Antibiotic Resistance and Microbiome in Children Aged 6-59 Months in Nouna, Burkina Faso
Phase: PHASE4
Conditions: Child Development
Sponsor: University of California, San Francisco

A nti bi otic R esist a nc e a n d Micr o bi o m e i n C hil dr e n A g e d 1- 5 9 m o nt hs  i n N o u n a, B ur ki n a F as o 
M a n u al of O p er ati o ns a n d Pr oc e d ur es 
V ersi o n 1. 2 , M a y 1 9, 2 0 1 7 
N C T n u m ber 0 3 1 8 7 8 3 4
C e ntr e d e R ec h erc h e e n S a nt é d e N o u n a 
U ni v ersit y of  H ei d el b er g 
Fr a ncis I. Pr oct or F o u n d ati o n, U ni v ersit y of C alif or ni a, S a n Fr a ncisc o 

 
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T a bl e of C o nt e nts  
 

 
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1.  O V E R VI E W 
 
1. 1  E x ec uti v e S u m m ar y  
 
T h e us e of a nti bi otics h as s a v e d milli o ns of h u m a n li v es, h o w e v er c o ns u m pti o n of a nti bi otics 
c a n s el ect f or a nti bi otic r esist a nt or g a nis ms a n d m a y l e a d t o c h a n g es i n c o m m e ns al 
micr o bi o m e. H er e, w e pr o p os e a s eri es of st u di es d esi g n e d t o esti m at e t h e eff ect of a nti bi otic 
c o ns u m pti o n o n a nti bi otic r esist a nc e a n d micr o bi o m e i n a r e gi o n of r ur al B ur ki n a F as o. I n St u d y 
O n e, w e will m e as ur e a nti bi otic c o ns u m pti o n at t h e c o m m u nit y l e v el a n d esti m at e t h e 
r el ati o ns hi p b et w e e n c o m m u nit y-l e v el a nti bi otic c o ns u m pti o n a n d c o m m u nit y-l ev el 
p n e u m oc occ al r esist a nc e. I n St u d y T w o, w e will m e as ur e c h a n g es i n t h e i nt esti n al a n d 
n as o p h ar y n g e al micr o bi o m e a n d r esist o m e f oll o wi n g a s h ort c o urs e of a nti bi otics. 
 
1. 2  O bj ecti v es  
 
O bj ecti v e 1. D et er mi n e w h et h er c o m m u niti es wit h hi g h er l e v els of a nti bi otic c o ns u m pti o n h a v e 
hi g h er pr e v al e nc e of p n e u m oc occ al r esist a nc e i n c hil dr e n u n d er 5 y e ars of a g e. 
 
O bj ecti v e 1 B. Esti m at e t h e pr e v al e nc e of acti v e tr ac h o m a i n a r ur al r e gi o n of N ort h w est er n 
B ur ki n a F as o. 
 
O bj ecti v e 2. D et er mi n e t h e eff ect of tr e at m e nt wit h a nti bi otics o n micr o bi o m e di v ersit y i n 
c hil dr e n f oll o wi n g a 5- d a y c o urs e of a nti bi otics. 
 
O bj ecti v e 2 A.  D et er mi n e t h e dir ect  eff ect of a 5- d a y c o urs e of azit hr o m yci n, a m oxicilli n, or 
c o-tri m ox az ol e o n i nt esti n al a n d n as o p h ar y n g e al b act eri al di v ersit y i n c hil dr e n a g e d 6-5 9 
m o nt hs c o m p ar e d t o n o tr e at m e nt.  
O bj ecti v e 2 B.  D et er mi n e t h e i n dir ect eff ect of a nti bi otic tr e at m e nt of c hil dr e n i n a 
h o us e h ol d o n i nt esti n al a n d n as o p h ar y n g e al b act eri al di v ersit y i n a n u ntr e at e d c h il d a g e d 6-
5 9 m o nt hs. 
O bj ecti v e 2 C.  Ass ess t h e ass oci ati o n b et w e e n i nt esti n al b act eri al di v ersit y a n d 
a nt hr o p o m etr y i n a p o p ul ati o n- b as e d s a m pl e of c hil dr e n. 
 
1. 3  St u d y Sit es  
 
T his st u d y will t a k e pl ac e wit hi n t h e N o u n a H e alt h a n d D e m o gr a p hic S ur v eill a nc e Sit e ( H D S S). 
T h e H D S S c o v ers a n ar e a of 1, 7 7 5 k m 2, c o v eri n g 1 0 3, 5 7 6 i n di vi d u als li vi n g i n 5 8 vill a g es as w ell 
as t h e t o w n of N o u n a. F or O bj ecti v e 1 , w e will i ncl u d e 2 0 vill a g es s er v e d b y 6 h e alt h c e nt ers: 
B o ur ass o, Si k or o, L a b ar a ni, L e k u y, N o k u y B o b o, a n d K o d o u g o u i n t h e s o ut h e ast of t h e H D S S. F or 
O bj ecti v e 2 , w e will i ncl u d e t h e vill a g es of D ar a a n d K a m a d e n a i n t h e w est er n r e gi o n of t h e 
H D S S. T o a v oi d c o nt a mi n ati o n b et w e e n st u di es, t h e t w o st u d y ar e as will n ot o v erl a p. 
 
2.  B A C K G R O U N D 
 

 
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Azit hr o m yci n a n d ot h er a nti bi otics ar e b ei n g c o nsi d er e d f or r o uti n e us e i n s etti n gs wit h hi g h 
b ur d e ns of c hil d m or bi dit y a n d m ort alit y. I n B ur ki n a F as o, t h e u n d er- 5 c hil d m ort alit y r at e is 
1 1 0 p er 1, 0 0 0 li v e birt hs, w hic h is a m o n g t h e hi g h est c hil d m ort alit y r at es i n t h e w orl d. 1 T h e 
m aj or c a us es of c hil d m ort alit y i n B ur ki n a F as o ar e i nf ecti o us: u p p er r es pir at or y tr act i nf ecti o n, 
di arr h e a, a n d m al ari a. 1, 2 F urt h er m or e, m al n utriti o n is a m aj or c o ntri b ut or t o c hil d m ort alit y a n d 
m or bi dit y, wit h a n esti m at e d 3 5 % of c hil dr e n m al n o uris h e d i n r ur al B ur ki n a F as o.
3 A nti bi otics 
m a y t h er ef or e r e d uc e c hil d m or bi dit y a n d m ort alit y b y tr e ati n g u n d erl yi n g ( oft e n s u bcli nic al) 
i nf ecti o ns. W e pr o p os e t o c o n d uct t his st u dy i n a r e gi o n of r ur al B ur ki n a F as o w h er e t h e b ur d e n 
of c hil d m ort ality a n d m or bi dity is hi g h, a n d c hil d d e at hs ar e attri b ut a bl e t o u p p er r es pir at ory 
tr act i nf ecti o n, di arr h e a, m al ari a, a n d m al n utriti o n. 
 
M ass azit hr o m yci n distri b uti o n f or tr ac h o m a c o ntr ol m a y l e a d t o s u bst a nti al d ecr e as e i n c hil d 
m ort alit y. 4 A r ec e nt st u d y i n r ur al Et hi o pi a n ot e d a n e arl y 5 0 % d ecr e as e i n c hil d m ort alit y i n 
c o m m uniti es t h at h a d r ec ei v e d a si n gl e m ass azit hr o m yci n distri b uti o n f or tr ac h o m a c o ntr ol. 
T h e us e of a nti bi otics m a y als o c o ntri b ut e t o i m pr o v e d gr o wt h a n d n utriti o n o utc o m es f or 
c hil dr e n i n r e gi o ns wit h a hi g h b ur d e n of m al n utriti o n. 5 A m oxicilli n is us e d r o uti n el y as p art of 
o ut p ati e nt tr e at m e nt of ac ut e m al n utriti o n. R a n d o miz e d c o ntr oll e d tri als of a m oxicilli n as 
a dj u ncti v e t h er a p y f or m al n utriti o n h as f o u n d t h at c hil dr e n r ec ei vi n g a nti bi otics g ai n m or e 
w ei g ht o v er a n 8- w e e k p eri o d c o m p ar e d t o c hil dr e n r ec ei vi n g pl ac e b o. 6, 7 Cl ust er-r a n d o miz e d 
tri als of c o m m u nit y- b as e d m a n a g e m e nt of p n e u m o ni a wit h c o-tri m ox az ol e f o u n d si g nific a ntl y 
l o w er all-c a us e u n d er- 5 m ort alit y i n c o m m u niti es r ec ei vi n g t h e c o-tri m ox az ol e i nt er v e nti o n.8 
A m oxicilli n a n d c o-tri m ox az ol e ar e als o us e d f or pr e v e nti o n of l o w er r es pir at or y tr act i nf ecti o ns 
i n hi g h-ris k c hil dr e n.9 A r ec e nt s yst e m atic r e vi e w a n d m et a- a n al ysis of a nti bi otic us e i n c hil dr e n 
i n l o w- a n d mi d dl e-i nc o m e s etti n gs f o u n d a c o nsist e nt, str o n g ass oci ati o n b et w e e n a nti bi otic 
us e a n d r e d uc e d pr e v al e nc e of w asti n g a n d st u nti n g, b ot h a m o n g c hil dr e n wit h a n d wit h o ut 
cli nic al i n dic ati o n f or a nti bi otics. 5 W e r ec e ntl y p erf or m e d a nt hr o p o m etric a n al ys es o n c hil dr e n 
r a n d o miz e d t o azit hr o m yci n v ers us n o tr e at m e nt f or tr ac h o m a a n d f o u n d a st atistic all y 
si g nific a nt i m pr o v e m e nt i n h ei g ht a m o n g c hil dr e n tr e at e d wit h a nti bi otics ( m a n uscri pt i n 
pr e p ar ati o n).  
 
Milli o ns of d os es of azit hr o m yci n ar e distri b ut e d i n pr e d o mi n a ntl y r ur al S u b- S a h ar a n Afric a n 
s etti n gs f or tr ac h o m a c o ntr ol as p art of m ass azit hr o m yci n distri b uti o n pr o gr a ms . I n a d diti o n 
t o dr astic all y r e d uci n g t h e pr e v al e nc e of tr ac h o m a gl o b all y1 0 -1 6, pr e vi o us w or k h as s h o w n t h at 
azit hr o m yci n distri b ut e d as p art of tr ac h o m a c o ntr ol pr o gr a ms h as l e d t o d ecr e as es i n 
di arr h e a 1 7, 1 8, s ki n i nf ecti o ns, l o w er r es pir at or y i nf ecti o n1 9, a n d m al ari a2 0, i n a d diti o n t o all-c a us e 
m ort alit y. 4 O n t h e ot h er h a n d, t h e ris ks of m ass azit hr o m yci n distri b uti o n ar e l ar g el y c o nfi n e d 
t o n o n-s eri o us a d v ers e e v e nts, i ncl u di n g a b d o mi n al p ai n, n a us e a, a n d v o miti n g 2 1, a n d tr a nsi e nt 
i ncr e as e i n c o m m u nit y r esist a nc e, a r esist a nc e w hic h h as n e v er b e e n s h o w n t o b e cli nic all y 
m e a ni n gf ul a n d r esist a nc e l e v els r et ur n t o n or m al l e v els w h e n a nti bi otic s el ecti o n pr ess ur e is 
r e m o v e d.2 2, 2 3 
 
T h er e is a p ot e nti al f or u ni nt e n d e d c o ns e q u e nc es wit h t h e us e of a nti bi otics, i n cl u di n g 
r e d uc e d b act eri al di v ersit y. A nti bi otics m a y alt er t h e h ost’s c o m m e ns al micr o bi ot a. 2 2, 2 4 -2 9 A 
r ec e nt st u d y a m o n g a nti bi otic- n aï v e c hil dr e n i n Ni g er f o u n d a si g nific a nt d ecr e as e i n b act eri al 

 
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di v ersit y i n c hil dr e n r ec ei vi n g a si n gl e d os e of azit hr o m yci n c o m p ar e d t o pl ac e b o 2 4, alt h o u g h 
t his st u d y di d n ot e v al u at e l o n g-t er m c h a n g es i n micr o bi o m e f oll o wi n g a n a nti bi otic c o urs e. T o 
d at e, t h er e h a v e b e e n v er y f e w r a n d o miz e d ass ess m e nts of c h a n g es i n i nt esti n al micr o bi o m e 
f oll o wi n g a c o urs e of a nti bi otics, a n d t h us t h e c a us al c h a n g es i n micr o bi o m e ar e u n d erst u di e d. 
F urt h er m or e, a n y f u ncti o n al diff er e nc es d u e t o alt er e d micr o bi o m e r e m ai n as- y et u n k n o w n, as 
micr o bi o m e st u di es ar e i n t h eir i nf a nc y a n d m ec h a nistic st u di es ar e l ac ki n g. C hil dr e n wit h 
m al n utriti o n h a v e b e e n s h o w n t o h a v e d ecr e as e d i nt esti n al b act eri al di v ersit y. 
3 0, T his 
ass oci ati o n is t h o u g ht t o b e a r es ult of p o or di et a n d i ncr e as e d s usc e pti bilit y t o i nf ecti o n t h at 
l e a ds t o str uct ur al c h a n g es i n t h e g ut, p o or a bs or pti o n of n utri e nts, a n d s u bs e q u e ntl y alt er e d 
micr o bi o m e. 3 1 F urt h er m or e, e n vir o n m e nt al e nt eric d ysf u ncti o n is t h o u g ht t o b e wi d es pr e a d i n 
r ur al S u b- S a h ar a n Afric a. T his c o n diti o n is m e di at e d i n p art b y c hr o nic i nf ecti o n wit h gr a m 
n e g ati v e b act eri a, a n d c a us es m or p h ol o gic c h a n g es i n t h e i nt esti n es t h at l e a d t o m al a bs or pti o n 
a n d s u bs e q u e nt m al n utriti o n a n d st u nti n g. 3 2 -3 4 Pr es u m pti v e tr e at m e nt wit h a nti bi otics m a y 
m o d ul at e t h e ass oci ati o n a n d l e a d t o i nt esti n al micr o bi o m e c h a n g es t h at ar e act u all y b e n efici al 
f or c hil d h e alt h. 
 
W e pr o p os e t w o st u di es d esi g n e d t o g ai n a b ett er u n d erst a n di n g of t h e m ec h a nis ms b e hi n d 
t h e p ot e nti al b e n efits a n d c o ns e q u e nc es of a nti bi otic us e i n s etti n gs wit h hi g h c hil d m ort alit y 
a n d m or bi dit y. W e s p ecific all y pr o p os e t o st u d y t h es e o utc o m es i n c hil dr e n a g e d 6- 5 9 m o nt hs, 
as t h e y ar e b y f ar at t h e gr e at est ris k of m ort alit y d u e t o i nf ecti o us c a us es a n d ar e at t h e 
hi g h est ris k of m al n utriti o n, a n d t h us w o ul d b e n efit t h e m ost fr o m c hil d m or bi dit y a n d 
m ort alit y i nt er v e nti o ns . T o d at e, t h er e h a v e b e e n v er y f e w r a n d o miz e d ass ess m e nts of 
c h a n g es i n t h e i nt esti n al micr o bi o m e f oll o wi n g a c o urs e of a nti bi otics, a n d t h us t h e c a us al 
c h a n g es i n micr o bi o m e ar e u n d erst u di e d. W e pr o p os e t w o st u di es t o 1) u n d erst a n d t h e eff ect 
of c o m m u nit y a nti bi otic c o ns u m pti o n o n c o m m u nit y a nti bi otic r esist a nc e a n d 2) t h e eff ect of a 
s h ort c o urs e of r o uti n e a nti bi otics o n c h a n g es i n i nt esti n al a n d n as o p h ar y n g e al micr o bi o m e a n d 
r esist o m e. W e h y p ot h esiz e t h at c o m m u niti es wit h hi g h er a nti bi otic c o ns u m pti o n will h a v e 
hi g h er pr e v al e nc e of p n e u m oc occ al r esist a nc e i n n as o p h ar y n g e al s a m pl es. F urt h er m or e, w e 
h y p ot h esiz e t h at a s h ort c o urs e of a nti bi otics will l e a d t o d ecr e as e d b act eri al di v ersit y s h ortl y 
aft er c o m pl eti o n of t h e a nti bi otic c o urs e, a n d hi g h er pr o b a bilit y of i d e ntific ati o n of b act eri al 
r esist a nc e g e n es i n r ect al a n d n as o p h ar y n g e al s a m pl es. 
 
3.  S T U D Y D E SI G N  
 
W e pr o p os e t w o st u di es t o a d dr ess t h e pr o p os e d o bj ecti v es. St u d y O n e  will b e a cr oss-
s ecti o n al, o bs er v ati o n al st u d y d esi g n e d t o ass ess t h e r el ati o ns hi p b et w e e n c o m m u nit y 
a nti bi otic c o ns u m pti o n a n d p n e u m oc occ al r esist a nc e pr e v al e nc e. St u d y T w o will b e a f o ur- ar m 
r a n d o miz e d c o ntr oll e d tri al d esi g n e d t o yi el d esti m at es of t h e eff ect of a s h ort c o urs e of 
a nti bi otics o n i nt esti n al a n d n as o p h ar y n g e al micr o bi o m e di v ersit y a n d r esist o m e. 
 
3. 1  St u d y O n e: C o m m u nit y a nti bi otic c o ns u m pti o n a n d pr e v al e nc e of p n e u m oc occ al 
r esist a nc e 
 

 
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St u d y O n e is a cr oss-s ecti o n al ass ess m e nt of c o m m u nit y a nti bi otic c o ns u m pti o n a n d 
n as o p h ar y n g e al p n e u m oc occ al r esist a nc e. T h e st u d y will occ ur i n t w o p h as es.  
 
