Viability Study of Nontuberculous Mycobacteria Commonly Causing Skin/Soft Tissue Infections Using the Copan ESwab Collection and Transport System

Abstract

Objective

The aim of this study was to evaluate the performance of Copan ESwab™ collection and transport system (Copan Diagnostics Inc.) for the viability of nontuberculous mycobacteria commonly causing skin and soft tissue infections.

Method

Viability tests were performed in triplicate using ESwabs and the roll plate method as per the CLSI M40-A2 document. ESwabs were prepared in duplicate with one set held at room temperature (RT) (20oC to 25oC) and the other held at refrigerator temperature (2°C to 8%C) for 0, 24 and 48 hours (hrs) prior to processing. Five selected mycobacterial isolates consisting of Mycobacterium abscessus, M. fortuitum, M. chelonae, M. marium, and M. haemophilum were planted onto Columbia blood agar plates with 5% sheep blood and incubated at 30°C in O2 for 5 to7 days. These organisms were then used to prepare three independent 0.5 McFarland suspensions. From this working suspension four 1:10 serial dilutions (107, 106, 105 and 104) were prepared in sterile saline. One hundred microliters of each organism suspension were transferred into wells of a round bottom microtiter plate. Each swab was then immersed into the organism suspension and allowed to absorb for 10 seconds before inserting into its corresponding ESwab tube. For each 0-hr set, the ESwab was removed from the transport device after 15 minutes. The swabs were discarded and the tubes vortexed again for 5 seconds. One hundred microliters of the suspension were added to Middlebrook 7H10 agar (MA), streaked and incubated at 30oC in O2 for 5 to 7 days. M.haemophilum was streaked on blood agar plates. Colonies were counted and averaged for three swabs for each time point and dilution. Average colony counts at 24 and 48 hrs were compared to the 0 hr inoculated swabs.

Results

All five organisms, were able to produce countable colonies at dilutions of 106, 105 and/or 104 and within ~300 to 500 CFU/mL at 24 and 48 hrs, respectively, compared to 0 hr counts.

Conclusion

Results suggest that the Copan ESwab collection and transport system is able to maintain and recover nontuberculous mycobacteria for up to 48 hrs at room and refrigeration temperatures. Routine swab collections are considered less than optimal for the recovery of mycobacterial isolates; however, this study indicates that mycobacterial organisms can, with high efficiency, be recovered from ESwabs. Further study is required to determine the utility of the ESwab in recovering nontuberculous mycobacterium from clinically infected patients.

Readers found these studies helpful