Talaromides A-C, Bioactive Cyclic Heptapeptides from Talaromyces siglerae Isolated from a Marine Sponge.

Three new cyclic heptapeptides, talaromides A-C (1-3), were isolated from cultures produced by the fungus Talaromyces siglerae (Ascomycota), isolated from an unidentified sponge. The structures, featuring an unusual proline-anthranilic moiety, were elucidated by analysis of spectroscopic data and chemical transformations, including the advanced Marfey's method and GITC derivatization. Talaromides A and B inhibited migration activity against PANC-1 human pancreatic cancer cells without significant cytotoxicity.

Cytotoxic and Antibacterial Meroterpenoids Isolated from the Marine-Derived Fungus Talaromyces sp. M27416.

Three novel meroterpenoids, taladrimanins B-D (1-3), were isolated from the marine-derived fungus Talaromyces sp. M27416, alongside three biogenetically related compounds (4-6). We delineated taladrimanin B's (1) structure using HRESIMS and NMR, confirmed its configuration via quantum chemical NMR analysis and DP4+ methodology, and verified it through X-ray crystallography. ECD calculations determined the absolute configuration of compound 1, while comparative NMR and ECD analyses elucidated the absolute configurations of 2 and 3. These compounds are drimane-type meroterpenoids with a C10 polyketide unit (8R-configuration). We proposed a biosynthetic pathway and noted that compound 1 showed cytotoxic activity against MKN-45 and 5637 cell lines and selective antibacterial effects against Staphylococcus aureus CICC 10384.

Pulmonary dimorphic fungal infections among HIV/AIDS non-TB patients with chronic cough in Kampala, Uganda.

INTRODUCTION: Dimorphic fungi cause infection following the inhalation of spores into the pulmonary system. In the lower respiratory tract, the conidia transform into yeasts, which are engulfed by alveolar macrophages and may be destroyed without disease manifestation. However, in some immunocompromised individuals, they may persist and cause active fungal disease characterized by formation of granulomas in the infected tissues, which may mimic Mycobacterium tuberculosis (MTB). OBJECTIVE: To determine the prevalence of pulmonary dimorphic fungal infections among HIV/AIDS patients with non-TB chronic cough at Mulago National Referral and Teaching Hospital in Kampala, Uganda. METHODS: Sputum samples were collected from 175 consented HIV/AIDS patients attending the immuno-suppression syndrome (ISS) clinic at the hospital. Upon Xpert MTB/RIF sputum testing, 21 patients tested positive for MTB, and these were excluded from further analysis. The other 154 sputum negative samples were then subjected to PCR for dimorphic fungi at MBN Clinical Laboratories. Singleplex PCR was used to detect the target sequences in selected respective genes of each dimorphic fungal species of interest. DNA amplicons were detected based on gel electrophoresis. RESULTS: Dimorphic fungi were detected in 16.2% (25/154) of the studied population. Of these 9.1% (14/154) had Blastomyces dermatitidis and 7.1% (11/154) had Talaromyces marneffei. The remaining 84% of the studied participants had no dimorphic fungi. Histoplasma capsulatum, Coccidioides immitis and Paracoccidioides brasiliensis were not detected in any of the participants. CONCLUSION: Dimorphic fungi (B. dermatitidis and T. marneffei) were found in 16.2% of the HIV/AIDS patients with non-TB chronic cough in Kampala, Uganda. We recommend routine testing for these pathogens among HIV/AIDS patients with chronic cough.

4-Hydroxy-2-pyridone derivatives with antitumor activity produced by mangrove endophytic fungus Talaromyces sp. CY-3.

OSMAC strategy is a useful tool for discovering series of metabolites from microorganism. Five new sambutoxin derivatives (1-2, 4, 8-9), together with seven known compounds (3, 5-7, 10-12), were isolated from Talaromyces sp. CY-3 under OSMAC strategy and guidance of molecular networking. Their planar structures and absolute configurations were determined by NMR, HRESIMS, ECD spectra and common biosynthetic pathway. In bioassay, compounds 1-12 showed cytotoxicity to tumor cell lines with IC50 values in the range of 1.76-49.13 µM. The antitumor molecular mechanism of 10 was also explored. In vitro compound 10 significantly inhibited the growth and proliferation of two lung cancer cell lines (A549 and H1703). Furthermore, colony formation, EdU analysis, flow cytometry and Western blot analysis showed that 10 could induce cell cycle arrest in G0/G1 phase by promoting the expression of p53 and p21. The molecular mechanism of its antitumor effects in vitro is that 10 arrests the cell cycle by activating the p21/CyclinD1/Rb signaling pathway and the p53 pathway. Our results identified a lead small molecule compound with efficient antitumor growth and proliferation activity.

