Neural tube defects (NTDs) are a common birth defect responsible for spina bifida and anencephaly. They are caused by genetic and environmental factors, including vitamin deficiencies, pollutants, and pharmaceuticals. In May of 2006, NTDs were observed in untreated control mice. The cause was traced to the drinking water provided to the mice. We investigated possible sources of the teratogen to narrow the potential list of contaminants. To determine if the teratogen formed in the water bottles as a result of bacterial metabolism, we compared NTD rates in mice provided sterilized water daily with mice receiving weekly water changes. NTD rates were not significantly different between the daily (17.4%) and the weekly (19.5%) groups. Thus, the teratogen is not likely caused by bacterial metabolism in the mouse's water bottle. To determine where the teratogen might enter the water supply; we compared NTD rates in mice provided water from 5 locations along the watercourse supplying water to the treatment plant. The stream begins in unpopulated National Forest land and runs through both rural and urban areas. We found no significant difference in NTD rates in mice provided water from the 5 locations, with rates ranging from 16 – 27%. From this, it appears that the teratogen is widely distributed in the environment. This work was funded by the VA‐MD College of Veterinary Medicine and NIH‐NIEHS R21ES016886.

Previous research has demonstrated increased rates of neural tube defects, cleft palates and rib malformation in CD‐1 mice exposed to tap water from a variety of municipalities. Exposure to tap water resulted in neural tube defect rates of 10–20%. Rates of cleft palates and rib malformations were 5% and 70% respectively. This research investigated whether analogous findings could be demonstrated in a second animal model, specifically Sprague Dawley and Wistar rats. Female rats were maintained on tap water for 12 weeks prior to breeding and throughout gestation. Resulting embryos were evaluated on day 18 of gestation for fetal weight, placenta weight, and neural tube, palate, facial, ocular, tail, limb and digital malformations. The fetal skeleton was then assessed using tissue clearing and staining with Alcian Blue – Alizarin Red. Both strains of tap water exposed rat embryos demonstrated no significant gross or skeletal malformations at gestational day 18. Lumbar ribs were observed in 8.3% of tap water exposed Sprague Dawley fetuses; however, this is similar to background rates for this strain. These results suggest that rats may be more resistant than mice to the possible tertogen present in tap water. This work was funded by the VA‐MD College of Veterinary Medicine and NIH‐NIEHS R21ES016886.

In May of 2006, we began to observe neural tube defects (NTDs ‐ a common birth defect resulting in spina bifida and anencephaly) in untreated mice. The longer the mice were housed, the greater the incidence of defects. The cause of these defects was eventually traced to the municipal tap water provided to the mice. Rearing mice for 2 generations (F2 mice) on distilled deionized (DDI) water eliminated the NTDs. When the mice were placed back on tap water from 4 different municipalities, their offspring developed NTDs again. We hypothesized that the contaminant in the tap water could induce other fetal malformations as well. To test the hypothesis, F2 mice were exposed to tap water or DDI for 8 weeks before breeding and throughout gestation. Gestational day 17 embryos were examined for eye, tail, limb, palate, and craniofacial malformations. The embryos were then cleared and stained to assess axial skeleton malformations such as number of ribs and vertebrae, fused, transitional, hemi, and supernumary vertebrae/ribs. Tap water exposure significantly increased cleft palates (5.0% vs. 1.6%), rib malformations (68% vs. 30%) and number of resorptions (7.5% vs. 5.0%) compared to DDI. Analysis of the tap water is underway to identify the teratogen and possible risks to humans. This work was funded through the Harvey Peter's Foundation; Institute for Critical Technology and Applied Science (ICTAS); and NIH grant K01RR16241.