Age, sex and breed effect on laboratory parameters in natural Babesia canis infection.

We tested the hypothesis that age, breed, and sex are related to hematology, biochemistry, acute phase proteins (APPs), seroreactivity and level of parasitemia in dogs with an acute phase response (APR) due to Babesia canis infection. The study enrolled 61 privately owned dogs that naturally acquired B. canis infection. Groups were formed according to the age: young dogs less than one year, and adult dogs more than one year old. Moreover, the group of males was compared to females and purebred to mixed breed dogs. Seroreactivity was tested with immunofluorescence antibody test, level of parasitemia with real-time polymerase chain reaction (real-time PCR), hematology, and biochemistry with automatic analyzers, serum amyloid A with enzyme-linked immunosorbent assay, fibrinogen with heat precipitation and ceruloplasmin and paraoxonase-1 with manual spectrophotometric methods. For protein separation agarose gel electrophoresis was used. The main changes in the whole population of B. canis-infected dogs were fever, pancytopenia, and change in APPs level. One-third of young, and 96% of adult dogs were seropositive (P < 0.001). The level of parasitemia was higher in the young dogs (P < 0.001). Erythroid lineage parameters (P < 0.01), and leukocytes (P < 0.05) were lower in the young, when compared to the adult dogs. Young dogs had lower total globulins (P < 0.001), ß- and γ-globulins (P < 0.001), and higher α-globulins (P = 0.022) than adult dogs. Young dogs had higher concentrations of phosphate (P = 0.003) and cholesterol (P < 0.001) and lower amylase (P = 0.014) and lipase activity (P = 0.020) than adult ones. Male dogs had lower neutrophil count than females (P = 0.035), and purebred dogs had more band neutrophils than mixed breed dogs (P = 0.004). In conclusion, dogs with natural Babesia canis infection at a young age have more severe anemia and APR including leukopenia than adults. Male and purebred dogs might also have more severe APR than females and mix-breeds, as they have more pronounced changes related to the myeloid lineage.

Consequences of Transport Conditions on the Welfare of Slaughter Pigs with Different Health Status and RYR-1 Genotype.

This study assessed the influence of transport conditions on welfare indicators of slaughter pigs with different health status and RYR-1 genotype. The group of pigs, predominantly consisting of Nn (56.67%) and subclinically diseased (60.00%) individuals, that were exposed to short transportation (<30 min) at high loading density (~235 kg/m2) had the highest slipping (p < 0.0001), falling (p = 0.0009), turning back (p < 0.0001), reluctance to move (p < 0.0001), panting (p < 0.0001) and shivering (p < 0.0001) frequencies at unloading. Subclinically diseased Nn pigs subjected to short transportation (<30 min) and high loading density (~235 kg/m2) had the highest lactate (p < 0.0001 and p < 0.0001), glucose (p = 0.0450 and p = 0.0002), CK (p < 0.0001 and p = 0.0010), LDH (p < 0.0001 and p = 0.0484), AST (p = 0.0208 and p = 0.0170), ALT (p = 0.0500 and p = 0.00081), ceruloplasmin (p = 0.0334 and p < 0.0001) and MDA (p = 0.0048 and p < 0.0001) concentrations, but the lowest sodium (p < 0.0001 and p < 0.0001), chloride (p = 0.0001 and p = 0.0432), albumin (p < 0.0090 and p < 0.0001), PON-1 (p = 0.0122 and p = 0.0500) and GSH (p = 0.0042 and p = 0.0340) levels, respectively. In the group consisting of of stress-resistant (100%) and predominantly healthy (60.00%) pigs subjected to short transportation (<30 min) at high loading density (~235 kg/m2), none of the individuals showed irregular behavioural reactions during unloading. Healthy NN pigs that underwent short transportation (<30 min) at high loading density (~235 kg/m2) had the lowest lactate (p < 0.0001 and p < 0.0001), glucose (p = 0.0450 and p = 0.0002), CK (p < 0.0001 and p = 0.0010), LDH (p < 0.0001 and p = 0.0484) and ceruloplasmin (p = 0.0334 and p < 0.0001) levels, but the highest sodium (p < 0.0001 and p < 0.0001) and chloride (p = 0.0001 and p = 0.0432) concentrations, respectively. In conclusion, the most compromised welfare was recorded in subclinically diseased Nn pigs exposed to short transportation (<30 min) and high loading density (~235 kg/m2), while under the same conditions, the welfare of healthy NN pigs was not compromised. Therefore, stress-carrier pigs with subclinical pathological lesions should not be considered fit for transportation, indicating that the health status and genotype are the key factors for optimising pig welfare.

