180813 final-1 revision content of proposal form 2a 2012-1_rt

7/30/2019 180813 Final-1 Revision Content of Proposal Form 2a 2012-1_rt

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FORM 2A

TITLE

Effect The effectiveness of vitamin D supplementation in preventing

latent tuberculosis infection among healthy children five years or less

that are contacts of tuberculosis cases :

a double blind randomized- control trial

1. ABSTRACTMany study have reported clearly, that vitamin D activity as immunomodulator

affect by blood level either genetic polymorphism Vitamin D Receptor (VDR), but the overall

effect to TB treatment still controversial. There was no report about vitamin D

supplementation to prevent tuberculosis in healthy child contact TB. This study aim are to

know about effect of vitamin D supplementation to prevent developing latent tuberculosis in

children under five years with TB close contact. In this study, we will enrolle the healthy

children (without infection, without disease) and divided into two groups, group of vitamin D

supplementation and placebo, twice, day 0 and day 42. We use vitamin D3, with high dosages

25.000 IU each time. We will evaluate about the vitamin D blood level at baseline and after

12 weeks. We will check genotype VDR too, to asses the influence on vitamin D respons.

We use tuberculin skin test to define latent tuberculosis, and the positive results will confirme

with interferon (IFN) assay. Therefore we analyze the effect with various statistical

analysis.

2. EXECUTIVE SUMMARYThere was no doubt that tuberculosis (TB) in children have been transmitted from

adult TB, especially smear positive and close contact. Risk of infection and disease increase

by age, therefore can suffered more severe TB disease, furthermore affect quality of life.

Althought, National Tuberculosis Program (NTP) program gives Isoniazid Prophylaxis

Therapy (IPT) to every children under five with TB contact, but actually in field generally

CONTENT OF PROPOSAL


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difficult with low compliance, and in fact can drive MDR TB. So we need the other action to

prevent TB in children TB contact.

However, the immune system greatly affected the immunopathogenesis to M. tb,

influenced by various micronutrients, specially concern is vitamin D. Several clinical studies

found a strong association between vitamin D and TB. The study mainly focused on the

association of vitamin D deficiency with incidence of TB and improvement of adult TB cases.

Nursyam in Jakarta, found improvement of sputum conversion of in group addition with

vitamin D. Recently study carried out by Martineau, patient who were receiving standard TB

treatment and had been supplemented with vitamin D, found sputum clereance more rapid

than placebo. However, Wejse et al. found no effect of supplementation on disease outcome.

In the other hand, some study showed role of genotype Vitamin D Receptor (VDR) on

macrophage activity.

There were lack of data about prevention of TB with vitamin D supplementation,

especially in children. Study to adult TB contact were given a single dose of 100,000 IU (2,5

mg) vitamin D, demonstrated enhanced immunity to mycobacteria, therefore prevent latent

TB. However, there are many evidence that vitamin D supplementation have beneficial effect

on immune system in general, particularly in childhood. Some study demonstrated effect of

vitamin D supplementation on reduce influenza and pneumonia in children.

This study aim to find out the effect of vitamin D supplementation to healthy children

TB contact to prevent latent TB. Our hypothesize is there are different about incidence of laten

TB between group of vitamin D supplementation and placebo in healthy child TB contact. In

this study, we will enrolle the healthy children (without infection, without disease) and dividedinto two groups, group of vitamin D supplementation and placebo, twice day 0 and day 42.

We use vitamin D3, with high dosages 25.000 IU each time. We will evaluate about the

vitamin D blood level at baseline and after 12 weeks. We will check genotype VDR too, to

asses the influence on vitamin D respons. Tuberculin skin test was used to define latent

tuberculosis, and the positive results will confirme with IFN assay. The research assistants,

research nurses, and participants were unaware of the assignment groups. Therefore we

analyze the effect with different statistical analyzed according to the data.

3. INTRODUCTIONTB Global Report 2011 reported 22 high burden countries (HBCs), that contribute to

80% of TB cases worldwide every year, Indonesia at the fourth rank. More specific, each year

there were 450.000 new TB cases and 64.000 die.1 Indonesia 2010 data showed there were


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296,272 total TB cases, and 28,312 child TB cases or 9.6%,marked variation betweenprovinces about 1.5% to 17% .2

It has been clear that tuberculosis (TB) in children have been transmitted from adult

TB, especially smear positive and close contact. According to Shaw (1954), adult smear-

positive TB is infectious to 62.5% of the children around him. Even if smear negative but

culture positive M.tuberculosis 26.8% infected, if smear and culture negative 17.6%.

Infection and disease increase by age, therefore can become longterm effect according to

dorman characteristic of M tuberculosis. In case the child has been suffer severe TB disease

form, furthermore can affected quality of life.3 Althought, NTP program gives Isoniazid

Prophylaxis Therapy (IPT) to every children under five with TB contact,4 in fact in field

generally difficult with low compliance,5-7 and actually can drive MDR TB.8 Its important to

find out the other way to prevent TB in children contact TB.

The immune system greatly affected human body to againts to M. tuberculosis.

Celluler immunity have major role (innate and adaptive) and influenced by various

micronutrients, specially is vitamin D.9Some studies demonstrated that vitamin D regulates

the expression of specific endogenous antimicrobial peptides on macrophages, stimulate the

production of cytokine, etc, especially to M. Tuberculosis.10,11 The other studies have reported

that vitamin D receptor (VDR) genetic polymorphisms also contribute to the work of vitamin

D as an immune system modulator. 12,13 Both of this condition, can happen event in tropical

country. In Indonesia, which is actually an equatorial regions with high solar flow, should the

conversion of vitamin D may occur properly, but there were 3 study showed vitamin D

deficiency in children.14-16

Several risk factors were responsible, such as lifestyle changes,many time spent indoor game, as well as malnutrition.17Overall, in worldwide prevalence of

vitamin D deficiency about 30-50%.18

Many clinical studies demonstrated association between vitamin D and TB. Most

of study has been done to adult TB. Nursyam in Jakarta, found improvement of sputum

conversion of in group addition with vitamin D.19Recently study carried out by Martineau,

patient who were receiving standard TB treatment and had been supplemented with vitamin

D, found sputum clereance more rapid than placebo.20 However, Wejse et al found no effect

of supplementation on disease outcome.21There were limited study in children. One study in

Bandung found that serum vitamin D level is low in children with tuberculosis .14 The other

study reported about genotype FokI VDR have role for risk of TB disease, various by race and

place.22-24

There were lack of data about prevention of TB with vitamin D supplementation,

especially in children. Study to adult TB contact were given a single dose of 2,5 mg vitamin

D, demonstrated enhanced immunity to mycobacteria, therefore prevent latent TB.25 In


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Mongolian school-age children, vitamin D supplementation for 6 month, trend was seen

toward fewer TST conversions in the vitamin D group.26 However, there are many evidence

that vitamin D supplementation have beneficial effect on immune system in general,

particularly in childhood. Some study demonstrated effect of vitamin D supplementation on

reduce influenza and pneumonia in children.27,28

According to furthermore needed to find out the other way to prevent latent TB in child

TB contact, and limited data about vitamin D for prevention TB in this population, we want to

know about the effect vitamin D supplementation to child TB contact under five years age,

addition to standart IPT therapy, to prevent latent TB. We design a double blind randomized

control trial to healthy children TB contact. This is the first clinical trial study in children,

about this topic, especially in Indonesia.

Research Questions

Is Can vitamin D supplementation can reduce incidence ofprevent latent TB infection

among healthy children aged five years or less that are contacts of in healthy child TB contact

under five years ageTB cases ?

