DISPATCHES

Molecular data were analyzed with SPOTCLUST (http://cgi2.cs.rpi.

edu/~bennek/SPOTCLUST.html).
Epidemiology of We performed 15-locus variable number tandem
repeat (VNTR) using a CEQ 8000 Genetic Analysis System
Mycobacterium (Beckman Coulter, Inc., Fullerton, CA, USA) (4). Cluster

tuberculosis,
analysis was performed by using Bionumerics software
(Applied Maths, St-Martens-Latem, Belgium). Strains

Buenos Aires,
lacking a unique pattern were subjected to further analysis
with an expanded set of VNTR loci (5).

Argentina In-house macroarrays were performed on MDR TB

strains (6) to identify mutations in the katG and the inhA
genes. Two regions of the rpoB gene of MDR TB strains
Ximena Gonzalo, Marta Ambroggi, were sequenced with the CEQ 8000 Genetic Analysis
Ezequiel Cordova, Tim Brown, Susana Poggi, System. Statistical analyses were conducted by using χ2
and Francis Drobniewski and Fisher exact tests.
To analyze the molecular epidemiology of
After we excluded duplicates, treatment follow-ups,
Mycobacterium tuberculosis strains at a hospital in Buenos and strains with susceptibility patterns other than MDR
Aires, Argentina, and mutations related to multidrug- TB or susceptibility to all drugs tested, 881 strains were
resistant and extensively drug-resistant tuberculosis, we susceptible to all drugs tested. Patients with a minimum
conducted a prospective case–control study. Our findings dataset (name, sex, date of birth or age, TB presentation
reinforce the value of incorporating already standardized [i.e., pulmonary or nonpulmonary], and at least 1 sign or
molecular methods for rapidly detecting resistance. symptom describing TB illness and treatment received)
were enrolled: 57 of 62 hospitalized patients with MDR
TB cultures (Table) and 100 fully susceptible unmatched
D uring the 1990s, an outbreak of multidrug-resistant
(MDR) tuberculosis (TB) in HIV-positive patients
occurred at the Muñiz Hospital in Buenos Aires, Argentina
inpatient controls, for a total of 157 patients. This
convenience sample (Figure 1) included only admitted
(1). Molecular analysis showed that a member of Haarlem2 patients because of the difficulty of obtaining clinical
family of Mycobacterium tuberculosis was responsible (1). information about outpatients.
We conducted a prospective case-control study during The most common spoligofamilies were T1 (18%),
June 1, 2006–April 30, 2007, at this 300-bed public hospital, Haarlem 1 (10%), 2 (16%), and 3 (11%), LAM 3 (10%),
which reports ≈40% of new TB cases in the city (2). Our and LAM 9 (13%). Initial 15-loci VNTR analysis showed
primary aims were to analyze the molecular epidemiology 73 strains with unique patterns. Further analysis using
of M. tuberculosis strains circulating at the hospital and 7-loci VNTR was performed on those that did not have
the mutations related to MDR TB and extensively drug- a unique pattern. Twenty-six isolates remained clustered
resistant TB. (Figure 2).
Of the 57 MDR TB strains, 43 had a mutation in the
The Study katG315 locus, and 6 had a mutation in the inhA region. No
The strains were isolated and tested for antimicrobial strain had mutations in both genes. In 8 strains, resistance
drug susceptibility according to the proportion method to isoniazid was not mediated by mutations in any of them.
(3) at the Muñiz Hospital Mycobacteria Laboratory. A Mutations in the rpoB region were detected by
proportion were tested for reserve drugs at the Health sequencing. The most frequent mutation was the S531L.
Protection Agency National Mycobacteria Reference Mutations at >1 site were rare. In 3 cases, we found 2 point
Unit Laboratory, London, UK. DNA was extracted mutations in the same codon. Only 1 MDR TB strain had no
from cultures at this UK laboratory; spoligotyping was mutation in the rpoB segment sequenced; it also had a katG
performed according to the manufacturer’s instructions and inhA wild type. Complete susceptibility profile for all
(Isogen Life Science, IJsselstein, the Netherlands); and 57 MDR TB strains is available in the online Appendix
Table (www.cdc.gov/EID/content/17/3/528-appT.htm).
In addition, we detected 5 extensively drug-resistant
Author affiliations: Hospital de Infecciosas Francisco Javier Muñiz,
TB strains. None were clustered by 22-loci VNTR typing.
Buenos Aires, Argentina (X. Gonzalo, M. Ambroggi, E. Cordova,
S. Poggi); and Clinical Sciences Research Center, London, UK (T.
Conclusions
Brown, F. Drobniewski)
Spoligotyping identified predominance of the Haarlem
DOI: 10.3201/eid1703100394 family among the MDR TB cases (family responsible for

