EnterococciEnterococci but not but not E.coliE.coli counts in counts in drinking water are positively associated drinking water are positively associated

with gastrointestinal disease risk in urban with gastrointestinal disease risk in urban Zambia Zambia

UNC water and Health Conference 2016UNC water and Health Conference 20161313thth October, 2016 October, 2016

Michelo SimuyandiMichelo SimuyandiPhD candidate: LSHTM-ITD-DCDPhD candidate: LSHTM-ITD-DCD

Research Fellow: CIDRZResearch Fellow: CIDRZ

Outline

• Background• Methods• Results• Discussion• Conclusion

Background• Studies on the association between faecal indicator bacteria in drinking

water and diarrhoeal disease risk have yielded mixed findings that range from no association to significant associations1,2

• This variation could be due to:– high variability of water quality over time3

– Indicator organisms being weakly correlated to enteric pathogens4

– Study design related: collecting water quality and health data at the same time may fail to determine direction of causality 5

– Acquired immunity to certain diarrhoea causing pathogens could affect generalisation of the associations found in some studies6

– Other transmission routes of infection7

1. Gundry et al 20042. Gruber et al, 20143. Levy et al., 20094. Wu et al., 2011,5. Khush et al., 20136. Frost et al., 20057. Luby et al 2015

Study design

• We conducted a prospective study collecting repeated measures of water quality and health indicators and related covariates at both household and individual level in a peri-urban area south of Lusaka– As part of the pilot study of a field trial of a point

of use HWTS intervention

Aim

• To determine the association between the levels of Enterococci, E. coli and total coliforms in household drinking water and self-reported highly credible gastrointestinal illness (HCGI) experienced by household members using a 7 day recall period.

Definitions • The exposure was defined as

– Safe = <1 MPN/100 mL– Unsafe >1 MPN/100 mL

• Outcome measure used was self-reported highly credible gastrointestinal illness (HCGI) experienced by household members using a 7 day recall period– HCGI : defined as a report of at least one of: 1) vomiting; 2) watery

diarrhoea; 3) soft diarrhoea and abdominal cramps both occurring during recall period; 4 ) nausea and abdominal cramps both occurring during recall period (Colford et al., 2005)

• The associations between Enterococci, E. coli, total coliforms in household drinking water and the HCGI experienced by household members were investigated using mixed-effects logistic regression

Methods• Data was collect by

– household interviews using a survey,– household observations

• Testing of household stored drinking water and source water for bacterial indicators of faecal contamination was done using the IDEXX quanti-tray, MPN Methods– Enterolert® Test for Enterococci– Colilert® Test for E coli and total coliforms

FACTS/stats

• Intermittent water supply• unsafe sanitation• high fertility• High Unemployment rate• poor drainage

5 CEA Household clusters(1206 HH)

300 households recruited (60/cluster)

244 households included in analysis

244 households contributing to analysis

832 Individuals466 females

196 children below 5yrs

10 households lost to follow-up•8 relocated to area outside study area.•2 withdrew without reason

44 households excluded due to missing/incomplete data

Inclusion criteriaAt least a child below 5yrs, written

consent, purposive sampling till agreed 60 hh /cluster

Household Characteristic Number of subjects Proportion(%) (% of total) with HCGI (95% CI)1

Water QualityEcoli count (n=635)

<1 268(42.2) 42/339 (12.4) (8.8, 17.1)1-10 127(20.0) 15/180 (8.3) (4.1, 16.3)

11-100 98(15.4) 13/131 (9.9) (4.8, 19.4)101 + 142(22.4) 19/185 (10.3) (5.9, 17.2)

Total coliforms (Ecoli-media) (n=635)<1 81(12.8) 12/98 (12.2) (6.6, 21.5)

1-10 48(7.6) 12/65 (18.5) (8.2, 36.5)11-100 18(2.8) 2/40 (5.0) (1.3, 17.7)101 + 488(76.9) 63/632 (10.0) (7.4, 13.3)

