Interdisciplinary treatment for an adolescent with fusion of the left maxillary lateral and central incisors: A clinical report.

The interdisciplinary treatment provided for a 12-year-old boy with fusion of his left central and lateral incisors and concomitant distal space loss is described. His treatment was initiated with orthodontic therapy followed by endodontic treatment of the fused tooth. A conservative zirconia and porcelain cantilevered bonded fixed dental prosthesis was then completed to replace his left lateral incisor.

Temporary Anosmia in Mice Following Nasal Lavage With Dilute Detergent Solution.

Olfactory sensory deprivation induces anosmia and reduces tyrosine hydroxylase and dopamine levels in the olfactory bulb. The behavioral consequences specific to the loss of olfactory bulb dopamine are difficult to determine because sensory deprivation protocols are either confounded by side effects or leave the animal anosmic. A new method to both induce sensory deprivation and to measure the behavioral and circuit consequences is needed. We developed a novel, recoverable anosmia protocol using nasal lavage with a dilute detergent solution. Detergent treatment did not damage the olfactory epithelium as measured by scanning electron microscopy, alcian blue histology, and acetylated tubulin immunohistochemistry. One treatment-induced anosmia that lasted 24 to 48 h. Three treatments over 5 days reduced olfactory bulb tyrosine hydroxylase and dopamine levels indicating that anosmia persists between treatments. Importantly, even with multiple treatments, olfactory ability recovered within 48 h. This is the first report of a sensory deprivation protocol that induces recoverable anosmia and can be paired with biochemical, histological, and behavioral investigations of olfaction.

Upon the Shoulders of Giants: Open-Source Hardware and Software in Analytical Chemistry.

Isaac Newton famously observed that "if I have seen further it is by standing on the shoulders of giants." We propose that this sentiment is a powerful motivation for the "open-source" movement in scientific research, in which creators provide everything needed to replicate a given project online, as well as providing explicit permission for users to use, improve, and share it with others. Here, we write to introduce analytical chemists who are new to the open-source movement to best practices and concepts in this area and to survey the state of open-source research in analytical chemistry. We conclude by considering two examples of open-source projects from our own research group, with the hope that a description of the process, motivations, and results will provide a convincing argument about the benefits that this movement brings to both creators and users.

Integrating single-cell and spatially resolved transcriptomic strategies to survey the astrocyte response to stroke in male mice.

Astrocytes, a type of glial cell in the central nervous system (CNS), adopt diverse states in response to injury that are influenced by their location relative to the insult. Here, we describe a platform for spatially resolved, single-cell transcriptomics and proteomics, called tDISCO (tissue-digital microfluidic isolation of single cells for -Omics). We use tDISCO alongside two high-throughput platforms for spatial (Visium) and single-cell transcriptomics (10X Chromium) to examine the heterogeneity of the astrocyte response to a cortical ischemic stroke in male mice. We show that integration of Visium and 10X Chromium datasets infers two astrocyte populations, proximal or distal to the injury site, while tDISCO determines the spatial boundaries and molecular profiles that define these populations. We find that proximal astrocytes show differences in lipid shuttling, with enriched expression of Apoe and Fabp5. Our datasets provide a resource for understanding the roles of astrocytes in stroke and showcase the utility of tDISCO for hypothesis-driven, spatially resolved single-cell experiments.

Integrated digital microfluidic platform for voltammetric analysis.

Digital microfluidics (DMF) is an emerging technique for manipulating small volumes of liquids. DMF is particularly well suited for analytical applications as it allows automated handling of discrete samples, and it has been integrated with several inline analysis techniques. However, examples of the integration of DMF with electroanalytical methods are notably scarce, and those that have been reported rely on external electrodes that impose limitations on complexity. To combine the full capabilities of DMF with voltammetry, we designed a platform featuring a three-electrode electrochemical cell integrated in a microfabricated DMF device, removing the need for external electrodes and allowing for complete droplet control. The performance of the DMF/voltammetry system is comparable to that of a commercial screen printed electrode, and the new platform was applied to generating a calibration series for acetaminophen with a limit of detection of 76 µM and good precision (4% average RSD), all with minimal human intervention. We propose that this platform and variations thereof may be a useful new tool for microscale electroanalysis and will be a complementary system to existing inline analysis techniques for DMF.

