Application of culture-independent methods to assess the bacteria removal efficiency of subsurface flow constructed wetlands

The bacteriologic treatment efficiency of vertical and horizontal subsurface flow constructed wetlands (SFCWs) was analysed in two multistage wastewater treatment systems by culture dependent and independent methods. When assessed with standard cultivation procedures, bacteria removal efficiency of the vertical and horizontal SFCWs was similar. However, microscopic enumerations of the wastewater bacteria after DNA staining revealed a completely different removal pattern: bacteria removal efficiency of the horizontal SFCWs was in general low and erratic, whereas the vertical SFCWs displayed high bacteria removal rates. The discrepancies in the results obtained by bacteria enumeration and cultivation was due to a strong decrease in bacterial culturability after treatment by the horizontal SFCWs, leading to overestimation of the real bacterial concentrations in these effluents. Additionally, a PCR based approach for the detection of the enteropathogenic bacteria Campylobacter jejuni and Yersinia enterocolitica was tested in the wastewater samples. The methods were specific and reproducible in the analysed samples and could be carried out within 12 h, proving very adequate as an alternative to cultivation. This work recommends a review of the current standard methodology for wastewater quality surveillance, as well as of the design of SFCW.