Fecal Source
Tracking in Water by Next-Generation
Sequencing Technologies Using Host-Specific <i>Escherichia coli</i> Genetic
Markers
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Abstract
High levels of fecal
bacteria are a concern for the aquatic environment,
and identifying sources of those bacteria is important for mitigating
fecal pollution and preventing waterborne disease. <i>Escherichia
coli</i> has been used as an indicator of fecal pollution, however
less success has been achieved using this organism for library-independent
microbial source tracking. In this study, using next-generation sequencing
technology we sequenced the whole genomes of 22 <i>E. coli</i> isolates from known sources (9 from humans, 2 from cows, 6 from
pigs, and 5 from chickens) and identified candidate host-specific
genomic regions. Specificity testing on the candidate regions was
performed using 30 <i>E. coli</i> isolates from each source.
Finally, we identified 4 human-, 2 cow-, 3 pig-, and 4 chicken-specific
genetic markers useful for source tracking. We also found that a combination
of multiplex PCR and dual index sequencing is effective for detecting
multiple genetic markers in multiple isolates at one time. This technique
was applied to investigating identified genetic markers in 549 <i>E. coli</i> isolates obtained from the Yamato River, Japan.
Results indicate that humans constitute a major source of water contamination
in the river. However, further work must include isolates obtained
from geographically diverse animal hosts to make this method more
reliable