Lipidomics Profiling Reveals Differential Alterations after FAS Inhibition in 3D Colon Cancer Cell Culture Models.

Cancerous cells synthesize most of their lipids de novo to keep up with their rapid growth and proliferation. Fatty acid synthase (FAS) is a key enzyme in the lipogenesis pathway that is upregulated in many cancers and has gained popularity as a druggable target of interest for cancer treatment. The first FAS inhibitor discovered, cerulenin, initially showed promise for chemotherapeutic purposes until it was observed that it had adverse side effects in mice. TVB-2640 (Denifanstat) is part of the newer generation of inhibitors. With multiple generations of FAS inhibitors being developed, it is vital to understand their distinct molecular downstream effects to elucidate potential interactions in the clinic. Here, we profile the lipidome of two different colorectal cancer (CRC) spheroids treated with a generation 1 inhibitor (cerulenin) or a generation 2 inhibitor (TVB-2640). We observe that the cerulenin causes drastic changes to the spheroid morphology as well as alterations to the lipid droplets found within CRC spheroids. TVB-2640 causes higher abundances of polyunsaturated fatty acids (PUFAs) whereas cerulenin causes a decreased abundance of PUFAs. The increase in PUFAs in TVB-2640 exposed spheroids indicates it is causing cells to die via a ferroptotic mechanism rather than a conventional apoptotic or necrotic mechanism.

Multi-glycomic analysis of spheroid glycocalyx differentiates 2- and 3-dimensional cell models.

A multi-glycomic method for characterizing the glycocalyx was employed to identify the difference between 2-dimensional (2D) and 3-dimensional (3D) culture models with two human colorectal cancer cell lines, HCT116 and HT29. 3D cell cultures are considered more representative of cancer due to their ability to mimic the microenvironment found in tumors. For this reason, they have become an important tool in cancer research. Cell-cell interactions increase in 3D models compared to 2D, indeed significant glycomic changes were observed for each cell line. Analyses included the N-glycome, O-glycome, glycolipidome, glycoproteome, and proteome providing the most extensive characterization of the glycocalyx between 3D and 2D thus far. The different glycoconjugates were affected in different ways. In the N-glycome, the 3D cells increased in high-mannose glycosylation and in core fucosylation. Glycolipids increased in sialylation. Specific glycoproteins were found to increase in the 3D cell, elucidating the pathways that are affected between the two models. The results show large structural and biological changes between the 2 models suggesting that the 2 are indeed very different potentially affecting individual outcomes in the study of diseases.

Enhancement of Lipid Signals in Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry with Ammonium Fluoride as a Matrix Additive.

Lipids are essential macromolecules that play a crucial role in numerous biological events. Lipids are structurally diverse which allows them to fulfill multiple functional roles. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is a powerful tool to understand the spatial localization of lipids within biological systems. Herein, we report the use of ammonium fluoride (NH4F) as a comatrix additive to enhance lipid detection in biological samples, with a signal increase of up to 200%. Emphasis was placed on anionic lipid enhancement with negative polarity measurements, with some preliminary work on cationic lipids detailed. We observed lipid signal enhancement of [M-H]- ions with the addition of NH4F additive attributed to a proton transfer reaction in several different lipid classes. Overall, our study demonstrates that the use of the NH4F comatrix additive substantially improves sensitivity for lipid detection in a MALDI system and is capable of being applied to a variety of different applications.

Association between Duddingtonia flagrans, dimethylsulfoxide and ivermectin for the control of Rhabditis spp. in cattle.

