Aleyrodidae fly is a vector carrier of various diseases in crops, plants, and vegetables. The present investigation evaluated the Aleyrodidae fly population for its relationship with cotton disease under different environmental conditions. Faisalabad, Chiniot, Sargodha, Jhang, Toba Tek Singh (T. T. Singh), and Burewala from Punjab, Pakistan were selected for this study. The Aleyrodidae fly population was found to be significantly different in selected regions, and endosymbionts of Hamiltonella, Rickettsia, Fritschea, Cardinium, Arsenophonus, and Wolbachia were recorded in promotions to the Aleyrodidae fly population. The Hamiltonella co-infection percentages in Faisalabad, Chiniot, Sargodha, Jhang, T. T. Singh, and Burewala were recorded as 51, 28, 75, 8, 23, and 25, respectively, whereas relative abundance percentages of Hamiltonella endosymbiont were recorded as 52, 35, 65, 5, 20, and 21 in Faisalabad, Chiniot, Sargodha, Jhang, T. T. Singh, and Burewala, respectively. A similar trend was observed for Rickettsia, Fritschea, Cardinium, Arsenophonus, and Wolbachia co-infection and relative abundances. Results revealed that the Aleyrodidae fly is a carrier of different endosymbionts, which were significantly different under different environmental conditions.