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Infectious haemolytic anaemia (IHA) in dogs share similar clinical signs including fever, lethargy, icterus, paleness of mucous membranes and splenomegaly. Postmortal findings are similar and, without additional diagnostic methods, an accurate aetiological diagnosis is difficult to achieve. In order to investigate causes of lethal IHA in Croatian dogs, we performed a retrospective study on archived formalin-fixed, paraffin-embedded tissue blocks (FFPEB) from dogs that died due to haemolytic crisis, using microscopic and molecular diagnostic tools to determine the aetiological cause of disease. Molecular analysis was performed on kidney, lung, myocardium and spleen on FFPEB from all dogs. The originally stated aetiological diagnosis of B. canis or leptospirosis was confirmed in only 53% of the dogs. PCR and sequencing revealed that, in addition to the expected pathogens, B. canis and Leptospira interrogans, the presence of previously undiagnosed "new" pathogens causing anaemia including Candidatus Neoehrlichia mikurensis and Anaplasma phagocytophilum. Furthermore, Theileria capreoli was detected for the first time in a dog with postmortal descriptions of lesions. Intensive extravascular hemolysis was noticeable as jaundice of the mucosa, subcutis and fat tissue, green or yellow discoloration of renal parenchyma caused by bilirubin excretion in the renal tubules and bile accumulation within the liver in 90% of the dogs. This work highlights the value of molecular diagnostics to complement traditional ante-mortem and post-mortem diagnostic protocols for the aetiological diagnosis of pathogens associated with IHA.

Background Classification of Babesia parasites has traditionally relied on morphological differentiation based on piroplasm size and shape. Molecular typing has subsequently revealed a more complex taxonomy for these piroplasms than previously thought. To evaluate the factors that influence the morphology of Babesia species upon microscopic examination and hence, their taxonomic classification, we performed detailed characterizations of piroplasms from archival and prospective collections of cytological samples of dogs with piroplasmosis before and after death. Merozoite morphology and time of parasite disappearance following imidocarb dipropionate was also investigated. Methods The study was divided into a (i) review of archived cytological slides from confirmed cases of canine piroplasmosis, and (ii) a prospective study of smears and tissue imprints from 15 recently necropsied dogs. The latter group could be further sub-divided into a non-treated group and an imidocarb dipropionate-treated group. Exact times of treatment before death were reviewed. Additional blood smears prepared from the live dogs and taken before therapy were also evaluated in the latter group. Parasite burden per each slide was determined in both studies. The shape and size of merozoites were described from blood smears taken while the dogs were alive and from different organs during necropsy. The results of all measurements were statistically analyzed. Results The morphology and size of merozoites from live dogs corresponded to that of previously described ‘large’ Babesia. The morphology and size of merozoites were significantly different (P

Pulmonary function tests (PFT’s) routinely implemented in clinics are the first step in the diagnosis of idiopathic pulmonary fibrosis. Evaluation of PFT’s in the mouse model of pulmonary fibrosis accompanied by histological readouts may improve clinical predictability of new therapeutic candidates. Forced expiration (FE) parameters are considered as the most predictive for restrictive pulmonary disorders. The aim of the study was to estimate the translational value of the PFT technique in the established mouse model by evaluating the effects of two approved therapies, pirfenidone and nintedanib. C57BL/6 mice were administered with bleomycin (BLM) intranasally and treated with pirfenidone or nintedanib from day 0 to day 14. Fourteen days after BLM challenge, PFT’s were assessed by using in vivo invasive lung function measurements. Histological evaluation was performed as modified Ashcroft score and digital analysis of de novo collagen deposition (CO1A1) and alpha-smooth muscle actin (αSMA) expression. Bleomycin challenge induced a significant decrease of forced vital capacity (FVC) and forced expiratory volume (FEV). Nintedanib treatment induced significant improvement of FE parameters, specifically 30% improvement of FVC and FEV100 in comparison to the vehicle control. Pirfenidone treatment showed no effect. These findings were confirmed with histological analysis. In conclusion, a good correlation of functional test results and clinical effect of nintedanib and pirfenidone was shown. Based on our findings, implementation of PFT could be a good platform to increase the translational value of the model.