Second-generation Anticoagulant Rodenticides Research Articles

Core-shell LC–MS/MS method for quantification of second generation anticoagulant rodenticides diastereoisomers in rat liver in relationship with exposure of wild rats

Second generation anticoagulant rodenticides (SGARs), pesticides used worldwide to control rodent populations, exist in two diastereoisomer chemical species because they own two stereogenic centers. A core-shell LC-MS/MS multi-residue method for comprehensive quantitative analysis of the diastereoisomers of five SGARs as well as three first generation anticoagulant rodenticide molecules has been fully validated in liver of rats according to a bioanalytical guideline. A core-shell column (superficially porous particles) has been chosen for its ability to separate the diastereomers of bromadiolone, difenacoum, brodifacoum, flocoumafen and difethialone and for its robustness to rat liver extracts. The highly selective chromatographic separation of the diastereoisomers contributes to good signal to noise ratios and then enhances the sensitivity of the method compared to the ones of fully porous columns. An elution gradient has been optimized with 10mM ammonium acetate and acetonitrile as aqueous/organic mobile phase respectively. Triple quadrupole mass detector has been used to achieve specifity and LLOQ from 0.92 to 2.2ng/g for each diastereoisomer, or first generation anticoagulant rodenticides. Then we evidenced diastereoisomeric ratios in liver of rats issued from not controlled exposure of wild rats (Rattus norvegicus) trapped in a French Parisian park through a campaign of rodent eradication. We compared them to diastereoisomeric ratios in SGARs commercial baits that contain both isomers, and showed that one of the two diastereoiomers had nearly disappeared in liver of rats. The proportions of cis-bromadiolone and trans-difenacoum were really lowered compared to the baits: 5/7 and 9/12 rats had only trans-bromadiolone and cis-difenacoum hepatic residues respectively. Liver persistence of the two diastereoisomers of bromadiolone and difenacoum was different due to differences in their pharmacokinetics in wild rats. The new core-shell LC-MS/MS method is particularly well adapted for further exploration of diastereoisomers ratios in rodent and predatory wildlife biological samples in order to evaluate ecological consequences of actual baits, to explore new formulated baits with a good balance between efficacity (ability to kill rodents) and diastereoisomers persistence, and hopefully to mitigate exposure of non-target species.

Use of anticoagulant rodenticides by pest management professionals in Massachusetts, USA.

Secondary exposure to chemical rodenticides, specifically second-generation anticoagulant rodenticides (SGARs), poses a threat to non-target wildlife including birds of prey. Federal regulations in the United States currently limit homeowner access to SGARs as a way of minimizing this threat. With legal access to SGARs, pest management professionals (PMPs) represent a potential linkage to non-target exposure. There is limited research focused on rodent control practices, chemical rodenticide preferences, level of concern and awareness, or opinions on rodenticide regulations as they relate to PMPs. An online survey was sent to PMP companies across Massachusetts, USA, between October and November 2015. Thirty-five responses were obtained, a 20 % response rate. The preferred rodent control method among responding PMP companies was chemical rodenticides, specifically the SGAR bromadiolone. Respondents varied in their level of concern regarding the impact of chemical rodenticides on non-target species and showed a low level of awareness regarding SGAR potency and half-life. All responding companies reported using integrated pest management (IPM) strategies, with nearly all utilizing chemical rodenticides at some point. Enhanced education focused on SGAR potency, bioaccumulation potential, exposure routes, and negative impacts on non-target wildlife may improve efforts made by PMPs to minimize risk to wildlife and decrease dependence on chemical rodenticide use. Future studies evaluating use of anticoagulant rodenticide (ARs) by PMPs and the association with AR residues found in non-target wildlife is necessary to determine if current EPA regulations need to be modified to effectively reduce the risk of SGARs to non-target wildlife.

Management of Rodent Populations by Anticoagulant Rodenticides: Toward Third-Generation Anticoagulant Rodenticides.

