Real-time PCR versus MALDI-TOF MS and culture-based techniques for diagnosis of bloodstream and pyogenic infections in humans and animals.
J Appl Microbiol
; 130(5): 1630-1644, 2021 May.
Article
em En
| MEDLINE
| ID: mdl-33073430
ABSTRACT
AIMS:
This study was applied to evaluate the usefulness of a high-throughput sample preparation protocol prior to the application of quantitative real-time PCR (qPCR) for the early diagnosis of bloodstream and pyogenic infections in humans and animals compared to matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and classical culture. METHODS ANDRESULTS:
Saponin-mediated selective host cell lysis combined with DNase-1 was applied for processing of whole blood and pus clinical samples collected from suspected cases of septicaemia and pyogenic infections in humans and animals. The pre-PCR processing strategy enabled the recovery of microbial cells with no changes in their colony forming units immediately after the addition of saponin. DNase-1 was efficient for removing the DNAs from the host cells as well as dead cells with damaged cell membranes. The metagenomic qPCR and MALDI-TOF MS could identify the bacterial community of sepsis at species level with a concordance of 97·37% unlike the conventional culture. According to qPCR results, Staphylococcus aureus (24·24%) was predominated in animal pyogenic infections, whereas Klebsiella pneumonia (31·81%) was commonly detected in neonatal sepsis.CONCLUSIONS:
Saponin combined with DNase-1 allowed the efficient recovery of microbial DNA from blood and pus samples in sepsis using qPCR assay. SIGNIFICANCE AND IMPACT OF THE STUDY Metagenomic qPCR could identify a broad range of bacteria directly from blood and pus with more sensitivity, higher discriminatory power and shorter turnaround time than those using MALDI-TOF MS and conventional culture. This might allow a timely administration of a prompt treatment.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Bactérias
/
Sepse
/
Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
/
Reação em Cadeia da Polimerase em Tempo Real
Tipo de estudo:
Diagnostic_studies
/
Guideline
/
Prognostic_studies
/
Screening_studies
Limite:
Animals
/
Humans
Idioma:
En
Revista:
J Appl Microbiol
Assunto da revista:
MICROBIOLOGIA
Ano de publicação:
2021
Tipo de documento:
Article
País de afiliação:
Egito