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Centre for Health & Population Research

RRC APPLICATION FORM









FOR OFFICE USE ONLY


R
ESEARCH PROTOCOL






RRC Approval:

Yes /
No Date: 21.06.06

NUMBER: 2006
-
027




ERC Appro
val:

Yes /
No Date: 11.07.06


AEEC Approval:

Yes /
No Date:







Project Title:

Use of Neem oil and insecticide
-
treated bednet (ITN) to control
visceral leishmaniasis (VL)

in an
endemic area of
Bangladesh

Short protocol title (in 50 characters
including space
):

Neem
oil
and ITN
for prevention of VL


Theme:

(
Check all that apply)


Nutrition







Environmental Health


Emerging and Re
-
emerging Infectious Diseases



Health Services


Population Dynamics






Child Health


Reproductive Health






Clinical Case Management


Vaccine evaluation






Social and Behavioural Sciences


HIV/AIDS



Key words:

Disease control, Epidemiology, Visceral l
eishmaniasis,
S
and fly,
Plebotomus argentipes
, Neem
, ITN

1


Relevance of the protocol:
Visceral leishmaniasis or Kala
-
azar is caused by a protozoan parasite,
Leishmania
donovani
and transmitted by a sandfly specie
s
Phlebotomus argentipes
. It is endemic in Bangladesh and more than
50% of kala
-
azar cases are reported from Mymensingh district. After DDT has been withdrawn from indoor
spraying from the country the incidence of kala
-
azar is on the rise. Throughout the
World delamethrin impregnated
bed nets (ITN) are consider a cost effective control from vectors like mosquitoes and sandflies. Neem oil has
microbiocidal activities against both adult and larval stages of various insects. In a preliminary study, Neem oil
was
found to be effective against
P. argentipes
. The proposed study will be conducted on 3400 individuals of 3400
households at two different union of Trishal Upazila at baseline. 2960 individuals of 2960 households expected to
be enrolled in the interven
tion study. Two different interventions (Neem oil and ITN) will be given. We will
randomly select 740 households in each intervention group and 1480 households for control. The objective of this
study is to

compare the incidence of visceral leishmasnias
is and density of vector
P. argentipes
in Neem oil
-
sprayed, ITN user and control houses in Trishal.


Health education for Neem oil application, Neem tree plantation
and Neem oil processing from Neem seed will be conducted in Trishal, if Neem oil is found t
o be effective in
reducing the incidence of visceral leishmaniasis and density of
P. argentipes

at the 3
rd

year of the project.
Similarly,
if ITN is found effective then a programme will be taken to promote ITN in the study areas.



Centre’s priority:

(a
s per strategic plan):
Code no: 22 : Epidemiology and burden of kala azar and identify
effective strategies for prevention and control.



Programmes


Child Health Programme





Health and Family Planning SystemsProgr
amme


Nutrition Programme






Population Programme


Programme on Infectious Diseases & Vaccine Science


Reproductive Health Programme


Poverty and Health Prog
ramme




HIV/AIDS Programme








Revised on: 6
th

January 2005


2



Principal Investigator:

Internal







Mohammad Shafiul Alam

Division:
Clinical Sciences Division

/Laboratory Sciences Division


Address:
ICDDR,B, Mohakhali, Dhaka 1212, Bangladesh

Phone:
880
-
2
-
8860523
-
32 Ext.2333/2411

Email:

sujon2u@yahoo.com







Gender:
Male


Co
-
Principal Investigator(s):

Internal

Rashidul Haque, MBBS, PhD

Division:
Laboratory Sciences Division


Address:
ICDDR,B, Mohakhali, Dhaka 1212, Bangladesh

Phone:
880
-
2
-
8860523
-
32 Ext.2333/2411

Email:

rhaque@icddrb.org







Gender:
Male

Co
-
Principal Investigator(s):

External

(Please provide full official address and Gender)

Yukiko Wagatsuma, M
.
D
.
, Dr.P
.
H
.
,

Address:
Graduate School of Comprehensive Human Sciences,

University o
f Tsukuba

1
-
1
-
1 Tennodai

1
-
1
-
1, Tsukuba, Ibaraki

305
-
8575
, Japan,



E
-
mail
: ywagats@md.tsukuba.ac.jp







Gender:
Female


Co
-
Investigator(s):

Internal

Dr. Kazi Mohammad Asif Jamil

Division:
Clinical Sciences Division


Address:
Clini
cal Sciences Division
,
ICDDR,B, Mohakhali, Dhaka 1212, Bangladesh

Phone:
880
-
2
-
8860523
-
3 Ext.2333

Email:

jamil@icddrb.org








Gender:
Male


Co
-
Investigator(s):

External

Makoto Itoh

Address:

Department of Parasi
tology


Aichi Medical University


Nagakute, Aichi 480
-
1195, Japan

E
-
mail:

machito@aichi
-
med
-
u.ac.jp


Gender:
Male

Co
-
Investigator(s):

External

Mohammad Zahidul Islam

Address:
Department of Parasitology


Aichi Medical University


Nagakute, Aichi 480
-
1195,

Japan








Gender:

Male

Student Investigator/Intern:

External: 2

1. Masaaki Takinami, School of Medicine, University of Tsukuba,


1
-
1
-
1 Tennodai 1
-
1
-
1, Tsukuba, Ibaraki 305
-
8575, Japan




Gender:
Male

2.
Mariko Doi, Doctoral student, Graduate Schoo
l of Comprehensive Human Sciences, University of Tsukuba,


1
-
1
-
1 Tennodai

1
-
1
-
1, Tsukuba, Ibaraki

305
-
8575
, Japan




Gender:

Female




Student Investigator/Intern:

None

Internal (Centre’s staff)




3


Collaborating Institute(s):

University of Tsukuba

and
Aichi Medical University


Please provide full address



Country: Japan




Contact persons:

1.
Yukiko Wagatsuma


Professor


Graduate School of Comprehensive Human Sciences


University of Tsukuba





2.
Makoto Itoh


A
ssociate P
rofessor, Aichi Medical University




School of Medicine, Nagakute, Aichi
-
ken, Japan






Population: Inclusion of special groups
(
Check all that apply)
:

Gender








Pregnant Women


Male







Fetuses


Females








Prisoners


Age








Destitutes


0


5 years







Service providers


5


9 years







Cognitively Impaired


10


19 years






CSW


20


64 years






Others (specify
____________________________)


65 +








Animal


NOTE:
It is the policy of the Centre to include men, women and children in all biomedical and behavioural
research projects involving human subjects unless a clear and compelling rationale and justification
(e.g. gender specific or inappropriate w
ith respect to the purpose of the research) is there.
Justification be provided in case inclusiveness of study participants is not proposed in the study.



Project / study Site (
Check all the apply)
:



Dhaka Hospital






Mirsarai


Matlab Hospital






Patyia


Matlab DSS area






Other areas in Bangladesh:
Trishal


Matlab non
-
DSS area






Outside Bangladesh


Mirzapur








name of country: ________________________


Dhaka Community






Multi centre trial


Chakaria






(Name other countries involved)

________________


Abhoynagar









Type of Study
(Check all that apply)
:



Case Control study






Cross sectional survey


Community based trial / intervention




Longitudinal Study (cohort
or follow
-
up)


Program Project (Umbrella)





Record Review


Secondary Data Analysis





Prophylactic trial


Clinical Trial (Hospital/Clinic)






Surveillance / monitoring


Family follow
-
up study





Others



NOTE:
All clinical studies/trials should be registered in appropriate websites, preferably at
www.clinic
altrials.gov
. When the study is registered in website(s), the PI should provide
website address, registration number, and date of registration to the Committee Coordination
Secretariat for entering these information into the Centre’s database of your res
earch
protocol.


4


Targeted Population
(Check all that apply)
:


No ethnic selection (Bangladeshi)




Expatriates


Bangalee








Immigrants


Tribal groups







Refugee


Consent Process
(Check all that apply)
:



Written







Bengali language


Oral








English language


None


Proposed Sample size:




To
tal sample size at base line: 3400 households (3400 people)


Total sample size for intervention study:

2960 households (2960 people)



Sub
-
group:

Neem oil Intervention: 740 households (740 people
)



ITN intervention:

740 households (740 people
)



Control: 1480 households (1480 people
)


For sandfly collection: 200 households

Neem oil Intervention: 50 households




ITN intervention: 50 households



Control: 100 households




D
etermination of Risk: Does the Research Involve
(Check all that apply)
:



Human exposure to radioactive agents?




Human exposure to infectious agents?


Fetal tissue or abortus?





Investigational new drug


Investigational new device?





Existing data available via public
archives/source (specify____________________)


Pathological or diagnostic clinical specimen only


Existing data available from Co
-
investigator




Observation of public behaviour









New treatment regime

Yes/No



Is the information recorded in such a manner that
study participants

can be identified from information
provided directly or through identifiers linked to the subjects?




Does the resea
rch deal with sensitive aspects of the
study participants

behaviour; sexual behaviour, alcohol
use or illegal conduct such as drug use?


Could the information recorded about the individual if it became known outside of the research:




a. place the
study participants

at risk of criminal or civil liability?



b. damage the
study participants

financial standing, reputation or employability; social rejection, lead



to stigma, divorce etc.



Do you consider this research
(Check one)
:



greater than minimal risk





no more than minimal risk



only part of the diagnostic test


Minimal Risk is "a risk

where the probability and magnitude of harm or discomfort anticipated in the proposed
research are not greater in and of themselves than those ordinarily encountered in daily life or during the
performance of routine physical, psychological examinations o
r tests. For example, the risk of drawing a small
amount of blood from a healthy individual for research purposes is no greater than the risk of doing so as a part of
routine physical examination".


5





Yes/No




Is the

proposal funded?


If yes, sponsor Name:

National Science Fund of Japan




Yes/No




Is the proposal being submitted for funding ?



If yes, name of funding agency: (1) ___________________________________________
____________








(2) _______________________________________________________


Do any of the participating investigators and/or their immediate families have an equity relationship (e.g.
stockholder) with the sponsor of the project or manufactu
rer and/or owner of the test product or device to
be studied or serve as a consultant to any of the above?


IF YES, submit a written statement of disclosure to the Executive Director.



Dates of Proposed Period of Support Cost Requir
ed for the Budget Period ($)



(Day, Month, Year
-

DD/MM/YY)


Direct Cost

Indirect Cost

Total Cost



Beginning date
-

01.07.06


Year
-
1



8,075


807

8,882







Year
-
2




12,614

1,261



13,876



End date
-


28.02.09


Year
-
3


10,339



1,034

11,373







TOTAL:

31,028


3,103


34,131


Approval of the Project by the Division Director of the Applicant


The above
-
mentioned project has been dis
cussed and reviewed at the Division level as well by the external
reviewers.

The protocol has been revised according to the reviewer’s comments and is approved.



_
________________________ _____________________ _________________
______

Name of the Division Director Signature Date of Approval



Certification by the Principal Investigator



Signature of PI

I certify that the statements herein are true, complete

and accurate to the best of my knowledge. I am aware



Date:

that any false, fictitious, or fraudulent s
tatements or

claims may subject me to criminal, civil, or administra
-



Name of Contact Person
(if applicable)

tive penalties. I agree to accept responsibility for the

scientific conduct of the project and to provide the re
-



quired progress reports

if a grant is awarded as a result



of this application.














6







Table of Contents







P
age Numbers

Face Page………………………………………………………………………………… 01

Project Summary……………………………………………………………………….... 07

Description of the Research Project
…………………………………………………… 08


Hypothesis to be tested……………………………………………………………… 08


Specific Aims ………………
.………………………………………………………… 08


Background of the Project Including Preliminary Observations……………………… 09


Research Design and Methods………………………………………………………… 12


Facilities Available…………………………………………………………………… 19


Data Analysis…………………………………
…………………………………………20


Ethical Assurance for Protection of Human Rights…………………………………… 21


Use of Animals………………………………………………………………………… 21


Literature Cited……………………………………………………………………….. 22


Dissemination and Use of Findings……………………………………
……………… 25
Collaborative Arrangements……………………………………………………..…… 25

Biography of the Investigators
………………………………………………………… 26

Deta
i
led Budget
…………………………………………….……………………………… 38

Budget Justifications
……………………………………………………………………… 39

Other Support
……………………………………………….…………………………… 39


Appendix…..
………………………………………………………………………………….

..................40


Consent Forms in English









Check here if appendix is included







7


P
ROJECT SUMMARY:

Describe in concise terms, the hypothe
sis, objectives, and the relevant background of the
project. Describe concisely the experimental design and research methods for achieving the objectives. This
description will serve as a succinct and precise and accurate description of the proposed resear
ch is required. This
summary must be understandable and interpretable when removed from the main application.
(TYPE TEXT WITHIN
THE SPACE PROVIDED).


