Resistant Populations Research Articles

Low expression of auxin receptor EcAFB4 confers resistance to florpyrauxifen-benzyl in Echinochloa crus-galli (L.) P. Beauv

Echinochloa crus-galli (L.) P. Beauv is a monocotyledonous weed that seriously infests rice fields. Florpyrauxifen-benzyl, a novel synthetic auxin herbicide commercialized in China in 2018, is an herbicide for controlling E. crus-galli. However, a suspected resistant population (R) collected in 2012 showed resistance to the previously unused florpyrauxifen-benzyl. Whole-plant dose-response bioassay indicated that the R population evolved high resistance to quinclorac and florpyrauxifen-benzyl. Pretreatment with P450 inhibitors did not influence the GR50 of E. crus-galli to florpyrauxifen-benzyl. The expression of target receptor EcAFB4 was down-regulated in the R population, leading to the reduced response to florpyrauxifen-benzyl (suppresses over-production of ethylene and ABA). We verified this resistance mechanism in the knockout OsAFB4 in Oryza sativa L. The Osafb4 mutants exhibited high resistance to florpyrauxifen-benzyl and moderate resistance to quinclorac. Furthermore, DNA methylation in the EcAFB4 promoter regulated its low expression in the R population after florpyrauxifen-benzyl treatment. In summary, the low expression of the auxin receptor EcAFB4 confers target resistance to the synthetic auxin herbicide florpyrauxifen-benzyl in the R- E. crus-galli.

Identification and expression patterns of voltage-gated sodium channel genes with intron retentions in different strains of Bactrocera dorsalis

Pyrethroid are the primary insecticides used for controlling of Bactricera dorsalis, a highly destructive and invasive fruit pest. Field populations have developed serious resistance, especially to β-cypermethrin. While mutations in the voltage-gated sodium channel (Vgsc) are a common mechanism of pyrethroid resistance, variations in BdVgsc associated with β-cypermethrin resistance remain unclear. Here, we reported the resistance levels of five field populations from China, with resistance ratio ranging from 1.54 to 21.34-fold. Cloning the full length of BdVgsc revealed no specific or known amino acid mutations between the most resistant population and the susceptible strain. However, three types of partial intron retention (IRE4–5, IRE19-f and IREL-24) were identified in BdVgsc transcripts, with these intron retentions containing stop codons. The expression of IRE4–5 transcripts and total BdVgsc showed different trends across developmental stages and tissues. Exposure to β-cypermethrin led to increased expression of IRE4–5. Comparison of genomic and transcriptional sequences reveled that IRE4–5 transcripts had two types (IRE4–5.5 T and IRE4–5.6 T) caused by genomic variations. Both field and congenic strains indicated that homozygotes for IRE4–5.5 T had lower IRE4–5 transcript levels than homozygotes for IRE4–5.6 T. However, congenic and field strains exhibited inconsistent results about the association of expression levels of IRE4–5 transcripts with sensitivity to β-cypermethrin. In summary, this study is the first to identify intron retention transcripts in the Vgsc gene from B. dorsalis and to examine their expression patterns across different developmental stages, tissues, and strains with varying sensitivities to β-cypermethrin. The potential role of the intron retentions of BdVgsc in insecticide toxicity is also discussed.

Using RNA-Seq Analysis to Select Key Genes Related to Seed Dormancy in ALS-Inhibiting Resistant Descurainia sophia with Pro-197-Thr Mutation.