Characterization and Effect of a Nematophagous Fungus Talaromyces cystophila sp. nov. for the Biological Control of Corn Cyst Nematode.

The dynamic of plant-parasitic nematode populations in soil is closely related to soil microorganisms. Fungi from Heterodera zeae cysts were isolated to explore the phenomenon of decline in the H. zeae population in the soil. Phylogenetic study of partial ITS, BenA, CaM, and RPB2 gene sequences, in addition to morphological investigations, was utilized to identify a nematode-destroying fungus. The nematicidal activity of a novel strain GX1 against H. zeae was assessed in vitro and in the greenhouse. Our findings revealed that strain GX1 is a new species of Talaromyces, named Talaromyces cystophila. It has a strong parasitic and lethal effect on H. zeae cysts, with 91.11% parasitism on cysts at 3 days after treatment. The contents of second-stage juveniles (J2s) and eggs inside the cysts were degraded and formed dense vacuoles, and the damaged eggs could not hatch normally. The spore suspension exhibited high nematophagous activity against nematodes, and fermentation filtrate exhibited marked inhibition of egg hatching and nematicidal activities on J2s. The hatching inhibition rates of eggs exposed to 1 × 108 CFU/ml spore suspensions or 20% 1-week fermentation filtrate (1-WF) for 15 days were 98.56 and 100%, respectively. The mortality of J2s exposed to 1 × 108 CFU/ml spore suspension reached 100% at 24 h; exposure to 50% 2-WF was 98.65 and 100% at 24 and 48 h, respectively. Greenhouse experiments revealed that the spore suspension and fermentation broth considerably decreased H. zeae reproduction by 56.17 to 78.76%. T. cystophila is a potential biocontrol strain with nematophagous and nematicidal activity that deserves attention and application.

New Dipyrroloquinones from a Plant-Derived Endophytic Fungus Talaromyces sp.

Two new dipyrroloquinones, namely talaroterreusinones A (1) and B (2), together with four known secondary metabolites, terreusinone A (3), penicillixanthone A (4), isorhodoptilometrin (5), and chrysomutanin (6), were isolated from the solid culture of the endophytic fungus Talaromyces sp. by integrating mass spectrometry-based metabolic profiling and a bioassay-guided method. Their planar structures and stereochemistry were elucidated by comprehensive spectroscopic analysis including NMR and MS. The absolute configuration at C-1″ of terreusinone A (1) was established by applying the modified Mosher's method. Compounds 1-6 were evaluated for anti-inflammatory activity and cytotoxicity. As a result, 1-3 inhibited the LPS-stimulated NO production in macrophage RAW264.7 cells, with IC50 values of 20.3, 30.7, and 20.6 µM, respectively. Penicillixanthone A (4) exhibited potent cytotoxic activity against Hep G2 and A549 cell lines, with IC50 values of 117 nM and 212 nM, respectively, and displayed significant antitumour effects in A549 cells by inhibiting the PI3K-Akt-mTOR signalling pathway.

Cytotoxic Ergosteroids from a Strain of the Fungus Talaromyces adpressus.

Nine new ergosteroids (1-9) and seven known ones (10-16) were isolated from Talaromyces adpressus. Their structures and absolute configurations were determined by the interpretation of NMR spectroscopic data, HRESIMS, ECD calculations, and single-crystal X-ray diffraction analyses. Structurally, compound 1 was an ergosteroid with two epoxy and a 3α-OH group at ring A, while compounds 8 and 9 had a contracted ring A with a peroxy bridge between C-3 and C-9, which were reported for the first time. Compounds 2-6, 9, 11, and 15 displayed cytotoxic activities with IC50 values ranging from 0.4 to 32 µM, and compound 7 exhibited an immunosuppressive effect against LPS-induced B lymphocyte proliferation with an IC50 value of 8.6 µM. The structure-activity relationships of these compounds are briefly discussed.

Endophytic Fungus Talaromyces sp. MR1 Promotes the Growth and Cadmium Uptake of Arabidopsis thaliana L. Under Cadmium Stress.

