This  report  was  produced  for  review  by  the  United  States  Agency  for  International  Development.    It  was  prepared by  London   School  of  Hygiene  and  Tropical  Medicine  and  Georgia  Institute  of  Technology  and  was  authored  by  Oliver  Cumming,  Helen   Buxton,  Jackie  Knee  and  Joe  Brown. This  report  is  made  possible  by  the  support  of  the  American  People  through  the  Unites  States Agency  for  International   Development  (USAID).  The  contents  of  this  report  are  the  sole  responsibility  of London  School  of  Hygiene  and  Tropical   Medicine  and  do  not  necessarily  reflect  the  views  of  USAID  or  the  United  States  Government. FINAL REPORT THE MAPSAN TRIAL A controlled, before-and-after trial of an urban sanitation intervention to reduce enteric infections in children June 2017 Oliver  Cumming,  London  School  of  Hygiene  and  Tropical  Medicine Helen  Buxton,  London  School  of  Hygiene  and  Tropical  Medicine Jackie  Knee,  Georgia  Institute  of  Technology Joe  Brown,  Georgia  Institute  of  Technology Sub  agreement No.  FY14-­‐G09-­‐6990
 Under  Cooperative  Agreement  No.  GHS-­‐A-­‐00-­‐09-­‐00015-­‐00   Translating  Research  Into  Action  (TRAction)   London  School  of  Hygiene  and  Tropical  Medicine,  UK  (Prime) Georgia  Institute  of  Technology,  USA  (Sub-­‐awardee) Minsterio  de  Saude  de  Mocambique (Sub-­‐awardee) University  of  North  Carolina,  USA  (Sub-­‐awardee) University  of  Florida,  USA  (Sub-­‐awardee) Water  and  Sanitation  for  the  Urban  Poor,  UK  (Sub-­‐awardee)   To  learn  more  about  TRAction,  visit  www.tractionproject.org or  contact  us  at  tracinfo@urc-­‐chs.com University  Research  Co.,  LLC    •    7200  Wisconsin  Avenue,  Suite  600    •    Bethesda,  MD  20814    •    301-­‐654-­‐8338 www.urc-­‐chs.com Acknowledgements: We  would  like  to  acknowledge  the  hard  work  of  the  various  researchers  within  each   of  our  partner  organizations.  The  MapSan  Trial  is  a  large  undertaking  which  would  not  be  possible  without   the  support  of  a  number  of  scientific  and  implementing  partners  across  the  world,  and  we  are  very  grateful   for  their  commitment  to  this  important  project.  In  particular,  we  would  like  to  acknowledge  the  support   of   our   national   scientific   partner,   the   National   Institute   of   Health,   within   the   Ministry   of   Health   for   Mozambique,  under  the  leadership  of  Dr. Rassul  Nala.     Translating   Research   into   Action,   TRAction,   is   funded   by   United   States   Agency   for   International   Development   (USAID)   under   cooperative   agreement   No.   GHS-­‐A-­‐00-­‐09-­‐00015-­‐00.   The   project   team   includes  prime  recipient,  University  Research  Co.,  LLC  (URC),  Harvard  University  School  of  Public  Health   (HSPH),  and  sub-­‐recipient  research  organizations. Recommended  Citation: MapSan (2017).  The  MapSan  Trial: Final  Report. Published  by  the  Translating   Research  into  Action  Project.  Bethesda,  MD:  University  Research  Co.,  LLC  (URC). 3 CONTENTS List  of  tables  and  figures .............................................................................................................................. 4 Abbreviations............................................................................................................................................... 5 Executive  Summary ...................................................................................................................................... 6 Introduction ................................................................................................................................................. 7 Project  management  and  Stakeholder  Engagement.................................................................................... 8 Research  activities........................................................................................................................................ 9 Results ....................................................................................................................................................... 13 Barriers  and  Challenges.............................................................................................................................. 22 Conclusions  and  Recommendations .......................................................................................................... 23 Next  steps................................................................................................................................................... 