Development and Application of a Most-Probable-Number–PCR Assay To Quantify Flagellate Populations in Soil Samples
AUTOR(ES)
Fredslund, Line
FONTE
American Society for Microbiology
RESUMO
This paper reports on the first successful molecular detection and quantification of soil protozoa. Quantification of heterotrophic flagellates and naked amoebae in soil has traditionally relied on dilution culturing techniques, followed by most-probable-number (MPN) calculations. Such methods are biased by differences in the culturability of soil protozoa and are unable to quantify specific taxonomic groups, and the results are highly dependent on the choice of media and the skills of the microscopists. Successful detection of protozoa in soil by DNA techniques requires (i) the development and validation of DNA extraction and quantification protocols and (ii) the collection of sufficient sequence data to find specific protozoan 18S ribosomal DNA sequences. This paper describes the development of an MPN-PCR assay for detection of the common soil flagellate Heteromita globosa, using primers targeting a 700-bp sequence of the small-subunit rRNA gene. The method was tested by use of gnotobiotic laboratory microcosms with sterile tar-contaminated soil inoculated with the bacterium Pseudomonas putida OUS82 UCB55 as prey. There was satisfactory overall agreement between H. globosa population estimates obtained by the PCR assay and a conventional MPN assay in the three soils tested.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=92777Documentos Relacionados
- Application of Most-Probable-Number Statistics to Direct Enumeration of Microorganisms
- Use of the Most-Probable-Number Technique To Detect Polymyxa betae (Plasmodiophoromycetes) in Soil
- Application of the Most-Probable-Number Procedure to Suspensions of Leptospira autumnalis Akiyami A
- Dynamics of Soil Denitrifier Populations: Relationships between Enzyme Activity, Most-Probable-Number Counts, and Actual N Gas Loss †
- A Most-Probable-Number Assay for Enumeration of Infectious Cryptosporidium parvum Oocysts