The causative agents of Babesiosis are intraerythrocytic protozoa of the genus Babesia. Babesia parasites are present around the world, affecting several mammals including humans, pets and livestock, hence its medical and veterinary relevance. Babesia spp. detection in its invertebrate host is a main point in understanding the biology of the parasite to acquire more knowledge on the host–Babesia–vector interactions, as increasing knowledge of the Babesia lifecycle and babesiosis epidemiology can help prevent babesiosis outbreaks in susceptible mammals. The aim of the present review is to highlight the newest findings in this field, based on a bibliographic compilation of research studies recently carried out for the detection of the main Babesia species found in tick vectors affecting mammalian hosts, including the different tick stages such as adult ticks, larvae, nymphs and eggs, as well as the detection method implemented: microscopic tools for parasite identification and molecular tools for parasite DNA detection by conventional PCR, nested-PCR, PCR-RFLP, PCR-RLB hybridization, real time-PCR, LAMP and RAP assays. Although molecular identification of Babesia parasites has been achieved in several tick species and tissue samples, it is still necessary to carry out transmission experiments through biological models to confirm the vectorial capacity of various tick species.

The indirect fluorescent antibody test (IFAT) is the most frequently used test to conduct seroepidemiological studies so far, and it is regarded as the "gold standard" test for the serological diagnosis of bovine babesiosis. The aim of the present study was to compare the enzyme-linked immunosorbent assay (ELISA) and the rapid immunochromatography test (ICT) for use in the serological diagnosis of cattle exposed to B. bovis in Mexico. The evaluation of test performance was carried out with 30 positive and 30 negative reference sera. A total of 72 bovine sera samples collected from cattle in a region with endemic bovine babesiosis were analyzed by ELISA and ICT, and the results were compared with those of IFAT. Kappa value (k) was also calculated to determine the agreement between tests. The sensitivity and specificity of ELISA for detecting antibodies against B. bovis were 87% (26/30) and 80% (24/30), respectively. The sensitivity and specificity of ICT for detecting antibodies against B. bovis were 90% (27/30) and 83.3% (25/30), respectively. The overall concordance determined for ELISA and ICT was 94.4% (68/72) and 98.6% (71/72), respectively, when the results were compared with those of IFAT. ICT was more sensitive and specific in this comparative study, showing good strength of agreement (k = 0.79) with respect to IFAT. ICT combines a strip-based assay system that is fast, practical, and sensitive for detection of antibodies to B. bovis, which suggests that it could be applied in the field without requiring any laboratory equipment for its use and interpretation of test results.

Canine monocytic ehrlichiosis (CME) is the most common tick-borne disease affecting domestic dogs and other wild canids. It has a worldwide distribution and is associated with the presence of the brown dog tick. Few studies have been conducted in Mexico to identify and characterize Ehrlichia canis genetic variability. In the present study, 111 dogs of different sex, breed, and age from three geographic regions in Mexico were included. All of them had a previous history of tick infestation and/or the presence of one or more clinical signs compatible with CME. All dogs were tested by a commercial ELISA and nested PCR assay for the detection of E. canis. In addition, we analyzed the E. canis genetic diversity from the 16S rRNA gene sequences obtained in this study, along with 15 additional sequences described for E. canis in Mexico and obtained from GeneBank. Serological detection by commercial ELISA results showed overall infection rates of 85.58% (95/111), including 73.1% (30/41) in samples from Guerrero state; 75% (15/20) in Morelos; and 100% (50/50) in Chihuahua. On the other hand, molecular detection (nPCR assay) showed 31.5% (35/111) overall infection rate, with 41.4% (17/41) in Guerrero state; 55% (11/20) in Morelos; and 14% (7/50) in Chihuahua. We observed a high 16S rRNA gene sequence conservancy in most of the E. canis isolates in the three geographical areas from Mexico, including those analyzed in this research, suggesting a common geographic origin among isolates.

