study Yellow Sand dust, a seasonal meteorological phenomenon affecting East Asia.
These storms often provide long-range transport to various microorganisms.
Microbiological air samples were collected using a PM2.5 cyclones, Yellow Sand
events, and non-Yellow Sand events. Total nucleic acids were also extracted, and the
16S rDNA was amplified by PCR and analyzed by denaturing gradient gel electrophoresis (DGGE). Dendrogram analysis, based on DGGE, indicated that the microbial profiles from the Yellow Sand were distinctive from those of the non-Yellow
Sand samples. These results suggest that, as a result of Yellow Sand events, humans
in the affected regions are exposed to communities of microorganisms that might
cause various adverse health effects. In DGGE, group-specific 16S rRNA primers
are useful to compare different microbial communities, as well as to monitor
microbial communities in function of time.
9.2.1.3 Pulsed Field Gel Electrophoresis (PFGE)
Pulsed field gel electrophoresis (PFGE) is an agarose gel electrophoresis technique
used for separating larger pieces of DNA by applying electrical current that periodically changes direction (three directions) in a gel matrix unlike the conventional gel
electrophoresis where the current flows only in one direction (Schwartz and Cantor
1984; Arbeit 1999; Trindade et al. 2003). In PFGE, intact chromosomes are digested
using restriction endonucleases to generate a series of DNA fragments of different
sizes and patterns specific for a particular species or strain (Shi et al. 2010). This
method has good reproducibility, discriminatory power, and typeability, but PFGE is
sensitive to genetic instability, has limited availability, and requires at least 3–4 days
to complete a test (Wassenaar and Newell 2000).
9.2.1.4 Multilocus Sequence Typing (MLST)
Multilocus sequence typing (MLST) is an unambiguous, portable, and nucleotidebased technique for typing bacteria using the DNA sequences of internal fragments
of multiple housekeeping genes (Maiden et al. 1998; Spratt 1999; Urwin and Maiden
2003). In MLST, approximately 450–500 bp internal fragments of each gene are
used and most bacteria have enough variation within the house-keeping genes to
provide many alleles per locus, thus allowing billions of distinct allelic profiles to be
differentiated utilizing the multiple house-keeping loci (Enright and Spratt 1999;
Urwin and Maiden 2003). The advantages of MLST are that it provides typing data
that are unambiguous, portable, more accurate, and more discriminatory for most
bacteria. These data are readily available, comparable, and accessible via the internet
in contrast to most typing procedures involving the comparison of DNA fragment
sizes on a gel (Dingle et al. 2005). Furthermore, MLST data can be used to
investigate evolutionary relationships among bacteria (Urwin and Maiden 2003).
9.2.1.5 Random Amplified Polymorphism Deoxyribonucleic Acid (RAPD)
Random amplified polymorphism deoxyribonucleic acid (RAPD) is a PCR-based
technique in which arbitrary primers (typically 10-mer primers) are used to randomly
amplify segments of target DNA under low-stringency PCR condition (Wassenaar
and Newell 2000). This process leads to the amplification of one or more DNA
192
S. Purwar and S. Srivastava
These storms often provide long-range transport to various microorganisms.
Microbiological air samples were collected using a PM2.5 cyclones, Yellow Sand
events, and non-Yellow Sand events. Total nucleic acids were also extracted, and the
16S rDNA was amplified by PCR and analyzed by denaturing gradient gel electrophoresis (DGGE). Dendrogram analysis, based on DGGE, indicated that the microbial profiles from the Yellow Sand were distinctive from those of the non-Yellow
Sand samples. These results suggest that, as a result of Yellow Sand events, humans
in the affected regions are exposed to communities of microorganisms that might
cause various adverse health effects. In DGGE, group-specific 16S rRNA primers
are useful to compare different microbial communities, as well as to monitor
microbial communities in function of time.
9.2.1.3 Pulsed Field Gel Electrophoresis (PFGE)
Pulsed field gel electrophoresis (PFGE) is an agarose gel electrophoresis technique
used for separating larger pieces of DNA by applying electrical current that periodically changes direction (three directions) in a gel matrix unlike the conventional gel
electrophoresis where the current flows only in one direction (Schwartz and Cantor
1984; Arbeit 1999; Trindade et al. 2003). In PFGE, intact chromosomes are digested
using restriction endonucleases to generate a series of DNA fragments of different
sizes and patterns specific for a particular species or strain (Shi et al. 2010). This
method has good reproducibility, discriminatory power, and typeability, but PFGE is
sensitive to genetic instability, has limited availability, and requires at least 3–4 days
to complete a test (Wassenaar and Newell 2000).
9.2.1.4 Multilocus Sequence Typing (MLST)
Multilocus sequence typing (MLST) is an unambiguous, portable, and nucleotidebased technique for typing bacteria using the DNA sequences of internal fragments
of multiple housekeeping genes (Maiden et al. 1998; Spratt 1999; Urwin and Maiden
2003). In MLST, approximately 450–500 bp internal fragments of each gene are
used and most bacteria have enough variation within the house-keeping genes to
provide many alleles per locus, thus allowing billions of distinct allelic profiles to be
differentiated utilizing the multiple house-keeping loci (Enright and Spratt 1999;
Urwin and Maiden 2003). The advantages of MLST are that it provides typing data
that are unambiguous, portable, more accurate, and more discriminatory for most
bacteria. These data are readily available, comparable, and accessible via the internet
in contrast to most typing procedures involving the comparison of DNA fragment
sizes on a gel (Dingle et al. 2005). Furthermore, MLST data can be used to
investigate evolutionary relationships among bacteria (Urwin and Maiden 2003).
9.2.1.5 Random Amplified Polymorphism Deoxyribonucleic Acid (RAPD)
Random amplified polymorphism deoxyribonucleic acid (RAPD) is a PCR-based
technique in which arbitrary primers (typically 10-mer primers) are used to randomly
amplify segments of target DNA under low-stringency PCR condition (Wassenaar
and Newell 2000). This process leads to the amplification of one or more DNA
192
S. Purwar and S. Srivastava