Abstract
Background
Infestations with fleas, ticks and intestinal nematodes are commonly identified in domestic dogs and have significant veterinary and public health importances. The study described in this manuscript was conducted to verify the natural parasitism of domestic dogs in two Brazilian regions, and to confirm the efficacy of the isoxazoline afoxolaner, formulated alone (NexGard®) or in combination with the macrocyclic lactone milbemycin oxime (NexGard Spectra®), for the control of fleas and ticks, and to confirm the efficacy of the afoxolaner/milbemycin oxime (MO) combination for the control of intestinal nematodes.
Methods
This trial was conducted in Mato Grosso (MT) and Maranhão (MA) states, Center-West and Northeast region. Three hundred and forty-five (345) domestic dogs, 170 in MT and 175 in MA, infested with at least one of these parasites, were studied. Dogs were treated once and evaluated approximately two and four weeks later for ectoparasites, and two weeks later for intestinal nematodes.
Results
Approximately half (47.5%) of the dogs were infested with more than one parasite, more than one third (38.6%) with intestinal nematodes. All fleas were identified as Ctenocephalides felis, ticks as Rhipicephalus linnaei (previously defined as Rhipicephalus sanguineus s.l.) and intestinal nematodes eggs as Ancylostoma, Toxocara or Trichuris. In Maranhão, 11 dogs were also infested with the chewing louse Heterodoxus spiniger. Both products had a comparable efficacy against fleas (98.4% to 100%), ticks (96.9% to 100%) and lice (100%). The MO formulation had an efficacy of ≥ 97.8% against Ancylostoma, ≥ 94.7% against Toxocara and ≥ 97.7% against Trichuris.
Conclusions
This study demonstrated a significant parasitic abundance in domestic dogs in two urban areas in Brazil, and confirmed a high efficacy of afoxolaner against fleas, ticks and lice and a high efficacy of MO against intestinal nematodes.
Graphical Abstract
Keywords: Dog, Efficacy, Afoxolaner, Milbemycin oxime, Ctenocephalides felis, Rhipicephalus linnaei, Heterodoxus spiniger, Ancylostoma, Toxocara, Trichuris
Background
Fleas, ticks, lice and intestinal nematodes are commonly identified in domestic dogs and have a significant veterinary and public health importance [1–3]. These parasites can have a direct pathogenic effect on the host, associated to the immune reactions caused by their presence [4], their life cycle, and their feeding on host tissues [5]. Many ectoparasites have an indirect pathogenic effect, being vector of filarial nematode, protozoan, bacterial and viral diseases. Importantly, several canine parasites and several vector-borne agents are zoonotic [3, 6–10], and the local awareness of their presence is important in a One-Health perspective.
Afoxolaner is an insecticide and acaricide isoxazoline compound formulated as an oral chewable ectoparasiticide tablet for dogs (NexGard®, Boehringer Ingelheim). Milbemycin oxime (MO) is a macrocyclic lactone nematicide compound formulated in combination with afoxolaner as an oral chewable endectoparasiticide tablet for dogs (NexGard Spectra®, Boehringer Ingelheim). Isoxazolines antagonize the GABA-gated chloride ion channels [11–13], while macrocyclic lactones mainly agonize the Glutamate-gated chloride ion channels [14, 15] of the parasites. Afoxolaner, alone or in combination with MO has been reported efficacious several times against fleas [16–18], ticks [18–21], mites [22–25], flies [26, 27] and lice [28, 29], in laboratory or in field conditions. Milbemycin oxime, in combination with afoxolaner has also been reported efficacious against several intestinal nematodes [30–32].
This manuscript describes a field study conducted in two cities in the Mato Grosso and Maranhão Brazilian states, located, respectively, in the Center-West and Northeast administrative regions of Brazil, for evaluating the efficacy and tolerance of NexGard® and NexGard Spectra® in dogs naturally infested with fleas, ticks and/or intestinal helminths. The objectives of this study were to verify the absence of local variations on sensitivity to the parasiticide molecules in the field and to evaluate the parasitism of domestic dogs in these regions. A recent epidemiological study on gastro-intestinal nematodes and protozoans conducted in Cuiaba Mato Grosso, in 353 healthy pet dogs, revealed that 22.7% were infected with at least one parasite, the main one being Ancylostoma sp., and that 5.9% of them were mix-infected with at least two parasites [33]. Another epidemiological study on gastro-intestinal nematodes and protozoans, and ectoparasite conducted in Rondônia, Amazon Biome, in disadvantaged urban communities, in 183 pet dogs, revealed that 74.2% were infected with at least one genus of endoparasite, the main one being Ancylostoma sp., and 43% with ticks and/or fleas [34].
