ABSTRACT
This study evaluates the growth of mycobacteria in samples from cystic fibrosis (CF) patients and tissue samples using the mycobacteria growth indicator tube (MGIT) incubated at 30°C in comparison to conventional MGIT cultures incubated at 37°C in a BACTEC MGIT 960 device and solid media incubated at 36°C and 30°C. A total of 1,549 samples were analyzed, of which 202 mycobacterial isolates were cultured from 197 positive specimens, including five mixed cultures. The highest detection rate was achieved from MGIT at 30°C, with 84.2% of mycobacterial isolates (170 of 202), which was significantly higher than any other culture condition (P < 0.0001 for any condition). MGIT at 37°C yielded 61.4% (124 of 202) of the recovered isolates, whereas Löwenstein Jensen (LJ) and Stonebrink at 36°C, and LJ and Stonebrink at 30°C retrieved 47.0% (95), 49.5% (100), 50.0% (101), and 53.0% (107) of the isolates, respectively. Of the 53 isolates that were grown exclusively under one culture condition, the highest number of isolates (36) was recovered from MGIT incubated at 30°C. MGIT at 37°C recovered eight of the 53 isolates, whereas LJ incubated at 30°C and Stonebrink incubated at 30°C and 36°C recovered five, three, and one isolate, respectively. No isolates were grown exclusively from LJ incubated at 36°C. In CF patients and tissue samples, MGIT cultivated at 30°C for 8 weeks increases the performance of mycobacterial culture.
IMPORTANCE
Our study shows that the addition of mycobacteria growth indicator tube (MGIT) liquid culture incubated at 30°C improves the detection of mycobacteria from CF and tissue samples. MGIT incubated at 30°C recovered significantly more mycobacterial isolates than MGIT incubated at 37°C and significantly more isolates than either Lowenstein Jensen or Stonebrink solid media incubated at either 36°C or 30°C. Of 202 mycobacterial isolates recovered from 1,549 specimens, 170 were recovered from MGIT incubated at 30°C, followed by MGIT incubated at 37°C with 124 isolates and solid media culture conditions that recovered between 95 and 107 mycobacterial isolates. All conventional culture conditions combined without MGIT incubated at 30°C recovered 166 isolates. MGIT incubated at 30°C recovered the highest number of isolates detected exclusively by a single culture condition and recovered mycobacterial isolates of highly relevant mycobacterial species, including Mycobacterium abscessus and Mycobacterium tuberculosis.
KEYWORDS: MGIT, non-tuberculous mycobacteria, culture, cystic fibrosis, temperature
INTRODUCTION
The genus Mycobacterium comprises more than 200 bacterial species that vary in pathogenicity and their ability to cause human disease (1). Mycobacterium tuberculosis and Mycobacterium leprae are obligate human-to-human transmitted pathogens, whereas non-tuberculous mycobacteria (NTM) are considered to be acquired from environmental sources (2). Several NTM species can cause a wide range of human diseases, including pulmonary, lymphatic, skin, tissue or disseminated disease. However, many other NTM species have not yet been linked to human disease. The genus Mycobacterium has a high level of genetic diversity and specific growth requirements (2, 3). Different species have different growth rates and optimal growth temperatures (2). Although M. tuberculosis and some clinically relevant NTM grow well at 35°C to 37°C, other clinically relevant NTM species, including Mycobacterium marinum and Mycobacterium ulcerans or individual strains from clinically relevant NTM species, such as Mycobacterium chelonae and Mycobacterium abscessus, exhibit optimal growth on culture media at lower temperatures (2). The addition of liquid culture to the traditionally used solid media has improved mycobacterial diagnostics by reducing the time to detect mycobacteria and by increasing the sensitivity of mycobacterial culture (4–7). Guidelines for mycobacterial diagnostics recommend the use of a liquid medium with automated growth detection, cultured at 37°C in addition to the use of solid media (8, 9). Cystic fibrosis (CF), a genetic disorder affecting the lungs, is one of the primary diseases with the highest prevalence of NTM lung disease and the highest rate of NTM recovered from respiratory specimens. Antibiotic treatments for recurrent lung infections in CF patients facilitate colonization of the respiratory tract with multidrug-resistant, predominantly Gram-negative bacteria, such as Pseudomonas aeruginosa. This complicates diagnostics and leads to higher contamination rates of mycobacterial cultures in this patient population. The implementation of sequential decontamination procedures, such as oxalic acid, is utilized to reduce contamination rates in CF specimens by killing microbial contaminants. However, this procedure may also reduce mycobacterial growth. A number of research groups have conducted studies to determine the optimal growth temperatures for mycobacterial isolates in vitro. These studies have assessed the extent and rate of growth on culture media under different conditions. Therefore, additional cultures have been recommended to be incubated at 30°C in parallel with the conventional cultures to improve recovery of mycobacteria with a lower optimal growth temperature (2). For solid media, the incubation at 30°C has been shown to be superior compared with 36°C–37°C for the recovery especially of rapidly growing mycobacteria (10, 11). To our knowledge, the performance of mycobacteria growth indicator tube (MGIT) liquid medium incubated at 30°C for the detection of mycobacteria from CF and tissue samples in comparison to conventional culture conditions has not been evaluated before. The aim of this study was to investigate whether MGIT incubation at 30°C improves mycobacterial culture in clinical samples from patients with CF and tissue and puncture fluid samples, which are anticipated to have a higher prevalence of NTM growing at lower optimal growth temperatures. To this end, the efficacy of MGIT incubation at 30°C was compared with that of MGIT incubation at 37°C and solid media incubation at 36°C and 30°C.
