3.1. The Concentration and Diversity of Fungal Isolates from Indoor Air
The total count across all the studied rooms is 971 CFU, reflecting the presence and distribution of 8 fungal genera including
Alternaria, Aspergillus,
Cladosporium,
Curvularia,
Fusarium,
Paecilomyces,
Penicillium and
Trichoderma shown in
Table 1. Classrooms exhibit the highest concentration of fungal isolates, with a total of 400 CFU, which is significantly higher compared to other locations (p < 0.05). Staff offices follow with a significant concentration of 239 CFU, showing a notable difference from classrooms (p < 0.05). Toilets rank third with 171 CFU, significantly lower than classrooms (p < 0.05) and staff offices (p < 0.05). Microbiology laboratories have the lowest concentration at 161 CFU, which is significantly lower compared to classrooms (p < 0.05), staff offices (p < 0.05), and toilets (p < 0.05). The distribution of fungal isolates indicates that classrooms, with the highest human activity and potential dust accumulation, are the most conducive environment for fungal growth, followed by staff offices. Toilets, likely due to higher moisture levels, also support significant fungal presence. Laboratories, despite being controlled environments, still harbor a notable number of fungi. Also, classrooms exhibit the greatest fungal diversity, hosting all 11 identified fungal species. Staff offices also show significant diversity with 8 different fungal species. Toilets contain 9 fungal species. Microbiology laboratories, despite their controlled environments, have a diverse presence of 9 fungal types.
In our study, the concentration and diversity of fungal isolates from indoor air have been extensively studied, revealing a wide range of species and varying levels of concentration. Aspergillus, Penicillium and Cladosporium species were identified as among the most prevalent fungi, found abundantly in various environments. These findings align with studies by Navale et al. (2021) and Belizario et al. (2021). A review of research from 2005 to 2019 highlighted the presence of Aspergillus, Penicillium and Cladosporium species in the indoor air of critical hospital areas. Also, among the common fungi identified in indoor environments are Cladosporium halotolerans, Paecilomyces variotii, and Trichoderma harzianum, each with its own unique characteristics and implications for indoor air quality. Cladosporium species is often found in indoor environments, particularly in areas with high humidityStudies have shown that Cladosporium species, including C. halotolerans, can contribute significantly to indoor air spore concentrations, particularly in damp or water-damaged buildings. This species is known for its tolerance to salt, which allows it to thrive in conditions where other fungi might not survive. (Sánchez Espinosa et al., 2022). Paecilomyces variotii has been reported as another common indoor fungus, frequently isolated from air samples in various studies. This species is known for its ability to grow in a wide range of environmental conditions, including those with limited nutrients and varying temperatures (Lu et al., 2021). Trichoderma harzianum is a well-known indoor fungus, often found in soil and decaying organic matter. However, it can also colonize indoor environments, particularly in areas with high moisture levels. Trichoderma species are known for their antagonistic properties against other fungi, making them a common contaminant in buildings with water damage or mold issues (Polizzi et al., 2011). A study in Havana, Cuba, sampled indoor from 44 bedrooms during 2018 and 2019. Results indicated poor indoor air quality in 18 bedrooms, with concentrations of fungal propagules between 20 and 1330 CFU/m³. The most frequent genera identified were Cladosporium, Aspergillus, Penicillium, and Curvularia. Another study using qPCR found Aspergillus, Penicillium, and Paecilomyces variotii among the detected species, highlighting seasonal variations and the presence of fungal fragments in indoor air (Lu et al., 2021; Sánchez Espinosa et al., 2022).
In contrast, Curvularia sp. was more restricted in this study, primarily occurring outdoors rarely present indoors. This distribution underscores the adaptability of fungal species and their varied prevalence in different indoor settings. Curvularia species, is primarily associated with plants and soil, thriving in outdoor environments. While it can occasionally be detected indoors, it is not typically a dominant mold. Its spores may enter buildings through ventilation systems or open windows, but its lower prevalence indoors minimizes its impact compared to more common indoor fungi like Aspergillus and Penicillium species (Tang, 2015). Aspergillus ustus is another species that is rarely present in indoor air. It is primarily found in soil and decaying organic matter, making it more common outdoors. Although it can occasionally be detected indoors, its prevalence is significantly lower compared to other Aspergillus species. This limited presence indicates that A. ustus is not a predominant indoor mold, further emphasizing the variability in fungal species distribution between indoor and outdoor environments (Andersson et al., 2022).