Endophytes play essential roles in plant growth under metal(loid)s stress. An endophytic fungus strain MR1 was isolated from the roots of Miscanthus floridulus collected from a lead-zinc mining area (Huayuan, China), which could produce indole-3-acetic acid and have Cadmium (Cd) tolerance. Further 18S rRNA sequencing analysis showed that it was highly similar (99.83%) to Talaromyces pinophilus. In pot experiments, we explored the effects of strain MR1 on the growth and Cd uptake of a wide-type Arabidopsis thaliana under low (LC) and high (HC) Cd concentrations. The results showed that MR1 effectively increased the dry weight of aboveground and underground tissues by 25.95-107.21% in both LC and HC groups. Due to MR1 inoculation, the Cd content in the underground tissues was significantly (p < 0.05) decreased by 39.28% under low Cd concentration, while it was significantly (p < 0.05) increased by 28.28% under high Cd concentration. Besides, MR1 inoculations significantly (p < 0.05) increased the total content of removed Cd (17.080 µg) and BCF (0.064) by 129.77% and 153.95% under high Cd concentration. Therefore, we speculated that MR1 might be selected as the effective microbial agent to increase crop yield and control Cd content in the crop in light Cd-contaminated soil. Besides, MR1 could potentially enhance the phytoremediation efficiency of extremely Cd-contaminated soil.

Identification of glutathione metabolic genes from a dimorphic fungus Talaromyces marneffei and their gene expression patterns under different environmental conditions.

Talaromyces marneffei is a human fungal pathogen that causes endemic opportunistic infections, especially in Southeast Asia. The key virulence factors of T. marneffei are the ability to survive host-derived heat and oxidative stress, and the ability to convert morphology from environmental mold to fission yeast forms during infection. Glutathione metabolism plays an essential role in stress response and cellular development in multiple organisms. However, the role of the glutathione system in T. marneffei is elusive. Here, we identified the genes encoding principal enzymes associated with glutathione metabolism in T. marneffei, including glutathione biosynthetic enzymes (Gcs1 and Gcs2), glutathione peroxidase (Gpx1), glutathione reductase (Glr1), and a family of glutathione S-transferase (Gst). Sequence homology search revealed an extended family of the TmGst proteins, consisting of 20 TmGsts that could be divided into several classes. Expression analysis revealed that cells in conidia, mold, and yeast phases exhibited distinct expression profiles of glutathione-related genes. Also, TmGst genes were highly upregulated in response to hydrogen peroxide and xenobiotic exposure. Altogether, our findings suggest that T. marneffei transcriptionally regulates the glutathione genes under stress conditions in a cell-type-specific manner. This study could aid in understanding the role of glutathione in thermal-induced dimorphism and stress response.

Clinical features of Talaromyces marneffei infection in HIV-positive and HIV-negative individuals: A retrospective study in southern China.

Talaromyces marneffei (TSM) is a temperature-dependent dimorphic fungus endemic to Southeast Asia and southern China. As the number of people at risk of TSM infection continues to increase, the clinical manifestations are becoming increasingly complex, posing challenges for clinical management. In this study, we analyzed the medical records of 99 patients (71 human immunodeficiency virus [HIV]-positive and 28 HIV-negative) diagnosed with TSM infection from January 1, 2017, to December 31, 2022, in southern China and compared the clinical manifestations in HIV-positive and HIV-negative patients. Most patients (83/99, 84%) were male. The incidence of skin and soft tissue involvement (48% vs. 21%, P = .016); disseminated infection with blood circulation, hematopoietic, lymphatic, alimentary, or central nervous system involvement (69% vs. 36%, P = .002); and gastrointestinal bleeding (33% vs. 9%, P = .023) was higher in the HIV-positive group than the HIV-negative group. The HIV-positive group also had significantly higher alanine aminotransferase (ALT) levels (31 [26-42] vs. 14 [11-16] U/l, P < .001) and ALT/aspartate transaminase ratio (1.9 [1.5-2.2] vs. 1.3 [1.1-1.6], P = .006) than the HIV-negative group. The time to diagnosis (5.5 ± 1.1 vs. 5.1 ± 1.4 days, P = .103), antifungal regimen (P = .278), case fatality rate (20% vs. 21%, P = .849), and relapse/reinfection rate (11% vs. 19%, P = .576) did not differ significantly between the HIV-positive and HIV-negative groups. Poor antiretroviral therapy adherence (OR = 26.19, 95%CI 3.26-210.70, P = .002), advanced age (OR = 1.13, 95%CI 1.03-1.23, P = .010), and Epstein-Barr virus co-infection (OR = 37.13, 95%CI 3.03-455.64, P = .005) were independent risk factors for all-cause mortality from TSM infection in HIV-positive patients. Overall, the predominant infection sites, clinical manifestations, and complications of TSM infection differed by HIV status. However, with prompt diagnosis and appropriate treatment, HIV-positive patients with TSM infection can have similar outcomes to HIV-negative patients.

