Mapping nanoscale dynamic properties of suspended and supported multi-layer graphene membranes via contact resonance and ultrasonic scanning probe microscopies

Graphene's (GR) remarkable mechanical and electrical properties - such as its Young's modulus, lowmass per unit area, natural atomic flatness and electrical conductance - would make it an ideal material for micro and nanoelectromechanical systems (MEMS and NEMS). However, the difficulty of attaching GR to supports coupled with naturally occurring internal defects in a few layer GR can significantly adversely affect the performance of such devices. Here, we have used a combined contact resonance atomic force microscopy (CR-AFM) and ultrasonic force microscopy (UFM) approach to characterise and map with nanoscale spatial resolution GR membrane properties inaccessible to most conventional scanning probe characterisation techniques. Using a multi-layer GR plate (membrane) suspended over a round hole we show that this combined approach allows access to the mechanical properties, internal structure and attachment geometry of the membrane providing information about both the supported and suspended regions of the system. We show that UFM allows the precise geometrical position of the supported membrane-substrate contact to be located and provides indication of the local variation of its quality in the contact areas. At the same time, we show that by mapping the position sensitive frequency and phase response of CR-AFM response, one can reliably quantify the membrane stiffness, and image the defects in the suspended area of the membrane. The phase and amplitude of experimental CR-AFM measurements show excellent agreement with an analytical model accounting for the resonance of the combined CR-AFM probe-membrane system. The combination of UFM and CR-AFM provide an beneficial combination for investigation of few-layer NEMS systems based on two dimensional materials

CLINICAL EVALUATION OF ENROFLOXACIN (BAYTRIL® 5% INJECTABLE SOLUTION, BAYER, ITALY) AGAINST STAPHYLOCOCCUS AUREUS CLINICAL MASTITIS IN SHEEP

Mastitis in ewes has been reported to occur in all the major sheep-breeding countries. The disease is economically important for both sheep and dairy farmers, because of premature culling of ewes and the reduced performance of lambs that nurse from mastitis ewes. The objective of this research were to evaluate the potential role of commercially available enrofloxacin (Baytril®, Bayer, Italy) in controlling the severity of the clinical signs, to improve cure rates against Staphylococcus aureus and to minimize the effects of the disease on the mammary gland.. These studies were conducted in commercial dairy herds where there was ongoing intensive monitoring of subclinical mastitis by Somatic Cell Count (SCC) and bacteriology. From these herds, 2 different groups namely Group A (n=34 animals) and Group B (n=39 animals) were selected and treated with enrofloxacin (Baytril® 5% injectable solution) 2.5mg/kg/bw and 5mg/kg/bw, respectively for three consecutive days (2 dose per day). The efficiency of the enrofloxacin in curing the S. aureus-induced clinical mastitis were monitored with SCC, rectal temperature, local mammary reaction and systemic reaction from the day 1 (D1) to 14th day (D14) of post treatment. The presence or absence of S. aureus in the sheep was confirmed by bacteriological examination and followed by PCR with specific primer, SA nuc, before and after the treatment. The percentage of bacteriological cure was 39% in Group A and the 82% in Group B. The SCC reduction from D1 to D14 was statistically significant (P<0.001) for both group and also between the groups. The mean rectal temperature also decreased significantly (P<0.001) from D1 to D14 in both groups. The local mammary reaction and systemic reaction were decreased significantly in both treated groups. In conclusion, both enrofloxacin concentration shows good response in reducing clinical signs of mastitis like reduction in SCC, rectal temperature, better attitude and bacteriological cure, but the 5mg/kg/bw concentration used in Group B shows best results when compared with 2.5mg/kg/bw concentration of Group A. The present study provides the evidence that Baytril® 5% injectable solution could helps in controlling the S. aureus-induced clinical mastitis in sheep

STUDY OF EPIGENETIC DEREGULATION OF A NOVEL DEUBIQUITYLASE IN MEDULLOBLASTOMA

A major goal of our work is to study medulloblastoma biology to facilitate the development of targeted therapy. Here, we have dissected the contribution of epigenetics to tumor development. Epigenetic perturbations are pharmacologically reversible and may have therapeutic potential. The RE1-Silencing Transcription Factor (REST) is an epigenetic modulator, and a repressor of neuronal differentiation genes. Its expression is significantly elevated in human medulloblastomas, and is associated with poor prognosis. REST is a driver of medulloblastoma in mouse orthotopic models. To understand REST's contribution to tumor development, we created a novel genetically engineered mouse model in which REST transgene is conditionally elevated in the cerebellar granule progenitors (GNPs), the cells of origin of some medulloblastoma. Transgene induction caused GNP hyperproliferation, accompanied by a failure to express the cell cycle regulator-p27. Using genetic and biochemical approaches, we attributed the lack of p27 expression to direct REST-mediated repression of a novel deubiquitylase, USP37. We also identified a role for USP37 in p27 protein stabilization, and terminal cell cycle exit. USP37 and p27 levels were significantly correlated in patient samples, and their loss was associated with poor overall patient survival. Importantly, constitutive USP37 expression blocked the tumorigenic potential of high-REST human medulloblastomas, suggesting a tumor suppressive function for USP37. REST binding to the USP37 promoter caused increased histone H3 lysine (K)-9 tri-methylation, an epigenetic mark associated with gene silencing. The histone methyl-transferase G9a, which promotes this modification, is a known REST co-repressor. Importantly, genetic ablation of G9a or pharmacological inhibition of its activity upregulated USP37 expression in a REST-dependent manner, and blocked tumor growth in mice. Our work has provided the first link between epigenetic deregulation of the proteasome and medulloblastoma development, and supports the manipulation of the REST-epigenome for future therapeutic application