PCR amplifications of DNA extracted from the 11 fungal species were performed using the ITS1/4 universal fungal primer pair. The sequencing data of the fungal strains were then aligned with sequences of closely related strains available in GenBank. The
Table 2 presents the fungal accession numbers, closest related species, and their similarity percentages based on sequencing data.
Alternaria alternata, with accession number OR533706.1, shows a 100% similarity to its closest related species
. Aspergillus flavus (MT635198.1) has an 89.29% similarity, while
A. niger (JX556221.1) has a 90% similarity.
Aspergillus ustus (KC800599.1) exhibits an 88.83% similarity, and
C. hawaiiensis (KY788103.1) has an 87.32% similarity.
Fusarium proliferatum (MT466521.1) shows a 91.21% similarity, and
P. variotii (MN547409.1) has an 88.57% similarity.
Penicillium chrysogenum (OK510242.1) exhibits an 89.69% similarity, while
P. citrinum (MH990629.1) shows a 91.24% similarity.
Cladosporium halotolerans (MW412494.1) has a 93.07% similarity, and
T. harzianum (MF108874.1) exhibits a 91.43% similarity. This data highlights the genetic relationships and diversity among the studied fungal isolates.
The provided phylogenetic tree illustrates the genetic relationships among various fungal isolates based on sequencing data as shown in
Figure 1.
Curvularia hawaiiensis (KY788103.1) stands as a distinct outgroup, indicating its unique genetic position.
Alternaria alternata (OR533706.1) and
C. halotolerans (MW412494.1) form a close cluster, showing a strong genetic similarity.
Fusarium proliferatum (MT466521.1) and
T. harzianum (MF108874.1) also cluster together, indicating a closer genetic relationship.
Paecilomyces variotii (MN547409.1) is positioned separately, suggesting moderate similarity to the others.
Penicillium species, including
P. chrysogenum (OK510242.1) and
P. citrinum (MH990629.1), show close genetic ties.
Aspergillus species such as
A. flavus (MT635198.1),
A. niger (JX556221.1), and
A. ustus (KC800599.1) form a tight cluster, highlighting their genetic relatedness. Similar studies have isolated
A. flavus,
A. niger, and
A. ustus from indoor air environments in research conducted (Nafis et al., 2023; Espinosa et al., 2021).
This tree emphasizes the diversity and evolutionary relationships among the studied fungal isolates, providing insights into their genetic affiliations. All 11 fungal species in the current study belong to the phylum Ascomycota. Most Ascomycota mycelium, composed of hyphae or filaments, forms the vegetative body of fungi. Spores, dispersed through the air, contribute to the anemophilous microbiota. These spores can be pathogenic, causing severe symptoms in immunocompromised individuals or those on long-term antimicrobial treatments. Variations in fungal isolate quantities and identities across studies are influenced by factors like exposure locations, collection techniques, identification methods (both conventional and molecular), and environmental conditions such as temperature and humidity (Tian et al., 2024).
3.2. Human Health Risks Associated with Indoor Fungi
Aspergillus species, such as Aspergillus niger, are commonly found in indoor environments and can pose serious health hazards to human health. Aspergillus niger is a saprophytic fungus that thrives in damp and poorly ventilated areas, making it a common indoor contaminant. Inhalation of Aspergillus spores can cause a variety of respiratory issues, including allergic reactions, asthma exacerbations, and allergic bronchopulmonary aspergillosis (ABPA). For immunocompromised individuals, the risk is significantly higher as A. niger can cause invasive aspergillosis, a severe and potentially fatal infection that can spread from the lungs to other parts of the body. The symptoms of invasive aspergillosis include fever, chest pain, cough, and hemoptysis (coughing up blood), and the infection requires prompt medical treatment (Agarwal et al., 2024). Penicillium species, including Penicillium chrysogenum, are commonly found in indoor environments, where they can become a significant source of allergens. These fungi thrive in conditions of high humidity and poor ventilation, often contaminating indoor air and surfaces. The spores released by Penicillium can contribute to various health issues, particularly for individuals with pre-existing respiratory conditions or compromised immune systems. Exposure to Penicillium spores is known to exacerbate asthma and other respiratory conditions. Individuals with asthma may experience increased frequency and severity of symptoms such as coughing, wheezing, and shortness of breath when exposed to these fungal spores. This exacerbation occurs because the allergens from Penicillium can trigger inflammatory responses in the airways, leading to heightened asthma symptoms and potentially more frequent asthma attacks. In individuals with chronic respiratory conditions or weakened immune systems, prolonged exposure can also lead to more severe health complications (Xing et al., 2022). Moreover, Penicillium chrysogenum has been associated with other allergic reactions and respiratory conditions beyond asthma. Studies have shown that inhalation of Penicillium spores can induce hypersensitivity pneumonitis, a condition where the lungs become inflamed due to an allergic reaction to inhaled organic dust, including fungal spores (Al Hallak et al., 2023). Cladosporium halotolerans, pose significant health risks to humans. Exposure to these fungi can lead to respiratory problems, allergic reactions, and exacerbate conditions like asthma. Cladosporium species, although less common, can cause phaeohyphomycosis, a serious fungal infection, particularly in immunocompromised individuals (Zhou et al., 2023).