Cattle parasitic otitis caused by the nematode Rhabditis spp. is a serious health problem in Brazil, a situation which is confounded by lack of effective control measures. In vitro studies associating biological and chemical control as an alternative method showed promising results. The objective was to evaluate the combined use of Duddingtonia flagrans (AC001), 10% dimethylsulfoxide (DMSO), and 1.9% ivermectin for the in vivo control of Rhabditis spp., in naturally infected Gyr cattle. For this purpose, 48 animals, whose infection in both ears was diagnosed, were randomly assigned to 6 groups: group 1 (ivermectin 1.9%); group 2 (10% DMSO); group 3 (AC001); group 4 (ivermectin 1.9% + 10% DMSO w/v); group 5 (1.9% ivermectin + AC001 w/v); group 6 (10% DMSO + AC001 v/v). The treatments were performed in a single dose, in the right ears, with the left ears remaining untreated, as a control group. There was a significant reduction (p < 0.01) in the number of nematodes in the treated groups in relation to the control, with the following best efficacies: groups 1 and 2, 47% and 52.9%, respectively, 7 days after treatment; groups 3, 4, and 5, 47.8%, 48.6% and 36.7%, respectively, 14 days post-treatment; group 6, 38.4%, 21 days post-treatment. It was concluded that the combination of chemical compounds and D. flagrans in a single application was effective for the in vivo control of Rhabditis spp. in naturally infected cattle.

Mass spectrometry imaging and LC/MS reveal decreased cerebellar phosphoinositides in Niemann-Pick type C1-null mice.

Niemann-Pick disease type C1 (NPC1) is a lipid storage disorder in which cholesterol and glycosphingolipids accumulate in late endosomal/lysosomal compartments because of mutations in the NPC1 gene. A hallmark of NPC1 is progressive neurodegeneration of the cerebellum as well as visceral organ damage; however, the mechanisms driving this disease pathology are not fully understood. Phosphoinositides are phospholipids that play distinct roles in signal transduction and vesicle trafficking. Here, we utilized a consensus spectra analysis of MS imaging data sets and orthogonal LC/MS analyses to evaluate the spatial distribution of phosphoinositides and quantify them in cerebellar tissue from Npc1-null mice. Our results suggest significant depletion of multiple phosphoinositide species, including PI, PIP, and PIP2, in the cerebellum of the Npc1-null mice in both whole-tissue lysates and myelin-enriched fractions. Additionally, we observed altered levels of the regulatory enzyme phosphatidylinositol 4-kinase type 2α in Npc1-null mice. In contrast, the levels of related kinases, phosphatases, and transfer proteins were unaltered in the Npc1-null mouse model, as observed by Western blot analysis. Our discovery of phosphoinositide lipid biomarkers for NPC1 opens new perspectives on the pathophysiology underlying this fatal neurodegenerative disease.

Considerations for MALDI-Based Quantitative Mass Spectrometry Imaging Studies.

Significant advances in mass spectrometry imaging (MSI) have pushed the boundaries in obtaining spatial information and quantification in biological samples. Quantitative MSI (qMSI) has typically been challenging to achieve because of matrix and tissue heterogeneity, inefficient analyte extraction, and ion suppression effects, but recent studies have demonstrated approaches to obtain highly robust methods and reproducible results. In this perspective, we share our insights into sample preparation, how the choice of matrix influences sensitivity, construction of calibration curves, signal normalization, and visualization of MSI data. We hope that by articulating these guidelines that qMSI can be routinely conducted while retaining the analytical merits of other mass spectrometry modalities.

Effect of silver nanoparticles (AgNP's) from Duddingtonia flagrans on cyathostomins larvae (subfamily: cyathostominae).

The in vitro effect of silver nanoparticles of the Duddingtonia flagrans filtrate enriched with chitin was evaluated on infective larvae of cyathostomins (L3). After biosynthesis, an assay was carried out with two experimental groups in microtubes, for a period of 24 h: G1 (AgNP's-D. flagrans (43.4 µg/mL) + 120 L3) and G2 (distilled water + 120 L3). At the end of this period, AgNP's-D. flagrans (G1) demonstrated an effect on L3 with a 43% reduction (p < 0.01) in relation to G2. Thus, the authors suggest new designs with AgNP's-D. flagrans for the control of cyathostomins.

Developing a Drug Screening Platform: MALDI-Mass Spectrometry Imaging of Paper-Based Cultures.

Many potential chemotherapeutics fail to reach patients. One of the key reasons is that compounds are tested during the drug discovery stage in two-dimensional (2D) cell cultures, which are often unable to accurately model in vivo outcomes. Three-dimensional (3D) in vitro tumor models are more predictive of chemotherapeutic effectiveness than 2D cultures, and thus, their implementation during the drug screening stage has the potential to more accurately evaluate compounds earlier, saving both time and money. Paper-based cultures (PBCs) are an emerging 3D culture platform in which cells suspended in Matrigel are seeded into paper scaffolds and cultured to generate a tissue-like environment. In this study, we demonstrate the potential of matrix-assisted laser desorption/ionization-mass spectrometry imaging with PBCs (MALDI-MSI-PBC) as a drug screening platform. This method discriminated regions of the PBCs with and without cells and/or drugs, indicating that coupling PBCs with MALDI-MSI has the potential to develop rapid, large-scale, and parallel mass spectrometric drug screens.