MATERIALS AND METHODS
Study design
All samples from CF patients, tissue and puncture fluid samples submitted to the University Hospital Essen for mycobacterial diagnostics between February 2021 and December 2023 were included in the study. All samples were decontaminated using the NALC-NaOH method with the BD BBL-Myco-Prep Kit (Becton Dickinson, Franklin Lakes, NJ, US) according to the manufacturer’s guidelines. Auramine staining was performed using the Aerospray TB device (Kreienbaum Neoscience, Langenfeld, Germany) for the detection of acid-fast bacilli in all decontaminated primary specimens included in the study. We inoculated 500 µL of the decontaminated sample into MGIT liquid medium supplemented with PANTA (polymyxin B 50 U/mL, amphotericin B 5 mg/L, nalidixic acid 20 mg/L, trimethoprim 5 mg/L, and azlocillin 10 mg/L) that was incubated in an automated Bactec MGIT 960 device (Becton Dickinson) at 37°C. A volume of 100 µL of each decontaminated sample was inoculated onto Lowenstein Jensen (LJ) with PACT (polymyxin 200,000 IU/L, amphotericin 10 mg/L, carbenicillin 50 mg/L, and trimethoprim 10 mg/L) and Stonebrink solid media with PACT (both from Oxoid, Wesel, Germany) and incubated at each 36°C and at 30°C. A volume of 500 µL of decontaminated sample was inoculated into an additional MGIT liquid medium, which was incubated together with solid media in a standard 30°C incubator. Solid media and MGIT incubated at 30°C were visually inspected weekly for growth. At week 8, MGIT liquid tubes incubated at 30°C were additionally assessed for growth using MicroMGIT fluorescence reader (Becton Dickinson). Ziehl–Neelsen staining was performed primarily on all positive cultures to confirm mycobacterial growth. All positive MGIT liquid cultures from CF patients incubated at 37°C that were contaminated with non-mycobacterial pathogens, which was confirmed by Ziehl–Neelsen staining, were sequentially decontaminated using an oxalic acid decontamination procedure with 5% oxalic acid (12) before continuation of the culture. Therefore, another MGIT tube was inoculated and incubated at 37°C. In addition, the oxalic acid-decontaminated sample was incubated on LJ and Stonebrink solid media at 36°C. All positive subcultures after oxalic acid decontamination were assigned to the MGIT tube incubated at 37°C. In the subcultures that were positive for mycobacterial growth, the oxalic acid-decontaminated MGIT tube was positive for all six NTM isolates. Moreover, the LJ and Stonebrink subcultures incubated at 36°C from the oxalic acid decontaminated MGIT incubated at 37°C were positive for four NTM isolates. The culture period of solid media and MGIT incubated at 30°C was prolonged for 4 weeks for those samples that were positive for acid fast bacilli from primary specimens but yielded negative culture after 8 weeks. A total of two MGIT cultures incubated at 30°C demonstrated positive results during the extended incubation period. One positive result was observed at week 9, and the other at week 12, both for M. tuberculosis.
Mycobacterial identification was primarily performed using GenoType CM, GenoType AS, GenoType NTM-DR, and GenoType MTBC (Bruker, Billerica, MA, US). For selected NTM isolates, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI ToF MS; MALDI Biotyper Sirius, Bruker) was used for mycobacterial identification. McNemar’s test, a paired sample statistical test that examines a dichotomous characteristic, was used to analyze differences in performance between MGIT incubated at 30°C and each of the alternative culture conditions and between MGIT incubated at 30°C and all other culture methods combined for recovery of mycobacterial isolates. Statistical significance was defined as P< 0.05.
