Please use this identifier to cite or link to this item: https://hdl.handle.net/1889/4777
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dc.contributor.advisorRoti, Giovanni-
dc.contributor.authorSpolzino, Angelica-
dc.date.accessioned2022-05-30T07:53:27Z-
dc.date.available2022-05-30T07:53:27Z-
dc.date.issued2022-
dc.identifier.urihttps://hdl.handle.net/1889/4777-
dc.description.abstractBACKGROUND AND AIMS OF THE STUDY Invasive fungal infections (IFI) are opportunistic diseases and the cause of significant morbidity and mortality, especially in haematological patients. The capacity to determine the actual risk for IFI is of fundamental importance to indicate optimal therapeutic choices for patients and individualize therapy to create a balance between the prevention of fungal infection and the occurrence of side effects. The importance of the microbiota, which collectively indicates all the microorganisms colonizing the human body is increasingly being recognized for its role in tissue homeostasis, and alterations in its composition, or dysbiosis, have been associated with a variety of pathological conditions. The airway microbiome is being recognized as an important player in tissue physiology and in protection against colonization by respiratory pathogens, including fungi. Hematological patients are at risk for dysbiotic changes, and it was recently confirmed that patients undergoing hematopoietic stem cell transplantation were characterized by a dysbiotic lung microbiome. In this study we aimed to investigate the possible role of pulmonary microbiota in pulmonary aspergillosis. In particular, the primary objective was the characterization of the lung microbiota in a cohorts of patients susceptible to fungal lung infections due to filamentous fungi, Aspergillus in particular, such as patients with malignant haematological diseases or those undergoing bone marrow transplantation. The secondary objective was the identification of a microbial signature of the airway microbiome predictive of the risk of fungal infection. PATIENTS AND METHODS We have designed a multicenter, prospective, observational study called SNIF (Survey of Nasal InFection) in which hematological patients were recruited and their nasal and pharyngeal swabs collected over a 6-month period for microbiome characterization. In order to gain further insight regarding the potential predictive role of the orophayngeal microbiome, each sample was associated with the overall risk of fungal infection of the patient at the time of collection, according to the SEIFEM (Epidemiological Surveillance of Infections in Haematological Diseases) recommendations. A total of 173 caucasian patients with different hematological diseases recruited from November 2015 to November 2017 from 7 Italian participating centers were enrolled. From each patient, up to six nasal and oropharyngeal swabs were collected over a six-month period. At moment, we focused on pharyngeal swabs. RESULTS In a first analysis of the SNIF database, we focused on the oropharyngeal microbiome owing to its higher richness, that may increase the likeliness of uncovering associations with the risk of IFI, for being the main source of the lung microbiota in adults tract and for the potential protective role in pulmonary infections. Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria and Fusobacteria represented the major phyla accounting for nearly the entire spectrum of bacteria detected in the oropharynx, followed by low-abundant phyla with a cumulative abundance of less than 1%. At the genus level, Streptococcus was found to be the most abundant, followed by Veillonella, Prevotella, Neisseria, Actinomyces, Haemophilus, Fusobacterium, and Rothia, common genera of the pharynx. Upon application of the SEIFEM algorithm, each samples were assigned to low- (LR) and high-risk (HR) groups at the time of collection, respectively. The analysis of alpha and beta diversities revealed that LR and HR samples were associated with distinct microbiota. Indeed, the LR group was characterized by a higher richness and evenness, as measured by Chao1 (LR, median 86, 95% CI 83.1, 88.9; HR, median 51, 95% CI 46.2, 55.9) and Shannon (LR, median 5, 95% CI 4.9, 5.1; HR, median 3.83, 95% CI 3.2, 3.4) indexes, than the HR group . This result was expected as the HR group is associated with a more intense pharmacological treatment that is predicted to impact the microbial diversity more severely. Moreover, the oropharyngeal microbiota of the two groups showed significant differences in compositional structure, as measured by Jaccard (LR, median 0.758, 95% CI 0.757, 0.79; HR, median 0.840, 95% CI 0.839, 0.841) and Bray-Curtis (LR, median 0.718, 95% CI 0.716, 0.719; HR, median 0.825, 95% CI 0.822, 