D uri n g P h as e I, w e will extr act d at a fr o m r o uti n el y-c oll ect e d Pr escri pti o n S h e et fr o m 6 h e alt h 
c e nt ers l oc at e d i n t h e N o u n a H D S S. T h es e pr escri pti o n s h e ets ar e c oll ect e d a n d st or e d at t h e 
h e alt h f acilit y f or all c hil dr e n u n d er t h e a g e of 5. T h e s h e ets ar e t a k e n t o t h e p h ar m ac y w h e n 
t h e c ar e gi v er fills a pr escri pti o n f or t h e c hil d. W e will extr act a nti bi otic pr escri pti o ns ( a n d 
dis p e ns i n g, as t h es e s h e ets ar e fill e d w h e n t h e c ar e gi v er pic ks u p t h e pr escri pti o n) f or a p eri o d 
c o v eri n g a p pr oxi m at el y 6 m o nt hs (J a n u ar y t hr o u g h J u n e, 2 0 1 7). W e will extr act t h e f oll o wi n g 
d at a o n a st a n d ar diz e d, el ectr o nic d at a c oll ecti o n f or m (s e e S ecti o n 8, D at a C oll ecti o n a n d 
M a n a g e m e nt): 
 
•  H e alt h f acilit y n a m e 
•  D at e of pr escri pti o n  
•  Vill a g e of r esi d e nc e of t h e c hil d 
•  C hil d’s a g e 
•  C hil d’s s ex  
•  A nti bi otic  
•  D os e a n d d ur ati o n of a nti bi otic 
 
T o c alc ul at e a nti bi otic c o ns u m pti o n, w e will c alc ul at e t h e t ot al d os es of a nti bi otics a n d 
st a n d ar diz e usi n g t h e siz e of t h e vill a g e p er t h e m ost r ec e nt H D S S c e ns us. 
 
D uri n g P h as e II, w e will d et er mi n e p n e u m oc occ al r esist a nc e i n a r a n d o m s a m pl e of 1 5 c hil dr e n 
a g e d 6- 5 9 m o nt hs i n e ac h vill a g e i n t h e c atc h m e nt ar e a f or e ac h h e alt h f acilit y ( e. g., e ac h vill a g e 
f or w hic h a nti bi otic d at a w er e c oll ect e d d uri n g P h as e I). C hil dr e n will b e r a n d o ml y s el ect e d f or 
i ncl usi o n b as e d o n t h e m ost r ec e nt H D S S c e ns us. N as o p h ar y n g e al s w a bs will b e c oll ect e d fr o m 
e ac h r a n d o ml y-s el ect e d c hil d, w hic h will b e pr oc ess e d vi a Kir b y- B a u er disc diff usi o n ass a y at 
t h e C R S N micr o bi ol o g y l a b or at or y (s e e S ecti o n 4 f or d et ails o n s p eci m e n c oll ecti o n a n d S ecti o n 
6 f or d et ails o n micr o bi ol o gic al m et h o ds). I n a d diti o n, w e will p erf or m c o nj u ncti v al 
p h ot o gr a p h y a n d cli nic al e v al u ati o n of t h e c o nj u ncti v a o n e ac h e nr oll e d c hil d f or r a pi d 
ass ess m e nt of tr ac h o m a pr e v al e nc e i n e ac h vill a g e, if a c hil d is di a g n os e d wit h acti v e tr ac h o m a 
t h e g u ar di a n will b e gi v e n a t u b e of t etr ac ycli n e. C ar e gi v ers of c hil dr e n r a n d o ml y s el ect e d f or 
i ncl usi o n i n t h e st u d y will a d diti o n all y c o m pl et e a bri ef a nti bi otic us e s ur v e y. 
 
3. 1. 1  S a m pl e Siz e  
 
A t ot al of 6 h e alt h f aciliti es a n d 2 0 vill a g es will b e i ncl u d e d i n St u d y O n e. W e will c oll ect 
n as o p h ar y n g e al s w a bs fr o m 3 0 0 c hil dr e n i n t h e 2 0 vill a g es. D u plic at e s w a bs will b e c oll ect e d o n 
2 c hil dr e n p er vill a g e, s o a t ot al of 3 2 0 s w a bs will b e c oll ect e d. 
 
3. 1. 2  H e alt h C e nt er a n d C o m m u nit y  Eli gi bilit y  
 

 
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All h e alt h c e nt ers i n t h e H D S S will b e eli gi bl e f or i ncl usi o n. W e s el ect h e alt h c e nt ers b as e d o n 
t h eir acc essi bilit y d uri n g t h e r ai n y s e as o n, a n d s el ect c o m m u niti es i n a c o nti g u o us ar e a of t h e 
H D S S. All vill a g es i n t h e c atc h m e nt ar e a of t h e h e alt h f acilit y t h at ar e i n t h e H D S S will b e 
i ncl u d e d. 
  
3. 1. 3  I n di vi d u al Eli gi bilit y  
 
C hil dr e n  r esi di n g i n t h e vill a g es i n t h e c atc h m e nt ar e a of i ncl u d e d h e alt h f aciliti es d uri n g t h e 
l ast c e ns us w h o ar e 6- 5 9 m o nt hs will b e eli gi bl e f or r a n d o m i ncl usi o n i n t h e st u d y. W e will us e 
t h e c e ns us t o r a n d o ml y s el ect 1 5 c hil dr e n a g e d 6- 5 9 m o nt hs of a g e fr o m e ac h vill a g e. If t h e 
c hil d h as m o v e d o ut of t h e vill a g e or di e d si nc e t h e l ast c e ns us, w e will r a n d o ml y s el ect a n ot h er 
c hil d fr o m t h e c e ns us. 
 
3. 2  St u d y T w o: A nti bi otic tr e at m e nt a n d micr o bi o m e c h a n g es i n c hil dr e n 
 
St u d y T w o is a r a n d o miz e d tri al of a s h ort c o urs e of a nti bi otics ( azit hr o m yci n, c o-tri m ox az ol e, or 
a m oxicilli n) c o m p ar e d t o n o tr e at m e nt. W e will ass ess t h e eff ect of a s h ort a nti bi otic c o urs e o n 
i nt esti n al a n d n as o p h ar y n g e al b act eri al di v ersit y a n d will t est f or a nti bi otic r esist a nc e i n t h e 
r esist o me. H o us e h ol ds wit h at l e ast 2 c hil dr e n a g e d 6- 5 9 m o nt hs will b e r a n d o miz e d i n a 
1: 1: 1: 1 f as hi o n t o o n e of f o ur st u d y ar ms: 1) azit hr o m yci n, 2) a m oxicilli n, 3) c o-tri m ox az ol e, or 
4) n o tr e at m e nt. All c hil dr e n mi n us o n e i n t h e h o us e h ol d will b e tr e at e d, a n d w e will ass ess t h e 
i n dir ect eff ect of a nti bi otic tr e at m e nt o n t h e u ntr e at e d c hil d r esi di n g i n t h e h o us e h ol d. W e will 
m e as ur e c h a n g es i n micr o bi o m e t hr e e d a ys f oll o wi n g c o m pl eti o n of t h e a nti bi otic c o urs e 
( pri m ar y o utc o m e). D uri n g t h e fi n al s a m pl e c oll ecti o n w e will als o c o m pl et e a bri ef s ur v e y 
a b o ut t h e c hil dr e n’s h e alt h p artic ul arl y a b o ut di arr h e a a n d r es pir at or y i nf ecti o ns. Fi n all y, 4 
w e e ks aft er t h e c o m pl eti o n of t h e tr e at m e nt w e will e v al u at e t h e c hil dr e n’s gr o wt h b y d oi n g 
a nt hr o p o m etric m e as ur e m e nts. 
 
3. 2. 1  S a m pl e Siz e  
 
W e will  e nr oll 1 2 0 h o us e h ol ds i nt o t h e f o ur st u d y ar ms ( 3 0/ ar m). 
 
3. 2. 2  B as eli n e M e as ur e m e nt 
 
Aft er e nr oll m e nt a n d i nf or m e d c o ns e nt b ut pri or t o r a n d o miz ati o n , c hil dr e n will u n d er g o 
a nt hr o p o m etr y ( h ei g ht, w ei g ht, a n d mi d- u p p er ar m circ u mf er e nc e m e as ur e m e nts). C ar e gi v ers 
will c o m pl et e a bri ef b as eli n e s ur v e y t h at will i ncl u d e i nf or m ati o n o n w h et h er t h e c hil d is 
c urr e ntl y excl usi v el y br e astf e e di n g, is c urr e ntl y ex p eri e nci n g di arr h e a ( m e as ur e d b y t h e 
st a n d ar d W H O q u esti o n, “ H as t h e c hil d h a d 3 or m or e l o os e or w at er y st o ols i n t h e pr e vi o us 2 4 
h o urs”), a n d, f or n o n- excl usi v el y br e astf e e di n g c hil dr e n, t h e c hil d’s di et ar y di v ersit y. 
 
3. 2. 3  R a n d o miz ati o n 
 

 
8  
A list of p ot e nti all y eli gi bl e h o us e h ol ds will b e g e n er at e d fr o m t h e m ost r ec e nt c e ns us. T his list 
will b e s e nt t o t h e st u d y bi ost atistici a n ( T C P), w h o will g e n er at e t h e r a n d o miz ati o n list. T h e 
r a n d o miz ati o n list will b e 1: 1: 1: 1 a n d will b e g e n er at e d wit h o ut str atific ati o n or bl oc ki n g. 
 
T h e r a n d o miz ati o n list will c o nt ai n t h e r a n d o miz ati o n ar m a n d h o us e h ol d i d e ntific ati o n n u m b er 
a n d will b e tr a ns mitt e d t o t h e fi el d c o or di n at or at C R S N.    
 
3. 2. 4  C o m m u nit y Eli gi bilit y  
 
W e will s el ect t w o c o m m u niti es t h at h a v e at l ast 1 2 0 h o us e h ol ds wit h 2 or m or e c hil dr e n u n d er 
a g e 6- 5 9 m o nt hs at t h e m ost r ec e nt c e ns us. C hil dr e n w h o ar e all er gic t o a n y of t h e st u d y 
m e dic ati o ns will n ot b e e nr oll e d i n t h e st u d y. C hil dr e n w h o ar e r ec ei vi n g a nti bi otics f or a n 
o n g oi n g dis e as e w o n’t b e i ncl u d e d i n t h e st u d y 
 
3. 2. 5  H o us e h ol d Eli gi bilit y  
 
H o us e h ol ds will b e eli gi bl e f or i ncl usi o n i n t h e st u d y if t h e y h a v e 2 or m or e c hil dr e n a g e 6- 5 9 
m o nt hs c urr e ntl y r esi di n g i n t h e h o us e h ol d.  
 
3. 2. 6  St u d y M e dic ati o n 
 
St u d y m e dic ati o n will b e pr oc ur e d l oc all y f or a m oxicilli n a n d c o-tri m ox az ol e. Azit hr o m yci n a n d 
pl ac e b o will b e pr oc ur e d i n t h e U nit e d St at es as t h e y ar e n ot r e a dil y a v ail a bl e l oc all y, a n d 
s hi p p e d t o t h e st u d y sit e. All m e dic ati o ns will b e pr e p ar e d as a n or al s us p e nsi o n.  
 
3. 2. 6. 1  D os a g e I nf or m ati o n  
 
All c hil dr e n will r ec ei v e 5 d a ys of tr e at m e nt. D os a g es will b e a d mi nist er e d o nc e p er d a y f or all 
m e dic ati o ns wit h t h e exc e pti o n of a m oxicilli n, w hic h will b e t wic e p er d a y. D os a g e will b e 
w ei g ht - b as e d, b as e d o n t h e b as eli n e w ei g ht ass ess m e nt as p art of a nt hr o p o m etr y 
m e as ur e m e nts, as f oll o ws: 
 
•  Azit hr o m yci n: 1 0 m g/ k g o nc e d ail y o n D a y 1, t h e n 5 m g/ k g o nc e d ail y D a ys 2- 5 
•  A m oxicilli n: 2 5  m g/ k g / d a y, di vi d e d i nt o t wic e d ail y d os es  f or D a ys 1- 5 
•  C o -tri m ox az ol e: 2 4 0 m g d ail y f or D a ys 1- 5 
•  Pl ac e b o: O nc e d os e d ail y f or D a ys 1- 5 
 
3. 2. 6. 2  Dir ectl y O bs er v e d Tr e at m e nt  
 
All d os es of st u d y m e dic ati o n will b e dir ectl y o bs er v e d b y a fi el d w or k er. St u d y m e dic ati o n will 
b e pr e p ar e d o n t h e d a y of a d mi nistr ati o n b y st aff at t h e C R S N a n d will b e pr e-l o a d e d i nt o 
s yri n g es wit h t h e c orr ect d osi n g b as e d o n t h e c hil d’s w ei g ht fr o m t h e b as eli n e ass ess m e nt. E ac h 
s yri n g e will b e l a b el e d wit h t h e c hil d’s i d e ntific ati o n a n d h o us e h ol d I D n u m b er. All d os es of 
m e dic ati o n will b e r ec or d e d el ectr o nic all y o n t h e t a bl et.  

 
9  
 
3. 2. 6. 3  A d v ers e E v e nts S yst e ms 
 
All st u d y m e dic ati o ns ar e g e n er all y w ell-t ol er at e d. T h e m ost c o m m o n si d e eff ects of 
azit hr o m yci n ar e di arr h e a, a b d o mi n al p ai n, a n d v o miti n g, e ac h of w hic h m a y occ ur i n f e w er 
t h a n o n e i n t w e nt y p ers o ns w h o r ec ei v e azit hr o m yci n. R ar er si d e eff ects i ncl u d e a b n or m al li v er 
f u ncti o n t ests, all er gic r e acti o ns, a n d n er v o us n ess. Di arr h e a d u e t o Cl ostri di u m difficil e  h as b e e n 
r e p ort e d i n r ar e c as es. 
E ac h d a y, t h e st aff m e m b er will scr e e n all e nr oll e d c hil dr e n f or a n y a d v ers e e v e nt. All a d v ers e 
e v e nts ( n o n-s eri o us a n d s e v er e) will b e c o m m u nic at e d t o t h e M e dic al M o nit or o n a d ail y b asis. 
 
T h e m ost c o m m o n si d e eff ects of a m oxicilli n ar e di arr h e a, a b d o mi n al p ai n, a n d v o miti n g. R ac h 
or ot h er h y p ers e nsiti vit y r e acti o ns h a v e b e e n r e p ort e d i n u p t o 1 0 % of p ati e nts. Di arr h e a d u e 
t o Cl ostri di u m difficil e  h as b e e n r e p ort e d i n r ar e c as es. 
 
T h e m ost c o m m o n si d e eff ects of c o-tri m ox az ol e (s ulf a m et h ox az ol e/tri m et h o pri m) i ncl u d e 
n a us e a, di arr h e a, a n d h e a d ac h e. S o m e p ati e nts m a y d e v el o p a r as h.  
 
D uri n g t h e c o ns e nt pr oc ess, t h e c o m m o n a d v ers e r e acti o ns t h at m a y occ ur will b e ex pl ai n e d t o 
p ar e nts/ g u ar di a ns, a n d t h e y will b e a d vis e d t o c o m m u nic at e a n y a d v ers e e v e nts t o t h e l oc al 
h e alt h f acilit y i m m e di at el y. If, f or a n y r e as o n, t h e p artici p a nt n e e ds f urt h er c ar e, t h e y will b e 
r ef err e d t o t h e n e ar est h e alt h c e nt er f or ex a mi n ati o n a n d tr e at m e nt. T h e h e alt h f acilit y w or ks 
cl os el y wit h t h e N o u n a H D S S, a n d p ati e nts w h o ar e e nr oll e d i n t h e st u d y will b e i d e ntifi e d 
b as e d o n a u ni q u e st u d y c ar d t h at will b e gi v e n t o e ac h p ar e nt/ g u ar di a n f or e ac h st u d y 
p artici p a nt. A d v ers e e v e nts will b e r e p ort e d t o t h e st u d y t e a m i m m e di at el y.    
 
3. 2. 7  St u d y Sc h e d ul e  
 
 
D a y 0  
D a y 1 - 5 
D a y 9  
4 w e e ks aft er 
tr e at m e nt 
c o m pl eti o n 
E nr oll m e nt  
X 
 
 
 
A nt hr o p o m etr y 
X 
 
 
X 
S w a bs 
X 
 
X 
 
R a n d o miz ati o n * 
X 
 
 
 
Tr e at m e nt 
 
X 
 
 
O utc o m e s ur v e y 
 
 
X 
 
* Aft er all b as eli n e ass ess m e nts ar e c o m pl et e 
 
4.  S P E CI M E N C OLL E C TI O N P R O C E D U R E S 
 
4. 1  N as o p h ar y n g e al S w a bs  
 

 
1 0  
N as o p h ar y n g e al s w a bs will b e c oll ect e d fr o m a r a n d o m s a m pl e of 1 5 c hil dr e n p er vill a g e 
i ncl u d e d i n St u d y O n e f or ass ess m e nt of a nti bi otic r esist a nc e a n d fr o m all c hil dr e n i n St u d y T w o 
f or ass ess m e nt of n as o p h ar y n g e al micr o bi o m e. 
 
