Phylogenetic analysis in the clinical risk management of an outbreak of hepatitis C virus infection among transfused thalassaemia patients in Italy.

BACKGROUND: Occurrence of hepatitis C virus (HCV) infection is reduced by effective risk management procedures, but patient-to-patient transmission continues to be reported in healthcare settings. AIM: To report the use of phylogenetic analysis in the clinical risk management of an HCV outbreak among 128 thalassaemia outpatients followed at a thalassaemia centre of an Italian hospital. METHODS: Epidemiological investigation and root-cause analysis were performed. All patients with acute hepatitis and known chronic infection were tested for HCV RNA, HCV genotyping, and NS3, NS5A, and NS5B HCV genomic region sequencing. To identify transmission clusters, phylogenetic trees were built for each gene employing Bayesian methods. FINDINGS: All patients with acute hepatitis were infected with HCV genotype 1b. Root-cause analysis, including a lookback procedure, excluded blood donors as the source of HCV transmission. The phylogenetic analysis, conducted on seven patients with acute infection and eight patients with chronic infection, highlighted four transmission clusters including at least one patient with chronic and one patient with acute HCV infection. All patients in the same cluster received a blood transfusion during the same day. Two patients with acute hepatitis spontaneously cleared HCV within four weeks and nine patients received ledipasvir plus sofosbuvir for six weeks, all achieving a sustained virological response. CONCLUSION: Combined use of root-cause analysis and molecular epidemiology was effective in ascertaining the origin of the HCV outbreak. Antiviral therapy avoided the chronic progression of the infection and further spread in care units and in the family environment.

Italian young doctors' knowledge, attitudes and practices on antibiotic use and resistance: A national cross-sectional survey.

OBJECTIVES: Antimicrobial resistance (AMR) is one of the major health issues worldwide. Clinicians should play a central role to fight AMR, and medical training is a pivotal issue to combat it; therefore, assessing levels of knowledge, attitudes and practices among young doctors is essential for future antimicrobial stewardship (AMS) programmes. METHODS: A nationwide, cross-sectional, multicentre survey was conducted in Italy. A descriptive analysis of knowledge and attitudes was performed, along with a univariate and multivariate analysis of their determinants. RESULTS: Overall, 1179 young doctors accessed the survey and 1055 (89.5%) completed all sections. Regarding the knowledge section of the questionnaire, almost all participants declared to know the different species of bacteria proposed, however the percentage of participants who correctly responded to clinical quizzes was 23% for the question on vancomycin-resistant enterococci (VRE), 42% on carbapenem-resistant Enterobacteriaceae (CRE), 32% on extended-spectrum ß-lactamase-producing enterobacteria (ESBL) and 27% on methicillin-resistantStaphylococcus aureus (MRSA). Similarly, 81% of participants disagreed in stating that AMR was adequately addressed during their medical training and 71% disagreed that they received the right example from their tutors. Finally, a high rate of agreement with the proposed actions to combat AMR was documented; in particular, the percentage agreement was 76% for respondents who agreed to be part of an active surveillance system or AMS programme. CONCLUSIONS: Tackling AMR should be a priority for politicians and for all health workers. Inclusion of competencies in antibiotic use in all specialty curricula is urgently needed.

Maternal caesarean section infection (MACSI) in Sierra Leone: a case-control study.

Sierra Leone is the country with highest maternal mortality and infections are the underlying cause in 11% of maternal deaths, but the real burden remains unknown. This study aims to determine the incidence and risk factors of surgical site infection (SSI) post-caesarean section (CS) in women admitted to Princess Christian Maternity Hospital (PCMH) in Freetown, Sierra Leone. A prospective case-control (1:3 ratio) study was implemented from 1 May 2018 to 30 April 2019 and 11 women presenting with suspected or confirmed infection post-CS were screened for inclusion as a case. For each case, three patients undergoing CS on the same day and admitted to the same ward, but not presenting with SSI, were selected as controls. The post-CS infection rate was 10.9%. Two hundred and fifty-four clinically confirmed cases were enrolled and matched with 762 control patients. By multivariable analysis, the risk factors for SSI were: being single (odds ratio (OR) 1.48, 95% confidence interval (CI) 1.36-1.66), low education level (OR 1.68, 95% CI 1.55-1.84), previous CS (OR 1.27, 95% CI 1.10-1.52), presenting with premature membranes rupture (OR 1.49, 95% CI 1.18-1.88), a long decision-incision time (OR 2.08, 95% CI 1.74-2.24) and a high missing post-CS antibiotic doses rate (OR 2.52, 95% CI 2.10-2.85).

