Acaricides Resistance in Ticks: Selection, Diagnosis, Mechanisms, and Mitigation

1Ticks are hematophagous ecto-parasites with global geographical distribution, especially in tropical and subtropical regions, where they parasitize terrestrial and semi-terrestrial vertebrates. Ticks also serve as vector reservoirs for the spread of pathogens among their vertebrate hosts. In the livestock industry, this results in significant losses due to host blood depletion, general discomfort and irritation, reduced dairy and meat production, suppression of immune functions, and damage to hides, among other negative impacts.

2Before the introduction of synthetic acaricides, different chemicals such as cotton-seed oil, fish oil, beaumont crude oil, the combination of lard oil with sulfur as well as lard oil with kerosene oil have been used on the host’s body surface to control tick infestations. In cattle, for example, the combination of kerosene oil with cotton-seed oil and sulfur, kerosene oil with cotton-seed oil, and crude petroleum alone have shown effectiveness against ticks. Arsenic, introduced in 1895, was the first acaricide widely used to control ticks and tick-borne diseases until the introduction of organochlorines (OCs), because it was cheap, stable, and water-soluble. Arsenic was used to control tick infestations in different countries like Australia, South Africa, Mexico, and the USA. In 1937, treatment failure due to the selection of arsenic resistant Rhipicephalus microplus was reported for the first time in Australia, and arsenic resistant Rhipicephalus decoloratus as well as Rhipicephalus appendiculatus and Rhipicephalus evertsi were documented in South Africa in 1940 and 1975, respectively.

3The OCs, organophosphates (OPs), amitraz, synthetic pyrethroids (SPs), macrocyclic lactones (MLs), and fipronil are commonly used acaricides that act on the tick central nervous system through various mechanisms, affecting gamma-aminobutyric acid (GABA)-gated chloride channels, inhibitors of acetylcholine-esterase (AChEs), octopamine tyramine receptors, voltage-gated sodium channels, glutamate gated chloride channels (Glu-Cl), and GABA-receptors, respectively. Different acaricides have specific targets and different modes of action, which affects the reproduction, growth and survival of various tick species. Spraying, washing, pouring, and injections are the different methods for the application of acaricides on host animals. Incorrect dilution, inappropriate application, persistent use and overdosing are the main factors that hasten acaricide resistance selection in ticks. Certain species may be even more prone to develop acaricide resistance, due to favorable conditions created by their broad distribution and/or particularities of their life cycle.

4Rhipicephalus annulatus, R. microplus, and Rhipicephalus australis undergo rapid selection in response to selective pressure and higher concentration of acaricides. Different populations of R. microplus have been found resistant to all acaricides that have been used for their control, including: arsenics, OCs e.g. dichloro-diphenyl-trichloroethane (DDT), OPs, MLs e.g. ivermectin, formamidines e.g. amitraz, SPs, fipronil, and fluazuron. The surveillance of acaricide resistance in various regions is essential in order to adopt adequate treatment approaches. Globally the acaricide-resistant ticks identified through different diagnostic tests have been reported ( Table 1 ).

5In recent years, a number of reviews have discussed different aspects of acaricide resistance, including identification of resistant populations, development of novel diagnostic methods, effects of resistance in tick metabolism, environment pollution, evolution and spread of resistant populations, and actions to mitigate risks related to acaricide resistance. Genetic mutations in ticks frequently result in the selection of acaricide resistance, causing conformational changes in the target site, enhanced acaricide metabolism, or reduced ability by the drug to penetrate the tick’s outer defensive layers. A comprehensive understanding on acaricides regarding their mode of action, resistance development, and single nucleotide polymorphisms (SNPs) in the target sites is needed to mitigate acaricides resistance in different tick species. This review paper aims to summarize and critically discuss different acaricides applied for tick control, with focus on mode of action, genetic polymorphisms on target sites, diagnostic approaches, and mitigation strategies regarding acaricides resistance.

6Tick resistance against acaricides is described as “the specific heritable trait(s) in a population of ticks selected as a result of the population’s interaction with an acaricide”. The number of ticks that remain alive after exposure to the specific applied concentration of acaricides increases significantly. The mutated genes, inherited from the survivor ticks, are initially uncommon and rarely occur in the tick population, but increase in frequency with time. Acquired resistance is thus defined as “a resistance that results from heritable declines in drug susceptibility with the passage of time”, which in turn results in phenotypic resistance. The term tolerance refers to the ability of a parasite to survive exposure to a specific drug dose that would usually be considered effective. Resistance across different active chemical ingredients having similar mechanisms of actions is known as cross-resistance.

