1. Introduction
Fungal infections have emerged as a major cause of morbidity and mortality in hospitals worldwide. Recent estimates suggest that approximately 6.5 million cases of invasive fungal infections occur each year, resulting in about 3.8 million deaths, of which 2.5 million (68%) are directly attributable to fungal diseases [1]. Aspergillus and Candida are the most common fungal genera responsible for severe infections, accounting for nearly 15% of hospital-acquired infections [2]. Among them, Aspergillus species, particularly A. fumigatus, represent the second leading cause of invasive fungal infections in healthcare settings, with reported mortality rates reaching up to 58% [3]. Another uncommon yet highly fatal infection is mucormycosis, caused by molds belonging to the order Mucorales [4].
The increasing emergence of antifungal-resistant fungi has further complicated the management of invasive fungal infections worldwide. Environmental molds, particularly Aspergillus fumigatus, may harbor intrinsic or acquired resistance to antifungal agents, especially triazoles, partly driven by the widespread use of agricultural azole fungicides and prolonged clinical antifungal exposure [5,6,7]. Antifungal resistance is associated with therapeutic failure, prolonged hospitalization, and increased mortality, especially among immunocompromised patients. In the context of the global antimicrobial resistance crisis, surveillance of environmental fungal reservoirs and their susceptibility profiles has become an important component of infection prevention and antimicrobial stewardship strategies [6,8].
The primary route of transmission of invasive fungal infections in susceptible individuals is exposure to mold spores, which are ubiquitous in soil, in air, and on surfaces [9]. These spores readily become airborne and can disseminate throughout hospital environments, where numerous mold genera, including Alternaria, Cladosporium, Aspergillus, Penicillium, Rhizopus, Fusarium, and Mucor, have been detected [3,4,10,11,12,13]. A recent systematic review of airborne fungal monitoring in healthcare facilities further identified Aspergillus, Penicillium, Alternaria, Cladosporium, and Rhizopus as the most frequently recovered genera and emphasized the absence of internationally standardized protocols and guidelines for fungal surveillance in hospital environments [14]. Assessing their presence and diversity in healthcare facilities is therefore essential, particularly in wards caring for immunocompromised or otherwise vulnerable patients [3,4].
Molds are frequently encountered in microbiology laboratories and are generally considered environmental contaminants [15]. To prevent contamination of fungal cultures, Sabouraud dextrose agar is commonly supplemented with cycloheximide. This compound inhibits eukaryotic protein synthesis, including that of microscopic fungi (eumycetes), by blocking both the initiation and elongation phases of protein synthesis translation [16]. However, data on the antifungal activity of cycloheximide remain limited. Furthermore, the existing studies are outdated, and contemporary evidence regarding its efficacy and use in sub-Saharan Africa is scarce [17,18,19,20].
In Burkina Faso, information on the fungal component of indoor air quality in hospital settings remains limited. The few available studies are outdated and were conducted exclusively at Sourô Sanou Teaching Hospital in Bobo-Dioulasso, the country’s second-largest city. Reported airborne fungal contamination rates ranged from 66.9% to 85.5%, with Aspergillus species identified as the predominant isolates [21,22,23]. The antifungal activity of cycloheximide, however, has not been investigated against these environmental fungi. This study was therefore undertaken to characterize the antifungal activity of cycloheximide on airborne fungal isolates collected from hospital indoor environments in Burkina Faso, thereby addressing an important knowledge gap.
4. Discussion
The overall prevalence of airborne fungal flora observed in this study (70.2%) was lower than the rates previously reported at the Sourô Sanou Teaching Hospital, which ranged from 83.5% to 88% in 2017 for the central operating room and intensive care unit [21,22], and 86.1% in 2019 for the medical emergency and neonatology units [23]. This difference may be attributed to the smaller number of samples analyzed in the present study (57 versus >200 in earlier investigations) and to temporal variations in environmental conditions influencing airborne fungal load.
Airborne fungal contamination rates reported in hospital environments worldwide vary widely, from 35% to 100% [13,24,25,26,27]. Such variability may reflect differences in climatic conditions, geographical location, hospital infrastructure, ventilation systems, occupancy, and infection prevention practices. A recent systematic review also highlighted substantial heterogeneity in airborne fungal monitoring methods, including differences in sampling techniques, culture media, incubation conditions, and interpretation criteria, emphasizing the need for standardized surveillance protocols in healthcare settings [14].
Because air sampling was conducted exclusively during the dry season (November 2023 to February 2024), the fungal distribution observed in this study should be interpreted within this seasonal context. Climatic factors such as temperature, humidity, rainfall, and wind are known to influence fungal growth, sporulation, and airborne spore dispersion. Consequently, seasonal fluctuations in airborne fungal contamination have been reported in healthcare environments [14]. Longitudinal studies covering both the dry and rainy seasons would provide a more comprehensive understanding of seasonal variations in indoor fungal communities in Burkina Faso.
The predominance of molds observed in this study is consistent with findings from previous investigations in the same hospital [21,22,23] and in other regions [11,12,13,24,25,27,28,29,30]. Molds are ubiquitous, filamentous fungi that produce numerous lightweight spores capable of dispersing through air currents and by human or animal carriers [9,10]. These characteristics facilitate their persistence and rapid colonization of indoor environments, including healthcare facilities.
