The aim of this study was to identify factors that play a key role in the epidemiology of bovine anaplasmosis by adapting a model primarily developed for cattle babesiosis. A cross-sectional observational study was conducted to study the proportion of calf herds in endemic stability/instability for A. marginale in a semi-arid area of Argentina. The A. marginale inoculation rate (h) was calculated from age-specific seroprevalence using double-antigen sandwich ELISA in 58 herds of 4.5-8.5-month-old calves. Herds were considered to be in endemic instability (EI) at h < 0.005 and, therefore, at risk of anaplasmosis outbreaks. A generalized linear model was performed to explore husbandry practices associated with differences in A. marginale transmission. Additionally, spatial clustering of herds with the same immunological status was analyzed using spatial scan statistics (SatScan, Bernoulli model). Spearman\'s correlation was used to explore a possible association between A. marginale h and Babesia bovis and B. bigemina h (data obtained in previous works). Almost half (43 %) of the herds were in the EI zone for A. marginale. Calves raised under forage combinations had a greater risk of being in EI (OR = 5.41, CI95 %OR = 1.43-20.41) than those reared exclusively on permanent pastures, where cattle density is higher (P = 0.01). Moreover, calves from herds treated only with pyrethroids to control ticks had more chances of being in EI (OR = 4.16, CI95 %OR = 1.12-15.38) than calves from herds receiving different acaricide combinations (P = 0.03). Calves from herds subjected to more than two treatments against Haematobia irritans had higher odds for EI (OR = 5.69, CI95 %OR = 1.24-26.11) than those from herds using fewer than two treatments (P = 0.02). The spatial analysis revealed no spatial clustering of the immune status of the herds (P = 0.67 and P = 0.74 for low and high incidence rates, respectively). A significant variation between farms was observed in A. marginale h (CV = 90.38 %). The correlation analysis revealed a strong epidemiological link of A. marginale h with B. bovis h (Rho=0.794, P<0.001) and B. bigemina h (Rho=0.839, P<0.001). Given that R. microplus is the only vector of B. bovis and B. bigemina in the region, the results of this work strongly suggest an active and significant role of R. microplus in the transmission of A. marginale.

Rhipicephalus microplus poses a substantial threat to livestock health and agricultural economies worldwide. Its remarkable adaptability to diverse environments and hosts is a testament to its extensive genetic diversity. This review delves into the genetic diversity of R. microplus, employing three pivotal genetic markers: the cytochrome c oxidase I (COX1) gene, ribosomal genes, and microsatellites. The COX1 gene, a crucial tool for genetic characterization and phylogenetic clustering, provides insights into the adaptability of ticks. Ribosomal genes, such as internal transcribed spacer regions (ITS-1 and2) as well as 18S and 28S, are routinely utilized for species differentiation. However, their use is limited due to indels (insertions and deletions). Microsatellites and minisatellites, known for their high polymorphism, have been successfully employed to study populations and genetic diversity across various tick species. Despite their effectiveness, challenges such as null alleles and marker variations warrant careful consideration. Bm86, a well-studied vaccine candidate, exhibits substantial genetic diversity. This diversity directly influences vaccine efficacy, posing challenges for developing a universally effective Bm86-based vaccine. Moreover, the review emphasizes the prevalence of genes associated with synthetic pyrethroid resistance. Identifying single nucleotide polymorphisms in the acaricide-resistant genes of R. microplus has facilitated the development of molecular markers for detecting and monitoring resistance against synthetic pyrethroids. However, mutations in sodium channels, the target site for synthetic pyrethroid, correlate well with the resistance status of R. microplus, which is not the case with other acaricide target genes. This study underscores the importance of understanding genetic diversity in developing effective tick management strategies. The choice of genetic marker should be tailored based on the level of taxonomic resolution and the group of ticks under investigation. A holistic approach combining multiple markers and integrating additional molecular and morphological data may offer a more comprehensive understanding of tick diversity and relationships. This research has far-reaching implications in formulating breeding programs and the development of vaccine against ticks and tick-borne diseases (TTBDs) as well as strategies for the management of resistant ticks.

Through a collaborative effort across six Sub-Saharan African countries, using recognized international assessment techniques, 23 stocks of three tick species (Rhipicephalus microplus, Rhipicephalus appendiculatus and Amblyomma variegatum) of economic importance for rural small holder farming communities from East and West Africa were collected from cattle, and evaluated in in vitro larval packet tests (LPT). The results demonstrated medium to high resistance to chlorfenvinphos and amitraz across species. Rhipicephalus microplus demonstrated high level alpha-cypermethrin and cypermethrin resistance. Stocks of A. variegatum (West Africa) and R. appendiculatus (Uganda) demonstrated medium level ivermectin resistance. The four least susceptible stocks (East and West African R. microplus, A. variegatum and R. appendiculatus) were taken into in vivo controlled cattle studies where fipronil was found effective against West and East African R. microplus isolates although persistent efficacy failed to reach 90%. Cymiazole and cypermethrin, and ivermectin based acaricides were partially effective against R. microplus without persistent efficacy. Flumethrin spray-on killed A. variegatum within 72 h for up to 10 days posttreatment, however product application was directly to tick attachment sites, which may be impractical under field conditions. A flumethrin pour-on formulation on goats provided persistent efficacy against A. variegatum for up to one-month. Therapeutic control was achieved against R. appendiculatus through weekly spraying cattle with flumethrin, amitraz or combined cymiazole and cypermethrin. A fipronil pour-on product offered four-week residual control against R. appendiculatus (with slow onset of action). Few studies have assessed and directly compared acaricidal activity in vitro and in vivo. There was some discordance between efficacy indicated by LPT and in vivo results. This observation calls for more research into accurate and affordable assessment methods for acaricide resistance. No single active or product was effective against all three tick species, emphasising the need for the development of alternative integrated tick management solutions.

