PREVELANCE OF HELICOBACTER PYLORI INFECTION IN

GALL BLADDER MUCOSA IN PATIENTS UNDERGOING

CHOLECYSTECTOMY

D R . AKSHAYA H B
R E G . N O : 19 011 01 0 0 6 | NO V EM BE R /2 01 9 | A UG U ST / 2 021

COURSE : MS GENERAL SURGERY , MGMCRI

CANDIDATE

 Candidate Name : DR AKSHAYA H B

 Course of Study : MS - GENERAL SURGERY

 University Identity No : 1901101006

 Mobile Phone No : +91 9008500180

 E-mail Address : hbakshaya29@gmail.com

 Month/Yr of Admission : NOVEMBER 2019

 Month/Yr of Examination : MAY 2021

GUIDES

 GUIDE:

 DR. RAMANATHAN M

 Professor

 General Surgery

 Contact Number-9842338502

 Email

 CO GUIDE:

 DR. ARUL KUMAR

 Associate Professor

 General Surgery

 Contact Number-7200708687

 Email

2
 Check list for submission to Ethics committee

Page
number in
S.No Element
which
written
1. Scientific background and explanation of rationale
2. Specific objectives or hypotheses
3. Study Population
4. How sample size was determined
5. Description of study design
6. Eligibility criteria for participants / volunteers
7. Settings and locations where the data will be collected
The interventions for each group with sufficient details
8. to allow replication, including how and when they will
actually be administered
For Randomised trial
Method that will be used to generate the random
9.
allocation sequence
Type of randomization; details of any restriction (such
10.
as blocking and block size)
Mechanism that will be used to implement the random
allocation sequence (such as sequentially numbered
11.
containers), describing any steps taken to conceal the
sequence until interventions were assigned
For blinded trials
Who will generate the random allocation sequence,
12. who will enrol the participants, and who will assign
participants to interventions
If done, who will be blinded after assignment to
13. interventions (for example, participants, care
providers, those assessing outcomes) and how
For Analytical / Observational Studies (STROBE Guidelines)
a) Cohort study—The eligibility criteria, and the
14. sources and methods of selection of participants.
b) Methods of follow-up
Case-control study—
a) The eligibility criteria, and the sources and
15.
methods of case ascertainment and control
selection.

3
 b) The rationale for the choice of cases and controls
Cross-sectional study—
16. a) The eligibility criteria, and the sources and
methods of selection of participants
Cohort study—
17. a) For matched studies, matching criteria and number
of exposed and unexposed
Case-control study—
18. a) For matched studies, matching criteria and the
number of controls per case
For Qualitative Studies(McMaster University)
19. A theoretical perspective is identified
20. The process of purposeful selection is described
21. Is sampling done until redundancy in data is reached?
22. Is Procedural rigor used in data collection strategies?
Is there evidence of the four components of
trustworthiness?
23.
Credibility, Transferability, Dependability,
Confirmability
Outcomes
Completely defined primary and secondary outcome
24. measures, including how and when they will be
assessed
Statistical methods
Statistical methods that will be used to compare groups
25.
for primary and secondary outcomes
Methods for additional analyses, such as subgroup
26.
analyses and adjusted analyses
Consort flow chart
27 Consort flow chart
28. Ethical issues
29. Consent form
30. Patient information sheet in Tamil

Signature of the PG Signature of Guide

4
PART II – THE PROTOCOL

1 INTRODUCTION

Helicobacter pylori (H. pylori), a causative agent of chronic gastric infections, is estimated

to be found in at least half the world population. A meta-analysis on global prevalence

shows an overall prevalence rate of 44.3% with prevalence in south-eastern population of

1,2
43.1 % (31.5−54.8) and a prevalence rate in India of 63.5% (53.4−73.5) . Kawaguchi et

al first detected H. pylori in the gallbladder’s mucosa of a patient with

calculus cholecystitis in 1996 3. The prevalence of H. pylori infection is found to be high in

4,5
patients with cholecystitis and cholelithiasis . Although the role of H. pylori in gastritis,

peptic ulcer disease and gastric malignancy has been established, the role of H. pylori in

cholelithiasis has been controversial with studies for and against the role of H. pylori as a
6-10
predisposing factor for cholelithiasis . Since very few studies have been done to detect

the presence of H. pylori infection in gastric mucosa and gall bladder mucosa, the role of

eradication of gastric H. pylori in prevention of cholelithiasis and the disease burden, it

necessitates the need for further study of H. pylori infection in cholelithiasis 11,12.

