Differential ABC transporter gene expression in adult Dirofilaria immitis males and females following in vitro treatment with ivermectin, doxycycline or a combination of both

BACKGROUND: Combination doxycycline/macrocyclic lactone (ML) protocols have been shown to provide a more rapid adulticidal and microfilaricidal effect than either MLs or doxycycline alone, although female worms were reported to have a higher tolerance to treatments compared to male worms. The present study aimed to evaluate how ABC transporters may be involved in the synergic effect of the combination treatment. Adult worms of D. immitis were treated in vitro for 24 hours with doxycycline (DOXY), ivermectin (IVM) and a combination of both, and changes in the modulation of ABC transporter genes were measured. Levels of doxycycline inside different treatment media, post-treatment, were determined through HPLC analysis. RESULTS: Quantitative RT-PCR analysis showed the presence of changes in the modulation of ABC transporter genes evaluated in this study. In particular, in female worms, the combination treatment induced a substantial increase in gene expressions, especially of Dim-pgp-10 and Dim-haf-4; whereas in male worms, the greatest increase in gene expression was observed for Dim-pgp-10 and Dim-pgp-11 when treated with DMSO\u2009+\u2009IVM and DMSO\u2009+\u2009DOXY/IVM. HPLC analysis of the DOXY concentrations in the media after in vitro treatments of male worms showed a slight difference between the DMSO\u2009+\u2009DOXY samples and the combination (DMSO\u2009+\u2009DOXY\u2009+\u2009IVM), while no difference was observed among females. CONCLUSIONS: Further studies are required to explain whether the modulation of cellular efflux plays a role, even partially, in the adulticide effect of doxycycline/macrocyclic lactone combinations in heartworm-infected dogs. To the authors' knowledge, this is the first study to evaluate P-gp expression in adult D. immitis

Electron and ion spectroscopy of azobenzene in the valence and core shells

Azobenzene is a prototype and a building block of a class of molecules of extreme technological interest as molecular photo-switches. We present a joint experimental and theoretical study of its response to irradiation with light across the UV to x-ray spectrum. The study of valence and inner shell photo-ionization and excitation processes combined with measurement of valence photoelectron-photoion coincidence and mass spectra across the core thresholds provides a detailed insight into the site- and state-selected photo-induced processes. Photo-ionization and excitation measurements are interpreted via the multi-configurational restricted active space self-consistent field method corrected by second order perturbation theory. Using static modeling, we demonstrate that the carbon and nitrogen K edges of azobenzene are suitable candidates for exploring its photoinduced dynamics thanks to the transient signals appearing in background-free regions of the NEXAFS and XPS

Characterization of the inflammatory cell infiltrate and expression of costimulatory molecules in chronic echinococcus granulosus infection of the human liver

Background: The local immune responses to chronic echinococcal infections in various organs are largely unknown. Since the liver is the most frequently involved organ in such infections in human we aimed to characterize the inflammatory as well as immune cell infiltrate around hydatid cysts in the liver and compared to common inflammatory processes of the liver. Method: Surgical samples from the liver of 21 cystic echinococcosis (CE) patients were studied and the distribution of different types of inflammatory and immune cells were determined by immunohistochemistry. Furthermore, expression levels of costimulatory CTLA4, CD28, CD80 and CD86 molecules were measured at RNA level by PCR. Liver biopsy samples from patients with steatohepatitis (SH, n = 11) and chronic hepatitis (CH, n = 11) were used as non-inflammatory and chronic inflammatory controls, respectively. The composition and density of the inflammatory and immune cell infiltrates have been compared by using morphometry. Results: CD3+ T cells predominated the inflammatory infiltrate in all pathological processes, while in CE samples CD20+ B cells, in CH samples CD68+ macrophages were also frequent. Both myeloperoxidase (MPO) + leukocytes and CD68+ macrophages were found to be significantly decreased in CE as compared to either SH or CH samples. Concerning T cell subtypes, only CD8+ T cells were found to be significantly decreased in SH samples. CD1a + dendritic cells were almost completely missing from CE biopsies unlike in any other sample types. There were no differences detected in the mRNA expression of costimulatory molecules except decreased expression of CD28 in CE samples. Conclusion: In the hydatid lesions of the liver of chronic echinococcal infections T cell-mediated immunity seems to be impaired as compared to other types of chronic inflammatory processes, suggesting an immunosuppressive role for Echinococcus granulosus, which deserve further attentions

Successful treatment of pleural and peritoneal larval mesocestoidosis with fenbendazole in two dogs

