Utility of accessible SARS-CoV-2 specific immunoassays in vaccinated adults with a history of advanced HIV infection.

Accessible SARS-CoV-2-specific immunoassays may inform clinical management in people with HIV, particularly in case of persisting immunodysfunction. We prospectively studied their application in vaccine recipients with HIV, purposely including participants with a history of advanced HIV infection. Participants received one (n = 250), two (n = 249) or three (n = 42) doses of the BNT162b2 vaccine. Adverse events were documented through questionnaires. Sample collection occurred pre-vaccination and a median of 4 weeks post-second dose and 14 weeks post-third dose. Anti-spike and anti-nucleocapsid antibodies were measured with the Roche Elecsys chemiluminescence immunoassays. Neutralising activity was evaluated using the GenScript cPass surrogate virus neutralisation test, following validation against a Plaque Reduction Neutralization Test. T-cell reactivity was assessed with the Roche SARS-CoV-2 IFNγ release assay. Primary vaccination (2 doses) was well tolerated and elicited measurable anti-spike antibodies in 202/206 (98.0%) participants. Anti-spike titres varied widely, influenced by previous SARS-CoV-2 exposure, ethnicity, intravenous drug use, CD4 counts and HIV viremia as independent predictors. A third vaccine dose significantly boosted anti-spike and neutralising responses, reducing variability. Anti-spike titres > 15 U/mL correlated with neutralising activity in 136/144 paired samples (94.4%). Three participants with detectable anti-S antibodies did not develop cPass neutralising responses post-third dose, yet displayed SARS-CoV-2 specific IFNγ responses. SARS-CoV-2 vaccination is well-tolerated and immunogenic in adults with HIV, with responses improving post-third dose. Anti-spike antibodies serve as a reliable indicator of neutralising activity. Discordances between anti-spike and neutralising responses were accompanied by detectable IFN-γ responses, underlining the complexity of the immune response in this population.

Stronger and durable SARS-CoV-2 immune response to mRNA vaccines in 5-11 years old children with prior COVID-19.

BACKGROUND AND OBJECTIVES: mRNA vaccines elicit a durable humoral response to SARS-CoV-2 in adults, whereas evidence in children is scarce. This study aimed to assess the early and long-term immune response to the mRNA vaccine in children with or without previous SARS-CoV-2 infection. METHODS: In a multicentre prospective observational study, we profiled the immune response to the Pfizer BioNTech (BNT162b2) vaccine in 5-11-year-old children attending the University Pediatric Hospital of Padua and Bambino-Gesù Hospital in Rome (Italy) from December-2021 to February-2023. Blood samples were collected pre-, 1-, and 6-months after vaccination. Neutralizing antibodies (NAbs) and anti-spike-receptor-binding-domain (anti-S-RBD) IgG titers were analyzed through Plaque Reduction Neutralization Test (PRNT) and chemiluminescent immune-enzymatic assay (CLIA), respectively. Immune cell phenotypes were analyzed by flow cytometry. RESULTS: Sixty children (26 [43 %] female, median age = 8 years [IQR = 7-10.7]) were enrolled in the study, including 46 children with a laboratory-confirmed previous COVID-19 (SARS-CoV-2-recovered) and 14 SARS-CoV-2-naïve participants defined as the absence of antigen-specific antibodies before vaccination. SARS-CoV-2-recovered participants recorded higher anti-S-RBD IgG and Wild-type and Omicron BA.2 NAbs titers than SARS-CoV-2-naïve participants at both 1- and 6-months after vaccination. Antibody titers correlated with T (Tregs) and B (Bregs) regulatory cell frequencies in SARS-CoV-2-recovered children. Both SARS-CoV-2-recovered and SARS-CoV-2-naïve participants decreased antibody titers by approximately 100 to 250 % from 1 to 6 months. While children with immunocompromising underlying conditions developed immune responses comparable to those of healthy children, solid organ transplant recipients exhibited lower levels of NAbs and anti-S-RBD IgG titers, as well as reduced frequencies of Tregs and Bregs. CONCLUSIONS: mRNA vaccination triggered a higher production of specific anti-SARS-CoV-2 antibodies along with increased levels of regulatory cells in children with previous SARS-CoV-2 infection up to the following 6 months. These findings provide insights into boosting pre-existing immunity.

