Large-scale identification of rodenticide resistance in Rattus norvegicus and Mus musculus in the Netherlands based on Vkorc1 codon 139 mutations

BACKGROUND

Resistance to rodenticides has been reported globally and poses a considerable problem for efficacy in pest control. The most-documented resistance to rodenticides in commensal rodents is associated with mutations in the Vkorc1 gene, in particular in codon 139. Resistance to anticoagulant rodenticides has been reported in the Netherlands since 1989. A study from 2013 showed that 25% of 169 Norway rats (Rattus norvegicus) had a mutation at codon 139 of the Vkorc1 gene. To gain insight in the current status of rodenticide resistance amongst R. norvegicus and house mice Mus musculus in the Netherlands, we tested these rodents for mutations in codon 139 of the Vkorc1 gene. In addition, we collected data from pest controllers on their use of rodenticides and experience with rodenticide resistance.

RESULTS

A total of 1801 rodent samples were collected throughout the country consisting of 1404 R. norvegicus and 397 M. musculus. In total, 15% of R. norvegicus [95% confidence interval (CI): 13–17%] and 38% of M. musculus (95% CI: 33–43%) carried a genetic mutation at codon 139 of the Vkorc1 gene.

CONCLUSION

This study demonstrates genetic mutations at codon 139 of the Vkorc1 gene in M. musculus in the Netherlands. Resistance to anticoagulant rodenticides is present in R. norvegicus and M. musculus in multiple regions in the Netherlands. The results of this comprehensive study provide a baseline and facilitate trend analyses of Vkorc1 codon 139 mutations and evaluation of integrated pest management (IPM) strategies as these are enrolled in the Netherlands. © 2022 The Dutch Pest and Wildlife. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Distribution of VKORC1 single nucleotide polymorphism in wild Rattus norvegicus in France

BACKGROUND: Anticoagulant rodenticides are commonly used to control rodent pests all over the world. These pesticides inhibit one enzyme of the vitamin K cycle, Vkorc1, and thus prevent blood clotting and cause death by haemorrhage. Resistance to anticoagulants was first observed in Scotland in 1958, and more potent anticoagulants have been developed to overcome this obstacle. Unfortunately, these chemicals are very toxic and cannot be used everywhere. Some authors have shown that resistance to anticoagulants seems closely linked with single nucleotide polymorphism (SNP) in the Vkorc1 gene. RESULTS: This study draws a map of SNP and haplotypes found in Vkorc1 in rats from different areas of France. Some of them had never been described before. Moreover, the Y139F mutation, described previously in France and Belgium, is the most frequent in France. This mutation is known to be associated with a strong resistance to anticoagulants, and it was found in 28% of the samples. CONCLUSION: This biomolecular approach to studying and detecting resistance is easier to carry out than the phenotypic approach measuring blood coagulation time because it can be conducted on biological samples from dead animals, and it is less dangerous for the operator. Copyright © 2009 Society of Chemical Industry     

Bromadiolone susceptibility in wild and laboratory Mus musculus L.(house mice) in Buenos Aires,Argentina

BACKGROUND: Rodents are major pests in many agricultural systems, where they can cause significant economic losses and involve a sanitary risk. The application of anticoagulant rodenticides for rodent control has showed a decrease in effectiveness through time because of the development of resistant populations and the development of aversion behaviour. The goal of the present study was to test the susceptibility to bromadiolone and the existence of anticoagulant resistance in Mus musculus L. (house mouse) in Argentina. We conducted a feeding test with wild animals captured in poultry farms and a laboratory strain that were fed with bromadiolone bait. RESULTS: Three animals of the field experimental group survived the 21 days study period, while for laboratory animals mortality was 100%. Control field animals which were fed without anticoagulant showed 100% survival. CONCLUSION: We found evidence of the presence of anticoagulant resistant M. musculus L. in the study area. Feeding behaviour may have contributed to increasing the time of survival, and may be a mechanism that allows metabolic clearance of the bromadiolone. Under field conditions control with anticoagulants would be less effective because animals have alternative food. Copyright © 2009 Society of Chemical Industry     

