Detection of viable antibiotic-resistant/sensitiveAcinetobacter baumanniiin indoor air by propidium monoazide quantitative polymerase chain reaction

Abstract

Acinetobacter baumannii represents a significant cause of nosocomial infections. Therefore, we combined real-time quantitative polymerase chain reaction (PCR) with the propidium monoazide (PMA-qPCR) to assess the feasibility of detecting viable, airborne A. baumannii. The biological collection efficiencies of three samplers for collecting airborne A. baumannii were evaluated by PMA-qPCR in a chamber study. After sampling, the effects of storage in collection fluid on A. baumannii were evaluated. The results showed that the culturable ratio of A. baumannii measured using the culture method was significantly correlated with the viable ratio measured using PMA-qPCR, but was not significantly correlated with the qPCR results. It was indicated that the AGI-30 impinger and the BioSampler were much more effective than the Nuclepore filter sampler for collecting airborne A. baumannii. The storage temperature was critical for aerosol samples, as the loss of viable A. baumannii was minimized when the PMA-bound DNA was stored at -20°C or if the collected cells were stored at 4°C and subsequently processed by PMA-qPCR within 1 month. The PMA-qPCR method was also to distinguish between colistin-sensitive and colistin-resistant A. baumannii, and no colistin-sensitive A. baumannii was detected by PMA-qPCR upon treatment of the BioSampler collection medium with 2 μg/ml colistin for 5 min.

Altered susceptibility to the bactericidal effect of photocatalytic oxidation by TiO2 is related to colistin resistance development in Acinetobacter baumannii

“…The minimum inhibitory concentration (MIC) of colistin for the colistin-resistant mutants was 32 μg/ml. This colistin-resistant strain was also evaluated in our previous study (Tseng et al 2015).…”

Non-selective Separation of Bacterial Cells with Magnetic Nanoparticles Facilitated by Varying Surface Charge

“…Currently, qPCR is still the simplest and most widely used method for the quantification of target genes of interest. According to its definition, the method detection limit (MDL) for the qPCR detection of bioaerosols, is determined by the instrumental detection limit (IDL) of qPCR, the DNA extraction efficiency, and the sampling volume (Hospodsky et al, 2010; Tseng et al, 2015). Among these three factors, the IDL (typically 0.5∼1 copy/μL for a SYBR Green dye based detection) and extraction efficiency (approximately 15.5∼43.3% for most commercial kits; Mumy and Findlay, 2004) are the most difficult to improve further.…”

Altered susceptibility to air sampling stress by filtration is related to colistin resistance development inAcinetobacter baumannii

“…The test chamber was the same as that in our previous study (Figure ). Our test chamber had a volume of 24.3 L. Airborne A. baumannii was generated by a Collison three‐jet nebulizer (BGI Collison Nebulizer; BGI Inc., Waltham, MA, USA), and the flow rate was regulated by a mass flow controller (Sierra Instruments, Monterey, CA, USA) to reach 3 L/min with an inlet pressure of 25 psi.…”

“…In this study, we conducted qPCRs to determine the total number of A. baumannii cells. The DNA extraction and qPCR processes and their related conditions were as they were in our previous study . DNA extraction was performed using a Chemagic DNA Bacteria Kit (PerkinElmer, Santa Clara, CA, USA) according to the manufacturer's instructions.…”

“…In our current study, the filtration stress on bacteria was evaluated because filtration can cause mechanical and nonmechanical stresses such as desiccation and elution . These sampling stresses resulted from filtration causing greater stress to airborne bacteria than other stresses, including impaction and impingement . If the susceptibilities of antibiotic‐resistant bacteria to this sampling stress are different from those of antibiotic‐sensitive bacteria, the validity of the air sampling strategy for quantifying these harmful bacteria may need to be verified.…”