Cholelithiasis is one of the most frequent surgical ailments encountered by general

surgeons in clinical practice. Cholelithiasis is usually asymptomatic; about 2-4% develop

symptoms with biliary colic being the most common. Biliary colic is characteristically

right upper quadrant abdominal pain lasting more than half an hour without fever, other

symptoms include upper quadrant pain epigastric pain, intolerance to fatty foods, nausea,

bloating, flatulence, and frothy, foul-smelling stools.

5
 2. AIMS AND OBJECTIVE

AIM: To determine the prevelance of helicobacter pylori infection in gallbladder mucosa

in patients who underwent cholecystectomy

OBJECTIVES:

1.To detect the presence of H. pylori organism in gall bladder mucosa using histology.

2. To measure the frequency of H. pylori infection among gall bladder diseases.

3. To find the composition of gallbladder stone in H.pylori infected patients.

4. To find the virulence of H.pylori stool antigen.

2 REVIEW OF LITERATURE

In a Prospective study by Silva C P, the presence of H. pylori DNA in biliary

epithelium was investigated in Brazilian population with a sample size of 64, 46 with

cholelithiasis and 18 subjects without cholelithiasis by identification of Helicobacter

species culture and nested 16S rRNA in gall bladder and bile. An association was found.

Strengths - prospective study design, accurate selection of controls, adjustment for

confounding factors and detection of Helicobacter DNA. Limitations - small number of

subjects studied, difficulty in obtaining a healthy control group, absence of controlling

for confounding factors or differences among populations 15.

In another study by Bulajic M, the objective of study was to determine whether there

is an association between H. pylori in bile and biliary tract carcinoma. A prospective study,

with a sample size of 89 patients, 63 patients had gallstones, 15 patients had biliary tumors,

6
and 11 patients had neither condition. Strengths - double positive samples from two

laboratories were taken. Limitations – confounding factors and sample size 7.

In a case control study of patients with dyspepsia by Shokry Shirvani J, who

underwent endoscopy ,72 patients with and 136 patients without gall stones were assigned

to case and control groups respectively. 31 (43.1%) out of 72 patients and 45(33.8%) out of

136 patients were positive for H. pylori infection. Mild gastritis- with gall stones is 10

(13.9%) and without stones is 28(20.6%) and moderate/severe gastritis – with stones

21(29.2%) and without stones is 17 (12.5%). H. pylori has been associated with increased

incidence of severe gastritis in comparison. This study shows the increased frequency of H.

pylori infection with stone in gall bladder, though statistically not significant. Limitations

include - sample size, confounding factors for choledocholithiasis 11.

In a study by Wafi Attaallah, the presence of H. pylori in gall bladder mucosa in

symptomatic gall stones were investigated. Out of the 94 patients, 35 patients (37%) gall

bladder mucosa tested positive for H. pylori by any of the methods. H. pylori was positive

in 47 (58.7%) gastric mucosa and in 21 (22%) gall bladder mucosa. In 15 patients (15.9%)

both gall bladder mucosa and gastric mucosa tested positive for H. pylori. Study

demonstrates presence of H. pylori in gall bladder of 37% of symptomatic cholelithiasis.

Strengths – three methods of detection, prospective study. Limitations – sample size and

confounding factors 16.

7
 RESEARCH QUESTION OR HYPOTHESIS

To show the positive association between H.pylori infection in gall bladder and
cholelithiasis using histolopathology.

4.SUBJECTS AND METHODS

STUDY SUBJECTS: All patients who are undergoing cholecystectomy.

TYPE OF STUDY: A cross-sectional study.

STUDY PERIOD: 1.5 years from November 2019 to June 2021

PLACE OF STUDY:DEPARTMENT OF GENERAL SURGERY, MAHATMA

GANDHI MEDICAL COLLEGE AND RESEARCH INSTITUTE

STUDY DESIGN- CROSS SECTIONAL STUDY

SAMPLING METHOD-CONSECUTIVE SAMPLING

STUDY GROUP-ONE

SAMPLE SIZE:

Based on previous study by Wafi Attaallah et al.The sample size is calculated as-

n = Z(1-alpha)2 x P x (1-P) / d2

n = (1-96)2 x 0.587 x 0.413 / (0.1)2

n = 94

Inclusion criteria

1. All patients who are undergoing cholecystectomy.

2. Patient willing to give informed consent.

Exclusion criteria

1. Patient not willing for informed consent.

2. Patients with present or prior treatment with Anti H. pylori regimen.

8
METHODOLOGY
The patients are diagnosed with cholelithiasis by ultrasonography and undergoing

laparoscopic or open cholecystectomy, the gall bladder specimen is subjected to histopathological

examination. The prevelance of H. pylori infection in gall bladder mucosa is detected. The

relationship of gall bladder histopathology and its association with H. pylori is assessed. The

frequency of H. pylori induced cholelithiasis is tabulated.

