First insights into the phylogenetic diversity of Mycobacterium tuberculosis in Kuwait and evaluation of REBA MTB-MDR assay for rapid detection of MDR-TB

Noura M Al-Mutairi 1, Suhail Ahmad 1, Eiman Mokaddas 1 2, Sahal Al-Hajoj 3

Affiliations

20 October 2022

-

doi: 10.1371/journal.pone.0276487

Abstract

Early detection of Mycobacterium tuberculosis (Mtb) in clinical specimens, its susceptibility to anti-TB drugs and disruption of infection transmission to new hosts are essential components for global tuberculosis (TB) control efforts. This study investigated major Mtb genotypes circulating in Kuwait and evaluated the performance of REBA MTB-MDR (REBA) test in comparison to GenoType MTBDRplus (gMTBDR+) assay for rapid detection of resistance of Mtb to isoniazid and rifampicin (MDR-TB). M. tuberculosis isolates (n = 256) originating predominantly from expatriate patients during a 6-month period were tested by spoligotyping and a dendrogram was created by UPGMA using MIRU-VNTRplus software. Phenotypic drug susceptibility testing (DST) was performed by MGIT 960 system. Genotypic DST for isoniazid and rifampicin was done by REBA and gMTBDR+ assays. Spoligotyping assigned 188 (73.4%) isolates to specific spoligotype international type (SIT) while 68 isolates exhibited orphan patterns. All major M. tuberculosis lineages were detected and EAI, CAS and Beijing families were predominant. Phylogenetic tree showed 131 patterns with 105 isolates exhibiting a unique pattern while 151 isolates clustered in 26 patterns. Fifteen isolates were resistant to one/more drugs. REBA and gMTBDR+ detected isoniazid resistance in 11/12 and 10/12 and rifampicin resistance in 4/5 and 4/5 resistant isolates, respectively. The diversity of SIT patterns are highly suggestive of infection of most expatriate patients with unique Mtb strains, likely acquired in their native countries before their arrival in Kuwait. Both, REBA and gMTBDR+ assays performed similarly for detection of resistance of Mtb to isoniazid and rifampicin for rapid detection of MDR-TB.

Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Fig 1. Pie chart showing the distribution of M. tuberculosis lineages (in percentages) prevalent in Kuwait on the basis of SITVIT2 WEB analysis. EAI, CAS and Beijing lineages were dominant.

Fig 1. Pie chart showing the distribution of M. tuberculosis lineages (in percentages) prevalent in Kuwait on the basis of SITVIT2 WEB analysis. EAI, CAS and Beijing lineages were dominant.

Fig 2. The dendrogram generated from the spoligotyping data using MIRU-VNTRplus database and unweighted pair group method with arithmetic mean (UPGMA) settings for 256 M. tuberculosis isolates from Kuwait is shown. The shared international type (SIT), M. tuberculosis family (Family), nationality of the TB patient yielding the isolate and the actual spoligotyping pattern for each isolate are shown in vertical columns. show full caption

Fig 2. The dendrogram generated from the spoligotyping data using MIRU-VNTRplus database and unweighted pair group method with arithmetic mean (UPGMA) settings for 256 M. tuberculosis isolates from Kuwait is shown. The shared international type (SIT), M. tuberculosis family (Family), nationality of the TB patient yielding the isolate and the actual spoligotyping pattern for each isolate are shown in vertical columns. show full caption

Fig 3. A minimum spanning tree (MST) showing evolutionary relationships based on spoligotypes in M. tuberculosis population (n = 256 isolates). The phylogenetic tree connects each genotype based on degree of changes required to go from one allele to another. The structure of the tree is represented by branches (continuous vs. dashed and dotted lines) and circles representing each individual pattern. The length of the branches represents the distance between patterns while the complexity of the lines (continuous, gray dashed and gray dotted) denotes the number of allele/spacer changes between two patterns: solid lines, 1 or 2 or 3 changes (thicker ones indicate a single change, while the thinner ones indicate 2 or 3 changes); gray dashed lines represent 4 changes; and gray dotted lines represent 5 or more changes. The size of the circle is proportional to the total number of isolates in the study, presenting unique isolates (smaller nodes) versus clustered isolates (bigger nodes). The color of the circles indicates the phylogenetic lineage to which the specific pattern belongs. The labels of nodes indicate SITs in the study. (N indicates orphan or new spoligotype pattern). show full caption

Fig 3. A minimum spanning tree (MST) showing evolutionary relationships based on spoligotypes in M. tuberculosis population (n = 256 isolates). The phylogenetic tree connects each genotype based on degree of changes required to go from one allele to another. The structure of the tree is represented by branches (continuous vs. dashed and dotted lines) and circles representing each individual pattern. The length of the branches represents the distance between patterns while the complexity of the lines (continuous, gray dashed and gray dotted) denotes the number of allele/spacer changes between two patterns: solid lines, 1 or 2 or 3 changes (thicker ones indicate a single change, while the thinner ones indicate 2 or 3 changes); gray dashed lines represent 4 changes; and gray dotted lines represent 5 or more changes. The size of the circle is proportional to the total number of isolates in the study, presenting unique isolates (smaller nodes) versus clustered isolates (bigger nodes). The color of the circles indicates the phylogenetic lineage to which the specific pattern belongs. The labels of nodes indicate SITs in the study. (N indicates orphan or new spoligotype pattern). show full caption

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