Comparison of Bacterial Communities in the Throat Swabs from Healthy Subjects and Pharyngitis Patients by Terminal Restriction Fragment Length Polymorphism

Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-la...

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Vydáno v:Applied biochemistry and biotechnology Ročník 167; číslo 5; s. 1459 - 1473
Hlavní autoři: Balaji, Kannan, Thenmozhi, Ramalingam, Sundaravadivel, Marimuthu, Pandian, Shunmugiah Karutha
Médium: Journal Article
Jazyk:angličtina
Vydáno: New York Springer-Verlag 01.07.2012
Springer Nature B.V
Témata:
Bacteria
Bacteria - classification
Bacteria - genetics
Bacteria - isolation & purification
Bacteria - pathogenicity
bacterial communities
Bacteriology
Biochemistry
Biodiversity
Biotechnology
Chemistry
Chemistry and Materials Science
classification
Community structure
computer software
Enzymes
Flora
genes
genetics
Genomes
Health
Humans
isolation & purification
Metagenome
Metagenome - genetics
metagenomics
microbiology
pathogenicity
Pathology
patients
Pharyngitis
Pharyngitis - microbiology
Pharynx
Pharynx - microbiology
Phylogeny
Polymerase chain reaction
Polymorphism
Polymorphism, Restriction Fragment Length
restriction fragment length polymorphism
ribosomal DNA
ribosomal RNA
RNA, Bacterial
RNA, Bacterial - genetics
RNA, Ribosomal, 16S
RNA, Ribosomal, 16S - genetics
Species diversity
Species richness
Streptococcus pyogenes
throat
uncertainty
Normal flora
Restriction enzymes
Pharyngitis
Metagenome
T-RFLP
ISSN:0273-2289, 1559-0291, 1559-0291
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Abstract Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites ( Msp I or Rsa I) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from Msp I and Rsa I digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though Rsa I digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [±2.1 (SD)] and 10 (±2.9) for Msp I and Rsa I digests, respectively, whereas healthy throats generated 6.3 (±1.2) and 6.1 (±1.5) in Msp I and Rsa I digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.
AbstractList Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites (MspI or RsaI) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from MspI and RsaI digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though RsaI digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [ plus or minus 2.1 (SD)] and 10 ( plus or minus 2.9) for MspI and RsaI digests, respectively, whereas healthy throats generated 6.3 ( plus or minus 1.2) and 6.1 ( plus or minus 1.5) in MspI and RsaI digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.
Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites (MspI or RsaI) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from MspI and RsaI digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though RsaI digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [±2.1 (SD)] and 10 (±2.9) for MspI and RsaI digests, respectively, whereas healthy throats generated 6.3 (±1.2) and 6.1 (±1.5) in MspI and RsaI digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.
Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites (MspI or RsaI) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from MspI and RsaI digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though RsaI digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [±2.1 (SD)] and 10 (±2.9) for MspI and RsaI digests, respectively, whereas healthy throats generated 6.3 (±1.2) and 6.1 (±1.5) in MspI and RsaI digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.
Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites (MspI or RsaI) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from MspI and RsaI digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though RsaI digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [±2.1 (SD)] and 10 (±2.9) for MspI and RsaI digests, respectively, whereas healthy throats generated 6.3 (±1.2) and 6.1 (±1.5) in MspI and RsaI digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites (MspI or RsaI) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from MspI and RsaI digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though RsaI digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [±2.1 (SD)] and 10 (±2.9) for MspI and RsaI digests, respectively, whereas healthy throats generated 6.3 (±1.2) and 6.1 (±1.5) in MspI and RsaI digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.
Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites ( Msp I or Rsa I) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from Msp I and Rsa I digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though Rsa I digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [±2.1 (SD)] and 10 (±2.9) for Msp I and Rsa I digests, respectively, whereas healthy throats generated 6.3 (±1.2) and 6.1 (±1.5) in Msp I and Rsa I digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.
