Novel Heptaplex PCR-Based Diagnostics for Enteric Fever Caused by Typhoidal Salmonella Serovars and Its Applicability in Clinical Blood Culture.

Enteric fever is caused by typhoidal Salmonella serovars (Typhi, Paratyphi A, Paratyphi B, and Paratyphi C). Owing to the importance of Salmonella serovars in clinics and public hygiene, reliable diagnostics for typhoidal serovars are crucial. This study aimed to develop a novel diagnostic tool for typhoidal Salmonella serovars and evaluate the use of human blood for clinically diagnosing enteric fever. Five genes were selected to produce specific PCR results against typhoidal Salmonella serovars based on the genes of Salmonella Typhi. Heptaplex PCR, including genetic markers of generic Salmonella, Salmonella enterica subsp. enterica, and typhoidal Salmonella serovars, was developed. Typhoidal Salmonella heptaplex PCR using genomic DNAs from 200 Salmonella strains (112 serovars) provided specifically amplified PCR products for each typhoidal Salmonella serovar. These results suggest that heptaplex PCR can sufficiently discriminate between typhoidal and nontyphoidal Salmonella serovars. Heptaplex PCR was applied to Salmonella-spiked blood cultures directly and provided diagnostic results after 12- or 13.5-h blood culture. Additionally, it demonstrated diagnostic performance with colonies recovered from a 6-h blood culture. This study provides a reliable DNA-based tool for diagnosing typhoidal Salmonella serovars that may be useful in clinical microbiology and epidemiology.

Identification of LAB and Fungi in Laru, a Fermentation Starter, by PCR-DGGE, SDS-PAGE, and MALDI-TOF MS.

Samples of Laru (a fermentation starter) obtained from the upper part of Borneo Island were analyzed for their lactic acid bacteria (LAB) and fungal diversity using both a culture-independent method (PCR-DGGE) and culture-dependent methods (SDS-PAGE and MALDI-TOF MS). Pediococcus pentosaceus, Lactobacillus brevis, Saccharomycopsis fibuligera, Hyphopichia burtonii, and Kodamaea ohmeri were detected by all three methods. In addition, Weissella cibaria, Weissella paramesenteroides, Leuconostoc citreum, Leuconostoc mesenteroides, Lactococcus lactis, Rhizopus oryzae/Amylomyces rouxii, Mucor indicus, and Candida intermedia were detected by PCR-DGGE. In contrast, Lactobacillus fermentum, Lactobacillus plantarum, Pichia anomala, Candida parapsilosis, and Candida orthopsilosis were detected only by the culture-dependent methods. Our results indicate that the culture-independent method can be used to determine whether multiple laru samples originated from the same manufacturing region; however, using the culture-independent and the two culture-dependent approaches in combination provides a more comprehensive overview of the laru microbiota.