Sylvatic plague, caused by the flea-borne bacterium Yersinia pestis, is an invasive disease in North America that causes reductions of native fauna and transforms ecosystems. Fipronil baits have shown promise in reducing flea loads on prairie dogs Cynomys spp. for plague mitigation. Many species depend on prairie dogs and their ecological influences, including the black-footed ferret Mustela nigripes (ferret), an obligate predator of prairie dogs. To better understand how fipronil affects ferrets, we offered carcass portions from black-tailed prairie dogs C. ludovicianus that had consumed fipronil bait (0.005% fipronil by weight) to captive ferrets and monitored their health. We fed carcass portions of three prairie dogs to four adult ferrets for 1 week. No ill effects were observed in the ferrets. We collected scat from the ferrets before, during, and after their feeding on treated prairie dogs. We evaluated potential effects of ferret scat on larval fleas, which feed on organic matter. Fipronil residues were not detected in ferret scat samples collected before treatment. During and shortly after treatment, ferret scat contained 3.76 ng/g fipronil and 13.75 ng/g fipronil sulfone, on average, demonstrating trophic transfer of the residues from prey to predator. We presented 0.5 mg of ferret scat to each of 96 larval Oropsylla montana (Siphonaptera: Ceratophyllidae) and assessed survival rates over 24 h. When exposed to ferret scat lacking fipronil residues, 85% of larvae survived. Survival was reduced to 61% and 35% for larvae contacting or consuming scat with fipronil residue, respectively. Fipronil residues in scat from a variety of species on prairie dog colonies, perhaps especially the prairie dogs, may assist in flea control and plague mitigation. Hosts eliminate fipronil residues, and fipronil residues in the environment degrade over time, reducing but not eliminating potential concerns with bioaccumulation.

The bacterium Yersinia pestis is relatively new to North America, having been unintentionally introduced by humans in the early 1900s (McCoy 1908). The wildlife disease caused by this bacterium, sylvatic plague, is widely understood to cause catastrophic die offs of some native wildlife species (Gage and Kosoy 2005). Rodents can be particularly susceptible, and species that depend on rodents for their own survival, such as the endangered (ESA 1973, as amended) black-footed ferret Mustela nigripes, can be directly and indirectly affected. The primary transmission route of Yersinia pestis between individual mammals is through fleabites (Hinnebusch et al. 2017) and the life habits of many rodents allow for a continuum of plague transmission from enzootic disease maintenance (with host killing) to epizootic outbreaks (with mass host mortality; Biggins and Eads 2019). To counteract the spread of this non-native disease, biologists often focus on reducing flea populations using insecticides. Treatments can be administered to rodent refugia, such as burrows, which may produce limited flea control in some cases due to the complexities of burrow systems (Biggins et al. 2010, 2021; Matchett et al. 2010), or treatments can be administered via systemic pulicides provided to rodents as baits, which allows the rodents to carry pulicide residues deep into their burrows or wherever else they may travel (Eads et al. 2019, 2021, 2023; Matchett et al. 2023).

An emerging tool in the science of plague management is fipronil-laced baits. When these baits are consumed by a mammal, fipronil (a phenylpyrazole) enters the blood stream and is then passed to ectoparasitic blood feeders when they feed on the host, causing hyperexcitation, paralysis, and mortality of some ectoparasites, including fleas (Page 2008; Pfister and Armstrong 2016; Eads et al. 2020). Fipronil grain (0.005% fipronil by weight; Poché et al. 2017, 2020) is a commercially available product that is used to control fleas that vector the plague bacterium in efforts to minimize plague in prairie dog Cynomys spp. colonies. Some of these prairie dog colonies are also home to the black-footed ferret (ferret), obligate predator of three prairie dog species. Scientists and site managers have seen encouraging results using fipronil-laced baits, with effective flea control up to 2 years in some cases after a single treatment (Eads et al. 2019;, 2020, 2021, 2022b; Matchett et al. 2023).

