The contribution of root border cells as a defense barrier against soil-borne pathogen Verticillium dahliae: Insights from the host cotton and the non-host corn.

ABSTRACT This study evaluated the antifungal potential of essential oils and plant extracts for the control of Monilinia fructicola , the causal agent of brown rot in peaches, under in vitro and postharvest conditions. Essential oils and cold aqueous, hot aqueous, and hydroalcoholic extracts of Allium sativum , Syzygium aromaticum , Origanum vulgare , Rosmarinus officinalis , Artemisia absinthium and Eugenia uniflora were evaluated using direct and indirect contact assays. In vitro assays showed complete inhibition of mycelial growth by the essential oils of A. sativum , S. aromaticum and O. vulgare , comparable to the chemical control (captan). The essential oil of A. sativum exhibited the lowest minimum inhibitory concentration (0.04%), whereas essential oils of S. aromaticum and O. vulgare showed minimum inhibitory concentrations of 0.16%. Plant extracts exhibited predominantly fungistatic effects. Scanning electron microscopy revealed structural damage to fungal hyphae induced by the essential oils, confirming their fungitoxic activity. In postharvest assays, only the essential oil of O. vulgare and the chemical treatment, when applied by direct contact, completely suppressed disease incidence and severity over 7 days of storage. Although the essential oils of A. sativum and S. aromaticum reduced disease development at early stages, their efficacy decreased over time, particularly under indirect contact. Biochemical analyses indicated that essential oil treatments, particularly O. vulgare , induced defence responses in the host fruit, as evidenced by increased protein content and enhanced phenylalanine ammonia‐lyase and chitinase activity. These results indicate that essential oils—particularly that of oregano—act through a dual mechanism involving direct antifungal activity and induction of fruit defence responses. These findings highlight their potential as sustainable alternatives for postharvest management of brown rot in peaches.

Objective: To assess the prevalence of allergic fungal rhinosinusitis among patients with nasal polyps and associate it with risk factors Methods: This cross-sectional study design was conducted at the ENT department of Combined Military Hospital Quetta, from January to December 2024. One hundred and sixty-three adult patients presenting with bilateral nasal polyps were enrolled in the study. All the patients underwent functional endoscopic sinus surgery, and the histopathology sample reports were evaluated. Patients were assessed according to the Bent and Kuhn diagnostic criteria for allergic fungal rhinosinusitis, which include the presence of nasal polyps with characteristic double-density CT findings, fungal hyphae on fungal microscopy, and the presence of eosinophilic mucin on histopathology samples, and were labelled as having allergic fungal rhinosinusitis. Results: Out of 163, there were 76 (46.6%) males and 87 (53.4%) females with a mean age of 49.10±9.4 years. Allergic fungal rhinosinusitis was found in 50 (30.7%) of the patients. Females were more commonly found to have allergic fungal rhinosinusitis as compared to males (34.0% vs 66.0%, p=0.032). A higher percentage of patients with allergic fungal rhinosinusitis were found to have a history of allergy (p<0.001), diabetes (p<0.001) and hypertension (p=0.001) as compared to those without allergic fungal rhinosinusitis. Conclusion: The frequency of allergic fungal rhinosinusitis was reported to be 30.7% among patients presenting with nasal polyps, where the mean nasal score was higher among such patients, along with various risk factors, including gender, history of allergy, diabetes and hypertension. Keywords: Nasal polyps, allergic fungal sinusitis, risk factors, allergy, fungal infection

Mycorrhizal fungi serve as fundamental agents in forest establishment and progression, underpinning critical ecosystem functions through symbiotic root associations. Drylands, which cover nearly half of Earth’s land, have limited forest establishment, and factors influencing mycorrhization in these stressful environments remain unclear. Here, we integrate large-scale field surveys along aridity gradients with greenhouse experiments and over 33,000 microscopic mycorrhizal observations, revealing that aridity significantly enhances mycorrhization. Mycorrhizal fungi undergo niche modification, whereby facilitative microbial interactions promote mycorrhization under aridity stress. We identify a core synthetic microbial community linked to mycorrhization and provide mechanistic evidence that this community facilitates mycorrhization through physical attachment to fungal hyphae and by alleviating soil metabolite inhibition that otherwise suppresses mycorrhization under arid conditions. In this work, our findings highlight the role of microbial interkingdom interactions in driving tree mycorrhizal colonization in arid regions, offering critical insights for guiding tree planting and restoration efforts in drylands.