Mass spectrometry imaging reveals ganglioside and ceramide localization patterns during cerebellar degeneration in the Npc1-/- mouse model.

Mass spectrometry imaging (MSI) is a powerful tool to perform untargeted mapping of biomolecules in situ. In the current study, we performed matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) to evaluate lipid changes during disease progression (asymptomatic to symptomatic time points) in Niemann-Pick disease, type C1 (NPC1), a cerebellar neurodegenerative, lipid storage disorder. Our data show that gangliosides GM2 and GM3 are elevated in NPC1 disease and localize in the posterior lobules of the cerebellum, which is enhanced over a time-course analysis of the disease. Further analysis of sphingolipids in negative ion mode indicated reduction of sulfatides in white matter of the cerebellum and patterned distribution and co-localization of ceramide species Cer(d36:1), HexCer(d36:1), and the ganglioside GM1(d36:1) during disease progression. Finally, a putative lipid of unknown structure demonstrated similar patterning during NPC1 cerebellar degeneration. These studies provide insight into lipid markers of neurodegeneration in NPC1 and link lipid alterations to altered pathways that lead to cell death.

Mass spectrometry imaging of lipids: untargeted consensus spectra reveal spatial distributions in Niemann-Pick disease type C1.

Mass spectrometry imaging (MSI) is a tool to rapidly map the spatial location of analytes without the need for tagging or a reporter system. Niemann-Pick disease type C1 (NPC1) is a neurodegenerative, lysosomal storage disorder characterized by accumulation of unesterified cholesterol and sphingolipids in the endo-lysosomal system. Here, we use MSI to visualize lipids including cholesterol in cerebellar brain tissue from the NPC1 symptomatic mouse model and unaffected controls. To complement the imaging studies, a data-processing pipeline was developed to generate consensus mass spectra, thereby using both technical and biological image replicates to assess differences. The consensus spectra are used to determine true differences in lipid relative abundance; lipid distributions can be determined in an unbiased fashion without prior knowledge of location. We show the cerebellar distribution of gangliosides GM1, GM2, and GM3, including variants of lipid chain length. We also performed MALDI-MSI of cholesterol. Further analysis of lobules IV/V and X of the cerebellum gangliosides indicates regional differences. The specificity achieved highlights the power of MSI, and this new workflow demonstrates a universal approach for addressing reproducibility in imaging experiments applied to NPC1.

Role of Side Chains in ß-Sheet Self-Assembly into Peptide Fibrils. IR and VCD Spectroscopic Studies of Glutamic Acid-Containing Peptides.

Poly(glutamic acid) at low pH self-assembles after incubation at higher temperature into fibrils composed of antiparallel sheets that are stacked in a ß2-type structure whose amide carbonyls have bifurcated H-bonds involving the side chains from the next sheet. Oligomers of Glu can also form such structures, and isotope labeling has provided insight into their out-of-register antiparallel structure [ Biomacromolecules 2013 , 14 , 3880 - 3891 ]. In this paper we report IR and VCD spectra and transmission electron micrograph (TEM) images for a series of alternately sequenced oligomers, Lys-(Aaa-Glu)5-Lys-NH2, where Aaa was varied over a variety of polar, aliphatic, or aromatic residues. Their spectral and TEM data show that these oligopeptides self-assemble into different structures, both local and morphological, that are dependent on both the nature of the Aaa side chains and growth conditions employed. Such alternate peptides substituted with small or polar residues, Ala and Thr, do not yield fibrils; but with ß-branched aliphatic residues, Val and Ile, that could potentially pack with Glu side chains, these oligopeptides do show evidence of ß2-stacking. By contrast, for Leu, with longer side chains, only ß1-stacking is seen while with even larger Phe side chains, either ß-form can be detected separately, depending on preparation conditions. These structures are dependent on high temperature incubation after reducing the pH and in some cases after sonication of initial fibril forms and reincubation. Some of these fibrillar peptides, but not all, show enhanced VCD, which can offer evidence for formation of long, multistrand, often twisted structures. Substitution of Glu with residues having selected side chains yields a variety of morphologies, leading to both ß1- and ß2-structures, that overall suggests two different packing modes for the hydrophobic side chains depending on size and type.

