Numerous health disorders are associated with human occupational exposure to organophosphate (OP) pesticides and these pesticides impose serious public health problems. Agricultural workers are at high risk of exposure to hazards of OP pesticides. Evaluation of the pesticide effects on hematological, serological and immunological parameters is necessary to determine the adverse effects of excessive use of pesticides. In the presentstudycomplete blood count, liver enzyme level, acetylcholinestrase activity, serum immunoglobulin, pesticides residues level, and molecular detection of tuberculosis among agricultural workers of District Gujranwala, Punjab, Pakistan, were investigated. Data were collected through questionnaires from 200 professional agricultural spray workers and 100 agricultural non-spray workers (control) ranging in age between 20 to 60 years. The most commonly reported signs and symptoms were feeling of vomiting (males 22%, females 42%) and skin irritation (males 32%, females 28%)after spraying. Generally, inadequate protective measures were observed by these pesticidal spray workers : only 16.5% male and 32% female workers used gloves while 12.5% males and 6% females used face masks during their field spray work.The blood parameters, such as hemoglobin, hematocrit, MCV, MCH, MCHC, of spray workers and non-spray workers were also investigated. WBC and RBC levels of male sprayers and non-sprayers were statistically significantly (P < 0.05) different. In female spray workers, the WBC and neutrophils levels were significantly different as compared to non-spray workers. Overall, in both male and female pesticides-exposed spray workers, mean values of ALT and AST were higher as compared to nonexposed workers. This research revealed abnormal level of acetylcholinestrase (AChE) due to the exposure to multiple pesticides; serum AChE activity level was reduced in male as well as female spray workers as compared to the corresponding control subjects. Mean unit value of AChE in male spray workers was 0.16 U/ mL (min = -0.10 U/ mL and max = 0.66 U/ mL) and in non-spray workers was 0.44 U/ mL (min =0.01 U/ mL and max = 0.93 U/ mL); these values were significantly (P < 0.05) different statistically. Similarly, highly significant differences (P< 0.05) was observed in female spray workers, i.e., 0.41 U/ mL (min = 0.06 U/ mL and max = 0.93 U/ mL) and non-spray workers : 0.65 U/ mL (min =0.32 U/ mL and max = 1.1 U/ mL). ix Immunotoxicity in spray workers with a significant difference of IgA and IgM (P = 0.033 and P = 0.000) in male spray workers and non-spray workers, respectively, was also observed. Significant differences of immunoglobulin IgM and IgA (P = 0.000) and P = 0.000) were also observed in female spray workers and non-spray workers. Non-significant difference was observed for IgE (P = 0.928) level in male spray workers and non-spray workers. Likewise, non-significant difference was also observed for IgE (P= 0.425) levels in female spray workers and non-spray workers.Molecular detection of Mycobacterium tuberculosis (M. tuberculosis) and Mycobacterium bovis (M. bovis) was carried out by targeting histone-like protein (hupB) gene in cultured sputum samples of spray workers.Mycobacteriumtuberculosis was detected in 15% male sprayers and 36% female sprayers, while M. bovis was detected in 5% male sprayers and 10% female sprayers. A chi-square test indicated thata significant difference (P < 0.05) existed between positive and negative M. tuberculosis and M. bovis in both male and female spray workers. The susceptibility of pesticides-exposed spray workers to tuberculosis and alterations in hematological and liver enzymes is crucial for health. Results indicated that blood serum samples of spray workers were contaminated with various OPs. Among spray workers (n=200) the OP insecticide, chlorpyrifos, was detected in 18 (12%) male blood serum samples and 7 (14%) female blood serum samples, whereas profenophos was detected in 9 (6%) male serum samples and 3(6%) blood serum samples of females spray workers. Pesticides residues of imidacloprid were detected in 25 (16.6%) males and malathion in 13 (8.6%) males and 3 (6%) females, whiledichlorovos was detected in 2 (4%) females. The OP pesticides detected in the serum samples of spray workers were chlorpyrifos (1.4μg/L), profenophos (1.29 μg/L), imidacloprid (0.14 μg/L) and malathion (1.40 μg/L); dichlorovos was not detected in male spray workers.In female spray workers, chlorpyrifos (1.39 μg/L) profenophos (1.23 μg/L), and malathion (1.40 μg/L) anddichlorovos (0.03 μg/L) were detected but imidacloprid was not detected in blood serum of spray workers, respectively.From the results of the present study it can be concluded that frequent use of highly toxic pesticides, inadequate or no use of personal protective equipment and poor hygienic practices are the main reasons for the symptoms of pesticides toxicity in spray workers.