Principal Investigator
: Mohammad Shafiul Alam

Project Name
:


Use of Neem oil and insecticide
-
treated bed
net (ITN) to control
visceral leishmaniasis (VL)

in an
endemic area of
Bangladesh

Total Budget Beginning Date
Ending Date


$
34,131


01/07/2006



28
/02/2009


Hypothesis.

R
egular

use of Neem oil and ITN will
reduce the risk of
visceral
leishmaniasis
(VL) and the
density of the vector
P. argentipes
in th
e VL endemic

area
s

of Bangladesh.


Objectives.

This study will make a contribution
for

prevention
of VL

in rural areas because
both ITN and
Neem oil spraying is
a
low
-
cost
measure
. For this purpose, we
will determine whether
regular
use of ITN
and
sprayin
g
of Neem oil
reduce
s

the
incidence

of
VL and its vector density
.



Background.

Although leishmaniasis
can be largely prevented
in the Indian subcontinent by building
concrete

house
s,

most of the affected population reside in the rural areas and cannot af
ford to do so
.
Spraying
insecticide

is
known to be
a cost
-
effective way to prevent this disease. However, insecticide
s

such
as DDT has been banned
in certain regions including Bangladesh
due to
its toxic effects causing
ecological
problem
s
.
ITN has shown a

low cost and effective control for Malaria in Africa and also shown promising
results for the control of leishmaniasis.
Neem oil,

a natural pesticide, extracted from Neem seed
s

contains

azadirachtin


that has gained the recognition as an agrichemical by

FDA in USA and Japanese Ministry of
Health, Labor and Welfare. Azadirachtin has an effect of preventing larvae from feeding. It has been shown
that many kinds of insect
s

such as
malaria

mosquitoes are sensitive
to

Neem oil. There are some
reports
suggest
i
ng a role of
Neem oil against
Phlebotomus argentipes
in
vitro
. Our preliminary study
conducted
in
a VL endemic

area of Bangladesh showed that Neem oil
could reduce vector density
in the targeted areas 7
days
after
spraying.


Methods.

Initially we will con
duct a baseline serosurvey among single individuals from 3400 households
which will be selected randomly. 2960 seronegative

individuals

expected to be enrolled after baseline and
will be
follow
ed

for a period of
two years to observe
the conversion to serop
ositive state
using the rapid
dipstick test

(rK39 dipstick).

We will also collect urine samples to conduct urine based antibody detection
(urine DAT). After baseline Neem oil will be sprayed in 740 households and ITN will be distributed
another 740 househ
olds. Remaining 1480 households will serve as control (equal to total interventions).
Follow
-
up serological survey of
the

subjects will be repeated in July
-
August 2007 and 2008. If a person
being tested develop
s

clinical visceral
leishmasniasis
,
he/she wil
l be referred

to the nearby
Government

treatment facility.
Sandfly will be collected from 200 randomly selected households by CDC light trap
immediately after baseline and prior to interventions. Then one month after intervention, and also during
Aug
-
Sep

p
eriod
in 2007 and

2008
.
This intervention study will evaluate the effect of
ITN and
spraying
Neem oil
at
the
field level

and allow us to identify

a cost
-
effective method
for prevention of VL
in
the
endemic area
s
.


KEY PERSONNEL (List names of all investi
gators including PI and their respective specialties)

Name


Professional Discipline/ Specialty Role in the Project

1. Mohammad Shafiul Alam



Parasitology





PI

2. Rasidul H
aque




Parasitology





Co
-
PI

3. Yukiko Wagatsuma



Epidemiology





Co
-
PI

4. Kazi M. Jamil




Medicine/nutrition





Co
-
investigator

5. Makoto Ito




Parasitology





Co
-
investigator

6. Mohammad Zahidul Islam



Parasitology





Co
-
investigator


8


DESCRIPTI
ON OF THE RESEARCH PROJECT

Hypothesis to be tested:


Concisely list in order, in the space provided
,

the hypothesis to be tested and the Specific Aims of the proposed
study. Provide the scientific basis of the hypothesis, critically examining the observat
ions leading to the formulation
of the hypothesis.



R
egular

use of Neem oil and ITN will
reduce the risk of
visceral
leishmaniasis
(VL) and the density of
the vector
P. argentipes
in the VL endemic

area
s

of Bangladesh.


Specific Aims:


Describe the spec
ific aims of the proposed study. State the specific parameters, biological functions/ rates/ processes
that will be assessed by specific methods

(TYPE WITHIN LIMITS).



Primary:


1.

To compare the incidence of visceral leishmasniasis and density of vector
P.

argentipes
between interventions (Neem oil sprayed and ITN user) and control houses in Trishal.



Secondary:


1.

If Neem oil is found to be effective in reducing the incidence of visceral leishmaniasis and the
density of the vector
P. argentipes
, then health

education for Neem oil application, Neem tree
plantation and Neem oil processing from Neem seeds will be conducted during the 3
rd

year of the
project.

Similarly, if ITN is found effective then a programme will be taken to promote ITN in the
study areas.

9


B
ackground of the Project including Preliminary Observations


Describe the relevant background of the proposed study. Discuss the previous related works on the subject by citing
specific references. Describe logically how the present hypothesis is support
ed by the relevant background
observations including any preliminary results that may be available. Critically analyze available knowledge in the
field of the proposed study and discuss the questions and gaps in the knowledge that need to be fulfilled to a
chieve
the proposed goals. Provide scientific validity of the hypothesis on the basis of background information. If there is
no sufficient information on the subject, indicate the need to develop new knowledge. Also include the

significance
and rationale

o
f the proposed work by specifically discussing how these accomplishments will bring benefit to
human health in relation to biomedical, social, and environmental perspectives.
(DO NOT EXCEED 5 PAGES, USE
CONTINUATION SHEETS).






Epidemiology

Leishmaniasis is a worldwide disease; affecting more than 12 million people. Leishman
iasis has some
kinds of pattern and each is distributed in limited areas.
1
Visceral Leishmaniasis (VL) is wide
-
spreading
in the east of Indian subcontinent, Bangladesh, India, and Nepal.
The reported incidence in Bangladesh
has increased in the last deca
de from 3,978 cases in 1993 to 8,920 cases in 2004. Official surveillance
figures thought to be substantial underestimates, with a true incidence of 15,000
-
30,000 per year. In
Bangladesh, 20 million people are considered to be at risk for visceral leishm
aniasis. The reported
incidence in Bangladesh has increased in the last decade from 3,978 cases in 1993 to 7,640 cases in
2000. Official surveillance figures thought to be substantial underestimates, with a true incidence of
15,000
-
30,000 per year. In Ba
ngladesh, 20 million people are considered to be at risk for visceral
leishmaniasis.
In Bihar, next to the border of Bangladesh, about 50,000 cases occurred in 1978. And the
epidemic has spread to West Bengal; approximately 7,500 cases were reported in fir
st 8 months of 1982.
In India, for 14 years; 1977
-
1990, three billion people are infected and 2% were dead.

2


Taxonomy

Leishmania

spp. are intermediated by two genus,
Phlebotomus

in the Old world and
Lutzomyia

in the
New World. Although there are three ge
nera in sandfly in the Old world;
Phlebotomus
,
Sergentomyia

and
Chinius
, genus

Phlebotomus

is the only vector of
Leishmania
. Although genus
Phlebotomus

has six
subgenera,
P. Euphlebotomus argentipes

is the only vector in the east of Indian subcontinent.

3

However
there are necessities to pay attention on vector, like

P. papatasi
being suspected to have
L. donovani

in
Bihar, India.

4



Geography

Sandflies are thought to be weak to aerial plankton that they can not fly high. They also have a liking
for high
humidity, best is 80
-
100%.

7

In Bangladesh, the geographic and climate conditions are at less
than 600m above sea level, heavy rainfall of 250
-
400cm a year (North West), a mean humidity above
70%, a maximum temperature of 29.0°C and a minimum temperature o
f 18.9° C which indicated an
optimum environment for sandflies.


Breeding site

Adult
P. argentipes

in India are detected in human house, horse shelter and cow shed, and immature
stages are recovered in domestic and peridomestic area such as abandoned build
ings, basements and
cellars of houses, cracks in mud floors and walls, soil in human dwellings, animal burrows, animal
shelters, caves, chicken coops, debris and soil cracks, dry excreta of small domestic animals, earth dyke,
embankments, latrines, rotted
manure, rubbish in the street, soil at the base of old walls, under stones
and wells.

8
-
11

But considering the number of detecting immature stages by soil incubation, the biology of
sandflies larva can be said not to be well known.







10



PKDL

Post kala
-
aza
r dermal leishmaniasis (PKDL) is a complications developed in a person who got
medical attention against kala
-
azar but was not completely cured. After incubation period of several
months
-

several years, PKDL appears with its symptoms, darkening of general
ized skin. Sometimes
PKDL has asymptomical incubation period and in such case most of the patients do not report to a
physician. Thus, PKDL has possibility to trigger the VL outbreak in India

12

and in Sudan

13
, although the
type of PKDL between India and
Sudan has a little difference.
14



Control

As above, the principles of leishmaniasis control are to stop the cycle of
L. donovani
; in the point of
sand fly vector or reservoir host. Four basic approaches of sand fly control are below.

15

(i) Residual
spra
ying of houses and animal shelters prevented blood
-
feeding by the insects, (ii) Such treatments
denied them resting and breeding sites, (iii) The effects of spraying were sharply localized, so that
control measures need only be undertaken within a limited
area of a few hundred meters, (iv) The
relatively long life cycle of sandflies delayed recovery of populations within a treated area.

As far as reservoir host goes, considering that human being is an only reservoir, it is the most
important method that det
ects kala
-
azar and cure. As above, curing PKDL is great, too.


DDT

Indoor residual spraying of DDT was very effective against various insect vectors especially
mosquitoes.
16, 17
During 1950’s, DDT was sprayed throughout Bangladesh (then East Pakistan) in o
rder
to eradicate malaria vectors (anopheline mosquitoes). As a multiplier effect, DDT also killed sandflies.

18

Especially between 1958 and 1964 the number of sandflies were decreased all but none. Moreover,
larval habitat modification by plastering walls

of human dwellings and cattle sheds was quite effective to
P. argentipes
.
19
But repeated DDT spraying induced resistant insects, for example, malaria mosquitoes
and also
P. argentipes

in Bihar, India. Within one month of DDT spraying, resistant sandflies

were
confirmed.

20

In Muzaffarpur and Vaishali district, India, that had long time DDT spraying, lead a few
percent of
P. argentipes

to death. In the opposite direction 100% mortality was observed in case of
P.
argentipes

of Patna, which was not treated f
or a long time.

21



Insecticide
-
treated bednet (ITN)

ITN is an effective, relatively cheap and sustainable method of malarial control throughout the
World.
52

The synthetic pyrethroids used for the treatment of the nets combine the properties of low to
mod
erate mammalian toxicity, low volatility and high insecticidal activity.
6, 55 ,56

ITNs have also been
evaluated against phlebotomine sandflies and leishmaniasis in several countries including Italy, Burkina
Faso, Syria, Kenya, Colombia and Venezuela.
6

In
Afghanistan ITN was proven protective against
cutaneous leishmaniasis.
53

In Venezuela significant reduction of sandfly vector and cutaneous
leishmaniasis had occurred in pyrethroid impregnated curtain houses in compare to control groups (both
non
-
impregna
ted curtains and no curtains at all).

54



Natural repellent

Complex structure of pesticide from natural product is difficult to make resistant insects.

22

And it is
easy to environment.


Neem

Neem tree (
Azadirachta indica
)
)


is called Indian meliaceou
s. It is also a native tree in India. Therefore,
the climate of Indian subcontinent is well suited for the growth of Neem tree. Moreover,
Neem
tree
thrives well in hot weather, where the maximum shade temperature is as high as 49° C and tolerates cold
up

t
o 0° C.
54
Neem

tree

grows on almost all types of soils including clayey, saline and alkaline soils, with
pH up

to 8.5
.

Neem

tree grows naturally in areas where the rainfall is
a lot such as Bangladesh.

Neem oil has six kinds of limonoids, azadirachtin (AZA)
, salannin, deacetylgedunin, gedunin, 17
-
hydroxyazadiradione and deacetylnimbin. One of them, azadirachtin is the most effective in Neem tree
and its seed.
54

Azadirachtin is regarded as an agricultural chemical by FDA (Food and Drug

11


Administration) in USA,

Japanese Health, Labor and Welfare Ministry and APVMA (The Australian
Pesticides and Veterinary Medicines Authority) in Australia.

