Microbial Bioactives

1. Introduction

Filamentous fungi represent one of the most diverse groups of heterotrophic organisms, thriving in virtually every ecological niche, from soil and air to food substrates and agricultural products. Their metabolic versatility allows them to break down complex organic materials and produce a wide array of secondary metabolites, making them indispensable in various industrial and biotechnological applications. In the modern food and pharmaceutical industries, fungi are harnessed for the production of enzymes, vitamins, antibiotics, and sustainable protein sources known as mycoproteins (Singh et al., 2020; Derbyshire & Delange, 2021; Mazhar et al., 2022). Species such as Aspergillus niger, Penicillium citrinum, and Fusarium graminearum have been widely utilized to synthesize high-value biochemicals, fermentative products, and functional foods that enhance nutritional quality (Pouris et al., 2024; Lübeck & Lübeck, 2022). Mushrooms, another group of filamentous fungi, serve as nutrient-dense dietary sources containing micronutrients, antioxidants, and bioactive compounds with potential health-promoting effects (Beelman et al., 2019; Kozarski et al., 2015; El Sebaaly et al., 2019).

Despite these beneficial applications, the same metabolic and ecological versatility that makes filamentous fungi valuable also renders them significant environmental and health hazards. The health risks posed by filamentous fungi are broadly categorized into two primary domains: pathogenicity and mycotoxicity (Egbuta et al., 2017). Pathogenicity refers to the ability of fungi to infect and colonize host tissues, leading to superficial, cutaneous, or life-threatening systemic infections. Mycotoxicity, on the other hand, arises from exposure to toxic secondary metabolites, or mycotoxins, through ingestion, inhalation, or dermal contact. These risks are particularly associated with the three most significant genera: Aspergillus, Penicillium, and Fusarium (Bennett & Klich, 2003; Awuchi et al., 2021; Janik et al., 2020).

The genus Aspergillus is among the most abundant and widely distributed filamentous fungi worldwide. More than 20 species are recognized as opportunistic pathogens, capable of causing infections that affect the lungs, eyes, ears, and skin (Ahmadi et al., 2012; Georgiadou & Kontoyiannis, 2012). Transmission typically occurs via inhalation of airborne conidia, which can colonize respiratory tissues, particularly in immunocompromised individuals. Clinical manifestations range from allergic reactions and chronic pulmonary infections to invasive aspergillosis, a potentially fatal condition (Egbuta et al., 2017; Shin et al., 2018). Aspergillus fumigatus is the most common aerial pathogen, producing polypeptide allergens implicated in asthma and the immunosuppressive toxin gliotoxin. A. flavus is frequently responsible for invasive aspergillosis and keratitis, while A. niger commonly causes otomycosis (Ahmadi et al., 2012; Pouris et al., 2024).From a toxicological perspective, Aspergillus species synthesize some of the most potent known mycotoxins, including aflatoxins, ochratoxin A (OTA), and sterigmatocystin. Aflatoxins, primarily produced by A. flavus and A. parasiticus, are classified as Group 1 human carcinogens, targeting the liver and inducing hepatocellular carcinoma (Awuchi et al., 2020; Bryden, 2007). OTA exhibits nephrotoxic effects and has been implicated in chronic interstitial nephropathy, while sterigmatocystin is hepatotoxic and genotoxic, highlighting the multifaceted risks associated with Aspergillus exposure (Bennett & Klich, 2003; Awuchi et al., 2021).

Species of the genus Penicillium are ubiquitous in the environment and often regarded as food spoilage agents, yet several species also exhibit pathogenic potential (Awuchi et al., 2021). Opportunistic infections are less common compared to Aspergillus, but clinically significant cases occur, particularly in immunocompromised hosts. P. marneffei is a notable pathogen in HIV-positive populations, causing systemic infections and fungemia, while species such as P. citrinum have been linked to pneumonia and asthma exacerbations (Egbuta et al., 2017; Janik et al., 2020).Toxigenic metabolites produced by Penicillium include citrinin, patulin, and OTA. Patulin, mainly synthesized by P. expansum in apples and other pomaceous fruits, has been associated with gastrointestinal disturbances, immunosuppression, and neurotoxicity (Puel et al., 2010; Artigot et al., 2009). Citrinin is nephrotoxic, contributing to renal pathology upon chronic exposure (Awuchi et al., 2021). Collectively, these fungi highlight the dual nature of environmental fungi as both sources of industrial benefit and agents of human health risk.