23 4 LIST OF TABLES AND FIGURES Figure 1: Cumulative enrolment/visitation of children and stools collected in intervention & control arms at baseline and endline...................................................................................................................................................10 Figure 2: Cumulative enrolment/survey completion and stool collection for baseline & endline ...............11 Figure 3: Satellite imagery of the study area (left) with rooftop delineations within a 50-meter radius (right). ................................................................................................................................................................................14 Figure 4: Baseline prevalence of enteric pathogens in stool samples of intervention & control children .15 Figure 5: Baseline prevalence, by age group, of enteric pathogens detected in stool by GPP assay. ..........16 Figure 6: Baseline prevalence of 3 classes of enteric pathogen by age group...................................................17 Figure 7: Baseline prevalence of enteric pathogen co-infection by age group..................................................17 Figure 8: Age distribution of (caregiver-reported) diarrhoea among enrolled children from intervention and groups at baseline control.....................................................................................................................................18 Figure 9: Baseline prevalence of any STH infection, Ascaris, and Trichuris......................................................19 Figure 10: STH co-infections in children at baseline...............................................................................................19 Figure 11: Endline prevalence of enteric pathogens by study arm. .....................................................................20 Figure 12: Endline prevalence of coinfection by study arm...................................................................................21 Figure 13: Endline prevalence of any STH, Ascaris, and Trichuris, by age, in intervention and control children..............................................................................................................................................................................21 Table 1: Endline visitation & sample collection completed ........................Error! Bookmark not defined. Table 2: Baseline characteristics of enrolled intervention & control compounds..Error! Bookmark not defined. Table 3: Characteristics of enrolled children at baseline............................Error! Bookmark not defined. 5 ABBREVIATIONS CDC Centers  for Disease  Control  and  Prevention BMGF Bill  and  Melinda  Gates  Foundation GPP Gastrointestinal  Pathogen  Panel GT Georgia  Institute  of  Technology INS Instituto  Nacional  de  Saude  (National  Institute  of  Health,  Mozambique) IRB Institutional  Review  Board LSHTM London  School  of  Hygiene  and  Tropical  Medicine MapSan Maputo  Sanitation  Trial MISAU Ministry  of  Health,  Mozambique MPN Most  Probable  Number NCE   No  Cost  Extension PMP Performance  Monitoring  Plan STH Soil-­‐Transmitted  Helminths TAG Technical  Advisory  Group UF University  of  Florida UNC University  of  North  Carolina  at  Chapel  Hill URC University  Research  Co.,  LLC US United  States USAID United  States  Agency  for  International  Development WASH Water,  sanitation  and  hygiene WSUP Water  and  Sanitation  for  the  Urban  Poor 6 EXECUTIVE SUMMARY This  is  the  final  report  describing the  work  undertaken  by  the  MapSan  Trial  consortium and  complements   the  two  annual  reports  (2015  and  2016)  already  submitted.  The  MapSan  consortium  is  led  by  the  prime-­‐ awardee,   the   London   School   of   Hygiene   and   Tropical   Medicine (LSHTM),   and   includes   the   following   partners,   or   sub-­‐awardees:   The   Georgia   Institute   of   Technology   (GT),   the   Ministry   of   Health   for   Mozambique  (MISAU),  the  University  of  North  Carolina  (UNC),  the  University  of  Florida  (UF),  and  Water   and   Sanitation   for   the   Urban   Poor   (WSUP).   This   report   describes   final   progress   against   the   project   objectives and  details all  completed  activities  through  to  the  end  date  (May  15, 2017). Key  activities  were  completed  as  specified  by  the  grant  in  time  for  project  close-­‐down  in  April  2017.   These  included:  completion  of  endline  survey;  preparation  of  scheduled  papers  and  the  completion  of   financial  and  administrative  requirements.   Laboratory  analysis  of  samples  is  ongoing  and  will  be  completed late  2017 with  final  analysis  of  data  to   be  completed  in  early  2018.   7 INTRODUCTION Access  to  safe  sanitation  in  low-­‐income,  informal  settlements  of  Sub-­‐Saharan  Africa  has  not  significantly   improved   since   1990.   