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Abstract Background This is the first report of Besnoitia bennetti in donkeys in Ireland. B. bennetti, an apicomplexan protozoan parasite specific to equids, is an emerging pathogen in Europe. This parasite forms chronic intracytoplasmic cysts in cells of the mesenchymal lineage, mainly fibroblasts, in the skin, sclera and mucosa. Clinical signs in affected equine hosts vary from mild to severe debilitating disease. Little is known of the phylogeny, epidemiology or transmission of B. bennetti infection in donkeys, mules or horses. Case presentation Two cases of besnoitiosis in donkeys are presented. Both donkeys were born and raised in theSouthwest of Ireland. The first case was a 2.5-year-old donkey that had a suspect sarcoid removed, while the second case,a 2-year-old donkey, had a biopsy of nodular dermatitis of the muzzle. Diagnosis was made by histopathology and the parasite species, B. bennetti, was confirmed by PCR followed by sequencing and microsatellite analysis. Both donkeys had high antibody titres against Besnoitia spp. Small (0.5 mm) scleral, conjunctival and dermal cysts over the muzzle were subsequently observed in both animals. Treatment with trimethoprim sulfadiazine for 30 days did not lead to clinical resolution. The findings are compared to the cases of besnoitiosis in donkeys reported in the past 10 years throughout Europe. Conclusions Besnoitiosis should be considered as a differential diagnosis for chronic skin disease particularly in cases of cutaneous masses, non-pruritic dermatitis, and dermatitis that is not responsive to treatment in donkeys and other equids. Future studies are needed to investigate the prevalence of the disease in Irish donkeys, the spread of the disease and the potential impact on the health and welfare of the donkeys.

Abstract Zoonotic diseases are like a sneaky game of “tag” between animals and humans, where the stakes are high and the consequences can be deadly. From the bubonic plague to COVID-19, zoonotic diseases have affected humanity for centuries, reminding us of our interconnectedness with the animal kingdom and the importance of taking proactive measures to prevent their spread. Whether it is avoiding contact with animals or practicing good hygiene, staying safe from zoonotic diseases is a game we all need to play.

Background: Few clinical data are available on thulium fiber laser (TFL) and conservative treatment of upper urinary tract urothelial carcinoma (UTUC). Objective: To assess the effectiveness and safety of TFL in the conservative treatment of UTUC in terms of both tumor ablation and complication rates in a short-term follow-up. Design, setting, and participants: Retrospective data were collected from all patients who underwent endoscopic management of UTUC between January 2021 and April 2022. All patients with nonmetastatic UTUC who were deemed suitable candidates (low- and high-grade disease) for conservative treatment were reviewed. Intervention: All patients underwent ureteroscopy with biopsy and at 2, 6, and 12 mo after the first surgery. UTUC ablation was achieved using TFL. Outcome measurements and statistical analysis: Clinical data were collected in a dedicated database. Intra- and postoperative outcomes were assessed. A descriptive statistical analysis was performed. Results and limitations: In total, 28 patients were evaluated. Thirteen patients (46.4%) were included in the low-risk UTUC treatment group and 15 (53.6%) in the high-risk group. The mean tumor size was 15.3 ± 5.7 mm. Biopsy showed low- and high-grade UTUCs in 19 and eight patients, respectively. Only one biopsy was inconclusive for achieving a diagnosis. At the second procedure biopsy, no tumor was found in 19 cases (70.4%), whereas seven had tumors confirmed (25.9%). To date, 23 and 17 out of 26 patients completed the 6- and 12-mo follow-up, respectively. UTUC recurrence was detected in five of 23 patients (21.7%) and in three of 17 patients (17.7%) at 6 and 12 mo, respectively. A total of 95 procedures were performed. No intraoperative complications were observed. In ten of the 95 procedures (10.5%), Clavien-Dindo grade I–II postoperative complications were experienced. Only one grade IIIB postoperative complication was noted. Conclusions: TFL is a safe and effective technique for conservative treatment of UTUC in a short-term follow-up. Optimal tumor ablation and fine hemostatic control were achieved without major complications. Patient summary: In this study, we looked at the outcomes of upper urinary tract tumors conservatively treated with the new thulium fiber laser (TFL). We conclude that TFL represents a safe and effective technique for the treatment of this kind of tumors in a short-term follow-up.

