Page 12 - Epidemiological studies on tuberculosis control and respiratory viruses

Chapter 2
Clonal complexes, defined as groups of at least two isolates differing in not more than 3/24 loci, were identified on a minimum-spanning tree with BioNumerics software (Applied Maths, Kortrijk, Belgium), using MIRU-VNTR data and the categorical distance, which scores the number of alleles shared or different over the 24 markers used.
Multiple imputation was used to account for 184 (5%) and 37 (1%) of 3,776 cases with missing data for the variables “time since immigration at TB diagnosis” and “gender,” respectively. All remaining variables were used to create five imputed data sets, and results are based on pooled statistics. Two different cluster definitions were used to investigate the interpretation of recent transmission; identical VNTR patterns and single-locus variants (SLVs). The theory behind this was that allowing SLVs to be clustered might involve genetically closely related strains that in fact share the same transmission chain. The analyses were performed separately for the Euro-American and non-Euro-American lineages and completed with SPSS 18.0 (SPSS, Chicago, IL) and statistical program R version 2.11.0.
A minimum-spanning tree was produced for all 3,776 isolates included in the analysis. In total 3,377 (89%) isolates were distributed over 83 clonal complexes (Fig. 1), whereas the remaining 399 (11%) isolates did not belong to any clonal complex. Within each complex, all the VNTR patterns represented the same lineage type, except the two largest complexes comprising 84% Haarlem strains and 67% T-specific strains.
Of the 3,776 isolates, 1,130 (30%) represented the non-Euro-American lineages (Table 1). We reasoned that recently arrived immigrant cases having nonclustered M. tuberculosis isolates most likely represent importation of foreign genotypes. Among the 504 nonclustered recent-immigrant cases, 239 (47%) were caused by isolates of the non-Euro-American lineages, of which EAI (45%), CAS (26%), and Beijing (18%) constituted the majority (Table 1). Cases caused by these non-Euro- American lineages originated from Asia (41%) and Africa (56%). In contrast, recent- immigrant nonclustered cases with Euro-American lineages had a higher diversity in geographical origin. Furthermore, 40 (8%) of the 504 recent-immigrant nonclustered cases originated from European countries, of which the majority (93%) were caused by the Euro-American lineages. Among the 564 native Dutch cases with nonclustered M. tuberculosis isolates, 459 (81%) had isolates of the Euro-American lineages, of which the majority were of the Haarlem (36%), T-specific (33%), and LAM (13%) lineages (Table 1).
22