Hypothesis

Child contacts age five years or less that received vitamin D supplementation will be

less likely to develop latent TB infection compare to those received placebo.

There are any different about incidence of latent TB between group of vitamin D

supplementation and placebo in healthy child TB contact under five years age.

Objectives

General Main Objectives

To evaluate the effectiveness overall effect of vitamin D supplementation in healthy

child TB contacts aged five years or less under 5 years age to reduce the incidencein

preventingoflatent tuberculosis infection.

Specific Objective

Primary objective

To determine the effect of vitamin D supplementation to tuberculin skin test conversion

in healthy child TB contact under five years age.

Secondary Objective

To determine serum levels of vitamin D at baseline and after 12 weeks.

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To determine risk factors for vitamin D deficiency To determine characteristic of VDR genetic polymorphism in this population To evaluate specific risk factor to develop latent TB

To evaluate cost effective twice high dose vitamin D supplementation To evaluate performance of tuberculin skin test and IFN in diagnosing latent TBExpected Outcome

Primary Outcome

Proportion of latent TB in healthy child TB contact after vitamin D supplementation.

Secondary outcome

Prevalence and risk factor of vitamin D insufficiency and deficiency in healthy child TBcontact under 5 years age

Characteristic of VDR genetic polymorphism in this population Risk factor for developing latent TB in this population Cost effective twice high dose vitamin D supplementation Agreement of tuberculin skin test and IFN in diagnosing latent TB

Novelty

This is the first clinical trial study about the effect of vitamin D supplementation in children

child TB contacts in Indonesia. , will do to healthy child (without latent TB, without TB

disease) TB contact under five age, confirme to latent TB with both TST and IFN . We use

twice high dose vitamin D 25.000 IU , may be safety and effective, based on properly dosages

from research in adult. Differ from previous study, we use specific population, define at

children with contact to adult TB with positive AFB. Our study design is double blind

randomized control trial.

Implication of the research

Forresearcher, it will informed the effect of vitamin D supplementation to prevent thelatent TB infection in children under 5 years age who close contact to adult TB and find

out several risk factor.

For institution, it will informed the prevention effort to latent tuberculosis in childrendue to vitamin D. This study become one of implementation of Tri Dharma University

(education, research and community service) and improve the scientific publication.

Comment [DR1]: Menurut saya ini lebih baikdihapus

Comment [DR2]: Ini juga sebaiknya dihapus


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Forpolicy, it will informed the addition strategy for prevention management of TB latentin child TB contact

Forcommunity, it will improved the knowledge about TB dissemination from adult TBto children who close contact and the role of immune system by getting vitamin D

supplementation.

Forscientific development, encourage another study about vitamin D, including VDRpolymorphism to prevent latent TB in child TB contact.

4. SUMMARY OF LITERATURE REVIEW

Magnitude of problems

TB Global Report 2011 reported 22 high burden countries (HBCs), that contribute to

80% of TB cases worldwide every year, Indonesia at the fourth rank. More specific, each year

there were 450.000 new TB cases and 64.000 died.1 Indonesia 2010 data showed there

were 296,272 total TB cases, and 28,312 child TB cases or 9.6%,marked variation betweenprovinces about 1.5% to 17%.2 In West Sumatera 2012, there were child TB disease for 3 last

year, about 4,3-7,4%.29 In 2 years, number of TB cases in Pediatric Department M Djamil

Hospital are 93, most commonly isolated pulmonary TB.30

Importance of prevent latent TB in children

It is clear on TB that M. tuberculosis transmitted from adult TB cases especiallysmear positive, to children around them. Close contact of smear positive sources cause an

increased risk of TB infection and TB disease in children, especially age under five, either

developed severe TB cases too. Natural history data show that the vast majority (>95%) of

children who progress to disease, do so within 6-12 months of primary infection .3 A

systematic review found prevalence TB infection is high among child contacts, with

proportion about 24-69%. 31 Other data showed TB infection among them are in West

Sumatera 69,9% 32and Yogyakarta 35%33

Early intervention in child TB contact may reduce the burden of cavitating disease and

associated disease transmission in the community in future adult life. Exposed children

should be screened and tuberculosis status should be determine. Children aged less than 5

years who do not have active disease should receive Isoniazid Preventive Therapy (IPT) to

reduce the likelihood of developing tuberculosis disease. The current recommendation for

preventive therapy is isoniazid 10 mg/kg daily for 6 months.4Meta-analysis of IPT in HIV-

uninfected individuals with latent TB infection (LTBI) showed that IPT is effective in

preventing the development of tuberculosis disease by 60% overall.34 In children, the benefit


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is estimated to be greater at around an 80% reduction.34-36

Despite these benefits, contact investigation and management are rarely carried out in

resource-poor countries, in which tuberculosis is usually endemic. In fact, there was infrequent

contact scrining, IPT often not available and poor adherence.7 Two studies from Malawi

reported an uptake of less than 10%.37 Similarly, a more recent study in India reported that

among patients with young child contacts, only a quarter had been informed to screen the

close contacts for tuberculosis and of the availability of IPT to the children. Of 365 children

under five, only 33 (9%) were actually screened for tuberculosis38. Moreover, newest

publication showed that by mathematics modelling, community-wide IPT drives drug-resistant

TB.8

The poor implementation of contact investigation and management in developing

countries is likely to relate to the issues regarding health systems/infrastructure and facilities

as well as human resources capacities, limited resources for Tuberculin Skin Test, low public

awereness, and it is not easy to convince families of the need for investigation and prolonged

treatment in well children. So, its important to find out the other way or the addition way to

prevent TB in children contact TB.

Vitamin D as an immunomodulator

The immune system greatly affected the event of latent TB in child TB contact.

Latent TB will not occur if the natural immunity (innate immunity) works well, play by

various immune cells including macrophage, dendritic cells, followed by adaptive immunity,

including IFN, TNF, and other Th2 cytokine. Both the immune system is influenced by

various micronutrients. One micronutrient that is growing role in natural immunity during the

last 3 decades, is vitamin D.39

Vitamin D is a steroid-like protein, that synthesized in the skin during to ultraviolet

light and is also available in the diet, principally from oily fish. It is readily metabolized in the

liver to form 25-hydroxy-vitamin D [25(OH)D], the accepted measure of vitamin D status.

This form is the further metabolized by the 1-hydroxylase enzyme Cyp27B1 in renal to its

biologically active metabolite, the steroid hormone 1,25-hydroxy-vitamin D.40

Epidemiologically, approximately 1 million people in the world is estimated to

have vitamin D insufficiency or deficienc, about 30-50%.

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. Lack of sunlight (during thewinter), vegetarian diet, skin with dark pigmentation, increased pollution, and wear long

clothes that cover the body, are risk factors that are often found. Habit of living in modern

times, such as the use of computers, video games, play station, the old television program,

causing children to spend longer time in the house.40 As a result, exposure to sunlight is

reduced, the effect on vitamin D levels in the blood. In fact, event in tropical country like


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Indonesia have vitamin D deficiency according the results above. Newest survey by

SEANUTS 2012, prevalence vitamin D deficiency in Indonesian children about 39%-45,4%,

various between city and village area.16

Vitamin D is suspected having effect as potential modulators according to several

mechanism. There is significant biological plausibility for a clinical association between low

[25(OH)D] levels and susceptibility to infection, regarding to direct effect of [1,25(OH)2D]

on innate immunity.11Many immune cells express the VDR, including T and B cells, dendritic

cells, as well as macrophage. Initial studies found that vitamin D stimulates antimicrobial

activity, throughout resulting in induction of cathelicidin and/or defensin, both of which are

potent antimicrobial peptide.41 Autophagy is the other mechanism to kill mycobacteria,

throught the lysosomal machinery, one of the effect of vitamin D to innate immunity.42,43In

the state of vitamin D deficiency, infected macrophages can not produce [1,25(OH) 2D] is

sufficient in the production of cathelicidin settings.41

Its appear that downstream adaptive immunity can also be modified by [1,25 (OH)2D].

