DNA adsorbs and binds on clay minerals, which provides protection to the DNA against degradation by nucleases but does not eliminate the ability of bound DNA to transform cells. These observations support the concept that ‘cryptic genes’ can persist in the environment when bound on particles and that the genes could subsequently be expressed if an appropriate host was transformed. The polymerase chain reaction (PCR) was used to amplify free and bound DNA from Bacillus subtilis and calf thymus. DNA bound on montmorillonite, but not on kaolinite, was amplified. However, amplification occurred when kaolinite was pretreated with sodium metaphosphate. DNA was not released from the clays during the amplification procedure. The type of clay (e.g. its structure and charges) affected amplification. Because DNA bound on clay is protected against biodegradation, the ability to amplify DNA bound on clay by the PCR has palaeontological, archaeological, and anthropological implications for the detection of ‘ancient’ DNA, as well as for monitoring the persistence of recombinant DNA introduced to the environment in genetically modified organisms.
The solid-phase PCR (SP-PCR) was compared with a culture-based technique for the detection of aerosolized Escherichia coli DH1. Results with SP-PCR showed an increase in detection sensitivity over that of culture methods. Therefore, SP-PCR may be useful for the detection of airborne microorganisms which may be nonculturable because of aerosolization or sampling stress.
The techniques in current use for detection of pathogens in environmental samples are restricted to those organisms whose replication in either culture media or cell culture is feasible. These methods lack the selectivity and sensitivity necessary for their unequivocal detection and identification. We have developed an assay for the detection of bacterial cells in large volumes of water. Low concentrations of cells containing target sequences were concentrated on membrane filters and were subjected to amplification directly using a stepwise polymerase chain reaction. This procedure, together with nucleic acid probes, has enhanced the limit of detection to the level of a single bacterial cell. This technique could be used for the detection of any bacteria or virus in water or air.
In this study, the survival of genetically engineered microorganisms (GEMs) and their interactions with the environmental microbiota of a tropical river was investigated. Diffusion chambers were used for the in situ survival experiments with a nonplasmid containing Escherichia coli DH7 strain and two model GEMs, E. coli JM103 containing a 2.6 kilobase plasmid (pUC9) and E. coli DH1 with a 4.8 kb plasmid (pWTAla5'). Pure culture survival studies indicated that after a week in the environment a 1.0 log,, decrease in bacterial numbers occurred for both E. coli DH1, while a 3.0 log,, reduction was observed for E. coli JM103. However, a reduction of 4.0 log,, was observed for the E. coli DH1 (pWTAla5') when placed in a chamber conjointly with the resident microbiota. The data suggest that the presence of a plasmid makes no difference on the survival time of GEMs, whereas the presence of competing bacteria is ultimately what limits the survival time of GEMs in the environment. 0 7996 by John Wiley & Sons, Inc.
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