The use of Kudoh method for culture of Mycobacterium tuberculosis and Mycobacterium africanum in The Gambia.

BACKGROUND: Mycobacterium tuberculosis culturing remains the gold standard for laboratory diagnosis of tuberculosis. Tuberculosis remains a great public health problem in developing countries like The Gambia, as most of the methods currently used for bacterial isolation are either time-consuming or costly. OBJECTIVE: To evaluate the Kudoh swab method in a West African setting in Gambia, with a particular focus on the method's performance when culturing Mycobacterium africanum West Africa 2 (MAF2) isolates. METHOD: 75 sputum samples were collected in the Greater Banjul Area and decontaminated in parallel with both the standard N-acetyl-L-Cysteine-NaOH (NALC-NaOH) and the Kudoh swab method in the TB diagnostics laboratory in the Medical Research Council Unit The Gambia between 30th December 2017 and 25th February 2018. These samples were subsequently cultured on standard Löwenstein-Jensen and Modified Ogawa media respectively and incubated at 37°C for mycobacterial growth. Spoligotyping was done to determine if the decontamination and culture methods compared could equally detect Mycobacterium tuberculosis, Mycobacterium africanum West Africa 1 and Mycobacterium africanum West Africa 2. RESULT: Among the 50 smear positives, 35 (70%) were culture-positive with Kudoh and 32 (64%) were culture positive with NALC-NaOH, whilst 7(28%) of the 25 smear negative samples were culture positive with both methods (Table 2). There was no significant difference in recovery between both methods (McNemar's test, p-value = 0.7003), suggesting that the overall positivity rate between the two methods is comparable. There were no differences in time-to-positivity or contamination rate between the methods. However, Kudoh yielded positive cultures that were negative on LJ and vice versa. All findings were irrespective of mycobacterial lineages. CONCLUSION: The Kudoh method has comparable sensitivity to the NALC-NaOH method for detecting Mycobacterium tuberculosis complex isolates. It is easy to perform and could be an add on option for mycobacterial culture in the field in The Gambia, since it requires less biosafety equipment.

Comparative genomics shows differences in the electron transport and carbon metabolic pathways of Mycobacterium africanum relative to Mycobacterium tuberculosis and suggests an adaptation to low oxygen tension.

The geographically restricted Mycobacterium africanum lineages (MAF) are primarily found in West Africa, where they account for a significant proportion of tuberculosis. Despite this phenomenon, little is known about the co-evolution of these ancient lineages with West Africans. MAF and M. tuberculosis sensu stricto lineages (MTB) differ in their clinical, in vitro and in vivo characteristics for reasons not fully understood. Therefore, we compared genomes of 289 MAF and 205 MTB clinical isolates from the 6 main human-adapted M. tuberculosis complex lineages, for mutations in their Electron Transport Chain and Central Carbon Metabolic pathway in order to explain these metabolic differences. Furthermore, we determined, in silico, whether each mutation could affect the function of genes encoding enzymes in these pathways. We found more mutations with the potential to affect enzymes in these pathways in MAF lineages compared to MTB lineages. We also found that similar mutations occurred in these pathways between MAF and some MTB lineages. Generally, our findings show further differences between MAF and MTB lineages that may have contributed to the MAF clinical and growth phenotype and indicate potential adaptation of MAF lineages to a distinct ecological niche, which we suggest includes areas characterized by low oxygen tension.

Performance comparison of a pair of Lowenstein-Jensen media supplemented with pyruvate or glycerol, and the combination of both supplements in a single Lowenstein-Jensen medium for the growth support of the Mycobacterium Tuberculosis complex.

OBJECTIVE/BACKGROUND: To evaluate the performance of Lowenstein-Jensen medium (LJ) supplemented with pyruvate and glycerol (LJPG), compared with LJ supplemented with pyruvate (LJP) or glycerol (LJG) for the support of mycobacterial growth. METHOD: This study used 100 Ziehl-Neelsen-confirmed positive mycobacterium growth indicator tube 960 culture samples that were obtained from clinical samples during routine diagnosis. All cultures were inoculated in parallel on LJG/LJP and on LJGP, which were incubated and read weekly for the evidence of growth. The mycobacterial recovery rate, contamination rate, and time to detection were compared. RESULT: The recovery rate for LJG/LJP and for LJPG was 90% (90 samples) and 88% (88 samples), respectively (kappa p-value, 0.9). There was no significant difference in the contamination rate, which was 8% (8 samples) for LJG/LJP and 9% (9 samples) for LJPG. Mycobacterial growth was faster in LJPG (1.6weeks) than in LJG/LJP (2weeks). CONCLUSION: A single LJPG slope was not significantly different, compared with the usual pair of LJG or LJP slopes. This is a promising new culturing approach that could be used in Mycobacterium africanum-endemic in West African countries. It significantly reduces labor time and consumable costs and more quickly detects the M. tuberculosis complex.