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Pathogenic isolates of Klebsiella pneumoniae (K. pneumoniae), particularly the extended-spectrum β-lactamase (ESBL) producing strains, are mostly associated with the failure of antibiotic therapy in nosocomial infections. The present work was designed to evaluate the impact of Mr. Trivedi’s biofield energy treatment on phenotypic and genotypic characteristics of K. pneumoniae. The strain of K. pneumoniae bearing ATCC 15380 (American Type Culture Collection) was procured from the Bangalore Genei, in sealed pack and divided into control and treated groups. Treated group was subjected to Mr. Trivedi’s biofield energy treatment and analyzed for the antimicrobial susceptibility, minimum inhibitory concentration (MIC), biochemical reactions, and biotyping using automated MicroScan Walk-Away®system. Further, the effect of biofield treatment was also evaluated using Random Amplified Polymorphic DNA (RAPD) in order to determine their epidemiological relatedness and genetic characteristics of biofield treated K. pneumoniaesamples. The antimicrobial susceptibility results showed an improve sensitivity (i.e. from intermediate to susceptible) of ampicillin/sulbactam and chloramphenicol, while altered sensitivity of cephalothin (i.e. from susceptible to intermediate) was also reported as compared to the control sample. The MIC value showed two-fold decrease in MIC value of ampicillin/sulbactam (i.e. 16/8 to ≤8/4 μg/mL) and chloramphenicol (i.e. 16 to ≤ 8 μg/mL) as compared to the control. The cephalothin showed two-folds change (i.e. ≤ 8 to 16 μg/mL) in the MIC value as compared with the control. Biofield treatment showed 9.09% alterations in biochemical reactions followed by a change in biotype number (7774 4272) in the treated group with respect to the control (7774 4274). Genetic fingerprinting was performed on control and treated samples using RAPD-PCR biomarkers, which showed an average range of 11 to 15% of polymorphism among the treated samples with respect to the control. These results suggested that Mr. Trivedi’s biofield energy treatment has a significant impact on K. pneumoniae. | Authors:\nMahendra Trivedi, Dahryn Trivedi, Alice Branton, Gopal Nayak

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Journal of health medical informatics

Journal of Health & Medical

Informatics

Trivedi, et al., J Health Med Inform 2015, 6:5

http://dx.doi.org/10.4172/2157-7420.1000206

Research article

Research article

Open Access

Open Access

Antimicrobial Susceptibility, Biochemical Characterization and Molecular

Typing of Biofield Treated Klebsiella pneumoniae

Mahendra Kumar Trivedi1, Alice Branton1, Dahryn Trivedi1, Mayank Gangwar2 and Snehasis Jana2*

1Trivedi Global Inc., 10624 S Eastern Avenue Suite A-969, Henderson, NV 89052, USA

2Trivedi Science Research Laboratory Pvt. Ltd., Hall-A, Chinar Mega Mall, Chinar Fortune City, Madhya Pradesh, India

*Corresponding Author: Dr. Snehasis Jana, Trivedi Science Research Laboratory Pvt. Ltd., Hall-A, Chinar Mega Mall, Chinar Fortune City, Hoshangabad Rd.,

Bhopal-462026, Madhya Pradesh, India, Tel: +91-755-6660006; E-mail: [email protected]

Received date: September 15, 2015; Accepted date: September 22, 2015; Published date: September 25, 2015

Copyright: © 2015 Trivedi MK, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted

use, distribution, and reproduction in any medium, provided the original author and source are credited.

