Bruno Dall’Agnol

First report of fluazuron resistance in Rhipicephalus microplus: A field tick population resistant to six classes of acaricides

The control of the cattle tick Rhipicephalus microplus is based mainly on the use of chemical acaricides, which has contributed to the emerging problem of selection of resistant tick populations. Currently, there are six main classes of acaricides commercially available in Brazil to control cattle ticks, with fluazuron, a tick growth regulator with acaricidal properties, being the only active ingredient with no previous reports of resistance. Ticks (designated the Jaguar strain) were collected in a beef cattle ranch located at Rio Grande do Sul state, Southern Brazil, after a complaint of fluazuron treatment failure. To characterise the resistance of this strain against acaricides, larval tests were performed and showed that the Jaguar strain was resistant to all of the drugs tested: cypermethrin (resistance ratio, RR=31.242), chlorpyriphos (RR=103.926), fipronil (RR=4.441), amitraz (RR=11.907) and ivermectin (3.081). A field trial was conducted to evaluate the efficacy of fluazuron treatment in heifers that had been experimentally infested with the Jaguar or a susceptible strain. Between 14 and 28 days after treatment, the average efficacy in cattle experimentally infested with the susceptible strain was 96%, while for the Jaguar strain the efficacy was zero. Additionally, the Jaguar strain response to fluazuron was evaluated in vitro using a modified adult immersion test (AIT) and the artificial feeding assay (AFA). With the AIT, 50 ppm of fluazuron inhibited 99% of larvae hatching in the susceptible strain (POA) and less than 50% in the Jaguar strain. Results of the AFA showed a larval hatching rate of 67% at 2.5 ppm of fluazuron with the Jaguar strain; conversely, only 3% of larvae of the susceptible strain hatched at the same fluazuron concentration. The results showed here demonstrated the first case of fluazuron resistance in R. microplus and the first tick population resistant to six classes of acaricides in Brazil.

Multiple resistance to acaricides in field populations of Rhipicephalus microplus from Rio Grande do Sul state, Southern Brazil

Acaricide resistance is a major obstacle to the control of Rhipicephalus microplus. Historically, the indiscriminate use of chemical compounds has contributed to the selection of populations resistant to different classes of acaricides. Therefore, multiple acaricide resistance is an important threat to the chemical control of the cattle tick. To investigate the occurrence and extent of multiple resistance to acaricides in Southern Brazil we performed larval tests with cypermethrin, chlorpyriphos, amitraz, fipronil and ivermectin on 104 cattle tick field samples from different ranches in Rio Grande do Sul, between the years 2013 and 2015. Adult immersion tests with a commercial formulation mixture of chlorpyriphos and cypermethrin were performed on 75 samples. Four levels of resistance were established according to the mortality of larvae: Level I: mortality between 82% and 95%; Level II: mortality between 57% and 82%; Level III: mortality between 25% and 57%; and Level IV: mortality lower than 25%. Resistance to cypermethrin was detected in 98.08% of the samples evaluated, mostly at resistance level IV. The frequency of samples resistant to amitraz, chlorpyriphos, ivermectin and fipronil was 76.92%, 60.58%, 60.58% and 53.85% respectively. Multiple resistance to three or more compounds was found in 78.85% of the samples. The results obtained in this study are alarming and reveal a new scenario for the challenge of tick control using chemicals. This is an issue of high importance to cattle production systems where this tick is responsible for a high economic impact.

Spotted Fever Group Rickettsia in the Pampa Biome, Brazil, 2015–2016

To the Editor: Several cases of tickborne rickettsiosis have been reported in South America in recent years (1,2). In Brazil, 2 spotted fever group (SFG) Rickettsia species, R. rickettsii and Rickettsia sp. strain Atlantic Rainforest, have been identified as causes of human disease. Rio Grande do Sul is the southernmost state of Brazil and the only part of the country located in the Pampa biome. Despite confirmed cases of human spotted fever in that state since 2005, little information is available regarding Rickettsia species. We report an eco-epidemiologic investigation of R. parkeri in Amblyomma tigrinum ticks on dogs from a household (and neighborhood) where a case of human spotted fever was diagnosed.

Integrated control of an acaricide-resistant strain of the cattle tick Rhipicephalus microplus by applying Metarhizium anisopliae associated with cypermethrin and chlorpyriphos under field conditions.

