.67%. The average incidence of C. jejuni in overall cattle and environmental samples, milk cows, heifers and diarrhoea patients was 7.77, 5.02, 8.70 and 4.84%, respectively. A higher prevalence was detected in outpatients than ward patients (P<0.01), and in patients aged <7 years than in older patients (P<0.01). The 265 isolates of C. jejuni were classified into 20 distinct types by PCR-RFLP analysis of the flaA gene, with the genotype distribution in humans overlapping that in poultry and cattle. This suggests that certain C. jejuni strains circulate between humans and domestic animals such as cattle and poultry.