Abstract

Pathogenic isolates of Klebsiella pneumoniae (K. pneumoniae), particularly the extended-spectrum β-lactamase

(ESBL) producing strains, are mostly associated with the failure of antibiotic therapy in nosocomial infections. The

present work was designed to evaluate the impact of Mr. Trivedi’s biofield energy treatment on phenotypic and

genotypic characteristics of K. pneumoniae. The strain of K. pneumoniae bearing ATCC 15380 (American Type

Culture Collection) was procured from the Bangalore Genei, in sealed pack and divided into control and treated

groups. Treated group was subjected to Mr. Trivedi’s biofield energy treatment and analyzed for the antimicrobial

susceptibility, minimum inhibitory concentration (MIC), biochemical reactions, and biotyping using automated

MicroScan Walk-Away® system. Further, the effect of biofield treatment was also evaluated using Random Amplified

Polymorphic DNA (RAPD) in order to determine their epidemiological relatedness and genetic characteristics of

biofield treated K. pneumoniae samples. The antimicrobial susceptibility results showed an improve sensitivity (i.e.

from intermediate to susceptible) of ampicillin/sulbactam and chloramphenicol, while altered sensitivity of

cephalothin (i.e. from susceptible to intermediate) was also reported as compared to the control sample. The MIC

value showed two-fold decrease in MIC value of ampicillin/sulbactam (i.e. 16/8 to ≤8/4 µg/mL) and chloramphenicol

(i.e. 16 to ≤ 8 µg/mL) as compared to the control. The cephalothin showed two-folds change (i.e. ≤ 8 to 16 µg/mL) in

the MIC value as compared with the control. Biofield treatment showed 9.09% alterations in biochemical reactions

followed by a change in biotype number (7774 4272) in the treated group with respect to the control (7774 4274).

Genetic fingerprinting was performed on control and treated samples using RAPD-PCR biomarkers, which showed

an average range of 11 to 15% of polymorphism among the treated samples with respect to the control. These

results suggested that Mr. Trivedi’s biofield energy treatment has a significant impact on K. pneumoniae.

Keywords: Klebsiella pneumoniae; Biofield energy treatment;

Antibiogram,

Biochemical

reactions,

Amplified Polymorphic DNA.

mortality [3]. It has acquired resistance against extended-spectrum

cephalosporins and penicillins, due to the production of extended-

spectrum β-lactamases (ESBLs) [4].

Polymorphism;

Random

Multidrug combination therapy and some alternate treatment

options are required to control the infections associated with this

microorganism. Due to the associated side effects and failure of drug

treatment therapy, alternate and complementary therapy approach are

the preferred treatment strategies. Recently, an alternate treatment

approach using healing therapy or therapeutic touch known as biofield

energy treatment, which has been widely reported in various research

field.Thebiofield therapies (putative energy fields) were reported to

alter the sensitivity of antimicrobial against treated microorganism [5],

inhibits the growth of bacterial cultures [6], effect on in vitro cells,

tissues [7], animals [8], and the clinical effects such as hematologic [9],

immunologic effects [10], healing rates of wounds [11], etc. Biofield is

the name given to the electromagnetic field that permeates and

surrounds living organisms [12]. It is referred as the biologically

produced electromagnetic and subtle energy field that provides

regulatory and communication functions within the human organism.

Specific environmental frequencies, are absorbed by the different

biomolecules, due to changes in the movements of component parts.

Therefore, the human or any living object, not only radiate but also

absorb and respond to these frequencies [13]. Mr. Mahendra Kumar

Trivedi is well known biofield treatment practitioners, and his unique

biofield energy treatment is known as The Trivedi Effect®. Mr. Trivedi’s

Abbreviations:

CAM: Complementary and Alternate Medicine; NHIS: National

Health Interview Survey; NCHS: National Center for Health Statistics;

ATCC: American Type Culture Collection; MIC: Minimum Inhibitory

Concentration; MEGA: Molecular Evolutionary Genetics Analysis;

NBPC 30: Negative Breakpoint Combo Panel 30; RAPD: Random

Amplified Polymorphic DNA; PCR: Polymerase chain reaction; ESBL:

Extended Spectrum β-Lactamase

Introduction

The increased medical practice for antibiotic usage creates selection

pressure and results emergence of nosocomial pathogens. Klebsiella

pneumoniae (K. pneumoniae) is a Gram-negative, facultative

anaerobic and rod-shaped bacterium of the Enterobacteriaceae family.

