This study evaluated the ability of a dextranase from a marine bacterium Catenovulum sp. (Cadex) to impede formation of Streptococcus mutans biofilms, a primary pathogen of dental caries, one of the most common human infectious diseases. Cadex was purified 29.6-fold and had a specific activity of 2309 U/mg protein and molecular weight of 75 kDa. Cadex showed maximum activity at pH 8.0 and 40 °C and was stable at temperatures under 30 °C and at pH ranging from 5.0 to 11.0. A metal ion and chemical dependency study showed that Mn2+ and Sr2+ exerted positive effects on Cadex, whereas Cu2+, Fe3+, Zn2+, Cd2+, Ni2+, and Co2+ functioned as inhibitors. Several teeth rinsing product reagents, including carboxybenzene, ethanol, sodium fluoride, and xylitol were found to have no effects on Cadex activity. A substrate specificity study showed that Cadex specifically cleaved the α-1,6 glycosidic bond. Thin layer chromatogram and high-performance liquid chromatography indicated that the main hydrolysis products were isomaltoogligosaccharides. Crystal violet staining and scanning electron microscopy showed that Cadex impeded the formation of S. mutans biofilm to some extent. In conclusion, Cadex from a marine bacterium was shown to be an alkaline and cold-adapted endo-type dextranase suitable for development of a novel marine agent for the treatment of dental caries.

Enhancing the limit of detection (LOD) is significant for crucial diseases. Cancer development could take more than 10 years, from one mutant cell to a visible tumor. Early diagnosis facilitates more effective treatment and leads to higher survival rate for cancer patients. Rolling circle amplification (RCA) is a simple and efficient isothermal enzymatic process that utilizes nuclease to generate long single stranded DNA (ssDNA) or RNA. The functional nucleic acid unit (aptamer, DNAzyme) could be replicated hundreds of times in a short period, and a lower LOD could be achieved if those units are combined with an enzymatic reaction, Surface Plasmon Resonance, electrochemical, or fluorescence detection, and other different kinds of biosensor. Multifarious RCA-based platforms have been developed to detect a variety of targets including DNA, RNA, SNP, proteins, pathogens, cytokines, micromolecules, and diseased cells. In this review, improvements in using the RCA technique for medical biosensors and biomedical applications were summarized and future trends in related research fields described.

The crystal structure of Dextranase from the marine bacterium Arthrobacter oxidans KQ11 (Aodex) was determined at a resolution of 1.4 Å. The crystal structure of the conserved Aodex fragment (Ala52−Thr638) consisted of an Nterminal domain N and a C-terminal domain C. The N-terminal domain N was identified as a β-sandwich, connected to a righthanded parallel β-helix at the C-terminus. Sequence comparisons, cavity regions, and key residues of the catalytic domain analysis all suggested that the Aodex was an inverting enzyme, and the catalytic acid and base were Asp439 and Asp420, respectively. Asp440 was not a general base in the Aodex catalytic domain, and Asp396 in Dex49A may not be a general base in the catalytic domain. The thermostability of the S357F mutant using semirational design based on B-factors was clearly better than that of wild-type Aodex. This process may promote the aromatic−aromatic interactions that increase the thermostability of mutant Phe357.

Vibrio anguillarum is a bacterial pathogen that causes serious damage to aquatic fish, and its rapid detection and prevention are critical. DNAzymes are DNA-based catalysts with excellent stability. In this study, in vitro selection of DNAzymes was performed using the crude extracellular matrix (CEM) of V. Anguillarum as the target. Different from previous selections targeting bacterial CEM, this work used an unmodified DNA library, allowing easier adoption of the technology. After seven rounds of selection, a DNAzyme named VAE-2 with high activity and specificity was obtained. It showed the highest activity toward V. Anguillarum among eight types of tested bacterial strains. Polyvalent metal ions are needed for its activity. Protease treatment of the CEM and filtration studies indicated that the target is a protein with a molecular weight between 50 k and 100 k Da. A fluorescent biosensor was designed for V. anguillarum with a detection limit down to 4000 cfu/mL, and detection was demonstrated for real fish tissue and feeding water samples. Being the first work of DNAzyme-based sensing of aquatic bacteria, this study indicates that unmodified DNA can be used for targeting bacterial CEM, and it provides a new framework for developing other RNAcleaving DNAzymes for rapid detection of pathogenic bacteria and water pollution.

Vibrio vulnificus is an important bacterial pathogen that causes serious infections in fish and is also highly pathogenic to humans. Many effective detection methods targeting this pathogen have previously been designed, but many of these methods are time‐consuming, complicated and expensive. Thus, these approaches cannot be widely used by small aqacultural concerns. Although DNA aptamers have been used to detect pathogenic bacteria, these have not been applied to marine bacteria, including V. vulnificus. Therefore, we developed a highly specific DNA aptamer for V. vulnificus detection using systematic evolution of ligands by exponential enrichment (SELEX), coupled with asymmetric PCR. After 13 rounds of cross‐selection, we identified a novel DNA aptamer (Vapt2). We evaluated the affinity, specificity and limit of detection (LOD) of this aptamer for V. vulnificus. We found that Vapt2 had a high affinity for V. vulnificus (Kd = 26.8 ± 5.3 nM) and detected this pathogen at a wide range of concentrations (8–2.0 × 108 cfu/ml). Vapt2 bound to V. vulnificus with high selectivity in the presence of other pathogenic bacteria. Our study increases our knowledge of the possible applications of aptamers with respect to marine bacteria. Moreover, our work might provide a framework for the rapid detection of pathogenic bacteria and water pollution.