The present work studies the environmental isotopes assess groundwater characteristics of the different parts of the main aquifer in the northeast Missan Province in south of Iraq. Water samples of groundwater and surface water were collected for two dry and wet seasons during the water year of 2011–2012. The study shows that most of the groundwater in the aquifer falls above the global meteoric water line, and all the samples fall below the Mediterranean meteoric water line, indicating that these samples are a mixture of two water types. The tritium content of these samples supports this conclusion. The overall conclusion of this study indicates that there are two sources of groundwater recharge in the studied area: the ephemeral streams (Teeb and Dewerge) and major precipitation sources. According to the tritium levels at or below one tritium unit (TU) obtained from the water, supply wells are highly confined or “not vulnerable”. Overall, the 3H results imply that recent recharge has taken place during the last four to five decades. In the recharge area, the high tritium content in the southern part of the Teeb area suggests that the recharge originates from rapid in?ltration of surface runoff. Therefore, the groundwater resources in the study area should be protected from contamination, because it will influence the aquifer in a relatively short period of time if any contamination enters the recharge areas of the aquifer.