Funnel plots and choropleth maps in cancer risk communication: a Delphi study.

BACKGROUND: In the last decades, the issues related to health risk communication to stakeholders and citizens involving health care practitioners and local political authorities have been increasingly debated. The study evaluated an alternative strategy to communicate cancer risk to local communities, involving an expert panel of public health operators in comparing two different graphic tools, Funnel Plot and Choropleth map. STUDY DESIGN: A Delphi method process was implemented to achieve a unified consensus on an expert panel of public health operators with regard to weaknesses and strengths of the Funnel Plot and the Choropleth map as tools for cancer risk communication to local communities and other stakeholders. METHODS: Participants were asked to score the efficacy of the two tools using a scale. Six properties were explored through two consecutive consensus rounds. Scales were used to calculate frequencies and the content validity ratio for each domain within the consensus rounds. RESULTS: After the two consecutive rounds, participants expressed their preference in favour of the Choropleth map for its ability to define the spatial location of the risk and to locate any potential cluster, while reaching a consensus with regard to the Funnel Plot properties to identify hot spots, displaying the scope of the phenomenon under investigation, and to show the precision of estimates and communicating the significance of estimates. CONCLUSIONS: The Delphi process allowed us to conclude that Funnel Plot could be used as a complement to the current and commonly used graphical and visual formats to effectively communicate cancer epidemiological data to communities and local authorities, representing a useful tool for empowering the general population.

Effectiveness of an educational intervention on seasonal influenza vaccination campaign adherence among healthcare workers of the Palermo University Hospital, Italy.

INTRODUCTION: Healthcare workers are continuously exposed to the risk of being infected by influenza viruses during their work, thus representing a threat especially for fragile patients. Although the Italian Ministry of Health strongly recommends influenza vaccination for all HCWs, coverage levels in Italy are still far from the expected. Several studies report that one of the preferred strategies to improve vaccination coverage among Healthcare Workers is improving vaccination knowledge through specific multidisciplinary courses. To assess the effectiveness of an educational intervention on influenza vaccination coverage among Healthcare Workers a study was conducted at "Paolo Giaccone" University Hospital of Palermo, in the occasion of the 2016/2017 seasonal influenza vaccination campaign. MATERIAL AND METHODS: Educational interventions on influenza infection and vaccination were organized involving personnel of the hospital units in which patients were more fragile. The Healthcare Workers who volunteered attend the course were considered as the intervention group, while two controls for each case, composed by Healthcare Workers not attending it, were randomly selected from the same unit. For both groups, a questionnaire was used to investigate attitude and behaviors toward influenza vaccination, while vaccination coverage data were obtained throughout the Hospital informational data system. RESULTS: Overall, out of the 125 participants, 38 (30.4%) followed the course (intervention group) and 87 (69.6%), not attending the course, represented the control group; later, only 43 Healthcare Workers out of 125 (34.4%) underwent vaccination during the season considered. In particular, after the educational intervention, 42% of the attending Healthcare Workers got vaccinated, while vaccination prevalence in the control group was 31%. The Healthcare Workers who underwent vaccination reported, before the intervention, a higher risk perception for contracting (transmitting) influenza compared to those not vaccinated (p<0.05), while no significant difference in risk perception of transmitting influenza to their patients was reported between the two groups. DISCUSSION: Despite the training provided, and an improvement in vaccination adherence by the Healthcare Workers involved, coverage obtained was lower than recommended to reduce influenza spread in hospital contexts. In conclusion, our data suggest that specific training alone may play a role in the improvement of influenza vaccination adherence, but it should be integrated by a wider range of public health measures, including mandatory vaccination.

