New methodological strategies for detecting bacterial indicators

The development of monitoring strategies for the early warning of seawater pollution, with particular reference to faecal and hydrocarbon contamination, has been the specific goal of research carried out within the Cluster 10-SAM (Advanced Systems for Coastal Marine Monitoring) Project, funded by the Italian Ministry for University and Scientific Research. Advanced analytical approaches have been designed and applied to detect bacterial species that have been selected as potential indicators of pollution in seawater samples. We report the results obtained using the fluorescent antibody and enzymatic assays for the detection of Escherichia coli, and a real-time PCR protocol for monitoring marine areas for hydrocarbon pollution. Immunofluorescence and enzymatic methods revealed the occurrence of different faecal pollution levels, reaching 105 E. coli cells 100 ml−1 in the Straits of Messina. Real-time PCR results corresponded to the different degree of oil pollution of the analysed samples. The specificity and speed make these methods promising for the detection and evaluation of marine pollution.

[1]  Mark W. LeChevallier,et al.  Using a Conceptual Framework for Assessing Risks to Health From Microbes in Drinking Water , 1993 .

[2]  S. Vesper,et al.  Evaluation of a Rapid, Quantitative Real-Time PCR Method for Enumeration of Pathogenic Candida Cells in Water , 2003, Applied and Environmental Microbiology.

[3]  J. T. Staley,et al.  Cycloclasticus pugetii gen. nov., sp. nov., an aromatic hydrocarbon-degrading bacterium from marine sediments. , 1995, International journal of systematic bacteriology.

[4]  Gabriella Caruso,et al.  Use of the indirect immunofluorescence method for detection and enumeration of Escherichia coli in seawater samples , 2000, Letters in applied microbiology.

[5]  C. Martín,et al.  Development of a PCR-Based Line Probe Assay for Identification of Fungal Pathogens , 2000, Journal of Clinical Microbiology.

[6]  Hans W. Paerl,et al.  Coastal eutrophication and harmful algal blooms: Importance of atmospheric deposition and groundwater as “new” nitrogen and other nutrient sources , 1997 .

[7]  S. Belkin,et al.  Survival of enteric bacteria in seawater. , 2001, FEMS microbiology reviews.

[8]  Gabriella Caruso,et al.  Combined fluorescent antibody assay and viability staining for the assessment of the physiological states of Escherichia coli in seawaters , 2003, Journal of applied microbiology.

[9]  S. Edberg,et al.  Escherichia coli: the best biological drinking water indicator for public health protection , 2000, Symposium series.

[10]  Thomas D. Schmittgen,et al.  Real-Time Quantitative PCR , 2002 .

[11]  A. Farnleitner,et al.  Rapid enzymatic detection of Escherichia coli contamination in polluted river water , 2001, Letters in applied microbiology.

[12]  Gabriella Caruso,et al.  IMMUNOFLUORESCENCE DETECTION OF ESCHERICHIA COLI IN SEAWATER: A COMPARISON OF VARIOUS COMMERCIAL ANTISERA , 2002, Journal of immunoassay & immunochemistry.

[13]  F. W. Gilcreas,et al.  Standard methods for the examination of water and waste water. , 1966, American journal of public health and the nation's health.

[14]  Nicholas J. Ashbolt,et al.  Indicators of microbial water quality , 2001 .

[15]  A. Huang,et al.  Rapid Identification of Yeasts in Positive Blood Cultures by a Multiplex PCR Method , 2001, Journal of Clinical Microbiology.

[16]  K. Porter,et al.  The use of DAPI for identifying and counting aquatic microflora1 , 1980 .

[17]  Darrell P. Chandler,et al.  Application of the 5′ Fluorogenic Exonuclease Assay (TaqMan) for Quantitative Ribosomal DNA and rRNA Analysis in Sediments , 2001, Applied and Environmental Microbiology.

[18]  M. Levin,et al.  Relationship of microbial indicators to health effects at marine bathing beaches. , 1979, American journal of public health.

[19]  R. Colwell,et al.  A Global Decline in Microbiological Safety of Water: A Call for Action , 1995 .

[20]  A. E. Greenberg,et al.  Standard methods for the examination of water and wastewater : supplement to the sixteenth edition , 1988 .

[21]  Gordon A. McFeters,et al.  Sensitive Detection of Escherichia coliO157:H7 in Food and Water by Immunomagnetic Separation and Solid-Phase Laser Cytometry , 1999, Applied and Environmental Microbiology.

[22]  J. J. Borrego,et al.  An intercalibration study of the use of 4-methylumbelliferyl-β-D-glucuronide for the specific enumeration of Escherichia coli in seawater and marine sediments , 1991 .

[23]  R. Colwell,et al.  Retention of enteropathogenicity by viable but nonculturable Escherichia coli exposed to seawater and sunlight , 1996, Applied and environmental microbiology.

[24]  G. Roberts,et al.  Molecular probes for diagnosis of fungal infections , 1995, Journal of clinical microbiology.

[25]  J. Tiedje,et al.  DNA recovery from soils of diverse composition , 1996, Applied and environmental microbiology.

[26]  M. Troussellier,et al.  Assessing terminal restriction fragment length polymorphism suitability for the description of bacterial community structure and dynamics in hydrocarbon-polluted marine environments. , 2005, Environmental microbiology.

[27]  K. Horikoshi,et al.  Rapid Detection and Quantification of Members of the Archaeal Community by Quantitative PCR Using Fluorogenic Probes , 2000, Applied and Environmental Microbiology.

[28]  M. Pace,et al.  Human influence on river nitrogen , 1991, Nature.

[29]  H. Mooney,et al.  Human Domination of Earth’s Ecosystems , 1997, Renewable Energy.

[30]  J. Watkins,et al.  Conventional culture for water quality assessment: is there a future? , 1998, Journal of applied microbiology.

[31]  E. Delong,et al.  Quantitative Analysis of Small-Subunit rRNA Genes in Mixed Microbial Populations via 5′-Nuclease Assays , 2000, Applied and Environmental Microbiology.

[32]  R. Colwell,et al.  Metabolic activity of bacterial cells enumerated by direct viable count , 1987, Applied and environmental microbiology.

[33]  K. Timmis,et al.  Thalassolituus oleivorans gen. nov., sp. nov., a novel marine bacterium that obligately utilizes hydrocarbons. , 2004, International journal of systematic and evolutionary microbiology.

[34]  Gabriella Caruso,et al.  Development of an enzyme assay for rapid assessment of Escherichia coli in seawaters , 2002, Journal of applied microbiology.