A radiometric analysis of soil samples from the studied areas of the exclusion zone, which have been under exposure to chronic irradiation for more than three decades, was carried out, from which micromycetes of different species of the genus Trichoderma were isolated, and their biological activity was studied. It was shown that the radial growth rate of these fungi exceeds the average values for species of this genus. It was found that the isolated strains of the species T. koningi and T. atroviride have very high (over 90 %) antagonistic activity towards the phytopathogenic fungi Rhizoctonia solani and Nectria inventa, and the species T. viride, T. harzianum, and Trichoderma sp. showed antagonistic activity towards the studied species of phytopathogenic hyphal fungi researched, but their magnitude was different.

Wildfire is a major ecological disturbance with profound implications for forest ecosystems, particularly through its effects on soil quality and structure. This study examined the impacts of a severe wildfire (2019) on soil physical, chemical, and micromorphological properties in a mountainous forest region of Yunnan Province, southwestern China. Soil samples were collected from post-fire and adjacent control sites at three depths (0-5, 5-15, and 15-30cm) and analyzed using a factorial design. Results indicated that the most pronounced fire-induced alterations occurred in the surface layer (0-5cm). Post-fire soils exhibited increased sand content but reduced clay content, porosity, and field capacity. Chemically, wildfire increased soil pH while reducing electrical conductivity (EC), soil organic carbon (SOC), available phosphorus (AP), and available potassium (AK) in the surface horizon, with limited changes in deeper layers. Micromorphological observations revealed modified aggregate structures, carbonized root fragments, dispersed fungal hyphae, and an abundance of charcoal particles, reflecting both physical and biological disturbances. The soil mass fractal dimension (D) also increased in post-fire soils, indicating greater structural fragmentation and aggregate breakdown. This increase in D is consistent with the observed decreases in porosity and aggregate stability, reflecting a more heterogeneous and degraded pore network. These findings demonstrate that wildfire predominantly affects surface soil horizons, altering infiltration capacity, nutrient dynamics, and microbial activity. Integrating physical, chemical, and micromorphological assessments offers a more comprehensive perspective on fire-induced soil changes, and may provide a useful foundation for post-fire management strategies aimed at supporting the resilience of forest ecosystems under increasing wildfire frequency and intensity.

This study assessed the efficacy of the entomopathogenic fungus Akanthomyces muscarius against the greenhouse whitefly Trialeurodes vaporariorum and the gray mold pathogen Botrytis cinerea. It was found that the area of B. cinerea infestation on tomato leaves was reduced after pre-treatment with conidia of two out of three A. muscarius strains. Strain Vl 61 demonstrated the greatest inhibitory effect on gray mold development. This strain, when applied to the whole tomato plants, most effectively suppressed disease development on the fourth day, with subsequent plant growth at the level of the control. When tomato leaves were infested with B. cinerea pre-inoculated with T. vaporariorum, whitefly mortality from treatment with Vl 61 spores was 69 % on the fourth day, 50 % higher than in the absence of B. cinerea. The efficacy of A. muscarius against the phytopathogen in the presence of whitefly larvae was significantly lower than on plants not inhabited by the whitefly. Fluorescence microscopy demonstrated the hyperparasitic properties of A. muscarius against B. cinerea. It was found that, when strain Vl 61* (labeled with a fluorescent tag) and the gray mold pathogen were grown together on agar medium, the fungal hyphae entwined B. cinerea hyphae, penetrating, growing, and emerging, destroying the pathogen’s cell walls. These results suggest that strain Vl 61 of A. muscarius holds promise for protecting tomatoes from B. cinerea and T. vaporariorum.

REPORTscraping showed hyaline septate fungal hyphae, consistent with rhinocerebral mucormycosis.He was started on intravenous (IV) amphotericin B (cumulative dose 3-4 gm), followed by oral posaconazole for 2 months.Endoscopic debridement and maxillectomy under general anesthesia were advised.However, due to high surgical risk associated with Eisenmenger syndrome, the family declined surgical consent and opted for medical management only.The patient gradually improved, although he had residual third nerve palsy.He was discharged after being prescribed 3 months of oral treatment.