Improvement in vaccination knowledge among health students following an integrated extra curricular intervention, an explorative study in the University of Palermo.

INTRODUCTION: Vaccination coverages threaten to decrease because of false beliefs in their unsafety and inefficacy. Therefore formation of future health-care workers on this topic is fundamental to deal with any doubt and to promote active immunization among general population. METHODS: In order to assess health-care students' knowledge about vaccination before an integrated seminar on this topic, and to evaluate their improvement after the educational intervention, an integrated educational intervention was held by a multidisciplinary team. Before and after the seminar, 118 students of medicine and biology schools at Palermo University were asked to answer 10 multiple-choice questions regarding vaccine history, mechanism of action, side effects, composition, use and nowadays issues (hesitancy). Two more questions investigating possible changes on students' attitudes towards vaccination and the usefulness of the formative intervention, were added at the post-test phase of the survey. RESULTS: Eighty-one out of 118 students (68.6%) answered to both pre- and post-test questions. 97.6% and 81.5% of the participating group also completed the two additional questions about their improvement in knowledge (question 11) and attitudes (question 12) towards vaccinations. The post-test results showed a significant improvement for all questions administered, except for number 3 (about a specific immunological content), with an overall percentage of correct answers increasing from 38.8% to 77.6% (p©< 0.001). CONCLUSIONS: The present explorative study put the basis for future studies, stronger in the methodology, and highlights the importance of educating health-care professions students by integrated extra-curricular intervention to be held early in their degree curricula and in order to improve knowledge and attitudes towards vaccinations and to prepare them to promote vaccines among the general population.

Current state of genomic policies in healthcare among EU member states: results of a survey of chief medical officers.

Background: A need for a governance of genomics in healthcare among European Union (EU) countries arose during an international meeting of experts on public health genomics (PHG). We have conducted a survey on existing national genomic policies in healthcare among Chief Medical Officers (CMOs) of the 28 EU member states, plus Norway. Methods: A questionnaire was sent to CMOs after a meeting on the policy implications of PHG held during the Italian presidency of the Council of EU in 2014. The survey was closed in November 2015. Results: CMOs response rate was 65.5% (19/29). Twelve (63.2%) reported that their countries had a policy for genomics in healthcare in place, and 15 (78.9%) reported that public funding existed. Public research facilities for the development of such policies were documented in 13 (68.4%) countries, and 15 (83.3%) had working groups devoted to policy development. National agencies carrying out Health Technology Assessment of genomic-based technologies were present in nine countries (50%). Sixteen (88.9%) countries reported having agencies dealing with ethical issues related to genomic technologies. About 55% of countries disclosed the lack of information campaigns aimed at citizens, and 44.4% reported they had a legal framework for direct-to-consumer genetic tests. Conclusion: Belgium, France, Italy, Spain and UK documented the presence of a policy on genomics in healthcare. While many caveats are necessary because of the methodology, results suggest a need for a co-ordinated effort to foster development and harmonization of dedicated policies across EU to responsibly integrate genomics policies into existing health systems.

Neurophysiologic exploration: a reliable tool in HIV-1 encephalopathy diagnosis in children.

BACKGROUND: HIV-1 related encephalopathy has a bad prognostic meaning in the course of AIDS disease, but the early association of different drugs can modify its course. For this reason it is very important to recognize CNS involvement as soon as possible. As shown in the literature, at least in adult studies, EEG and Evoked Potentials (EP) are good tools in evaluating CNS alterations. In children data are rare. METHODS: A ten-year prospective study of 44 infected children out of 142 born from HIV-1 positive mothers has been done. The children have been submitted to EEG recording every six months in the first 18 months of life and then every year, to multimodal EP every six months. A total of 357 EEG, 47 P-VEP, 62 F-VEP and 98 BAEP have been performed. RESULTS: EEG: we found no pathologic results in patients belonging to category A; results were pathologic in 17.7% in category B, in 47.7% in C and in 77% of encephalopathic patients. It seems that EEG alterations are parallel to disease progression, with a relative risk of developing encephalopathy (R.R. = 1.15) and of death (R.R. = 2.33) for patients belonging to category C. We obtained a statistically significant lengthening in BAEP interpeak latency of left ear in all groups. For patients in category C the risk of developing encephalopathy is statistically significant (p = 0.045; R.R. = 6.75) and risk of death is high (R.R. = 4). CONCLUSIONS: Neurophysiologic exams are a reliable tool for the diagnosis of encephalopathy, in addition to clinical evidence.

