Introduction
Malassezia furfur (M. furfur) is a lipophilic yeast that is part of the normal human skin microbiota. Under specific conditions, it can transition to a pathogenic form (1,2). This organism possesses unique biological characteristics, notably a lipid-rich cell wall structure that requires exogenous long-chain fatty acids for growth (3). In immunocompetent individuals, M. furfur typically causes superficial infections such as pityriasis versicolor and folliculitis (4). However, in certain populations, particularly neonates, its infection can lead to life-threatening systemic disease (5). The pathogenic mechanisms involve biofilm formation, phospholipase secretion and immune evasion, rendering it a notable opportunistic pathogen in nosocomial infections (1,6,7).
Neonates are susceptible to M. furfur infections because of multiple interacting factors (6), including an underdeveloped skin barrier, characterized by a thin stratum corneum and incomplete intercellular junctions, which provide a physical pathway for pathogen invasion (8). Concurrently, an immature immune system, including reduced neutrophil activity and low complement levels, impairs pathogen clearance (9). Clinical interventions further exacerbate infection risk. For instance, lipid emulsions in parenteral nutrition serve as ideal media for fungal proliferation and central venous catheters (CVCs) offer surfaces for biofilm formation (10). These risk factors are synergistically amplified in preterm infants, positioning them as a high-risk group for invasive Malassezia infections.
Traditional fungal detection methods for M. furfur have notable limitations (11). Routine cultures require lipid supplementation, such as olive oil, and exhibit slow growth, typically taking 5-7 days, thereby resulting in a clinical laboratory detection rate <30%. Morphological identification relies on the characteristic ‘spaghetti and meatballs’ appearance under microscopy, which is often absent in systemic infection samples (12). Serological tests are hindered by immature antibody responses in neonates, leading to low sensitivity (13). These technical constraints contribute to missed or delayed diagnosis in >40% of invasive cases, highlighting key gaps in current diagnostic frameworks (14).
Metagenomic next-generation sequencing (mNGS) offers a promising solution for these diagnostic challenges (15). This technology captures all nucleic acid sequences in a sample without bias, thus eliminating the need for pathogen-specific primers or prior culture (16). Theoretically, mNGS can detect the entire microbial genome of a clinical specimen. Compared with traditional methods, mNGS has distinct advantages for detecting fastidious organisms, as it does not rely on pathogen viability, can identify unculturable microbes and enables simultaneous species identification and resistance gene profiling via bioinformatic analysis (17). Furthermore, the turn-around time is typically shortened to 24-48 h. Multiple studies have demonstrated the efficacy of mNGS in detecting bacterial pathogens in neonatal bloodstream infections (18,19); however, its utility in diagnosing fungal infections, especially lipid-dependent yeasts, remain underexplored (20).
The present study analyzed a representative M. furfur infection case and conducted a systematic review to illustrate the need to incorporate M. furfur detection into the routine fungal pathogen screening spectrum for neonatal bloodstream infections. The technical value of mNGS was evaluated for the detection of this organism. The present case report highlights the diagnostic improvements offered by mNGS over conventional methods, which include timely detection, enhanced capability for identifying polymicrobial infections and the beneficial impact on clinical decision-making. These findings are expected to provide evidence-based guidance for optimizing the diagnostic pathways involved in neonatal invasive fungal infections and to inform revisions of infection control strategies in intensive care units. The present case report provides specific attention to technological innovations in mNGS for detecting lipid-dependent fungi, with the aim of addressing current diagnostic blind-spots in neonatal fungal bloodstream infections.
Discussion
M. furfur is a lipophilic yeast commonly residing on human skin, especially in sebum-rich regions such as the scalp, face and upper torso (72). It exhibits strict requirements for exogenous fatty acids owing to the absence of the acyl-CoA synthase genes necessary for endogenous long-chain fatty acid biosynthesis (73,74). Although M. furfur is typically commensal and contributes to skin microbiota homeostasis, it can undergo pathogenic transformations under certain conditions.
The transition of M. furfur from a benign skin commensal to opportunistic pathogen involves a multifactorial interplay between host immunity, skin and gut barrier integrity, microbial dysbiosis and nutrient-rich external environments (72). In extremely preterm infants, underdeveloped skin, immature immune systems, frequent antibiotic administration and invasive procedures such as CVC insertion are the key drivers of pathogenic conversion. Both the present case report and a previous report underscore these risk factors (8,75). In particular, CVCs serve as entry points for M. furfur into the bloodstream and are recognized as risk factors for catheter-associated Malassezia fungemia (31).
