Canine Babesiosis Research Articles

Complications and multiple organ dysfunctions in canine babesiosis: A review

Complications and multiple organ dysfunctions in canine babesiosis: A review

Genetic diversity and molecular evolution of the internal transcribed spacer regions (ITS1–5.8S-ITS2) of Babesia vogeli

Canine babesiosis, a severe haemoparasitic disease caused by Babesia species, has a significant global presence and can be fatal if left untreated. The current study was aimed to perform the population genetic characterization of B. vogeli on the basis of the internal transcribed spacer regions (ITS1–5.8S-ITS2). A maximum likelihood tree constructed with the Hasegawa-Kishino-Yano model grouped all sequences into a single major clade (BvG1), with the exception of a Taiwanese isolate (EF186914), which branched separately. This Taiwanese isolate represented a novel genotype (BvG2) identified in the present study. Nucleotide sequences (n = 62) exhibited 92.5–100 % nucleotide identity among themselves. However, the BvG1 and BvG2 genotypes shared a lower identity of 92.5–93.8 % between them. Notably, the newly generated Indian sequences (n = 21) demonstrated a high degree of homology, with 98.3–100 % identity. Alignment of the nucleotide sequences revealed 58 variations across the dataset. Additionally, 32 sites exhibited variation within the BvG1 genotype, while 56 sites differed between BvG1 and BvG2 genotypes. Within different B. vogeli populations, the nucleotide diversity (π) was low, but the haplotype diversity (Hd) was high. The haplotype diversity of the Indian population, BvG1 genotype and the combined dataset was ~0.8 suggesting a high haplotype diversity. The median-joining haplotype network displayed a total of 21 haplotypes, out of which six haplotypes consisted of more than one sequence (2–25 sequences). Haplotype distribution showed significant geographical structuring, with most haplotypes confined to a single country. Only two haplotypes (9.52 %; Hap_1 and Hap_4) were shared between countries, whereas 19 haplotypes (90.48 %) were country-specific. Hap_1, Hap_6, and Hap_4 were the most representative haplotypes, comprising 25, 10, and four sequences, respectively. India exhibited the highest number of haplotypes (h = 13) followed by China (h = 4), the United States of America (h = 3), Taiwan and Tunisia (h = 2), and Thailand (h = 1). Both location-wise and genotype-wise median joining haplotype networks clustered the haplotypes in two groups, representing two distinct genotypes (BvG1 and BvG2). The B. vogeli populations between Thailand and Tunisia exhibited the highest genetic differentiation (FST = 0.80) with a low gene flow (Nm = 0.125) between them. Results of AMOVA revealed a higher genetic variation within populations (69.43 %) as compared to the variation between them (30.57 %). Neutrality indices and the mismatch distributions of the Indian population and the overall dataset of B. vogeli indicated a constant population size to population expansion and population expansion, respectively, with the presence of two distinct genotypes. These data provide information about parasite population genetics and highlight the importance of starting a long-term molecular surveillance program. In conclusion, a high genetic diversity along with the presence of two distinct genotypes of B. vogeli were observed on the basis of internal transcribed spacer regions (ITS1–5.8S-ITS2).

Diagnosis and Treatment of Canine Babesiosis in a Female German Shepherd in Tandojam, Pakistan

Diagnosis and Treatment of Canine Babesiosis in a Female German Shepherd in Tandojam, Pakistan

Clinical pathology and molecular examination of Babesia spp. infection in dogs; Mashhad, Northeast Iran