SARS-CoV-2 in Animal Companions: A Serosurvey in Three Regions of Southern Italy.

Several animal species have been found to be susceptible to SARS-CoV-2 infection. The occurrence of infection in dogs and cats living in close contact with owners deserves particular attention from public health authorities in a One Health approach. In this study, we conducted serological screening to identify SARS-CoV-2 exposure in the sera from dogs and cats in three regions of southern Italy sampled during the years 2021 and 2022. We collected 100 serum samples in 2021 (89 from dogs and 11 from cats) and 640 in 2022 (577 from dogs and 63 from cats). Overall, the ELISA positivity rate was found to be 2.7% (20/740), with higher seroprevalence in dogs. Serum neutralization tests confirmed positivity only in two samples collected from dogs, and the assays, performed with serologically distinct SARS-CoV-2 variants, showed variant-specific positivity. This paper shows that monitoring SARS-CoV-2 exposure in animals might be affected by the viral antigenic evolution, which requires continuous updates to the serological tests used. Serological surveys are useful in understanding the true extent of exposure occurring in specific animal populations, not suffering the same limitations as molecular tests, and could help in identifying the infecting virus if tests able to characterize the immune response are used. The use of variant-specific validated serological methods should always be considered in serosurvey studies in order to determine the real impact of emerging variants on animal populations and its implications for veterinary and human health, as well as to identify potential reservoirs of the virus and its evolutionary changes.

Interplay of Modified Sialic Acid Inhibitors and the Human Parainfluenza Virus 1 Hemagglutinin-Neuraminidase Active Site.

In the search for effective antivirals against Paramyxoviridae, the dynamics of human parainfluenza virus type 1 hemagglutinin-neuraminidase (hPIV1-HN) inhibition offers a promising perspective. This study focuses on the potential of C5- and C4-modified 2,3-unsaturated sialic acid (DANA) inhibitors and highlights their interaction with the hPIV1-HN enzyme. We show that a strategic substitution, replacing the C5 isopropyl group in BCX 2798 with a trifluoroacetyl function, increases inhibitory potency 3- to 4-fold. At the same time, we explore the special properties of the catalytic site of hPIV1-HN, which harbors only small substituents and favors a C4 sulfonylamido function over a carbonyl function, in contrast to the C4 pocket of Newcastle disease virus hemagglutinin-neuraminidase (NDV-HN). Based on these findings, we present a newly identified potent inhibitor that has the preferred C5 trifluoroacetamido and C4 trifluorosulfonylamide groups. The results of this study pave the way for a deeper understanding of the C4 and C5 binding pockets of hPIV1-HN and promote the development of new, more selective inhibitors.

Host Response of Syrian Hamster to SARS-CoV-2 Infection including Differences with Humans and between Sexes.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has highlighted the importance of having proper tools and models to study the pathophysiology of emerging infectious diseases to test therapeutic protocols, assess changes in viral phenotypes, and evaluate the effects of viral evolution. This study provided a comprehensive characterization of the Syrian hamster (Mesocricetus auratus) as an animal model for SARS-CoV-2 infection using different approaches (description of clinical signs, viral load, receptor profiling, and host immune response) and targeting four different organs (lungs, intestine, brain, and PBMCs). Our data showed that both male and female hamsters were susceptible to the infection and developed a disease similar to the one observed in patients with COVID-19 that included moderate to severe pulmonary lesions, inflammation, and recruitment of the immune system in the lungs and at the systemic level. However, all animals recovered within 14 days without developing the severe pathology seen in humans, and none of them died. We found faint evidence for intestinal and neurological tropism associated with the absence of lesions and a minimal host response in intestines and brains, which highlighted another crucial difference with the multiorgan impairment of severe COVID-19. When comparing male and female hamsters, we observed that males sustained higher viral RNA shedding and replication in the lungs, suffered from more severe symptoms and histopathological lesions, and triggered higher pulmonary inflammation. Overall, these data confirmed the Syrian hamster as a suitable model for mild to moderate COVID-19 and reflected sex-related differences in the response against the virus observed in humans.