Fipronil baits consumed by prairie dogs are thought to suppress flea populations long-term (sometimes >2 years) because they can reduce flea populations in at least two life stages: adult blood feeders and larval scat feeders (Eads et al. 2023a). After rodents such as prairie dogs eat fipronil bait, the active ingredient is metabolized into fipronil sulfone and the residues are temporarily stored in the animal’s fat tissues, from which the residues are released into the bloodstream (Cravedi et al. 2013). Until those residues are fully eliminated, they can pass to predators, parasites and scavengers that feed on treated animals. Preliminary data from our research indicates black-tailed prairie dogs C. ludovicianus can retain fipronil sulfone residues in the body for an estimated 40–60 d. Prairie dogs eliminate fipronil and fipronil sulfone in scat and urine (Cravedi et al 2013). Prairie dogs commonly drop scat in their dark burrows (Wilcomb 1954; Hoogland 1995). Fipronil residues degrade fairly quickly in sunlight, but the half-life in soils of dark tunnel systems may range from 3–7.3 mo (Ying and Kookana 2002; U.S. Environmental Protection Agency 1996; Bonmatin et al. 2015). In a laboratory setting, black-tailed prairie dog scat with fipronil residues killed most larval Oropsylla montana fleas within 24 h as they contacted and often consumed the scat (Eads et al. 2023a).

The ferret is highly susceptible to plague (Godbey et al. 2006; Abbott and Rocke 2012) and, therefore, most reintroduction sites for the species are intensively managed to minimize plague (USFWS 2016a). Where fipronil baits are used and ferrets are present, ferrets will likely prey on prairie dogs (Figure 1) that previously consumed fipronil bait, including prairie dogs still harboring fipronil residues. Consumption of treated prey might allow for transfer of fipronil residues to ferrets, potentially providing temporary protection from fleas and plague or perhaps posing unintended negative health consequences, such as behavioral changes that have been observed in other species under laboratory conditions (e.g., Udo et al. 2014).

Figure 1.

Prairie dogs Cynomys spp. are the primary food source of the black-footed ferret Mustela nigripes. Ferrets are therefore affected by diseases that affect prairie dog populations, such as flea-borne sylvatic plague, as well as treatments applied to prairie dogs to manage those diseases. Fipronil-laced baits can be used to control flea populations on prairie dogs and residues from fipronil consequently pass into ferrets when a treated prairie dog is consumed. Those treatments can then possibly affect ferrets and further assist with flea control and disease management. Here, a ferret dam returns to her burrow and kits with a freshly killed black-tailed prairie dog C. ludovicianus. Larimer County, Colorado, 2017. Photo credit: T.N. Tretten, U. S. Fish and Wildlife Service, National Black-footed Ferret Conservation Center.

Figure 1.

Prairie dogs Cynomys spp. are the primary food source of the black-footed ferret Mustela nigripes. Ferrets are therefore affected by diseases that affect prairie dog populations, such as flea-borne sylvatic plague, as well as treatments applied to prairie dogs to manage those diseases. Fipronil-laced baits can be used to control flea populations on prairie dogs and residues from fipronil consequently pass into ferrets when a treated prairie dog is consumed. Those treatments can then possibly affect ferrets and further assist with flea control and disease management. Here, a ferret dam returns to her burrow and kits with a freshly killed black-tailed prairie dog C. ludovicianus. Larimer County, Colorado, 2017. Photo credit: T.N. Tretten, U. S. Fish and Wildlife Service, National Black-footed Ferret Conservation Center.

