Influence of pancreatic status on circulating plasma sterols in patients with cystic fibrosis.

Background Patients with cystic fibrosis (CF) have a reduced intestinal absorption of cholesterol and in a preliminary study we observed differences in plasma sterol profile between patients with pancreatic sufficiency (PS) and those with pancreatic insufficiency (PI). Therefore, we hypothesized that the sterol analysis may contribute to study the digestion and absorption state of lipids in patients with CF. To this aim we evaluated plasma sterols in a significant number of adult patients with CF in relation to the pancreatic status. Methods Beside cholesterol, we measured phytosterols and lathosterol as markers of intestinal absorption and hepatic biosynthesis, respectively, by gas-chromatography in plasma of adult CF patients with pancreatic sufficiency (PS-CF, n = 57), insufficiency (PI-CF, n = 97) and healthy subjects (control group, CT, n = 71). Results PI-CF patients had cholesterol and phytosterols levels significantly lower than PS-CF and CT (p < 5 × 10-10) suggesting a reduced intestinal absorption of sterols related to PI. Instead, lathosterol was significantly higher in PI-CF patients than PS-CF and CT (p < 0.0003) indicating an enhanced cholesterol biosynthesis. In PI-CF patients, phytosterols positively correlate with vitamin E (p = 0.004). Both the classes of molecules need cholesterol esterase for the intestinal digestion, thus the reduced levels of such lipids in serum from PI-CF patients may depend on a reduced enzyme activity, despite the pancreatic enzyme supplementation in all PI-CF patients. Conclusions A plasma sterols profile may be useful to evaluate the metabolic status of lipids in adult patients with CF and could help to manage the pancreatic enzyme supplementation therapy.

Disinfection by-Products and Ecotoxic Risk Associated with Hypochlorite Treatment of Tramadol.

In recent years, many studies have highlighted the consistent finding of tramadol (TRA) in the effluents from wastewater treatment plants (WTPs) and also in some rivers and lakes in both Europe and North America, suggesting that TRA is removed by no more than 36% by specific disinfection treatments. The extensive use of this drug has led to environmental pollution of both water and soil, up to its detection in growing plants. In order to expand the knowledge about TRA toxicity as well as the nature of its disinfection by-products (DBPs), a simulation of the waste treatment chlorination step has been reported herein. In particular, we found seven new by-products, that together with TRA, have been assayed on different living organisms (Aliivibrio fischeri, Raphidocelis subcapitata and Daphnia magna), to test their acute and chronic toxicity. The results reported that TRA may be classified as a harmful compound to some aquatic organisms whereas its chlorinated product mixture showed no effects on any of the organisms tested. All data suggest however that TRA chlorination treatment produces a variety of DBPs which can be more harmful than TRA and a risk for the aquatic environment and human health.

The principles of Health Technology Assessment in laboratory medicine.

The Health Technology Assessment (HTA) is a multi-professional and multidisciplinary evaluation approach designed to assess health technology in the broadest sense of the term, from its instruments to the rearranging of its organizational structures. It is by now an established methodology at national and international levels that involves several medical disciplines thanks to its versatility. Laboratory medicine is one of these disciplines. Such specialization was subjected, in recent years, to deep changes even from an organizational standpoint, in order to meet the health needs of the population, making them as effective and cost-effective as possible. In this regard, HTA was the tool used to assess implications in different areas.

Twelve Novel Mutations in the SLC26A3 Gene in 17 Sporadic Cases of Congenital Chloride Diarrhea.

OBJECTIVES: We aimed to improve the knowledge of pathogenic mutations in sporadic cases of congenital chloride diarrhea (CCD) and emphasize the importance of functional studies to define the effect of novel mutations. METHODS: All member 3 of solute carrier family 26 (SLC26A3) coding regions were sequenced in 17 sporadic patients with CCD. Moreover, the minigene system was used to analyze the effect of 2 novel splicing mutations. RESULTS: We defined the SLC26A3 genotype of all 17 patients with CCD and identified 12 novel mutations. Using the minigene system, we confirmed the in silico prediction of a complete disruption of splicing pattern caused by 2 of these novel mutations: the c.971+3_971+4delAA and c.735+4_c.735+7delAGTA. Moreover, several prediction tools and a structure-function prediction defined the pathogenic role of 6 novel missense mutations. CONCLUSIONS: We confirm the molecular heterogeneity of sporadic CCD adding 12 novel mutations to the list of known pathogenic mutations. Moreover, we underline the importance, for laboratories that offer molecular diagnosis and genetic counseling, to perform fast functional analysis of novel mutations.

