Evaluation of the Xpert Carba-R assay for quantifying carbapenemase-producing bacterial load in stool samples

by Jie Yin Chua, Ze Qin Lim, Song Qi Dennis Loy, Vanessa Koh, Natascha May Thevasagayam, Xiaowei Huan, Kyaw Zaw Linn, Kalisvar Marimuthu, Oon Tek Ng

Background

The spread of Carbapenemase-producing Organisms (CPO) remains a major threat globally. Within clinical settings, the existing method of determining gene load involves traditional culture to determine bacterial load and polymerase-chain-reaction-based Xpert Carba-R Assay to determine carbapenemase gene type. However, there is a need for a fast and accurate method of quantifying CPO colonisation to study the risk of persistent CPO carriage.

Objective

This study evaluated the accuracy of Xpert Carba-R Ct value in estimating carbapenamase producing bacterial loads in stool samples.

Methods

Stool samples were obtained from an ongoing study investigating the household transmission of CPO in Singapore. Stool samples lacking carbapenemase producing organisms were spiked with organism carrying a single carbapenemase gene (bla_KPC, bla_NDM, bla_VIM, bla_{OXA-48(-like)} or bla_IMP-1) and serially diluted before being subjected to Xpert Carba-R assay and traditional culture. Standard curves with regression lines showing correlation between C_t values and plate counts were generated. The standard curves were validated with stool samples collected from patients.

Results

The limit of detection of bla_NDM, bla_KPC, and bla_OXA-48 was approximately 10³ cfu/mL, while that of bla_IMP-1 and bla_VIM was approximately 10⁴ cfu/mL. Validation of the bla_NDM and bla_OXA-48 curves revealed average delta values of 0.56 log(cfu/mL) (95% CI 0.24–0.88) and 0.80 log(cfu/mL) (95% CI 0.53–1.07), respectively.

Conclusions

Our validation data for stool positive for bla_NDM and bla_OXA-48-type suggests that bacterial loads can be estimated within a reasonable range of error.