Effects of Influenza Vaccine on the Immune Responses to SARS-CoV-2 Vaccination.

A number of studies have suggested that influenza vaccination can provide protection against COVID-19, but the underlying mechanisms that could explain this association are still unclear. In this study, the effect of the 2021/2022 seasonal influenza vaccination on the immune response to the booster dose of anti-SARS-CoV-2 vaccination was evaluated in a cohort of healthy individuals. A total of 113 participants were enrolled, 74 of whom had no prior COVID-19 diagnosis or significant comorbidities were considered for the analysis. Participants received the anti-influenza tetravalent vaccine and the booster dose of the anti-SARS-CoV-2 vaccine or the anti-SARS-CoV-2 vaccine alone. Blood was collected before and 4 weeks after each vaccination and 12 weeks after SARS-CoV-2 vaccination and analyzed for anti-flu and anti-spike-specific antibody titers and for in vitro influenza and SARS-CoV-2 neutralization capacity. Results indicated an increased reactivity in subjects who received both influenza and SARS-CoV-2 vaccinations compared to those who received only the SARS-CoV-2 vaccine, with sustained anti-spike antibody titers up to 12 weeks post-vaccination. Immune response to the influenza vaccine was evaluated, and individuals were stratified as high or low responders. High responders showed increased antibody titers against the SARS-CoV-2 vaccine both after 4 and 12 weeks post-vaccination. Conversely, individuals classified as low responders were less responsive to the SARS-CoV-2 vaccine. These data indicate that both external stimuli, such as influenza vaccination, and the host's intrinsic ability to respond to stimuli play a role in the response to the vaccine.

Detection and full genomic sequencing of rare hepatitis E virus genotype 4d in Italian wastewater, undetected by clinical surveillance.

Hepatitis E is a liver disease caused by the hepatitis E virus (HEV), primarily transmitted through contaminated water or food. There are four different HEV genotypes in humans, with genotypes 1 and 2 being the most widespread. Genotypes 3 and 4 are found in animals and can also infect humans. Genotype 4 is prevalent in Asia, mainly in China. In Italy, only one outbreak of HEV-4 has been documented, which occurred in 2011, involving five patients. In 2013, HEV G4 was also detected in a pig farm. Since then, no further evidence of HEV genotype 4 has been found in the country. This study describes the first detection of HEV genotype 4, subtype d, in wastewater in central Italy, despite a lack of any clinical case reported in the area. By using a multiplex PCR protocol and two sequencing strategies, Illumina and ONT, the virus's complete genome was sequenced and characterized as subtype 4d. These findings shed light on the potential of environmental surveillance for infectious agents to improve our understanding of epidemiology and support public health efforts.

Epidemiological and molecular characterization of HBV and HCV infections in HIV-1-infected inmate population in Italy: a 2017-2019 multicenter cross-sectional study.

HBV/HCV co-infection is common in HIV-1-infected prisoners. To investigate the characteristics of HIV co-infections, and to evaluate the molecular heterogeneity of HIV, HBV and HCV in prisoners, we carried-out a multicenter cross-sectional study, including 65 HIV-1-infected inmates enrolled in 5 Italian detention centers during the period 2017-2019. HIV-1 subtyping showed that 77.1% of inmates were infected with B subtype and 22.9% with non-B subtypes. Italian nationals were all infected with subtype B (93.1%), except two individuals, one infected with the recombinant form CRF72_BF1, and the other with the HIV-1 sub-subtype A6, both previously not identified in inmates of Italian nationality. Non-Italian nationals were infected with subtype B (52.6%), CRFs (36.8%) and sub-subtypes A1 and A3 (5.2%). HIV variants carrying resistance mutations to NRTI, NNRTI, PI and InSTI were found in 7 inmates, 4 of which were never exposed to the relevant classes of drugs associated with these mutations. HBV and/or HCV co-infections markers were found in 49/65 (75.4%) inmates, while 27/65 (41.5%) showed markers of both HBV and HCV coinfection. Further, Italian nationals showed a significant higher presence of HCV markers as compared to non-Italian nationals (p = 0.0001). Finally, HCV phylogenetic analysis performed in 18 inmates revealed the presence of HCV subtypes 1a, 3a, 4d (66.6%, 16.7% and 16.7%, respectively). Our data suggest the need to monitor HIV, HBV and HCV infections in prisons in order to prevent spreading of these viruses both in jails and in the general population, and to implement effective public health programs that limit the circulation of different genetic forms as well as of viral variants with mutations conferring resistance to treatment.