0.827) indexes. Collectively, these results indicate that an algorithm based on demographic and clinical characteristics of patients may provide a useful framework to identify potential signatures of the oropharyngeal microbiome that associate with the risk of IFI. In order to identify signatures of the oropharyngeal microbiome that could be associated with the risk of IFI, we performed high dimensional class comparisons using linear discriminant analysis of effect size (LEfSe) by which we could detect significant differences in the bacterial communities between LR and HR groups. In particular, at the phylum level, the LR group was associated with a higher predominance of Bacteroidetes, Fusobacteria and Proteobacteria, while Firmicutes were predominant in the HR group. Collectively, these results indicate that microbiome signatures can be identified in oropharyngeal samples of hematological patients that are associated with distinct risks of IFI. To improve the predictive value of microbial signatures we have assessed microbial functional activity in dynamic-HR and dynamic-LR groups. In particular, genes involved in the biosynthesis of tryptophan (trp), glycolysis and homolactic fermentation were more abundant in HR than LR patients, while genes involved in the fatty acid elongation and the starch degradation pathways (abundantly present in oral bacteria25) were more abundant in LR than HR patients . The KEGG database confirmed the association of trp biosynthesis with the HR group. The increased trp auxotrophy would also predict different availability of trp in the HR and LR groups. We measured these levels in a subgroup of pharyngeal samples from 14 consecutive patients undergone HSCT during the study period and found that trp levels were indeed lower in HR than LR patients. Moreover, and importantly, while the levels of L-kynurenine (kyn), resulting from host catabolism of trp, were not different between the two groups, the production of indole-3-aldehyde (3-IAld), an indole that reflects the microbial consumption of trp, was significantly lower in HR than LR patients. These findings suggest an apparent restricted availability of trp for microbial consumption in HR patients. Consistently, amino acid biosynthetic pathways were abundantly present in HR samples Collectively, these results reveal the existence of a transcriptionally active oropharyngeal microbiota that may impact on lung immune status and suggest that not only microbial composition but also active functional activity characterizes the pharyngeal microbiota. CONCLUSIONS Overall, these results, while consistent with previous findings in gut, provide evidence for significant differences in oral microbial composition of patients at different risk for IFI. Supported by the high number of patients enrolled, the results clearly highlight the major contribution of neutropenia, the associated antibiotic usage and the occurrence of mucositis in the differences observed between HR and LR patients. As such, these results may pay the way for further studies to uncover associations between the many different risk factors for IFI and the changes in the microbiome. In addition, considering the tolerability and rapidity of the oropharyngeal swabbing, including the advantage of self-collection211, this sampling method appears highly feasible and relatively at low cost. Thus, if corroborated by further studies, the loss of alpha diversity associated with the loss of beneficial Clostridiales and Bacteroidetes could help delineate patients at risk of IFI, thereby providing information for antimicrobial therapy optimization. Indeed, HR patients would benefit from an antifungal prophylactic-based approach, as opposed to LR patients, for whom a fungal diagnostic-based approach is recommended to reduce overtreatment and unintended collateral damage to beneficial commensals. Discriminating patients who will benefit or not from antimicrobial prophylaxis will also help reducing the antimicrobial resistance crisis.en_US
dc.language.isoItalianoen_US
dc.publisherUniversità degli Studi di Parma. Dipartimento di Medicina e chirurgiaen_US
dc.relation.ispartofseriesDottorato di ricerca in Scienze mediche e chirurgiche traslazionalien_US
dc.rights© Angelica Spolzino, 2022en_US
dc.rightsAttribution-NonCommercial-NoDerivatives 4.0 Internazionaleen_US
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/*
dc.subjectairwayen_US
dc.subjectmicrobiomeen_US
dc.subjectinfectionen_US
dc.subjecthematologicalen_US
dc.subjectpatienten_US
dc.titleThe airway microbial landscape of hematologic patients at risk for invasive fungal infections: a prospective, observational, multicenter study (SNIF)en_US
dc.typeDoctoral thesisen_US
dc.subject.miurMED/15en_US
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