7Three main forms of acaricide resistance are generally known ( Figure 1 ). Metabolic resistance is achieved through acaricide detoxification by the metabolic activity of enzymes such as cytochrome P-450s (CYP), esterases, and glutathione S-transferase (GST). Resistance developed through conformational changes in the acaricide target site in neuronal enzymes and receptors, leading to impaired interaction between the drug and the target, is termed “target site modification resistance”. The decreased access of acaricides to the internal body environment due to modifications in the tick outer layer (exoskeleton) is called reduced penetration resistance.

8Figure 2 presents a general timeline showing different acaricides regarding their year of introduction and the first reports of resistance. Organochlorines (OCs) were the first commercially viable synthetic acaricides, and were introduced in 1946. Dichloro-diphenyl-trichloroethane (DDT) and benzene hexachloride (BHC) were the first OCs used as acaricides, and resistance against both drugs was documented in R. microplus as early as 1960 in Australia. The organophosphates (OPs) were introduced approximately in 1955 (dioxathion), and resistance against this acaricide was recorded for the first time in 1987 in R. decoloratus and Amblyomma variegatum from Zambia. Formamidines were introduced in the early 1960s, while amitraz was registered for the first time in 1975, just 2 decades before the first amitraz-resistant R. microplus ticks were reported in Brazil. Amitraz-resistant R. microplus ticks were confirmed in 2001 in Mexico. Synthetic pyrethroids (SPs) were introduced in 1977, while resistance against SPs was observed for the first time just 2 years later, in R. microplus from Australia. Macrocyclic lactones (MLs), in turn, were introduced in 1981, and resistance against MLs was recorded for the first time in 2001 in R. microplus from Brazil. Fipronil was introduced in 1987, and resistant R. microplus ticks were reported for the first time in 2007 in Uruguay. The growth regulator fluazuron was introduced in 1994, and similar to other acaricides, first reports of resistant ticks arised 20 years later, in 2014, in R. microplus from Brazil which were already resistant to all six types of available acaricides.

9Monitoring acaricide resistance in the field tick population is necessary for optimal and strategic use, slowing resistance selection, and surveillance of different acaricide resistant ticks. Diagnosis of resistance against different acaricides is essential and urgently required through efficient and inexpensive tests which should be easily followed.

10The larval packet test (LPT), developed in 1962, uses acaricide-treated filter paper packets to incubate the tick larvae. This test has been used to survey and diagnose resistance against OPs and SPs. The LPT takes 5–6 weeks to be completed, thus it is restricted by the lead time until results are available. The larval immersion test (LIT) was developed in 1966. In this assay, larvae are immersed in the test acaricide solution prepared at the applicable concentrations, to check the mortality rate and to obtain resistant ticks. This test is usually employed to characterize resistance against amitraz and MLs. In adult immersion test (AIT), which has been developed in 1973 and is currently applied in different studies, engorged female ticks are immersed in acaricide solutions prepared according to the tested concentrations. Despite broadly used for different acaricides, the AIT is not suitable to test for resistance against amitraz, which is one of the most commonly used acaricides in Mexico. The larval tarsal test (LTT) was established for the first time in Switzerland in 2011. It is a highly sensitive and time-efficient in vitro test, and has been used to determine resistance levels in R. microplus, as well as other ixodid ticks.

11Organochlorines include ethane-derived chlorates such as DDT, cyclodienes (chlordane, aldrin, dieldrin, endrin, heptachlor, and toxaphene), and compounds related to hexa-chlorocyclohexane such as lindane. Organochlorines were demonstrated to cause mortality, suppression of fertility as well as inhibition of egg hatching in R. annulatus and Haemaphysalis bispinosa engorged female ticks. Resistance against toxaphene in R. decoloratus, R. appendiculatus, and R. evertsi has been documented. Organochlorines have a wide range of activity against arthropods (ticks and insects), but have harmful impacts as may be found in the environment, milk, meat as well as in the fat of animal hosts after their application. This lipophilic property was the main drawback of OCs, for which its use against ticks was banned.

