Review of the Microbial Spectrum of Mixed Respiratory Fungal Infections.

Background and Purpose: The healthcare system in India collapsed during the second wave of the COVID-19 pandemic. A fungal epidemic was announced amid the pandemic with several casesof COVID-associated mucormycosis and pulmonary aspergillosis being reported. However, there is limited data regarding mixed fungal infections in COVID-19 patients.Therefore, we present a series of ten consecutive COVID-19 patients with mixed invasive fungal infections (MIFIs).Materials and Methods: Among COVID-19 patients hospitalized in May 2021 at a tertiary care center in North India, 10 cases of microbiologically confirmed COVID-19-associated mucormycosis-aspergillosis (CAMA) were evaluated.Results: All patients had diabetes and the majority of them were infected with severe COVID-19 pneumonia (6/10, 60%) either on admission or in the past month while two were eachof moderate (20%) and mild (20%) categories of COVID-19; and were treated with steroid and cocktail therapy. The patients were managed with amphotericin-B along with surgical intervention.In total, 70% of all CAMA patients (Rhizopus arrhizus with Aspergillus flavus in seven and Aspergillus fumigatus complex in three patients) survived.Conclusion: The study findings reflected the critical importance of a high index of clinical suspicion and accurate microbiological diagnosis inmanaging invasive dual molds and better understanding of the risk and progression of MIFIs among COVID-19 patients. Careful scrutiny and identificationof MIFIs play a key role in the implementation of effective management strategies.

Metagenomic next-generation sequencing (mNGS) is a promising tool for improving antimicrobial therapy and infection control decision-making in complex infections. Secondary infection surveillance using mNGS in COVID-19 patients has rarely been reported. Respiratory pathogen and antibiotic resistance prediction were evaluated by BALF mNGS for 192 hospitalized COVID-19 patients between December 2022 and February 2023. Secondary infection was confirmed in 83.3% (160/192) of the COVID-19 patients, with bacterial infections (45%, 72/160) predominating, followed by mixed bacterial and fungal infections (20%, 32/160), and fungal infections (17.5%, 28/160). The incidence of bacterial or viral secondary infection was significantly higher in patients who were admitted to the ICU, received mechanical ventilation, or developed severe pneumonia (all p<0.05). Klebsiella pneumoniae (n=30, 8.4%) was the most prevalent pathogen associated with secondary infection followed by Acinetobacter baumannii (n=29, 8.1%), Candida albicans (n=29, 8.1%), Aspergillus fumigatus (n=27, 7.6%), human herpes simplex virus type 1 (n=23, 6.4%), Staphylococcus aureus (n=20, 5.6%) and Pneumocystis jiroveci (n=14, 3.9%). The overall concordance between the resistance genes detected by mNGS and the reported phenotypic resistance in 69 samples containing five clinically important pathogens (ie, K. pneumoniae, A. baumannii, S. aureus, P. aeruginosa and E. coli) that caused secondary infection was 85.5% (59/69). mNGS can detect pathogens causing secondary infection and predict antimicrobial resistance for COVID19 patients. This is crucial for initiating targeted treatment and rapidly detect unsuspected spread of multidrug-resistant pathogens.

Critically ill COVID-19 patients are at high risk for invasive fungal infections (IFIs). Data on IFI prevalence in severe COVID-19 patients in Latin America are scarce. This study aimed at analyzing the prevalence and outcomes of IFIs in COVID-19 patients from Argentina. For this purpose, a retrospective study was conducted on COVID-19 patients admitted to the intensive care unit of a hospital in Buenos Aires between 2020 and 2022, with mycological evidence of IFI. A total of 86 cases of IFIs were reported, including 50 cases of COVID-19-associated candidiasis (CAC), 29 of COVID-19-associated pulmonary aspergillosis (CAPA), 10 of COVID-19-associated histoplasmosis (CAH), two cases of cryptococcemia, and one case of invasive fusariosis. Mixed fungal infections were also detected: two cases of Pneumocystis jirovecii pneumonia with CAPA, two cases of CAC with CAPA, one case of cryptococcemia with CAPA, one case of CAPA with CAH, and one case of CAC with CAPA and CAH. The overall mortality was 67.4%, with mortality of 59.6%, 72.7%, and 62.5% for CAC, CAPA, and CAH, respectively. All cases with mixed fungal infections were fatal. The most frequent underlying comorbidities were arterial hypertension, type-2 diabetes mellitus, obesity, smoking, oncohematological disease, chronic kidney disease, and chronic obstructive pulmonary disease. Candida parapsilosis, C. albicans, and C. tropicalis were the most common species in CAC. Aspergillus fumigatus, A. flavus, A. terreus, and A. niger were predominant in CAPA. In conclusion, this study highlights the high prevalence and mortality of CAC, CAPA, and CAH in severe COVID-19 patients from Argentina.