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Continued study is underway to better understand effects of fipronil bait treatments, be they positive or negative, on wildlife and the ecosystems they inhabit (Eads et al. 2021, 2022a; Matchett et al. 2023). Here, we concentrate on trophic transfer of fipronil residues from treated prairie dogs to ferrets and potential implications for the control of larval fleas. We fed carcass portions from prairie dogs that had consumed fipronil grain to captive ferrets and monitored consumption and their overall health. Additionally, we collected scat from ferrets to investigate transfer of fipronil residues from prey to predator, and we tested the effects of this form of secondary fipronil treatment on larval fleas exposed to ferret scat containing fipronil and fipronil sulfone. Like prairie dogs, ferrets primarily deposit scat in burrows where flea larvae forage and develop (Sheets et al. 1972). We hypothesized that fipronil residues in ferret scat would assist in controlling larval fleas, as found previously with scat from black-tailed prairie dogs treated with fipronil grain (Eads et al. 2023a).

To acquire food portions for ferrets, we fed four adult black-tailed prairie dogs fipronil-treated grain (0.005% fipronil by weight; Scimetrics Ltd. Corp., Wellington, Colorado, United States; EPA Reg. Num.: 72500-28) for 5 consecutive days under animal use and care guidelines of the American Society of Mammalogists (Sikes et al. 2016). Please see Wang et al. (2019) and Eads et al. (2022b) for details on black-tailed prairie dog care and feeding. After 5 days, the prairie dogs were humanely euthanized and frozen at -18°C. We chose three of the four prairie dogs for subsequent feeding to ferrets. These three prairie dogs had consumed the largest amounts of fipronil-treated grain: 38, 54, and 63 g (mean 48 g) or 0.00175–0.00315 g (mean 0.002 g) of fipronil (Wang et al. 2019). In addition, their combined carcass weight of approximately 2,000 g equaled the amount that four adult ferrets would normally consume over a 7-d period.

We trialed four adult captive-raised ferrets, two 4-year-old males and two 3-year-old females, at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center (NBFFCC), Carr, Colorado, United States. We cared for ferrets under the conditions described in the 2016 Intra-Service Section 7 Consultation on Region 6’s Section 10(a)(1)(A) Permitting Program and Fish and Wildlife Service Initiated Recovery Actions (USFWS 2016b). We housed ferrets individually in standard ex situ cages (USFWS 2017) at ∼18°C and provided them drinking water ad libitum.

From 16 through 22 July 2017, we fed the fipronil-exposed prairie dog carcasses to the four trial ferrets. Prairie dogs were portioned into appropriate sizes for a ferret meal, 60 g/female and 80 g/male. Portions of each carcass were stored in resealable plastic freezer bags and kept separate from portions of other carcasses. Once daily, during feeding time, biological science technicians arbitrarily selected a carcass portion of the appropriate size out of a freezer bag and placed it in the cage with each ferret. In accordance with standard protocols for all animals housed at NBFFCC, we visually observed ferrets at least once daily for signs of distress or illness such as lethargy, lack of appetite, high water consumption, vomit, diarrhea, and any obvious physical or behavioral changes. We monitored the ferrets in this method for the duration of their stay at NBFFCC, which ended 20 November 2017.

To investigate trophic transfer of efficacious fipronil residues, we collected multiple droppings of scat per ferret per day before we offered fipronil-treated prairie dog carcasses, and again on days 1, 3, 5, and 7 of feeding when cages were cleaned and all remaining scat and soiled bedding were removed. We stored scat from individual ferrets in labeled write-on sealable bags and froze them at −18°C.

Using the stored samples in preparation for the larval flea exposure study, we selected one pre-exposure scat sample from each of two ferrets to serve as controls. We then, by placing all scats from a single ferret from a single day into a bag and shaking the bag, randomly selected one scat per feeding days 1, 5, and 7 from each of the four ferrets. We used these 14 samples to analyze for pre-feeding and post-feeding fipronil residues. Scat samples from day 3 of feeding were saved for future study. We hypothesized that if samples from days 1 and 5 had lethal effects on flea larvae then day 3 should as well. Fipronil and sulfone metabolite concentrations in the 14 ferret scats were quantified by liquid chromatography–tandem mass spectrometry at the Analytical Toxicology Laboratory, Colorado State University, Fort Collins, Colorado, United States. We composited the remaining ferret scats by combining scats taken from the same ferret on the same day into a single sample. We then ground these samples into fine particles for flea larvae to eat (Bland et al. 2017) using a disposable polypropylene pestle. We separated each sample as 0.5 mg subsamples into prelabeled centrifuge tubes. We stored these subsamples frozen at −18°C.

