Comparison of DNA-fingerprinting (T-RFLP) and high-throughput sequencing (HTS) to assess the diversity and composition of microbial communities in groundwater ecosystems

Abstract

The majority of usable freshwater is stored as groundwater in the subsurface. Pristine groundwater ecosystems are characterised as oligotrophic environments which facilitate low energy yield, activity, growth, and reproduction for numerous and highly adapted organisms living in these environments. Degradation of groundwater quality and quantity are hence reflected in the structural changes of groundwater species communities. Despite an increasing awareness of this problem, current assessment methods for groundwater ecosystems are solely based on the analysis of abiotic parameters. However, this approach is insufficient to detect changes in microbial communities and their related metabolic functions. In recent years, the development of culture-independent molecular techniques to analyse microbes has vastly improved our knowledge concerning the diversity and composition of microbial communities in various environments. High-throughput sequencing (HTS) techniques enable the detection of single bacterial species in a sample and thus provide a high resolution of the composition and diversity of microbial communities in various environments. Furthermore, the taxonomic information obtained allows for the inference of metabolic functions of a given community. However, since the method is labour intensive and costly it is not necessarily the method of choice for analysing numerous samples. By comparison, DNA-fingerprinting is a less elaborate and inexpensive method that is able to detect changes in microbial communities, although identification of species present in a community is not possible, and therefore represents a valuable supplement to HTS. The present paper intends to render information about the applicability of this method as a monitoring tool against this background, by directly comparing results of DNA-fingerprinting with the results of HTS. Despite the fact that the analysis of bacterial communities using HTS captured significantly higher diversity estimates in our study, results of both methods were positively associated. And even though HTS produced more accurate and detailed results regarding composition and diversity of bacterial communities, patterns of community composition captured by DNA-fingerprinting were similar in comparison to HTS. We thus can suggest DNA-fingerprinting as a cost efficient alternative for community assessment and diversity estimation, specifically as a promising methodological approach in environmental assays.