Background: Melanoma is a highly heterogeneous disease, and a deeper molecular classification is essential for improving patient stratification and treatment approaches. Here, we describe the histopathology-driven proteogenomic landscape of 142 treatment-naïve metastatic melanoma samples to uncover molecular subtypes and clinically relevant biomarkers. Methods: We performed an integrative proteogenomic analysis to identify proteomic subtypes, assess the impact of BRAF V600 mutations, and study the molecular profiles and cellular composition of the tumor microenvironment. Clinical and histopathological data were used to support findings related to tissue morphology, disease progression, and patient outcomes. Results: Our analysis revealed five distinct proteomic subtypes that integrate immune and stromal microenvironment components and correlate with clinical and histopathological parameters. We demonstrated that BRAF V600-mutated melanomas exhibit biological heterogeneity, where an oncogene-induced senescence-like phenotype is associated with improved survival. This led to a proposed mortality risk-based stratification that may contribute to more personalized treatment strategies. Furthermore, tumor microenvironment composition strongly correlated with disease progression and patient outcomes, highlighting a histopathological connective tissue-to-tumor ratio assessment as a potential decision-making tool. We identified a melanoma-associated SAAV signature linked to extracellular matrix remodeling and SAAV-derived neoantigens as potential targets for anti-tumor immune responses. Conclusions: This study provides a comprehensive stratification of metastatic melanoma, integrating proteogenomic insights with histopathological features. The findings may aid in the development of tailored diagnostic and therapeutic strategies, improving patient management and outcomes.

Objective: To compare the specific IgE positive rates of the patients between allergic respiratory diseases and respiratory infectious diseases in Guangzhou, the relationship between the co-sensitization of house dust mite (HDM) allergen and Aspergillus fumigatus (AF) allergen and asthma, allergic rhinitis with asthma, pneumonia, upper respiratory infections, bronchitis, serum total immunoglobulin E (total Immunoglobulin E, tIgE) and age were analyzed, to provide the basis for the prevention and treatment of respiratory allergic diseases and respiratory infectious diseases in this area. Methods: A total of 2 535 patients with confirmed respiratory allergic diseases and respiratory infectious diseases were selected retrospectively from the outpatient or inpatient department of the First Affiliated Hospital of Guangzhou Medical University from April 2017 to June 2021 and detected HDM and AF specific IgE (sIgE) by the ImmunoCAP system. The age range was 1 to 89 years. The median age was 5 years. The average age was 9. ≤3 years old group n=894, 4-6 years old group n=721, 7-18 years old group n=615, 19-49 years old group n=207, >49 years old group n=98. There were 1 596 males (62.96%) and 939 females (37.04%). There were 1 279 cases of allergic diseases and 1 256 cases of respiratory infectious diseases. The different disease groups were divided into asthma group (411 cases), allergic rhinitis group (458 cases), allergic rhinitis combined with asthma group (410 cases), pneumonia group (463 cases), upper respiratory tract infection group (299 cases) and bronchitis group (494 cases). The difference of specific IgE (sIgE) and tIgE between HDM and AF was analyzed. For statistical analysis, continuous variables were tested by Mann-Whitney U. Classification data by chi-square test or Fisher's exact test. Results: 1 313 (51.79%) patients were sIgE positive for HDM allergen, 65 (2.56%) were sIgE positive for AF allergen, and 50 (1.97%) were both positive. In the respiratory allergic disease group, 877 cases (68.57%,877/1 279) were positive for HDM allergen sIgE, 57 cases (4.46%,57/1 279) were positive for AF allergen sIgE, and 44 cases (3.44%,44/1 279) were both positive; 436 cases (34.71%,436/1 256) of respiratory infectious diseases were positive for HDM allergen sIgE, 8 cases (0.64%,8/1 256) were positive for AF allergen sIgE, and 6 cases (0.48%,6/1 256) were both positive. In monosensitization, the HDM allergen sIgE sensitization rate was the highest in the allergic rhinitis & asthma group, at 80.24% (329/410). The positive rate of HDM allergen sIgE in male patients was 53.76%(858/1 596), and the positive rate in female patients was 46.22%(434/939), and the difference between the two was statistically significant (χ2=13.449, P<0.001). In polysensitization, asthma patients (5.35%,22/411) had the highest positive rate of HDM sensitization with AF, followed by allergic rhinitis patients (3.06%,14/458), allergic rhinitis with asthma (1.95%,8/410). The positive rate of respiratory infectious diseases such as pneumonia (0.43%,2/463), upper respiratory infections (0.33%,1/299), and bronchitis (0.61%,3/494) with AF was extremely low. The positive rate of HDM combined with AF in infants(≤3 years old group,0.34%, 3/894; 4-6 years old group, 0.97%, 7/721)was significantly lower than that in teenagers and adults(7-18 years old group,3.58%, 22/615; 19-49 years old group,6.28%, 13/207;>49 years old group,5.10%, 5/98).In the patients with HDM and AF combined sensitization, HDM sIgE levels were distributed in all grades, and AF sIgE levels were mainly in grades 1, 2, and 3. Conclusion: The positive rate of HDM combined with AF was higher in patients with respiratory allergic diseases such as asthma, allergic rhinitis, and allergic rhinitis combined with asthma, suggesting that clinical attention should be paid to the combination of HDM and AF in patients with asthma and allergic rhinitis, especially adults, more likely to be combined with AF.