Second-generation anticoagulant rodenticides (SGARs) have been used since the 1980s for pest management. They are highly efficient even in warfarin-resistant rodents. Nevertheless, because of their tissue persistence, nontarget poisoning by SGARs is commonly described in wildlife. Due to this major problem, a new generation of anticoagulants must be developed to limit this risk. This study proposes a method of developing a new generation of anticoagulant rodenticides by revisiting the old SGARs based on the concept of stereochemistry. Each current SGAR is a mixture of diastereomers. Diastereomers of each compound were purified, and their biologic properties were compared by determining their ability to inhibit vitamin K epoxide reductase (VKOR) activity involved in the activation of vitamin K-dependent clotting factors and their toxicokinetic properties. Systematically, for each SGAR, both diastereomers are as effective in inhibiting VKOR activity. However, their toxicokinetic properties are very different, with one of the two diastereomers always more rapidly cleared than the other one. For all SGARs except flocoumafen, the less persistent diastereomer is always the less predominant isomer present in the current mixture. Therefore, the development of baits containing only the less persistent diastereomer would avoid the ecotoxicological risk associated with their use without decreasing their efficacy.

A negative association between bromadiolone exposure and nestling body condition in common kestrels: management implications for vole outbreaks

Vole outbreaks have been extensively described, along with their impacts on humans, particularly in agricultural areas. The use of rodenticides is a common legal practice to minimise crop damage induced by high vole density for biocidal use. However, rodenticides can have negative direct and indirect impacts on non-target species that feed on voles. We studied whether the use of a second-generation anticoagulant rodenticide (SGAR), bromadiolone, can be detected in the blood of fledglings of wild common kestrels Falco tinnunculus in two areas of central Spain, exploring its possible indirect effects. We found that 16.9% of fledglings had a detectable concentration of bromadiolone in their blood, with an average concentration of 0.248 ± 0.023 ng mL-1 . Fledglings with bromadiolone in their blood, regardless of the concentration, had 6.7% lower body mass than those without detectable bromadiolone. The use of bromadiolone was detectable in the blood of alive non-target species. Detected bromadiolone in blood may reduce the body condition of nestlings, potentially reducing their fitness. The source of bromadiolone found in nestlings needs to be determined in future studies to derive accurate management advice. However, we urge the discontinuation of official SGAR distribution to farmers and their use in agrarian lands to minimise damage of voles on crops, particularly where common kestrels breed, and encourage the use of alternative effective practices. © 2016 Society of Chemical Industry.

Mortality of Western Burrowing Owls ( Athene cunicularia hypugaea) Associated with Brodifacoum Exposure.

Western Burrowing Owls ( Athene cunicularia hypugaea) frequently occupy periurban areas, where they may be exposed to pest control agents. This short communication describes necropsy findings and detected brodifacoum rodenticide levels for four Western Burrowing Owls in Lake Havasu City, Arizona, US, 2013-15. Levels detected ranged from 0.077 mg/kg to 0.497 mg/kg. Brodifacoum, one of several second-generation anticoagulant rodenticides recently removed from the general consumer market, is still available for use by licensed pesticide applicators. Despite recent regulatory actions, second-generation anticoagulant pesticides continue to threaten predatory species in periurban areas.

Increased rodenticide exposure rate and risk of toxicosis in barn owls (Tyto alba) from southwestern Canada and linkage with demographic but not genetic factors.

Among many anthropogenic drivers of population decline, continual rapid urbanization and industrialization pose major challenges for the survival of wildlife species. Barn owls (Tyto alba) in southwestern British Columbia (BC) face a multitude of threats ranging from habitat fragmentation to vehicle strikes. They are also at risk from secondary poisoning of second-generation anticoagulant rodenticides (SGARs), a suite of toxic compounds which at high doses results in a depletion of blood clotting factors leading to internal bleeding and death. Here, using long-term data (N=119) for the hepatic residue levels of SGAR, we assessed the risk of toxicosis from SGAR for the BC barn owl population over the past two decades. We also investigated whether sensitivity to SGAR is associated with genetic factors, namely Single Nucleotide Polymorphisms (SNPs) found in the CYP2C45 gene of barn owls. We found that residue concentration for total SGAR was significantly higher in 2006-2013 (141ng/g) relative to 1992-2003 (57ng/g). The proportion of owls exposed to multiple SGAR types was also significantly higher in 2006-2013. Those measures accordingly translate directly into an increase in toxicosis risk level. We also detected demographic differences, where adult females showed on average lower concentration of total SGAR (64ng/g) when compared to adult males (106ng/g). Juveniles were overall more likely to show signs of toxicosis than adults (33.3 and 6.9%, respectively), and those symptoms were positively predicted by SGAR concentrations. We found no evidence that SNPs in the CYP2C45 gene of barn owls were associated with intraspecific variation in SGAR sensitivity. We recommend several preventative measures be taken to minimize wildlife exposure to SGAR.

