# The Perpetual Vector Mosquito Threat and Its Eco-Friendly Nemeses
## 5 Eco-Friendly _Lysinibacillus sphaericus_ and _Bacillus thuringiensis_ subsp. _israelensis_
### Brief History of Lysinibacillus (Formerly Bacillus) sphaericus (Ls)
#### 5.1.2 Mechanism of Toxicity of Tpp1A1/Tpp2 Aa1  
The mechanism of toxicity of Tpp1 Aa1/Tpp2 Aa1 is unclear. Studies show that after exposure to the binary toxin, (i) _Culex quinquefasciatus_ larvae stop feeding within 4 h, and body paralysis occurs at 36 h, most likely due to neural and muscular tissue damage; (ii) the binding of Tpppp2A1 to the Cqm1 receptor is essential for internalization of Tpp1 Aa1/Tpp2 Aa1, likely mediated by an endocytic pathway; and (iii) a number of cytotoxic effects follow, including an increase in the lysosomal number and size, damage to the mitochondria, intense cytoplasmic vacuolization, destruction of the endoplasmic reticulum, and apoptosis, collectively leading to the destruction of the midgut microvilli [233; 234; 235; 236; 237; 238]. Transcriptome analyses of _Culex quinquefasciatus_ following intoxication with the binary toxin show a downregulation of genes related to metabolism and mitochondrial function, and an induction of genes coding for proteins linked to mitochondrial-mediated apoptosis, autophagy, and lysosomal compartments [239]. The collective data suggest that diverse pathways are involved in cytotoxic and tissue tropic (neuromuscular) malfunctions that culminate in larval death. Brief Historical Account of Bacillus thuringiensis Strains--A Collective of Highly Specific Insect Larvicides  
The first isolate of _Bacillus thuringiensis_ was discovered over 120 years ago in Japan by [PERSON], a sericultural engineer, in larvae of the silkworm moth (_Bombyx mori_), in which it caused a sudden lethal disease, bacillary paralysis [240]. Fourteen years later, [PERSON] [241] described a similar disease in larvae of the flour moth, _Ephesia kuhniella_, in Thuringia, Germany, hence the species name \"_thuringiensis_\". By the mid-1970s, based on the biological profiles of hundreds of isolates, at least thirteen subspecies were characterized. These Bt subspecies exhibited larvicidal activities against a broad range of lepidopterous (moth) and coleopterous (beetle) larvae, and a few were developed commercially as formidable eco-friendly biopesticides. These include Bt subsp. _kurstaki_ (Btk; Garden Dust, Caterpillar Killer, Dipel); _Bt_ subsp _aizauai_ (BtA; XenTari BT DF, Certan B 401), which targets lepidopteran pests; and Bt subsp. _morrisoni_ strain _tenebrionis_ (Btm), which is toxic to coleopteran pests [242]. Like Ls, _Bacillus thuringiensis_ is an aerobic Gram-positive spore-forming rod that is naturally present in many ecological niches, including soil, plants, stored products, aquatic environments, and insects and their habitats [243]. The bacterium is classified in the _Bacillus cereus sensu lato (sl)_, which contains at least 22 non-pathogenic and pathogenic species, with the most notable of the latter being _B. anthracis_[244]. Although these bacteria share a high degree of genetic identity, Bt is distinguished from other members of _B. cereus ls_ by the parasporal crystalline inclusions. These inclusions are composed of larvicidal protein protivas that are produced during the sporulation phase of growth. In addition to the established subspecies and strains, a growing list of new Bt isolates has led to the identification of novel protein toxins, which complicates their classification. Nonetheless, [PERSON] et al. [126; 220] have developed an informative conserved structure-based nomenclature system that includes over 1100 Bt crystalliferous and other bacteria-derived pesticidal proteins produced during sporulation and vegetative growth, including VIPs (vegetative insecticidal proteins) and Mtx (mosquitocidal toxins) initially isolated from _Lysinibacillus sphaericus_. Fifteen classes (Cry, Cyt, Vip, Tpp, Mpp, Gpp, App, Spp, Mcf, Mtx, Vpa, Vpb, Pra, Prb, and Mpf) have been defined based on conserved domain similarities, and a separate class (Xpp) has been set aside for pesticidal proteins with unknown or uncharacterized structures.