Azadirachtin

A
zadirachtin

(AZA)

is a tetranortriterpenoid, and is too complex to synthesize artificially.
23

AZA
has
3 effec
ts,
strong antifeedant, insect growth regulatory and reproductive effects. Its mode of action
includes (i) effects on deterrent and other chemoreceptor resulting in antifeedant, (ii) effects on
ecdysteroid and juvenile hormone titres through a blockage of
morphogenetic peptide hormone release
and (iii) direct effects on most other tissues studied resulting in an overall loss of fitness of the insect.

24


Neem oil protects from various infections including
nematodes, fungi, viruses, and protozoa
. Most
remark
ably AZA showed efficacy and sensitivity to reduce
T
rypanosoma
,
which

is also a kinotoplast
protozoa like
Leishmania

spp.


25, 26

Researchers nowadays, thinking of an insecticide which is friendly to environment for the control of
malaria mosquitoes, prefe
rring Neem oil.
27
-
29

Malaria vector (Diptera: Culicidae), different from
sandflies, do not breed in the cracks of the house wall. Burning of Neem oil in Kerosene in the living
rooms and mosquitoes resting walls is often used to control mosquitoes.

31

In la
boratory condition, it is
known that first to third larval instars are more susceptible to Neem oil.

32
Throughout the world various
Neem products (leaves, twigs, seed oil etc) have been found to be effective for protecting human from
various pathogens and

parasites where no side effectes were observed.
57


Neem oil

AZA is apt to dissolve chemically.
32
AZA and Neem extracts often show superiority to certain
commercially available pesticides.

33

Neem
-
seed extract (NSE) are used as insecticide
.

34
-
36


Extrac
tion and frequency of spraying

Extraction method from Neem seed is according with Dhar R.
et al
, 1998.

28
Although this varies
between insect species and insect’s behavior
,

the protection of Neem may persist for a number of
months.

36
10.0 microg/gm. of az
adirachtin/mg of diet inhibit larvae of
Lutzomyia longipalpis

growing.
37

10.0 microg/mg is equal to 500ppm.
2% (40ppm) Neem oil inhibits adult
Phlebotomus argentipes
.

38

5%
(100ppm) Neem oil inhibits adult
Phlebotomus papatasi.

39
Our pilot test in 2005 su
ggests that Neem oil
spraying works against
Phlebotomus argentipes

after 7 days.
40




12


Research Design and Methods


Describe in detail the methods and procedures that will be used to accomplish the objectives and specific aims of the
project. Discuss the
alternative methods that are available and justify the use of the method proposed in the study.
Justify the scientific validity of the methodological approach (biomedical, social, or environmental) as an
investigation tool to achieve the specific aims. Dis
cuss the limitations and difficulties of the proposed procedures
and sufficiently justify the use of them. Discuss the ethical issues related to biomedical and social research for
employing special procedures, such as invasive procedures in sick children,
use of isotopes or any other hazardous
materials, or social questionnaires relating to individual privacy. Point out safety procedures to be observed for
protection of individuals during any situations or materials that may be injurious to human health. Th
e methodology
section should be sufficiently descriptive to allow the reviewers to make valid and unambiguous assessment of the
project.

(DO NOT EXCEED TEN PAGES, USE CONTINUATION SHEETS).



A. Study sites



Mymensingh is the most endemic area for kala
-
azar in Bangladesh and 50% of country’s total kala
-
azar
cases are reported from there. Fulbaria and Trishal Upazila reported maximum of these cases where
more than 30 cases are reported per 10,000 people per year. But actual incidence rate is higher than
r
eported now. Trishal consist of 12 unions with an area of 339 Sq.K.M.
The study
will be conducted

at

Harirumpur, Sakhua and Trishal

union
of
Tris
h
al Upazila of
Mymensingh

District
.

They have a
combined land area of 19,321 acres and estimated population nu
mber is 96,600 (projected from 1991
census).

From, 2003 Hospital data it has been seen that Harirumpur got the highest incidence rate (59.2
per 10,000 population) for kala
-
azar followed by Sakhua (50.2 per 10,000 population) and Trishal (50.2
per 10,000 po
pulation) within Trishal Upazila.
However, if the number of person being tested
ascertained in the initial survey in th
ese

unions
is not sufficient for our study size requirements due to
difficult to get around, we will add a second choice, either Bhaluka

or Gaffargaon Upazila.
At present
ICDDR,B is conducting a hospital based passive kala
-
azar surveillance at Trishal together with IEDCR,
NIPSOM and DGHS.



B.
Study design



At the beginning of the study we will enumerate the total number of and househol
ds in the study areas.
We will consider individual household as a cluster in this study. Area boundary will be marked in the area
map by using
Geographic

Information System

(GIS). Then we will randomly select 3400 households/
clusters and single individua
l from each for baseline serosurvey. Children less than three years of age will be
excluded from this study.
After informed consent, field workers will complete a household roster and
record persons treated for VL in the household over the past 3 years an
d persons considered to have
current VL based on a simplified history. Untreated VL suspected patients
would

be referred to the
Upazila health complex for appropriate case management. The household and patient information data
will
also
be mapped by
GIS
(u
sing ArcGIS softwere).

For deaths that have occurred in the household in
the previous 3 years, a brief series of questions will be asked to classify the cause of death as (i) define
VL; (ii) probable VL; or (iii) not VL

Selected individuals
will be asked t
o provide a

few drops of finger
prick

blood and

their ur
ine at baseline
.

Rapid rk39 dipstick test will be performed using collected blood
samples in the field. Serum and urine DAT will also be performed to confirm anti
-
leishmania antibody at
Parasitology

laboratory of ICDDR,B or at the Aichi Medical University, Japan. Seropositive individuals
at the base line will not be followed up for the study purpose. After base line we will be able to recruit
2960 seronegative individuals from 2960 households whom wi
ll be assigned randomly to interventions
and control groups.

Two different interventions will be made in this study. In intervention 1, Neem oil
solution (Neem oil and soap water) will be sprayed all of the houses (740 households). Similarly in
interventio
n 2, ITN will be distributed in 740 households. Remaining 1480 households will serve as
control where no intervention will be given. Neem oil will be applied bi
-
weekly in summer months
(April to September) and monthly in other months for two years.

All
these seronegative subjects will be again tested by these blood and urine antibody detection tests in
year 2 and year 3. At the end of the study the seropositivity rates among seronegative subjects in

13


intervention groups will be compared with control group
. Baseline serological survey will be conducted
in July
-
August 2006.

Follow
-
up serological survey will be repeated in July
-
August in 2007 and 2008.



Sandflies will be collected from 200 households
; 50 households in each intervention and 100 households
for

control
. These households will be selected randomly immediately after baseline but prior to
intervention (Aug
-
Sep 2006). Then a
fter one month of interventions
and will be repeated in
Aug
-
Sep

in
2007 and 2008
.



C. Methods


C.
1. Laboratory testing for the
household survey


C.1.1. Blood test


Blood specimen will be tested by rK39 dipstick test and The
D
irect Agglutination Test (DAT). We use
rK39 dipstick kit,

InSure Rapid Test for Visceral Leishmaniasis® from InBios International, Seattle,
USA


and DAT (Th
e Direct Agglutination Test, KIT Biomedical Research, USA); both tests are
currently recommended by WHO.

We will follow standard method for rK39 dipstick test and serum
DAT.

42
-
45



C.1.2. Urine test

Collected urine samples will bring back to Dhaka. DAT w
ith urine samples will be conducted at
Parasitology Lab of ICCDR,B followed by the method developed by the co
-
investigators of this study
50

or send to Aichi Medical University.


C.2. Study case definitions, inclusion and exclusion criteria


Household memb
ers
: comprise those people who eat from the same cooking pot.


Clinical kala
-
azar (past case)
: patient diagnosed with illness characterized by ≥2 weeks of fever and at
least one of the following: splenomegaly, skin darkening, and/or weight loss, plus a history of treatment
with either stibogluconate sodium or pentamidine with clinical resolution o
f symptoms; or a patient with
Leishmania

amastigotes demonstrated in bone marrow or splenic aspirate or tissue in the past and
documented in medical records.


Clinical kala
-
azar (current case)
: patient diagnosed with illness characterized by ≥2 weeks of fe
ver and
at least one of the following: splenomegaly, skin darkening, and/or weight loss; with
Leishmania

amastigotes demonstrated in relevant aspirate or tissue, and/or positive rK39 dipstick result, and/or
positive DAT result.


Subclinical
L. donovani

inf
ection
: person without fever, skin darkening, weight loss, or splenomegaly,
but with positive results on at least one of the following tests: rK39 dipstick, DAT results. Intervention
data will be analyzed both separately and in aggregate for persons positi
ve by each individual test and
combination of tests.


Control households
: One of the half of the households made by mud wall and no object of roof material,
tin or mud. These control households will be allocated randomly.


Control individuals
: community s
urvey participants with no history of disease compatible with VL, and
with negative serology results and living in control household.


Target households
: Households made with mud wall including any kind of roof material, tin or mud.



14


Target individuals
: c
ommunity survey participants with no history of disease compatible with VL, and
with negative serology results and living in target household.


VL case households
: households with at least one current VL case (may include members with
subclinical infection
) at present or households in which VL was detected within three years.

Subclinical
L. donovani

infection case households
: households with at least one subclinical infection
(but no VL cases).


Inclusion

criteria
: All persons residing in the chosen sect
ions of the study villages who give informed
consent will be included.


Exclusion criteria:

Children younger than 3 years will be listed in the household roster but will not be
included in the serologic survey.

Similarly person treated for VL in past three

years, current VL patients
and seropositive person at the baseline will also be excluded from our study.


C
.
3
.
1.

Distribution of ITN


We will distribute second
-
generation deltamethrin impregnated bed net PermaNet
®

(origin: Vestargaurd &
Frandsen) among

all the inhabitants of an arm in the study area. This net has a mesh size of 161holes/inch
2

with long lasting impregnation in knitted polyester. We will also inform the household members how to use
this ITN in proper way.


C
.
3
.
2.

Spraying Neem oil


We
wi
ll
use the commercially
available

standardized

1500ppm Neem oil (they prepared for the organic
farming purposes). Neem oil will be dilute
d

to
5% solution
with soap powder

(
Jet
®
, P&G, Bangladesh used
as emulsifier
)
.
40

Using knack sprayer, we coat

the mud wa
ll with Neem oil dilution solution

up to
the height of
3

meter
from the floor
.

Each house will be sprayed bi
-
weekly in summer months (April to September) and
monthly in the rest of the months (October to March).


C
.
3
.
3.

Control houses


C
ontrol houses w
il
l be twice as much as each intervention group or equal to the sum of two intervention
groups. After baseline survey control houses will remain undisturbed except for scheduled serum and sandfly
collection for next two years.


C
.
4
. Entomologic
methods


Al
though sticky trap with castor oil

has been suggested for the estimate of sandfly population, the data of
capturing shows weak trend
46

Our

previous study in Bangladesh, however,
indicate
d that collected sandfly
numbers

by light trap

were well correlated wi
th the risk of leishmaniasis.
40

CDC Miniature light trap Model
-
512 will be
set

at 17:00
-
19:00, and recover 6:00
-
8:00

for sandfly collection
.

Collected sandflies will be
preserved in 70% ethanol and will be dissected under dissecting microscope after bring
back to the
Parasitology Lab of ICDDR,B. PVA mounting medium (origin: BioQuip Products, Inc) or Hoyer’s medium
will be used for temporary slide preparation. These slides will be then observed under compound microscope
for sandfly species identification by
using Key’s of Lewis.
51




C
.5. Planting Neem tree


Although this is not
a

primary objective of the study, we will assess the willingness of Neem tree planting,
harvesting and preparing Neem oil among village
rs
.
40

We
will made necessary arrangement to
pla
nt
Neem

trees and facilitate adequate training for harvesting and preparing Neem oil among villager
s
.



C
.6. Sample size calculations


From a previous community based kala
-
azar study in an endemic area of Mymensingh district we
have experienced that the cu
mulative prevalence rate for seropositivity could be 14% with a serconversion of
4%.

47

In absence of any intervention for a period two years we assume that in control area seropositive rate

15


could remain 4%. We also assume that in each intervention group s
eropositive rate could become 2%. A
sample size of 617 for each group thus be required to detect this difference with a power of 80% and α = 0.05.
Considering a drop out rate of around 20%, we estimated a sample size of 740 seronegative subjects for each

group of the study. Thus, a total of 2960 individuals will be required for this intervention study. In order to
recruit these 2960 seronegative individuals we will require 3400 individuals at baseline survey.



