Vibrio mediterranei, a potential emerging pathogen of marine fauna: investigation of pathogenicity using a bacterial challenge in Pinna nobilis and development of a species-specific PCR.

AIMS: Extreme mortality events affecting Pinna nobilis, some associated to Vibrio mediterranei, have depleted many populations of this bivalve. The objective of this study was to demonstrate pathogenicity of V. mediterranei in the host P. nobilis by performing a bacterial challenge in P. nobilis to understand if V. mediterranei has specific virulence in this host. To assist this objective, a secondary objective was to develop a species-specific DNA diagnostic test. METHODS AND RESULTS: Pinna nobilis collected from local bays were used in a challenge experiment with V. mediterranei (strain IRTA18-108). The virulence in the host background of P. nobilis was demonstrated at doses of 103 CFUs per animal. An alignment of published Vibrio sp. atpA sequences was used to design V. mediterranei-specific primers. Furthermore, data mining of published literature and V. mediterranei genomes identified multiple virulence-related genes (vir genes) from which specific primers were designed for PCR detection of selected genes. CONCLUSION: Vibrio mediterranei strain IRTA18-108 is pathogenic in the host P. nobilis. The virulence genes sod, rtx and mshA were identified in this strain. Temperatures of 24°C or higher appear to trigger onset of virulence. Sensitivity and specificity of the Vm atpA PCR is useful for diagnosis of Vibriosis in shellfish. SIGNIFICANCE AND IMPACT OF THE STUDY: The presence of previously described virulence genes have been confirmed in this strain. The specific Vm atpA PCR assay will aid management of future epizootics of this emerging pathogen of aquatic fauna, and improve surveillance capabilities for mortality events where Vibrios are suspect.

Baseline pathological data of the wedge clam Donax trunculus from the Tyrrhenian Sea (Mediterranean Basin).

In recent years, a collapse in Donax trunculus fishing yields has occurred in the Tyrrhenian Sea (Mediterranean Basin). There is little information available on the impact disease may have had on D. trunculus populations. For the first time, a pathological survey was performed on the natural beds of the bivalve on the Campania and Lazio coasts, western Italy. Detected pathogens and related diseases were analysed, and their prevalence and mean intensity values were calculated. Viral particles, Chlamydia-like organisms, ciliates, coccidians, microcells and trematodes were observed. An unknown ciliate was linked to severe inflammatory and necrotic lesions in the digestive gland. Metacercariae of the trematode Postmonorchis sp. were also strongly represented in almost all samples, reaching high levels of infection; however, none of the pathogens described required the World Organisation for Animal Health to be notified. Initial results indicated that further surveys related to environmental data are necessary in order to assess the relevance of these early observations in managing the declining D. trunculus population in the Tyrrhenian Sea.

Comparative pathology in bivalves: Aetiological agents and disease processes.

Comparative pathology as a scientific discipline studies animal diseases in relation to their aetiology, pathogenesis and prognosis. Among the main aspects of this discipline, regressive changes, host defense responses with pathological implications and progressive changes, represent the majority of the possible responses of cells and tissues to pathogens and exposure to chemicals. One of the most persistent issues in the field of invertebrate pathology is the variability in terminology and definition, which has led to confusion in scientific communication. The aim of this paper is to provide an overview of the pathological basis of bivalve disease (defensive, regressive and progressive phenomena) and contribute to the standardised terminology for bivalve molluscan disease in the context of comparative pathology.

Quantitative histopathology of the Mediterranean mussel (Mytilus galloprovincialis L.) exposed to the harmful dinoflagellate Ostreopsis cf. ovata.

