Up-regulation of the monocyte chemotactic protein-3 in sera from bone marrow transplanted children with torquetenovirus infection

Torquetenovirus (TTV) represents a commensal human virus producing life-long viremia in approximately 80% of healthy individuals of all ages. A potential pathogenic role for TTV has been suggested in immunocompromised patients with hepatitis of unknown etiology sustained by strong proinflammatory cytokines

The Bacterial DNA Profiling of Chorionic Villi and Amniotic Fluids Reveals Overlaps with Maternal Oral, Vaginal, and Gut Microbiomes

The in utero microbiome hypothesis has been long debated. This hypothesis will change our comprehension of the pioneer human microbiome if proved correct. In 60 uncomplicated pregnancies, we profiled the microbiome of chorionic villi (CV) and amniotic fluids (AF) in relation to maternal saliva, rectum, and vagina and the soluble cytokines cascade in the vagina, CV and AF. In our series, 12/37 (32%) AF and 10/23 (44%) CV tested positive for bacterial DNA. CV and AF harbored bacterial DNA of Streptococcus and Lactobacillus, overlapping that of the matched oral and vaginal niches, which showed a dysbiotic microbiome. In these pregnant women, the immune profiling revealed an immune hyporesponsiveness in the vagina and a high intraamniotic concentration of inflammatory cytokines. To understand the eventual role of bacterial colonization of the CV and AF and the associated immune response in the pregnancy outcome, further appropriate studies are needed. In this context, further studies should highlight if the hematogenous route could justify the spread of bacterial DNA from the oral microbiome to the placenta and if vaginal dysbiosis could favor the likelihood of identifying CV and AF positive for bacterial DNA

Genital Dysbiosis and Different Systemic Immune Responses Based on the Trimester of Pregnancy in SARS-CoV-2 Infection

Respiratory infections are common in pregnancy with conflicting evidence supporting their association with neonatal congenital anomalies, especially during the first trimester. We profiled cytokine and chemokine systemic responses in 242 pregnant women and their newborns after SARS-CoV-2 infection, acquired in different trimesters. Also, we tested transplacental IgG passage and maternal vaginal–rectal microbiomes. IgG transplacental passage was evident, especially with infection acquired in the first trimester. G-CSF concentration—involved in immune cell recruitment—decreased in infected women compared to uninfected ones: a beneficial event for the reduction of inflammation but detrimental to ability to fight infections at birth. The later the infection was acquired, the higher the systemic concentration of IL-8, IP-10, and MCP-1, associated with COVID-19 disease severity. All infected women showed dysbiosis of vaginal and rectal microbiomes, compared to uninfected ones. Two newborns tested positive for SARS-CoV-2 within the first 48 h of life. Notably, their mothers had acute infection at delivery. Although respiratory infections in pregnancy are reported to affect babies’ health, with SARS-CoV-2 acquired early during gestation this risk seems low because of the maternal immune response. The observed vaginal and rectal dysbiosis could be relevant for neonatal microbiome establishment, although in our series immediate neonatal outcomes were reassuring

Clinical, anamnestic, and sociodemographic predictors of positive SARS-CoV-2 testing in children: A cross sectional study in a tertiary hospital in Italy

ObjectivesWe aimed to identify clinical, anamnestic, and sociodemographic characteristics associated with a positive swab for SARS-CoV2, and to provide a predictive score to identify at risk population in children aged 2-14 years attending school and tested for clinical symptoms of COVID-19.DesignCross sectional study.SettingOutpatient clinic of the IRCCS Burlo Garofolo, a maternal and child health tertiary care hospital and research centre in Italy.Data collection and analysisData were collected through a predefined form, filled out by parents, and gathered information on sociodemographic characteristics, and specific symptoms, which were analysed to determine their association with a positive SARS-CoV-2 swab. The regression coefficients of the variables included in the multivariate analysis were further used in the calculation of a predictive score of the positive or negative test.ResultsBetween September 20th and December 23rd 2020, from 1484 children included in the study, 127 (8.6%) tested positive. In the multivariate analysis, the variables retained by the model were the presence of contact with a cohabiting, non-cohabiting or unspecified symptomatic case (respectively OR 37.2, 95% CI 20.1-68.7; 5.1, 95% CI 2.7-9.6; 15.6, 95% CI 7.3-33.2); female sex (OR 1.49, 95% CI 1.0-2.3); age (6-10 years old: OR 3.2, 95% CI 1.7-6.1 p<0.001; >10 years old: OR 4.8, 95% CI 2.7-8.8 p<0.001); fever (OR 3.9, 95% CI 2.3-6.4); chills (OR 1.9, 95% CI 1.1-3.3); headache (OR 1.45, 95% CI 0.9-2.4); ageusia (OR 1.3, 95% CI 0.54.0); sore throat (OR 0.48, 95% CI 0.3-0.8); earache (OR 0.4, 95% CI 0.1-1.3); rhinorrhoea (OR 0.8, 95% CI 0.5-1.3); and diarrhoea (OR 0.52, 95% CI 0.2-1.1). The predictive score based on these variables generated 93% sensitivity and 99% negative predictive value.ConclusionsThe timely identification of SARS-CoV2 cases among children is useful to reduce the dissemination of the disease and its related burden. The predictive score may be adopted in a public health perspective to rapidly identify at risk children