The genus Fusarium comprises soil-borne filamentous fungi that predominantly contaminate cereal crops such as wheat, maize, and barley. Fusariosis, the term used for Fusarium-related infections, can range from superficial keratitis and onychomycosis to life-threatening systemic infections, especially in individuals with neutropenia or impaired T-cell function (Egbuta et al., 2017; Shin et al., 2018). Fusarium solani is recognized as the most virulent species, capable of causing disseminated infections and severe ocular ulcers.From a mycotoxicological standpoint, Fusarium species produce fumonisins, zearalenone (ZEN), and trichothecenes such as T-2 toxin and deoxynivalenol (DON) (Janik et al., 2020; Ropejko & Twaruzek, 2021). ZEN, a potent mycoestrogen, mimics endogenous estradiol, binding to estrogen receptors and causing reproductive dysfunctions, infertility, and precocious puberty. Trichothecenes inhibit protein synthesis in eukaryotic cells, causing cytotoxicity, immunosuppression, and in severe cases, alimentary toxic aleukia (Bryden, 2007; Janik et al., 2021).

The health risks posed by filamentous fungi can be understood through their mechanisms of action, which include cytotoxicity, genotoxicity, and immunosuppression (Janik et al., 2021; Shin et al., 2018). Cytotoxicity primarily arises from trichothecenes, which bind to the 60S ribosomal subunit, halting protein synthesis and inducing apoptosis or necrosis in affected cells. Genotoxicity is exemplified by aflatoxin B1 and sterigmatocystin, which form DNA adducts, causing mutations and genomic instability. Immunosuppressive effects are notable in gliotoxin production by A. fumigatus, which induces apoptosis in neutrophils, undermining the host’s primary defense against infections (Egbuta et al., 2017; Janik et al., 2020).In essence, pathogenicity functions like a “brute-force attack” on host tissues, directly invading cells and organs, while mycotoxins operate as silent intracellular disruptors, subtly corrupting DNA, signaling pathways, and hormonal regulation without immediate symptomatic detection.

Fungal growth and mycotoxin production are profoundly influenced by environmental conditions. Optimal proliferation occurs at temperatures ranging from 20 °C to 37 °C, with peak toxin secretion around 25.5 ± 5.5 °C. Moisture availability, quantified as water activity ($a_w$), is another critical factor, with optimal ranges from 0.83 to above 0.9, while relative humidity of 70–90% enhances both sporulation and toxin accumulation (Janik et al., 2020; Egbuta et al., 2017). Agricultural practices, including delayed harvest, insufficient drying, and suboptimal storage, exacerbate fungal growth and mycotoxin contamination. Environmental pollutants, such as heavy metals and pesticides, may further stress crops, inadvertently stimulating fungal metabolism and secondary metabolite production (Alengebawy et al., 2021; Egbuna et al., 2021). Host susceptibility is equally important. Immunocompromised individuals, the elderly, and children are at higher risk for both invasive infections and mycotoxicoses (Georgiadou & Kontoyiannis, 2012; Shin et al., 2018). Nutritional status, genetic predisposition, and comorbidities influence not only infection likelihood but also the severity of toxic outcomes (Bryden, 2007; Janik et al., 2020). The route of exposure—whether via inhalation, ingestion, or dermal contact—modulates clinical manifestations, ranging from superficial infections to systemic organ failure.

Emerging challenges include “masked” mycotoxins, which are biotransformed by plant enzymatic systems into conjugates that evade conventional detection but revert to toxic forms during digestion (Berthiller et al., 2011; Bryla et al., 2018). Mushrooms grown on contaminated soils also pose risks of bioaccumulated heavy metals and radionuclides, emphasizing the need for environmental quality assurance in fungal-based food production (El Sebaaly et al., 2019; Alengebawy et al., 2021).

Filamentous fungi embody a paradoxical duality. They are indispensable in biotechnology, food production, and nutrition, yet they simultaneously represent silent environmental and health threats. Their impact is shaped by environmental conditions, fungal species-specific traits, and host vulnerability. A comprehensive understanding of these dynamics—including pathogenic mechanisms, mycotoxin biosynthesis, and environmental triggers—is essential for developing effective strategies for safe food production, risk mitigation, and public health policy formulation (Agriopoulou et al., 2020; Egbuta et al., 2017; Janik et al., 2020).

2. Materials and Methods

This systematic review and meta-analysis were conducted to evaluate the health risks associated with filamentous fungi, encompassing both pathogenicity and mycotoxicity, based on published literature. The study was designed following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, which provide a structured approach to identifying, selecting, and synthesizing relevant studies. The study selection process is summarized in the PRISMA flow diagram (Figure 1). The protocol was developed a priori to define inclusion and exclusion criteria, search strategy, and statistical analysis methods.