Ostreopsis cf. ovata is a benthic dinoflagellate that produces palytoxin-like compounds that adversely affect both marine vertebrates and invertebrates and are reported to be responsible for human intoxication in aerosol form. In this work, a histopathological analysis accompanied by quantitative evaluation of tissue injury in mussels (Mytilus galloprovincialis) exposed to O. cf. ovata cells under natural and experimental conditions, provided baseline data on the health status of the mussels in terms of defensive and regressive pathological changes. We recorded a total of 15 health parameters in the digestive system, muscle, kidney and gills in mussels exposed to O. cf. ovata both in the laboratory and at sea. Animals exposed to different concentrations of O. cf. ovata cells (300, 500 and 1000cellsml(-)(1)) for 48h showed activation of the inflammatory response, which increased with the cell concentration, mainly characterized by haemocyte aggregates actively enclosing the algae, while mussel mortality was also recorded in some cases. Moreover the use of image analysis for the evaluation of digestive tubule damage revealed a pronounced increase in the lumen in terms of its area, perimeter and circularity, with a shift in a high percentage of tubules from an adsorbing profile to an atrophic profile. Animals collected from the natural environment during a summer bloom of O. cf. ovata in the Gulf of Naples (Italy) showed comparable lesions in terms of types and severity. This is the first quantitative study assessing damage to the digestive epithelia in terms of lumen modifications in mussels exposed to O. cf. ovata. The presented methodology provides a new technique for automating the evaluation of epithelial tubule modifications. Our results highlight the importance of monitoring the presence of O. cf. ovata in this area, taking into account the effects on the residing marine species.

Gorgonian disease outbreak in the Gulf of Naples: pathology reveals cyanobacterial infection linked to elevated sea temperatures.

In recent years, mass mortality events of benthic invertebrates in the Mediterranean Sea have been documented to coincide with the increased seawater temperatures associated with global climate change. Following a disease outbreak in gorgonians during the summer seasons of 2008 and 2009 in the Gulf of Naples (Tyrrhenian Sea), we conducted gross and microscopic analyses of healthy and diseased specimens of Eunicella cavolinii and E. singularis using both light and electron microscopy (SEM). Macroscopically, diseased colonies exhibited evident tissue thinning, and dead colonies showed a complete loss of polyps and coenenchyme, exposing their skeletons to settlement by fouling organisms. Histopathology revealed chronic inflammatory lesions at the polyp and axial level, characterized by amoebocyte infiltration of tissue accompanied by new apposition of melanin/gorgonin sheets. We interpreted this response as a defense against different kinds of pathogens-identified as mainly a heterogeneous consortium of filamentous cyanobacteria-and which gradually led to enlargement and hardening of the coral axis, which resembled a wood-like structure at the final stage of the disease. These processes elicited the formation of multiple inflammatory nodules and capsules, some of which were macroscopically visible. A parallel 16S rRNA and ITS analysis of the diseased tissue identified Synechococcus, Arthrospira and other uncultured cyanobacteria grouped within the Oscillatoriales. These results suggest that a cyanobacterial consortium is involved in the pathogenesis of the inflammatory disease leading to the mortality of Gorgoniaceae in the area. Finally, there were anomalously high temperatures (up to 25°C) between 10 and 20 m depth during the sampling period, particularly in June 2009. This supports the hypothesis that the coral skeleton may serve as a reservoir for the pathogens in cooler seasons, with warmer conditions leading to pathogen reactivation and recurring mortality events.

Postmonorchis sp. inq. (Digenea: Monorchiidae) metacercariae infecting natural beds of wedge clam Donax trunculus in Italy.

The wedge clam Donax trunculus Linnaeus, 1758 is one of the most common bivalve molluscs inhabiting the sandy shores of the Mediterranean Sea and is considered an important commercial resource. In this study, we report the first molecular, morphological and histopathological descriptions of metacercariae from a trematode belonging to the genus Postmonorchis (Digenea: Monorchiidae) that infects D. trunculus in natural beds of the Italian Tyrrhenian coast (Campania, Lazio and Tuscany). Morphological analysis of the parasite revealed a combination of features that exist in the 3 previously identified species of Postmonorchis, viz. P. donacis, P. variabilis and P. orthopristis, with the addition of new, distinctive morphological characteristics. The pathogen exhibited a predilection for the gill; however, it was also present in the labial palp and mantle in addition to the gut, kidney epithelium and foot. The inflammatory response was characterised by either a focal or diffuse haemocyte infiltration followed by the formation of multiple, large multi-layered capsules associated with tissue destruction. The prevalence of the pathogen ranged from 75 to 100%, while the infection intensity fluctuated among the study areas. Further studies regarding the life cycle of this parasite and the identification of other larval and adult stages and their respective hosts may confirm the identification of a new species of Postmonorchis that infects wedge clams in Mediterranean waters. The study of the parasite is completed by molecular analysis of the ITS1 and ITS2 rDNA sequences.

