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Microbial Identification (16S rDNA)            SECTION - B
 
 
Staphylococcus aureus Treated - A
PROJECT REPORT
NAME OF SCIENTIST:
Mr. Mahendra Kumar Trivedi 		SERVICE:
Bacterial Identification: 3A
 SERVICE NO.:
BIS-08-003.1
 
Project Title Bacterial Identification Service based on 16S rDNA data
Cat#: CSER 47
Name of Scientist: Mr. Mahendra Kumar Trivedi
A 4/14, Kanwal & Kanwal apartments, Four Bungalows, Opposite
Fish Market, Near Church, Andheri , Mumbai
India- 400053
thelifeenergy@gmail.com
Institute: The Life Energy, Mumbai
Sample name: Staphylococcus aureus Treated A (Given Name: 3A)
Source of the Sample: On 23rd January 2008, Mr. Mahendra Kumar Trivedi visited our R&D facility at Bangalore to observe the impact of his treatment modality on various groups of bacteria at the level of DNA fingerprinting. On the same day, he subjected Staphylococcus aureus, ATCC no-49831 to his said treatment modality that were handed over to Ms. Bindu in two separate tubes named as Series A & Series B.
Following the treatment, after few days, we sub cultured the inoculums of treated Series A & Series B into Treated Sample A1 and Treated Sample B1 respectively.
All four treated samples (Treated sample A, Treated sample B, Treated Sample A1 And Treated Sample B1) along with Control were analyzed for DNA fingerprinting by RAPD analysis. And the comparative results are as follows.
 
 
  • The percentage of true polymorphism observed between Control & Treated sample A of Staphylococcus aureus sample is on an average 79%.
  • The percentage of true polymorphism observed between Control & Treated sample A1 of Staphylococcus aureus sample is on an average 45%.
  • The percentage of true polymorphism observed between Control & Treated sample B of Staphylococcus aureus sample is on an average 74%.
  • The percentage of true polymorphism observed between Control & Treated sample B1 of Staphylococcus aureus sample is on an average 52%.
  The details of the same report have been issued to him. As per his request, we have preserved all the treated samples. Out of which, preserved Treated Sample A was requested to analyse for Bacterial Identification Service/ Test based on 16S rDNA data by Mr. Trivedi on 9th April 2008.The original source (before the treatment) of preserved Treated Sample A is Staphylococcus aureus, ATCC no-49831.
Steps Involved: 1. Genomic DNA was isolated from the pure culture pellet after treatment provided by Mr. Mahendra Kumar Trivedi

2. Using consensus primers, the ~1.5 kb 16S rDNA fragment was amplified using high-fidelity PCR Polymerase.

3. The PCR product was cloned and plasmid DNA was bi-directionally sequenced using the forward, reverse and an internal primer.

4. Sequence data was aligned and analyzed for finding the closest homologs for the microbe.
 
Result:
  1. Based on nucleotides homology and phylogenetic analysis the Microbe (Sample: 3A) was detected to be Staphylococcus aureus (GenBank Accession Number: L37597).

  2. Nearest homolog species was found to be Staphylococcus simiae (Accession No. AY727530).

  3. Information about other close homologs for the microbe can be found from the Alignment View table.

Materials Provided to the Scientist:

1. Final Report


Date: 28.04.2008


*The sample has been indicated as 3A

Aligned Sequence Data: (1490 bp)

AGGATGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAGCGAACGGACGAGAAGCTTGCTTCTCTGATGTTAGCG
GCGGACGGGTGAGTAACACGTGGATAACCTACCTATAAGACTGGGATAACTTCGGGAAACCGGAGCTAATACCGGAT
AATATTTTGAACCGCATGGTTCAAAAGTGAAAGACGGTCTTGCTGTCACTTATAGATGGATCCGCGCTGCATTAGCT
AGTTGGTAAGGTAACGGCTTACCAAGGCAACGATGCATAGCCGACCTGAGAGGGTGATCGGCCACACTGGAACTGAG
ACACGGTCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGGCGAAAGCCTGACGGAGCAACGCCGC
GTGAGTGATGAAGGTCTTCGGATCGTAAAACTCTGTTATTAGGGAAGAACATATGTGTAAGTAACTGTGCACATCTT
GACGGTACCTAATCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTATCCG
GAATTATTGGGCGTACAGCGCGCGTAGGCGGTTTTTTAAGTCTGATGTGAAAGCCCACGGCTCAACCGTGGAGGGTC
ATTGGAAACTGGAAAACTTGAGTGCAGAAGAGGAAAGTGGAATTCCATGTGTAGCGGTGAAATGCGCAGAGATATGG
AGGAACACCAGTGGCGAAGGCGACTTTCTGGTCTGTAACTGACGCTGATGTGCGAAAGCGTGGGGATCAAACAGGAT
TAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGGGGGTTTCCGCCCCTTAGTGCTGCAGCTA
ACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTAAAAGGAATTGACGGGGACTCGCAGAATCG
GTGGAACATGTGGTTTAATTCGAAGCAACGCGAAGAGCCTTACCAAATCTTGACATCCTTTGACAACTCTAGAGATA
GAGCCTTCCCCTTCGGGGGACAAAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAA
GTCCCGCAACGAGCGCAACCCTTAAGCTTAGTTGCCATCATTAAGTTGGGCACTCTAAGTTGACTGCCGGTGACAAA
CCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGATTTGGGCTACACACGTGCTACAATGGACAATA
CAAAGGGCAGCGAAACCGCGAGGTCAAGCAAATCCCATAAAGTTGTTCTCAGTTCGGATTGTAGTCTGCAACTCGAC
TACATGAAGCTGGAATCGCTAGTAATCGTAGATCAGCATGCTACGGTGAATACGTTCCCGGGTCTTGTACACACCGC
CCGTCACACCACGAGAGTTTGTAACACCCGAAGCCGGTGGAGTAACCTTTTAGGAGCTAGCCGTCGAAGGTGGGACA
AATGATTGGGGTGAAGTCGTAACAGCT

Alignment View using combination of NCBI GenBank and RDP database:

Alignment View
ID
Alignment results
Sequence description
0.97
Sample studied
1.00
Staphylococcus aureus strain: MPU99
1.00
Staphylococcus aureus
1.00
Staphylococcus aureus strain ATCC 14458
1.00
Staphylococcus aureus strain SMKV-2
0.99
Staphylococcus aureus strain F136
0.99
Staphylococcus aureus strain F155
0.97
Staphylococcus simiae strain CCM 7229
0.97
Staphylococcus simiae strain CCM 7213
0.99
Staphylococcus epidermidis strain CU22
1.00
Staphylococcus epidermidis strain S09

Distance Matrix based on Nucleotide Sequence Homology (Using Kimura-2 Parameter):

Distance Matrix
 
 
1
2
3
4
5
6
7
8
9
10
11
DQ997837
 1 
  ---   0.999   1   0.995   1   0.995   1   0.986   0.986   0.999   0.995 
DQ997836
 2 
  0.001   ---   0.999   0.996   0.999   0.996   0.999   0.985   0.985   0.998   0.994 
L37597
 3 
  0.000   0.001   ---   0.995   1   0.995   1   0.986   0.986   0.999   0.995 
DQ127902
 4 
  0.005   0.004   0.005   ---   0.995   1   0.995   0.985   0.985   0.994   0.990 
DQ306891
 5 
  0.000   0.001   0.000   0.005   ---   0.995   1   0.986   0.986   0.999   0.995 
AY727530
 6 
  0.005   0.004   0.005   0.000   0.005   ---   0.995   0.985   0.985   0.994   0.990 
AB353073
 7 
  0.000   0.001   0.000   0.005   0.000   0.005   ---   0.986   0.986   0.999   0.995 
EF522128
 8 
  0.014   0.015   0.014   0.015   0.014   0.015   0.014   ---   1   0.987   0.981 
AY741152
 9 
  0.014   0.015   0.014   0.015   0.014   0.015   0.014   0.000   ---   0.987   0.981 
DQ997835
 10 
  0.001   0.002   0.001   0.006   0.001   0.006   0.001   0.013   0.013   ---   0.995 
3A
 11 
  0.005   0.006   0.005   0.010   0.005   0.010   0.005   0.019   0.019   0.005   --- 

Table indicates nucleotide similarity (above diagonal) and distance (below diagonal) identities between the studied sample '3A' and ten other closest homologs microbe.

Phylogenetic Tree made in MEGA 3.1 software using Neighbor Joining method:

 

Approved by: Dr. Avijit Roy

Manager R & D
Bangalore Genei
 
 
 
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