Background SARS-CoV-2 predisposes patients to secondary infections; however, a better understanding of the impact of coinfections on the outcome of hospitalized COVID-19 patients is still necessary. Aim To analyse death risk due to coinfections in COVID-19 patients. Methods We evaluated the Odds of death of 212 severely ill COVID-19 patients, with detailed focus on the risks for each pathogen, site of infection, comorbidities and length of hospitalization. Findings The mortality rate was 50.47%. Fungal and/or bacterial isolation occurred in 89 patients, of which 83.14% died. Coinfected patients stayed hospitalized longer and had an increased Odds of dying (OR = 13.45, R 2 =0.31). The risk of death was increased by bacterial (OR=11.28) and fungal (OR=5.97) coinfections, with increased levels of creatinine, leukocytes, urea and C-reactive protein. Coinfections increased the risk of death if patients suffer from cardiovascular disease (OR= 11.53), diabetes (OR=6.00) or obesity (OR=5.60) in comparison with patients with these comorbidities but without pathogen isolation. The increased risk of death was detected for negative-coagulase Staphylococcus (OR=25.39), Candida non- albicans (OR=11.12), S. aureus (OR=10.72), Acinetobacter spp. (OR=6.88), Pseudomonas spp. (OR=4.77) and C. albicans (OR=3.97). The high-risk sites of infection were blood, tracheal aspirate and urine. Patients with coinfection undergoing invasive mechanical ventilation were 3.8 times more likely to die than those without positive cultures. Conclusions Severe COVID-19 patients with secondary coinfections required longer hospitalization and had higher risk of death. The early diagnosis of coinfections is essential to identify high-risk patients and to determine the right interventions to reduce mortality.

Extracellular vesicles (EVs) has been considered an alternative process for intercellular communication. EVs release by filamentous fungi and the role of vesicular secretion during fungus-host cells interaction remain unknown. Here, we identified the secretion of EVs from the pathogenic filamentous fungus, Aspergillus fumigatus. Analysis of the structure of EVs demonstrated that A. fumigatus produces round shaped bilayer structures ranging from 100 to 200 nm size, containing ergosterol and a myriad of proteins involved in REDOX, cell wall remodeling and metabolic functions of the fungus. We demonstrated that macrophages can phagocytose A. fumigatus EVs. Phagocytic cells, stimulated with EVs, increased fungal clearance after A. fumigatus conidia challenge. EVs were also able to induce the production of TNF-α and CCL2 by macrophages and a synergistic effect was observed in the production of these mediators when the cells were challenged with the conidia. In bone marrow-derived neutrophils (BMDN) treated with EVs, there was enhancement of the production of TNF-α and IL-1β in response to conidia. Together, our results demonstrate, for the first time, that A. fumigatus produces EVs containing a diverse set of proteins involved in fungal physiology and virulence. Moreover, EVs are biologically active and stimulate production of inflammatory mediators and fungal clearance.

Fifty clinical isolates ofwere included as quality controls. All isolates produced clearly detectable growth only after 7 days of incubation. MICs were significantly independent of the incubation temperature (28 or 35°C) (P < 0.05). Different incubation periods resulted in MICs which were consistently different for each medium when azoles and griseofulvin were tested (P < 0.05). MICs obtained from different media at the same incubation time for the same isolate were significantly different when azoles and griseofulvin were tested (P < 0.05). MICs were consistently higher (usually 1 to 2 dilutions) with RPMI than with MVM or SDB (P < 0.05). When terbinafine was tested, no parameter had any influence on MICs (P < 0.05). RPMI standard medium appears to be a suitable testing medium for determining the MICs for T. rubrum. MICs obtained at different incubation times need to be correlated with clinical outcome to demonstrate which time has better reliability.

Skin mycoses are caused mainly by dermatophytes, which are fungal species that primarily infect areas rich in keratin such as hair, nails, and skin. Significantly, there are increasing rates of antimicrobial resistance among dermatophytes, especially for Trichophyton rubrum, the most frequent etiologic agent worldwide. Hence, investigators have been developing new therapeutic approaches, including photodynamic treatment. Photodynamic therapy (PDT) utilizes a photosensitive substance activated by a light source of a specific wavelength. The photoactivation induces cascades of photochemicals and photobiological events that cause irreversible changes in the exposed cells. Although photodynamic approaches are well established experimentally for the treatment of certain cutaneous infections, there is limited information about its mechanism of action for specific pathogens as well as the risks to healthy tissues. In this work, we have conducted a comprehensive review of the current knowledge of PDT as it specifically applies to fungal diseases. The data to date suggests that photodynamic treatment approaches hold great promise for combating certain fungal pathogens, particularly dermatophytes.

A total of 92 clinical isolates of dermatophytes (52 of Trichophyton rubrum and 40 of Trichophyton mentagrophytes) were selected for testing with six antifungal drugs (terbinafine, griseofulvin, clotrimazole, miconazole, isoconazole, and fluconazole) and two pairs of drug combinations (ketoconazole-cyclopiroxolamine and itraconazole-cyclopiroxolamine). Two methods of inoculum preparation for susceptibility testing were evaluated that used (i) inocula consisting only of microconidia of dermatophytes filtered in Whatman filter model 40 and (ii) unfiltered inocula consisting of hyphae and microconidia. We followed the recommendations of approved document M38-A of CLSI (formerly NCCLS) with some adaptations, including an incubation period of 7 days and an incubation temperature of 28°C. Reference strains of Candida parapsilosis, Candida krusei, Trichophyton rubrum, and Trichophyton mentagrophytes were included as quality-control strains. MICs were consistently higher (usually 1 to 2 dilutions for drugs tested individually) when nonfiltered inocula were tested (P < 0.01) except for terbinafine. Larger MICs were seen when testing drugs with nonfiltered inocula. The curves of drug interaction were used to analyze the reproducibility of the test, and it was shown that high levels of reproducibility were achieved using the methodology that included the filtration step. The standardization of methodologies is the first step to yield reliability of susceptibility testing and to proceed with clinical laboratory studies to correlate MICs with clinical outcomes.Dermatophytoses are among the world's most common diseases, and dermatophytes constitute an important public health problem as yet unresolved (7). Because dermatophytes require keratin for growth, they are commonly restricted to hair, nails, and superficial skin. Transmission can occur by direct contact or from exposure to desquamated cells. Direct inoculation through breaks in the skin often occurs in individuals with depressed cell-mediated immunity. The choice of appropriate treatment is determined by the site and extent of the infection and the species involved as well as by the efficacy, safety profile, and kinetics of the available drugs (8). Dermatophytoses generally respond well to topical antifungal therapy, although local therapy may be inappropriate for extensive infections or infections affecting the nails or the scalp (9). Onychomycosis is a common condition that represents up to 50% of all nail problems and 30% of all cases of dermatophytoses (6).The prevalence of fungal infections in humans and the development of new antifungal agents have increased the interest in antifungal susceptibility testing for dermatophytes. Despite much effort, there are still some methodological problems (14). Work on the development of standardized procedures for testing filamentous fungi has led to the publication by CLSI (formerly NCCLS) (15) of the approved reference document M38-A, which recommends the use of standard RPMI 1640 broth, nongerminated conidial inoculum suspe...