Abstract Background Although a satisfactory disease control is nowadays achievable in most patients with JIA, a substantial proportion of them still do not respond adequately or reach long-term drug-free remission. According to current recommendations, treatment should be escalated in subsequent steps. A different approach is based on the assumption that the initial start of an aggressive therapy may take advantage of the “window of opportunity” and could alter the biology of the disease, leading to an improvement of long-term outcomes, including the prevention of cumulative joint damage. Objectives This randomised clinical trial aims to compare the effectiveness of a conventional therapeutic regimen, based on treatment escalation and driven by the treat-to-target approach, with that of an early aggressive intervention based on the initial start of a combination of conventional and biological DMARDs. Methods JIA patients with oligoarthritis or RF negative polyarthritis aged more than 2 years and with less than 4 months of disease course will be included in the study. Children will be randomised into two arms: patients in Step-up arm with less severe oligoarthritis will undergo an intra-articular corticosteroid injection (IACI) in all affected joints; patients with polyarthritis or severe oligoarthritis will receive IACI and methotrexate. Subsequent treatment will follow a standardised protocol based on the patients’ level of disease activity measured with the JADAS, according to a treat-to-target strategy. Patients in Step-down arm will receive a 6-month early combined treatment (methotrexate plus IACI for less severe oligoarthritis, methotrexate plus etanercept for severe oligoarthritis and polyarthritis). The primary endpoint is the frequency of achievement of the status of clinical remission (i.e. persistence of inactive disease for at least 6 months) at the 12-month visit. Safety events, physician-centred measures and parent/patient-reported outcomes will be collected through the Paediatric Rheumatology International Trials Organisation on line database. Expected results The STARS trial aims to provide important evidence supporting the first-line treatment choices in the care of children with oligoarticular and polyarticular JIA. If the superiority of an early aggressive therapy will be demonstrated, this will demand further studies on the biological definition of the window of opportunity for JIA. Trial registration The Trial is registered on the ClinicalTrials.gov registry (NCT03728478) on the 31st October 2018 and EU Clinical Trials Register on the 14th May 2018 (EudraCT Number: 2018–001931-27).

Abstract Background Besnoitia besnoiti is an Apicomplexan protozoa causative of bovine besnoitiosis, a chronic and debilitating disease of cattle, with a variety of pathological findings that could alter some laboratory parameters. A study was conducted in a bovine besnoitiosis endemically infected dairy herd located in Italy characterized by high intra-herd seroprevalence and cattle with clinical signs of the disease. In the study, alterations in laboratory parameters, i.e. hematological and biochemical parameters, enzyme activities and serum cortisol levels, in Besnoitia besnoiti naturally infected cows were investigated in depth. Methods Laboratory parameters in 107 cows, of which 61 were seronegative and 46 were seropositive to B. besnoiti, including 27 with clinical signs of bovine besnoitiosis, were compared. Generalized linear models were used to evaluate the effect of Besnoitia infection on the considered laboratory parameters. Results Hematological analyses revealed that B. besnoiti infection determined a significant alteration to the leukocyte differential, with a higher percentage of granulocytes and a lower percentage of lymphocytes in seropositive and clinically affected animals (Mann–Whitney U-test, P = 0.022); erythrocyte and platelet counts did not show any difference between the considered groups of cows. Biochemistry tests evidenced that the parasite infection influenced serum protein values in seropositive cows and glutamate dehydrogenase values in clinically affected animals. No or only slight differences were revealed for all of the other biochemical and enzyme activity parameters in B. besnoiti-infected animals. In addition, despite the lack of statistical significance, seropositive and clinically affected cows evidenced higher concentrations of serum cortisol values compared to seronegative animals. Conclusions Although physiological, pathological and farm-related factors could have influenced the results in investigated animals, further studies involving more animals from different farms would be advisable to infer the role of B. besnoiti on these alterations, since laboratory parameters could help veterinarians in the diagnosis of bovine besnoitiosis in cattle.

Due to the increasing expansion into urban and rural areas, wild boars represent a potential source of infection with zoonotic and animal-specific parasites for both humans and animals. Therefore, the aim of this study was to investigate the serological prevalence to Toxoplasma gondii, Neospora caninum, and Sarcoptes scabiei var. suis in blood samples from wild boars (Sus scrofa) hunted in an anthropized area in Italy. Enzyme-linked immunosorbent assay (ELISA) tests were used to detect antibodies anti-T. gondii and anti-S. scabiei and an immunofluorescence antibody test (IFAT) for antibodies anti-N. caninum. 81 out of 128 wild boars (P = 63.3%) resulted positive for at least one of the three parasites. 68 of them were seropositive to T. gondii (P = 53.1%) and 14 to N. caninum (P = 10.9%). 9 wild boars resulted seropositive to S. scabiei var. suis (P = 7.0%). Sampling season was the only significant risk factor related to S. scabiei var. suis seroprevalence (OR = 7.8). The high occurrence of T. gondii supports the role of this species as a source of infection for other animals and humans. Furthermore, the serological prevalence of N. caninum and S. scabiei var. suis in wild boars from the study area characterized by the presence of numerous dairy cattle and pig farms is relevant to demonstrate its suitability for the circulation of these parasites both in domestic and wild species.

