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ABSTRACT Aeromonas spp. are emerging human pathogens causing intestinal and extra-intestinal infections. Since their relevance in Western Europe as gastrointestinal pathogens is not well established, we investigated Aeromonas spp. prevalence in diarrheal fecal samples in an Italian University Hospital and characterized the virulence mechanisms of the isolates. Aeromonas spp. isolated from diarrheic stools using standard culture methods were identified by molecular techniques. Antimicrobial resistance was assessed by the micro broth dilution. Toxins, flagella, and type III secretion system genes were evaluated by polymerase chain reaction. Biofilm was quantified by crystal-violet staining. Interaction with human intestinal epithelial cells (Caco-2) was assessed by quantifying adhesion, interleukin (IL)-8 secretion, and epithelial barrier integrity. Aeromonas spp. represented 20.6% of bacterial pathogens isolated from diarrheic feces, the second most common enteropathogen. A. cavieae constituted 75% of the identified species, showing a relatively low clustering value. About 52% of Aeromonas isolates showed resistance to amikacin, whereas only 7.5% showed multiple drug resistance. Four or more virulence genes were identified in 66.7% of A. cavieae isolates and 100% of A. dakensis. Aeromonas isolates (82.5%) showed moderate or important biofilm-producing ability. Adhesion to Caco-2 cells correlated to fla+ gene, whereas ascV+ and aexU+ strains significantly induced IL-8 release from Caco-2. Aeromonas aer+ strains caused ZO1 and occluding redistribution and a significative reduction in trans-epithelial resistance. Aeromonas spp. emerge as relevant human intestinal pathogens with a disparate arsenal of pathogenicity factors causing diarrhea through different mechanisms. IMPORTANCE In this work, we demonstrate the epidemiologic relevance of the Aeromonas genus as the cause of infective diarrhea in North East Italy, both in children and adult subjects, with the significative presence of highly pathogenic strains. Aeromonas strains possess a heterogeneous armamentarium of pathogenicity factors that allows the microbe to affect a wide range of human intestinal epithelial cell processes that justify the ability to induce diarrhea through different mechanisms and cause diseases of variable severity, as observed for other gastrointestinal pathogens. However, it remains to be determined whether specific genotype(s) are associated with clinical pictures of different severity to implement the diagnostic and therapeutic approaches for this relevant enteric pathogen.

During the COVID-19 pandemic, immunosuppressed patients showed prolonged SARS-CoV-2 infections, with several studies reporting the accumulation of mutations in the viral genome. The weakened immune system present in these individuals, along with the effect of antiviral therapies, are thought to create a favourable environment for intra-host viral evolution and have been linked to the emergence of new viral variants which strongly challenged containment measures and some therapeutic treatments. To assess whether impaired immunity could lead to the increased instability of viral genomes, longitudinal nasopharyngeal swabs were collected from eight immunocompromised patients and fourteen non-immunocompromised subjects, all undergoing SARS-CoV-2 infection. Intra-host viral evolution was compared between the two groups through deep sequencing, exploiting a probe-based enrichment method to minimise the possibility of artefactual mutations commonly generated in amplicon-based methods, which heavily rely on PCR amplification. Although, as expected, immunocompromised patients experienced significantly longer infections, the acquisition of novel intra-host viral mutations was similar between the two groups. Moreover, a thorough analysis of viral quasispecies showed that the variability of viral populations in the two groups is comparable not only at the consensus level, but also when considering low-frequency mutations. This study suggests that a compromised immune system alone does not affect SARS-CoV-2 within-host genomic variability.