Salmonella survival
High-throughput screening to determine the genetic requirements for Salmonella survival under different growth conditions
from Mollie Megan Reynolds, Rocio Canals, Michael McClelland and Helene Andrews-Polymenis writing in Salmonella: From Genome to Function
Salmonella species are capable of survival in a wide range of niches, both in the environment and in an infected host. Genetic requirements for survival of Salmonella in different niches have traditionally been identified using gene expression and forward genetics. The availability of complete genome sequences, microarray technology, and cost-effective new sequencing capabilities enabled increasingly efficient high-throughput analyses of Salmonella genomes to identify elements that contribute to survival in these niches. A recent review describes many of the high-throughput tools that have been developed over the past two decades, and the genetic requirements for Salmonella survival that have been identified using these techniques.
Further reading: Salmonella: From Genome to Function
from Mollie Megan Reynolds, Rocio Canals, Michael McClelland and Helene Andrews-Polymenis writing in Salmonella: From Genome to Function
Salmonella species are capable of survival in a wide range of niches, both in the environment and in an infected host. Genetic requirements for survival of Salmonella in different niches have traditionally been identified using gene expression and forward genetics. The availability of complete genome sequences, microarray technology, and cost-effective new sequencing capabilities enabled increasingly efficient high-throughput analyses of Salmonella genomes to identify elements that contribute to survival in these niches. A recent review describes many of the high-throughput tools that have been developed over the past two decades, and the genetic requirements for Salmonella survival that have been identified using these techniques.
Further reading: Salmonella: From Genome to Function
Getting The Most Out of PCR
We think you will be interested in an online seminar series entitled "Getting The Most Out of PCR", which is being broadcast by the popular life science blog, Bitesize Bio. Bitesize Bio is headed by Nick Oswald and Suzanne Kennedy, who co-edited our recent title "PCR Troubleshooting and Optimization".
The series lineup includes many of the authors from this book and kicks off on 18 May with a talk from LightCycler co-inventor, Carl Wittwer, entitled "Magic in Solution: An Introduction and Brief History of PCR". This will be a great learning experience with an opportunity to ask questions and learn from experts and pioneers in the PCR field. The full program is shown below.
Click here to book your place on these excellent events.
Recommended reading: PCR publications
The series lineup includes many of the authors from this book and kicks off on 18 May with a talk from LightCycler co-inventor, Carl Wittwer, entitled "Magic in Solution: An Introduction and Brief History of PCR". This will be a great learning experience with an opportunity to ask questions and learn from experts and pioneers in the PCR field. The full program is shown below.
Click here to book your place on these excellent events.
- Magic in Solution: An Introduction and Brief History of PCR
Speaker: Carl Wittwer
18 May 2010 / 9am Pacific / 12pm Eastern / 5pm GMT / 6pm CET - Obtaining Maximum PCR Sensitivity and Specificity
Speaker: Cameron N. Gundry
25 May 2010 / 9am Pacific / 12pm Eastern / 5pm GMT / 6pm CET - Significance of Controls and Standard Curves in PCR
Speaker: Ian Kavanagh
01 June 2010 / 9am Pacific / 12pm Eastern / 5pm GMT / 6pm CET - The MBD2-based Enrichment Approach for Analyzing DNA methylation
Speaker: Chris Adams
08 June 2010 / 9am Pacific / 12pm Eastern / 5pm GMT / 6pm CET - The MIQE Guidelines Uncloaked
Speaker: Greg Shipley
15 June 2010 / 9am Pacific / 12pm Eastern / 5pm GMT / 6pm CET - High Resolution Melting Analysis - Beyond the SNP
Speaker: John Mackay
22 June 2010 / 9am Pacific / 12pm Eastern / 5pm GMT / 6pm CET
Recommended reading: PCR publications
Detection of Viable Organisms Using Molecular Techniques
from Paul A. Rochelle, Anne K. Camper, Andreas Nocker and Mark Burr in Environmental Microbiology: Current Technology and Water Applications
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. Many molecular-based tools are available for assessing viability and functional gene expression, and have 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.
Further reading:
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. Many molecular-based tools are available for assessing viability and functional gene expression, and have 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.
Further reading:
Identity of Single Microbial Cells
from Daniel S. Read and Andrew S. Whiteley in Environmental Microbiology: Current Technology and Water Applications
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. Various single-cell techniques are currently in use for the study of microbial ecology, an important one being Raman spectroscopy. This technique can be used for studying different features of microbial systems. Raman spectroscopy can be used 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.
Further reading:
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. Various single-cell techniques are currently in use for the study of microbial ecology, an important one being Raman spectroscopy. This technique can be used for studying different features of microbial systems. Raman spectroscopy can be used 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.
Further reading:
Amoebae as a Tool
from Julia Lienard and Gilbert Greub in Environmental Microbiology: Current Technology and Water Applications
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 have been 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 available, 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 possible.
Further reading:
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 have been 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 available, 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 possible.
Further reading:
Detection of Pathogens in Water Using Microarrays
from Timothy M. Straub in Environmental Microbiology: Current Technology and Water Applications
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.
Further reading:
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.
Further reading:
Low Cost Screening of Multiple Waterborne Pathogens
from Seyrig et al in Environmental Microbiology: Current Technology and Water Applications
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. Loop-mediated isothermal amplification (LAMP) is 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.
Further reading:
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. Loop-mediated isothermal amplification (LAMP) is 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.