We used the flea species Oropsylla montana (Siphonaptera: Ceratophyllidae) as a proxy for Oropsylla hirsuta, which parasitizes prairie dogs and ferrets under natural conditions (Harris et al. 2014; Eads et al. 2018). We found no evidence in the literature to suggest the efficacy of fipronil would vary significantly between these species. The O. montana were colonized and cared for as described in Eisen et al. (2007). We used flea larvae that appeared to be first or second instars with empty alimentary canals.

We exposed 48 larval fleas to ferret scat with fipronil residues and 48 larvae to ferret scat lacking fipronil residues. We held and observed flea larvae in Corning® 12-well microplates (Eads et al. 2023a). We made small holes in the microplate caps, immediately above each well, to allow for air exchange. We filled each well (∼1/4 to 1/2 of well depth) with sterilized fine sand substrate for larvae locomotion and refuge. Using a random number generator, we supplied four of the plates with randomly selected scat that was collected from ferrets that were fed fipronil-treated prairie dogs (exposed) and four with scat collected before ferrets were exposed to fipronil (control). Each well received 0.5 mg of scat. Separation of the wells among plates should have eliminated any potential cross-contamination. One larval flea was randomly selected for placement in each well. All plates were tested under the same environmental conditions, with at least one treated and one nontreated plate tested on the same days and no more than two treated or two nontreated on the same day. Oropsylla montana larvae have limited or no exposure to light during their development in rodent burrows (Bland et al. 2017). Thus, in our study, we lidded the microplates, loosely covered them with aluminum foil, and stored them in a dark location for 24 h at 23°C and 85% relative humidity.

After 24 h, we uncovered and opened the microplates. We used probes to prod each larva for 2 s. Live larva responded by coiling and moving away from the prod. Larvae that did not respond within 2 s were prodded for an additional 5 s and we considered them dead if no movement was observed (Panella et al. 2005). We used a 60 to 120 x pocket microscope to determine if each larva consumed ferret scat (yes = visible meal, colored like ferret scat, in the gut; no = no visible meal in the gut; Figure 2). Fleas that had consumed scat were categorized separately from fleas that had no evidence of consuming scat. At the end of each trial, we placed larvae in ethanol and transferred them to a biohazard waste container.

Figure 2.

A meal visible in the alimentary canal (gut) of an Oropsylla montana flea larva. Larval fleas live within nesting refugia of burrowing mammals, such as prairie dogs Cynomys spp., and often consume scat material of host species. Residues from fipronil-laced food items consumed by mammals can pass into their scat and affect larval flea survival. Photo credit: B.Joseph Hinnebusch, Laboratory of Bacteriology, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institute of Health, Hamilton, Montana, 2022.

Figure 2.

A meal visible in the alimentary canal (gut) of an Oropsylla montana flea larva. Larval fleas live within nesting refugia of burrowing mammals, such as prairie dogs Cynomys spp., and often consume scat material of host species. Residues from fipronil-laced food items consumed by mammals can pass into their scat and affect larval flea survival. Photo credit: B.Joseph Hinnebusch, Laboratory of Bacteriology, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institute of Health, Hamilton, Montana, 2022.

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We used logistic regression to examine the effect of fipronil residues in ferret scat on 24-h survival of flea larvae (‘glm’ function in ‘stats’ package, R x64 version 4.1.2, R Core Team 2021). We concentrated on an interaction between categorical predictor variables for treatment and evidence of scat consumption (α = 0.050). Consumption of scat with fipronil residues was expected to be more lethal than consumption of scat lacking fipronil residues (as found with larval fleas feeding on black-tailed prairie dog scat; Eads et al. 2023a). We selected a parsimonious model via backward elimination (type III χ2 likelihood-ratio tests, ‘Anova’ function in ‘car’ package; Fox et al. 2023) and assessed model goodness-of-fit using a Hosmer and Lemeshow test (‘hoslem.test’ function in ‘ResouceSelection’ package; Lele et al. 2023).