Mesocestoides spp. are common intestinal tapeworms of dogs and cats. The presence of numerous larval stages in the pleural or peritoneal cavities causes larval mesocestoidosis, that is a potentially life-threatening disease in massive infection. In vivo diagnosis of larval mesocestoidosis is challenging and confirmed diagnosis is often made post-mortem. For this reason, therapeutic recommendations are empirical and there is currently no data on the best treatment options in dogs and cats. Both fenbendazole and praziquantel have been used to treat canine larval mesocestoidosis but failure of complete clearance of larvae has been described for both. Treatment with fenbendazole at 100 mg/kg orally twice a day, for 1–3 months has been reported as being effective. However, it is known that administration of high, off-label doses of fenbendazole can lead to adverse reactions. Here, we report the efficacy of fenbendazole at a lower dosage (50 mg/kg twice a day) in two dogs with pleural and peritoneal mesocestoidosis

IMMUNOPHENOTYPING OF T CELL POPULATIONS IN OVINE CYSTIC ECHINOCOCCOSIS (ECHINOCOCCUS GRANULOSUS)

Introduction: Ovine hydatidosis (OH; Echinococcus granulosus) is endemic in several European countries surrounding the Mediterranean basin. It has been reported that early, establishment-phase cysts stimulate a Th1-type immune response, while established phase cysts are associated with a Th2, immuno-suppressive type profile. The aim of the present study was to differentiate and enumerate T cell sub-populations present in the adventitia in established cysts from E. granulosus-naturally infected sheep. Materials and Methods: A total of 18 established cysts were obtained from naturally infected sheep. Samples were frozen and 5 μm-thick serial sections were stained with anti-CD4, anti-CD8 and anti-Foxp3 monoclonal antibodies. Stained tissue sections were analyzed at 40× magnification. Positive cells were enumerated in five consecutive fields of the adventitial layer of the cyst. Three co-authors performed blinded, independent enumeration and mean values ± standard deviations, were calculated. Wilcoxon and Mann–Whitney tests for non-parametric distribution between CD4+ and CD8+ cells were performed and p-Values less than 0.01 were considered to be significant. Results: In all except 5 cyst, the number of CD4+ T lymphocytes was significantly higher (p = 0.009) than CD8+ cells. Foxp3+ cells were present in 17 cysts and made up approximately 35% of total T cell counts in those cysts. Conclusions: The results of the present study may contribute to identifying the immunological features involved during the different developmental stages of Cystic Echinococcosis (CE), including early establishment. Furthermore, the understanding of established-phase associated immunity may offer insights into immunotherapeutic strategies in human CE

A survey on canine leishmaniosis: Best practice and guideline awareness among Italian veterinary practitioners

Canine leishmaniosis (CanL) is a severe, potentially life-threatening disease of dogs and is considered one of the most important vector-borne zoonoses in Europe, where there is evidence that it is currently spreading into previously unaffected areas. Veterinary practitioners represent the first-line defence against CanL and in recent years, several guidelines have been developed to assist veterinarians in the correct diagnosis, management and prevention of the disease. The present study was aimed at surveying Italian practitioners on how they manage CanL and compare results to recommendations from guidelines developed by the European Scientific Council for Companion Animal Parasites (ESCCAP®), by the Leishvet® organization and by the Canine Leishmaniasis Working Group (C.L.W.G.). Approximately 14.0 % of veterinarians contacted replied to the survey. Results suggest that Italian practitioners are aware of currently available guidelines for the management of CanL and adherence to recommendations is good. Interestingly, 16.0 % carry out year-long prevention, even though recent data suggest that transmission in Italy continues to be seasonal. In conclusion, the high awareness among practitioners reported here should be considered an excellent opportunity for guideline developers to continue updating their recommendations to include any new information that can support the correct management of CL

Escherichia coli strains isolated from retail meat products: Evaluation of biofilm formation ability, antibiotic resistance, and phylogenetic group analysis

Escherichia coli is a ubiquitous organism capable of forming a biofilm. This is an important virulence factor and is critical in certain diseases and in the development of antibiotic resistance, which is increased by biofilm synthesis. In the present study, the potential health risk associated with handling and consumption of foods of animal origin contaminated with E. coli- producing biofilm was evaluated. We analyzed the ability of 182 E. coli strains isolated from pork, poultry, and beef, purchased in three different supermarkets in the area of the "Italian Food Valley" (Parma, northern Italy), to form biofilms. Positive strains were also tested for the presence of 12 biofilm-associated genes. Moreover, the 182 E. coli were characterized for antibiotic resistance, presence of multidrug resistance, extended-spectrum β-lactamase strains, and phylogenetic diversity through PCR. Twenty-five percent of the isolates produced biofilm. The majority showed weak adherence, five were moderate, and three were strong producers. E. coli with a strong adherence capability (three of three) harbored eight biofilm-associated genes, while weak and moderate producers harbored only five (frequencies ranging from 80 to 100%). Multidrug resistance was observed in 20 biofilm-producing E. coli, and 15 of these belonged to phylogenetic group D. Among nonbiofilm producers, the percentage of strains belonging to phylogenetic groups B2 and D was approximately 40%, highlighting a potential health risk for consumers and people handling contaminated products. The present study underlines the importance of monitoring the prevalence and characteristics of E. coli contaminating retail meat in relation to the potential virulence highlighted here