12The target site for DDT is the voltage-gated sodium channels of the axon, and resistance is caused by mutations in the amino acid at the binding site. The targets for dieldrin are GABA-gated chloride channel receptors. The drug modifies the channel properties, causing hyperexcitation by inhibiting the entry of chloride ions into the nerve, ultimately leading to the death of ticks. Cyclodienes cause mutations (single nucleotide polymorphisms, or SNPs within the GABA-gated chloride channel gene that result in the selection of tick resistance. Nucleotide mutations (non-synonymous and synonymous) in R. microplus genes targeted by or associated with specific acaricides have been identified. Mutations related to OCs use in R. microplus were different from the observed for certain insects, however they were located in the same second transmembrane protein domain of the GABA-gated channel (TM-2). The effect of this mutation was unknown, but the modified TM-2 may alter the channel properties, inhibiting the entry of chloride ions into the nerve, and resulting in the insensitivity of the dieldrin’s target site. Polymorphisms had also been identified in the voltage-gated sodium channels of nerve axons, a target of DDT, which causes the channel’s closure after depolarization and results in a consecutive leakage of sodium ions across the membrane of the neuron ( Table 2 ).

13Organophosphates acaricides were introduced in the market mainly for the control of OC-resistant Rhipicephalus ticks, which were alarmingly increasing in numbers. This class of acaricides include coumaphos, chlorpyrifos, chlorfenvinphos, diazinon, dioxathion, and ethion. From 1950s to the early 1970s, the OPs were used for tick control in Australia. They were used in large amounts in the national tick eradication program in Mexico between 1974 and 1984.

14Rhipicephalus microplus resistance against different OPs (coumaphos, chlorpyrifos, and chlorfenvinphos) have been recorded, with the first observations coming from South Africa in 1979, then Mexico in 1980. Organophosphates resistance in R. microplus is attributed to increased esterase enzyme activity, and different microarray techniques have been used to detect resistance against OPs, including coumaphos, in R. microplus. Resistant homozygous R. microplus (strain Tuxpan) were experimentally obtained after the application of OP over seven successive generations.

15Organophosphates do not accumulate in fatty tissues of animals like OCs, however, OPs are more toxic to humans than OCs. The use of OPs poses health risks especially to agricultural and manufacturing workers, and children. Organophosphates target the central nervous system (CNS) and cause restlessness, headache, drowsiness, confusion, slurred speech, emotional lability, psychosis, ataxia. Cardiovascular, respiratory, gastrointestinal, sensory, CNS, and motor systems are the most affected by OPs, with life-threatening outcomes.

16A key enzyme (AChEs), necessary for nervous system functions in R. microplus, is the target site for OPs ( Figure 3 ), as well as carbamates (carbaryl and promacyl). In the presence of a cholinesterase inhibitor, the enzyme would not be available to break down acetylcholine, resulting in a continuous neuron firing, which in turn causes overstimulation of the nervous system and lead to death.

17Mechanisms of resistance to OPs include the modifications of the target site in the AChEs gene, carboxylesterase (CE) gene, and metabolic detoxification. Organophosphates may interact with esterases found in the integument layers of R. microplus, resulting in the overexpression of esterases in larvae as well as in adult R. microplus, and consequent resistance to OPs. Even though tick resistance to OPs is known to occur mostly through changes in metabolic activities and conformational changes in AChEs. Post-translational modifications in AChEs proteins have also been suggested as resistance mechanisms.

18The SNPs associated with OPs could be detected in three genes potentially coding for AChEs in R. microplus ticks: BmAChE-1, BmAChE-2, and BmAChE-3. Searches for mutations in BmAChE-1 and BmAChE-2 associated with OP-resistance could not reveal a clear link to any of the identified amino acid substitutions. All six detected mutations were found in combination with each other except R-86-Q. These mutations are responsible for the insensitivity of R. microplus ticks to OPs ( Table 2 ).

19Formamidines were introduced as alternatives to OPs after their failure. This class of acaricides includes chlordimeform, clenpyrin, chloromethiuron, amitraz, and cymiazole. Chlordimeform was used in cattle’s vats to restore the effectiveness of OPs against OP-resistant R. microplus in Australia. Formamidine acaricides have been suggested to have a harmful impact on tick CNS, by targeting octopamine tyramine receptors, and also inhibit the activity of monoamine oxidase enzymes in R. microplus. The effectiveness of amitraz as an acaricide has been proven for several tick species including R. microplus and R. annulatus. The control efficacy of amitraz was more than 97%, and resulted in weight loss as well as reduced egg production in female ticks. Amitraz was used for rapid removal (within hours) of larvae, nymphs as well as adult ticks from the host.

