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Funding: MFB’s participation at the meeting of the ACCOBAMS-ASCOBANS Joint Bycatch Working Group was in the framework of the Life DELFI project, co-financed by the European Community under the LIFE programme. Information on movements and connectivity among populations of animals is important for the delineation of units to conserve, so that demographic parameters, such as abundance, fecundity and mortality, can be placed in an appropriate population and conservation context. Common bottlenose dolphins (Tursiops truncatus) are often considered relatively ‘resident’ and demonstrating strong site fidelity to specific areas. However, this perception may partly be an artefact of the distribution and ‘habitat use’ of cetacean researchers, rather than animals themselves, and bottlenose dolphins have been shown to be capable of substantial movements, often in relatively short periods of time. Here, we report on two long-distance movements of a common bottlenose dolphin within the Mediterranean Sea, across the Tyrrhenian, Ionian and Adriatic Seas, and subsequently back across all three seas to Ligurian Sea, making these the two longest recorded movement for this species in the Mediterranean Sea to date and some of the longest in the world. We also review published records of long-distance movements in this species worldwide. This study highlights the utility of photo-identification and the importance of regional data sharing. We argue that photo-identification comparisons are always worthwhile and the results are informative regardless of the presence or absence of matches, especially with the ongoing advances in automated matching software. Publisher PDF

The loggerhead sea turtle, Caretta caretta (Linnaeus, 1758), is the most common sea turtle species in the Mediterranean Sea, where it can experience severe anthropogenic impacts. Although C. caretta is known to host more than 200 epibiotic taxa (crustaceans, algae and cyanobacteria), no reports have included a detailed evaluation of the microbial community of its carapace scutes. Thus, this study aimed to determine the diversity and composition of the visible and invisible communities on the carapace scutes of wild loggerhead turtles from the Aeolian Archipelago (Southern Tyrrhenian Sea, Italy) by using a combined approach of morphological/spatial examination and molecular analyses. Altogether, our results displayed a higher abundance of crustaceans, macroalgae and Proteobacteria on the posterior carapace scutes, while Firmicutes were more abundant on the anterior scutes. For the first time, this study showed the complexity of the microbial (invisible) and visible epibionts of the loggerhead sea turtles from the Mediterranean Sea and suggests the importance of including evaluation of the microbial components when studying epibiont communities.

The reproductive output of wild bottlenose dolphinsTursiops truncatushas extensively been investigated; nevertheless, it remains widely unexplored in the Mediterranean Sea. We investigated the reproductive history of female bottlenose dolphins in the Aeolian Archipelago (Italy). Thirteen years of photo-identification data were used to track the reproductive pattern of 7 females and their 16 calves. The birth period was accurately estimated within 1 mo for 6 calves (37.5%), peaking in June/July. The mean number of calves for females was 2.3 ± 1.1 (SD), ranging from 1 to 4. The mean interbirth interval was 3.5 ± 1.6 yr, ranging from 2 to 7 yr. The mean weaning age was 3.2 ± 0.6 yr, although 1 calf may have been weaned as late as its fifth birthday. Mother-calf postweaning associations were tracked for 9 calves and ranged from 4 to 10 yr of residence in the maternal group of the calf. The age of first pregnancy was 7.5 yr. The number of calves was influenced by the mean maternal group size, with females that were sighted in larger groups having more calves. However, the number of calves was not influenced by the strength of the associations of mothers with other females. Calf mortality by age 1 was high (12.5%). Two juveniles also died postweaning within 6 yr of their birth. Finally, 2 females may have died during the study, as they were not re-sighted at the end of the study. Our results highlight the need to implement urgent conservation actions to protect this bottlenose dolphin population.

The ingestion of anthropogenic plastic debris by marine wildlife is widespread in the Mediterranean Sea. The endangered status (in the IUCN Red List) of Loggerhead turtle (Caretta caretta, Linnaeus, 1758) is a consequence of its vulnerability. In this study, macro-/meso-plastics (5-170 mm) collected from faeces of twelve loggerhead turtles rescued (live) in the Aeolian Archipelago (Southern Tyrrhenian Sea, Italy) were analyzed by size, weight, shape, color and polymer type through Attenuated Total Reflectance Fourier-Transform Infrared Spectroscopy (ATR-FTIR). The defecation rate during hospitalization (7-14 days) varied among turtles (from 0.08 to 0.58). The mean number of plastic expulsions (2.7 ± 1.8 items for turtle) was higher during the 5th day of hospitalization (Kruskal-Wallis test, P = 0.01). However, the mean number of plastic-like items defecated during the common days of hospitalization did not vary among turtles (Kruskal-Wallis test, P 0.05). All turtles were found to have ingested plastic. A total of 114 debris items were recovered from their faeces, 113 of which were identified as plastic. Their color was mostly white-transparent (64.9%) and light (19.3%). Shape was mainly fragments (52.6%), sheets (38.6%), followed by nylon, net-fragments, elastic plastic, foamed plastic and industrial granules (8.8%). Meso-plastics (5-25 mm) represented 72% of the total number of debris and were found more frequently in turtle with Curved Carapace Length (CCL) ≤ 60 cm (CCL = 30-60 cm, n = 5) than those with CCL60 cm (CCL = 60-71 cm, n = 7). Plastic items were composed mainly of polyethylene (48.2%) and polypropylene (34.2%). Polypropylene (R

