Subacute infective endocarditis due to Lodderomyces elongisporus: a case report and review of the literature

Lodderomyces elongisporus, a rare emerging pathogen, can cause fungemia often related to immunosuppression or intravenous devices. Herein, we report the case of a 58-year-old woman with subacute infective endocarditis due to Lodderomyces elongisporus identified by blood fungal culture and whole-genome sequencing, who was treated with antifungals, mitral replacement and endocardial vegetation removal surgery.


Case report
A 58-year-old woman was admitted to our hospital on September 8, 2022 (Day 1) with a 2-month history of repeated low fever, headache, dizziness, weakness in both lower limbs and excessive weight loss (10 kg).Before admission, she had been given a variety of antiinfective agents and antipyretic drugs (β-lactam/β-lactamase inhibitors, quinolones, and macrolides) at the local hospital to no avail.The patient gave no recent travel history.She had a history of rheumatoid arthritis without treatment, type 2 diabetes, and hypertension.This patient did not have intravenous devices or any complications of diabetes.Body temperature was 38.8 • C, a systolic blowing noise was audible in the mitral valve auscultation area, and no superficial lymph nodes in the body were found.Laboratory tests showed a normal white blood cell count of 6.86 × 10 9 /L (normal 3.5-9.5 × 10 9 /L), with 67.2% neutrophils, rheumatoid factor (13.2 IU/mL, normal) and anti-streptolysin O (127.0 IU/mL, normal).Random blood glucose (19.45 mmol/L, normal <7.8), high-sensitivity C-reactive protein (33.96 mg/L; normal 0-5 mg/L), erythrocyte sedimentation rate (78 mm/h; normal 0-20), procalcitonin (0.30 ng/L; normal 0-0.1), and 1,3-β-D-glucan (245.33 pg/mL; normal 0-100) were elevated.Low levels of CD3 + CD45 + T lymphocytes (919 cells/µL; normal 955-2860), CD3 − CD19 + B lymphocytes (103 cells/µL; normal 95-560), CD3 − CD16 + CD56 + NK cells (35 cells/µL; normal 150-1,100), and CD3 + CD16 + CD56 + NKT cells (20 cells/µL; normal 40-300) were observed.The Aspergillus antigen test for serum galactomannan, CrAg lateral flow assay, T-SPOT TB, and parasite and tumor marker tests were negative.No other primary immunodeficiency diseases, human immunodeficiency virus infection, or antibody spectrum of autoimmune diseases was detected.Transthoracic echocardiography showed a vegetation lesion adherent to the patient's anterior mitral leaflet, which was concerning for infectious endocarditis (Day 3) (Figure 1A).Chest computed tomography (CT) revealed left atrial enlargement.Bacterial infective endocarditis was suspected.The patient was started on empirical antibacterial therapy with piperacillin-tazobactam, moxifloxacin and vancomycin.However, the patient's condition did not improve.Five days into the patient's hospital stay, her blood was positive for fungal culture (Day 5) (Figures 1B-E).Using wholegenome sequencing, the organism was identified as Lodderomyces elongisporus (L.elongisporus) (Supplemental Figure 1) from the blood.Antifungal susceptibility testing showed that L. elongisporus is sensitive to fluconazole, flucytosine, itraconazole, caspofungin, amphotericin B and voriconazole (Supplementary Table 1).Then, anti-infectious therapy was increased to caspofungin plus voriconazole for 2 weeks and sequential voriconazole treatment for 2 weeks.Subsequently, the patient's symptoms, including fever, headache, dizziness, and weakness, improved.However, transthoracic echocardiography showed that the vegetation lesion in the anterior mitral leaflet became larger.Thus, mitral replacement and endocardial vegetation removal surgery were performed (Day 30).
The patient's condition worsened after surgery.The white blood cell count of 14.89 × 10 9 /L (normal 3.5-9.5)with 78.4% neutrophils, procalcitonin (3.57ng/L; normal 0-0.1), and 1,3-β-D-glucan (393.46 pg/mL; normal 0-100) increased rapidly.Blood for fungal culture was still positive for L. elongisporus (Day 39), and the result of antifungal susceptibility testing was the same as before.Then, liposomal amphotericin B (5 mg/kg.d,intravenous drip) was administered for 2 weeks, and fluconazole (400 mg/d, per os) continued for 4 weeks.The patient's blood culture turned negative, and the symptoms and signs improved observably (Day 57).After discharge from the hospital (Day 73), the patient received fluconazole for sequential antifungal therapy for the duration of the study (Figure 1F).

