Fitness in a malathion resistant Tribolium castaneum strain; feeding, growth and digestion

https://doi.org/10.1016/j.jspr.2021.101814Get rights and content

Highlights

  • Fitness in a malathion resistant strain based on underlying physiological processes.

  • Wheat flour ingestion in the resistant Tribolium castaneum was significantly reduced compared to the susceptible strain.

  • Gut tract size and protein contents were lower in the resistant strain than in both the susceptible strain and T.confusum.

  • Inhibition of gut proteolysis was less in the resistant than in the susceptible strain.

  • Resistant larvae had lower total soluble protein, while resistant adults had higher levels.

Abstract

Extensive historical use of malathion in grain stores has led to the universal development of resistant strains of the red flour beetle, Tribolium castaneum Herbst. Adaptation to insecticide stress often reduces general fitness. To investigate non-defensive attributes physiological and biochemical properties (that may underlie fitness) of a field-derived malathion-resistant strain (larval resistance factor of 490) were measured against those of a susceptible strain and also with susceptible T. confusum. Further inhibition in vitro of gut proteolysis was determined for the resistant and susceptible strains of T. castaneum.

The weights of whole gut tracts of the susceptible T. castaneum strain and T. confusum were similar but the resistant strain had significantly lower weight and width of whole gut tract. The gut of the resistant strain had more, and more intense, electrophoretic protein staining bands. Food consumed through the life cycle was significantly less for the resistant strain. Inhibitors for cysteine proteases had less effect in vitro in the resistant strain than in the susceptible strain. An inhibitor for serine proteases had almost no effect on proteolysis in the resistant strain.

The reduced gut dimensions and food consumed, together with lower larval protein, appear to indicate a fitness cost that may be associated with malathion resistance. However, this may be offset by enhanced gut proteins and a relative greater protein content of the adult stage. The latter may be related to the reported increased reproductive capacity of the resistant adult and consequent superior fitness, compared to the susceptible. The lowered response of the resistant strain to cysteine and serine protease inhibitors may interact with adaptations to dietary inhibition in cases where resistant strains need managing, if this result is confirmed to be widespread.

Introduction

The wide use of insecticides to control stored insect pests has resulted in resistance to many common insecticides. Tribolium castaneum (Herbst.) is a serious and cosmopolitan insect pest of stored grain products (Champ and Dyte, 1977) and has widely developed resistance to malathion, the main insecticide historically used for control of this pest (Champ and Dyte, 1977). Even though malathion is no longer used in stores resistance remains widespread (Zettler, 1991; Haubruge and Arnaud, 2001; Rossi et al., 2010). It has generally been replaced with phosphine to which resistance has also appeared (Jagadeesan et al., 2012).

Insecticide resistant strains in an insecticide free environment are reportedly at a disadvantage compared to the corresponding susceptibles due to the costs of the resistance mechanisms (Guedes et al., 2006). However, many studies of this have been carried out in low stress artificial situations, where the effects of other stressors cannot be determined (ffrench-Constant and Bass, 2017). Other changes associated with insecticide resistance, the pleiotropic genes, may or may not have cost implications (Lenormand et al., 2018). Some resistant strains of T. castaneum feeding on a standard diet have reduced fecundity, decreased survival and prolonged developmental time of certain life stages when compared with susceptible strains (White and Bell, 1988; Bughio and Wilkins, 2004; Djihinto et al., 2013). This has also been observed in cases of resistance to phosphine in this species (Pimentel et al., 2007; Jagadeesan, 2012; also Sousa et al., 2009). However, in the presence of other stressors a fitness advantage may be observed (Steinbach et al., 2017). For example, a highly malathion-resistant strain, of T. castaneum feeding on rice grain (which is less suitable than wheat) of an insect resistant cultivar gained significantly higher weight than a susceptible strain (Bughio and Wilkins, 2004).

Adaptation to toxins in the insect's environment is an example of changes in response to stressors in general and often result in a lack of fitness in the absence of such stressors. These pleiotropic effects can involve non-defensive changes having no obvious function in the stressed environment. In the case of T. castaneum resistance to malathion has become widespread and persistent. For this population resistance has no cost and lifecycle parameters are better than the corresponding susceptibles in insecticide free conditions (Haubruge and Arnaud, 2001). Malathion-specific resistant strains had an increase in biotic potential, having fitness independent of the genetic background with no relationship to the resistance mechanism. Increased female fecundity and enhanced male reproductive ability have been shown by these resistant strains (Arnaud et al., 2002; Arnaud and Haubruge, 2002).