Flixweed (Descurainia sophia) is a weed that seriously affects wheat fields in China. Over the past 20 years, it has evolved resistance to the herbicide tribenuron-methyl. In the present study, a resistant D. sophia population with a Pro-197-Thr mutation of acetolactate synthetase (ALS) was found to have a resistance index of 457.37 for tribenuron-methyl. Under the same growth conditions, the seeds of resistant (R) and susceptible (S) populations exhibited similar vitality but the germination rates of R seeds were higher than those of S seeds. This result demonstrated that seed dormancy periods were shorter in the R seeds. RNA-Seq transcriptome analysis was then used to choose candidate genes that could regulate seed dormancy pathways in the R population. A total of 504,976,046 clean reads were selected from nine RNA-Seq libraries and assembled into 79,729 unigenes. Among these, 33,476 unigenes were assigned to 51 GO subgroups, and 26,117 unigenes were assigned to 20 KEGG secondary metabolic pathways. Next, 2473 differentially expressed genes (DEGs) were divided into three groups, as follows: G-24 h (germinating seeds) vs. D (dormant seeds); G-48 h (germinated seeds) vs. D; and G-48 h vs. G-24 h. From these 2473 DEGs, 8 were selected as candidate dormancy unigenes for the R population if their expression levels continuously decreased during the seed germination progress and their functional annotations were related to plant seed dormancy. One candidate unigene was annotated as CYP707A2; two unigenes were annotated as the transcription factors TGA4 and TGA2; one unigene was annotated as the cystathionine beta-synthase gene; and four unigenes could not be annotated as any gene listed in the six public databases. However, qRT-PCR-validated results showed that, during the germination of R seeds, the expression of the three candidate unigenes first decreased and then increased, indicating that they may have other growth-regulating functions in R populations. In brief, the dormancy function of the eight candidate dormancy unigenes needs to be further studied.

Common consumer residual insecticides lack efficacy against insecticide-susceptible and resistant populations of the German cockroach (Blattodea: Ectobiidae).

The German cockroach, Blattella germanica (L.) (Blattodea: Ectobiidae), is a ubiquitous pest in affordable housing. They represent a major threat to human health due to their contribution of asthma-exacerbating allergens and the potential to transfer pathogenic microorganisms indoors. Despite well-documented pyrethroid resistance, pyrethroid-based broadcast residual insecticide products are often used by residents to control cockroaches in their homes. Additionally, there is little empirical independent testing of these products. Thus, it remains unclear how effective these commonly used do-it-yourself products are at controlling German cockroaches. This study represents a comprehensive examination of the efficacy of these products with direct, limited, and continuous exposure assays on a variety of common household surfaces on field populations of cockroaches with varying levels of pyrethroid resistance. While most products performed well when applied directly to test insects, mortality was substantially lower across all surfaces with limited exposure (30min). In continuous exposure assays on a nonporous surface, products took at least 24hr to cause 100% mortality in a field population, with some products taking up to 5d to achieve 100% mortality. The findings of this study demonstrate a lack of residual efficacy from common pyrethroid-based consumer-use pesticides products. Given that it is not feasible to find and treat every cockroach in a home directly, the residuality of spray-based formulations is critical for products designed to control German cockroaches. Without residual efficacy, as shown in the consumer aerosol and spray products tested, we expect these products to add little to no value to cockroach control.

Metabolic resistance mechanism to glufosinate in Eleusine indica

Eleusine indica is one of the most troublesome weeds in farmland worldwide, especially in Citrus Orchard of China. Glufosinate, as an efficient non-selective broad-spectrum herbicide, has been widely utilized for the control of E. indica in Citrus Orchard. The E. indica resistant population (R) was collected from a Citrus Orchard in Yichang City in Hubei province, China. Bioassay experiments showed that the R plants exhibited 3-fold resistance to glufosinate compared with the E. indica susceptible population (S). No known glutamine synthetase (GS) gene mutation associated with glufosinate resistance was found in R plants. And there was also no significant difference in GS activity between R and S plants. Those results indicated that the resistance to glufosinate in R did not involve target-site resistance. However, glutathione S-transferase (GST) inhibitor 4-chloro-7-nitrobenzoxadiazole (NBD-Cl) plus glufosinate gave a better control of R plants compared with glufosinate treatment alone. Moreover, both before and after glufosinate treatment, the GST activity in R plants was significantly higher than that in S plants. By RNA-seq, the expression of GSTU6 and GST4 up-regulated in R plants relative to S plants with or without glufosinate treatment. They were also significantly up-regulated expression in E. indica field resistant populations compared with S population. In summary, the study elucidated that R plants developed metabolic resistance to glufosinate involving GST. And GSTU6 and GST4 genes may play an important role in this glufosinate metabolic resistance. The research results provide a theoretical basis for a deeper understanding of resistance mechanism to glufosinate in E. indica.

Lipid Metabolism Modulatory Cisplatin Prodrug Sensitizes Resistant Prostate Cancer toward Androgen Deprivation Therapy.