T h e ex a mi n er will: 
1.  Pl ac e a p e di atric fl oc k e d s w a b wit h a n yl o n ti p t hr o u g h t h e ri g ht n ostril a n d d o w n 
t h e n as o p h ar y nx of e ac h p artici p a nt. N ot e t h at if t h e s w a b is n ot p er p e n dic ul ar t o 
t h e fr o nt al pl a n e of t h e f ac e, it is li k el y n ot i n t h e i nf eri or t ur bi n at e. 
2.  O nc e y o u r e ac h t h e n as o p h ar y nx, r ot at e t h e s w a b 1 8 0° as y o u r e m o v e t h e s w a b 
fr o m t h e n os e.  
3.  Pl ac e t h e s w a b i n a t u b e c o nt ai ni n g 1. 0 mL D N A/ R N A s hi el d m e di a b y Z y m o or S T G G 
(s ki m mil k, tr y pt o n e, gl uc os e, a n d gl yc eri n) m e di a, c ut t h e h a n dl e off usi n g st eril e 
sciss ors, a n d cl os e t h e c a p of t h e t u b e wit h t h e s w a b i m m ers e d. 
4.  T h e n as o p h ar y n g e al s w a b s a m pl es i n S T G G will i niti all y b e st or e d i n t h e fi el d at 4° C 
usi n g a n i ns ul at e d st or a g e b a g wit h Fis h er br a n d ic e g el p ac ks, a n d t h e n tr a nsf err e d 
t o - 2 0° C. T h e n as o p h ar y n g e al s w a b s a m pl es i n D N A/ R N A s hi el d m e di a will b e st or e d 
i n a m bi e nt t e m p er at ur e i n t h e fi el d. T h e n tr a nsf err e d t o a r efri g er at or or fr e ez er. 
5.  T h e sciss ors us e d t o c ut c alci u m al gi n at e s w a bs will b e st eriliz e d wit h alc o h ol p a ds or 
cl e a n e d wit h bl e ac h wi p es b et w e e n p artici p a nts. W h e n c oll ecti n g s p eci m e ns i n 
D N A/ R N A s hi el d, sciss ors will b e cl e a n e d b et w e e n p artici p a nts - first wit h bl e ac h 
wi p es, a n d t h e n wit h alc o h ol p a ds.  
 
D o n ot att e m pt t o c oll ect t h e N P s w a b if y o u ar e n ot s ucc essf ul aft er t hr e e att e m pts.  
N as o p h ar y n g e al s w a bs will b e st or e d i n D N A/ R N A s hi el d m e di a b y Z y m o or S T G G m e di a, a n d 
st a n d ar d micr o bi ol o gic t ec h ni q u es will b e us e d t o is ol at e S. p n e u m o ni a e  a n d t est f or r esist a nc e 
t o er yt hr o m yci n, t etr ac ycli n e, s ulf a-tri m et h o pri n e, oxicilli n , a n d cli n d a m yci n. R esist a nt is ol at es 
will b e ass ess e d f or t h e m ost c o m m o n g e n etic r esist a nt d et er mi n a nts ( er m B  a n d m ef A ) usi n g a 
P C R - b as e d ass a y. S er ot y p e will b e ass ess e d usi n g a n est e d P C R r e acti o n f or t h e m ost c o m m o n 
s er ot y p es, f oll o w e d b y t h e Q u ell u n g r e acti o n f or a n y u nt y p e d is ol at es.  
 
M at eri als f or S w a b C oll ecti o n f or R esist a nc e T esti n g  
 
S w a bs  
N P s p eci m e ns will b e c oll ect e d usi n g st eril e, i n di vi d u all y- wr a p p e d p e di atric fl oc k e d s w a bs wit h 
a pl astic s w a b s h aft ( m a n uf act ur e d b y C o p a n). N as al sit es will b e s w a b b e d wit h a st eril e, D acr o n 
p ol y est er-ti p p e d s w a b wit h a pl astic s h aft ( m a n uf act ur e d b y Fis h er br a n d).   
 
S a m pl e T u b es  
All fi el d s a m pl es f or D N A t esti n g will b e c oll ect e d i nt o st eril e 2. 0 ml micr oc e ntrif u g e t u b es, 
m a n uf act ur e d b y S arst e dt ®. ( D N A -fr e e t u b es will b e us e d f or c oll ecti o n i n D N A/ R N A s hi el d.) 
 
 
C o ol er B a gs wit h Fr oz e n Ic e P ac ks 

 
1 1  
I ns ul at e d c o ol er b a gs will b e us e d t o c arr y s a m pl es t o a n d fr o m t h e fi el d.  I n a d diti o n, fr oz e n g el 
ic e p ac ks d esi g n e d t o t h a w sl o wl y will b e us e d t o m ai nt ai n t h e t e m p er at ur e i n t h e c o ol er b a gs 
d uri n g tr a ns p ort. 
 
- 8 0° C Fr e ez er 
A st a n d ar d - 8 0° C fr e ez er l oc at e d at t h e C R S N micr o bi ol o g y l a b or at or y will b e us e d f or t h e 
st or a g e a n d fr e ezi n g of ic e p ac ks a n d s a m pl es. T his fr e ez er is k e pt i n a l oc k e d r o o m o n t h e 
gr o u n ds of t h e C R S N , w hic h is u n d er 2 4- h o ur s ec urit y g u ar d s u p er visi o n. 
 
Pr ot oc ol f or T u bi n g a n d H a n dli n g of S a m pl es  
 
T h e t u bi n g a n d h a n dli n g pr ot oc ol m ust b e c ar ef ull y f oll o w e d i n or d er t o pr e v e nt c o nt a mi n ati o n 
a n d e ns ur e t h e s af e tr a ns p ort of t h e s a m pl es b ac k t o t h e C R S N micr o bi ol o g y l a b or at or y a n d/ or 
t o t h e U S f or pr oc essi n g. T h e p ers o n i n c h ar g e of l a b eli n g, t u bi n g, arr a n gi n g, a n d h a n dli n g t h e 
s a m pl es n e e ds t o p erf or m t his t as k i n t h e m ost or d erl y a n d att e nti v e m a n n er. 
 
1.  B ot h h a n ds of t h e t u b er s h o ul d b e gl o v e d at all ti m es. T h e t u b er's gl o v es o nl y n e e d t o b e 
c h a n g e d w h e n a n y p ot e nti al c o nt a mi n ati o n of t h e gl o v es occ urs. T h e t u b er o p e ns t h e 
c a p p e d, hi n g e d li d of a micr oc e ntrif u g e t u b e, w hic h h as b e e n l a b el e d wit h t h e 
p artici p a nt’s r a n d o m i d e ntific ati o n n u m b er. 
2.  T h e s w a b is i ns ert e d b y t h e ex a mi n er i nt o t h e micr oc e ntrif u g e t u b e h el d b y t h e t u b er.  
T h e s w a b s h aft s h o ul d o nl y b e i ns ert e d u ntil t h e s w a b h e a d is f ull y i n t h e t u b e. T h e 
t u b er will c ut t h e s w a b s h aft wit h st eril e sciss ors.  
3.  T h e t u b er s h o ul d scr e w t h e c a p of t h e micr oc e ntrif u g e t u b e ti g htl y, flic k t h e t u b e t o mix 
t h e s a m pl e wit h t h e m e di a (f or t u b es wit h D N A/ R N A s hi el d m e di a), a n d pl ac e it i n t h e 
s a m pl e c oll ecti o n b ox, l oc at e d i n t h e c o ol er b a g fill e d wit h fr oz e n ic e p ac ks. T h e fl a p of 
t h e c o ol er b a g s h o ul d b e cl os e d b et w e e n e ac h p ati e nt.  T h e c o ol er b a g s h o ul d b e i n as 
c o ol a pl ac e as p ossi bl e i n t h e fi el d, i n a s h a d e d ar e a o ut of t h e s u n. 
 
U p o n r et ur ni n g fr o m t h e fi el d e ac h d a y, t h e s a m pl es i n S T G G will b e i m m e di at el y t a k e n t o a 
l oc al h e alt h c e nt er a n d st or e d i n a c o m m erci al - 8 0° C fr e ez er, r es er v e d s ol el y f or st or a g e of 
s p eci m e ns a n d ic e p ac ks.  All s a m pl es will b e i n s a m pl e b ox es, l a b el e d wit h t h e vill a g e n a m e f or 
e as y f ut ur e i d e ntific ati o n.  
 
4. 2  R ect al S w a bs 
 
R ect al s w a bs will b e c oll ect e d a n d pl ac e d i nt o A mi es tr a ns p ort m e di a or N or g e n St o ol 
Pr es er v ati v e. T h e t est will r e q uir e t h at t h e c hil d’s p ar e nt a n d ex a mi n ers w or k t o g et h er t o 
o bt ai n a g o o d s a m pl e.  Is it i m p ort a nt t o d escri b e t h e t est t o t h e p ar e nt s o t h at t h e y c a n b est 
assist wit h k e e pi n g t h e c hil d still d uri n g t h e pr oc e d ur e, if n ec ess ar y. 
    
I n pl ac e of st o ol s p eci m e ns, r ect al s w a bs c a n b e c oll ect e d i n t h e f oll o wi n g w a y:  
1.  P ut o n a cl e a n p air of gl o v es. 

 
1 2  
2.  P arti all y o p e n t h e f ec al s w a b p ac k a g e a n d r e m o v e t h e t o p s ecti o n of t h e c oll ecti o n vi al 
(t his c a n b e disc ar d e d). 
3.  P ositi o n t h e c hil d: 
•  Li e t h e c hil d o n his/ h er b ac k, h ol d l e gs i n t h e air (it is us ef ul t o h a v e assist a nc e). 
•  Or h a v e t h e c hil d l a y o n his/ h er st o m ac h acr oss t h e m ot h er/ g u ar di a n’s l a p 
4.  R e m o v e t h e s w a b fr o m t h e p ac k a g e. T a k e c ar e t h at t h e c ott o n ti p is n ot t o uc h e d.  If it is 
t o uc h e d, t hr o w t h e s w a b a w a y a n d b e gi n wit h a n e w o n e. 
5.  I ns ert t h e ti p of t h e s w a b i nt o t h e c hil d’s a n us o nl y as f ar as n e e d e d t o c o nt act f ec al 
m at eri al ( 1- 3c m) a n d r ot at e 1 8 0 d e gr e es.  T h e ti p s h o ul d b e a br o w nis h c ol or w h e n 
r e m o v e d. 
6.  Pl ac e s w a b i nt o t h e pr es er v ati v e i n t h e c oll ecti o n t u b e. M a k e s ur e t h e s w a b is f ull y 
s u b m er g e d i n t h e li q ui d pr es er v ati v e a n d t h e n br e a k t h e s w a b off usi n g t h e pr e-sc or e d 
br e a ki n g p oi nt.  
7.  Scr e w t h e c a p b ac k o n t h e t u b e a n d m a k e s ur e t h at it’s ti g ht e n e d. Wr a p t h e ar e a w h er e 
t h e c a p m e ets t h e t u b e wit h P ar afil m t o e ns ur e t h at t h e s a m pl e will n ot l e a k, a n d t h e n 
pl ac e t h e t u b e i nt o t h e a p pr o pri at e s a m pl e b ox.  
•  If t h e s w a b c a n n ot b e br o k e n off w hil e t h e ti p is f ull y s u b m er g e d i n t h e li q ui d, tr y 
t wirli n g t h e s w a b i n t h e li q ui d first (t o r el e as e t h e c o nt e nts of t h e s a m pl e i nt o 
t h e pr es er v ati v e) b ef or e br e a ki n g it off. A v oi d r u b bi n g t h e s a m pl e o n t h e ti p of 
t h e s w a b off o n t h e si d e of t h e t u b e w h er e t h er e is n o li q ui d. 
8.  Pl ac e a r a n d o m n u m b er l a b el o n t h e c oll ecti o n t u b e. 
9.  Pl ac e t h e t u b e t h e r ect al s w a b c o nt ai n er. 
1 0.  S w a b st or a g e f or G e n etic a n al ysis:  St or e s a m pl es at r o o m t e m p er at ur e. Acc or di n g t o 
t h e m a n uf act ur er, t h e pr es er v ati v e i n t h e t u b e will pr es er v e D N A f or 5 m o nt hs at r o o m 
t e m p er at ur e ( 7 d a ys f or R N A), a n d t h er e aft er c a n b e fr oz e n (-2 0° C or -8 0° C) f or l o n g -
t er m st or a g e. 
 
R ect al S w a b C oll ecti o n f or C ult uri n g/ Micr o bi ol o gic al T esti n g  
 
S w a b  
A n i n di vi d u all y- wr a p p e d C o p a n fl oc k e d s w a b wit h a pl astic s h aft will b e us e d t o c oll ect t h e 
r ect al s w a b a n d t h e n pl ac e d i nt o a St o ol N ucl eic Aci d C oll ecti o n a n d Tr a ns p ort T u b e c o nt ai ni n g 
N or g e n St o ol Pr es er v ati v e or A mi es Tr a ns p ort M e di u m.   
 
S a m pl e T u b e wit h M e di a 
T h e s p eci m e n will b e i n a st eril e St o ol N ucl eic Aci d C oll ecti o n a n d Tr a ns p ort T u b e c o nt ai ni n g 
N or g e n St o ol Pr es er v ati v e or A mi es Tr a ns p ort M e di u m wit h a c a p t h at will b e ti g ht e n e d fir ml y.  
 
Q u alit y C o ntr ol M e as ur es f or S p eci m e n C oll ecti o n 
 
N e g ati v e Fi el d C o ntr ols  
N e g ati v e fi el d c o ntr ol s w a bs f or N P a n d st o ol will b e t a k e n i n e ac h c o m m u nit y t o ass ess f or 
c o nt a mi n ati o n: o n e co ntr ol s w a b e ac h ( N P a n d st o ol/r ect al) ar e t a k e n b ef or e s p eci m e n 
c oll ecti o n b e gi ns i n a c o m m u nit y; a n d a n ot h er ( N P a n d st o ol/r ect al) u p o n c o m pl eti o n of 

 
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s p eci m e n c oll ecti o n. F or e ac h n e g ati v e fi el d c o ntr ol, t h e ex a mi n er will o p e n a n e w s w a b as 
d escri b e d a b o v e. W a v e t h e s w a b i n t h e air, wit h o ut m a ki n g c o nt act wit h a n y o n e/ a n yt hi n g.  
T u b e t h e s w a b i n m e di a, as d escri b e d a b o v e.  
 
D u plic at e s w a bs  
D u plic at e N P s w a bs a n d r ect al s w a bs/st o ol s p eci m e n will b e c oll ect e d fr o m t w o c hil dr e n p er 
c o m m u nit y.  
 
St o ol s a m pl es  
 
E ac h c ar e gi v er will b e s u p pli e d wit h a b a g a n d pl astic c hil d’s p ott y c h air a n d i nstr uct e d t o h a v e 
t h eir c hil d d ef ec at e i n a b a g a n d r et ur n t h e b a g t o t h e st o ol c oll ecti o n st ati o n. C hil dr e n t h at 
c a n n ot pr o d uc e a st o ol wit hi n t w o h o urs will b e gi v e n s u p pli es t o c oll ect t h e st o ol at h o m e. 
C ar e gi v ers will b e i nstr uct e d t o c oll ect st o ol t h e f oll o wi n g m or ni n g, aft er w hic h a st u d y t e a m 
m e m b er w o ul d c o m e t o t h eir h o us e t o pic k u p t h e s p eci m e n. Fr o m a si n gl e st o ol s p eci m e n, w e 
will pr es er v e t w o s a m pl es: 0. 5 ml of st o ol i n 0. 5 ml et h a n ol a n d 1 gr a m of st o ol i n 1 0 ml s o di u m 
ac et at e- ac etic aci d f or m ali n ( S A F) f or micr osc o p y. St o ol s a m pl es pr es er v e d i n et h a n ol a n d 
p ot assi u m dic hr o m at e s a m pl es will b e pl ac e d o n ic e i m m e di at el y aft er c oll ecti o n a n d 
tr a ns p ort e d t o a - 2 0º C fr e ez er b y t h e e n d of t h e d a y. S A F- pr es er v e d s a m pl es will b e k e pt at 
r o o m t e m p er at ur e i n t h e s h a d e. 
 