A review of biochemical mechanisms of warfarin resistance in the house mouse

The biochemical mechanisms of warfarin resistance in the house mouse (Mus musculus L.) have not been fully investigated. In some populations of warfarin-resistant mice there is a reduction in sensitivity of hepatic vitamin K-epoxide reductase to inhibition by warfarin. This is similar to a proposed mechanism of anticoagulant resistance in the Norway rat (Rattus norvegicus Berk.). The same enzyme in other warfarin-resistant populations is, however, sensitive to warfarin inhibition. Other studies have indicated that detoxification may play a role in conferring warfarin resistance in house mice.     

The impact of resistance on the use of second-generation anticoagulants against rats on farms in Southern England

Resistance amongst rats to second-generation anticoagulants, notably difenacoum, has been recognised since the 1970s. Although initially considered to be of practical significance, doubts were subsequently expressed that the degree of resistance was sufficient to explain ineffective rodenticide treatments. Research published elsewhere demonstrated the overriding importance of poor poisoned bait consumption, caused by the influence of certain ecological factors on rat behaviour, in reducing the effectiveness of second-generation anticoagulant treatments on farms in southern England. In this paper we consider, however, the subtle effects of resistance to anticoagulants on treatment outcome. The prevalence of resistance was increased amongst survivors of treatments and bait consumption by resistant survivors was higher than amongst susceptible survivors. The overall prevalence and degree of resistance to second-generation anticoagulants does not, however, currently represent a practical problem. Nevertheless, selection favouring increased resistance remains a long-term threat to the effectiveness of both difenacoum and bromadiolone.     

Field evaluation of capsaicin as a rodent aversion agent for poultry feed

Developing additional techniques for reducing animal feed contamination by rodents and controlling rodent populations is critical to efforts aimed at reducing the occurrence of Salmonella spp infection on poultry farms. Capsaicin, a compound found in chili peppers of the genus Capsicum, produces a burning sensation in the mouth of mammals and is used effectively as an animal deterrent for some pest species. Applied to poultry feed, capsaicin may be effective as an aversive agent to deter rodent feeding and enhance acceptability of rodenticide baits. We tested capsaicin-treated poultry diets (2000 and 3000 Scoville Heat Units, SHU) in no-choice feeding trials at four active New York farms in the winter of 1997-1998. At all farms, consumption of the 2000 SHU diet by rodents (Norway rats, Rattus norvegicus (Berk), and house mice, Mus musculus L) was significantly less than consumption of a control diet. Consumption of the 3000 SHU diet by rodents was significantly less than consumption of a control diet at three of the four farms. Overall, consumption of treated diets was 58-97% and 55-98% less than consumption of the control diet, for the 2000 and 3000 SHU diets, respectively. These reductions appeared to be related closely to the availability of alternative feed sources at these farms. Two-choice feeding trials involving a rodenticide bait (0.05 g kg(-1) brodifacoum) and the 3000 SHU diet demonstrated that Norway rats preferred the rodenticide to the capsaicin-treated poultry feed. Overall, rodenticide bait acceptance was high (95.6%) when offered simultaneously with capsaicin-treated poultry feed. Although poultry managers must utilize several techniques to manage rodent pests, the use of capsaicin-treated diets to reduce feed losses and increase rodenticide bait acceptance appears promising. Use of capsaicin-treated feed on poultry farms may substantially reduce feed contamination by rodents and ultimately the incidence of Salmonella infection in poultry.     