Mainstream treatment modalities which dominate the therapeutic landscape of prostate cancer (PCa) are prostatectomy, radiation therapy, and androgen deprivation therapy (ADT) or castration. These therapeutic options can extend the life expectancy of the patients but eventually fail to completely cure the disease. Despite undergoing ADT, patients still experience disease recurrence. One of the reasons for this recurrence is the binding of the basal androgens present in blood plasma to the androgen receptor (AR). At this stage, the disease becomes castration-resistant prostate cancer (CRPC) showing resistance to ADT promoting progression, and there is no effective treatment available. Although another male cancer such as testicular cancer responds to cisplatin-based therapy very well, PCa is resistant to cisplatin. In our continued effort to find the pathways that are important for such resistance, we link in this report, tumor metabolism driven androgen regulation and PCa resistance toward cisplatin-based therapy. To delve deeper into understanding how metabolic modulatory cisplatin prodrugs can be used to target the ADT resistant population, we demonstrate that metabolic inhibition by a cisplatin prodrug, Platin-L has the potential to modulate AR activity and resensitize ADT resistant cells toward cisplatin-based chemotherapy as well as ADT. The mode of action for Platin-L is inhibition of fatty acid oxidation (FAO) of prostate cancer cells. We demonstrated that FAO inhibition by Platin-L in PCa cells contribute to AR regulation resulting in altered tumorigenicity of androgen sensitive prostate cancer.

Characterisation of low-level pyrasulfotole resistance and the role of herbicide translocation in wild radish (Raphanus raphanistrum)

The synthetic auxin 2,4-D and the 4-hydroxyphenylpyruvate dioxygenase inhibitor pyrasulfotole are phloem-mobile post-emergence herbicides, the latter applied in co-formulation with either bromoxynil (a contact herbicide causing leaf desiccation) or MCPA (another synthetic auxin). Previous studies have shown a wide range of 2,4-D translocation phenotypes in resistant populations of the agricultural weed Raphanus raphanistrum, but it was hypothesised that enhanced movement out of the apical meristem could contribute to resistance. Little is known about pyrasulfotole translocation or the effect of bromoxynil on pyrasulfotole movement. Therefore, the behaviour of pyrasulfotole and 2,4-D applied to the growing point of susceptible and resistant R. raphanistrum seedlings was assessed, along with the effect of bromoxynil on pyrasulfotole translocation. The small amount of herbicide directly contacting the growing point after spraying was sufficient to induce herbicide symptoms, and there was no enhancement of translocation away from the growing point in either pyrasulfotole- or 2,4-D-resistant populations. Bromoxynil had a slightly inhibitory effect on pyrasulfotole translocation in some populations, somewhat negating the minor differences observed among populations when pyrasulfotole was applied alone. Resistance to pyrasulfotole could not explained by enhanced metabolism or vacuolar sequestration of the herbicide. Overall, differential translocation in either the treated leaves or apical meristems does not appear to be a major determinant of resistance to pyrasulfotole or 2,4-D.

Acaricide effect of plants from the Brazilian savanna on a population of Rhipicephalus microplus with phenotypic resistance to cypermethrin and trichlorfon

Rhipicephalus microplus is among the most important ectoparasites for livestock. The use of synthetic acaricides has raised some concerns due to the selection of tick populations that are resistant to acaricides and environmental contamination. Therefore, plant extracts have been used as alternatives for the treatment of animals infested with ticks. In this study, R. microplus populations from seven different dairy farms were collected and assessed for their resistance to the acaricides cypermethrin or trichlorfon. Larvae of the most resistant population were used in assays to evaluate the acaricide effect of leaf extracts from plants of the Brazilian savanna. The most active extracts were also tested against fully engorged females. Among seven tick populations, five and three showed resistance level ≥ III for cypermethrin or trichlorfon, respectively. The most resistant tick population was evaluated in mortality assays with the plants Piptadenia viridiflora, Annona crassiflora, Caryocar brasiliense, Ximenia americana, and Schinopsis brasilienses. The ethanolic extracts of C. brasiliense, X. americana and S. brasilienses showed higher larvicidal effects in comparison to the other extracts and cypermethrin. The ethanolic extract of X. americana showed 60.79 % efficacy against fully engorged females of the acaricide resistant tick strain. The ethanolic extracts of C. brasiliense, X. americana, and S. brasilienses showed peaks in HPLC-DAD, indicating the presence of tannins and flavonoids. Three of the plants showed promising results and should be explored in further studies to develop novel tools to control R. microplus in cattle.