Systematic literature review search strategy
We searched PubMed, Web of Science, Embase, and BIOSIS Library for original case reports and cohort studies published in English on Lodderomyces elongisporus (L.elongisporus) between January 1, 1985, and December 31, 2022.We used the keywords "Lodderomyces elongisporus" and "L.elongisporus".The references of the retrieved articles were reviewed for additional relevant citations.The abstracts of all identified articles were viewed, and the full-text versions of relevant articles were retrieved for data extraction and analysis.
The inclusion criteria for articles and cases in the systematic literature review were as follows: (1) original case reports or cohort studies on L. elongisporus published in English between January 1, 1985, and December 31, 2022; and (2) articles describing definitive etiological evidence for L. elongisporus based on pathological and culture proof.

Clinical outcome definitions
The clinical course of infection was divided into the following two categories: (1) survival (complete or partial improvement of clinical symptoms after antifungal treatment); and (2) death.

Data extraction
The following data were extracted: geographical distribution, demographics, pathogen proof, clinical signs and symptoms, involvement sites, diagnosis, treatment, outcomes and minimum inhibitory concentration (MIC) values for antifungal drugs for L. elongisporus.If a case was reported in more than one publication, the most recent article was used for data extraction.

Discussion
L. elongisporus, a rare emerging pathogen that is an ascomycetous yeast, can cause fungemia often related to immunosuppression or intravenous devices (4).To date, 23 invasive L. elongisporus infection patients have been reported, including 22 with fungemia and 1 with meningitis (Table 1) (4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16).Notably, only one patient was a neonate, and the other 22 cases occurred in adult patients.In addition to bloodstream infections, L. elongisporus was also present in the central nervous system and intracranial vascular system in 7 patients.Only 3 patients (patients 1 and 11 in the systematic literature review and the present patient) had infective endocarditis (4,14).All 3 patients presented with cardiac valve vegetations adherent to the prosthetic aortic valve, anterior aorta and mitral valve.Two patients were diagnosed with blood-positive fungal cultures and identified as having L. elongisporus by PCR amplification or PCR amplification, followed by DNA sequencing (PCR sequencing) of the internal transcribed spacer (ITS) region and/or the D1/D2 domains of ribosomal R DNA or by matrix-assisted laser desorption ionization timeof-flight (MALDI-TOF).In our present patient, diagnosis with blood-positive fungal culture and identification of L. elongisporus was achieved through mNGS.
In this case report and review of the literature, only 2 patients were not clearly identified for the existence of underlying disease/risk factors (15,16).The underlying disease/risk factors were as follows: surgery and trauma (6 patients), cardiovascular and cerebrovascular diseases (6 patients), central venous catheterization (3 patients), injection drugs (2 patients), cancer (2 patients), diabetes mellitus (2 patients), broad-spectrum antibiotics (2 patients), and mental illness (2 patients).Each case of these diseases included high-dose dexamethasone, nasogastric tube, stoma and total parenteral nutrition, mechanical ventilation, aorto-esophageal fistula, end-stage renal disease, rheumatoid arthritis, premature neonates, hyaline membrane disease, chronic lymphopenia, hepatitis C virus and chronic obstructive pulmonary disease.These results suggested that, in addition to the previously discussed immunosuppression and intravenous devices underlying diseases, surgery and trauma, cardiovascular and cerebrovascular diseases, cancer, diabetes mellitus and broad-spectrum antibiotics, vigilance against L. elongisporus infection is also needed.
There is currently a lack of consensus and guidelines for recognized treatment of L. elongisporus infection.The crude mortality rate in this study for invasive L. elongisporus infection was 27.27% (6/22).The minimum inhibitory concentration values for 23 strains of L. elongisporus indicated that they were sensitive to common antifungal drugs.It is worth noting that, when regular antifungal therapy for L. elongisporus endocarditis is not effective, surgical intervention should be considered.In addition, it is essential to incorporate prospective future projects or study subjects pertaining to the proper identification of L. elongisporus, as well as risk factors, surveillance, epidemiological investigations and standardized diagnosis and treatment.