Characterization of chorionic villus microbiome in the first trimester and of the amniotic fluid microbiome in the second trimester of pregnancy: relationship with the vaginal, rectal and oral maternal microbiome.

La presenza di un microbioma nell'utero sano è un argomento molto discusso. Per lungo tempo si è ritenuto che la placenta fosse un organo sterile. Recenti studi condotti utilizzando tecniche di sequenziamento di nuova generazione su tessuto uterino e placentale hanno dimostrato che sia l’utero che la placenta ospitano un “proprio microbioma”. Pertanto, lo scopo di questo studio è di caratterizzare, utilizzando la tecnica del 16S rRNA Next-Generation Sequencing, il batterioma feto-placentare nelle prime fasi della gravidanza e di confrontarlo con quello di altri distretti materni (rettale, vaginale, orale). Inoltre, è stato studiato il profilo immunitario del complesso feto-placentare e dell'ambiente vaginale. In questo studio sono state arruolate 60 donne, afferenti al Burlo Garofolo per l'esecuzione di villocentesi o amniocentesi. È stato analizzato un totale di 240 campioni biologici, composti da campioni di villi coriali (CVS, n= 23), e campioni di liquido amniotico (AF, n=37), e i corrispondenti tamponi vaginali, tamponi rettali e campioni di saliva. Dall'analisi del microbioma, sono risultati positivi per la presenza di DNA batterico 12 (32%) campioni AF e 10 (44%) campioni CVS. I batteri identificati nei campioni positivi di CVS e di AF appartenevano a patogeni commensali e opportunisti del tratto riproduttivo e del cavo orale (Lactobacillus e Streptococcus). Nell’evidenziare un possibile microbioma predisponente alla colonizzazione di CVS e AF negli altri distretti materni, è stata osservata una diminuzione del probiotico S. salivarius nei campioni di saliva abbinati ai CVS e AF risultati positivi per la presenza di batteri. È noto che questa specie probiotica è in grado di inibire la risposta infiammatoria indotta da disbiosi orale e dalla presenza di patogeni associati a malattie parodontali. Nei campioni vaginali, il risultato più evidente è stato la diminuzione di abbondanza relativa di Lactobacillus crisptaus (L. crispatus) nei tamponi vaginali abbinati ai campioni CVS/AF risultati positivi per la presenza di batteri. Come dimostrato da studi precedenti, L. crispatus mostra un potenziale ruolo nell'inibire la disbiosi vaginale, e la diminuzione di questo batterio nell'ambiente vaginale durante la gravidanza è stata associata a un rischio maggiore di infezione e parto pre-termine. Per confrontare il profilo immunitario del complesso feto-placentare con quello dell'ambiente vaginale, è stata misurata la concentrazione di 27 proteine immunitarie solubili, comprese citochine Th1/proinfiammatorie e Th2/antinfiammatorie, chemochine e fattori trofici, nei campioni CVS/AF e nei tamponi vaginali. É stato osservato un significativo aumento di concentrazione di due fattori pro-infiammatori, IL-8 e G-CSF, in presenza di DNA batterico nei campioni di liquido amniotico. Al contrario, è stata osservata un'iporeattività immunitaria nei tamponi vaginali abbinati ai campioni di liquido amniotico risultati positivi alla presenza di batteri, suggerendo una diversa risposta immunitaria nei due ambienti. In conclusione, questo studio conferma, per la prima volta, la presenza di DNA batterico nel complesso feto-placentare nelle prime fasi della gravidanza, supportando l'ipotesi di un microbioma in utero. I risultati del nostro studio pilota dimostrano che la placenta può essere colonizzata non solo dalla via urogenitale ma anche dalla via orale, suggerendo un accesso ematogeno. Inoltre, ipotizziamo che l'iporeattività immunitaria osservata nell'ambiente vaginale possa contribuire alla traslocazione del DNA batterico nel liquido amniotico dove, in assenza di un'infezione in corso, è stata rivelata