20In insects, octopamine is present in peripheral and central nervous systems, being involved in proprioception, vision and complex behaviors like flying, walking, and learning. In ticks, the putative octopamine receptor has been identified, and classified as OCT/Tyr receptor in phylogenetic studies. The examined gene was initially described as an octopamine-like G-protein-coupled receptor (GPCR). The GPCRs are known to have a wide range of important roles in eukaryotes, including physiology, reproduction, growth and development, stress responses, feeding, and behavior. They have seven-transmembrane proteins which relay signals in the extracellular environment to the intracellular second messenger system by binding to heterotrimeric G-proteins, adenylate cyclase, cAMPs, and protein kinases. Thus, these receptors transfer signals to second messenger pathways upon interaction with octopamine. Among all acaricides used for tick control in Uganda, 37% of the formulations were composed of amitraz, while this acaricide was the control method of choice in 27% of the surveyed farms in Zambia. Amitraz was effective in controlling ticks, however, resistance against amitraz was detected in R. microplus and other Rhipicephalus species in Australia, South America, and South Africa. The selection of tick resistance against amitraz is a serious problem that attracted urgent attention. It has been observed that the chances of resistance selection become greater due to the excessive use of amitraz against R. microplus, and Hyalomma anatolicum. Research on the ovicidal activity of amitraz against R. sanguineus eggs has shown that one-day old eggs are more sensitive to amitraz than 10-12-day old eggs.

21Drowsiness in animals, dry skin, and a harsh hair coat are frequent side effects of amitraz applications. It can cause toxicity in both animals and humans when swallowed, inhaled, or applied to the animal’s skin in high doses. It is carcinogenic in nature and was removed from the market in 1976.

22Octopamine receptors are found in acari, which makes it a suitable specific target site for acaricides ( Figure 3 ), but no octopamine receptors have been described in mammalian cell membrane. The mechanism of action of amitraz is based on its toxicity-inducing interaction with octopamine receptors. Resistance in R. microplus might occur via conformational changes caused by mutations in the gene encoding octopamine receptor. Four putative amitraz resistance mechanisms have been described, including (1) insensitivity of octopamine tyramine receptor, (2) insensitive β-adrenergic octopamine receptors (BAOR), (3) enhanced expression of monoamine oxidase, and (4) the activity of ATP binding transporters (ABC-transporters). The ABC-transporters act by pumping toxins out of the cell, and serving as a defense mechanism against acaricides in R. microplus.

23Amitraz resistance has been described as a complicated multigenic trait, with recessive resistance alleles. Nucleotide mutations were identified in the octopamine tyramine receptor gene among resistant R. microplus ticks. All the polymorphisms in GPCR target sites were connected with amitraz resistance in R. microplus, and resulted in the insensitivity of the target ( Table 2 ).

24Synthetic pyrethroids are chlorinated or brominated halogenated esters of a chrysanthemic acid isomer and synthetic alcohol molecule. Synthetic pyrethroids were introduced in the late 1970s, after the identification of resistance against formamidines in R. microplus. Permethrin was registered and authorized for use by the US Environmental Protection Agency (US Environmental Protection Agency [USEPA], 2006). Synthetic pyrethroids comprised about 17% of the total acaricides used at the beginning of the 21st century. Different SPs can be categorized as type-I and type-II, according to the presence or absence of a cyano group in the alcohol portion. Cypermethrin, flumethrin, cyhalothrin, and cyfluthrin are other common SPs used to control ticks. Permethrins were the first SPs used as acaricides, followed by cypermethrin and deltamethrin.

25The CYP enzyme family controls the metabolism and detoxification of endogenous and exogenous cell damaging chemicals such as plant toxins, drugs, and pesticides in many arthropods. A study regarding gene expression of CYP and CE related to detoxification of SPs in acaricide-resistant R. microplus showed higher levels of CYP expression in pyrethroid-resistant ticks as compared to susceptible ticks. Resistance against pyrethroid and propetamphos was observed in R. bursa in Iran, and found to be related to the metabolic activities of esterases, CYP, and GST enzymes.

