Abstract:
BACKGROUND
In South Africa and other high prevalence countries, transmission is a significant contributor
to rising rates of multidrug resistant tuberculosis (MDR-TB). Thus, there is a need to
develop an early detection system for transmission clusters suitable for high burden settings.
We have evaluated the discriminatory power and clustering concordance of a novel
and simple genotyping approach, combining spoligotyping with pncA sequencing (SpoNC),
against two well-established methods: IS6110-RFLP and 24-loci MIRU-VNTR.
METHODS
A total of 216 MDR-TB isolates collected from January to June 2010 from the NHLS Central
TB referral laboratory in Braamfontein, Johannesburg, representing a diversity of strains
from South Africa, were included. The isolates were submitted for genotyping, pncA
sequencing and analysis to the Centre for Tuberculosis in South Africa and the Public
Health Research Institute Tuberculosis Center at Rutgers University in the United States.
Clustering rates, Hunter-Gaston Discriminatory Indexes (HGI) and Wallace coefficients
were compared between the methods.
RESULTS
Overall clustering rates were high by both IS6110-RFLP (52.8%) and MIRU-VNTR (45.8%),
indicative of on-going transmission. Both 24-loci MIRU-VNTR and IS6110-RFLP had similar HGI (0.972 and 0.973, respectively), with close numbers of unique profiles (87 vs. 70), clustered
isolates (129 vs. 146), and cluster sizes (2 to 26 vs. 2 to 25 isolates). Spoligotyping
alone was the least discriminatory (80.1% clustering, HGI 0.903), with 28 unique types.
However, the discriminatory power of spoligotyping was improved when combined with pncA sequencing using the SpoNC approach (61.8% clustering, HGI 0.958). A high proportion
of MDR-TB isolates had mutations in pncA (68%, n = 145), and pncA mutations were
significantly associated with clustering (p = 0.007 and p = 0.0013 by 24-loci MIRU-VNTR
and IS6110-RFLP, respectively), suggesting high rates of resistance to pyrazinamide
among all MDR-TB cases and particularly among clustered cases.
CONCLUSION
We conclude that SpoNC provides good discrimination for MDR-TB surveillance and early
identification of outbreaks in South Africa, with 24-loci MIRU-VNTR applied for pncA wildtype
strains as needed.
