Impact of GeneXpert MTB/RIF on patients and tuberculosis programs in a low-burden setting. a hypothetical trial.

RATIONALE: Guidelines recommend routine nucleic-acid amplification testing in patients with presumed tuberculosis (TB), but these tests have not been widely adopted. GeneXpert MTB/RIF (Xpert), a novel, semiautomated TB nucleic-acid amplification test, has renewed interest in this technology, but data from low-burden countries are limited. OBJECTIVES: We sought to estimate Xpert's potential clinical and public health impact on empiric treatment, contact investigation, and housing in patients undergoing TB evaluation. METHODS: We performed a prospective, cross-sectional study with 2-month follow-up comparing Xpert with standard strategies for evaluating outpatients for active pulmonary TB at the San Francisco Department of Public Health TB Clinic between May 2010 and June 2011. We calculated the diagnostic accuracy of standard algorithms for initial empiric TB treatment, contact investigation, and housing in reference to three Mycobacterium tuberculosis sputum cultures, as compared with that of a single sputum Xpert test. We estimated the incremental diagnostic value of Xpert, and the hypothetical reductions in unnecessary treatment, contact investigation, and housing if Xpert were adopted to guide management decisions. MEASUREMENTS AND MAIN RESULTS: A total of 156 patients underwent Xpert testing. Fifty-nine (38%) received empiric TB treatment. Thirteen (8%) had culture-positive TB. Xpert-guided management would have hypothetically decreased overtreatment by 94%, eliminating a median of 44 overtreatment days (interquartile range, 43-47) per patient and 2,169 total overtreatment days (95% confidence interval, 1,938-2,400) annually, without reducing early detection of TB patients. We projected similar benefits for contact investigation and housing. CONCLUSIONS: Xpert could greatly reduce the frequency and impact of unnecessary empiric treatment, contact investigation, and housing, providing substantial patient and programmatic benefits if used in management decisions.

Molecular epidemiology of tuberculosis in foreign-born persons living in San Francisco.

RATIONALE: In San Francisco, 70% of the tuberculosis cases occur among foreign-born persons, mainly from China, the Philippines, and Mexico. We postulate that there are differences in the characteristics and risk factors for tuberculosis among these populations. OBJECTIVES: To determine the clinical, epidemiological and microbiological characteristics of tuberculosis caused by recent infection and rapid evolution in the major groups of foreign-born and the U.S.-born populations. METHODS: We analyzed data from a 20-year prospective community-based study of the molecular epidemiology of tuberculosis in San Francisco. We included all culture-positive tuberculosis cases in the City during the study period. MEASUREMENTS AND MAIN RESULTS: We calculated and compared incidence rates, clinical and microbiological characteristics, and risk factors for being a secondary case between the various foreign-born and U.S.-born tuberculosis populations. Between 1991 and 2010, there were 4,058 new cases of tuberculosis, of which 1,226 (30%) were U.S.-born and 2,832 (70%) were foreign-born. A total of 3,278 (81%) were culture positive, of which 2,419 (74%) had complete data for analysis. The incidence rate, including the incidence rate of tuberculosis due to recent infection and rapid evolution, decreased significantly in the U.S.-born and the major foreign-born populations. The clinical and microbiological characteristics and the risk factors for tuberculosis due to recent infection differed among the groups. CONCLUSIONS: There are differences in the characteristics and the risk factors for tuberculosis due to recent transmission among the major foreign-born and U.S.-born populations in San Francisco. These differences should be considered for the design of targeted tuberculosis control interventions.

Factors associated with mortality in patients with drug-susceptible pulmonary tuberculosis.

BACKGROUND: Tuberculosis is a leading cause of death worldwide, yet the determinants of death are not well understood. We sought to determine risk factors for mortality during treatment of drug-susceptible pulmonary tuberculosis under program settings. METHODS: Retrospective chart review of patients with drug-susceptible tuberculosis reported to the San Francisco Tuberculosis Control Program from 1990-2001. RESULTS: Of 565 patients meeting eligibility criteria, 37 (6.6%) died during the study period. Of 37 deaths, 12 (32.4%) had tuberculosis listed as a contributing factor. In multivariate analysis controlling for follow-up time, four characteristics were independently associated with mortality: HIV co-infection (HR = 2.57, p = 0.02), older age at tuberculosis diagnosis (HR = 1.52 per 10 years, p = 0.001); initial sputum smear positive for acid fast bacilli (HR = 3.07, p = 0.004); and experiencing an interruption in tuberculosis therapy (HR = 3.15, p = 0.002). The association between treatment interruption and risk of death was due to non-adherence during the intensive phase of treatment (HR = 3.20, p = 0.001). The median duration of treatment interruption did not differ significantly in either intensive or continuation phases between those who died and survived (23 versus 18 days, and 37 versus 29 days, respectively). No deaths were directly attributed to adverse drug reactions. CONCLUSIONS: In addition to advanced age, HIV and characteristics of advanced tuberculosis, experiencing an interruption in anti-tuberculosis therapy, primarily due to non-adherence, was also independently associated with increased risk of death. Improving adherence early during treatment for tuberculosis may both improve tuberculosis outcomes as well as decrease mortality.