Enterococci count (n=570)<1 529(92.8) 51/676 (7.5) (5.3, 10.6)

1-10 23(4.0) 5/23 (21.7) (6.2, 5.4)11-100 12(2.1) 3/12 (25.0) (9.1, 52.6)101 + 6(1.1) 2/7 (28.6) (8.2, 64.3)

Household Water Treatment (n=829)  No 661 (79.73) 89/692 (12.86) (9.92, 16,51)Yes 168 (20.27) 21/280 (6.98) (4.45, 10.77)

Primary Water Source (n=569) Tap inside house 53 (9.31) 5/67 (7.46) (3.94, 13.70)

Stand pipe 244 (42.88) 52/371 (14.02) (10.12, 19.09)Other 272 (47.80) 52/406 (12.81) (9.17, 17.60)

Containers per daya mean(SD) 3.16 (2.00)    Containers per day category (n=766)  

0-2 347 (45.30) 40/472 (8.47) (5.62, 12.59)3-5 307 (40.08) 62/468 (13.25) (9.86, 17.58)6-9 112 (14.62) 22/181 (12.15) (7.86, 18.33)

Distance to water source (n=791) <200 meters 762 (96.33) 128/1101 (11.63) (9.42, 14.26)200+ metres 29 (3.67) 1/47 (2.13) (0.26, 15.25)

How long to collect water (n=371)  5-14 minutes 306 (82.48) 40/468 (8.55) (5.74, 12.55)15+ minutes 65 (17.52) 11/85(12.94) (7.70, 20.99)

Household and individual baseline characteristics for the study population

Household Characteristic Number of subjects Proportion(%) (% of total) with HCGI (95% CI)1

How long to collect water (n=371)  5-14 minutes 306 (82.48) 40/468 (8.55) (5.74, 12.55)15+ minutes 65 (17.52) 11/85(12.94) (7.70, 20.99)

Safe Sanitationb (n=695) No 622 (89.50) 107/920 (11.63) (9.23, 14.55)Yes 73 (10.5) 11/112 (9.82) (4.50, 20.11)

Good Hygienec (n=821)  No 622 (75.76) 102/900 (11.33) (8.91, 14.30)Yes 199 (24.24) 29/290 (10.00) (6.56, 14.96)

Place to Wash hands in house (n=827) No 640 (77.39) 95/934 (10.17) (8.05, 12.78)Yes 187 (22.61) 37/262 (14.12) (9.15, 21.16)

Place to Wash hands near toilet (n=829)  No 721 (86.97) 106/1038 (10.21) (8.10, 12.80)Yes 108 (13.03) 26/160 (16.25) (10.00, 25.31)

Seasone Wet 86/928(9.27) (7.21, 11.83)Dry 46/273(16.85) (11.54,23.94)

Zone (n=832) Zone A 113 (13.58) 9/145(6.21) (2.67, 13.76) Zone B 172 (20.67) 41/229(17.90) (13.03, 24.09) Zone C 249 (29.93) 41/366(11.20) (7.49, 16.44) Zone D 176 (21.15) 17/254(6.69) (4.07, 10.82)Zone E 122 (14.66) 24/207(11.59) (7.22, 18.10)

Individual CharacteristicsSex (n=824)    

Female 358 (43.45) 82/677(12.11) (9.56, 15.23)Male 466 (56.55) 48/510(9.41) (7.09, 12.40)

Age Mean(SD) 19.47 (16.35) Age Category (n=825)

<5 years 196 (23.76) 37/268 (13.81) (10.03, 18.70)>= 5 years 629 (76.24) 95/925 (10.27) (8.10, 12.92)

Total 832 (100%) 132/1201 (10.99) (8.93, 13.45)

Water quality: A=Source vs HH stored and B =Storage container type

VariableVariable Number of Obs.Number of Obs.