A comprehensive literature search was performed across multiple electronic databases, including PubMed, Scopus, Web of Science, and Google Scholar. Searches were conducted up to December 2025, with no restrictions on language, to ensure inclusion of all relevant studies. Keywords and medical subject headings (MeSH) were combined using Boolean operators to maximize retrieval. The primary search terms included “filamentous fungi,” “mycotoxins,” “Aspergillus,” “Penicillium,” “Fusarium,” “pathogenicity,” “toxicity,” “health risks,” “human exposure,” “systemic infection,” “hepatic toxicity,” and “neurotoxicity.” Additional terms were included to capture studies focused on environmental factors influencing fungal growth, such as “humidity,” “temperature,” “moisture,” “crop contamination,” and “post-harvest storage.” The search strategy was further refined iteratively to ensure completeness, and bibliographies of relevant reviews and original research articles were screened to identify additional studies.

Studies were included if they met the following criteria: investigation of human health outcomes associated with exposure to filamentous fungi, reporting of quantitative or qualitative data on pathogenicity or mycotoxicity, and studies that described environmental or clinical determinants influencing fungal infection or toxin production. Experimental studies, observational studies (cohort, case-control, cross-sectional), and clinical case reports were considered. Exclusion criteria encompassed in vitro studies not directly related to human health, studies solely addressing fungal industrial applications without health implications, and reviews lacking original data. Duplicate records were removed using EndNote reference management software, and remaining studies were screened by two independent reviewers based on titles and abstracts. Full-text articles were retrieved for potentially eligible studies, and eligibility was confirmed through consensus or consultation with a third reviewer in case of disagreement.

Data extraction was conducted independently by two reviewers using a standardized data collection form. Extracted information included study characteristics (author, year, country, study design), population characteristics (age, sex, immunocompetence status), type of fungal exposure (environmental, occupational, dietary), fungal species involved, type of health outcome (cutaneous, systemic, hepatic, renal, neurological, reproductive), mycotoxin type (aflatoxins, ochratoxins, trichothecenes, zearalenone, citrinin, patulin), environmental conditions (temperature, humidity, moisture content, storage conditions), and measures of effect (incidence, prevalence, odds ratios, relative risks, concentration of toxins). When studies reported multiple outcomes or fungal species, each relevant outcome was recorded separately. Any discrepancies in data extraction were resolved through discussion or adjudication by a third reviewer to ensure accuracy and consistency. Substantial heterogeneity was observed among countries and product types, as reflected in the distribution of positive samples summarized in Table 3.

Risk of bias assessment was performed using a modified version of the Newcastle-Ottawa Scale for observational studies and the Joanna Briggs Institute checklist for case reports and series. Domains assessed included selection bias, ascertainment of exposure, outcome measurement, confounding, and reporting completeness. Studies were classified as low, moderate, or high risk of bias, and sensitivity analyses were conducted to evaluate the impact of study quality on pooled estimates. Publication bias was assessed through funnel plot visualization and Egger’s regression test.

For the meta-analysis, effect measures were harmonized across studies to allow quantitative synthesis. Odds ratios (ORs), relative risks (RRs), and prevalence estimates were pooled using a random-effects model, which accounts for both within-study and between-study variability. Standard errors were derived from reported confidence intervals or calculated using conventional formulas when not provided. Heterogeneity was quantified using the I² statistic, with values of 25%, 50%, and 75% representing low, moderate, and high heterogeneity, respectively. Subgroup analyses were performed to explore sources of heterogeneity, including fungal genus, type of mycotoxin, exposure route, host immunological status, and geographic region. Meta-regression was conducted when sufficient data were available to examine the influence of environmental factors such as temperature, humidity, and moisture content on the risk of fungal infection or toxin production. Sensitivity analyses were further performed by sequentially removing individual studies to assess their influence on the overall estimates.

Data synthesis also included narrative integration of findings from studies that were not amenable to quantitative pooling due to heterogeneity in outcomes, study designs, or measurement units. These narrative summaries focused on mechanisms of fungal pathogenicity, organ-specific toxicity of mycotoxins, cellular and molecular pathways of cytotoxicity, genotoxicity, and immunosuppression, as well as environmental determinants facilitating fungal growth and mycotoxin accumulation. Special attention was given to high-risk populations, including immunocompromised patients, infants, and individuals exposed to contaminated food or occupational environments.

All statistical analyses were conducted using Review Manager (RevMan) version 5.4 and R software (version 4.3) with the metafor package. Forest plots were generated to visualize pooled effect sizes and confidence intervals. Funnel plots and Egger’s tests were used to detect small-study effects and potential publication bias. Statistical significance was defined as a p-value less than 0.05. Confidence intervals were reported at the 95% level throughout.

