Comparison of four agar media for the enumeration of the Mycobacterium abscessus complex

Background: Traditional culture of nontuberculous mycobacteria (NTMs) has involved egg-based formulations (Lowenstein-Jensen medium) or defined media (Middlebrook formulations), which have disadvantages of composition complexity, availability, and cost. This study quantitatively compared three non-selective, non-blood based basal agars with Columbia blood agar (CBA), to enumerate Mycobacterium abscessus complex organisms in pure culture.

Methods: M. abscessus subsp. massiliense, M. abscessus subsp. bolletii, and M. abscessus subsp. abscessus were employed. Inocula of each of these were counted on three basal agar media, including (i) standard plate count agar (SPCA), (ii) tryptone soya agar (TSA), and (iii) Mueller-Hinton agar (MHA) and compared to counts on CBA.

Results: All NTM isolates of all subspecies grew successfully on all four media examined. The growth was most profuse on SPCA, with a mean colony diameter of 3 mm, whereas the mean colony diameter on all other media was 1 mm. Statistically, there was no significant difference in counts when comparing CBA with SPCA or MHA (P > 0.05), whereas there was a statistically significant difference between CBA and TSA (P = 0.01). There was no statistically significant difference between SPCA and MHA (P = 0.53).

Conclusion: This study indicates that SPCA and MHA are equally effective as CBA, when enumerating of M. abscessus complex organisms. Employment of TSA gave significantly lower counts than CBA (P = 0.01) and therefore should not be employed when enumerating these organisms. SPCA yielded the most profuse growth of all media examined. In addition to these advantages, given that SPCA does (i) not require blood as a medium constituent, (ii) is simple to reconstitute, (iii) is relatively cheap, and (iv) is widely available commercially, this study endorses employment of SPCA for the nonselective culture of M. abscessus complex organisms, including enumeration.