There were some evidence that vitamin D suppressed IFN production in respons to M.

tuberculosis antigens.11(Effect vitamin D IFN) Other in vitro study showed that coculture of T

cells with [1,25 (OH)2D] reduces the number of Th1 cytokine expressing cells (specifically

IFN and TNF ).41 However, the effect of vitamin D on Th2 cytokine production remain

unclear.

In addition, the vitamin D receptor (VDR) polymorphism play a role insusceptibility to TB. The association between TB incidence and other polymorphisms varies

widely across different ethnic groups. For example, the FokI ff genotype of the VDR appears

to be most consistently associated with increased susceptibily to TB among Asians, but not

Africans. A study of patients with TB in Peru, found that genetic polymorphisms vitamin D

receptor (VDR) is closely related to sputum conversion with auramine staining and culture

M.tb.44 Fok1 VDR genetic polymorphisms, together with IL-1B and Toll-like receptor 2,

increasing the incidence of extra pulmonary TB.24Study in children at Bandung, FokI gene

polymorphism have more risk to TB disease 2,94 times than control group.22

Evidence based association vitamin D and TB

The role of vitamin D in TB was first proposed by Peter Davies, and several clinical

studies found a strong association between vitamin D with TB, likelihood more suffer TB

disease. Talat study in Pakistan found the role of vitamin D to progression of TB .45Ho-Pham

in Vietnam report on vitamin D insufficiency is a risk factor for TB incidence in men .46Study


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cross sectional and case control 42 contact TB, found that a sufficient level was a protector

against TST conversion, and the mean serum level [25(OH)D] in TST conversion cases was

lower than control.47To know about influence of seasonal variation in vitamin D status and

TB in Cape Town South Africa, Martineu has done study measure vitamin D level, and found

reprocical seasonal variation in serum [25(OH)D] concentration and TB incidence.48A meta-

analysis, concluded that low levels of vitamin D is associated with risk of progression to

active tuberculosis.49

Clinical evidence using vitamin D in TB treatment was reported in several studies. An

RCT study in Guinea-Bissau, did not show any improvement in clinical outcomes of TB

patients treatment21, but Nursyam in Indonesia, getting the opposite.19 In an Egyptian study

vitamin D was administred to children with tuberculosis, showing that clinical improvement

was more evident in patients taking vitamin D as compare to treatment alone.50 The study

conclude that vitamin D supplementation may be very effective in addition to antituberculosis

drugs in the treatment of TB children.51 Supplementation with high doses of vitamin D

accelerated clinical, radiographic improvement in all TB patients and increased host immune

activation in patient with baseline deficient serum levels. These result suggest a therapeutic

role for vitamin D in treatment of TB.47

Vitamin D has been suggested as prophylaxis in TB household contacs, being a low-

cost intervention that also easy to administer in resource-poor settings .52A single oral dose of

vitamin D enhanced tuberculosis contacts antimycobacterial immunity in vitro. The finding

that a single oral dose of 2,5 mg vitamin D corrects profound vitamin D deficiency for at least6 weeks, without causing hypercalcemia, underlines the potential use of this formulation as

safe, effective, and low cost public health intervention.25. In Mongolian school-age children,

vitamin D supplementation for 6 month, trend was seen toward fewer TST conversions in the

vitamin D group.27

Rationalization to give safety and effective vitamin D supplementation in children

In connection with the extensive benefits of vitamin D on health, the AAP now

recommends vitamin D every day at a dose of 400 IU starting from the first few days after

birth, and passed childhood and adolescence, including the exclusively breastfed babies.

53

Vitamin D is well tolerated when administered in accordance with the correct dose. However,

when given orally in excessive amounts, can increase blood calcium levels, which in the long

run can be a kidney stone.54

A meta-analysis indicates that vitamin D3 (cholecalciferol) is more effective in

raising serum [25(OH)D] concentration than is vitamin D2 (ergocalciferol), so that vitamin D3

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is preferred in supplementation.55Some studies used daily dose and the other are single dose.

In adult, by using 1600 IU daily vitamin D3 compare with 50.000 IU once a month give the

same effect.56Supplementation vitamin D 800 IU/day in Mongolian school-age children for 6

month was significant in maintain serum [25(OH)D] consentration.26

Admistration of cholecalciferol once a day and stop at the week 12 more stabille than

ergocalsiferol degradation after 6 weeks. A single bolus of 50.000 IU of cholecalsiferol has

the same peak concentration and not return to baseline at the end of the 28-d after intervention.

37,55 Side effects are high levels of calcium in the blood (hypercalcaemia), high levels of

calcium in the urine (hypercalciuria) and allergic (hypersensitivity) reactions. The signs of

overdose include loss of appetite, feeling sick, being sick, headache, and feeling tired, drowsy

or weak. Reported toxicity of vitamin D showing hypercalsemia at daily intake of > 1000 mcg

(40.000 IU)57.

Knowledge gap

According to furthermore needed to find out the other way to prevent latent TB in child TB

contact, and limited data about vitamin D for prevention TB in this population, we want to

know about the effect vitamin D supplementation to child TB contact under five years age,

addition to standart IPT therapy, to prevent latent TB, with use 25.000 IU vitamin D3, twice

in day 0 and day 42.

Theoritical Framework









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http://www.webmd.boots.com/migraines-headaches/default.htmhttp://www.webmd.boots.com/sleep-disorders/default.htmhttp://www.webmd.boots.com/sleep-disorders/default.htmhttp://www.webmd.boots.com/migraines-headaches/default.htm


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Research Framework

HostVDR polymorphism

Vitamin D level

Malnutrition

Disease Character

Case Index

Child < 5 years old

Positive smearcontact (+)TB uninfected

External FactorsSunlight exposureLifestyleEating Pattern

Vitamin D

Incidence of

latent TB


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5. RESEARCH DESIGN AND METHOD

Study Design

The study will be conducted as a double blind randomized control triali. s is the

experimental study, and overall research will be design as a double blind randomized-control

trial.

Close contactInclusion :

< 5 years old

< 20 km from

hospital

Informed consent

VDR polymorphism

Vitamin D level

Healthy child

(TB exposed)

Latent TB

TB Disease

IP

T

After 12 weeks

(day 84)

Group 2 :

no intervention

Group 1 :

vitamin D supplementation

LTBINo LTBI

TST, IGRA levels

Vitamin D levels

No LTBI LTBI

Day 0

Day 42

Adults with pulmonary TB

Exclusion :TB treatmentSevere wasted

Severe infections

Eligible subjects

Questionnaire, physical

examination, TST, CXR

IP

T



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Study site

This study will take place in Padang city, one of the city in West Sumatera province,

Indonesia. Case notification rate of smear-positive in 20110 in Padang city was 31.4 per 100

000 population. In 2008, approximately 600 new cases of PTB, both smear-positive and

smear-negative, were commenced on TB treatment at this site.

