Claus Henn et al. development than expected based on effects of exposure to either metal alone. Interestingly, this interaction was observed for 12-month but not for 24-month blood metal concentra- tions. A possible interpretation of the speci- ficity of the interaction of 12-month blood manganese–lead levels is that this is a sensitive developmental window specific to this inter- action. From repeated measures models, for a one-unit increase in lead at 12 months, MDI scores were 2.16 points lower among chil- dren with high manganese (–0.89 + –0.07 + –1.27 = –2.23), compared with children with midrange manganese [–0.07 (95% CI: –0.39, 0.25)]. PDI scores for a one-unit increase in lead at 12 months were 0.97 points lower among children with high manganese (–0.05 + –0.27 + –0.92 = –1.24), compared with children with midrange manganese [–0.27 (95% CI: –0.56, 0.02)]. Although these dif- ferences represent a small decrease in MDI and PDI scores for an individual, they may have large implications for the population level, because the entire distribution of neuro- development scores may shift, increasing the number of children with neurodevelopmental deficits that are of clinical concern. The association between lead and Bayley scores was similar among children with low versus midrange manganese levels (i.e., no evidence of interaction). Although children in the lowest quintile of blood manganese had lower 12- and 18-month MDI scores than did children in quintiles 2–4, this association was observed only in the absence of a change in blood lead [manganese quintile 1, adjusted β = –2.86 (95% CI: –5.52, –0.21) and –3.36 (95% CI: –5.98, –0.74) at 12 and 18 months, respectively]. An independent effect of low manganese is consistent with its function as an essential nutrient, because manganese is a key component of various metalloenzymes needed for normal central nervous system function and protection against oxidative injury (Aschner and Aschner 1991). The lack of an interaction with lead at low manga- nese levels may be because the mechanisms by which manganese deficiency causes adverse effects may not overlap with those for lead, unlike manganese toxicity. Alternatively, the lowest levels observed in our population may not be low enough to produce an effect. In another epidemiologic study that exam- ined manganese–lead coexposure, Kim et al. (2009) reported significant inverse associations of lead with full-scale and verbal IQ among school-age children with manganese concen- trations > 14 μg/L but not among children with levels < 14 μg/L. Although manganese and lead concentrations were lower than in our study [mean levels of 1.7 gμ/dL lead and 14.3 gμ/L manganese in Kim et al. (2009) vs. 5.0 gμ/dL lead and 21.5 gμ/L manganese for 24-month blood metals in the present study], 130 the results are consistent across the two stud- ies. An interaction between manganese and lead has also been seen in laboratory studies. In adult rats, exposure to manganese and lead resulted in synergistic effects on in vivo dop- amine release (Rodriguez et al. 1998). In other laboratory studies, manganese exposure poten- tiated lead effects on learning and behavioral patterns and increased accumulation of met- als in rat brains (Chandra et al. 1981, 1983; Shukla and Chandra 1987). Kalia et al. (1984) proposed that the presence of excess manga- nese in the brain may increase the binding affinity of brain proteins to lead or may induce expression of a lead-binding protein, resulting in more toxicity than expected based on effects of exposure to lead alone. A direct comparison of manganese levels in our study with other studies is not possible because, to our knowledge, our study is the first to measure blood manganese in 12- and 24-month-olds. The average blood manganese levels among our participants are lower than average cord blood manganese reported previ- ously by Takser et al. (2003) (38.5 gμ/L) and by Zota et al. (2009) (40 μg/L) and higher than levels reported in 10-year-old children (12.8 gμ/L) (Wasserman et al. 2006). This is consistent with evidence of declining manga- nese with increasing age (Chan et al. 1992; Collipp et al. 1983; Spencer 1999). Our previous work, which reported lower MDI scores among children with both low and high manganese after adjusting for lead, found the strongest manganese associations at 12 months (Claus Henn et al. 2010). This asso- ciation was not seen at later ages, suggesting a diminution over time. When lead and man- ganese exposures occur jointly at 12 months; however, the detrimental impact may be more enduring, because we see decrements in Bayley scores of similar magnitude at all ages stud- ied. When results from both of our studies are taken together, it appears that, although blood manganese is associated with deficits in concurrent 12-month neurodevelopment test scores, a subset of children with lead coex- posure have adverse manganese effects that persist. Because environmental manganese and lead frequently occur together in urban and/or contaminated areas (Aelion et al. 2009; Joselow et al. 1978), these results, if confirmed in other studies, may have implications for including the joint impact of chemicals in risk assess- ment and public health interventions rather than focusing on the independent effects of individual toxicants. Limitations. We did not conduct a formal exposure assessment of these children and are thus unable to identify sources of metals expo- sure. However, blood lead levels among chil- dren residing in Mexico City during the time of this study (1997–2000) have been asso- ciated most strongly with the use of leaded gasoline and lead-glazed ceramics (Romieu et al. 1995). Elevated blood manganese levels likely arise from exposure to manganese in air, drinking water, or dietary sources, but differences in manganese levels among chil- dren with the same environmental exposures may also arise from variability in metabolic function that alters retention/excretion of internal manganese. Although it is unclear which biological matrix best represents man- ganese exposure, blood manganese is believed to be a reasonable indicator of environmental exposure and body burden (Agency for Toxic Substances and Disease Registry 2008). Bayley scores may seem low relative to the expected mean score of 100 in a standard U.S. population. However, the BSID-IIS used in this study are not normalized to the Mexican population. Scores should therefore not be interpreted as low or be compared with other populations. Test results are internally valid and may be used to compare scores within this population but are not generalizable to other populations, which is a limitation of the study. We lack data on socioeconomic status (SES) and quality of the home environment, two potential confounders of the relation- ship between manganese–lead and neuro- development. In the context of the present study, if social environment is associated with lead–manganese exposure and devel- opment, it may confound the “interaction” of these two metals. However, adjusting for maternal education, which is strongly cor- related with SES among families in Mexico (Bronfman et al. 1988), likely addresses, at least in part, potential confounding by this variable. Furthermore, both SES and home environment are social influences that may also represent exposure opportunity. If these exposures and these variables are intertwined, adjusting for SES or home environment may mask the complete effects of exposure and may be inappropriate (Bellinger 2008b, 2009). Nonetheless, unmeasured and residual confounding remains possible. We also lack data on prenatal blood metals concentrations, which prevented us from assessing whether exposure before birth is more predictive of neurodevelopment in early childhood. There may be concerns about selection bias due to the imbalance in the numbers of children for whom blood samples were avail- able. Although some children provided only one sample whereas others provided two, we found no difference in major characteristics of these two groups (data not shown). However, we did observe a difference between 24-month metals interaction effect estimates for PDI based on all children with a 24-month sam- ple and those based on children with both a 12- and 24-month sample. Although no differences in effect estimates were observed for MDI or for 12-month metals, we cannot ovlume 120 | number 1 | January 2012 • Environmental Health Perspectives