FLOW-CHART TO SUMMARIZE THE SEQUENCE OF EVENTS

PATIENTS WHO ARE UNDERGOING LAPAROSCOPIC/OPEN

CHOLECYSTECTOMY ARE INCLUDED

SAMPLE TAKEN FROM GALLBLADDER MUCOSA ARE SUBJECTED TO

HISTOPATHOLOGICAL EXAMINATION

THE PREVELANCE OF H. PYLORI IN GALLBLADDER MUCOSA OF

CHOLECYSTECTOMY PATIENTS IS ESTABLISHED

COMPOSITION OF GALLBLADDER STONE IN H PYLORI INFECTED

PATIENTS IS DETERMINED

VIRULENCE AND STOOL ANTIGEN FOR H. PYLORI IS TESTED

9
 POSITIVE ASSOCIATION BETWEEN PRESENCE OF H.PYLORI IN GALL

BLADDER MUCOSA AND CHOLELITHIASIS IS ESTABLISHED.

3 STUDY VARIABLES

Data will be analized by descriptive statistics – Mean, median, standard

deviation, interquartile range, percentages . Chi square test, sensitivity ,

specificity will be used to see the association between Qualitative variables.

P<0.05 will be considered statistically significant. Data will tabulated using

Microsoft excel and analysed.

1. HISTOLOGY

2. STOOL ANTIGEN

3. COMPOSITION OF GALL STONE

4. VIRULENCE

Name of the dependent [ Scale of measurement ( Descriptive Inferential

independent variables. Quantitative / Qualitative) Starter to be used

Histology Qualitative Sensitivity, specificity,

Gall bladder mucosa mean, median, standard
deviation, interquartile
range, percentage, chi
square chart

10
Type of gallbladder stone Qualitative Sensitivity, specificity,
mean, median, standard
deviation, interquartile
range, percentage, chi
square chart
Virulence of H pylori Qualitative Sensitivity, specificity,
mean, median, standard
deviation, interquartile
range, percentage, chi
square chart
H pylori stool antigen Qualitative Sensitivity, specificity,
mean, median, standard
deviation, interquartile
range, percentage, chi
square chart

4 ETHICAL ISSUES

Gall bladder sample from all patients who underwent cholecystectomy are subjected to

histology and stool sample for H pylori antigen after taking informed consent for the same

in the language they best understand. This involves minimal risk according to ICMR

guidelines.

5 INFORMED CONSENT PROCEDURE

Participant will be explained in detail , in the language they best understand, regarding all

details that will be obtained from them for the study.

11
 6 REFERENCES

1.Zamani M, Ebrahimtabar F, Zamani V, Miller WH, Alizadeh-Navaei R, Shokri-Shirvani

J, et al. Systematic review with meta-analysis: the worldwide prevalence of Helicobacter

pylori infection. Aliment. Pharmacol. Ther. 2018 Apr;47(7):868–76.

2. Hooi JKY, Lai WY, Ng WK, Suen MMY, Underwood FE, Tanyingoh D, et al. Global

Prevalence of Helicobacter pylori Infection: Systematic Review and Meta-Analysis.

Gastroenterology. 2017 Aug;153(2):420–9.

3. Moricz A de, Melo M, Castro AM, Campos T de, Silva RA, Pacheco Jr AM. Prevalence

of Helicobacter spp in chronic cholecystitis and correlation with changes on the

histological pattern of the gallbladder. Acta Cir Bras. 2010 Jun;25(3):218–24.

4. Cen L, Pan J, Zhou B, Yu C, Li Y, Chen W, et al. Helicobacter Pylori infection of the

gallbladder and the risk of chronic cholecystitis and cholelithiasis: A systematic review

and meta-analysis. Helicobacter. 2018 Feb;23(1):e12457.

5. Chen DF, Hu L, Yi P, Liu WW, Fang DC, Cao H. H.pylori exist in the gallbladder

mucosa of patients with chronic cholecystitis. World J Gastroenterol. 2007;13:1608–11.

6. Bhandari A, Crowe SE. Helicobacter pylori in Gastric Malignancies.Curr Gastroenterol

Rep. 2012 Dec;14(6):489–96.