The relevance of prothrombotic genetic variants in women who experienced pregnancy loss or embryo implantation failure: A retrospective analysis of 1922 cases.

OBJECTIVE: The aim of our study was that to assess the allelic and genotype frequencies of nine prothrombotic gene variants in patients with a history of pregnancy loss and recurrent pregnancy loss (RPL). Women who underwent assisted reproductive technology (ART) with ongoing pregnancy and those with recurrent implantation failure (RIF) were also included. METHODS: Nine prothrombotic gene variants were evaluated: factor V Leiden (FVL), factor V, H1299R variant (FVR2), factor II (FII) G20210A, methylene-tetrahydrofolate reductase (MTHFR) C677T and A1298C, beta-fibrinogen -455G>A, factor XIII (FXIII) V34L, human platelet antigen-1 (HPA-1) L33P variants, and plasminogen activator inhibitor-1 (PAI-1) 4G/5G. The following study groups were assessed: (1) women who experienced one (n = 334) or two (n = 264) episodes of pregnancy loss; (2) 468 women who experienced RPL; (3) 214 women who underwent ART followed by ongoing pregnancies; and (4) 282 women who experienced RIF after ART, that is, three or more consecutive implantation failures following high-quality embryo transfers to the uterus with an appropriate endometrium. As control group, 430 subjects from the general population were enrolled. RESULTS: FVL, the -455G>A variant of beta-fibrinogen, and PAI-1 4G were associated with a higher risk of developing RPL compared with the general population. Furthermore, FVL, FVR2, FII G20210A and MTHFR C677T conferred a significantly higher risk of RIF in women who performed ART compared with the general population. No statistical differences between the general population and other study groups were observed. CONCLUSIONS: Specific prothrombotic genetic variants are more frequently expressed in women with RPL and RIF, supporting their role in the development of polimicrothrombosis and impairing the invasion during embryo implantation.

Contamination of Hotel Water Distribution Systems by Legionella Species: Environmental Surveillance in Campania Region, South Italy.

The COVID-19 pandemic period was marked by the absence or reduced circulation of some infectious diseases. Legionellosis may have been affected by the prevention measures adopted to counter COVID-19. Legionellosis is a form of pneumonia interstitial that is normally transmitted via aerosol-containing bacteria (genus Legionella), that could be present in contaminated water sources and is often associated with travel and with staying in hotels. In this work, the data of the environmental surveys carried out by ARPA Campania in accommodation facilities since 2019 were analyzed for a better understanding of the dispersion patterns of L. pneumophila associated with the environment and to evaluate the variation of the data during the pandemic period. The aim was to provide a better understanding of Legionella at different geographic scales and to define a predictive epidemiological method. Results: In 2019, the Legionella genus contaminated 37.7% of all tourist facilities evaluated. In 2020, the Legionella genus contaminated 44.4% of all tourist facilities evaluated. In 2021, the Legionella genus contaminated 54.2% of all tourist facilities evaluated. Conclusions: Legionella pneumophila was the most prevalent species in our community, serogroup 1 was the most frequently isolated and the most implicated risk factor of contamination was the temperature of water in circulation.

Intralesional TLR4 agonist treatment strengthens the organ defense against colonizing cancer cells in the brain.