26Commercially available SPs like deltamethrin and cypermethrin did not strongly induce resistance in Hy. anatolicum and R. microplus in India. Reduction in egg masses of R. microplus upon the use of different concentrations of deltamethrin and cypermethrin was observed. A low to moderate level of resistance to λ-cyhalothrin and cypermethrin, respectively, in R. annulatus from Iran. Further, R. sanguineus eggs were more sensitive to cis-cypermethrin and cypermethrin than to coumaphos, although when nymphs and unfed females were treated with coumaphos, cis-cypermethrin, deltamethrin, amitraz, or cypermethrin in a comparative study, they showed sensitivity to cis-cypermethrin and cypermethrin only. Ovicidal activity of cis-cypermethrin, deltamethrin, and cypermethrin was assessed, and results showed that one day old eggs were more sensitive to these acaricides than 10-12 days old eggs. Additionally, different life stages (eggs, larvae, nymph, and adult) of R. sanguineus were also evaluated, with larval stages showing almost 100% mortality.

27Synthetic pyrethroids are poorly soluble in water and highly lipid soluble. They have limited toxic effects against terrestrial animals because of the short life-span in the environment. SPs have been shown to be carcinogenic when animals were exposed to them on a daily basis.

28Voltage-gated sodium channels are integral transmembrane proteins, responsible for electrical propagation in most excitable cells. Sodium ions enter the cell by keeping the voltage-gated sodium channels open (activated) and cause membrane depolarization. The closure of voltage-gated sodium channels for just a few milliseconds after channel activation (fast inactivation) is responsible for the action potential’s falling phase. The mechanism of action of different SPs have been suggested to involve adhesion to the target site (voltage-gated sodium channels), causing these channels to remain open for a longer period, and resulting in a higher influx of sodium ions into the cell, with aperiodic neuronal discharge. The voltage-gated sodium channel (containing four domains (I–IV) each having six segments) is a known SP target, where point mutations may be the cause of resistance in many tick species.

29Type-I SPs cause the repetitive release of sodium ions in response to even a single stimulus, while SPs type-II cause membrane depolarization by interfering with membrane action potential. Type-II pyrethroids also delay the inactivation of voltage-gated sodium channels for longer periods compared with type-I SPs.

30Synthetic pyrethroids resistance is linked to mutations in voltage-gated sodium channel genes of R. microplus, with polymorphisms being identified in two of the four domains (domain-II and domain-III) as shown in Table 2 ( Table 2 ).

31Due to the unusual ability to eliminate both external and internal parasites, macrocyclic lactones are referred to as “endectocide”. Avermectin, doramectin, selamectin, abamectin, ivermectin, eprinomectin, milbemycins, moxidectin, milbemycin oxime, and spinosyns are the MLs most commonly used in cattle. Different MLs have been used for the control of internal (gastrointestinal nematodes and microfilariae) and external (ticks, mites, and mange) parasites. The use of a generic version of MLs, or excessive use of MLs in tropical and subtropical countries have influenced the selection of resistance in R. microplus.

32Environmental, biological, and management factors influence the selection of resistance against these acaricides in R. microplus populations. The rise in potential resistance against ivermectin in R. microplus was observed after 10 generations in four different settings: larvae treated with ivermectin, larvae obtained from ivermectin-treated engorged female ticks, larvae obtained from ivermectin-treated engorged female ticks on an ivermectin-treated host, and larvae obtained from ivermectin-treated engorged female ticks that had high oviposition potential. In R. microplus ticks collected from red deers, resistance against ivermectin was recorded for the first time in Mexico. In R. annulatus, resistance against ivermectin was studied for the first time in Egypt. Rhipicephalus microplus ticks resistant against ivermectin have been reported from India, while ivermectin-resistant Hy. anatolicum ticks were documented from Pakistan. Different MLs have different lipophilic properties, and human handlers should be careful to avoid contact with these drugs. Local swelling or pruritis at the injection site in animals occurs in response to subcutaneous administration of MLs. Additionally, the MLs treated animals suffer weight loss, increased body temperature and muscle tremors. Macrocyclic lactones, specifically ivermectin, become harmful to aquatic life when applied in water or when ivermectin-treated animals enter lakes, ponds, or streams shortly after application.