This study aimed to fill a crucial gap in our understanding of Babesia infection in dogs in Mashhad, northeast Iran. We not only investigated the prevalence of Babesia species among dogs but also undertook a comprehensive comparison of clinical, hematological, and clinicopathological findings between infected and non-infected cases, a unique aspect of our research. Materials and methodsOur research was conducted with meticulous attention to detail. We randomly collected blood specimens from a diverse population of 150 dogs, including owned pets (n = 47), stray dogs (n = 66), and shelter dogs (n = 37), to ensure the reliability and representativeness of our findings. We then used microscopy and PCR to investigate Babesia spp. infection and analyzed various biochemical and hematological variables. ResultsThe overall prevalence of babesiosis was 15.3 % (23/150) by PCR and 2 % (3/150) by microscopy. Upon microscopic examination, two cases of large Babesia and one case of small-sized Babesia were identified. The sequencing results confirmed that the two dogs testing positive for large-sized Babesia species in this study were both infected with B. vogeli, exhibiting 100 % sequence identity.There was no association between infection and gender, while housing status (k = 37.294, p = 0.000) and age (k = 6.897, p = 0.021) significantly related to infection rate. Among laboratory variables, infection with Babesia spp. showed a remarkable association with Hct (k = 4.749, p = 0.025) and RBC count (k = 14.669, p = 0.000), which were significantly lower in infected dogs compared to non-infected dogs (p < 0.05). Aside from severe non-regenerative anemia observed in all three clinically infected cases, the most clinicopathological changes were observed in one B. vogeli-infected dog, including pancytopenia, azotemia, hyperphosphatemia, hyperkalemia, hypoglycemia, hypocholesterolemia, hyponatremia. ConclusionThis study reveals a higher-than-expected prevalence of canine babesiosis in Northeastern Iran, necessitating further investigation of tick vectors and Babesia spp. distribution. Notably, many infected dogs were asymptomatic, raising concerns about silent spread via carriers. Moreover, the high prevalence of infection in shelters highlights the need for more effective control strategies in these centers.

Canine Babesiosis in Austria in the 21st Century – A Review of Cases

Canine Babesiosis in Austria in the 21st Century – A Review of Cases

Canine Babesiosis and Therapy Options – A Review

Canine Babesiosis and Therapy Options – A Review

A global systematic review and meta-analysis on the babesiosis in dogs with special reference to Babesia canis.

Canine babesiosis is a clinically significant tick-transmitted disease caused by several species of the intraerythrocytic protozoan parasite Babesia, which result in a wide range of clinical manifestations, from mild, transient infection to serious disease and even death. The current study aimed to estimate the global prevalence and associated risk factors of Babesia in dogs. Multiple databases (PubMed, Scopus, ProQuest, Web of Science and Google Scholar) were searched for relevant literature published from January 2000 up to December 2022. The statistical analyses were performed based on the R software (version 3.6) meta-package. Out of 23,864 publications, 229 studies met the inclusion criteria. The pooled prevalence of canine babesiosis was 0.120 (95% CI; 0.097-0.146). The highest pooled prevalence was found in Europe (0.207, 95% CI; 0.097-0.344). Among several species, Babesia canis was the most prevalent parasite (0.216, 95% CI; 0.056-0.441). The highest pooled prevalence of Babesia in dogs was observed in the summer season (0.097, 95% CI; 0.040-0.174). Regular screening and appropriate control strategies are recommended for the prevention of transmission of tick-borne disease transmission among dogs.

Characterization and LC-MS/MS based proteomic analysis of extracellular vesicles separated from blood serum of healthy and dogs naturally infected by Babesia canis. A preliminary study

Canine babesiosis is a rapidly spreading tick-borne disease in Europe, which entails protozoan parasites invading red blood cells. Small extracellular vesicles (EVs) (< 200 nm) were isolated from the serum of 15 healthy and 15 by Babesia canis naturally infected dogs aimed to distinguish EV characteristics and protein profiles. There were no significant differences (P = 0.05) observed in the mean sizes and concentrations of serum EVs between the healthy and canine babesiosis groups. Despite a higher number of Canis lupus proteins detected in EVs from serum of diseased dogs, there were no statistically significant differences (P < 0.05) in the number of protein IDs between the experimental groups. We successfully identified 211 Canis lupus proteins across both experimental groups, of which 147 Canis lupus proteins were validated as being EV-associated. This data set is accessible via the ProteomeXchange PXD047647. EVs isolated from serum of B. canis infected dogs were Cd9+, Cd63+, Cd81+, and Cd82+. Furthermore, 73 Canis lupus proteins were validated as EV-associated and specific for EVs isolated from serum of B. canis-infected dogs. These were predominantly membrane and cytosolic proteins, and innate and adaptive immune system-related proteins, especially those involved in adhesion and proteoglycan mechanisms like integrins. Enrichment was also observed for proteins involved in vascular and cellular responses, including signalling pathways such as VEGF, VEGFR, and the LKB1 network. When only blood-related sites of EV expression were evaluated, the origins of EV proteins were mostly cells of immune system. These were dendritic cells, neutrophils, B cells, monocytes and platelets. In general, proteins were enriched in pathways that collectively regulate various cellular processes, including immune responses, communication, signal transduction, membrane trafficking, and apoptosis. Serum EVs and their protein cargo may have an important role in both the invasion of B. canis and the host's response to the parasitic infection, nevertheless, additional experimental research is warranted. The overall count of identified EV proteins of parasitic origin, meeting cut off criteria of two peptides and 1 % FDR, was relatively low.

