RESEARCH REPORT © 2014

INFLUENCE OF DIETARY AFLATOXIN CONTAMINATION ON AFLATOXIN LEVELS IN BREAST MILK AMONG LACTATING MOTHERS IN KIKUMBULYU LOCATION, KIBWEZI DISTRICT OF MAKUENI COUNTY PRESENTER:OTIENO ISAAC OGALLO REG:A56/83054/2012 SUPERVISORS: DR.ALICE M. MWANGI MR.P.LAMUKA PROF.G.CHEMININGWA

INTRODUCTION Aflatoxins is of fungal origin (mycotoxins). Geographic, climatic, agricultural and susceptibility of food commodities. Passed to body tissues and fluids eg breast milk, urine and blood. Dietary exposure of lactating mothers also puts breast feeding infants at equal risk of aflatoxin exposure. Severe outbreaks have been reported in eastern region of Kenya, hence selected study area.

Aflatoxin Contamination PROBLEM STATEMENT Eastern region of Kenya prone to aflatoxin contamination. Makueni County reported the highest and severest incidences of contamination in maize, Kenya’s main staple Kilonzi, 2011 found the following in Kibwezi: *Recommended limit = 10µg/kg Little still known on aflatoxin contamination in human breast milk in Kenya. And the influence of these high levels of dietary contamination on the levels in human breastmilk is not clear Hence dietary maternal to child aflatoxin exposure through breast milk could be an under-evaluated risk factor in Kenya. Food Item Aflatoxin Contamination % of Samples Levels* (ng/kg) Maize kernels 45 18000 - 480000 Muthokoi 20 12000-123000 Maize 30 6000–30000

JUSTIFICATION Recurrent incidences of aflatoxin contamination, outbreaks of aflatoxicosis in Eastern region of Kenya. There is severe toxicity even when taken at lower levels both in diet and milk. As a consequence, deleterious health effect such as carcinoma, hemorrhage, stunting, and immune-suppression has been reported in children. Hence the need to carry out the study.

Aim of the study Purpose of the study To contribute towards improvement of food safety as well as maternal and infant and young child health during the lactation period. Purpose of the study To generate information on the current situation on the levels of aflatoxins in food and breast milk among lactating mothers from Makueni County as a basis for emphasizing the constant need of monitoring aflatoxin occurrence in at risk regions.

Main objective To determine the influence of dietary aflatoxin contamination on aflatoxin levels in breast milk among lactating mothers in Kkumbulyu Location, Kibwezi District of Makueni County. SPECIFIC OBJECTIVES To describe the demographic and socio-economic status of the lactating mothers in the study. To determine the food consumption patterns and dietary diversity of the lactating mothers in the study. To determine the breast feeding practices of the breast fed children between 0 to 6 months

Cont’d 4.To determine the levels and types of aflatoxins in foods susceptible to aflatoxin contamination and are mostly consumed by the lactating mothers in the study. 5.To determine the types and levels of aflatoxins in the breast milk of lactating mothers, and in the urine of the breast fed children between 0-6 months in the study.

Study Hypotheses No significant association; Socio-demographic characteristics of the lactating mothers and the levels of aflatoxin in the breast milk. Food consumption pattern and dietary diversity of the lactating mothers and the levels of aflatoxin in the breast milk and urine of breastfeeding children. Levels of aflatoxin among foods that are susceptible to aflatoxin contamination.

LITERATURE REVIEW General overview of aflatoxins Types, Structures and Properties of Aflatoxins Factors Promoting Aflatoxin Occurrence in the World Incidences of Aflatoxin Outbreak Global Occurrence Aflatoxin Occurrence in Kenya Food, Dietary Diversity and Aflatoxin Aflatoxin in Human Breast Milk Maximum Set Levels for Aflatoxins Nutritional and Health Consequences Related to Aflatoxin Contamination

METHODOLOGY Study Design Study Setting Kikumbulyu Location, Kibwezi District, Makueni County. Makueni County is located in Eastern region of Kenya and covers an area of 8008.9 sq Km Study Design A descriptive and analytical cross-sectional study.

Study Population Lactating mothers and breastfeeding children below 6 months old Sample size Fischer et al.(1992) formula N=Zpq⁄d where; Z=1.96, p= 50.0%, d=5% =384 + 4 % attrition rate N=399≈400 200 paired lactating mothers and breastfeeding children below 6 months of age

Sampling Procedure: Multistage

Inclusion Criteria Available and willing lactating mothers with children between 0-6 months Exclusion Criteria Lactating mothers with disease of the breast or with breast complications such as breast pain, breast engorgement, nipple pain, milk stasis, mastitis and an overactive let down Eligible but unavailable and/or unwilling mothers

Data Collection Tools and Equipment The main study tool used was a semi-structured questionnaire Equipment for aflatoxin analysis ; MaxSignal ® ELISA kit High Performance Liquid Chromatography (HPLC). Data Collection Recruitment and training of research assistants Pretesting of the data collection tools and equipment

Data Collection Procedure Demographic and socio-economic status Semi-structured pretested questionnaire administered to 200 lactating mother. This was done through interview. Dietary diversity, food consumption patterns, consumption levels and breast feeding practices Semi-quantitative food frequency questionnaires administered. 24 hour Dietary Diversity for the lactating mothers was also done.

Sample collection Food samples Purposive sampling . Foods susceptible to aflatoxin contamination (maize ugali, githeri, muthokoi and maize porridge) collected. Available during data collection were collected. A total of 400 food samples were collected.

Cont’d-Sample collection Breast milk samples 100 Breast milk samples . Hand expressed 10ml of breast milk. Urine samples 100 Urine samples collected from breast feeding children between 0-6 months belonging to mothers’ whose breast milk samples were collected. Mainstream urine, wet their under wrappings (napkins or clothing). wrung the under-wrappings, let the urine drip into plastic urine containers .

Data Management and Analysis Data Quality Assurance Data quality assurance across all the stages of study Adequate training of field assistants (community health workers) Pretesting of data collection tools and standardizing data collection procedures. Close supervision of field assistants. Contamination was minimized during sample and specimen collection by using sterilized gloves and air tight cryovial tubes. Strict hygiene observed during specimen collection

Data entry into SPSS version 20 is still on progress. Statistical Analyses Data entry into SPSS version 20 is still on progress. Objective 1: Socio-demographic characteristics of the lactating mothers Descriptive statistical analyses will be done. ANOVA Chi-square Regression test (inference statistical analysis)

Descriptive statistical analyses. Objective 2/3: Food consumption patterns and dietary diversity of the lactating mothers, and breast feeding practices of children between 0-6 months in the study Descriptive statistical analyses. Multiple –ANOVA . Regression analysis .

Objective 4: Types and levels of aflatoxin in foods that are mostly consumed and are likely to be contaminated with aflatoxin among lactating mothers in the study. Descriptive analysis. ANOVA. One sample t-test/one way ANOVA. Multiple correlations between food and corresponding aflatoxin levels.

Descriptive statistical analysis. Objective 5:Types and levels of aflatoxins in breast milk of lactating mothers and urine of breastfeeding children below 6 months of age. Descriptive statistical analysis. ANOVA. One sample t-test/one way ANOVA. Correlations.

Generate a predictor model of aflatoxin levels in breast milk from independent variables which significantly contribute to the influence of aflatoxin in breast milk.

Thank You