Correction to: Genetic Diversity Among Genogroup II Noroviruses and Progressive Emergence of GII.17 in Wastewaters in Italy (2011-2016) Revealed by Next-Generation and Sanger Sequencing.

The original version of this article unfortunately contained a mistake. The presentation of Table 1 was incorrect. The corrected table is given below. The original article has been corrected.

Genetic Diversity Among Genogroup II Noroviruses and Progressive Emergence of GII.17 in Wastewaters in Italy (2011-2016) Revealed by Next-Generation and Sanger Sequencing.

Noroviruses (NoV) are a major cause of gastroenteritis worldwide. Recently, a novel variant of NoV GII.17 (GII.P17_GII.17 NoV), termed Kawasaki 2014, has been increasingly reported in NoV outbreaks in Asia, and has also been described in Europe and North America. In this study, sewage samples were investigated to study the occurrence and genetic diversity of NoV genogroup II (GII) along a 6-year period. Moreover, the spread of GII.17 strains (first appearance and occurrence along time) was specifically assessed. A total of 122 sewage samples collected from 2011 to 2016 from four wastewater treatment plants in Rome (Italy) were initially tested using real-time RT-(q)PCR for GII NoV. Positive samples were subsequently subjected to genotypic characterization by RT-nested PCRs using broad-range primes targeting the region C of the capsid gene of GII NoV, and specific primers targeting the same region of GII.17 NoV. In total, eight different genotypes were detected with the broad-range assay: GII.1 (n = 6), GII.2 (n = 8), GII.3 (n = 3), GII.4 (n = 13), GII.6 (n = 3), GII.7 (n = 2), GII.13 (n = 2), and GII.17 (n = 3), with the latter two genotypes detected only in 2016. Specific amplification of GII.17 NoV was successful in 14 out of 110 positive samples, spanned over the years 2013-2016. The amplicons of the broad-range PCR, pooled per year, were further analyzed by next-generation sequencing (NGS) for a deeper analysis of the genotypes circulating in the study period. NGS confirmed the circulation of GII.17 NoV since 2013 and detected, beyond the eight genotypes identified by Sanger sequencing, three additional genotypes regarded as globally uncommon: GII.5, GII.16, and GII.21. This study provides evidence that GII.17 NoV Kawasaki has been circulating in the Italian population before its appearance and identification in clinical cases, and has become a major genotype in 2016. Our results confirm the usefulness of wastewater surveillance coupled with NGS to study the molecular epidemiology of NoV and to monitor the emergence of NoV strains.

Molecular characterization of human adenoviruses in urban wastewaters using next generation and Sanger sequencing.

Human adenoviruses (HAdVs) are of major public health importance and are associated with a variety of clinical manifestations, including gastroenteritis, respiratory, ocular and urinary tract infections. To study the occurrence, prevalence and diversity of HAdV species and types circulating in Italy, we conducted a large-scale molecular-epidemiological investigation, a yearlong monitoring of 22 wastewater treatment plants, covering 10 Italian regions, representative of northern, central, and southern Italy. A total of 141 raw sewage samples were collected from January to December 2013, and processed to detect and characterize by phylogenetic analysis a fragment of the hexon coding region of HAdVs. Nested PCR results showed the presence of HAdVs in 85 out of 141 samples (60% of samples). Fifty-nine samples were characterized by conventional Sanger sequencing as belonging to four HAdV species and four types: A (type 12, 5 samples), B (type 3, 8 samples), C (type 5, 1 sample) and F (type 41, 45 samples). The remaining 26 samples could not be characterized because of uninterpretable (mixed) electropherograms suggesting the presence of multiple species and/or types. Pools of characterized and uncharacterized PCR amplicons were further analyzed by next-generation sequencing (NGS). NGS results revealed a marked HAdV diversity with 16 additional types detected beyond the four types found by Sanger sequencing. Overall, 19 types were identified, belonging to HAdV species A-F: types 12 and 31 (species A), type 3 (species B), types 1, 2, and 5 (species C), types 9, 17, 24, 26, 37, 38, 42, 44, 48, and 70 (species D), type 4 (species E), and types 40 and 41(species F). An untypeable HAdV was also detected, showing similar percentages of identity with more than one prototype (types 15, 30, 56, and 59). Our findings documented the circulation of a wide variety of species and types in raw sewage, potentially able to affect other surface water environments and hence human health. Next-generation sequencing proved to be an effective strategy for HAdV genotyping in wastewater samples. It was able to detect a wide range of "less prevalent" types unidentified by conventional Sanger sequencing, confirming that studies based on conventional technologies may grossly underestimate the existence of some, possibly less common, types. Knowledge of the distribution of HAdV species and types would improve our understanding of waterborne HAdV-related health risks.