The efficacy of the fungus Metarhizium anisopliae to control ticks has been shown in several in vitro experiments. However, few studies have been undertaken in field conditions in order to demonstrate the applicability of its use as a biological control of ticks and its combination with chemical acaricides. The aim of the present study was to evaluate the efficacy of M. anisopliae to control an acaricide-resistant strain of Rhipicephalus microplus under laboratory and field conditions. First, the compatibility of M. anisopliae strain (TIS-BR03) with commercial acaricides and its potential to control the cattle tick were evaluated in vitro. In general, acaricide treatments had mild effects on fungus viability. In the field experiment, the median of treatment efficacy with acaricide only, M. anisopliae only and combination of M. anisopliae with acaricide were 71.1%, 56.3% and 97.9%, respectively. There is no statistical difference between groups treated with M. anisopliae and acaricide alone. Thus, in this work we have demonstrated the applicability of M. anisopliae use associated or not with chemical acaricides on field conditions in order to control an acaricide-resistant strain of the cattle tick R. microplus.

Rickettsia parkeri in Amblyomma dubitatum ticks in a spotted fever focus from the Brazilian Pampa

Spotted fever is an acute febrile illness, which is considered severely underreported and misdiagnosed in the Brazilian Pampa, caused by tick-borne Rickettsiae. Here, we report an eco-epidemiological investigation of Rickettsia spp. in ticks from a spotted fever focus in Toropi, southern Brazil. Ticks were collected from capybara carcasses and processed individually to obtain genomic DNA. Rickettsia was investigated using PCR that amplified the rickettsial fragments of the gltA, ompA and htrA genes. DNA from Rickettsia parkeri was found in four of 14 Amblyomma dubitatum ticks collected from capybara carcasses in Toropi and the nearby municipality of Quevedos. We also tested 210A. dubitatum ticks obtained from road-killed capybaras of other localities from the Pampa biome; none of them were positive for Rickettsiae. Thus, in Rio Grande do Sul, two Rickettsia species can be potentially associated to spotted fever: Rickettsia sp. strain Atlantic Rainforest, associated with Amblyomma ovale ticks in the Atlantic Rainforest biome, and R. parkeri, associated both with Amblyomma tigrinum and A. dubitatum ticks in the Pampa biome. Our results reinforce that R. parkeri may be the agent associated with spotted fever in the Brazilian Pampa.

Borrelia burgdorferi sensu lato in Ixodes longiscutatus ticks from Brazilian Pampa

Borrelia burgdorferi sensu lato (s.l.) complex includes the agents of Lyme disease/borreliosis in North America, Europe, and Asia, such Borrelia burgdorferi sensu stricto, Borrelia afzelii, Borrelia garinii, Borrelia bavariensis, Borrelia spielmanii, Borrelia bissettiae, and Borrelia mayonii. In 2013 B. burgdorferi s.l. was reported for the first time in the Neotropical region, from Ixodes aragaoi ticks in Uruguayan Pampa. In addition, from 2011 to 2016, 17 suspected human cases of borreliosis-like syndrome were reported in Rio Grande do Sul (RS) state, Brazil, which contains only part of country in the Pampa biome. The goal of this work is to report the results of a state surveillance program conducted in order to investigate the presence of B. burgdorferi s.l. in its classic vector, Ixodes spp. ticks, from the Brazilian Pampa. For this, we searched for Ixodes spp. ticks in 307 rodents from 11 municipalities of RS state. We then tested the ticks for the presence of B. burgdorferi s.l. DNA using PCR analysis. Of 35 Ixodes spp. ticks tested, one larva and one nymph of Ixodes longiscutatus ticks tested positive for Borrelia sp. DNA. The phylogenetic analysis of the flaB fragment grouped our samples (referred as Borrelia sp. haplotype Pampa) into B. burgdorferi s.l. group in a particular branch with other South American haplotypes, and this group was close to Borrelia carolinensis, B. bissettiae, and Borrelia californiensis. This is the first evidence of B. burgdorferi s.l. circulation in ticks of the genus Ixodes in Brazil. These results highlight the need for the implementation of public health policies for the diagnosis and prevention of potential cases of human borreliosis in Brazil. Further studies are needed to fill the gaps in our knowledge of the distribution, pathogenicity, reservoirs, and vectors of these emerging South American B. burgdorferi s.l. haplotypes.

Evaluation of the vector competence of six ixodid tick species for Rangelia vitalii (Apicomplexa, Piroplasmorida), the agent of canine rangeliosis