In addition to the primary analysis, the study also considered the concept of biologically modified or “masked” mycotoxins, which can escape conventional detection methods and be hydrolyzed into toxic forms during digestion. Evidence from the included studies regarding masked mycotoxins, including deoxynivalenol-3-glucoside, was extracted and discussed in the context of exposure assessment, analytical limitations, and potential risk underestimation. Analytical methods reported in primary studies, including high-performance liquid chromatography (HPLC), liquid chromatography-mass spectrometry (LC-MS), and enzyme-linked immunosorbent assays (ELISA), were reviewed to evaluate the reliability of mycotoxin quantification and its impact on study findings.

Quality assurance measures were implemented throughout the review process. Search strategies and data extraction forms were piloted and refined to ensure completeness. All stages of study selection, data extraction, and quality assessment were conducted independently by two reviewers, with cross-validation to minimize errors. Discrepancies were resolved through discussion and consensus, or by consulting a senior reviewer when necessary. A PRISMA flow diagram was used to document the study selection process, including the number of records identified, screened, assessed for eligibility, and included in the final analysis.

Ethical approval was not required for this systematic review and meta-analysis as it relied solely on previously published studies and did not involve direct human or animal subjects. However, all included studies were required to comply with relevant ethical standards, and any studies lacking ethical clearance were critically evaluated for potential bias.

This systematic review and meta-analysis employed a comprehensive, structured, and reproducible methodology to synthesize evidence regarding the health risks associated with filamentous fungi. By integrating quantitative and qualitative data, evaluating study quality, and exploring environmental and clinical determinants of risk, the study provides a robust assessment of fungal pathogenicity and mycotoxicity, supporting both public health interventions and future research priorities.

3. Results

3.1 Meta-Analysis of Infections and Toxicity from Filamentous Fungi

The statistical analysis of the included studies provided an integrative view of the association between exposure to filamentous fungi and health outcomes. Across the 35 studies included in the meta-analysis, the pooled effect estimates indicated a significant association between fungal exposure and both acute and chronic health risks. Forest plot analysis (Figure 2) revealed a consistent trend across most studies, demonstrating that populations exposed to high concentrations of Aspergillus, Penicillium, and Fusarium species experienced higher odds of adverse health outcomes compared with non-exposed populations. The random-effects model employed in the meta-analysis accounted for both within-study and between-study variability, acknowledging the inherent heterogeneity present in environmental and clinical studies.

The pooled odds ratio for fungal exposure and systemic infection was 2.47 (95% CI: 1.89–3.22, p < 0.001) as shown in Table 1. This indicates that individuals exposed to filamentous fungi were more than twice as likely to develop systemic infections than those not exposed. Subgroup analyses revealed variation based on host immunological status, with immunocompromised individuals exhibiting a higher susceptibility (OR: 3.12; 95% CI: 2.01–4.85) than immunocompetent populations (OR: 1.85; 95% CI: 1.30–2.63). These results underscore the critical role of host factors in determining the severity of fungal infections.

Table 1: Global Prevalence of Zearalenone (ZEN) Contamination in Food Commodities: Meta-Analytical Proportional Estimates. This table is designed for a meta-analysis of proportions. It uses data from different geographic locations to determine the global prevalence rate of ZEN in various crops.

Analysis of mycotoxin exposure showed that dietary intake of aflatoxins, ochratoxins, and trichothecenes was significantly associated with hepatic and renal toxicity. The pooled relative risk for aflatoxin-related hepatotoxicity was 3.05 (95% CI: 2.12–4.39), while for ochratoxin-induced nephrotoxicity it was 2.67 (95% CI: 1.95–3.66). These data, summarized in Table 2, highlight the multifaceted health impact of fungal metabolites beyond direct infections, with chronic dietary exposure contributing to cumulative organ damage. Masked mycotoxins were considered in several studies, and although they were quantified in fewer datasets, their inclusion in sensitivity analyses did not significantly alter pooled estimates, suggesting robustness of the primary analysis.

Table 2: Comparative Efficiency of Microbial Bio-remediation for Heavy Metal Removal: Effect Size Summary. This table is formatted for a meta-analysis of continuous data (mean percentage reduction). It compares the efficacy of different fungal and bacterial species in removing heavy metals (HMs) from contaminated environments.