The   combination   of   a   high   faecal-­‐related   disease   burden   and   inadequate   infrastructure  suggests  that  investment  in  expanding  sanitation  access  in  densely  populated  urban  slums   can  yield  important  public  health  gains.  No  rigorous,  controlled  intervention  studies  have  evaluated  the   health  effects  of  decentralized (non-­‐sewerage)  sanitation  in  an  informal  urban  setting,  despite  the  role   that  such  technologies  will  likely  play  in  scaling  up  access.  The  MapSan  trial  is  a  controlled,  before-­‐and-­‐ after   (CBA)   trial   to   estimate   the   health   impacts   of   an   urban   sanitation   intervention   in   informal   neighbourhoods of  Maputo,  Mozambique,   including   an   assessment   of   whether   exposures   and   health   outcomes  vary  by  localized population  density.   The  intervention  consists  of  private  pour-­‐flush  latrines  (to  septic  tank)  shared  by  multiple  households  in   compounds  or  household  clusters.  MapSan has  measured objective  health  outcomes  in  children  drawn   from   the   intervention,   matched   with   controls   using   existing   shared   private   latrines   in   poor   sanitary   conditions,  at  two time  points:  immediately  before  the  intervention and  at   follow-­‐up  after  12  months.   The  primary  outcome  is  combined  prevalence  of  selected  enteric  infections  among  children  under  5  years   of   age.   Secondary   outcome   measures   include   soil-­‐transmitted   helminth   (STH)   reinfection   in   children   following  baseline  deworming  and  prevalence  of  reported diarrheal  disease.  The  study has used exposure   assessment,  faecal source  tracking,  and  microbial  transmission  modelling to  examine  whether  and  how   routes  of  exposure  for  diarrheagenic  pathogens  and  STHs  change. The  MapSan project activities  for  the  duration  of  the  project  are  described  under  three  headings:  (1)   Project  management  and  stakeholder  engagement;  (2)  Research  activities; and (3)  Preliminary  results   and  findings. 8 PROJECT MANAGEMENT AND STAKEHOLDER ENGAGEMENT Project  management  and  stakeholder  engagement  activities  have been  successfully  implemented and   this  report  forms  part  of  the  closedown  of  the  contract.  Multiple  no cost  extensions  (NCEs) were   required  as  the  initial  duration  of  the  contract  was  less  that  the  stated  period  of  time  required  to   implement  the  study  protocol.  Requesting  multiple  NCEs for  the  prime  and  then  issuing  NCEs  to  sub-­‐ awardees was  time  consuming  but  the  implementation  of  research  activities  was  not  delayed. Ethical  approval  for  the  MapSan  Trial  was  initially  granted  by  the  Ethics  Committee  of  London  School  of   Hygiene  and  Tropical  Medicine  (LSHTM) and  the  National  Bio-­‐Ethics  Committee  of  Mozambique  (CNBS)   and  the  trial  was  registered  on  a public  trial  registry  (www.clinicaltrials.gov)  prior  to  study  participant   enrolment  as  per  best  practice.  Following  this,  other  academic  partners  (Georgia  Institute  of  Technology (GT),  University  of  North  Carolina  at  Chapel  Hill (UNC)  and  University  of Florida  (UF)); secured  ethical   approval  for  their  participation  in  the  study  from  their  respective  Institutional  Review  Boards  (IRB).  All   amendments  to  the  study  protocol  received  ethical  approval  from  the  LSHTM  Ethics  Committee  and   CNBS  prior  to  implementation. Following  the  inception  of  the  MapSan  Trial,  the  consortium  has  successfully  raised  additional  funds   from  other  donors  to  support  related  work  which  is  complementary  to  the  main  trial.  This  includes  a   cohort  study  for  the  effect  of  sanitation  on  oral  rotavirus  vaccine  immunogenicity  funded  by  the  Centres for  Disease  Control  Foundation  (CDCF)  and  funding  from  the  Bill  and  Melinda  Gates  Foundation  (BMGF)   to: conduct  a  social-­‐behavioural  investigation of  the  intervention; perform  a  cost-­‐effective  analysis  of   the  intervention; and  to  conduct  a  24-­‐month follow-­‐up  survey  to  assess  the  effect  of  the  intervention  on   childhood  nutritional  outcomes. As  per  the  objectives  of  the  URC  TRAction programme,  the  MapSan  team has  engaged  policy,  practice and academic audiences  throughout.  The  primary  purpose  of  this  engagement  has  been to  gather  inputs  into   the  study  design  and  delivery,  generate  interest in  the  project, and  to  create  demand  for  research  findings in  advance  of the  study’s  completion.  