Acaricides are the most widely used method to control the cattle tick Rhipicephalus microplus. However, its use increases production costs, contaminates food and the environment, and directly affects animal and human health. The intensive use of chemical control has resulted in the selection of genes associated with resistance to acaricides, and consumers are increasingly less tolerant of food contamination. This scenario has increased the interest of different research groups around the world for anti-tick vaccine development, in order to reduce the environmental impact, the presence of residues in food, and the harmful effects on animal and human health. There is enough evidence that vaccination with tick antigens induces protection against tick infestations, reducing tick populations and acaricide treatments. Despite the need for an anti-tick vaccine in Mexico, vaccination against ticks has been limited to one vaccine that is used in some regions. The aim of this review is to contribute to the discussion on tick control issues and provide a reference for readers interested in the importance of using anti-tick vaccines encouraging concerted action on the part of Mexican animal health authorities, livestock organizations, cattle producers, and academics. Therefore, it is suggested that an anti-tick vaccine should be included as a part of an integrated tick management program in Mexico.

Interactions within the tick microbiome involving symbionts, commensals, and tick-borne pathogens (TBPs) play a pivotal role in disease ecology. This study explored temporal changes in the microbiome of Rhipicephalus microplus, an important cattle tick vector, focusing on its interaction with Anaplasma marginale. To overcome limitations inherent in sampling methods relying on questing ticks, which may not consistently reflect pathogen presence due to variations in exposure to infected hosts in nature, our study focused on ticks fed on chronically infected cattle. This approach ensures continuous pathogen exposure, providing a more comprehensive understanding of the nesting patterns of A. marginale in the R. microplus microbiome. Using next-generation sequencing, microbiome dynamics were characterized over 2 years, revealing significant shifts in diversity, composition, and abundance. Anaplasma marginale exhibited varying associations, with its increased abundance correlating with reduced microbial diversity. Co-occurrence networks demonstrated Anaplasma\'s evolving role, transitioning from diverse connections to keystone taxa status. An integrative approach involving in silico node removal unveils the impact of Anaplasma on network stability, highlighting its role in conferring robustness to the microbial community. This study provides insights into the intricate interplay between the tick microbiome and A. marginale, shedding light on potential avenues for controlling bovine anaplasmosis through microbiome manipulation.

Rhipicephalus (Boophilus) microplus, also known as the cattle tick, causes severe parasitism and transmits different pathogens to vertebrate hosts, leading to massive economic losses. In the present study, we performed a functional characterization of a ribosomal protein from R. microplus to investigate its importance in blood feeding, egg production and viability. Ribosomal protein S18 (RPS18) is part of the 40S subunit, associated with 18S rRNA, and has been previously pointed to have a secondary role in different organisms. Rhipicephalus microplus RPS18 (RmRPS18) gene expression levels were modulated in female salivary glands during blood feeding. Moreover, mRNA levels in this tissue were 10 times higher than those in the midgut of fully engorged female ticks. Additionally, recombinant RmRPS18 was recognized by IgG antibodies from sera of cattle naturally or experimentally infested with ticks. RNAi-mediated knockdown of the RmRPS18 gene was performed in fully engorged females, leading to a significant (29 %) decrease in egg production. Additionally, egg hatching was completely impaired, suggesting that no viable eggs were produced by the RmRPS18-silenced group. Furthermore, antimicrobial assays revealed inhibitory activities against gram-negative Escherichia coli and gram-positive Staphylococcus aureus bacteria, affecting bacterial growth. Data presented here show the important role of RmRPS18 in tick physiology and suggest that RmRPS18 can be a potential target for the development of novel strategies for tick control.

Tick-borne viruses (TBV) have gained public health relevance in recent years due to the recognition of human-associated fatal cases and the increase in tick-borne disease and transmission. However, many tick species have not been studied for their potential to transmit pathogenic viruses, especially those found in Latin America. To gain better understanding of the tick virome, we conducted targeted amplification using broadly-reactive consensus-degenerate pan-viral targeting viruses from the genera Flavivirus, Bandavirus, Uukuvirus, and Orthonairovirus genus. Additionally, we conducted unbiased metagenomic analyses to investigate the presence of viral RNA sequences in Amblyomma cajennense, A. patinoi and Rhipicephalus microplus ticks collected from a horse slaughter plant in Medellín, Colombia. While no viral products were detected by PCR, results of the metagenomic analyses revealed the presence of viral genomes belonging to the genera Phlebovirus, Bandavirus, and Uukuvirus, including Lihan Tick Virus (LTV), which was previously reported in Rhipicephalus microplus from Colombia. Overall, the results emphasized the enormous utility of the next-generation sequencing in identifying virus genetic diversity presents in ticks and other species of vectors and reservoirs.