There are certain differences in the clinical features, sites of infection, and associated complications of Talaromyces marneffei infection between individuals with and without human immunodeficiency virus. It is necessary to accurately identify individuals at high risk to enable prompt diagnosis and standardized treatment.

Extracellular vesicles derived from Talaromyces marneffei contain immunogenic compounds and modulate THP-1 macrophage responses.

Pathogenic eukaryotes including fungi release extracellular vesicles (EVs) which are composed of a variety of bioactive components, including peptides, nucleic acids, polysaccharides, and membrane lipids. EVs contain virulence-associated molecules suggesting a crucial role of these structures in disease pathogenesis. EVs derived from the pathogenic yeast phase of Talaromyces (Penicillium) marneffei, a causative agent of systemic opportunistic mycoses "talaromycosis," were studied for their immunogenic components and immunomodulatory properties. Some important virulence factors in EVs including fungal melanin and yeast phase specific mannoprotein were determined by immunoblotting. Furthermore, fluorescence microscopy revealed that T. marneffei EVs were internalized by THP-1 human macrophages. Co-incubation of T. marneffei EVs with THP-1 human macrophages resulted in increased levels of supernatant interleukin (IL)-1ß, IL-6 and IL-10. The expression of THP-1 macrophage surface CD86 was significantly increased after exposed to T. marneffei EVs. These findings support the hypothesis that fungal EVs play an important role in macrophage "classical" M1 polarization. T. marneffei EVs preparations also increased phagocytosis, suggesting that EV components stimulate THP-1 macrophages to produce effective antimicrobial compounds. In addition, T. marneffei EVs stimulated THP-1 macrophages were more effective at killing T. marneffei conidia. These results indicate that T. marneffei EVs can potently modulate macrophage functions, resulting in the activation of these innate immune cells to enhance their antimicrobial activity.

Case report: Diagnosis of Talaromyces marneffei infection in an HIV-negative patient with septic shock and high-titer anti-interferon gamma autoantibodies by metagenomic next-generation sequencing.

Background: Sepsis is a life-threatening condition caused by a dysfunctional response to infection from the host. Septic shock, a subset of sepsis, caused by Talaromyces marneffei infection (talaromycosis) has rarely been reported. Owing to its slow culture and low yield, talaromycosis is typically misdiagnosed in HIV-negative patients as other infections, such as tuberculosis, bacterial pneumonia, and lung cancer, especially in non-endemic regions. Early and accurate diagnosis as well as efficient treatment options are required to improve prognosis. Method: A 30-year-old HIV-negative Chinese woman from a non-endemic area of T. marneffei was initially misdiagnosed with tuberculosis. She had a poor response to anti-tuberculosis treatment. On July 16, 2022, she was admitted to our hospital; the patient developed septic shock on the third day after hospitalization and was ultimately diagnosed with talaromycosis via metagenomic next-generation sequencing (mNGS). Result: The condition of the patient improved after appropriate treatment with amphotericin B. Furthermore, enzyme-linked immunosorbent assay results confirmed that the patient had a high-titer of anti-interferon gamma (IFN-γ) autoantibodies. Conclusion: HIV-negative individuals with anti-IFN-γ autoantibodies typically have relapsing, refractory, and fatal infections, such as talaromycosis, which is typically misdiagnosed in the initial course of the disease. This can lead to septic shock. Clinicians should be aware that they may encounter HIV-negative patients with T. marneffei infection in non-endemic areas. Thus, mNGS is an effective technology for detecting T. marneffei infection. Additionally, the detection of anti-IFN-γ autoantibodies in these patients would aid in knowing their susceptibility to fatal infections.

Retracted: Isolation and purifications of an ambuic acid derivative compound from marine algal endophytic fungi Talaromyces flavus that induces apoptosis in MDA-MB-231 cancer cells.