Background: The majority of skin conditions in Ayurveda fall under "Kushta". Dadru is a sub-classification of the Kushta. Dadru (Tinea corporis), a skin condition, has become more prevalent in our society in recent years. It is a tridoshaja vyadhi, with a predominance of pitta and kapha doshas. It is a skin condition marked by papules with dark pigmentation and a tendency to spread. Clinical findings: A 55-year-old male presented to the skin OPD with a circular, dry, hyperpigmented active lesion over the chest region and associated with severe itching, symptoms had persisted for the past six months. The lesion had gradually increased in size despite topical cream use. Microscopic examination of skin scrapings revealed fungal elements, confirming the diagnosis of Dadru (Tinea corporis). The condition was managed with only external application of Bilwadi agada and Nalpamradi taila, both applied twice a day for 10 days, followed by only application of Nalpamradi taila twice a day for the next 10 days; thus, had two follow-ups on the 11th and 21st days. Possessing krimighna (antimicrobial), kandughna (anti-pruritic), and vishaghna (detoxifying), Twak-prasadana (skin-brightening) synergistically relieves symptoms of Dadru. By the 21st day, Kandu (itching) and Raga (erythema) reduced to grade 1 (mild), while Pidaka (papular eruptions) and Rookshata (dryness) and Utsanna Mandala (elevated lesions) completely resolved to grade 0. Outcome: Notable improvement in symptoms and the size of the lesion was reduced, restoring the skin to its normal texture and colour and on KOH mount for presence of fungal hyphae microscopic examination of the lesion showed Hyphae absent and no spores indicating clearing of the fungal elements. Conclusion: The external application of Bilwadi agada and Nalpamradi taila significantly reduced symptoms and achieved microbial clearance, thus managing Dadru effectively in 21 days.

Rationale:Endogenous fungal endophthalmitis (EFE) is a rare but sight- and life-threatening condition that often arises in immunocompromised patients. While Aspergillus fumigatus is the predominant pathogen, Aspergillus niger-associated endocarditis leading to bilateral EFE has not been previously reported.Patient concerns:A 51-year-old woman with a history of two renal transplants over the past 30 years, along with diabetes mellitus, hypertension, and ongoing immunosuppressive therapy, presented with rapidly progressive bilateral visual impairment, beginning with acute vision loss and ocular pain in the right eye, followed by involvement of the left eye within two days. She had no systemic symptoms such as fever, chest pain, or dyspnea.Diagnosis:Initial aqueous humor analysis, including Gram staining, fungal culture, and viral PCR, yielded negative results. As the disease progressed to bilateral involvement within 2 days, a diagnostic vitrectomy was performed. Vitreous sampling demonstrated fungal hyphae, and culture confirmed Aspergillus niger. Transesophageal echocardiography further revealed mitral valve vegetation, establishing the diagnosis of fungal infective endocarditis with bilateral endogenous endophthalmitis.Interventions:The patient received empiric intravitreal antibiotics and antivirals initially, followed by diagnostic pars plana vitrectomy and systemic antifungal therapy after identification of Aspergillus. Mitral valve replacement surgery was subsequently performed.Outcomes:Despite surgical intervention and systemic antifungal treatment, the patient developed disseminated fungal sepsis and died 2 weeks after valve replacement.Lessons:This case highlights the diagnostic challenges of EFE and emphasizes the importance of considering fungal endocarditis as a source of infection in immunosuppressed patients presenting with severe vitritis, even in the absence of systemic symptoms. Early recognition, prompt vitrectomy with intravitreal antifungal therapy, and thorough systemic evaluation are essential to improve visual and systemic outcomes.

Fungus-growing ants maintain an ectosymbiotic microbial garden, an intertwined mesh of fungal symbiont hyphae and microbiota growing through plant substrates. Here, we investigate how different plant diets influence the garden lignocellulosic profile, and whether the microbiota respond to dietary changes. Colonies of Atta sexdens were provided with four different dietary regimens, varying in fiber composition and nutritional content. Diet changed the garden lignocellulosic profile, also influencing the microbial taxonomic composition. The diet of only leaves enriched the garden in Bacillus and Weissella, while a diet of only fruits/cereals lead to a Carnimonas and Mesoplasma enrichment; diets mixing leaves and fruits/cereals intermittently and alternatively enriched the garden in Bacillus, Mesoplasma, and Weissella. The fungal crop development and the spatial distribution of the microbiota and biofilm also varied according to the diet. Our findings suggest that diet has a pivotal role in determining whether ant colonies function optimally and remain healthy.