Design, Synthesis, and Antiviral Evaluation of Sialic Acid Derivatives as Inhibitors of Newcastle Disease Virus Hemagglutinin-Neuraminidase: A Translational Study on Human Parainfluenza Viruses.

Global infections with viruses belonging to the Paramyxoviridae, such as Newcastle disease virus (NDV) or human parainfluenza viruses (hPIVs), pose a serious threat to animal and human health. NDV-HN and hPIVs-HN (HN hemagglutinin-neuraminidase) share a high degree of similarity in catalytic site structures; therefore, the development of an efficient experimental NDV host model (chicken) may be informative for evaluating the efficacy of hPIVs-HN inhibitors. As part of the broad research in pursuit of this goal and as an extension of our published work on antiviral drug development, we report here the biological results obtained with some newly synthesized C4- and C5-substituted 2,3-unsaturated sialic acid derivatives against NDV. All developed compounds showed high neuraminidase inhibitory activity (IC50 0.03-13 µM). Four molecules (9, 10, 23, 24) confirmed their high in vitro inhibitory activity, which caused a significant reduction of NDV infection in Vero cells, accompanied by very low toxicity.

SARS-CoV-2 intra-host evolution during prolonged infection in an immunocompromised patient.

OBJECTIVES: Intra-host SARS-CoV-2 evolution during chronic infection in immunocompromised hosts has been suggested as being the possible trigger of the emergence of new variants. METHODS: Using a deep sequencing approach, we investigated the SARS-CoV-2 intra-host genetic evolution in a patient with HIV over a period of 109 days. RESULTS: Sequencing of nasopharyngeal swabs at three time points demonstrated dynamic changes in the viral population, with the emergence of 26 amino acid mutations and two deletions, 57% of them in the Spike protein. Such a combination of mutations has never been observed in other SARS-CoV-2 lineages detected so far. CONCLUSION: Our data confirm that persistent infection in certain immunocompromised individuals for a long time may favor the dangerous emergence of new SARS-CoV-2 variants with immune evasion properties.

Detailed epitope mapping of SARS-CoV-2 nucleoprotein reveals specific immunoresponse in cats and dogs housed with COVID-19 patients.

Since the initial emergence in December 2019, the novel Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has been reported in over 200 countries, representing an unprecedented challenge related to disease control worldwide. In this context, cases of human to animal transmission have been reported, raising concern about the potential role of companion animals in the pandemic and stressing the need for reliable animal testing. In the study, a detailed epitope mapping of SARS-CoV-2 nucleoprotein, using both human and pet sera, allowed the identification of the most antigenic region in the C-terminus domain of the protein, which was used to develop an experimental double antigen-based ELISA. A panel of pre-pandemic sera and sera of animals immunized against (or naturally infected with) related coronaviruses was used to assess assay specificity at 99.5%. Positive sera belonging to animals housed with COVID-19 patients were confirmed with the experimental double-antigen ELISA using Plaque Reduction Neutralization test (PRNT) test as gold standard. The availability of a serological assay that targets a highly specific viral antigen represents a valuable tool for multispecies monitoring of Coronavirus Disease 2019 (COVID-19) infection in susceptible animals.

Neutralizing antibody titers six months after Comirnaty vaccination: kinetics and comparison with SARS-CoV-2 immunoassays.