In this study, we utilized the mNGS method for pathogen detection in a particular sample.The high-quality reads obtained were assembled using the default pipeline of SPAdes software, version 3.15.2(1).We evaluated the genome assembly using QUAST software, version 5.0.2, and assessed the completeness of the genome assembly using BUSCO software, version 5.4.3 (2), against the fungi_odb10 (or saccharomycetes_odb10) database from OrthoDB (OrthoDB | E Zdobnov lab).We performed genome annotations using Funannotate software, version 1.8.13 (3).
To determine the closest match to the sample, we aligned the assembled scaffolds to the abvf (archaea, bacteria, viruses, fungi) database from the NT and ITS databases.In both cases, the results showed that the sample was closest to Lodderomyces elongisporus.To confirm our conclusion, we calculated the values of ANI (average nucleotide identity) and dDDH (digital DNA-DNA hybridization) among the species, which also indicated a close match to Lodderomyces elongisporus.
Furthermore, we used reads to map the abvf database directly with blastn (17) and Kraken (18)-Bracken (19).The classification results revealed that most of the reads were labeled Lodderomyces elongisporus, providing additional support for our conclusion.
We conducted a phylogenetic analysis to determine the relationships of the sample L. elongisporus in the Saccharomycetales order.The results provided important insights into the evolutionary relationships of L. elongisporus within the Saccharomycetales order and its position among other fungi.This information could be useful for understanding the genetic diversity and evolutionary history of fungal species, as well as for developing effective classification schemes and identifying potential targets for drug development.Survive * * P23 was the present case.* An extremely low birth weight (ELBW, 710 g), preterm (23 weeks of gestational age), cannulated with an umbilical arterial catheter and an umbilical venous catheter using a vacuum-assisted closure technique and started on total parenteral nutrition.# Chronic lymphopenia: lymphocyte count <0.3 × 109 g/L.HMD, hyaline membrane disease; HCV, hepatitis C virus; LE, Lodderomyces elongisporus; TPN, total parenteral nutrition; CNS, Central nervous system; ESRD, End-stage renal disease; COPD, chronic obstructive pulmonary disease.PCR amplification or PCR amplification followed by DNA sequencing (PCR sequencing) of the internal transcribed spacer (ITS) region and/or the D1/D2 domains of ribosomal R DNA or by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI TOF-MS).FLU, fluconazole; VOR, voriconazole; ITR, itraconazole; POS, posaconazole; AFG, anidulafungin; MFG, micafungin; CAS, caspofungin; AMB, amphotericin B; 5-FC, 5-fluorocytosine; ASP, aspofungin.

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The use of mNGS technology in clinical pathogen detection demonstrated high accuracy in detecting fungi.This technology allowed for a comprehensive and precise analysis of the sample's genome assembly, as well as identification of the closest match to Lodderomyces elongisporus.The findings suggested that mNGS has great potential as a valuable tool for clinical pathogen detection and research.
The results of our study suggested that mNGS could be a valuable tool for clinical pathogen detection and open new avenues for research in the field.Moreover, mNGS could provide important data on pathogen epidemiology and evolution for infectious disease surveillance.By analyzing large amounts of pathogen genomic data, a deeper understanding of the pathogen's evolutionary history, pattern, and interaction with the host can be achieved.This information is significant for developing effective prevention control strategies and researching new vaccines.

FIGURE( A )
FIGURE (A) Transthoracic echocardiography showed a vegetation lesion adherent to the patient's anterior mitral leaflet, which was concerning for infectiousendocarditis. (B-D) Culture and microscopic characteristics of L. elongisporusfrom the patient.Blood culture growth of cream-colored colonies on Sabouraud dextrose agar (B), chocolate agar (C), and Columbia CNA bloodagar plates (D) after h of incubation at • C. (E) Microscopy of the blood smear revealed ellipsoidal to elongated blastoconidia of L. elongisporus (Gram stain, × , ). (F) The patient's disease development process.