During each day of the treatment period, all four ferrets consumed their provided portion of fipronil-treated prairie dog and displayed no aversion to the taste despite the presumed presence of fipronil residues. No signs of illness, distress, or behavioral changes were observed in any of the ferrets during the 7-d feeding trial in July 2017. Moreover, no effects were observed for the duration of the ferrets’ stay at NBFFCC, into November 2017, approximately four months after the feeding trial.

Neither fipronil nor fipronil sulfone were detected in ferret scat collected before they were provided with fipronil-treated prey. In contrast, fipronil and fipronil sulfone were detected in all scat samples collected post-consumption. Fipronil residues (0.70 to 26.22 ng/g ferret scat) declined during the 7-d feeding trial, whereas fipronil sulfone residues (2.07 to 34.29 ng/g ferret scat) tended to increase (Table 1). We found that ferret scat containing fipronil residues killed larval fleas at a higher rate than ferret scat without fipronil residues. When exposed to ferret scat lacking fipronil residues, 85% of larvae survived. Survival was reduced to 61% and 35% for larvae contacting or consuming fipronil-residue scat, respectively (Table 2).

Table 1.

Fipronil and fipronil sulfone concentrations (ng/g) in individual scats collected from four adult black-footed ferrets Mustela nigripes (ferrets) on days 1, 5, and 7 of feeding. Food consisted of black-tailed prairie dogs Cynomys ludovicianus fed 0.005% fipronil grain for five days. Scat samples were collected from ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center in July 2017. Two ferret scat samples, collected before treatment, were used for calibration (with “ND” indicating “none detected” for fipronil or fipronil sulfone in those samples and “n.a.” indicating “not applicable” for collections that occurred before treatment).

Fipronil and fipronil sulfone concentrations (ng/g) in individual scats collected from four adult black-footed ferrets Mustela nigripes (ferrets) on days 1, 5, and 7 of feeding. Food consisted of black-tailed prairie dogs Cynomys ludovicianus fed 0.005% fipronil grain for five days. Scat samples were collected from ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center in July 2017. Two ferret scat samples, collected before treatment, were used for calibration (with “ND” indicating “none detected” for fipronil or fipronil sulfone in those samples and “n.a.” indicating “not applicable” for collections that occurred before treatment).
Fipronil and fipronil sulfone concentrations (ng/g) in individual scats collected from four adult black-footed ferrets Mustela nigripes (ferrets) on days 1, 5, and 7 of feeding. Food consisted of black-tailed prairie dogs Cynomys ludovicianus fed 0.005% fipronil grain for five days. Scat samples were collected from ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center in July 2017. Two ferret scat samples, collected before treatment, were used for calibration (with “ND” indicating “none detected” for fipronil or fipronil sulfone in those samples and “n.a.” indicating “not applicable” for collections that occurred before treatment).
Table 2.

Larval flea Oropsylla montana survival after 24 h of being exposed to black-footed ferret Mustela nigripes (ferret) scat samples. Scat samples were collected from ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center in July 2017 before and after ferrets had been fed carcass of black-tailed prairie dogs Cynomys ludovicianus that had been fed 0.005% fipronil grain for 5 days.

Larval flea Oropsylla montana survival after 24 h of being exposed to black-footed ferret Mustela nigripes (ferret) scat samples. Scat samples were collected from ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center in July 2017 before and after ferrets had been fed carcass of black-tailed prairie dogs Cynomys ludovicianus that had been fed 0.005% fipronil grain for 5 days.
Larval flea Oropsylla montana survival after 24 h of being exposed to black-footed ferret Mustela nigripes (ferret) scat samples. Scat samples were collected from ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center in July 2017 before and after ferrets had been fed carcass of black-tailed prairie dogs Cynomys ludovicianus that had been fed 0.005% fipronil grain for 5 days.