[Pilot study: Apricena Marble District quarry workers and COPD]. / Studio pilota: addetti alle cave di pietra nel distretto del Marmo di Apricena e BPCO.

The aim of the present article was to verify the Chronic Obstructive Pulmonary Disease (COPD) prevalence in a cohort of quarry workers who belong to the Apricena Marble District. We studied 70 workers. They received a questionnaire about the disease and confounding factors. The spirometry showed that the FEV1 was normal in 95% of workers, instead 5% showed values lower than former (Average: 73%). TNF alpha and IL-1 Beta in Exhaled breath condensate (EBC) were lower than the method limit in all workers. Our cohort is limited, but we could retain that the lung disease is not present in workers taken into consideration. Our results are in according to Rushton who demonstrated that only a prolonged occupation, higher than thirty years, is able to induce lung disease.

Training the teachers for improving primary schoolchildren's fruit and vegetables intake: a randomized controlled trial.

Effectiveness comparison of different school-based nutrition education interventions to improve fruit and vegetable (F&V) consumption in schoolchildren not yet well researched. We evaluated the effectiveness of two school-based interventions promoting fruit and vegetable intake among Italian schoolchildren. In this randomized intervention trials, the first intervention was conducted by schoolteachers who attended a nutritionist-led training course (group-1), the second intervention conducted by schoolteachers who performed a self-training course (group-2). Thirty-two second to fifth-grade elementary classes enrolling 96 schoolteachers and 813 schoolchildren were randomized, and assigned to the two different nutrition education interventions. 804 students completed the study. By the end of the study, the group-1 (n = 409) followed by the teachers who attended a nutritionist-led course successfully increased the consumption of fruit and vegetables: in 183 (44.7%) and 157 (38.3%) schoolchildren respectively; the group-2 (n = 395) with teachers who performed a self-training course reported an increased consumption of fruit and vegetables in 81 (20.5%) and 76 (19.2%), respectively. This study indicates that a school-based nutrition education conducted by teachers who attended a nutritionist-led training course has a significant impact on primary school-age children's F&V intake.

Intraoperative microrecordings in the posterior hypothalamus of anaesthetized humans with aggressive behaviour.

Two patients with uncontrollable aggressive behaviour underwent the placement of stimulating leads in the posterior hypothalamus (pHyp). One patient had also multifocal refractory epilepsy. Microrecordings were obtained in both patients during surgery under general anaesthesia. Firing rate, interspike intervals and oscillatory discharge patterns were analysed in 14 neurons. A mean discharge rate of 19 Hz, and oscillatory activity at 7-8 Hz were recorded in the first patient with aggressiveness and epilepsy. In the second patient the mean firing rate was 10 Hz, with evidence of both tonic and random firing patterns. Previous studies in patients with cluster headache showed that a discharge rate around 20 Hz and lack of a specific rhythmic pattern were the most consistent characteristics of neuronal discharge in this area. Our present findings therefore would suggest that the pattern of discharge of neurons in the pHyp should be evaluated with reference to the presence of concurrent pathology.

Effects of acute changes in salinity and temperature on routine metabolism and nitrogen excretion in gambusia (Gambusia affinis) and zebrafish (Danio rerio).

Acute stress may affect metabolism and nitrogen excretion as part of the adaptive response that allows animals to face adverse environmental changes. In the present paper the acute effects of different salinities and temperatures on routine metabolism, spontaneous activity and excretion of ammonia and urea were studied in two freshwater fish: gambusia, Gambusia affinis and zebrafish, Danio rerio, acclimated to 27 degrees C. The effects on gill morphology were also evaluated. Five salinities (0 per thousand, 10 per thousand, 20 per thousand, 30 per thousand and 35 per thousand) were tested in gambusia, while four salinities were used in zebrafish (0 per thousand, 10 per thousand, 20 per thousand and 25 per thousand). Each salinity acute stress was tested alone or in combination with an acute temperature reduction to 20 degrees C. In gambusia, both salinity and temperature acute stress strongly stimulated urea excretion. Routine oxygen consumption was barely affected by acute salinity or temperature stress, and was reduced by the combined effects of temperature and high salinity. Gills maintained their structural integrity in all stressing conditions; hyperplasia and hypertrophy of mitochondria-rich cells were observed. In zebrafish, temperature and salinity acute changes, both alone and in combination, scarcely affected any parameter tested. The major effect observed was a reduction of nitrogen excretion at 20 degrees C-25 per thousand; under these extreme conditions a significant structural disruption of gills was observed. These results confirm the high tolerance to acute salinity and temperature stress in gambusia, and demonstrate the involvement of urea excretion modulation in the stress response in this species.