(N=1,201) Odds Ratio 95% CI P-valueAge1 < 5 years 1,903 1.52 (0.95,2.44) 0.08Sex (female) 1,187 1.40 (0.92,2.11) 0.12Water quality

Unsafe E. coli count 270/835 0.59 (0.33,1.06) 0.08 Unsafe total coliforms 270/835 0.79 (0.35,1.79) 0.57

Unsafe Enterococci count 250/718 15.25 (2.27,102.27) 0.01

Treat Water1 993 0.41 (0.22,0.75) <0.01

Containers collected per day2 1,121 1.09 (0.96, 1.25) 0.19

Distance to Source > 200 meters2 1,148 0.16 (0.02,1.51) 0.11

>14 minutes to collect water2 553 1.85 (0.66, 5.14) 0.24

Narrow mouth container1 1,023 0.98 (0.53,1.84) 0.96

Good Hygiene2 1,190 0.85 (0.47, 1.54) 0.60

Place to wash hands in house2 1,196 1.46 (0.83, 2.60) 0.20

Place to wash hands near toilet2 1,198 2.12 (1.08, 4.17) 0.03

Safe Sanitation2 1,032 1.03 (0.40, 2.64) 0.95

Containers collected per day= number of 20 litre containers of water collected per day.Good Hygiene=Have soap (shown to interviewer) and household respondent asserts washes with soap. Safe sanitation=Household respondent asserts that the household uses a private latrine.1 As reported at visit corresponding to HCGI outcome. (i.e. “Did you treat this water you are currently drinking?”)2 As reported at baseline (first available visit for which response was available). Mixed effects logistic regression accounting for household and individual-level clustering. All associations with p-value < 0.05 are underlined

Univariate associations with Highly Credible Gastrointestinal Illness

(Unadjusted) Adjusted

Variable n Odds Ratio 95%CI P-value Odds Ratio 95%CI P-value

Unsafe Enterococci count 528 26.55 (1.45, 486.04) 0.03 31.33 (2.13, 461.73) 0.01

Age < 5 years 528 2.36 (0.95, 5.84) 0.06 2.92 (0.85, 10.03) 0.09

Treat Water 528 0.41 (0.14, 1.22) 0.11

Place to wash hands near toilet 528 1.87 (0.46, 7.52) 0.38

Mixed effects logistic regression (accounting for household and individual-level clustering) using a backwards selection procedure; nested models were evaluated using likelihood ratio tests with a p<0.1 threshold. ‘Treat water’ and ‘Place to wash hands near toilet’ were dropped from final model. P-values <0.05 are underlined.

Mixed effects logistic regression accounting for household and individual-level clustering

Discussion• Within the limitations of the study design, data and analysis

– Enterococci counts and not Ecoli and total coliforms is potentially associated with HCGI.

• Risebro et al (2012) reported a strong association between Enterococci counts and infectious intestinal disease as opposed to E.coli counts in England

– Reported household treatment of drinking water was negatively associated with HCGI only in the unadjusted analysis 0.41 (0.22,0.75) <0.01

– Place to wash hands near toilet was positively associated with HCGI only in the unadjusted analysis (OR 2.12 (1.08, 4.17)

Conclusion• Given higher percentage of samples that were

unsafe (E.coli and total coliforms) makes these potentially poor predictors (i.e., there is E. coli everywhere!

• In this urban setting, Enterococci counts have a potential association with HCGI than E.coli or total coliform counts.

• Need to validate Enterococci counts in this and other settings as stronger indicator for risk of HCGI

Acknowledgements • Funding

– Research funding NIH-through my supervisor: Joe Brown– Travel scholarship- BMGF through UNC Water & Health

Institute• Tropical Gastroenterology group

– Research host site/study research team• LSHTM-EHG

– Host research group for my PhD• Centre for Infectious Disease Research in Zambia

– Enteric Disease and Vaccines Research Unit• Employers, local PhD advisory team

• Misisi community– Study participants

Comments and Questions?