Attentional control of gait and falls: Is cholinergic dysfunction a common substrate in the elderly and Parkinson's disease?

The aim of this study was to address whether deficits in the central cholinergic activity may contribute to the increased difficulty to allocate attention during gait in the elderly with heightened risk of falls. We recruited 50 participants with a history of two or more falls (33 patients with Parkinson's Disease and 17 older adults) and 14 non-fallers age-matched adults. Cholinergic activity was estimated by means of short latency afferent inhibition (SAI), a transcranial magnetic stimulation (TMS) technique that assesses an inhibitory circuit in the sensorimotor cortex and is regarded as a global marker of cholinergic function in the brain. Increased difficulty to allocate attention during gait was evaluated by measuring gait performance under single and dual-task conditions. Global cognition was also assessed. Results showed that SAI was reduced in patients with PD than in the older adults (fallers and non-fallers) and in older adults fallers with respect to non-fallers. Reduction in SAI indicates less inhibition i.e., less cholinergic activity. Gait speed was reduced in the dual task gait compared to normal gait only in our faller population and changes in gait speed under dual task significantly correlated with the mean value of SAI. This association remained significant after adjusting for cognitive status. These findings suggest that central cholinergic activity may be a predictor of change in gait characteristics under dual tasking in older adults and PD fallers independently of cognitive status

Discovery of parvovirus-related sequences in an unexpected broad range of animals

Our knowledge of the genetic diversity and host ranges of viruses is fragmentary. This is particularly true for the Parvoviridae family. Genetic diversity studies of single stranded DNA viruses within this family have been largely focused on arthropod- and vertebrate-infecting species that cause diseases of humans and our domesticated animals: a focus that has biased our perception of parvovirus diversity. While metagenomics approaches could help rectify this bias, so too could transcriptomics studies. Large amounts of transcriptomic data are available for a diverse array of animal species and whenever this data has inadvertently been gathered from virus-infected individuals, it could contain detectable viral transcripts. We therefore performed a systematic search for parvovirus-related sequences (PRSs) within publicly available transcript, genome and protein databases and eleven new transcriptome datasets. This revealed 463 PRSs in the transcript databases of 118 animals. At least 41 of these PRSs are likely integrated within animal genomes in that they were also found within genomic sequence databases. Besides illuminating the ubiquity of parvoviruses, the number of parvoviral sequences discovered within public databases revealed numerous previously unknown parvovirus-host combinations; particularly in invertebrates. Our findings suggest that the host-ranges of extant parvoviruses might span the entire animal kingdom

Effect of the 2020/21 season influenza vaccine on SARS-CoV-2 infection in a cohort of Italian healthcare workers

Objectives: Healthcare workers (HCWs) are a priority group for seasonal influenza vaccination (SIV). The 2020/21 SIV campaign was conducted during the second wave of the COVID-19 pandemic. Vaccines, including SIV, may exert non-specific protective effects on other infectious diseases which may be ascribable to the concept of trained immunity. The aim of this study was to explore the association between 2020/21 SIV and SARS-CoV-2 positivity in a cohort of Italian HCWs. Methods: In this observational study, a cohort of HCWs employed by a large (ca 5000 employees) referral tertiary acute-care university hospital was followed up retrospectively until the start of the COVID-19 vaccination campaign. The independent variable of interest was the 2020/21 SIV uptake. Both egg-based and cell culture-derived quadrivalent SIVs were available. The study outcome was the incidence of new SARS-CoV-2 infections, as determined by RT-PCR. Multivariable Cox regression was applied in order to discern the association of interest. Results: The final cohort consisted of 2561 HCWs who underwent ≥1 RT-PCR test and accounted for a total of 94,445 person-days of observation. SIV uptake was 35.6%. During the study period, a total of 290 new SARS-CoV-2 infections occurred. The incidence of new SARS-CoV-2 was 1.62 (95% CI: 1.22–2.10) and 3.91 (95% CI: 3.43–4.45) per 1000 person-days in vaccinated and non-vaccinated HCWs, respectively, with an adjusted non-proportional hazard ratio of 0.37 (95% CI: 0.22–0.62). E-values suggested that unmeasured confounding was unlikely to explain the association. Conclusions: A lower risk of SARS-CoV-2 infection was observed among SIV recipients

Prevalence of Pneumococcal Serotypes in Community-Acquired Pneumonia among Older Adults in Italy: A Multicenter Cohort Study.