From our previous experience we have see
n that the density of vector
P. argentipes
would become 14
per trap. In absence of any intervention for a period of two years we assume that in control area this density
will remain same. We also assume that vector density might reduce to 7 per trap in any

of the interventions.
Therefore, 34 households will be required for each of the group with a power of 90% and α = 0.05.
Considering 20% drop out, we estimated the total number of houses required for the collection of vector
is 41
for each group but we wil
l collect sandflies from 50 houses for each arm. Thus, a total of 200 households
will be required for sandfly collection. We will select these households randomly for collection of
sandflies .



Table
-
1
: Sample size estimation for serosurvey

Indicator

Po
wer

α
=
o卓⁩渠敡捨c
gr潵o
=
q卓
1

adjusted
for group
comparison
plus 20 %
drop out

TSS
2

adjusted
for baseline
(plus 14% of
TSS
1
)

Intervention 1 (Neem oil)

80%

0.05

617

740

850

Intervention 2 (ITN)

80%

0.05

617

740

850

Control

80%

0.05

1234

1480

1700

Total




2960

3400




Table
-
2
: Sample size estimation for sandfly collection


Indicator

Power

α
=
o卓⁩渠敡捨c
gr潵o
=
q卓⁡摪畳u敤⁦潲⁧r潵瀠
捯c灡pi獯渠灬畳′〠u⁤=潰畴=
=
f湴敲v敮ei潮‱
k敥m=lF
=
㤰V
=
〮〵
=

=

=
f湴敲v敮ei潮′†ofqkF
=
㤰V
=
〮〵
=

=

=
C潮or潬
=
㤰V
=


=

=
㄰N
=
q潴慬
=
=
=
=
㈰O
=
=
=
o卓W⁒敱畩r敤⁳慭灬攠eize
=
q卓W⁔潴慬⁳=m灬攠eize
=

16


Flow chart for sampling



























3400 households

Estimated population
3400 individuals

(After randomization)




Intervention 1:

Neem oil spray

740 households

740 seronegative
individuals




Intervention 2:

Impregnated bed net
(ITN)

740 households

740 seronegative
individual
s




Control:

Blank (no intervention)

1480 households

1480 seronegative
individuals


Intervention 1:

Neem oil spraying will be
conducted.

Follow up study to
determine seroconversion
rate will be determine once
in a year for a period of two
year. Sandfly will be
collected a
fter baseline
from 50 randomly selected
households and then one
month after intervention
and again once in a year for
a period of two years


Intervention 2:

Impregnated bed net (ITN)
will be distributed.

Follow up study to
determine seroconversion
rate will be conducted once
in a year for a period of two
year. Sandfly will be
collected after baseline
from 50 randomly selected
households and then one
month afte
r intervention
and again once in a year for
a period of two years


Control:

Blank (no intervention)

Follow up study to
determine seroconversion
rate will be conducted once
in a year

for a period of two
year. Sandfly will be
collected after baseline
from100 randomly selected
households and then one
month after intervention
and again once in a year for
a period of two years


Baseline serosurvey

Randomization


17




D
. Time line for activities


Time period

Activity

Jul, 2006

Preparation for
fieldwork
,
obtain
ing
supplies
, hiring personnel.

Jul
-
Aug, 2006

Baseline serological and entomological survey;
questionnaire

and selection of
households for
ITN distribution and
Neem oil application.

Aug
-
Sep 2006

ITN distribution and
Neem oil application.

Sandfly
collection

Aug
, 2006
-
Jul, 2008

Supervised standardized Neem oil spraying (bi
-
weekly in summer months of April to
September; monthly for other months)

and monitoring ITN’s activity.

Jul
-
Aug, 2007

Serosurvey and
questionnaire
.
S
andfly collection

Jul
-
Aug
, 2008

Serosurvey and
questionnaire
. Sandfly collection

Sep,2008
-
Mar,2009

Analysis and preparation of reports and papers.

Jul, 2006
-
Mar, 2009

Neem tree planting and Neem oil application promotion



18




Fig. 1
: Map of Trishal Upazila showing study areas

19



Facilities Available

Describe the availability of physical facilities at the place where the study will be carried out. For clinical and
laboratory
-
based studies, indicate the provision of hospital and other types of patient’s care facilities and adequate

laboratory support. Point out the laboratory facilities and major equipment that will be required for the study. For
field studies, describe the field area including its size, population, and means of communications.

(TYPE WITHIN
THE PROVIDED SPACE).



We will conduct the study in t
hree
unions of Trishal Upazila

in Mymensingh District
.

If the number of kala
-
azar cases ascertained in
the selected
villages is insufficient for our study size requirements, we will add
more
unions.

Most of the villages of
these two above mentioned unions are accessible by vehicle without four
-
drive,
and are within ½ hour by vehicle from the respective Upazila Health Complex (UZHC). All are within 1 hour
by vehicle from the district capital and within
2

hours of Dhaka. UZH
C personnel have expressed
commitment to VL treatment and control, and the willingness to collaborate on the study. Patients diagnosed
with VL or other diseases requiring treatment will be referred to the
Trishal

UZHC
.



S
pecimens
, e.g.,
blood, and
urine
,

will be transported to Dhaka, during the serological survey, and ELISA will
be run in the ICDDR,B Parasitology laboratory
and/or conducted at the Department of Parasitology, Aichi
Medical University for examination
. The Parasitology Laboratory has all t
he equipment necessary to run
these tests. Study subjects needing treatment for VL will be referred to the UZHC. We will work closely with
UZHC personnel to ensure appropriate diagnosis, treatment, and clinical follow
-
up.





20


Data Analysis


Describe pla
ns for data analysis. Indicate whether data will be analyzed by the investigators themselves or by other
professionals. Specify what statistical software packages will be used and if the study is blinded, when the code will
be opened. For clinical trials,
indicate if interim data analysis will be required to monitor further progress of the
study.

(TYPE WITHIN THE PROVIDED SPACE).


Data will be analyzed by the investigators in Epi
-
Info,
R,
SPSS and SAS software packages.

The analysis will
be in three parts
, the initial community
-
wide cross
-
sectional survey, the geographic analysis, and the cohort
analysis for the follow
-
up of seropositive persons. Data will not be blinded.


W
e will use regression models incorporating Generalized Estimating Equations to ass
ess both individual and
household level risk factors, and to account for household clustering. We will also measure inter cluster
correlation (ρ).


Epidemiologic and entomologic data will be plotted to the household level to provide a detailed Geographic
Information System map of VL cases, asymptomatic infections, environmental features, and sand

fly vector
densities. Phlebotomine sandflies fly relatively short distances and it is therefore likely that proximity to
current and previous VL cases is an impo
rtant determinant of newly apparent cases of
L. donovani

infection
and VL. GIS analysis will allow us to address this hypothesis and search for relevant geographic patterns.



21


Ethical Assurance for Protection of Human Rights


Describe in the space provide
d the justifications for conducting this research in human subjects. If the study needs
observations on sick individuals, provide sufficient reasons for using them. Indicate how subject’s rights are
protected and if there is any benefit or risk to each sub
ject of the study.



In the Indian subcontinent, VL is anthroponotic. The only way to effective prevention and control methods
is to improve our understanding of the disease in humans.


So far we know currently there is no intervention programme is going
on for the control of VL or vector in
our study area. If the authority starts any kind of intervention (i.e. residual insecticidal spraying) as a part of
National Policy we will immediately suspend our intervention.


Risks and measures taken to minimize th
em:


The risks to subjects are minimal and consist primarily of the discomfort of blood sampling. Blood
sampling will cause transient pain at the site of the finger stick lancet or needle entry.
Rarely taking blood
from finger may cause infection, but we w
ill try to prevent this by using sterile lancet and cleaning the
skin before taking the blood sample

There will be the inconvenience of answering a
questionnaire
, a brief physical examination for symptomatic
persons, and baseline and follow
-
up
questionnair
es
.

But the responded would be allowed not to answer any
question if he/she does not fill comfort.


Neem oil may create bad odor few days after spraying but soon it will disappear. There is no risk of having
light trap hanging inside bedrooms.
It is not
recommended to wash ITN in open water like ponds, cannels
and rivers because of its toxic effects on the aquatic organisms. For minimize this risk we will teach the
participant how to wash this ITN in a proper way
. CDC light trap runs by normal torch light

batteries and
creates very little light, which is sufficient to attract insects. It creates no or very little sound and will not
disturb sleeping.


Benefits:



The benefits to study subjects include detection of undiagnosed VL cases with referral for
tre
atment, which

may be life
-
saving. For future VL patients, these data should allow the
implementation

of better prevention
method for VL. For the community as a whole, we hope that this study will become good strategies of VL
prevention.



Use of Animals

D
escribe in the space provided the type and species of animal that will be used in the study. Justify with reasons the
use of particular animal species in the experiment and the compliance of the animal ethical guidelines for conducting
the proposed procedu
res.



No use of animals.



22


Literature Cited


Identify all cited references to published literature in the text by number in parentheses. List all cited references
sequentially as they appear in the text. For unpublished references, provide complete info
rmation in the text and do
not include them in the list of Literature Cited. There is no page limit for this section, however exercise judgment in
assessing the “standard” length.


1.

WHO

Technical

Report Series, No 793, 1990, (
Control of the leishmaniases
: report of a WHO Expert
Committee).

2.

Thakur C.P.
and
Kumar K. 1992. Post kala
-
azar dermal leishmaniasis: a neglected aspect of kala
-
azar
control programmes.
Ann Trop Med Parasitol
, Aug;
86
(4)
:
355
-
359.

3.

Swaminath CS, Shortt HE. and Anderson LAP.

1942.

1942. Transmission of Indian kala
-
azar to man by
the bites of
Phlebotomus argentipes
, Ann.
a
nd Brun
.

Indian J Med Res
;
30
:473
-
7.

4.

Sandeep M, Mohammed QH, Anil G, Kashi NG, Amal B and Samit A., 1
997. L
eishmania
DNA
in

P
hlebotomus

and

S
ergentomyia

species during a kala
-
azar epidemic.

Am J Trop Med Hyg
,
57
(4):423
-
5.

5.

Lane RP, 1991. The contribution of sandfly control to leismaniasis control.
Ann Soc Belg Med Trop
,
71

Suppl 1:65
-
74.

6.

A
lexander B and

M
a
roli

M. 2003. Control of phlebotomine sandflies.
Med Vet Entomol.

17
(1):1
-
18.

7.

F
eliciangeli

M
.
D., 2004. Natural breeding places of phlebotomine sandflies.

Med Vet Entomol
,
18
(1):71
-
80.

8.

Modi GB and Tesh RB. 1983. A
simple

T
echnique

for

mass

R
earing

L
utzomy
ia

longipalpis

and

P
hlebotomus papatasi

(D
iptera
: P
sychodidae
)
in the laboratory
.
J Med Entomol
, Oct 5;
20
(5):568
-
9.

9.

Pandya AP
and
Niyogi AK. 1980. Ecological study on immature stages of phlebotomid sandflies in
Gujarat.
Indian J Med Res
, Sep;
72
, 355
-
8.


10.

Dh
iman RC, Shetty PS

and
Dhanda V. 1983. Breeding habitats of phlebotomine sandflies in Bihar, India.
Indian J Med Res
, Jan;
77
,29
-
32.


11.

Ghosh KN
and
Bhattacharya A. 1991. Breeding places of Phlebotomus argentipes Annandale and
Brunetti (Diptera: Psychodidae)
in West Bengal.
India Parassitologia
;

33
Suppl:267
-
72.

12.

Addy M

and

Nandy A. 1992. Ten years of kala
-
azar in West Bengal, Part I. Did post
-
kala
-
azar dermal
leishmaniasis initiate the outbreak in 24
-
Parganas?,
Bull World Health Organ
;

70

(3): 341
-
6.

13.

El Hassan

AM

and

Khalil EA. 2001. Post
-
kala
-
azar dermal leishmaniasis: does it play a role in the
transmission

of
Leishmania donovani

in the

Sudan?,
Trop Med Int Health
;

6
(9):743
-
4.

14.

Zijlstra EE, Musa AM, Khalil EA
,
el
-
Hassan IM and el
-
Hassan AM. 2003. Review Post
-
k
ala
-
azar
leishmaniasis,
Lancet Infect Dis
;

3
(2):87
-
98.

15.

Hertig M

and

Fairchild GB.1948. The control of Phlebotomus in Peru with DDT.
Ann Entomol Soc Am
;

63
(5):1460
-
1.

16.

