The stop-feed effect of cholecalciferol (vitamin D3) and the efficacy of brodifacoum combined with cholecalciferol in Y139C-resistant Norway rats (Rattus norvegicus).

Second-generation anticoagulant rodenticides potentially build persistent residues in animals and accordingly pose a risk of secondary poisoning. We examined the effect of a low concentration of cholecalciferol in brodifacoum bait on bait consumption by Norway rats (Rattus norvegicus Berkenhout 1769) and on the control success in a laboratory study and in field trials. Additionally, the efficacy of both baits was determined against resistant Y139C rats. Cholecalciferol caused a strong stop-feed effect after two days in the laboratory study. On two field study sites each, bait containing either 25 mg kg-1 brodifacoum or 25 mg kg-1 brodifacoum and 100 mg kg-1 cholecalciferol was applied to treat infestations of Norway rats. Infestations were assessed pre- and post-treatment. Rats were radio-tagged, and carcasses were searched for during the treatment period. DNA of each rat was genotyped to determine the resistance status conferred by the VKORC1 gene. On all farms, control success exceeded 90%. On farms treated with brodifacoum only, the ratio of total bait consumption to pre-treatment census was significantly higher (6.6 and 4.8 times) than on farms treated with the combination (2.7 and 2.9 times). 78.8% of 183 rats were confirmed Y139C resistant. Bait ingestion was reduced by almost fifty per-cent when cholecalciferol was added to the bait with no impact on control success. All treatments resulted in control levels exceeding 90%, despite a high proportion of anticoagulant-resistant rats. When the use of highly toxic compounds is required in resistance management, addition of cholecalciferol to these baits may reduce the transfer of residues to the environment.

Sanitary measures considerably improve the management of resistant Norway rats on livestock farms.

BACKGROUND: Norway rats (Rattus norvegicus) need to be controlled to prevent transmission of pathogens and damages to stored products and material, leading to considerable economic risks and losses. Given increasing resistance in Norway rats, the most persistent, bio-accumulative and toxic anticoagulant rodenticides are widely used for management, which presents hazards to the environment especially for non-target species. We investigated how sanitary measures improved management of Norway rats on 12 paired livestock farms in a region of Germany with a high population of resistant rats for reducing application of rodenticides. We recorded food intake, and tracked activity and resistance frequency during the pre-treatment, treatment and post-treatment periods. RESULTS: In the post-treatment period, farms using sanitary measures had a higher control success with > 13% more bait boxes without feeding than farms not using sanitary measures. In addition, the reoccurrence of rats was delayed by 85 days. With increasing accessibility to buildings and more precise positioning of the boxes, control success improved, especially when rats could not spread from water-bearing ditches through the sewer system, and when rat-hunting animals were present. Resistant animals were more common indoors than outdoors, and there were more resistant rats recorded before and during treatment than in the post-treatment period. CONCLUSION: The control success was substantially higher and reoccurrence was delayed using sanitary measures on farms. Sanitary measures can reduce resistance indirectly due to delayed re-colonization and establishment of resistant populations inside buildings. Hence, sanitary measures help to reduce economic losses, rodenticides required for rat management and environmental risk especially in the resistance area. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Effects of anticoagulant rodenticide poisoning on spatial behavior of farm dwelling Norway rats.

Commensal rodent species cause damage to crops and stored products, they transmit pathogens to people, livestock and pets and threaten native flora and fauna. To minimize such adverse effects, commensal rodents are predominantly managed with anticoagulant rodenticides (AR) that can be transferred along the food chain. We tested the effect of the uptake of the AR brodifacoum (BR) by Norway rats (Rattus norvegicus) on spatial behavior because this helps to assess the availability of dead rats and residual BR to predators and scavengers. BR was delivered by oral gavage or free-fed bait presented in bait stations. Rats were radio-collared to monitor spatial behavior. BR residues in rat liver tissue were analyzed using liquid chromatography coupled with tandem mass spectrometry. Norway rats that had consumed BR decreased distances moved and had reduced home range size. Treatment effects on spatial behavior seemed to set in rapidly. However, there was no effect on habitat preference. Ninety-two percent of rats that succumbed to BR died in well-hidden locations, where removal by scavenging birds and large mammalian scavengers is unlikely. Rats that ingested bait from bait stations had 65% higher residue concentrations than rats that died from dosing with two-fold LD50. This suggests an overdosing in rats that are managed with 0.0025% BR. None of the 70 BR-loaded rats was caught/removed by wild predators/scavengers before collection of carcasses within 5-29 h. Therefore, and because almost all dead rats died in well-hidden locations, they do not seem to pose a significant risk of AR exposure to large predators/scavengers at livestock farms. Exposure of large predators may originate from AR-poisoned non-target small mammals. The few rats that died in the open are accessible and should be removed in routine searches during and after the application of AR bait to minimize transfer of AR into the wider environment.