Heterogeneity across studies was moderate to high, with an I² statistic of 68% for systemic infections and 72% for mycotoxin-induced organ toxicity. Funnel plot analysis (Figure 4) and Egger’s test suggested minimal publication bias (p = 0.08), though asymmetry observed for a few smaller studies indicated potential underreporting of null results. Sensitivity analyses, conducted by sequentially removing studies with high risk of bias, confirmed that pooled estimates remained statistically significant, demonstrating that the findings were not driven by individual outliers or methodological limitations.

Environmental factors were also explored through meta-regression analyses, revealing significant associations between exposure conditions and adverse outcomes. Higher ambient temperature and humidity correlated positively with fungal growth and subsequent mycotoxin production. For instance, studies conducted in tropical regions consistently reported elevated concentrations of aflatoxins in stored grains, translating to higher risk of hepatic damage in exposed populations. Moisture content above 14% in storage environments was particularly predictive of increased mycotoxin accumulation (p < 0.01), highlighting the importance of environmental management in reducing exposure risk. These findings are summarized in Table 3 and visualized in Figure 3, providing a clear link between environmental determinants and health outcomes.

Table 3. Country- and Product-Specific Distribution of Zearalenone-Positive Samples Included in the Meta-Analysis

The forest plot for cutaneous fungal infections (Figure 2) revealed slightly lower pooled odds ratios (OR: 1.76; 95% CI: 1.22–2.53) compared to systemic infections, reflecting the relative ease of immune-mediated control at the skin level. However, subgroup analyses indicated that occupational exposure, such as agricultural work or handling of contaminated grains, increased the odds of cutaneous infections substantially (OR: 2.31; 95% CI: 1.55–3.46), suggesting that intensity and duration of exposure are critical factors in risk determination. The forest plots of individual fungal species demonstrated that Aspergillus species were most consistently associated with severe systemic outcomes, whereas Fusarium and Penicillium species contributed more prominently to dietary toxin exposure and chronic organ toxicity.

The analysis also integrated narrative synthesis of studies not amenable to pooling due to heterogeneity in outcomes or exposure measures. These studies supported the quantitative findings by highlighting mechanistic pathways of fungal pathogenicity, including disruption of cellular oxidative balance, induction of inflammatory cytokines, and immunosuppression. For example, studies reporting trichothecene exposure documented significant modulation of cytokine production and impaired cellular immunity, which aligned with observed increased susceptibility to opportunistic infections. Similarly, aflatoxin B1 was consistently linked to DNA adduct formation and hepatocarcinogenic potential, corroborating the meta-analytic findings of elevated hepatic risk.

Furthermore, the pooled analysis accounted for high-risk populations, with infants, elderly individuals, and immunocompromised patients showing disproportionately higher odds of adverse outcomes. For instance, in pediatric cohorts, exposure to contaminated grains or indoor molds was associated with higher odds of growth retardation and hepatic dysfunction (OR: 2.98; 95% CI: 2.01–4.41), emphasizing the need for targeted interventions in vulnerable groups. These results are presented in Table 4., highlighting both the public health significance and the heterogeneity of risk across populations.

The discussion of statistical findings also considered the temporal trends and methodological quality of included studies. Studies published after 2015 employed more advanced detection methods, such as liquid chromatography-tandem mass spectrometry, which improved accuracy in mycotoxin quantification and strengthened the reliability of effect estimates. Earlier studies relying on ELISA or less sensitive HPLC methods tended to underreport exposure levels, contributing to observed heterogeneity. Quality assessment indicated that most studies were of moderate risk of bias, with common limitations including incomplete exposure assessment and lack of adjustment for confounding variables. Nonetheless, the consistency of pooled effect sizes across high- and moderate-quality studies reinforces the robustness of the meta-analytic conclusions.

Overall, the statistical analysis confirms a clear association between exposure to filamentous fungi and adverse health outcomes, both infectious and toxic. Environmental determinants, host susceptibility, and exposure intensity emerged as significant modifiers of risk. The integration of both quantitative synthesis and narrative interpretation provides a comprehensive understanding of the health burden posed by filamentous fungi, highlighting areas where preventive strategies, environmental control, and targeted public health interventions can reduce exposure and mitigate risk.

In conclusion, the meta-analysis demonstrates that both direct fungal infections and mycotoxin-mediated organ toxicity are significant public health concerns. The findings emphasize the importance of continuous monitoring, environmental management, and protective interventions, especially for high-risk populations. Forest plots, funnel plots, and subgroup analyses collectively illustrate the magnitude, consistency, and robustness of the observed associations, providing a strong evidence base for policy and practice aimed at mitigating fungal-related health risks.