la sovra-regolazione di due citochine infiammatorie. Comunque, sono necessari ulteriori studi per comprendere i cambiamenti dei processi metabolici indotti dal microbioma feto-placentale e il loro ruolo nell'esito della gravidanza.The presence of a microbiome in healthy uterus has long been a matter of debate. Until a few years ago, the placenta was thought to be a sterile tissue, therefore, the amniotic cavity and the fetus were also supposed to be sterile. A multitude of recent studies exploiting high-throughput sequencing technologies has challenged this paradigm, proposing that the placenta harbors a unique microbiome, neither the fetus, therefore, nor the amniotic fluid are sterile, and that acquisition of microbes begins in utero. In this context, the aim of this study is to characterize, using the Next-Generation Sequencing technique, fetus-placental bacteriome in the early stages of pregnancy and to compare it with that of other maternal districts (rectal, vaginal, oral). In addition, the immune profile of the fetus-placental complex and vaginal environment was investigated. In this study, 60 women, afferent to IRCCS Burlo Garofolo for the execution of villocentesis or amniocentesis, were enrolled. A total of 240 biological samples was analyzed, including chorionic villi (CVS, n= 23) and amniotic fluid samples (AF, n=37), and the matched samples including vaginal swabs, rectal swabs, and saliva samples. From the microbiome analysis, 12 (32%) AF samples and 10 (44%) CVS samples tested positive for the presence of bacterial DNA. The identified bacteria in the positive CVS and AF samples belonged to commensal and opportunistic pathogens of the reproductive tract and of the oral cavity (Lactobacillus and Streptococcus). When looking at a possible predisposing microbiome of maternal body districts to the colonization of CVS and AF, we found a decrease of probiotic Streptococcus salivarius in saliva samples matched to CVS and AF tested positive for the presence of bacterial DNA. To note, this probiotic species is able to inhibit immune activation by oral dysbiosis and periodontal disease pathogens. In vaginal samples, the most evident result was the decrease of Lactobacillus crisptaus in the samples matched to the CVS/AF samples that tested positive for the presence of bacteria compared to the samples matched to the negative CVS/AF samples. As shown by previous studies, L. crispatus shows a potential role to inhibit dysbiotic vaginal microbiome and infectious inflammation. Lastly, to compare the immune profile of fetus-placental complex and that of vaginal environment, the concentration of 27 soluble immune proteins, was measured in the AF/CVS samples and vaginal swabs. In particular, markers of intraamniotic inflammation, such as IL-8 and G-CSF, were increased in presence of bacterial DNA in the AF samples. Conversely, an immune hyporesponsiveness in the vaginal swabs matched to the positive AF samples was observed, suggesting, in according to previous studies, that the mother, placenta, and fetus all possess unique innate immune systems. To conclude, this study, confirms, for the first time, the presence of bacterial DNA in fetus-placental complex in the early stages of pregnancy, supporting the hypothesis of an in utero microbiome. The results from our pilot study show that the placenta can be colonized not only from the urogenital route but also from the oral route, suggesting hematogenous access. In addition, we speculate that the immune hyporesponsiveness in the vaginal milieu could contribute to the bacterial DNA translocation in the amniotic fluid where, in absence of an ongoing infection, the up-regulation of inflammatory cytokines was revealed. Further studies are needed to understand the variations in placental microbiome-induced metabolic pathways and their role in pregnancy outcomes