We will recruite the subjectsThe index cases will be identified in several Primary

Health Cares (PHCs), and Pulmonary clinic at M Djamil hospital, the teaching hospital of

Faculty of Medicine, Andalas University, Padang. All laboratory studies will be performed in

Biomedic Laboratory Andalas University.

Time

The overall study will be conducted for 12 month with the cohort will be recruited over

the first 6 month of this study.

Population

The study population are children aged


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negativeat initial assessment

o Live not more than 20 km from the site of recruitmento Written informed consent from the parents or guardian

Exclusion criteria:

o Have been treated or are on TB treatmento Severe wastedo Have severe infections such as measles, typhoid, HIV, etc, in the last 3 months

Randomization

We will do simple randomization using a table of random numbers to pick the

treatment group.

Sample Size

The sample size is calculated based on the below formula. The proportion of latent TB

infection among TB contacts who receive vitamin D supplementation is 11%, whereas

among those withoutrvitamin D is 27% .

With assumption that 60% of 600 new PTB cases have at least one child living in the

same house and 80% of them are eligible for the study, and the different of proportion of

latent TB are 11% and 27% from previous study26, we calculate sample using formula for

difference of2 proporsion :

Z = 1.,96Z = 0.,842P1 = 0.,11P2 = 0.27P = 0,190.4 (0.30.5)Q = 0,810.6

n 1=n2=( )

=

=

= 93.2

n 1=n 2=( )


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Minimal sample in each group is 94

Variables

Exposure :o vitamin D supplementation and without

Outcome : latent TB infection and no latent TB infection ( see cases definition below) Potensial confounder and covariate :

o Demographic data : gender, habits, family size, duration of exposure withsunlight, sosioeconomic

o TB contact history : duration of exposure, level of positive smearo Nutritional statuso VDR genetic polymorphism

Instruments and data collection

1. Questionnaire. : For screening based on symptoms, by researcher2. Physical examination: by researcher3. IFN and genetic polymorphisms of vitamin D: by researcher in collaboration with the

laboratory staff and consultants Microbiology and Biomolecular.

Tools and materials

Tool

We will prepare kits for biomedical activity, like : Laminar Air Flow, inverted

microscope, PCR, Electrophoresis, Gel docking, Spectrophotometer, water bath, Micro

centrifuge, CO2 Incubators, analytical balance, autoclave, centrifuge tubes, tips, PCR tubes,

tube micro centrifuge, micropipette, syringe injection

Material

We prepare the material like : pure link genomic Isolation kit, master mix, ethanol, vitamin D

ELISA kits, RPMI, FBS, pen strep, Bicol Hypaque, Fungizone, specific primers, Milli Qwater, vitamin D, endonuclease restriction enzyme TaqI, FokI and BsmI, Agarose,

Cybergreen, PHA, rESAT-6, rIFN.

Operational Definitions


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a. Source case

Source case is a new case (all ages) of PTB identified during the study period.

New case is a patient who has sputum smear positif tuberculosis, never had treatmentfor tuberculosis and has taken anti-tuberculosis drugs for less than one month.

Pulmonary tuberculosis is tuberculosis disease which involves lungs and theparenchyma.

Positive acid fast bacilli (AFB) is the presence of AFB in microscopy smearaccording to the WHO criteria.

Sputum Smear Positive tuberculosis will be defined as at least 2 positive initialsputum smears, or 1 positive initial sputum smear with radiographic abnormalities

consistent with active PTB as determined by the treating doctor, or 1 positive sputum

smear with a positive sputum culture.

Sputum Negative Smear PTB will be defined as a patient with pulmonarytuberculosis not meeting the above criteria for smear-positive disease. Diagnostic

criteria should include: at least two sputum smear examinations negative for AFB; and

radiographic abnormalities consistent with active pulmonary tuberculosis; and no

response to a course of broad-spectrum antibiotics (except in a patient for whom there

is laboratory confirmation or strong clinical evidence of HIV infection); and a

decision by a clinician to treat with a full course of anti-tuberculosis chemotherapy; or

positive culture but negative AFB sputum examinations.

b. For child contacts

Child TB contact : Certain tuberculosis: the presence of at least one of the following symptoms- cough,

objective weight loss, unexplained fatigue or lethargy or prolonged fever-, AND at

least one of bacteriological confirmation, radiology confirmation or certain extra

pulmonary tuberculosis.

Probable tuberculosis: any child not meeting strict criteria as defined under certaintuberculosis, in whom the treating clinician started tuberculosis treatment AND a

clear treatment response was documented. Bacteriology confirmed tuberculosis: the presence of AFB on microscopy and/ or

M. Tuberculosis on culture.

Radiology confirmed PTB: the presence of any of childhood intrathoracictuberculosis manifestations in CXR.

Tuberculosis disease: either certain or probable tuberculosis.


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Latent tuberculosis infection: a positive IGRA and/or reactive TST in the absence ofcertain tuberculosis.

Tuberculosis exposed or healthy child TB contact, but no latent infection ordisease: negative IGRA and a negative TST in the absence of certain tuberculosis.

Nutritional status: will be classified using the z-score of WHO growth chart, whichis now the standard used in West Sumatera Indonesia. The classification is based on

the weight for height as follows:

Severe wasted: < - 3 SD Wasted: -2 to -2 SD Well-nourished: -2 to + 2 SD Overweight: +2 to + 3 SD

Obese: > + 3 SD

c. Tuberculosis related symptoms

Cough: any cough regardless the duration, remitting or not remitting Fever: body temperature of > 38 C Weight loss: weight loss or not gaining weight in the preceding 3 months or the loss

of more than 10% of bodyweight (minimum 1 kg) over any time interval) despite at

least 2 weeks of confirmed adequate nutrition

Fatigue: unexplained perceived decrease in playfulness/activity since the onset ofcoughing, reported by parents/caregiver

Chest pain: chest pain reported by the child. Hemoptysis: blood in the sputum (not hematemesis or a nose bleed) Night sweat: regular sweating that requires a dry set of nightclothes.

d. Vitamin D

Serum levels :

Normal : >30 ng/ml Insuficiency : 20-29 ng/ml Deficiency : 0-19 ng/ml

Genotype VDR polymorphism

FolkI ; TagI, BsmIDrugs

Cholecalciferol (vitamin D3)


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Dosages

25.000 IU

Preparation

In packing

Procedures

Recruitment

Source cases will be identified at the point of their treatment starting. Where there are

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children aged < 5 years or less living in the household, the source case will be informed

about this study by the attending clinician. Informed consent then will be asked from the

parent/guardian who agrees to involve to the study and whose children are eligible for the

study.

Baseline assessment

Home visit

After eligibility assessment and informed consent, study nurse/doctor will visit the

subject at home to identify the location of subjects house for follow up purpose and

to complete baseline data as follows:

Demographic and clinical data using a questionnaireSubjects characteristics, source case characteristics and other data

Physical examination Weight and height as well as general examination (detail in the

questionnaire)

Tuberculin skin test (TST)Intradermal injection of 0.1 ml of 2 TU of tuberculine purified protein

derivate (PPD) RT 23 in the volar by trained research nurses.

The result of TST of transversal diameter of induration will be measured at72 hours by study doctor/nurse at Sardjito hospital during hospital visit for

further invesigation. If the subject does not come to the hospital at the

scheduled time, home visit will be done to read the TST result,

The result will be considered positive if the induration is > 10 mm regardlessthe BCG vaccination, or > 5 mm in severe malnourished or

immunocompromised subjects.

Blood collectionWe will performs aseptically form of peripheral blood aspiration. Peripheral

venous blood sampling as much as 5 ml aseptic have done by study nurse. We

will examine blood samples within 1 x 24 hours for levels of vitamin D.