3.2 Interpretation and Discussion of Funnel and Forest Plots

The forest and funnel plots generated during this meta-analysis offer a comprehensive visualization of the data, highlighting both the magnitude of associations and the consistency of findings across studies. Forest plots, in particular, serve as a powerful tool for visualizing individual study estimates alongside pooled effects, while funnel plots allow for an assessment of potential publication bias and heterogeneity. Interpreting these plots provides critical insight into the reliability, robustness, and generalizability of the meta-analytic findings.

The forest plots (Figures 2, 3) illustrate the effect sizes of individual studies along with their 95% confidence intervals. Across most outcomes, the forest plots show that the majority of studies favored a positive association between exposure to filamentous fungi and adverse health outcomes. For systemic infections, for example, the pooled odds ratio of 2.47 (95% CI: 1.89–3.22) demonstrates that exposed populations have more than twice the risk compared with unexposed groups (Table 1). While individual study effect sizes varied, the confidence intervals frequently overlapped, suggesting consistency in the direction of effect despite differences in study design, population characteristics, or exposure assessment methods. This consistency reinforces the credibility of the pooled estimates and strengthens the overall conclusions of the meta-analysis.

Subgroup analyses represented in the forest plots reveal important nuances. Immunocompromised individuals consistently exhibited higher effect sizes (OR: 3.12; 95% CI: 2.01–4.85) than immunocompetent populations (OR: 1.85; 95% CI: 1.30–2.63), highlighting the influence of host susceptibility on outcomes. Similarly, forest plots for mycotoxin-related organ toxicity show that aflatoxins and ochratoxins contribute disproportionately to hepatic and renal dysfunction, with pooled relative risks exceeding 2.5. These subgroup analyses not only clarify risk stratification but also illustrate the forest plot’s utility in visualizing differential effects across populations and exposure types.

Heterogeneity, as indicated by the forest plots and corresponding I² statistics, ranged from moderate to high (68–72%), reflecting variability in study populations, geographic regions, exposure measurement, and outcome definitions. While high heterogeneity could raise concerns regarding the comparability of studies, the use of a random-effects model mitigates this by accounting for between-study variability. The forest plots visually confirm that even with heterogeneity, most studies align in the direction of effect, supporting the overall validity of the meta-analytic conclusions.

Funnel plots (Figure 4) were examined to assess publication bias, which occurs when studies with significant results are more likely to be published than those with null or negative results. In this meta-analysis, the funnel plots were largely symmetrical, and Egger’s regression test yielded a p-value of 0.08, suggesting minimal publication bias. Some asymmetry was observed among smaller studies, indicating that small studies with null findings may have been underreported. However, sensitivity analyses, conducted by excluding these small studies sequentially, demonstrated that the pooled effect estimates remained significant and largely unchanged. This indicates that any potential bias had limited impact on the overall conclusions, enhancing confidence in the robustness of the findings.

The funnel plots also provided insights into the precision of studies. Larger studies with smaller standard errors clustered around the pooled effect estimate at the top of the funnel, while smaller studies with wider confidence intervals were distributed more broadly along the base. This pattern is consistent with expectations in meta-analytic datasets and reinforces the reliability of the pooled estimates derived primarily from larger, high-quality studies.

Interpreting forest and funnel plots together allows for a nuanced understanding of both effect magnitude and study quality. Forest plots quantify the strength of associations and highlight heterogeneity, while funnel plots help identify potential biases that could distort conclusions. For example, the forest plot for cutaneous fungal infections (Figure 4) shows a slightly lower pooled odds ratio (OR: 1.76; 95% CI: 1.22–2.53) compared to systemic infections, but the direction of effect remains consistent. Corresponding funnel plot analysis indicated minimal bias, suggesting that the observed lower effect size reflects a true difference in risk rather than selective reporting.

The interpretation of these plots also underscores the importance of environmental and occupational factors in shaping risk. Forest plots for studies conducted in tropical or high-humidity regions show consistently higher effect sizes, reflecting elevated fungal proliferation and mycotoxin production in these climates. Similarly, occupational exposure, such as handling stored grains or working in agricultural environments, amplifies the risk of cutaneous and systemic infections, as visualized through larger effect estimates in forest plots. By integrating these visualizations, the meta-analysis elucidates both the biological and environmental determinants of fungal-associated health outcomes.

Finally, the plots highlight the strength and limitations of the current evidence base. While the forest plots indicate strong and consistent associations across multiple studies, the observed heterogeneity and minor funnel plot asymmetry emphasize the need for careful interpretation. Future research should aim to reduce variability by standardizing exposure assessment, improving outcome definitions, and including larger, high-quality studies across diverse populations. Despite these limitations, the integration of forest and funnel plots provides a clear, comprehensive view of the evidence, reinforcing the conclusion that exposure to filamentous fungi and their metabolites is significantly associated with adverse health outcomes.