Joshi RD
and
Rai RN. 1994.
Impact of DDT spraying on population of P argentipes and P. pap
atasi in
Varanasi district, Uttar Pradesh.
J Com Dis
;

26
(1), 56
-
8

17.

Kaul SM, Sharma RS, Dey KP, Rai RN
and

Verghese T. 1994. Impact of DDT indoor residual spraying
on Phlebotomus argentipes in a kala
-
azar endemic village in eastern Utter Pradesh.
Bull World
Health
Organ
;

72
(1):79
-
81.

18.

Pandya AP. 1983. Impact of antimalaria house spraying on phlebotomid population in Surat District,
Gujarat.
Indian J Med Res
;
78
:354
-
60.

19.

Kumar V, Kesari SK, Sinha

NK, Palit A, Ranjan A, Kishore K, Saran R

and

Kar SK. 1995.
Field t
rial of
an ecological approach for the control of
Phlebotomus argentipes

using mud & lime plaster.
Indian J Med
Res
;

101
:154
-
6.

20.

Mukhopadhyay AK, Hati AK, Chakraborty S

and

Saxena NB. 1996. Effect of DDT on Phlebotomus
sandflies in Kala
-
Azar endemic foci i
n West Bengal.
J Commun Dis,

Sep;

28
(3):171
-
5.

21.

Kishore K, Kumar V, Kesari S, Bhattacharya SK

and

Das P, 2004. Susceptibility of Phlebotomus
argentipes against DDT in endemic Districts of North Bihar,
India.

J Commun dis
;

36
(1):41
-
4.

22.

Jacobson M.

1986. Natur
al pesticides. Pages 144
-
148 in
Natural Resources: The 1986 Yearbook of
Agriculture.
U.S. Government Printing Office, Washington, D.C.

23.

Medina P, Budia F, del Estal P and Vinuela E.

2004.
Influence of azadirachtin, a botanical insecticide, on
Chrysoperla car
nea (Stephens) reproduction: toxicity and ultrastructural approach.

J Econ Entomol
;
97
(1):

43
-
50.


23


24.

Mordue AJ and Blackwell A.

1993. Azadirachtin: an Update,
J Insect Physiol,

39
(11):903
-
24

25.

de Azambuja P

and

Garcia ES., 1992. Effects of azadirachtin on
Rhod
nius prolixus
: immunity and
Trypanosoma
interaction.

Mem Inst Oswaldo Cruz
;

87

Suppl 5:69
-
72.

26.

Garcia ES, Gonzales MS

and

Azambuja P. 1991.
Effects of azadirachtin in Rhodnius prolixus: date and
hypotheses.,
Mem Inst Oswaldo Cruz
;
86

Suppl 2:107
-
11

27.

Garg S,
Talwar GP and Upadhyay SN.

1994.
Comparison of extraction procedures on the
immunocontraceptive activity of Neem seed extracts
,
J Ethnopharmacol
;

44
(2):87
-
92.

28.

Dhar R, Zhang K, Talwar GP, Garg S and Kumar N
. 1998.
Inhibition of the growth and development of

asexual and sexual stages of drug
-
sensitive and resistant strains of the human malaria parasite
Plasmodium falciparum by Neem (Azadirachta indica) fractions
,
J Ethnopharmacol
;
61
(1):31
-
9.

29.

Nathan SS
, Ka
laiva
n
i

K and
Murugan
K. 2005.
Effects of Neem limonoi
ds on the malaria vector

Anopheles stephensi

Liston (Diptera: Culicidae)
.
Acta. Trop
;

96
(1):47
-
55.

30.

Sharma VP, Ansari MA. 1994.
Personal Protection from Mosquitoes (Diptera: Culicidae) by Burning
Neem Oil in Kerosene
.,
J. Med. Entomol
., May;
31
(3):505
-
7.

31.

Sen
gottayan SN,

Kandaswamy K, Paul GC

and

Kadarkarai M. 2005. Effect of Neem limonoids on
lactate dehydrogenase (LDH) of the rice leaffolder, Cnaphalocrocis medinalis (Guenee)(Insecta:
Lepidoptera: Pyralidae)
.

Chemosphere
;
62
(8):1388
-
93.

32.

Kirsh K and Schmuttere
r H. 1988. Low efficacy of a
Baccilus thuringensis
formulation in controlling the
diamondback moth,
Plutella xylostella
(L.), in the Philippines.
J
A
ppl
E
nt

;
195
, 249
-
55.

33.

Stark JD, Vargas RI and Thalman RK. 1990. Azadirachtin: effects on metamorphosis, lo
ngevity, and
reproduction of three tephritid fruit fly species (
Diptera
: Tephritidae).
J
E
con Ent
;

83
:

2168
-
74.

34.

Stark JD, Wong TTY, Vargas RI

and Thalman RK. 1992., Survival, longevity and reproduction of
tephritid fruit fly parasitoids (Hymenoptera: Brac
onidae) reared from fruit flies exposed to azadirachtin.
J
E
con En
t
;

85
:

1125
-
9.

35.

Hough
-
Goldstein J. and Keil CB.1991. Prospects for integrated control of the Colorado potato beetle
(Coleoptera: Chrysomelidae) using
Perillus bioculatus
(Hemiptera: Pentatom
idae) and various pesticides.
J Econ Ent
;
.
84
, 1645
-
51.

36.

Makanjuola

WA.

1989
. Evaluation of extracts of Neem (
Azadirachta indica

A. Juss) for the control of
some stored prodct pests.
J. Stored Prod. Re
s
;
.

25
, 231
-
8.

37.

Andrade Coelho CA, de Souza NA, Feder MD,

da Silva CE, Garcia Ede S, Azambuja P, Gonzalez MS
and Rangel EF.

2006
.

Effects of azadirachtin on the development and mortality of
Lutzomyia longipalpis

larvae (Diptera: Psychodidae: Phlebotominae)
,
J Med Entomol
;
43
(2):262
-
6.

38.

Sharma VP

and

Dhiman RC
.

1
993. Neem oil as a sand fly (Diptera: Psychodidae) repellent,
J Am Mosq
Control Assoc
;

9
(3):364
-
6.

39.

Srivansan R

and

Kalyanasundaram M. 2001. Relative efficacy of DEPA and Neem oil for repwllent
activity against Phlebotomus papatasi, the vector of leishmania
sis,
J Commun Dis
, Sep;
33
(3):180
-
4.

40.


W
agatsuma Y
, D
har I
,
Alam MS
,
Khanum H,

Washed M.A. and
Haque R.

2006. Neem oil as biological
control against Phlebotomine sandfly,
Tropical Medicine and Health
;
34
(1):53.

41.

Kurkjian KM, Vaz LE, Haque R, Cetre
-
Sossah C, A
khter S, Roy S, Steurer F, Amann J, Ali M,
Chowdhury R, Wagatsuman Y, Williamson J, Crawford S, Breiman RF, Maguire JH, Bern C, Secor WE,
2005. Application of an improved method for the recombinant k 39 enzyme
-
linked immunosorbent assay
to detect visceral
leishmaniasis disease and infection in Bangladesh,
Clin Diagn Lab Immunol
;
12
(12):1410
-
5.

42.

Sarker CB, Momen A, Jamal MF, Siddiqui NI, Siddiqui FM, Chowdhury KS, Rahman S

and

Talukder SI.
2003. Immunochromatographic (rK39) strip test in the diagnosis of visc
era leishmaniasis in Bangladesh,
Mymensingh Med J
.
12
(2):9307.

43.

Bern C
, Shambhu NJ, Anand BJ, Thakur GD and Mahendra BB., 2000. U
se of recombinant
K39

dipstick
test and the direct agglutination test inn a setting endemic for

visceral leishmaniasis in Nepal.


Am J Trop
Med Hyg
;

63
(3,4):153
-
7.

44.

Chappuls F, Rijal S, Singh R, Acharya P, Karki BMS, Bovier PA, Desjeux P, Le Ray D, Koirala S

and

Loutan L
.

2003. Prospective evaluation and comparison of the direct agglutination test and an rK39
-
antigen
-
based dipstick
test for the diagnosis of suspected kala
-
azar in Nepal,
Trop Med Inter Health
;

8
(3):277
-
85.


24


45.

Guernaoui S, Boussaa S, Pesson B, Boumezzough A.
, 2006.
Nocturnal activity of phlebotomine sandflies
(Diptera: Psychodidae) in a cutaneous leishmaniasis focus in Ch
ichaoua, Morocco
,
Parasitol Res
;
98
(3):184
-
8.

46.

Koirala S, Parija SC, Karki P

and

Das ML
.

1998.
Knowledge, attitudes, and practices about kala
-
azar and
its sandfly vector in rural communities of Nepal, Bull World Health Organ
;

76
(5):485
-
90.

47.

Bern C, Hightowe
r AW, Chowdhury R, Ali M, Amann J, Wagatsuma Y, Haque R, Kurkjian K, Vaz LE,
Begum M, Akter T, Cetre
-
Sossah CB, Ahluwalia IB, Dotson E, Secor WE, Breiman RF and Maguire JM,
2005. Risk Factors for Kala
-
Azar in Bangladesh.
Emerg Infect Dis
;
11
(5):655
-
62.

48.

Ber
n C, Joshi AB, Jha SN, Das ML, Hightower A, Thakur GD, Bista MB., 2000. Factors associated with
visceral leishmaniasis in Nepal: bed
-
net use is strongly protective, Am J Trop Med Hyg ;
63
(3
-
4):184
-
8.

49.

Islam MZ, Itoh M, Shamsuzzaman SM, Mirza R, Matin F, Ahme
d I, Choudhury AKMS, Hossain MA,
Qiu XG, Begam N, Furuya M, Leafasia JL, Hashiguchi Y, Kimura E, 2002. Diagnosis of Visceral
Leishmaniasis by ELISA Using Urine Samples
.
Clin Diagn Lab Immunol
;

9
:

789
-
794.

50.

Islam MZ, Itoh M, Mirza R, Ahmed I, Ekram ARMS, S
arder AH, Shamsuzzaman SM, Hashiguchi Y,
Kimura E, 2004. Direct agglutination test with urine samples for the diagnosis of visceral leishmaniasis
.
Am J Trop Med Hyg
;
70
:

78
-
82.

51.

Lewis DJ. 1978. The Phlebotomine sandflies (Diptera: Psychodidae) of the Orien
tal Region.
Bulletin
of the British Museum (Natural History)
;
37:
217
-
343.

52.

Rayburn H, Ashford R, Mohsen M, Hewitt S and Rowland M. 2000. A randomized controlled trial
of insecticide
-
treated bednets and
chaddar

or top sheets and residual spraying of interio
r rooms for
the prevention of cutaneous leishmaniasis in Kabul, Afghanistan.
Trans R Soc Trop Med Hyg
;
94
:
361
-
66.

53.

Koreger A, Avila EV and Morison L. 2002. Insecticide impregnated curtains to control domestic
transmission of cutaneous leishmanisis in Vene
zuela: cluster randomized trial.
BMJ
;
325
: 810
-
13.

54.

Schmutterer S. 1990. Properties and potential of natural pesticides from the neem tree,
Azadirachta
indica
.
Annu Rev Entomol
;
35
: 271
-
97.

55.

Curtis CF. 1991. Control of disease vectors in the community. Lond
on: Wolfe

56.

Kishore K, Kumar V, Kesari S, Dinesh DS, Kumar AJ, Das P and Bhattacharya. 2006. Vector
control of leishmaniasis.
Indian J Med Res
;
123
: 467
-
472.


57.

Bostid FRR (ed.). 1992. Neem: a tree for solving global problems: A report of an ad hoc panel of th
e
board of Science and technology for international development national research council. National
academy press, Washington, D. C. USA.





25



Dissemination and Use of Findings


Describe explicitly the plans for disseminating the accomplished results. D
escribe what type of
publication is anticipated: working papers, internal (institutional) publication, international
publications, international conferences and agencies, workshops etc. Mention if the project is
linked to the Government of Bangladesh throu
gh a training programme.



We anticipate that this research will result in several international publications. If appropriate, this work will
facilitate to launch a VL prevention
campaign

in Bangladesh.



Collaborative Arrangements


Describe briefly if
this study involves any scientific, administrative, fiscal, or programmatic arrangements with other
national or international organizations or individuals. Indicate the nature and extent of collaboration and include a
letter of agreement between the applic
ant or his/her organization and the collaborating organization.

(DO NOT
EXCEED ONE PAGE)



This study is
collaboration

among the ICDDR,B and Japanese
universities

(University of Tsukuba and
Aichi Medical University)
.

Financial support will be from the National Science Fund of Japan allocated in
these two universities. These funding will promote further career development of young Bangladesh scientists
through PhD studies in Japan.