Hepatitis E Virus (Genotype 3) in Slurry Samples from Swine Farming Activities in Italy.

Hepatitis E virus (HEV) is an emergent causative agent of acute hepatitis, transmitted by fecal-oral route. Infection with HEV is a global cause for morbidity and mortality throughout the world: it mainly causes large outbreaks in endemic areas and sporadic autochthonous cases in industrialized countries where HEV infections seem to be an emergent zoonotic disease. Infection of porcine livestock and its relationship with the human cases have been demonstrated. The present study describes an investigation on the prevalence and diversity of HEV in pig slurry in Italy. Slurry samples (24) were collected from ten farms located in North Italy during 2015 and analyzed for HEV, using four broad-range nested PCR assays targeting ORF1 (MTase), ORF2 (capsid) genes, and ORF2/3 regions. Overall, 18 samples (75%) were positive for HEV RNA, and characterized as genotype 3. Nine samples could be subtyped by ORF2 sequencing: Eight belonged to subtype 3f, while one sequence could not be characterized by blast analysis and phylogenetic analysis and may actually represent a new subtype. Furthermore, similarity of 99% was found between 3f Italian HEV sequences of human and swine origins. Real-Time PCR assay was also performed, in order to obtain quantitative data on positive samples. Two swine slurry samples were positive, containing 600 and 1000 UI per mL of sewage. The results of this study show that HEV strains belonging to zoonotic genotype 3 are widely present in swine excreta, and have high degree of identity with strains detected in autochthonous HEV cases. Improving swine farming operations safety and increasing operators' awareness of the zoonotic potential connected with the handling of swine effluents turn out to be key points in order to reduce the environmental and sanitary problem represented by the possible dissemination of HEV to water bodies.

Large and prolonged food-borne multistate hepatitis A outbreak in Europe associated with consumption of frozen berries, 2013 to 2014.

In May 2013, Italy declared a national outbreak of hepatitis A, which also affected several foreign tourists who had recently visited the country. Molecular investigations identified some cases as infected with an identical strain of hepatitis A virus subgenotype IA. After additional European Union/European Economic Area (EU/EEA) countries reported locally acquired and travel-related cases associated with the same outbreak, an international outbreak investigation team was convened, a European outbreak case definition was issued and harmonisation of the national epidemiological and microbiological investigations was encouraged. From January 2013 to August 2014, 1,589 hepatitis A cases were reported associated with the multistate outbreak; 1,102 (70%) of the cases were hospitalised for a median time of six days; two related deaths were reported. Epidemiological and microbiological investigations implicated mixed frozen berries as the vehicle of infection of the outbreak. In order to control the spread of the outbreak, suspected or contaminated food batches were recalled, the public was recommended to heat-treat berries, and post-exposure prophylaxis of contacts was performed. The outbreak highlighted how large food-borne hepatitis A outbreaks may affect the increasingly susceptible EU/EEA general population and how, with the growing international food trade, frozen berries are a potential high-risk food.

Hepatitis A and E Viruses in Wastewaters, in River Waters, and in Bivalve Molluscs in Italy.

Several studies have reported the detection of hepatitis A (HAV) and E (HEV) virus in sewage waters, indicating a possibility of contamination of aquatic environments. The objective of the present study was to assess the occurrence of HAV and HEV in different water environments, following the route of contamination from raw sewage through treated effluent to the surface waters receiving wastewater discharges . Bivalve molluscan shellfish samples were also analyzed, as sentinel of marine pollution. Samples were tested by RT-PCR nested type in the VP1/2A junction for HAV, and in the ORF1 and ORF2 regions for HEV. Hepatitis A RNA was detected in 12 water samples: 7/21 (33.3%) raw sewage samples, 3/21 (14.3%) treated sewage samples, and 2/27 (7.4%) river water samples. Five sequences were classified as genotype IA, while the remaining 7 sequences belonged to genotype IB. In bivalves, HAV was detected in 13/56 samples (23.2%), 12 genotype IB and one genotype IA. Whether the presence of HAV in the matrices tested indicates the potential for waterborne and foodborne transmission is unknown, since infectivity of the virus was not demonstrated. HEV was detected in one raw sewage sample and in one river sample, both belonging to genotype 3. Sequences were similar to sequences detected previously in Italy in patients with autochthonous HEV (no travel history) and in animals (swine). To our knowledge, this is the first detection of HEV in river waters in Italy, suggesting that surface water can be a potential source for exposure .