Evaluation of a therapeutic protocol against Neospora caninum-induced abortion in domestic buffalo (Bubalus bubalus) in Italy

Neospora caninum is a coccidian protozoa that very closely resembles Toxoplasma gondii and causes abortion in a variety of farm animals. This organism was first identified in 1988 as a cause of abortion in dogs and, shortly after a different strain of Neospora was described as causing abortions in dairy cows. Neospora has been found worldwide and is the most common cause of abortions and congenital disease in ruminants. N. caninum has a two-host life cycle in which the infection is acquired through ingestion of coccidial oocysts shed by the definitive host (wild or domestic canids). Antibodies against this agent have been reported in cattle, horses, goats, sheep, deer, hosts. A field evaluation of treatment against N. caninum induced abortions in domestic buffaloes was undertaken in three selected dairy herds of Central Italy (named A,B,C) with a history of high number of abortions. 430 animals, which were serologically negative for BoHV-1, BoHV-4, BVDV, Leptospira hardjo and Chlamydophila abortus were included in the study. In the farm A all animals were treated and sera were collected after 1 year. In the farm B all animals were treated following the same protocol of farm A, but the control sera were collected after six months and, finally, in the farm C only a little group of animals were treated and maintained in the infected farm. Sulphadiazine 200mg/ml and Trimethoprim 40mg/ml was given to the animals at 15mg/kg body weight following different age-depending protocols. Toltrazuril 50mg/ml was also given at 20mg/kg body weight to newborn buffalo and once every three months to the dogs present in the farms. The environment of the farm was disinfected with phenolic disinfectants periodically. A significant reduction of serological values was observed in farms A and B but not in C. Similarly, the percentage of abortions decreased from 24.7 to 3.2 in farm A and from 13 to 1 in farm B (p>0.001), while no difference was observed in farm C. The treatment with Sulphadiazine/Trimethoprim and/or Toltrazuril resulted in drastic reduction in both abortions and seroprevalence since after six months from treatment in farm A and B, while when treated animals were maintained in an infected environment and in contact with infected animals, the proposed therapeutic treatment failed

EFFECT OF LEVAMISOLE ADMINISTRATION ON BLUETONGUE VACCINATION IN SHEEP

Levamisole is an anthelmintic drug with immunostimulant properties when administered at repeated doses of 2.5 mg/kg prior to a vaccine being administered. In order to assess the effect of levamisole administration on bluetongue (BT) vaccination in sheep, four groups of unvaccinated pregnant sheep (8 sheep per group) were used. Group A received vaccine only; Group B received levamisole+vaccine; Group C received Levamisole only; Group D was a non-treated control. Levamisole (Citarin L-10%) was administered three times weekly at an initial dose of 5.0 mg/kg of body weight and subsequently at 2.5 mg/kg of body weight. There was a significant decrease in faecal egg count of gastrointestinal strongyles in Groups B and C. At the beginning of the trial, all animals were serologically negative for BT antibodies; after vaccination, there was a difference in antibody response in animals in the treated groups. Significantly, more animals in Group B developed BT antibodies following vaccination than those in Group A. In conclusion, levamisole appeared to have an immunostimulating effect on the response of sheep to BT vaccination

Effect of levamisole administration on bluetongue vaccination in sheep

Levamisole is an anthelmintic drug with immunostimulant properties when administered at repeated doses of 2.5 mg/kg prior to a vaccine being administered. In order to assess the effect of levamisole administration on bluetongue (BT) vaccination in sheep, four groups of unvaccinated pregnant sheep (8 sheep per group) were used. Group A received vaccine only; Group B received levamisole + vaccine; Group C received Levamisole only; Group D was a non-treated control. Levamisole (Citarin L \u2014 10%) was administered three times weekly at an initial dose of 5.0 mg/kg of body weight and subsequently at 2.5 mg/kg of body weight. There was a significant decrease in faecal egg count of gastrointestinal strongyles in Groups B and C. At the beginning of the trial, all animals were serologically negative for BT antibodies; after vaccination, there was a difference in antibody response in animals in the treated groups. Significantly, more animals in Group B developed BT antibodies following vaccination than those in Group A. In conclusion, levamisole appeared to have an immunostimulating effect on the response of sheep to BT vaccination