Comparison of the initial and residual speed of Ixodes scapularis kill on dogs treated with a single dose of Bravecto® Chew (25 mg/kg fluralaner) or Simparica TRIO® (1.2 mg/kg sarolaner, 24 µg/kg moxidectin, 5 mg/kg pyrantel).

BACKGROUND: Compliant ectoparasiticide product use is a comprehensive way to control ticks and reduce the risk of tick-borne pathogen transmission to dogs. Because the systemically acting isoxazoline ectoparasiticides require tick attachment for drug delivery, fast speed of kill is essential to minimize tick-borne pathogen transmission risk. METHODS: Dogs of satisfactory tick-carrying capacity were randomly allocated to treatment groups and administered, per label instructions, Bravecto® Chews (minimum 25 mg/kg fluralaner), Simparica TRIO® (minimum 1.2 mg/kg sarolaner, 24 µg/kg moxidectin, 5 mg/kg pyrantel), or no treatment. Dogs were infested with approximately 50 unfed adult (35 female, 15 male) Ixodes scapularis on Day -2, 21 and 28. Live tick counts were performed at 4, 8, 12 and 24 h post-treatment (Day 0) and post-infestation on Day 21 and 28. Tick control efficacy was determined by comparing live tick means for each product-treated group to the untreated control group and each other at all time points using a linear mixed model. The percent of dogs free of live ticks was analyzed using the Fisher's exact test for treatment group comparison. RESULTS: The untreated control group maintained adequate tick infestations throughout the study. Using geometric means, an existing I. scapularis infestation was controlled by 99.7% and 93.0% 12 h post-treatment and by 100% and 99.5% 24 h post-treatment, for Bravecto® and Simparica TRIO®-treated dogs, respectively. Ixodes scapularis infestations were controlled more quickly for Bravecto®- compared to Simparica TRIO®-treated dogs on Day 21 at 8 h (efficacy 74.0% vs. 0.0%, p = 0.003) and 12 h (efficacy 99.2% vs. 39.4%, p < 0.001) post-infestation and Day 28 at 8 h (efficacy 92.2% vs. 0.0%, p < 0.001) and 12 h (efficacy 99.6% vs. 27.7%, p < 0.001) post-infestation. On Day 28 post-treatment, the efficacy of Bravecto® and Simparica TRIO® to control a new I. scapularis infestation was 100% and 96.6%, respectively, by 24 h post-infestation. Of product-treated dogs, 100% of Bravecto®-treated dogs were free of live ticks by 24 h post-treatment or post-infestation. No treatment-related adverse reactions occurred during the study. CONCLUSIONS: Ixodes scapularis infestations are controlled more quickly 21 and 28 days post-treatment for dogs administered a single dose of Bravecto® compared to dogs administered a single dose of Simparica TRIO®.

Structural properties of metal-free apometallothioneins.

The metalated forms of metallothionein are well studied (particularly Zn-MT, Cu-MT and Cd-MT), but almost nothing is known about the chemical and structural properties of apometallothioneins despite their importance in initial metalation and subsequent demetalation. Electrospray ionization mass spectrometry was used to provide a detailed view of the structural properties of the metal-free protein. Mass spectra of Zn(7)-MT and apo-MT at pH 7 exhibit the same charge state distribution, indicating that apo-MT is tightly folded like the metallated protein, whereas apo-MT at pH 3 exhibits a charge state spectrum associated with unfolding or denaturation. Benzoquinone was used to modify the cysteines in the ß-MT (9 Bq), and α-MT (11 Bq) fragments, and the full ßα-MT (20 Bq) protein. ESI-MS showed that the overall volume and, therefore, the extent of folding for the modified proteins is similar to that of Zn-MT. Molecular modeling using MM3-MD methods provided the volume of each modified protein. The volumes of the partially modified proteins follow the same trend as the charge states, showing that ESI-MS is an excellent method with which to follow small changes in protein folding as a function of applied chemical stress. The data suggest that the structure of apo-ßα-MT is more organized than previously considered.

Ophthalmomyiasis externa in a puppy due to Cuterebra infestation.

An 8 wk old Scottish terrier was evaluated for a 2×2 cm mass located directly adjacent to the lateral canthus of the right eye. Examination revealed a well-developed Cuterebra larva at the lateral aspect of the orbital cavity, anchored at the lateral canthus. The larva was manually removed under general anesthesia, and the eyelid margin was repaired. Systemic and topical antibiotics along with systemic anti-inflammatory treatment were instituted. The lesion healed completely, revealing a normal functional right eye 10 days after removal of the Cuterebra larva. Careful manual removal of the larva is recommended along with correction of any resultant conformational defects. The remaining cavity can be left to heal; however, antibiotic therapy is recommended.

