Phosphine gas, or hydrogen phosphide (PH3), is the most common insecticide applied to durable stored products worldwide and is routinely used in the United Stat

Field experiments were conducted in steel bins containing 13,600 kg of hard red winter wheat, Triiticum aestivum L. One bin was treated with ozone an

Phosphine is a valuable fumigant to control pest populations in stored grains and grain products. However, recent studies indicate a substantial increase in phosphine resistance in stored product pests worldwide. To understand the molecular bases of phosphine resistance in insects, we used RNA-Seq to compare gene expression in phosphine-resistant and susceptible laboratory populations of the red flour beetle, Tribolium castaneum. Each population was evaluated as either phosphine-exposed or no phosphine (untreated controls) in triplicate biological replicates (12 samples total). Pairwise analysis indicated there were eight genes differentially expressed between susceptible and resistant insects not exposed to phosphine (i.e., basal expression) or those exposed to phopshine (>8-fold expression and 90 % C.I.). However, 214 genes were differentially expressed among all four treatment groups at a statistically significant level (ANOVA, p < 0.05). Increased expression of 44 cytochrome P450 genes was found in resistant vs. susceptible insects, and phosphine exposure resulted in additional increases of 21 of these genes, five of which were significant among all treatment groups (p < 0.05). Expression of two genes encoding anti-diruetic peptide was 2- to 8-fold reduced in phosphine-resistant insects, and when exposed to phosphine, expression was further reduced 36- to 500-fold compared to susceptible. Phosphine-resistant insects also displayed differential expression of cuticle, carbohydrate, protease, transporter, and many mitochondrial genes, among others. Gene ontology terms associated with mitochondrial functions (oxidation biological processes, monooxygenase and catalytic molecular functions, and iron, heme, and tetrapyyrole binding) were enriched in the significantly differentially expressed dataset. Sequence polymorphism was found in transcripts encoding a known phosphine resistance gene, dihydrolipoamide dehydrogenase, in both susceptible and resistant insects. Phosphine-resistant adults also were resistant to knockdown by the pyrethroid deltamethrin, likely due to the increased cytochrome P450 expression. Overall, genes associated with the mitochondria were differentially expressed in resistant insects, and these differences may contribute to a reduction in overall metabolism and energy production and/or compensation in resistant insects. These data provide the first gene expression data on the response of phosphine-resistant and -susceptible insects to phosphine exposure, and demonstrate that RNA-Seq is a valuable tool to examine differences in insects that respond differentially to environmental stimuli.

Phosphine resistance in stored-product insects occurs worldwide and is a major challenge to continued effective use of this fumigant. We determined resistance frequencies and levels of resistance in Tribolium castaneum and Plodia interpunctella collected from California almond storage and processing facilities. Discriminating doses of phosphine were established for eggs and larvae of P. interpunctella and eggs of T. castaneum using laboratory susceptible strains of the two species. For T. castaneum and P. interpunctella eggs, discriminating doses were 62.4 and 107.8 ppm, respectively, over a 3-d fumigation period, and for P. interpunctella larvae, discriminating dose was 98.7 ppm over a 20-h fumigation period. Discriminating dose tests on adults and eggs showed that 4 out of 11 T. castaneum populations tested had resistance frequencies that ranged from 42 to 100% for adults and 54 to 100% for eggs. LC99 values for the susceptible and the most resistant adults of T. castaneum were 7.4 and 356.9 ppm over 3 d, respectively. LC99 values for T. castaneum eggs were 51.5 and 653.9 ppm, respectively. Based on adult data, the most resistant T. castaneum beetle population was 49× more resistant than the susceptible strain. Phosphine resistance frequencies in P. interpunctella eggs ranged from 4 to 20%. Results show phosphine resistance is present in both species in California. Future research will investigate phosphine resistance over a wider geographic area. In addition, the history of pest management practices in facilities where insects tested in this study originated will be determined in order to develop phosphine resistance management strategies for California almond storage and processing facilities.

A field experiment was conducted in eight 13.6-MT steel bins containing 6.8 MT each of wheat to assess efficacy of sulfuryl fluoride or SF fumigant to control phosphine-resistant and susceptible Rhyzopertha dominica (F.) and Tribolium castaneum (Herbst). Approximately 400 adults of each type of beetle were added to each bin. Additionally, muslin bags containing immature stages and adults, with their respective diets, were also placed in bins. Four bins were fumigated with SF and others were untreated control bins. The SF dosages in treated bins ranged from 1,196–1,467 mg-h/liter. Mortality of adults in each bag was assessed 5 d postfumigation; diet minus adults was incubated in a jar, and number of adults counted after 8 wk. No significant change occurred in number of insect-damaged kernels in SF-treated bins. In trier samples from SF-treated bins, R. dominica numbers declined from 24 prefumigation to 0 at 3- and 6-wk postfumigation; T. castaneum numbers were unchanged. In WBII traps from SF-treated bins, numbers R. dominica and T. castaneum declined from 25 and 33, respectively, prefumigation to 0 or near 0 at 3- and 6-wk postfumigation. Mortalities of resistant and susceptible adult R. dominica, and adult and large larvae of T. castaneum in SF-treated bags was 100%. For all four types of beetles, adult numbers in jars associated with SF-treated bins were 0 or near 0. Results show SF is effective against all life stages of phosphine-resistant R. dominica and T. castaneum, and can be used for phosphine resistance management.

Phosphine gas (PH₃) is one of the most commonly used fumigants for controlling stored-grain pests worldwide. We estimated the discriminating dose for Cryptolestes ferrugineus (Stephens) (Coleoptera: Laemophloeidae) adult insects using a laboratory susceptible strain. This discriminating dose was then used to determine presence or absence of PH₃ resistance (resistance frequencies) in 19 field-collected populations of C. ferrugineus from Oklahoma, United States. The discriminating dose was estimated as 56.2 ppm of PH₃ over a 20-h exposure period at 25 °C. Discriminating dose bioassay results showed that PH₃ resistance was present in all 19 populations of C. ferrugineus tested. However, five populations—Stillwater (Stil), Enid Terminal 1 (ET-1), Enid Terminal 2 (ET-2), Johnson-Enid population (JE), and DK Farm 20 population (DK Farm 20) had ≥90% resistance frequencies. LC₉₉ values estimated by probit analyses of dose–response mortality data for the laboratory susceptible strain (Lab-S), JE, and DK Farm 20 were 7.3, 636.4, and 968.6 ppm, respectively, over a 3-d exposure period. The level of resistance in DK Farm 20, the most resistant population, was 133.5 times that of the susceptible laboratory strain. This study shows that PH₃ resistance in C. ferrugineus may be widespread in Oklahoma. Based on this study, there is a need for a wider PH₃ resistance survey in grain-growing regions of Oklahoma and United States. Furthermore, results show there is a need to develop PH₃ resistance management strategies for C. ferrugineus and other stored-product insect pest species to combat resistance and ensure continued effective future use of PH₃.