Anticoagulant resistance in Europe: Appraisal of the data from the 1992 EPPO questionnaire

Resistance to the first successful anticoagulant rodenticide, warfarin, was detected in the Norway rat (Rattus norvegicus Berk.) in 1958, and it has since expanded to cover other commensal rodent species (R. rattus L., Mus musculus L.) and the majority of later-developed anticoagulant compounds. The purpose of this paper is to give an up-to-date picture of the current distribution of anticoagulant resistance in Europe, as revealed by the questionnaire dispatched by EPPO (European and Mediterranean Plant Protection Organization) to its member countries in 1992. Replies were received from 13 countries (43% of EPPO members), and it was found that broad-spectrum resistance occurred in practically all those countries that reported testing activity. A likely conclusion based on this finding is that true distribution of resistance is certainly more extensive than documented by the questionnaire replies. Another conclusion drawn by the EPPO Rodent Control Panel was that the existing resistance detection protocols were no longer adequate to cover the whole array of existing anticoagulant rodenticides. Consequently, a new guideline including novel methods particularly for detecting resistance to single-dose anticoagulant compounds was urgently needed. This guideline is currently under review.     

The influence of bait presentation on bait uptake by mice (Mus domesticus) in infested urban domestic dwellings

Several researchers have established a link between the presence of house mouse (Mus domesticus) infestations and detrimental effects on human health. Controlling mouse infestations therefore requires methods that are quick, safe and effective. A tamper-resistant bait station with a wax block which is manufactured to fit within it so that it cannot be shaken out is considered the safest way to present the bait. However, some authors have expressed concern about the efficacy of this method. Research to examine the effects of bait stations on the feeding of mice infesting domestic dwellings was undertaken in the city of Manchester, UK. Working in conjunction with Manchester City Council, suitable sites were found. A balanced Latin Square experimental design was employed to overcome any operational biases that could have occurred. Non-toxic baits were presented in one of four ways: whole wheat in a cardboard box; whole wheat in a open tray; whole wheat in a tamper-resistant box; and a non-toxic wax block in a tamper-resistant box. Baits were placed in four different locations within dwellings and during the feeding trial; the bait stations were rotated, so that each bait type was presented in each location. The bait take from 12 dwellings within 3 housing blocks was recorded. Three-quarters of the bait consumed during the trial was taken from the cardboard box. No bait was taken from the tamper-resistant box containing the wax block.

Risk assessments should drive the approach adopted in presenting rodenticide. Dwellings where vulnerable adults/children/pets are present may represent a risk of accidental poisoning may require a regime with tamper-resistant bait boxes containing wax blocks, to prevent baits being accessed. However, a balance between the safest and the most effective presentation of bait is needed and in areas where there is unlikely to be any access to baits (e.g. behind kitchen kickboards), use of alternatives must remain to ensure the shift resolution of infestations. Where bait can be placed safely and there are no children or pets present, cardboard boxes provide an effective means of ensuring that bait is consumed by the mice infesting domestic dwellings.     

Warfarin-based rodenticides: Mode of action and mechanism of resistance

Warfarin and related substances have been in use as rodenticides for fifty years. They act by binding to the enzyme vitamin K 2,3-epoxide reductase, thereby interrupting the cellular recycling of vitamin K. Vitamin K in its hydroquinone form is an essential cofactor for the synthesis of functional prothrombin and related blood-clotting factors. The binding with the reductase is essentially irreversible, indicating these compounds have a prolonged half-life in target tissues; 7–10 days for warfarin and congeners and over 100 days for the second generation rodenticides or ‘superwarfarins’ such as difenacoum, brodi-facoum, and flocoumafen. Rat liver contains 1–2 nmole of enzyme per gram tissue which is a 4–5 fold overcapacity for maintaining effective vitamin K recycling. The use of warfarin as a rat poison has resulted in the natural selection of warfarin-resistant rats. The resistance is inheritable. Two distinct warfarin resistance genotypes, Welsh and Scottish, have been identified, clearly differing in their biochemistry of vitamin K epoxide reductase. In the Welsh strain, resistance arises from an altered enzyme expressing reduced reactivity to warfarin, whereas the reductase from the Scottish strain is as sensitive as the normal enzyme, but the interaction with warfarin is now readily reversible. The altered enzyme leaves the Welsh rat in need of higher dietary vitamin K intake. The superwarfarins cope with the resistance by having their structures firmly bind with the altered enzymes. Reduced sensitivity to warfarin-based rodenticides may also be pharmacokin-etically based, arising from increased warfarin biotransformation. This mechanism may be responsible for resistance to some of the superwarfarins such as difenacoum. A third resistance mechanism may arise from an enhanced capacity to synthesize vitamin K from menadione, a commonly used additive in animal foods on farms. The choice of rodenticide in the case of warfarin resistance should be guided by the underlying mechanism of resistance.     