New Aspects on Human Rhinovirus Infections

Pneumonia in the third year of the pandemic: one eye on the pathogens, the other on the host.

COVID-19 associated invasive candidiasis

Targeting the PI3K/Akt/mTOR pathway: A therapeutic strategy in COVID-19 patients

Under natural environmental settings or in the human body, the majority of microorganisms exist in complex polymicrobial biofilms adhered to abiotic and biotic surfaces. These microorganisms exhibit symbiotic, mutualistic, synergistic, or antagonistic relationships with other species during biofilm colonization and development. These polymicrobial interactions are heterogeneous, complex and hard to control, thereby often yielding worse outcomes than monospecies infections. Concerning fungi, Candida spp., in particular, Candida albicans is often detected with various bacterial species in oral biofilms. These Candida-bacterial interactions may induce the transition of C. albicans from commensal to pathobiont or dysbiotic organism. Consequently, Candida-bacterial interactions are largely associated with various oral diseases, including dental caries, denture stomatitis, periodontitis, peri-implantitis, and oral cancer. Given the severity of oral diseases caused by cross-kingdom consortia that develop hard-to-remove and highly drug-resistant biofilms, fundamental research is warranted to strategically develop cost-effective and safe therapies to prevent and treat cross-kingdom interactions and subsequent biofilm development. While studies have shed some light, targeting fungal-involved polymicrobial biofilms has been limited. This mini-review outlines the key features of Candida-bacterial interactions and their impact on various oral diseases. In addition, current knowledge on therapeutic strategies to target Candida-bacterial polymicrobial biofilms is discussed.

IntroductionOpportunistic bacterial and fungal infections are the major cause of morbidity and mortality among immune suppressed HIV-positive patients. The main objective of this study was to determine bacterial and fungal organisms causing respiratory infections and their susceptibility to commonly prescribed antimicrobials among HIV patients attending a tertiary infectious disease hospital in Kathmandu.MethodsSputum samples were collected from the HIV-positive patients attending Sukraraj Tropical and Infectious Disease Hospital (STIDH) from August 2017 to March 2018. A total of 100 sputum samples were cultured on conventional bacterial and fungal culture media. Bacterial and fungal isolates were identified based on their colony characteristics, microscopic morphology and various biochemical tests. Antibiotic susceptibility test (AST) of bacterial isolates was performed by modified Kirby Bauer disc diffusion method.ResultsOut of 100 sputum samples cultured, 24% (n=24) showed bacterial growth, 42% (n=42) showed fungal growth and 10% (n=10) had both bacterial and fungal growth. Among bacteria, 91.6% (n=22) were monomicrobial and 8.4% (n=2) were polymicrobial in growth, of which, Klebsiella pneumoniae (37.5%) were predominant isolates, followed by Pseudomonas aeruginosa (29.2%), and Escherichia coli (16.7%). The antibiotic susceptibility test (AST) showed 68% (17/25) of bacterial isolates were multi-drug resistant (MDR) and among them 41.2% (7/17) were found to be extended spectrum β lactamase (ESBL) producers. Fungal growth was observed in 42% of samples (42/100). A total of six different species of Candida and four different genera of molds were identified. On species differentiation, Candida albicans (20%) were followed by Candida parapsilosis (4%), and Candida dubliniensis (3%); and various molds were Aspergillus fumigatus (4%), Aspergillus flavus (2%), and Penicillium species (5%). CD4 count was inversely associated with bacterial and fungal infections. Fifty percent of the patients with the fungal infections had a CD4 count below 200. No fungal organisms were isolated from HIV-positive patients under antifungal drug treatment.ConclusionHIV-positive patients with a CD4 count less than 200 cells/µL are more vulnerable to opportunistic infections of bacterial and fungal origin. Early isolation, identification and appropriate treatment can reduce mortality due to co-infections. Routine screening of opportunistic pathogens is critical to contain the disease progression.