Hepatitis E virus genotypes 1 and 3 in wastewater samples in Tunisia.

Hepatitis E represents an important public-health concern throughout the world. It is one of the leading causes of hepatitis in North Africa, Asia and the Middle East. In Tunisia, the true burden of HEV infection is still unknown. The objectives of the present study were to assess the occurrence of hepatitis E virus in Tunisia through the monitoring of urban sewage and to characterize the strains identified using molecular assays. A total of 150 sewage samples (raw and treated) were collected from three wastewater treatment plants located in the regions of Monastir and Mahdia and analyzed by nested RT-PCR using a qualitative assay targeting the methyltransferase gene in ORF1. Of these, only three samples (2 %) were found to be positive for HEV, one belonging to genotype 1 and two to genotype 3. The results of the present study indicate a low level of virus excretion among the Tunisian population. Both genotypes 1 and 3 are circulating in this country, however, possibly causing sporadic infections. The presence of the zoonotic genotype 3, known to be transmitted to humans mainly by swine and demonstrated in Tunisia for the first time in this work, raises the question of possible reservoir species, since pork products are not consumed in this country, pigs are not bred, and wild boar is not endemic. Further studies will be needed to gather information on the occurrence and diversity of HEV strains circulating among humans and animals in Tunisia, and on possible animal reservoirs.

Qualitative and Quantitative Assessment of Hepatitis A Virus in Wastewaters in Tunisia.

Hepatitis A causes substantial morbidity in both industrialized and non-industrialized countries and represents an important health problem in several southern Mediterranean countries. The objectives of the study were as follows: (a) to assess the occurrence of hepatitis A virus (HAV) in Tunisia through the monitoring of urban wastewaters collected at wastewater treatment plants (WTPs); (b) to characterize environmental strains; and (c) to estimate the viral load in raw and treated sewages, in order to evaluate the potential impact on superficial waters receiving discharges. A total of 150 raw and treated wastewaters were collected from three WTPs and analyzed by both qualitative (RT-PCR/nested) and quantitative (qRT-PCR) methods. Of these, 100 (66%) were found to be positive for HAV by the qualitative assay: 68.3% in influents and 64.7% in effluents. The vast majority of HAV sequences belonged to sub-genotype IA, with 11 different strains detected found to be identical to clinical strains isolated from Tunisian patients with acute hepatitis. Five unique variants were also detected, not previously reported in clinical cases. Only two IB strains were found, confirming the rarity of this sub-genotype in this country. The results of the present study indicate a wide circulation of the pathogen in the population, most probably in the form of asymptomatic infections, a finding consistent with the classification of the country as having intermediate/high endemicity. Quantitative data showed high viral loads in influents (3.5E+05 genome copies/liter, mean value) as well as effluents (2.5E+05 genome copies/liter, mean value), suggesting that contaminated water could be a critical element in transmission.

Strong CD8+ T cell antigenicity and immunogenicity of large foreign proteins incorporated in HIV-1 VLPs able to induce a Nef-dependent activation/maturation of dendritic cells.

Virus-like particles (VLPs) are excellent tools for vaccines against pathogens and tumors. They can accommodate foreign polypeptides whose incorporation efficiency and immunogenicity however decrease strongly with the increase of their size. We recently described the CD8(+) T cell immune response against a small foreign antigen (i.e., the 98 amino acid long human papilloma virus E7 protein) incorporated in human immunodeficiency virus (HIV)-1 based VLPs as product of fusion with an HIV-1 Nef mutant (Nef(mut)). Here, we extended our previous investigations by testing the antigenic/immunogenic properties of Nef(mut)-based VLPs incorporating much larger heterologous products, i.e., human hepatitis C virus (HCV) NS3 and influenza virus NP proteins, which are composed of 630 and 498 amino acids, respectively. We observed a remarkable cross-presentation of HCV NS3 in dendritic cells challenged with Nef(mut)-NS3 VLPs, as detected using a NS3 specific CD8(+) T cell clone as well as PBMCs from HCV infected patients. On the other hand, when injected in mice, Nef(mut)-NP VLPs elicited strong anti-NP CD8(+) T cell and CTL immune responses. In addition, we revealed the ability of Nef(mut) incorporated in VLPs to activate and mature primary human immature dendritic cells (iDCs). This phenomenon correlated with the activation of Src tyrosine kinase-related intracellular signaling, and can be transmitted from VLP-challenged to bystander iDCs. Overall, these results prove that Nef(mut)-based VLPs represent a rather flexible platform for the design of innovative CD8(+) T cell vaccines.