Canine schistosomiasis in Kansas: five cases (2000-2009).

This is a retrospective case series consisting of five dogs diagnosed with schistosomiasis. The purpose of this article is to report the presence of naturally occurring canine schistosomiasis in Kansas and to provide clinical details regarding schistosomiasis. Medical records of dogs diagnosed with schistosomiasis from 2000 to 2009 were reviewed, and information extracted included signalment, history, clinical signs, diagnostic test results, treatment, and outcome. Affected dogs were primarily medium to large breed and young to middle aged. All dogs were considered outdoor dogs, with three having known access to surface water. Common clinical signs included gastrointestinal disease and signs associated with hypercalcemia. Fecal flotation was negative in all dogs in contrast to fecal saline sedimentation and fecal polymerase chain reaction, which were both positive in all dogs in which it was performed. All dogs treated specifically for schistosomiasis fully recovered. This article describes the first reported cases of canine schistosomiasis in the Midwest and the first reported case of intestinal intussusception secondary to schistosomiasis. Recognizing that canine schistosomiasis is present in Kansas and possibly other Midwestern states should prompt veterinarians to perform appropriate diagnostic investigation in suspect animals as the diagnosis is straight forward and relatively inexpensive.

Arsenic transfer between metallothionein proteins at physiological pH.

As³+ bound to the two-domain, recombinant human metallothionein (isoform 1a) is stable at pH 7 and translocates via protein-protein interactions to other metallothionein proteins. The data show As³+ transfer from the two-domain ß-α-hMT to binding sites in the isolated apo-ß-hMT and apo-α-hMT. Under conditions of equilibrium, apo- and partially-metallated species coexist indicating that noncooperative demetallation of the As(6)-ßα-hMT occurrs. As³+ transfer under conditions (pH 7) where the free As³+ ion is not stable, provides evidence that Cd²+ and Zn²+ transfer may also take place through protein-protein interactions and that partially metallated Cd-MT and Zn-MT would be stable.

Digital microfluidic isolation of single cells for -Omics.

We introduce Digital microfluidic Isolation of Single Cells for -Omics (DISCO), a platform that allows users to select particular cells of interest from a limited initial sample size and connects single-cell sequencing data to their immunofluorescence-based phenotypes. Specifically, DISCO combines digital microfluidics, laser cell lysis, and artificial intelligence-driven image processing to collect the contents of single cells from heterogeneous populations, followed by analysis of single-cell genomes and transcriptomes by next-generation sequencing, and proteomes by nanoflow liquid chromatography and tandem mass spectrometry. The results described herein confirm the utility of DISCO for sequencing at levels that are equivalent to or enhanced relative to the state of the art, capable of identifying features at the level of single nucleotide variations. The unique levels of selectivity, context, and accountability of DISCO suggest potential utility for deep analysis of any rare cell population with contextual dependencies.

Evaluation of a topical sarolaner-selamectin combination to control flea populations on naturally infested cats in private residences in West Central Florida.

Historic data show that home flea infestations can be managed by treating all animals on the premises with a highly effective flea control product. The use of effective products has also been shown to reduce pruritus and minimize dermatologic lesions in both cats and dogs. Therefore, an in-home study was conducted in West Central Florida USA to evaluate the efficacy of a topically applied selamectin-sarolaner formulation to control fleas in naturally infested cats over a 12-week period. Thirty-seven cats in 21 households were treated once monthly with the selamectin-sarolaner topical solution. In the topical fluralaner treatment (positive control) group, forty-three cats in 20 households were treated once on day 0. A combined total of thirty dogs in both groups were treated once monthly with oral sarolaner. Fleas on cats were counted by flea combing, fleas on dogs were counted using visual area counts and fleas in the indoor premises were assessed using intermittent-light flea traps. Blinded-assessments of feline dermatologic lesions (modified-SCORFAD) were conducted monthly by a boarded-dermatologist and pruritus severity was evaluated by pet owners. Three consecutive monthly treatments of selamectin-sarolaner reduced flea populations on cats by 96.3 % within 7 days and by 100% from week 6 to the end of the 12-week study. The topical application of fluralaner reduced flea populations by 98.1 % within 7 days and efficacy reached 100% by week 12. At the end of the study, fleas were completely eradicated (from cats, dogs and homes) in every home regardless of treatment group. Owner reported cat pruritus was reduced by > 87 % in both treatment groups by week 12. Significant improvements in dermatologic lesion scores (> 81 %) were achieved by both products by the end of the study. Monthly applications of topical selamectin-sarolaner or topical fluralaner to cats living in the heavy flea challenge environment of West Central Florida USA were effective in eradicating flea infestations, reducing pruritus and improving dermatologic lesions.