33Protostome invertebrates, including ticks, express Glu-Cl channels in neurons and muscle, the target sites of MLs ( Figure 3 ). The mechanism of action of ivermectin was characterized by its high binding affinity to Glu-Cl receptors. These proteins control chloride ions movement across the channels located in the muscle and nervous cells, and thus their impairment can cause muscular paralysis and death of ticks by inhibition of neuronal impulses. When chloride gated channels remain open for a long period, conductivity of the membrane becomes irreversibly higher, which causes hyperpolarization as well as paralysis of tick muscles. Several species express efflux transporters called ABC-transporters (a super-family of integral membrane proteins). Extracellular and intracellular membranes can pump out the xenobiotic (drug, chemical, toxins) and endogenous metabolites through these protein channels, which reduces the concentrations of harmful chemicals in the cell. In R. microplus, these efflux pumps work as a defense mechanism against ivermectin, while esterases can also have a role in the detoxification of ivermectin.

34Macrocyclic lactones have been shown to be more effective against single host ticks than multi-host ticks. Different studies reported that R. microplus ticks are resistant to MLs, and mutations were observed in the Glu-Cl gene of R. microplus ticks ( Table 2 ) ( Table 2 ).

35Fipronil is a member of the phenylpyrazolic family of insecticides and has been used to control R. microplus infestations in cattle. While the use of fipronil was introduced in the late 1990s, resistance was only recently detected after the development of stable diagnostic tests. It was persistently used in many countries and has been shown to be highly effective against ticks.

36Fipronil was not widely utilized at first because of its higher costs relative to other chemicals. Its usage and demand increased in the early 2000s due to a decrease in costs and an increase in R. microplus tick populations resistant to other acaricides. Rhipicephalus microplus ticks collected in Mexico have developed resistance against fipronil, however, the extensive use of permethrin in the 1980s may have contributed to the selection of this resistance. Rhipicephalus microplus ticks resistant against fipronil have been reported from different regions such as Uruguay, Switzerland, and Brazil. Other species of the genus Rhipicephalus (R. sanguineus) were also found to be resistant to fipronil. Fipronil-resistant Hy. anatolicum ticks were reported from Pakistan. Resistance against fipronil may have developed at lower rates in R. microplus compared with Hy. anatolicum. Fipronil is effective for tick control, but it is also toxic for honeybees, birds, fish and some other beneficial invertebrates. During fipronil applications, animals including humans (workers) can be exposed through their skin, eyes, or by breathing, resulting in skin irritation, sweating, nausea, vomiting, headache, stomach pain, dizziness, weakness, and seizures.

37Similar to OCs (e.g. lindane, dieldrin), fipronil acts by blocking GABA-receptors, and inhibiting the passage of chloride ions through GABA-gated chloride channels ( Figure 3 ). It may cause mortality when used in high concentrations, while smaller doses may disrupt nervous functions. Fipronil is classified as a selective GABA antagonist because it binds effectively to the GABA-receptor chloride channels of ticks.

38Mutations were also identified within the second transmembrane (TM-2) and third transmembrane (TM-3) domain of the GABA-gated chloride channel gene in fipronil resistant R. microplus ticks ( Table 2 ).

39The fluazuron is a benzoylphenyl urea compound that inhibits chitin formation in R. microplus. Other benzoylphenyl urea compounds such as diflubenzuron, lufenuron, and flufenoxuron are effective against a wide range of insects except the fluazuron, which is effective against tick species. Because of its narrow spectrum of activity and lack of lethal impact on established tick infestations, the fluazuron utilization was low to moderate after its introduction. However, its utilization has steadily increased due to the increasing resistance of R. microplus and R. decoloratus to numerous acaricides such as synthetic pyrethroids and amitraz, ivermectin, and fipronil.

40Fluazuron has shown higher efficacy against ticks than against other arthropods. In Brazil, the field trials of fluazuron resulted in 99% effectiveness after treatment for 21 days. Only larval stages of R. microplus ticks could be observed after the host was treated with fluazuron, and no ticks were found that had reached further developmental stages. Reduction in egg viability also occurred due to fluazuron application. Fluazuron has the ability to accumulate in fat, and may be released from the body through milk, thus protecting calves from ticks without direct application. It has been suggested that fluazuron affected solely the hatchability of larvae from treated female ticks, and that a synergistic effect of fluazuron with MLs negatively impacted reproductive parameters in R. microplus. Fluazuron affected nymphs and larvae, when they were attempting molting, and did not harm any adult ticks.

