Canine Babesiosis: Where Do We Stand? Petra, Bilić; Josipa, Kuleš; Renata, Barić Rafaj ...
Acta veterinaria,
06/2018, Volume:
68, Issue:
2
Journal Article
Peer reviewed
Open access
Canine babesiosis is a tick-borne disease caused by protozoal haemoparasites of different
species. Babesiosis is one of the most important globally extended and quickly spreading tick-borne ...infections of dogs. This comprehensive review gives an in-depth overview of
species currently identified in dogs together with relevant vector tick species and their geographical distribution, life cycle and transmission of parasite. The main mechanisms in the pathogenesis of babesiosis are described and elucidated by recent literature overview. As
infection causes a disease with very variable clinical manifestations, special attention is given to clinical signs, laboratory features and clinicopathological findings. The diagnosis of canine babesiosis by microscopy, serological and molecular methods is reviewed, together with recent advances in mass spectrometry based assays. Accurate detection and species recognition are important for the selection of the appropriate therapy, monitoring and prediction of the outcome of the disease. Finally, guidelines for the treatment and prevention of canine babesiosis are given.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Over the past two decades, library-based display technologies have been staggeringly optimized since their appearance in order to mimic the process of natural molecular evolution. Display ...technologies are essential for the isolation of specific high-affinity binding molecules (proteins, polypeptides, nucleic acids and others) for diagnostic and therapeutic applications in cancer, infectious diseases, autoimmune, neurodegenerative, inflammatory pathologies
etc.
Applications extend to other fields such as antibody and enzyme engineering, cell-free protein synthesis and the discovery of protein-protein interactions. Phage display technology is the most established of these methods but more recent fully
in vitro
alternatives, such as ribosome display, mRNA display, cis-activity based (CIS) display and covalent antibody display (CAD), as well as aptamer display and
in vitro
compartmentalization, offer advantages over phage in library size, speed and the display of unnatural amino acids and nucleotides. Altogether, they have produced several molecules currently approved or in diverse stages of clinical or preclinical testing and have provided researchers with tools to address some of the disadvantages of peptides and nucleotides such as their low affinity, low stability, high immunogenicity and difficulty to cross membranes. In this review we assess the fundamental technological features and point out some recent advances and applications of display technologies.
Display technology is a collection of methods for creating libraries of modularly coded biomolecules and submit them to directed evolution to obtain arrays of desired properties with multiple applications.
Babesiosis is a disease of significant medically and veterinary importance with worldwide distribution. It is caused by intra-erythrocyte protozoal parasites, with Babesia rossi causing the most ...severe clinical signs of all the large Babesia parasites infecting dogs. The disease can be clinically classified into uncomplicated and complicated forms with a wide range of clinical presentations from a mild, subclinical illness to complicated forms and death. The aim of this study was to assess serum proteomic profiles from dogs with babesiosis and healthy dogs using a label-based proteomics approach. Altogether 32 dogs naturally infected with B. rossi (subdivided into 18 uncomplicated cases and 14 complicated cases of babesiosis) and 20 healthy dogs were included. There were 78 proteins with significantly different abundances between the three groups of dogs. Elucidation of proteins and pathways involved in canine babesiosis caused by B. rossi have revealed key differences associated with haemostasis, innate immune system, lipid metabolism and inflammation. Shotgun proteomic profiling allowed identification of potential serum biomarkers for differentiation of disease severity in canine babesiosis caused by B. rossi. These findings may be applicable to the study of host-parasite interactions and the development of novel therapeutic targets.
