Gambusia holbrooki Girard, 1859 has established very strong populations in freshwater resources in Turkey after being introduced into Amik Lake (Hatay/Antakya) at the beginning of the 1920s. In this study, it was aimed to determine the female:male ratios and length-weight relationships of G. holbrooki populations in Turkey, which are considered as a threat, especially for endemic species. G. holbrooki specimen were sampled at total of 66 locations, including 39 lentic and 27 lotic locations, between 2016 and 2017. The female:male ratio was 1:0.39 for all of the investigated localities. Length-weight relationships were investigated at 44 sampling locations for the females and 34 sampling locations for the males. The sampled specimens ranged between 12.7 and 57.2 mm in total length, and 0.03 and 2.88 gin total weight. The length-weight relationship of the female and male G. holbrooki specimens indicated mostly isometric growth, whereas the b values varied between 1.755 and 4.009, and 2.154 and 3.665 for the females and males, respectively, in terms of the sampling locations.