Mixed-sex clusters on grass blades: breeding strategy of the ornate dog tick, Dermacentor reticulatus

BackgroundThe ornate dog tick Dermacentor reticulatus is second only to the hard tick Ixodes ricinus in terms of importance as a vector of infectious organisms, especially of Babesia canis, the agent of canine babesiosis. Both the geographical range and local densities of D. reticulatus are steadily increasing in many regions of Europe. In the present study, we tested the hypothesis that D. reticulatus possesses an efficient breeding strategy that allows for a rapid increase in tick numbers and densities through the formation of mixed-sex clusters/aggregations while questing in the environment.MethodsAn observational study was carried out in the spring of 2023, at three sites in two regions in Central and North-Eastern Poland, both characterised by high tick densities. At each site, a 400-m-long transect was inspected for questing ticks. All noted ticks were collected, and tick numbers and sexes per stem were recorded. Differences in tick distribution by site and sex were analysed statistically.ResultsA total of 371 D. reticulatus (219 females, 152 males) ticks were collected from 270 grass stems over a combined 1200 m of transect. The majority of grass stems (74.4%) were occupied by just a single individual, with two-tick clusters the second most common category. The maximum number of D. reticulatus individuals observed on a single grass stem was six.Mixed-sex clusters were significantly more common than single-sex clusters at all three sites. With study sites combined, mixed-sex clusters accounted for 17.4% (95% confidence limit [95% CL] 13.9–21.6%) of observations, while for multiple males and multiple females, the values were 2.6% (95% CL: 1.4–4.7%) and 5.6% (95% CL: 3.7–8.3%), respectively.ConclusionsMixed-sex clusters of D. reticulatus ticks were significantly more common than single-sex clusters, which we hypothesise reflects an efficient, likely pheromone-mediated breeding strategy of this expansive tick species.Graphical

Evaluation of the efficacy of Simparica Trio® in the prevention of the transmission of Babesia canis by infected Dermacentor reticulatus to dogs

BackgroundBabesia canis is a clinically relevant vector-borne pathogen in dogs, and its presence is expanding. The efficacy of Simparica Trio® (Zoetis) in the prevention of B. canis transmission was evaluated at the minimum recommended label dose of 1.2 mg/kg sarolaner, 24 µg/kg moxidectin and 5 mg/kg pyrantel per kg bodyweight.MethodsTwenty-four (24) dogs were randomly allocated to either a placebo-treated group or one of two treatment groups with Simparica Trio. Dogs were infested with B. canis-infected Dermacentor reticulatus ticks 21 or 28 days after treatment administration. Blood samples for antibody and DNA detection were collected from each dog prior to tick infestation until 28 days after infestation. A dog was defined as being B. canis positive if it tested positive by both an indirect immunofluorescence assay (IFA) and PCR at any time during the study.ResultsNo treatment-related adverse reactions were recorded during the study. All placebo-treated animals displayed clinical signs due to babesiosis and tested positive on both IFA and PCR. None of the Simparica Trio-treated animals displayed any clinical symptoms or tested positive, resulting in a 100% efficacy in the prevention of canine babesiosis (P < 0.0001).ConclusionsA single treatment with Simparica Trio at the minimum recommended label dose of 1.2 mg/kg sarolaner, 24 µg/kg moxidectin and 5 mg/kg pyrantel per kg bodyweight prevents the transmission of B. canis by infected D. reticulatus to dogs for at least 28 days.Graphical

Tick-borne pathogens in questing adults Dermacentor reticulatus from the Eastern European population (north-eastern Poland)

Dermacentor reticulatus is tick species with an expanding geographical range in Europe, which creates the possibility of spreading microorganisms of significant veterinary and medical importance. The study aimed to investigate the prevalence and genetic diversity of Rickettsia spp., Babesia spp., Borrelia spp. and Anaplasma phagocytophilum in adult D. reticulatus ticks from the Eastern European population in the urban and the natural biotopes of north-eastern Poland. Microorganisms were detected by PCR and identified by DNA sequencing. The overall infection rate of at least one of the pathogens was 29.6%. The predominantly was Rickettsia spp. (27.1%) (with R. raoultii—9.1%) followed by Babesia spp. (2.4%) with B. canis (1.5%) as the most frequent. Based on 18S rRNA gene sequence, three B. canis genotypes were revealed. The prevalence of R. raoultii and B. canis was significantly higher in ticks from natural biotopes. The infection rates of B. afzelii and A. phagocytophilum were determined at 0.9% and 0.3%, respectively. Co-infections were detected in 3.8% of infected ticks. In diagnosing tick-borne diseases in humans, tick-borne lymphadenopathy should not be excluded. The prevalence of different genotypes of B. canis suggests differences in the clinical picture of canine babesiosis in the area.

