BACKGROUND: Bactrocera correcta is a quarantine pest that negatively impacts the fruit and vegetable industry. Differentiating B. correcta from similar species, especially in non-adult stages, remains challenging. Rapid molecular identification techniques, such as recombinase polymerase amplification (RPA) combined with CRISPR/Cas12a and multienzyme isothermal rapid amplification with lateral flow dipstick (MIRA-LFD), play a crucial role in early monitoring and safeguarding agricultural production. Our study introduces two methods for the rapid visual identification of B. correcta.
RESULTS: Bactrocera correcta specific RPA primers, CRISPR RNA (crRNA), and the LFD probe were designed based on the cox1 genes. The RPA reaction conditions were optimized (at 37 °C for 8 min) for effective template DNA amplification. Two nucleic acid detection methods were established to visualize RPA. In the RPA-CRISPR/Cas12a system, the optimal LbCas12a/crRNA concentration ratio was 200:400 nmol L-1. Successful amplification was determined by the presence or absence of green fluorescence following 15 min incubation at 37 °C. The MIRA-LFD system achieved precise identification of the target species within 4 min at 37 °C. Both methods exhibited high specificity and sensitivity, allowing for detection from 1.0 × 10-1 ng μL-1 of DNA. Combined with rapid DNA extraction, rapid identification of individual B. correcta at different developmental stages was achieved, enhancing the practicality and convenience of the established methods.
CONCLUSIONS: Our research findings demonstrate that both the RPA-CRISPR/Cas12a and MIRA-LFD methods for B. correcta detection was accurate and rapid (within 30 min and 10 min, respectively), at 37 °C. Our methods do not rely on expensive equipment, thus possess high practical value, providing improved identification solutions for port quarantine pests and field applications. © 2024 Society of Chemical Industry.

The guava fruit fly, Bactrocera correcta, is one of the most destructive pests in the genus Bactrocera and detects environmental odorants mainly through antennal olfactory sensilla phenotypes with nanopores. However, it is unclear whether there are naturally occurring abnormal antennal olfactory sensilla phenotypes that affect olfaction. Here, we found that there were abnormal bulges besides nanopores on the surface of trichoid and basiconic olfactory sensilla in the antennal flagellum of long-term laboratory rearing colony (LTC), and that nanopore number in these olfactory sensilla was also remarkably reduced. Notably, the electroantennogram (EAG) responses of LTC insects to methyl eugenol or β-caryophyllene were inhibited, and their behavioral responses elicited by the same odorants were also impaired. These results revealed naturally occurring abnormal antennal olfactory sensilla phenotypes which were involved in olfactory deficit in B. correcta, providing a platform to further study nanopore-targeted pest control technologies in the future.

In insects, imaginal disk growth factors (IDGFs), an important component of the glycoside hydrolase 18 (GH18) family of chitinases, have been reported to be associated with the maintenance of the cuticle and molting. However, there is little knowledge of their function. In this study, imaginal disk growth factor 6 (Idgf6), which is an Idgf, was first identified and cloned from the guava fruit fly Bactrocera correcta (Bezzi) (Diptera: Tephritidae), one of the most serious pest insects in South China and surrounding Southeast Asian countries. This gene encodes IDGF6 protein with a conserved domain similar to ChiA chitinases, the glycoside hydrolase 18 (GH18) family of chitinases, according to NCBI BLAST. Phylogenetic analysis indicated that all Idgf6s were highly conserved among similar species. Subsequent temporal expression profiling revealed that Idgf6 was highly expressed in both the late-pupal and mid-adult stages, suggesting that this gene plays a predominant role in pupal and adult development. Furthermore, RNA interference experiments against Idgf6 in B. correcta, which led to the specific decrease in Idgf6 expression, resulted in larval death as well as adult wing malformation. The direct effects of Idgf6 silencing on B. correcta indicated its important role in development, and Idgf6 might be further exploited as a novel insecticide target in the context of pest management.

Bactrocera correcta (Diptera: Tephritidae) is one of the most serious pest insects in south China and surrounding Southeast Asian countries. The family Tephritidae includes over 4257 species distributed worldwide, so the complete mitochondrial genome would be helpful for bio-identification, biogeography and phylogeny. The B. correcta genome consists of 15 936 bp. Annotation indicated that the structure and orientation of 13 protein-coding genes (PCGs), 22 tRNA and 2 rRNA sequences were typical of, and similar to, the ten closely related tephritid species. The nucleotide composition shows heavily biased toward As and Ts accounting 73.2% and exhibits a slightly positive AT skew, which is similar to other known tephritid species and other insects. The phylogenetic tree indicated the presence of three distinct families (Tephritidae, Muscidae, Drosophilidae) in Order Diptera.

This laboratory-based study examined the effects of low-temperature phosphine fumigation on the survival of the eggs and larvae of the guava fruit fly, Bactrocera correcta (Bezzi). Individual flies at different developmental stages, from 6-h-old eggs to third instars, were exposed to 0.92 mg/liter phosphine for 1-7 d at 5°C. We found that 12-h-old eggs and third instars were the most tolerant to phosphine. Increasing phosphine concentrations from 0.46 to 4.56 mg/liter increased mortality in these two stages. However, increased exposure times were required to achieve equal mortality rates in 12-h-old eggs and third instars when phosphine concentrations were ≥4.56 and ≥3.65 mg/liter, respectively. C(n)t = k expression was obtained at 50, 90, and 99% mortality levels, and the toxicity index (n) ranged from 0.43 to 0.77 for the two stages. The synergistic effects of a controlled atmosphere (CA) with elevated CO(2) levels were also investigated, and we found that a CO(2) concentration between 10% and 15% under CA conditions was optimal for low-temperature phosphine fumigation.

It is generally believed that widely distributed species differ in their thermal plasticity from narrowly distributed species, but how differences in thermal plasticity are regulated at the molecular level remains largely unknown. Here, we conducted a comparative study of two closely related invasive fruit fly species, Bactrocera correcta and Bactroceradorsalis, in China. The two species had overlapping distributions, but B. dorsalis had a much wider range throughout the country and a longer invasive history than B. correcta. We first examined the effects of thermal acclimation on the ability of the two fruit flies to survive heat stress. The heat shock tolerance of B. dorsalis was significantly enhanced by heat hardening at 35, 37, 39 and 41°C, but that of B. correcta was only enhanced by heat hardening at 39°C and 41°C. Thus, the more widespread species has a higher thermal plasticity than the narrowly distributed species. We then determined the expression of Hsp70 and Hsp90 during different developmental stages and their responses to thermal hardening. The expression of both Hsp70 and Hsp90 in larvae was upregulated in response to heat hardening, starting at 35°C for B. dorsalis and at 39°C for B. correcta. The two species exhibited a highly consistent pattern of thermal response in terms of their heat shock survival rates and levels of Hsp gene expression. The results suggest that the difference in thermal plasticity may be responsible for the different distributions of the two species and that Hsp expression may be involved in the regulation of thermal plasticity. Our findings have important implications for the prediction of the thermal limits and ecological responses of related species in nature.