In recent years, there has been a lot of buzz about the possibilities of marine microflora as a source of new therapeutic drugs. The strong anti-tumor potency of compounds found in marine resources reflects the ocean's enormous potential as a source of anticancer therapeutics. In this present investigation, an ambuic acid derivative anticancer compound was isolated from Talaromyces flavus, and its cytotoxicity and apoptosis induction potential were analyzed. T. flavus was identified through morphological and molecular analysis. The various organic solvent extracts of T. flavus grown on different growth mediums were evaluated for cytotoxicity on different cancer cell lines. The potent cytotoxicity was shown in the ethyl acetate extract of a fungal culture grown in the M1-D medium for 21 days. Furthermore, the anticancer compound was identified using preparative thin layer chromatography, followed by its purification in significant proportions using column chromatography. The spectroscopic and chromatographic analysis revealed that the structure of the purified molecules was an ambuic acid derivative. The ambuic acid derivative compound showed potent cytotoxicity on MDA-MB-231 (breast cancer cells) with an IC50 value of 26 µM and induced apoptosis in the MDA-MB-231 cells in a time-dependent and reactive oxygen species-independent manner.

Thalidomide promotes NLRP3/caspase-1-mediated pyroptosis of macrophages in Talaromyces marneffei infection.

Macrophage-derived inflammatory cytokines are critical for host defense against Talaromyces marneffei (T. marneffei) infection among HIV/AIDS patients, and excessive inflammatory cytokines are associated with poor outcomes of AIDS-associated talaromycosis. However, the underlying mechanisms of macrophage-caused pyroptosis and cytokine storm are poorly understood. Here, in the T. marneffei-infected mice and macrophages, we show that T. marneffei induced pyroptosis in macrophages through the NLRP3/caspase-1 pathway. The immunomodulatory drug thalidomide could promote the pyroptosis of macrophages infected T. marneffei. In T. marneffei-infected mice, the splenic macrophages underwent increasing pyroptosis as talaromycosis deteriorated. Thalidomide ameliorated inflammation of mice, while amphotericin B (AmB) in combination with thalidomide did not improve overall survival compared with AmB alone. Taken together, our findings suggest that thalidomide promotes NLRP3/caspase-1-mediated pyroptosis of macrophages in T. marneffei infection.

Red biocolorant from endophytic Talaromyces minnesotensis: production, properties, and potential applications.

Fungal colorants are gradually entering the global color market, given their advantages of being less harmful to human health, as well as having greater stability and biotechnological potential, compared to other natural sources. The present work concerns the isolation and identification of an endophytic filamentous fungus, together with the chemical characterization and assessment of the fluorescence, toxicity, stability, and application potential of its synthesized red colorant. The endophytic fungus was isolated from Hymenaea courbaril, a tree from the Brazilian savannah, and was identified as Talaromyces minnesotensis by phenotypic and genotypic characterization. Submerged cultivation of the fungus resulted in the production of approximately 12 AU500 of a red biocolorant which according to LC-DAD-MS analysis is characterized by being a complex mixture of molecules of the azaphilone class. Regarding cytotoxicity assays, activity against human hepatoblastoma (HepG2) cells was only observed at concentrations above 5.0 g L-1, while antimicrobial effects against pathogenic bacteria and yeast occurred at concentrations above 50.0 g L-1. The biocolorant showed high stability at neutral pH values and low temperatures (10 to 20 °C) and high half-life values (t1/2), which indicates potential versatility for application in different matrices, as observed in tests using detergent, gelatin, enamel, paint, and fabrics. The results demonstrated that the biocolorant synthesized by Talaromyces minnesotensis has potential for future biotechnological applications. KEY POINTS: • An endophytic fungus, which was isolated and identified, synthesize a red colorant. • The colorant showed fluorescence property, low toxicity, and application potential. • The red biocolorant was highly stable at pH 8.0 and temperatures below 20°C.

Integration of metabolomics and transcriptomics analyses reveals sphingosine-1-phosphate-mediated S1PR2/PI3K/Akt pathway involved in Talaromyces marneffei infection of macrophages.

Talaromycosis is a fatal mycosis caused by the thermally dimorphic fungus Talaromyces marneffei (T. marneffei). The pathogenic mechanisms of talaromycosis are still poorly understood. This work combined metabolomics, transcriptomics, and verification experiments in vivo and in vitro to detect metabolic profiles and differentially expressed genes (DEGs) in T. marneffei infected and uninfected macrophages to explore possible pathogenesis and underlying mechanisms. A total of 256 differential metabolites (117 up-regulated and 148 down-regulated) and 1320 DEGs (1286 up-regulated and 34 down-regulated) were identified between the two groups. Integrative metabolomics and transcriptomics analysis showed sphingolipid signaling pathway is the most influential. Verification experiments showed that compared with the control group, the production of sphingosine-1-phosphate (S1P) and the expression of the S1PR1, S1PR2, phosphor-PI3K, and phosphor-Akt genes involved in the sphingolipid signaling pathway have significantly increased in the T. marneffei infection group (p < 0.05). T. marneffei activates the S1PR2/PI3K/Akt pathways in J774A.1 macrophage, regulation of the S1P singling might serve as a promising therapeutic strategy for talaromycosis.