After 24 h of exposure to ferret scat, most flea larvae were found submerged in substrate adjacent to the edges of wells. No larva escaped from the well-plates and pupation was not observed. Overall, 33% (n = 32 of 96) of larvae exhibited evidence of consuming ferret scat. Fipronil treatment had no detectable effect on the proportion of larvae with meals, of any size, in their guts (χ2 test P = 0.665). For those larvae with evidence of scat consumption, qualitative observations suggested that <25% of the gut was filled for most larvae that had consumed ferret scat with fipronil residues, whereas >50% of the gut was commonly filled for larvae that had consumed ferret scat lacking fipronil residues.

In the logistic regression analysis of larvae survival, the two-way statistical interaction between treatment and evidence of scat consumption was unsupported and therefore eliminated (P = 0.256), which reduced the model to the individual variables for treatment and evidence of scat consumption. We failed to detect any difference in survival for larvae with or without evidence of ferret scat consumption, so that variable was eliminated (P = 0.256). In the current study, the treatment variable was supported; survival was 61% lower (P < 0.001) for larvae exposed to ferret scat with fipronil residues (25/48 survived) vs. larvae exposed to ferret scat lacking fipronil residues (41/48 survived; Table 2; Figure 3). We failed to detect disparity between laboratory results and model predictions; there was good statistical correspondence between the observed laboratory data and model predictions (P = ∼1).

Figure 3.

Survival probabilities (95% confidence intervals) for larval fleas Oropsylla montana exposed to 0.50 mg of black-footed ferret Mustela nigripes scat. From 16 through 22 July 2017, ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center were offered carcass portions of black-tailed prairie dogs Cynomys ludovicianus that had consumed 0.005% fipronil grain. Survival was assessed 24 h after the larval fleas were exposed to scat in well plates and estimated using logistic regression. Sample sizes are denoted by treatment. The P-value (< 0.001) is for the treatment variable in a logistic regression model.

Figure 3.

Survival probabilities (95% confidence intervals) for larval fleas Oropsylla montana exposed to 0.50 mg of black-footed ferret Mustela nigripes scat. From 16 through 22 July 2017, ferrets at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center were offered carcass portions of black-tailed prairie dogs Cynomys ludovicianus that had consumed 0.005% fipronil grain. Survival was assessed 24 h after the larval fleas were exposed to scat in well plates and estimated using logistic regression. Sample sizes are denoted by treatment. The P-value (< 0.001) is for the treatment variable in a logistic regression model.

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Previously, we detected lethal effects of fipronil residues in black-tailed prairie dog scat on larval O. montana (Eads et al. 2023a). In that study, approximately 59% of larvae (196 of 331) consumed prairie dog scat vs. 33% of larvae (32 of 96) consuming ferret scat herein. O. montana, an important vector of Y. pestis, parasitizes a variety of ground squirrels (Eisen et al. 2009). Perhaps scat from rodents such as ground squirrels and prairie dogs is more attractive to larval O. montana than scat from carnivores due to the various chemical and physical differences that exist between samples. Factors such as taste or processing and excretion costs might be influential. It is possible that our samples included small organic paper fibers from the ferret bedding material used at NBFFCC. These fibers could have influenced the food preferences of flea larvae, either encouraging or discouraging their consumption of ferret scat. However, we feel this was largely avoided by grinding the samples before they were offered to fleas and proportions of these fibers should be comparable between ferret scat with or without fipronil residues.