SlCarE054 in Spodoptera litura (Lepidoptera: Noctuidae) showed direct metabolic activity to β-cypermethrin with stereoselectivity.

Carboxylesterases (CarEs) is an important detoxification enzyme system in phase Ⅰ participating in insecticides resistance. In our previous study, SlCarE054, a CarEs gene from lepidoptera class, was screened out to be upregulated in a pyrethroids and organophosphates resistant population. Its overexpression was verified in two field-collected populations of Spodoptera litura (Lepidoptera: Noctuidae) resistant to pyrethroids and organophosphates by qRT-PCR. Spatiotemporal expression results showed that SlCarE054 was highly expressed in the pupae stage and the digestive tissue midgut. To further explore its role in pyrethroids and organophosphates resistance, its metabolism activity to insecticides was determined by UPLC. Its recombinant protein showed significant metabolism activity to cyhalothrin and fenvalerate, but not to phoxim or chlorpyrifos. The metabolic activity of SlCarE054 to β-cypermethrin showed stereoselectivity, with higher metabolic activity to θ-cypermethrin than the enantiomer α-cypermethrin. The metabolite of β-cypermethrin was identified as 3-phenoxybenzaldehyde. Further modelling and docking analysis indicated that β-cypermethrin, cyhalothrin and fenvalerate could bind with the catalytic triad of the 3D structure of SlCarE054. The interaction of β-cypermethrin with SlCarE054 also showed the lowest binding energy. Our work provides evidence that SlCarE054 play roles in β-cypermethrin resistance in S. litura.

Hormetic dose response induced by sublethal-dose sulfoxaflor leads to reproductive stimulation of Aphis gossypii

Aphis gossypii Glover is one of the most agriculturally important phloem-feeding economic pests, causing tremendous loss in crop yield annually. The hormesis is an important cause of A. gossypii resistance formation, population resurgence, and re-outbreak. However, whether the hormesises induced by different insecticides interact mutually remain largely unclear. In the study, four-generation A. gossypii experiment found that the 24-h sublethal-dose (LC20) sulfoxaflor treatment on G0 significantly increased the net reproductive rate (R0) and fecundity of G1 and G2 generation A. gossypii, but it did not significantly affect the fecundity of G3 and G4 individuals. Transcriptomic analyses revealed that the insecticide-induced significant up-regulation of pathways ribosome, energy metabolism, and the DNA replication and reparation might be responsible for the enhancement of fecundity in G1 and G2 A. gossypii. Notably, G0 exposure to LC20 sulfoxaflor followed by G1 exposure to LC30 deltamethrin resulted in a stronger reproductive stimulation than sulfoxaflor or deltamethrin exposure alone. Our findings provide valuable reference for optimizing sulfoxaflor application in integrated pest management strategies.

Insecticidal activity of Schinus terebinthifolius essential oil for the management of permethrin-resistant Sitophilus zeamais (Coleoptera: Curculionidae)

Maize weevil, Sitophilus zeamais, is a major cause of damage to corn. The preventive control of this agricultural pest is performed with permethrin. However, due to the intensive use of insecticides, the population of S. zeamais has developed resistance to more than 10 synthetic insecticides, including the active ingredient permethrin. In the search for techniques aiming to reduction of the use of synthetic pesticides for the integrated management of S. zeamais, the present study was evaluate the effects of Schinus terebinthifolius essential oil – by fumigation, contact and ingestion methods – against populations of S. zeamais, both susceptible and resistant to the insecticide permethrin. Chemical profile identification showed α-pinene, β-myrcene, α-phellandrene, δ-3-carene and β-phellandrene were the major components in S. terebinthifolius essential oil. Permethrin susceptibility of the S. zeamais revealed four resistant populations (STPR, STP1, STP2 and STP3). But, in the Fumigant bioassay, STPR and STP1 permethrin-resistant populations were susceptible to the essential oil. When the insects were put in direct contact with treated maize grains, only STPR permethrin-resistant population was susceptible this essential oil. Resistance ratio of the permethrin on S. zeamais populations revealed differences of permethrin susceptibility among the groups tested. S terebinthifolius essential oil exhibited high insecticidal activity, reducing the populations of S. zeamais through fumigation and contact/ingestion methods. The present findings suggest that use of permethrin can be substituted with the preparation of botanical insecticide with S. terebinthifolius essential oil.

