Laboratory of Environmental Microbiology
School of Molecular and Theoretical Biology, 2013
Microorganisms play important roles in the environment. In this study we
characterized the composition of the microbial community in the Oka River near
Pushchino. Our particular focus was to evaluate the impact of wastewater influx
on the diversity of microorganisms.
Previously, a study of samples from rivers near Chicago had reported that
wastewater released into the rivers had a detectable impact on the downstream
ecosystems.1,2 This was despite the wastewater had been treated and met quality
standards. Treated wastewater contained higher concentrations of nutrients such
as nitrate and phosphate, which likely caused the observed decrease in bacterial
abundance and diversity in the downstream sediments. This recent study
suggested that wastewater influx caused a homogenization of microbial
communities in rivers, which otherwise had distinct microbial composition.
For our study we have collected water and sediment samples at four sites along
the Oka River in Pushchino: First, a site lacking obvious human impact
(‘Upstream’). Second, a site near wastewater influx (‘Dump’). Third, a control site
about 30 meters upstream the wastewater influx (‘Swamp’). Fourth, a site about
100 meters downstream of the wastewater influx (‘Beach’).
Immediately after sampling we measured several chemical parameters of the
water samples using calorimetric assays (Figure 1). We found that water near the
site of wastewater influx had increased levels of phosphate and nitrate, while
values for the other sites were comparable.
We then went on to determine the abundance of bacteria in the water and
sediment samples by counting the number of colony forming units (CFU) on agar
plates or direct counting of cells using fluorescence microscopy and DAPI
staining. Overall, no statistically significant difference for the abundance of
microorganisms in the four sites was found. Several interesting looking strains
from these plates were isolated and characterized by microscopy, colony PCR of
the 16S gene and measuring growth curves.
Figure 1. Chemical parameters of the water samples collected from Oka River.
Figure 2. Composition of the microbial communities from the four sites in OKA River
based on 16S amplicon sequencing.
Finally, the microbial communities were characterized by metagenomic analysis of
16S gene sequences. For this DNA from soil samples was isolated and used as
template for polymerase chain reaction (PCR) to amplify the V45 region of the
16S gene. The PCR products were purified from primers and other contaminants
on an agarose gel and subsequently used for next-generation sequencing (NGS)
on the Illumina MiSeq platform. The composition of the microbial communities is
illustrated in Figure 2.
Interestingly, the nutrient input into the Oka River showed a slight increase of
bacteria from the phylum chloroflexi and a decrease of oxygenic photosynthesis
(chloroplasts from algae) but otherwise no major changes were detected using a
crude taxonomic clustering. This preliminary analysis suggests that at these
particular sites the wastewater influx has mainly a local effect on the microbial
community in the Oka River.
Further reading:
1.
Drury, B., Rosi-Marshall, E. & Kelly, J. J. Wastewater Treatment Effluent Reduces the
Abundance and Diversity of Benthic Bacterial Communities in Urban and Suburban
Rivers. Appl Environ Microbiol 79, 1897-1905 (2013).
2.
Wigginton, N. S. Downstream Discharge. Science (2013).