S u p pli es f or st o ol s a m pl e c oll ecti o n 
 
•  C hil d p ott y  
•  W o o d e n d o wls  
•  Di git al s p eci m e n sc al e 
•  St o ol s p eci m e n t u b e c o nt ai ni n g 1 0 ml S A F pr es er v ati v e 
•  2 ml t u b es f or P C R pr es er v ati o n 
•  A p plic at or stic k 
•  Alc o h ol wi p e  
•  Fi n e p er m a n e nt M ar k er 
•  Tr as h b a g 
•  Gl o v es  
•  B att eri es 
•  S a nitizi n g h a n d wi p es  
 
L a b or at ory m et h o ds 
 
a. R e al-ti m e t ar g et e d P C R  
D N A  will  b e  extr act e d  fr o m  r ect al  s w a bs  a n d  st o ol  s a m pl es  usi n g  t h e  t h e  Z R- 9 6  D N A  Cl e a n  & 
C o nc e ntr at or ™- 5  Kit  ( Z y m o  R es e arc h  C or p or ati o n,  Ir vi n e,  C A).  D N A  c o nc e ntr ati o n  will  b e 
m e as ur e d  usi n g  t h e  Q u bit  1. 0  Fl u or o m et er  (Lif e  T ec h n ol o gi es,  C arls b a d,  C A)  a n d  t h e  Q u bit 

 
1 4  
ds D N A  Br o a d  R a n g e  Ass a y  Kit  (Lif e  T ec h n ol o gi es).  Q u a ntit ati v e  r e al-ti m e  p ol y m er as e  c h ai n 
r e acti o n ( q P C R) will b e p erf or m e d t o f or s p eci ati o n a n d q u a ntific ati o n of t h e p ar asit e b ur d e n as 
pr e vi o usl y d escri b e d b y Pil o ett e et al 1 8. S p eci es-s p ecific q P C R a m plific ati o n will b e c o n d uct e d i n 
7 μl v ol u m es, c o nt ai ni n g 3. 5 μl of 2 X T a q M a n F ast U ni v ers al P C R M ast er Mix (Lif e T ec h n ol o gi es), 
1 2 5 n m ol of e ac h ass a y’s r es p ecti v e pr o b e, 2 μl of t e m pl at e D N A at a c o nc e ntr ati o n of 1 n g/ μl, 
a n d r es p ecti v e c o nc e ntr ati o ns of e ac h s p eci es-s p ecific P C R r e acti o n d e v el o p e d b y Pil ott e et al 
(s e e  T a bl e  2). T h er m oc ycli n g  c o n diti o ns  will  b e  as  f oll o ws:  a n  i niti al  2  mi n  i nc u b ati o n  st e p  at 
5 0° C, f oll o w e d b y a 1 0 mi n i nc u b ati o n at 9 5° C. T h es e i nc u b ati o ns will b e f oll o w e d b y 4 0 c ycl es 
of 9 5° C f or 1 5 s ec f or d e n at ur ati o n, f oll o w e d b y 1 mi n at 5 9° C f or a n n e ali n g a n d ext e nsi o n. All 
r e acti o ns will b e c o n d uct e d usi n g t h e St e p O n e Pl us R e al- Ti m e P C R S yst e m (Lif e T ec h n ol o gi es). 
 
b. Micr osc o py  
All  s o di u m  ac et at e- ac etic  aci d  f or m ali n  pr es er v e d  s a m pl es  will  b e  pr oc ess e d  at  t h e  C R S N 
l a b or at or y  usi n g  t h e  di et h yl  et h er-c o nc e ntr ati o n  m et h o d  a n d  ex a mi n e d  micr osc o p y  f or  s oil-
tr a ns mitt e d  h el mi nt h  e g gs  a n d  l ar v a2 4.  Bri efl y, t h e  pr es er v e d  s p eci m e n  will  b e  g e ntl y  s h a k e n, 
t h e n  filt er e d  a n d  c e ntrif u g e d  f or  o n e  mi n ut e  at  2 0 0 0  r e v ol uti o ns  p er  mi n ut e  (r p m).  T h e 
s u p er n at a nt  will  b e  d ec a nt e d  a n d  t h e  pr eci pit at e  mix e d  wit h  7  ml  of  0. 8 5 %  N a Cl  a n d  3  ml  of 
di et h yl et h er. T h e w h ol e s a m pl e will b e c e ntrif u g e d a g ai n f or fi v e mi n ut es at 2 0 0 0 r p m 2 4, t h e 
s u p er n at a nt  d ec a nt e d  a g ai n,  a n d  t h e  s e di m e nt  ex a mi n e d  wit h  a  micr osc o p e  at  1 0x  a n d  4 0x 
m a g nific ati o n  f or  h el mi nt h  e g gs  a n d  i nt esti n al  pr ot oz o a  c ysts.  F or  h el mi nt hs,  t h e  n u m b er  of 
e g gs  will  b e  c o u nt e d  a n d  r ec or d e d,  wit h  a  p ossi bl e  r a n g e  of  1  u p  t o  > 1 0 0  e g gs.  As  q u alit y 
c o ntr ol, a n i n d e p e n d e nt ex p ert l a b or at or y t ec h nici a n will c o nfir m all p ositi v e sli d es, as w ell as 
e v er y 1 0 t h n e g ati v e s p eci m e n.  
 
4. 3  C o nj u ncti v al P h ot o gr a p h y  a n d Cli nic al e v al u ati o n.  
 
4. 3. 1 E x a mi n ati o n p ositi o n  
T h e ex a mi ni n g p ositi o n t o b e us e d i n t h e fi el d f or y o u n g c hil dr e n will b e t h e cl assic p e di atric 
o p ht h al mic ex a mi n ati o n t ec h ni q u e. Wit h t h e ai d of a h el p er s e at e d dir ectl y o p p osit e t o t h e 
ex a mi n er, t h e c hil d will b e p ositi o n e d wit h his/ h er h e a d b et w e e n t h e ex a mi n er's k n e es, wit h 
t h e c hil d's f ac e l o o ki n g u p w ar ds t o w ar d t h e ex a mi n er.  T h e l e gs of t h e c hil d will b e str a d dl e d 
acr oss t h e h el p er a n d t h e ar ms h el d g e ntl y acr oss t h e c hil d’s c h est.  C ar e s h o ul d b e t a k e n t o 
k e e p t h e c hil d’s e y es a b o v e t h e l e v el of t h e ex a mi n er’s k n e es, i n or d er t o pr o p erl y t a k e t h e 
c o nj u ncti v al s w a b. 
 
4. 3. 1 E v erti n g t h e u p p er e y eli d  
F or all p artici p a nts, o nl y t h e ri g ht u p p er e y eli d will b e ex a mi n e d f or t his st u d y.  T h e o nl y 
exc e pti o n t o t his is if t h e ri g ht e y e is diffic ult t o ex a mi n e d u e t o e y e dis e as e or i nj ur y, i n w hic h 
c as e t h e l eft e y e will b e ex a mi n e d. I n or d er t o a v oi d p assi n g c o nt a mi n ati o n fr o m t h e c hil d i nt o 
t h e e y e, o nc e t h e ex a mi n er d o ns a n e w p air of gl o v es, t h eir gl o v e d h a n ds s h o ul d n ot b e us e d t o 
p osit i o n t h e c hil d. T h e ex a mi n er us es t h eir fi n g erti ps t o gr as p t h e c e ntr al p orti o n of t h e 
p artici p a nt’s u p p er li d e y el as h es. T h e u p p er li d is t h e n e v ert e d, usi n g a fi n g er of t h e ex a mi n er's 
ot h er h a n d ( or t h e e n d of a st eril e s w a b) as a f ulcr u m, p ositi o n e d s u p e rior t o t h e t ars al pl at e. 

 
1 5  
T h e e v ert e d li d is h el d i n pl ac e b y t h e ex a mi n er's n o n- d o mi n a nt h a n d h ol di n g t h e e y el as h es 
a g ai nst t h e or bit al ri m. 
 
4. 3. 3 C o nj u ncti v al p h ot o gr a p h y  
S p ecific  t e a m  m e m b ers  will  b e  d esi g n at e d  as  p h ot o gr a p h ers  f or  t h e  e ntir e  tri al  i n  or d er  t o 
e ns ur e hi g h q u alit y p h ot o gr a p hs b y a n ex p eri e nc e d p h ot o gr a p h er. T h e p h ot o gr a p h er will t a k e 
p h ot o gr a p hs  of  t h e  c o nj u ncti v a  wit h  a  h a n d h el d  Ni k o n  D 9 0  di git al  c a m er a  wit h  a  m acr o  l e ns 
( 1: 1). C o nj u ncti v al p h ot o gr a p h y c a us es n o d a m a g e t o t h e e y e, is w ell t ol er at e d b y c hil dr e n, a n d 
is  a  st a n d ar d  cli nic al  pr oc e d ur e  at  U C S F.  P h ot o gr a p hs  will  b e  i d e ntifi e d  b y  first  t a ki n g  a 
p h ot o gr a p h of t h e 6- di git r a n d o m i d e ntific ati o n stic k er t h at w as assi g n e d t o t h e c hil d d uri n g t h e 
r e gistr ati o n p h as e of t h e st u d y visit. B y c o n v e nti o n, all p h ot os t a k e n aft er t his r a n d o m n u m b er 
( u p t o t h e n ext p h ot o gr a p h e d r a n d o m n u m b er) b el o n g t o t h e pr ec e di n g r a n d o m n u m b er. If a 
r a n d o m n u m b er is mist a k e nl y n ot p h ot o gr a p h e d b ef or e t a ki n g a p h ot o gr a p h of t h e c o nj u ncti v a, 
it m a y b e p h ot o gr a p h e d aft er w ar ds; i n t his c as e, a n ot e s h o ul d b e i ncl u d e d t h at s a ys “ pr ec e di n g 
p h ot os.” P h ot o gr a p hs ar e st or e d o n t h e m e m or y c ar d i n t h e c a m er a, a n d t h e n tr a nsf err e d t o a n 
ext er n al h ar d dri v e e ac h ni g ht. B att eri es ar e c h ar g e d e ac h ni g ht; e ac h c a m er a b a g h as at l e ast 2 
s p ar e b att eri es. 
 
P h ot o gr a p h y Pr ot oc ol  
§  E q ui p m e nt n e e d e d: c a m er a, l a b els, p h ot o l o g b o o k, a n d C D b ur n er 
§  Misc ell a n e o us e q ui p m e nt: Extr a b att er y, c ar d r e a d er, l e ns filt er, extr a m e di a c ar d  
 
C a m er a S et u p  
A  h a n d h el d  Ni k o n  D-s eri es  c a m er a  a n d  l e ns  will  b e  us e d  f or  all  p h ot o gr a p hs.  If  p ossi bl e,  us e 
n at ur al li g ht r at h er t h a n a fl as h, as t h e fl as h i ntr o d uc es artif acts i nt o t h e i m a g e.  
§  M o d el 
o  Ni k o n D -s eri es c a m er a: D-4 0, 5 0, 7 0, 8 0, 9 0, 1 0 0 or 2 0 0  
o  T h er e  ar e  mi n or  diff er e nc es  b et w e e n  t h e  diff er e nt  m o d els,  m ostl y  r el at e d  t o 
s et u p 
§  L e ns 
o  Ni k o n M acr o A ut of oc us 1 0 5 m m f/ 2. 8 
§  S etti n gs 
o  M a n u al s etti n g- t ur n off a ut of oc us 
o  Dis e n g a g e “li mit”  e n g a g e “f ull” 
o  Ext e n d l e ns t o 1: 1; n ot e wit h ol d er v ersi o ns of t his l e ns it is 1: 1 at f ull ext e nsi o n; 
n e w er v ers i o ns c a n g o t o sli g htl y hi g h er m a g nific ati o n t h a n 1: 1 s o c a uti o n m ust 
b e ex ert e d 
o  I m a g e siz e/r es ol uti o n 
§  J p g n or m al- s et t hr u “s et u p” m e n u 
§  L ar g e-  s et  t hr u  “s et u p”  m e n u  (“s m all  a n d  m e di u m”  ar e  pr o b a bl y 
a d e q u at e) 
§  W hit e b al a nc e- a ut o m atic or fl as h; s et t hr u “s et u p” m e n u 
§  S h utt er- A p ert ur e Pri orit y 
§  f 5 7- s et t hr u m e n u; n ot e if l e ns is n ot ext e n d e d t o 1: 1, f 5 7 will n ot b e all o w e d 

 
1 6  
§  I S O 4 0 0 
 
P h ot o gr a p h Pr oc e d ur e  
§  Pl ac e p ati e nt i nt o p ositi o n t h at will all o w m axi m u m st a bilit y; st a n di n g, sitti n g or “ h e a d-
cl a m p” p ositi o n. E m pl o yi n g vill a g e v ol u nt e er t o h el p is v er y us ef ul. 
§  T a k e p h ot o of p ati e nt I D 
§  I m a g e is br o u g ht i nt o f oc us b y c h a n gi n g t h e w or ki n g dist a nc e, n ot b y t ur ni n g t h e l e ns. 
T his  is  b ec a us e  t h e  l e ns  is  fix e d  i n  its  m a n u al  s etti n g.  T h e  w or ki n g  dist a nc e  is 
a p pr oxi m at el y 2 0 c m fr o m t h e e y e wit h o ur c urr e nt s etti n gs 
§  T a k e mi ni m u m of 2 p h ot os. If t h er e is a n y d o u bt of t h e q u alit y of t h e p h ot o w hil e t h e 
p ati e nt  is  i n  p ositi o n  it  is  b ett er  t o  c o nti n u e  t o  t a k e  m or e  p h ot o gr a p hs  b ef or e  t h e 
p ati e nt is all o w e d t o l e a v e. It is e as y t o d el et e p h ot os if t h e y ar e n ot n e e d e d. 
§  C h ec k  p h ot os  b ef or e  all o wi n g  c hil d  t o  l e a v e.  If  t h e y  ar e  n ot  acc e pt a bl e,  r e p e at 
pr oc e d ur e. O nl y st o p if t h e p ati e nt or g u ar di a n r e q u ests t h at w e st o p, or if is d e e m e d 
i m p ossi bl e,  e v e n  wit h  f urt h er  att e m pts.    N ot e  w e  h a v e  o bt ai n e d  > 9 5 %  acc e pt a bl e 
p h ot os i n pr e vi o us st u di es. 
§  If t h e c hil d c a n n ot b e p h ot o gr a p h e d f or s o m e r e as o n, it d o es n ot aff ect t h e eli gi bilit y of 
t h e  c hil d.  A  n ot ati o n  is  m a d e  o n  t h e  oc ul ar  f or m  of  w h y  t h e  p h ot o gr a p h  c a n n ot  b e 
t a k e n, b ut n o r e pl ac e m e nt is s o u g ht.  A g ai n, t his is ex p ect e d t o b e a r ar e e v e nt. 
 
P h ot o  Tr o u bl es h o oti n g G ui d e  
 
N ot e  all  c a m er a  s etti n gs  ar e  p er m a n e ntl y  e m b e d d e d  i n  e v er y  p h ot o  t h at  is  t a k e n  a n d  c a n  b e 
vi e w e d wit h t h e c a m er a or wit h a n y st a n d ar d c o m m erci al p h ot o- vi e wi n g s oft w ar e ( e. g., A d o b e 
P h ot os h o p; P h ot o m ec h a nic, Ni k o n Vi e w, C a n o n, etc.) 
§  P h ot os t o o D A R K, t o o LI G H T or O U T O F F O C U S 
o  C h ec k c a m er a s etti n gs: I S O 4 0 0, A p ert ur e pr ef err e d, +/- o n z er o, M a n u al s etti n g 
o n l e ns, fl as h el e v at e d; b att er y f ull y c h ar g e d 
§  If  all  p h ot os  ar e  aff ect e d  i n  t h e  s a m e  w a y  t h a n  it  is  m ost  li k el y  t h e  s etti n gs  o n  t h e 
c a m er a or l e ns 
o  C h a n g e c a m er a s etti n gs fr o m d ef a ult r ec o m m e n d ati o ns 
o  D ecr e as e F st o p t o li g ht e n, i ncr e as e t o d ar k e n 
o  C o ul d b e t a k e n d o w n t o 3 2 or l o w er b ut d e pt h of fi el d w ill b e l ess e n e d 
o  +/- c a n b e i ncr e as e d or d ecr e as e d t o c h a n g e ex p os ur e ( + t o li g ht e n; - t o d ar k e n) 
o  C h ec k t h at l e ns is o n M a n u al N O T A ut of oc us 
o  I S O- i ncr e as e t o li g ht e n p h ot o, d ecr e as e t o d ar k e n 
o  V er y hi g h I S O will pr o d uc e gr ai ni n ess 
o  M et er- tr y “s p ot” i nst e a d of “ p att er n 
o  H a v e a n ot h er p ers o n o bs er v e p h ot o gr a p h y i n r e al ti m e 
o  M a k e s ur e fl as h is n ot d e pr ess e d b y p h ot o gr a p h er or s o m e o n e els e w hil e p h ot o 
is  t a k e n.  ( T his  c o m m o nl y  h a p p e ns  if  p h ot o gr a p h er  h as  l o u p es).  Is  t h e  l e ns  w ell 
s u p p ort e d ? 
o  M a k e s ur e b att er y is f ull y c h ar g e d 

 
1 7  
§  If  first  p h ot os  i n  s eri es  ar e  acc e pt a bl e  a n d  t h e n  t h e y  gr a d u all y  b ec o m e  l ess  ex p os e d 
( d ar k er)  it  mi g ht  b e  b ec a us e  t h e  b att er y  is  gr a d u all y  l osi n g  p o w er  d uri n g  t h e  s essi o n. 
N ot e t h at t h e fl as h r e acti o n ti m e i ncr e as es as t h e b att er y p o w er d ecr e as es. 
§  O ut of f oc us 
o  F st o p m a y b e t o o l o w; m o v e m e nt artif act; fl as h d o es n’t w or k; l e ns is dirt y 
§  I m a g e n ot c e nt er e d 
o  M o v e m e nt  of  c hil d  or  c a m er a  ( h ol d  c hil d’s  h e a d  b et w e e n  k n e es  of  “ h el p er”; 
st a biliz e c a m er a wit h s ec o n d h a n d) 
§  R efl ecti o n artif act 
o  M o v e  c a m er a  sli g htl y  b et w e e n  first  a n d  s ec o n d  p h ot os  t o  ac hi e v e  diff er e nt 
a n gl e;  g e ntl y  d a b  c o nj u ncti v a  wit h  s w a b-  m ust  b e  d o n e  at  p eri p h er y  t o  a v oi d 
cr e ati n g i nfl a m m ati o n 
 
Gr a di n g of cli nic al c o nj u ncti v al p h ot o gr a p hs  
T hr e e ex p eri e nc e d o p ht h al m ol o gists will gr a d e t h e p h ot o gr a p hs at t h e c o ncl usi o n of t h e st u d y . 
Gr a d ers  will  gr a d e  e ac h  p h ot o gr a p h  i n  t h e  s et  f or  f ollicl es  a n d  i nfl a m m ati o n  acc or di n g  t o  t h e 
W H O si m plifi e d gr a di n g s yst e m.  
 