Predictive biomarkers for response to TGF- β inhibition in resensitizing chemo(radiated) esophageal adenocarcinoma

Epithelial-mesenchymal transition (EMT) has been identified as a driver of therapy resistance, particularly in esophageal adenocarcinoma (EAC), where transforming growth factor beta (TGF-β) can induce this process. Inhibitors of TGF-β may counteract the occurrence of mesenchymal, resistant tumor cell populations following chemo(radio)therapy and improve treatment outcomes in EAC. Here, we aimed to identify predictive biomarkers for the response to TGF-β targeting. In vitro approximations of neoadjuvant treatment were applied to publicly available primary EAC cell lines. TGF-β inhibitors fresolimumab and A83–01 were employed to inhibit EMT, and mesenchymal markers were quantified via flow cytometry to assess efficacy. Our results demonstrated a robust induction of mesenchymal cell states following chemoradiation, with TGF-β inhibition leading to variable reductions in mesenchymal markers. The cell lines were clustered into responders and non-responders. Genomic expression profiles were obtained through RNA-seq analysis. Differentially expressed gene (DEG) analysis identified 10 positively- and 23 negatively-associated hub genes, which were bioinformatically identified. Furthermore, the correlation of DEGs with response to TGF-β inhibition was examined using public pharmacogenomic databases, revealing 9 positively associated and 11 negatively associated DEGs. Among these, ERBB2, EFNB1, and TNS4 were the most promising candidates. Our findings reveal a distinct gene expression pattern associated with the response to TGF-β inhibition in chemo(radiated) EAC. The identified DEGs and predictive markers may assist patient selection in clinical studies investigating TGF-β targeting.

Involvement of P450s in the metabolic resistance of Digitaria sanguinalis (L.) Scop. To ALS-inhibiting herbicides

Weed resistance to a range of herbicides has rapidly evolved, often with different mechanisms of action. The resulting uninhibited growth of weeds poses demonstrable threats to crop production and sustainable agriculture. Digitaria sanguinalis (L.) Scop., a troublesome weed in corn and other agricultural fields, has developed resistance to herbicides that inhibiting ALS (Acetolactate Synthase), such as nicosulfuron. Understanding the weed's resistance patterns and mechanisms is crucial. However, little is known of the non-target site resistance (NTSR) mechanisms of D. sanguinalis owing to a lack of relevant genome sequences and other materials. Therefore, in this study, a population of D.sanguinalis presenting multiple resistance was tested and found that its high level of resistance to ALS-inhibiting herbicides was not associated with target-related alterations.Administration of P450 inhibitors reversed the resistance to ALS-inhibiting herbicides. Following the application of ALS-inhibiting herbicides, the activities of NADPH-P450 reductase and p-nitroanisole O-demethylase (PNOD) were notably greater in the resistant population of D. sanguinalis than those in the susceptible population. The results suggested P450 enzyme familyplays a major role in the metabolic resistance mechanism, that increased P450 enzyme activity promote cross-resistance in D. sanguinalis to ALS-inhibiting herbicides. RNA-seq analysis showed that five genes from the P450 family (CYP709B2, CYP714C2, CYP71A1, CYP76C2, and CYP81E8) were upregulated in resistant D. sanguinalis. In conclusion, the upregulation of several P450 genes is responsible for establishing resistance to ALS-inhibiting herbicides in D. sanguinalis.

Transgenerational phenotypic responses to herbicide stress are more rapid than to shade and simulated herbivory in Arabidopsis.

