High-throughput 16S rDNA sequencing assisting in the detection of bacterial pathogen candidates: a fatal case of necrotizing fasciitis in a child

Qu, Dong; Qiao, Dongfang; Klintschar, Michael; Qu, Zhi; Xia, Yunlong

doi:10.1007/s00414-020-02421-x

Cited by 3 publications

(1 citation statement)

References 22 publications

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“…However, 16S rRNA Sanger sequencing does not discriminate between mixed flora in polybacterial samples (Stavnsbjerg et al, 2017). 16S rRNA metagenomic sequencing enables the identification of all bacteria present in the sample, irrespective of their relative amounts and vitality, by increasing the number of reads per sample (Kommedal et al, 2014;Qu et al, 2021).…”

Section: Introductionmentioning

confidence: 99%

Identification of microbial agents in culture-negative brain abscess samples by 16S/18S rRNA gene PCR and sequencing

et al. 2022

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Despite clinical suspicion of an infection, brain abscess samples are often culture-negative in routine microbiological testing. Direct PCR of such samples enables the identification of microbes that may be fastidious, non-viable, or unculturable. Brain abscess samples (n = 217) from neurosurgical patients were subjected to broad range 16S rRNA gene PCR and sequencing for bacteria. All these samples and seven formalin-fixed paraffin-embedded tissue (FFPE) samples were subjected to species-specific 18S rRNA PCR for neurotropic free-living amoeba that harbour pathogenic bacteria. The concordance between smear and/or culture and PCR was 69%. One-third of the samples were smear-and culture-negative for bacterial agents. However, 88% of these culture-negative samples showed the presence of bacterial 16S rRNA by PCR. Sanger sequencing of 27 selected samples showed anaerobic/fastidious gram negative bacteria (GNB, 38%), facultative Streptococci (35%), and aerobic GNB (27%). Targeted metagenomics sequencing of three samples showed multiple bacterial species, including anaerobic and non-culturable bacteria. One FFPE tissue revealed the presence of Acanthamoeba 18S rRNA. None of the frozen brain abscess samples tested was positive for 18S rRNA of Acanthamoeba or Balamuthia mandrillaris. The microbial 16/18S rRNA PCR and sequencing outperformed culture in detecting anaerobes, facultative Streptococci and FLA in brain abscess samples. Genetic analyses of 16S/18S sequences, either through Sanger or metagenomic sequencing, will be an essential diagnostic technology to be included for diagnosing culture-negative brain abscess samples. Characterizing the microbiome of culture-negative brain abscess samples by molecular methods could enable detection and/or treatment of the source of infection.

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Section: Introductionmentioning

confidence: 99%

Identification of microbial agents in culture-negative brain abscess samples by 16S/18S rRNA gene PCR and sequencing

et al. 2022

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Post-mortem diagnosis of malaria in decomposed and embalmed body

Kusior,

Piniewska-Róg,

Wojtas

et al. 2024

Forensic Sci Med Pathol

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The diagnosis of malaria during the autopsy of a decomposed corpse may prove challenging. Macroscopic changes are non-specific and may include, among others, cerebral oedema, pulmonary oedema, hepatosplenomegaly and, on occasion, the presence of petechiae. The most effective diagnostic tools for malaria are the examination of blood smears and the use of rapid immunochromatographic tests. As a result of the progressive putrefaction of the corpse and blood hemolysis, classical tests are no longer viable. Consequently, the sole remaining option is the utilisation of real-time reaction (RT-PCR) to ascertain the presence of plasmodium DNA in specific organs. This study concerns the diagnosis of a fatal form of cerebral malaria in a 23-year-old Caucasian male who had travelled to Africa. The autopsy was conducted at a local hospital, after which the body was embalmed and stored in cold storage for a period of 8.5 months. Subsequently, the corpse was transported to Poland, where a further forensic autopsy was conducted. A significant challenge was to confirm the presence of malaria in a corpse that had been embalmed several months prior to the investigation. Samples were obtained from internal organs for genetic analysis to determine the presence of parasite DNA. An RT-PCR test was conducted on genetic material obtained from the brain, heart, lungs, kidney, liver, and spleen. The presence of Plasmodium falciparum genetic material was identified in samples obtained from the brain, lungs, kidney, liver, and spleen. These findings substantiated the post-mortem diagnosis of a severe form of cerebral malaria, which was the underlying cause of death.

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NGS plus bacterial culture: A more accurate method for diagnosing forensic-related nosocomial infections

Xie

Zhao

et al. 2021

Legal Medicine

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High-throughput 16S rDNA sequencing assisting in the detection of bacterial pathogen candidates: a fatal case of necrotizing fasciitis in a child

Cited by 3 publications

References 22 publications

Identification of microbial agents in culture-negative brain abscess samples by 16S/18S rRNA gene PCR and sequencing

Identification of microbial agents in culture-negative brain abscess samples by 16S/18S rRNA gene PCR and sequencing

Post-mortem diagnosis of malaria in decomposed and embalmed body

NGS plus bacterial culture: A more accurate method for diagnosing forensic-related nosocomial infections

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