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Neospora caninum
is one of the most prevalent Apicomplexa parasites that causes abortion in cattle, as it infects dogs as its definitive host, causing subclinical disease or active neosporosis, ...marked by meningoencephalitis, and myopathies with muscle and neuromuscular signs of disease. This study aimed to evaluate the acute phase protein response in dogs seropositive and seronegative for
N. caninum
. Serum samples of 72 dogs were tested by an immunofluorescence antibody test using
N. caninum
NC-1 strain, and the study population was divided into four groups: symptomatic — muscular and/or neuromuscular signs — and seropositive (
n
= 16); symptomatic and seronegative (
n
= 9); asymptomatic and seropositive (
n
= 34); and asymptomatic and seronegative (
n
= 13). C-reactive protein (CRP) was measured via immunoturbidimetric assay and serum haptoglobin (Hp) via hemoglobin-binding capacity assay. In the symptomatic groups, seropositive dogs had higher levels of Hp, but not CRP, while seronegative dogs had higher CRP levels. There was no difference in CRP concentration in asymptomatic dogs. Dogs with neuromuscular signs had higher concentrations for Hp in the group seropositive. Hp concentration did not differ between dogs seropositive and seronegative dogs for each group. Serum Hp and CRP could not sufficiently alone flag subclinical infections. Measurement of CRP and Hp concentrations could be clinically valuable to the diagnosis of neurological diseases, and their relative change may indicate the stage of the infection, although their sole use is not able to support the diagnosis of canine neosporosis. Further studies are encouraged to evaluate the specific dynamics of acute phase proteins in canine neosporosis.
Graphical abstract
Serum samples of 72 dogs were tested by an immunofluorescence antibody test using
N. caninum
NC-1 strain, and the study population was divided into four groups: (1) dogs with muscular and/or neuromuscular signs and seropositive for
N. caninum
; (2) dogs with muscular and/or neuromuscular signs and seronegative for
N. caninum
; (3) dogs seropositive for
N. caninum
with no neuromuscular signs; and (4) healthy dogs and seronegative for
N. caninum
. The study evaluated if
N. caninum
infection could have pathophysiological changes activating the acute phase response and an increase in the concentration of acute phase proteins in serum.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Exosomes mediate near and long-distance intercellular communication by transferring their molecular cargo to recipient cells, altering their biological response. Milk exosomes (MEx) are internalized ...by immune cells and exert immunomodulatory functions in vitro. Porcine MEx can accumulate in the small intestine, rich in macrophages. No information is available on the immunomodulatory ability of porcine MEx on porcine monocytes, which are known precursors of gut macrophages. Therefore, this study aims at (1) assessing the in vitro uptake of porcine MEx by porcine monocytes (CD14+), and (2) evaluating the in vitro impact of porcine MEx on porcine monocytes immune functions. MEx were purified by ultracentrifugation and size exclusion chromatography. The monocytes' internalization of PKH26-labeled MEx was examined using fluorescence microscopy. Monocytes were incubated with increasing exosome concentrations and their apoptosis and viability were measured. Lastly, the ability of MEx to modulate the cells' immune activities was evaluated by measuring monocytes' phagocytosis, the capacity of killing bacteria, chemotaxis, and reactive oxygen species (ROS) production. MEx were internalized by porcine monocytes in vitro. They also decreased their chemotaxis and phagocytosis, and increased ROS production. Altogether, this study provides insights into the role that MEx might play in pigs' immunity by demonstrating that MEx are internalized by porcine monocytes in vitro and exert immunomodulatory effects on inflammatory functions.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Canine myxomatous mitral valve disease (MMVD) is similar to Barlow's form of MMVD in humans. These valvulopathies are complex, with varying speeds of progression. We hypothesized that the relative ...abundances of serum proteins would help identify the consecutive MMVD stages and discover new disease pathways on a systemic level. To identify distinction-contributing protein panels for disease onset and progression, we compared the proteomic profiles of serum from healthy dogs and dogs with different stages of naturally occurring MMVD. Dogs were divided into experimental groups on the basis of the left-atrium-to-aorta ratio and normalized left ventricular internal dimension in diastole values. Serum was collected from healthy (N = 12) dogs, dogs diagnosed with MMVD in stages B1 (N = 13) and B2 (N = 12) (asymptomatic), and dogs diagnosed with MMVD in chronic stage C (N = 13) (symptomatic). Serum biochemistry and selected ELISAs (galectin-3, suppression of tumorigenicity, and asymmetric dimethylarginine) were performed. Liquid chromatography-mass spectrometry (LC-MS), tandem mass tag (TMT) quantitative proteomics, and statistical and bioinformatics analysis were employed. Most of the 21 serum proteins with significantly different abundances between experimental groups (