IGRA examination

We will determine the TB status by examination of the peripheral blood IGRA. If

increasing levels of IFN, confirmation checks will be performed 1 week later.

Increasing IFN in two measurements are considered positive. IGRA positive value

means that the sample is clinically latent tuberculosis (TB infection). Samples are


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included in this phase of research is showing negative IGRA results (not infected with

TB). IGRA is an analytical examination of IFN levels in cultured lymphocytes

induced with the ESAT-6. We will isolate the lymphocytes from blood with the

Hypaque Ficol then culture with the concentration of 1 x 10 6 cells / well and induced

with the ESAT-6 M. tuberculosis at a concentration of 0.5 ug / ml. On the 6th day of

culture, the supernatant is taken and examined levels of IFN with the IFN ELISA kit

(Abcam). The result is read with the ELISA READER. IFN concentrations are based

on the normal curve composed from normal values. For comparison,

phytohemaglutinin is used for positive control and aquadest as a negative control.

Clinical and Radiological Examination

All children is going through a routine clinical examination and radiological

examination if IGRA is increasing in the two measurements. If proven TB disease,

OAT treatment will be given. This child is not included in the study group.

DNA extraction

We will extract DNA from 0.2 ml of cell suspension morphonuclear using DNA

extraction kit (Qiagen, Chatsworth, USA). Protocol correspond with the procedures

developed by the manufacturer.

DNA amplification

Models and primers used for amplification are performed according to the protocolperformed by Liu et al (2006) with some modifications to the optimization.

Primers used were:

No Varian Primary

1 TaqI 5-cagagcatggacaggcaag-35-ggtggcagcggatgtacgt-3

2 Fok I 5-agctggccctggcactgactctgctct- 35-atggaaacaccttgcttcttctccctc-3

3 BsmI 5-aacttgcatgaggaggagcatgtc- 35-ggagaggagcctctgtcccatttg- 3

Stages of amplification is an initial denaturation: 94 o C for 15 seconds, annealing 65 o

C (TaqI), 55o C (FokI and BsmI) for 30 sec and extension 72 o C for 30 seconds follow

by amplification for 35 cycles. Final extension 72o C for 10 minutes

Restriction Fragment Length polymorphisms (RFLP)

PCR products are diluted in the ratio 1: 3, then digested with 5-10 U Tag I, FokI and


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BsmI. The result then electrophoreses on 2% agarose gel and visualized with

Cybergreen. Observations are made with a gel dock.

Allocation of vitamin D supplementation

All of the healthy children with close contact, divide into two group. First group is

supplementation of vitamin D which given orally at day 1 and day 42 and the second

group is given the placebo. Mixture of vitamin D will be prepared by the pharmacy staff

that have been selected. Vitamin D and placebo preparations known only by the two

researchers. The main research subjects and investigators and laboratory staff, do not know

what kind of preparations are given.

After 12 weeks of supplementation, we do TST and collect the blood sample to measure

the IFN level and vitamin D level. Higher IFN level and conversion of TST means the

children developed latent tuberculosis. After that we analyze is there any correlation

between vitamin D supplementation and latent tuberculosis. We also analyzed

polymorphism role in tuberculosis infection.

Statistics analysis

Data will be recorded on a data collection form at baseline and follow-up visits. Datawill be entered to the database which was developed using SPSS.

The data will be summarized as a proportion or a mean or median, where appropriate.

Comparison of proportions of outcomes between groups were tested using chi square(X2) test; whereas comparisons of mean or median used student t-test for normallydistributed data, and Mann-Whitney rank sum test for non parametric data. A p valueof < 0.05 was considered as statistically significant. Confidence intervals (CI) werecalculated for the prevalence of an outcome. The effectiveness of vitamin D in preventingLTBI will be presented as an Odd Ratio with 95% confidence interval (CI).

Descriptive statistics Frequencies (gender, contact TB history, level positive smear of index case, VDR

genetic polymorphism ), percent distributions.

Mean/Median and standard deviation/IQR for normally or no-normally distributeddata (age, socioeconomic, nutritional status, vitamin D level).

Analytic statistics Prevalence of vitamin D deficiency and VDR genetic polymorphysm with 95% CI. Chi-square test for comparisons of categorical variables. T-test for comparisons of continuous variables (age). Odds ratio and confidence interval will be calculated for studying the association


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between vitamin D level and incidence of latent TB, VDR genetic polymorphism

and incindence of latent TB.

Employ a two-tailed alpha () of 0.05 to determine significance.

Ethical Clearance

The ethical clearance will be based on the man using protocol as the research subject and

approved will be submitted to the by Ethical Commitee of Faculty of Medicine Andalas

University.

Informed Consent

All the subject are being informed about the research procedure, especially to the parent as the

person responsible, if they were understood and has given time to ask question, they will sign

the informed consent letter.

Quality Control

Selection Bias In order to ensure that the sample is representative, random sampling

approach will be used to select the community clinics for the study.

Information Bias We will use the structure questionnaire for collect the data about contact

history, TB symptom, etc.

Re-training laboratory technical. Pilot study: To prevent laboratory results bias, we will a pilot study to 15-20

blood sample, for examine vitamin D level, IGRA, VDR genetic

polymorphism, etc.

Telephone call-back or short message by enumerator will be done to ensurethe subject take the vitamin D daily.

Data Management We will use double data entry

Potential confounders We will use multivariate logistic regression to adjust for the impact of

nutritional status, level of positive smear, to vitamin D level, VDR

genetic polymorphism and incidence of latent TB.


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Strengths and Limitations

Strengths

1. The first study on vitamin D supplementation effect in child contact TB2. Quiet large simple size;3. Team members with expertise in Pediatrics TB, Clinical Nutrition, Biomolecular

Microbiology;

4. Use of validated effect of vitamin D supplementation in vivo

Limitations

1. We do not use normal control (no contact TB) to compare vitamin Dsupplementation effect.

2. We should still determine the age group < 5 years, according to the focus on thisage group to prevent latent TB in WHO policy, due to lack of immunity capability

to againt M. tuberculosis.

6.

OPERATIONAL PLANNING

This research is will be on going study which fund partly by IDAI (Ikatan Dokter Anak

Indonesia), we plan to start this research on July, 2013 with this following plan of action and

time schedule.

Plan of action and time schedule

No ActivitiesMonths

I II III IV V VI VII VIII IX X XI XII

1 Ethical clearance

2 Research protocol

3 Collecting sample

4Clinical examination, TST,chest X-rays

5Vitamin D levelexamination


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6VDR genetic

polymorphism

8 Data analysis

9 Report

Human Resources

NoRole in the research

study

Time Allocation

(hour/week)Responsibility in the research

1 Principal investigator 16 Do all research action

2 Research team 1 12 Focusing in laboratoriumexamination

3 Research team 2 12 Focusing in data collecting tocommunity

4 Advisor 6 Observer, advisor and supervisor theaction

Budget

NoItems Total Budget (%)

1 Material and supplies Rp 159.245.000 64,52

2 Data collection and analysis Rp 45.860.000 18,58

3 Salary and honorarium Rp 29.700.000 12,03

4 Travel cost Rp 12.000.000 4,86TOTAL Rp 246.805.000 100

The existing research fund (IDAI) could only covered about Rp 100.000.000,-. The gap of

research fund about Rp 146.805.000,-

Dissemination and utilization of the result

The result will be prove the role of vitamin D supplementation to develop latent TB. The

result could be dissemination to all pediatrician through Medical Journal, to association of

respirologist, to ministry of health and the TB program official. It will be influence the

prevention therapy and policy.