OBJECTIVES: mRNA vaccines, including Comirnaty (BNT162b2 mRNA, BioNTech-Pfizer), elicit high IgG and neutralizing antibody (NAb) responses after the second dose, but the progressive decrease in serum antibodies against SARS-CoV-2 following vaccination have raised questions concerning long-term immunity, decreased antibody levels being associated with breakthrough infections after vaccination, prompting the consideration of booster doses. METHODS: A total number of 189 Padua University-Hospital healthcare workers (HCW) who had received a second vaccine dose were asked to collect serum samples for determining Ab at 12 (t12) and 28 (t28) days, and 6 months (t6m) after their first Comirnaty/BNT162b2 inoculation. Ab titers were measured with plaque reduction neutralization test (PRNT), and three chemiluminescent immunoassays, targeting the receptor binding domain (RBD), the trimeric Spike protein (trimeric-S), and surrogate viral neutralization tests (sVNT). RESULTS: The median percentages (interquartile range) for decrease in antibodies values 6 months after the first dose were 86.8% (67.1-92.8%) for S-RBD IgG, 82% (58.6-89.3%) for trimeric-S, 70.4% (34.5-86.4%) for VNT-Nab, 75% (50-87.5%) for PRNT50 and 75% (50-93.7%) for PRNT90. At 6 months, neither PRNT titers nor VNT-Nab and S-RBD IgG bAb levels correlated with age (p=0.078) or gender (p=0.938), while they were correlated with previous infection (p<0.001). CONCLUSIONS: After 6 months, a method-independent reduction of around 90% in anti-SARS-CoV-2 antibodies was detected, while no significant differences were found between values of males and females aged between 24 and 65 years without compromised health status. Further efforts to improve analytical harmonization and standardization are needed.

SARS-CoV-2 infection and replication in human gastric organoids.

COVID-19 typically manifests as a respiratory illness, but several clinical reports have described gastrointestinal symptoms. This is particularly true in children in whom gastrointestinal symptoms are frequent and viral shedding outlasts viral clearance from the respiratory system. These observations raise the question of whether the virus can replicate within the stomach. Here we generate gastric organoids from fetal, pediatric, and adult biopsies as in vitro models of SARS-CoV-2 infection. To facilitate infection, we induce reverse polarity in the gastric organoids. We find that the pediatric and late fetal gastric organoids are susceptible to infection with SARS-CoV-2, while viral replication is significantly lower in undifferentiated organoids of early fetal and adult origin. We demonstrate that adult gastric organoids are more susceptible to infection following differentiation. We perform transcriptomic analysis to reveal a moderate innate antiviral response and a lack of differentially expressed genes belonging to the interferon family. Collectively, we show that the virus can efficiently infect the gastric epithelium, suggesting that the stomach might have an active role in fecal-oral SARS-CoV-2 transmission.

SARS-CoV-2 neutralizing antibodies after one or two doses of Comirnaty (BNT162b2, BioNTech/Pfizer): Kinetics and comparison with chemiluminescent assays.

BACKGROUND: Studies evaluating neutralizing antibody (NAb) after BNT162b2 vaccine are scarce. We therefore compared NAb using the plaque reduction neutralization test (PRNT) in vaccinated subjects, with those from five chemiluminescent (CLIA) assays, two targeting ACE and S-RBD interaction. METHODS: Sera from 174 completely Comirnaty/BNT162b2 vaccinated healthcare workers (HCW) were evaluated at t12 and t28. NAb titers at low (PRNT50) or high (PRNT90) stringency were compared with: Liaison SARS-CoV-2 Trimeric-S IgG, Elecsys S-RBD Ab, Maglumi SARS-CoV-2 S-RBD IgG and SARS-CoV-2 Nab; iFlash 2019-nCoV NAb. RESULTS: Neither PRNT50 nor PRNT90 correlated with age (range, 24-65 years); no significant differences were found for gender. PRNT50 and PRNT90 seropositive titers (≥1:20) were 43 (24.7%) and 15 (8.6%) at t12 and 167 (95.9%) and 149 (85.6%) at t28. CLIA results at t28 were uncorrelated with age, apart from Elecsys S-RBD Ab (r = -0.164, p = 0.046). Gender differences were found for Maglumi SARS-CoV-2 S-RBD IgG (p = 0.037) and Maglumi NAb (p = 0.046). Considering PRNT50 at thresholds of 1:20 (or 1:40) and 1:160 (or 1:320), corresponding to different immune protective levels, CLIA cut-offs have been identified. CONCLUSIONS: Comirnaty/BNT162b2 elicits strong NAb production, especially 28 days after first inoculum. Differences in correlation between Nab titers and circulating antibodies measured by 5 immunoassays have been found, being stronger the correlation for Maglumi Nab.