4. 3. 4 Cli nic al e v al u ati o n of t h e c o nj u ncti v a  
 
O n sit e cli nic al ex a mi n ati o n will b e p erf or m e d b y m e dic al st aff m e m b er pr e vi o usl y tr ai n e d. T h e 
ex a mi n er will us e t h e 2. 5 X m a g nif yi n g oc ul ar l o u p es ( Wils o n o p ht h al mics) t o ass ess t h e t ars al 
c o nj u ncti v a of t h e e v ert e d u p p er ri g ht e y eli d. T h e ex a mi n er will gr a d e t h e c o nj u ncti v a 
acc or di n g t o t h e W H O si m plifi e d Tr ac h o m a gr a di n g sc al e (s e e t a bl e b el o w). If n ec ess ar y, a 
h a n d- h el d t orc h li g ht will b e us e d b y t h e ex a mi n er f or ill u mi n ati o n of t h e c o nj u ncti v a.  T h e 
ex a mi n er t ells t h e Tr ac h o m a Gr a d e t o t h e t u b er, w h o t h e n r ec or ds t h e gr a d e o nt o t h e 
el ectr o nic d e vic e. Fi el d st aff will o nl y b e c ertifi e d t o gr a d e f or t h e st u d y if t h e y exc e e d s uffici e nt 
a gr e e m e nt wit h a n ex p ert tr ac h o m a gr a d er ( k a p p a ≥0. 6 o n a s eri es of 2 5 pict ur es) d uri n g t h e 
tr ai ni n g s essi o n. 
 
Defi niti o ns of t he W H O Si m plifie d Tr ac h o m a Gr a di n g Sc ale 
 
T F  ( Trac h o mat o us I nfla m mati o n – F ollic ular):  t he prese nce of fi ve or m ore f ollicles i n t he 
u p per tarsal c o nj u ncti va. 
TI  ( Trac h o mat o us I nfla m mati o n – I nte nse):  pr o n o u nce d i nfla m mat or y t hic ke ni n g of t he u p per 
tarsal c o nj u ncti va t hat o bsc ures m ore t ha n half of t he n or mal dee p tarsal vessels. 
T S  ( Trac h o mat o us Scarri n g):  t he prese nce of scarri n g i n t he tarsal c o nj u ncti va. 
T T  ( Trac h o mat o us Tric hiasis):  at least o ne e yelas h r u bs o n t he e ye ball. 
C O  ( C or neal O pacit y):  easil y visi ble c or neal o pacit y o ver t he p u pil. 
Cli nic ally active tr ac h o m a:  defi ne d as eit her T F or TI.  If W H O g ui deli nes rec o m me n d t h at T F 
al o ne is t he m ost a p pr o pri ate si g n t o f oll o w, t he n we will e asily be a ble t o re p ort t his.  
 
 
 

 
1 8  
5.  A N T H R O P O M E T R Y 
 
A nt hr o p o m etr y will b e c oll ect e d f or all c hil dr e n i n St u d y T w o at b as eli n e a n d 4 w e e ks aft er t h e 
c o m pl eti o n of t h e tr e at m e nt. W e will m e as ur e l e n gt h or h ei g ht, w ei g ht, a n d M U A C.  
 
M e as uri n g L e n gt h a n d H ei g ht 
A li g ht w ei g ht m e as uri n g b o ar d will b e us e d t o m e as ur e t h e p artici p a nt’s h ei g ht t o t h e n e ar est 
0. 1 c m. H ei g ht will b e ass ess e d wit h a S h orr B o ar d. D e p e n di n g o n a c hil d’s a g e a n d a bilit y t o 
st a n d, m e as ur e t h e c hil d’s l e n gt h or h ei g ht.   
 
L e n gt h: If a c hil d is l ess t h a n 2 y e ars ol d, or c a n n ot st a n d al o n e, m e as ur e r ec u m b e nt l e n gt h. A 
c hil d’s l e n gt h is m e as ur e d l yi n g d o w n (r ec u m b e nt).   
H ei g ht: If a c hil d is a bl e t o st a n d, w e will m e as ur e st a n di n g h ei g ht.  
 
Pr oc e d ur e  
If t h e c hil d h as br ai ds or h air or n a m e nts t h at will i nt erf er e wit h l e n gt h/ h ei g ht m e as ur e m e nts, 
r e m o v e t h e m if p ossi bl e.  C h ec k t h at a n y s a n d als, s h o es, or s oc ks h a v e als o b e e n r e m o v e d.  
W h et h er m e as uri n g l e n gt h or h ei g ht, t h e m ot h er/ g u ar di a n is n e e d e d t o h el p wit h 
m e as ur e m e nts a n d t o s o ot h e a n d c o mf ort t h e c hil d.  Ex pl ai n t o t h e m ot h er t h e r e as o ns f or t h e 
m e as ur e m e nts, a n d d escri b e t h e st e ps i n t h e pr oc e d ur e. A ns w er a n y q u esti o ns s h e mi g ht h a v e. 
S h o w h er a n d t ell h er h o w s h e c a n h el p y o u. Ex pl ai n t h at it is i m p ort a nt t o k e e p t h e c hil d still 
a n d c al m t o o bt ai n t h e b est m e as ur e m e nt.  
 
S h orr B o ar d S et -U p  
1)  R e m o v e S h orr B o ar d fr o m b a g. 
2)  St a n d t h e S h orr B o ar d u pri g ht.  Y o u c a n st e p o n t h e b as e of t h e b o ar d t o k e e p it st a bl e. 
3)  As y o u f ac e t h e b o ar d, t ur n t h e n o n-r e m o v a bl e b olt c o u nt ercl oc k wis e t o r el e as e t h e 
ext e nsi o n pi ec e.  N ot e: t h e b olt r e m ai ns att ac h e d t o t h e b ac k of t h e ext e nsi o n pi ec e – 
d o N O T r e m o v e it. 
4)  Sli d e t h e ext e nsi o n pi ec e i nt o t h e t o p e n d of t h e m ai n b o ar d a n d f ast e n t h e cl as p o n t h e 
b ac k of t h e b o ar d.  M a k e s ur e t h e cl as p is f ast e n e d pr o p erl y.   
5)  T h e a ut o-l oc k sli di n g h e a d/f o ot pi ec e is st or e d i n t h e b as e of t h e m ai n b o ar d a n d c a n b e 
m o v e d u p a n d d o w n t h e l e n gt h of t h e m e as uri n g b o ar d. It s h o ul d st a y i n pl ac e o n its 
o w n w h er e v er y o u p ositi o n it.   
6)  T h e m e as uri n g b o ar d m ust b e pl ac e d a g ai nst a fir m s urf ac e f or st a n di n g h ei g ht ( e. g. 
w all, t a bl e, tr e e, a g ai nst a v e hicl e, etc.). M a k e s ur e t h e b o ar d is st a bl e. If n ec ess ar y, 
pl ac e it e ms s uc h as s m all r oc ks u n d er n e at h t h e h ei g ht b o ar d t o st a biliz e it d uri n g t h e 
m e as ur e m e nt.  
7)  Cl e a n t h e e q ui p m e nt wit h alc o h ol s w a bs at t h e b e gi n ni n g of e ac h d a y.  
 
N ot e: w h e n y o u s et u p t h e b o ar d e ac h d a y, ex a mi n e e ac h of t h e pi ec es t o c h ec k f or d a m a g e. 
 
M e as ur e L e n gt h  
 

 
1 9  
C o v er t h e l e n gt h b o ar d wit h a c h uc k f or h y gi e n e a n d f or t h e b a b y’s c o mf ort. Ex pl ai n t o t h e 
m ot h er t h at s h e will n e e d t o pl ac e t h e b a b y o n t h e l e n gt h b o ar d a n d t h e n h el p t o h ol d t h e 
b a b y’s h e a d i n pl ac e w hil e t h e m e as ur e m e nt is t a k e n. S h o w h er w h er e t o st a n d w h e n pl aci n g 
t h e b a b y d o w n (i. e. o p p osit e y o u, o n t h e si d e of t h e l e n gt h b o ar d a w a y fr o m t h e t a p e).  Als o 
s h o w h er w h er e t o pl ac e t h e b a b y’s h e a d ( a g ai nst t h e fix e d h e a d b o ar d) s o t h at s h e c a n m o v e 
q uic kl y a n d s ur el y wit h o ut distr essi n g t h e b a b y. W h e n t h e m ot h er u n d erst a n ds y o ur 
i nstr ucti o ns a n d is r e a d y t o assist: As k h er t o l a y t h e c hil d o n his b ac k wit h his h e a d a g ai nst t h e 
fix e d h e a d b o ar d, c o m pr essi n g t h e h air.  
Q uic kl y p ositi o n t h e h e a d s o t h at a n i m a gi n ar y v ertic al li n e fr o m t h e e ar c a n al t o t h e l o w er 
b or d er of t h e e y e s oc k et is p er p e n dic ul ar t o t h e b o ar d.  ( T h e c hil d’s e y es s h o ul d b e l o o ki n g 
str ai g ht u p.) As k t h e m ot h er t o m o v e b e hi n d t h e h e a d b o ar d a n d h ol d t h e h e a d i n t his p ositi o n.   
S p e e d is i m p ort a nt. St a n di n g o n t h e si d e of t h e l e n gt h b o ar d w h er e y o u c a n s e e t h e m e as uri n g 
t a p e a n d m o v e t h e f o ot b o ar d: C h ec k t h at t h e c hil d li es str ai g ht al o n g t h e b o ar d a n d d o es n ot 
c h a n g e p ositi o n. S h o ul d ers s h o ul d t o uc h t h e b o ar d, a n d t h e s pi n e s h o ul d n ot b e arc h e d. As k t h e 
m ot h er t o i nf or m y o u if t h e c hil d arc h es t h e b ac k or m o v es o ut of p ositi o n. H ol d d o w n t h e 
c hil d’s l e gs wit h t h e o n e h a n d a n d m o v e t h e f o ot b o ar d wit h t h e ot h er. A p pl y g e ntl e pr ess ur e t o 
t h e k n e es t o str ai g ht e n t h e l e gs as f ar as t h e y c a n g o wit h o ut c a usi n g i nj ur y or distr ess.   
 
N ot e: it is n ot p ossi bl e t o str ai g ht e n t h e k n e es of n e w b or ns t o t h e s a m e d e gr e e as ol d er 
c hil dr e n. T h eir k n e es ar e fr a gil e a n d c o ul d b e e asil y i nj ur e d, s o a p pl y o nl y mi ni m u m pr ess ur e.   
 
If a c hil d is extr e m el y a git at e d a n d b ot h l e gs c a n n ot b e h el d i n p ositi o n, m e as ur e wit h o n e l e g i n 
p ositi o n. W hil e h ol di n g t h e k n e es, p ull t h e f o ot b o ar d a g ai nst t h e c hil d’s f e et. U p o n r e a di n g t h e 
m e as ur e m e nt, t h e ex a mi n er will cl e arl y c all o ut t h e n u m b er t o t h e r ec or d er. R ec or d t h e c hil d’s 
l e n gt h i n c e nti m et ers t o t h e l ast c o m pl et e d 0. 1 c m.  ( 1. 0 m m). K e e pi n g t h e c hil d i n pl ac e, 
r el e as e t h e sli di n g f o ot b o ar d, a n d pr e p ar e t o r e p e at t h e m e as ur e m e nt. R e- p ositi o n t h e c hil d f or 
a s ec o n d a n d t hir d m e as ur e m e nt.  
 
 
 
 
 

 
2 0  
M e as ur e St a n di n g H ei g ht 
 
E ns ur e t h at t h e h ei g ht b o ar d is o n l e v e l gr o u n d. W or ki n g wit h t h e m ot h er, a n d k n e eli n g i n 
or d er t o b e at t h e l e v el of t h e c hil d. H el p t h e c hil d st a n d o n t h e b as e b o ar d wit h t h e w ei g ht of 
t h e c hil d e v e nl y distri b ut e d o n b ot h f e et.  T h e h e els of t h e f e et ar e pl ac e d t o g et h er wit h b ot h 
h e els t o uc hi n g t h e b as e of t h e v ertic al b o ar d.  Pl ac e t h e f e et p oi nt e d sli g htl y o ut w ar d at a 6 0 
d e gr e e a n gl e. T h e b ac k of t h e h e a d, s h o ul d er bl a d es, b utt oc ks, c al v es, a n d h e els s h o ul d all 
t o uc h t h e v ertic al b o ar d. Ar ms s h o ul d h a n g fr e el y b y t h e si d es of t h e b o d y with t h e p al ms f aci n g 
t h e t hi g hs.  
 
N ot e: St a n di n g wit h all b o d y p arts t o uc hi n g t h e b o ar d m a y b e diffic ult f or s o m e c hil dr e n, i n 
w hic h c as e, h el p t h e c hil d t o st a n d o n t h e b o ar d wit h o n e or m or e c o nt act p oi nts t o uc hi n g t h e 
b o ar d. 
 
As k t h e m ot h er t o h ol d t h e c hil d’s k n e es a n d a n kl es t o h el p k e e p t h e l e gs str ai g ht a n d f e et fl at, 
wit h h e els a n d c al v es t o uc hi n g t h e v ertic al b o ar d. As k h er t o f oc us t h e c hil d’s att e nti o n, s o ot h e 
t h e c hil d as n e e d e d, a n d i nf or m y o u if t h e c hil d m o v es o ut of p ositi o n. P ositi o n t h e c hil d’s h e a d 
s o t h at a h oriz o nt al li n e fr o m t h e e ar c a n al t o t h e l o w er b or d er of t h e e y e s oc k et r u ns p ar all el 
t o t h e b as e b o ar d. As k t h e c hil d t o i n h al e d e e pl y a n d t o st a n d f ull y er ect wit h o ut alt eri n g t h e 
p ositi o n of t h e h e els.  If n ec ess ar y, p us h g e ntl y o n t h e b ell y t o h el p t h e c hil d st a n d t o f ull h ei g ht.  
Still k e e pi n g t h e h e a d i n p ositi o n, us e y o ur ot h er h a n d t o p ull d o w n t h e h e a d b o ar d t o r est 
fir ml y o n t o p of t h e h e a d a n d c o m pr ess t h e h air. U p o n r e a di n g t h e m e as ur e m e nt, t h e ex a mi n er 
will cl e arl y c all o ut t h e n u m b er t o t h e r ec or d er.  R ec or d t h e c hil d’s h ei g ht i n c e nti m et ers t o t h e 
l ast c o m pl et e d 0. 1 c m ( 1. 0 m m). K e e pi n g t h e c hil d i n pl ac e, r el e as e t h e sli di n g h e a d b o ar d, a n d 
pr e p ar e t o r e p e at t h e m e as ur e m e nt. R e- p ositi o n t h e c hil d f or a s ec o n d a n d t hir d m e as ur e m e nt.    

 
2 1  
 
 
 

 
2 2  
 
Dis m a ntli n g t h e S h orr B o ar d  
1)  St a n d t h e b o ar d u pri g ht: f ac e t h e b o ar d a n d st e p o n t h e b as e wit h o n e f o ot t o k e e p it 
st a bl e. 
2)  Sli d e t h e h e a d/f o ot pi ec e i nt o t h e b as e of t h e m ai n b o ar d. 
3)  R el e as e t h e cl as p o n t h e b ac k of t h e ext e nsi o n pi ec e a n d r e m o v e it.  P us h t h e cl as p FL A T 
a g ai nst t h e ext e nsi o n pi ec e. 
4)  T o att ac h t h e ext e nsi o n pi ec e t o t h e m ai n b o ar d, t ur n t h e fr o nt of t h e ext e nsi o n pi ec e 
i n w ar d a n d pl ac e it a g ai nst t h e fr o nt of t h e m ai n b o ar d.  M a k e s ur e t h at all si d es of t h e 
ext e nsi o n pi ec e ar e str ai g ht a n d i n li n e wit h t h e m ai n b o ar d. 
5)  P us h o n t h e b olt t h at is o n t h e b ac k of t h e ext e nsi o n pi ec e a n d scr e w it i nt o t h e m ai n 
b o ar d.   
6)  P ut t h e b o ar d b ac k i nsi d e of t h e c arr yi n g c as e f or st or a g e u ntil y o ur n ext us e.   
 
M e as uri n g W ei g ht 
 
T h e S E C A 8 7 4 sc al e will b e us e d t o w ei g h i nf a nts a n d c hil dr e n t o t h e n e ar est 0. 0 1 k g. I nf a nts 
a n d y o u n g c hil dr e n c a n als o b e w ei g h e d si m ult a n e o usl y wit h t h eir p ar e nt or g u ar di a n b y t h e 
u ni q u e “ m ot h er- b a b y” f u ncti o n ( p ar e nt or g u ar di a n is w ei g h e d a n d t h e n t h e i nf a nt or c h il d is 
w ei g h e d w hil e h el d b y t h e p ar e nt). Ex pl ai n t o t h e m ot h er t h at w e w a nt t o w ei g h h er c hil d t o 
s e e h o w h e or s h e is gr o wi n g.  If s h e h as a b a b y or a c hil d w h o is u n a bl e t o st a n d, s h e will h ol d 
t h e c hil d o n t h e sc al e.  If t h e c hil d is 2 y e ars or ol d er/c a n st a n d al o n e, t he c hil d will b e w ei g h e d 
al o n e. C hil dr e n s h o ul d b e w e ari n g o nl y li g ht cl ot hi n g, n o s h o es or s a n d als, n o h air or n a m e nts, 
a n d n o j e w elr y. Ex pl ai n t h at t h e c hil d n e e ds t o r e m o v e o ut er cl ot hi n g a n d s h o es/s a n d als i n 
or d er t o o bt ai n a n acc ur at e w ei g ht.  If t h e b a b y is w e ari n g a di a p er, t h e di a p er s h o ul d b e 
r e m o v e d.  If a n y h e a v y cl ot h es r e m ai n o n t h e c hil d, m a k e a n ot e i n t h e N ot es s ecti o n. 
 