Methods
This field trial was conducted during the period October 2018–February 2019, in urban areas, in two states in Brazil, Mato Grosso (Sinop and Ipiranga do Norte cities) and Maranhão (metropolitan region of São Luís). Although both São Luís and Sinop have tropical climates with high temperatures throughout the year, their environmental conditions differ markedly due to their geographic separation (~ 1,600 km) and distinct climatic influences. São Luís experiences a humid tropical coastal climate characterized by high relative humidity and a short dry winter (Aw Köppen classification), whereas Sinop, located inland in the Amazon–Cerrado transition zone, presents a tropical monsoon climate influenced by the South American monsoon system, with a pronounced dry season (Am Köppen classification) [35].
Study design
This study was conducted in accordance with the relevant WAAVP (World Association for the Advancement of Veterinary Parasitology) guidelines for the evaluation of efficacy of parasiticides on dogs and cats against fleas and ticks [36] and anthelmintics [37]; and the relevant VICH (Veterinary Internation Conference in Harmonization) International Cooperation on Harmonisation of Technical Requirements for Registration of Veterinary Medicinal Products guidelines: VICH GL7, “Efficacy of Anthelmintics: General Requirements”, VICH GL19 “Efficacy of Anthelmintics: Specific Recommendations for Canines”, and VICH GL9 “Good Clinical Practices”.
This study was designed to test the efficacy of NexGard® and NexGard Spectra® against natural infestations with fleas, ticks and intestinal nematodes (NexGard Spectra® only) in the field in two regions of Brazil, and to confirm their safety.
The study was fully conducted in households. The households were recruited on voluntary basis, without other veterinary-related purposes. To qualify for the study, a dog was naturally infested with fleas, ticks or intestinal nematodes. To avoid situations too different from a standard domestic dog household (e.g. a shelter with high density of dogs), a maximum of three dogs per household were included for efficacy assessment, and a maximum of five dogs per household were acceptable (households with dogs and cats were excluded). The same treatment was administered to all dogs in a household. Dogs infected with intestinal nematodes were assigned to the NexGard Spectra® group, the nematicide endectoparasiticide product. Dogs only infested with fleas and/or ticks were assigned to the NexGard® group, the ectoparasiticide product (except for dogs within a household of a dog assigned to NexGard Spectra®, in which case treated with the same treatment). Animal characteristics, assignment and treatment details are described in Table 1. This assignment strategy was necessary to evaluate the parasiticide efficacy of the two products and include all diagnosed parasites in the evaluations, it did not influence the group sizes as visible in Table 1. In a household, dogs that were not included for efficacy evaluation received the same treatment and were evaluated for safety. No masking procedure was implemented during this study.
Table 1.
Animal characteristics and treatment assignments
| Treatment | n | Sex (n) | Bodyweight (kg) | Age | Location (n) | ||||
|---|---|---|---|---|---|---|---|---|---|
| Male | Female | Min | Max | Min | Max | MT | MA | ||
| Nexgard | 149 | 78 | 71 | 2.0 | 38.6 | 2 months | 15 years | 86 | 63 |
| NexGard Spectra | 196 | 99 | 97 | 2.0 | 46.3 | 2 months | 13 years | 84 | 112 |
Data are presented as number of animals (n) or minimum–maximum values, as indicated
MT: Mato Grosso state; MA: Maranhão state
Following treatment, dogs were assessed for ectoparasites and/or intestinal nematodes based on their diagnosed parasitism at inclusion day.
No parasiticide compound with remanent activity within the three previous months of inclusion had been used on dogs or in their environment.
Animals
Three-hundred and forty-five (345) domestic dogs (177 males, 168 females) of various age (2 months to 15 years), bodyweight (2 to 46 kg) and breed (85% mongrels) were included for efficacy assessment. These dogs belonged to 84 households in Mato Grosso, and 130 households in Maranhão. The sample size was defined based on the minimum number of evaluable dogs required per parasite and per geographic region for the calculation of antiparasitic efficacy. All dogs were infested with at least six live fleas and/or four live ticks (nymphs and/or adults) and/or were infected with intestinal nematode (hookworm, roundworm, whipworm) through identification of eggs in feces (there was no defined inclusion criterion for lice, as these parasites were not expected to be found). Dogs were maintained at home during the study in their usual routine, environmental and husbandry conditions.
