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Brucella, the etiological agent of brucellosis, is an important zoonosis pathogen worldwide. Brucella infects humans and various domestic and wild animals, and represents a great threat to public health and animal husbandry. In the present study, we developed a real-time recombinase polymerase amplification (RPA) assay for the detection of Brucella. The assay targeted the bcsp31 gene of Brucella, and an RPA exo probe and a pair of primers were selected for assay validation. RPA sensitivity and specificity were evaluated using plasmid standards, Brucella representative strains, and non-Brucella strains. The RPA assay achieved a detection limit of 17 molecules in 95% of cases based on probit analysis, and could successfully distinguish 18 representative Brucella strains (B. abortus biovars 1, 2, 3, 4, 5, 6, 7 and 9, B. melitensis biovars 1, 2 and 3, B. suis biovars 1, 2, 3 and 4, B. canis, B. neotomae and B. ovis), and four Brucella vaccine strains (A19, S19, S2 and M5). A total of 52 Brucella field strains were detected by real-time PCR and RPA in parallel, and compared with real-time PCR, the sensitivity of the RPA assay was 94% (49/52). Thus, this RPA assay may be a rapid, sensitive, and specific tool for the prevention and control of Brucellosis.

Brucellosis is a widespread zoonotic disease caused by Brucella spp. Immunization with attenuated vaccines has proved to be an effective method of prevention; however, it may also interfere with diagnosis. Brucella abortus strain A19, which is homologous to B. abortus strain S19, is widely used for the prevention of bovine brucellosis in China. For effective monitoring of the control of brucellosis, it is essential to distinguish A19 from field strains. Single-nucleotide polymorphism–based assays offer a new approach to such discrimination studies. In the current study, we developed a cycleave PCR assay that successfully distinguished attenuated vaccine strains A19 and S19 from 22 strains of B. abortus and 57 strains of 5 other Brucella species. The assay gave a negative reaction with 4 non- Brucella species. The minimum sensitivity of the assay, evaluated using 10-fold dilutions of chromosomal DNA, was 7.6 fg for the A19 strain and 220 fg for the single non-A19/non-S19 Brucella strain tested ( B. abortus 104M). The assay was also reproducible (intra- and interassay coefficients of variation: 0.003–0.01 and 0.004–0.025, respectively). The cycleave assay gave an A19/S19-specific reaction in 3 out of 125 field serum samples, with the same 3 samples being positive in an alternative A19/S19-specific molecular assay. The cycleave assay gave a total of 102 Brucella-specific reactions (3 being the A19/S19-specific reactions), whereas an alternative Brucella-specific assay gave 92 positive reactions (all also positive in the cycleave assay). Therefore, this assay represents a simple, rapid, sensitive, and specific tool for use in brucellosis control.

Bioelectrochemical system is an emerging technology for the remediation of soils contaminated by petroleum hydrocarbons. However, performance of such systems can be limited by the inefficient mass transport in soil. Here we report a new method of sand amendment, which significantly increases both oxygen and proton transports, resulting to increased soil porosity (from 44.5% to 51.3%), decreased Ohmic resistance (by 46%), and increased charge output (from 2.5 to 3.5Cg(-1)soil). The degradation rates of petroleum hydrocarbons increased by up to 268% in 135d. The degradation of n-alkanes and polycyclic aromatic hydrocarbons with high molecular weight was accelerated, and denaturing gradient gel electrophoresis showed that the microbial community close to the air-cathode was substantially stimulated by the induced current, especially the hydrocarbon degrading bacteria Alcanivorax. The bioelectrochemical stimulation imposed a selective pressure on the microbial community of anodes, including that far from the cathode. These results suggested that sand amendment can be an effective approach for soil conditioning that will enhances the bioelectrochemical removal of hydrocarbons in contaminated soils.

BACKGROUND: Bioelectrochemical remediation of soils contaminated by petroleum hydrocarbons has received increasing attention as an effective approach to remove pollution with simultaneous electricity generation. RESULTS: Here, glucose was added as a co-substrate in this system. The charge output of soil microbial fuel cells (MFCs) was enhanced by 262%, and the total petroleum hydrocarbon degradation rate increased by 200%. According to the increased dehydrogenase and polyphenol oxidase activities, the exogenetic glucose substantially activated hydrocarbon degradation bacteria (such as Alcanivorax). The Shannon–Wiener Index and richness of soil microbial community decreased after glucose addition, indicating that a selective enrichment of specific communities was imposed by glucose. Correlation analysis showed the biodiversity in soil far from the cathode was mainly determined by the concentration of petroleum hydrocarbons, while that in soil close to the cathode was dominated by the current. CONCLUSION: This study demonstrated that an exogenetic carbon source enhanced the bioelectrochemically assisted degradation of aged petroleum hydrocarbons in saline soils, providing an effective approach to remove contaminations from soil in barren areas or extreme environments. © 2015 Society of Chemical Industry Supporting information may be found in the online version of this article.

The electrical conductivity (EC) of soil is generally measured after soil extraction, so this method cannot represent the in situ EC of soil (e.g., EC of soils with different moisture contents) and therefore lacks comparability in some cases. Using a resistance measurement apparatus converted from a configuration of soil microbial fuel cell, the in situ soil EC was evaluated according to the Ohmic resistance (Rs) measured using electrochemical impedance spectroscopy. The EC of soils with moisture content from 9.1% to 37.5% was calculated according to Rs. A significant positive correlation (R2 = 0.896, p < 0.01) between the soil EC and the moisture content was observed, which demonstrated the feasibility of the approach. This new method can not only represent the actual soil EC, but also does not need any pretreatment. Thus it may be used widely in the measurement of the EC for soils and sediments.