Abstract
Aim:
To countermeasure the global spread of β-lactamases, we developed a rapid molecular test for the highly variable OXA-β-lactamases that allows minimizing the time to effective treatment.
Methods:
OXA-mRNA was specifically enriched from total RNA using group-specific biotinylated DNA probes and streptavidin-coated magnetic beads. Phylogenetic OXA groups were distinguished by PCR product size.
Results:
This mRNA fishing method is highly sensitive, yielding specific results from as little as 1 ng total RNA. It enables discrimination of OXA-extended substrate spectrum β-lactamases and carbapenemases and the semi-quantitative detection of highly expressed ISAba1-controlled variants.
Conclusion:
Targeting mRNA with specific probes on magnetic beads will allow for adaptation to automated systems, such as point-of-care diagnostics.
Keywords:
Acinetobacter baumannii; CHDL; Dynabeads; ESBL; carbapenem resistance; multiplex; sequence homology.
MeSH terms
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Acinetobacter Infections / diagnosis*
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Acinetobacter Infections / microbiology
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Acinetobacter baumannii / classification
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Acinetobacter baumannii / enzymology*
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Acinetobacter baumannii / genetics
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Acinetobacter baumannii / isolation & purification
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Bacterial Proteins / genetics*
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Bacterial Proteins / metabolism
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Enterobacteriaceae / classification
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Enterobacteriaceae / enzymology*
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Enterobacteriaceae / genetics
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Enterobacteriaceae / isolation & purification
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Enterobacteriaceae Infections / diagnosis*
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Enterobacteriaceae Infections / microbiology
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Humans
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Magnetics / instrumentation
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Magnetics / methods*
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Phylogeny
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RNA, Bacterial / genetics
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RNA, Bacterial / isolation & purification*
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RNA, Bacterial / metabolism
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RNA, Messenger / genetics
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RNA, Messenger / isolation & purification*
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RNA, Messenger / metabolism
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beta-Lactamases / genetics*
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beta-Lactamases / metabolism
Substances
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Bacterial Proteins
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RNA, Bacterial
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RNA, Messenger
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beta-Lactamases