Treatment
On Day 0, dogs were treated with NexGard® (when they had been included on the basis of ectoparasite infestation, only) or with NexGard Spectra® (when they or any other dog from their household had been included on the basis of endoparasite infestation, namely). The products were administered per label recommendations (orally and with a tablet strength corresponding to their bodyweight, delivering 2.5 to 7.1 mg/kg afoxolaner and 0.5 to 1.1 mg/kg milbemycin oxime in the combined product). In Mato Grosso, 86 and 84 dogs and in Maranhão, 63 and 112 dogs were treated with NexGard® and NexGard Spectra®, respectively (exclusive of the 21 dogs that were not evaluated for efficacy).
Parasite evaluations
Live flea counts were performed by combing all parts of the animal’s haircoat using a fine-tooth comb (11 to 13 teeth per centimeter) for at least 5 min after the encounter of the last ectoparasite on the animal’s skin.
Live nymph and adult ticks found during the flea combing process or haircoat search were counted and removed manually with fingertips or forceps. The interdigital and pad areas, internal face of ears, pubic, perineal and axillary areas were also individually checked.
All removed ectoparasites from each dog were collected and kept stored in ethanol 70% solution for species identification based on morphological characteristics. Fleas and ticks were examined under stereomicroscope and identified using dichotomous keys according to Linardi and Guimarães [38], as well as taxonomic references described by Aragão and Fonseca [39] subsequently updated by Martins et al. [40]. Lice specimens were mounted on glass slides using a drop of lactophenol solution and identified following the criteria established by Werneck [41, 42].
Intestinal nematodes, i.e. hookworms (Ancylostoma sp.), roundworms (Toxocara sp.) and whipworms (Trichuris sp.) were evaluated from fecal samples by copromicroscopy using Centrifugal Flotation and McMaster techniques, for identification at genus level per morphological characteristics and calculation of eggs per gram (EPG). The species of roundworm and whipworm were probably T. canis and T. vulpis, but were not identified at molecular level, which is a limitation of this study. Fecal material was not obtained from all the dogs that were included for ectoparasite efficacy evaluation due to lack of feces available from a few animals, by the time of the inclusion visit.
Within seven days of treatment (on the day of treatment for most of the dogs), dogs were evaluated for ectoparasites and intestinal nematodes. After treatment, dogs were evaluated for the parasites for which they had been included, i.e. fleas and/or ticks and/or intestinal nematodes. For ectoparasites the post-treatment evaluations were performed on Days 14 (± 4) and 30 (± 4), for intestinal nematodes on Days 14 (± 4).
Tolerance evaluations
Owners were requested to observe their dog(s) for two hours after treatment and daily throughout the study and report any abnormality to the veterinarian in charge. At the scheduled visits (on Days 14 (± 4) and 30 (± 4) when applicable), a veterinarian performed a physical examination, which together with any adverse reaction reported by owners (resulting or not in an unplanned veterinary consultation, veterinary care, concurrent medication) was considered for an evaluation of the tolerance. Relatedness to treatment for all adverse reactions and abnormalities were evaluated by the Investigator. This involved the 345 dogs included for efficacy assessment and another 21 dogs from multiple dog households that were treated but not assessed for efficacy, as not diagnosed positive for a parasite at inclusion or being a 4th or 5th dog in the household.
Statistical analysis
No untreated control group was used in this study, each dog served as its own control, the parasite count obtained before treatment at inclusion serving as control value. For each dog, the number of live fleas, live attached ticks and the EPG by nematode genus were transformed to the natural logarithm of “count + 1” for analysis and calculation of the geometric means. The percent efficacy was calculated using the formula 100 × [(B − T)/B], where B = geometric mean of the Day 0 (baseline) visit count and T = geometric mean of the appropriate visit day count. The geometric mean was calculated by taking the anti-logarithm of the average of the log-counts and then subtracting 1, or was computed by taking the anti-logarithm of the least square mean minus 1 from the analysis model. The flea and tick efficacies were also calculated using arithmetic means.
