Recommendations for microbial source tracking: Lessons from a methods comparison study

Abstract

The methods comparison study described in accompanying manuscripts demonstrated the potential value of microbial source tracking (MST) techniques, but also identified a need for method refinement. This paper provides three classes of recommendations to improve MST technology: optimization, development and evaluation. Optimization recommendations focus on library-dependent methods and include improved selection of restriction enzymes or antibiotics, better definition of appropriate library size, selection of target species and choice of statistical pattern-matching algorithms. Methods development recommendations focus on identifying new genomic targets and quantification procedures for library-independent methods. Longer-term methods development recommendations include integration of microarrays and other direct pathogen detection technology with MST. Studies defining host specificity and population dynamics should aid selection of target species during methods development. Evaluation recommendations include enhancements that should be incorporated into future methods comparison studies, along with studies to assess the value of MST results for risk characterization.

Detection of Genetic Markers of Fecal Indicator Bacteria in Lake Michigan and Determination of Their Relationship to Escherichia coli Densities Using Standard Microbiological Methods

“…appear promising, since these markers are a sensitive measure of fecal pollution, are relatively simple to detect, and have been shown to be host specific (10,11). Further work is necessary to assess the informational value of detecting these markers in the environment and to determine their association with pollution sources (8,11,24,32,34). Extensive field testing will be necessary to determine the geographical distribution of source-specific genetic markers and their correlation with standard fecal indicators (2).…”

“…Zoonotic reservoirs (including cattle, swine, and chickens) of pathogens are also of high concern (24). There are a wide variety of microbial-source tracking approaches under development, and a single approach may not be adequate, since all these methods have limitations (30,31,34). Culture-independent, molecular methods detect genetic targets of organisms found in a specific host.…”

Method for Isolation of Bacteroides Bacteriophage Host Strains Suitable for Tracking Sources of Fecal Pollution in Water

“…In contrast, other strains, such as RYC 2056, detect similar numbers of phages in different geographical areas but do not discriminate between the sources of fecal pollution (5,7,18,22). Strains tested in the United States to date appear to behave like RYC 2056 (15).Limitations of existing source tracking methods (19,24,25,26,27), combined with the good source tracking performance of strain HSP40 in certain geographical areas (4,9,12,28,30), along with increasing information about the specificity between the animal host and the bacteria of the Bacteroides group (11, 32) and the narrow host ranges reported for phages infecting Bacteroides (6,8,16,22,30), prompted our search for new Bacteroides host strains.We describe here a rapid method for isolating and further testing Bacteroides host strains potentially useful for source tracking.Isolation of new hosts for phages infecting Bacteroides. Four trials for isolation of Bacteroides strains from raw municipal sewage from Spain (two trials), Colombia (one trial), and the United Kingdom (one trial) were carried out by two independent operators.…”

Microbial Source Tracking in a Watershed Dominated by Swine

“…The ARA approach is based on the premise that fecal indicator bacteria (FIB) from hosts exposed to antibiotics will develop resistance to those antibiotics, and on the hypothesis that this selective pressure would be a mechanism for discriminating among fecal indicator bacteria from a variety of hosts [30][31][32][33][34][35]. A library or database composed of antibiotic resistance profiles (ARP) of FIB from known host sources is developed and serves as a point of reference for identifying unknown source isolates from water sources.…”

“…Two source tracking methods comparison studies reported issues with accuracy for numerous source tracking methods, including ARA. However, both studies also described approaches that could compensate for potential problems [35,36]. Such approaches include the application of cross-validation analysis of known-source isolates as well as the removal of clonal isolates to ensure a reliable library.…”