12
7. Bulajic M, Maisonneuve P, Schneider-Brachert W, Müller P, Reischl U, Stimec B, et al.

Helicobacter pylori and the risk of benign and malignant biliary tract disease: Helicobacter

pylori in Biliary Carcinoma. Cancer. 2002 Nov 1;95(9):1946–53.

8. Abayli B, Colakoglu S, Serin M, Erdogan S, Isiksal YF, Tuncer I, et al. Helicobacter

pylori in the etiology of cholesterol gallstones. J Clin Gastroenterol. 2005 Feb;39(2):134–

7.

9. Roosendaal R, Kuipers EJ, Vandenbroucke-Grauls CM, Kusters JG. Helicobacter

species are not detectable by 16S rDNA PCR in bile from Dutch patients with common

bile duct stones. Digestion. 2002;66:89-91.

10. Méndez-Sánchez N, Pichardo R, González J, Sánchez H, Moreno M, Barquera F, et al.

Lack of association between Helicobacter sp colonization and gallstone disease. J Clin

Gastroenterol. 2001 Feb;32(2):138–41.

11. Shirvani, J.S, S. Siadati, and M. Molai, The Frequency of Helicobacter pylori Infection

in Gastric Biopsies of Patients with Gallbladder Stones. Govaresh, 2014. 19(3): p. 208-

211.

12. Javaherzadeh M, Shekarchizadeh A, Sabet B, Mousavi-Almaleki SA, Mirafsharieh A.

Simultaneous Helicobacter Pylori Infection in Gastric Mucosa and Gallbladder Mucosa in

Patients with Cholecystitis; Is There Any Relationship?. AJS. 3(1-2):12-4.

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13. James S.Dooley , Kurinchi S. Guruswamy ,Brian r. Davidson .Gallstones and Benign

Biliary Disease. James S .Dooley, editor. Sherlock’s Diseases Of Liver and Biliary System

, 13th ed . West Sussex, UK :John Wiley & Sons Ltd ;2018 .14. p. 264-65.

14. Dzierżanowska-Fangrat K, Lehours P, Megraud F, Dzierżanowska D. Diagnosis of

Helicobacter pylori Infection. Helicobacter. 2006 Oct;11(s1):6–13.

15. Silva CP, Pereira-Lima JC, Oliveira AG, Guerra JB, Marques DL, Sarmanho L, et al.

Association of the Presence of Helicobacter in Gallbladder Tissue with Cholelithiasis and

Cholecystitis. J. Clin. Microbiol.2003 Dec 1;41(12):5615–8.

16. Attaallah W, Yener N, Ugurlu MU, Manukyan M, Asmaz E, Aktan AO. Gallstones

and Concomitant Gastric Helicobacter pylori Infection. Gastroenterol Res Pract.

2013;2013:1–4.

 QUALITY CONTROL

Name of Officer designated by the department for quality control:

Designation:

14
Telephone No:

E-mail:

 SPONSORSHIPS

NONE

15
  INVESTIGATORS DECLARATION

This is to certify that the protocol entitled ―PREVELANCE OF HELICOBACTER

PYLORI INFECTION IN GALL BLADDER MUCOSA IN PATIENTS

UNDERGOING CHOLECYSTECTOMY‖ was reviewed by us for submission to the

SBV Institutional Ethics Committee and certified that this protocol represents an accurate

and complete description of the proposed research. We have read the ICMR guidelines,

ICP-GCP guidelines/CPCSEA guidelines/and other applicable guidelinesand undertake to

ensure that the rights and welfare of the study subjects are protected.

The study will be performed as per the approved protocol only. If any deviation is

warranted, the same will be presented to the ethical committee and permission will be

sought. We assure that the study will be terminated immediately in case of any unforeseen

adverse consequences and we will inform the same to the ethical committee immediately.

Dr.RAMANATHAN M
Professor
Department of General Surgery
Guide DD/MM/YYYY

Dr. ARUL KUMAR

Associate Professor
Department of General Surgery
Co-guide DD/MM/YYYY

Dr.AKSHAYA H B
Department of General Surgery
CANDIDATE DD/MM/YYYY

Dr.GANESH BABU
Head ofDepartment
Department of General Surgery
DD/MM/YYYY

16
Appendix

 Blood investigations: complete hemogram, random blood sugar, liver function

test, Renal function test, HbsAg, HIV, HCV, fasting lipid profile.

