Abstract
Urban sewer systems consist of wastewater and stormwater sewers, of which only wastewater is processed before being discharged. Occasionally, misconnections or damages in the network occur, resulting in untreated wastewater entering natural water bodies via the stormwater system. Cultivation of faecal indicator bacteria (e.g. Escherichia coli; E. coli) is the current standard for tracing wastewater contamination. This method is cheap but has limited specificity and mobility. Here, we compared the E. coli culturing approach with two sequencing-based methodologies (Illumina MiSeq 16S rRNA gene amplicon sequencing and Oxford Nanopore MinION shotgun metagenomic sequencing), analysing 73 stormwater samples collected in Stockholm. High correlations were obtained between E. coli culturing counts and frequencies of human gut microbiome amplicon sequences, indicating E. coli is indeed a good indicator of faecal contamination. However, the amplicon data further holds information on contamination source or alternatively how much time has elapsed since the faecal matter has entered the system. Shotgun metagenomic sequencing on a subset of the samples using a portable real-time sequencer, MinION, correlated well with the amplicon sequencing data. This study demonstrates the use of DNA sequencing to detect human faecal contamination in stormwater systems and the potential of tracing faecal contamination directly in the field.
Highlights
Many urban areas use separate sewer systems to transport wastewater and stormwater, with the two pipes often buried adjacent to each other
® The E. coli count data generated with the Colilert -18 test varied from
We have compared the performance of the classical E. coli culturing method with two DNA sequencing-based approaches for tracking wastewater contamination in urban stormwater systems
Summary
Many urban areas use separate sewer systems to transport wastewater and stormwater, with the two pipes often buried adjacent to each other. Faecal coliforms (e.g., E. coli) and enterococci have been extensively used as indicators for assessing the contamination level of water samples due to their high prevalence in human faeces, high growth rates, and ease of cultivation[4] They are not in any way perfect because closely related strains, which are hard to distinguish via culturing, exist in the intestines of other animals. Only the Covalently Linked Immunomagnetic Separation/Adenosine Triphosphate (Cov-IMS/ATP) technique can quantify faecal indicator bacteria in the field, though being restricted to the microorganisms E. coli and Enterococcus spp.[20,21] It is less sensitive than the culture-dependent methods and of no avail with regard to the detection of host-specific E. coli or Enterococcus. Its high sequencing error rate has made metagenomic sequencing problematic, but the latest upgrade brought about a drop in its error rate from 38% to approximately 10%, rendering this methodology more attractive[30,31]
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