41Fluazuron showed the highest efficacy (100%) than any other used acaricides against R. microplus ticks, and no resistance was reported against fluazuron, although it was suggested that its inappropriate use may lead to resistance development. In a later study, resistance against fluazuron was recorded in R. microplus ticks.

42Fluazuron inhibits tick growth by interfering with the formation of chitin through inhibition of enzymes involved in the molting process, and results in the hardening of the tick exoskeleton. Fluazuron treatment causes damage to several chitinous structures (smaller hypostome and chelicerae, scutum, sensilla, pores, anal plaque) in R. sanguineus nymphs, structures that play essential roles in tick survival.

43Although the use of fluazuron was necessary for tick control and for cattle management, the careless use of this drug may weaken the immune system of the host, which in turn carries huge risks for diseases. Although SNPs associated with fluazuron have not been reported, further research is essential to characterize its target site.

44Cross-resistance between dieldrin (an OC) and fipronil has been reported, which affects the GABA-gated chloride channels of many insects. The selection of cross-resistance against OCs led to a reduction in the use of these acaricides against different tick populations including R. microplus, R. decoloratus, and R. appendiculatus. It has been reported that there was a significant pattern of cross-resistance in many R. microplus populations against OCs (DDT) and SPs (cypermethrin). Cross-resistance between two OPs (diazinon and coumaphos) and carbamate acaricide (carbaryl) has also been observed in R. microplus ticks. Cross-resistance between fipronil and lindane was examined in R. microplus ticks in Brazil and Uruguay, because both acaricides have the same target. Rhipicephalus microplus populations from Mexico were studied with multiple resistance to OCs, OPs and other acaricides.

45Strategies must be developed to minimize the selection of acaricide resistance in ticks. Different acaricides have different modes of action, thus alternate use may help in the reduction of resistance selection. Rotation of acaricides such as pyrethroids with coumaphos (OPs) has been used, and rotation of amitraz with other groups of acaricides may delay a further spread of amitraz resistance. The alternate use of acaricides, especially amitraz with spinosad, may help in the minimization of resistance selection. It has been recommended to perform further trials of experiments to finalize the conclusions about beneficial aspects of acaricide rotation in the field.

46Synergistic use of acaricide combinations may reduce the chances of resistance selection, however, due to the lack of appropriate field work, this assumption remains to be confirmed. Nevertheless, the combination of fipronil with permethrin was recommended after showing effectiveness against Dermacentor reticulatus, Ixodes ricinus and R. sanguineus ticks. The combination of OPs (chlorfenvinphos and ethion) with SPs (deltamethrin and cypermethrin) has been successfully applied for the control of R. microplus. The combination of OPs (e.g. dichlorvos) with tetrachlorvinphos was successfully used against OP-resistant R. microplus. The combination of different acaricides such as flumethrin with cyfluthrin, chlorpyriphos with permethrin, and cypermethrin with cymiazole were available in the market, and were used for tick control in Mexico. Satisfactory results (high larvae mortality rate) were observed from the combined effects of permethrin and amitraz, while there was zero larvae mortality when permethrin was used alone. The application of combined acaricides may help in delaying the resistance selection, because individual ticks are very unlikely to have resistant alleles of two or more genes for different acaricides with different modes of action. Deltamethrin, thymol, and essential oil derived from Eucalyptus have shown synergistic acaricidal efficacy against R. microplus.