Phthalic acid esters (PAEs) are classified as endocrine disruptors, but it remains unclear if they can enter the marine food-web and result in severe health effects for organisms. Loggerhead turtles (Caretta caretta) can be chronically exposed to PAEs by ingesting plastic debris, but no information is available about PAEs levels in blood, and how these concentrations are related to diet during different life stages. This paper investigated, for the first time, six PAEs in blood of 18 wild-caught Mediterranean loggerhead turtles throughout solid-phase extraction coupled with gas chromatography-ion trap/mass spectrometry. Stable isotope analyses of carbon and nitrogen were also performed to assess the resource use pattern of loggerhead turtles. DEHP (12-63 ng mL

The presence of phthalic acid esters (PAEs) in marine environments is an important issue. These chemicals are able to affect marine organisms, particularly marine turtles, and to act as endocrine disrupters. In this paper, for the first time, a simple and reproducible analytical method based on solid-phase extraction (SPE) coupled with gas chromatography—ion trap/mass spectrometry (GC-IT/MS) was developed for the extraction of phthalates from the blood of marine turtles. The extraction was obtained by using C18 phthalates-free as the stationary phase. In order to individuate the best working conditions for the extraction, the adsorption isotherms and breakthrough curves were studied. The overall analytical methodology was validated in terms of limit of detection (LOD, 0.08–0.6 ng mL−1), limit of quantification (LOQ, 0.4–0.8 ng mL−1), and correlation coefficients (>0.9933). By using this procedure, percentage recoveries ranging from 89 to 103% were achieved. The precision parameters (intra-day and inter-day) were studied, and the obtained values were smaller than 12.5%. These data confirm the goodness of the proposed analytical methodology, which is applied to real samples.

Annual and seasonal encounter rates, life stages and the main threats to loggerhead sea turtles (Caretta caretta) in the Aeolian Archipelago (southern Italy) were studied. Dedicated boat surveys resulted in 258 surface observations and 138 captures of healthy (n = 309), ailing (n = 66) and dead (n = 21) turtles from 2009 to 2013. Loggerheads were encountered at the sea water surface while resting (87%) or feeding on pelagic prey (13%). The loggerhead encounter rate (observations per km surveyed) was higher during the autumn, suggesting the presence of potential foraging/overwintering habitats in the area. The mean (± SD) curved carapace length (CCL) was 48.8 ± 10.7 cm, with 65% of the individuals ranging from 40–70 cm in size. Smaller turtles were encountered more frequently during the spring months, probably as a result of the abundant pelagic prey within the coastal area. During the colder season, larger turtles were more common in the afternoon than in the morning, suggesting that the time needed for rewarming might increase with turtle size. Ingestion of anthropogenic debris was reported in 48.5% of the rescued turtles. Individual mortality was mainly related to longline fishing (70.6%), with debris entanglements/ingestion frequently associated with these records. Longline bycatch and boat collisions were higher in summer, whereas debris ingestion was highest in spring. Different threats might affect particular life stages because the longline bycatch was more frequent for larger turtles, whereas boat collisions were more frequent with smaller individuals. Migratory patterns, habitat characteristics and seasonal changes in sea temperature and currents might influence the seasonal occurrence of loggerhead turtles in this area. These results increase the current ecological knowledge of the factors driving loggerhead turtle life and are important for implementing management plans for its conservation in the Mediterranean Sea.