Evaluation of quantitative IFN-gamma response for risk stratification of active tuberculosis suspects.

RATIONALE: The contribution of interferon-gamma release assays (IGRAs) to appropriate risk stratification of active tuberculosis suspects has not been studied. OBJECTIVES: To determine whether the addition of quantitative IGRA results to a prediction model incorporating clinical criteria improves risk stratification of smear-negative-tuberculosis suspects. METHODS: Clinical data from tuberculosis suspects evaluated by the San Francisco Department of Public Health Tuberculosis Control Clinic from March 2005 to February 2008 were reviewed. We excluded tuberculosis suspects who were acid fast-bacilli smear-positive, HIV-infected, or under 10 years of age. We developed a clinical prediction model for culture-positive disease and examined the benefit of adding quantitative interferon (IFN)-gamma results measured by QuantiFERON-TB Gold (Cellestis, Carnegie, Australia). MEASUREMENTS AND MAIN RESULTS: Of 660 patients meeting eligibility criteria, 65 (10%) had culture-proven tuberculosis. The odds of active tuberculosis increased by 7% (95% confidence interval [CI], 3-11%) for each doubling of IFN-gamma level. The addition of quantitative IFN-gamma results to objective clinical data significantly improved model performance (c-statistic 0.71 vs. 0.78; P < 0.001) and correctly reclassified 32% of tuberculosis suspects (95% CI,11-52%; P < 0.001) into higher-risk or lower-risk categories. However, quantitative IFN-gamma results did not significantly improve appropriate risk reclassification beyond that provided by clinician assessment of risk (4%; 95% CI, -7 to +22%; P = 0.14). CONCLUSIONS: Higher quantitative IFN-gamma results were associated with active tuberculosis, and added clinical value to a prediction model incorporating conventional risk factors. Although this benefit may be attenuated within highly experienced centers, the predictive accuracy of quantitative IFN-gamma levels should be evaluated in other settings.

First performance report of QIAreach™ Anti-SARS-CoV-2 Total Test, an innovative nanoparticle fluorescence digital detection platform.

In 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a global pandemic. Disease diagnosis, appropriate clinical management and infection control are all important factors in controlling the spread of SARS-CoV-2. The QIAreach™ Anti-SARS-CoV-2 Total Test (Anti-CoV2) is a rapid, qualitative serological test, using proprietary nanoparticle fluorescence technology to detect total antibody (IgA, IgM, and IgG) against SARS-CoV-2. Here we report the results of the US Food and Drug Administration (FDA) clinical agreement study. Thirty positive plasma or serum samples were taken from consenting individuals with polymerase chain reaction (PCR)-confirmed SARS-CoV-2 infection ≥14 days from symptom onset. Seventy-five samples from before the believed circulation of SARS-CoV-2 (November 1, 2019) were used to assess specificity. Positive percent agreement (PPA) and negative percent agreement (NPA) were calculated along with the corresponding exact two-sided 95 % confidence intervals (CI) using an FDA Emergency Use Authorized PCR test as the reference method. Anti-CoV2 was shown to have 100 % sensitivity (PPA; 95 % CI 88.4-100 %) and 100 % specificity (NPA; 95 % CI 95.2-100 %). Against 157 pre-pandemic samples, no cross-reactivity was observed with seasonal coronaviruses or other respiratory pathogens tested. Additionally, no interference was observed when samples were spiked with: conjugated bilirubin 0.4 mg/ml; unconjugated bilirubin 0.4 mg/ml; hemoglobin 5 mg/ml; prednisolone 0.12 mg/ml; triglycerides 15 mg/ml. In conclusion, Anti-CoV2 provides accurate qualitative detection of total antibodies against SARS-CoV-2.

Clinical characteristics and treatment outcomes of patients with isoniazid-monoresistant tuberculosis.