Activation of the ER stress gene gadd153 by hepatitis C virus sensitizes cells to oxidant injury.

HCV induces endoplasmic reticulum (ER) stress which correlates with transcriptional induction of ER stress genes. Previously, we reported that expression of HCV structural proteins activates the ER stress and pro-apoptotic gene gadd153 which plays a relevant role in cell death induced by oxidative stress. In the present study, using human hepatic cell lines Huh7 carrying a full-length HCV replicon, we demonstrated that replication and expression of the complete set of HCV proteins were associated with elevated expression of gadd153. Analysis of gadd153 promoter activity revealed that both the ATF4 and the ATF6 pathways, which are typically induced during ER stress response, contribute to the induction of gadd153 in HCV replicon cells. Activation of the ATF4 pathway was confirmed by identification of increased levels of ATF4 protein in replicon cells. Importantly, we showed that, following H2O2 treatment, gadd153 gene reached higher levels of expression in replicon cells. Consistent with the marked induction of the pro-apoptotic gene gadd153, HCV replicon cells showed an increased vulnerability to oxidant injury. Treatment of replicon cells with a specific small interfering RNA, targeted to gadd153 gene, reduced basal expression of gadd153 and decreased cell death following H2O2. These findings suggest that gadd153 may play a major role in sensitivity of HCV replicon cell to oxidative stress.

Activation of endoplasmic reticulum stress response by hepatitis C virus proteins.

Flaviviruses utilize the endoplasmic reticulum (ER) as the main site for replication and protein synthesis and cause some level of ER stress. In the present study, we evaluated the ability of HCV proteins to induce ER stress response by using a tetracycline-regulated cell line expressing a region of HCV genome containing the structural genes. In this system different levels of HCV protein expression could be obtained by varying the concentration of tetracycline in the medium. Real Time PCR and Western blotting assay demonstrated that HCV mRNA and protein levels reach a maximum value at 24-48 h and decrease at 72 h postinduction. Cell proliferation analysis indicated that HCV synthesis causes cell growth inhibition. The effect was also observed in cells expressing lower levels of HCV proteins. The expression profile of specific genes, which are markers of ER stress response, revealed the upregulation of the chaperone GRP78 and the transcription factor GADD153. Induction of GADD153 correlates with the downregulation of the antiapoptotic Bcl-2 gene suggesting that synthesis of HCV proteins may influence cell fate through the activation of ER stress signaling pathway.

The transmembrane domain of hepatitis C virus E1 glycoprotein induces cell death.

The E1 protein of hepatitis C virus (HCV) shows the ability to induce cell lysis by the alteration of membrane permeability when expressed in Escherichia coli cells. This function seems to be an intrinsic property of a C-terminal hydrophobic region of E1 as permeability changes and cell lysis can be blocked by mutagenesis of specific amino acids in this domain. To establish whether the expression of E1 protein and its C-terminal domain was able to induce cell death also in eukaryotic cell, we cloned HCV sequences expressing the full-length E1 (E383), the C-terminal domain (SVP) and a mutant lacking the C-terminal region (E340) in the pRC/CMV expression vector. HepG2 cell line was co-transfected with empty vector or HCV expression plasmids and a reporter vector that expressed beta-galactosidase (beta-gal) to visualize co-transfected blue cells. At 60 h after transfection, the loss of blue cells, considered as a measure of cell death, was 31.5 and 64.3% for the E1 and SVP clones. On the contrary, the number of blue cells after transfection with E340 plasmid was similar to that observed with the control vector. The analysis by the terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling (TUNEL) assay revealed an increased number of apoptotic cells at 48 h after transfection with E1 and SVP clones. Furthermore, cells transfected with SVP revealed a typical internucleosomal DNA fragmentation and the activation of caspase-3-like proteases as the specific inhibitor Ac-DEVD-CHO peptide partially blocked SVP apoptosis. These data indicate that the intracellular expression of HCV E1 protein and its C-terminal domain induces an apoptotic response in human hepatoma cell line.