LC-MS-MS Analysis of Brodifacoum Isomers in Rat Tissue.

Brodifacoum (BDF) is a second-generation anticoagulant rodenticide structurally related to warfarin but containing two chiral centers. Highly stable, BDF can contaminate food and water supplies causing accidental poisoning of humans and nontarget animals. To determine the distribution of BDF isomers in serum and tissues, a quantitative method was developed and validated according to FDA guidelines based on high-performance liquid chromatography-tandem mass spectrometry. A single liquid-liquid extraction step provided recoveries exceeding 93%. Reversed-phase chromatographic separations required <6 min, and quantitative analysis utilized a triple-quadrupole mass spectrometer equipped with negative ion electrospray and selected reaction monitoring. The standard curve had a linear regression coefficient of 0.999 and intra- and inter-assay variations of <10%. The chromatographic method enabled the resolution and measurement of pairs of BDF diastereomers in commercial materials as well as in rat tissues. This method is suitable for measuring BDF exposure as well as basic science studies of the distribution and elimination of BDF diastereomers to various tissues.

Usefully Persistent? Anticoagulants as Quantitative Markers of Bait Uptake in Brushtail Possums

Prolonged persistence of second-generation anticoagulant rodenticides (SGARs) in animal tissues facilitates trophic transfer of residues, with exposure of predatory and scavenging non-target wildlife now widely reported. In many instances, antico-agulant residue levels measured in wildlife are apparently sublethal, although longer-term effects of such exposures are currently not well understood. Conversely, prolonged metabolic persistence is of practical utility when compounds are used as biological markers to determine food uptake by animals. We required two effective and distinct marker compounds to progress field-based research on optimal baiting strategies to manage introduced brushtail possums in New Zealand. Two SGARs (flocoumafen and bromadiolone) were evaluated, as neither are currently registered for application as rodenticides in areas typically subject to possum management. All captive possums ingesting small, sublethal (2-6 g) quantities of food containing 0.0005% (by weight) of either SGAR were reliably marked by the presence of residual concentrations in liver as measured by HPLC analysis. This marking persisted for at least six weeks, and liver concentrations in marked possums did not decline between three and six weeks after the marker was ingested. Marking was also quantitative for both SGARs with a correlation between liver residue concentration and the amount of marker ingested. Using such marker baits would provide at least a six-week period in which to recover possums in field research. By recording bodyweight and testing liver samples from such possums for both markers, regression equations generated from the work reported here will enable back-estimation, with confidence intervals, of the amounts of marker bait eaten by each possum.

New Rodenticide on the Block: Diagnosing Bromethalin Intoxication in Wildlife

Bromethalin is being used more widely for commensal rodent control because of increased regulation on second-generation anticoagulant rodenticides. Wildlife losses in California are tracked by the California Department of Fish and Wildlife. Bromethalin is a neurotoxicant which is not thought to cause secondary poisoning. From August 2014 to January 2016, 24 cases of bromethalin intoxication were investigated in California. These include 11 raccoons, 11 striped skunks, one gray fox, and one fox squirrel. Most of these occurred in Marin County, where active surveillance of wildlife for rodenticide exposure is occurring. Bromethalin exposure should be evaluated when a wild animal that may have accessed bait is showing neurological signs. Trauma and distemper should be ruled out. Histological changes may be found in the central nervous system but are not always present. The tissue of choice for toxicological analysis is adipose. It is likely that bromethalin intoxication is under-reported in the rest of the state and may be mistaken for distemper infection or trauma. Primary exposure of wildlife to bromethalin could be prevented by placing baits in tamper-resistant bait stations.