M. furfur should not be regarded solely as a commensal skin infection in the NICU setting. Despite being a part of the normal human skin microbiota, M. furfur is currently recognized worldwide as an opportunistic fungal pathogen in the NICU, particularly among preterm and extremely low birth-weight infants exposed to lipid-containing parenteral nutrition, prolonged CVC placement, broad-spectrum antimicrobial therapy or prolonged hospitalization (75,76). In this high-risk population, M. furfur may cause catheter-related fungemia, persistent bloodstream infections, pulmonary involvement, thrombocytopenia and sepsis-like clinical deterioration. Clinical manifestations are often subtle and non-specific and include temperature instability, respiratory distress, feeding intolerance, apnea, bradycardia, poor peripheral perfusion, lethargy and reduced responsiveness. Laboratory findings may include leukocytosis, elevated CRP levels and thrombocytopenia (8). Because these features overlap with those of Gram-negative bacterial sepsis or other invasive fungal infections, missed or delayed diagnoses remain common. Thus, M. furfur should be actively considered as a differential diagnosis in neonatal bloodstream infections, particularly when routine blood cultures are negative and risk factors such as lipid parenteral nutrition or prolonged catheter use are present.
For the patient of the present case report, M. furfur was confirmed as the causative pathogen using mNGS, highlighting the limitations of currently available conventional microbiological diagnostic methods for neonatal infections.
Although conventional culture remains essential for obtaining viable isolates and performing antifungal susceptibility testing, the capacity to recover M. furfur from clinical specimens is limited (77). Because M. furfur is a lipid-dependent yeast, routine blood culture systems and standard fungal media may fail to support its growth unless appropriate lipid supplementation is provided (50). Lipid-enriched media, such as Dixon's or Sabouraud dextrose agar supplemented with olive oil or long-chain fatty acids, can improve recovery and allow for downstream phenotypic testing. However, culture sensitivity is limited by strict lipid requirements, slow growth and the possibility of overgrowth by faster-growing microorganisms. These limitations contribute to false-negative conventional culture results and the delayed recognition of bloodstream Malassezia infections (53,78).
Microscopic techniques, such as potassium hydroxide preparation combined with India ink staining or Gram staining, offer rapid and convenient diagnostic options. However, their sensitivity is limited, particularly for the detection of organisms in sterile body fluids. These methods cannot provide species-level identification and rely heavily on the skills and experience of laboratory personnel (53).
The (1→3)-β-D-glucan (BDG) assay is commonly employed to evaluate systemic fungal infections (79). However, because the cell wall of M. furfur contains little to no β-D-glucan, BDG testing in such infections frequently yields false-negative results. Therefore, BDG assays should not be solely used to exclude the diagnosis of Malassezia-associated infections (80).
Conventional PCR and reverse-transcription-quantitative PCR exhibit high sensitivity and specificity and can directly detect pathogen DNA or RNA in clinical samples. Nonetheless, these approaches require optimized PCR primer design and specialized technical platforms and currently lack widely available commercial assays or standardized operating procedures (53,56).
Although matrix-assisted laser desorption ionization time-of-flight mass spectrometry theoretically enables rapid and accurate species-level identification, its clinical application for Malassezia species remains limited. Challenges include difficulties in protein extraction from lipid-rich yeasts and inadequate coverage of Malassezia sequences in current databases, preventing their widespread adoption in routine diagnostics (53,61).
As a rapidly advancing untargeted detection technology, mNGS is particularly advantageous for identifying pathogens in difficult, low-abundance infections (81). In the present study, mNGS achieved etiological confirmation when traditional methods failed to yield positive results. However, mNGS application faces several challenges, including its high cost, the need for specialized personnel to interpret the results and incomplete representation of Malassezia sequences in current databases, which increases the risk of misinterpretation due to background contamination.
