We compared the effectiveness of three PCR protocols for the detection of Bifidobacterium adolescentis and one PCR protocol for detecting Bacteroidales as indicators of human fecal pollution in environmental samples. Quantitative PCR indicated that a higher concentration of B. adolescentis DNA was recovered from sewage samples on the 0.2？ m filters compared to the 0.45？ m filters, and there was no evidence of qPCR inhibitors in the DNA extracts. With the Matsuki method (1999),B. adolescentis was detected only in undiluted sewage samples. The King method (2007) performed well and detected B. adolescentis in all of the sewage dilutions (from undiluted to ). In contrast, the Bonjoch approach (2004) was effective at detecting B. adolescentis at lower dilutions ( ) of sewage samples and it gave false positive results with some (3/8) pig fecal samples. Human-specific Bacteroidales (HuBacs) were detected in the lower diluents of sewage samples but was positive in pig (6/8) and cattle fecal samples. PCR detection of B. adolescentis in marine samples from Puerto Rico and freshwater samples from Georgia indicated that the PCR method of King et al. (2007) and the modified Layton method for HuBac were in agreement in detecting human fecal pollution in most sites. 1. Introduction Fecal contamination can degrade the water quality in estuaries, beaches, lakes, and rivers to such an extent that these environments may become impaired for recreational, agricultural, and industrial uses. A major concern for resource managers is to determine the source of fecal pollution in order to apply appropriate corrective measures. In recent years, several molecular PCR-dependent approaches have been developed and used for detecting diagnostic sequences of the 16S rRNA gene of human fecal indicator bacteria as a marker for human fecal pollution. Many researchers use the amplicons from Bifidobacteria and Bacteroidales as molecular markers for indicating the presence of human fecal pollution [1–4]. While some studies have used the molecular detection of Bifidobacterium adolescentis to indicate the presence of human fecal pollution in environmental samples [1, 2, 4, 5], other researchers have relied on the detection of human-associated Bacteroides (HuBac) as a marker of human fecal pollution [6–8]. Currently, there are conflicting reports on which fecal bacterial group provides the most reliable marker for the presence of human fecal pollution in the environment. In addition, the lack of uniform methods of DNA extraction from environmental samples has added to the inconsistencies among
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