Evolution of HIV-1 encephalopathy in children.

BACKGROUND: This study has been conducted on a series of HIV-1 infected children, with the aim of illustrating the features of encephalopathy onset, its evolution and its influence on life expectancy. The most useful exams for diagnosis are also outlined. METHODS: The perspective study lasted from January 1989 to June 1997. Forty six symptomatic patients, out of 142 seropositive children, were followed up in the Department of Paediatric and Adolescence Sciences of the University of Turin. The patients, now between 1 yr 2 mth and 13 yr 9 mth old, were born from HIV-1 seropositive mothers; seroreverters have been excluded. Scheduled neuropsychiatric consultations were used, consisting of a neurologic exam and an interview with parents, cognitive evaluations, EEGs, Evoked Potentials and CT scans. The results have been evaluated with log-rank test for the analysis of the survival curves. RESULTS: We found a significantly higher mortality rate in encephalopathic versus non encephalopathic patients; encephalopathic patients, in whom neurologic signs began in the first year of life, have a worse prognosis than the other patients, in whom encephalopathy appeared later. We did not find a statistical correlation between clinical course and immunological deficit. The clinical features of encephalopathy are mainly characterized by pyramidal signs and cognitive deterioration. Clinical sign evolution is linked to the age of encephalopathy onset: plateau pattern encephalopathy, characterized by an early onset, severe motor signs and cognitive delay from the very beginning, shows a greater severity and a shorter survival than progressive encephalopathy, characterized by a slowly progressive evolution of pyramidal signs, to which a cognitive deterioration may be added. CONCLUSIONS: Neuropsychological exams can be helpful in the diagnosis and follow-up of encephalopathy.

Caregivers of persons with Alzheimer's Disease: cultural differences in perceived caregiver burden in Guatemala and Rhode Island.

The intent of this study is to illustrate cultural differences in the amount of perceived burden for primary caregivers of persons with Alzheimer's Disease. Caregivers in Guatemala and Rhode Island were given a questionnaire exploring: caregiver well-being, available supports, traditional ideology, and perceived burden. The data indicate that Guatemalans have less institutional and more informal supports available, as compared with USA caregivers. Guatemalan caregivers brought patients to a doctor sooner after the appearance of their first symptoms (0.9+/-1.0 years versus 1.6+/-1.8 years) and had poorer perceived health than USA subjects, suggesting a higher level of caregiver burden. Cultural response bias however may account for the difference in perceived health.

Neuroimaging of vessel amyloid in Alzheimer's disease.

Despite extensive recent advances in understanding Alzheimer's disease (AD) we are unable to noninvasively establish a definite diagnosis during life and cannot monitor the cerebral deposition of amyloid beta protein (A beta) in living patients. We evaluated the use of 10H3, a monoclonal antibody Fab targeting A beta protein 1-28 labeled with Tc-99m. Six subjects with probable AD were studied using single-photon emission computed tomography (SPECT) at times from 0-24 hours following injection. Curves of radioactivity in blood demonstrate a half-life of the injected Fab of 2-3 hours. Images show uptake around the head in the scalp or bone marrow in all subjects. There is no evidence of cerebral uptake of the antibody. Scalp biopsies in all six patients demonstrate diffuse staining with 10H3 of the scalp, a pattern indistinguishable from that found in controls. Evidence of amyloid deposition in the scalp in AD is not seen with other anti-A beta antibodies, suggesting that 10H3 is cross-reacting with another protein. Further studies with anti-A beta antibodies will require longer-lived radionuclides to detect cerebral uptake at later times after injection to allow for complete clearance from the blood. Alternately, imaging using labeled A beta itself may provide a means for noninvasive targeting of cerebral amyloid.