Asymptomatic and Mild SARS-CoV-2 Infections Elicit Lower Immune Activation and Higher Specific Neutralizing Antibodies in Children Than in Adults.

Background: The immune response plays a pivotal role in dictating the clinical outcome in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected adults, but it is still poorly investigated in the pediatric population. Methods: Of 209 enrolled subjects, 155 patients were confirmed by PCR and/or serology as having coronavirus disease 2019 (COVID-19). Blood samples were obtained at a median of 2.8 (interquartile, 2.1-3.7) and 6.1 (5.3-7.2) months after baseline (symptom onset and/or first positive virus detection). The immune profiles of activation, senescence, exhaustion, and regulatory cells were analyzed by flow cytometry. Neutralizing antibodies (nAbs) were detected by a plaque reduction neutralization test. In available nasopharyngeal swabs at baseline, SARS-CoV-2 levels were quantified by digital droplet PCR (ddPCR). Results: Overall, COVID-19 patients had higher levels of immune activation, exhaustion, and regulatory cells compared to non-COVID-19 subjects. Within the COVID-19 group, activated and senescent cells were higher in adults than in children and inversely correlated with the nAbs levels. Conversely, Tregs and Bregs regulatory cells were higher in COVID-19 children compared to adults and positively correlated with nAbs. Higher immune activation still persisted in adults after 6 months of infection, while children maintained higher levels of regulatory cells. SARS-CoV-2 levels did not differ among age classes. Conclusions: Adults displayed higher immune activation and lower production of anti-SARS-CoV-2 nAbs than children. The different immune response was not related to different viral load. The higher expression of regulatory cells in children may contribute to reduce the immune activation, thus leading to a greater specific response against the virus.

Mild SARS-CoV-2 Infections and Neutralizing Antibody Titers.

BACKGROUND: Recent evidence suggests that neutralizing antibodies (nAbs) to severe acute respiratory syndrome coronavirus 2 may persist over time; however, knowledge regarding pediatric subjects is limited. METHODS: A single-center, prospective observational study was conducted on 57 family clusters of coronavirus disease 2019, including children of neonatal and pediatric age attending the University Hospital of Padua (Italy). For each patient, blood samples were collected for both the quantification of nAbs through a plaque reduction neutralizing test and the detection of antinucleocapsid-spike protein immunoglobulin G and/or immunoglobulin M. RESULTS: We analyzed 283 blood samples collected from 152 confirmed coronavirus disease 2019 cases (82 parents and 70 children or older siblings of median age of 8 years, interquartile range: 4-13), presenting asymptomatic or with mildly symptomatic disease. Despite the decrease of immunoglobulin G over time, nAbs were found to persist up to 7 to 8 months in children, whereas adults recorded a modest declining trend. Interestingly, children aged <6 years, and, in particular, those aged <3 years, developed higher long-lasting levels of nAbs compared with older siblings and/or adults. CONCLUSIONS: Mild and asymptomatic severe acute respiratory syndrome coronavirus 2 infections in family clusters elicited higher nAbs among children.

Analytical and clinical performances of a SARS-CoV-2 S-RBD IgG assay: comparison with neutralization titers.