Aspergillus was the predominant genus in all hospital departments, accounting for 57.9% to 86.9% of recovered isolates. Although Aspergillus predominated across all departments, the relative abundance of other fungal genera varied. Rhizopus spp., for example, were proportionally more frequent in the pediatric emergency (31.6%) and surgical emergency (25.0%) departments, whereas Penicillium spp. were recovered from four of the five departments. These differences may reflect variations in patient turnover, ventilation, environmental conditions, cleaning practices, and the specific activities performed within each department. Similar spatial heterogeneity of airborne fungal communities has been reported in hospital environments elsewhere [14].
The predominance of the Aspergillus genus observed here aligns with reports from Burkina Faso [21,22,23] and other regions [11,12,13,29]. Aspergillus species are cosmopolitan fungi capable of surviving under diverse environmental conditions. Their spores originate from multiple reservoirs, including soil, organic matter, dust, and ventilation systems, and can remain airborne for prolonged periods [9,31]. Their ecological adaptability and efficient spore dispersal likely explain their predominance in the present study.
In contrast, Cladosporium and Penicillium were the predominant genera in studies from Senegal and Portugal, respectively [27,30]. Such regional variation in fungal communities reflects differences in local climate, architecture, and ventilation, as well as variable standards of environmental control [9,27,30,31].
Among the Aspergillus isolates, A. niger was the most frequently identified species (48.7%), consistent with previous findings in Burkina Faso [23] and Tunisia [25]. A. niger is a common environmental contaminant found in food, soil, and indoor air owing to its ability to colonize diverse substrates [9,31]. Although widely distributed, A. niger is less pathogenic to humans than other Aspergillus species [32], typically affecting tissues compromised by infection, trauma, or cerumen accumulation [31]. Notably, A. fumigatus predominated in the same hospital in 2017 [21,22], suggesting temporal shifts in the composition of airborne fungal communities. Similarly, A. fumigatus was predominant in studies from Brazil [12] and Italy [29], whereas Penicillium spp. predominated in Iran [28] and Portugal [30]. These discrepancies may result from environmental and geographic differences, as well as variations in hospital infrastructure and infection control measures.
The overall inhibition rate of cycloheximide in this study (71.4%) was higher than that reported by Fuente et al. in the United Kingdom (40%) [19], likely due to the higher concentration used here (0.4 mg/mL vs. 0.1 mg/mL) and differences in fungal species composition. Conversely, the inhibition rate was lower than that reported in the United States (US) (93.3%) [18]. This may be explained by methodological differences, including a smaller sample size (15 isolates vs. 224), a slightly higher cycloheximide concentration (0.5 mg/mL), and the specific fungal species tested. In the U.S. study, Aspergillus, Mucor, Penicillium, and Rhizopus isolates were all inhibited, whereas only one of the two Cladosporium isolates showed susceptibility [18]. In the present study, inhibition rates were 69.6% against Aspergillus, 100% against Mucor, 33.3% against Penicillium, and 82.1% against Rhizopus.
A complete inhibition rate (100%) was also reported in the United States in 1950 for Aspergillus, Cladosporium, Penicillium, and Rhizopus [17], possibly reflecting the smaller number of isolates tested. Similarly, cycloheximide demonstrated complete inhibitory activity against Absidia, Chrysosporium, Cladosporium, Curvularia, and Mucor isolates in this study, consistent with findings from Chabasse et al. in France [20].
Although two Candida isolates were recovered in the present study, they were identified only to the genus level. Therefore, their response to cycloheximide could not be interpreted at the species level. This is relevant because cycloheximide susceptibility varies considerably among Candida species: some exhibit intrinsic tolerance, whereas others are inhibited at concentrations commonly used in selective culture media, as reported by Dal Pizzol et al. [33]. Species-level identification would therefore be necessary to interpret the cycloheximide growth response of Candida isolates accurately.
This study has several limitations. First, the passive sedimentation method, although simple and cost-effective, has limited sensitivity in highly controlled environments and lacks standardized international reference values, limiting comparisons across studies [34]. Second, environmental assessment was restricted to air sampling; hospital surfaces, medical materials, and reusable medical instruments were not sampled, providing only a partial characterization of the hospital fungal environment [34]. Third, fungal identification relied solely on conventional macroscopic and microscopic methods, which have limited discriminatory power for closely related fungal species and do not allow reliable species-level identification of yeasts such as Candida. Consequently, yeast isolates were identified only to the genus level, and some filamentous fungi may have been misidentified. Fourth, air sampling was not standardized with respect to routine cleaning and disinfection schedules, precluding differentiation between residual contamination and environmental recontamination. Finally, the effect of cycloheximide was evaluated using a single concentration (0.4 mg/mL) from a single manufacturer, limiting the generalizability of the findings. Future studies should integrate air, surface, material, and instrument sampling, employ advanced fungal identification techniques (e.g., chromogenic media, MALDI-TOF mass spectrometry, automated identification systems such as Vitek 2, and molecular methods), standardize sampling in relation to cleaning procedures, and evaluate multiple cycloheximide concentrations.