S ec a 8 7 4 Sc al e  
1)  R e m o v e sc al e fr o m b a g. 
2)  B e s ur e t h at t h e sc al e is pl ac e d o n a fl at, h ar d, e v e n s urf ac e. All 4 l e gs of t h e sc al e s h o ul d 
m a k e c o nt act wit h t h e gr o u n d s urf ac e, wit h o ut w o b bli n g. It m a y b e h el pf ul t o pl ac e a 
pi ec e of pl y w o o d o n t h e gr o u n d u n d er n e at h t h e sc al e.  
3)  W h e n b att  a p p e ars i n t h e dis pl a y, y o u s h o ul d c h a n g e t h e b att eri es.  R e m o v e t h e ol d 
b att eri es a n d i ns ert 6 n e w b att eri es.   
 
T ur n t h e p o w er o n t h e sc al e w h e n y o u ar e r e a d y t o b e gi n w ei g hi n g. If t h e c hil d is u n a bl e t o 
st a n d o n t h e sc al e, y o u will us e t h e 2 i n 1 w ei g hi n g f u ncti o n (c all e d t ar e d w ei g hi n g). T h e 2 i n 1 
f u ncti o n e n a bl es t h e w ei g ht of b a bi es a n d s m all c hil dr e n t o b e d et er mi n e d w hil e a n a d ult h ol ds 
t h e m. I d e ntif y a s uit a bl e ar e a t h at is fl at f or h oriz o nt al pl ac e m e nt of t h e sc al e. Pr ess t h e st art 
k e y wit h n o l o a d o n t h e sc al e. T h e sc al e is r e a d y f or us e w h e n it s ets t o 0. 0 0. If n ec ess ar y, 
s witc h t h e w ei g ht dis pl a y t o K G: h ol d d o w n t h e 2 i n 1 k e y f or a b o ut 3 s ec o n ds. Pr ess t h e st art 
k e y wit h n o l o a d o n t h e sc al e. W ait u ntil t h e dis pl a y s h o ws 0. 0 0. 

 
2 3  
As k t h e a d ult t o r e m o v e his/ h er s h o es a n d st a n d i n t h e mi d dl e of t h e sc al e wit h o ut t h e c hil d. 
S/ h e s h o ul d r e m o v e a n y l o n g g ar m e nts, as t h es e c a n c o v er t h e dis pl a y a n d als o l e a d t o v ari a bl e 
m e as ur e m e nts. Aft er t h e a d ult’s w ei g ht a p p e ars o n t h e dis pl a y, t ell hi m/ h er t o r e m ai n st a n di n g 
o n t h e sc al e. Pr ess t h e 2 i n 1 k e y t o acti v at e t h e f u ncti o n. T h e sc al e st or es t h e w ei g ht of t h e 
a d ult a n d t h e dis pl a y r et ur ns t o z er o. W h e n 0. 0 0 a n d N E T a p p e ar i n t h e dis pl a y, h a n d t h e c hil d 
t o t h e a d ult.  T h e sc al e will d et er mi n e t h e w ei g ht of t h e c hil d.  O nc e t h e v al u e is st a bl e f or 
a b o ut 3 s ec o n ds, t h e w ei g ht is m e as ur e d.   
 
N ot e: If a n a d ult is v er y h e a v y ( e. g. m or e t h a n 1 0 0 k g) a n d t h e b a b y’s w ei g ht is r el ati v el y l o w 
( e. g. l ess t h a n 2. 5 k g), t h e b a b y’s w ei g ht m a y n ot r e gist er o n t h e sc al e.  I n s uc h c as es, h a v e a 
li g ht er p ers o n h ol d t h e b a b y o n t h e sc al e. 
 
T h e p ositi o n er will cl e arl y c all o ut t h e c hil d’s w ei g ht t o t h e r ec or d er. R ec or d t h e c hil d’s w ei g ht 
t o t h e n e ar est 0. 0 1 k g. R e p e at t h e m e as ur e m e nt 2 m or e ti m es. N ot e t h at o nl y t h e b a b y n e e ds 
t o r e m o v e d fr o m t h e sc al e; t h e a d ult s h o ul d r e m ai n o n t h e sc al e t h e e ntir e ti m e. T o t ur n off t h e 
2 i n 1 f u ncti o n, pr ess t h e 2 i n 1 k e y. T h e 2 i n 1 f u ncti o n r e m ai ns o n u ntil y o u pr ess t h e 2 i n 1 k e y 
a g ai n, or u ntil t h e sc al e s witc h es off a ut o m atic all y. If s e v er al c hil dr e n ar e t o b e w ei g h e d 
c o ns ec uti v el y wit h t h e s a m e a d ult h ol di n g t h e b a bi es, it is i m p ort a nt t h at t his p ers o n’s w ei g ht 
d o es n ot c h a n g e ( e. g. d u e t o a pi ec e of cl ot hi n g b ei n g r e m o v e d/ a d d e d). If t h e c hil d is a bl e t o 
st a n d o n t h e sc al e, y o u will w ei g h t h e c hil d al o n e. T al k wit h t h e c hil d a b o ut t h e n e e d t o st a n d 
still.  C o m m u nic at e wit h t h e c hil d i n a s e nsiti v e, n o n-fri g ht e ni n g w a y. Pr ess t h e st art k e y wit h n o 
l o a d o n t h e sc al e.  T h e sc al e is r e a d y f or us e w h e n it s ets t o 0. 0 0. As k t h e c hil d t o st a n d i n t h e 
mi d dl e of t h e sc al e. O nc e o n t h e sc al e, t h e c hil d m ust st a n d still. T h e H OL D f u ncti o n is 
a ut o m atic all y acti v at e d f or w ei g hts o v er 1. 5 k g/ 3. 3 l bs. T h e dis pl a y fl as h es u ntil a st a bl e w ei g ht 
h as b e e n m e as ur e d. T h e dis pl a y is t h e n fr oz e n u ntil t h e n ext w ei g hi n g o p er ati o n.   
 
N ot e: If t h e c hil d j u m ps o n t h e sc al e or w o n’t st a n d still, y o u will n e e d t o us e t h e t ar e d w ei g hi n g 
pr oc e d ur e i nst e a d.  
 
T h e p ositi o n er will cl e arl y c all o ut t h e c hil d’s w ei g ht t o t h e r ec or d er. R ec or d t h e c hil d’s w ei g ht 
t o t h e n e ar est 0. 0 1 k g. R e p e at t h e m e as ur e m e nt 2 m or e ti m es. H a v e t h e c hil d m o v e c o m pl et el y 
off t h e sc al e a n d t h e n st a n d a g ai n o n t h e sc al e. If n o f urt h er w ei g hi n g o p er ati o ns ar e 
p erf or m e d, t h e sc al e s witc h es off a ut o m atic all y aft er 2- 3 mi n ut es. 
 
Sc al e C ali br ati o n  
I n or d er t o m o nit or t h e c ali br ati o n of t h e sc al es o v er ti m e, e ac h t e a m will w ei g h a 5 k g t est 
w ei g ht at t h e b e gi n ni n g a n d e n d of t h e d a y. T his m e as ur e m e nt will t a k e pl ac e at t h e sit e of t h e 
sc al e st or a g e s o t h at t h e w ei g hts d o n ot n e e d t o b e c arri e d t o t h e fi el d. I n s o m e c as es, t h es e 
w ei g hts will b e k e pt i n a pr oj ect v e hicl e.  
 
 
 

 
2 4  
M e as uri n g Mi d- U p p er Ar m Circ u mf er e nc e ( M U A C) 
 
T h e c hil d’s M U A C will b e m e as ur e d o n e ti m e. M U A C m e as ur e m e nts will b e t a k e n at t h e 
mi d p oi nt of t h e l eft ar m b et w e e n t h e ti p of t h e s h o ul d er a n d t h e ti p of t h e el b o w usi n g n o n-
str etc h M U A C t a p es.  
 
First, fi n d t h e a p pr oxi m at e mi d p oi nt of t h e u p p er ar m. H a v e t h e c hil d st a n d 
u p str ai g ht wit h f e et t o g et h er, a n d t h e ri g ht ar m b e nt 9 0 d e gr e es at t h e 
el b o w, p al m f aci n g u p.  T h e ex a mi n er is p ositi o n e d b e hi n d t h e c hil d. If it is 
h el pf ul, m ar k t h e mi d p oi nt wit h a p er m a n e nt m ar k er. T o m e as ur e mi d- u p p er 
ar m circ u mf er e nc e, h a v e t h e c hil d st a n d u p str ai g ht wit h t h e ar ms r el ax e d at 
t h e si d es.  T h e ex a mi n er will st a n d f aci n g t h e c hil d’s ri g ht si d e.  T h e m e as uri n g t a p e is pl ac e d 
ar o u n d t h e u p p er ar m at t h e m ar k e d p oi nt. Wr a p t h e t a p e ar o u n d t h e ar m, p ulli n g it t o li e fl at 
a g ai nst t h e s urf ac e of t h e s ki n.  B e c ar ef ul n ot t o p ull t h e t a p e t o o ti g htl y (t o c o m pr ess t h e s ki n).   
 
T h e li n e f or r e a di n g m e as ur e m e nts is cl e arl y l a b el e d o n t h e M U A C 
t a p e ( R E A D (c m)). R e a d t h e n u m b er ali g n e d wit h t h e m e as ur e m e nt 
li n e o n t h e t a p e U p o n m e as uri n g, t h e ex a mi n er will cl e arl y c all o ut 
t h e n u m b er t o t h e r ec or d er.  R ec or d t o t h e n e ar est 0. 1 c m ( 0. 1 c m = 
1 m m). K e e pi n g t h e c hil d i n pl ac e, r el e as e t h e M U A C stri p. 
 
If t h e M U A C is < 1 1. 5c m r ef er t h e c hil d t o t h e n e ar est h e alt h c e nt er.  
 
 
6.  C O G NI TI V E D E V EL O P M E N T A S S E S S M E N T  
 
T h e D e v el o p m e nt al Mil est o n es C h ec klist ( D M C-III) will b e us e d t o e v al u at e t h e d e v el o p m e nt of 
m ot or a n d l a n g u a g e s kills i n 2 4 0 c hil dr e n at b as eli n e i n t his st u d y. T h e D M C-III i ncl u d es t hr e e 
s u bsc al es: gr oss m ot or, fi n e m ot or, a n d l a n g u a g e. T h e ori gi n al v ersi o n of t h e D M C-III w as 
d esi g n e d f or c hil dr e n a g e d 3 t o 2 4 m o nt hs. T h e u p d at e d D M C-III i ncl u d es a d diti o n al it e ms t o 
ass ess c hil dr e n a g e d 2 t o 8 y e ars. T h e D M C-III r e q uir es p ar e nt i nt er vi e w f or all it e ms a n d 
i ncl u d es o pti o n al dir ect o bs er v ati o n of t h e c hil d f or all it e ms. F or t his st u d y, d at a c oll ect ors will 
i ncl u d e b ot h p ar e nt i nt er vi e w a n d dir ect o bs er v ati o n f or all it e ms w h er e p ossi bl e. 
 
M at eri als f or o bs er v ati o n it e ms: 
•  P e ns  
•  W hit e p a p er 
•  A b all (c hil d-siz e f o ot b all/s occ er b all) 
•  A s m all t o y 
•  S m all gr ai ns i n a b ox 
•  A p ull -t o y 
•  A c ol or s h e et pri nt e d wit h b ox es of t h e f oll o wi n g c ol ors : bl u e, gr e e n, y ell o w, br o w n, 
p ur pl e, w hit e, gr e y, or a n g e, r e d, bl ac k, a n d pi n k 

 
2 5  
•  5 c oi ns   
 
Pr e p ari n g t h e ass ess m e nt s essi o n f or o bs er v ati o n it e ms:  
2. 1  B ef or e t h e ass ess m e nt s essi o n 
a)  C h ec k  t h at y o u h a v e all of t h e e q ui p m e nt  a n d r ec or d f or ms r e q uir e d usi n g t h e 
c h ec klist.  
 
2. 2  W h e n y o u arri v e at t h e c hil d’s h o us e 
a)  Gr e et  t h e m ot h er or c ar e gi v er acc o m p a n yi n g t h e c hil d. 
b)  I ntr o d uc e y o urs elf a n d o utli n e t h e p ur p os e of t h e visit  
c) A p pr o ac h  t h e c hil d f or t h e first ti m e i n t h e pr es e nc e of t h e m ot h er / c ar e gi v er  
d)  O bs er v e c ar ef ull y t h e c hil d’s r e acti o n t o y o urs elf a n d t h e sit u ati o n as it u nf ol ds.  
e)  C o nfir m  t h at t h e c hil d is i n g o o d h e alt h . D O N O T A S S E S S A C HIL D W H O I S 
U N W ELL . If t h e c hil d is sic k, c o n d uct t h e i nt er vi e w p orti o n of t h e visit a n d m a k e 
a n a p p oi nt m e nt t o r et ur n at a l at er d at e t o c o n d uct t h e o bs er v ati o n it e ms. 
 
2. 3  G ui d eli n es f or t h e t esti n g l oc ati o n 
a)  T h e t esti n g l oc ati o n s h o ul d b e a pl ac e w h er e t h er e is s h a d e, fl at gr o u n d, 
m axi m u m 
   
pri v ac y, a n d s p ac e f or m o v e m e nt. 
b)  T a k e ti m e t o s el ect t h e l ar g est, q ui et est l oc ati o n t h at all o ws y o u t o r u n ar o u n d. 
Y o u c a n sit 
i n o n e pl ac e a n d r u n ar o u n d els e w h er e.  
c) R e m o v e p ossi bl e  distr act ers  b ef or e st arti n g t o ass ess. 
d)  If p ossi bl e, p ositi o n t h e c hil d f aci n g a w all t o mi ni miz e distr acti o ns. 
e)  M a k e s ur e t h e c hil d a n d m ot h er ar e s ettl e d a n d c o mf ort a bl e. 
f) T a k e g o o d c ar e t o e ns ur e t h e s af et y of t h e c hil dr e n. L o o k o ut f or p ot e nti al 
h az ar ds. 
g)  If ot h ers g at h er t o w atc h, as k t h e m p olit el y t o l e a v e. Ex pl ai n t h at t h e c hil d mi g ht 
b e s h y if  
h e or s h e is b ei n g w atc h e d b y ot h ers. If n ec ess ar y, as k f or t h e m ot h er’s h el p t o 
dis p ers e t h e cr o w d. 
 
2. 4 S etti n g u p t h e e q ui p m e nt.  
a)  Arr a n g e  e q ui p m e nt w h er e it will b e e asil y acc essi bl e t o y o u.  
b)  E x pl ai n  t o t h e f a mil y m e m b er w h o acc o m p a ni es t h e c hil d t h eir r ol e. T o  
•  pr o vi d e c o mf ort t o t h eir c hil d b y t h eir pr es e nc e  
•  e nc o ur a g e t h e c hil d t o p artici p at e 
•  (if n ec ess ar y) t o h el p c o m m u nic at e t h e i nstr ucti o ns t o t h e c hil d  H el p 
t h e m t o u n d erst a n d t h e g a m e, b ut N O T t o assist t h e c hil d t o c o m pl et e 
t as ks.  
 
2. 5 S ettli n g t h e c hil d 
a)  Us e t h e n a m e t h e c hil d is c all e d b y t hr o u g h o ut t h e s essi o n. W h e n e v er y o u n e e d 

 
2 6  
t o g et t h e 
c hil d’s att e nti o n t o as k a q u esti o n or gi v e a n i nstr ucti o n, s a y his or h er n a m e.  
b)  R e - i ntr o d uc e y o urs elf t o t h e c hil d  
c) I n vit e t h e c hil d t o p artici p at e i n a n acti vit y. 
d)  E n g a g e  t h e c hil d wit h s o m e of t h e e q ui p m e nt/ acti viti es. T his m a y i ncl u d e a w ar m 
u p  
acti viti es, s e p ar at e fr o m t h e m ai n ass ess m e nt.  
e)  I n g e n er al, c hil dr e n r es p o n d w ell w h e n w e s mil e, ar e e nt h usi astic, pl a yf ul, a n d 
aff ecti o n at e. 
 
2. 6  G e n er al i nstr ucti o ns f or a d mi nist eri n g fi n e a n d gr oss m ot or it e ms: 
a)  First , d e m o nstr at e t h e t as k. 
b)  S ec o n d , h el p t h e c hil d t o d o t h e t as k. Gi v e s u p p ort w h er e n ec ess ar y u ntil y o u ar e 
c ert ai n t h at t h e c hil d c a n c o m pl et e t h e t as k u ns u p p ort e d. R e m o v e a n y s u p p ort 
pr o vi d e d i n st a g es.   
c) T hir d , t ell t h e c hil d t o d o t h e t as k al o n e, wit h o ut assist a nc e. 
 
