Diversity and population structure of sewage-derived microorganisms in wastewater treatment plant influent

Abstract

The release of untreated sewage introduces non-indigenous microbial populations of uncertain composition into surface waters. We used massively parallel 454 pyrosequencing of hypervariable regions in rRNA genes to profile microbial communities from eight untreated sewage influent samples of two wastewater treatment plants (WWTPs) in metropolitan Milwaukee. The sewage profiles included a discernible human faecal signature made up of several taxonomic groups including multiple Bifidobacteriaceae, Coriobacteriaceae, Bacteroidaceae, Lachnospiraceae and Ruminococcaceae genera. The faecal signature made up a small fraction of the taxa present in sewage but the relative abundance of these sequence tags mirrored the population structures of human faecal samples. These genera were much more prevalent in the sewage influent than standard indicators species. High-abundance sequences from taxonomic groups within the Beta- and Gammaproteobacteria dominated the sewage samples but occurred at very low levels in faecal and surface water samples, suggesting that these organisms proliferate within the sewer system. Samples from Jones Island (JI--servicing residential plus a combined sewer system) and South Shore (SS--servicing a residential area) WWTPs had very consistent community profiles, with greater similarity between WWTPs on a given collection day than the same plant collected on different days. Rainfall increased influent flows at SS and JI WWTPs, and this corresponded to greater diversity in the community at both plants. Overall, the sewer system appears to be a defined environment with both infiltration of rainwater and stormwater inputs modulating community composition. Microbial sewage communities represent a combination of inputs from human faecal microbes and enrichment of specific microbes from the environment to form a unique population structure.

Fig. 1

(A) Sewage, (B) human, and (C) surface water taxonomic composition by class. Clostridia is expanded to the order level, with Clostridiales further expanded to the family level. Taxa represented occur at ≥ 1% abundance in at least one sample.

Figure 2

Rarefaction curves of OTUs defined by 3% and 6% sequence variation in sewage, human, and surface water samples. The 6% curves of sewage and surface water overlap.

Figure 3

Venn diagram of the overlap of OTUs from sewage, human, and surface water. OTUs represent clusters with 3% sequence variation. Within a random sub-sampling of 38,368 tag sequences from the total datasets, sewage (S) contained 3045 OTUs, human (H) contained 1950 OTUs, and surface water (W) contained 2833 OTUs. The numbers of overlapping OTUs were as follows: S vs. H = 611, S vs. W = 479, H vs. W = 27 and S vs. H vs. W = 20.

Fig. 4

Histogram of Bacteroidetes (A), and Firmicutes (B and C) tags in humans compared with surface water (designated Junction) and eight WWTP samples. The x-axis represents the ordination of sequence tags: sorted first alphabetically by taxomonic name then by decreasing tag frequency. The major taxonomic designations are labeled along the human sample.

Fig. 5

UPGMA tree based on Bray-Curtis distances of eight sewage samples, surface water, and human datasets (Dethlefsen et al., 2008 and Turnbaugh et al., 2009). Sewage samples clustered by collection date and were distinct from surface water and human fecal samples.