Further reading:
- Environmental Microbiology: Current Technology and Water Applications
- Nanotechnology in Water Treatment Applications
- Metagenomics: Theory, Methods and Applications
- Environmental Molecular Microbiology
Biosensors for the Detection of Waterborne Pathogens
from Sen Xu and Raj Mutharasan in Environmental Microbiology: Current Technology and Water Applications
The detection of waterborne pathogens and toxins by biosensor-based methods are becoming increasingly important. Optical, electrochemical and electromechanical sensors are available and surface chemistries are being used for immobilizing biorecognition molecules on sensor surfaces. Topics that are important include representative sensor responses, limit of detections (LOD) and time to results (TTR).
Further reading:
The detection of waterborne pathogens and toxins by biosensor-based methods are becoming increasingly important. Optical, electrochemical and electromechanical sensors are available and surface chemistries are being used for immobilizing biorecognition molecules on sensor surfaces. Topics that are important include representative sensor responses, limit of detections (LOD) and time to results (TTR).
Further reading:
Detection of Microbes in Water
from Keya Sen in Environmental Microbiology: Current Technology and Water Applications
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. From the vast panorama of available molecular techniques, some are finding a place in the water industry: Quantitative PCR, protein detection and immunological approaches, loop-mediated isothermal amplification (LAMP), microarrays.
Further reading:
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. From the vast panorama of available molecular techniques, some are finding a place in the water industry: Quantitative PCR, protein detection and immunological approaches, loop-mediated isothermal amplification (LAMP), microarrays.
Further reading:
Risk assessment of nanoparticles and nanomaterials
from Michele de Kwaadsteniet and Thomas Eugene Cloete in Nanotechnology in Water Treatment Applications
The risk assessment of nanoparticles and nanomaterials is of key importance for the continous development in the already striving new field of nanotechnology. Humans are increasingly being exposed to nanoparticles and nanomaterials, placing stress on the development and validation of reproducible toxicity tests. Tests currently used include genotoxicity and cytotoxicity tests, and in vivo toxicity models. The unique characteristics of nanoparticles and nanomaterials are responsible for their toxicity and interaction with biological macromolecules within the human body. This may lead to the development of diseases and clinical disorders. A loss in cell viability and structure can also occur in exposed tissues as well as inflammation and granuloma formation. The future of nanotechnology depends on the responsible assessment of nanoparticles and nanomaterials.
Further reading: Nanotechnology in Water Treatment Applications
The risk assessment of nanoparticles and nanomaterials is of key importance for the continous development in the already striving new field of nanotechnology. Humans are increasingly being exposed to nanoparticles and nanomaterials, placing stress on the development and validation of reproducible toxicity tests. Tests currently used include genotoxicity and cytotoxicity tests, and in vivo toxicity models. The unique characteristics of nanoparticles and nanomaterials are responsible for their toxicity and interaction with biological macromolecules within the human body. This may lead to the development of diseases and clinical disorders. A loss in cell viability and structure can also occur in exposed tissues as well as inflammation and granuloma formation. The future of nanotechnology depends on the responsible assessment of nanoparticles and nanomaterials.
Further reading: Nanotechnology in Water Treatment Applications
Detection of Microbial Pathogens
from Jacques Theron, Thomas Eugene Cloete and Michele de Kwaadsteniet in Nanotechnology in Water Treatment Applications
Detection of pathogens often involves time-consuming culture methods. Newer enzymatic, immunological and genetic methods are being developed to replace and/or support classical approaches to microbial detection. Innovations in nanotechnology and nanosciences are having a significant impact in biodiagnostics, where a number of nanoparticle-based assays and nanodevices have been introduced for biomolecular detection.
Waterborne disease is still a major cause of death in many parts of the world, particularly in young children, the elderly, or those with compromised immune systems. As the epidemiology of waterborne diseases is changing, there is a growing global public health concern about new and reemerging infectious diseases that are occurring through a complex interaction of social, economic, evolutionary, and ecological factors. An important challenge is therefore the rapid, specific and sensitive detection of waterborne pathogens.
Further reading: Nanotechnology in Water Treatment Applications
Detection of pathogens often involves time-consuming culture methods. Newer enzymatic, immunological and genetic methods are being developed to replace and/or support classical approaches to microbial detection. Innovations in nanotechnology and nanosciences are having a significant impact in biodiagnostics, where a number of nanoparticle-based assays and nanodevices have been introduced for biomolecular detection.
Waterborne disease is still a major cause of death in many parts of the world, particularly in young children, the elderly, or those with compromised immune systems. As the epidemiology of waterborne diseases is changing, there is a growing global public health concern about new and reemerging infectious diseases that are occurring through a complex interaction of social, economic, evolutionary, and ecological factors. An important challenge is therefore the rapid, specific and sensitive detection of waterborne pathogens.
Further reading: Nanotechnology in Water Treatment Applications
Nanotechnology and Water Microbiology
Nanotechnology is the engineering and art of manipulating matter at the nanoscale (1-100 nm) level. Nanotechnology offers the potential of novel nanomaterials for the treatment of surface water, groundwater and wastewater contaminated by toxic metal ions, organic and inorganic solutes and microorganisms. At the present time many nanomaterials are under active research and development.
Further reading: Nanotechnology in Water Treatment Applications
Further reading: Nanotechnology in Water Treatment Applications