In T. castaneum populations resistant to phosphine there are usually fitness costs, but as pest evolution has continued this is changing (Nayak et al., 2020). This was true for the red flour beetle (T. castaneum, the lesser grain borer (Rhyzopertha dominica (F.) and the sawtoothed grain beetle (Oryzaephilus surinamensis (L.). However, phosphine-resistant populations, of O. surinamenis and R. dominica displayed no such fitness costs (Sousa et al., 2009). Further genetic studies on resistance in T. castaneum have revealed two genes acting together to give high levels of resistance to phosphine (Jagadeesan et al., 2012). High levels of resistance to deltamethrin were associated with enhanced activities of cellulases and amylases, compared to a susceptible strain (Riaz et al., 2018).

Fitness costs can be described through growth, development or reproduction rates (Maroufpoor et al., 2018). Other authors consider a wider range of life cycle parameters (Belinato and Martins, 2016) such as wing length/body size and feeding in the case of mosquitoes (Huho et al., 2007). This approach can incorporate physiological properties, particularly alteration and synthesis of soluble protein (Ahmed et al., 1998) in various stages (larval, pupal and adult) as this is involved in growth and development of the insect. The most pronounced physiological precursors to observed fitness can occur with metamorphosis in the holometabolous orders (Kanost et al., 1990), such as beetles, as this is the time when storage proteins accumulate in the haemolymph (Telfer and Kunkel, 1991; Telang et al., 2002). Contributing factors are the gut digestive efficiency and tolerance of the enzymes (to dietary inhibitors, pathogens and toxicants) used by insects for the digestion of their dietary proteins and other substrates (Terra et al., 2019).

To investigate many such factors in resistant strains, the larval instars are most relevant, as these are the main food-ingesting stages of Tribolium species. Protein concentrations of whole digestive gut tracts and other loci of soluble protein such as haemolymph and lipids (liquid fraction) and the remaining carcass of the larva (Telang et al., 2002) can thus usefully be compared.

In the current study a selected range of physiological parameters likely to underlie fitness costs were chosen, using a highly malathion-resistant, alongside a susceptible, strain of T. castaneum and a susceptible strain of a closely related species, Tribolium confusum Jacquelin Du Val. T. confusum was included as an example of a susceptible strain of another species, to allow contrast with the resistant strain of T. castaneum. The body weights and protein concentrations of last instar larvae, 24-h old pupae and 2 day-old adults, the larval carcass, liquid fraction (haemolymph and lipids) and whole gut tract of the three Tribolium species/strains were investigated. Food ingestion, gut size, and proteolytic activity were determined. Electrophoresis studies were carried out to compare the bands and the molecular weights of proteins from the gut tracts of the strains of the Tribolium species. Responses of gut proteolysis to inhibitors in vitro was also determined for the resistant and the susceptible strains of T. castaneum.

Section snippets

Insect species/strains

Two strains of T. castaneum, Ph-1, highly resistant and FSS-II, susceptible to malathion (Rauf et al., 1997) and a T. confusum strain, susceptible to malathion were used. The field-originating resistant strain, Ph-1, was obtained from the Natural Resources Institute, Chatham, UK, and susceptible strains of T. castaneum, FSS-II and of T. confusum were originally from the Central Science Laboratory, Slough, UK. The susceptible strain FSS-II has been further characterised for genetic variation (

Food consumption and digestive tract dimensions

The greatest amount of the whole meal wheat flour was consumed by FSS-II and least by Ph-1 (measured from first instar to adult) (Table 1). A comparison of the two strains of T. castaneum, showed FSS-II consumed a significantly higher amount than Ph-1 (ANOVA; F = 11, df = 2, LSD; P < 0.05). T. confusum also ingested a significantly higher amount than Ph-1 (ANOVA; F = 11, df = 2, LSD; P < 0.05), but was not significantly different to FSS-II (LSD; NS).

The lengths of the last instar larval whole

Discussion

The Ph-1 strain was used in the present study as it represents resistant insects with no fitness costs (which include up to 40% of reported resistance fitness cases, Freeman et al., 2020), is stable and similar malathion resistant strains are most likely present in most grain stores (Anusree et al., 2019; Attia et al., 2020). Thus, information on the biology and fitness attributes of such a strain are useful in pest management strategies, even though malathion is no longer used. For example, if

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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