< 0.05, FDR ˂ 0.05) were classified as matrix metalloproteinases, protease inhibitors, scaffold/adaptor proteins, complement components, anticoagulants, cytokine, and chaperone. LC-MS TMT proteomics results obtained for haptoglobin, clusterin, and peptidase D were further validated analytically. Canine MMVD stages, including, for the first time, asymptomatic B1 and B2 stages, were successfully distinguished in dogs with the disease and healthy dogs on the basis of the relative abundances of a panel of specific serum proteins. Most proteins with significantly different abundances were involved in immune and inflammatory pathways. Their role in structural remodeling and progression of canine MMVD must be further investigated. Further research is needed to confirm the resemblance/difference with human MMVD. Proteomics data are available via ProteomeXchange with the unique dataset identifier PXD038475.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Abstract
The coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV 2), is usually associated with a wide variety of clinical presentations from ...asymptomatic to severe cases. The use of saliva as a diagnostic and monitoring fluid has gained importance since it can be used to investigate the immune response and to direct quantification of antibodies against COVID-19. Additionally, the use of proteomics in saliva has allowed to increase our understanding of the underlying pathophysiology of diseases, bringing new perspectives on diagnostics, monitoring, and treatment. In this work, we compared the salivary proteome of 10 patients with COVID-19, (five patients with mild and five patients with severe COVID-19) and ten control healthy patients. Through the application of proteomics, we have identified 30 proteins whose abundance levels differed between the COVID-19 groups and the control group. Two of these proteins (TGM3 and carbonic anhydrase-CA6) were validated by the measurement of gGT and TEA respectively, in 98 additional saliva samples separated into two groups: (1) COVID-19 group, integrated by 66 patients who tested positive for COVID-19 (2) control group, composed of 32 healthy individuals who did not show any sign of disease for at least four weeks and were negative for COVID-19 in RT-PCR. In the proteomic study there were observed upregulations in CAZA1, ACTN4, and ANXA4, which are proteins related to the protective response against the virus disturbance, and the upregulation of TGM3, that is correlated to the oxidative damage in pulmonary tissue. We also showed the downregulation in cystatins and CA6 that can be involved in the sensory response to stimulus and possibly related to the presence of anosmia and dysgeusia during the COVID-19. Additionally, the presence of FGB in patients with severe COVID-19 but not in mild COVID-19 patients could indicate a higher viral aggregation and activation in these cases. In conclusion, the salivary proteome in patients with COVID-19 showed changes in proteins related to the protective response to viral infection, and the altered sensory taste perception that occur during the disease. Moreover, gGT and TEA could be potential biomarkers of respiratory complications that can occurs during COVID 19 although further larger studies should be made to corroborate this.
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Canine babesiosis caused by the intraerythrocytic protozoan parasite Babesia canis is a tick-borne disease characterized by a host response that involves both cellular and humoral immunity. This ...study focuses on the secretion of cytokines Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF), Keratinocyte Chemotactic-like (KC-like), Interleukins (IL)-2, IL-7, IL-8, IL-10, IL-15, IL-18 and Monocyte Chemotactic Protein-1 (MCP-1) in babesiosis caused by Babesia canis upon treatment with Imizol®. We assessed time dependent changes in cytokine levels and tested whether these changes correlate with pathogenesis of the disease. Sixteen healthy dogs and 31 dogs infected with Babesia canis, of which 18 showed complications, were treated with Imizol®. One dog died during the study (3.2%). Longitudinal study was perfomed by monitoring dogs at the first day of presentation (day 1) and 6 days later (day 7). Our results show that higher MCP-1 levels on day 1 are positively associated with the occurrence of complications, (complicated vs. uncomplicated; p = 0.00016). A similar pattern was observed for KC-like on day 1 (p = 0.0326) and day 7 (p = 0.044). Moreover, babesiosis caused by B. canis produced a steady increase in IL-8 levels with a moderate to strong negative correlation with