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DNA strand

Microbial Response to Crude Oil in Ocean Sediments Using ‘Omic Technologies

Challenge

Microbes are vital to ecosystems; they are critical to essential processes such as oxygen cycling and nitrogen fixation. Through carbon utilization and bioremediation, microbes are instrumental to removal of pollutants from the environment. Microbes from complex environmental samples are difficult, if not impossible, to culture and identify in the laboratory, efforts which consume valuable time and resources while often yielding ambiguous results.

Solution

Battelle's combined metagenomic and metaproteomic analytical approaches demonstrated how microorganisms respond to changes in the environment such as exposure to crude oil. The microbial response to crude oil was investigated using genetic sequence profiling (metagenomics) and functional protein profiling (metaproteomics) to observe perturbations in microbial biodiversity and functional protein content. The results showed an increase in the abundance of microbial hydrocarbon-degrading species upon exposure to crude oil, and increased levels of hydrocarbon-degrading enzymes and other proteins. Battelle’s analytical methods allow for high resolution detection of ecologically important and diverse microbial species, and are powerful tools for application to monitoring environmental changes.

Outcome

  • Systems biology approach incorporating multidisciplinary assays
  • Extensive evaluation of microbial communities with depth previously unattainable
  • Rapid screening of complex environmental samples
  • Unbiased detection relative to culturing in laboratory
  • Important gene and biochemical pathway identification
  • Key protein identification/quantification
  • Discovery tool, capable of detecting the unexpected

Integrating next-generation sequencing (NGS) data with functional protein profiling approaches allows for comprehensive identification of diverse and difficult to culture microbes in environmental samples. The types and frequencies of detection of hydrocarbon-degrading bacteria identified by Battelle scientists using this approach are tabulated below.

Table 1. Frequency of detection of bacterial species in different environmental samples with and without exposure to crude oil. Microbial metagenomics analyses were conducted on sediment samples from two geographically distinct nearshore sites. The samples were then incubated with 10% crude oil and the frequencies of bacterial sequence detection compared. A ten-fold increase in detection of genomic sequences from bacterial species with known hydrocarbon-degrading capabilities was observed in the samples from both locations when the samples were incubated with 10% crude oil [6.9% (no-oil control) to 68% (with oil) in the Site 1 sediment, and 4.3% (no-oil control) to 48% (with oil) in the Site 2 sample]. Interestingly, while the same hydrocarbon degrading species were initially present in both sediment samples, incubation with 10% crude oil led to increased abundance of Methylophaga thiooxidans, Pseudoalteromonas spp., and Marinobacter hydrocarbonoclasticus which were dominant in Sample 2, while Pseudomonas spp. became most abundant in Sample 2. The test illustrates how indigenous microbial communities respond to crude oil exposure in sediments from different geographic locations.

Hydrocarbon- Degrading Genus or Species

% of Total Bacterial Sequences

Site 1

Site 2

0% crude oil

10% crude oil

0% crude oil

10% crude oil

Methlyophaga thiooxydans

0.03%

19%

0.6%

4%

Pseudoalteromonas spp.

0.1%

13%

1%

9%

Marinobacter hydrocarbonoclasticus

0.1%

19%

1%

4%

Pseudomonas spp.

6%

1%

1%

30%

Vibrio spp.

0.4%

8%

0.1%

0.2%

Denitrovibrio acetiphilus

0%

5%

0.04%

0%

Shewanelia spp.

0.3%

3%

0.6%

0.4%

Total

6.9%

68%

4.3%

48%


Visualization

Figure 1.
A 3D image of the protein methylene-tetrahydrofolate dehydrogenase, an oxioreductase, which was only identified from oi-treatment sediments. This enzyme is involved in the degradation of organic materials.