Land Use as a Driver of Patterns of Rodenticide Exposure in Modeled Kit Fox Populations.

Although rodenticides are increasingly regulated, they nonetheless cause poisonings in many non-target wildlife species. Second-generation anticoagulant rodenticide use is common in agricultural and residential landscapes. Here, we use an individual-based population model to assess potential population-wide effects of rodenticide exposures on the endangered San Joaquin kit fox (Vulpes macrotis mutica). We estimate likelihood of rodenticide exposure across the species range for each land cover type based on a database of reported pesticide use and literature. Using a spatially-explicit population model, we find that 36% of modeled kit foxes are likely exposed, resulting in a 7-18% decline in the range-wide modeled kit fox population that can be linked to rodenticide use. Exposures of kit foxes in low-density developed areas accounted for 70% of the population-wide exposures to rodenticides. We conclude that exposures of non-target kit foxes could be greatly mitigated by reducing the use of second-generation anticoagulant rodenticides in low-density developed areas near vulnerable populations.

Toxicity and bioaccumulation of bromadiolone to earthworm Eisenia fetida

Bromadiolone, a potent second-generation anticoagulant rodenticide, has been extensively used worldwide for the field control of rodents. Invertebrates may be at risk from primary poisoning as a result of bromadiolone bait applications. However, there are few data regarding the toxicity and bioaccumulation of bromadiolone to earthworms. In this study, we reported that bromadiolone was toxic to earthworms at 1mg/kg soil, which is a likely concentration in the field following application of bromadiolone baits. Exposure to bromadiolone resulted in a significant inhibition of earthworm growth. The antioxidant activities of superoxide dismutase and catalase were slightly increased in earthworms, while malondialdehyde content (as a molecular marker indicative of the damage to lipid peroxidation) was dominantly elevated over the duration of exposure. Bromadiolone in soil is bioaccumulative to earthworms. The biota to soil accumulation factors (BSAFs) of bromadiolone were concentration dependent and BSAFs decreased as the level of bromadiolone in soil increased. These results suggest earthworms are not only the potential subject to primary poisoning but also the source of secondary exposure for insectivores and scavengers following application of bromadiolone.

Assessment of toxicity and coagulopathy of brodifacoum in Japanese quail and testing in wild owls.

Based on detection of hepatic residues, scavenging and predatory non-target raptors are widely exposed to second generation anticoagulant rodenticides (SGARs). A small proportion, generally <10%, of tested birds are diagnosed as acutely poisoned. Little is known, however, of sub-lethal effects of SGARs, such as interaction of clotting capacity with traumatic injury. Assessment of coagulation function of birds submitted live to wildlife rehabilitators or veterinarians may provide a means of establishing the proportion of animals suffering sub-lethal coagulopathies, as well as identifying individuals requiring treatment. As a first step in exploring the potential of this approach, we dosed Japanese quail (Coturnix japonica) with the SGAR, brodifacoum, at 0, 0.8, 1.4, 1.9, and 2.5 mg/kg and sampled birds at 1, 3, 5 and 7 days post-dosing. Prothrombin time (PT), which measures the extrinsic coagulation pathway, was significantly prolonged in 98% of brodifacoum-exposed quail in a dose- and time-dependent manner. 50-fold prolongation of PT occurred at higher brodifacoum dosages and correlated to hemorrhage found at necropsy. Activated clotting time (ACT), a measure of the intrinsic pathway also increased with dose and time. Hemoglobin (Hb) and hematocrit (Hct) decreased dose- and time-dependently at doses ≥1.4 mg/kg with no significant change at 0.8 mg/kg. Reference intervals for PT (10.0-16.2 s), ACT (30-180 s), Hb (9.6-18.4 g/dl), and Hct (34-55%) were established in Japanese quail. Species-specific reference intervals are required as barn owl PT (17-29 s) and quail PT were different. The proportion of brodifacoum-exposed quail with hemorrhage was not correlated with liver residues, but was correlated with PT, suggesting that this assay is a useful indicator of avian anticoagulant rodenticide exposure. PTs measured in free-living barn owls sampled between April 2009 and August 2010 in the lower Fraser Valley of BC do not suggest significant exposure to SGARs.

