Isolation, characterization and larvicidal potential of indigenous soil inhabiting bacteria against larvae of southern house mosquito (Culex quinquefasciatus Say)

• Published in: International journal of tropical insect science Vol. 43; no. 3; pp. 781 - 791
• Main Authors: Iftikhar, Sohaib, Riaz, Muhammad Asam, Majeed, Muhammad Zeeshan, Afzal, Muhammad, Ali, Anam, Saadia, Mubshara, Ali, Zulfiqar, Ahmed, Sohail
• Format: Journal Article
• Language: English
• Published: Cham Springer International Publishing 01.06.2023; Springer Nature B.V
• Subjects: Apoptosis; Aquatic insects; Autophagy; Bacteria; Binding; Bioassays; Biomedical and Life Sciences; Culex quinquefasciatus; Developing countries; Entomology; Infectious diseases; Insecticide resistance; Insecticides; Insects; Larvae; LDCs; Life Sciences; Microorganisms; Microscopic analysis; Midgut; Mosquitoes; Original Research Article; Pore formation; Protein purification; Proteins; Public health; Receptors; Septicemia; Similarity; Soil bacteria; Soil microorganisms; Soils; Toxicity; Toxins; Trypsin; Vectors; Entomopathogenic bacteria; Cqm1; Binary toxins; Insecticidal toxin; Mosquito control
• ISSN: 1742-7592; 1742-7584; 1742-7592
• DOI: 10.1007/s42690-023-00992-x

Mosquitoes vector many infectious diseases and pose a serious threat to public health in developing countries. A wide array of synthetic insecticides is employed against these vectors. As the mosquitoes have got resistance to most of these chemical insecticides, there is a dire need to explore alternative control strategies such as entomopathogens. This laboratory study was aimed at isolating, purifying and evaluating the insecticidal proteins from indigenous strains of soil-inhabiting pathogenic bacteria against the larvae of southern house mosquito ( Culex quinquefasciatus ) and to identify the larval midgut receptor of bacterial toxin. One hundred bacterial isolates were isolated and screened. Only two isolates (AA-11 and AA-19) exhibited considerable larval mortality (> 80%) and were further identified by sequencing and blasting their 16  S rDNA gene fragments. Isolate AA-11 was identified as Pseudomonas sp. with 96% similarity, while AA-19 as Lysinibacillus sphaericus with 98% similarity. Microscopic analysis and gram staining of AA-19 confirmed its rod-shape structure and gram positive nature, respectively. It is likely that the bacterial isolate AA-19 was L. sphaericus . Furthermore, protein purification of this bacterial isolate revealed two proteins of approximately 51 and 43 kDa which reduced to 43 and 39 kDa respectively after digestion with trypsin indicating that these were Bin B and Bin A toxin proteins respectively of L. sphaericus . Results of toxicity bioassay revealed that the purified and activated toxin was highly toxic to C. quinquefasciatus larvae with a LC 50 value of 191 ppb. Pull-down assays demonstrated that one protein of 66.5 kDa was pulled down by CNBr-beads conjugated with insecticidal proteins (Bin B and A). The pulled down protein was considered as C. quinquefasciatus maltase 1 (Cqm1), a receptor of Bin toxin in the midgut of mosquito larvae. After binding with its receptor, it is probable that Bin toxin induced vaculation, pore formation, autophagy and apoptosis in the midgut cells of C. quinquefasciatus larvae to cause septicemia. Conclusively, the purified and activated insecticidal protein (Bin toxin) can be a key part of integrated vector management and has potential to be developed as commercial microbial formulation against mosquito larvae with a low risk to environment and non-target insects. The present study can be helpful to control the vectors of important diseases by understanding the interaction and binding of toxin and receptor.