The study was reviewed and approved by the Ethics Committee of the Medical Faculty of the University of Duisburg-Essen (no. 23–11655-BO). All samples were collected as a part of standard care. The recommendations of the ICH-GCPC guidelines were followed, and the study was conducted in accordance with the latest version of the Declaration of Helsinki.
RESULTS
All mycobacterial diagnostic samples collected from CF patients, tissues and puncture fluids between February 2021 and December 2023 were included in the study. All samples were inoculated in a MGIT incubated at 30°C, in addition to conventional culture conditions that included MGIT incubated at 37°C in a BD Bactec MGIT 960 device (Becton Dickinson), LJ incubated at both 36°C and 30°C, and Stonebrink solid media incubated in regular incubators at 36°C and 30°C.
A total of 1,549 samples were analyzed, including 1,158 respiratory samples from CF patients, 381 tissue samples, five puncture fluids, four heparin blood samples, and one catheter sample from a CF patient (Table 1). Acid-fast bacilli were observed by microscopy in 42 samples. A total of 202 mycobacterial isolates were cultured from 197 samples. Five of the positive cultures were mixed cultures with two mycobacterial species recovered in each sample. Six of the 202 mycobacterial isolates (four M. intracellulare subsp. chimaera, one M. abscessus, and one M. gordonae) were recovered from MGIT incubated at 37°C after a second decontamination step with oxalic acid treatment. Of the 202 grown mycobacterial isolates, 143 were cultured from respiratory samples, 48 from tissue samples, and six from blood, puncture fluid, or catheter samples (Table 1). The 143 respiratory samples were comprised of 138 sputum samples, four bronchial secretion samples and one bronchoalveolar lavage sample.
TABLE 1.
Sample specimens and number of positive cultures
| Specimen | No. of cultures | No. of cultures positive for mycobacteria | No. of smears positive for AFB |
|---|---|---|---|
| Respiratory CF specimens | |||
| Sputum | 1,077 | 138 | 26 |
| Bronchoalveolar lavage fluid | 12 | 1 | 0 |
| Bronchial wash | 14 | 4 | 0 |
| Throat lavage/swab | 55 | 0 | 0 |
| Other specimens | |||
| Tissue | 263 | 32 | 12 |
| Lymph node | 70 | 11 | 2 |
| Tissue swab | 48 | 5 | 1 |
| Puncture fluid | 5 | 1 | 0 |
| Heparin blooda | 4 | 4 | 0 |
| Cathetera | 1 | 1 | 1 |
| Total | 1,549 | 197 | 42 |
The four heparin blood and the catheter sample were from a CF patient with a venous port catheter infection due to M. intracellulare subsp. chimaera.
The highest detection rate of mycobacterial isolates, 84.2% (170 of 202) was observed in MGIT incubated at 30°C (Table 2). MGIT at 37°C recovered 61.4% (124 of 202) of the isolates, whereas LJ at 36°C, Stonebrink at 36°C, LJ at 30°C, and Stonebrink at 30°C recovered 47.0% (95), 49.5% (100), 50.0% (101), and 53.0% (107) of the isolates, respectively. The total number of positive mycobacterial cultures from MGIT incubated at 30°C was significantly higher than that of any of the other conventional culture conditions (P < 0.0001 for any comparison of MGIT incubated at 30°C with any individual culture condition). The total number of mycobacteria that were successfully grown from MGIT incubated at 30°C was greater (170) than the number of all conventional culture conditions combined (166 isolates); however, this was not statistically significant (P = 0.39). MGIT incubated at 30°C detected the highest number of isolates from all specimens included in the study for M. abscessus complex (41), M. tuberculosis (24), M. intracellulare subsp. chimaera (22), M. gordonae (22), M. avium (20), M. chelonae (6), M. mucogenicum (3), M. kansasii (2), M. celatum (1), M. fortuitum (1), M. intermedium (1), M. nebraskense (1), M. simiae complex (1), and M. spec. (6). The sole exceptions for which other culture conditions recovered more mycobacterial isolates than MGIT incubated at 30°C were MGIT at 37°C for M. intracellulare (6) and M. intracellulare complex (10) and Stonebrink at 30°C and LJ at 30°C for M. marinum (4).
TABLE 2.