Weeds in agricultural settings continually adapt to stresses from ecological and anthropogenic sources, in some cases leading to resistant populations. However, consequences of repeated sub-lethal exposure of these stressors on fitness and stress "memory" over generations remain poorly understood. We measured plant performance over a transgenerational experiment with Arabidopsis thaliana where plants were exposed to sub-lethal stress induced by the herbicides glyphosate or trifloxysulfuron, stresses from clipping or shading in either one (G1) or four successive generations (G1-G4), and control plants that never received stress. We found that fourth-generation (G4) plants that had been subjected to three generations of glyphosate or trifloxysulfuron stress produced higher post-stress biomass, seed weight, and rosette area as compared to that produced by plants that experienced stress only in the first generation (G1). By the same measure, clipping and shade were more influential on floral development time (shade) and seed weight (clipping) but did not show responsive phenotypes for vegetative metrics after multiple generations. Overall, we found that plants exhibited more rapid transgenerational vegetative "stress memory" to herbicides while reproductive plasticity was stressor dependent and similar between clipping/shade and anthropogenic stressors. Our study suggests that maternal plant stress memory aids next-generation plants to respond and survive better under the same stressors.

The cytochrome P450 subfamilies CYP392A and CYP392D are key players in acaricide metabolism in Tetranychusurticae

The spider mite Tetranychus urticae is a major agricultural pest with a global distribution, extremely diverse host range and a remarkable ability to develop resistance to a wide variety of acaricides. P450 mono‐oxygenases have been frequently associated with resistance development in this species. In particular enzymes of the CYP392A-subfamily were shown to metabolize a number of key acaricides, including abamectin, amitraz, fenpyroximate and the active metabolite of pyflubumide. However, transcriptomic studies comparing highly resistant and susceptible populations have often revealed high expression of members of the CYP392D-subfamily, but these have been only poorly studied. Here, we conducted a meta-analysis of gene expression data of 20 populations and identified two key enzymes of this family, CYP392D2 and CYP392D8, whose expression is associated with resistance. We subsequently functionally expressed these enzymes, together with CYP392A11 and CYP392A16 as known metabolizers, and compared their potential to accept a wide diversity of acaricides as substrate. This study overall confirms previous discovered substrates for CYP392A11 and CYP392A16, but also reveals unreported metabolic activity towards new acaricides. These include carbaryl, chlorpyrifos and etoxazole for CYP392A16 and carbaryl, chlorpyrifos and NNI-0711-NH pyflubumide for CYP392A11. For the newly studied CYP392D-family, we show that CYP392D2 metabolizes pyridaben, fenpyroximate, etoxazole and chlorpyrifos, while CYP392D8 metabolizes carbaryl, fenazaquin and tebufenpyrad. Last, we observed that both CYP392A- and CYP392D-subfamily enzymes activate chlorpyrifos to its corresponding oxon. Our study indicates that there is both overlap and specificity in the activity of A- and D-subfamily enzymes against acaricides and model substrates. With the recent advent of highly efficient CRISPR/Cas9 gene editing protocols in T. urticae, the way is now paved to conduct further genetic experiments revealing and quantifying the role of these enzymes in the resistance phenotype in field populations.

Boric acid toxic sugar bait suppresses male Aedes aegypti (Diptera: Culicidae): wing beat frequency and amplitude, flight activity, fecundity, insemination, and mate-finding Allee effect.

Controlling the spread of arboviral diseases remains a considerable challenge due to the rapid development of insecticide resistance in Aedes mosquitoes. This study evaluated the effects of boric acid-containing toxic sugar bait (TSB) on field populations of resistant Aedes aegypti mosquitoes. In addition, this study examined the flight activity and wing beat frequency and amplitude of males and the flight activity, fecundity, and insemination of females after pairing with males exposed to TSB. The population dynamics of Aedes mosquitoes under imbalanced sex ratios were examined to simulate realistic field conditions for male suppression under the effect of TSB. The mortality of male mosquitoes was consistently high within 24 h after exposure. By contrast, the mortality of female mosquitoes was inconsistent, with over 70% mortality observed at 168 h. The flight activity and wing beat amplitude of treated males were significantly lower than those of controls, but no significant difference in wing beat frequency was detected. The fecundity and insemination of treated female mosquitoes were lower than those of controls. A simulation study indicated that considerably low male population densities led to mating failures, triggering a mate-finding Allee effect and resulting in persistently low population levels. Boric acid-containing TSB could effectively complement current chemical intervention approaches to control resistant mosquito populations. TSB is effective in reducing field male populations and impairing male flight activity and female-seeking behavior, resulting in decreased fecundity and insemination. Male suppression due to TSB potentially results in a small mosquito population. © 2024 Society of Chemical Industry.