Growth inhibition of Staphylococcus aureus and escherichia coli strains by neutralizing IgY antibodies from ostrich egg yolk.

Ostrich raising around the world have some key factors and farming profit depend largely on information and ability of farmers to rear these animals. Non fertilized eggs from ostriches are discharged in the reproduction season. Staphylococcus aureus and Escherichia coli are microorganisms involved in animal and human diseases. In order to optimize the use of sub products of ostrich raising, non fertilized eggs of four selected birds were utilized for development of polyclonal IgY antibodies. The birds were immunized (200ug/animal) with purified recombinant staphylococcal enterotoxin C (recSEC) and synthetic recRAP, both derived from S. aureus, and recBFPA and recEspB involved in E. coli pathogenicity, diluted in FCA injected in the braquial muscle. Two subsequent immunization steps with 21 days intervals were repeated in 0,85% saline in FIA. Blood and eggs samples were collected before and after immunization steps. Egg yolk immunoglobulins were purified by precipitation with 19% sodium sulfate and 20% ammonium sulphate methodologies. Purified IgY 50µL aliquots were incubated in 850µL BHI broth containing 50µL inoculums of five strains of S. aureus and five strains of E.coli during four hours at 37°C. Growth inhibition was evaluated followed by photometry reading (DO550nm). Egg yolk IgY preparation from hiperimmunized birds contained antibodies that inhibited significantly (p<0,05) growth of strains tested. Potential use of ostrich IgY polyclonal antibodies as a diagnostic and therapeutic tool is proposed for diseased animals.

Lipidomic comparison of 2D and 3D colon cancer cell culture models.

Altered lipid metabolism is one of the hallmarks of cancer. Cellular proliferation and de novo synthesis of lipids are related to cancer progression. In this study, we evaluated the lipidomic profile of two-dimensional (2D) monolayer and multicellular tumor spheroids from the HCT 116 colon carcinoma cell line. We utilized serial trypsinization on the spheroid samples to generate three cellular populations representing the proliferative, quiescent, and necrotic regions of the spheroid. This analysis enabled a comprehensive identification and quantification of lipids produced in each of the spheroid layer and 2D cultures. We show that lipid subclasses associated with lipid droplets form in oxygen-restricted and acidic regions of spheroids and are produced at higher levels than in 2D cultures. Additionally, sphingolipid production, which is implicated in cell death and survival pathways, is higher in spheroids relative to 2D cells. Finally, we show that increased numbers of lipids composed of polyunsaturated fatty acids (PUFAs) are produced in the quiescent and necrotic regions of the spheroid. The lipidomic signature for each region and cell culture type highlights the importance of understanding the spatial aspects of cancer biology. These results provide additional lipid biomarkers in colon cancer cells that can be further studied to target pivotal lipid production pathways.

Electroblotting through Enzymatic Membranes to Enhance Molecular Tissue Imaging.

MALDI-TOF mass spectrometry imaging (MSI) is a powerful tool for studying biomolecule localization in tissue. Protein distributions in tissue provide important histological information; however, large proteins exhibit a high limit of detection in MALDI-MS when compared to their corresponding smaller proteolytic peptides. As a result, several techniques have emerged to digest proteins into more detectable peptides for imaging. Digestion is typically accomplished through trypsin deposition on the tissue, but this technique increases the complexity of the tissue microenvironment, which can limit the number of detectable species. This proof-of-principle study explores tryptic tissue digestion during electroblotting through a trypsin-containing membrane. This approach actively extracts and enzymatically digests proteins from mouse brain tissue sections while simultaneously reducing the complexity of the tissue microenvironment (compared to trypsin deposition on the surface) to obtain an increased number of detectable peptide fragments. The method does not greatly compromise spatial location or require expensive devices to uniformly deposit trypsin on tissue. Using electrodigestion through membranes, we detected and tentatively identified several tryptic peptides that were not observed after on-tissue digestion. Moreover, the use of pepsin rather than trypsin in digestion membranes allows extraction and digestion at low pH to detect peptides from a complementary subset of tissue proteins. Future studies will aim to further improve the method, including changing the substrate membrane to increase spatial resolution and the number of detected peptides.