Toxicity reference values for chlorophacinone and their application for assessing anticoagulant rodenticide risk to raptors.

Despite widespread use and benefit, there are growing concerns regarding hazards of second-generation anticoagulant rodenticides to non-target wildlife which may result in expanded use of first-generation compounds, including chlorophacinone (CPN). The toxicity of CPN over a 7-day exposure period was investigated in American kestrels (Falco sparverius) fed either rat tissue mechanically-amended with CPN, tissue from rats fed Rozol(®) bait (biologically-incorporated CPN), or control diets (tissue from untreated rats or commercial bird of prey diet) ad libitum. Nominal CPN concentrations in the formulated diets were 0.15, 0.75 and 1.5µg/g food wet weight, and measured concentrations averaged 94% of target values. Kestrel food consumption was similar among groups and body weight varied by less than 6%. Overt signs of intoxication, liver CPN residues, and changes in prothrombin time (PT), Russell's viper venom time (RVVT) and hematocrit, were generally dose-dependent. Histological evidence of hemorrhage was present at all CPN dose levels, and most frequently observed in pectoral muscle and heart. There were no apparent differences in toxicity between mechanically-amended and biologically-incorporated CPN diet formulations. Dietary-based toxicity reference values at which clotting times were prolonged in 50% of the kestrels were 79.2µg CPN consumed/kg body weight-day for PT and 39.1µg/kg body weight-day for RVVT. Based upon daily food consumption of kestrels and previously reported CPN concentrations found in small mammals following field baiting trials, these toxicity reference values might be exceeded by free-ranging raptors consuming such exposed prey. Tissue-based toxicity reference values for coagulopathy in 50% of exposed birds were 0.107µg CPN/g liver wet weight for PT and 0.076µg/g liver for RVVT, and are below the range of residue levels reported in raptor mortality incidents attributed to CPN exposure. Sublethal responses associated with exposure to environmentally realistic concentrations of CPN could compromise survival of free-ranging raptors, and should be considered in weighing the costs and benefits of anticoagulant rodenticide use in pest control and eradication programs.

Nontarget mortality of New Zealand lesser short-tailed bats (Mystacina tuberculata) caused by diphacinone.

Primary and secondary poisoning of nontarget wildlife with second-generation anticoagulant rodenticides has led to restrictions on their use and to increased use of first-generation anticoagulants, including diphacinone. Although first-generation anticoagulants are less potent and less persistent than second-generation compounds, their use is not without risks to nontarget species. We report the first known mortalities of threatened New Zealand lesser short-tailed bats (Mystacina tuberculata) caused by diphacinone intoxication. The mortalities occurred during a rodent control operation in Pureora Forest Park, New Zealand, during the 2008-09 Austral summer. We observed 115 lesser short-tailed bat deaths between 9 January and 6 February 2009, and it is likely that many deaths were undetected. At necropsy, adult bats showed gross and histologic hemorrhages consistent with coagulopathy, and diphacinone residues were confirmed in 10 of 12 liver samples tested. The cause of mortality of pups was diagnosed as a combination of the effects of diphacinone toxicity, exposure, and starvation. Diphacinone was also detected in two of 11 milk samples extracted from the stomachs of dead pups. Eight adults and 20 pups were moribund when found. Two adults and five pups survived to admission to a veterinary hospital. Three pups responded to treatment and were released at the roost site on 17 March 2009. The route of diphacinone ingestion by adult bats is uncertain. Direct consumption of toxic bait or consumption of poisoned arthropod prey could have occurred. We suggest that the omnivorous diet and terrestrial feeding habits of lesser short-tailed bats make them susceptible to poisoning with the bait matrix and the method of bait delivery used. We recommend the use of alternative vertebrate pesticides, bait matrices, and delivery methods in bat habitat.