26


Biography of the Investigators


Give biographical data in the following table for key personnel including the Principal Investigator. Use a
photocopy of this page for each investigator.



(Note: Biography of the external Investigators may, however, be submitted in the format as
convenie
nt to them)



1. Name: Mohammad Shafiul Alam (PI)


2. Present Position: Field Research Officer, Clinical Sciences Division/Laboratory Sciences


Division, ICDDR,B


3. Educational background (last degree an
d diploma & training relevant to the present research
proposal)

MS in Zoology (Parasitology), University of Dhaka, Bangladesh, 2002

B. Sc in Zoology, University of Dhaka, Bangladesh, 2001




4. List of ongoing research protocols

(start and end

dates; and percentage of time)



4.1 As Principal Investigator


Protocol Number

Starting date

End date

Percentage of
time




















As Co
-
Investigator





Protocol Number

Starting date

Ending date

Percentage of
time

















27




5. Publications



Types of publications

Numbers

a) Original scientific papers in peer
-
review journals

5

b) Peer reviewed articles and book chapters

0

c) Papers in conference proceedings

3

d) Letters, editorials, annotations, and abstracts in peer
-
reviewed
journals

0

e) Working papers

0

f) Monographs

0




6. Five recent publications including publications relevant to the pre
sent research protocol:


Khanum H, Sultana R,
Alam MS
, Zaman RF. 2002. Endoparasitic helminth infection in
Hemidactylus flaviviridis


(Ruppel, 1835).
Univ. J. Zool. Rajshahi Univ.

21:17
-
19

Alam MS
, Khanum H, Nessa Z. 2002. Rat pinworm
Syphacia muris

Yamag
uti, 1941 from
laboratory rat in Bangladesh.
Univ. J. Zool. Rajshahi Univ.

21: 95
-
96.

Alam MS
, Khanum , Nessa Z. 2003. Helminth infection in laboratory rat strain, long
-
evans
(
Rattus norvegicus

Barkenhout, 1769).
Bangladesh J. Zool
. 31(2): 221
-
25.

Alam MS
,

Khanum H. 2005. Infection of
Ascaris lumbricoides
and
Trichuris trichura
among the children of two slum areas in Dhaka city. Accepted for published at
Bangladesh
J. Zool
. 33(1): 89
-
94.

Alam MS
, Khanum H. 2005. Educational status of mother as a factor f
or roundworm
infection in children.
Dhaka Univ J Biol Sci
14(2): 199
-
201.



28



Biography of the Investigators


Give biographical data in the following table for key personnel including the Principal Investigator. Use a
photocopy of this page for each inves
tigator.



1. Name: Dr. Rashidul Haque (Co
-
PI)


2. Present Position: Scientist & Head, Parasitology Laboratory, Laboratory Sciences


Division, ICDDR,B


3. Educational background (last degree and dipl
oma & training relevant to the present research
proposal)

M.B
-

Sofia Medical Academy, Sofia, Bulgaria, 1985

Ph.D
-

Institute of Parasitology, Bulgarian Academy of Sciences, Sofia,


Bulgaria, 1988

Certificate Course on Laboratory Diag
nosis of Parasitic Diseases, London School of Hygiene of
Tropical Medicine, London, 1991.



4. List of ongoing research protocols

(start and end dates; and percentage of time)



4.1 As Principal Investigator


Protocol Number

Starting date

E
nd date

Percentage of
time

2003
-
010


01/07/03

31/12/07

40%

2003
-
022


01/10/03

30/03/08

15%

2003
-
021


02/07/03

01/07/08

10%

2004
-
021

01/09/04

01/08/09


5%



As Co
-
Investigator





Protocol Number

Starting date

Ending date

Percentage of
time


















29



5. Publications



Types of publications

Numbers

a) Original scientific papers in peer
-
review journals

47

b) Peer reviewed articles and book chapters


3

c) Papers in conference proceedings

18

d) Letters, editorials, annotations, and abstracts in peer
-
reviewed
journals

5

e) Working papers

0

f) Monographs

0




6. Five recent publications including publications re
levant to the present research protocol:


Haque R
, Duggal P, Ali IM, Hossain MB, Mondal D, Sack RB, Farr BF, Beaty TH, Petri WA
Jr. (2002). Innate and Acquired Resistance to Amebiasis in Bangladeshi Children.
J Infect Dis

186:547
-
552


Haque R,

Huston C
D, Hughes M, Houpt E and Petri WA Jr. (2003) Current Concepts:
Amebiasis.
New England Journal of Medicine
. 348: 1565
-
1573.


Haque R
, Mondal D, Kirkpatrick BD, Akther S, Farr BM, Sack RB, Petri WA Jr. (2003).
Epidemiological and clinical characteristics of
acute diarrhea with emphasis on
Entamoeba
histolytica

infections in preschool children in an urban slum of Dhaka, Bangladesh.
Am J Trop
Med Hyg

69: 398
-
405.


Haque R
, Mondal D, Duggal P, Kabir M, Roy S, Farr BM, Sack RB, Petri WA Jr.( 2006).
Entamoeba

histolytica

infection in children and protection from subsequent amebiasis.
Infect
Immun
74: 904
-
909.


Threimer K,
Haque R
, Wagatsuma Y, Salam MA, Akther S, Attlmaayer B , Fukuda M,
Schaecher K, Miller RS, Nodel H. (2006) Therapeutic efficacy of Qunine
plus sulfadoxine
-
pyremethamine for the treatment of uncomplicated malaria in Bangladesh.

Am J Trop Med Hyg

( submitted).






30



Biography of the Investigators


Give biographical data in the following table for key personnel including the Principal Inves
tigator. Use a
photocopy of this page for each investigator.


1 Name



: Kazi Mohammad Asif Jamil
(Co
-
PI)


2 Present position


: Associate Scientist


3 Educational background

:

1988:

M.B.B.S. (Bachelor of Medicine and Surgery)

Institute


Chitt
agong Medical College, Chittagong, Bangladesh


1995:

Ph.D. in Medical Sciences



Institute


University of Tokyo, Japan

2001:

Postdoctoral training in Nutrition at the Department of Nutrition of University
of California Davis, USA


4 List of ongoing re
search protocols



4.1.


As Principal Investigator


Protocol Number

Starting date

End date

Percentage of time


2002
-
032


1
-
1
-
2003

31
-
12
-
2005

20






4.2.

As Co
-
Principal Investigator


Protocol Number

Starting date

End date

Percentage of time










4.3.


As Co
-
Investigator





Protocol Number

Starting date

Ending date

Percentage of time

2002
-
009

1
-
9
-
2003

31
-
12
-
2005

5

2003
-
009

1
-
3
-
2004

31
-
12
-
2005

10


5 Publications



Types of publications

Numbers

a) Original scientific papers in peer
-
review j
ournals

6

b) Peer reviewed articles and book chapters


c) Papers in conference proceedings


d) Letters, editorials, annotations, and abstracts in peer
-
re
viewed
journals

5








31



6

Recent publications including publications relevant to the present research protocol:


(i)

Daily consumption of Indian Spinach (
Basella alba
) or sweet potatoes has a positive
impact on total body vitamin A stores of Bangladeshi men.

Marjorie J. Haskell,
Kazi M.
Jamil
, Ferdaus Hassan , Janet M. Peerson, M. Iqbal Hossain, George J. Fuchs, and
Kenneth H. Brown.
Am J Clin Nutr
. 2004 Sep;80 (3):705
-
14.

(ii)


Human milk as a source of ascorbic acid: no enhancing effect on iron bioavailability
f
rom a traditional complementary food consumed by Bangladeshi infants and young
children. Lena Davidsson,
Kazi Asif Jamil
, Shafiqual Alam Sarker, Christophe Zeder,
George Fuchs, Richard Hurrell.
Am J Clin Nutr.

2004 Jun;79(6):1073
-
7.

(iii)

Detection of endotoxin
in sera from children hospitalized for treatment of diarrhea in
Bangladesh. Ahmed T, Azam MA, Armed N,
Jamil KM
, Hassan F, Ogura N, Tamura H,
Yokochi T.
J Endotoxin Res
. 2004;10(4):223
-
8.

(iv)

Low temperature hemodialysis prevents hypotensive episodes by reduci
ng nitric oxide
synthesis. Nephron.
KM Jamil
, K Yokoyama, F Takemoto, S Hara, and A Yamada. 2000
Mar; 84(3): 284
-
6.

(v)

Distinct mechanisms of action of V1 antagonists OPC
-
21268 and
[d(CH
2
)
5
Tyr(Me)AVP] in mesangial cells.
KM Jamil
, T Watanabe, A Nakao, T Okuda

and K Kurokawa. Biochem. Biophys. Res. Commun. 1993; 193: 738
-
743.

(vi)


Expression of platelet activating factor receptor in renal tubular cell line (LLC
-
PK1).
KM Jamil
, T Takano, A Nakao, Z Honda, T Shimizu, T Watanabe and K Kurokawa.
Biochem. Biophys. Res.C
ommun. 1992; 187: 767
-
772.






























32


Biography of the Investigators


Give biographical data in the following table for key personnel including the Principal Investigator. Use a
photocopy of this page for each investigator.



1 Name



:

Yukiko Wagatsuma

(Co
-
investigator)


2 Present position

:

Professor, Graduate School of Comprehensive
Human

Sciences, University of Tsukuba, Tsukuba,
Japan


3 Educational background

:




(last degree and diploma & training relevant to

the present research proposal)

M.D., University of Tsukuba, Japan, 1987

D.T.M., Tropical Medicine, University of Tokyo, Japan, 1987

M.P.H., International Health, Johns Hopkins University, USA, 1990

Dr. P.H., International Health, Johns Hopkins University,

1996


4.
List of ongoing research protocols (start and end dates; and percentage of time)


#2002
-
031:
Combined
i
nterventions to
p
romote
m
aternal and
i
nfant
h
ealth:
e
ffects
o
ver a

p
regnancy
c
ycle and on
c
hildren 0
-
24
m
onths

#2003
-
021: Antimalarial drug

resistance in Bangladesh


4.4.


As Principal Investigator

Protocol Number

Starting date

End date

Percentage
of time










4.5.

As Co
-
Principal Investigator

Protocol Number

Starting date

End date

Percentage
of time










4.6.


As Co
-
Investigator


Protocol

Number

Starting date

Ending date

Percentage
of time

#2002
-
031

01/10/2002

01/12/2006

10%

#2003
-
021

01/07/2003

30/06/2008

5%




33


5 Publications



Types of publications

Numbers

a)
Original scientific papers in peer
-
review journals


21

b)
Peer reviewed articles and book chapters

4

c)

Papers in conference proceedings

15

d)
Letters, editorials, annotations, and abstracts in peer
-
reviewed
jo
urnals

2

e)

Working papers

6

f)

Monographs

1



6 Five recent publications including publications relevant to the present research protocol


i.

Anoopa Sharma D, Bern C, Varghese B, Chowdhury R, Haque R, Ali M, Amann J,

Ahluwalia
IB, Wagatsuma Y, Breiman RF,
Maguire JH, McFarland DA. The economic impact of visceral
leishmaniasis on households in Bangladesh.

Trop Med Int Health. 2006;11(5):757
-
64.


ii.

Bern C, Hightower AW, Chowdhury R, Ali M, Amann J,
Wagatsuma Y
, Haque R, Kurkjian K,
Vaz LE, Begum M, Akter T,
Cetre
-
Sossah CB, Ahluwalia IB, Dotson E, Secor WE, Breiman
RF, Maguire JH. Risk Factors for Kala
-
Azar in Bangladesh.
Emerging Infectious Diseases

1
1
(
5
):
655
-
662
;200
5
.


iii.

Kurkjian KM, Vaz LE, Haque R, Cetre
-
Sossah C, Akhter S, Roy S, Steurer F,

Amann J, Ali M,

Chowdhury R, Wagatsuma Y, Williamson J, Crawford S, Breiman RF,

Maguire JH, Bern C,
Secor WE. Application of an improved method for the recombinant k 39 enzyme
-
linked

immunosorbent assay to detect visceral leishmaniasis disease and infection in

Banglades
h.

iv.

Clin Diagn Lab Immunol. 2005 Dec;12(12):1410
-
5.


v.

Ahluwalia IB, Bern C, Wagatsuma Y , Costa C, Chowdhury R, Ali M, Amann J, Haque R,
Breiman RF, Maguire JH. Visceral leishmaniasis: Consequences to women in a Bangladesh
community.
Journal of Women’s Healt
h

13(4):360
-
364;2004
.


vi.