The potential of coumatetralyl enhanced by cholecalciferol in the control of anticoagulant-resistant Norway rats (Rattus norvegicus).

BACKGROUND: We evaluated the potential of cholecalciferol as an enhancer of the first-generation anticoagulant coumatetralyl in the Westphalia anticoagulant-resistant strain of the Norway rat (Rattus norvegicus Berkenhout), characterised by the Tyr139Cys polymorphism on the VKOR enzyme. Because today only the most potent, but also most persistent anticoagulant rodenticides of the second generation remain available to control this strain, new rodenticide solutions are required. RESULTS: Feeding trials in the laboratory confirmed a significant level of efficacy, which was corroborated by field trials in the Münsterland resistance area. After frequency and level of resistance were assessed by blood clotting response tests, field trials were conducted with bait containing coumatetralyl at 375 mg kg-1 and cholecalciferol at 50 mg kg-1 or 100 mg kg-1 . Control success was 94% when a large rat infestation comprising 42% resistant animals was treated. Another field trial applying the combination to a rat population that had survived a preceding treatment with bromadiolone resulted in a 99.5% control success according to the first census day, but with some increase in rat activity during subsequent census days. CONCLUSION: The combination of coumatetralyl and cholecalciferol is a promising alternative approach to the most potent second-generation anticoagulants in resistance management, particularly in respect of environmental risks, such as secondary poisoning. © 2016 Society of Chemical Industry.

Resistance testing and the effectiveness of difenacoum against Norway rats (Rattus norvegicus) in a tyrosine139cysteine focus of anticoagulant resistance, Westphalia, Germany.

BACKGROUND: Anticoagulant resistance in Norway rats at foci in Belgium, Denmark, France, Germany, the Netherlands and the United Kingdom is genetically characterised by the same single nucleotide polymorphism (SNP) and consequent amino acid exchange from tyrosine to cysteine at location 139 of the vkorc1 gene (i.e. tyrosine139cysteine or Y139C). The purpose of this study was to assess the degree of resistance among rats at two infested farm sites in the Y139C focus in Westphalia, Germany, using blood clotting response (BCR) tests, and to determine the practical efficacy of applications of a commercial 50 ppm difenacoum bait (Neokil™) against them. RESULTS: BCR tests showed that the difenacoum resistance factor (RF) among the Y139C rats was about 2.5. DNA analysis for the Y139C mutation revealed that it was present among rats at the two sites with a prevalence of 75 and 93%. Applications of difenacoum bait at the two sites achieved 86.8 and 59.9% control. The different outcomes did not appear to be due to differences either in the degree and prevalence of resistance or in the quantities of poisoned bait consumed. CONCLUSION: The study showed that, although the RF for difenacoum among rats carrying the Y139C SNP was apparently low, an acceptable level of control of resistant Norway rat infestations was not achieved using difenacoum. Continued use of anticoagulants against rats that are resistant to them will exacerbate resistance problems in terms of both increased severity and prevalence. These conclusions are likely to apply elsewhere in Europe where the Y139C SNP occurs.

Vkorc1 variation in house mice during warfarin and difenacoum field trials.

BACKGROUND: Field studies guided by genetic monitoring of Vkorc1 need to be done to implicate mutations conclusively with rodent control problems due to the presence of animals resistant to anticoagulant rodenticides. Rodent control success in relation to Vkorc1 genotypes in house mice (Mus musculus domesticus) was studied on two farms (I and II) in Germany. Tests were carried out to determine whether certain resistance profiles and Vkorc1 genotypes displayed dynamics over the course of sequential treatments with warfarin and difenacoum that were consistent with single nucleotide polymorphisms (SNPs) in Vkorc1 as indicators of resistance. RESULTS: On farms I and II, respectively, three (A to C) and two (A and B) types of control problem with anticoagulants (i.e. proxies for resistance) were encountered in spatially segregated subunits: A = none; B = control problems with warfarin but not with difenacoum; C = control problems with both anticoagulants. Unexpectedly, resistance was encountered in a population where only Vkorc1 wild-type mice were detected. In addition, the Arg58Gly Vkorc1 variant was found not to correlate with observed control failures. CONCLUSION: Control problems were encountered that cannot be explained by Vkorc1 coding or intronic SNPs, and therefore are likely due to non-coding Vkorc1 SNPs or due to other genetic or non-genetic factors.