The present case represents the first neonate with bloodstream infection caused by M. furfur in mainland China identified using mNGS. One culture-confirmed case of neonatal M. furfur fungemia has previously been reported in Taiwan, China (47). The present case differs from the adult pulmonary case reported by Zhang et al (76) in terms of host background, infection site and diagnostic metho. The study by Zhang et al (76) showed that M. furfur infection mainly occurred in preterm infants and immunocompromised adults, that fungemia was the most common clinical form, and that invasive procedures and total parenteral nutrition were the main risk factors. Adult infection is commonly associated with acquired immunosuppression, transplantation, hematological disease or severe gastrointestinal disease, whereas neonatal infection is mainly driven by extreme prematurity, extremely low birth weight, immature skin and mucosal barriers, prolonged NICU hospitalization, central venous catheterization, broad-spectrum antimicrobial exposure and lipid-containing parenteral nutrition. In addition, neonatal manifestations are usually non-specific and may resemble bacterial sepsis, while routine culture may fail to detect M. furfur because of its lipid-dependent growth requirements. The clinical significance of the present case lies in the timely identification of M. furfur in an extremely preterm neonate with sepsis-like deterioration, multiple predisposing factors and negative or inconsistent conventional microbiological findings. This case highlights the value of mNGS in detecting rare lipid-dependent fungal pathogens that may be missed in routine cultures.
Through an in-depth analysis of the present case, combined with a retrospective review of the literature, the present case report summarized all known risk factors associated with M. furfur bloodstream infection in neonates. These include extremely low birth weight (<1,500 g), extreme prematurity (gestational age <28 weeks), immature immune function, CVC placement, prolonged parenteral nutrition containing lipid emulsions, exposure to broad-spectrum antibiotics and compromised skin and intestinal mucosal barrier integrity. These risk factors are prevalent among patients in NICUs, creating a favorable clinical environment for the transition of M. furfur from a commensal organism to a pathogen (76).
Notably, the patient of the present case report exhibited nearly all of the aforementioned high-risk factors as well as typical clinical characteristics, including persistently negative blood cultures, difficulty in localizing the source of infection and lack of response to conventional antibiotic therapy. When facing diagnostic challenges, mNGS enabled rapid and definitive identification of M. furfur, elucidated the causative agent and directly guided antifungal treatment strategies. As a result, the patient's clinical symptoms improved rapidly, culminating in a successful recovery and hospital discharge, thereby affirming the importance of early pathogen identification and timely intervention for improving clinical outcomes.
This case highlights that although invasive M. furfur infections remain under-recognized in routine clinical practice, their pathogenic potential in high-risk NICU patients should not be underestimated. The present case report suggests that clinical infection guidelines should consider including M. furfur in the spectrum of potential pathogens, particularly in cases of neonatal sepsis characterized by negative blood cultures. This inclusion can help achieve a more comprehensive diagnostic framework. Furthermore, the present study reinforces the core principles of the early detection, diagnosis and treatment of neonatal infections. Proactive assessment of infection-related risk factors, coupled with the timely application of advanced diagnostic techniques, such as mNGS, can help overcome the limitations of conventional methods and significantly enhance diagnostic efficiency and precision.
At present, no standardized treatment guidelines specifically targeting M. furfur bloodstream infections are available. Management generally follows the principles established for invasive fungal diseases and is individualized based on the antifungal susceptibility profiles and clinical status of the neonate. Commonly employed antifungal agents include amphotericin B, itraconazole, voriconazole and fluconazole. Amphotericin B demonstrates sensitivity against the majority of Malassezia species but carries a high risk of toxicity, particularly nephrotoxicity, necessitating cautious use (82). Itraconazole and voriconazole exhibit good in vitro activity against M. furfur, although clinical experience with their use in neonates remains limited (51). Fluconazole has been effective in certain cases, although some studies report lower susceptibility of Malassezia species to fluconazole. Given its favorable safety profile, stable plasma concentration and relatively low side effect burden, fluconazole is more commonly used in neonatal populations (47).
In the present case, the patient, an extremely preterm infant with multiple infection-related risk factors, was promptly managed upon diagnosis by discontinuing lipid emulsion support and initiating antifungal therapy with fluconazole. Despite reported variability in M. furfur susceptibility to fluconazole, clinical decision-making in this case was guided by the limited extent of infection, absence of deep organ involvement and preserved hepatic and renal function (47). Fluconazole was selected as the initial treatment owing to its safety and controllability. Following treatment initiation, the patient's body temperature normalized, inflammatory markers decreased, no recurrence of infection was observed and the patient was successfully discharged. This outcome indicates that under specific clinical circumstances, fluconazole may be a viable therapeutic option for neonatal M. furfur bloodstream infections.
As a lipophilic yeast, M. furfur lacks the ability to synthesize long-chain fatty acids and relies entirely on exogenous lipid sources for growth. In the context of parenteral nutrition, especially when lipid emulsions are used, the organism can colonize catheter surfaces and extensively proliferate, thereby leading to fungemia. Therefore, immediate discontinuation of lipid emulsions upon diagnosis or strong suspicion of M. furfur infection is key for halting ongoing pathogen expansion and controlling infection progression. Combining antifungal therapy with cessation of lipid supplementation has been reported to significantly shorten the disease course and improve clinical outcomes (8).