BACKGROUND: Risk factors and treatment outcomes under program conditions for isoniazid (INH)-monoresistant tuberculosis have not been well described. METHODS: Medical charts were retrospectively reviewed for all cases of culture-confirmed, INH-monoresistant tuberculosis ( n = 137) reported to the San Francisco Department of Public Health Tuberculosis Control Section from October 1992 through October 2005, and those cases were compared with a time-matched sample of drug-susceptible tuberculosis cases (n = 274) RESULTS: In multivariate analysis, only a history of treatment for latent tuberculosis (odds ratio [OR], 3.1; 95% confidence interval [CI], 1.5-6.4; P = .003) or for active tuberculosis (OR, 2.7; 95% CI, 1.4-5.0; P = .002) were significantly associated with INH-monoresistant tuberculosis. Of the 119 patients who completed treatment, 49 (41%) completed a 6-month treatment regimen. Treatment was extended to 7-12 months for 53 (45%) of the patients and to >12 months for 17 (14%). Treatment was most commonly extended because pyrazinamide was not given for the recommended 6-month duration (35 patients [29%]). Despite variation in treatment regimens, the combined end point of treatment failure or relapse was uncommon among patients with INH-monoresistant tuberculosis and was not significantly different for patients with drug-susceptible tuberculosis (1.7% vs. 2.2%; P = .73). CONCLUSIONS: A history of treatment for latent or active tuberculosis was associated with subsequent INH monoresistance. Treatment outcomes for patients with INH-monoresistant tuberculosis were excellent and were no different from those for patients with drug-susceptible tuberculosis. However, new, short-course regimens are needed because a small proportion of patients completed the 6-month treatment regimen recommended by the American Thoracic Society, Centers for Disease Control and Prevention, and Infectious Diseases Society of America, primarily because of pyrazinamide intolerance.

Reaching the limits of tuberculosis prevention among foreign-born individuals: a tuberculosis-control program perspective.

Analysis of whether assiduous implementation of American Thoracic Society/Centers for Disease Control and Prevention/Infectious Diseases Society of America guidelines for targeted testing and treatment of latent tuberculosis infection could have prevented any of 223 cases of active tuberculosis in foreign-born persons in San Francisco during the period 2002-2003. We report that 62% of these cases were not preventable and conclude that a further reduction in the incidence of tuberculosis among foreign-born persons will be modest without modification of current guidelines.

Quantitative impact of human immunodeficiency virus infection on tuberculosis dynamics.

RATIONALE: Human immunodeficiency virus (HIV) infection has a major but unquantified impact on the risk of tuberculosis. OBJECTIVES: To quantify the impact of HIV infection on the number of tuberculosis cases in San Francisco. METHODS: We studied all patients reported with tuberculosis in San Francisco from 1991 to 2002. The initial isolates of Mycobacterium tuberculosis were genotyped using IS6110 restriction fragment-length polymorphism genotyping as the primary method, and clustered cases (identical genotype patterns) were identified. MEASUREMENTS AND MAIN RESULTS: We determined the case number, case rate, and the fraction of tuberculosis attributable to HIV infection. Of 2,991 reported tuberculosis cases, 2,193 (73.3%) had a genotype pattern of M. tuberculosis available. Genotypic clusters with at least one HIV-positive person were larger, lasted longer, and had a shorter time between successive cases relative to clusters with only HIV-uninfected persons (P < 0.00005, P = 0.0009, P = 0.018, respectively). Overall, 13.7% of the tuberculosis cases were attributable to HIV infection and an estimated 405 excess tuberculosis cases occurred. CONCLUSIONS: During a period encompassing the resurgence and decline of tuberculosis in San Francisco, a substantial number of the tuberculosis cases were attributable to HIV infection. Coinfection with HIV amplified the local tuberculosis epidemic.

Too little too late: Waiting for TB to come.

There is a new paradigm that preventing tuberculosis (TB) and addressing the reservoir of latent TB infection in combination with curing all TB cases is essential to accelerate the decline of TB rates and ending TB by 2050. However, complacency and incremental change eludes radical policy transformation needed to meet global targets. This essay explores current attitudes, policy disparities between high and lower burden settings, and what changes are needed to remove the obstacles to progress.

Low sensitivity of a whole-blood interferon-gamma release assay for detection of active tuberculosis.