Brain metastasis in breast cancer remains difficult to treat and its incidence is increasing. Therefore, the development of new therapies is of utmost clinical relevance. Recently, toll-like receptor (TLR) 4 was correlated with IL6 expression and poor prognosis in 1 215 breast cancer primaries. In contrast, we demonstrated that TLR4 stimulation reduces microglia-assisted breast cancer cell invasion. However, the expression, prognostic value, or therapeutic potential of TLR signaling in breast cancer brain metastasis have not been investigated. We thus tested the prognostic value of various TLRs in two brain-metastasis gene sets. Furthermore, we investigated different TLR agonists, as well as MyD88 and TRIF-deficient microenvironments in organotypic brain-slice ex vivo co-cultures and in vivo colonization experiments. These experiments underline the ambiguous roles of TLR4, its adapter MyD88, and the target nitric oxide (NO) during brain colonization. Moreover, analysis of the gene expression datasets of breast cancer brain metastasis patients revealed associations of TLR1 and IL6 with poor overall survival. Finally, our finding that a single LPS application at the onset of colonization shapes the later microglia/macrophage reaction at the macro-metastasis brain-parenchyma interface (MMPI) and reduces metastatic infiltration into the brain parenchyma may prove useful in immunotherapeutic considerations.

Microbial retention and resistances in stormwater quality improvement devices treating road runoff.

Current knowledge about the microbial communities that occur in urban road runoff is scarce. Road runoff of trafficked roads can be heavily polluted and is treated by stormwater quality improvement devices (SQIDs). However, microbes may influence the treatment process of these devices or could lead to stress resistant opportunistic microbial strains. In this study, the microbial community in the influent, effluent and the filter materials used to remove dissolved heavy metals from two different SQIDs were analyzed to determine microbial load, retention, composition, and mobile resistance genes. Although the microbes were replaced by new taxa in the effluent, there was no major retention of microbial genera. Further, the bacterial abundance of the SQIDs effluent was relatively stable over time. The heavy metal content correlated with intl1 and with microbial genera. The filter media itself was enriched with Intl1 gene cassettes, carrying several heavy metal and multidrug resistance genes (e.g. czrA, czcA, silP, mexW and mexI), indicating that this is a hot spot for horizontal gene transfer. Overall, the results shed light on road runoff microbial communities, and pointed to distinct bacterial communities within the SQIDs, which subsequently influence the microbial community and the genes released with the treated water.

Screening and isolation of microbes from a Mud Community of Ischia Island Thermal Springs: preliminary analysis of a bioactive compound.

INTRODUCTION: Balneotherapy centers of Ischia island (Italy) offer treatments for different dermatological diseases (psoriasis, acne, atopic dermatitis) and upper respiratory tract infections. In this study, we integrated morphological and molecular approaches to give a focus on isolation and screening of extremophile bacteria from Ischia thermal mud for potential antimicrobial applications. METHODS: Samples were collected during 2019 at four sites. Some bacterial strains ATCC for antibacterial and antibiofilm activity were tested. After morphological characterization, screening for antagonistic isolates was made. The colonies isolated from thermal mud samples were submitted to molecular characterization. Susceptibility testing by dilution spotting was carried out and antibacterial efficacies of most active isolate were evaluated with a Minimal inhibition concentration assay. Biofilm formation, inhibition, eradication were examined. Statistical analyses were carried out utilizing Microsoft® Excel 2016/XLSTAT©-Pro. RESULTS: We isolated a natural compound with antimicrobial and antibiofilm activities. CONCLUSIONS: The results obtained in this study are discussed in the context of how hydrothermal systems are important environmental source of uncharted antimicrobial and antibiofilm compounds. In conclusion, to the most effective of our knowledge, this work presents the primary report on the preliminary investigation of thermophile microbial diversity and their antimicrobial and antibiofilm activities for future biotechnological interest.

Cerium, gadolinium, lanthanum, and neodymium effects in simplified acid mine discharges to Raphidocelis subcapitata, Lepidium sativum, and Vicia faba.

The alteration of rare earth elements (REEs) biogeochemical cycles has increased the potential effects related to their environmental exposure in a one-health perspective. Cerium (Ce), gadolinium (Gd), lanthanum (La), and neodymium (Nd) are frequently related to technological applications and their environmental concentrations are already in the µg/kg - mg/kg (i.e., or L) range depending on the considered matrices. The effect of Ce, Gd, La, and Nd was investigated in a simulated AMD (0.01-10.22 mg/L) at pH 4 and 6 considering a battery of photosynthetic organisms (Raphidocelis subcapitata, Lepidium sativum, and Vicia faba) according to a multiple-endpoint approach (growth inhibition, germination index, and mutagenicity). According to modelled chemical speciation, the considered elements were mostly in the trivalent free form (86-88%) at pH 4. Gd, La, and Nd exerted the most relevant toxic effect at pH 4. The pH 6 scenario evidenced a reduction in REEs toxicity level. Mutagenicity was detected only at pH 4 by Gd (up to 3-fold compared to negative controls), La and Nd, while Ce did not show any adverse effect. Toxic effects due to Ce, Gd, La, and Nd can be reduced by controlling the pH, but several gaps of knowledge still remain about their uptake and trophic transfer, and long-term effects on targeted species.