Flea and tick control in the 21st century: challenges and opportunities.

Historically, veterinarians have told their clients that one flea is all that is necessary to produce and maintain the clinical signs of flea allergy dermatitis (FAD). Newer adulticides, such as fipronil, imidacloprid, nitenpyram and selamectin, have had a positive clinical effect on dogs and cats with FAD. However, data on flea feeding and the effect of these products on flea feeding bring into question the once perceived dogma of the single flea bite concept. Current data would indicate that the primary role of these insecticides in managing FAD is in rapidly reducing flea numbers and reducing flea feeding rather than preventing flea bites. Controlling tick infestations is important not only because ticks are nuisance parasites of dogs and cats, but also because they are vectors of a variety of bacterial and protozoal diseases. Achieving satisfactory tick control is often difficult due to unrealistic expectations of pet owners, to residual acaricidal properties of products that are often less than 100% and because of constant re-infestation pressure. Some of the most important factors veterinarians must be aware of are regional changes in tick distributions, our inability to control wildlife tick hosts and expectation differences between flea and tick control. These factors probably cause most real and perceived product failures.

In-home assessment of flea control and dermatologic lesions in dogs provided by lotilaner (Credelio®) and spinosad (Comfortis®) in west central Florida.

Post-launch field investigations of recently-approved flea control products establish an efficacy baseline and in subsequent years can detect any efficacy decline suggestive of emerging resistance. As part of a continuing program of yearly assessment of flea control products in west central Florida, this study, using client-owned dogs, investigated the efficacy of lotilaner and spinosad in controlling fleas and in alleviating dermatologic signs likely associated with flea infestations. Forty-four qualifying households were randomized to either a lotilaner (Credelio®) (minimum dose rate 20 mg/kg) or a spinosad (Comfortis®) (30 mg/kg) group, with 33 and 36 dogs in each group, respectively. On Days 0 and 28 (±2) all dogs in each household were treated with the allocated product according to label directions, and all household cats received spinetoram (Cheristin®). On Day 0 and at weekly intervals through Day 56 (±2), on-animal and premises flea burdens were enumerated, a veterinary dermatologist scored integumental changes using canine atopic dermatitis extent and severity index (CADESI)-4 and flea allergy dermatitis (FAD) scales, and owners scored pruritus using the validated canine pruritus severity scale (CPSS). At study entry geometric mean flea counts were 33.2 and 29.9 in the lotilaner and spinosad groups, respectively. For both groups, reductions in flea counts were > 99% at the first post-treatment assessment (Week 1), and 100% from Week 6 through the final assessment (Week 8) when all study dogs were flea-free. For both groups, at each timepoint, flea counts on dogs and in traps were significantly reduced compared to the initial assessment (p < 0.001), as were improvements in median CADESI-4, FAD and CPSS scores (p ≤ 0.001). At Week 4, the geometric mean flea count on dogs in the lotilaner group (0.1) was significantly lower than that of dogs in the spinosad group (0.6) (p = 0.027), significantly fewer dogs in the lotilaner group were found to have fleas (p = 0.034), and mean owner-rated pruritus scores were significantly lower (p = 0.025). Under field conditions favoring heavy flea challenge, two consecutive monthly treatments of dogs with either lotilaner or spinosad produced a 100% reduction in canine flea infestations and dramatic improvements in dermatologic lesions and pruritus, based on scoring by a veterinary dermatologist and by dog owners. Household flea burdens were driven to extinction in all but one home in each treatment group.

In-home assessment of flea control and dermatologic lesions in dogs provided by lotilaner (Credelio®) and spinosad (Comfortis®) in west central Florida.