Eads et al. (2023a) exposed larval fleas to prairie dog scat that was estimated to contain 296.20 ng/g fipronil and 158.19 ng/g fipronil sulfone, on average. In the present study, we exposed larval fleas to ferret scat that contained 3.76 ng/g fipronil and 13.75 ng/g fipronil sulfone, on average. In addition, sample sizes were smaller and, thus, statistical power was weaker than we reported in Eads et al. (2023a), in which we did detect significantly lower survival of larvae with evidence of prairie dog scat consumption compared to those without. Both compounds can be lethal to insects (Hainzl et al. 1998), but perhaps unsurprisingly, given these differences in fipronil residue concentrations, prairie dog scat killed more larval fleas (79%) than ferret scat (48%). Even so, the ferret scat was often lethal to larval fleas; and even trace amounts of some insecticidal compounds can have measurable effects on flea larvae contacting or consuming them (Marchiondo et al. 2013). It is also possible that with more than 24 h of exposure to fipronil-laced ferret scats, larval fleas would experience higher mortality.

Scat from prairie dogs is likely to be abundant in many prairie dog burrows (Hoogland 1995), perhaps especially the burrows they use as nesting chambers (Wilcomb 1954). In contrast, ferret scat may be relatively scarce in most burrows within a prairie dog colony given their uncommon status and the influences of intrasexual territoriality on habitat carrying capacity (Biggins et al. 2006; Livieri and Anderson 2012; Eads et al. 2014). That said, fipronil residues in ferret scat might assist in flea control in burrows most commonly used by ferrets for nesting sites and refuges, with potential benefits for plague mitigation, and perhaps ferret body condition and reproduction (Eads et al. 2022b). We also presume that fipronil and its metabolites in ferret blood could affect localized populations of adult fleas, as indicated by the results in Eads et al. (2023b), but we did not investigate this possibility. Even though this relatively small, localized level of flea control on a landscape may seem trivial, the possibility remains for benefits to ferrets. Ferrets are extraordinarily susceptible to plague, often succumbing to the equivalent of a single bite from a plague-positive flea under laboratory conditions (E. Williams pers comm 1999 inGodbey et al. 2006). By reducing flea numbers in burrows commonly used by ferrets, which presumably are not commonly used by prairie dogs during ferret occupancy, it is possible that even this localized effect on flea populations could have positive effects on ferret survival.

The concept of multiple individuals of many different species consuming fipronil and excreting fipronil and fipronil sulfone in their scat may raise concerns that fipronil residues will bioaccumulate in prairie dog colonies, and because prairie dogs occur at higher densities than other rodents in the colony sites, prairie dog scat can be the primary contributing factor. Prairie dog burrows are dark (Wilcomb 1954) and may function as “light shields” for fipronil residues (Gunasekara et al. 2007), perhaps allowing them to persist for 200 d or more underground (Gunasekara et al. 2007; Simon-Delso et al. 2015). Such persistence could facilitate long-term control of flea larvae; potential effects on non-target arthropods are under investigation.

Regarding scat and potential bioaccumulation aboveground, fipronil degrades by a variety of pathways in environment, including oxidation, hydrolysis, biodegradation, and photodegradation (Fent 2014). Photodegradation is the only pathway that produces fipronil disulfide, which is more persistent in the environment than the parent product (Singh et al. 2021). Encouragingly, some studies suggest fipronil residues may fully degrade under ultraviolet light (e.g., Mianjy and Niknafs 2013). However, potential effects on other predators, such as long-lived raptors, as well as other prairie dog colony cohabitants, warrant further study (Eads et al. 2023a).

To date, our research suggests fipronil baits are effective tools for controlling adult and larval fleas in the prairie dog-ferret system, with conservation implications for ferrets and other species (Eads et al. 2019, 2020, 2021, 2022a, b, 2023; Matchett et al. 2023; see also Poché et al. 2017, 2020). In this study, we treated black-tailed prairie dogs with fipronil grain at 0.005% fipronil by weight; similar outcomes are expected with fipronil “FipBit” pellets (∼0.84 mg fipronil/pellet; Eads et al. 2023a). According to the World Association for the Advancement of Veterinary Parasitology (Marchiondo et al. 2013), fipronil bait treatments kill fleas already on animals at time of treatment when they blood-feed, the treatments kill fleas newly acquired by hosts for a period of time after treatment, and the treatments progressively reduce or eliminate off-host flea stages in the environment. In these ways, fipronil baits allow for rapid onset, immediate flea kill, and long-term persistent and curative efficacy for plague mitigation and wildlife conservation. We did not investigate unobtrusive health consequences of fipronil on ferrets in this study. We suggest further investigations into the effect of fipronil on ferrets; behavioral studies of easy-to-observe ferrets in captivity and survival and reproductive success rates in the wild all seem useful.