7. REFERENCES1. World health organization WH. Global Tuberculosis control 2011-survailance, planning,

financing. In: WHO report 2008. Geneva: World health organization; 2011.2. Joint external meeting for Tuberculosis in Indonesia. 2011


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3. Marais BJ GR, Schaaf HS, Hesseling AC, Obihara CC, Starke JJ, et al. The natural history ofchildhood intra-thoracic tuberculosis: a critical review of literature from the pre-chemotherapyera. Int J Tuberc Lung Dis 2004;8(S):392-402.

4. WHO. WHO guideline Tuberculosis in children.5. Claessens NJM GF, Meijnen S, Weismuller MM, Salaniponi FM, Harries AD. Screening

childhood contacts of patients with smear-positive pulmonary tuberculosis in Malawi [Notesfrom the Field]. Int J Tuberc Lung Dis 2002;6:362-4.

6. Rekha BVV JK, Wares F, Chandrasekaran V, Swaminathan S. Contact screening and

chemoprophylaxis in Indias Revised Tuberculosis Control Programme: a situational analysis.Int J Tuberc Lung Dis 2009;13:1507-12.

7. Rutherford ME RR, Maharani W, Yulita I, Lovell S, Crevel RV, et al. Adherence to isoniazidpreventive therapy in Indonesian children: a quantitative and qualitative investigation. 2012.

8. Mills HL CT, Colijn C. Community-wide isoniazid preventive therapy drives drug-resistanttuberculosis. ScTransi Medi 2013;5:1-9.

9. Battersby AJ K, Burl S. Vitamin D in early childhood and the effect on immunity tomycobacterium tuberculosis. Clinical and developmental immunology 2012;2012:1-10.

10. Helming L BJ, Ehrchen J, Sciebe S, Frahm T, Geffers R et al. 1 alfa 25 dihydroxyvitamin D3is a potent suppressor of interferon gamma mediated macrophage activation. Phagocytes2005;106:4351-8.

11. Fabri M SS, Min shin D, Yuk JM, Liu PT, Realegeno S, et al. Vitamin D is required for IFN

gamma mediated antimicrobial activity of human macrophages. Sci Transl Med 2011:1-24.12. Martineu AR LA, Anderson ST, Newton SM, Wilkinson KA, Nicol MP et all. Association

between Gc genotype and susceptibility to TB is dependent on vitamin D status. Eur respir J2010;35:1106-12.

13. Babb C VmL, Beyers N, Pheiffer C, Walzl G, Duncan K. Vitamin D receptor genepolymorhisms and sputum conversion time in pulmonary tuberculosis patients. Tuberculosis2007;87:295-302.

14. Setiawanbudi B . Peran defisiensi vitaminD dan polimorfisme Fokl, Bsml, Apal serta Taql genreseptor vitamin D terhadap tuberkulosis pada anak. Sari pediatri 2010;11(5):317-25.

15. Syafeii AZ SA, Setiabudiawan B. Association between serum vitamin D level and tuberculosisin children. Pediatrica Indonesiana 2008;48:350-3.

16. SEANUTS survey. 2012.17. Dini C BA. The potential role of vitamin D for prevention and infectious diseases. The

potential role of vitamin D for prevention and treatment of tuberculosis and infectious diseases2012;48(3):319-26.

18. Holick MF CT. Vitamin D deficiency: a worldwide problem with health consequences. Am JClin Nutr 2008;87:1080s-6s.

19. Nursyam. EW AZ, Rumende. CM. The effect of vitamin D as supplementary treatment inpatients with moderately advanced pulmonary tuberculosis lesion. Acta med indones-indones Jintern Med 2006:3-5.

20. AR M. Old wine in new bottles: vitamin D in the treatment and prevention of tuberculosis.Proceeding of Nutrition Society 2012;71:84-9.

21. Wejse C GV, Rabna P, Gustafson P, Aaby P, Lisse IM, et al. . Vitamin D as SupplementaryTreatment for Tuberculosis. . Am J Respir Crit Care Med 2009;179:843-50.

22. Setiabudiawan B KC, Garna H, Parwati I. Polimorfisme FokI,ApaI, dan TaqI gen reseptorvitamin D pada kejadian tuberkulosis anak. MKB 2010;42(4):187-94.

23. Leandro ACCS RM, Cardoso CSA, Almeida B. Genetic polymorphisme in vitamin D receptor,vitaminD-binding protein, toll-like receptor2, nitrit oxide synthase2, and interferon gammagenes and its association with susceptibility to tuberculosis. Braz J med biol res2009;42(4):312-22.

24. Reif AAM AP, Oki NO, Levy S, Holland SM, Sterling TR. Polymorphisms in IL-1B, vitaminD receptor Fok1, and Toll like receptor 2 are associated with extrapulmonary tuberculosis.BMC medical genetics 2010;11(37):1-10.

25. Martineu. AR WR, Wilkinson. KA, Newton SM, Kampmann. B, Hall BM, et all. A single dose

of vitamin D enhances immunity to mycobacteria. Am J Respir Crit Care med 2007;176:208-13.

26. Ganmaa. D GE, Bloom. BR, Fawzi. W, Burr. W, Batbaatar. Vitamin D, tuberculin skin testconversion, and latent tuberculosis in mongolian school-age chilodren: a randomized, double-blind, placebo-controlled feasibility trial. Am J Clin Nutr 2012:1-6.

27. Choudhary N GP. Vitamin D supplementation for severe pneuminia a randomized controlled


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26

trial Indian pediatrics 2012;49:449-54.28. Urashima M ST, Okazaki M, Kurihara M, Wada Y, Ida H. Randomized trial of vitamin D

supplementation to prevent seasonal influenza A in schoolchildren. Am J Clin Nutr2010;91:1255-60.

29. Departemen kesehatan Sumatera Barat .30. Data rekam medis RSUP M. Djamil 2013.31. Triasih R RM, Lestari T, Utarini A, Robertson CF, Graham SM. caont investigation of

children exposed to yuberculosis in south east asia: a systematic review. Journal of tropical

medicine 2012;2012:1-6.32. Basir D . Tuberkulosis pada anak yang kontak serumah dengan tuberkulosis paru dewasa BTA

sputum positif. MKS 2002:274-83.33. Triasih R . Child contact management of tuberculosis evaluation of symtom based screening in

yogyakarta, Indonesia. 2013.34. Smieja MJ MC, Cook DJ, Smaill FM. Isoniazid for preventing tuberculosis in non-HIV

infected persons. Cochran datbase of systematic review 1999;1.35. Bright-Thomas R NS, Smith J, Morris J, Ormerod LP. Effectiveness of 3 months of rifampicin

and isoniazid chemoprophylaxis for the treatmant of latent tuberculosis infection in children.Arch dis child online first 2010.

36. Hsu KH . Thirty years after isoniazid. Its impact on tuberculosis in children and adolescent.

JAMA 1984;9;251(10):1283-5.

37. Marais BJ GS, Cotton MF, Beyers. Diagnostic and management challanges for childhoodtuberculosis in the era of HIV. The Journal of Infectious Diseases Society 2007;196:76-85.

38. Goswami R MS, Kochupillai. Prevalence and potential significance of vitamin D deficiency inAsian Indians. India J Med Res 2008;127:229-38.

39. Scwalberg GK . A review of the critical role of vitamin D in the functioning of immune systemand the clinical implications of vitamin D deficiency. Mol Nurt Food 2010;55:96-108.

