Environmental Microbiology: Current Technology and Water Applications
"very helpful to researchers" (SciTech Book News)
"a valuable resource for many years to come" (Microbiol. Today)
"comprehensive and useful" (Quar. Rev. Biol.)
Caister Academic Press
Keya Sen and Nicholas J. Ashbolt
US EPA, 26 West Martin Luther King Drive, Cincinnati, OH 45268, USA
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With contributions from a broad range of leading researchers this book focuses on current technology and its applications. Although aimed primarily at research scientists and graduate students in water microbiology, the topics and techniques are equally applicable to all branches of environmental microbiology. The initial chapters cover the concentration, detection and characterization of microbes in drinking water, other chapters are technology focused and cover topics such as geochips and microarrays and their applications, Raman microspectroscopy and related single cell techniques, the use of amoebae hosts, bacteria and bacteriophage as bioreporters, viability of detected microbes and fecal source tracking.
Authors have included comparative tables and figures that address detection sensitivity, specificity, ease of use and other criteria. The value of the book emanates from its explanation of the principles of techniques and the comparison of related techniques. In addition the reader is introduced to key references and important web sites where further details of the technology can be found.
An essential book for water microbiologists, environmental microbiologists and regulators and recommended reading for all microbiologists and environmental scientists.
"For researchers and graduate students in environmental microbiology, related scientists, and regulators ... The mix of conventional and novel techniques should be very helpful to researchers." from SciTech Book News
"Both the content and the quality of the writing exceeded my expectations ... carefully written and explained ... a valuable resource for many years to come ... an excellent resource for senior undergraduates, researchers and academics" from Microbiology Today
"This volume provides a good overview of how newer techniques are being used to study environmental microbial populations, primarily in water. It is a very useful starting point for those who are looking for an introduction to some of the methods or need to come up to speed on developments over the last decade or so ... The chapters are very well referenced ... it provides quite a comprehensive and useful look at the applications of a range of methodologies to aquatic microbiology in recent years. " from The Quarterly Review of Biology (2011) 86: 354-355.
Table of contents
1. The Needle in a Haystack: Detection of Microbes in Source and Drinking Water by Molecular Methods
Molecular techniques based on genomics, proteomics and transcriptomics are rapidly growing as complete microbial genome sequences are becoming available, and advances are made in sequencing technology, analytical biochemistry, microfluidics and data analysis. While the clinical and food industries are increasingly adapting these techniques, there appear to be major challenges in detecting health-related microbes in source and treated drinking waters. This is due in part to the low density of pathogens in water, necessitating significant processing of large volume samples. In this chapter, the state-of-the-art techniques available for pathogen detection and characterization from water are discussed. From the vast panorama of available techniques, only those that are finding a place in the water industry are presented. Quantitative PCR is the prime focus of this chapter, along with protein detection and immunological approaches, with other molecular techniques such as loop-mediated isothermal amplification (LAMP) and microarrays being the focus of other chapters. A detailed section on future trends is also included.
2. Taking the Hay Out of the Haystack: Collecting and Processing Water Samples
H. D. Alan Lindquist
Methods for the detection of specific microorganisms in drinking or source water invariably involve the collection of a water sample, and some treatment to reduce the volume of that sample. Microbial concentration is one of the defining steps in a complete method for the detection of microorganisms in water samples. Numerous approaches have been developed for this task, including filtration, sedimentation, and attraction of the microorganisms of interest to various molecules or substrates. Once the sample volume has been reduced, it is imperative to remove some of the non-biological elements of the sample as these substances generally interfere with the detection step. Finally, one of the primary elements of many molecular technologies involves a further step of extraction of genetic material from whole microorganisms. Although mechanisms and techniques have been devised for each of these tasks, there is still considerable room for improvement, as discussed in this chapter.
3. The Coming Together of the Sciences: Biosensors for the Detection of Waterborne Pathogens Using Antibodies and Gene-Based Recognition Chemistries
Sen Xu and Raj Mutharasan
The detection of waterborne pathogens and toxins by biosensor-based methods are reviewed in this chapter. Descriptions of optical, electrochemical and electromechanical sensors are included to provide a broad perspective on these emerging devices. Surface chemistries for immobilizing biorecognition molecules on sensor surfaces are summarized. Further, biosensor-based detection methods for microbial contaminants are analyzed and discussed. Finally, representative sensor responses, limit of detections (LOD) and time to results (TTR) are compared and discussed.