Preliminary monitoring of knockdown resistance (kdr) mutation in Anopheles stephensi: insights from a malarious area in Southeastern Iran

BackgroundAnopheles stephensi is recognized as the main malaria vector in Iran. In recent years, resistance to several insecticide classes, including organochlorine, pyrethroids, and carbamate compounds, has been reported for this medically important malaria vector. The main objective of the present study was to evaluate the insecticide susceptibility status of An. stephensi collected from the southern part of Iran, and to clarify the mechanism of resistance, using bioassay tests and molecular methods comparing the sequence of susceptible and resistant mosquitoes.MethodsMosquito larvae were collected from various larval habitats across six different districts (Gabrik, Sardasht, Tidar, Dehbarez, Kishi and Bandar Abbas) in Hormozgan Provine, located in the southern part of Iran. From each district standing water areas with the highest densities of Anopheles larvae were selected for sampling, and adult mosquitoes were reared from them. Finally, the collected mosquito species were identified using valid keys. Insecticide susceptibility of An. stephensi was tested using permethrin 0.75%, lambdacyhalothrin 0.05%, deltamethrin 0.05%, and DDT 4%, following the World Health Organization (WHO) test procedures for insecticide resistance monitoring. Additionally, knockdown resistance (kdr) mutation in the voltage-gated sodium channel (vgsc) gene was sequenced and analysed among resistant populations to detect possible molecular mechanisms of observed resistance phenotypes.ResultsThe susceptibility status of An. stephensi revealed that resistance to DDT and permethrin was found in all districts. Furthermore, resistance to all tested insecticides in An. stephensi was detected in Gabrik, Sardasht, Tidar, and Dehbarez. Analysis of knockdown resistance (kdr) mutations at the vgsc did not show evidence for the presence of this mutation in An. stephensi.ConclusionBased on the results of the current study, it appears that in An. stephensi from Hormozgan Province (Iran), other resistance mechanisms such as biochemical resistance due to detoxification enzymes may be involved due to the absence of the kdr mutation or non-target site resistance. Further investigation is warranted in the future to identify the exact resistance mechanisms in this main malaria vector across the country.

Determination of target site mutations and detoxification enzymes in the abamectin resistance of spider mite Tetranychus urticae collected from cotton fields of Türkiye

The two-spotted spider mite, Tetranychus urticae (Koch, 1836) (Acari: Tetranychidae) causes significant problems in cotton fields. The pest has developed resistance to abamectin, which is used intensely against spider mite populations. A discriminating dose was applied to T. urticae populations collected from Aydın, Adana, Şanlıurfa, and Diyarbakır provinces between 2019 and 2021, where there is significant cotton production and intensive acaricide applications. Populations with a mortality rate of less than 80% at the discriminating dose applied to field populations were included in the bioassay study. Then, the 10 most resistant populations were determined based on their LC50 and LC90 values. It was observed that there was high resistance (HR) in two T. urticae populations collected from Diyarbakır (DIY28 and DIY2) with LC50 values of 24.5 and 33.81 mg L-1, respectively 113 and 156-fold and one population collected from the province of Aydın (AYD4) LC50 value of 23.5 mg L-1 and 109-fold. As a result of the comparisons with a susceptible population in biochemical studies, it was determined that the carboxylesterase (CarE) enzyme of these populations (except for SAN6, SAN7, SAN8, ADA9, and ADA11) had more than 2-fold higher activity compared to the glutathione enzyme (GST), and they were in statistically different groups. When the GST enzyme activities were examined, it was found that the enzyme activities were generally low, except for the ADA11 and ADA16 populations (3.73 and 2.3-fold). In terms of the target site resistance, it was observed that there was no target site mutation (G314D and I321T-G326E) in the glutamate-gated chloride channels (Units GluCl 1 and 3) in these highest 10 resistant populations. It can be said that the resistance levels determined for abamectin from the obtained populations were determined to be low. In brief, out of the total 44 populations brought in and tested, only 30 populations were considered resistant with a mortality rate below 80% after the applied discriminating dose. It was determined that 6.8% of these 30 populations had HR, 31.8% had medium resistance (MR), and 29.5% had low resistance (LR). When examining by regions, it was found that high resistance populations were more prevalent in Diyarbakır and Aydın (15.4% and 10%, respectively) provinces, while the highest percentage of MR populations was found in Şanlıurfa, at 90%. Abamectin still acts as an effective acaricide. However, it is recommended to use acaricides with different modes of action to prevent possible emergence of resistance cases.