The clinical outcome of COVID-19 is strongly associated with microbiome dynamics in the upper respiratory tract

Fungal infections are common complications of respiratory viral infections and are associated with the increased need for intensive care and elevated mortality. Data regarding microbiological and molecular characteristics of such infections in COVID-19 patients are scarce. Here, we performed a comprehensive analysis, including species identification, antifungal susceptibility testing, molecular resistance determinants analysis, typing, and retrospective clinical data review, of fungal isolates recovered from 19 COVID-19 patients, who were hospitalized at the Hackensack University Medical Center (HUMC) in Hackensack, New Jersey, USA, in the initial phase of the pandemic from April–May 2020. In total, 17 Candida albicans, two C. parapsilosis, and two Aspergillus fumigatus were analyzed. All Candida spp. isolates were susceptible to micafungin and azole drugs (fluconazole, voriconazole, posaconazole, itraconazole, isavuconazole). A. fumigatus isolates were susceptible to micafungin and all triazole drugs except fluconazole (intrinsic resistance). Multilocus sequence typing (MLST) of C. albicans isolates revealed 15 different sequence types (STs), which clustered below the clade-defining limit of p-distance < 0.04. Pulsed-field gel electrophoresis (PFGE) karyotyping revealed no chromosomal rearrangements in these isolates. A. fumigatus isolates were of different, non-related genotypes. We speculate that virus- and drug-induced immunosuppression (94.7% of the patients received corticosteroids), together with prolonged hospital stay (median duration of 29 days) and mechanical ventilation (median duration of 24 days) likely increased the susceptibility to secondary respiratory and bloodstream infections in the studied patient population. The presence of fungi in blood or respiratory tract fluid was a prognosticator for poor clinical outcome, which presented as an 89.5% 30-day mortality in our patient cohort.

Objective To investigate the effect of fungus Aspergillus fumigatus and Candida albicans on the expression of endocellular interferon-γ (IFN-γ) and interleukin-4 (IL-4) in cytokine-induced natural killer (NK) cells. Methods NK cells were cultured with Aspergillus fumigatus or Candida albicans by non-contact or direct-contact methods with a ratio of NK cells to fungus of 10∶1. The expressions of IFN-γ and IL-4 in NK cells were evaluated by flow cytometry after co-cultured for 6 h. Analysis of variance or SNK-q test was used to compare the expressions of IFN-γ and IL-4 among different groups. Results The IFN-γ expression rates in NK cells with direct contacting to Aspergillus fumigatus hyphae, or to different morphotypes of Candida albicans were (20.12±0.53)%, (20.69±0.34)% and (20.8±0.37)% respectively, while IFN-γ expression in NK cells with indirect contacting to fumigatus hyphae, or to different morphotypes of Candida albicans were (21.40±0.53)%, (20.57±1.09)% and (20.20±0.51)% respectively, and all were significantly higher than that in the blank group [(15.11±2.60)%, all P>0.05]. The IFN-γ expression rates in the Aspergillus fumigatus spores direct and indirect contacting groups were (14.33±0.98)% and (14.97±1.53)%, which were not of significant difference compared with the blank group (P>0.05). The IL-4 expression rates in NK cells with direct contacting to different morphotypes of Aspergillus fumigatus and Candida albicans were (1.25±0.06)%, (1.21±0.03)%, (1.22±0.46)% and (1.26±0.11)%, while those in indirect contacting groups were (1.21±0.06)%, (1.25±0.04)%, (1.27±0.03)% and (1.26±0.1)%, which were not of significant difference compared with the blank group [(1.23±0.05)%, all P>0.05]. Conclusion Fungus stimuli can reduce the secretion of IFN-γ in NK cells, but have not significant influence on the secretion of IL-4. Key words: Aspergillus fumigatus; Candida albicans; Natural killer T-cells; Interferon-γ; Interleukin-4

Respiratory infectious diseases, particularly those caused by respiratory viruses, have the potential to lead to global pandemics, thereby posing significant threats to public and human health. Historically, the primary treatment for respiratory bacterial infections has been antibiotic therapy, while severe cases of respiratory viral infections have predominantly been managed by controlling inflammatory cytokine storms. Ferroptosis is a novel form of programmed cell death that is distinct from apoptosis and autophagy. In recent years, Recent studies have demonstrated that ferroptosis plays a significant regulatory role in various respiratory infectious diseases, indicating that targeting ferroptosis may represent a novel approach for the treatment of these conditions. This article summarized the toxic mechanisms underlying ferroptosis, its relationship with respiratory infectious diseases, the mechanisms of action, and current treatment strategies. Particular attentions were given to the interplay between ferroptosis and Mycobacterium tuberculosis, Epstein-Barr virus, severe acute respiratory syndrome coronavirus-2, Pseudomonas aeruginosa, dengue virus, influenza virus and herpes simplex virus type1infection. A deeper understanding of the regulatory mechanisms of ferroptosis in respiratory infections will not only advance our knowledge of infection-related pathophysiology but also provide a theoretical foundation for the development of novel therapeutic strategies. Targeting ferroptosis pathways represents a promising therapeutic approach for respiratory infections, with significant clinical and translational implications.