RNA-seq technology has been widely used for the characterization of the transcriptome profile induced by several diseases in both humans and animals. In the present study, RNA-seq was used to identify the differential expression of genes associated with the immune response in cattle infected with two different strains of Babesia bigemina, both derived from the same Mexican field isolate, which exhibit distinct phenotypic characteristics: the virulent strain, capable of producing acute clinical signs, and the attenuated strain, capable of stimulating a protective immune response when used as an immunogen with an efficacy greater than 80%. The differential gene expression analysis performed revealed a total of 620 differentially expressed genes (DEGs). However, the intersection of the edgeR and DESeq2 programs used in the bioinformatics analysis only identified 247 DEGs, of which 108 genes were enriched to be closely correlated with the bovine immune response based on gene ontology terms; most of the DEGs obtained encode proteins associated with the major histocompatibility complex, immunoglobulins, and T-cell surface receptors. The infection caused by the attenuated strain induced higher transcription of immune response genes compared to the infection caused by the virulent strain; nonetheless, in both infections, a greater down-regulation than up-regulation was observed. Different immunoglobulin-associated genes were found to be up-regulated in the group inoculated with the attenuated strain, whereas these were down-regulated in the virulent strain-inoculated group. In addition, an up-regulation of the HSPA6, CD163, and SLC11a1 genes was observed in the group inoculated with the virulent strain, previously reported in other Apicomplexan infections. The findings provide relevant information that could contribute to clarifying the immune response associated with an acute bovine babesiosis infection by B. bigemina.

Two hundred and thirty-three blood samples of water buffalo were collected on four farms in Veracruz state and Tabasco state, Mexico, to detect and confirm the identities of Babesia and Anaplasma spp. sequences. Nested PCR assays were used for the amplification of specific genes encoding B. bovis rhoptry-associated protein (RAP-1), B. bigemina SpeI-AvaI restriction fragment, and Anaplasma marginale major surface protein 5 (MSP5). Using DNA sequencing and BLASTn analysis for DNA homology hemoparasite identification, the identities of the hemoparasites were established by comparing the nucleotide sequences obtained in this study with those available in the GenBank database at the National Center for Biotechnology Information (NCBI). Water buffalo infection with at least one of the hemoparasites under study was detected in 45% (105/233) of the blood samples, while a mixed infection with B. bovis and B. bigemina was detected in 6.4% (15/233) of samples. For this cross-sectional study, mixed infections with the three hemoparasites were not detected. BLASTn analysis revealed that the nucleotide sequences of the water buffalo isolates shared sequence identity values ranging from 88 to 100% with previously published gene sequences of B. bovis, B. bigemina, and A. marginale. The current results confirm that water buffalo, as cattle, are also carriers of hemoparasite infections that are tick-transmitted, and suggest that they probably have an important role in the epidemiology of bovine babesiosis in Mexico.

Bovine babesiosis, caused by the protozoan Babesia bigemina, is one of the most important hemoparasite diseases of cattle in Mexico and the world. An attenuated B. bigemina strain maintained under in vitro culture conditions has been used as a live attenuated vaccine; however, the biological mechanisms involved in attenuation are unknown. The objective of this study was to identify, through a comparative transcriptomics approach, the components of the B. bigemina virulent parasites that are differentially expressed in vivo, as opposed to those expressed by B. bigemina attenuated vaccine parasites when inoculated into naïve cattle. The biological material under study was obtained by inoculating spleen-intact cattle with infected erythrocytes containing either the attenuated strain or a virulent field strain. After RNA extraction, transcriptomic analysis (RNA-seq) was performed, followed by bioinformatic Differential Expression (DE) analysis and Gene Ontology (GO) term enrichment. The high-throughput sequencing results obtained by analyzing three biological replicates for each parasite strain ranged from 9,504,000 to 9,656,000, and 13,400,000 to 15,750,000 reads for the B. bigemina attenuated and virulent strains, respectively. At least 519 differentially expressed genes were identified in the analyzed strains. In addition, GO analysis revealed both similarities and differences across the three categories: cellular components, biological processes, and molecular functions. The attenuated strain of B. bigemina derived from in vitro culture presents global transcriptomic changes when compared to the virulent strain. Moreover, the obtained data provide insights into the potential molecular mechanisms associated with the attenuation or pathogenicity of each analyzed strain, offering molecular markers that might be associated with virulence or potential vaccine candidates.