Rhipicephalus microplus, the cattle fever tick, is the most important ectoparasite impacting the livestock industry worldwide. Overreliance on chemical treatments for tick control has led to the emergence of acaricide-resistant ticks and environmental contamination. An immunological strategy based on vaccines offers an alternative approach to tick control. To develop novel tick vaccines, it is crucial to identify and evaluate antigens capable of generating protection in cattle. Chitinases are enzymes that degrade older chitin at the time of moulting, therefore allowing interstadial metamorphosis. In this study, 1 R. microplus chitinase was identified and its capacity to reduce fitness in ticks fed on immunized cattle was evaluated. First, the predicted amino acid sequence was determined in 4 isolates and their similarity was analysed by bioinformatics. Four peptides containing predicted B-cell epitopes were designed. The immunogenicity of each peptide was assessed by inoculating 2 cattle, 4 times at 21 days intervals, and the antibody response was verified by indirect ELISA. A challenge experiment was conducted with those peptides that were immunogenic. The chitinase gene was successfully amplified and sequenced, enabling comparison with reference strains. Notably, a 99.32% identity and 99.84% similarity were ascertained among the sequences. Furthermore, native protein recognition was demonstrated through western blot assays. Chitinase peptide 3 reduced the weight and oviposition of engorged ticks, as well as larvae viability, exhibiting a 71% efficacy. Therefore, chitinase 3 emerges as a viable vaccine candidate, holding promise for its integration into a multiantigenic vaccine against R. microplus.

Rhipicephalus (Boophilus) microplus is one of the most successful ticks infesting cattle around the world. This highly-invasive species transmits cattle parasites that cause cattle fever leading to a high socio-economic burden. Tick eradication programs have often failed, due to the development of acaricide resistance. Here we characterize acaricide resistance in a large number of tick isolates from regions in South Africa (KwaZulu Natal, Mpumalanga, Western & Eastern Cape provinces) and two Brazilian regions. By means of Larval Packet Tests (LPT\'s) acaricide resistance was evaluated against five commonly used acaricides (chlorfenvinphos, fipronil, deltamethrin, amitraz, and ivermectin). Furthermore, the coding region containing the knock down resistance (kdr) mutation, known to result in pyrethroid resistance, was sequenced. Resistance to at least one acaricide class was reported in each of the five regions, and a high proportion of tick isolates exhibited multi-resistance to at least two acaricide classes (range: 22.2-80.0%). Furthermore, resistance ratios (RR) showed high spatial variation (intercontinental, as well as regional) but low regional spatial autocorrelation. Previous and current acaricide use correlated with current RR, and several combinations of acaricide RR were positively correlated. Moreover, fipronil resistance tended to be higher in farms with more intense acaricide use. The kdr-mutations provided the ticks a fitness advantage under the selection pressure of synthetic pyrethroids based on population (kdr-allele frequency) and individual level data (genotypes). The data show the threat of acaricide (multi-)resistance is high in Brazil and South Africa, but acaricide specific levels need to be assessed locally. For this purpose, gathering complementary molecular information on mutations that underlie resistance can reduce costs and expedite necessary actions. In an era of human-caused habitat alterations, implementing molecular data-driven programs becomes essential in overcoming tick-induced socio-economic losses.

Stochastic models are valuable tools to describe and analyze tick population dynamic in a given area, and to evaluate different control schemes. The objective of this study was to provide a tool to predict a priori how a control scheme could affect the abundance of Rhipicephalus microplus in an area highly favourable for its development, through the building of a stochastic model of the population dynamic of this tick. The dynamic was stochastically modelled using field data of the parasitic and non-parasitic phases of R. microplus. The host susceptibility to tick infestations was also considered by including representative values of natural resistance of three different breeds (Bos indicus, B. taurus and B. indicus x B. taurus). Two different control schemes (strategic and threshold methods) using three annual applications of synthetic acaricides in different moments were evaluated. Furthermore, we have evaluated the impact of different moments of pasture spelling as a tool for tick control on the dynamic of the non-parasitic phases of R. microplus. The results showed that the model appropriately fits to field data and can simulate the annual variability in female ticks count in animals from different ecological characteristics. The model has also captured the variability that host susceptibility has on tick abundance. This fact was expressed in the different levels of tick abundance obtained for the three breeds of bovines. According to the model, strategic control applications of chemical acaricides have more efficacy than the threshold control method. The model indicates that a pasture spelling started in early or mid-summer generates a higher reduction in the number of available eggs in pastures. Analysis of the results allows to conclude that the model developed in this study was useful to predict a priori the performance of different control methods based on the strategic application of chemical acaricides or by applying pasture spelling.