 Urine: Albumin, sugar, Microscopy,

 Radiological investigations: X ray Chest PA view, x ray erect abdomen,

Ultrasound abdomen and pelvis.

 Histopathological examination of cholecystectomy specimens

 Composition of gallbladder stone, virulence of H pylori and H. pylori stool antigen

test

 No animals are involved as part of study.

INFORMATION SHEET FOR THOSE WHO PLAN TO PARTICIPATE IN THE

RESEARCH PROJECT

NAME OF THE RESEARCH PROJECT

- THE PREVELANCE OF HELICOBACTER PYLORI INFECTION IN GALL

BLADDER MUCOSA IN PATIENTS UNDERGOING CHOLECYSTECTOMY.

We welcome you and thank you for having accepted our request to consider whether you
can participate in our study. This sheet contains details the details of the study; the possible
risks , discomfort and benefits for the participants are also given.

You can read and understand by yourself; if you wish , we are ready to read and explain the
same to you.

If you do not understand anything or if you want any more details we are ready to provide
the details.

INFORMATION TO THE PARTICIPANTS

What is the purpose of this study?

17
 This study is being conducted not only as a part of the syllabus of post graduate course, but
also to effectively come out with results favourable in diagnostic and therapeutic work up of
patients in the future in this institution.

Who/where this study is being conducted?

This study is being conducted by DR.AKSHAYA H B a post graduate medical student

belonging to GENERAL SURGERY department under the guidance of DR.
RAMANATHAN M

Why I am being considered as one of the participant?

You have been diagnosed to have gallbladder stones and undergoing surgery for the same.
We want to know whether you have concomitant H pylori infection in your gallbladder
mucosa which would have predisposed to form gallbladder stones. So that we can treat H
pylori in the early stages and prevent the formation of gallstones in other patients.

Should I definitly have to take part in this study?

No. If you do not wish to participate you will not be included in the study. Also you will
continue to get medical treatment without any prejudice.

If I am participating in this study, what are my responsibilities?

You may have to follow some simple rules.

These are- To cooperate in some basic clinical examinations and giving consent for taking
your gallbladder sample and stool sample for study.

Will there be any discomfort /risks to me?

No, gall bladder sample will be subjected to histological examination and this will involve
minimal risk.

Will I paid for the study?

No.I will not be paid.

If I participate in this study, my personal details will be kept confidentially?

Yes, confidentially will be maintained.

Will I be informed of this study results and findings?

Yes, you will be informed about the results and findings.

18
Can I withdraw from this study at any time during the study period?

Yes. You can withdraw at any time during the study period.

19
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n`spNfhgf;NlhH igNyhhp Neha;jhf;fj;ij fz;lwpAk; XH Ma;T.

,e;j Muha;r;rp gw;wpa tptuq;fs; vdf;Fj; njhptpf;fg;gl;lJ. ,J Fwpj;J vd;Dila

nrhe;j nkhopapNyNa vdf;F tpthpf;fg;gl;lJ. ,J Fwpj;J re;Njfq;fspypUe;J njspT
ngWtjw;fhd tha;g;Gk; vdf;F mspf;fg;gl;lJ. kUj;Jt Ma;TfSf;Fg; gpd;G chpa
rpfpr;ir mspf;fg;gLk; vd;gij mwpNtd;. vd;Dila rpfpr;irf;Fg; ghjfk; VJk; ,y;yhky;>
,e;j Muha;r;rpf;fhd rk;kjj;ij kWg;gjw;Fk; vdf;F chpikAz;L vd;gjid ehd; mwpNtd;.
,e;j Muha;r;rpapd; tptuq;fs; kw;Wk; KbTfs; kUj;Jt mwptpay; Nehf;fj;jpw;fhf kl;LNk
gad;gLj;jg;gLk; vd;gjw;F cl;gl;L> mj;jifa tptuq;fs; kw;Wk; KbTfSf;F jilNaJk;
nra;a khl;Nld; vd cld;gLfpNwd;.

……………………………………..Mfpa ehd; ,e;j Muha;r;rpapy; gq;Nfw;gjw;Fr;

rk;kjk; mspf;fpNwd;.

gq;FngWgthpd;
ifnahg;gk;…………………………………….Njjp…………….…………………….
Kfthp…………………………………………………………………………………
………………………………………………………………………………………
rhl;rpahshpd;
ifnahg;gk;……………………………….Njjp……………………………..
Kfthp…………………………………………………………………………………
………………………………………………………………………………………
Muha;r;rpahshpd; ifnahg;gk;………………………………………….….Njjp……………………………..

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