OBJECTIVES: SARS-CoV-2 serology presents an important role in several aspects of COVID-19 pandemic. Immunoassays performances have to be accurately evaluated and correlated with neutralizing antibodies. We investigated the analytical and clinical performances of a SARS-CoV-2 RBD IgG assay, automated on a high throughput platform, and the correlation of the antibodies (Ab) levels with the plaque reduction neutralization (PRNT50) Ab titers. METHODS: A series of 546 samples were evaluated by SARS-CoV-2 RBD IgG assay (Snibe diagnostics), including 171 negative and 168 positive SARS-CoV-2 subjects and a further group of 207 subjects of the COVID-19 family clusters follow-up cohort. RESULTS: Assay imprecision ranged from 3.98 to 12.18% being satisfactory at low and medium levels; linearity was excellent in all the measurement range. Considering specimens collected after 14 days post symptoms onset, overall sensitivity and specificity were 99.0 and 92.5%, respectively. A total of 281 leftover samples results of the PRNT50 test were available. An elevated correlation was obtained between the SARS-CoV-2 RBD IgG assay and the PRNT50 titer at univariate (ρ=0.689) and multivariate (ρ=0.712) analyses. CONCLUSIONS: SARS-CoV-2 S-RBD IgG assay shows satisfactory analytical and clinical performances, and a strong correlation with sera neutralizing activity.

Virological and immunological features of SARS-CoV-2-infected children who develop neutralizing antibodies.

As the global COVID-19 pandemic progresses, it is paramount to gain knowledge on adaptive immunity to SARS-CoV-2 in children to define immune correlates of protection upon immunization or infection. We analyzed anti-SARS-CoV-2 antibodies and their neutralizing activity (PRNT) in 66 COVID-19-infected children at 7 (±2) days after symptom onset. Individuals with specific humoral responses presented faster virus clearance and lower viral load associated with a reduced in vitro infectivity. We demonstrated that the frequencies of SARS-CoV-2-specific CD4+CD40L+ T cells and Spike-specific B cells were associated with the anti-SARS-CoV-2 antibodies and the magnitude of neutralizing activity. The plasma proteome confirmed the association between cellular and humoral SARS-CoV-2 immunity, and PRNT+ patients show higher viral signal transduction molecules (SLAMF1, CD244, CLEC4G). This work sheds lights on cellular and humoral anti-SARS-CoV-2 responses in children, which may drive future vaccination trial endpoints and quarantine measures policies.

Analytical and clinical performances of five immunoassays for the detection of SARS-CoV-2 antibodies in comparison with neutralization activity.

BACKGROUND: Reliable high-throughput serological assays for SARS-CoV-2 antibodies are urgently needed for the effective containment of the COVID-19 pandemic, as it is of crucial importance to understand the strength and duration of immunity after infection, and to make informed decisions concerning the activation or discontinuation of physical distancing restrictions. METHODS: In 184 serum samples from 130 COVID-19 patients and 54 SARS-CoV-2 negative subjects, the analytical and clinical performances of four commercially available chemiluminescent assays (Abbott SARS-Cov-2 IgG, Roche Elecsys anti-SARS-CoV-2, Ortho SARS-CoV-2 total and IgG) and one enzyme-linked immunosorbent assay (Diesse ENZY-WELL SARS-CoV-2 IgG) were evaluated and compared with the neutralization activity achieved using the plaque reduction neutralization test (PRNT). FINDINGS: Precision results ranged from 0.9% to 11.8% for all assays. Elecsys anti-SARS-CoV-2 demonstrated linearity of results at concentrations within the cut-off value. Overall, sensitivity ranged from 78.5 to 87.7%, and specificity, from 97.6 to 100%. On limiting the analysis to samples collected 12 days after onset of symptoms, the sensitivity of all assays increased, the highest value (95.2%) being obtained with VITRO Anti-SARS-CoV-2 Total and Architect SARS-CoV-2 IgG. The strongest PRNT50 correlation with antibody levels was obtained with ENZY-Well SARS-CoV-2 IgG (R2adj = 0.569). INTERPRETATION: The results confirmed that all immunoassays had an excellent specificity, whereas sensitivity varied across immunoassays, depending strongly on the time interval between symptoms onset and sample collection. Further studies should be conducted to achieve a stronger correlation between antibody measurement and PRNT50 in order to obtain useful information for providing a better management of COVID-19 patients, effective passive antibody therapy, and developing a vaccine against the SARS-CoV-2 virus. FUNDING: None.

maxRatio improves the detection of samples with abnormal amplification profiles on QIAgen's artus HIV-1 qPCR assay.