Canine babesiosis: A report of two different cases

Canine babesiosis: A report of two different cases

Germany-wide citizen science study reveals spread of Babesia canis-infected Dermacentor reticulatus ticks by dogs travelling within the country

The ornate dog tick Dermacentor reticulatus, vector of Babesia canis, has shown a considerable range expansion in several European countries. Previously, only few areas in Germany were recognised as endemic for B. canis, but a marked increase in autochthonous canine babesiosis cases and spread to new areas has been noted recently. To better assess the current risk for dogs, the present study screened 5913 specimens of D. reticulatus from all over Germany, collected in the frame of a Citizen Science study during 2019–2023. Moreover, 343 Dermacentor marginatus ticks were also included. Babesia detection was achieved by quantitative real-time PCR (qPCR). Positive samples were confirmed by sequencing. Moreover, a MGB-probe-based triplex qPCR was established to detect and distinguish between the canine Babesia spp. relevant in Europe, i.e. B. canis, Babesia vogeli and Babesia gibsoni. Overall, B. canis DNA was detected in five D. reticulatus specimens (0.08%). Two of the B. canis-positive ticks originated from areas previously known as endemic for canine babesiosis, namely from the area of Freiburg im Breisgau, federal state of Baden-Wuerttemberg, and from the district St. Wendel, federal state of Saarland. Three further B. canis-positive ticks were detected in districts not yet recognised as endemic, one each in the district of Mansfeld-Suedharz, federal state of Saxony-Anhalt, the district of Ravensburg, federal state of Baden-Wuerttemberg and in the city of Fürth, federal state of Bavaria. However, the tick in Fürth was found on a dog who had returned from a trip to the Breisgau region on the previous day, indicating translocation of the specimen out of this well-known endemic focus. The geographical distribution of the positive samples shows that B. canis is currently spreading in Germany, particularly via dogs travelling within the country, increasing the infection risk throughout the country. Important measures to contain a further spread of the pathogen include comprehensive year-round tick prophylaxis with licensed acaricides, not only to protect the individual pet, but also the entire dog population. Moreover, screening of dogs entering Germany from B. canis-endemic countries is required and any treatment should aim at pathogen elimination by use of appropriate imidocarb dosages.

What's Eating You? Rhipicephalus Ticks Revisited.

Rhipicephalus ticks are vectors of disease in humans and animals. Rhipicephalus sanguineus sensu lato (the brown dog tick) is one of the most geographically widespread tick species worldwide, likely due to its ability to colonize human and canine dwellings over a range of habitats. They transmit a variety of diseases to dogs and humans, including canine babesiosis, canine monocytic ehrlichiosis, hepatozoonosis, Mediterranean spotted fever, and Rocky Mountain spotted fever. Tick bites manifest as intensely pruritic, erythematous papules at the site of tick attachment; symptomatic relief usually can be achieved with topical antipruritics. Prevention of tick bites is best achieved through a combination of veterinary and environmental control; protective clothing; repellents, such as N,N-diethyl-meta-toluamide (DEET) and permethrin; and prompt identification and removal of ticks.

Analysis of Genetic Diversity of cytb gene from Babesia gibsoni Isolates from Naturally Infected Dogs in Karnataka, India.