Long-term multi-endpoint exposure of the microalga Raphidocelis subcapitata to lanthanum and cerium.

Significant release of rare earth elements (REEs) into the environment is mainly due to active or abandoned mining sites, but their presence is globally increasing due to their use in several industrial sectors. The effects on primary producers as Raphidocelis subcapitata are still limited. This research focused on La and Ce as the two most widespread REEs that can be currently found up to hundreds of µg/L in water and wastewater. Microalgae were exposed to La and Ce for 3 days (pH = 7.8) (short-term exposure) to derive the effective concentrations inhibiting the growth on 10% (EC10) of the exposed population. EC10 values (0.5 mg/L of La and 0.4 mg/L of Ce) were used for the 28 days long-term exposure (renewal test) to observe after 7, 14, 21, and 28 days on a multi-endpoint basis microalgae growth inhibition (GI), biomarkers of stress (reactive oxygen species (ROS), superoxide dismutase (SOD), and catalase (CAT)), and bioconcentration. Results evidenced that La and Ce EC10 increased GI (day 28) up to 38% and 28%, respectively. ROS, CAT, and SOD activities showed differential responses from day 7 to day 14, 21, and 28, suggesting, in most of the cases, that La and Ce effects were counteracted (i.e., being the values at day 28 not significantly different, p > 0.05, from the relative negative controls), except for La-related ROS activities. La and Ce significantly bioconcentrated in microalgae populations up to 2- and 5-fold (i.e., at day 28 compared to day 7), in that order. Bioconcentrated La and Ce were up to 3157 and 1232 µg/g dry weight (day 28), respectively. These results suggested that low La and Ce concentrations can be slightly toxic to R. subcapitata having the potential to be bioaccumulated and potentially transferred along the food web.

Mediterranean Diet, Physical Activity and Gut Microbiome Composition: A Cross-Sectional Study among Healthy Young Italian Adults.

BACKGROUND: This cross-sectional study aimed to explore the microbial composition of the gut and its possible association with the Mediterranean diet (MD) after adjusting for demographic and anthropometric characteristics in a sample of healthy young Italian adults. METHODS: Gut microbiota, demographic information, and data on adherence to MD and physical activity (PA) habits were collected in a sample of 140 university students (48.6% males, mean age 22.5 ± 2.9) with a mean body mass index (BMI) of 22.4 ± 2.8 kg/m2 (15.2-33.8) and a mean PA level of 3006.2 ± 2973.6 metabolic equivalent (MET)-minutes/week (148-21,090). RESULTS: A high prevalence of Firmicutes and Bacteroidetes was found in all the fecal samples. Significant dissimilarities in the microbiota composition were found on the basis of MD adherence and PA levels (p = 0.001). At the genus level, Streptococcus and Dorea were highly abundant in overweight/obese individuals, Ruminococcus and Oscillospira in participants with lower adherence to MD, and Lachnobacterium in subjects with low levels of PA (p = 0.001). A significantly higher abundance of Paraprevotella was shown by individuals with lower BMI, lower MD adherence, and lower PA levels (p = 0.001). CONCLUSIONS: This study contributes to the characterization of the gut microbiome of healthy humans. The findings suggest the role of diet and PA in determining gut microbiota variability.

Salivary Cytokines and Airways Disease Severity in Patients with Cystic Fibrosis.