4. Simple, Powerful, and Smart: Using LAMP for Low Cost Screening of Multiple Waterborne Pathogens
Grégoire Seyrig, Farhan Ahmad, Robert D. Stedtfeld, Dieter M. Tourlousse and Syed A. Hashsham
A vast array of low cost, simple, rugged, and rapid molecular approaches are emerging for the detection of indicators and pathogens, along with the collection of relevant genotypic information. This chapter focuses on loop-mediated isothermal amplification (LAMP), a relatively new DNA amplification technique, which due to its simplicity, ruggedness, and low cost could provide major advantages to the water industry. In LAMP, the target sequence is amplified at a constant temperature using either two or three sets of primers and a polymerase with high strand displacement activity. Due to the specific nature of the action of these primers, the amount of DNA produced in LAMP is considerably higher than PCR based amplification. The corresponding release of pyrophosphate results in visible turbidity due to precipitation, which allows easy visualization by the naked eye, especially for larger reaction volumes or via simple detection approaches for smaller volumes. The reaction can be followed in real-time either by measuring the turbidity or the signals from DNA produced via fluorescent dyes that intercalate or directly label the DNA, and in turn can be correlated to the number of copies initially present. Hence, LAMP can also be quantitative. While LAMP is already the method of choice in organizations engaged in combating infectious diseases such as tuberculosis, malaria, and sleeping sickness in developing regions, it has yet to be extensively validated for commonly known waterborne pathogens.
5. Challenges of Multiplexed Detection: Detection of Pathogens in Water and Wastewater Using Microarrays
Timothy M. Straub
For waterborne pathogen monitoring, regulatory agencies have traditionally focused on developing a single method for an existing or emerging pathogen in water supplies. However, the ability to use a single method to determine all potential pathogens or indicators in a water supply would be particularly advantageous. Such an approach has three major hurdles: 1) sensitive detection of highly dilute pathogens in a water supply, 2) specific detection of pathogens from non-pathogenic near-neighbors, and 3) multiplexed strategies that preserve the sensitivity and specificity of the assay. This chapter describes how microarrays, a series of unique nucleic acid sequences arrayed on a suitable surface, allow through hybridization, the detection of multiple genetic sequences from different viruses, bacteria, protozoa, and other emerging pathogens. Through the context of a complete method, including the recovering of pathogens from water sources, this chapter is designed to introduce the reader to microarrays, challenges of labeling pathogen nucleic acids recovered from water samples, and specific microarray applications related to waterborne pathogen monitoring.
6. Discovering New Pathogens: Amoebae as Tools to Isolate Amoeba-resisting Microorganisms from Environmental Samples
Julia Lienard and Gilbert Greub
Obligate intracellular microorganisms are unculturable by classic axenic culture methods. As a result they have largely been overlooked, despite many being significant human and animal pathogens. Resistance of amoeba-resisting microorganisms (ARM) to amoebal destruction may predict ability to also resist mammalian macrophages, which are somehow similar to amoebae and represent one of the first cellular immune defenses in mammals. Thus, general approaches are described for the growth of strict intracellular microorganisms, using amoebae as hosts in a cell culture system. Such an approach has been shown to be advantageous, since amoebal co-culture will selectively grow microorganisms that resist these professional phagocytes. An alternative approach for the isolation of novel ARM is also described, which requires the isolation of new amoebal strains by amoebal enrichment on a suitable prey (such as Escherichia coli), and then to search for intra-amoebal microorganisms within the isolated amoebae. Once new potentially pathogenic ARM has been isolated, one should then further assess the potential infectivity of these intracellular microorganisms. The application of macrophages, as an in vitro model to test microbial virulence is also described.
7. Identity and Function of Single Microbial Cells Within a Community by Raman Microspectroscopy and Related Single-cell Techniques
Daniel S. Read and Andrew S. Whiteley
Linking both identity and function within microbial communities has long been seen as essential for understanding the role that bacteria play in the environment. Techniques based on the study of single microbial cells offer a unique approach that provides information about heterogeneity within populations, and the role of spatial organization within the environment. This chapter details some of the prominent single-cell techniques currently in use for the study of microbial ecology, with a particular focus on Raman spectroscopy. A general overview of this technique is provided, with examples of its applicability for studying different features of microbial systems. Special attention is given to the use of Raman spectroscopy in combination with Fluorescence in situ Hybridization (FISH) and Stable Isotope Probing (SIP), which together can be utilized to gain an insight into the identity and function of single bacterial cells in situ.
