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Correlation between quantitative PCR and Culture-Based methods for measuring Enterococcus spp. over various temporal scales at three California marine beaches
Citation:
Converse, R. R., J. F. Griffith, R. T. Noble, R. A. HAUGLAND, K. C. Schiff, AND S. B. Weisberg. Correlation between quantitative PCR and Culture-Based methods for measuring Enterococcus spp. over various temporal scales at three California marine beaches. APPLIED AND ENVIRONMENTAL MICROBIOLOGY. American Society for Microbiology, Washington, DC, 78(4):1237-1242, (2012).
Impact/Purpose:
1) Develop and evaluate qPCR assays and test methods for the detection and quantification of genetic markers from indicator bacteria that are associated with human fecal waste and from two new groups of general fecal indicator bacteria (E. coli and Clostridia) that historically have been widely used or are favored in specific regions 2) Determine the occurrence and densities of genetic markers detected by new qPCR assays developed under objective 1 and compare with occurrence and densities of genetic markers detected by previously developed qPCR assays for enterococci and total Bacterioidalesin waste waters and fecal material from different animal sources. 3) Determine stability of fecal indicator bacteria target DNA sequences in freezer archived filter retentates of ambient surface water samples 4) Determine the densities of human and general fecal indicator markers in a wide range of surface and recreational waters including archived samples from previous NEEAR studies.
Description:
Several studies have examined how fecal indicator bacteria (FIB) measurements compare between quantitative polymerase chain reaction (QPCR) and the culture methods it is intended to replace. Here we extend those studies by examining the stability of that relationship within a beach, as affected by time of day and seasonal variations in source. Enterococcus spp. were quantified at three southern California beaches in the morning and afternoon using two QPCR assays, membrane filtration, and defined-substrate testing. While QPCR and culture-based measurements were consistently and significantly correlated, strength of the correlation varied both among and within beaches. Correlations were higher in the morning (0.45<ρ<0.74) than in the afternoon (0.18< ρ <0.45) and higher when the fecal contamination was concentrated (0.38< ρ <0.83) than when it was diffuse (0.19< ρ <0.34). The ratios between culture-based and QPCR results (colony forming units (CFU) or most probable number (MPN) per calibrator cell equivalents (CCE)) also varied spatially and temporally. Ratios ranged between 0.04 and 0.85 CFU or MPN per CCE, and were lowest at the beach affected by diffuse pollution. Patterns in the ratios over the course of the day were dissimilar across beaches, increasing with time at one beach and decreasing at another. The spatial and temporal variability we observed indicate that the empirical relationship between culture-based and QPCR results is not universal, even within a beach.