Rangelia vitalii is the etiologic agent of canine rangeliosis, a severe piroplasmosis that affects domestic dogs in Brazil, Uruguay and Argentina. While R. vitalii is one of the most pathogenic tick-borne pathogens for dogs in the world, its tick vector has remained unknown. The present study evaluated the vector competence of Rhipicephalus sanguineus sensu lato (both tropical and temperate species), Amblyomma aureolatum, Amblyomma ovale, Amblyomma tigrinum, and Amblyomma sculptum for R. vitalii. These six tick species were selected for the study because they comprise the main tick species infesting dogs within the distribution area of canine rangeliosis in South America. Acquisition feeding of the above six tick species was performed on domestic dogs showing clinical signs of canine rangeliosis, after being experimentally infected through intravenous inoculation or infestation with R. vitalii-infected ticks. Thereafter, engorged ticks were evaluated for transstadial and transovarial passages of R. vitalii through molecular analysis after molting or oviposition and egg hatching. The resultant ticks were evaluated for their competence to transmit R. vitalii to susceptible dogs. Among the six tick species, only A. aureolatum was able to acquire and perpetuate R. vitalii by transstadial and transovarial passages, as demonstrated by >5% infection rates of ticks after hatching or molting. When exposed to transmission feeding, only A. aureolatum ticks were competent to transmit R. vitalii to dogs, which became severely ill, and the results confirmed by molecular methods and blood smear examination to have acquired rangeliosis. Results of the present study, coupled with epidemiological data, indicate that A. aureolatum is a natural vector of R. vitalii. Our results also indicate that R. vitalii is the first Piroplasmorida agent to be transovarially transmitted in Amblyomma ticks.

Records of ticks on humans in Rio Grande do Sul state, Brazil

More than seventy tick species have been reported in Brazil. Despite the emergence of tick-borne diseases in Neotropical region, there are still limited data available on tick species parasitizing humans in Brazil. Rio Grande do Sul is the southernmost state of Brazil, comprising the only part of Brazilian territory inside the Pampa biome, as well as the transition between subtropical and temperate zones. Here, we report on human parasitism by ticks in Rio Grande do Sul state between 2004 and 2017. Seventy cases of human parasitism by ticks were recorded, with a total of 81 tick specimens collected. These included 11 tick species belonging to three genera of Ixodidae (hard-ticks), Amblyomma, Haemaphysalis and Rhipicephalus; and one genus of Argasidae, Ornithodoros. The most prevalent tick species associated to cases of human parasitism were Amblyomma parkeri (24%), Rhipicephalus sanguineus sensu lato (22%), Amblyomma aureolatum (15%) and Amblyomma ovale (12%). A spatial analysis showed two major hot spots of human parasitism by ticks in Rio Grande do Sul state. The findings of this study highlight the need for permanent monitoring of human parasitism by ticks in order to provide a better understanding of tick and tick-borne disease eco-epidemiology, and the early identification of potential cases of tick-borne diseases, particularly in spotted fever endemic regions.

Molecular survey of Rickettsia spp. in the Neotropical deer tick Haemaphysalis juxtakochi from Brazilian Pampa

Spotted fever (SF) is a tick-borne disease associated with Rickettsia spp.. In the Pampa biome, Southern Brazil, cases of SF seem to be strongly linked with the practice of hunting wild animals. An investigation of rickettsiae in tick species found on wild animals could provide more information regarding the rickettsiosis enzootic cycle. The aim of this study is to describe the results of a molecular survey of Rickettsia spp. in the Neotropical deer tick, Haemaphysalis juxtakochi Cooley, 1946 (Acari: Ixodidae), from the Brazilian Pampa. Ticks were obtained from 14 road-killed gray brocket deer, Mazama gouazoubira (Artiodactyla: Cervidae), found in nine different municipalities of Rio Grande do Sul state, Southern Brazil. Ticks were processed individually to obtain genomic DNA, and then Rickettsia spp. was investigated using a set of PCR reactions that amplified the rickettsial fragments of the gltA, ompA, and htrA genes. Of the 24 tick samples tested, DNA of Rickettsia parkeri sensu stricto (s.s.) was found in 11 H. juxtakochi specimens collected in two different areas of the Brazilian Pampa. This is the first report of R. parkeri s.s. (the main agent associated with SF in the Uruguayan, Argentinian, and Brazilian Pampa) in H. juxtakochi ticks. These findings indicate that R. parkeri s.s. may be much more common and widely distributed in the Pampa biome than previously assumed. Moreover, H. juxtakochi ticks and gray brocket deer could participate in the potential spillover of R. parkeri s.s. from endemic to non-endemic areas in the South American Pampa.

Re-emergence of Chorioptes bovis (Acari: Psoroptidae) in cattle in the state of Rio Grande do Sul, Brazil

Here we describe an outbreak of chorioptic mange in cattle, 56 years after its first identification in Brazil. Between the months of June and July 2011, dermatitis characterized by alopecia and crusted and thickened skin at the insertion of the tail and in the ischiorectal fossa was recognized in 40 (35.7%) out of 112 Holstein cows on a farm in the northeastern mesoregion of the state of Rio Grande do Sul, Brazil. After diagnosing mange caused by Chorioptes bovis, the cows were weighed and treated with 0.5% ivermectin, as a pour-on single dose, and were separated into two groups: cows in early lactation and those in late lactation. The survival rate of C. bovis and the healing rate in the two groups of infested cows were monitored every seven days through skin scrapings. After 28 days of evaluation, the cure rate through treatment was greater among cows in early lactation (p <0.0001). The survival rate of C. bovis was higher in cows in late lactation.