It is regarded as an opportunistic pathogen that is associated with the

hospital-acquired urinary tract infections, septicemia, pneumonia, and

soft tissue infections [1]. K. pneumoniae is responsible for the

nosocomial outbreaks worldwide, due to its ability to spread rapidly in

the hospital environment [2], and results in high morbidity and

J Health Med Inform

ISSN:2157-7420 JHMI, an Open access journal

Volume 6 • Issue 5 • 206


Citation

Citation:

Trivedi MK, Branton A, Trivedi D, Gangwar M, Jana S (2015) Antimicrobial Susceptibility, Biochemical Characterization and Molecular

Typing of Biofield Treated Klebsiella pneumoniae. J Health Med Inform 6: 206. doi:10.4172/2157-7420.1000206

Page 2 of 7

biofield energy treatment has been well known and studied in the field

of materials science research [14 -16], agricultural science research

[17,18], and microbiology research [19,20].

miniaturized of the broth dilution susceptibility test that had been

dehydrated. Briefly, 100 μL of the standardized suspension of K.

pneumoniae was pipetted into 25 mL of inoculum water using pluronic

and inverted 8-10 times and inoculated, rehydrated, and then

subjected to incubation for 16 hours at 35°C. Rehydration and

inoculation were performed using the RENOK® system with

inoculators-D (B1013-4). The detailed experimental procedures and

conditions were followed as per the manufacturer's instructions.

Briefly,after inoculation and rehydration with a standardized

suspension of K. pneumoniae, it was incubated at 35°C for 16 hours.

MIC and a qualitative susceptibility like susceptible (S), intermediate

(I), and resistant (R) were determined by observing the lowest

antimicrobial concentration showing growth inhibition [24].

Due to the clinical importance of K. pneumoniae and outstanding

results of biofield treatment, the present study was designed to evaluate

the impact of Mr. Trivedi’s biofield energy treatment on K.

pneumoniae with respect to the antimicrobial susceptibility,

biochemical study, and biotype number. Further, in order to study the

phenotypic characteristics of biofield treated K. pneumoniae,

molecular typing using arbitrary amplification of polymorphic DNA

sequences, termed as random amplified polymorphic DNA (RAPD)

analysis was used [21]. RAPD is a preferred technique used in different

studies for typing and discriminating the epidemiology of

microorganism [22]. RAPD has an advantage over other traditional

phenotypic typing methods as it is rapid, relatively inexpensive and

technically feasible [23]. The aim of this study was to evaluate the

impact of Mr. Trivedi’s biofield energy treatment on K. pneumoniae

with respect to antibiogram characteristics and genotyping using

RAPD of the organism.

Biochemical studies

The biochemical reactions of K. pneumoniae were determined by

MicroScan Walk-Away® where, interpretation of biochemical reactions

for microbial identification of Gram-negative organisms [24].

Biotype number

Materials and Methods

The biotype number of K. pneumoniae was determined by

MicroScan Walk-Away® processed panel data utilizing data of

biochemical reactions [24].

K. pneumoniae ATCC 15380 [American Type Culture Collection]

was procured from Bangalore Genei, in sealed pack, and stored as per

the recommended storage conditions for further use. The antimicrobial

susceptibility, minimum inhibitory concentration (MIC), biochemical

reactions, and biotype number were evaluated using automated

MicroScan Walk-Away® system (Dade Behring Inc., West Sacramento,

CA) using Negative Breakpoint Combo 30 (NBPC 30) panel. RAPD

was carried out using Ultrapure Genomic DNA Prep Kit; Cat KT 83

(Bangalore Genei, India). All the tested antimicrobials, biochemicals,

media, and reagents were procured from Sigma-Aldrich, India.

Random Amplified Polymorphic DNA (RAPD) analysis

Three series of inoculums (one for control and other two for

treatment named as treated A and B) were prepared from K.