The sensitivity of an interferon-gamma assay (Quantiferon-TB Gold; Cellestis) was evaluated for the detection of tuberculosis among 242 persons with suspected tuberculosis in San Francisco, California. Thirty-seven subjects had culture-confirmed tuberculosis. Excluding 1 indeterminate result, 23 (64%; 95% confidence interval, 48%-78%) of 36 subjects had positive results using the QuantiFERON-TB Gold assay. The 64% sensitivity suggests that the QuantiFERON-TB Gold assay should not be used alone to exclude active tuberculosis.

Tuberculosis in healthcare workers: a molecular epidemiologic study in San Francisco.

OBJECTIVE: Healthcare workers (HCWs) are at risk of becoming infected with Mycobacterium tuberculosis through occupational exposure. To identify HCWs who became infected and developed tuberculosis as a result of their work, we studied the molecular epidemiology of tuberculosis in HCWs. DESIGN: Eleven-year prospective cohort molecular epidemiology study. SETTING: City and County of San Francisco, California. PATIENTS: All persons reported with tuberculosis between 1993 and 2003. HCWs were identified from the San Francisco Tuberculosis Control Section's database, and mycobacterial isolates from culture-positive subjects were analyzed by IS6110-based genotyping. RESULTS: Of 2510 cases of tuberculosis reported during the study period, 31 (1.2%) occurred in HCWs: the median age of the HCWs was 37 years, and 11 (35%) were male. HCWs were more likely than non-HCWs to be younger (P=.0036), born in the United States (P=.0004), and female (P=.0003) and to not be homeless (P=.010). The rate of tuberculosis among HCWs remained constant during the study period, despite a significant decrease in the overall case rate in San Francisco. Work-related transmission was documented in at least 10 (32%) of 31 HCWs, including 4 of 8 HCWs whose isolates were part of genotypically determined clusters. Only 1 of 7 cases of tuberculosis in HCWs after 1999 was documented as being work-related. CONCLUSIONS: Although most cases of tuberculosis in HCWs, as in non-HCWs, developed as a result of endogenous reactivation of latent infection, at least half of clustered cases of tuberculosis in HCWs were related to work. The number of work-related cases of tuberculosis in HCWs decreased during the study period.

Feasibility, acceptability, and cost of tuberculosis testing by whole-blood interferon-gamma assay.

BACKGROUND: The whole-blood interferon-gamma release assay (IGRA) is recommended in some settings as an alternative to the tuberculin skin test (TST). Outcomes from field implementation of the IGRA for routine tuberculosis (TB) testing have not been reported. We evaluated feasibility, acceptability, and costs after 1.5 years of IGRA use in San Francisco under routine program conditions. METHODS: Patients seen at six community clinics serving homeless, immigrant, or injection-drug user (IDU) populations were routinely offered IGRA (Quantiferon-TB). Per guidelines, we excluded patients who were <17 years old, HIV-infected, immunocompromised, or pregnant. We reviewed medical records for IGRA results and completion of medical evaluation for TB, and at two clinics reviewed TB screening logs for instances of IGRA refusal or phlebotomy failure. RESULTS: Between November 1, 2003 and February 28, 2005, 4143 persons were evaluated by IGRA. 225(5%) specimens were not tested, and 89 (2%) were IGRA-indeterminate. Positive or negative IGRA results were available for 3829 (92%). Of 819 patients with positive IGRA results, 524 (64%) completed diagnostic evaluation within 30 days of their IGRA test date. Among 503 patients eligible for IGRA testing at two clinics, phlebotomy was refused by 33 (7%) and failed in 40 (8%). Including phlebotomy, laboratory, and personnel costs, IGRA use cost $33.67 per patient tested. CONCLUSION: IGRA implementation in a routine TB control program setting was feasible and acceptable among homeless, IDU, and immigrant patients in San Francisco, with results more frequently available than the historically described performance of TST. Laboratory-based diagnosis and surveillance for M. tuberculosis infection is now possible.

Interferon-Gamma Release Assays and Pediatric Public Health Tuberculosis Screening: The San Francisco Program Experience 2005 to 2008.