47Besides resistance development, excessive use of acaricides has negative impacts on the environment, animal health, and animal-derived products. For these reasons, environmentally friendly alternative control strategies are needed, which would also be important for the mitigation of acaricide resistance. Numerous plant-derived products have been used to aid tick control. Rhipicephalus microplus ticks were killed at a nearly 100% rate by applying the essential oils of Eucalyptus citriodora and Eu. staigeriana at 10% concentration, while Eu. globulus showed 100% efficacy at 20% concentration. In another study, Eucalyptus spp. leaf extracts showed 96% effectiveness against female R. microplus ticks. Larval mortality of 100% and 98%, and reproductive efficiency inhibition of 100% and 91.5%, were observed in R. microplus treated with Melia azedarach chloroformic and hexanic extracts, respectively. The acaricidal activity of environment friendly essential oils from different plant species has been tested against different ticks. Essential oils obtained from Origanum bilgeri and Cunila species have shown acaricidal activity against Rhipicephalus ticks. Thymol, a major component of essential oils, has shown 100% acaricidal efficacy against R. microplus and R. sanguineus ticks. Essential oils extracted from Allium sativum were 90-100% effective in killing R. microplus larvae. Essential oils extracted from Tagetes minuta have shown 95% efficacy against larvae of Amblyomma cajennense and Argas miniatus. The application of essential oils extracted from Azadirachta indica seeds affected egg hatching and caused 100% larval mortality in Hy. anatolicum ticks. Oils extracted from leaves of Hesperozygis ringens caused 95% inhibition of larval hatchability and 100% larval mortality in R. microplus. Two monoterpenes (carvacrol and thymol) were used alone or in combination with cypermethrin to control R. microplus. It was observed that thymol enhances cypermethrin toxicity, and that both monoterpenes increased the activity of antioxidant and detoxifying enzymes (glutathione-S-transferase, catalase, superoxide dismutase, and glutathione peroxidase) in the R. microplus ticks. The combination of essential oils from Cinnamomum zeylanicum, Cuminum cyminum, and Pimenta dioica in emulsion preparations showed a synergic effect against R. microplus. Furthermore, the combination of essential oils extracted from Laurus nobilis and Copaifera officinalis showed higher efficacy than the isolated oils against R. microplus, indicating that the synergic action could increase the effect of environment-friendly compounds. Some of the mechanisms of acaricidal activity by plant products include inhibition of egg development, suppression of chitin formation, inhibition of growth regulator hormones, and disruption of the mating communications. These products have different modes of action, thereby slowing down the selection of resistant ticks. Factors such as identification of suitable phyto-metabolites, extraction time of phytochemicals, the concentration of extracts, odors of plant-derived extracts, extracted plant age, specie of ticks, and exposure time of ticks may have undermined the research on phytotherapeutic alternatives. During many years, various studies were focused on the identification of natural resources, but recent research has focused on chemical standardization, quality of phytochemicals for tick control, and other issues like long-term stability, storage, and transportation of different plant extracts, which are essential for the commercial availability of these products.

48There are reports of 58 species of fungi found infecting 73 different acari, and several fungi species have shown potential anti-tick efficacy. Beauveria bassiana, Metarhizium anisopliae, Isaria farinosa, Isaria famosorosea, and Lecanicillium lecanii have been shown to infect various ixodid ticks. Environmental factors that affect the persistence and virulence of fungi against ticks include high or low environmental temperature, low humidity, UV radiation, precipitation, soil type, composition and pH (in case of direct application on soil). The stability of some fungal products is very limited, which is why its large-scale production can be expensive. Most manufacturers would recommend exactly the same application method for fungal products as used for other chemical pesticides.

49To minimize the need for acaricides and delay resistance selection, the strategic use of anti-tick vaccines has been strongly and preferably recommended as suitable alternatives for tick control. The first commercial anti-tick vaccine was developed by expressing gut glycoproteins (Bm86) of diversely distributed R. microplus. The effectiveness of this anti-tick vaccine varies in different regions against different ticks, therefore the identification of novel antigens as vaccine candidates are underway. Since Bm86, numerous novel antigens from several tick species have been tested for potential anti-tick vaccine development, however, none has been approved for the market so far.

50The development of alternative approaches to mitigate tick resistance against available acaricides, such as the implementation of non-chemical control methods, application of biosafety protocols for the prevention of resistant ticks, development of novel environment-friendly acaricides, suitable diagnostic methods, and anti-tick vaccines, are in continuous and pressing demand.

51All of the available acaricides have faced failure due to the selection of tick resistance. The repeated use of a single acaricide is often responsible for the selection of fast-developing acaricide resistance. Regular surveillance of ticks for resistance against different acaricides may play a vital role in the minimization of resistance selection. Livestock owners and workers must be trained on tick control and resistance mitigation measures. Environmentally friendly, stable, cheaper, easily accessible, and novel acaricides, as well as anti-tick vaccines should be developed to overcome tick resistance against acaricides.

52MKO, NI, AAlo, AZK and AA searched and collected the literature. ISV, TT, AA supervised the overall investigations. All authors equally contributed to the formulation of the whole manuscript.

53We acknowledge the financial support provided by Pakistan Science Foundation and Higher Education Commission Islamabad Pakistan, researchers supporting project number (RSP2022R494), and the CNPq and CAPES – Brazil for providing research facilities. This work was supported by JSPS KAKENHI Grant Number JP22H02522.

54The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

55All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.