Chinese Medical Journal 2010;123(2):184-187
Nontuberculous mycobacteria: susceptibility pattern and prevalence rate in Shanghai from 2005 to 2008

Correspondence to:YUE Jun,Key Laboratory of Mycobacteria Tuberculosis, Shanghai Pulmonary Hospital Affiliated to Tongji University, Shanghai 200433, China (Tel: 86-21- 65115006-3027. Fax:86-21- 65115006-3027.
prevalence rate; nontuberculous mycobacteria; susceptibility pattern

Background  An increasing incidence of disease caused by nontuberculous mycobacteria (NTM) is being reported. The purpose of this study was to determine the isolation rates of NTM from various clinical specimens, and their antimicrobial susceptibility patterns, over a 4-year period in Shanghai.
Methods  All NTM isolated between 2005 and 2008 at Shanghai Pulmonary Hospital, a key laboratory of mycobacteria tuberculosis in Shanghai, China, were identified with conventional biochemical tests and 16S rRNA gene sequencing. Antimicrobial susceptibility for all NTM was determined using the BACTEC MGIT 960 system.
Results  A total of 21 221 specimens were cultured, of which 4868 (22.94%) grew acid fast bacilli (AFB), and 248 (5.09%) of the AFB were NTM. The prevalence rate of NTM was determined as 4.26%, 4.70%, 4.96% and 6.38% among mycobacteria culture positive samples in years 2005, 2006, 2007 and 2008 respectively. These data indicated that the prevalence rate has continuously increased. Sixteen different species of NTM were identified, the most commonly encountered NTM in Shanghai were M. chelonae (26.7%), followed by M. fortuitum (15.4%), M. kansasii (14.2%), M. avium-intracellulare complex (13.1%) and M. terrae (6.9%). The rare species identified were M. marinum, M. gastri, M. triviale, M. ulcerans, M. smegmatis, M. phlci, M. gordonae, M. szulgai, M. simiae, M. scrofulaceum and M. xenopi. The five most commonly identified NTM species showed high drug resistance to general anti-tuberculosis drugs, particularly, M. chelonae and M. fortuitum appear to be multi-drug resistance.
Conclusions  The prevalence of NTM in Shanghai showed a tendency to increase over the course of the study. The five most commonly isolated NTM species showed high drug resistance to first line anti-tuberculosis drugs.

The rising incidence of human infection by nontuberculous mycobacteria (NTM) is a serious public-health concern. NTM are widely distributed in the environment with high isolation rates worldwide; they can be found in soil, water, animals, and dairy products.1-6 NTM can cause a broad spectrum of diseases: (1) pulmonary infections resembling tuberculosis; (2) extra pulmonary infections affecting lymph nodes, skin and soft tissue; (3) multifocal disseminated infections; (4) infections in immunocompromised individuals, such as AIDS and transplant patients,7 and (5) nosocomial infections with outbreaks related to inadequate disinfection/ sterilization of medical devices.8,9 NTM are catched increasing attention worldwide.

In this study, we have reviewed the isolation rate of NTM from various clinical specimens at Shanghai Pulmonary Hospital over a period of four years. We have also investigated the prevalence of NTM in Shanghai and determined the drug-resistance susceptibility pattern for treatment.


Clinical isolates
This study included the isolation of NTM from 21 221 clinical specimens from suspected pulmonary and extra pulmonary tuberculosis received between 2005 and 2008. Early morning, well coughed out sputum specimens and broncho-alveolar lavage fluid (BALF) were collected from patients with clinical and radiological findings suggestive of pulmonary tuberculosis. A punch biopsy was obtained from patients with lymphadenitis or with abscesses. Entire collective, 24-hour urine specimens were received from patients suspected of urinary tuberculosis. Sterile body fluids, such as cerebrospinal fluid (CSF) or pleural fluid, were collected from patients suspected of disseminated mycobacteria infection. All specimens were collected with aseptic precautions in sterile containers and transported to the laboratory.

Laboratory procedures
Clinical specimens were digested with 4% sodium hydroxide. The processed specimens were inoculated into a mycobacteria Growth Indicator Tube and placed into the BACTECTM MGITTM 960 system (Becton Dickinson Company, USA) for continuous monitoring until they tested positive or until the end of the testing period.10,11 An instrument-positive sample was determined by the system and confirmed by an acid-fast smear. NTM can be differentiated from mycobacteria tuberculosis complex by P-Nitrobenzoic acid (PNB) and 2-Thiophene carboxylic acid hydrazide (TCH) tests, and further identified with conventional biochemistry tests and 16SrRNA gene sequencing to the species level.

The BACTECTM MGITTM 960 SIRE kit was used for antimycobacterial susceptibility testing (AST) of NTM. Critical concentrations of isoniazid (INH, 0.1 µg/ml), rifampin (RFP, 1.0 µg/ml), ethambutol (EMB, 5.0 µg/ml) and streptomycin (SM, 1.0 µg/ml) were tested.12,13


Of the 21 221 specimens received for mycobacteria culture during the four year study period, 4868 (22.94%) grew acid fast bacilli (AFB). Two hundred and forty-eight (5.09%) of the AFB were NTM from various clinical specimens with a presumptive diagnosis of mycobacterial infection. The prevalence rate of NTM was determined as 4.26% (48/1127), 4.70% (56/1191), 4.96% (65/1311), 6.38% (79/1239) among mycobacteria culture positive samples in years 2005, 2006, 2007 and 2008 respectively. The mean prevalence rate of NTM is 5.09% (248/4868).