BACKGROUND: Interferon-gamma release assay utilization in pediatric tuberculosis (TB) screening is limited by a paucity of longitudinal experience, particularly in low-TB burden populations. METHODS: We conducted a retrospective review of QuantiFERON (QFT)-TB Gold results in San Francisco children from 2005 to 2008. Concordance with the tuberculin skin test (TST) was analyzed for a subset of children. Progression to active disease was determined through San Francisco and California TB registry matches. RESULTS: Of 1092 children <15 years of age, 853 (78%) were foreign-born, and 136 (12%) were exposed to active TB cases (contacts). QuantiFERON tests were positive in 72 of 1092 (7%) children; 15 of 136 (11%) recent contacts; 53 of 807 (7%) foreign-born noncontacts; and 4 of 149 (3%) US-born noncontacts. QuantiFERON-negative/TST-positive discordance was seen more often in foreign-born/bacille Calmette-Guerin (BCG)-vaccinated children <5 years of age (52 of 56, 93%) compared to those ≥ 5 years of age (90 of 123, 73%; P = .003). Foreign-born, BCG-vaccinated children were more than twice as likely to have a discordant (79%) result as US-born, non-BCG-vaccinated children (37%; P < .0001). During 5587 person-years of follow-up of untreated children, including 146 TST-positive/QFT-negative children, no cases of active TB were identified, consistent with a negative predictive value of 100%. CONCLUSIONS: Our experience supports the use of QFT to evaluate latent TB infection in children, particularly young BCG-vaccinated children. The proportion of QFT-positive results correlated with risk of exposure, and none of the untreated QFT-negative children developed TB. The low QFT-positive rate highlights the need for more selective testing based on current epidemiology and TB exposure risk.

Treatment of multidrug-resistant tuberculosis in San Francisco: an outpatient-based approach.

BACKGROUND: Treatment of patients with multidrug-resistant tuberculosis requires prolonged therapy, often involving long hospital stays. Despite intensive and costly therapy, cure rates are relatively low. METHODS: We reviewed the outcomes for all patients with multidrug-resistant tuberculosis treated in San Francisco, California, during 1982-2000 and identified billing charges for patients treated during 1995-2000. Mycobacterium tuberculosis isolates were genotyped by IS6110-based restriction fragment-length polymorphism analysis. RESULTS: Forty-eight cases were identified with resistance to a median of 3 drugs (range, 2-9 drugs). The median age of the patients was 49.5 years (range, 22-78 years); 36 (75%) of 48 patients were foreign born, 11 (23%) were human immunodeficiency virus (HIV) seropositive, and 45 (94%) had pulmonary tuberculosis. Thirty-two (97%) of the 33 HIV-seronegative patients were cured, with only 1 relapse occurring 5 years after treatment. All 11 HIV-seropositive patients died during observation. Twenty-one patients (44%) required hospitalization, with a median duration of stay of 14 days (range, 3-74 days). The estimated inpatient and outpatient aggregate cost for the 11 patients treated after 1994 was $519,928, with a median cost of $27,752 per patient. No secondary cases of multidrug-resistant tuberculosis were identified through population-based genotyping. CONCLUSIONS: Treatment of multidrug-resistant tuberculosis in HIV-seronegative patients largely on an outpatient basis was feasible and was associated with high cure rates and lower cost than in other published studies. Patients with underlying HIV infection had very poor outcomes.

Incidence of TB in inmates with latent TB infection: 5-year follow-up.

BACKGROUND: Inmates are a high-risk population for tuberculosis (TB) control efforts, including treatment for latent tuberculosis infection (LTBI). Completion of therapy after release has been poor. The goal of this study was to evaluate therapy completion and active disease over 5 years in a cohort of inmates. METHODS: The sample was from a completed randomized trial in 1998-1999 of education or incentive versus usual care to improve therapy completion after release from the San Francisco County Jail. Records from the jail, the County Tuberculosis Clinic, and the California TB Registry were used to measure therapy completion and development of active TB. Analyses were conducted in 2005. RESULTS: Of a total 527 inmates, 31.6% (n=176) completed therapy, of whom 59.7% (n=105) completed it in jail. Compared with the U.S.-born, foreign-born inmates residing in the United States for < or =5 years were less likely to complete the therapy (adjusted odds ratio [AOR]=0.49, 95% confidence interval [CI]=0.28-0.85), and those with more education were more likely to complete the therapy (AOR=1.06, 95% CI=1.01-1.12). Three subjects developed active TB in the 5 years of follow-up, resulting in an annual rate of 108 per 100,000. Compared with California rates, subjects were 59 times as likely to develop active TB (standardized morbidity ratio of 59.2, 95% CI=11.2-145.1). None had completed therapy, none were new immigrants, and two were known to be HIV-positive at diagnosis. CONCLUSIONS: Completion of therapy for LTBI is a challenge, but the active TB seen in this jail cohort emphasizes the importance of continued efforts to address TB risk in this population.

Short-course rifampin and pyrazinamide compared with isoniazid for latent tuberculosis infection: a multicenter clinical trial.