Effect of acute and chronic administration of the GABA B agonist baclofen on 24 hour pH metry and symptoms in control subjects and in patients with gastro-oesophageal reflux disease.

BACKGROUND AND AIMS: The gamma-aminobutyric acid (GABA(B)) agonist baclofen has been shown to reduce reflux episodes during the first three postprandial hours in patients with gastro-oesophageal reflux disease (GORD) and in normal controls. The aim of the study was to assess the effect of acute (one day) and chronic (four weeks) administration of baclofen on 24 hour pH metry and symptoms in GORD patients and normal controls. PATIENTS AND METHODS: Acute study: 28 patients with GORD with none or mild oesophagitis at endoscopy and 15 controls underwent oesophageal and gastric 48 hour pH metry in which baclofen or placebo was given for 24 hours in a double blinded manner. Chronic study: 16 GORD patients received baclofen (10 mg four times daily) or placebo for four weeks. Twenty four hour oesophageal pH metry and reflux symptom scores were evaluated before and at the end of treatment. RESULTS: Acute study: the number of reflux episodes and per cent time with pH <4 was significantly lower after baclofen in GORD patients and controls (p<0.003; p<0.0007). Gastric pH increased significantly in GORD patients and controls (p<0.001; p<0.05). Chronic study: four weeks after initial administration of baclofen, the number of reflux episodes and percentage of time with pH <4 significantly decreased in all GORD patients (p<0.003; p<0.02). Symptom scores significantly improved after treatment with baclofen (p<0.0007). CONCLUSIONS: The GABA(B) agonist baclofen reduces 24 hour gastro-oesophageal reflux and increases gastric pH in GORD patients and controls. When given for one month to GORD patients, baclofen reduces oesophageal acid refluxes and significantly improves symptoms. Baclofen may be useful in the therapy of GORD.

Effect of the 5-HT1 agonist sumatriptan on oesophageal motor pattern in patients with ineffective oesophageal motility.

The 5-HT1 agonist sumatriptan (SUM) elicits an increase in amplitude of oesophageal motor waves and of lower oesophageal sphincter (LOS) tone in healthy subjects. The aim of the study was to evaluate whether such an effect occurs also in patients with ineffective oesophageal motility (IOM). 16 patients (nine males and seven females, age range 34-55 years) with chest pain and mild to moderate dysphagia were studied; all had undergone previous cardiologic, radiologic and upper gastrointestinal endoscopic exams that were normal. An oesophageal manometry was performed using an electronic probe to record swallows, oesophageal, LOS and gastric motility. The patients whose motor pattern were compatible with IOM (>30% of motor waves with amplitude <30 mmHg and/or non-transmitted) received SUM or placebo 6 mg s.c., injected in the morning and in the afternoon in a random order. The data analysis was limited to 1 h before and 1 h after the drug injections. Ten out of the 16 patients showed an IOM motor pattern. The administration of SUM caused a significant increase in the number of swallows (SUM 99.5 +/- 15.4 vs 78.6 +/- 16.1 basal, P = 0.03) and of primary oesophageal motor waves (SUM 89.6 +/- 13.4 vs 67.2 +/- 12.9 basal, P = 0.04) with no significant changes in the percentage of swallows associated with propagation. Placebo was not associated with increase in the number of swallows (80.3 +/- 14.6, P = 0.9) or of primary oesophageal motor waves (70.1 +/- 12.3, P = 0.7). The amplitude and the percentage of propagated oesophageal motor waves as well as the mean basal LOS tone were unaltered by SUM. There was no change in the symptoms reported after SUM. Although effective in healthy subjects, SUM 6 mg s.c. improves only the numbers but not the amplitude or propagation of oesophageal motility of patients with IOM. The 5-HT1 pathway and its acute stimulation seem to play only a minor role in the pathogenesis of such a disease.

5-HT1-receptor agonist sumatriptan modifies gastric size after 500 ml of water in dyspeptic patients and normal subjects.