Normality was first assessed using the D’Agostino–Pearson test, which indicated non-normal distribution of parasite count data; therefore, pre- and post-treatment comparisons were performed using the Wilcoxon signed-rank test for paired samples with statistical significance set at P < 0.05.
Results
Parasitism at inclusion
The parasites detected at inclusion are detailed in Table 2 and the types of (co)-infestations and are described in Table 3.
Table 2.
Parasitism per individual dog at inclusion by infestation pattern and state
| Infestations | Brazilian State | Total | |
|---|---|---|---|
| MA (n = 175) | MT (n = 170) | ||
| Single Infestation | 181 (52.5%) | ||
| Ancylostoma sp. | 14 | 6 | 20 |
| Toxocara sp. | 1 | 0 | 1 |
| Flea | 32 | 47 | 79 |
| Tick | 37 | 44 | 81 |
| Dual Infestations | 100 (29.0%) | ||
| Ancylostoma sp., Toxocara sp. | 10 | 0 | 10 |
| Ancylostoma sp., Trichuris sp. | 1 | 0 | 1 |
| Ancylostoma sp., Toxocara sp., Trichuris sp. | 0 | 2 | 2 |
| Flea, Ancylostoma sp. | 7 | 8 | 15 |
| Flea, Lice | 2 | 0 | 2 |
| Tick, Ancylostoma sp. | 8 | 5 | 13 |
| Tick, Flea | 37 | 12 | 49 |
| Tick, Toxocara sp. | 0 | 7 | 7 |
| Tick, Lice | 1 | 0 | 1 |
| Multiple Infestations (≥ 3 parasites) | 64 (18.5%) | ||
| Flea + Intestinal nematodes | 5 | 4 | 9 |
| Tick + Intestinal nematodes | 1 | 13 | 14 |
| Flea, Tick + Intestinal nematodes | 11 | 22 | 33 |
| Flea, Lice, Ancylostoma sp. | 1 | 0 | 1 |
| Tick, Flea, Lice | 7 | 0 | 7 |
MT: Mato Grosso state; MA: Maranhão state.To qualify for inclusion, a dog was infested with at least six live fleas and/or four live ticks and/or was diagnosed infected with intestinal nematode (Ancylostoma sp., Toxocara sp., Trichuris sp.). It was not possible to evaluate feces from all included animals during inclusion, so the co-infection with helminths by dogs harboring ectoparasites is underestimated
Table 3.
Overall distribution of parasitism categories at inclusion (n = 345)
| N | %* | |
|---|---|---|
| Ectoparasite | 311 | 90.1 |
| Endoparasite | 133 | 38.6 |
| Ectoparasite only | 212 | 61.4 |
| Endoparasite only | 34 | 9.9 |
| Ecto and endoparasite | 99 | 28.7 |
| Flea | 195 | 56.5 |
| Tick | 205 | 59.4 |
| Flea and tick | 40 | 11.6 |
*Percentage relative to the 345 included dogs. It was not possible to evaluate feces from all included animals during inclusion, so occurrence of intestinal helminths in dogs harboring ectoparasites and co-infection by helminths is underestimated
Among the 311 dogs infested with ectoparasites, 66% were infested with ticks and 63% with fleas. All the ticks were determined to be Rhipicephalus linnaei (previously defined as Rhipicephalus sanguineus s.l.) and the fleas Ctenocephalides felis.
Among the 133 dogs harboring endoparasites, 110 (83%), 45 (34%) and 21 (19%) were shedding hookworm, roundworm and/or whipworm eggs, respectively.
Interestingly, 11 dogs belonging to nine different households in the Maranhão state were also infested with lice identified as Heterodoxus spiniger.
Parasiticide efficacy
All the results described below are inclusive of all dogs in both regions and of both treatments for ectoparasites. No significant regional or treatment effects were observed.
The flea and tick efficacy results are detailed in Table 4. For the 204 dogs evaluated for ticks, the efficacy (based on geometric means) ranged from 96.9% to 100% on Day 14, and from 97.7% to 100% on Day 30.
Table 4.