Induction of connexin43 and gap junctional communication in PC12 cells overexpressing the carboxy terminal region of amyloid precursor protein.

Previous studies have shown that PC12 cells overexpressing beta/A4 amyloid peptide display altered morphology characterized by pronounced membrane ruffling and extensive intercellular appositions. Having observed other cell types in which these features accompany increased connexin43 (Cx43) production and gap junctional communication, we examined Cx43 in normal and beta/A4-transfected PC12 cells. Studies of two beta/A4-transfected PC12 clones revealed an induction of Cx43 expression by Western blotting, intracellular and plasma membrane-associated Cx43 in some cells of cultures processed by immunofluorescence, dye-transfer between some cells microinjected with Lucifer Yellow, and gap junctions between cells examined by EM. Normal and vector-transfected PC12 cells exhibited none of these properties. Increased immunofluorescence in some clusters of beta/A4-transfected cells was also observed with a monoclonal antibody against connexin32. The results suggest that beta/A4 amyloid peptide may cause aberrant intercellular communication and gap junction formation through induction or increased expression of connexins in cells that are not normally coupled or only poorly coupled by gap junctions.

Elevated connexin43 immunoreactivity at sites of amyloid plaques in Alzheimer's disease.

The distribution of the astrocytic gap junctional protein, connexin43 (Cx43) was compared immunohistochemically with that of amyloid plaques in Alzheimer's Disease (AD) brain. By light microscopy, cortical areas containing numerous beta/A4 amyloid plaques exhibited increased immunostaining density for Cx43 and some plaques corresponded exactly to sites of intensified Cx43 immunoreactivity. By electron microscopy, Cx43 was localized to astrocytic gap junctions in AD brain. Increased Cx43 expression in AD may represent an attempt to maintain tissue homeostasis by augmented intercellular communication via gap junction formation between astrocytic processes that invest senile plaques, or alternatively, an aberrant induction of astrocytic Cx43 expression which may further compromise homeostasis and exacerbate pathological conditions in the microenvironment of amyloid plaques.

Propagation of intercellular calcium waves in PC12 cells overexpressing a carboxy-terminal fragment of amyloid precursor protein.

We have recently found that PC12 cells overexpressing a C-terminal 97 amino acid fragment containing the beta/A4 amyloid peptide of amyloid precursor protein (APP) exhibit an induced production of the gap junctional protein connexin43 (Cx43) and an induction of gap junctional communication as assessed by dye-transfer. In studies of two beta/A4-transfected PC12 clones, we show here that these cells, unlike normal PC12 cells or those transfected with vector alone, have the capacity for intercellular transmission of calcium waves as determined by imaging of calcium with fura-2. Intercellular wave propagation occurred in the absence of extracellular calcium and was blocked by known inhibitors of gap junctional communication (octanol and 12-O-tetradecanoyl-phorbol-13-acetate), suggesting mediation by gap junctions. The results indicate a disruptive influence of the C-terminal region of APP or of beta/A4 amyloid peptide on intercellular signaling via gap junctions, which may be relevant to the normal functions of APP or to pathology in Alzheimer's disease.

Modulation of the PC12 cell response to nerve growth factor by antisense oligonucleotide to amyloid precursor protein.

1. The amyloid precursor protein (APP) is widely distributed among eukaryotic cells, however, its precise role in cellular functioning is not fully clarified. APP is glycoprotein membrane constituent and it may facilitate membrane associated functions. 2. The aim of the present study was to examine the possibility that APP may play a role in mediating cellular trophic responses. The methods made use of an antisense oligonucleotide that was prepared to the 5' terminus of APP and shown specifically to reduce the level of APP isoforms. 3. In sequential mixing experiments it was observed that the APP antisense oligonucleotide did not significantly modify the trophic response of PC12 cells pretreated with nerve growth factor (NGF). However, pretreatment of cells with the antisense oligonucleotide diminished NGF-induced increases in cellular size and neurite length. 4. These observations suggest that APP may play a role in modulating the trophic response. The combined use of APP antisense oligonucleotides and neurotrophic agents may find clinical utility in the treatment of Alzheimer-type dementia since it is known that NGF normally causes increases in APP levels.