Pneumococcal community-acquired pneumonia (CAP) is a leading cause of mortality. Following the introduction of pneumococcal conjugate vaccines (PCVs) in children, a decrease in the burden of the disease was reported. In parallel, an increase in non-vaccine serotypes was also noted. The objective of this study was to assess the current serotype-specific epidemiology of pneumococci among Italian older adults hospitalized for CAP. A prospective study was conducted between 2017 and 2020 in four Italian regions. Subjects aged ≥65 years hospitalized with confirmed CAP were tested for pneumococci using both pneumococcal urinary antigen and serotype-specific urine antigen tests able to identify all 24 serotypes included in the available vaccines. Of the 1155 CAP cases, 13.1% were positive for pneumococci. The most prevalent serotypes were 3 (2.0%), 8 (1.7%), 22F (0.8 %) and 11A (0.7%). These serotypes are all included in the newly licensed PCV20. The serotypes included in PCV13, PCV15 and PCV20 contributed to 3.3%, 4.4% and 7.5% of the CAP cases, respectively. In the context of a low PCV13 coverage among older adults and a high PCV coverage in children, a substantial proportion of CAP is caused by PCV13 serotypes. Higher valency PCV15 and PCV20 may provide additional benefits for the prevention of CAP in vaccinated older adults

Strategie adottate da un densovirus per superare la barriera intestinale di un lepidottero permissivo

I densovirus sono letali per lo stadio larvale di molte specie di insetto, incluse numerose dannose all\u2019agricoltura, e sono perci\uf2 oggi considerati possibili agenti di controllo biologico. In questo studio abbiamo utilizzato il densovirus di Junonia coenia (JcDNV) e il suo ospite permissivo, la larva del lepidottero Spodoptera frugiperda, con l\u2019obbiettivo di chiarire il meccanismo utilizzato dal virus per superare la barriera intestinale. Studi precedenti avevano dimostrato che le particelle virali, introdotte per ingestione di cibo contaminato, superano l\u2019epitelio intestinale senza replicarsi in questo tessuto e che nelle fasi iniziali dell\u2019infezione il virus \ue8 osservabile all\u2019interno delle cellule colonnari, ma non nelle cellule a coppa o nelle cellule staminali, gli altri due tipi cellulari che costituiscono l\u2019epitelio dell\u2019intestino medio larvale di lepidottero. Isolando l\u2019intestino di S. frugiperda in camere di Ussing, abbiamo ora osservato che dopo 10 minuti dall\u2019aggiunta di JcDNV nel compartimento luminale, il virus \ue8 localizzato all\u2019interno delle cellule colonnari, mentre dopo 30 o 60 minuti \ue8 presente negli spazi intercellulari. Utilizzando cellule colonnari isolate, abbiamo osservato che il virus viene internalizzato con diversi meccanismi di endocitosi, e che induce un aumento della concentrazione citosolica di calcio, un mediatore intracellulare che modula la permeabilit\ue0 della via paracellulare. In effetti, l\u2019aggiunta di JcDNV al lato luminale dell\u2019intestino montato in camere di Ussing determina una riduzione significativa della resistenza elettrica paracellulare, un parametro indice della permeabilit\ue0 della giunzione settata. Possiamo quindi affermare che il legame del virus al recettore di membrana e/o la sua internalizzazione nelle cellule colonnari attiva un pathway intracellulare non ancora identificato, che determina un aumento della concentrazione citosolica di calcio seguita da un aumento della permeabilit\ue0 della via paracellulare, la principale via di permeazione utilizzata dal virus per superare l\u2019intestino di S. frugiperda e raggiungere in grande quantit\ue0 i tessuti target. Il virus presente nelle cellule colonnari viene invece indirizzato verso il pathway di degradazione intracellulare

Cell penetration and electrophysiological effects of an insect parvovirus in the midgut of Spodoptera frugiperda

Densoviruses (DNV) are insect parvoviruses sharing with that group a non enveloped 20-25 nm icosaedric capsid and a single stranded DNA genome. Since they are lethal for several insects at larval stages, including agronomical pests and insects vector-borne diseases, the question of their use as biopesticides is revisited. As a model, we studied the interaction of Junonia coenia Densovirus (JcDNV) and one permissive host, Spodoptera frugiperda (Lepidoptera, Noctuidae). The larvae get infected by the oral route, ingesting viral particles contaminating food. The success of the infection first depends on the penetration of the virus through the midgut epithelium. In order to study this early step, we performed a cellular and physiological analysis of the entry of a Cy3-labelled JcDNV through S. frugiperda midgut in vitro. Primary cultures of midgut cells were developed to detect the sites of penetration and the intracellular pathway followed by Cy3-JcDNV. The virus was unable to infect stem cells and differentiated goblet cells, while in incubated columnar cells, whether at an early phase of differentiation or fully differentiated, the virus was visible after 10 minutes in the basolateral membrane and in microvilli, and after 30 minutes within the cytoplasm. Virus particles were apparent as spots, suggesting that they could be distributed in intracellular compartments. S. frugiperda midguts were then isolated from fifth instar larvae, mounted between Ussing chambers and incubated with JcDNV added to the luminal side of the epithelium. After 10 minutes of exposition, the virus induced a significant decrease of the paracellular electrical resistance, an indication that ions moved more rapidly through the aqueous channels formed by the intercellular junctions. The confocal image of whole-mount midguts, isolated and incubated for 10 minutes with luminal Cy3-JcDNV, clearly showed the presence of the virus in the enterocytes\u2019 cytoplasm