Bacterial-fungal interactions have traditionally been attributed to secondary metabolites, but the role of the bacterial extracellular matrix in shaping these relationships has remained unclear. Here, we demonstrate that the extracellular matrix protein TasA is a key mediator in the antagonistic interaction between Bacillus subtilis and Botrytis cinerea. TasA enables Bacillus to tightly adhere to fungal hyphae, disrupts the β-glucan layer, and compromises fungal cytoskeletal integrity synergistically with fengycin, which causes cytological damage. Additionally, TasA acts as a carrier for bacillaene, amplifying its fungistatic activity. In response, Botrytis mounts a multifaceted defense, enzymatically degrading fengycin, producing antibacterial oxylipins, and activating adaptive programs such as hyphal branching and chlamydospore formation. Our findings reveal the previously unrecognized role of extracellular matrix components in fungal suppression and the modulation of fungal adaptive responses. This study reveals the complex interplay between microbial aggression and defense, providing new insights into the ecological dynamics of microbial competition and coexistence.

Hyphal forms of human pathogenic fungi cause invasive disease in humans, but the hyphal response to antifungals is understudied. In the major fungal pathogens Aspergillus fumigatus and Candida albicans, we used microfluidic-coupled, fluorescence-mediated live-cell imaging to capture the real-time responses of fungal hyphae to clinical concentrations of AmBisome or Caspofungin. In both fungi, AmBisome exposure caused rapid growth arrest (<15 min) and subcellular reorganisation and, in C. albicans, localised expansions of lipid-like structures from the hyphal perimeter. Responses to Caspofungin exposure were slower, with initial lytic effects occurring after 1.5 or 4 h in A. fumigatus and C. albicans hyphae, respectively. While C. albicans hyphae undergo unsalvageable hyphal lysis in response to Caspofungin, A. fumigatus exhibits several compensatory growth behaviours, including a novel resuscitative growth response, that circumvents lytic events to maintain apical and sub-apical hyphal growth. This study reveals how the differing biologies of the two pathogens affected outcomes and contributes to the highly disparate rates of antifungal efficacy amongst commonly used drugs, where spore/yeast-derived inhibitory doses may be underestimated to arrest/kill the invasive hyphal morphotypes in vitro. Human pathogenic cause >2M deaths per year and we have a limited number of antifungals in the clinic to combat these infections. Those drugs are increasingly meeting resistance in killer fungi and our understanding of drug responses are limited. Our main assessment of antifungal resistance comes from end-point 48 h drug culture of the fungus and is performed by eye for presence of absence of growth in a well. Further, these tests are performed on the pre-invasive form of the fungus, the yeast or spore (for speed and simplicity), not the invasive filamentous form present during systemic infection of organs in humans. This work shows how the invasive filamentous form of two major human fungal pathogens respond to two frontline clinical antifungal drugs with real time microscopy. We describe the cell death and/or adaptive growth responses via live-cell microscopy to understand the morphological and cellular responses over time. We were able to do this by coupling fluorescently engineered pathogens and live-cell 4D microscopy to microfluidic delivery of culture media and/or antifungal drug. We used the triggerable microfluidics to first establish invasive filamentous growth without drug, and keep the cells in the same focal plane (with shallow roof that kept the filaments growing up out of the focal plane). We then triggered the switch to media with drug(s) to perfuse drug while observing comparative cellular responses live, up to 10h in two pathogenic fungi. In both pathogens (Aspergillus fumigatus and Candida albicans) we saw similar immediate responses to one drug (Ambisome), which inhibited filamentous growth almost immediately. In stark contrast, Caspofungin induced different but continued forms of growth to known inhibitory doses against invasive fungal filaments. A. fumigatus (a filamentous mould fungus) appeared hard-wired to continue filamentous growth by various compensatory regenerative growth forms, including a novel 'resuscitative' form we describe which occurs after the drug causes filament bursting. However, C. albicans filaments (a polymorphic yeast) respond by transitioning from filamentous to yeast growth (all filaments eventually burst, leaving only budding yeast). This study details the short- and long-term responses of invasive pathogenic fungal filaments to drugs and highlights that the traditional spore/yeast-derived inhibitory doses for these fungi may be insufficient for the invasive form of the fungus, where more attention to the filamentous form is needed.