3.3. Monitoring and Controlling Indoor Air Quality
To manage indoor fungal contamination and maintain a healthy environment, it is essential to implement regular monitoring and control measures. Proper ventilation is a critical factor in reducing indoor mold growth. Ensuring adequate airflow can help prevent the accumulation of moisture, which is a key contributor to fungal proliferation. Regular maintenance of HVAC systems and the use of air purifiers can also improve indoor air quality by reducing the concentration of airborne spores (Chen et al., 2023).
Moisture control is another crucial aspect of managing indoor fungal contamination. Leaks, condensation, and high humidity levels create favorable conditions for mold growth. Addressing these issues promptly through repairs and dehumidification can help prevent fungal proliferation. In areas prone to moisture, such as bathrooms and kitchens, the use of exhaust fans and moisture-resistant materials can further reduce the risk of mold growth (Engel et al., 2024).
Regular cleaning and maintenance are essential for minimizing fungal contamination. Dust and organic matter provide a substrate for fungal growth, so routine cleaning of surfaces, carpets, and ventilation systems is necessary to reduce the availability of these materials. Using cleaning agents with antifungal properties can also help eliminate mold spores from surfaces. The diversity and concentration of these and other fungal species in indoor air highlight the complexity of indoor air quality management. Factors such as humidity, ventilation, building materials, and human activity all influence the types and levels of fungi present. This includes regular inspection and maintenance of HVAC systems, controlling humidity levels, and promptly addressing water damage to prevent the proliferation of fungi like Cladosporium halotolerans, Paecilomyces variotii, and Trichoderma harzianum (Loukou et al., 2024).
Monitoring indoor air quality is vital for early detection of fungal contamination. Air sampling and spore counts can provide valuable information about the types and concentrations of fungi present in indoor environments. This data can inform targeted interventions to address specific sources of contamination and improve overall air quality. The use of molecular techniques for pathogen identification has enhanced the accuracy of fungal monitoring, allowing for more precise identification and tracking of fungal species (Tang et al., 2015).
3.4. Airborne Fungi Treatment
Effective treatment of fungal infections requires an understanding of the specific pathogens involved, as A. niger, P. chrysogenum, and C. halotolerans each present unique challenges and necessitate targeted antifungal therapies.
For infections caused by A. niger and P. chrysogenum, the primary treatment strategies involve similar antifungal agents due to their overlapping efficacy against various molds. Voriconazole is often the treatment of choice for both fungi. This newer antifungal agent is effective against a broad spectrum of fungi by inhibiting fungal cytochrome P450 enzymes, disrupting ergosterol synthesis, and thereby compromising fungal cell membrane integrity. Amphotericin B is another option for these infections; it works by binding to ergosterol in the fungal cell membrane, causing membrane disruption. It is particularly used in severe cases or when voriconazole is not tolerated. Lipid formulations of amphotericin B may be preferred to minimize renal and hepatic toxicity. Both voriconazole and amphotericin B can have serious side effects and may interact with other medications, so careful monitoring for adverse effects and drug interactions is essential. Accurate diagnosis and susceptibility testing are crucial for guiding appropriate antifungal therapy (Avilés-Robles et al., 2016).
Cladosporium halotolerans, a less common cause of phaeohyphomycosis, requires specific treatment approaches. Itraconazole, an azole antifungal, is used for Cladosporium infections. It inhibits ergosterol synthesis in the fungal cell membrane, similar to voriconazole but with a broader spectrum. For refractory cases, especially those involving skin lesions, aminolevulinic acid photodynamic therapy (ALA-PDT) has proven effective. This treatment involves applying a photosensitizing agent followed by light exposure to target and destroy fungal cells. In cases where lesions are localized and accessible, surgical intervention can be effective, though it may not always be feasible, particularly for periorbital or other challenging locations (Zhou et al., 2023).
Each treatment approach must be tailored to the specific fungal pathogen and patient condition, with careful monitoring for side effects and drug interactions. For all these infections, a multidisciplinary approach involving infectious disease specialists is often beneficial to optimize outcomes.