Serum oxidant-antioxidant status and butyrylcholinesterase activity in dogs with idiopathic epilepsy - A pilot study.

Oxidative stress plays an important role in pathogenesis of idiopathic epilepsy (IE). Although IE is the most common neurological condition, oxidant-antioxidant status in epileptic dogs is still unknown. The aim of this study is to evaluate the serum oxidant-antioxidant status in dogs with newly diagnosed IE. The status in 15 dogs with IE and 15 healthy dogs is estimated through spectrophotometric determination of two oxidant markers: advanced oxidation protein products-albumin index (AOPP) and thiobarbituric acid reactive substances (TBARS); and three antioxidant markers: total thiols (R-SH) level, glutathione (GSH) level, and paraoxonase-1 (PON-1) activity. Also, butyrylcholinesterase (BChE) activity is assessed in both groups of dogs. Higher AOPP is observed in the dogs with newly diagnosed IE, while TBARS level shows no difference when compared to the healthy dogs. In contrast, lower levels of antioxidants (R-SH, GSH, and PON-1) and BChE activity are found in the dogs with IE. No significant differences are observed in the oxidant and antioxidant markers and BChE activity across the investigated IE cases with focal and generalized seizures. Our findings provide evidence that dogs with IE are characterized by an impaired serum oxidant-antioxidant balance and lower BChE activity, which may contribute to a better understanding of IE pathogenesis.

Seasonal differences in the intensity of acute phase response in dogs infected with Babesia canis.

The highest number of acute Babesia canis cases in dogs is recorded over the February-May (Feb-May) period, which also represents the optimal climate conditions for tick activity in Belgrade, Serbia. A possibility that the acute phase response is more intense in dogs developing the disease in the Feb-May period compared with the response in other time periods of the year was tested. A total of 63 client-owned dogs with acute B. canis infection were enrolled and the routine hematology and biochemistry parameters-serum amyloid A (SAA), IgG against B. canis, level of parasitemia, ceruloplasmin (CER), paraoxonase-1 (PON-1), and fibrinogen-were measured. Acute phase indexes (API) were calculated as (SAA×CER)/(Iron×PON-1) and (SAA×CER)/(Albumin×Iron). Statistics included Kruskal-Wallis test and logistic regression analysis. The results showed that in the Feb-May period, the following parameters were lower: creatinine, albumin, iron, and level of parasitemia. Furthermore, increased API values were more probable in the Feb-May than in the other periods. Together, higher acute phase response intensity and presumptive hemodilution in the Feb-May period indicate a more acute course of B. canis infection than in other time periods of the year.

Low serum levels of promatrix metalloproteinase-2 and -9 occur during acute Babesia canis infection in dogs.

Inflammation is a hallmark of the acute Babesia canis infection. Promatrix metalloproteinase (proMMP)-2 and -9 are involved in inflammation, but their levels have not been analyzed in canine babesiosis. We hypothesized that in dogs infected with B. canis, serum proMMP-2 and -9 levels change between presentation and recovery. Degree of the change differs if dogs develop systemic inflammatory response syndrome (SIRS). This study included 24 dogs with an acute B. canis infection, at presentation and after two weeks. We used routine hematology and biochemistry methods, spectrophotometry for the acute-phase proteins, microscopy for parasitemia and zymography for (pro)MMPs. In vitro endothelial cells and leukocyte short-term cultures, and platelet lysates were used to detect specific MMP activity. Statistical analyses included Wilcoxon test for paired samples, Mann-Whitney U test and Spearman's rank correlation. Our results showed that endothelial cells, leukocytes and platelets are the source of proMMP-2 and proMMP-9. Furthermore, both proMMPs were lower at presentation than after recovery (p < 0.001). At presentation, proMMP-9 levels correlated with parasitemia (rho = -0.616, p = 0.009), total leukocyte (rho = 0.704, p < 0.001) and neutrophil counts (rho = 0.741, p < 0.001). Extent of alterations in proMMP-2 levels between presentation and recovery was lower (p = 0.038) in dogs with SIRS than in non-SIRS dogs, while levels of proMMP-9 were comparable between these groups. Our conclusion is that during the acute B. canis infection, low serum levels of proMMP-2 and proMMP-9 at presentation reflect thrombocytopenia and leukopenia. Decreased proMMP-2 level could be associated with SIRS.