Overall, the combined interpretation of forest and funnel plots confirms the robustness, reliability, and consistency of the meta-analytic findings. Forest plots demonstrate significant effect sizes across both systemic and organ-specific outcomes, highlight population subgroups at higher risk, and quantify heterogeneity. Funnel plots indicate minimal publication bias and verify that pooled estimates are not unduly influenced by small or selective studies. Together, these visualizations validate the statistical conclusions, provide clarity regarding the strength and consistency of the evidence, and emphasize the need for continued monitoring and preventive interventions in high-risk populations.

4. Discussion

The findings from this systematic review and meta-analysis provide compelling evidence regarding the health risks associated with filamentous fungi, emphasizing both the pathogenic and toxicological implications for humans. Filamentous fungi, particularly species from the Aspergillus, Penicillium, and Fusarium genera, have long been recognized as significant contributors to human disease, yet their prevalence and the breadth of their effects remain underappreciated in public health discourse (Egbuta et al., 2017). These fungi can cause direct infections in immunocompromised populations, while their mycotoxins—secondary metabolites—pose substantial risks even at low exposure levels, primarily through contaminated food and environmental sources (Awuchi et al., 2021; Agriopoulou et al., 2020).

Aspergillus species emerged as the most studied genus, with Aspergillus candidus and A. clavatus repeatedly implicated in opportunistic infections and mycotoxin production (Ahmadi et al., 2012; Artigot et al., 2009). The clinical significance of Aspergillus is particularly notable in individuals with hematologic malignancies or compromised immune systems, where invasive pulmonary aspergillosis remains a leading cause of morbidity and mortality (Georgiadou & Kontoyiannis, 2012). Beyond infection, aflatoxins produced by Aspergillus spp. demonstrate hepatotoxic and carcinogenic properties, corroborating earlier evidence linking chronic exposure to liver dysfunction and hepatocellular carcinoma (Awuchi et al., 2020; Bennett & Klich, 2003). Forest plot analyses in the included studies consistently illustrated elevated odds ratios for both infection and toxin-related health outcomes, confirming the broad health implications of Aspergillus exposure. Funnel plots suggested minimal publication bias, enhancing confidence in the robustness of these associations.

Penicillium species, while often overshadowed by Aspergillus, present a dual risk as agents of food spoilage and opportunistic infections (Awuchi et al., 2021). Penicillium toxins, such as patulin, have been shown to induce cytotoxicity and genotoxicity in human cells (Artigot et al., 2009). Although the frequency of clinical infections is lower than that of Aspergillus, the consistent presence of Penicillium in processed foods raises concerns about chronic exposure and cumulative health effects. Meta-analytic data revealed moderate heterogeneity in health outcomes associated with Penicillium, which can be partly attributed to variability in detection methods, environmental conditions, and dietary habits across study populations.

Fusarium species, and their associated trichothecene and zearalenone mycotoxins, were consistently linked to systemic toxicities and reproductive health disruptions (Bryla et al., 2018; Janik et al., 2021). Trichothecenes exhibit potent inhibition of protein synthesis, leading to gastrointestinal disturbances, immunosuppression, and neurotoxicity, whereas zearalenone exerts estrogenic effects, disrupting endocrine function and reproductive health (Janik et al., 2020). The meta-analysis indicated that Fusarium exposure is often underreported due to diagnostic limitations, yet forest plots confirmed significant associations between contaminated cereals and adverse outcomes. Notably, the funnel plots for Fusarium-related studies demonstrated asymmetry, suggesting potential underrepresentation of small studies reporting null findings, highlighting the need for comprehensive reporting and standardized monitoring protocols.

An important observation from this review is the multifaceted routes of human exposure to filamentous fungi. Inhalation of spores, ingestion of contaminated foodstuffs, and dermal contact all contribute to the overall burden of disease (Egbuta et al., 2017; Awuchi et al., 2021). Mycotoxins are particularly concerning due to their stability during food processing, which allows them to persist in diets despite conventional food safety measures (Bryden, 2007; Berthiller et al., 2011). The interplay between environmental factors, such as humidity and temperature, and agricultural practices including pesticide and heavy metal use, may exacerbate fungal proliferation and toxin production, further amplifying human health risks (Alengebawy et al., 2021).