Wagatsuma Y, Breiman RF , Hossain A, Rahman M. Dengue Fever Outbreak in a Recreation
Club in Dhaka, Bangladesh.
Emerging Infectious Diseases

10(4):747
-
750;2004.


-----------------------------------------------------------------------
-------------------------------------



34



Biography of the Investigators


Give biographical data in the following table for key personnel including the Principal Investigator. Use a
photocopy of this page for each invest
igator.


1 Name



:

Makoto Itoh
(Co
-
investigator)

2 Present position

:

Associate Professor, Aichi Medical University


School of Medicine, Nagakute, Aichi
-
ken, Japan

3

Educational background:

(last degree and diploma &

training relevant to the present research proposal)

M.Sc., Nagoya University, Japan, 1977

Ph.D., Magoya City University, Japan, 1990


4
List of ongoing research protocols
at ICDDR,B
(start and end dates; and percentage of time)



4.7.


As Principal Inv
estigator

Protocol Number

Starting date

End date

Percentage
of time














4.8.

As Co
-
Principal Investigator

Protocol Number

Starting date

End date

Percentage
of time














4.9.


As Co
-
Investigator


Protocol Number

Starting date

Ending date

P
ercentage
of time














5 Publications



Types of publications

Numbers

a)
Original scientific papers in peer
-
review journals

38

b)
Peer reviewed articles and book chapters


2

c)

Papers in conference proceedings

0

d)
Letters, editorials, annotations, and abstracts in peer
-
reviewed
journals

1

g)

Working papers

3

h)

Monographs

0


6

Five recent publications including publications rel
evant to the present research protocol


i.

Islam M. Z., Itoh M., Mirza R., Matin F., Ahmed I., Ekram A.R.M.S., Sarder A.H.,
Shamsuzzaman S.M., Hashiguchi Y. & Kimura E. Direct agglutination test (DAT) with urine
samples for the diagnosis of visceral leishman
iasis. Am J Trop Med Hyg, 70: 78
-
82, 2004
.



35


ii.

Weerasooriya M.V., Itoh M., Islam M.Z., Qiu X
-
G., Fujimaki Y. & Kimura E. Prevalence and
levels of
filarial
-
specific urinary IgG4 among children less than five years of age and the
association of antibody positiv
ity between children and their mothers. Am J Trop Med Hyg, 68:
465
-
468, 2003.


iii.

Itoh M., Ohta N., Kanazawa T., Nakajima Y., Sho M., Minai M., Daren Z., Yan C., Hongbin H.,
Yong
-
Kang H. & Zhinan Z. Sensitive enzyme
-
linked immunosorbent assay with urine samp
les:
a tool for surveillance of schistosomiasis japonica. Southeast Asian J. Trop. Med. Public Health,
34: 469
-
472, 2003


iv.

Watanabe K., Itoh M., Matsuyama H., Hamano S., Kobayashi S., Shirakawa T., Suzuki A.,
Sharma S., Acharya G.P., Itoh K., Kawasaki T., K
imura E. & Aoki Y. Bancroftian filariasis in
Nepal: A survey for circulating antigenemia of
Wuchereria bancrofti

and urinary IgG4 antibody
in two rural areas of Nepal. Acta Tropica, 88: 11
-
15, 2003.


v.

Islam M. Z., Itoh M. Shamsuzzaman S.M. , Mirza R., Matin

F., Ahmed I. , Choudhury A.K.M.S.,
Hossain M.A.,
Qiu X
-
G
., Begam N., Furuya M., Leafasia J.L., Hashiguchi Y. & Kimura E.
Diagnosis of visceral leishmaniasis by enzyme
-
linked immunosorbent assay using urine samples.
Clin Diagn Lab Immunol , 9: 789
-
794, 2
002.



































36


Biography of the Investigators


Give biographical data in the following table for key personnel including the Principal Investigator. Use a
photocopy of this page for each investigator.


1 Name



:

Mohammad Zahid
ul Islam

(Co
-
investigator)

2 Present position


:

Assistant Professor

3 Educational background

:

(last degree and diploma & training relevant to the present research proposal)

JSPS Postdoctoral Fellow, Aichi Medical University, Aichi, Japan, 2004
-
2006
.

Ph.D., Medicine, Aichi Medical University, Aichi, Japan, 2004

Research Student, Dept. of Parasitology, Aichi Medical University, Aichi, 1998
-
2000.

Inter Doctor, Rajshahi Medical College Hospital, Rajshahi, Bangladesh, 1998

M.B.B.S., Rajshahi Medical Coll
ege, Rajshahi, Bangladesh, 1997


List of ongoing research protocols
at ICDDR,B
(start and end dates; and percentage of time)



4.10.


As Principal Investigator

Protocol Number

Starting date

End date

Percentage
of time














4.11.

As Co
-
Principal Inve
stigator

Protocol Number

Starting date

End date

Percentage
of time














4.12.


As Co
-
Investigator


Protocol Number

Starting date

Ending date

Percentage
of time











5 Publications



Types of publications

Numbers

a)
Original scientifi
c papers in peer
-
review journals

4

b)
Peer reviewed articles and book chapters

0

c)

Papers in conference proceedings

1

d)
Letters, editorials, annotation
s, and abstracts in peer
-
reviewed
journals


i)

Working papers

1

j)

Monographs








37


7

Five recent publications including publications relevant to the present research protocol



Shintoku Y, Kimura E, Kadosaka T, Kondo S, Itoh M, and
Islam MZ. 2005
. Strongyloides ratti infection in
the large intestine of wild rats, rattus norvegicus.

J. Parasitol.

:

1116
-
1121



Islam MZ
, Itoh M, Ekram ARMS, Islam MAU, Hossain MS, Ali MM, Takagi H, Hashiguchi Y, and Kimura
E.
2004.

Detection of
Leishmania(Leishmania) d
onovani
antigen in urine by antigen capture enzyme
-
linked immunosorbent assay.
Research Report Series

(Studies on New and Old World Leishmaniases and
their Transmission, with Particular Referance to Ecuador, Argentina and Pakistan. Edited by Prof. Yoshihis
ha
Hashiguchi) No.
7:

78
-
84



Islam MZ
, Itoh M, Mirza R, Ahmed I, Ekram ARMS, Sarder AH, Shamsuzzaman SM, Hashiguchi Y, and
Kimura E.
2004, January.

Direct agglutination test with urine samples for the diagnosis of visceral
leishmaniasis.
Am. J. Trop. Med. H
yg.

70(1):

78
-
82



Weerasooriya MV, Itoh M,
Islam MZ,
Qiu X
-
G, and Kimura E.
2003. Prevalence and intensity of filaria
-
specific urinary IgG4 among children under 5 years old, and the association of the antibody positivity
between the children and their mothe
rs.
Am. J. Trop. Med. Hyg.
68 (4):

465
-
468



Islam MZ
, Itoh M, Shamsuzzaman SM, Mirza R, Matin F, Ahmed I, Choudhury AKMS, Hossain MA, Qiu
X
-
G, Begam N, Furuya M, Leafasia JL, Hashiguchi Y, and Kimura E.
2002, July
.
Diagnosis of visceral
leishmaniasis by ELI
SA using urine Samples.
Clin. Diagn. Lab. Immunol.
9 (4):
789
-
794






































38





39



Budget Justifications


Please provide one page statement justifying the budgeted amount for each major item. Justify u
se of man power,
major equipment, and laboratory services.


Man power
-

Three field attendants will be hired for 26 months when regular spraying of Neem oil will
be done. They
will take part Neem oil spraying all the year round and take part in ITN distrib
ution. They
will also help our appointed field research assistant during July and August when sero
-
survey and
sandfly collection will be conducted simultaneously. Field research assistant will fill up consent forms
and study questionnaire, and will collect

blood through finger pricks. Two external intern students will
also take part during this period every year in blood collection and rK39 dipstick test. Principal
investigator of this study will regularly monitor their activities through intensive field
visit. He will
identify the collected sandflies in the Parasitology lab and if possible he will also take part during
laboratory analysis of urine samples.



Field supplies
-

These includes CDC miniature light traps, batteries for light trap, cryo vials for

sandfly
preservation, rK39 dipstick test kits, Neem oil etc will be supplied by Aichi Medical University and
Tsukuba University, Japan at free of cost.


Lab supplies


The

supplies are those needed for collection, processing, and storage of clinical
spe
cimens. Required lab supplies for this study will be provided free of cost by Aichi Medical
University and Tsukuba University, Japan.


Lab analyses



rK39 dipstick test for kala
-
azar detection will be done in the field settings. Urine samples
will be pres
erved and bring back to the Parasitology Lab, ICDDR,B for storage. DAT with the urine and
blood samples may be performed at the Parasitology Lab of ICDDR,B or will be performed at the Aichi
Medical University, Japan.




Other Support


Describe sources, a
mount, duration, and grant number of all other research funding currently granted to PI or under
consideration. (DO NOT EXCEED ONE PAGE FOR EACH INVESTIGATOR)







40


APPENDIX

International Centre for Diarrhoeal Disease Research, Bangladesh

Voluntary Consen
t Form


Protocol Number:

2006
-
27

Title of the Research Project:

Use of Neem oil and insecticide
-
treated bednet (ITN) to control
visceral
leishmaniasis (
VL)

in an endemic area of
Bangladesh

Principal Investigator:
Mohammad Shafiul Alam

Organization:
Inter
national Centre for Diarrhoeal Disease Research, Bangladesh


Before recruiting into the study, the study subject must be informed about the objectives, procedures,
and potential benefits and risks involved in the study. Details of all procedures must be p
rovided
including their risks, utility, duration, frequencies, and severity. All questions of the subject must be
answered to his/ her satisfaction, indicating that the participation is purely voluntary. For children,
consents must be obtained from their p
arents or legal guardians. The subject must indicate his/ her
acceptance of participation by signing or thumb printing on this form.


[If you do not understand any part of this informed consent please ask any one of the investigators of the
study regarding

it. If you agree to participate in this proposed study, a copy of this form will be provided
to you.]

Introduction


Visceral leishmaniasis or kala
-
azar is caused by a protozoan parasite,
Leishmania donovani
.
This illness
causes fevers, swelling in the be
lly, weight loss and weakness. People can die from kala
-
azar if they do
not receive treatment.
Kala
-
azar is transmitted by a sandfly species,
Phlebotomus argentipes
in
Bangladesh. Every year more than 7000 people are officially reported to have kala
-
azar a
mong them
more than 50% are from Mymensingh district. Children are the most vulnerable to this disease.
The
International Centre for Diarrhoeal Disease Research, Bangladesh is conducting a research to learn more
about the control of kala
-
azar in Banglades
h. We are
inviting
you/your child

to help us by participating
in this study

because there are people with kala
-
azar in your village.



Purpose


We are conducting a research in your village to see the impact of Neem oil and insecticide
-
treated bednet
(ITN)

on reducing kala
-
azar incidence and vector density in your area. This will help us to find a cost
effective and environment friendly method to control kala
-
azar in Bangladesh.


What we will ask you to do?


If your house is selected for this study we will

ask you some questions about you and your household
members, their health and past illness. You do not have to answer any questions if you do not want to. If
anyone in your house is ill and we think it is kala
-
azar, we will

immediately refer

him/her to th
e nearest
government

hospital to get proper diagnosis and treatment. The drug to treat kala
-
azar is given for free of cost
at Government hospitals. Kala
-
azar has long incubation period (2 months to more than a year), so sometimes it
may happen that you got

the parasite inside your blood but sign and symptoms do not develop or develop later.
For this

reason you or your child will be asked to provide few drops of blood (300μl only) from fingers
and also to give little volume of urine in our supplied tube if your/your child’s name is selected by
randomization. We will perform a rapid test for kala
-
azar
diagnosis by using rK39 dipstick test with this
blood sample and inform you its result within few days. From collected blood and urine sample we will
also perform DAT (direct agglutination test) for kala
-
azar detection. We will have these results within
on
e month and will inform you then. We will work in your village for more than two years. In this
duration you/your child will be asked to give finger’s blood and urine specimen for a total of 3 times. If
we found positive reports form our tests at any time
of our study we will refer you/your child to the
nearest Upazila Health Complex where treatment of kala
-
azar is provided free of cost. In case they could
not provide drug for you/your child we will provide them from our project.



41


We will randomly select f
ew houses for spraying Neem oil. If your house is selected we will spray Neem
oil in the floor of your cattle shed (if any) and inside your bed room’s and cattle shed’s mud wall (
up to
the height of 3

meter
from the floor
)
.
Neem
oil

contains
a chemical com
ponent name “
azadirachtin


which
is very effective to kill different kind of insects including kala
-
azar vector
P. argentipes
. If vector of
kala
-
azar is controlled the kala
-
azar incidence rate will automatically decline. We prepare this neem oil
solution w
ith soap powder (white detergent powder Jet®) and water because Neem oil does not mix with
mud wall unless we add soap water in it. This solution contains 5% “
azadirachtin
”.