Introduction: Penetrating ocular trauma complicated by secondary fungal keratitis presents a fundamental inflammatory paradox in ophthalmology. While corticosteroids are clinically imperative to suppress trauma-induced fibrinoid syndrome and prevent phthisis bulbi, they are traditionally contraindicated in active mycotic infections due to the risk of potentiating fungal proliferation. This report evaluates a dissociated therapeutic protocol utilizing systemic corticosteroids alongside topical antifungals to navigate this therapeutic impasse. Case presentation: We present the case of a 26-year-old male sustaining a penetrating corneal injury from a metallic nail. Following primary repair, the patient developed a severe anterior chamber reaction with Standardization of Uveitis Nomenclature (SUN) Grade 3+ cells and a corneal infiltrate exhibiting feathery margins. Potassium Hydroxide 10% wet mount confirmed the presence of fungal hyphae. A dissociated therapeutic regimen was initiated: intensive topical Natamycin 5% administered hourly to target the ocular surface infection, synchronized with oral Methylprednisolone (24 mg/day; 0.4 mg/kg) to control intraocular inflammation via the systemic circulation. The regimen successfully decoupled the immune response. Intraocular inflammation resolved within 14 days, preventing the formation of permanent synechiae or cyclitic membranes. The fungal infection was eradicated, confirmed by negative serial corneal smears and the clinical resolution of the infiltrate into a stable leucoma. Visual acuity improved from Hand Motion (LogMAR 2.30) at presentation to 6/45 (LogMAR 0.88) at Day 35. No fungal recurrence was observed during the follow-up period. Conclusion: A dissociated strategy using intermediate-dose systemic corticosteroids to manage internal inflammation, while reserving the ocular surface for aggressive topical antifungal therapy, may offer a viable strategy for globe salvage in complex traumatic fungal keratitis cases where standard protocols are undefined.

Endophytic bacteria play an important role in inhibiting plant pathogens. This study aimed to screen endophytic bacteria from upland rice with antagonistic activity against Fusarium graminearum, evaluated their antagonistic potential against F. graminearum, assessed their anti-fungal substances, and elucidated the underlying mechanisms. Some methods were performed, including dual-culture antagonism assays, lipopeptide extraction, identification of antifungal compounds via LC-MS and HS-GC-MS, metabolomic analysis, and microscopic observation. Two endophytic bacterial strains, URR1 and URR2, were identified as Pseudomonas sp. and Bacillus subtilis, respectively. Dual-culture antagonism assays demonstrated that both strains exhibited strong inhibitory activity against F. graminearum, with inhibition rates of 69.73% and 76.33%, respectively. In vitro experiments further revealed that bacterial suspensions at approximately 3.3 × 10 8 CFU·mL-1 significantly alleviated stress in upland rice seedlings infected with F. graminearum after 7 days. Both crude lipopeptides and volatile organic compounds (VOCs) markedly suppressed the hyphal growth of the pathogen. The maximum inhibition rate of crude lipopeptides reached 63.86% after 96 hours of treatment, while VOCs showed a peak hyphal inhibition rate of 30.38% after 48 hours of exposure. Antimicrobial lipopeptides, comprising 10 distinct surfactin isoforms and 7 fengycin variants, as well as VOCs such as acetone, ethanol, trichloromethane, pyruvic acid, and propadiene, were identified. After antagonism with lipopeptides extracted from endophytic bacteria URR2, the fungal hyphae of F. graminearum exhibited morphological abnormalities. Notably, treatment with URR2 resulted in the upregulation of metabolites and activation of key metabolic pathways. Metabolomic analysis indicated that the differentially upregulated metabolites encompassed a wide range of classes, including organic acids and derivatives, lipids and lipid-like molecules, organoheterocyclic compounds, organic oxygen compounds, benzenoids, organic nitrogen compounds, nucleosides, nucleotides and analogues, phenylpropanoids and polyketides, as well as lignans and neolignans. The analysis revealed that the associated molecules were significantly concentrated in multiple metabolic pathways, primarily involving ABC transporters, protein digestion and absorption, amino acid biosynthesis, aminoacyl-tRNA biosynthesis, the phosphotransferase system (PTS), starch and sucrose metabolism, among others. These results conclusively demonstrate the strong antifungal activity of strain URR2 against F. graminearum. The antagonistic mechanism of B. subtilis against F. graminearum appears to be multifaceted. Overall, our findings indicate that URR2 has promising potential to be developed as a novel biocontrol agent for the development of sustainable agriculture.