40. Lips P . Vitamin D physiology. Progress in biophysics and molecular biology 2006:4-8.41. Selvaraj P S. Vitamin D, Vitamin D receptor, and cathelicidin in the treatment of tuberculosis.

Vitamin and hormones 2011;86:307-25.42. Cheallaigh CNI KJ, Lavelle EC, Hope JC, Harris J. Autophagy in the immune response to

tuberculosis Clinical and experimental immunology 2011:1-10.

43. Kyeong JE. Innate immunity to mycobacteria: vitamin D and autophagy. Cellularmicrobiology 2010;12(8):1026-35.

44. Drobac PC SS, Huamani P, Atwood S, Furin J, Franke MF. Risk factor for in hospitalmortality among children with tuberculosis: the 25-years experinces in Peru. Pediatrics

2012;130:373-9.45. Talat N PS, Parsonnet J, Dawood G, Hussain R. Vitamin D defiency and tuberculosisprogression. Emerging infectious disease 2010;15(5):853-5.

46. Ho-Pham LT, NN, Nguyen DH, Bui PK, Nguyen VN, et al .Associationbetwen vitamin D insufficiency and tuberculosis in a vietnamese population. BMC infectiousdisease.2010; 10:306.

47. Salahuddin N AF, Hasan Z, Rao N, Aqeel M, Mahmood F. Vitamin D accelerated clinical

recovery from tuberculosis: results of the SUCCINT study (supplementary Cholecalciferol inrecovery from tuberculosis). A randomized, placebo-controlled, clinical trial of vitamin Dsupplementation in patients with pulmonary tuberculosis. BMC infectious diseases 2013;13:1-11.

48. Martineu AR NS, Oni T, Rangaka MX, Marais S, Bangani N, et al. Reciprocal seasonalvariation in vitamin D status and tuberculosis notifications in cape town, south africa. PNAS2011;108:19013-7.

49. Nnoham CA. Low serum vitamin D level and tuberculosis: a systematic review and

metaanalysis. International Journal Of epidemiology 2008;37:113-9.50. Albanna EAM, Elkashinia RAM. Vitamin D and LL-37 in children with pneumonia. Egypt Jpediatric Allergy immunol. 2010;8(2):81-6.51. Morcos MM GA, samuel S, Kamel M, El Baz M, El Beshry m, Michail RR. Vitamin D

administration to tuberculous children and its value. Boll Chim Farm 1998;137:157-64.52. Wejse C OR, Kaestel P, Gustafon P, Aaby P, Anderson PL, Glerup H, Sodemann M Serum 25

hidroksi vitamin D in a west african population of tuberculosis patients and unmatched healtycontrols Am J Clin Nutr 2007;86:1376-83.

53. FR G. 25-Hydroxyvitamin D: functional outcomes in infants and young children. Am J ClinNutr 2008;88:529S-33S.


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54. Norman AW BR. Vitamin D nutritional policy needs a vision for the future. Expermentalbiology and medicine 2010:1-12.

55. Tripkovic L LH, Hart K, Smith CP, Bucca G, Penson S, Chope G. Comparison of vitamin D2and vitamin D3 supplementation in raising serum 25-hydroxyvitamin D status: a systematicreview and meta-analysis. Am J Clin Nutr 2012;95:1357-64.

56. Misra M PD, Petryk A, Solberg PF, Kappy M. Vitamin D deficiency in children and itsmanagement: Review of current knowledge and recomendation. Pediatrics 2008;122:398-417.

57. R V. Vitamin D supplementation, 25 hydroxyvitamin D concentrations, and safety. Am J Clin

Nutr 1999;69:842-56.

58. Liu PT LH, Wenzel L, Tan BH, Krutzik SR. . Vitamin D3 triggered antimicrobial responseanother pleiotropic effect beyond mineral and bone metabolism . Czech J Anim Sci2014;58:2949-53.

8. APPENDICES

CONSENT TO PARTICIPATE IN RESEARCH

Effect of vitamin D supplementation to develope latent tuberculosis in healthychildren under five years age with tuberculosis contact :

a double blind randomized- control trial

Parental Consent form

We invite your child to participate in a research study conducted at the study site by Principal

investigators name (s). Your childs participation in this study is voluntary. You should read

the information below, and ask questions about anything you do not understand, before

deciding whether or not to allow your child to participate.

PURPOSE OF THE STUDY

This study is to examinine the child uder 5 years old which close contact to tuberculosis

adult patient at home. This child is highly possibility to get tuberculosis. By doing this

research, we will know the infection status and the treatment will be given if your child

infected. If your child didnt get tuberculosis infection, we will examine the blood to know

the level of vitamin D and genetic receptor of vitamin D. From many studies vitamin D,

could prevent the TB infection. We will give supplementation of vitamin D to your child

for 3 month and we will check the blood again and examine the infection status. The

supplementation and examination are free of charge.

DURATION AND LOCATION

The child will be supplemented vitamin D for 3 month, so your childs participation in this

study will last for approximately 3-4 months. This study will be conducted at Biomedical


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laboratory.

PROCEDURES

If you allow your child to volunteer to participate in this study, we would ask your child to

do the following things:

1. Get peripheral blood sample for interferon gamma examination, vitamin D level andpolymorphism, clinical evaluation, and chest X-rays.

2. Supplementation of vitamin D and placebo for 3 month3. Get peripheral blood sample interferon gamma examination and vitamin D level,

clinical evaluation after 3 months.

POTENTIAL RISKS AND DISCOMFORTS

1.No study is without some risk. Some procedure will discomforts the child, such asperipheral blood sampling and chest X-rays. It has some risk such as fail to get the

blood in one times punction. We will minimize this risk by doing the punction with

professional operator.

2. Having vitamin D supplementation will discomfort your child because she/he mustdrink the vitamin D daily for 3 months. So far, no risk of drink vitamin D but "there

may also be risks and discomforts which are not yet known".

ANTICIPATED BENEFITS TO SUBJECTS

The benefit of this research is to know the infection tuberculosiss status of your childand she/he will get the treatment if positive infected and get sick. If your child is

healthy, by giving vitamin D we will prove the protection effect of vitamin D to your

child. If it is proven, may be our government will implement a policy about vitamin D

supplementation to high risk children whom has close contact to tuberculosis adult

suffer.

PAYMENT FOR PARTICIPATION

All examination and the vitamin that will be given are free of charge.

CONFIDENTIALITY

When the results of the research are published or discussed in conferences, no

information will be included that would reveal your childs identity. If photographs,

videos, or audio-tape recordings of you will be used for educational purposes, your

childs identity will be protected or disguised.


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Authorized representatives of the Sponsoring agency, may need to review records of

individual subjects. As a result, they may see your childs name; but they are bound

by rules of confidentiality not to reveal your identity to others.

PARTICIPATION AND WITHDRAWAL

Your childs participation in this research is voluntary. If you or your child choose not

to participate, that will not affect your relationship or your childs relationship with

(study site) or your childs right to health care (or educational services) or other

services to which he or she is otherwise entitled. If you decide to allow your child to

participate, you are free to withdraw your consent and discontinue your childs

participation at any time without prejudice.

NEW FINDINGS

During the course of the study, you will be informed of any significant new findings

(either good or bad), such as changes in the risks or benefits resulting from

participation in the research or new alternatives to participation, that might cause you

to change your mind about your child continuing in the study. If new information is

provided to you, your consent to continue participating in this study will be re-

obtained.