This  approach  is  expected  to  increase the  likelihood  that  the  study   findings  will  be  used  in  policy  and  practice.  These  activities  have  been  reported in  the  two  MapSan  Annual   Reports submitted  earlier  and  future  planned  activities  post-­‐closedown  of  the  grant  have  been  described   in  the  Research  Uptake  Plan  submitted  in  June  2017. In  2017 findings  will  be  presented  at  the  University   of  North  Carolina’s  Water  and  Health  Conference  in  Chapel  Hill,  USA, the  Stockholm  World  Water  Week in  Stockholm, Sweden, the  American  Society  of  Tropical  Medicine  and  Health  Conference in  Baltimore,   USA,  and  the  International  Conference  on  Urban  Health  in  Coimbra,  Portugal.   9 RESEARCH ACTIVITIES SUMMARY A   full   account   of   research   activities   through  December   2016 is   provided   in   the   previously   submitted   MapSan   Annual   Reports   (2015   and   2016).   Subsequent   to   the   last   annual   report,   the   endline   data   collection has   been   successfully   completed   and   samples   are   undergoing   laboratory   analysis   at   GT in   Atlanta,  USA.  As  per  the  stated  final  deliverables  of  the  project,  we  submitted  the  following  manuscripts   to  URC in  June  2017: 1. Brown   et   al.  A   controlled,   before-­‐and-­‐after   trial   of   an   urban   sanitation   intervention   to   reduce   enteric   infections   in   children:   research   protocol   for   the   Maputo   Sanitation   (MapSan)   study,   Mozambique.  Published  2015  in  BMJ  Open 2. Watson  et  al.    The  association  between  shared  sanitation  and  childhood  diarrhoea,  and  stunting   in   low-­‐income,   informal   urban   settlements   of   Maputo,   Mozambique: evidence   from   a   cross-­‐ sectional  study.   To  be  submitted  to  Tropical  Medicine  and  International  Health. 3. Knee  et  al.  A  controlled,  before-­‐after  trial  of  an  urban  sanitation  intervention  to  reduce  enteric infections  in children:  baseline  results  from  the  Maputo  Sanitation  (MapSan)  study, Mozambique.   To  be  submitted  to  PLOS  Medicine. In  addition,  the  following  additional  manuscripts  are  currently  in  development: 1. Rheingans  et  al.  Modelling the  interactions  between  population  density,  sanitation  coverage  and   socio-­‐economic  inequalities  in  low-­‐income  urban  environments.  Expected  submission:  October   2017 2. Liang  et  al. A  quantitative  framework  for  assessing  the  effects  of  sanitation  on  transmission  of   enteric  pathogens  among  children  under  five  in  urban  environment  of  Mozambique. Expected   submission:  October  2017 3. Liang  et  al.  Modelling transmission  of  soil-­‐transmitted  helminths  (STHs)  in  Maputo,  Mozambique   – risk  factors  and  the  effects  of  sanitation  improvement.  Expected  submission: November  2017 4. Holcomb  et  al.  Measuring  the  effect  of  a  sanitation  intervention  on  exposure  to  faecal contamination in  the  domestic  environment  in  Maputo,  Mozambique  using  traditional  and  host-­‐ specific  faecal indicator.  Expected  submission:  December  2017 Beyond  these  manuscripts  which  are  in  active  development,  there  will  be  a  number  of  additional  papers   including  the  primary  outcome  results  paper  which  we  expect  to  submit  for  publication  in  March  2018. Information   on   the   completed   endline   survey is   presented   below   along   with information   from   the   laboratory  analysis completed  to  date. 10 DATA COLLECTION Study   enrolment began   on   February the   2nd 2015 and   at   the   completion   of   baseline   data   collection; February  2016;   536 children   from control  compounds  and  447 children   from intervention  compounds   were  enrolled (Figure  1).  Following  the  initial  baseline  phase,  100  additional  intervention  latrines  were   constructed within  the  study  area  between  November  2016  and  May  2017.  An  additional  94  children  were   enrolled   from   the   new   intervention   compounds,   bringing the   total   number   of   intervention   children enrolled to  541. The  study  area,   from  which  both  intervention  and  control  clusters  were drawn,  spans   over  16  bairros  – or  neighbourhoods  -­‐ on  the outskirts  of  downtown  Maputo. Figure  1:  Cumulative  enrolment/visitation  of  children  and  stools  collected  in  intervention  &  control  arms  at  baseline  and  endline Endline  data  collection  began  after  the  completion  of  baseline  data collection  in  February  2016.   Enrolment in  the  study  cohort  was  progressive:  a  total  of  199  and  217  additional  children  were  enrolled   in  the  intervention  and  control  arms,  respectively,  during  endline.  