8. Are They Alive? Detection of Viable Organisms and Functional Gene Expression Using Molecular Techniques
Paul A. Rochelle, Anne K. Camper, Andreas Nocker and Mark Burr
The ultimate measure of microbial viability and biological activity is growth in some form of culture system. Unfortunately, due to many limitations, growth is usually not the most sensitive or rapid detection method. This chapter describes many of the molecular-based tools for assessing viability and functional gene expression, and their applications for specific microbes in environmental samples. Methods include fluorescent nucleic acid binding dyes, enzymatic conversion of substrates to fluorescent compounds (often in conjunction with nucleic acid-based methods), various techniques based on amplification and detection of nucleic acids, nucleic acid amplification linked to biosensors and microarray detection platforms, detection and characterization of proteins, and molecular detection coupled with culturing. Principles supporting each of these techniques are discussed along with applications to bacteria, protozoa, and viruses, focusing primarily on microbes of concern to the drinking water and wastewater industries.
9. Characterization of Microbial Community Structures in Recreational Waters and Primary Sources of Fecal Pollution with a Next-Generation Sequencing Approach
Orin C. Shanks, Sandra McLellan, Susan M. Huse and Mitchell L. Sogin
The invention of new approaches to DNA sequencing commonly referred to as next generation sequencing technologies is revolutionizing the study of microbial diversity. In this chapter, we discuss the characterization of microbial population structures in surface waters and potentially contributing fecal sources using a GS-FLX pyrosequencer. An overview of the principals of the technology, workflow, sequencing strategies, and data analysis challenges are included. In addition, the bacterial population structure of an untreated wastewater sewage sample is reported and discussed to demonstrate a pyrosequencing approach for studying a microbial community. Emphasis is placed on the application of pyrosequencing technology for recreational water quality management including the discovery of alternative indicators of fecal pollution and improving the link between fecal pollution and the transmission of waterborne disease.
10. Microbial Source Tracking: Current and Future Molecular Tools in Microbial Water Quality Forensics
Jorge W. Santo Domingo, Regina Lamendella and Nicholas Ashbolt
Regulations in the United States and elsewhere stipulate that drinking and recreational waters should be regularly monitored for microbial indicators of fecal pollution. Hence, the health risks associated with these waters are determined using microbial indicators rather than by direct pathogen detection. Detecting pathogens may seem more appropriate, however, pathogens are difficult to enumerate since they often are found in environmental samples in very low numbers that still present health concerns and may take several days to weeks to detect, lessening their value for risk management. In this chapter, we discuss the use of molecular methods that attempt to track fecal sources of pollution in environmental waters and discuss their role in environmental monitoring and management. A general description of the most commonly used methods is provided, and where appropriate, some of the advantages and limitations are highlighted. The use of genomic technologies to develop new methods and to fill existing research gaps in microbial source tracking (MST) is also discussed. As clear from other chapters of this book, the field of environmental microbiology is undergoing a fundamental change through the development of tools that can describe the molecular diversity of microbial populations relevant to environmental fecal pollution. While this is a dynamic field, emphasis is placed on culture-independent methods relying on DNA-based targets that currently dominate the scientific literature.
11. Dynamics of Microbes in the Natural Setting: Development of the Geochip
Joy D. Van Nostrand, Zhili He and Jizhong Zhou
This chapter provides an overview of various microarrays available for environmental analysis, and building on previous chapters of this book, has the primary focus of describing functional gene arrays (FGAs). While many FGAs have been designed, the most comprehensive FGA available to date is the GeoChip, which targets thousands of key functional genes involved in the geochemical cycling of carbon (C), nitrogen (N), phosphorus (P), and sulphur (S), metal resistance and reduction, energy processing and contaminant degradation. Many studies have been performed using the GeoChip, showing it to be a powerful tool for rapid, sensitive and specific examination of microbial communities in a high-throughput manner. As such, the GeoChip is well-suited for linking geochemical processes with microbial community function and structure. However, challenges still remain with the use of FGAs and other microarrays for microbial community analysis, which one day may aid in revolutionizing how we assess microbial water quality.
12. The Microbe as a Reporter: Microbial Bioreporter Sensing Technologies for Chemical and Biological Detection
Steven Ripp, Alice C. Layton and Gary S. Sayler
Bioreporters function as living whole cell sensors of environmental contaminants. These sensors have been widely applied in water quality control and assessment to establish water toxicity profiles and for the specific identification of pollutant chemicals. Innovative bacteriophage (bacterial virus) bioreporters have also recently become available for the specific identification of contaminating organisms of human health concern (i.e., bacterial pathogens). All of these bioreporter systems take advantage of measured cellular signal outputs such as colorimetric, bioluminescent, or fluorescent emissions to signify target chemical or biological presence. When associated with a device capable of measuring these emissions, the resulting bioreporter assay or hybrid biosensor becomes an easy to use, rapid, and versatile water quality monitoring tool that can be integrated into continuous, on-line, near real-time sensing regimens that rival conventional analytical methods.
How to buy this book
(EAN: 9781904455707 9781912530779 Subjects: [bacteriology] [virology] [microbiology] [molecular microbiology] [environmental microbiology] )