15-week long school-based nutritional education program to promote Italian primary schoolchildren's fruit and vegetable intake.

A short-version (15-week long) of school-based nutrition education program called "Bring Fruit to School" to enhance primary schoolchildren's fruit and vegetable (F&V) intake was evaluated. We recruited 199 primary schoolchildren from 8 classes (grade 2-5). The primary end-point was an increase in the children's F&V intake. The intervention period lasted 15 weeks, and was divided into three phases: weeks 1-5, weeks 6-10 and weeks 11-15. The F&V intake of the schoolchildren was also monitored in a subsequent follow-up period (16th-32nd week). By week 15, 92 (46.2%, P < .001) schoolchildren increase fruit intake and 91 (45.7%, P < .001) increase vegetable intake. The F&V intake increased during the intervention period and was constant in follow-up. The BFtS program is an effective means to promote primary schoolchildren's F&V intake, and was implemented in Italy by the Food and Nutrition Service of Foggia's Local Health Authority. It could be extended to many other countries by local nutrition agencies, such as associations for nutrition.

A mycobacterial disease is associated with the silent mass mortality of the pen shell Pinna nobilis along the Tyrrhenian coastline of Italy.

Disease is an increasing threat for marine bivalves worldwide. Recently, a mass mortality event (MME) impacting the bivalve Pinna nobilis was detected across a wide geographical area of the Spanish Mediterranean Sea and linked to a haplosporidian parasite. In 2017-2018, mass mortality events affecting the pen shell Pinna nobilis were recorded in two different regions of Italy, Campania and Sicily, in the Tyrrhenian Sea (Mediterranean Sea). Histopathological and molecular examinations of specimens showed the presence of Haplosporidium sp. in only one specimen in one area. Conversely, in all of the surveyed moribund animals, strong inflammatory lesions at the level of connective tissue surrounding the digestive system and gonads and linked to the presence of intracellular Zhiel-Neelsen-positive bacteria were observed. Molecular analysis of all of the diseased specimens (13) confirmed the presence of a Mycobacterium. Blast analysis of the sequences from all of the areas revealed that they were grouped together with the human mycobacterium M. sherrisii close to the group including M. shigaense, M. lentiflavum and M. simiae. Based on pathological and molecular findings, it is proposed that a mycobacterial disease is associated with the mortality episodes of Pinna nobilis, indicating that, at this time, Haplosporidium sp. is not responsible for these events in Campanian and Sicilian waters.

Lessons from 8 years' experience of hypothalamic stimulation in cluster headache.

Neuroimaging studies in cluster headache (CH) patients have increased understanding of attack-associated events and provided clues to the pathophysiology of the condition. They have also suggested stimulation of the ipsilateral posterior inferior hypothalamus as a treatment for chronic intractable CH. After 8 years of experience, stimulation has proved successful in controlling the pain attacks in almost 60% of chronic CH patients implanted at various centres. Although hypothalamic implant is not without risks, it has generally been performed safely. Implantation affords an opportunity to perform microrecordings of individual posterior hypothalamic neurons. These studies are at an early stage, but suggest the possibility of identifying precisely the target site by its electrophysiological characteristics. Autonomic studies of patients undergoing posterior hypothalamic stimulation provide further evidence that long-term stimulation is safe, revealing that it can cause altered modulation of the mechanisms of orthostatic adaptation without affecting the baroreflex, cardiorespiratory interactions or efferent sympathetic and vagal functions. Chronically stimulated patients have an increased threshold for cold pain at the site of the first trigeminal branch ipsilateral to the stimulated side; when the stimulator is switched off, changes in sensory and pain thresholds do not occur immediately, suggesting that long-term stimulation is required to induce sensory and nociceptive changes. Posterior inferior hypothalamic stimulation is now established as a treatment for many chronic CH patients. The technique is shedding further light on the pathophysiology of the disease, and is also providing clues to functioning of the hypothalamus itself.