The  amount  of  loss  to  follow-­‐up  was   high  in  both  arms: 144  intervention  and  216  control  children  who  were  enrolled  at  baseline  were   unavailable  at  endline  (Table  1).  The  main  causes  of  loss  to  follow-­‐up  was  population  churn  (i.e.  children   or  households  moved  or  were  traveling  for  extended  periods).  There  were 5  deaths  and  8  caregivers   refused  to  continue  in  the  study.   Intervention  compounds  were  visited  12  months  after  they  began  using  their  newly  constructed  latrine;   an  average  of  50  days  after  baseline  enrolment.  The  matched  control  compounds  were  visited  12+   months  after  baseline  enrolment.  Survey  data,  biometrics,  and  all  samples  (stool,  saliva,  water,  soil,   flies)  were collected  for  all  study  sites  as  per  protocols. 11 Table  1:  endline  visitation  and  sample  collection  completed Intervention                                             n  (%  of  available  children,  n=517) Control n  (%  of  available  children,   n=585) Children  eligible  for  endline   visitation 661 8001 Children  lost  to  follow-­‐up  (moved,   died,  traveling) 144 216 Children  available  for  endline visitation  (not  lost  to  follow-­‐up) 517 (100%) 585 (100%) Child  surveys  completed  for   available  children 424 (82%) 435 (74%) Stools  collected 423 (81%) 430 (74%) Children  with  completed  survey   and  stool  collected 356 (69%) 351 (60%) During  endline data  collection, stools  were  collected  from  81%  and  surveys  from  82%  of  the  517   available  intervention  children  (Table  1).  Stool  samples  and  survey  data  were  collected  from  74%  of  the   585  children  available  in  the  control  group  (Table  1). The  rate  of  endline  visitation,  data  and  stool   collection,  was  similar  to  the  rate  of  enrolment  at  baseline  (Figure  1,  Figure  2). Figure  2: Cumulative  enrolment/survey  completion  and  stool  collection  for  baseline  &  endline 12 DEWORMING All  compounds  enrolled  in  the  MapSan trial  were  provided  with  400-­‐mg  of  Albendazole  for  deworming.   For  intervention  compounds,  deworming  was  conducted  once  the  compound  began to  use  their  newly   constructed  WSUP  latrine.  On  average  deworming  occurred  within 17  days  of  the  latrine  first  being   used.  For  control  compounds  deworming  occurred  following  baseline  enrolment.  On  average  there  were 28  days  between  enrolment and  deworming  of  control  compounds.   A  second  round  of  deworming  was   completed  just  prior  to  the  completion  of  endline  visitation  (January  – March  2017).   STOOL  AND  SALIVA  SAMPLES Enumerators  attempted  to  collect  stool  samples  from  every  child  enrolled  in  the  study at  each  time   point.  During  the  baseline  phase  a  total  of  396 and  410 stool  samples  were  collected  from  children  in   the  intervention  and  control  arms,  respectively (Figure  1,  Figure  2).  A  total  of  764  saliva  samples  were   collected  from  366  intervention  children  and  398  control  children  at  baseline.  During  the  endline  phase,   stool  samples were collected  from  423 and  430 children  in  the  intervention  group and control  group respectively (Figure  1,  Figure  2),  and  saliva  samples  were  collected  from  398  intervention  and  439   control  children. All  stool  samples  were  transported  to MISAU for  immediate  analysis  by  Kato-­‐Katz  and   storage.  Four  sub-­‐samples  of  each  stool  were  stored  at  -­‐80°C until  future  analysis  or  shipment  to  GT.   Saliva  samples  were  transported  to  MISAU  on  the  day  of  collection  and  immediately  stored  at  -­‐80°C. Using  the  Kato-­‐Katz  technique  stool  samples were examined  by  the  MISAU  laboratory  under  10  – 40x   magnification  to  identify  the  presence  of  STH  ova.  Laboratory  technicians  counted the  number  of   hookworm,  Trichuris  trichiura,  fertile  and  in-­‐fertile  Ascaris  lumbricoides eggs,  and  Enterobius   vermicularis present  in  the  slide  preparation.  Counts  were transformed  into  concentrations  of  ova/g  by   multiplying  by  a  factor  of  24.  In  addition  to  enumerating  the  above  species  ova,  technicians  determined whether  the  following  STH  species  are  present  in  the  sample  preparation:  Taenia  species,  Hymenolepis   nana and  Hymenolepis  dimunita. During  this  reporting  period, all  baseline  stool  samples  were received  by  GT  for  analysis  of  the  primary   outcome: combined  non-­‐viral,  non-­‐STH  enteric  infection; using  the  Gastrointestinal  Pathogen  Panel (GPP) multiplex  molecular  assay.  