Interestingly, the review also underscores the paradoxical role of fungi in human nutrition and health. Edible fungi and mycoprotein products offer significant nutritional benefits, including essential micronutrients, proteins, and bioactive compounds that contribute to healthy aging and metabolic health (Beelman et al., 2019; Derbyshire & Delange, 2021; El Sebaaly et al., 2019). However, the protective versus harmful effects of fungi hinge on species, strain, processing methods, and exposure levels. Filamentous fungi, particularly Aspergillus and Penicillium species, showed moderate to high bio-remediation capacity compared with bacterial strains (Table 4). This duality highlights the necessity for rigorous monitoring and targeted interventions that maximize nutritional benefits while minimizing pathogenic and toxic exposures.

Table 4. Heavy Metal Removal Performance of Selected Microorganisms Categorized by Efficiency Level

The molecular mechanisms underlying fungal toxicity further elucidate the health risks observed in epidemiological studies. Cytochrome P450 enzymes in Aspergillus species, for example, mediate patulin biosynthesis, contributing to the cytotoxicity observed in human epithelial cells (Artigot et al., 2009). Fusarium trichothecenes disrupt ribosomal function, leading to apoptosis and immune dysregulation (Janik et al., 2021). These mechanistic insights are crucial for developing targeted detoxification strategies and therapeutic interventions, as they enable the identification of vulnerable molecular pathways and the design of inhibitory compounds to mitigate toxicity.

From a public health perspective, the cumulative evidence emphasizes the urgent need for robust exposure mitigation strategies. Preventive measures include improved agricultural practices, food storage and processing protocols, and regulatory limits for mycotoxins in food and feed (Agriopoulou et al., 2020; Awuchi et al., 2020). Additionally, awareness campaigns targeting both healthcare professionals and the general public are vital to recognize early signs of fungal exposure and contamination. Surveillance programs employing molecular detection methods, alongside conventional culturing techniques, can enhance the accuracy and timeliness of fungal identification, reducing the likelihood of severe infections and chronic toxin exposure.

Despite the comprehensive nature of this review, several limitations must be acknowledged. Heterogeneity across studies, including differences in diagnostic approaches, geographic regions, and population characteristics, may introduce variability in the reported outcomes. Moreover, many studies relied on observational designs, limiting causal inference, and some meta-analytic estimates were derived from relatively small sample sizes, particularly for Penicillium and Fusarium species. Nevertheless, the overall trends and statistically significant associations underscore the relevance of filamentous fungi as a public health concern.

In summary, this systematic review and meta-analysis affirms that filamentous fungi represent a multifaceted threat to human health, encompassing both infectious and toxicological pathways. Aspergillus, Penicillium, and Fusarium species are consistently implicated in adverse outcomes ranging from opportunistic infections to hepatotoxicity, immunosuppression, and reproductive disruption. While fungi contribute valuable nutrients in food products, the balance between beneficial and harmful effects is precarious and contingent upon species, exposure, and processing practices. The findings underscore the importance of integrated approaches to fungal risk management, combining surveillance, public health education, regulatory oversight, and mechanistic research to mitigate exposure and safeguard human health (Egbuta et al., 2017; Agriopoulou et al., 2020; Awuchi et al., 2021). Ultimately, advancing our understanding of fungal pathophysiology and mycotoxin dynamics is critical for informed policy-making and the development of effective interventions that protect populations from the pervasive risks posed by filamentous fungi.

5. Limitations

Despite the rigorous methodology employed in this systematic review and meta-analysis, several limitations must be acknowledged. First, heterogeneity among included studies was moderate to high, reflecting differences in study populations, geographic locations, exposure assessments, and outcome definitions. While random-effects models were used to account for this variability, residual heterogeneity may still influence the precision of pooled estimates. Second, although funnel plot analysis and Egger’s test suggested minimal publication bias, smaller studies with null or negative results may have been underreported, potentially affecting the overall findings. Third, the reliance on observational studies for most outcomes limits causal inference, as confounding factors such as underlying health conditions, environmental exposures, and socioeconomic variables may have influenced results. Fourth, data on specific fungal species and mycotoxin types were often incomplete or inconsistently reported, restricting the granularity of subgroup analyses. Finally, variability in diagnostic methods and reporting standards across studies could have introduced measurement bias. These limitations underscore the need for well-designed, prospective studies with standardized exposure and outcome assessments to validate and expand upon the current findings.

6. Conclusion

This meta-analysis demonstrates a significant association between fungal exposure and adverse health outcomes, with higher risks observed in immunocompromised populations. Forest and funnel plot analyses confirm the robustness and consistency of these findings. Despite heterogeneity and minor potential biases, the evidence underscores the need for targeted preventive strategies, improved monitoring, and further high-quality research to mitigate health risks associated with filamentous fungi and their metabolites.

References