Bacillus thuringiensis

Nascimento, Tatiane Aparecida, Cruz, Igor Luiz Souza da, Carvalho, Karine Silva de, Valicente, Fernando Hercos, Santos, Maria José Paes & Queiroz, Margareth Maria de Carvalho, 2024, Screening for new Bacillus thuringiensis (Caryophanales: Bacillaceae) strains effective against Aedes aegypti larvae (Diptera: Culicidae), Revista Brasileira de Entomologia (e 20240021) 68 (4), pp. 1-11 : 3

Pathogenicity of Bacillus thuringiensis

Among the 13 Bt strains tested, 1644, 1608A, and 1656 resulted in a mortality rate of over 70% for Ae. aegypti larvae . A Bti strain was used as the positive control in this study due to its proven efficacy against Culicidae of medical importance and its presence in various commercial products for mosquito control ( Davidson and Sweeney, 1983; Mohammad, 2022). Consequently, the BtiJAB strain caused 100% mortality in Ae. aegypti larvae . Similarly, strain 1644 was capable of causing 100% mortality in immatures within 24 hours. Up to 72 hours, strains 1608A and 1656 killed 95 and 85% of the larvae , respectively. The other strains exhibited low efficiency in controlling the target insect, with average mortality ranging from 0 to 53% ( Table 2).

During the evaluation, a distinct behavior was observed between the larvae in the control group and those treated with the BtiJAB strain (positive control) ( Figures 1C and 1D ), as well as strains 1644, 1608A, and 1656.The larvae contaminated by these Bt strains lost their agility, spending most of their time at the bottom of the plastic container without feeding, and rarely surfacing to breathe. The larvae slowed down their movements to a standstill, and their tegument changed color, becoming matte. The larvae became flaccid, and upon death, there was a darkening of the muscles and fatty body, indicating the onset of tissue deterioration. These changes may indicate the toxic effect caused by ingesting the spores and crystals of the Bt strains ( Figures 1C, 1E, 1G ).

This evaluation showed that the observed changes were dependent on the Bt strains tested. The Ae. aegypti larvae in the negative control group maintained a normal elongated and vermiform appearance, with a body visually divided into the head, thorax, and abdomen. The thorax was wider than the head and had tufts of bristles, which were also present on the abdomen ( Figures 1A and 1B ). According to Serra-Freire and Mello (2006), the general characters of the first body parts of Ae. aegypti larvae are globose, and the abdomen consists of 10 segments, with the eighth segment having a pair of spiracles located at the end of the siphon (tubular organ).

During exposure to the Bt strains, we observed that the Ae. aegypti larvae showed reduced agility, spending most of their time at the bottom of the container without feeding and rarely rising to the surface to breathe. Their movements gradually slowed to a complete stop, and their integument changed color, adopting a matte appearance. The larvae became flaccid, and, upon death, their muscles and fatty bodies darkened, initiating a process of tissue deterioration.This indicates the toxic effect resulting from ingesting the spores and crystals of the Bt strains ( Figures 1E, 1F, 1G, 1H, 1I, 1J ).

The main changes in larvae treated with Bt strains include a narrowing of the mesenteron, a decrease in body fat, thickening of the peritrophic membrane, leakage from the intestine, elongation of the cervix, and spacing of the anal papillae. The morphology of the anal papillae can affect the regulation of osmotic functions, influencing the survival of Ae. aegypti larvae ( Chaithong et al., 2006) . These observations are consistent with those reported by Fujiwara et al. (2017) and Lobato Rodrigues et al. (2021), who, in addition to observing lethargic movement of Culicidae larvae exposed to natural substances extracted from plants, noted the shortening and darkening of the abdomen and morphological changes in the anal papillae of Ae. aegypti .

Figures

Figure 1. Light photomicrographs of the morphological aspects of 3rd instar Ae. aegypti larvae. Larva from the control treatment showing highlighted structures, with no alterations to the head, segments of the abdomen, respiratory siphon and anal papilla (A and B). Larva exposed to the standard BtiJAB strain with changes in the segments of the abdomen (dark pigmentation) (black arrow), loss of external hairs,and slight spacing in the anal papillae (hollow arrow) (C and D).Larva exposed to strain 1644, showing a strong darkening in the trunk region and along the extremities of the body, accentuating the darkening in the anal papillae (black arrows), the toxic effect of this strain also induced spacing of the anal papillae (hollow arrow) (E and F). Larva treated with strain 1608A showing darkening of the intestine and extremities of the body (black arrows), in addition to the translucent tracheal system and spacing of the anal papillae (G and H). Larva exposed to strain 1656 showing a damaged intestinal tract (black arrow), spacing of the anal papillae (hollow arrow),partial extrusion of the intestinal contents, a desiccated larva and the exuvia of the instar attached to the respiratory siphon region (dotted arrow) (I and J). Note: A, abdomen; H, head; T, thorax; RS, respiratory siphon; AP, anal papillae; LH,lateral hairs.