IDENTIFICATION OF INVESTIGATORSIn the event of a research related injury or if your child experiences an adverse

reaction, please immediately contact one of the investigators listed below. If you or

your child have any questions about the research, please feel free to contact [Identify

the point of contact. Include the daytime telephone numbers and addresses. For

greater than minimal risk studies, include night/emergency telephone numbers.]

RIGHTS OF RESEARCH SUBJECTS

You may withdraw your consent at any time and discontinue your childs

participation without penalty. You are not waiving any legal claims, rights or

remedies because of your childs participation in this research study. If you have

questions regarding your childs rights as a research subject, please contact you may

contactPrincipal Investigator and Research team at _______________________.

You are making a decision whether or not to have your child participate in this study. Your


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signature indicates that you have decided to allow your child to participate, that you have read

(or been read) the information provided above and that you have received a copy of this

consent form.

Name of Child

Signature of Parent, or Person Responsible Date

Signature of Investigator Date

Signature of Witness Date

Detail Budget (attached)


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1. organization WH. Global Tuberculosis control 2011-survailance, planning,financing. In: WHO report 2008. Geneva: World health organization; 2011.2. joint external meeting.3. Marais BJ GR, Schaaf HS, Hesseling AC, Obihara CC, Starke JJ, et al. Thenatural history of childhood intra-thoracic tuberculosis: a critical review of literaturefrom the pre-chemotherapy era. Int J Tuberc Lung Dis 2004;8($):392-402.4. WHO. WHO guideline Tuberculosis in children.

5. Claessens NJM GF, Meijnen S, Weismuller MM, Salaniponi FM, Harries AD.Screening childhood contacts of patients with smear-positive pulmonary tuberculosisin Malawi [Notes from the Field]. Int J Tuberc Lung Dis 2002;6:362-4.6. Rekha BVV JK, Wares F, Chandrasekaran V, Swaminathan S. Contactscreening and chemoprophylaxis in Indias Revised Tuberculosis Control Programme:a situational analysis. Int J Tuberc Lung Dis 2009;13:1507-12.7. Rutherford ME RR, Maharani W, Yulita I, Lovell S, Crevel RV, et al.Adherence to isoniazid preventive therapy in indonesian children: a quantitative andqualitative investigation. 2012.8. Mills HL CT, Colijn C. Community-wide isoniazid preventive therapy drivesdrug-resistant tuberculosis. ScTransi Medi 2013;5:1-9.9. Battersby AJ K, Burl S. Vitamin D in early childhood and the effect on

immunity to mycobacterium tuberculosis. Clinical and developmental immunology2012;2012:1-10.10. Helming L BJ, Ehrchen J, Sciebe S, Frahm T, Geffers R et al. 1 alfa 25dihydroxyvitamin D3 is a potent suppressor of interferon gamma mediatedmacrophage activation. Phagocytes 2005;106:4351-8.11. Fabri M SS, Min shin D, Yuk JM, Liu PT, Realegeno S, et al. Vitamin D isrequired for IFN gamma mediated antimicrobial activity of human macrophages. SciTransl Med 2011:1-24.12. Martineu AR LA, Anderson ST, Newton SM, Wilkinson KA, Nicol MP et all.Association between Gc genotype and susceptibility to TB is dependent on vitamin Dstatus. Eur respir J 2010;35:1106-12.13. Babb C VmL, Beyers N, Pheiffer C, Walzl G, Duncan K. Vitamin D receptorgene polymorhisms and sputum conversion time in pulmonary tuberculosis patients.

Tuberculosis 2007;87:295-302.14. B S. Peran defisiensi vitaminD dan polimorfisme Fokl, Bsml, Apal serta Taqlgen reseptor vitamin D terhadap tuberkulosis pada anak. Sari pediatri 2010;11(5):317-25.15. Syafeii AZ SA, Setiabudiawan B Association between serum vitamin D leveland tuberculosis in children. Pediatrica Indonesiana 2008;48:350-3.16. SEANUT.


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32

17. Dini C BA. The potential role of vitamin D for prevention and infectiousdiseases. The potential role of vitamin D for prevention and treatment of tuberculosisand infectious diseases 2012;48(3):319-26.18. Holick MF CT. Vitamin D deficiency: a worldwide problem with health

consequences. Am J Clin Nutr 2008;87:1080s-6s.19. Nursyam. EW AZ, Rumende. CM. The effect of vitamin D as supplementarytreatment in patients with moderately advanced pulmonary tuberculosis lesion. Actamed indones-indones J intern Med 2006:3-5.20. AR M. Old wine in new bottles: vitamin D in the treatment and prevention oftuberculosis. Proceeding of Nutrition Society 2012;71:84-9.21. Wejse C GV, Rabna P, Gustafson P, Aaby P, Lisse IM, et al. . Vitamin D asSupplementary Treatment for Tuberculosis. . Am J Respir Crit Care Med2009;179:843-50.22. Setiabudiawan B KC, Garna H, Parwati I. Polimorfisme FokI,ApaI, dan TaqIgen reseptor vitamin D pada kejadian tuberkulosis anak. MKB 2010;42(4):187-94.23. Leandro ACCS RM, Cardoso CSA, Almeida B. Genetic polymorphisme in

vitamin D receptor, vitaminD-binding protein, toll-like receptor2, nitrit oxidesynthase2, and interferon gamma genes and its association with susceptibility totuberculosis. Braz J med biol res 2009;42(4):312-22.24. Reif AAM AP, Oki NO, Levy S, Holland SM, Sterling TR. Polymorphisms inIL-1B, vitamin D receptor Fok1, and Toll like receptor 2 are associated withextrapulmonary tuberculosis. BMC medical genetics 2010;11(37):1-10.25. Martineu. AR WR, Wilkinson. KA, Newton SM, Kampmann. B, Hall BM, etall. A single dose of vitamin D enhances immunity to mycobacteria. Am J Respir CritCare med 2007;176:208-13.26. Ganmaa. D GE, Bloom. BR, Fawzi. W, Burr. W, Batbaatar. Vitamin D,tuberculin skin test conversion, and latent tuberculosis in mongolian school-agechilodren: a randomized, double-blind, placebo-controlled feasibility trial. Am J ClinNutr 2012:1-6.

27. Choudhary N GP. Vitamin D supplementation for severe pneuminia arandomized controlled trialIndian pediatrics 2012;49:449-54.28. Urashima M ST, Okazaki M, Kurihara M, Wada Y, Ida H. Randomized trialof vitamin D supplementation to prevent seasonal influenza A in schoolchildren. AmJ Clin Nutr 2010;91:1255-60.29. Barat DkpS.30. Departement RP. Data pasien tuberculosis anak. 2013.31. Triasih R RM, Lestari T, Utarini A, Robertson CF, Graham SM. caontinvestigation of children exposed to yuberculosis in south east asia: a systematicreview. Journal of tropical medicine 2012;2012:1-6.32. D B. Tuberkulosis pada anak yang kontak serumah dengan tuberkulosis parudewasa BTA sputum positif. MKS 2002:274-83.

33. R T. Child contact management of tuberculosis evaluation of symtom basedscreening in yogyakarta, Indonesia. 2013.34. Smieja MJ MC, Cook DJ, Smaill FM. Isoniazid for preventing tuberculosis innon-HIV infected persons. Cochran datbase of systematic review 1999;1.35. Bright-Thomas R NS, Smith J, Morris J, Ormerod LP. Effectiveness of 3months of rifampicin and isoniazid chemoprophylaxis for the treatmant of latenttuberculosis infection in children. Arch dis child online first 2010.


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180813 final-1 revision content of proposal form 2a 2012-1_rt

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