Mucormycosis is an angioinvasive fungal infection caused by ubiquitous saprophytic fungi of the order Mucorales. In humans, the infection is generally caused by inhalation of spores or traumatic inoculation in cutaneous wounds, typically affecting immunocompromised patients. In animals, Mucorales infection is common in cattle, but is rare in the other species, including felids. No cases of mucormycosis have been reported to date in non-domestic felids, to our knowledge. We describe a case of mucormycosis in a 5-y-old female captive tiger (Panthera tigris). The affected tiger was part of a group of 9 subjects, 7 of which had serohemorrhagic nasal discharge, anorexia, and depression, followed by hemorrhagic diarrhea. Five of the tigers died and organs from one animal were sent for histologic examination, which revealed severe necrohemorrhagic enteritis, with periodic acid-Schiff- and Grocott methenamine silver-positive fungal hyphae. Mucor colonies were cultured from sawdust, which served as litter for the animals, indicating a possible source of infection through inhalation of spores and/or ingestion of contaminated sawdust. Mucormycosis is a possible differential for necrohemorrhagic enteritis in captive felids; careful storage and periodic assessment of the materials used as litter for these animals is recommended.

Extracting fungal hyphae with their naturally associated microbiota from soil samples presents a significant challenge due to their small size, typically in the micrometer range, and the formation of dynamic fungal networks. We combined elements of previous protocols and automated the wet-sieving steps of the methodology to efficiently extract fungal hyphae from various soil types, including natural loamy soils. This approach reduces manual handling, minimizes operator-dependent variability, and shortens processing time by up to 2.5-fold. Unlike earlier methods that require sand or glass bead supplementation, which can introduce artificial conditions and limit large-scale field applications, our Sieving and Sucrose Centrifugation (SSC) method avoids these drawbacks. The SSC technique enables both quantification of hyphal length density (HLD) and, importantly, preserves surface-associated microbes for downstream analyses. Among the tested methods, SSC yielded the highest hyphal length density. Using a combination of microscopy, molecular techniques, and next-generation sequencing (NGS), we demonstrate that this method allows targeted study of bacteria tightly attached to fungal hyphae. Furthermore, the SSC approach effectively enriched fungal hyphae from a highly diverse soil community, establishing a dependable tool for advancing research on fungal hyphae as microbial hotspots in soil ecosystems.

Fungal pleural effusion (FPE) is an uncommon cause of pleural infection, accounting for only 1%–3% of all cases. It usually occurs in immunocompromised individuals, such as those with malignancy, diabetes, human immunodeficiency virus (HIV), or chronic steroid use. We report a rare case of left-sided Candida-associated pleural effusion in an 85-year-old man with no known comorbidities or immunosuppressive condition. The patient presented with left-sided chest pain and weakness for two weeks. The pleural fluid analysis showed an exudative, lymphocyte-predominant effusion with low adenosine deaminase (ADA) levels. Medical thoracoscopy with pleural biopsy revealed fungal hyphae consistent with Candida species, confirming the diagnosis. The patient was treated with oral fluconazole 200 mg twice daily for eight weeks, showing marked clinical improvement and radiological resolution without the need for surgical intervention. This case highlights that FPE can occur even in immunocompetent individuals, emphasising the need for early pleural biopsy and timely antifungal therapy in undiagnosed exudative effusions