Development of an anti-A beta monoclonal antibody for in vivo imaging of amyloid angiopathy in Alzheimer's disease.

We evaluated the efficacy of murine monoclonal antibodies (MAbs) targeted to the A beta amyloid of Alzheimer's disease for development of procedures for the in vivo identification of amyloid angiopathy (AA). MAbs to A beta were prepared and screened for effectiveness in visualizing AA and neuritic plaques in postmortem AD brain sections. They were assessed again after enzymatic cleavage to produce Fab fragments and after labeling with technetium-99m (99mTc) using a diamide dimercaptide ligand system. Modified and radiolabeled Fab fragments retained activity and specificity toward amyloid-laden blood vessels and neuritic plaques. A highly specific murine MAb, 10H3, was identified and characterized that fulfills criteria necessary for the development of an in vivo diagnostic imaging agent. Toxicity studies in rats showed the MAb to be safe. Biodistribution studies in mice demonstrated desirable properties for use as an imaging agent. Expansion and adaptation of these strategies may provide the methods and materials for the noninvasive analysis of AA in living patients, and permit assessment of the contribution of AA to the clinical and pathological features of AD.

Intercellular interactions in PC12 cells overexpressing beta/A4 amyloid.

The amyloid precursor protein (APP) is an integral membrane component of eukaryotic cells. A variety of research approaches have addressed the contribution of the beta amyloid peptide region of the APP to neuritic plaque structure and formation in the Alzheimer disease brain as well as the relationship between beta amyloid accumulation and the occurrence of dementia. However, there is limited information available concerning the cellular consequences of amyloid deposition. The present studies were undertaken to investigate the relationship between beta amyloid and intercellular junctions. Transfected PC12 cell lines, that overexpress the beta amyloid peptide, exhibit structural and functional alterations at the cell surface and tend to form aggregates more readily than normal control cells. Intermediate junctions were the most common intercellular interactions of both normal and transfected cells. However, the control and transfected cells differed since areas of continuous and extensive junctions were readily seen in transfected cells and infrequently seen in control cells. The data suggest that excess accumulation of beta amyloid is associated with the junctional apparatus and may be related to increased intercellular adhesion.

Membrane surface ruffling in cells that over-express Alzheimer amyloid beta/A4 C-terminal peptide.

Deposition of beta/A4 amyloid in brain is a defining characteristic of Alzheimer disease (AD); however, the extent to which amyloid deposits may interfere with normal cellular processes is incompletely understood. We examined this issue by means of PC12 cells. After transfection with DNA coding for 97 amino acids of the beta/A4 C-terminal region of the amyloid precursor protein, beta/A4 antigen was visible at the cell membrane. We report that normal unstimulated PC12 cells exhibit ruffling activity at the cell surface when plated on a plastic substrate. Relative to control cells, however, those that over-expressed the beta/A4 C-terminal peptide had significantly higher levels of ruffling activity, suggesting a structural and/or functional membrane modification. Similar cellular alterations, if present, in Alzheimer brain cells, may indicate disturbances in membrane-associated functions, including intercellular communication.

PC12 cells release stimulatory factors after transfection with beta/A4-C-terminal DNA of the Alzheimer amyloid precursor protein.

The beta/A4 region of the amyloid precursor protein (APP) accumulates in brains of victims of Alzheimer disease (AD) where it is a major component of senile plaques. We examined the pathophysiological consequences of overexpression of the beta/A4-C-terminal DNA in PC12 cells. Serum-free conditioned media (SFCM) from positive transfectants stimulated control PC12 cells to extend neurites and increase in size. Unlike the factor that affected cell size, neurite lengthening activity was significantly decreased after immunoabsorption with anti-beta/A4 monoclonal antibodies (MAb) and changes in pH. The data support the view that among the consequences of beta/A4-C-terminal DNA overexpression in PC12 cells is the release of factors that stimulate nontransfected cells to undergo morphological transformations that include differentiation to a neuronal phenotype. It is hypothesized that similar activities that may contribute to the molecular pathophysiology of the disorder may be present in the AD brain.