Residue profiles of brodifacoum in coastal marine species following an island rodent eradication

The second-generation anticoagulant rodenticide brodifacoum is an effective tool for the eradication of invasive rodents from islands and fenced sanctuaries, for biodiversity restoration. However, broadcast application of brodifacoum bait on islands may expose non-target wildlife in coastal marine environments to brodifacoum, with subsequent secondary exposure risk for humans if such marine wildlife is harvested for consumption. We report a case study of monitoring selected marine species following aerial application of brodifacoum bait in August 2011 to eradicate Norway rats (Rattus norvegicus) from Ulva Island, New Zealand. Residual concentrations of brodifacoum were detected in 3 of 10 species of coastal fish or shellfish sampled 43–176d after bait application commenced. Residual brodifacoum concentrations were found in liver, but not muscle tissue, of 2 of 24 samples of blue cod (0.026 and 0.092µg/g; Parapercis colias) captured live then euthanized for tissue sampling. Residual brodifacoum concentrations were also found in whole-body samples of 4 of 24 mussels (range=0.001–0.022µg/g, n=4; Mytilus edulis) and 4 of 24 limpets (range=0.001–0.016µg/g, n=4; Cellana ornata). Measured residue concentrations in all three species were assessed as unlikely to have eventually caused mortality of the sampled individuals. We also conducted a literature review and determined that in eleven previous accounts of residue examination of coastal marine species following aerial applications of brodifacoum bait, including our results from Ulva Island, the overall rate of residue detection was 5.6% for marine invertebrates (11 of 196 samples tested) and 3.1% for fish (2 of 65 samples tested). Furthermore, our results from Ulva Island are the first known detection of brodifacoum residue in fish liver following an aerial application of brodifacoum bait. Although our findings confirm the potential for coastal marine wildlife to be exposed to brodifacoum following island rodent eradications using aerial bait application, the risk of mortality to exposed individual fish or shellfish appears very low. There is also a very low risk of adverse effects on humans that consume fish or shellfish containing residual concentrations in the ranges reported here. Furthermore, any brodifacoum residues that occur in marine wildlife decline to below detectable concentrations over a period of weeks. Thus potential human exposure to brodifacoum through consumption of marine wildlife containing residual brodifacoum could be minimized by defining ‘no take’ periods for harvest following bait application and regular monitoring to confirm the absence of detectable residues in relevant marine wildlife.

Dispatches

Dispatches

A specialist in the city: the diet of barn owls along a rural to urban gradient

We investigated the variations in the diets of barn owls (Tyto alba) along a rural to urban gradient in the Lower Fraser Valley of southwestern British Columbia, Canada. Our objectives were to determine the food preferences of barn owls in different habitat types and to assess both seasonal and annual variations in their diets. In particular, given the increased incidences and concentrations of second generation anticoagulant rodenticide (SGAR) residues in barn owls, we were interested in determining whether there was any relationship between the degree of urbanization surrounding their nest/roost sites and the proportion of commensal rodents (Rattus norvegicus, Rattus rattus, Mus musculus) in their diet. In total, 8,941 individual prey remains were identified. The proportion of rats consumed increased significantly with the amount of urbanization within home ranges. However, voles (primarily field voles, Microtus townsendii) were the main prey item for all sites irrespective of surrounding land use within home ranges. Shrews were the second most consumed prey species (10.8 ± 8 %), and were found predominantly in the diet of barn owls nesting in more rural landscapes. The dominance of field voles in the diet was also reflected in the food-niche breadth, which was consistently low for all sites across the rural to urban gradient. Interestingly, barn owls were found nesting in highly urban environments (66–95 % urbanization), where bait stations containing SGARs were regularly used. This, combined with the increased consumption of rats in more urban environments, illustrates why the SGAR exposure rate in barn owls is substantial and rising.

Adverse outcome pathway and risks of anticoagulant rodenticides to predatory wildlife.