Tick and flea efficacy of NexGard and NexGard Spectra in dogs from Mato Grosso and Maranhão states (Brazil) at Inclusion, Day 14 (± 4), and Day 30 (± 4) based on geometric (Geo mean) and arithmetic (A mean) means of live attached tick or flea counts (minimum–maximum)
| Tick efficacy—NexGard | Tick efficacy—NexGard Spectra | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Mato Grosso (n = 50) | Maranhão (n = 46) | Mato Grosso (n = 53) | Maranhão (n = 55) | |||||||||
| Inclusion | Day 14 (± 4) | Day 30 (± 4) | Inclusion | Day 14 (± 4) | Day 30 (± 4) | Inclusion | Day 14 (± 4) | Day 30 (± 4) | Inclusion | Day 14 (± 4) | Day 30 (± 4) | |
|
Geo mean (min, max) |
10.2 (4, 119)a |
0.2 (0, 14)b |
0.2 (0, 5)b |
18.1 (4, 313)a |
0.1 (0, 16)b |
0.0 (0, 6)b |
16.0 (4, 326)a |
0.5 (0, 12)b |
0.3 (0, 9)b |
17.4 (4, 496)a |
0.0 (0, 0)b |
0.0 (0, 0)b |
| % efficacy1 | – | 97.7 | 97.7 | – | 99.4 | 99.8 | – | 96.9 | 98.4 | – | 100.0 | 100.0 |
| A mean | 15.1 | 0.6 | 0.4 | 40.1 | 0.5 | 0.1 | 28.3 | 1.0 | 0.7 | 43.4 | 0.0 | 0.0 |
| % efficacy1 | – | 96.0 | 97.2 | – | 98.9 | 99.7 | – | 96.3 | 97.6 | – | 100.0 | 100.0 |
| Flea efficacy—NexGard | Flea efficacy—NexGard Spectra | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Mato Grosso (n = 47) | Maranhão (n = 46) | Mato Grosso (n = 46) | Maranhão (n = 57) | |||||||||
| Inclusion | Day 14 (± 4) | Day 30 (± 4) | Inclusion | Day 14 (± 4) | Day 30 (± 4) | Inclusion | Day 14 (± 4) | Day 30 (± 4) | Inclusion | Day 14 (± 4) | Day 30 (± 4) | |
|
Geo mean (min, max) |
20.7 (7, 46)a |
0.3 (0, 3)b |
0.3 (0, 6)b |
23.1 (6, 390)a |
0.0 (0, 0)b |
0.0 (0, 0)b |
21.0 (6,195)a |
0.2 (0, 4)b |
0.1 (0,6)b |
27.2 (6, 633)a |
0.02 (0, 1)b |
0.1 (0, 66)b |
| % efficacy1 | – | 98.4 | 98.8 | – | 100.0 | 100.0 | – | 98.9 | 99.6 | – | 99.9 | 99.6 |
| A mean | 23.9 | 0.5 | 0.5 | 52.2 | 0.0 | 0.0 | 31.3 | 0.4 | 0.2 | 59.0 | 0.0 | 1.2 |
| % efficacy1 | – | 97.8 | 97.8 | – | 100.0 | 100.0 | – | 98.7 | 99.4 | – | 99.9 | 97.9 |
Different letters within the same row indicate statistically significant differences among evaluation time points. All comparisons were statistically significant (P < 0.001)
1Percent efficacy = 100 × [(C − T)/C], where C represents the mean parasite count at Inclusion and T represents the mean count at Day 14 or Day 30 (geometric or arithmetic, as indicated)
For the 196 dogs evaluated for fleas, the efficacy ranged from 98.4% to 100% on Day 14, and from 98.8% to 100% on Day 30.
The intestinal nematode efficacy results are detailed in Tables 5, 6. For the 110 dogs evaluated for hookworm the efficacy was ≥ 97.8%, for the 44 dogs evaluated for roundworm the efficacy was ≥ 94.7%, and for the 23 dogs evaluated for whipworm, the efficacy was ≥ 97.7%.
Table 5.
