HPB-Chip: An accurate high-throughput qPCR-based tool for rapidly profiling waterborne human pathogenic bacteria in the environment.

Waterborne pathogens are threatening public health globally, but profiling multiple human pathogenic bacteria (HPBs) in various polluted environments is still a challenge due to the absence of rapid, high-throughput and accurate quantification tools. This work developed a novel chip, termed the HPB-Chip, based on high-throughput quantitative polymerase chain reactions (HT-qPCR). The HPB-Chip with 33-nL reaction volume could simultaneously complete 10,752 amplification reactions, quantifying 27 HPBs in up to 192 samples with two technical replicates (including those for generating standard curves). Specific positive bands of target genes across different species and single peak melting curves demonstrated high specificity of the HPB-Chip. The mixed plasmid serial dilution test validated its high sensitivity with the limit of quantification (LoD) of averaged 82 copies per reaction for 25 target genes. PCR amplification efficiencies and R2 coefficients of standard curves of the HPB-Chip averaged 101 % and 0.996, respectively. Moreover, a strong positive correlation (Pearson' r: 0.961-0.994, P < 0.001) of HPB concentrations (log10 copies/L) between HPB-Chip and conventional qPCR demonstrated high accuracy of the HPB-Chip. Subsequently, the HPB-Chip has been successfully applied to absolutely quantify 27 HPBs in municipal and hospital wastewater treatment plants (WWTPs) after PMA treatment. A total of 17 HPBs were detected in the 6 full-scale WWTPs, with an additional 19 in the hospital WWTP. Remarkably, Acinetobacter baumannii, Legionella pneumophila, and Arcobacter butzler were present in the final effluent of each municipal WWTP. Overall, the HPB-Chip is an efficient and accurate high-throughput quantification tool to comprehensively and rapidly quantify 27 HPBs in the environment.

Cultivation of Chlorella vulgaris in a pilot-scale photobioreactor using real centrate wastewater with waste glycerol for improving microalgae biomass production and wastewater nutrients removal.

To improve nutrients removal from real centrate wastewater and enhance the microalgae biomass production, cultivation of Chlorella vulgaris in lab and a pilot-scale photobioreactor with waste glycerol was studied. The results showed the optimal concentration of the crude glycerol was 1.0gL-1 with the maximum biomass productivity of 460mgL-1d-1 TVS, the maximum lipid content of 27%, the nutrient removal efficiency of all above 86%, due to more balanced C/N ratio. The synergistic relationship between the wastewater-borne bacteria and the microalgae had significant good influence on nutrient removal. In pilot-scale wastewater-based algae cultivation, with 1gL-1 waste glycerol addition, the average biomass production of 16.7gm-2d-1, lipid content of 23.6%, and the removal of 2.4gm-2d-1 NH4+-N, 2.7gm-2d-1 total nitrogen, 3.0gm-2d-1 total phosphorous, and 103.0gm-2d-1 of COD were attained for 34days semi-continuous mode.