The study aimed to investigate genetic diversity in Babesia gibsoni, the causative agent of canine babesiosis, and to assess the presence of atovaquone-resistant isolates in naturally infected dogs. A total of 24 blood samples confirmed for B. gibsoni infection was subjected to PCR amplification and sequencing based on cytb gene. Genetic characterization of B. gibsoni as well as attempts to detect the point mutation rendering atovaquone resistance was carried out based on the analysis of nucleotide sequence of cytb gene using bioinformatics software. The findings indicated that the B. gibsoni isolates in the investigation exhibited a high nucleotide identity with the Asian genotype, ranging from 98.41 to 98.69%. Notably, none of the isolates carried cytb gene variants associated with atovaquone resistance. Phylogenetic analysis revealed clustering of most isolates with those from Japan and China, except for one isolate forming a distinct subclade. Haplotype network analysis indicated a high diversity with 22 distinct haplotypes among the B. gibsoni isolates, emphasizing the genetic variability within the studied population. In conclusion, the cytb gene exhibited remarkable conservation among the twenty-four B. gibsoni isolates studied and the study represents the first genetic diversity assessment of B. gibsoni using the cytb gene in dogs from India. These findings shed light on the genetic characteristics of B. gibsoni in the region and provide valuable insight for addressing the challenges posed by this life-threatening disease in dogs.

Babesia negevi infection in dogs and response to treatment

Canine babesiosis is an important protozoan tick-borne disease associated with anemia and thrombocytopenia and caused by several different Babesia spp. Babesia negevi was first reported to infect dogs in the Middle East in 2020. This study describes the presentation, clinical signs, parasitemia levels quantified by molecular techniques, laboratory findings and treatment of dogs infected with B. negevi following the first description of this species. Clinical findings in the infected dogs, a 3-year old female and two 8-week old male and female pups, included extreme lethargy and pale mucous membranes, anemia and thrombocytopenia found in all three animals. Fever was present in the older female and icterus in the female pup. Babesia parasites resembling B. negevi were detected by microscopy of blood smears from the dogs. PCR of blood targeting the 18S rRNA and cox1 genes confirmed that babesiosis was caused by B. negevi and PCR targeting the Borrelia flagellin gene indicated co-infection with Borrelia persica in two dogs. Treatment of the dogs with imidocarb dipropionate resulted in clinical improvement and initial decrease in the B. negevi parasite load as detected by quantitative PCR in two dogs, however the female pup continued to deteriorate and died. The parasite load in the 3-year old female decreased from 43,451 parasites/µl blood pre-imidocarb dipropionate treatment to 803 parasites/µl within two weeks. In the surviving pup, it decreased from 3,293,538 parasites/µl pre-treatment to 20,092 parasites/µl after two weeks. Babesia negevi DNA was still recovered from blood samples by PCR despite repeated treatment with imidocarb dipropionate one-month post-treatment in the surviving pup and up to seven months post-treatment in the 3-year old female. Only treatment with atovaquone and azithromycin for ten days eliminated B. negevi in both dogs as confirmed by negative PCR two weeks later. In conclusion, treatment with imidocarb dipropionate was helpful for recovery from clinical disease but did not facilitate parasite elimination, and it is therefore recommended to treat canine B. negevi infection with the combination of atovaquone and azithromycin.

Molecular detection of Babesia spp. in dogs in Germany (2007–2020) and identification of potential risk factors for infection

BackgroundIn Europe, canine babesiosis is most frequently caused by Babesia canis and Babesia vogeli, and occasionally by Babesia gibsoni.. In Germany, B. canis is recognized as endemic. The aims of this study were to assess how often Babesia spp. infections were diagnosed in a commercial laboratory in samples from dogs from Germany, and to evaluate potential risk factors for infection.MethodsThe database of the LABOKLIN laboratory was screened for Babesia spp.-positive polymerase chain reaction (PCR) tests for dogs for the period January 2007–December 2020. Sequencing was performed for positive tests from 2018 and 2019. Binary logistic regression analysis was performed to determine the effects of sex, season, and year of testing. Questionnaires were sent to the submitting veterinarians to obtain information on travel abroad, tick infestation, and ectoparasite prophylaxis of the respective dogs. Fisher’s exact test was used to calculate statistical significance and P < 0.05 was considered statistically significant.ResultsIn total, 659 out of 20,914 dogs (3.2%) tested positive for Babesia spp. by PCR. Of 172 sequenced samples, B. canis was identified in 156, B. vogeli in nine, B. gibsoni in five, and B. vulpes in two. Season had a statistically significant impact on test results when summer/winter (1.6% tested positive) was compared to spring/autumn (4.7%), with peaks in April (5.2%) and October (7.4%) [P < 0.001, odds ratio (OR) = 3.16]. Sex (male 3.5%, female 2.8%; P = 0.012, OR = 1.49) and age (< 7 years old 4.0%, ≥ 7 years old 2.3%; P < 0.001, OR = 1.76) of the tested dogs also had a statistically significant effect. A statistically significant impact was demonstrated for observed tick attachment (P < 0.001, OR = 7.62) and lack of ectoparasite prophylaxis (P = 0.001, OR = 3.03). The frequency of positive Babesia spp. tests did not significantly differ between the 659 dogs that had never left Germany and the 1506 dogs with known stays abroad (P = 0.088).ConclusionsThe possibility of canine infection with B. canis needs to be especially taken into consideration in spring and autumn in Germany as the activity of the tick Dermacentor reticulatus, a potential vector for canine babesiosis, is highest in these seasons. Travel and importation of dogs are considered major factors associated with canine babesiosis in Germany. However, autochthonous Babesia spp. infections also occur in a considerable number of dogs in Germany.Graphical