Efficacy of NexGard and NexGard Spectra against intestinal nematodes (Ancylostoma sp., Toxocara sp., and Trichuris sp.) in dogs from Mato Grosso and Maranhão states, Brazil, at inclusion and Day 14 (± 4), based on geometric mean egg counts per gram of feces (Geo mean EPG)
| Ancylostoma sp. | ||||
|---|---|---|---|---|
| Mato Grosso (n = 52) | Maranhão (n = 58) | |||
| Inclusion | Day 14 (± 4) | Inclusion | Day 14 (± 4) | |
| Geo mean | 359.4ª | 7.9b | 492.3a | 1.5b |
| % efficacy1 | – | 97.8 | – | 98.9 |
| Max eggs | 16,400 | 3200 | 10,200 | 700 |
| Min eggs | 100 | 0 | 1 | 0 |
| Toxocara sp. | ||||
|---|---|---|---|---|
| Mato Grosso (n = 27) | Maranhão (n = 17) | |||
| Inclusion | Day 14 (± 4) | Inclusion | Day 14 (± 4) | |
| Geo mean | 383.8ª | 20.5b | 520.1a | 2.7b |
| % efficacy1 | – | 94.7 | – | 99.5 |
| Max eggs | 3700 | 1200 | 1753 | 2 |
| Min eggs | 100 | 0 | 1 | 0 |
| Trichuris sp. | ||||
|---|---|---|---|---|
| Mato Grosso (n = 20) | Maranhão (n = 3)2 | |||
| Geo mean | 289.8a | 6.6b | 2.8 | 0.0 |
| % efficacy1 | – | 97.7 | – | 100.0 |
| Max eggs | 1200 | 300 | 8 | 0 |
| Min eggs | 100 | 0 | 1 | 0 |
Different letters within the same row indicate statistically significant differences between Inclusion and Day 14 (± 4). All comparisons were statistically significant (P < 0.001)
1Percent efficacy = 100 × [(C − T)/C], where C is the geometric mean EPG at Inclusion and T is the geometric mean EPG at Day 14
2P-value not calculated due to insufficient sample size
Table 6.
Efficacy of NexGard and NexGard Spectra against Heterodoxus spiniger lice in dogs from Maranhão state (n = 11)
| Heterodoxus spiniger | |||
|---|---|---|---|
| Inclusion | Day 14 (± 4) | Day 30 (± 4) | |
| Geo mean (min, max) | 12.7 (3, 105)a | 0.0b | 0.0b |
| % efficacy1 | – | 100.0 | 100.0 |
| A mean | 23.8a | 0.0b | 0.0b |
| % efficacy1 | – | 100.0 | 100.0 |
Different letters within the same row indicate statistically significant differences among evaluation time points. All comparisons were statistically significant (P < 0.001)
1Percent efficacy = 100 × [(C − T)/C], where C represents the mean parasite count at Inclusion and T represents the mean count at Day 14 or Day 30 (geometric or arithmetic, as indicated)
The lice efficacy results are detailed in Table 6. In Maranhão, Heterodoxus spiniger lice were found on eleven dogs from nine households. Seven were treated with NexGard® and four with NexGard Spectra®. The efficacy was 100% on Days 14 and 30.
All efficacy evaluations were significant (P < 0.001).
Safety
There was no adverse event related to treatment reported by the owners, post-treatment or during the study. No significant abnormality deemed related to treatment was observed during this study.
Discussion
This study allowed the confirmation that, in two regions in Brazil and in field conditions, NexGard® and NexGard Spectra® were efficacious against C. felis and R. linnaei natural infestations, and that NexGard Spectra® is also highly efficacious against Ancylostoma, Toxocara, and Trichuris infestations.
Total elimination of the chewing louse H. spiniger, two and four weeks after the treatment was also demonstrated with the two products on a sample of eleven infested dogs and confirms a previous report of efficacy of afoxolaner in Europe [29]. Even though statistically significant, it may be interesting to test this hypothesis on a higher sample size.
This study did not use a negative control group, but instead the number of parasites counted on each animal at inclusion served as own control values. This is a method recognized by WAAVP and VICH in field trials, it allows treatment of all animals at the time of diagnosis and, providing adequate number of subjects, allows robust efficacy testing.
