Abstract:

Objective To report laboratory diagnosis of a rare primary amebic meningoencephalitis case. Methods The clinical data and the cerebrospinal fluid (CPF) sample was collected from the patient, and the CPF sample was smeared and stained with Wright Giemsa for microscopy examination. The genomic DNA was extracted from the CPF for amplification of internal transcribed spacer 1 (ITS1)-5.8S-ITS2 sequence using Naegleria genus- specific and N. fowleri species-specific primers by PCR, and then the amplicon sequenced. The sequence obtained was aligned with sequences in GenBank using BLAST, with which phylogenetic tree was constructed by neighbour-joining method using MEGA5. The CPF sample was sent to Hangzhou Kingdomain Medical Laboratory Co., LTD for DNA-pathogenic microorganism metagenomic detection. Results The patient, a 42-year-old man from Xiangshan, Ningbo, had been bedridden with severe burns covering whole body for 20 years. On July 21, 2022, he was sent to the First People’s Hospital of Xiangshan, Ningbo, Zhejiang Province, for fever, chills, headache, sore throat, nausea and vomiting. The physical examination showed that the weight of the patient was 45 kg, the body temperature was 40.5 ℃, with obnubilation, eyes on the turn, conjunctival congestion. Both eye pupils were 3 mm in diameter, equal in size and circle. The patient also had jaw clenching, neck stiffness, stertorous breathing, limb convulsion, muscle spasm, urinary incontinence, skin and mucous membrane flushing and other clinical symptoms. The head and chest CT showed no obvious abnormalities. The CPF pressure of lumbar puncture was 39 cmH₂O (1 cmH₂O = 0.098 kPa). Amoeba trophozoites with rapid and continuous amoeba-like movement could be seen on the smear of CPF under microscope. The amoeba trophozoites were also seen after Wright Giemsa staining. The genus- and species-specific gene fragments, which were 183 bp and 311 bp, respectively, were amplified. A total of 21 534 sequences of N. fowleri with a relative abundance of 99.8% were found by DNA-pathogenic microorganism metagenomic testing. No sequence of other pathogenic microorganisms was found. The sequence alignment showed that the amplified ITS1-5.8S-ITS2 sequence was 99% identical to the N. fowleri Na 420c strain sequence (Accession no. AJ132028) recorded in GenBank, and clustered with Amoeba Negri on the phylogenetic tree. The case was confirmed to be infected with N. fowleri. The patient was given meropenem for anti-bacteria, metronidazole for anti-amoeba, fluconazole for anti-fungal treatment, and 20% mannitol dehydration to reduce intracranial pressure. However, due to the rapid deterioration, which lead to respiratory and heart failure, the patient died on July 23, 2022. Conclusion The patient was diagnosed with N. fowleri infection based on the clinical symptoms, pathogenic examination of CPF and molecular biological identification.

Key words: Naegleria fowleri, Primary meningoencephalitis, Infection, Diagnosis