pneumoniae sample. Two inoculums (treated samples A and B) were

subjected to Mr. Trivedi's biofield energy treatment. Whilst handing

over treated groups to Mr. Trivedi for biofield treatment, optimum

precautions were taken to avoid the contamination. After that, the

treated samples (A and B) were sub-cultured by taking 1% inoculum

and inoculated to fresh 5 mL medium and labeled as treatment A-1

and treatment B-1 respectively. Control and treated samples were

incubated at 37°C with 160 rpm for 18 h. Subsequently, the cultures

were spun down, and genomic DNA was isolated for control and

treated samples using the genomic DNA Prep Kit (Bangalore Genei,

India). The RAPD was performed with all samples of K. pneumoniae

using five RAPD primers, which were labelled as RBA 5A, RBA 10A,

RBA 15A, RBA 21A, and RBA 22A. The PCR mixture contained 2.5 μL

each of buffer, 4.0 mM each of dNTP, 2.5 μM each of primer, 5.0 μL

each of genomic DNA, 2U each of Taq polymerase, 1.5 μL of MgCl2

and 9.5 μL of nuclease-free water in a total of 25 μL mixture. PCR

amplification protocol was followed with initial denaturation at 94ºC

for 7 min, followed by 8 cycles of denaturation at 94ºC for 1 min,

annealing at 35ºC for 1 min, and extension at 72ºC for 2 min; and 35

cycle of denaturation at 94ºC for 1 min, annealing at 38ºC for 1 min,

and extension at 72ºC for 1.5 min; and the final extension at 72ºC for 7

min. Amplified PCR products (12 μL) from all the samples (control

and treated) were separated on 1.5% agarose gels at 75 volts, stained

with ethidium bromide and visualized under UV illumination [25].

Biofield treatment modalities

K. pneumoniae strain was divided into two groups i.e. control and

treated. The treated group was in sealed pack and handed over to Mr.

Trivedi for the biofield energy treatment under laboratory conditions.

Mr. Trivedi provided the treatment through his energy transmission

process to the treated group that includes bioenergy emission of

certain wavelength, which has the ability to do the changes in the

microbes without touching the sample. Mr. Trivedi’s unique energy

treatment is known as The Trivedi Effect®. Mr. Trivedi visited the

laboratory individually over a period of treatment and for control

experiments, nobody entered the experimental room during the

treatment period. Whilst handing over these cultures to Mr. Trivedi for

treatment purposes, optimum precautions were taken to avoid

contamination. After treatment, control and treated groups were

assessed on day 10 for the antimicrobial susceptibility, minimum

inhibitory concentration (MIC), biochemical reactions, biotype, and

genotyping using RAPD analysis. The result of treated sample was

compared with respect to the control.

Investigation of antimicrobial susceptibility assay

The percentage of polymorphism was calculated using following

equation-

Investigation of antimicrobial susceptibility of K. pneumoniae was

carried out with the help of automated instrument, MicroScan Walk-

Away® using Negative Breakpoint Combo 30 (NBPC30) panel as per

the manufacturer’s instructions. The panel was allowed to equilibrate

to room temperature prior to rehydration. All opened panel were used

on the same day. The tests were carried out on MicroScan, which were

Percent polymorphism = A/B×100;

Where, A = number of polymorphic bands in treated sample; and B

= number of polymorphic bands in control.

J Health Med Inform

ISSN:2157-7420 JHMI, an Open access journal

Volume 6 • Issue 5 • 206


Citation 1

Citation:

Trivedi MK, Branton A, Trivedi D, Gangwar M, Jana S (2015) Antimicrobial Susceptibility, Biochemical Characterization and Molecular

Typing of Biofield Treated Klebsiella pneumoniae. J Health Med Inform 6: 206. doi:10.4172/2157-7420.1000206

Page 3 of 7

Results and Discussion

1

Amikacin

≤ 16

≤ 16

2

Amoxicillin/k-clavulanate

≤ 8/4

≤ 8/4

Antimicrobial susceptibility assay

3

Ampicillin/sulbactam

16/8

≤ 8/4

The results of biofield treatment on K. pneumoniae with respect to

antimicrobials susceptibility pattern and MIC are summarized in Table

1 and 2, respectively.

