The genotyping services process is summarized in the . The number of eligible isolates by year is shown in the . As of March 2004, isolates for 2,600 (96.8%) of 2,685 patients with a diagnosis of culture-positive TB from January 1, 2001, to December 31, 2003, were submitted. Of 85 patient isolates not submitted to the public health laboratory, 78.8% were processed at commercial laboratories, mostly outside of New York City. For patient isolates with incomplete genotyping (n = 163), RFLP could not be performed because of inadequate growth or overgrowth with other mycobacteria or fungi. Spoligotype and RFLP results were available for 2,437 (93.7%) of the 2,600 isolates submitted (90.7% of all culture-positive patients). The median days from specimen collection to reporting of spoligotype decreased from 84 days in 2001 to 53 days in 2003, and the reporting time for RFLP patterns decreased from 127 days in 2001 to 78 days in 2003 (). Fifty-nine (2.4%) isolates were false-positive cultures; 37% of investigations of these false-positive cultures were initiated through matching genotyping results or a spoligotype suggestive of contamination with a laboratory TB strain. Outside requests initiated 8.5% of investigations; 24.0% were initiated from the single positive culture list, and 30.5% by request from staff within the Bureau of Tuberculosis Control. The median time to complete investigations of false-positive cultures decreased from 178 days in 2001 to 85 days in 2003. In 2003, patients with a false-positive culture were treated unnecessarily for a median of 7 days (range 0–145). This median number of days is considerably lower than that seen before universal genotyping; in 1999, patients identified by retrospective surveillance (i.e., the single-positive culture list) as having false-positive cultures completed a median of 7 months of treatment.
Genotyping of isolates, New York City, 2001–2003*
Median days for submission and turnaround time by laboratory, New York City, 2001–2003. PHL, public health laboratory; RFLP, restriction fragment length polymorphism.
Among 2,378 isolates with a complete genotype (true-positive cultures), 565 spoligotype patterns and 1,600 RFLP patterns were identified; 2,009 (84.5%) of 2,378 patient isolates clustered in 196 spoligotype clusters, and 1,002 (42.1%) of 2,378 patient isolates in 224 RFLP clusters. Eight hundred thirty-seven (35.2%) of the 2,378 isolates had RFLP and spoligotype patterns that matched >1 other isolate pattern; these isolate patterns were grouped into 205 genotype clusters ranging in size from 2 to 81 cases (mean 4 cases/cluster; median 2 cases/cluster). The percentage of clustered patient isolates remained stable during the 3-year period (χ2 for trend p = 0.3652). While most patient isolates had 9–13 copies of IS6110 (median 11 copies, range 1–23), strains with a lower copy number (<6 bands) were more likely to be clustered. From 2001 to 2003, two large outbreaks occurred that involved strains of 1 and 3 IS6110 copies. After these strains were excluded, the percentage clustered remained higher for patterns with lower numbers of IS6110.
A total of 278 (33.2%) of 837 clustered cases had epidemiologic links identified; of these, 105 (37.8%) had links established through traditional contact investigations. Genotype cluster investigations established links for the remaining 62%: 15% of the links were definite, 11% probable, and 36% possible. For 66.4% of clustered cases (556 cases), no epidemiologic links were identified. Time to completion of cluster investigations decreased from a median of 176 days in 2001 to 37 days in 2003. The delay in completing investigations at the beginning of the project was mostly due to staff vacancies. Cluster investigations uncovered 57 additional links among cases with matching genotypes and 17 additional sites of transmission. Links established through genotype cluster investigations led to 4 expanded contact investigations in congregate settings (2 in homeless shelters, 1 in a single-room-occupancy hotel, and 1 in a local grocery store). These investigations identified additional infected contacts and 4 additional TB patients at a homeless shelter. These sites are now monitored closely for additional patient isolates with these genotypes. Transmission between TB patients was ruled out in >5 site investigations because the genotypes were unrelated, avoiding more extensive case-finding efforts that are needed once transmission is seen.
Four quality assurance exercises were performed from 2001 to 2003 on 216 isolates. The result was 94.4% concordance for spoligotyping and 93.5% for RFLP. Of retyped spoligotype patterns that did not exactly match the original patterns, 50% differed by ±1 spacer, 8% differed in multiple successive spacers, and 42% differed for other reasons. Among retyped RFLP patterns, 57% differed from the original patterns because of the existence or absence of >1 bands, 36% differed because of pattern shifts, and 7% differed for other reasons.