Acquisition of Resistance to PEGylated Branched Polyethylenimine Increases Pseudomonas Aeruginosa Susceptibility to Aminoglycosides.

PEGylated branched polyethylenimine (PEG-BPEI) has antibacterial and antibiofilm properties. Exposure to PEG-BPEI through serial passage leads to resistant P. aeruginosa strains. The minimum inhibitory concentration (MIC) of 600 Da BPEI and PEGylated 600 Da BPEI (PEG-BPEI) in the wild-type PAO1 strain is 16 μg/ml while, after 15 serial passages, the MIC increased to 1024 μg/mL. An additional 15 rounds of serial passage in the absence of BPEI or PEG-BPEI did not change the 1024 μg/mL MIC. Gentamicin, Neomycin, and Tobramycin, cationic antibiotics that inhibit protein synthesis, have a 16-32 fold reduction of MIC values in PEG350-BPEI resistant strains, suggesting increased permeation. The influx of these antibiotics occurs using a self-mediated uptake mechanism, suggesting changes to the outer membrane Data show that resistance causes changes in genes related to outer membrane lipopolysaccharide (LPS) assembly. Mutations were noted in the gene coding for the polymerase Wzy that participates in the assembly of the O-antigen region. Other mutations were noted with wbpE and wbpI of the Wbp pathway responsible for the enzymatic synthesis of ManNAc(3NAc)A in the LPS of P. aeruginosa. These changes suggest that an altered gene product could lead to PEG-BPEI resistance. Nevertheless, the increased susceptibility to aminoglycosides could prevent the emergence of PEG-BPEI resistant bacterial populations.

Control of a phytopathogenic infection in an apple orchard in the forest-steppe of Samara Oblast

The study evaluates a protection system by selecting and alternating fungicides having different mechanisms of action to exclude the emergence of resistant populations of apple tree pathogens. The experiments were started at Koshelevsky Posad LLC (Syzransky District, Samara Oblast) in 2022-2023. The orchard was planted in 2013-2014, with trees arranged in a 3.5×1.5 m grid; the 62-396 rootstock was used. The biological effectiveness of fungicide protection against apple scab (Venturia inaequalis (Cooke) Wint.) and fruit rot (Monilia fructigena Pers.) was analyzed using ‘Berkutovskoye,’ ‘Kulikovskoye,’ ‘Kuibyshevskoye,’ ‘Lobo,’ and ‘Northern Sinap’ varieties. The protection system including contact and systemic agents having different mechanisms of action was developed according to the FRAC diagram for assessing the combined risk of fungicide resistance developing in phytopathogens, which takes into account the adaptation rate of organisms (emergence of new resistant types) to changing environmental conditions. An analysis of the correlation between the growth rate of apple-specific phytopathogens and weather conditions shows no significant relationship (0.032-0.245). In these studies, the anthropogenic factor-chemical treatment with fungicides – had a predominant effect on the development of scab and monilia in apple orchards along with weather factors. The yield of marketable apples in the experimental quarter of the orchard differed insignificantly from year to year. In 2023, the yield was lower; however, this fact can be probably attributed to the return of late frosts during apple blossom in Samara Oblast. Noteworthy is that for the ‘Berkutovskoye’ and ‘Lobo’ varieties, unmarketable apples did not exceed 5 and 12 %, respectively, in the studied quarter of the orchard. The protection system using fungicides (GC Shans) helped to optimize the phytosanitary condition of the orchard, thus reducing the amount of fallen fruit and providing a stable harvest of marketable apples.