About 50% of patients with cystic fibrosis (CF) have sinonasal complications, which include inferior turbinate hypertrophy (NTH) and/or nasal polyposis (NP), and different degrees of lung disease, which represents the main cause of mortality. Monitoring of sinonasal disease requires complex instrumental procedures, while monitoring of lung inflammation requires invasive collection of bronchoalveolar lavage fluid. The aim of this study was to investigate the associations between salivary cytokines levels and CF-related airway diseases. Salivary biochemical parameters and cytokines, i.e., interleukin-6 (IL-6), IL-8, and tumor necrosis factor alpha (TNF-α), were analyzed in resting saliva from healthy subjects and patients with CF. Patients with CF showed significantly higher levels of salivary chloride, IL-6, IL-8, and TNF-α and lower calcium levels than healthy subjects. Among patients with CF, IL-6 and IL-8 were significantly higher in patients with NTH, while TNF-α was significantly lower in patients with NP. A decreasing trend of TNF-α in patients with severe lung disease was also observed. On the other hand, we did not find significant correlation between cytokine levels and Pseudomonas aeruginosa or Stenotrophomonas maltophilia colonization. These preliminary results suggest that salivary IL-6 and IL-8 levels increase during the acute phase of sinonasal disease (i.e., NTH), while the end stages of pulmonary disease and sinonasal disease (i.e., NP) show decreased TNF-α levels.

Molecular Analysis of Prothrombotic Gene Variants in Venous Thrombosis: A Potential Role for Sex and Thrombotic Localization.

Background: Requests to test for thrombophilia in the clinical context are often not evidence-based. Aim: To define the role of a series of prothrombotic gene variants in a large population of patients with different venous thromboembolic diseases. Methods: We studied Factor V Leiden (FVL), FVR2, FII G20210A, Methylenetetrahydrofolate reductase (MTHFR) C677T and A1298C, beta-fibrinogen -455 G>A, FXIII V34L, and HPA-1 L33P variants and PAI-1 4G/5G alleles in 343 male and female patients with deep vein thrombosis (DVT), 164 with pulmonary embolism (PE), 126 with superficial vein thrombosis (SVT), 118 with portal vein thrombosis (PVT), 75 with cerebral vein thrombosis (CVT) and 119 with retinal vein thrombosis (RVT), and compared them with the corresponding variants and alleles in 430 subjects from the general population. Results: About 40% of patients with DVT, PE and SVT had at least one prothrombotic gene variant, such as FVL, FVR2 and FII G20210A, and a statistically significant association with the event was found in males with a history of PE. In patients with a history of PVT or CVT, the FII G20210A variant was more frequent, particularly in females. In contrast, a poor association was found between RVT and prothrombotic risk factors, confirming that local vascular factors have a key role in this thrombotic event. Conclusions: Only FVL, FVR2 and FII G20210A are related to vein thrombotic disease. Other gene variants, often requested for testing in the clinical context, do not differ significantly between cases and controls. Evidence of a sex difference for some variants, once confirmed in larger populations, may help to promote sex-specific prevention of such diseases.

Impact of 13Valent Vaccine for Prevention of Pneumococcal Diseases in Children and Adults at Risk: Possible Scenarios in Campania Region.

BACKGROUND: Pneumonias are the most frequent infectious diseases, characterized by a high prevalence especially among children and adults at risk. The socio-economic impact caused by Streptococcus pneumoniae is evaluated in terms of morbidity, death rate and hospitalizations. OBJECTIVE: The aim of the study was to demonstrate the potential economic advantages by implementation of an active anti-pneumococcal 13-valent vaccine strategy in Campania region (Southern Italy) in two different categories of subjects, children (aged 0-12), and adults (aged 50- 79) at risk (hypertension, nephropathies, COPD and heart diseases). METHODS: Vaccination costs were compared with costs necessary to treat avoidable diseases in the presence and absence of a vaccination program. RESULTS: Offering anti-pneumococcal 13-valent vaccine to the paediatric population was quantified as saving one million euros for Italian national health service in two years. In addition, offering anti-pneumococcal vaccine to adults at risk would generate a return of around 29 million euros. CONCLUSION: In both cases, offering anti-pneumococcal 13-valent vaccine was proven to be a helpful political health strategy, not only in consideration of a reduction of cases but also in view of the favourable economic impacts.

Prevention of Pseudomonas aeruginosa Biofilm Formation on Soft Contact Lenses by Allium sativum Fermented Extract (BGE) and Cannabinol Oil Extract (CBD).