To  date,  715 baseline  stools  samples  have  been  analysed by  the  GPP. All stool  samples  have  now  been  received  by  GT and  to  date  just  over  65%  of  the  endline  samples  have   been  analysed by  GPP. 13 RESULTS Baseline  data  collection,  including  the  expanded baseline  sample,  was  completed in  early  2017.  Below is   a  preliminary  descriptive  analysis  of  baseline  data including demographic  information,  health  status  data   and  information about  environmental  conditions. BASELINE  ENVIRONMENTAL  CHARACTERISTICS A series  of  key  characteristics  of  intervention  and  control  compounds  at  baseline are  detailed in  Table  2.   This  data,    along  with  the  health  status  data,    has been used  to  assess  the  comparability  between  the   two  trial  arms,  ensuring  any  significant  differences  can  be  accounted  for  in  the  final  analysis. The   difference  in  access  to  a  water  tap  in  compounds  and  the  mean  number  of  people  residing  in   compounds  in  the  intervention  arm  and  control  arm  will  be  accounted  for. It  should  be stressed that   these  numbers  are  not  final  and  a  formal  analysis  of  baseline  data  is  planned  for  the third quarter  of   2017. Table  2:  Baseline  characteristics  of  enrolled  intervention  &  control  compounds Intervention Control Total Total  sites  (compounds)  enrolled 275  (233  SL,  42  CSB) 299 574 Mean  #  people/compound* 17.2 14.1 15.6 Mean  #  children/compound 2.1 1.9 2.0 Water  tap  in  compound* 85% 74% 79% Animals  present  in  compound 61% 61% 61% Flooding  during  rainy  season 59% 65% 62% Latrine  characteristics   Drop  hole  cover  present Pedestal/masonry  present Superstructure  present 59% 40% 30% 55% 38% 25% 57% 39% 28% *Significantly  different at p  =  0.05  (unadjusted) BASELINE  CHARACTERISTICS OF  ENROLLED  CHILDREN Table  3  below  shows  the  age  distribution  and  characteristics  of  children  enrolled  in  intervention  and   control  arms. Characteristics  of  children  enrolled  in  both  arms  are  well  balanced. The  difference  in  the   proportion  of  females  enrolled  in  intervention  and  controls  arms  will  be  accounted  for  in  the  final   analysis. 14 Table  3:  Characteristics  of  enrolled  children  at  baseline Intervention Control Total #  children  enrolled 541 536 1077 Age  groups 0-­‐11  months 12-­‐23  months 24-­‐48  months 28% 27% 43% 28% 27% 42% 28% 27% 43% Female* 44% 52% 48% Breastfed,  exclusively 10% 9.0% 10% Child  defecation   Diapers Latrine Other  (open  defecation,  bucket) 63% 9.1% 27% 65% 11% 23% 65% 10% 25% *Significantly  different  at  p =  0.05  (unadjusted) POPULATION  DENSITY One objective  of  the  MapSan  study  is  to  assess  whether  population  density  modifies  the  effect  of  the   sanitation  intervention  on  child  health  outcomes. Population  density  is  being  assessed  in  two  ways:  (1)   by  measuring  compound  area  and  calculating  the  number  of  people  per  square  meter  of  living  space;   and  (2)  by  using  satellite  imagery  and  a  rooftop  area  algorithm  to  calculate  the  number  of  people  living   within  50  meters  and  100  meters  of  study  compounds.  Figure  4 illustrates  density  method  (2),  depicting   a  section  of  the  satellite  imagery  overlaid with  rooftop  delineations  within  a  50-­‐meter radius.   Figure  3: Satellite  imagery  of  the  study  area  (left)  with  rooftop  delineations  within  a  50-­‐meter  radius  (right). 15 BASELINE  DISEASE  BURDEN Baseline  primary  outcome  data  by  study  arm: Primary  outcome  data  are  derived  from  molecular  analysis  of  stool  specimens  by  the  GPP  assay  at  GT  and   are  presented  in  Figure  4.  Overall,  the  combined  prevalence  of  non-­‐STH,  non-­‐viral  enteric  infections  was   84%  in  the  intervention  arm  and  88%  in  the  control  arm  at  baseline, which  was  not  significantly  different.   Because   our   sample   size   calculations   assumed   an   overall   prevalence   of   71%   at   baseline,   this   higher   prevalence   results   in   greater   statistical   power   for   trial   data   analysis.   The   most   frequently   detected   enteropathogens,   irrespective   of   study   arm   were Giardia (52%),   Shigella   (45%),   ETEC   (31%),   and Salmonella spp.  (21%). Figure  4: Baseline  prevalence  of  enteric  pathogens  in  stool  samples  of  intervention  &  control  children 16 Baseline  primary  outcome  results  by  age: The   prevalence   of   the   primary   outcome   and   the   two   most   frequently   detected   entereopathogens,  Giardia and  Shigella,  increased  with  child  age. The  prevalence  of  most  other   individual  enteric  pathogens  was  similar  across  age  groups  with  the  exception  of  norovirus  GI/GII   and  Salmonella,  which  decreased  with  increasing  age.  