Issue Title: NEW HORIZONS IN BIOTECHNOLOGY Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and pharyngitis patients. The 16S rRNA genes of bacteria present in throat metagenome were amplified by PCR with 6-carboxy-fluorescein (6-FAM)-labeled universal forward primer (27 F) and a universal reverse primer (1513R). The 16S rDNAs were digested with restriction enzymes with 4-bp recognition sites (MspI or RsaI) and analyzed by using an automated DNA sequencer. T-RFLP patterns were numerically analyzed using computer programs. From analysis of the throat bacterial community, patterns derived from MspI and RsaI digested samples of healthy subjects and pharyngitis patients were grouped into different clusters, though RsaI digested samples showed some uncertainty. Pharyngitis throats generated an average species richness of 9 [±2.1 (SD)] and 10 (±2.9) for MspI and RsaI digests, respectively, whereas healthy throats generated 6.3 (±1.2) and 6.1 (±1.5) in MspI and RsaI digests, respectively. These results suggest that samples from pharyngitis patients contain an unexpected diversity of causative bacteria. The pharyngitis throats were colonized with a rich diversity of bacterial species than that of healthy throats. Using T-RFLP, we are able to detect a model bacterium, Streptococcus pyogenes SF370, and T-RF patterns were consistent with the Streptococcal T-RFLP patterns. Our study indicates that T-RFLP analysis is useful for the assessment of diversity of throat bacterial flora and rapid comparison of the community structure between subjects with and without pharyngitis.[PUBLICATION ABSTRACT]
Author Balaji, Kannan
Pandian, Shunmugiah Karutha
Thenmozhi, Ramalingam
Sundaravadivel, Marimuthu
Author_xml – sequence: 1
  givenname: Kannan
  surname: Balaji
  fullname: Balaji, Kannan
  organization: Department of Biotechnology, Alagappa University
– sequence: 2
  givenname: Ramalingam
  surname: Thenmozhi
  fullname: Thenmozhi, Ramalingam
  organization: Department of Biotechnology, Alagappa University
– sequence: 3
  givenname: Marimuthu
  surname: Sundaravadivel
  fullname: Sundaravadivel, Marimuthu
  organization: Department of Biotechnology, Alagappa University
– sequence: 4
  givenname: Shunmugiah Karutha
  surname: Pandian
  fullname: Pandian, Shunmugiah Karutha
  email: sk_pandian@rediffmail.com
  organization: Department of Biotechnology, Alagappa University
BackLink https://www.ncbi.nlm.nih.gov/pubmed/22322827$$D View this record in MEDLINE/PubMed
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Issue 5
Keywords Normal flora
Restriction enzymes
Pharyngitis
Metagenome
T-RFLP
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Snippet Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora present in the throats of healthy subjects and...
Issue Title: NEW HORIZONS IN BIOTECHNOLOGY Terminal restriction fragment length polymorphism (T-RFLP) analysis was applied to characterize bacterial flora...
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StartPage 1459
SubjectTerms Bacteria
Bacteria - classification
Bacteria - genetics
Bacteria - isolation & purification
Bacteria - pathogenicity
bacterial communities
Bacteriology
Biochemistry
Biodiversity
Biotechnology
Chemistry
Chemistry and Materials Science
classification
Community structure
computer software
Enzymes
Flora
genes
genetics
Genomes
Health
Humans
isolation & purification
Metagenome
Metagenome - genetics
metagenomics
microbiology
pathogenicity
Pathology
patients
Pharyngitis
Pharyngitis - microbiology
Pharynx
Pharynx - microbiology
Phylogeny
Polymerase chain reaction
Polymorphism
Polymorphism, Restriction Fragment Length
restriction fragment length polymorphism
ribosomal DNA
ribosomal RNA
RNA, Bacterial
RNA, Bacterial - genetics
RNA, Ribosomal, 16S
RNA, Ribosomal, 16S - genetics
Species diversity
Species richness
Streptococcus pyogenes
throat
uncertainty
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Title Comparison of Bacterial Communities in the Throat Swabs from Healthy Subjects and Pharyngitis Patients by Terminal Restriction Fragment Length Polymorphism
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