Systemic inflammatory response syndrome in dogs naturally infected with Babesia canis: Association with the parasite load and host factors.

The common signs of canine babesiosis caused by an infection with Babesia canis are fever, anorexia, lethargy, pulse alterations, anemia, and occasionally mild icterus. Dogs with these clinical signs can be divided into two groups: those with acute-phase reaction and those with systemic inflammatory response syndrome (SIRS). Factors associated with the occurrence of SIRS in canine babesiosis have not been thoroughly researched. This article outlines a cross-sectional study of 54 client-owned dogs with an acute B. canis infection, and evaluates the differences in age, gender, laboratory findings, parasite load, and seroreactivity against B. canis between the SIRS and the SIRS-free dogs. We have analyzed a complete blood count, serum biochemistry, serum amyloid A, ceruloplasmin, paraoxonase-1, serology, and PCR testing using standard methodologies. The frequency of SIRS among the investigated dogs reached 0.59. Male dogs and those seronegative against B. canis, were more frequent in the SIRS group, whilst age and parasite load could not be associated with the presence of SIRS. Dogs with SIRS had a lower count of total leukocytes, neutrophils, lymphocytes, and monocytes, and a lower concentration of iron and bilirubin compared with SIRS-free dogs. No significant differences in the concentration of acute-phase proteins have been noticed to exist between the groups of dogs. Further, the seronegative dogs had a lower count of lymphocytes and monocytes and a higher parasite load than the seroreactive dogs. Multivariate logistic regression analysis has identified leukopenia (<6 × 109/L) and monocytopenia (<0.2 × 109/L) as independent associates of SIRS in the investigated dogs, thus implying that these routine tests could be used as reliable markers for SIRS.

Evidence of acute phase reaction in asymptomatic dogs naturally infected with Babesia canis.

Asymptomatic outdoor dogs can be carriers of Babesia canis, but data describing the development of an acute phase response (APR) are not available. We hypothesised that these dogs have a moderate APR that could be detected by hematological and biochemical changes. Two groups of Babesia-exposed dogs were represented by nine B. canis PCR-positive and twenty B. canis PCR-negative, seroreactive dogs. The control group consisted of ten Babesia-naïve dogs. Serum amyloid A (SAA), paraoxonase-1 (PON-1), complete blood count, and biochemistry parameters were analysed by standard methodologies. Protein and lipoprotein fractions were separated using agarose gel electrophoresis (GE), and the dominant diameters of lipoproteins were assessed on gradient GE. Results were evaluated using non-parametric tests and the Receiver Operating Characteristic curve. SAA (median 39.0 µg/mL, range 2.2-48.8 µg/mL), total protein (median 74.7 g/L, range 57.1-98.3 g/L) and the dominant diameter of α-lipoproteins (median 13.31 nm, range 12.09-14.17 nm) in B. canis PCR-positive dogs were higher relative to dogs in the control group or dogs that were PCR-negative but seroreactive (p < 0.001 for both groups). Mild to moderate anemia (4/29), thrombocytopenia (7/29), and leukocyte counts that were close to the upper limit of the reference range were encountered in both Babesia-exposed groups. When compared to controls, Babesia-exposed dogs displayed decreased a PON-1 activity and protein GE pattern consistent with low-grade chronic inflammation (p < 0.001 for both groups). Dogs with detectable amounts of B. canis DNA in blood contain increased levels of SAA and total protein along with α-lipoproteins that display an increased diameter relative to those dogs with positive Babesia serology but undetectable levels of B. canis DNA in blood.