Abstract Background Neospora caninum, a coccidian protozoan, represents an important cause of bovine abortion. Available N. caninum strains show considerable variation in vitro and in vivo, including different virulence in cattle. To which extent sexual recombination, which is possible in the intestines of domestic dogs and closely related carnivores as definitive hosts, contributes to this variation is not clear yet. Methods Aborted bovine foetuses were collected between 2015 and early 2019 from Italian Holstein Friesian dairy herds suffering from reproductive problems. A total of 198 samples were collected from 165 intensive farms located in Lombardy, northern Italy. N. caninum samples were subjected to multilocus-microsatellite genotyping using ten previously established microsatellite markers. In addition to our own data, those from a recent study providing data on five markers from other northern Italian regions were included and analysed. Results Of the 55 samples finally subjected to genotyping, 35 were typed at all or 9 out of 10 loci and their individual multilocus-microsatellite genotype (MLMG) determined. Linear regression revealed a statistically significant association between the spatial distance of the sampling sites with the genetic distance of N. caninum MLMGs (P

Abstract Background Leishmania infantum is a vector-borne pathogen endemic in countries in the Mediterranean basin, including Italy. Dogs act as the primary reservoir for this parasite, but other animal species may also be infected. Low-to-moderate seroprevalence levels of infection have been reported in apparent healthy equine populations in southern Europe, reinforcing the importance of exploring those species, including horses, that act as a food source for vectors and may thus participate in the epizoological scenario of canine leishmaniosis (CanL) and zoonotic visceral leishmaniosis (ZVL). Since little is known regarding the exposure to L. infantum in horses in Italy, we assessed the seroprevalence in healthy equine populations from different CanL endemic areas. Methods The survey was conducted on 660 apparently healthy horses distributed throughout central and northern regions of Italy between 2016 and 2019. Blood samples were collected and the presence of anti-Leishmania antibodies (IgG) was investigated by the immunofluorescence antibody test. Information on the location and altitude of the stables, along with the horses’ breed, age, sex, and reproductive status was obtained by filling in a questionnaire. This was then used for statistical analysis by generalized linear models to explore risk factors associated with seroreactivity to L. infantum. Results An average seroprevalence of 13.9% was detected for L. infantum in the equine populations investigated, with statistically significant associations between seroprevalence, geographical variables (northern vs central Italy, origin and altitude) and individual factors (i.e. age and breed morphotype). Conclusions Our results highlight that horses are frequently exposed to L. infantum. Further prevalence surveys in horses, also using direct methods (e.g. PCR), are warranted to clarify the role of these hosts in the epidemiology of Leishmania in Italy.

Gastrointestinal nematodes (GINs) is a limiting health factor for dairy goats, and the integration of the diet with fodder containing condensed tannins (CT) is becoming increasingly important to control GINs. To preliminary evaluate their potential role as part of GIN control in goat breeding, the in vitro anthelmintic efficacy of the CTs of Silvafeed BYPRO (SBP), Silvafeed Q powder (SQ), and sainfoin hay (SH) was evaluated, and the untargeted metabolomics profiling of the selected formulations was performed. CTs were extracted in water and in ethanol, their concentration was determined, and their chemical characterization was conducted using High-Performance Liquid Chromatography (HPLC) coupled to High-Resolution Mass Spectrometry (HRMS) platform. The in vitro anthelmintic activity of the extracts was evaluated using the Eggs Hatch Test (EHT) and the Larval Migration Inhibition Test (LMIT) using different extract concentrations (150, 300, 600, and 1200 μg/mL). The metabolomic profile of the ethanol extract showed a high number of flavonoids, while the water extract showed higher levels of hydrolysable tannins. The ethanol extracts were effective on both eggs hatching and larvae migration at low concentrations (150 μg/mL) for the three analyzed samples, while the water extracts showed more varied results: SH showed the greatest ovicidal efficacy (concentration 150 μg/mL, %IH = 40.9), while SBP and SQ were more effective against the larvae migration (concentration 600 μg/mL, %LMI = 69.7% and 88%), respectively. The integration of CT-rich fodder into the diet may be considered for the control of GIN infection in goats.