Please note: The Journal of Fish and Wildlife Management is not responsible for the content or functionality of any supplemental material. Queries should be directed to the corresponding author for the article.

Data S1. Scat samples were collected from black-footed ferrets Mustela nigripes at the U.S. Fish and Wildlife National Black-footed Ferret Conservation Center in July 2017 before and after they were fed carcass portions of black-tailed prairie dogs Cynomys ludovicianus that had been fed 0.005% fipronil grain for 5 days. Scat samples from ferrets were stored frozen and then placed in well plates with larval Oropsylla montana fleas at the Centers for Disease Control and Prevention, Fort Collins, Colorado in 2018. Scat consumption and flea survival was documented after 24 h. Eads, D.A., T.N. Tretten. 2024. Data on flea larvae survival following exposure to black-footed ferret scat with or without fipronil residues. U.S. Geological Survey data release, https://doi.org/10.5066/P90ZLDH6.

Available: https://doi.org/10.3996/JFWM-23-069.S1 (86 KB PDF)

Reference S1. U.S. Fish and Wildlife Service, Black-footed Ferret Recovery Program. June 15, 2016a. Black-footed Ferret Field Operations Manual. 89pp. https://www.blackfootedferret.org/uploads/1/2/7/7/127791157/usfws_2016_bff-field-operations-manual_oct2016-.pdf.

Available: https://doi.org/10.3996/JFWM-23-069.S2 (10,639 KB PDF)

Reference S2. U.S. Fish and Wildlife Service, Black-footed Ferret Recovery Program. January 16, 2017. Black-footed Ferret Managed Care Operations Manual. 228 pp. https://www.fws.gov/sites/default/files/documents/black-footed-ferret-managed-care-operations-manual-2017.pdf.

Available: https://doi.org/10.3996/JFWM-23-069.S3 (5,929 KB PDF)

Reference S3. U.S. Fish and Wildlife Service, Intra-Service Programmatic Section 7 Consultation on Region 6’s Section 10(a)(1)(A) Permitting Program and Fish and Wildlife Service Initiated Recovery Actions. 2016b. 160 pp.

Available: https://doi.org/10.3996/JFWM-23-069.S4 (12,979 KB PDF)

Financial and logistical support were provided by U.S. Geological Survey, Centers for Disease Control and Prevention, U.S. Fish and Wildlife Service, and Colorado State University. Additional logistical support was provided by Scimetrics Limited Corporation and Genesis Laboratories, Inc. We thank J. Montenieri, R. Enscore and K. Gage, who provided guidance on flea larvae sampling; R. Poché, who kindly provided fipronil treated grain; the biological science technicians at NBFFCC who completed the daily tasks of feeding and caring for the ferrets; and Smith Environmental and Engineering, who humanely donated the prairie dogs. We thank our many colleagues for discussions on fleas and plague, particularly M. Antolin, E. Childers, P. Dobesh, T. Livieri, P. Roghair, M. Schwarz, and especially M.R. Matchett for his reviewing of this manuscript and providing valuable input. Finally, we thank the three anonymous reviewers and the Associate Editor that all provided instrumental input that greatly improved earlier versions of this manuscript. Any use of trade, product, or firm names is for descriptive purposes only and does not imply endorsement by the U.S. Government.

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Author notes

The findings and conclusions in this article are those of the author(s) and do not necessarily represent the views of the U.S. Fish and Wildlife Service.