4

Ampicillin

>16

> 16

5

Aztreonam

≤8

≤ 8

S. No.

Antimicrobial

Control

Treated

6

Cefazolin

≤8

≤ 8

1

Amikacin

S

S

7

Cefepime

≤ 8

≤ 8

2

Amoxicillin/k-clavulanate

S

S

8

Cefotaxime

≤ 8

≤ 8

3

Ampicillin/sulbactam

I

S

9

Cefotetan

≤ 16

≤ 16

4

Ampicillin

R

R

10

Cefoxitin

≤ 8

≤ 8

5

Aztreonam

S

S

11

Ceftazidime

≤ 8

≤ 8

6

Cefazolin

S

S

12

Ceftriaxone

≤ 8

≤ 8

7

Cefepime

S

S

13

Cefuroxime

≤ 4

≤ 4

8

Cefotaxime

S

S

14

Cephalothin

≤ 8

16

9

Cefotetan

S

S

15

Chloramphenicol

16

≤ 8

10

Cefoxitin

S

S

16

Ciprofloxacin

≤ 1

≤ 1

11

Ceftazidime

S

S

17

ESBL-a Scrn

≤4

≤ 4

12

Ceftriaxone

S

S

18

ESBL-b Scrn

≤ 1

≤ 1

13

Cefuroxime

S

S

19

Gatifloxacin

≤ 2

≤ 2

14

Cephalothin

S

I

20

Gentamicin

≤ 4

≤ 4

15

Chloramphenicol

I

S

21

Imipenem

≤ 4

≤ 4

16

Ciprofloxacin

S

S

22

Levofloxacin

≤ 2

≤ 2

17

Gatifloxacin

S

S

23

Meropenem

≤ 4

≤ 4

18

Gentamicin

S

S

24

Moxifloxacin

≤ 2

≤ 2

19

Imipenem

S

S

25

Nitrofurantoin

≤ 32

≤ 32

20

Levofloxacin

S

S

26

Norfloxacin

≤ 4

≤ 4

21

Meropenem

S

S

27

Piperacillin/tazobactam

≤ 16

≤ 16

22

Moxifloxacin

S

S

28

Piperacillin

64

64

23

Piperacillin/tazobactam

S

S

29

Tetracycline

≤ 4

≤ 4

24

Piperacillin

I

I

30

Ticarcillin/k-clavulanate

≤ 16

≤ 16

25

Tetracycline

S

S

31

Tobramycin

≤ 4

≤ 4

26

Ticarcillin/k-clavulanate

S

S

32

Trimethoprim/sulfamethoxazole

≤ 2/38

≤ 2/38

27

Tobramycin

S

S

MIC values are presented in µg/mL; ESBL: Suspected extended-spectrum β-

lactamases a, b screen

28

Trimethoprim/sulfamethoxazole

S

S

R: Resistant; I: Intermediate; S: Susceptible

Table 2:

Table 2: Minimum inhibitory concentration (MIC) of tested

antimicrobials against K. pneumoniae

Table 1:

Table 1: Effect of biofield treatment on K. pneumoniae for its

antimicrobial susceptibility

The antimicrobial sensitivity result of three antimicrobials namely

ampicillin/sulbactam, cephalothin, and chloramphenicol showed the

alteration afterbiofield treatment with respect to control among

S. No.

Antimicrobial

Control

Treated

J Health Med Inform

ISSN:2157-7420 JHMI, an Open access journal

Volume 6 • Issue 5 • 206


Citation 2

Citation:

Trivedi MK, Branton A, Trivedi D, Gangwar M, Jana S (2015) Antimicrobial Susceptibility, Biochemical Characterization and Molecular

Typing of Biofield Treated Klebsiella pneumoniae. J Health Med Inform 6: 206. doi:10.4172/2157-7420.1000206

Page 4 of 7

twenty-eight tested antimicrobials. The sensitivity of ampicillin/

sulbactam and chloramphenicol was improved i.e. from intermediate

(I) to susceptible (S), while cephalothin showed an altered sensitivity

nature from S to I. Other tested antimicrobials did not show any

alterations of sensitivity pattern as compared to the control.