Dermacentor reticulatus is one of the most important vectors of tick-borne pathogens (TBPs) in Europe causing diseases in animals and humans. A longitudinal study was planned, aimed to detect the molecular prevalence of tick-borne pathogens, i.e., Babesia spp. and the spotted fever group Rickettsiae, and its seasonal variation in D. reticulatus questing ticks to define the temporal infection risk. Ticks were collected monthly over a period of 15 months in a peri-urban park in Lombardy, Italy. DNA extraction and molecular analyses were performed. Statistical analysis was carried out. Out of 488, 53 (P = 10.9%) adult questing ticks were positive for Babesia DNA. A higher prevalence was revealed in male (32/241, P = 13.3%) than in female (21/247, P = 8.5%) ticks. Positive ticks were mostly collected in winter months (P = 13.3%) compared to early (P = 7.9) and late (P = 12.8) spring months. A similar percentage of positive ticks was evidenced in transects 1 and 3 (5.8% and 6.5%, respectively); instead, a significant higher prevalence was recorded in transect 2 (P = 16.0%). Obtained sequences confirmed a homology of 100% with B. canis sequences deposited in GenBank. No ticks tested positive for Rickettsia spp. DNA (0/488, P = 0%). The conspicuous circulation of B. canis infection in D. reticulatus adult questing ticks confirms their role in the epidemiology of canine babesiosis and requires preventive measures for dogs in this recreational area. Even if no tick was positive for the spotted fever group Rickettsia, its capacity as a vector of zoonotic pathogens should not be neglected.

Bovine besnoitiosis (BB) is a chronic and debilitating parasitic disease in cattle caused by the protozoan parasite Besnoitia besnoiti. South European countries are affected and have reported clinical cases of BB. However, BB is considered as emerging in other countries/regions of central, eastern and northern Europe. Yet, data on drivers of emergence of BB in Europe are scarce. In this study, fifty possible drivers of emergence of BB in cattle were identified. A scoring system was developed per driver. Then, the scoring was elicited from eleven recognized European experts to: (i) allocate a score to each driver, (ii) weight the score of drivers within each domain and (iii) weight the different domains among themselves. An overall weighted score was calculated per driver, and drivers were ranked in decreasing order of importance. Regression tree analysis was used to group drivers with comparable likelihoods to play a role in the emergence of BB in cattle in Europe. Finally, robustness testing of expert elicitation was performed for the seven drivers having the highest probability to play a key role in the emergence of BB: i.e., (i) legal/illegal movements of live animals from neighbouring/European Union member states or (ii) from third countries, (iii) risk of showing no clinical sign and silent spread during infection and post infection, (iv) as a consequence, difficulty to detect the emergence, (v) existence of vectors and their potential spread, (vi) European geographical proximity of the pathogen/disease to the country, and (vii) animal density of farms. Provided the limited scientific knowledge on the topic, expert elicitation of knowledge, multi-criteria decision analysis, cluster and sensitivity analyses are very important to prioritize future studies, e.g., the need for quantitative import risk assessment and estimation of the burden of BB to evidence and influence policymaking towards changing (or not) its status as a reportable disease, with prevention and control activities targeting, firstly, the top seven drivers. The present methodology could be applied to other emerging animal diseases.

Local livestock breeds are pivotal to ensure sustainable agriculture and represent a real genetic resource in the context of genetic variability reduction. This study aimed at broadening the knowledge about hematological values in female Verzaschese and Camosciata delle Alpi goats (an Italian local and a cosmopolitan goat breed, respectively) and investigating factors affecting them (i.e., breed, age, season). Blood samples were collected monthly from 34 Verzaschese and 37 Camosciata delle Alpi female goats kept under the same semi-extensive farming system for a whole year. The main hematological parameters were evaluated, and descriptive as well as inferential statistical analyses were performed. Reference intervals for complete blood cell count were established and several inter-breed differences were found. In particular, most of the red blood cell parameters were higher in Verzaschese than Camosciata delle Alpi (p < 0.05) suggesting a greater gastrointestinal parasites’ resilience of the local breed. The age effect (p < 0.05) was consistent with the literature while the season effect (p < 0.05) could be explained by the melatonin immunostimulant action and gastrointestinal parasitism influence. Overall, differences in blood values could be attributed to physiological changes and adaptive strategies developed through centuries highlighting the remarkable rusticity and adaptation to the environment and farming system of the local breed.