Nematicidal activity of silver nanoparticles from the fungus Duddingtonia flagrans.

BACKGROUND: Helminth parasites cause morbidity and mortality in both humans and animals. Most anthelmintic drugs used in the treatment of parasitic nematode infections act on target proteins or regulate the electrical activity of neurons and muscles. In this way, it can lead to paralysis, starvation, immune attack, and expulsion of the worm. However, current anthelmintics have some limitations that include a limited spectrum of activity across species and the threat of drug resistance, which highlights the need for new drugs for human and veterinary medicine. PURPOSE: Present study has been conducted to determine the anthelmintic activity of silver nanoparticles (AgNPs) synthesized from the extract of nematophagous fungus, Duddingtonia flagrans, on the infecting larvae of Ancylostoma caninum (L3). METHODS: The nanoparticles were characterized by visual, ultraviolet, Fourier-transform infrared spectroscopy, transmission electron microscopy (TEM) analysis, and X-ray diffraction. The in vitro study was based on experiments to inhibit the motility of infective larvae (L3), and the ultrastructural analysis of the nematode was performed by images obtained by TEM. RESULTS: The XRD studies revealed the crystalline nature of the nanoparticles, and FTIR results implied that AgNPs were successfully synthesized and capped with compounds present in the extract. The results showed that the green synthesis of AgNPs exhibited nematicidal activity, being the only ones capable of penetrating the cuticle of the larvae, causing changes in the tegmentum, and consequently, the death of the nematode. CONCLUSION: The extract of the fungus D. flagrans is able to synthesize AgNP and these have a nematicidal action.

Rhabditis spp., in the Espírito Santo, State of Brazil and evaluation of biological control.

The objectives of this study were to describe occurrences of Rhabditis spp. causing parasitic otitis in dairy cattle of Gir breed in the state of Espírito Santo, southeastern Brazil, and to evaluate the biological control of this nematode using the nematophagous fungi Duddingtonia flagrans (AC001) and Monacrosporium thaumasium (NF34). After nematode detection and collection, three groups were formed: two groups that were treated, respectively, with the fungal isolates; and a control group, without fungus. The treatments were as follows: (a) Petri dishes containing the culture medium 2% water agar (WA) + 250 nematodes + AC001; (b) Petri dishes containing 2% WA + 250 nematodes + NF34; and (c) Petri dishes containing only 2% WA + 250 nematodes. After seven days at 27 °C the treatments with fungi were able to capture and destroy the nematodes, with percentages of 82.0% (AC001) and 39.0% (NF34) in relation to the control group. The results demonstrate the occurrence of Rhabditis spp. after animals physical examination and that there was efficacy of the in vitro predatory activity of both fungal isolates. Thus, these results are important because they can assist in future in vivo control of this nematode in cattle.

Combined use of ivermectin, dimethyl sulfoxide, mineral oil and nematophagous fungi to control Rhabditis spp.

Rhabditis spp., is a nematode known to cause otitis externa, an infection difficult to control, in cattle reared within tropical regions. The objective of this study was to assess the combined use of ivermectin 1%, dimethyl sulfoxide 1% and mineral oil 100% containing nematophagous fungi of both Duddingtonia flagrans (AC001) and Monacrosporium thaumasium (NF34) species to control in vitro Rhabditis spp. Thus, 12 experimental groups were designed with eight replicates each: G1 (nematodes + AC001); G2 (nematodes + NF34); G3 (nematodes + ivermectin 1%/positive control); G4 (nematodes + dimethyl sulfoxide 1%/positive control); G5 (nematodes + mineral oil 100%/positive control); G6 (nematodes + AC001 + ivermectin 1%); G7 (nematodes + NF34 + ivermectin 1%); G8 (nematodes + AC001 + mineral oil 100%); G9 (nematodes + NF34 + mineral oil 100%); G10 (nematodes + AC001 + dimethyl sulfoxide 1%); G11 (nematode + NF34 + dimethyl sulfoxide 1%); G12 (nematode + distilled water/negative control). The results demonstrated that all experimentally treated groups differed statistically (p < 0.01) from the control group. In the present study, the use of dimethyl sulfoxide 1% and mineral oil 100% in conjunction with conidia fungi portrayed noteworthy outcomes, which represents a future premise for the combined use of nematophagous fungi within these vehicles in both controlling Rhabditis spp.