With rapid advancements in mNGS, its application in the diagnosis of infectious diseases has become increasingly widespread. The core advantages of mNGS include its high sensitivity. mNGS can detect extremely low-abundance microbial DNA or RNA, providing supplementary diagnostic yield in scenarios where traditional methods exhibit low detection rates. Unlike targeted PCR or culture-based methods, the broad detection spectrum of mNGS does not rely on predefined pathogenic hypotheses. Instead, it enables unbiased and comprehensive screening of all potential pathogens within a sample. An important utility of this method is the rapid identification of novel or rare pathogens in clinical situations where conventional diagnostic techniques are limited. In such cases, mNGS facilitates the discovery of rare, fastidious or even novel pathogens, thereby providing key information for guiding clinical decision-making (16,83).
Despite these advantages, some limitations of mNGS must not be overlooked. Currently, the cost of mNGS is substantially higher when compared with that of traditional culture or PCR methods, restricting its widespread adoption in frontline clinical practice. Owing to factors such as sample handling, reagent impurities and environmental microbial DNA contamination, mNGS may detect non-pathogenic microorganisms, complicating data interpretation. Additionally, interpretation of mNGS results requires integration of sequencing read counts, relative microbial abundance and clinical context. It demands sophisticated bioinformatic platforms and highly trained personnel for accurate analysis. Furthermore, owing to the lack of antimicrobial susceptibility information as provided by culture-based methods, mNGS cannot directly provide antimicrobial susceptibility profiles, necessitating either additional testing or empiric therapeutic decision-making (17,84). In the present case report, additional lipid-supplemented fungal cultures using Dixon's or Sabouraud agar with an olive oil overlay were not performed after a positive mNGS result and no viable M. furfur isolate was available for phenotypic antifungal susceptibility testing. This represents a limitation of the present case, although the patient displayed favorable clinical and laboratory responses after fluconazole therapy. The mNGS results should not be interpreted in isolation, particularly for M. furfur, a common skin-associated commensal organism that may theoretically be introduced during blood collection or sample processing. Nonetheless, in this case, M. furfur was not detected in the corresponding NTC, whereas the blood sample yielded 148 reads assigned to M. furfur with a relative abundance of 72.6%, arguing against NTC-related background contamination. Moreover, the patient had multiple risk factors for invasive Malassezia infection, including extreme prematurity, extremely low birth weight, prolonged NICU hospitalization, broad-spectrum antimicrobial exposure, lipid exposure and catheter-related risks. The clinical course was also consistent with bloodstream infection, including fever, clinical deterioration, markedly elevated CRP levels, thrombocytopenia, negative or inconsistent conventional cultures and subsequent improvement after antifungal therapy. Collectively, these findings support a true bloodstream infection rather than incidental skin contamination or overinterpretation of the isolated mNGS results. Therefore, mNGS results must not be interpreted in isolation during clinical practice. Comprehensive evaluation combining patient clinical presentations, laboratory findings, imaging studies and results from conventional microbiological tests is essential. For any detected pathogen, a scientific judgment must be made based on infection site, host immune status, sequencing abundance and pathogenic relevance to avoid misdiagnosis, missed diagnosis or overtreatment.
In the present case report, by integrating mNGS findings within the clinical context, M. furfur was successfully identified as the causative pathogen. Targeted antifungal therapy was subsequently implemented, leading to a favorable clinical outcome. This case demonstrates the value of adopting mNGS technology in the management of complex infections in neonates.
The present case report describes the case of a neonatal bloodstream infection caused by M. furfur in mainland China that was identified directly from blood samples utilizing mNGS. Through a systematic review of the literature, the epidemiological characteristics, clinical manifestations, outcomes, predisposing factors and diagnostic approaches associated with neonatal M. furfur infection were summarized. Based on the diagnostic and therapeutic experience from this case, we propose that M. furfur should be considered in the differential diagnosis of neonatal sepsis, particularly among extremely low-birth-weight infants with persistent clinical suspicion of infection despite negative conventional microbiological tests. The application of mNGS may provide important diagnostic support in such cases, but its results should be interpreted in conjunction with overt clinical manifestations, conventional microbiology techniques and patient-specific risk factors.