erythrocyte counts and hematocrit in uncomplicated diseased dogs only (Spearman's rank correlation coefficient rs = -0.582 and rs = -0.598 respectively). Like for MCP-1, KC-like levels also differed in complicated and uncomplicated diseased dogs on day 1 (p = 0.03236) and day 7 (p = 0.044). Furthermore, KC-like levels were strongly correlated with IL-8 levels (rs = 0.663-0.7) and non-segmented neutrophil counts (rs = 0.572-0.732) in both diseased groups. Analysis of ROC suggests the use of serum levels of MCP-1 and IL-7 as predictors of the occurrence of complications with an AUC of 0.906 and 0.896 respectively and linear combinations of MCP-1, KC-Like, IL-7 and GM-CSF with values up to AUC = 0.983. Cytokine cluster analysis presented in this study can contribute to a better understanding of the pathogenesis of babesiosis and serve as a prognostic tool for the early detection of cases with highest likelihood of developing complications. Overall, our studies show that infection by B. canis elicits a cytokine pattern that is distinct from that observed with B. rossi, and that some of the inflammatory mediators can be useful to predict complications. Our results also suggest targets for the development of novel therapeutic strategies in babesiosis caused by B. canis.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Sepsis is a systemic inflammatory response triggered by an infectious agent and is recognized by the World Health Organization as a global concern, since it is one of the major causes of severe ...illness in humans and animals. The study of the changes that can occur in saliva and serum in sepsis can contribute to a better understanding of the pathophysiological mechanisms involved in the process and also to discover potential biomarkers that can help in its diagnosis and monitoring. The objective of this study was to characterize the changes that occur in the salivary and serum proteome of pigs with experimentally-induced sepsis. The study included five pigs with sepsis induced by LPS administration and five pigs with non-septic inflammation induced by turpentine for comparative purposes. In saliva, there were eighteen salivary proteins differentially expressed in the sepsis condition and nine in non-septic inflammation. Among these, significant increments in aldolase A and serpin B12 only occurred in the sepsis model. Changes in aldolase A were validated in a larger population of pigs with sepsis due to Streptococcus suis infection. In serum, there were 30 proteins differentially expressed in sepsis group and 26 proteins in the non-septic group, and most of the proteins that changed in both groups were related to non-specific inflammation. In the saliva of the septic animals there were some specific pathways activated, such as the organonitrogen compound metabolic process and lipid transport, whereas, in the serum, one of the main activated pathways was the regulation of protein secretion. Overall, saliva and serum showed different proteome variations in response to septic inflammation and could provide complementary information about the pathophysiological mechanisms occurring in this condition. Additionally, salivary aldolase A could be a potential biomarker of sepsis in pigs that should be confirmed in a larger population.
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Babesiosis, the disease caused by tick-borne hematozoan parasites of the genus
Babesia, is particularly common in dogs, and is caused by several “large” species of
Babesia, as well as by an ...increasing number of “small” species of
Babesia, some of which appear to be more closely related to members of the genus
Theileria. In this work, blood samples were collected from 848 randomly selected, asymptomatic dogs and from 81 symptomatic dogs, microscopically positive for
Babesia, and characterised by PCR and sequence analysis of a fragment of the ssrRNA gene. A prevalence of 3.42% (29 of 848) was found in asymptomatic dogs and sequence analysis revealed the presence of
Babesia canis canis in 20 dogs (69%),
Babesia gibsoni in six dogs (21%),
Babesia canis vogeli in two dogs (7%) and
Theileria annae in one dog (3%). In the group of symptomatic dogs, which were all positive by PCR,
B. canis canis was the predominant species (78 dogs, or 96%), followed by single infections with
B. canis vogeli,
Babesia caballi and
Theileria equi. Our study has confirmed that dogs are infected with a wide range of both large and small piroplasm species and subspecies, including
B. caballi and
T. equi, two parasites usually found in horses. The detection of the pathogenic species
B. canis canis and
B. gibsoni in asymptomatic dogs indicates that the relationship between parasite species/subspecies and clinical signs of infection in dogs deserves further investigation. Finally, the identities of the tick vectors transmitting
T. annae and
B. caballi remain to be elucidated.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
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