Two natural mixtures, Allium sativum fermented extract (BGE) and cannabinol oil extract (CBD), were assessed for their ability to inhibit and remove Pseudomonas aeruginosa biofilms on soft contact lenses in comparison to a multipurpose Soft Contact Lens-care solution present on the Italian market. Pseudomonas aeruginosa (ATCC 9027 strain) and Pseudomonas aeruginosa clinical strains isolated from ocular swabs were tested. Quantification of the biofilm was done using the microtiter plate assay and the fractional inhibitory concentration index was calculated. Both forms of Pseudomonas aeruginosa generated biofilms. BGE at minimal inhibitory concentration (MIC) showed inhibition percentages higher than 55% for both strains, and CBD inhibited biofilm formation by about 70%. The care solution at MIC inhibited biofilm formation by about 50% for both strains tested. The effect of BGE on the eradication of the microbial biofilm on soft contact lenses at MIC was 45% eradication for P. aeruginosa ATCC 9027 and 36% for P. aeruginosa clinical strain. For CBD, we observed 24% biofilm eradication for both strains. For the care solution, the eradication MICs were 43% eradication for P. aeruginosa ATCC 9027 and 41% for P. aeruginosa clinical strain. It was observed that both the test soft contact lenses solution/BGE (fractional inhibitory concentration index: 0.450) and the test soft contact lenses solution/CBD (fractional inhibitory concentration index: 0.153) combinations exhibited synergistic antibiofilm activity against most of the studied bacteria. The study showed that BGE and CBD have good effect on inhibition of biofilm formation and removal of preformed biofilms, which makes them promising agents that could be exploited to develop more effective care solutions.

Melittin Inhibition and Eradication Activity for Resistant Polymicrobial Biofilm Isolated from a Dairy Industry after Disinfection.

The emerging concern about the increase of antibiotic resistance and associated biofilm has encouraged scientists to look for alternative antibiotics such as antimicrobial peptides (AMPs). This study evaluated the ability of melittin to act as an antibacterial biofilm inhibitor and biofilm remover considering isolates from dairy industry. Minimum inhibitory concentrations (MICs), minimum bactericidal concentrations (MBCs), minimum biofilm inhibitory concentrations (MBICs), and biofilm removal activities were studied in polymicrobial biofilms produced from isolates. MIC and MBC were set at 1-3 µg/mL and 25-50 µg/mL for Gram-positive and Gram-negative bacteria, respectively. Results demonstrated a good MBIC reaching 85% inhibition ability and a good activity and better penetration in deeper layers against the mixed preformed biofilm, thereby increasing its activity against all isolates also at the lowest tested concentrations. Melittin showed interesting characteristics suggesting its potential to act as an antimicrobial agent for polymicrobial biofilm from dairy industry even in environmental isolates.

Two cases of microvillous inclusion disease caused by novel mutations in MYO5B gene.

Microvillous inclusion disease (MVID) typically appears with severe chronic diarrhea in the few days after birth and rapidly causes dehydration and metabolic acidosis. In this context, presenting two novel cases, we underline the crucial importance of mutation analysis for the diagnosis of this disease that may be easily misdiagnosed.

Supervised physical exercise improves clinical, anthropometric and biochemical parameters in adult cystic fibrosis patients: A 2-year evaluation.

OBJECTIVE: Cystic fibrosis (CF) is the most common inherited, life limiting condition among Caucasians. No healing therapy is currently available for patients with CF. The aim of the study was to define clinical, anthropometric and biochemical effects of regular, supervised physical exercise in a large cohort of patients with CF. MATERIALS AND METHODS: Fifty-nine adult patients with CF that performed regularly supervised physical exercise in the last 3 years in comparison to 59 sex and age matched sedentary patients with CF were included in the study. RESULTS: Physical exercise had significantly beneficial effects on: (a) FEV1% decline; (b) anthropometric parameters (lower number of cases with altered BMI, waist and arm circumferences); (c) lipid and glucose metabolism; (d) vitamin D serum levels. Of course, some of this improvement may be because of the better adherence to therapy typical of patients with CF that perform physical activity. CONCLUSIONS: Such clinical and metabolic effects make supervised physical activity one of the hubs in managing patients with CF.