Background: Accurate viral load (VL) determination is paramount to determine the efficacy of anti-HIV-1 therapy. The conventional method used, fit-point (FP), assumes an equal efficiency in the polymerase chain reaction (PCR) among samples that might not hold for low-input templates. An alternative approach, maxRatio, was introduced to compensate for inhibition in PCR. Methods: Herein, we assessed whether maxRatio could improve VL quantification using 2,544 QIAgen artus HI virus-1 RT-PCR reactions. The assay's standard dilutions were used to build external standard curves with either FP or maxRatio that re-calculated the VLs. Results: FP and maxRatio were highly comparable (Pearson's ρ=0.994, Cohen's  κ=0.885), and the combination of the two methods identified samples (n=41) with aberrant amplification profiles. Conclusions: The combination of maxRatio and FP could improve the predictive value of the assay.

Helicobacter pylori Affects the Antigen Presentation Activity of Macrophages Modulating the Expression of the Immune Receptor CD300E through miR-4270.

Helicobacter pylori (Hp) is a Gram-negative bacterium that infects the human gastric mucosa, leading to chronic inflammation. If not eradicated with antibiotic treatment, the bacterium persists in the human stomach for decades increasing the risk to develop chronic gastritis, gastroduodenal ulcer, and gastric adenocarcinoma. The lifelong persistence of Hp in the human stomach suggests that the host response fails to clear the infection. It has been recently shown that during Hp infection phagocytic cells promote high Hp loads rather than contributing to bacterial clearance. Within these cells Hp survives in "megasomes," large structures arising from homotypic fusion of phagosomes, but the mechanism that Hp employs to avoid phagocytic killing is not completely understood. Here, we show that Hp infection induces the downregulation of specific microRNAs involved in the regulation of transcripts codifying for inflammatory proteins. miR-4270 targets the most upregulated gene: the immune receptor CD300E, whose expression is strictly dependent on Hp infection. CD300E engagement enhances the pro-inflammatory potential of macrophages, but in parallel it affects their ability to express and expose MHC class II molecules on the plasma membrane, without altering phagocytosis. This effect compromises the possibility for effector T cells to recognize and activate the killing potential of macrophages, which, in turn would become a survival niche for the bacterium. Taken together, our data add another piece to the complicate puzzle represented by the long-life coexistence between Hp and the human host and contribute with new insights toward understanding the regulation and function of the immune receptor CD300E.

Helicobacter pylori antigenic Lpp20 is a structural homologue of Tipα and promotes epithelial-mesenchymal transition.

BACKGROUND: Helicobacter pylori is a bacterium that affects about 50% of the world population and, despite being often asymptomatic, it is responsible of several gastric diseases, from gastritis to gastric cancer. The protein Lpp20 (HP1456) plays an important role in bacterium survival and host colonization, but the possibility that it might be involved in the etiology of H. pylori-related disorders is an unexplored issue. Lpp20 is a lipoprotein bound to the external membrane of the bacterium, but it is also secreted inside vesicles along with other two proteins of the same operon, i.e. HP1454 and HP1457. RESULTS: In this study we determined the crystal structure of Lpp20 and we found that it has a fold similar to a carcinogenic factor released by H. pylori, namely Tipα. We demonstrate that Lpp20 promotes cell migration and E-cadherin down-regulation in gastric cancer cells, two events recalling the epithelial-mesenchymal transition (EMT) process. Differently from Tipα, Lpp20 also stimulates cell proliferation. CONCLUSIONS: This identifies Lpp20 as a new pathogenic factor produced by H. pylori that promotes EMT and thereby the progression of cancer to the metastatic state.