In pathology (surgical pathology/cytopathology) specimens, Aspergillus species can be challenging to differentiate from other fungi that produce hyaline septate hyphae by morphology alone. It has been suggested that fruiting bodies-if present-indicate Aspergillus. The aim of this study was to determine whether the presence of fruiting bodies in pathology specimens is specific for Aspergillus. Specimens containing fungal hyphae with fruiting bodies were identified and fungal culture and PCR results were reviewed to determine the identity of the fungi. To determine whether fruiting bodies can be formed in tissue by other fungi, non-Aspergillus fungi were included for analysis if cultures or PCR confirmed a non-Aspergillus fungus. Fruiting bodies were present in specimens from 13 patients (12 surgical pathology, 1 cytology). In 11/13, the identity of the fungus was confirmed (10 Aspergillus fumigatus, 1 Rhizopus species). In 6/13, A. fumigatus was confirmed by microbiologic cultures. In 4/13, A. fumigatus was confirmed by PCR on formalin-fixed paraffin-embedded tissue. In 2/13 specimens, cultures/PCR were not performed. The one Rhizopus sp. was confirmed by culture. In most (10/13) specimens, fruiting bodies of Aspergillus consisted of yellow vesicles containing a row of radiating phialides, occasional stalks/conidiophores and detached yellow conidia (spores) in the background. In contrast, fruiting bodies of the Rhizopus sp. were sporangia (large spore-containing sacs) lacking phialides. To evaluate fruiting bodies in non-Aspergillus fungi, 18 specimens were identified with hyphal structures in tissue with available microbiologic culture or PCR results. No fruiting bodies were identified in the 18 non-Aspergillus fungal infections (11 Fusarium, 6 Mucorales order genera, 1 Acrophialophora). In pathology specimens, fruiting bodies with the morphologic features described in this study (conidiophores with yellow-brown vesicles, attached phialides, detached yellow conidia) are specific for Aspergillus and can be differentiated from Rhizopus, whose fruiting bodies feature sporangia (large spore-containing sacs lacking phialides).

BackgroundAlthough pulmonary mucormycosis is rare, it is highly invasive and carries a significant mortality rate. Due to its nonspecific clinical manifestations, it is often misdiagnosed as other invasive fungal diseases. Bronchoalveolar lavage fluid metagenomic next-generation sequencing is a rapid, precise, and comprehensive method for pathogen detection, showing great potential in the early diagnosis of pulmonary mucormycosis in a single-center retrospective series. It provides clinicians with faster and more accurate etiological information, thereby improving patient outcomes and reducing mortality rates.MethodsThis study conducted a retrospective analysis of the clinical data from 14 patients diagnosed with pulmonary mucormycosis between 1/6/2021 and 30/6/2024. Peripheral blood samples were collected to perform a complete blood count, measure C-reactive protein levels, and conduct 1,3-β-D-glucan and Galactomannan tests. Lung tissue samples were sent to the pathology laboratory for histological examination. Bronchoalveolar lavage fluid was subjected to fungal culture and metagenomic next-generation sequencing. Additionally, a three-month follow-up on the patients’ survival status was carried out via telephone.ResultsMales accounted for 57.14% of the cases. Diabetes mellitus was present in 12 patients (85.71%, 12/14), and fever was observed in 12 patients (85.71%, 12/14). The 14 patients were categorized as proven cases (4 cases), probable cases (4 cases), and possible cases (6 cases). Two patients (14.29%, 2/14) were diagnosed with disseminated mucormycosis. Chest Computed Tomography scans revealed cavities in half of the patients (50.00%, 7/14). Fungal hyphae were identified in all the histopathological examinations (100%, 4/4). Metagenomic next-generation sequencing detected Mucorales pathogens in all the (100%, 14/14) cases, which is higher positivity than the positive rates of the 1,3-β-D-glucan test (35.71%, 5/14), Galactomannan test (42.86%, 6/14) and fungal culture (7.14%, 1/14). The turnaround time for metagenomic next-generation sequencing reports is 1–3 days, which is much shorter than the time required to obtain results from fungal culture (2–5 days). Additionally, metagenomic next-generation sequencing identified bacterial and viral co-infections, with 11 patients diagnosed as having mixed infections. All patients were treated with antifungal agents targeting Aspergillus species, such as voriconazole, posaconazole, isavuconazole, or amphotericin B, resulting in 9 patients improving, 2 patients being transferred to higher-level hospitals, and 3 patients discontinuing treatment. The 90-day follow-up revealed a mortality rate of 28.57%.ConclusionMetagenomic next-generation sequencing can serve as an important complement to traditional diagnostic methods, enabling rapid and accurate differentiation of Mucorales from other fungi. This allows patients to receive timely and targeted antifungal therapy, playing a critical role in early intervention and improving prognosis.