6

CF8

Cephalothin

-

+

7

CIT

Citrate

+

+

8

CL4

Colistin

+

-

9

ESC

Esculin hydrolysis

+

+

MIC results were well supported with antimicrobial sensitivity data,

as ampicillin/sulbactam and chloramphenicol showed decrease value

of MIC after the biofield treatment. Ampicillin/sulbactam (i.e. 16/8 to

≤8/4 µg/mL) and chloramphenicol (i.e. 16 to ≤8 µg/mL) showed two

folds change in MIC values as compared to the control. Cephalothin

showed an alteration in MIC value i.e. from ≤8 to 16 µg/mL after

biofield treatment. The rest of the tested antimicrobials did not show

any alteration in MIC values with respect to the control.

10

FD64

Nitrofurantoin

-

-

11

GLU

Glucose

+

+

12

H2S

Hydrogen sulfide

-

-

13

IND

Indole

-

-

14

INO

Inositol

+

+

Efficacy of sulbactam, a β-lactamase inhibitor, in combination with

ampicillin, was well reported and a preferred treatment option against

β-lactam-resistant K. pneumoniae infections. According to Hoffman et

al. the combination of ampicillin/sulbactam was found to have

synergistic effects, which significantly decreased the severity of

pneumonia. Bronchoalveolar lavage cytologic findings, and extent of

macroscopic lesions in lung tissue of the noninoculated regions were

reported as compared to the individual ampicillin or sulbactam [26].

The resistance pattern of ampicillin against K. pneumoniae is because

of two types of chromosomal β-lactamase enzymes being SHV-1 and

LEN-1 [27], and results suggest that biofield treatment might alter

these enzymes and alter the sensitivity pattern. Biofield treatment on

K. pneumoniae has improved the sensitivity profile of ampicillin/

sulbactam and showed decreased MIC value by two-folds, which might

be useful in the future treatment strategy against pneumoniae lung

infection. Multi-drug therapies are another approach against β-

lactamase producing strain of K. pneumoniae, as synergy has been

frequently reported in vitro between β-lactams and aminoglycosides.

According to Jones, a synergistic effect was frequently reported in a

combination of ertapenem and ciprofloxacin [28]. Besides synergistic

effect, antibiotics such as amikacin, gatifloxacin, gentamicin, and

chloramphenicol are the preferred drug of choice with respect to cost,

side effects and many other factors in K. pneumoniae associated

infections [29]. Mr. Trivedi’s biofield energy treatment on K.

pneumoniae showed improved sensitivity pattern of chloramphenicol

and simultaneously decreased the MIC value as compared to the

control. β-lactamases are enzymes that inactivates the β-lactam

containing antibiotic which is present in almost all Gram-negative

bacilli such as E. coli and Klebsiella spp. [30]. Biofield treatment might

alter the production of these enzymes which may result in the

improved sensitivity of antimicrobials.

15

K4

Kanamycin

-

-

16

LYS

Lysine

+

+

17

MAL

Malonate

+

+

18

MEL

Melibiose

+

+

19

NIT

Nitrate

+

-

Oxidation-fermentation/

glucose

20

OF/G

+

+

21

ONPG

Galactosidase

+

+

22

ORN

Ornithine

-

-

23

OXI

Oxidase

-

-

24

P4

Penicillin

+

+

25

RAF

Raffinose

+

+

26

RHA

Rhamnose

+

+

27

SOR

Sorbitol

+

+

28

SUC

Sucrose

+

+

29

TAR

Tartrate

-

-

30

TDA

Tryptophan deaminase

-

-

31

TO4

Tobramycin

-

-

32

URE

Urea

+

+

33

VP

Voges-Proskauer

-

-

Organism identification by biochemical reactions

Table 3:

Table 3: Effect of biofield treatment on K. pneumoniae to the vital

processes occurring in living organisms -: negative; +: positive.

Biochemical study results of control and biofield treated groups are

summarized in Table 3.

The results showed overall 9.09% of tested biochemical altered

reactions as compared to the control. The colistin and nitrate

biochemical showed negative reaction i.e. from (+) positive to (-)

negative as compared to the control. Cephalothin showed positive

reaction i.e. from (-) negative to (+) positive reaction. The rest of tested

biochemicals did not show any alteration in their reaction pattern after

biofield treatment. The biochemical reactions of control K.

pneumoniae were well supported with literature data [31].

