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Published on: October 18, 2019
Reliability of mCP method for identification of Clostridium perfringens from faecal polluted aquatic environments
S R Mueller-Spitz1, L B Stewart, S L McLellan
1Great Lakes WATER Institute, University of Wisconsin-Milwaukee, Milwaukee, WI 53204, USA.
Aims:
The purpose of the work was to evaluate the mCP method to correctly identify and enumerate Clostridium perfringens that are present in surface waters impacted by a mixture of faecal pollution sources.
Methods:
Clostridium perfringens were enumerated and isolated from sewage influent, surface water and suspended sediments using the mCP method. Molecular characterization of isolates was performed using species-specific PCR, along with full-length sequencing of the 16S rRNA gene for a subset of isolates.
Results:
The environmental isolates were presumptively identified as C. perfringens based on utilization of sucrose, inability to ferment cellobiose and a positive action for acid phosphatase activity. All isolates (n = 126) were classified as C. perfringens based on positive results with species-specific PCR with a subset confirmed as C. perfringens based on the 16S rRNA gene identity.
Conclusions:
The molecular results indicated all of the presumptive positive isolates were C. perfringens regardless of the source, e.g. sewage influent or environmental water samples. Sequencing revealed that C. perfringens obtained from sewage and the aquatic environment were nearly identical (c. 99.5% similarity).
Significance And Impact Of The Study:
From this study we conclude that the mCP method is a robust approach to enumerate and isolate C. perfringens from aquatic environments that receive diverse sources of faecal pollution.
Insights
The mCP method reliably identifies and quantifies Clostridium perfringens in diverse aquatic environments. Molecular analysis confirmed all isolates were C. perfringens, regardless of pollution source.
Area of Science:
- Environmental microbiology
- Water quality assessment
- Bacterial identification
Background:
- Clostridium perfringens is an indicator of faecal pollution in water.
- Diverse pollution sources complicate accurate enumeration of C. perfringens.
- Reliable methods are needed to assess water contamination.
Purpose of the Study:
- To evaluate the mCP method for identifying and enumerating Clostridium perfringens.
- To assess the method's efficacy in surface waters with mixed faecal pollution.
- To confirm isolate identity using molecular techniques.
Main Methods:
- Enumeration and isolation of Clostridium perfringens using the mCP method.
- Sampling from sewage influent, surface water, and suspended sediments.
- Molecular characterization including species-specific PCR and 16S rRNA gene sequencing.
Main Results:
- Presumptive identification based on biochemical tests (sucrose fermentation, cellobiose fermentation, acid phosphatase).
- All 126 isolates confirmed as C. perfringens via species-specific PCR.
- 16S rRNA gene sequencing confirmed high similarity (approx. 99.5%) between sewage and environmental isolates.
Conclusions:
- The mCP method is a robust tool for enumerating and isolating C. perfringens.
- Molecular confirmation validates the method across various faecal pollution sources.
- Identical C. perfringens strains found in sewage and aquatic environments.