Tick-borne diseases in horses are considered an emergent problem worldwide; the geographical redistribution of ticks, due to climatic and ecological changes, and the movements of infected horses between different nations play important roles in the spread of tick-borne diseases affecting these hosts. In this study, a survey was planned to estimate the seroprevalence of the Gram-negative bacterium Anaplasma phagocytophilum and the piroplasmid protozoa Babesia caballi and Theileria equi in Italian horses, as well as to evaluate possible risk factors associated with seropositive cases. Serum samples from 261 horses reared in northern Italy were collected and analyzed by indirect immunofluorescence antibody test for the detection of A. phagocytophilum-, B. caballi- and T. equi-specific antibodies. The overall seroprevalence to at least one of the investigated pathogens was 51%; sixty-one horses were seropositive to A. phagocytophilum (23.4%), forty-nine to B. caballi and the same number to T. equi (18.8% each). Seropositivity for more than one of the investigated agents was detected in thirty-two horses and the most common co-infection was observed between B. caballi and T. equi (5.7%). A significant risk factor for all the three pathogens was the elevation above sea level; indeed, the risk of infection was higher with an increase and decrease in elevation for A. phagocytophilum and for B. caballi and T. equi, respectively. Tick control in horses is strongly recommended considering the high seroprevalence values of transmitted pathogens.

Nematode infections of mammals can spread in zoos and faunistic parks and lead to disease in humans and animals. Group treatment strategies with anthelminthic drugs are common. Still, their effectiveness should be verified by sensitive and specific copromicroscopic analyses. This study assessed longitudinal parasitological monitoring, by FLOTAC® dual technique, in mammals housed in an Italian faunistic park, in order to verify the effectiveness of the two adopted ivermectin prophylactic treatments. Twenty-one species of herbivorous mammals from ten families were treated twice per year with ivermectin in an in-feed formulation (medicated feed containing 1.7 g/ton ivermectin daily, for 30 days in March and November), while 13 species of carnivores and primates from five families were treated once a month with oral or subcutaneous administrations of ivermectin (200 μg/kg body weight (b.w.), from March to November). Fecal samples were collected in June–July and October 2019 (late spring–early summer and autumn sampling groups, respectively). All nematode infections, sustained by Nematodirus spp., Capillaria spp., Trichuris spp., Parascaris spp. and Strongylida, were detected in samples collected from herbivores, presenting prevalence rates of infection of 17.3% (9/52), 15.4% (8/52), 15.4% (8/52), 5.8% (3/52), and 3.8% (2/52), respectively. All carnivores and primates tested negative. The general linear mixed model showed that nematode eggs’ excretion in herbivores were influenced by sampling and sampling-host family interaction. Results showed that frequency and dose of prophylactic treatments in herbivores should be improved according to host and parasite taxonomic groups. The treatment adopted in carnivores and primates, together with hygienic management, was effective in nematode control.

Neospora caninum is recognized as a major cause of abortion in cattle, determining economic losses, particularly in dairy industries. To evaluate the impact of neosporosis on herd efficiency, and to understand how the serological analysis for N. caninum is explicative of the farm picture, an epidemiological study was designed in two dairy farms recruited as a case study. Blood samples were collected from 540 animals, including cows and heifers over 12 months, and analyzed by an indirect immunofluorescent antibody test with subsequent antibody titration. Overall, 94 animals (17.4%) were identified as positive to N. caninum antibodies (15.5% and 18.5% in Farm 1 and Farm 2), with differences between the farms concerning the antibody titers (Chi-square, p-value = 0.04), particularly in cows (Chi-square, p-value = 0.018). Consequently, a different pattern of abortion episodes was depicted in the two investigated farms. Considering reproductive and productive performances, the number of inseminations necessary to make an animal pregnant was higher in seropositive cows (2.4 and 2.9 in Farm 1 and 2, respectively) than in seronegative ones (2.1 and 2.4 in Farm 1 and 2, respectively). Similarly, particularly in Farm 1, the number of days in milk of not-pregnant cows was higher in seropositive (167.7) than seronegative animals (133.4). Moreover, although the association between N. caninum infection and milk production is still unclear, both the daily milk production and the 305-mature equivalent milk yield were lower in seropositive (31.02 and 11,838.94) than seronegative cows (33.59 and 12,274.88) in Farm 1; an opposite pattern was otherwise depicted in Farm 2. The study showed that even if N. caninum circulated equally in the two herds, the dynamics of the parasite infection and its outcome may be different, also depending on the specific situation of the farm. In this way, the integration of serological analysis for N. caninum, the reproductive and productive parameters, and the information on herd performances, could give specific indications for the application of control strategies.