ß2-Type Amyloidlike Fibrils of Poly-l-glutamic Acid Convert into Long, Highly Ordered Helices upon Dissolution in Dimethyl Sulfoxide.

Replacing water with dimethyl sulfoxide (DMSO) completely reshapes the free-energy landscapes of solvated proteins. In DMSO, a powerful hydrogen-bond (HB) acceptor, formation of HBs between backbone NH groups and solvent is favored over HBs involving protein's carbonyl groups. This entails a profound structural disruption of globular proteins and proteinaceous aggregates (e.g., amyloid fibrils) upon transfer to DMSO. Here, we investigate an unusual DMSO-induced conformational transition of ß2-amyloid fibrils from poly-l-glutamic acid (PLGA). The infrared spectra of ß2-PLGA dissolved in DMSO lack the typical features associated with disordered conformation that are observed when amyloid fibrils from other proteins are dispersed in DMSO. Instead, the frequency and unusual narrowness of the amide I band imply the presence of highly ordered helical structures, which is supported by complementary methods, including vibrational circular dichroism and Raman optical activity. We argue that the conformation most consistent with the spectroscopic data is that of a PLGA chain essentially lacking nonhelical segments such as bends that would provide DMSO acceptors with direct access to the backbone. A structural study of DMSO-dissolved ß2-PLGA by synchrotron small-angle X-ray scattering reveals the presence of long uninterrupted helices lending direct support to this hypothesis. Our study highlights the dramatic effects that solvation may have on conformational transitions of large polypeptide assemblies.

Colonization and destruction of ants of the genus Camponotus sp. (Hymenoptera: Formicidae) in vitro by the fungus Pochonia chlamydosporia in the southeast region of Brazil.

The objective of this study was to evaluate, in vitro, the colonization and destruction of ants of the genus Camponotus sp. by the ovicidal fungus Pochonia chlamydosporia (VC4 isolate), in the southeast region of Brazil. The insects used in the experiment were worker ants of the genus Camponotus sp., collected periodically in the environment and immediately transported to the laboratory in test tubes. Then, VC4 growth was promoted in 2% chitin agar medium (2% WQ) to obtain a fungal solution containing conidia and/or chlamydospores. Two experimental groups were formed. Treated group consisted of Petri dishes containing 2% agar-water culture medium (2% WA) with nine live insects and 20 µL of fungal solution at the concentration of 15,000 conidia/chlamydospores. Control group consisted of Petri dishes containing 2% WA culture medium and nine live insects. The dishes in the treated and control groups were incubated in BOD at 25 ± 1 °C and 80 ± 10% relative humidity for 4 days. After 4 days, it was observed that the VC4 had grown, colonized, and caused the destruction of the ants. The fungus P. chlamydosporia was efficient at colonizing and destroying the urban ants collected on an experimental basis. Thus, it could open up new ways to reduce the use of chemical compounds in the future, decreasing health and environmental problems.

Ocorrência de Babesia sp, Ehrlichia canis e Hepatozoon canis em cães domiciliados, em dois municípios do estado do Espírito Santo - Brasil / Occurrence of Babesia sp, Ehrlichia canis and Hepatozoon canis in domiciled dogs in two municipalities of the state of Espírito Santo - Brazil / Ocurrencia de Babesia sp, Ehrlichia canis y Hepatozoon canis em perros domiciliados em dos municipios del estado de Espírito Santo - Brasil

