ABSTRACT
Studies have shown an association between bacterial load and virulence; however, not much is known about the diversity in this phenotypic characteristic of Mycobacterium tuberculosis complex (MTBC). This study was therefore aimed to determine the differences in bacterial load of the three most prevalent MTBC genotypes (L4, L5, and L6) in West Africa at the time of diagnosis. A total of 170 paired fresh sputum samples were collected; one part in guanidinium thiocyanate (GTC) was used for RNA extraction and tuberculosis molecular bacterial load assay (TB-MBLA), and the other part without GTC was confirmed for TB positivity using GeneXpert MTB/RIF, smear microscopy grading, and culture on Löwenstein-Jensen media slants. The 170 sputum samples comprised 155 new cases, three follow-up cases, and 12 TB negative sputum samples. The time-to-culture positivity (TTP) and degree of culture positivity (DCP) were recorded. All 122 isolates obtained were spoligotyped for lineage (L) classification, but spoligotypes were obtained from 120 isolates. Of the typed isolates, 70.0, 10.8, 10.8, 4.2, 2.5, 0.8, and 0.8% were lineages 4, 5, 6, 2, 3, 1, and Mycobacterium bovis, respectively. Further analysis of the three most prevalent lineages showed significantly shorter TTP and higher DCP by L4 compared to L5 and L6, respectively: TTP 20.8, vs. 26.5, and 28.2 days; p-value = 0.005 and DCP 1.27, vs. 0.81 and 0.29, p < 0.001. The average TB-MBLA measured bacterial load of L4 was 3.82 Log10eCFU/ml which was not significantly different from 3.81 and 3.80 Log10eCFU/ml of L5 and L6, respectively, p = 0.84. Degrees of smear microscopy L4 = 1.20, L5 = 1.20, and L6 = 0.92 and GeneXpert Cq values L4 = 17.08, L5 = 18.37, and L6 = 17.59 showed no significant difference between the lineages, p = 0.72 and p = 0.48, respectively. Retrospective analysis of a larger sample confirmed the difference in TTP, p < 0.001. In conclusion, the observed shorter TTP and high DCP of L4 could signify high growth rate in culture that is independent of total bacterial load at diagnosis.
ABSTRACT
BACKGROUND: Early access to diagnosis is crucial for effective management of any disease including tuberculosis (TB). We investigated the barriers and opportunities to maximise uptake and utilisation of molecular diagnostics in routine healthcare settings. METHODS: Using the implementation of WHO approved TB diagnostics, Xpert Mycobacterium tuberculosis/rifampicin (MTB/RIF) and Line Probe Assay (LPA) as a benchmark, we evaluated the barriers and how they could be unlocked to maximise uptake and utilisation of molecular diagnostics. RESULTS: Health officers representing 190 districts/counties participated in the survey across Kenya, Tanzania and Uganda. The survey findings were corroborated by 145 healthcare facility (HCF) audits and 11 policy-maker engagement workshops. Xpert MTB/RIF coverage was 66%, falling behind microscopy and clinical diagnosis by 33% and 1%, respectively. Stratified by HCF type, Xpert MTB/RIF implementation was 56%, 96% and 95% at district, regional and national referral hospital levels. LPA coverage was 4%, 3% below culture across the three countries. Out of 111 HCFs with Xpert MTB/RIF, 37 (33%) used it to full capacity, performing ≥8 tests per day of which 51% of these were level five (zonal consultant and national referral) HCFs. Likewise, 75% of LPA was available at level five HCFs. Underutilisation of Xpert MTB/RIF and LPA was mainly attributed to inadequate-utilities, 26% and human resource, 22%. Underfinancing was the main reason underlying failure to acquire molecular diagnostics. Second to underfinancing was lack of awareness with 33% healthcare administrators and 49% practitioners were unaware of LPA as TB diagnostic. Creation of a national health tax and decentralising its management was proposed by policy-makers as a booster of domestic financing needed to increase access to diagnostics. CONCLUSION: Our findings suggest higher uptake and utilisation of molecular diagnostics at tertiary level HCFs contrary to the WHO recommendation. Country-led solutions are crucial for unlocking barriers to increase access to diagnostics.