Skin marks occur frequently in many cetacean species across the globe revealing a broad spectrum of causes, including social interactions, infectious diseases and injuries produced by anthropogenic factors. The current study used photo-id data from 2005-2014 to estimate the skin mark pattern on resident bottlenose dolphins (Tursiops truncatus) from the Aeolian Archipelago (Italy). Thirteen skin mark types were identified and their origin, prevalence and permanence time were examined. The pattern of skin marks was assessed for the abundance, richness, distribution and severity in six body regions and compared among age classes, sex and degree of dolphins' interaction with trammel nets (DIN). Our results showed higher prevalence, abundance, richness and distribution of skin marks in adults than in the younger age classes, with the exception of black marks and white ring lesions. The prevalence and abundance of skin marks were higher in males than females, with the exception of scratches and white patches. Moreover, gunshot wounds, mutilations and irregular dorsal fin edges were found only on adult males. Since males showed higher DIN than females and, in dolphins with higher DIN, skin marks were more abundant and frequently distributed in different body regions, the skin mark pattern in regard to DIN seems to be sex-related. The more severe marks were observed on adults, males and dolphins with higher DIN, namely skin disorder, tooth rake marks, small shallow indentations, deep indentations and mutilations. On the contrary, the severity of scratches, white patches and dark ring lesions was higher in females than males, but not significantly related to DIN and age of the individuals. Our results showed that photo-id data provide an efficient and cost-effective approach to document the occurrence of skin marks in free-ranging bottlenose dolphin populations, a critical step toward understanding the cause and supporting the conservation strategies.

Climatic changes are associated with heat waves, droughts, and floods which have significant impacts on human health. Floods enhance the effects of water-borne pathogens increasing the concentration of biological agents in surface water. On the basis of national statistics collected by the Italian Ministry of Health, this work illustrates preliminary observations concerning outbreaks of water-borne diseases after flood events in Italy and presents options for an administrative response. The water-borne infectious diseases taken into account are Legionellosis, Salmonellosis, Hepatitis A, cutaneous and visceral Leishmaniasis, Leptospirosis, and Infectious Diarrhea. An association between these infectious diseases and flood events seems to exist in Italy, although further analysis should be carried out to confirm this relationship. Flood impacts can be distinguished into: immediate, medium term, and long term. Several factors are involved in determining the intensity and the severity of effects. Some vector-...

We describe a new approach to investigate alterations in the human MLH1 mismatch repair (MMR) gene. This is based on complementation of the phenotype of a MLH1-defective subclone of the ovarian carcinoma A2780 cells by transfection of vectors encoding altered MLH1 proteins. Measurements of resistance (tolerance) to methylating agents, mutation rate at HPRT, microsatellite instability (MSI), and steady-state levels of DNA 8-oxoguanine were used to define the MMR status of transfected clones. The approach was validated by transfecting cDNA of wild-type (WT) MLH1, cDNAs bearing two previously identified polymorphisms (I219V and I219L) and two with confirmed hereditary nonpolyposis colorectal cancer (HNPCC) syndrome mutations (G224D and G67R). A low-level expression of two MLH1 polymorphisms partially reversed methylation tolerance and the mutator phenotype, including MSI. Higher levels of I219V resulted in full restoration of these properties to WT. Increased expression of I129L did not fully complement the MLH1 defect, because there was a simultaneous escalation in the level of oxidative DNA damage. The findings confirmed the important relationship between deficient MMR and increased levels of oxidative DNA damage. Mutations from Italian HNPCC families (G224D, G67R, N635S, and K618A) were all ineffective at reversing the phenotype of the MLH1-defective A2780 cells. One (K618A) was identified as a low penetrance mutation based on clinical and genetic observations. (Cancer Res 2006; 66(18): 9036-44)

Oxidation is a common form of DNA damage to which purines are particularly susceptible. We previously reported that oxidized dGTP is potentially an important source of DNA 8-oxodGMP in mammalian cells and that the incorporated lesions are removed by DNA mismatch repair (MMR). MMR deficiency is associated with a mutator phenotype and widespread microsatellite instability (MSI). Here, we identify oxidized deoxynucleoside triphosphates (dNTPs) as an important cofactor in this genetic instability. The high spontaneous hprt mutation rate of MMR-defective msh2(-/-) mouse embryonic fibroblasts was attenuated by expression of the hMTH1 protein, which degrades oxidized purine dNTPs. A high level of hMTH1 abolished their mutator phenotype and restored the hprt mutation rate to normal. Molecular analysis of hprt mutants showed that the presence of hMTH1 reduced the incidence of mutations in all classes, including frameshifts, and also implicated incorporated 2-oxodAMP in the mutator phenotype. In hMSH6-deficient DLD-1 human colorectal carcinoma cells, overexpression of hMTH1 markedly attenuated the spontaneous mutation rate and reduced MSI. It also reduced the incidence of -G and -A frameshifts in the hMLH1-defective DU145 human prostatic cancer cell line. Our findings indicate that incorporation of oxidized purines from the dNTP pool may contribute significantly to the extreme genetic instability of MMR-defective human tumors.