Histological, histochemical and morphometric changes of splenic melanomacrophage centers (SMMCs) in Sparicotyle-infected cultured sea breams (Sparus aurata).

Diseases caused by parasites are much more frequently described in cultured fish, which suffer from artificial conditions and numerous stress factors. This study investigates the histological, histochemical and morphometric modifications of splenic melanomacrophage centers (SMMCs) infected by Sparicotyle chrisophrii (Monogenea, ectoparasite of the gills) in sea breams (Sparus aurata), cultured in floating cages in the Gulf of Gaeta (Italy). Infected fish swam near the water surface, showing severe signs of anemia. Several spleens were collected from both healthy and dead fish (70-100 gr. body weight). A spleen histopathology was evaluated by using traditional stainings, such as Haematoxylin and Eosin (HE), Periodic Acid-Schiff reaction (PAS), Perl's reaction for haemosiderin and Schmorl's reaction for lipofuscins. Furthermore, SMMCs morphometry was performed on PAS-stained sections to study 7 morphometric parameters [Mean SMMCs profile area (MPA), Mean SMMCs maximum diameter (Media), Mean SMMCs minimum diameter (media), Mean SMMCs diameter (Dia), Mean SMMCs Perimeter (P), Mean SMMCs Form Factor (FF) and Mean SMMCs number per square millimeter of spleen tissue (MN)]. A light microscope of HE stained sections of spleen revealed a dramatic increase in the size and number of SMMCs in parasitized animals. Morphometric data illustrated statistically significant differences (p < 0.01) of all studied parameters between healthy and diseased fish. This study emphasizes the importance of using histopathological investigations to unravel the complex biological host/parasite interaction, which results in systemic lesions affecting reared marine species.

[Prevalence of upper limb work-related musculoskeletal disorders (UL-WMSDs) in workers of the upholstered furniture industry]. / Prevalenza delle patologie da sovraccarico biomeccanico degli arti superiori negli addetti del comparto del mobile imbottito.

BACKGROUND: The upholstered furniture industry, the so-called "triangle of the sofa industry", is a geographic area of national and strategic economic importance in southern Italy. The single tasks are carried out mostly manually, with the characteristics of a handicraft approach. OBJECTIVES: The aim of the survey was to assess the prevalence of upper limb work-related musculoskeletal disorders (UL-WMSDs) in 30 factories of the sofa industry located in a large geographic area of the Puglia and Basilicata Regions. METHODS: In the period 1 January-31 December 2003 a network of occupational physicians investigated a population of 5.477 subjects (exposed n=3481, controls n=1996, M=3865, F=1612) in 30 different factories of the area. More than 60 percent of the total workforce studied was employed in large-sized companies (>500 employees). The following work tasks were considered: filling preparation workers, leather-cutting operators, sewing and upholstery-assembly workers. Case-definition was assessed through standardized procedures: symptoms by questionnaire plus physical and laboratory/imaging findings. RESULTS: Cumulative prevalence rates of UL-WMSDs as at 31 December 2003 reached values of up to 30% in high risk groups. Prevalence rates showed good correlation with the concise OCRA index used for assessment of exposure to repetitive strain and movements of the upper limb. The most frequently occurring disorders were tendon-related cysts and wrist tendonitis. Shoulder disorders were more frequent in male and female leather-cutting operators. CONCLUSIONS: This survey showed a significantly high prevalence of UL-WMSDs in sofa industry workers. It did not seem to be confirmed in this study that there was a greater female susceptibility to UL-WMSDs with the exception of carpal tunnel syndrome: gender difference seems to be less relevant at increasing levels of occupational exposure to repetitive movements and exertion of the upper limbs.

Imaging degeneration of the substantia nigra in Parkinson disease with inversion-recovery MR imaging.