S. No.

Code

Biochemical

Control

Treated

1

ACE

Acetamide

-

-

2

ADO

Adonitol

+

+

3

ARA

Arabinose

+

+

4

ARG

Arginine

-

-

5

CET

Cetrimide

-

-

J Health Med Inform

ISSN:2157-7420 JHMI, an Open access journal

Volume 6 • Issue 5 • 206


Citation 3

Citation:

Trivedi MK, Branton A, Trivedi D, Gangwar M, Jana S (2015) Antimicrobial Susceptibility, Biochemical Characterization and Molecular

Typing of Biofield Treated Klebsiella pneumoniae. J Health Med Inform 6: 206. doi:10.4172/2157-7420.1000206

Page 5 of 7

Organism identification by biotype number

Based on the biochemical results, alteration in biotype number was

also observed in the biofield treated K. pneumoniae as compared to the

control. The control group showed biotype number as 7774 4274, while

after treatment altered biotype number was reported as 7774 4272.

However, no change in organism was reported afterbiofield treatment

with respect to the control. Our research group recently reported the

significant alterations in biochemical reactions followed by the change

in biotype number that was also supported with published data

[19,20]. Biofield treatment might alter some enzymatic reactions in K.

pneumoniae, which resulted in alterations in characteristic

biochemical reactions and biotype number.

Random Amplified Polymorphic DNA (RAPD) analysis

Figure 1

Figure 1: Random amplified polymorphic-DNA fragment patterns

of K. pneumoniae generated using five RAPD primers, RBA 5A,

RBA 10A, RBA 15A, RBA 21A and RBA 22A. 1: Control; 2: Treated

A; 3: Treated A-1; 4: Treated B; 5: Treated B-1; M: 100 bp DNA

Ladder.

RAPD has been used as a genotyping tool to that is being used to

correlate the genetic similarity or mutations between species of K.

pneumoniae [32]. DNA fingerprinting analysis of the control and

treated K. pneumoniae was identified on the basis of their different and

discriminative RAPD patterns. The simplicity and wide applicability of

RAPD analysis mainly depend on the use of short nucleotide primers,

which were unrelated to known DNA sequences of the target organism

[21]. DNA polymorphism can be efficiently detected using the PCR

primers and identify the inter-strain variations among species in the

treated samples. The degree of relatedness and genetic mapping can be

correlated between similar or different treated sample species [33].

The polymorphic bands are marked by arrows in the gel image. The

RAPD patterns of treated samples showed some unique, dissimilar,

common, and polymorphic patterns. DNA polymorphism among the

different treated samples compared with the control were analyzed and

presented in Table 4.

In this experiment, DNA fingerprinting of control and treated

samples using RAPD are shown in Figure 1.

Unique band

Nucleotide

(5’-3’)

sequence

Total

polymorphic bands

no.

of

Common bands in

control and treated

S. No.

Primer

Control

TSA

TSA-1

TSB

TSB-1

1

RBA 5A

GTTTCGCTCC

18

11

1

2

1

1

2

2

RBA 10A

GTGGATCCGA

15

5

3

1

0

0

0

3

RBA 15A

GCGATCCCCA

12

8

1

1

0

0

0

4

RBA 21A

CCGCAGCCAA

15

13

0

0

0

0

0

5

RBA 22A

AAGAGCCCGT

14

5

1

1

0

2

0

Table 4:

Table 4: DNA polymorphism of K. pneumoniae analyzed by random amplified polymorphic DNA (RAPD) analysis, TSA: treated sample A;

TSA-1: treated sample A-1; TSB: treated sample B; TSB-1: treated sample B-1.

The level of polymorphism between control and treated samples (A,

A1, B, and B1) are summarized in Table 5.

