PMID- 36748452
OWN - NLM
STAT- MEDLINE
DCOM- 20230209
LR  - 20230209
IS  - 1473-5644 (Electronic)
IS  - 0022-2615 (Linking)
VI  - 71
IP  - 12
DP  - 2022 Dec
TI  - Diagnostic yield of broad-range 16s rRNA gene PCR varies by sample type and is 
      improved by the addition of qPCR panels targeting the most common causative 
      organisms.
LID - 10.1099/jmm.0.001633 [doi]
AB  - Introduction. Molecular techniques are used in the clinical microbiology 
      laboratory to support culture-based diagnosis of infection and are particularly 
      useful for detecting difficult to culture bacteria or following empirical 
      antimicrobial treatment.Hypothesis/Gap Statement. Broad-range 16S rRNA PCR is a 
      valuable tool that detects a wide range of bacterial species. Diagnostic yield is 
      low for some sample types but can be improved with the addition of qPCR panels 
      targeting common bacterial pathogens.Aim. To evaluate the performance of a 
      broad-range 16S rRNA gene PCR and the additional diagnostic yield of targeted 
      qPCR applied to specimens according to a local testing algorithm.Methodology. In 
      total, 6130 primary clinical samples were collected as part of standard clinical 
      practice from patients with suspected infection during a 17 month period. 
      Overall, 5497 samples were tested by broad-range 16S rRNA gene PCR and a panel of 
      targeted real-time qPCR assays were performed on selected samples according to a 
      local testing algorithm. An additional 633 samples were tested by real-time qPCR 
      only. The 16S rRNA gene PCR was performed using two assays targeting different 
      regions of the 16S rRNA gene. Laboratory developed qPCR assays for seven common 
      bacterial pathogens were also performed. Data was extracted retrospectively from 
      Epic Beaker Laboratory Information Management System (LIMS).Results. Broad-range 
      16S rRNA gene PCR improves diagnostic yield in culture-negative samples and 
      detects a large range of bacterial species. Streptococcus spp., Staphylococcus 
      spp. and the Enterobacteriaceae family are detected the most frequently in 
      samples with a single causative organism, but mixed samples frequently contained 
      anaerobic species. The highest diagnostic yield was obtained from abscess, pus 
      and empyema samples; 44.9 % were positive by 16S and 61 % were positive by the 
      combined 16S and targeted qPCR testing algorithm. Samples with a particularly low 
      diagnostic yield were blood, with 3.3 % of samples positive by 16S and CSF with 
      4.8 % of samples positive by 16S. The increased diagnostic yield of adding 
      targeted qPCR is largest (~threefold) in these two sample types.Conclusion. 
      Broad-range PCR is a powerful technique that can detect a very large range of 
      bacterial pathogens but has limited diagnostic sensitivity. The data in this 
      report supports a testing strategy that combines broad-range and targeted 
      bacterial PCR assays for maximizing diagnosis of infection in culture-negative 
      specimens. This is particularly justified for blood and CSF samples. Alternative 
      approaches, such as metagenomic sequencing, are needed to provide the breadth of 
      broad-range PCR and the sensitivity of targeted qPCR panels.
FAU - Harris, Kathryn A
AU  - Harris KA
AD  - Microbiology, Virology and Infection Prevention & Control, Great Ormond Street 
      Hospital NHS Foundation Trust, Great Ormond Street, London, WC1N 3JH, UK.
AD  - Queen Mary University of London, Mile End Road, London, E1 4NS, UK.
FAU - Brown, Julianne R
AU  - Brown JR
AD  - Microbiology, Virology and Infection Prevention & Control, Great Ormond Street 
      Hospital NHS Foundation Trust, Great Ormond Street, London, WC1N 3JH, UK.
LA  - eng
PT  - Journal Article
PL  - England
TA  - J Med Microbiol
JT  - Journal of medical microbiology
JID - 0224131
RN  - 0 (DNA, Bacterial)
RN  - 0 (RNA, Ribosomal, 16S)
SB  - IM
MH  - Humans
MH  - *Bacteria/genetics
MH  - DNA, Bacterial/genetics/analysis
MH  - Genes, rRNA
MH  - Polymerase Chain Reaction/methods
MH  - Retrospective Studies
MH  - RNA, Ribosomal, 16S/genetics
MH  - *Bacterial Infections/diagnosis
OTO - NOTNLM
OT  - 16SPCR
OT  - bacterial PCR
OT  - broad-range PCR
OT  - clinical microbiology
OT  - molecular diagnostics
EDAT- 2023/02/08 06:00
MHDA- 2023/02/09 06:00
CRDT- 2023/02/07 04:54
PHST- 2023/02/07 04:54 [entrez]
PHST- 2023/02/08 06:00 [pubmed]
PHST- 2023/02/09 06:00 [medline]
AID - 10.1099/jmm.0.001633 [doi]
PST - ppublish
SO  - J Med Microbiol. 2022 Dec;71(12). doi: 10.1099/jmm.0.001633.