All the parasites diagnosed and eliminated in this study have a significant veterinary importance. The ectoparasites, have a direct pathogenic effect on their host with a level of severity correlated to their number, duration of infestation and inherent host conditions (e.g. age, immune status). Ticks, fleas and lice may provoke mild to severe inflammatory cutaneous reactions in relation to their bite wounds that may also cause allergic reactions, such as the Flea Allergic Dermatitis, often leading to pruritus-related damages and secondary infections [4, 43]. Systemic effects such as anemia may occur because of spoliation caused by the hematophagous actions of fleas and ticks in high number. Importantly ectoparasites have a significant indirect effect on their vertebrate host, through their ability of vector borne disease (VBD) transmission. This is especially true for ticks but also a reality with fleas and lice [6, 44]. Rhipicehalus linnaei is a recognized vector of the protozoan Babesia vogeli, the bacteria Rickettsia rickettsii (an agent of spotted fever), Anaplasma platys, Ehrlichia canis (the agent of the canine monocytic ehrichiosis), Mycoplasma haemocanis [1, 45]. Ctenocephalides felis and H. spiniger namely carry and transmit Rickettsia, Bartonella, Dipylidium [46, 47]. Identically, the intestinal nematodes identified in this study provoke pathogenic effects by spoliation of nutrients (roundworms and whipworms) and hematophagous action (hookworms), enteritis through irritation or damage of intestinal mucosa, and very importantly tissue damage associated to their migratory cycle (hookworms and roundworms), namely in the liver or in lungs [7, 8].
Several of these vector-borne agents have a high public health significance, namely R. rickettsii Bartonella, D. caninum. Ancylostoma have a high zoonotic potential through cutaneous larva migrans properties and Toxocara have a severe zoonotic potential namely through visceral, cerebellar or ocular tissue larva migrans properties [9, 48].
It is thus important in a One-Health perspective to control ecto- and endoparasites, both for veterinary and public health purposes [49]. The parasiticide effect of afoxolaner, a systemic molecule, requires a blood meal from the arthropod vector. Thus, the direct effect of afoxolaner for blocking of transmission is related to the speed of transmission of the VBD agent. Afoxolaner has been demonstrated efficacious for the prevention of slowly transmitted VBD agents such as Borrelia, and Babesia [50–53]. The efficacy of afoxolaner to block agents transmitted more quickly (e.g. Anaplasma, Rickettsia, Ehrlichia) is unclear, nevertheless the hypothesis that its use in endemic areas may decrease the local biomass of these VBD agents, through a decrease of their vectors is sound and supports a beneficial effect for a decrease of these VBD presence and transmission.
Conclusions
This study demonstrated a significant level of parasitism in domestic dogs in two municipalities in Brazil, and a high efficacy of afoxolaner formulated alone or combined with milbemycin oxime, against local ectoparasites (R. linnaei, C. felis and H. spiniger). Comparably, a high efficacy of milbemycin oxime against the intestinal hookworms, roundworm and whipworms was demonstrated. The veterinary and public health importance of these parasites [47, 54] advocate consistent treatment of the domestic canine population with these products.
Acknowledgements
We gratefully acknowledge the National Institute of Science and Technology in Innovation in Animal Parasitology (INCT-IPA) sponsored by Brazil’s National Council for Scientific and Technological Development (CNPq), grant no. 408812/2024-9 for the high standard of involvement they provided to the study, and staff from R&D Boehringer Ingelheim Animal Health Brazil for the support on the logistics and study data review.
Author contributions
Andre A. Cutolo: study conceptualization, study protocol, study monitoring, funding, writing and language editing, general supervision; Luis. G. R. Pelissoni: study protocol, funding, writing and language editing; Tássia L. Vale: data gathering; Livio M. Costa-Junior: investigation, validation, data analysis, writing and language editing; Isabella C. Sousa: data gathering; Naylene C. S. Silva: data gathering; Cristiano Grisi do Nascimento: study coordination; Maycon Junior Heidmann: data gathering; Bruno Gomes de Castro: investigation, validation, data analysis, writing and language editing; Eric Tielemans: Manuscript drafting.
Funding
The work reported herein was funded by Boehringer Ingelheim Animal Health GA, USA.
Data availability
All relevant data are provided within the paper.
Declarations
Ethics approval and consent to participate
All dog owners were duly informed about study protocol before inclusion, agreeing by writing with all procedures performed after signing the informed consent. The study was approved by Universidade Federal de Mato Grosso (23108.954157/2018-91) and Universidade Federal do Maranhao Institutional (CIAEP: 01.0341.2014) Animal Care and Use Committees (IACUC) with the concurrence of Boehringer Ingelheim.
Consent for publication
All authors have reviewed this document and consent to publish the data.
Competing interests
A.C, L.P and E.T. are employees of Boehringer Ingelheim, manufacturer of the investigated products (NexGard® and NexGard Spectra®) and sponsor of this study.
Footnotes
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
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