Despite a long history of successful use, routine application of some anticoagulant rodenticides (ARs) may be at a crossroad due to new regulatory guidelines intended to mitigate risk. An adverse outcome pathway for ARs was developed to identify information gaps and end points to assess the effectiveness of regulations. This framework describes chemical properties of ARs, established macromolecular interactions by inhibition of vitamin K epoxide reductase, cellular responses including altered clotting factor processing and coagulopathy, organ level effects such as hemorrhage, organism responses with linkages to reduced fitness and mortality, and potential consequences to predator populations. Risk assessments have led to restrictions affecting use of some second-generation ARs (SGARs) in North America. While the European regulatory community highlighted significant or unacceptable risk of ARs to nontarget wildlife, use of SGARs in most EU member states remains authorized due to public health concerns and the absence of safe alternatives. For purposes of conservation and restoration of island habitats, SGARs remain a mainstay for eradication of invasive species. There are significant data gaps related to exposure pathways, comparative species sensitivity, consequences of sublethal effects, potential hazards of greater AR residues in genetically resistant prey, effects of low-level exposure to multiple rodenticides, and quantitative data on the magnitude of nontarget wildlife mortality.

Assessment of anticoagulant rodenticide exposure in six raptor species from the Canary Islands (Spain)

Anticoagulant rodenticides are highly toxic compounds that are widely used for pest control of rodents, but that also may threaten the wildlife's health. This work aimed to assess the exposure to first- and second-generation anticoagulant rodenticides (ARs) in six birds of prey species from the Canary Islands (Spain). The concentrations of seven widely used ARs were determined by LC–MS/MS in 104 liver samples of six species of birds of prey (Buteo buteo, Accipiter nisus, Falco pelegrinoides, Falco tinnunculus, Asio otus, and Tyto alba). We determined that 61% of the livers had detectable residues of at least one AR. The most frequently detected AR was bromadiolone, which was detected in 60.3% of the positive cases. The detection frequencies of these compounds varied widely, depending on the species. More than 75% of the A. nisus, T. alba, and A. otus individuals had detectable rodenticide residues in the liver. However, F. tinnunculus exhibited the highest concentrations of AR, with median values above 100ng/g w.w. We did not detect first-generation ARs in any of the samples. When grouped, nocturnal species exhibited higher AR concentrations than diurnal species (P<0.001). The residue levels were higher among small mammal-eaters than bird-eaters (P<0.01). While most animals exhibited no macroscopic signs of coagulation disorders, approximately 35% exceeded the threshold levels of toxicity, which suggests that these compounds could weaken these animals in their natural environment. In conclusion, the control of rodent populations by ARs suggests that these compounds will enter the food chain and thus threaten the vulnerable populations of raptors on the Canary Islands. Our findings require authorities to ban or strictly control the use of these rodenticides in the natural environment for the conservation of raptors and other predatory species.

Evaluation of a point‐of‐care anticoagulant rodenticide test for dogs

To evaluate a point-of-care anticoagulant rodenticide lateral flow analyzer for the detection of various rodenticide compounds. Prospective, laboratory study. University teaching hospital. The study utilized a serum sample from one healthy canine donor. Samples were centrifuged and serum samples were aliquoted and either used within 4 hours or frozen at -70°C for further quantitative analysis. Samples were spiked with clinically relevant concentrations of 1 of 6 rodenticide compounds (warfarin, pindone, chlorphacinone, brodifacoum, bromethalin, and its metabolite desmethylbromethalin). Seventy-five microliters of spiked serum (or unaltered serum) was introduced into the lateral flow test. Three readers who were blinded to the sample preparation interpreted the lateral flow test as either positive or negative for the presence of anticoagulant rodenticide. All readers were in agreement for the results of each serum sample. The point-of-care test kit was able to detect a single anticoagulant rodenticide (warfarin) at concentrations below the manufacturer's recommended limit of detection, but was unable to detect any other anticoagulant rodenticide. The results of this test and therapeutic interventions must be considered in light of history, physical examination, and other clinical data. Based on results from this study, the test kit only detects warfarin and not other more common second-generation anticoagulant rodenticides.