Post-launch field investigations of recently-approved flea control products establish an efficacy baseline and in subsequent years can detect any efficacy decline suggestive of emerging resistance. As part of a continuing program of yearly assessment of flea control products in west central Florida, this study, using client-owned dogs, investigated the efficacy of lotilaner and spinosad in controlling fleas and in alleviating dermatologic signs likely associated with flea infestations. Forty-four qualifying households were randomized to either a lotilaner (Credelio®) (minimum dose rate 20 mg/kg) or a spinosad (Comfortis®) (30 mg/kg) group, with 33 and 36 dogs in each group, respectively. On Days 0 and 28 (±2) all dogs in each household were treated with the allocated product according to label directions, and all household cats received spinetoram (Cheristin®). On Day 0 and at weekly intervals through Day 56 (±2), on-animal and premises flea burdens were enumerated, a veterinary dermatologist scored integumental changes using canine atopic dermatitis extent and severity index (CADESI)-4 and flea allergy dermatitis (FAD) scales, and owners scored pruritus using the validated canine pruritus severity scale (CPSS). At study entry geometric mean flea counts were 33.2 and 29.9 in the lotilaner and spinosad groups, respectively. For both groups, reductions in flea counts were > 99% at the first post-treatment assessment (Week 1), and 100% from Week 6 through the final assessment (Week 8) when all study dogs were flea-free. For both groups, at each timepoint, flea counts on dogs and in traps were significantly reduced compared to the initial assessment (p < 0.001), as were improvements in median CADESI-4, FAD and CPSS scores (p ≤ 0.001). At Week 4, the geometric mean flea count on dogs in the lotilaner group (0.1) was significantly lower than that of dogs in the spinosad group (0.6) (p = 0.027), significantly fewer dogs in the lotilaner group were found to have fleas (p = 0.034), and mean owner-rated pruritus scores were significantly lower (p = 0.025). Under field conditions favoring heavy flea challenge, two consecutive monthly treatments of dogs with either lotilaner or spinosad produced a 100% reduction in canine flea infestations and dramatic improvements in dermatologic lesions and pruritus, based on scoring by a veterinary dermatologist and by dog owners. Household flea burdens were driven to extinction in all but one home in each treatment group.

Efficacy of a topically applied spot-on formulation of a novel insecticide, metaflumizone, applied to cats against a flea strain (KS1) with documented reduced susceptibility to various insecticides.

A spot-on metaflumizone formulation was evaluated in adult domestic short hair cats to determine its adultidical efficacy against a flea strain that has reduced susceptibility to a number of insecticides. Eight cats served as non-treated controls, eight cats were treated with a metaflumizone formulation at 0.2 ml/kg (40 mg metaflumizone/kg) and eight cats were treated with fipronil 10% w/v-(s)-methoprene 12%w/v at 0.075 ml/kg (7.5-7.7 mg fipronil/kg:9.0-9.2 mg (s)-methoprene/kg). On days -1, 7, 14, 21, 28, 35, and 42 each cat was infested with approximately 100 unfed KS1 cat fleas, Ctenocephalides felis. At approximately 48 h after treatment or infestation, each cat was combed to remove and count live fleas. Treatment with metaflumizone provided > or = 99.3% efficacy for 3 weeks post-treatment and then 97.4, 91.4 and 86.2% efficacy at 4, 5 and 6 weeks post-treatment, respectively. Fipronil-(s)-methoprene provided 99.6% efficacy at 1 week post-treatment and then 97.6, 96.4, 71.3, 22.0 and 13.1% efficacy at weeks 2, 3, 4, 5 and 6, respectively. The reductions in flea numbers were significantly greater for the metaflumizone treatment than for fipronil-(s)-methoprene from 3 to 6 weeks after treatment.

In-home assessment of either topical fluralaner or topical selamectin for flea control in naturally infested cats in West Central Florida, USA.

