The identification of NTM is important because positive microscopy, a main stay of TB diagnosis in developing countries, cannot differentiate M. tuberculosis complex from NTM infection, while pulmonary TB shares clinical signs with NTM disease, causing diagnostic and clinical dilemmas . NTM isolated from humans can be found in ecosystems shared between humans and animals, and in soil and natural open water sources, all of which play a key role as sources of human infections . Our study area is a rural agricultural setting, similar to other rural areas where studies have shown a high concentration of NTM in domesticated animals and the environment [4, 14]. The current study has isolated and characterized NTM from respiratory secretions in more than 90 children, with a majority exhibiting symptoms suggestive of pulmonary TB.
We have identified seven species comprising of 65/95 (68%) of the NTM isolates in children from a rural Ugandan community. The most prevalent of the identified isolates, M. fortuitum (63.5%), is a ubiquitous contaminant and colonizer that can be isolated from environmental sources such as potable water systems and soil . The other major species isolated were M. szulgai (14.3%), and M. gordonae (9.5%). In rural agro-pastoral Uganda, a recent study of 310 samples from soil, water and fecal matter from cattle and pigs isolated 48 NTM . The major species identified in that study were 15 (31.2%) M. avium complex, 12 (25%) M. fortuitum-peregrinum complex, five (10.4%) M. gordonae, and five (10.4%) M. nonchromogenicum. Although there were no human samples analyzed in that study, the two sets of results clearly support an environmental link to the infection in the children in the current study. Our findings are in further agreement with those in other studies which observed that children in rural agricultural communities might be at greater risk of exposure to environmental NTM than their urban counterparts . Furthermore, our study showed that there were more NTM isolated from adolescents (69/95) compared to infants (26/95), a finding in agreement with results from a similar study in South Africa , and this is thought to be due to increased environmental exposure in older children. While we isolated more M. fortuitum, the study in agro-pastoral ecosystems of Uganda showed that M. avium intraceullare complex was the predominant NTM isolated . In our study 33.7% (32/95) of the isolates could not be identified; we thus recommend that new probes for the correct identification of more NTM species be sought.
Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR was used to type all 63 identifiable NTM species in order to assess intra species clustering, hence commonality of sources of NTM infection in the study community. The nine M. szulgai, six M. gordonae and three M. intracellualre isolates did not cluster within the species, each isolate showing a distinct fingerprint pattern. Analysis of the 40 M. fortuitum isolates, on the other hand, showed unique fingerprint patterns in 35 isolates while the remaining five clustered into two, comprising of three and two isolates each. The cluster of three involved two infants and one adolescent sharing M. fortuitum 1 while the cluster of two involved adolescents with M. fortuitum 2 subspecies. However, the low fragment number of the isolates in the two clusters above is not sufficiently discriminative to evaluate these M. fortuitum clones. A similar study investigating M. avium mycobacterial lymphadenitis in children in The Netherlands using IS1245 Restriction Fragment length Polymorphisms (RFLP) analysis did not reveal any geographical clustering, with the 34 isolates in that study scattered over several clades . The environment is the most likely reservoir for these infections, as there is no evidence of human-to- human or animal-to-human transmission, and the only aerosol transmission of NTM infection recorded to date was from shower water .
The exact main clinical relevance of these mycobacteria is that colonization may induce non-specific immune response and thereby leading to false positive reactions in the Mantoux test . However, the infrequency of smear positivity relatively negates the concern for misdiagnosis of TB when using microscopy alone. Furthermore, if culture alone is used without genotypic identification, then M. fortuitum and other rapid growers are less likely to be confused for M. tuberculosis, thus the clinical implication may be less profound than previously stated.