Pusillimonas thiosulfatoxidans sp. nov., a thiosulfate oxidizer isolated from activated sludge.

A Gram-stain-negative, motile bacterium, designated strain YE3T, was isolated from activated sludge obtained from a municipal wastewater treatment plant in Daejeon Metropolitan City, Republic of Korea. The cells were oxidase- and catalase-positive, and grew under aerobic conditions at 10-40 °C (optimum, 30 °C), with 1.0-8.0 % (w/v) NaCl (1.0 %) and at pH 5.5-9.0 (pH 7.0). Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain YE3T was most closely related to Pusillimonasharenae KACC 14927T (98.2 % sequence similarity) and Pusillimonasginsengisoli KCTC 22046T (98.0 %). DNA-DNA relatedness values for strain YE3T and P. harenae KACC 14927T, P. ginsengisoli KCTC 22046T and P. soli KCTC 22455T were 28.7±2.27 %, 21.3±1.16 %, and 14.0±0.67 %, respectively. The genomic G+C content of the type strain YE3T was 59.3 mol%, as determined by whole-genome sequencing. The dominant fatty acids were C16 : 0 (39.2 %) and C17 : 0cyclo (37.5 %). The major polar lipids of strain YE3T were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. Two aminophospholipids and four unidentified lipids were also detected. Furthermore, strain YE3T was able to oxidize thiosulfate under heterotrophic conditions. Based on the phenotypic, genotypic, chemotaxonomic and phylogenetic analyses, strain YE3T represents a novel species of the genus Pusillimonas, for which the name Pusillimonas thiosulfatoxidans sp. nov. is proposed. The type strain is YE3T (=KCTC 62737T=NBRC 113113T).

Aptamer-Immobilized Surface Plasmon Resonance Biosensor for Rapid and Sensitive Determination of Virulence Determinant.

Shigella sonnei isolate invasion plasmid antigen protein, IpaH, was successfully expressed in recombinant overexpression bacterial system. The soluble expression IpaH was enhanced with molecular chaperon co-expressed environment. Specific aptamer IpaH17 was isolated through the SELEX process and showed fM binding affinity. IpaH17-SPR biosensor platform was involved to verify the binding sensitivity and specificity. The IpaH concentration dependent IpaH17-SPR sensor response was highly linear with a linear regression constant of 99.4% in the range between 0 and 100 ng/mL. In addition, S. sonnei revealed the specific RU value and detected in a real-time manner within 1 hour. Our study indicated that IpaH17-SPR sensor can allow for rapid, sensitive and specific determination of Shigella sonnei virulent factor.

Detecting and Discriminating Shigella sonnei Using an Aptamer-Based Fluorescent Biosensor Platform.

In this paper, a Whole-Bacteria SELEX (WB-SELEX) strategy was adopted to isolate specific aptamers against Shigella sonnei. Real-time PCR amplification and post-SELEX experiment revealed that the selected aptmers possessed a high binding affinity and specificity for S. sonnei. Of the 21 aptamers tested, the C(t) values of the SS-3 and SS-4 aptamers (Ct = 13.89 and Ct = 12.23, respectively) had the lowest value compared to other aptamer candidates. The SS-3 and SS-4 aptamers also displayed a binding affinity (KD) of 39.32 ± 5.02 nM and 15.89 ± 1.77 nM, respectively. An aptamer-based fluorescent biosensor assay was designed to detect and discriminate S. sonnei cells using a sandwich complex pair of SS-3 and SS-4. The detection of S. sonnei by the aptamer based fluorescent biosensor platform consisted of three elements: (1) 5'amine-SS-4 modification in a 96-well type microtiter plate surface (N-oxysuccinimide, NOS) as capture probes; (2) the incubation with S. sonnei and test microbes in functionalized 96 assay wells in parallel; (3) the readout of fluorescent activity using a Cy5-labeled SS-3 aptamer as the detector. Our platform showed a significant ability to detect and discriminate S. sonnei from other enteric species such as E. coli, Salmonella typhimurium and other Shigella species (S. flexneri, S. boydii). In this study, we demonstrated the feasibility of an aptamer sensor platform to detect S. sonnei in a variety of foods and pave the way for its use in diagnosing shigellosis through multiple, portable designs.