We might request you to use
Insecticide
-
treated bed (ITN) if your house is selec
ted randomly, which will
be distributed by us.
This net is a second
-
generation deltamethrin impregnated bed net PermaNet
®

(origin:
Vestargaurd & Frandsen) with a mesh size of 161holes/inch
2

and has long lasting impregnation in knitted
polyester. If you are

selected for ITN then you will be asked to use it during your sleeping time.


Sandflies will be collected from some randomly selected houses. For this regard
CDC Miniature light trap
Model
-
512

will

be
used. This trap will be set

at 17:00
-
19:00

hrs inside
sleeping room
and
will be
recover
ed at

0
6:00
-
0
8:00

hrs on following day
.

It may also happen that your house has not been selected for Neem oil
spraying or ITN use but still selected for blood, urine and sandfly collection.



Risk and benefits

You/your chil
d may get a small bruise where the lancet goes in but this goes away soon. Rarely taking
blood from finger may cause infection, but we will try to prevent this by using sterile lancet and cleaning
the skin before taking the blood sample. You may smell odor

of Neem oil few days after we spray in
your house but soon it will disappear. However, Neem oil spraying inside your bed room and cattle shed
will not create any harm to your, your child or pets health. It is not recommended to wash ITN in open
water like

ponds, cannels and rivers because of its toxicity to the aquatic organisms. To minimize this
risk we will teach you how to wash this ITN in a proper way. But ITN will not affect your health. There
is no risk of having light trap hanging inside your house.

This trap is run by normal torch light batteries
and creates very little light, which is sufficient to attract insects.

This study will help you to get diagnosed in case you/your child have kala
-
azar. You and your child will
receive proper treatment free

of cost if we find you/your child are infected with kala
-
azar but Government
hospital could not provide it.


If any health problem occurs as a result of your/your child’s participation in the study, we will arrange
for the best treatment available in Ban
gladesh at our own costs.


Alternatives


Your doctor will decide the treatment if you/your child will have kala
-
azar. Bangladesh has national
guidelines for treatment of kala
-
azar. The study will not change your treatment.


Confidentiality

We do hereby af
firm that confidentiality of data identifying you and your child will strictly be
maintained. We would keep all medical information and results of the laboratory tests performed on you
and your child confidential, under lock and key. None other than the in
vestigators of this research;
possible study monitor, and any law
-
enforcing agency in the event of necessity would have an access to
the information. The samples related to the study may be sent outside the country for analysis; however,
any personal ident
ifiable information will be held and processed under secured conditions, with access
to limited appropriate staff of that organization.


We would be obliged to provide you/your child’s information related to the medical condition(s) of
you/your child’s, tr
eatment, or results of any or all tests performed on you/your child’s, and would be
happy to answer your questions about the study. You would be able to freely communicate with any
investigator of this study (contact address is given below). We would, howe
ver, like to inform you that
some tests would be performed in batches, at the end of the study, and thus their results would be
available only then.


42



Right not to participate and withdraw

Your/your child’s participation in the study is voluntary, and you a
re the sole authority to decide for and
against you/your child’s participation in this study. You would also be able to withdraw your
participation any time during the study. Refusal to take part in or withdrawal from the study will involve
no penalty or l
oss of benefits or attention.


Storage of blood and urine samples


If you agree, we will store the remaining blood and urine sample after the study is over. We may use the
sample to work with other tests for kala
-
azar. We may also look in your/your child’s

blood sample for
other infections, or for blood factors that make people more likely to get sick from kala
-
azar. After our
study ends, we will take your/your child’s name off the sample when we store it so we will not be able
to report the results of fut
ure tests to you. We will store the samples for about 20 years. If you are
willing to have us to test your/your child’s samples now, but you do not want us to store the samples,
you can still take part in the study.


For further information

If you have a
ny question regarding the study and its procedures, you may ask those to our study staff
now or in future. You will also be able to contact the principal investigator or other investigators of this
study at the address given below for asking questions. If
you want to know more about your rights as a
participant in a research study, you may contact with Mr. B. R. Saha, personally at the Training and
Education Department of ICDDR,B, Mohakhali, Dhaka 1212, or over telephone at PABX: 8860523
through 8860532, Ex
tension No. 3508.


Participant’s/parent’s/guardians Declaration
: The investigators/study staff has provided me all
information about the background, the purpose, and the methods of the study and their alternatives, and
risks and benefits. Based on this in
formation, I am voluntarily agreeing for my/my child’s participation
in the study. I understand that I will receive a copy of this signed consent form.



Name of the participant: ______________________________________________


Name of the parent/legal guar
dian of the child: ___________________________





______________________





___/___/___



Signature / Left thumbprint






Date

of the participant/ guardian












_____________________





___/___/___

Signature of the Investigator






D
ate





_____________

______






___/___/___

Witness’s signature







Date


43



Contact address of the Investigators:

Mohammad Shafiul Alam; Principal Investigator, Laboratory Sciences Division

ICDDR,B: Centre for Health and Population Research, Mohakha
li, Dhaka 1212

Telephone: PABX: 8860523 through 8860532, Extension 2411.



Dr. Rashidul Haque; Co
-
Investigator, Laboratory Sciences Division

ICDDR,B: Centre for Health and Population Research, Mohakhali, Dhaka 1212

Telephone: PABX: 8860523 through 8860532
, Extension 2411.


Dr. Kazi Mohammad Asif Jamil: Co
-
Investigator, Clinical Sciences Division

ICDDR,B: Centre for Health and Population Research, Mohakhali, Dhaka 1212

Telephone: PABX: 8860523 through 8860532, Extension 2333.



44



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Avš
R©vwZK D`ivgq M‡
elYv †K›`ª, evsjv‡`k


Hw”QK m¤§wZ cÎ


M‡elYv cÖK‡íi b¤^i:
2006
-
27

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evsjv‡`‡ki GKwU KvjvR¡icÖeb GjvKvq KvjvR¡i wbqš
•Y

cÖavb M‡elKt
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R©vwZK D`ivgq M‡elYv †K›`ª,

evsjv‡`k


M‡elYvq Aš
f©w³i c‡e© cÖZ¨K e¨w³‡K M‡elYvi D‡Ïk¨, c×wZ, Ges m¤¢ve¨ jvf I SuywK m¤^‡Ü
Rvbv‡Z n‡e| we¯
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mKj cÖ‡kœi DËi AskMÖnbKvix Zvi wb‡Ri gwR© Abyhvqx w`‡eb, †Kbbv A
skMÖnb m¤•
Y© Hw”QK|
wkï‡`i †¶‡Î m¤§wZ w`‡eb Zv‡`i wcZv
-
gvZv A_ev AvBbMZ AwffveK| M‡elYvq AskMÖYKvix GB di‡g ¯^v¶i
A_ev e„×v½yjwi Qvc w`‡q m¤§wZ cÖKvk Ki‡eb|



[Avcwb hw` †Kvb kã wKsev A_© bv ey‡S _v‡Kb Z‡e AbyMÖn K‡i Avgv‡`i Kg©x‡K wR‡Ám Ki“b| Avcwb
hw` m
¤§wZ cÖKvk K‡ib Z‡e Avgiv Avcbv‡K GB †¯^”Qv m¤§wZ c‡Îi GKwU Abywjwc †`e|
]


f‚wgKv

wfwmi¨vj wjmwgwbqvwmm ev KvjvR¡i cÖ‡Uv‡Rvqvce© f
•³
wjmwgwbqv †Wvbfvwb

(
Leishmania donovani
)
bvgK ciRxexi gva¨‡g Qovq|
GB †iv‡Mi d‡j R¡i, †cU dz‡j hvIqv, IRb n«vm Ges `
•e©jZv
cÖf„wZ DcmM©
†`Lv hvq| wPwKrmv MÖnY bv Ki‡j Avµvš
 e¨w³ gvivI †h‡Z cv‡ib| evsjv‡`‡k KvjvR¡i GKwU we‡kl
cÖRvwZi †e‡jgvwQ
wd¬‡ev‡Uvgvm Avi‡Rw›U‡cm

(
Phlebotomus argentipes
)
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• nb Ges Gi g
‡a¨ kZKiv 50 fvM gvbyl evm K‡ib
gqgbwmsn †Rjvq| wkïiv G †iv‡M ‡ekx Avµvš
• nq|

Avš
•R©vwZK

D`ivgq M‡elYv †K›`ª, evsjv‡`‡k KvjvR¡i
wbqš
Y m¤^‡Ü Rvb‡Z GB M‡elYv Kvh©µg cwiPvjbv Ki‡Q| Avgiv Avcbv‡K Ges Avcbvi mšvb‡K
M‡elYvq AskMÖnY Ki‡Z ejwQ †Kbbv Avcbv‡`i MÖ
v‡g KvjvR¡i Avµvš
 e¨w³ i‡q‡Qb|


D‡Ïk¨

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e¨envi K‡i KvjvR¡‡ii cÖ‡Kvc Ges Gi evnK †e‡j gvwQi msL¨v
wbqš
•Y

Kiv hvq wKbv Zv †`Le| hv fwel¨‡Z
evsjv‡`‡k KvjvR¡i I Zvi evn‡Ki
wb

‡Y

GKwU mnRjf¨ Ges cwi‡ek evÜe gva¨g wba©viY Ki‡Z mnqZv
Ki‡e|


Avgiv Avcbv‡K wK Ki‡Z ej‡ev ?


GB M‡elYvq hw` Avcbvi evwo wbe©vwPZ nq Z‡e Avgiv Avcbvi I Avcbvi evwoi m`m¨‡`i ¯^v¯’¨ I
c
e©eZ©x Amy¯’Zv m¤^‡Ü K‡qKwU cÖkœ Kie| Avcwb bv PvB‡j †h †Kvb cÖ‡kœi D
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vb‡K mywPwKrmvi Rb¨ wbKU¯’ Dc‡Rjv

45


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46


webv evavq GB M‡elYv msw¯­ó ‡h †Kvb M‡elK‡`i mv‡_ †hvMv‡hvM Ki‡Z cv‡ib (‡hvMv‡hv‡Mi wVKvbv
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Avcbvi/Avcbvi

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bgybv ¸‡jv 20 eQ‡ii Rb¨ msi¶Y Kie| Avcwb hw` Avgv‡`i‡K Avcbvi bgybv cix¶v Kiv‡Z ivwR nb wKš‘ bgybv
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1212 †Uwj‡dvb bs
8860523
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AskMÖn‡bi Rb¨

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AskMÖnbKvixi bvg:
______________________________________________


AskMÖnbKvixi AwffveK/AvBbvbyM Awffve‡Ki bvg:
___________________________



______________________





___/___/___




AskMÖnbKvix/Awffve‡Ki


¯^v¶i/e„×v½ywji Qvc









____________
_________





___/___/___



M‡el‡Ki ¯^v¶i








ZvwiL



_____________________





___/___/___




¯^v¶xi ¯^v¶i








ZvwiL




ZvwiL


47


M‡elK‡`i mv‡_ †hvMv‡hvM Kievi wVKvbv:


†gvnv¤§` kwdDj Avjg; cÖavb M‡elK, GjGmwW

AvBwmwWwWAvi,we: †m›Uvi d
i †nj_ GÛ ccy‡jkb wimvP©, gnvLvjx, XvKv
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1212

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32, G·‡Ubkb: 2411


Wvt ivwk`yj nK; mn
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M‡elK, GjGmwW

AvBwmwWwWAvi,we: †m›Uvi di †nj_ GÛ ccy‡jkb wimvP©, gnvLvjx, XvKv
-
1212

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32, G·‡Ubkb: 2411


Wv: KvRx †gvnv
¤§` Avwmd Rvwgj, mnKvix M‡elK wmGmwW

AvBwmwWwWAvi,we: †m›Uvi di †nj_ GÛ ccy‡jkb wimvP©, gnvLvjx, XvKv
-
1212

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32, G·‡Ubkb: 2333




48







Check List



After completing the protocol, please check tha
t the following selected items have been included.


1.

Face Sheet Included



2.

Approval of the Division Director on Face Sheet



3.

Certification and Signature of PI on Face Sheet, #9 and #10



4.

Table on Contents



5. Project S
ummary



6.

Literature Cited



7. Biography of Investigators



8.

Ethical Assurance



9.

Consent Forms



10.
Detailed Budget

APPENDIX
-
3


49



To :







Date:



From :




Subject :