The median age of the study patients with positive NTM cultures was 60.5 years. Most (67%, n=166) patients with disease were >56 years of age; 131 specimens (53%) were isolated from males and 117 specimens (47%) were isolated from females.

Distribution of infected NTM species isolated from all specimens between 2005 and 2008 is shown in the Table.

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Table. Distribution of infected NTM species isolated in Shanghai between 2005 and 2008

Sixteen different species of NTM were identified here. The most commonly encountered NTM in Shanghai was M. chelonae (n=66, 26.7%), followed by M. fortuitum (n=38, 15.4%), M. kansasii (n=35, 14.2%), M. avium-intracellulare complex (MAC) (n=32, 12.9%) and M. terrae (n=17, 6.9%), the rare species were M. marinum, M. gastri, M. triviale, M. ulcerans, M. smegmatis, M. phlci, M. gordonae, M. szulgai, M. simiae, M. scrofulaceum and M. xenopi. The majority (46.8%) of isolates were rapidly growing mycobacteria (RGM) (Figure 1).

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Figure 1. Isolation of NTM from clinical specimens.

Antibiotic susceptibility tests were done for the five most common NTM species, namely, M. chelonae, M. fortuitum, M. kansasii, MAC and M. terrae. MAC showed the highest susceptibility to RFP (99.6%), only one isolate of MAC were resistant to RFP. Susceptibility of MAC to SM, INH, and EMB was 58.8%, 24.6% and 66.5% respectively. But M. chelonae and M. fortuitum showed much less susceptibility to these drugs and showed multi-drug resistance. (Figure 2)

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Figure 2. Antibiotic susceptibility patterns of NTM.


Our study reveals a clear increase in the prevalence of NTM infection in Shanghai during the past four years. More recent data have shown an increase in the number and distribution of NTM infections. Data from two national surveys of NTM isolates among mycobacteria positive cultures done in 1990 and in 2000 showed that the isolate rate had increased from 4.5% to 11.1%. Possible reasons are as follows. (1) Better culture methods, primarily using broth-based systems, have been employed in more laboratories. There is evidence that broth-based systems are more sensitive than solid media-based systems for recovering NTM.14,15 Thus, to a certain extent, the increase in detected cases of disease due to NTM may reflect an improvement in methodology in the mycobacteriology laboratory, resulting in enhanced isolation and more rapid and accurate identification of NTM. (2) Due to an aging population, older patients (defined here arbitrarily as persons 56 years old and older) account for a significant number of NTM cases in our study. It is true that immune function, and in particular cellular immune function, declines with age, and there is every reason to think that older people are more susceptible to developing NTM than younger people.16-18 (3) There are increased numbers of HIV infections and it is well known that these patients have a greatly increased risk of nontuberculous mycobacterial infections. Mycobacterium avium infections account for almost 50% of mycobacterial infections among AIDS patients in certain geographical areas. So, NTM infections have increased in number and in clinical significance over the past decade.

NTM infections vary geographically by species. We can see from Figure 1 that the most commonly encountered NTM in Shanghai were M. chelonae (26.7%), followed by M. fortuitum (15.4%), M. kansasii (14.2%), MAC (13.1%) and M. terrae (6.9%). The majority (46.8%) of isolates were RGMs. The isolation rate of RGMs among all NTM in this study was similar to that reported in past studies in China. As was reported by the Beijing Tuberculosis Control Research Institute, among 52 strains of NTM isolated from sputum specimens, 48.1% were MAC, followed by M. gordonae (15.4%), M. chelonae (11.5%), M. scrofulaceum (7.7%), and M. flavescens (5.8%). RGMs accounted for 51.5% of isolates. M. avium-intracellulare is the most commonly encountered NTM in the United States, accounting for 61% of isolates, but in Africa it rarely causes disseminated infection in HIV/AIDS patients.19 In Africa, 19% of isolates were M. fortuitum complex and 1% were M. kansasii. M. xenopi, which is rarely found in the United States, is the second most commonly isolated organism in Canada and Europe.20

The management of NTM infections includes medical treatment with various antimicrobial agents based on susceptibility patterns and surgical treatment, as in the case of lymphadenitis, and skin or soft tissue infections. In our study, most of the NTMs showed high drug resistance to general anti-tuberculosis drugs. Since most of these organisms are resistant to commonly used antimicrobial agents, and their drug-resistance rate varies according to the NTM species, treating NTM infections is difficult and requires extended courses of multi-drug therapy. Susceptibility testing and accurate, rapid identification of mycobacterium becomes mandatory before instituting an effective therapy. In China, NTM infection, as well as tuberculosis, is still a major health care problem, especially in impoverished mountain areas. These infections can not be diagnosed in many laboratories due to the lack of facilities and expertise. Newer molecular methods such as gene-probes, PCR and DNA fingerprinting may be better diagnostic tools and have become a hot-topic public health care issue.

In conclusion, we found an increased incidence of NTM-positive cultures and disease compared with results in previous reports. RGM, especially M. fortuitum, appeared multi-drug resistant, posing a therapeutic challenge. New antibiotics, with excellent activity against NTM and multi-resistance tuberculosis, have been studied in recent years and may offer more successful and safe treatment options in the future.

Acknowledgments: We thank all study participants for their cooperation, for technical help and sample collection. We are grateful to all members of the Mycobacteriology Laboratory at Shanghai Pulmonary Hospital Affiliated to Tongji University for their knowledge, helpfulness and willingness to share.


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  1. Great Research Program of China,No. 2008zx10003-009;