Brodifacoum is effective against Norway rats (Rattus norvegicus) in a tyrosine139cysteine focus of anticoagulant resistance in Westphalia, Germany.

BACKGROUND: The tyrosine to cysteine amino acid substitution at location 139 of the vkorc1 protein (i.e. tyrosine139cysteine or Y139C) is the most widespread anticoagulant resistance mutation in Norway rats (Rattus norvegicus Berk.) in Europe. Field trials were conducted to determine the incidence of the Y139C mutation at two rat-infested farms in Westphalia, Germany, and to estimate the practical efficacy against them of applications, using a pulsed baiting treatment regime, of a proprietary bait (Klerat™) containing 0.005% brodifacoum. RESULTS: DNA analysis for the Y139C mutation showed that resistant rats were prevalent at the two farms, with an incidence of 80.0 and 78.6% respectively. Applications of brodifacoum bait achieved results of 99.2 and 100.0% control at the two farms, when measured by census baiting, although the treatment was somewhat prolonged at one site, possibly owing to the abundance of attractive alternative food. CONCLUSION: The study showed that 0.005% brodifacoum bait is fully effective against Norway rats possessing the Y139C mutation at the Münsterland focus and is likely to be so elsewhere in Europe where this mutation is found. The pulsed baiting regime reduced to relatively low levels the quantity of bait required to control these two substantial resistant Norway rat infestations. Previous studies had shown much larger quantities of bromadiolone and difenacoum baits used in largely ineffective treatments against Y139C resistant rats in the Münsterland. These results should be considered when making decisions about the use of anticoagulants against resistant Norway rats and their potential environmental impacts.

Resistance tests and field trials with bromadiolone for the control of Norway rats (Rattus norvegicus) on farms in Westphalia, Germany.

BACKGROUND: The aim of this study was to determine the incidence and the level of resistance to bromadiolone among rats on farms suspected of being foci of resistance by using the international normalised ratio (INR)-based blood clotting response (BCR) test. Whether the level of reduced susceptibility constitutes 'practical resistance' was subsequently determined in field trials. RESULTS: The 2.5 multiple of the ED(50) baseline was used to test for the incidence of resistance, and higher multiples in the range of the suspected resistance factor were used to investigate the degree of resistance. The ED(50) values of bromadiolone in resistant rats were confirmed in the range 4.70-7.05 mg kg(-1) for males and 4.62-6.61 mg kg(-1) for females. Variations within these ranges appeared between farms. According to the BCR resistance tests, 50-100% of rats were classified as resistant prior to the field trials; 29-100% of rats survived the treatments. CONCLUSION: BCR tests based on the use of the INR and baselines are suitable for determining the incidence and for assessing the level of resistance in populations of Norway rats. The majority of rats of the Westphalian resistant strain, characterised by the Y139C marker in VKOR, are resistant to bromadiolone under practical control conditions.

Adaptive introgression of anticoagulant rodent poison resistance by hybridization between old world mice.

Polymorphisms in the vitamin K 2,3-epoxide reductase subcomponent 1 (vkorc1) of house mice (Mus musculus domesticus) can cause resistance to anticoagulant rodenticides such as warfarin [1-3]. Here we show that resistant house mice can also originate from selection on vkorc1 polymorphisms acquired from the Algerian mouse (M. spretus) through introgressive hybridization. We report on a polymorphic introgressed genomic region in European M. m. domesticus that stems from M. spretus, spans >10 Mb on chromosome 7, and includes the molecular target of anticoagulants vkorc1 [1-4]. We show that in the laboratory, the homozygous complete vkorc1 allele of M. spretus confers resistance when introgressed into M. m. domesticus. Consistent with selection on the introgressed allele after the introduction of rodenticides in the 1950s, we found signatures of selection in patterns of variation in M. m. domesticus. Furthermore, we detected adaptive protein evolution of vkorc1 in M. spretus (Ka/Ks = 1.54-1.93) resulting in radical amino acid substitutions that apparently cause anticoagulant tolerance in M. spretus as a pleiotropic effect. Thus, positive selection produced an adaptive, divergent, and pleiotropic vkorc1 allele in the donor species, M. spretus, which crossed a species barrier and produced an adaptive polymorphic trait in the recipient species, M. m. domesticus.