ABSTRACT
Clean freshwater lies at the heart of human society and monitoring its quality is paramount. In addition to chemical controls, traditional microbiological water tests focus on the detection of specific bacterial pathogens. The direct tracing of all aquatic DNA poses a more profound alternative. Yet, this has hitherto been underused due to challenges in cost and logistics. Here we present a simple, fast, inexpensive and comprehensive freshwater diagnostics workflow centred around portable nanopore DNA sequencing. Using defined bacterial compositions and spatiotemporal microbiota from surface water of an example river in Cambridge (UK), our study shows how nanopore sequencing can be readily integrated for the assessment of aquatic bacterial diversity and pollution. We provide a computational benchmark that features more than ten taxonomic classification tools to derive guidelines for bacterial DNA analyses with nanopore data. Through complementary physicochemical measurements, we find that nanopore metagenomics can depict fine temporal gradients along the main hydrological axis of an urban-rural interface and yield high-resolution pathogen maps that address concerns of public health.