Isolation, structural determination, and antiviral activities of metabolites from vanitaracin A-producing Talaromyces sp.

Vanitaracin A, an anti-hepatitis B virus polyketide, has been previously isolated from Talaromyces sp. In the present study, we searched for novel compounds in the culture broth obtained from a vanitaracin A-producing fungus under various conditions. Three novel compounds (vanitaracin C, vanitaraphilone A, and 2-hydroxy-4-(hydroxymethyl)-6-methylbenzaldehyde) were isolated, and their structures were determined using spectroscopic methods (1D/2D NMR and MS). In addition, the antiviral spectrum of vanitaracin A was examined by measuring its antiviral activities against rabies virus, Borna disease virus 1, and bovine leukemia virus. This compound exhibited antiviral activity against bovine leukemia virus, which is the causative agent of enzootic bovine leukosis. The anti-bovine leukemia virus effects of other compounds isolated from the vanitaracin A-producing fungus, namely, vanitaracins B and C, vanitaraphilone A, and 2-hydroxy-4-(hydroxymethyl)-6-methylbenzaldehyde, were also evaluated. Vanitaracin B, vanitaraphilone A and 2-hydroxy-4-(hydroxymethyl)-6-methylbenzaldehyde were also found to exhibit activity against bovine leukemia virus. These findings reveal the broad-spectrum antiviral activity of the vanitaracin scaffold and suggest several candidates for the development of anti-bovine leukemia virus drugs.

Two previously undescribed phthalides from Talaromyces amestolkiae, a symbiotic fungus of Syngnathus acus.

Amestolkins A (1) and B (2), two previously undescribed phthalides sharing the same planar structure of (1, 5-dihydroxyhexyl)-7-hydroxyisobenzofuran-1(3H)-one were isolated from Talaromyces amestolkiae. Their absolute configurations were elucidated by comprehensive analyses of spectroscopic evidences in high-resolution electrospray mass spectra (HRESIMS) and nuclear magnetic resonance (NMR) combined with electronic circular dichroism (ECD) and NMR calculations. 1 and 2 showed anti-neuroinflammatory activity by inhibiting the gene expressions of proinflammatory factors including C-C motif chemokine ligand 2 (CCL-2), tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6), as well as attenuating the excretion of inducible nitric oxide synthase (iNOS) in BV-2 microglial cells at the concentration of 30 µM.

Case report: Hemophagocytic lymphohistiocytosis in a child with primary immunodeficiency infected with Talaromyces marneffei.

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening immune-mediated disease that affects patients with known genetic defects and is increasingly found among those with autoimmune diseases and persistent infections. Talaromyces marneffei (TM) is a human opportunistic fungus that commonly infects immunodeficient or immunosuppressed individuals. Few TM-associated secondary HLH cases resulting from autoimmune deficiency have been reported previously. The current case study describes a pediatric patient hospitalized with recurrent fever and lymphadenopathy. The child had abnormal blood cell classification, and microscopy revealed mature granulocytes that phagocytized fungal spores. It was speculated that the patient was infected with TM. The pathogen was detected earlier than the blood culture and confirmed by metagenomic next-generation sequencing. Whole-exome sequencing revealed that the patient had complex mutations associated with immunodeficiency. This included a mutation in exon 3 of the CD40LG gene, c.346G>A, which may be linked to hyper-IgM syndrome, a primary immunodeficiency disease with immunoglobulin conversion recombination defects that could explain the patient's increased susceptibility to serious opportunistic infections. In addition, a heterozygous frameshift variant, c.820dup (p.Asp274GlyfsTer61), was detected in exon 6 of CARD9, a key gene associated with fungal immune surveillance. After 4 days of fungal treatment, the abnormal blood cell clusters disappeared, but other infections occurred in succession for 6 months after rehabilitation. The patient was followed with the aim of providing subsequent immunotherapy. This study found that infection can trigger HLH in HIV-negative individuals, highlighting the importance of early definitive identification of the causative agent and investigation of potential immunodeficiency.