Primer

C and TSA

C and TSA-1

C and TSB

C and TSB-1

TSA and TSA-1

TSB and TSB-1

TSA and TSB

TSA-1 and TSB-1

RBA 5A

18%

12%

12%

18%

18%

18%

6%

6%

RBA 10A

30%

15%

15%

23%

10%

0%

15%

8%

RBA 15A

18%

27%

27%

9%

11%

0%

9%

15%

RBA 21A

0%

6%

0%

0%

0%

0%

0%

6%

RBA 22A

13%

6%

20%

6%

10%

20%

5%

0%

J Health Med Inform

ISSN:2157-7420 JHMI, an Open access journal

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Citation 4

Citation:

Trivedi MK, Branton A, Trivedi D, Gangwar M, Jana S (2015) Antimicrobial Susceptibility, Biochemical Characterization and Molecular

Typing of Biofield Treated Klebsiella pneumoniae. J Health Med Inform 6: 206. doi:10.4172/2157-7420.1000206

Page 6 of 7

Average polymorphism

15%

13%

14%

11%

9%

7%

7%

7%

Table 5:

Table 5: Level of polymorphism between control and treated K. pneumoniae samples. C: Control; TSA: treated sample A; TSA-1: treated sample

A-1; TSB: treated sample B; TSB-1: treated sample B-1.

The level of polymorphism was found in an average range of 11 to

15% in the treated samples as compared to control after the biofield

treatment. The highest change in DNA sequence was observed in

treated samples with RBA 10A primer as compared to control; whereas

no change was found in treated sample with RBA 21A primer as

compared to control. Thus, results indicates that treatment samples has

genetic variability among organism. RAPD also explains the relevant

degree of genetic diversity, however this technique has the potential to

detected genetic polymorphism throughout the genome [34].

interspecific polymorphic relationship with K. pneumoniae after

biofield treatment. Overall, it seems that Mr. Trivedi’s unique biofield

energy treatment on pathogenic microbes might be used as an

alternate approach to alter the antimicrobial sensitivity.

Acknowledgement

This work was supported by Trivedi Science™, Trivedi Master

Wellness™ and Trivedi Testimonials. Authors acknowledge the

generosity and cooperation of all participating members from PD

Hinduja National Hospital and MRC, Mumbai, Microbiology Lab for

conduction antimicrobial studies. Authors are thankful to Bangalore

Genei Private Limited, for conducting RAPD analysis.

Biofield energy as the complementary medicine is well documented

and considered as alternate medicine approach worldwide. According

to the report of National Health Interview Survey (NHIS), conducted

by the Centers for Disease Control and Prevention's (CDC) and

National Center for Health Statistics (NCHS) till 2007, energy

medicine was practiced almost 4 out of 10 adults in the past 12 months

[35]. Current experiment was designed to demonstrate the impact of

Mr. Trivedi’s biofield treatment on K. pneumoniae for its antimicrobial

susceptibility testing. Further, the molecular methods was performed

to study the genetic alterations and similarities using RAPD

sequencing methods. Increased infection of K. pneumoniae and other

Gram-negative pathogen associated with nosocomial infections have

become a global health problem. Results suggest that biofield

treatment on microorganism can alter the in vitro sensitivity of the

antimicrobials and it might be correlated with acetylation of

antimicrobials that may happen via active drug efflux mechanism.

Increased incidence of nosocomial infections and broad resistance

against broad spectrum antibiotics would be a serious global threat.

Mr. Trivedi’s biofield treatment showed a significant decrease in MIC

values of ampicillin/sulbactam and chloramphenicol along with

improved sensitivity, which suggested an alteration at enzymatic/

genetic level that may modify ligand-receptor interaction. Hence a

cascade of intra-cellular signals may be initiated, accelerated or

inhibited [36], afterbiofield treatment on pathogenic microbes.

Further, biofield treatment on K. pneumoniae, possible involve

alterations at receptor level due to energy transfer via. biofield

treatment, which may change the receptor drug interactions, which in

turn alter the internal state of the microbe. However, it was reported

that electromagnetic fields might alter the transmembrane

concentration of cell, and will alter the receptor protein molecule.

Biofield treatment might alter the receptor interaction, and results in

altered antibiogram of K. pneumoniae with respect to the control [37].

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Typing of Biofield Treated Klebsiella pneumoniae. J Health Med Inform 6: 206. doi:10.4172/2157-7420.1000206

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ISSN:2157-7420 JHMI, an Open access journal

Volume 6 • Issue 5 • 206


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