RANTES: una chemochina utile come marcatore diagnostico del mesotelioma

Immune mediators are likely to be relevant for the biological response to asbestos exposure. The aim of this study was to investigate the association between immune mediators involved in inflammation, cell survival and angiogenesis, and asbestos-related diseases in workers from a coastal area of North-East Italy with a high incidence of pleural malignant mesothelioma (PMM). Materials and methods: A selected custom set of 12 soluble mediators was evaluated with a Luminex platform in sera, pleural fluid and mesothelioma biopsies from 123 asbestos-exposed workers (38 free from pleural-pulmonary disorders, 46 with non-malignant asbestos diseases, 39 with PMM) and in sera from 33 healthy controls from the same territorial area. Results: Increased immune mediator concentrations were observed in the sera of the asbestos-exposed workers compared to controls for human fibroblast growth factor (FGF-b), vascular endothelial growth factor (VEGF), CCL5 (RANTES), CXCL10 (IP-10), CLEC11A (SCGF-b), CCL27 (CTACK), CCL11 (EOTAXIN), IL-5 and IL-6 (p < 0.001). The chemokines IP-10 and RANTES were associated with the severity of asbestosrelated diseases. In the workers with PMM, the immune proteins secreted by mesothelioma biopsies showed detectable levels of RANTES, VEGF, and IP-10. In the same workers with PMM, a significant relationship between serum and pleural fluid concentrations was found for RANTES alone. Conclusions: Occupational exposure to asbestos seems to drive the production of specific growth factors dually involved in the early inflammatory response and in pro-tumoral activity before clinical evidence of related disorders, suggesting that their over-expression may precede the onset of asbestos-related diseases. These findings suggest that some chemokines may have a prognostic role in the progression of asbestos-related diseases and could be used for the health surveillance of either workers with an occupational history of asbestos exposure or patients affected by non-malignant asbestos-related diseases

CV/BKV and SV40 viral load in lymphoid tissues of young immunocompetent children from an area of North-East Italy

Polyomavirus infection occurring during childhood is followed by a lifelong latency in immunocompetent subjects. The major site of polyomavirus persistence are the uroepithelial cells which leads to oral transmission. It has recently been hypothesized that tonsils could be a possible reservoir. The role of tonsil, adenoid, and peripheral blood mononuclear cells (PBMCs) as possible sites of JCV, BKV, and SV40 latency in young healthy children was assessed. Two hundred fifteen fresh specimens, including 57 tonsil, 80 adenoid, and 78 PBMC samples from 80 immunocompetent children (mean age 4.8 years) were analyzed to determine the viral load by quantitative real-time PCR. The human herpes virus 6 (HHV-6) was tested as a lymphotropic reference virus. Polyomavirus was detected in 5/ 80 (6.2%) children while HHV-6 infection affected 27/80 children (33.7%) (P<0.001). SV40 was detected in one adenoid sample, while footprints of BKV were found in one adenoid and three tonsil samples. JCV was never found in all samples. Polyomavirus sequences were not detected in the 78 blood samples. One adenoid and two tonsils from three children (1.4%) were positive for both polyomavirus and HHV-6. Infections were characterized by low replication rates ranging typically from 110e2/5.510e4 to 6.810e3/8.510e4 viral copies/number of cells. In conclusion, tonsils and adenoids of children could effectively harbor BKV and SV40, although only very few cells proved to be infected. Nevertheless, the low prevalence of polyomavirus, in comparison with the lymphotropic HHV-6, suggests that these tissues are unlikely to be the preferred site of polyomavirus latency, at least in younger children

Influence of Vaginal Lactoferrin Administration on Amniotic Fluid Cytokines and its Role Against Inflammatory Complications of Pregnancy

