Comparative Proteomics Analysis of Phosphine-Resistant and Phosphine-Susceptible Sitophilus oryzae (Coleoptera: Curculionidae)

A proteomic method combining two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF) was used to compare the protein expression profiles of phosphine-resistant (PH3-R) and -susceptible (PH3-S) strains of Sitophilus oryzae. Thirty-nine differentially expressed protein spots were identified between the PH3-R and PH3-S strains; 20 protein spots were upregulated, and 19 protein spots were downregulated in the PH3-R strain compared with their expression in the PH3-S strain. In particular, cytochrome oxidase subunit I showed 15-fold higher expression in the PH3-R strain than in the PH3-S strain. Additionally, citrate synthase 2, delta-1-pyrolline-5-carboxylate dehydrogenase, and triose-phosphate isomerase were highly expressed in the PH3-R strain. In summary, our study has improved understanding of the molecular mechanisms of phosphine resistance in the rice weevil.

Detection of Phosphine Resistance in Field Populations of Four Key Stored-Grain Insect Pests in Pakistan

In Pakistan, the control of stored-product insect pests mainly relies on the use of phosphine gas along with other control tactics. The aim of this study was to determine the level of phosphine resistance among ten differently located populations of the lesser grain borer, Rhyzopertha dominica (F.) (Coleoptera: Bostrychidae), the granary weevil, Sitophilus granarius (L.) (Coleoptera: Curculionidae), the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) and the khapra beetle, Trogoderma granarium Everts (Coleoptera: Dermestidae). Laboratory-susceptible populations of all insect species were also considered in the experiments. Concentration–response bioassays were conducted for each species. All of the tested populations (10 out of 10) of each species were found to be resistant to phosphine, but varied in their level of resistance. Probit analysis estimated LC50 at 2.85, 1.90, 2.54 and 2.01 ppm for laboratory-susceptible populations of R. dominica, S. granarius, T. castaneum and T. granarium, respectively. Against R. dominica, the highest and lowest resistance levels were observed in the Rahim Yar Khan (LC50 at 360.90 ppm) and Rawalpindi (LC50 at 210.98 ppm) populations, respectively. These resistant populations were 126.67- and 74.02-fold more resistant than the laboratory population. The Multan and Lahore populations of S. granarius exhibited the maximum (LC50 at 122.81 ppm) and minimum (LC50 at 45.96 ppm) resistance levels, respectively, i.e., they were 64.63- and 24.18-fold more resistant than the laboratory population. The Layyah population of T. castaneum showed the maximum resistance level (LC50 at 305.89 ppm) while the lowest was observed in the Lahore population (LC50 at 186.52 ppm), corresponding to 120.42- and 73.43-fold more resistant than the laboratory population, respectively. Regarding T. granarium, the Layyah population showed the maximum resistance level (LC50 at 169.99 ppm) while the Lahore population showed the minimum resistance (LC50 at 74.50 ppm), i.e., they were 84.57- and 37.06-fold more resistant than the laboratory population, respectively. Overall, R. dominica presented the highest resistance level, followed by T. castaneum, T. granarium and S. granarius. The current study suggests that the application of phosphine may not be an adequate control strategy for the management of the above tested insect pests in Pakistan.

RNA-seq Analysis Reveals Mitochondrial and Cuticular Protein Genes Are Associated with Phosphine Resistance in the Rusty Grain Beetle (Coleoptera:Laemophloeidae)

The rusty grain beetle, Cryptolestes ferrugineus (Stephens), is a serious pest of stored grain, which has developed high levels of resistance to phosphine. In this study, five geographically distant populations of C. ferrugineus had been collected in China, specifically in granaries where phosphine fumigant is used for pest control, and they showed a high resistance ratio up to 1,907 (LC50 = 21.0 mg/liter). Then, a reference transcriptome was constructed to use as a basis for investigating the molecular mechanisms of phosphine resistance in this species, which consisted of 47,006 unigenes with a mean length of 1,090. Subsequently, the RNA-Seq analysis of individuals from the most susceptible and resistant populations led to the identification of 54 genes that are differentially expressed. GO and KEGG analysis demonstrated that genes associated with mitochondrial and respiration functions were significantly enriched. Also, the ‘structural constituent of cuticle’ term was annotated in the GO enrichment analysis and further qRT-PCR confirmed that the expression levels of nine cuticular protein genes were significantly increased in the resistant population. In conclusion, we present here a transcriptome-wide overview of gene expression changes between resistant and susceptible populations of C. ferrugineus, and this in turn documents that mitochondria and cuticular protein genes may play together a crucial role in phosphine resistance. Further gene function analysis should enable the provision of advice to expedite resistance management decisions.

Fumigation Activity against Phosphine-Resistant Tribolium castaneum (Coleoptera: Tenebrionidae) Using Carbonyl Sulfide

Phosphine resistance is occurring among stored-grain pests worldwide. This study investigated the fumigation activity of phosphine (PH3) and carbonyl sulfide (COS) against domestic strain (d-strain) Tribolium castaneum, resistance strain (r-strain) T. castaneum and Oryzaephilus surinamensis. All developmental stages of the pests were exposed to two fumigants (PH3 and COS), and the fumigation activity according to the dose and exposure time was evaluated in a 12-L desiccator and 0.5 m3 fumigation chamber. The rice sorption rate and quality following exposure to thetwofumigants were evaluated. The mortality was 2.9% in r-strain T. castaneum, 49.5% in d-strain T. castaneum and 99.2% in O. surinamensis when 2 mg/L PH3 was used in a 12-L desiccator for 4 h. However, all pest developmental stages showed 100% mortality after 24 h of exposure in the 0.5 m3 fumigation chamber, except for the r-strain T. castaneum. A mortalityof 100% was observed in all of the r-strain T. castaneum developmental stages at an exposure time of 192 h. For COS applied at 40.23 mg/L and 50 g/m3 in the 12-L desiccator and the 0.5 m3 fumigation chamber, respectively, 100% mortality was observed across all developmental stages regardless of species and strain. The sorption of COS was 10% higher than that of PH3, but there was no significant difference in rice quality compared to that in the control. Therefore, this study suggests that COS can be used for controlling T. castaneum resistant to PH3.