BACKGROUND: An investigation was conducted in West Central Florida, USA to evaluate the efficacy of either topically applied fluralaner or topically applied selamectin to control flea infestations, minimize dermatologic lesions and reduce pruritus in naturally flea infested cats over a 12-week period. When dogs were present in the households, they were treated with either oral fluralaner (if household cats were treated with topical fluralaner) or oral sarolaner (if household cats were treated with topical selamectin). METHODS: Thirty-one cats in 20 homes were treated once with fluralaner topical solution on day 0 and 18 dogs in these homes were administered a single fluralaner chewable. Twenty-nine cats in 18 homes were treated once monthly with a selamectin topical solution for 3 treatments and 13 dogs in these same homes were treated once monthly for 3 treatments with a sarolaner chewable. Fleas on cats were counted by flea combing, fleas on dogs were estimated using visual area counts and fleas in the indoor premises were assessed using intermittent-light flea traps. Blinded-assessments of feline dermatologic lesions were conducted monthly and pruritus severity was evaluated by pet owners. RESULTS: A single topical application of fluralaner reduced flea populations on cats by 96.6% within 7 days and by 100% at 12 weeks post-treatment. This efficacy was significantly greater than selamectin treatment where single topical application reduced flea populations on cats by 79.4% within 7 days of initial treatment and 3 consecutive monthly treatments reduced flea populations by 91.3% at the end of 12 weeks. At the end of the 12-week study, all fluralaner-treated cats were flea-free and this was significantly greater than the 38.5% of selamectin treated cats that were flea-free. At the end of the study, fleas were completely eradicated (from cats, dogs and homes) in 95.0% of fluralaner treatment group homes, significantly greater than the 31.3% of selamectin/sarolaner treatment group homes with complete flea eradication. Owner reported cat pruritus was reduced similarly in both treatment groups. Significant improvements in dermatologic lesion scores were achieved by day 30 in fluralaner treated cats and by day 60 in selamectin treated cats. CONCLUSIONS: An in-home investigation in subtropical Florida found that 1 application of topical fluralaner eliminated flea infestations on cats and in homes significantly more effectively than 3 consecutive monthly doses of selamectin.

A digital microfluidic system for serological immunoassays in remote settings.

Serosurveys are useful for assessing population susceptibility to vaccine-preventable disease outbreaks. Although at-risk populations in remote areas could benefit from this type of information, they face several logistical barriers to implementation, such as lack of access to centralized laboratories, cold storage, and transport of samples. We describe a potential solution: a compact and portable, field-deployable, point-of-care system relying on digital microfluidics that can rapidly test a small volume of capillary blood for disease-specific antibodies. This system uses inexpensive, inkjet-printed digital microfluidic cartridges together with an integrated instrument to perform enzyme-linked immunosorbent assays (ELISAs). We performed a field validation of the system's analytical performance at Kakuma refugee camp, a remote setting in northwestern Kenya, where we tested children aged 9 to 59 months and caregivers for measles and rubella immunoglobulin G (IgG). The IgG assays were determined to have sensitivities of 86% [95% confidence interval (CI), 79 to 91% (measles)] and 81% [95% CI, 73 to 88% (rubella)] and specificities of 80% [95% CI, 49 to 94% (measles)] and 91% [95% CI, 76 to 97% (rubella)] (measles, n = 140; rubella, n = 135) compared with reference tests (measles IgG and rubella IgG ELISAs from Siemens Enzygnost) conducted in a centralized laboratory. These results demonstrate a potential role for this point-of-care system in global serological surveillance, particularly in remote areas with limited access to centralized laboratories.

Efficacy of a topically applied formulation of metaflumizone on cats against the adult cat flea, flea egg production and hatch, and adult flea emergence.

A spot-on metaflumizone formulation was evaluated to determine its adulticidal efficacy, effect upon egg production, and ovicidal activity when applied to flea infested cats. Eight male and eight female adult domestic shorthair cats were randomly assigned to either serve as non-treated controls or were treated topically with a minimum of 40mg/kg metaflumizone in single spot-on Day 0. On Days -2, 7, 14, 21, 28, 35, 42, 49, and 56, each cat was infested with approximately 100 unfed cat fleas, Ctenocephalides felis felis. On Days 1, 2, and 3, and at 48 and 72h after each post-treatment reinfestation, flea eggs were collected and counted. At approximately 72h after treatment or infestation, each cat was combed to remove and count live fleas. Egg viability was determined by examining hatched eggs after 5 days and adult emergence was determined 28 days after egg collection. Metaflumizone provided >/=99.6% efficacy against adult fleas from Days 3 to 45 following a single application. Following treatment, egg production fell by 51.6% within 24h and 99.2% within 48h. Following subsequent weekly infestations egg production from treated cats was negligible out to Day 38, with >/=99.5% reduction relative to non-treated cats. Where there were eggs to evaluate, metaflumizone treatment did not have any apparent effect on the hatching of eggs or on the development and emergence of adult fleas from the eggs produced by fleas from treated animals.