Sumatriptan, a 5-HT1-receptor agonist has been shown to delay gastric emptying of liquids and solids in humans. However, no data are available of the effect of sumatriptan on gastric adaptation after distension with liquids and on symptoms induced by gastric distension. In 23 normal subjects and 30 dyspeptic patients with normal upper gastrointestinal endoscopy and real-time ultrasonography, the transverse gastric proximal and distal area and sagittal axis of the proximal stomach were determined by real-time ultrasonography and computed tomography after 500 ml of water. The area was determined by real-time ultrasonography and computed tomography twice at times 48 hr apart. Thirty minutes before real-time ultrasonography, placebo or sumatriptam were give subcutaneously in a double-blind fashion. Epigastric pain, bloating, heartburn, and nausea were also monitored through an intensity score from zero to 10 performed during the test. In six dyspeptic patients, the gastric distension was performed also with real-time ultrasonography and computed tomography after placebo and hyoscine butyl-bromide, a quaternary anticholinergic agent. Real-time ultrasonography and computed tomography demonstrated that after sumatriptan there is a reduction in proximal and distal transverse area and an increase in the sagittal axis of the proximal stomach. Hyoscine butyl-bromide increased all gastric measurements. Among the symptoms evaluated, only nausea was significantly reduced by sumatriptan (P < 0.01). Sumatriptan modifies gastric size, with a reduction in the transverse section and an increase of the sagittal axis of the proximal stomach and improves the nausea induced by gastric distension in dyspeptic patients.

Transient lower oesophageal sphincter relaxations play an insignificant role in gastro-oesophageal reflux to the proximal oesophagus.

Gastro-oesophageal reflux to the proximal oesophagus may cause atypical symptoms of gastro-oesophageal reflux disease (GORD). The motor abnormalities underlying reflux into the proximal oesophagus are still unclear. The aim of this study was to analyse the oesophageal motility during reflux into the proximal oesophagus in a group of healthy subjects and in patients with atypical symptoms of GORD. We concentrated particularly on lower oesophageal sphincter (LOS) activity and transient lower oesophageal sphincter relaxations (TLOSRs). Ten patients (7M, 3F, age 25-51 years) with mild oesophagitis (Savary-Miller grade I-II) and 10 healthy subjects (6M, 4F, age 23-54 years) underwent a 24-h dual pH-metric and manometric recording, using an electronic portable device. This recorded distal and proximal oesophageal pH values, oesophageal body and LOS motility. GORD patients had more distal and proximal reflux (DR and PR) compared with healthy controls (DR P < 0.001; PR P < 0.05). TLOSRs were the most frequent event during reflux into the distal oesophagus, whereas TLOSR frequency was much lower during reflux to the proximal oesophagus in GORD patients and in healthy controls (P < 0.05 and P < 0.01 vs. distal reflux, respectively). A significant relationship between TLOSRs and distal refluxes was present but no relationship with proximal reflux was detected. We conclude that TLOSRs are much less frequent during reflux to the proximal oesophagus than distal oesophageal reflux in patients with mild GORD suffering from atypical manifestations. The mechanism of acid reflux to the proximal oesophagus is unclear.

Mutagenesis of hepatitis C virus E1 protein affects its membrane-permeabilizing activity.

The E1 glycoprotein of hepatitis C virus is a transmembrane glycoprotein with a C-terminal anchor domain. When expressed in Escherichia coli, E1 induces a change in membrane permeability that is toxic to the bacterial cell. The C-terminal hydrophobic region (aa 331-383) of E1 is mainly responsible for membrane association and for inducing changes in membrane permeability. These observed changes are similar to those produced in E. coli by influenza virus M2, human immunodeficiency virus gp41 and poliovirus 3AB proteins, whose hydrophobic domains are thought to cause pore formation in biological membranes. To further characterize the activity of E1 at a molecular level, the membrane-permeabilizing ability of a second internal hydrophobic region (aa 262-291) was examined by expressing different deletion mutants of E1 in an E. coli system that is widely used for analysing membrane-active proteins from other animal viruses. Moreover, highly conserved amino acids in the C-terminal hydrophobic region were mutated to identify residues that are critical for inducing changes in membrane permeability. Analysis of cell growth curves of recombinant cultures and membrane-permeability assays revealed that synthesis of this fragment increased the flux of small compounds through the membrane and caused progressive cell lysis, suggesting that this domain has membrane-active properties. Furthermore, analysis of C-terminal mutants indicated that the conserved amino acids Arg(339), Trp(368) and Lys(370) play a critical role in protein function, as both cell lysis and changes in membrane permeability induced by the wild-type clone could be blocked by substitutions in these positions.