As doenças transmitidas por carrapatos são afecções de grande importância na clínica médica de pequenos animais, devido à alta casuística e ampla distribuição vetorial no território brasileiro. Os principais agentes responsáveis pelas infecções em cães são Babesia sp., Ehrlichia canis e Hepatozoon canis. Os animais infectados são assintomáticos ou apresentam sinais clínicos inespecíficos, sendo necessário a utilização de testes diagnósticos para definição do agente etiológico, e diagnóstico seguro. O objetivo do presente estudo foi determinar a ocorrência desses micro-organismos em cães naturalmente infectados, domiciliados nos municípios de Vila Velha e Anchieta, Espírito Santo, utilizando diferentes testes de detecção: Reação em cadeia polimerase (PCR), sorologia para detecção de anticorpos anti Ehrlichia canis e pesquisa de hematozoários em esfregaço sanguíneo. Foram analisadas 65 amostras de sangue obtidas por venopunção de veia cefálica de cães. No teste de PCR, 4,62% dos animais foram positivos para Babesia vogeli e 1,54% para Ehrlichia canis sendo os resultados para Hepatozoon canis negativos. No teste sorológico para E. canis 90,77% dos animais foram positivos para a presença de anticorpos, e na pesquisa em lâminas de esfregaço sanguíneo 3,02% apresentavam outros hemoparasitas. Os resultados indicam a dispersão desses hemoparasitas na população canina da região de estudo, entretanto com baixa ocorrência. O teste de PCR demonstrou-se como o mais sensível no qual Babesia vogeli foi o agente mais observado.(AU)

Tick-borne diseases are diseases of great importance in the medical practice of small animals, due to the high casuistry and wide vectorial distribution in the Brazilian territory. The main agents responsible for infections in dogs are Babesia sp., Ehrlichia canis and Hepatozoon canis. Infected animals are asymptomatic or present nonspecific clinical signs, requiring the use of diagnostic tests to define the etiologic agent, and safe diagnosis. The objective of the present study was to determine the occurrence of these microorganisms in naturally infected dogs domiciled in the municipalities of Vila Velha and Anchieta, Espírito Santo, using different detection tests: polymerase chain reaction (PCR), serology to detect antibodies against Ehrlichia canis and research of hematozoa in blood smears. Sixty-five blood samples obtained by venipuncture of the cephalic vein of dogs were analyzed. In the PCR test, 4.62% of the animals were positive for Babesia vogeli and 1.54% for Ehrlichia canis, and the results for Hepatozoon canis were negative. In the serological test for E. canis, 90.77% of the animals were positive for the presence of antibodies, and in the research in blood smear slides, 3.02% presented other hemoparasites. The results indicate the dispersion of these hemoparasites in the canine population of the study region, however with low occurrence. The PCR test proved to be the most sensitive, in which Babesia vogeli was the most observed agent.(AU)

Las enfermedades transmitidas por garrapatas son enfermedades de gran importancia en la práctica médica de los pequeños animales, debido a la alta casuística y amplia distribución vectorial en el territorio brasileño. Los principales agentes responsables de las infecciones en los perros son Babesia sp., Ehrlichia canis y Hepatozoon canis. Los animales infectados son asintomáticos o presentan signos clínicos inespecíficos, siendo necesario el uso de pruebas diagnósticas para la definición del agente etiológico, y el diagnóstico seguro. El objetivo del presente estudio fue determinar la ocurrencia de estos microorganismos en perros infectados naturalmente, domiciliados en los municipios de Vila Velha y Anchieta, Espírito Santo, utilizando diferentes pruebas de detección: reacción en cadena de la polimerasa (PCR), serología para detectar anticuerpos anti Ehrlichia canis e investigación de hematozoos en frotis de sangre. Se analizaron sesenta y cinco muestras de sangre obtenidas por venopunción de la vena cefálica de los perros. En la prueba PCR, el 4,62% de los animales fueron positivos para Babesia vogeli y el 1,54% para Ehrlichia canis, y los resultados para Hepatozoon canis fueron negativos. En la prueba serológica para E. canis, el 90,77% de los animales fueron positivos a la presencia de anticuerpos, y en la investigación en láminas de frotis de sangre el 3,02% presentaron otros hemoparásitos. Los resultados indican la dispersión de estos hemoparásitos en la población canina de la región de estudio, aunque con una baja presencia. La prueba PCR resultó ser la más sensible, en la que Babesia vogeli fue el agente más observado.(AU)