Dogs’ general response to babesiosis infection of various severities

Global climate change in recent decades has led to an increase in the activity and expansion of the range of many diseases. One of them is canine babesiosis. Therefore, it is becoming increasingly important to monitor the functional state of the organism in sick dogs and timely detection of kidney, liver, spleen, cardiovascular system, anaemia, and other disorders that complicate the course of babesiosis. The research aims to determine the peculiarities of metabolic and functional changes in dogs with different intensities of babesiosis infection. During the laboratory diagnostics, general and special research methods were used: light microscopy, centrifugation, spectrophotometry, and mathematical statistics. Based on the results of comprehensive microscopic, morphological, and biochemical studies of blood in dogs with different intensities of babesiosis infection, the most characteristic functional and metabolic changes in their body were identified. Thus, at different intensities of babesiosis infection in diseased animals, the features of the haematological profile are leukocytopenia against the background of eosinopenia (at a mild degree of infection) and lymphocytopenia with a simultaneous compensatory increase in the number of monocytes and neutrophils, as well as erythrocytopenia, hypochromemia, thrombocytopenia with a decrease in thrombocrit and haematocrit. In addition, these patients developed hyperfermentemia of aspartate and alanine aminotransferases, alkaline phosphatase, and γ-glutamyl transpeptidase, indicating structural and functional changes, primarily in the myocardium, liver, skeletal muscle, brain and kidneys due to their toxic damage by babesia waste products. At the same time, hypoproteinaemia, hypoalbuminemia and hyperazotemia were observed in the affected animals, indicating an increase in the intensity of catabolic processes in functional cells of organs and tissues, especially the liver. The most pronounced changes in the studied parameters were observed at high intensity of babesiosis infection. The established regularities allow the use of these haematological parameters as markers of functional and metabolic changes in the body of dogs at different intensities of babesiosis infection

Babesia spp. en derrame peritoneal canino

Canine babesiosis is one of the most relevant hemoparasitoses affecting dogs, its typical clinical signs being lethargy, anorexia, weight loss, weakness, splenomegaly, hemolytic anemia, and thrombocytopenia. The conventional definitive diagnosis of canine babesiosis is made by identification of the intraerythrocytic protozoan in a blood smear. In the present report, the patient had ascites and cytologic analysis of the drained peritoneal effusion identified Babesia spp.

Proteomics identification of overexpressed serum proteins in dogs with Babesiacanis infection.

Canine babesiosis, caused by the protozoan parasite Babesia canis, is characterized by clinical manifestations, including hemolytic anemia, thrombocytopenia, multiple organ failure, and may result in death. This disease is detected using conventional blood smears, which are time-consuming and have low sensitivity. This study aimed to investigate a more rapid and sensitive method for detecting B. canis infection in dogs by examining the expressed serum protein profiles using proteomics. We collected six sera samples from three healthy and three B. canis-infected dogs diagnosed using blood smear and polymerase chain reaction. We analyzed the proteins using two-dimensional gel electrophoresis. The candidate spots from the gel were subjected to protein identification using a nano-liquid chromatography system coupled to an ion-trap mass spectrometer equipped with an electrospray ionization nano-sprayer. We found that 10 protein spots were overexpressed in the serum samples from infected dogs compared with healthy dogs, which corresponded to three proteins: serotransferrin, serotransferrin isoforms X1, and hemopexin. Furthermore, analysis of the protein-protein interaction network confirmed that they strongly interacted with each other. This study suggests that high levels of serotransferrin and hemopexin are related to B. canis infection, making these proteins potential candidates for the development of diagnostic molecules or vaccines.