Previous studies from our laboratory demonstrated the existence of at least two levels of structural complexity in E. coli 70S ribosomes. Ribosomal RNA seems to be principally involved in the overall stability of these structures. In this paper we present an investigation of ribosomes subjected to treatment with RNase. The study is based on both differential scanning microcalorimetry and dielectric spectroscopy. In the thermograms obtained on treated ribosomes only the low temperature peak of the two typical denaturation events observed in native ribosomes, is promptly eliminated by the enzyme treatment. Dielectric spectroscopy measurements carried out on the same samples indicate an alteration of the dielectric behavior previously shown to consist of two subsequent relaxation processes. In fact, only the low frequency relaxation is affected by the treatment. The second one, observed at higher frequency, remains unaltered. The same effect on the dielectric parameters is observed if the ribosome particles are heated and then cooled prior to measurement. These results are consistent with the idea that two different structures are present within the ribosome. One is very stable and withstands both temperature and RNase treatment while the second is promptly abolished by both treatments. Data presented here strongly suggest that the RNA domains exposed to the solvent play a fundamental role in the stability of the 3-D structure of the ribosome particle.

Modelling cetacean distributions is crucial to understanding their ecology and to relating use patterns to environmental changes. In the present study, a combination of statistical methods was applied to model the distribution of bottlenose dolphin Tursiops truncatus with 18 physiographic variables around the Island of Filicudi, southern Italy. Principal components and clustering analyses were used to describe the habitat structure derived from mutually correlated predictor variables. Multivariate regression and canonical correlation analyses were used to find critical habitats and core use areas by combining the contribution of 2 response variables: the encounter rate and an index of use calculated according to the spatial behaviour of the dolphin groups. Three critical habitats were identified as distinct combinations of physiographic vari- ables at a 1 km 2 spatial scale. Two of these were associated with opportunistic and natural feed- ing activities. A third, a highly variable topographic habitat located in shallow waters at greater distance from the coastline, appears to serve as a resting/calving habitat by providing defence from anthropogenic pressures. The analysis also estimated an 80% shift from feeding to resting habitats associated with physiographic changes. Since the bottlenose dolphin encounter rate has decreased in recent years, the identification of core areas is useful in the preparation of local marine protected areas for the Aeolian archipelago. This analytical approach to studying dolphin−habitat relationships is relevant for conservation planning as it shows how environmen- tal variability can modify dolphin distribution on a local scale according to the response variables relevant to the species.

Water-related diseases continue to cause a high burden of mortality and morbidity in the countries of the European Region. Parties to the Protocol on Water and Health are committed to the sustainable use of water resources, the provision of safe drinking water and adequate sanitation to all people of the European Region, and to the reduction of the burden of water-related diseases. A specialized Task Force is implementing a work plan aimed at strengthening the capacity for water-related disease surveillance, outbreak detection and contingency planning. Parties to the Protocol are obliged to set targets, and report on progress on water-related disease surveillance. The present paper aims to provide a baseline assessment of national capacities for water-related disease surveillance on the basis of the replies to a questionnaire. This was prepared in English and Russian and administered to 53 countries, 15 of which replied. The results confirm the heterogeneity in surveillance systems, the weakness of many countries to adequately survey emerging water-related diseases, and the need for specific remedial action. The findings of the exercise will form the basis for future action under the Protocol on Water and Health.

Gastric cancers with mismatch repair (MMR) inactivation are characterised by microsatellite instability (MSI). In this study, the transcriptional profile of 38 gastric cancers with and without MSI was analysed. Unsupervised analysis showed that the immune and apoptotic gene networks efficiently discriminated these two cancer types. Hierarchical clustering analysis revealed numerous gene expression changes associated with the MSI phenotype. Amongst these, the p53-responsive genes maspin and 14-3-3 sigma were significantly more expressed in tumours with than without MSI. A tight immunosurveillance coupled with a functional p53 gene response is consistent with the better prognosis of MSI cancers. Frequent silencing of MLH1 and downregulation of MMR target genes, such as MRE11 and MBD4, characterised MSI tumours. The downregulation of SMUG1 was also a typical feature of these tumours. The DNA repair gene expression profile of gastric cancer with MSI is of relevance for therapy response.