BACKGROUND AND PURPOSE: Visualizing with MR imaging and obtaining quantitative indexes of degeneration of the substantia nigra in Parkinson disease have been long-sought goals. We investigated the potential role of area and T1 contrast measurements in differentiating patients from controls and their age-related changes. METHODS: Eight patients with Parkinson disease, 8 age-matched controls, and 8 young controls were imaged. We obtained the pixel-wise difference between 2 sets of inversion-recovery images, acquired parallel to the bicommissural plane, with different inversion times. Pixel-intensity ratios between lateral and medial nigral regions, and nigral area and substantia-nigra/midbrain area ratios were computed. RESULTS: Compared with that of controls, loss of substantia nigra was evident in patients, its borders taking a smoother and more irregular appearance. Patients were characterized by a lateral-to-medial gradient, due to reduced hypointensity of the lateral portion of the substantia nigra and relative sparing of its medial portion. The visible nigral area was significantly smaller in patients compared with matched controls (P = .04). The substantia nigra/midbrain area ratio enabled considerably better separation (P = .0001). The lateral/medial pixel-intensity ratio was significantly higher in patients compared with matched controls (P = .01) and in young controls compared with age-matched controls (P = .01). CONCLUSION: Inversion-recovery sequences may provide a convenient way to visualize nigral degeneration. Relative area and pixel-intensity measurements may integrate other techniques (such as diffusion-tensor imaging on nigrostriatal pathways) in the neuroradiologic diagnosis and follow-up of Parkinson disease by quantitatively assessing the degeneration of the substantia nigra.

Omalizumab and allergen immunotherapy for respiratory allergies: A mini-review from the Allergen-Immunotherapy Committee of the Italian Society of Pediatric Allergy and Immunology (SIAIP)

Although currently approved to treat severe asthma and chronic spontaneous urticaria, omalizumab has also been an effective and safe add-on treatment for other allergic diseases. Namely, omalizumab has been proposed to be used as add-on therapy in patients with allergic rhinitis and asthma and undergoing specific allergen immunotherapy (AIT). AIT is the only treatment that modifies the natural history of IgE-mediated diseases. This brief review summarizes the available evidence and controversies on the efficacy and safety of omalizumab combined with specific AIT (AU)

Frontotemporal dementia and corticobasal degeneration in a family with a P301S mutation in tau.

The tau gene has been found to be the locus of dementia with rigidity linked to chromosome 17. Exonic and intronic mutations have been described in a number of families. Here we describe a P301S mutation in exon 10 of the tau gene in a new family. Two members of this family were affected. One individual presented with frontotemporal dementia, whereas his son has corticobasal degeneration, demonstrating that the same primary gene defect in tau can lead to 2 distinct clinical phenotypes. Both individuals developed rapidly progressive disease in the third decade. Neuropathologically, the father presented with an extensive filamentous pathology made of hyperphosphorylated tau protein. Biochemically, recombinant tau protein with the P301S mutation showed a greatly reduced ability to promote microtubule assembly.

Factors affecting course and survival in Alzheimer's disease. A 9-year longitudinal study.

OBJECTIVE: To evaluate mean survival and to identify prognostic factors in a cohort of patients with Alzheimer's disease (AD). DESIGN: Multicentric 9-year cohort analytic study. SETTING: Seven neurology departments throughout Italy between April 1982 and January 1984. PATIENTS: We recruited a consecutive sample of 145 patients affected by probable AD (Multicenter Italian Study on Dementia protocol, National Institute of Neurological Disorders and Stroke-Alzheimer's Disease and Related Disorders Association criteria). Five were misdiagnosed, and 21 could not participate in the longitudinal study. The clinicodemographic characteristics of the 119 enrolled patients (49 men, 70 women; mean age, 64.7 years; SD, 4.1 years; mean duration of disease, 3.1 years; SD, 1.8 years) did not differ from those of the 26 excluded patients. All underwent extensive cliniconeuropsychological testing every 6 months for at least 2 years until the patient died or our survey ended (April 30, 1991). Mean follow-up was 5.1 years (SD, 2.5 years). MAIN OUTCOME MEASURES: Death, severe functional impairment (a score > or = 17 on the Blessed Dementia Scale), and severe cognitive impairment (a score of < or = 7 on the Information-Memory-Concentration Test). RESULTS: Survival curves obtained by the Kaplan-Meier method indicated that (1) patients with early- and late-onset disease (ie, before or after age 65 years) showed no difference either in relative survival or in time to reach predetermined functional and cognitive end points; (2) severely aphasic patients became profoundly demented significantly sooner than those with mild to moderate aphasia (P < .0001). Among clinicodemographic variables analyzed by a Cox model, severe language disability and functional loss proved to be the best predictors of death independent of age at onset or degree of dementia. CONCLUSIONS: Age at onset did not influence course and survival in AD. Severe aphasia appears to be the best predictor of death and unfavorable course.