Toxoplasma gondii and Neospora caninum (Apicomplexa, Sarcocystidae) are protozoan parasites infecting a wide range of intermediate hosts worldwide, including birds. Raptors acquire the infections through the ingestion of both infected preys and oocysts in the environment suggesting they might be used as indicators of the spread of these pathogens. Here, we report an epidemiological survey with the aim of determining the prevalence of T. gondii and N. caninum infections in wild birds of prey, hospitalized in two Wildlife Recovery Centres (WRCs) in Northern Italy. Genomic DNA extracted from brain tissue samples was submitted to Real Time PCR targeting T. gondii B1 and N. caninum Nc5 genes. T. gondii genotyping was then performed by multilocus sequence typing (MLST) analysis, targeting three polymorphic genes (GRA6, BTUB, and altSAG2). T. gondii DNA was found in 35 (62.5%) out of 56 examined samples; concerning genotyping, it was possible to amplify at least one gene for 26 animals, and obtained sequences belonged to Type II. N. caninum DNA was only detected in two (3.6%) common kestrels (Falco tinnunculus), adding a new species to the list of suitable intermediate hosts for this pathogen. Data obtained in the present study thus confirmed the spread of both T. gondiiand N. caninum in wild bird of prey, endorsing the role of WRCs in the epidemiological surveillance of wildlife.

The objectives of this study were: (i) To investigate possible geographical or environmental factors influencing the infections by Toxoplasma gondii and Neospora caninum in sheep and goats in northern Italy; (ii) to identify areas at risk of infection to set up preventive measures. Forty-three sheep and goat farms were included. Their locations were plotted and associated with T. gondii and N. caninum seroprevalence, then the distribution of farms’ prevalence was evaluated by spatial analysis. Significant clusters for both low and high prevalence were obtained, and a generalized linear model with ordinal logistic regression was implemented to verify if spatial clustering could be due to climate factors (temperature, rainfall, and their interaction). Clusters of high (80.0%) and low prevalence (28.12%) resulted for T. gondii seroprevalence in sheep farms. No significant clusters resulted for goat farms. Clusters of high (38.68%) and low prevalence (21.23%) resulted for N. caninum seroprevalence in sheep farms. One high-prevalence cluster (15.62%) resulted for goat farms. For goats, spatial analysis and analysis on climatic data showed the absence of environmental significant risk factors associated with T. gondii or N. caninum infection. On the contrary, for sheep, annual temperature, rainfall, and their association affected the risk of T. gondii and N. caninum infection. Particularly, high temperatures and abundant rainfalls were related to T. gondii seroprevalence, while low temperatures and scarce rainfalls were related to N. caninum seroprevalence.

Warm renal ischemia performed during partial nephrectomy has been found to be associated with kidney disease. Since endogenous ouabain (EO) is a neuro-endocrine hormone involved in renal damage, we evaluated the role of EO in renal ischemia-reperfusion injury (IRI). We measured plasma and renal EO variations and markers of glomerular and tubular damage (nephrin, KIM-1, Kidney-Injury-Molecule-1, α1 Na-K ATPase) and the protective effect of the ouabain inhibitor, rostafuroxin. We studied five groups of rats: (1) normal; (2) infused for eight weeks with ouabain (30 µg/kg/day, OHR) or (3) saline; (4) ouabain; or (5) saline-infused rats orally treated with 100 µg/kg/day rostafuroxin for four weeks. In group 1, 2–3 h after IRI, EO increased in ischemic kidneys while decreased in plasma. Nephrin progressively decreased and KIM-1 mRNA increased starting from 24 h. Ouabain infusion (group 2) increased blood pressure (from 111.7 to 153.4 mmHg) and ouabain levels in plasma and kidneys. In OHR ischemic kidneys at 120 h from IRI, nephrin, and KIM-1 changes were greater than those detected in the controls infused with saline (group 3). All these changes were blunted by rostafuroxin treatment (groups 4 and 5). These findings support the role of EO in IRI and suggest that rostafuroxin pre-treatment of patients before partial nephrectomy with warm ischemia may reduce IRI, particularly in those with high EO.