Water related disease outbreak (WRDO) statistics in Italy from 1998 to 2005 have been discussed in this paper. The true incidence of WRDO is not reflected in the National Surveillance System (NSS), although this study has provided information on pathogens associated to different water sources, incidence in Regions and inadequacy of regulations. 192 outbreaks and 2546 cases of WRD were reported to the NSS, an average of 318 cases per year. Cases were associated to shellfish (58.79%), drinking water (39.94%) and agricultural products (1.25%). WRDs have been detected in 76% of Regions: central and southern Regions showed lower percentage of cases (35.4%) due to under-reporting. Most of WRD cases in the North were related to drinking water; WRDs in marine coastal Regions were mostly related to shellfish. 49% of Districts (Province) notified WRDs, including only 101 Municipalities. Pathogenic microorganisms were identified in a few cases from clinical investigations. They included enteric viruses, Norwalk viruses, Salmonella, Shigella, Giardia and Campylobacter. There is the need to improve the existing NSS in relation to WRDs. An adequate WRDs Surveillance System should be based on connection between health and environmental authorities, priority pathogens and critical areas identification, response capability and contingency plans.

Acute promyelocytic leukemia (APL) is a clonal expansion of hematopoietic precursors blocked at the promyelocytic stage. Gene expression profiles of APL cells obtained from 16 patients were compared to eight samples of CD34+-derived normal promyelocytes. Malignant promyelocytes showed widespread changes in transcription in comparison to their normal counterpart and 1020 differentially expressed genes were identified. Discriminating genes include transcriptional regulators (FOS, JUN and HOX genes) and genes involved in cell cycle and DNA repair. The strong upregulation in APL of some transcripts (FLT3, CD33, CD44 and HGF) was also confirmed at protein level. Interestingly, a trend toward a transcriptional repression of genes involved in different DNA repair pathways was found in APL and confirmed by real-time polymerase chain reactor (PCR) in a new set of nine APLs. Our results suggest that both inefficient base excision repair and recombinational repair might play a role in APLs development. To investigate the expression pathways underlying the development of APL occurring as a second malignancy (sAPL), we included in our study eight cases of sAPL. Although both secondary and de novo APL were characterized by a strong homogeneity in expression profiling, we identified a small set of differentially expressed genes that discriminate sAPL from de novo cases.

The activity of the cell is often coordinated by the organisation of proteins into regulatory circuits that share a common function. Genome-wide expression profiles might contain important information on these circuits. Current approaches for the analysis of gene expression data include clustering the individual expression measurements and relating them to biological functions as well as modelling and simulation of gene regulation processes by additional computer tools. The identification of the regulative programmes from microarray experiments is limited, however, by the intrinsic difficulty of linear methods to detect low-variance signals and by the sensitivity of the different approaches. Here we face the problem of recognising invariant patterns of correlations among gene expression reminiscent of regulation circuits. We demonstrate that a recursive neural network approach can identify genetic regulation circuits from expression data for ribosomal and genome stability genes. The proposed method, by greatly enhancing the sensitivity of microarray studies, allows the identification of important aspects of genetic regulation networks and might be useful for the discrimination of the different players involved in regulation circuits. Our results suggest that the constitutive regulatory networks involved in the generic organisation of the cell display a high degree of clustering depending on a modular architecture.

We investigated social structure and association patterns for a small population of Mediterranean bottlenose dolphins, Tursiops truncatus, inhabiting the Aeolian Archipelago (southern Italy). Specifically we evaluate the role of sex and age composition, residency patterns and interaction with trammel nets on this social organization. Association data for 23 regularly sighted individuals were obtained from summer photoidentification surveys collected from 2005-2012. Using a combined cluster and social network analysis approach, we found associations between dolphins were hierarchically structured, where two mixed-sex social units were subdivided into smaller temporarily dynamic groups. We found non-random and long-term preferred associations in the population; however, the degree of social cohesion, residence pattern and interaction with trammel nets differed considerably between the two social units. Six of eight females occurred in the more resident social unit-1; in addition, social unit-1 individuals had significantly stronger associations, higher preferred associates, lived in larger groups and occurred less frequently with trammel nets. Nine of eleven males were clustered in social unit-2 and five of these males, interacting with trammel nets, formed small groups and preferred associations. We propose that female and male groups associate in the study area during the breeding season and that some males choose to interact with reproductive females forming a distinct but interrelated social unit. Other males may be associating in a larger fission-fusion network, which consists of dolphins that appear to temporarily join the network from the coastal population. We cannot exclude that some males specialized in trammel net foraging, suggesting that this foraging technique may favor a solitary lifestyle. Large group sizes and high degree of social cohesion for females could be an indication of greater protection and more efficiency in detecting, deterring or repelling anthropogenic pressures. Most likely dolphins' social organization depends on a combination of socio-ecological, demographic and anthropogenic factors.