Resistance to anticoagulant rodenticides in Germany and future strategies to control Rattus norvegicus

Widespread anticoagulant resistance was discovered in populations of the brown rat (Rattus norvegicus Berk.) in an area of roughly 8000 km² in north-west Germany. Resistance testing was performed by feeding tests and/or by measuring blood clotting response after intraperitoneal injection of a sublethal testing solution. A hierarchical resistance system was found with warfarin resistance at the base followed by bromadiolone/coumatetralyl resistance to difenacoum resistance at the top. Warfarin resistance was spread over the whole area with reduced incidence towards the edges. Difenacoum resistance represented the highest level found so far and was restricted to the inner zone of the resistance area where it occurred with a frequency of 6% of all individuals tested. Breeding experiments with bromadiolone-resistant rats showed that expression of the bromadiolone resistance gene differed in the two sexes, suggesting additional sex-linked modifying effects to the resistance gene. Control strategy within the resistance area with respect to prevention of further selection for resistance is discussed.     

珠江三角洲地区黄毛鼠抗药性的发生趋势及其Vkorc1基因多态性

为阐明珠江三角洲地区黄毛鼠Rattus losea对第1代抗凝血灭鼠剂的抗性发生趋势及其遗传机制,以杀鼠灵为标准药物,采用致死期食毒法对2017—2021年在广东省江门市捕获的165只黄毛鼠进行生理抗性检测,并测定每只试鼠的维生素K环氧化物还原酶复合物亚单位1(vitamin K epoxide reductase complex subunit 1,Vkorc1)的编码基因序列,分析其突变情况。结果显示,江门市黄毛鼠对第1代抗凝血灭鼠剂杀鼠灵的抗性率为27.03%～50.00%,在黄毛鼠Vkorc1基因中检测到6个不同的突变位点,包括2个错义突变位点Arg58Gly及Tyr139Cys和4个沉默突变位点Ala41Ala、Cys96Cys、Arg98Arg及Ala143Ala,突变率分别为87.27%、0.61%、1.21%、0.61%、1.21%和0.61%,其中Ala143Ala是在黄毛鼠中新发现的沉默突变位点。表明珠江三角洲地区黄毛鼠已对第1代抗凝血灭鼠剂产生了群体抗性并呈上升趋势,第58位的精氨酸突变成甘氨酸(Arg58Gly)是黄毛鼠抗性基因Vkorc1的主要突变位点。     

Allelism of theFcu-1 andFoc genes conferring resistance to fusarium wilt in cucumber

The inheritance of resistance toFusarium oxysporum f.sp.cucumerinum race 1 was determined in the cucumber cv. WIS-248 by analyzing segregation of F₁, F₂, and BC populations of crosses with the susceptible cv. Straight-8. Resistance was conferred by a single dominant gene. In an allelism test, it was proven that theFcu-1 gene, which confers resistance toF. oxysporum f.sp.cucumerinum races 1 and 2 in cucumber cv. SMR-18 and theFoc gene, which confers resistance toF. oxysporum f.sp.cucumerinum race 2 in cucumber cv. WIS-248, are indistinguishable.