Background: An altered amniotic cytokine profile has been reported in inflammatory pregnancy complications with a leading role for IL-6, a marker of the foetal systemic inflammatory response. Up to this date there is no exhaustive information neither on the foetal cytokine balance nor on the best method for its modulation. We aimed to evaluate the influence of vaginal lactoferrin administration on amniotic fluid concentration of 47 cytokines, chemokines and growth factors. Methods: Sixty women undergoing genetic amniocentesis were enrolled in an open-label clinical trial. 300 mg of vaginal lactoferrin (Florence, Italy) were randomly administered to obtain 3 groups: A, 20 untreated patients; B and C (20 patients in each) respectively treated 4 and 12 h before amniocentesis. Cytokines, chemokines and growth factors concentrations were quantified by a magnetic bead Luminex multiplex immunoassays panel technology. Data analysis was performed with the software Stata (v. 13.1) and GraphPad Prism (v. 5). Group comparisons were performed using Kruskal–Wallis followed by Mann–Whitney U tests, with Bonferroni correction for multiple comparisons. A p < 0.05 was considered significant. Results: Among the 47 tested mediators, 24 (51.06%) were influenced by lactoferrin. 11 (23.4%), showed a highly significant difference (p <0.001); among these IL-9, IL-15, IFN-γ, IP-10, TNF-α, IL-1α and MCP-3 underwent a down-regulation, while IL-17 and FGF-basic, G-CSF, GM-CSF an up-regulation. Difference between group C and both B and A was small for IL-15, IP-10, IL-1α, MCP-3, while it was negligible for IL-9, IFN-γ and TNF-α. IL-17 and the 3 growth factors were strongly enhanced in B and C groups. IL-17, FGF-basic and GM-CSF showed increasing concentrations in both B and C groups, while G-CSF resulted up-regulated only in group C. Significance was intermediate (p < 0.01) for the down regulated IL-2RA, IL-12p40 and IFNα2 (6.38%) while it was small for 10 mediators (21.27%) 7 of which (IL-2, IL-4, eotaxin, PDGF-BB, RANTES, IL-18 and MIF) down-regulated and 3 (MCP-1, IL-3, and SDF-1α) up-regulated. Conclusion: Lactoferrin down-regulates 17 pro-inflammatory amniotic mediators while up-regulating 7 anti- inflammatory amniotic mediators, 5 of which definitively belonging to an anti-inflammatory profile. These findings open to clinical investigation on its use against inflammatory complications of pregnancy

Secondary lymphoid tissue as an important site for WU polyomavirus infection in immunocompetent children.

International audienceThe polyomaviruses KI and WU (KIPyV and WUPyV) have been identified in respiratory specimens from children with acute respiratory infections, which suggests the respiratory tract as a possible site of infection. However, the persistence of infection in the lymphoid system is unknown. Fresh samples (n=211) of tonsils, adenoids, and peripheral blood mononuclear cells (PBMCs) from 83 immunocompetent children (mean age 4.8 years) were tested for amplification of the KIPyV VP1 and WUPyV VP2 genes. The known BK and JC polyomaviruses and the lymphotropic human herpesvirus HHV-6 were also investigated by quantitative real-time PCR and direct sequencing. In addition, 98 nasopharyngeal swabs collected from children (mean age 6.2 years) affected by seasonal influenza-like illness were tested. Of the lymphoid tissues, 34.9% were positive for WUPyV, 4.8% for BK virus, and 33.8% for HHV-6. KIPyV and JC virus were not detected in these specimens. None of the polyomaviruses were detected in PBMCs. Among the nasopharyngeal samples, the prevalence of WUPyV was 27.5%, although 70% of the positive samples were co-infected with at least one of the following respiratory viruses: influenza virus, adenovirus, and respiratory syncytial virus. Phylogenetic analysis revealed high sequence homology (99%) between lymphoid- and nasopharynx-derived WUPyV strains. These results suggest that the tonsils and adenoids of immunocompetent children are a reservoir for WUPyV infection; probably due to the respiratory route of transmission. In addition, the prevalence of WUPyV was high among the children, and the virus was identified more frequently in older children than during the first years of life

In vivo microbiome and associated immune markers: New insights into the pathogenesis of vaginal dysbiosis

Abstract The microbiota fulfils a key role in the training and function of the immune system, which contributes to the symbiosis between the host and complex microbial communities. In this study, we characterized the interplay between vaginal bacteria and local immune mediators during dysbiosis in selected women of reproductive age who were grouped according to Nugent’s criteria. The abundance of Gardnerella vaginalis and Bifidobacterium breve was increased in the intermediate dysbiotic status, while the presence of a plethora of non-resident bacteria characterized the group with overt vaginosis. In response to these increases, the anti-inflammatory IL1ra and pro-inflammatory IL2 increased, while the embryo trophic factors FGFβ and GMCSF decreased compared to the healthy milieu. A specific pattern, including IL1α, IL1β, IL8, MIG, MIP1α and RANTES, distinguished the intermediate group from the vaginosis group, while IL5 and IL13, which are secreted by Th2 cells, were significantly associated with the perturbation of the commensals Lactobacilli, Gardnerella and Ureaplasma. Summarizing, we postulate that although the dysbiotic condition triggers a pro-inflammatory process, the presence of a steady state level of Th2 may influence clinical manifestations. These results raise clinically relevant questions regarding the use of vaginal immunological markers as efficacious tools to monitor microbial alterations