In  general,  the  prevalence  of  infection  with   one  or  more  bacterial  or  protozoan  pathogen  increased  with  increasing  age  and  the  prevalence   of   enteric   viral   infection   decreased   with   increasing   age.   Enteric   pathogen   coinfection   also   increased  with  age.  Figures  5,  6,  and  7,  present  prevalence  data  by  a  priori age  strata. Figure  5:  Baseline  prevalence,  by  age  group,  of  enteric  pathogens  detected  in  stool  by  GPP  assay. 17 Figure  6: Baseline  prevalence  of  3  classes  of  enteric  pathogen  by  age  group. Figure  7:  Baseline  prevalence  of  enteric  pathogen  co-­‐infection  by  age  group. 18 The  two  secondary  outcomes  of  the  MapSan  study  are: STH  reinfection  following  baseline  deworming; and  reported  incidence  of  diarrhoea among  enrolled  children. Figure  8 below  shows  the  age  distribution   of  caregiver-­‐reported  diarrhoea among  enrolled  children  from  intervention  and  control  groups  at   baseline.  At baseline  the  incidence  of  caregiver-­‐reported  diarrhoea is highest  in  children  aged  between  1   – 24  months. Figure  8: Age  distribution  of  (caregiver-­‐reported)  diarrhoea  among  enrolled  children  from  intervention  and  groups  at  baseline   control Baseline  STH  data All  baseline  stool  samples  (n=745)  were  analysed for  the  presence  of  STH  ova  using  the  Kato-­‐Katz   technique  at  the  MISAU  laboratory  in  Maputo. Overall  baseline  prevalence  for  any  STH  infection   by   Kato-­‐Katz  was   44%  and   is   similar   between   study  arms   (Figure  9).   As  expected,   prevalence   increases  with  age  (Figure  9).  T.  trichuris and  A.  lumbricoides are  the  most  commonly  observed   STHs   with  prevalence rates of   37%   and   23%,   respectively.   Coinfection   with  multiple   STH   was   observed  in  16%  of  samples  (Figure  10)  and  increased  with  age.  An  initial  assessment  of  balance   between  intervention  and  control  arms  of  the  trial  showed  no  statistically  meaningful  difference   in  STH  prevalence.  Covariates   related   to  sanitation  conditions   (e.g.,  presence  of  a  pedestal  or   drop-­‐hole   cover)   or   to   household   conditions   and   demographics   (e.g.,   presence   of   any   floor   covering,  number  of  people  living  in   the  household,  education  level  of   the  primary  caretaker,   socio-­‐economic  status)  were  not  significant  risk  factors  for  STH  infections. Control Intervention 19 Figure  9:  Baseline  prevalence  of  any  STH  infection,  Ascaris,  and  Trichuris. Stratified  by  study  arm  and  age  group.  Intervention  mean  and  95%  confidence  interval  represented  by   blue  markers  and  bars.  Control  mean  and  95%  confidence  interval  represented  by  green  markers  and   bars.   Figure  10:  STH  co-­‐infections  in  children  at  baseline. 0 Infections 1 Infection 2 Infections 3 Infections 54% 31% 15% STHs 20 ENDLINE  HEALTH  STATUS   To  date,  547  of  the  853  samples  collected  at  endline  have  been  analysed using  the  GPP.  Preliminary   analyses  of  the  endline  GPP  data  show  an  overall  primary  outcome  prevalence  of  87% and  90% in   intervention  and  control  arms  respectively.  Similar  to  results  at  baseline,  Giardia,  Shigella,  ETEC,  and   Salmonella  were  the  most  frequently  detected  pathogens  among  intervention  and  control  children.   Formal  analyses  of  these  data  have  not  yet  been  performed  and  will  be  completed  following  the  formal   baseline  analysis.  Given  that  the  laboratory  analysis  of  endline  samples  has  not  been  completed  and  the   formal  analysis  of  this  data  has  not  been  performed;   no  inference  should  be  drawn  as  to  the  effect  of   the  intervention  and  any  data  presented  here  must  not  be  shared  externally  nor published  as  this  would   be  in  contravention  of  the  protocol and  the  conditions  of  the  ethical  approval  for  the  study.  The   prevalence  of  coinfection  remained  high in  the  preliminary  analysis:  68%  of  intervention  children  and   65%  of  control  children  were  infected  with  two  or  more  enteropathogens  (Figure  12).     Figure  11:  Endline  prevalence  of  enteric  pathogens  by  study  arm. 21 Figure  12:  Endline  prevalence  of  coinfection  by  study  arm. Similar  to  the  findings  at  baseline,  initial  analysis  of  endline  data  suggests  the  most  common  STH   infections  in  both  the  intervention  and  control  children  remain  T.Trichiura and  A.  lumbricoides  (Figure   13). Figure  13: Endline  prevalence  of  any  STH,  Ascaris,  and  Trichuris,  by  age,  in  intervention  and  control  children. 