528 Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 17, No. 3, March 2011
 Molecular Epidemiology of Tuberculosis

the 1990s [1] outbreak) as well as the LAM and T families. the hospital or it could be that the MDR TB version has
A similar strain family distribution was reported for the become predominant in the population because of the low
French Departments of the Americas (7) and Turkey (8). fitness cost of its 2 mutations, katG315 and S531L (10,11).
The Beijing family was seldom encountered in these areas, In addition, the presence of clusters suggests that even
which is in line with recent observations in 7 countries in though new technologies have reduced the time taken to
South America, including Argentina (9). diagnose drug resistance, more rapid initial diagnosis of
The MDR TB Haarlem2 strain appears to be more MDR TB to reduce transmission still is needed (12).
successful than other circulating MDR TB strains and than All except 1 of the rpoB mutations in the MDR TB
its susceptible counterpart (of 25 Haarlem2 strains, 20 were strains were at nt positions 1303–1375. This finding
MDR TB). This phenomenon could be associated with a reinforces the value of incorporating already standardized
bias in the sample resulting from the specialized nature of molecular methods for rapidly detecting resistance. Cost

Table. Demographic information for 157 patients in a study of the molecular epidemiology of TB, Buenos Aires, Argentina, June 1,
2006–April 30, 2007*
Patients with MDR TB, Patients with non–MDR TB,
Demographic characteristic n = 57, no. (%)† n = 100, no. (%)‡ p value, OR (95% CI)§
Sex
M 35 (61) 70 (70) 0.271, 1.4667 (0.7404–2.9055)
F 22 (39) 30 (30)
Location
Buenos Aires area 46 (81) 95 (95) 0.004, 4.5435 (1.491–13.845)
Other 11 (19) 5 (5)
Country of birth
Argentina 43 (75) 66 (66) 0.2176, 0.632 (0.3041–1.3133)
Bolivia 6 (11) 20 (20)
Peru 7 (12) 8 (8)
Paraguay 0 3 (3)
Uruguay 1 (2) 1 (1)
Chile 0 1 (1)
Missing data 0 1 (1)
Education
Illiterate or some primary 16 (28) 32 (32) 0.2059, 0.5185 (0.1860–1.4456)
Some secondary or tertiary 7 (12) 27 (27)
Missing data 34 (59) 41 (41)
Occupation
Unemployed 7 (12)
Construction and manual worker 20 (35)
Factory worker 4 (7) 14 (14)
Health care worker 1 (2) 1 (1)
Education, i.e., student and teacher 2 (4) 4 (4)
Housewife 6 (11) 5 (5)
Missing data 17 (30) 23 (23)
HIV infection
Positive 25 (44) 27 (27) 0.04, 0.4737 (0.2308–0.9722)
Negative 25 (44) 57 (57)
Missing data 7 (12) 16 (16)
Nature of TB contact
Close (i.e., household, family, co-worker) 10 (18) 32 (32)
Institution (i.e., hospital, prison) 2 (4) 3 (3)
Casual (e.g., acquaintance) 5 (9) 3 (3)
Missing data 40 (70) 62 (62)
TB presentation
Pulmonary 36 (63) 61 (61) 0.7184, 1.1553 (0.5323–2.5073)
Nonpulmonary 15 (26) 22 (22)
Missing data 6 (11) 17 (17)
*TB, tuberculosis; MDR, multidrug resistant; OR, odds ratio; CI, confidence interval; IQR, interquartile range.
†Median age, y (IQR) for patients with MDR TB: 34 (27–40).
‡Median age, y (IQR) for patients with non-MDR TB: 28.5 (23.0–37.0).
§Ȥ2 test.

Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 17, No. 3, March 2011 529
DISPATCHES


Acknowledgments
Patients’culturepositivefor Monoresistance
MDRandfully
M.tuberculosis,excluding toanydrugor
We thank Juan Metrebian and Edward Hogg for their
susceptibleTB
(n=881)
duplicatesandtreatment polyresistance constant support and encouragement and Preya Vekaria for her
 followͲups(N=996) (n=115)
advice and help.
MDRTB FullysusceptibleTB Dr Gonzalo is a clinician and microbiologist working
(n=62) (n=819) Excluded
in the private and public health sector in Argentina on HIV,
tuberculosis, and clinical microbiology. Her research interests
Outpatientand
Clinicalrecords
incompletedataset include tuberculosis, mycobacteria, molecular epidemiology,
and HIV.
Complete*

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