The strategies adopted by a densovirus to cross the insect midgut barrier

Densoviruses (DNVs) are parvoviruses highly pathogenic for arthropods, mostly insect at larval stages, including agronomical pest and insect vector borne disease. They have limited host range and are not pathogenic to vertebrates, characteristic that make them particularly interesting as potential pest control agents alternative to chemical pesticides. As a model, we studied the Junonia coenia Densovirus (JcDNV) infection of the Lepidopteran pest Spodoptera frugiperda. In natural condition JcDNV infection is initiated after ingestion of viral particles. The virus does not replicate in intestinal cells, so the midgut epithelium represents a barrier that the virus has to overcome to reach the internal target tissue. To understand the early step of infection, we first analyzed the JcDNV entry in midgut cells in culture. We showed that JcDNV enters specifically columnar cells, but not stem and goblet cells, the three cells types that form the midgut epithelium. Mounting isolated midgut in a Ussing chamber in the presence of JcDNV in the luminal compartment, we showed that after 10 min of incubation viral particles are detectable in the cytoplasm of midgut cells and after 30 min or more of exposition the virus appears in the intercellular spaces. We also demonstrated that JcDNV internalization induces a raise of cytosolic Ca2+ concentration in columnar cells, an intracellular mediator that causes an increase of the epithelium permeability through the paracellular pathway. We effectively verified, in Ussing chamber, that JcDNV causes a significant decrease of the paracellular electrical resistance, i.e an increase of the septate junction (SJ) permeability to ions. Using the same apparatus we also observed that after 10 min of incubation the amount of virus able to cross the epithelium is lower compared to that measured after 30 min. Therefore our results indicate that the internalization of JcDNV into midgut cells causes the raise of intracellular calcium concentration and, consequently, the increase of the permeability of the paracellular route, the pathway used by the virus to cross the midgut epithelium to reach in a massive amount the internal target tissues. To characterized the mechanism involved in JcDNV internalization into midgut cells, isolated midguts were mounted in a Ussing chamber and incubated with the virus in the absence or in the presence of drugs able to inhibit specific endocytic pathway. Our results show that clathrin-coated vesicle formation and lipid rafts-dependent endocytosis are essential for JcDNV entry into columnar cells. Using specific molecular markers for vesicle trafficking we also demonstrated that early and late endosomes are involved in JcDNV intracellular trafficking

Mechanisms involved in the absorption of bioinsecticides and strategies to enhance their passage through the insect midgut

During the past decade a number of products have emerged from the effort to develop alternative biopesticidal technologies. The most recent approach to integrated pest management is based on the detection of new genes encoding for polypeptides with potential insecticidal activity, with a particular attention to biopesticides derived from viruses, microorganisms, fungi, plants and insects. Bacteria and viruses themselves can be used for biological control purpose. In most cases, biopesticides are molecules that have haemocoelic targets and must pass undegraded the gut barrier in order to exert their activity. Therefore for an effective oral delivery of these molecules it will be crucial to develop basic information on the molecular mechanisms mediating the absorption of proteins and peptides by the insect gut and to develop strategies to facilitate their passage through the midgut barrier. These issues have been the subject of my research project during these three years. In particular, I characterized the strategies adopted by Junonia coenia densovirus (JcDNV) to cross the midgut barrier of the lepidopteran pest Spodoptera frugiperda. I also studied a possible mechanism to increase the protein uptake in insect midgut characterizing the mechanisms involved in the internalization of selected Cell Penetrating Peptides (Tat, Penetratin, L-oligoarginine and D-oligoarginine) through the plasma membrane of lepidopteran larveae midgut cells. Even though these two aspects represent the main part of my PhD project, I have also collaborated to the study devoted to the identification of strategies to increase the permeability of the paracellular route