2. 7  D uri n g ass ess m e nt. 
a.  C arr y o ut all acti viti es o n fl at a n d e v e n gr o u n d. 
b.  C hil d s h o ul d b e i n a cl e ar s p ac e a w a y fr o m f ur nit ur e / w alls. 
c. T h e i ncl usi o n of m ot h er a n d si bli n gs i n t h es e acti viti es oft e n ai ds a n d e nc o ur a g es t h e 
t ar g et c hil d t o f e el l ess i n hi bit e d. 
d.  Gi v e  pr ecis e a n d cl e ar i nstr ucti o ns t o t h e c hil d, f oll o wi n g m a n u al g ui d eli n es. 
e.  M o v e  fr o m o n e t as k t o t h e ot h er i n a s yst e m atic w a y.  
f. P a y att e nti o n  t o t h e c hil d at all ti m es.  
g.  Pr ais e  t h e c hil d f or eff ort, N O T s ucc ess i n a n it e m. 
h.  R et ur n ass ess m e nt m at eri al t o t h e b ox o n c o m pl eti o n of t h e it e m. T a k e o ut t h e 
m at eri al f or o n e it e m at a ti m e a n d r e pl ac e it w h e n y o u h a v e fi nis h e d. S o m eti m es y o u 
m a y t a k e o ut t h e m at eri al f or t h e n ext it e m b ef or e r e pl aci n g t h e m at eri al f or t h e 
pr e vi o us it e m, if t his is n ec ess ar y t o k e e p t h e c hil d’s att e nti o n. H o w e v er, i n t his c as e, 
r e pl ac e t h e m at eri al f or t h e pr e vi o us it e m i m m e di at el y aft er e n g a gi n g t h e c hil d’s 
att e nti o n wit h t h e n e w it e m. 
i. T a k e a br e a k  if t h e c hil d g ets tir e d or b ec o m es distr ess e d. I n g e n er al, it is a g o o d i d e a 
t o t a k e a br e a k at l e ast e v er y 3 0 mi n ut es. 
j. If n ec ess ar y, r e mi n d t h e acc o m p a n yi n g f a mil y m e m b er of t h eir r ol e i n t h e 
ass ess m e nt.  
 
D o N O T:  
1)  Cr o w d t h e ass ess m e nt ar e a wit h us e d it e ms, t his will distr act t h e c hild.  
2)  H urr y  t h e c hil d. 
 
2. 8  Str at e gi es f or h a n dli n g c hil dr e n w h o r ef us e 
a)  H a v e t h e m ot h er d e m o nstr at e t h e t as k a n d e nc o ur a g e t h e c hil d t o d o it 

 
2 7  
b)  I n v ol v e ot h er c hil dr e n w h o ar e ar o u n d, p er h a ps a n ol d er si bli n g. H a v e t h e ot h er c hil d d o 
it a n d s e e if t h e c hil d will d o t h e t as k al o n g wit h a n ot h er c hil d. 
c) Gi v e t h e c hil d a dri n k a n d a br e a k, l et t h e c hil d r est or pl a y f or a f e w mi n ut es 
d)  Pr o mis e t o gi v e t h e c hil d a s w e et if h e/s h e d o es t h e t as k 
 
2. 9  E n di n g t h e ass ess m e nt 
a)  F or m all y  e n d t h e s essi o n b y t h a n ki n g e v er y o n e. 
b)  B ef or e y o u l e a v e t h e h o us e  
i. R e a d  t hr o u g h y o ur f or m c ar ef ull y.  
ii. C o nfir m e ac h s ecti o n/ c h ec klist is fill e d f ull y.  
c) C h ec k all e q ui p m e nt is p ac k e d pr o p erl y.  ( Us e c h ec klist t o h el p y o u.) 
 
2. 1 0  
B ef or e t h e n ext s essi o n  
CL E A N A N D DI SI N F E C T T H E E Q UI P M E N T E V E R Y D A Y.  
        
A d mi nistr ati o n pr oc e d ur es  
I n e ac h s u bsc al e, t h e it e ms ar e arr a n g e d i n or d er fr o m t h e e arli er ac q uir e d s kill (f or ex a m pl e, 
sits s u p p ort e d) t o t h e l at er ac q uir e d s kill (f or ex a m pl e, sits al o n e o n t h e fl o or). T h e arr o ws i n 
t h e l eft- h a d m ar gi n s h o w t h e st art p oi nts f or t h e c hil d’s a g e. St art at t h e it e m a p pr o pri at e f or 
t h e c hil d’s a g e. If t h e c hil d sc or es 0, g o t o t h e pr e vi o us st art p oi nt. If t h e c hil d sc or es 1 or 2, 
c o nti n u e t o t h e n ext it e m. St o p aft er 4 c o ns ec uti v e sc or es of 0. R ef er t o t h e D M C-III M a n u al f or 
a d et ail e d d escri pti o n of h o w t o a d mi nist er e ac h it e m, i ncl u di n g pr o bi n g q u esti o ns t o us e if t h e 
i nt er vi e w e e h as diffic ult y u n d erst a n di n g t h e q u esti o n. T h e D M C-III fli pc h art i ncl u d es pict ur es, 
w hic h s h o ul d b e us e d i n c o nj u ncti o n wit h t h e q u esti o ns f or t h e t w o m ot or s u bsc al es. 
 
7.  MI C R O BI OL O GI C AL M E T H O D S    
 
P artici p a nt n a m es will n ot b e i ncl u d e d i n l a b or at or y r ec or ds a n d s a m pl es will b e d e i d e ntifi e d.  
T h er ef or e, l a b or at or y p ers o n n el will b e m as k e d, pr e v e nti n g i d e ntific ati o n of t h e i n di vi d u als 
i nf ect e d.  L a b r es ults will n ot b e a v ail a bl e f or w e e ks, if n ot m o nt hs. T h us, all i n di vi d u als will b e 
tr e at e d acc or di n g t o t h eir st u d y ar m, w h et h er or n ot t h eir l a b t ests r e v e al e vi d e nc e of i nf ecti o n. 
 
Pr e p ar ati o n of Z y m o D N A/ R N A S hi el d T M 
 
M a n uf act ur er’s Pr o d uct D escri pti o n 
D N A/ R N A S hi el d T M e ns ur es n ucl eic aci d st a bilit y d uri n g s a m pl e st or a g e/tr a ns p ort at a m bi e nt 
t e m p er at ur es, pr es er vi n g t h e g e n etic i nt e grit y a n d ex pr essi o n pr ofil es of s a m pl es. T h er e is n o 
n e e d f o r r efri g er ati o n or s p eci aliz e d e q ui p m e nt. D N A/ R N A S hi el dT M eff ecti v el y l ys es c ells a n d 
i n acti v at es n ucl e as es a n d i nf ecti o us a g e nts ( vir us), a n d it is c o m p ati bl e wit h v ari o us c oll ecti o n 
a n d st or a g e d e vic es ( v ac u u m t u b es, s w a bs, n as al, b ucc al, f ec al, etc.). 
 
M at eri als R e q uir e d  

 
2 8  
 
Pr o d uct  
S u p pli er  
C at al o g N o.  
D N A/ R N A S hi el d T M 
1 X s ol uti o n  
Z y m o R es e arc h C or p.  
R 1 1 0 0 -5 0 ( 5 0 ml)  
R 1 1 0 0 -2 5 0 ( 2 5 0 ml)  
2 ml L o o p Scr e w C a p Micr o T u b e, P C R P erf or m a nc e 
T est e d  
( D N A-, D N as e-, R N as e-, & P C R i n hi bit or-fr e e) 
S arst e dt I nc.  
7 2. 6 9 4. 4 1 6  
I ns ert f or L o o p Scr e w C a p, Vi ol et 
S arst e dt I nc.  
6 5. 7 1 3. 0 0 7  
 
Pr oc e d ur e  
1.  Pr e p ar e t u b es b y a d di n g vi ol et c a p i ns erts t o t h e t o p of e ac h t u b e (t h es e ar e f or c ol or 
c o di n g p ur p os es t o h el p disti n g uis h t h e D N A/ R N A S hi el d s ol uti o n fr o m ot h er m e di a 
b ei n g us e d i n t h e fi el d). 
2.  B ef or e ali q u oti n g t h e D N A/ R N A S hi el d, g e ntl y mix t h e s ol uti o n b y eit h er s wirli n g it or 
i n v erti n g t h e b ottl e a f e w ti m es (tr yi n g n ot t o cr e at e t o o m a n y b u b bl es) t o e ns ur e t h at 
t h e s ol uti o n is h o m o g e n o us. I n a cl e a n bi os af et y c a bi n et, us e st eril e t ec h ni q u e t o ali q u ot 
1 ml of t h e s ol uti o n i nt o e ac h 2 ml t u b e. 
3.  W h e n y o u h a v e fi nis h e d ali q u oti n g, m a k e s ur e t h at t h e c a p o n e ac h t u b e h as b e e n 
ti g ht e n e d t o pr e v e nt t u b es fr o m l e a ki n g d uri n g tr a ns p ort. 
 
St or a g e a n d S h elf Lif e  
St or e pr e p ar e d t u b es at r o o m t e m p er at ur e ( 2 0° C- 2 5° C) u ntil us e; d o n ot fr e ez e s ol uti o n pri or t o 
c oll ecti n g s a m pl es. Z y m o g u ar a nt e es t h e s h elf-lif e of D N A/ R N A S hi el d T M  u p t o 1 y e ar fr o m t h e 
d at e of r ec ei pt. T h er ef or e, if t h e s ol uti o n is ali q u ot e d ri g ht a w a y, t h e n pr e p ar e d t u b es s h o ul d 
b e g o o d f or 1 y e ar. T h e s h elf-lif e m a y b e l o n g er, d e p e n di n g o n st or a g e c o n diti o ns. 
 
M et h o ds f or R esist a nc e T esti n g 
 
Is ol ati o n & I d e ntific ati o n of Or g a nis ms Usi n g Micr o bi ol o gic al T ec h ni q u es 
 
S. p n e u m o ni a e will b e is ol at e d fr o m n as o p h ar y n g e al s w a bs usi n g bl o o d a n d str e p s el ect pl at e, 
c h oc ol at e, a n d c h oc ol at e wit h b acitr aci n m e di a. W e will i d e ntif y or g a nis ms usi n g gr a m st ai n. 
St a n d ar d micr o bi ol o gic al h a n d b o o ks s h o ul d b e r ef er e nc e d f or m or e d et ail e d i nf or m ati o n.  
 
L a b or at or y R es ults R e p orti n g 
 
All l a b r es ults will b e k e pt i n c o m p ut er fil es as w ell as i n h ar d-c o p y f or m b y t h e Micr o bi ol o gist. 
 
D escri pti o n of I d e ntific ati o n T ests:  

 
2 9  
W e will f oll o w t h e st a n d ar d q u alit y c o ntr ol m et h o ds alr e a d y i n pl ac e at t h e l a b or at or y. F or 
ex a m pl e, l a b st aff will p erf or m p ositi v e a n d n e g ati v e gr o wt h c o ntr ols f or all m e di a, a n d p ositi v e 
c o ntr ols f or all st ai ns a n d i d e ntific ati o n t ests at a pr e- d et er mi n e d sc h e d ul e. 
 
C at al as e T est:  
A f e w dr o ps of 3 % h y dr o g e n p er oxi d e ar e a d d e d t o c ol o ni es r e m o v e d fr o m a g ar a n d s m e ar e d 
o nt o a cl e a n gl ass sli d e, a n d i m m e di at el y o bs er v e d f or t h e r el e as e of ox y g e n or “ b u b bli n g.” 
B u b bli n g is i nt er pr et e d as t h e pr es e nc e of c at al as e. L ac k of b u b bli n g is a n e g ati v e t est. 
 
T u b e C o a g ul a s e: 
C o a g ul as e pl as m a is i n oc ul at e d wit h a n is ol at e d st a p h yl oc occ al c ol o n y a n d i nc u b at e d f or u p t o 
2 4 h o urs at 3 7° C. A n y d e gr e e of cl ot f or m ati o n is a p ositi v e r e acti o n a n d n o cl ot is a n e g ati v e 
r e acti o n. 
 
L at e x A g gl uti n ati o n: 
A f e w st a p h yl oc occ al c ol o ni es ar e mix e d wit h t h e l at ex a g gl uti n ati o n r e a g e nt a n d o bs er v e d f or 
a g gl uti n ati o n. A p ositi v e r e acti o n is a g gl uti n ati o n of t h e l at ex p articl es, w hil e n o a g gl uti n ati o n is 
a n e g ati v e r e acti o n. 
 
O pt oc hi n T est:  
A n o pt oc hi n disc is pl ac e d o nt o a bl o o d a g ar s u bc ult ur e d pl at e t h at h as b e e n i n oc ul at e d wit h a 
p ur e c ult ur e of t h e t est or g a nis m. Pl at es ar e i nc u b at e d f or 2 4 h o urs at 3 5° C i n C O 2. If a z o n e of 
i n hi biti o n of 1 4 m m or m or e is f or m e d, t h e n it’s S. p n e u m o ni a e . 
 
D es o x yc h ol at e ( 1 0 %) T est: 
A dr o p of r e a g e nt is a d d e d t o a w ell-is ol at e d c ol o n y o n a bl o o d a g ar pl at e. C ol o ni es of S. 
p n e u m o ni a e  will disi nt e gr at e aft er 3 0 mi n ut es. 
 
S at elliti n g Gr o wt h o n Bl o o d A g ar: 
C ol o ni es r es e m bli n g H a e m o p hil us ar e s u bc ult ur e d t o a n o n-s el ecti v e bl o o d a g ar pl at e a n d cr oss 
str e a k e d wit h C o a g ul as e n e g ati v e St a p h yl oc occ us. Pl at es ar e i nc u b at e d i n C O 2 o v er ni g ht a n d 
ex a mi n e d f or s at ellit e gr o wt h ar o u n d St a p h yl oc occ us . 
 
Micr o bi ol o gic al R esist a nc e T esti n g f or S w a bs 
R esist a nc e t esti n g will b e p erf or m e d usi n g t h e Kir b y B a u er disc diff usi o n ass a y or E-t est stri ps 
(t o d et er mi n e MI C’s). 
 
Q u alit y C o ntr ol 
E. c oli  A T C C 2 5 9 2 2, S. p n e u m o ni a e  A T C C 4 9 6 1 9, H. i nfl u e nz a e  A T C C 4 9 2 4 7, S. a ur e us  A T C C 
2 5 9 2 3 a n d 2 9 2 1 3 ar e us e d f or q u alit y c o ntr ol t esti n g of a nti bi otic discs a n d E-t est stri ps. 
 
Kir b y B a u er Ass a y 
A  s us p e nsi o n of t h e is ol at e is pr e p ar e d a n d t ur bi dit y a dj ust e d u ntil it r e ac h es t h e 0. 5 Mc F arl a n d 
st a n d ar d. T h e s us p e nsi o n is us e d t o i n oc ul at e t h e a p pr o pri at e s usc e pti bilit y a g ar pl at e f or 

 
3 0  
c o nfl u e nt gr o wt h. T h e pl at e is all o w e d t o dr y. A nti micr o bi al dis ks ar e pl ac e d o n t h e a g ar pl at es 
wit h st eril e f orc e ps or t w e ez ers; dis ks m a y n ot b e m o v e d o nc e t h e y h a v e t o uc h e d t h e pl at es. 
Aft er t h e dis ks ar e pl ac e d o n t h e pl at e, t h e pl at es ar e i n v ert e d a n d i nc u b at e d at 3 5° C f or 1 6 t o 
1 8 h o urs. Aft er i nc u b ati o n, t h e di a m et er of t h e z o n es of c o m pl et e i n hi biti o n (i ncl u di n g t h e 
di a m et er of t h e dis k) ( Fi g ur e 2) ar e m e as ur e d a n d r ec or d e d i n milli m et ers. T h e m e as ur e m e nts 
c a n b e m a d e wit h a r ul er or sli di n g c ali p er o n t h e u n d ers urf ac e of t h e pl at e wit h o ut o p e ni n g t h e 
li d. T h e z o n es of gr o wt h i n hi biti o n will b e c o m p ar e d wit h t h e z o n e-siz e i nt er pr et ati v e t a bl e a n d 
r ec or d e d as s usc e pti bl e, i nt er m e di at e, or r esist a nt t o e ac h dr u g t est e d acc or di n g t o CL SI 
g ui d eli n es (s e e A p p e n dix).  
 
 
Fi g ur e 2.  R es ults of t h e dis k diff usi o n ass a y. T his S hi g ell a  is ol at e is r esist a nt t o tri m et h o pri m-
s ulf a m et h ox az ol e a n d is gr o wi n g u p t o t h e dis k ( S X T), t h e z o n e of w hic h is r ec or d e d as 6 m m. 
 
E- t est stri ps. 
I n a d diti o n, azit hr o m yci n Mi ni m u m I n hi bit or y C o nc e ntr ati o ns ( MI C) ar e d et er mi n e d usi n g E-t est 
stri ps. A  s us p e nsi o n of t h e is ol at e is pr e p ar e d a n d t ur bi dit y a dj ust e d u ntil it r e ac h es t h e 0. 5 
Mc F arl a n d st a n d ar d. T h e s us p e nsi o n is us e d t o i n oc ul at e t h e a p pr o pri at e s usc e pti bilit y a g ar 
pl at e f or c o nfl u e nt gr o wt h. T h e pl at e is all o w e d t o dr y. E-t est stri ps ar e pl ac e d o n t h e a g ar 
pl at es wit h st eril e f orc e ps or t w e ez ers; stri ps m a y n ot b e m o v e d o nc e t h e y h a v e t o uc h e d t h e 
pl at es. Aft er t h e stri ps ar e pl ac e d o n t h e pl at e, t h e pl at es ar e i n v ert e d a n d i nc u b at e d at 3 5° C f or 
2 0  t o 2 4 h o urs. T h e MI C is r e a d w h er e t h e elli ps e i nt ers ects t h e MI C sc al e o n t h e stri p. T h e MI C 
will b e c o m p ar e d wit h t h e i nt er pr et ati v e t a bl e acc or di n g t o CL SI g ui d eli n es (s e e A p p e n dix).  
 
