ABSTRACT
The literature on microbial source tracking (MST) suggests that DNA analysis of fecal samples leads to more reliable determinations of bacterial sources of surface water contamination than antibiotic resistance analysis (ARA). Our goal is to determine whether the increased reliability, if any, in library-based MST developed with DNA data is sufficient to justify its higher cost, where the bacteria source predictions are used in TMDL surface water management programs. We describe an application of classification trees for MST applied to ARA and DNA data from samples collected in the Potomac River Watershed in Maryland. Conclusions concerning the comparison of ARA and DNA data, although preliminary at the current time, suggest that the added cost of obtaining DNA data in comparison to the cost of ARA data may not be justified, where MST is applied in TMDL surface water management programs.
Subject(s)
Bacterial Typing Techniques/methods , Classification/methods , DNA/analysis , Drug Resistance, Bacterial , Feces/microbiology , Water Pollutants/analysis , Animals , Bacteria/isolation & purification , Bacterial Typing Techniques/economics , Bacterial Typing Techniques/standards , DNA/economics , Humans , Maryland , Models, Statistical , Rivers/microbiology , Sewage/microbiology , Species Specificity , Water MicrobiologyABSTRACT
The triplex PCR of Clermont et al. [Clermont, O., Bonacorsi, S., Bingen, E., 2000. Rapid and simple determination of the Escherichia coli phylogenetic groups. Appl. Environ. Microbiol. 66, 4555-4558.] was used to genotype E. coli isolates from the Mid-Atlantic region of the USA, obtained from freshwater, animal internal organs, and feces. Of 445 isolates subjected to genotyping, 118 isolates (26%) were genotype A, 111 (25%) genotype D, 140 (31%) genotype B1, and 76 (17%) genotype B2. All four genotypes were present in three sets of freshwater stream samples. When isolates from chicken cecal ingesta, cecal mucosa, and tracheal mucosa were screened, there was selective distribution of genotypes in these organs. Genotype D was rarely encountered in feces, milk, and intestinal tissues of dairy cows, while all four genotypes were represented in goose feces. Isolates from the feces of zoo animals reared in the US demonstrated a predominance of genotype B1. Thirty-six of the A isolates in our overall collection were subgenotype A(0), in which none of the three amplicons are observed; confirmation that these isolates were E. coli was done using an ancillary lacZ PCR assay. We conclude that the genotyping triplex PCR assay, used in combination with traditional culture methods, can be useful in categorizing E. coli from environmental and veterinary sources in the Mid-Atlantic region of the USA.
Subject(s)
Escherichia coli Infections/veterinary , Escherichia coli/classification , Escherichia coli/genetics , Fresh Water/microbiology , Animals , Animals, Zoo/microbiology , Cattle , Cecum/microbiology , DNA, Bacterial/genetics , DNA, Ribosomal/genetics , Escherichia coli/isolation & purification , Escherichia coli Infections/microbiology , Feces/microbiology , Geese , Genotype , Intestinal Mucosa/microbiology , Polymerase Chain Reaction/methods , RNA, Ribosomal, 16S/genetics , Respiratory Mucosa/microbiology , United StatesABSTRACT
Various statistical classification methods, including discriminant analysis, logistic regression, and cluster analysis, have been used with antibiotic resistance analysis (ARA) data to construct models for bacterial source tracking (BST). We applied the statistical method known as classification trees to build a model for BST for the Anacostia Watershed in Maryland. Classification trees have more flexibility than other statistical classification approaches based on standard statistical methods to accommodate complex interactions among ARA variables. This article describes the use of classification trees for BST and includes discussion of its principal parameters and features. Anacostia Watershed ARA data are used to illustrate the application of classification trees, and we report the BST results for the watershed.
Subject(s)
Bacteria , Bacterial Typing Techniques , Drug Resistance, Bacterial , Models, Biological , Rivers/microbiology , Water Pollution , Animals , Animals, Domestic/microbiology , Animals, Wild/microbiology , Anti-Bacterial Agents/pharmacology , Bacteria/classification , Bacteria/drug effects , Bacteria/isolation & purification , Humans , MarylandABSTRACT
Spiroplasma sp. strain GNAT3597T was isolated from the biting midge genus Atrichopogon (Diptera: Ceratopogonidae). It was serologically distinct from other Spiroplasma species, groups or subgroups. Dark-field microscopy of the cells revealed the classical helical shape and subsequent transmission electron microscopy revealed cells surrounded by only a cell membrane (i.e. lacking a cell wall). Growth of strain GNAT3597T occurred in M1D medium at 30 degrees C. Strain GNAT3597T catabolized both glucose and arginine, but did not hydrolyse urea. The DNA G+C content of strain GNAT3597T was 29+/-1 mol%. Only one strain, SMCAT (Spiroplasma mirum), is serologically related to strain GNAT3597T, although the relationship is weak (positive reaction to only a 1 : 80 dilution). It is therefore proposed that strain GNAT3597T (=ATCC BAA-520T=NBRC 100390T) represents a novel species, Spiroplasma atrichopogonis sp. nov. (class Mollicutes: order Entomoplasmatales: family Spiroplasmataceae).