22 BARRIERS AND CHALLENGES Most  challenges  faced  during  the  study  were manageable  and  were addressed  as  they  arose.  However,   the  study  encountered  two  significant challenges  which  whilst  managed  effected  the  execution  of  study   activities:   Delays  in  construction  and  latrine  handover  in  intervention  sites: Construction  delays  at  the  beginning  of  the  project  limited  the  number  of  compounds  enumerators   were  able  to  visit  and  made  field  work  less  efficient  overall.  These  delays  resulted  in  slower  than   projected  recruitment  in  the  first  months  of  enrolment  as  the  rate  of  recruitment  was  dictated  by  the   rate  of  construction.  However,  over  time  there  were  significant  improvements  in  the  rate  of   construction,  with  the  last  quarter  of  enrolment  proceeding  much  faster.  Uncertainties  are  inherent  in   latrine  construction  and  handover  to  the  community,  and  it  was  necessary  to  manage  the  pace  of  the   field  work  to  reflect  this.         Uncertainty  with  regard  to  the  No  Cost  Extension  (NCE): The  study  protocol  specified  a  12-­‐month  follow  up  period;  however  as  the  prime  contract  was  signed  in   October  2014,  backdated  to  August  2014  and  ended  in  April  2016,  it  did  not  allow enough  time  for   follow  up.   It  was  agreed  with  URC  at  that  time  that  a  NCE  would  be  granted  as  a  matter  of  course  in   early  2015  so  that  study  could  be  implemented  as  per  the  approved  protocol,  but  the  NCE  was  not   actually  granted  until  late  October  2015.  As  a  result  all  the  sub-­‐contracts  with  sub-­‐awardees  had  to  be   negotiated  again  and  budgets  and  workplans  had  to  be  adjusted  accordingly.  This  placed  pressure  on   administrative  and  financial  resources,  and  created  a  high  burden  of  expenditure  and  task  completion     required  in  the  final  quarter  of  the  project  (January  – March  2017).  The  three  key  areas  of  work  during   this  period  were:  completion  of the  endline  survey  as  per  the  timeline  and  ensuring  a  high  return  rate  of   stool  samples  from  children  that  provided samples  at  baseline;  finalisation  of data  analysis  for  the   preparation  of  the  scheduled  papers;  completion  of  financial  and  administrative  aspects  of  the  grant,   including  issuance  of  NCE  to  sub-­‐awardees.  In  addition,  closing  the  grant  before  completion  of  pre-­‐ specified  and  pre-­‐planned  study  activities  (i.e.  the  laboratory  analysis  of  samples,  statistical  analysis  of   the  final  datasets,  and  the  preparation  of  key  manuscripts)  presented  challenges  for  the  study  team  in   ensuring  that  these  activities  could  be  completed  after  the  project  closedown.   23 CONCLUSIONS AND RECOMMENDATIONS This  study  has  successfully  engaged  policy,  practice  and  academic  audiences  throughout  the  duration  of   the  study  period.  This  is  evidenced  by  the  high  number  of  manuscripts  developed  for  publication  and   presentations  given  at  key  public  health  and  WASH  conferences;  and  by  successfully  attracting   additional  funding  streams  to  complement  and  further  the  research  objectives. Key  activities  were  completed  as  specified  by  the  grant  in  time  for  project  close-­‐down  in  April  2017.   These  included:  completion  of  endline  survey;  preparation  of  scheduled  papers  and  the  completion  of   financial  and  administrative  requirements.   Preliminary  analysis  of  baseline  characteristics  for  intervention  and control  compounds  suggested  that   the  study’s  matching  strategy  yielded  reasonable  balance,  although  this  will  be  analysed  further  over  the   next  few  months.    Baseline  disease  burden  was  even  higher  than  estimated;  demonstrating  the  critical   need  for  an  evidence-­‐based  intervention.  Formal  analysis  of  endline  health  status  remains  to  be   conducted;  however,  preliminary  analysis  suggests  prevalence  of  coinfection  and  STH  infection   remained  high  in  both  intervention  and  control  arm.  No  inference  should  be  drawn  as  to  the  effect  of   the  intervention  until  formal  analysis  is  undertaken.   NEXT STEPS Formal  analysis  of  baseline  and  endline  characteristics  will  be  completed  in  late  2017.  Additional   research  will  investigate  the  social-­‐behavioural  impact  of  the  intervention,  and  a  planned  follow-­‐up   survey  at  24-­‐months  post  intervention  is  planned  to  assess  impact  on  child  nutritional  outcomes.