8.  P R O T E C TI O N O F H U M A N S U BJ E C T S 
 
B ef or e t h e st u d y b e gi ns, e ac h sit e will o bt ai n f or m al et hic al a p pr o v al fr o m t h eir r es p ecti v e 
n ati o n al et hics c o m mitt e e. I n a d diti o n, l oc al st aff will a p pr o ac h c o m m u nit y l e a d ers t o d escri b e 
t h e st u d y a n d a ns w er a n y q u esti o ns. St u d y st aff will pr oc e e d o nl y if l oc al l e a d ers hi p c o ns e nts t o 
p artici p at e. F or St u d y O n e, w e will o bt ai n v er b al c o ns e nt fr o m a p ar e nt or g u ar di a n of all st u d y 
p artici p a nts. F or St u d y T w o, w e will o bt ai n writt e n c o ns e nt fr o m a p ar e nt or g u ar di a n f or all 

 
3 1  
st u d y acti viti es, i ncl u di n g tr e at m e nt a n d ex a mi n ati o ns. C hil dr e n will o nl y b e i ncl u d e d i n t h e 
st u d y f oll o wi n g r ec ei pt of c o ns e nt fr o m t h e p ar e nt or g u ar di a n. If, at a n y ti m e, a p ar e nt or 
g u ar di a n el ects t o wit h dr a w a f a mil y m e m b er fr o m t h e st u d y, t h e y will b e fr e e t o d o s o. 
C hil dr e n wit h w asti n g or st u nti n g will b e r ef err e d f or a p pr o pri at e tr e at m e nt b y tr ai n e d st u d y 
p ers o n n el t o t h e n e ar est h e alt h c e nt er. 
 
Ris ks a n d B e n efits of St u d y Pr oc e d ur es 
 
S w a b bi n g Pr oc e d ur es  
T h er e ar e mi ni m al ris ks t o t h e p artici p a nt w h o r ec ei v es n as o p h ar y n g e al, a n d n ar es s w a b bi n g. 
P artici p a nts m a y ex p eri e nc e s o m e t e m p or ar y disc o mf ort, b ut t h e s w a b bi n g i n v ol v es mi ni m al 
ris k. A n y a d v ers e eff ects, s uc h as n os e- bl e e ds, will b e tr e at e d i m m e di at el y b y t h e ex a mi n ers. 
Ot h er h e alt h c ar e will b e pr o vi d e d at n o c ost t o t h e st u d y p artici p a nt if n ec ess ar y t o a d dr ess a 
st u d y-r el at e d a d v ers e h e alt h e v e nt. 
 
R ect al s w a bs a n d st o ol s a m pl es: R ect al s w a bs ar e a si m pl e pr oc e d ur e wit h v er y mi ni m al ris k.  
St o ol s a m pl e c oll ecti o n i n a n o n-i n v asi v e pr oc e d ur e wit h n o ass oci at e d ris ks.  
 
A nt hr o p o m etric M e as ur e m e nts  
T h er e ar e mi ni m al ris ks ass oci at e d wit h t h e m e as uri n g b o ar d, sc al e, or M U A C t a p es asi d e fr o m 
a nxi et y d uri n g t h e m e as ur e m e nts. Ex a mi n ers will d o t h eir b est t o e ns ur e t h at t h e 
p ar e nt/ g u ar di a n of t h e c hil d u n d erst a n ds t h e pr oc ess of ass essi n g a nt hr o p o m etric 
m e as ur e m e nts.  T h e ex a mi n ers will att e m pt t o mi ni miz e disc o mf ort f or all st u d y p artici p a nts 
b ef or e, d uri n g, a n d aft er t h e m e as ur e m e nts ar e t a k e n. C hil dr e n wit h w asti n g or st u nti n g will b e 
r ef err e d f or a p pr o pri at e tr e at m e nt at t h e n e ar est h e alt h c e nt er. 
 
9.  A P P E N DI C E S 
 

 
3 2  
 
1 0.  R E F E R E N C E S 
 
1  
W a n g H, B h utt a Z A, C o at es M M, et al.  Gl o b al, r e gi o n al, n ati o n al, a n d s el ect e d s u b n ati o n al 
l e v els of still birt hs, n e o n at al, i nf a nt, a n d u n d er - 5 m ort alit y, 1 9 8 0 – 2 0 1 5: a s yst e m atic 
a n al ysis f or t h e Gl o b al B ur d e n of Dis e as e St u d y 2 0 1 5. T h e L a nc et  2 0 1 6; 3 8 8 : 1 7 2 5– 7 4.  
2  
M ull er O, G ar e n n e M, K o u y at e B, B ec h er H. T h e ass oci ati o n b et w e e n pr ot ei n – e n er g y 
m al n utriti o n, m al ari a m or bi dit y a n d all-c a us e m ort alit y i n W est Afric a n c hil dr e n. Tr o pic al 
M e dici n e & I nt er n ati o n al H e alt h  2 0 0 3; 8 : 5 0 7– 1 1.  
3  
B ei ers m a n n C, B o u nt o g o M, Ti e n dr é b e o g o J, et al.  M al n utriti o n i n y o u n g c hil dr e n of r ur al 
B ur ki n a F as o: c o m p aris o n of s ur v e y d at a fr o m 1 9 9 9 wit h 2 0 0 9. Tr o pic al M e dici n e & 
I nt er n ati o n al H e alt h 2 0 1 2; 1 7 : 7 1 5– 2 1.  
4 
P orc o T C, G e br e T, A y el e B, et al.  Eff ect of M ass Distri b uti o n of Azit hr o m yci n f or Tr ac h o m a 
C o ntr ol o n O v er all M ort alit y i n Et hi o pi a n C hil dr e n: A R a n d o miz e d Tri al. J A M A 2 0 0 9; 3 0 2 : 
9 6 2 – 8.  
5  
G o u g h E K, M o o di e E E M, Pr e n d er g ast AJ, et al.  T h e i m p act of a nti bi otics o n gr o wt h i n 
c hil dr e n i n l o w an d mi d dl e i nc o m e c o u ntri es: s yst e m atic r e vi e w a n d m et a- a n al ysis of 
r a n d o mis e d c o ntr oll e d tri als. B MJ  2 0 1 4; 3 4 8 : g 2 2 6 7– 7.  
6  
Is a n a k a S, L a n g e n d orf C, B ert h é F, et al.  R o uti n e A m oxicilli n f or U nc o m plic at e d S e v er e 
Ac ut e M al n utriti o n i n C hil dr e n. N E n gl J M e d  20 1 6; 3 7 4 : 4 4 4– 5 3.  
7  
Tr e h a n I, G ol d b ac h H S, L a Gr o n e L N, et al.  A nti bi otics as P art of t h e M a n a g e m e nt of S e v er e 
Ac ut e M al n utriti o n. N E n gl J M e d  2 0 1 3; 3 6 8 : 4 2 5– 3 5.  
8  
B a n g A T, B a n g R A, T al e O, et al.  R e d ucti o n i n p n e u m o ni a m ort alit y a n d t ot al c hil d h o o d 
m ort alit y b y m e a ns of c o m m u nit y- b as e d i nt er v e nti o n tri al i n G a dc hir oli, I n di a. T h e L a nc et  
1 9 9 0; 3 3 6 : 2 0 1– 6.  
9  
O n a k p o y a IJ, H a y w ar d G, H e n e g h a n CJ. A nti bi otics f or pr e v e nti n g l o w er r es pir at or y tr act 
i nf ecti o ns i n hi g h-ris k c hil dr e n a g e d 1 2 y e ars a n d u n d er. C oc hr a n e D at a b as e Syst R ev  2 0 1 5; 
9 : C D 0 1 1 5 3 0. 
1 0   M el es e M, Al e m a y e h u W, L a k e w T, et al.  C o m p aris o n of A n n u al a n d Bi a n n u al M ass 
A nti bi otic A d mi nistr ati o n f or Eli mi n ati o n of I nf ecti o us Tr ac h o m a. J A M A 2 0 0 8; 2 9 9 : 7 7 8– 8 4.  
1 1   M el es e M, C hi d a m b ar a m J D, Al e m a y e h u W, et al.  F e asi bilit y of Eli mi n ati n g Oc ul ar 
C hl a m y di a tr ac h o m atis Wit h R e p e at M ass A nti bi otic Tr e at m e nts. J A M A 2 0 0 4; 2 9 2 : 7 2 1– 5.  
1 2   G e br e T, A y el e B, Z eri h u n M, et al.  C o m p aris o n of a n n u al v ers us t wic e- y e arl y m ass 
azit hr o m yci n tr e at m e nt f or h y p er e n d e mic tr ac h o m a i n Et hi o pi a: a cl ust er-r a n d o mis e d 
tri al. L a nc et 2 0 1 2; 3 7 9 : 1 4 3– 5 1.  

 
3 3  
1 3   H ar di n g- Esc h E M, Sill a h A, E d w ar ds T, et al.  M ass Tr e at m e nt wit h Azit hr o m yci n f or 
Tr ac h o m a: W h e n Is O n e R o u n d E n o u g h ? R es ults fr o m t h e P R E T Tri al i n T h e G a m bi a. PL o S 
N e gl Tr o p Dis  2 0 1 3;  7 : e 2 1 1 5– 1 2.  
1 4   A mz a A, K a dri B, N assir o u B, et al.  A cl ust er-r a n d o miz e d tri al t o ass ess t h e effic ac y of 
t ar g eti n g tr ac h o m a tr e at m e nt t o c hil dr e n. Cli n I nf ect Dis  2 0 1 6; D ec e m b er 1 2, 2 0 1 6 . 
1 5   H o us e JI, A y el e B, P orc o T C, et al.  Ass ess m e nt of h er d pr ot ecti o n a g ai nst tr ac h o m a d u e t o 
r e p e at e d m ass a nti bi otic distri b uti o ns: a cl ust er-r a n d o mis e dtri al. T h e L a nc et  2 0 0 9; 3 7 3 : 
1 1 1 1 – 8.  
1 6   C hi d a m b ar a m J D, Al e m a y e h u W, M el es e M, et al.  Eff ect of a Si n gl e M ass A nti bi otic 
Distri b uti o n o n t h e Pr e v al e nc e of I nf ecti o us Tr ac h o m a. J A M A 2 0 0 6; 2 9 5 : 1 1 4 2– 6.  
1 7   Fr y A M, J h a H C, Li et m a n T M, et al.  A d v ers e a n d B e n efici al S ec o n d ar y Eff ects of M ass 
Tr e at m e nt wit h Azit hr o m yci n t o Eli mi n at e Bli n d n ess D u e t o Tr ac h o m a i n N e p al. cli nic al 
i nf ecti o us dis e as es 2 0 0 2; 3 5 : 3 9 5– 4 0 2.  
1 8   C ol es CL, S ei d m a n J C, L e v e ns J, M k oc h a H, M u n oz B, W est S. Ass oci ati o n of M ass 
Tr e at m e nt wit h Azit hr o m yci n i n Tr ac h o m a- E n d e mic C o m m u niti es wit h S h ort- T er m 
R e d uc e d Ris k of Di arr h e a i n Y o u n g C hil dr e n. A m eric a n J o ur n al of Tr o pic al M e dici n e a n d 
Hy gi e n e  2 0 1 1; 8 5 : 6 9 1– 6.  
1 9   C ol es CL, L e v e ns J, S ei d m a n J C, M k oc h a H, M u n oz B, W est S. M ass Distri b uti o n of 
Azit hr o m yci n f or Tr ac h o m a C o ntr ol Is Ass oci at e d Wit h S h ort-t er m R e d ucti o n i n Ris k of 
Ac ut e L o w er R es pir at or y I nf ecti o n i n Y o u n g C hil dr e n. T h e P e di atric I nf ecti o us Dis e as e 
J o ur n al 2 0 1 2; 3 1 : 3 4 1– 6.  
2 0   G a y n or B D, A mz a A, K a dri B, et al.  I m p act of m ass azit hr o m yci n distri b uti o n o n m al ari a 
p ar asit e mi a d uri n g t h e l o w-tr a ns missi o n s e as o n i n Ni g er: a cl ust er-r a n d o miz e d tri al. A m J 
Tr o p M e d Hy g  2 0 1 4; 9 0 : 8 4 6– 5 1.  
2 1  A y el e B, G e br e T, H o us e JI, et al.  A d v ers e E v e nts aft er M ass Azit hr o m yci n Tr e at m e nts f or 
Tr ac h o m a i n Et hi o pi a. A m eric a n J o ur n al of Tr o pic al M e dici n e a n d Hy gi e n e  2 0 1 1; 8 5 : 2 9 1– 4.  
2 2   S k al et A H, C e v all os V, A y el e B, et al.  A nti bi otic S el ecti o n Pr ess ur e a n d M acr oli d e R esist a nc e 
i n N as o p h ar y n g e al Str e pt oc occ us p n e u m o ni a e: A Cl ust er- R a n d o miz e d Cli nic al Tri al. PL o S 
M e dici n e  2 0 1 0; 7 : e 1 0 0 0 3 7 7– 9.  
2 3   H a u g S, L a k e w T, H a bt e m ari a m G, et al.  T h e d ecli n e of p n e u m oc occ al r esist a nc e aft er 
c ess ati o n of m ass a nti bi otic distri b uti o ns f or tr ac h o m a. Cli n I nf ect Dis  2 0 1 0; 5 1 : 5 7 1– 4.  
2 4   D o a n T, Arzi k a A, R a y KJ, et al.  G ut micr o bi al di v ersit y i n a nti bi otic- n aï v e c hil dr e n aft er 
s yst e mic a nti bi otic ex p os ur e: a r a n d o miz e d c o ntr oll e d tri al. cli nic al i nf ecti o us dis e as es 
2 0 1 7; I n pr ess: 1– 1.  

 
3 4  
2 5   Y ass o ur M, V at a n e n T, Silj a n d er H, et al.  N at ur al hist or y of t h e i nf a nt g ut micr o bi o m e a n d 
i m p act of a nti bi otic tr e at m e nt o n b act eri al str ai n di v ersit y a n d st a bilit y. Sci e nc e 
Tr a nsl ati o n al M e dici n e  2 0 1 6; 8 : 3 4 3r a 8 1. 
2 6   K or p el a K, S al o n e n A, Virt a LJ, et al.  I nt esti n al micr o bi o m e is r el at e d t o lif eti m e a nti bi otic 
us e i n Fi n nis h pr e -sc h o ol c hil dr e n. N at ur e C o m m u nic ati o ns  2 0 1 6; 7 : 1– 8.  
2 7   D et hl efs e n L, H us e S, S o gi n ML, R el m a n D A. T h e P er v asi v e Eff ects of a n A nti bi otic o n t h e 
H u m a n G ut Micr o bi ot a, as R e v e al e d b y D e e p 1 6 S r R N A S e q u e nci n g. PL O S Bi ol o gy  2 0 0 8; 6 : 
e 2 8 0 – 1 8.  
2 8   R utt e n N B M M, Rij k ers G T, M eijss e n C B, et al.  I nt esti n al micr o bi ot a c o m p ositi o n aft er 
a nti bi otic tr e at m e nt i n e arl y lif e: t h e I N C A st u d y. B M C P e di atrics  2 0 1 5; : 1 – 8.  
2 9   L a n g d o n A, Cr o o k N, D a nt as G. T h e eff ects of a nti bi otics o n t h e micr o bi o m e t hr o u g h o ut 
d e v el o p m e nt a n d alt er n ati v e a p pr o ac h es f or t h er a p e utic m o d ul ati o n. G e n o m e M e dici n e  
2 0 1 6; : 1 – 1 6.  
3 0   S mit h MI, Y ats u n e n k o T, M a n ar y MJ, et al.  G ut micr o bi o m es of M al a wi a n t wi n p airs 
disc or d a nt f or k w as hi or k or. Sci e nc e  2 0 1 3; 3 3 9 : 5 4 3– 8.  
3 1   Pr e ntic e A M, N a b w er a H, K w a m b a n a B, A nt o ni o M, M o or e S E. Micr o b es a n d t h e 
M al n o uris h e d C hil d. Sci e nc e Tr a nsl ati o n al M e dici n e  2 0 1 3; 5 : 1– 3.  
3 2   O wi n o V, A h m e d T, Fr e e m ar k M, et al.  E n vir o n m e nt al E nt eric D ysf u ncti o n a n d Gr o wt h 
F ail ur e/ St u nti n g i n Gl o b al C hil d H e alt h. P e di atrics  2 0 1 6; 1 3 8 : e 2 0 1 6 0 6 4 1– 1.  
3 3   
W at a n a b e K, P etri W A Jr. E n vir o n m e nt al E nt er o p at h y: El usi v e b ut Si g nific a nt S u bcli nic al 
A b n or m aliti es i n D e v el o pi n g C o u ntri es. E BI O M  2 0 1 6; 1 0 : 2 5– 3 2.  
3 4   L oc ks L, M wir u R, Mtisi E, et al.  I nf a nt N utriti o n al St at us a n d M ar k ers of E n vir o n m e nt al 
E nt eric D ysf u ncti o n ar e Ass oci at e d wit h Mi dc hil d h o o d A nt hr o p o m etr y a n d Bl o o d Pr ess ur e 
i n T a nz a ni a. T h e J o ur n al of P e di atrics  2 0 1 7; : 1 – 1 0.