BACKGROUND: Rifampin and pyrazinamide are recommended for treatment of latent tuberculosis infection in adults without HIV infection, but reports of severe hepatotoxicity have raised concerns about its safety. Clinical trials have not compared this treatment with isoniazid in adults without HIV infection. OBJECTIVE: To compare the safety and tolerance of a 2-month regimen of rifampin and pyrazinamide with that of a 6-month regimen of isoniazid for treatment of latent tuberculosis infection. DESIGN: Multicenter, prospective, open-label trial. SETTING: Three urban public health tuberculosis clinics in the United States. PATIENTS: 589 adults with latent tuberculosis infection who met U.S. criteria for treatment. INTERVENTION: Patients were assigned in alternate weeks to receive rifampin and pyrazinamide daily for 2 months (n = 307) or isoniazid daily for 6 months (n = 282). MEASUREMENTS: Primary end points were hepatotoxicity, other adverse events, and percentage of patients who completed treatment. RESULTS: Sixteen of 207 (7.7%) patients assigned to rifampin and pyrazinamide developed grade 3 or 4 hepatotoxicity compared with 2 of 204 (1%) patients assigned to isoniazid (odds ratio, 8.46 [95% CI, 1.9 to 76.5]; P = 0.001). The rifampin plus pyrazinamide regimen was more likely than the isoniazid regimen to be discontinued because of hepatotoxicity (odds ratio, 5.19; P = 0.033). The overall percentage of nonhepatotoxic adverse events was 20% in the rifampin-pyrazinamide group and 16% in the isoniazid group. The proportion of patients who completed the study treatment was 61% and 57%, respectively. CONCLUSIONS: A 2-month regimen of rifampin and pyrazinamide was associated with an increased risk for grade 3 or 4 hepatotoxicity compared with a 6-month regimen of isoniazid. Liver enzymes should be measured routinely during treatment to screen for liver injury and prevent progression to severe toxicity.

Short-course rifampin and pyrazinamide compared with isoniazid for latent tuberculosis infection: a cost-effectiveness analysis based on a multicenter clinical trial.

Two months of treatment with rifampin-pyrazinamide (RZ) and 9 months of treatment with isoniazid are both recommended for treatment of latent tuberculosis infection in adults without human immunodeficiency virus infection, but the relative cost-effectiveness of these 2 treatments is unknown. We used a Markov model to conduct a cost-effectiveness analysis to assess the impact on life expectancy and costs based on the results of a recent clinical trial that compared the rates of adverse events and completion of the 2 treatment regimens. Compared with no treatment, both regimens increased life expectancy by 1.2 years, but RZ cost 273 dollars more per patient. Sensitivity analyses showed that, assuming equal efficacy between the 2 regimens, there was no threshold completion rate for RZ at which the 2 treatments would be of equal net cost. Under most circumstances, treatment of latent tuberculosis infection with isoniazid is cost-saving than treatment with RZ.

Use of whole genome sequencing to determine the microevolution of Mycobacterium tuberculosis during an outbreak.

RATIONALE: Current tools available to study the molecular epidemiology of tuberculosis do not provide information about the directionality and sequence of transmission for tuberculosis cases occurring over a short period of time, such as during an outbreak. Recently, whole genome sequencing has been used to study molecular epidemiology of Mycobacterium tuberculosis over short time periods. OBJECTIVE: To describe the microevolution of M. tuberculosis during an outbreak caused by one drug-susceptible strain. METHOD AND MEASUREMENTS: We included 9 patients with tuberculosis diagnosed during a period of 22 months, from a population-based study of the molecular epidemiology in San Francisco. Whole genome sequencing was performed using Illumina's sequencing by synthesis technology. A custom program written in Python was used to determine single nucleotide polymorphisms which were confirmed by PCR product Sanger sequencing. MAIN RESULTS: We obtained an average of 95.7% (94.1-96.9%) coverage for each isolate and an average fold read depth of 73 (1 to 250). We found 7 single nucleotide polymorphisms among the 9 isolates. The single nucleotide polymorphisms data confirmed all except one known epidemiological link. The outbreak strain resulted in 5 bacterial variants originating from the index case A1 with 0-2 mutations per transmission event that resulted in a secondary case. CONCLUSIONS: Whole genome sequencing analysis from a recent outbreak of tuberculosis enabled us to identify microevolutionary events observable during transmission, to determine 0-2 single nucleotide polymorphisms per transmission event that resulted in a secondary case, and to identify new epidemiologic links in the chain of transmission.