Recovery of Mycobacterium species in each of the culture media conditions studied from a total of 1,549 specimens
| Species isolated | Total no. | MGIT 30°C (n [%]) | MGIT 37°C (n [%]) | LJ 30°C (n [%]) | LJ 36°C (n [%]) | Stonebrink 30°C (n [%]) | Stonebrink 36°C (n [%]) |
|---|---|---|---|---|---|---|---|
| M. abscessus complex | 47 | 41 (87) | 35 (74) | 32 (68) | 31 (66) | 31 (66) | 30 (64) |
| M. tuberculosis | 30 | 24 (80) | 24 (80) | 8 (27) | 22 (73) | 14 (47) | 22 (73) |
| M. intracellulare subsp. chimaera | 28 | 22 (79) | 17 (61) | 14 (50) | 6 (21) | 14 (50) | 8 (29) |
| M. gordonae | 28 | 22 (79) | 2 (7) | 4 (14) | 1 (4) | 9 (32) | 2 (7) |
| M. avium | 21 | 20 (95) | 20 (95) | 16 (76) | 18 (86) | 15 (71) | 17 (81) |
| M. intracellulare | 6 | 5 (83) | 6 (100) | 5 (83) | 5 (83) | 5 (83) | 5 (83) |
| M. intracellulare complexa | 11 | 9 (82) | 10 (91) | 7 (64) | 3 (27) | 6 (55) | 5 (45) |
| M. chelonae/immunogenum | 8 | 6 (75) | 3 (38) | 5 (63) | 1 (13) | 3 (38) | 1 (13) |
| M. marinum | 4 | 3 (75) | 0 (0) | 4 (100) | 2 (50) | 4 (100) | 3 (75) |
| M. mucogenicum | 3 | 3 (100) | 1 (33) | 1 (33) | 0 (0) | 0 (0) | 0 (0) |
| M. kansasii | 2 | 2 (100) | 2 (100) | 2 (100) | 2 (100) | 2 (100) | 2 (100) |
| M. bovis | 2 | 2 (100) | 2 (100) | 0 (0) | 2 (100) | 2 (100) | 2 (100) |
| M. celatum | 1 | 1 (100) | 0 (0) | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
| M. fortuitum | 1 | 1 (100) | 0 (0) | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
| M. intermedium | 1 | 1 (100) | 0 (0) | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
| M. nebraskense | 1 | 1 (100) | 0 (0) | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
| M. simiae complex | 1 | 1 (100) | 0 (0) | 0 (0) | 0 (0) | 0 (0) | 0 (0) |
| M. spec. | 7 | 6 (86) | 2 (29) | 3 (43) | 2 (29) | 2 (29) | 3 (43) |
| Total | 202 | 170 (84) | 124 (61) | 101 (50) | 95 (47) | 107 (53) | 100 (50) |
Of the 11 M. intracellulare complex isolates, 10 were from patients with previously positive cultures for M. intracellulare subsp. chimaera and one for M. intracellulare. Bold values indicate the culture condition recovering the highest number of isolates of each species.
Of the 53 mycobacterial isolates recovered by a single culture condition only, 68.0% (36 isolates) were detected by MGIT at 30°C (Table 3). MGIT at 37°C recovered 15.1% (8 isolates), LJ at 30°C recovered 9.4% (5), Stonebrink at 30°C recovered 5.7% (3), Stonebrink at 36°C recovered 1.9% (1), and LJ at 36°C recovered 0.0% (0) of isolates that were not recovered by any other culture condition. M. gordonae was the most frequently cultured species with 21 isolates, followed by M. abscessus with seven isolates, and M. intracellulare subsp. chimaera with six isolates of the mycobacterial isolates that were recovered by only one culture condition. Of the seven M. abscessus isolates grown from only one culture condition, five were cultivated using MGIT at 30°C. One isolate was grown using MGIT at 37°C, and another was cultivated using LJ at 30°C. Notably, four M. tuberculosis isolates were grown exclusively from one culture condition, including three isolates that were recovered solely from MGIT incubated at 30°C and one from MGIT incubated at 37°C. Two samples from which M. tuberculosis was exclusively grown using MGIT at 30°C demonstrated positive results for acid-fast bacilli and exhibited a positive MTB complex PCR from primary specimens. With the exception of M. intracellulare, which demonstrated optimal recovery when incubated with MGIT at 37°C, MGIT incubated at 30°C yielded the greatest number of mycobacteria isolates exclusively recovered from one culture condition.
TABLE 3.
Isolates recovered exclusively by one culture conditiona
| Species | No. of isolates | MGIT 30°C | MGIT 37°C | LJ 30°C | LJ 36°C | Stonebrink 30°C | Stonebrink 36°C |
|---|---|---|---|---|---|---|---|
| M. gordonae | 21 | 15 | 1 | 2 | 0 | 3 | 0 |
| M. abscessus complex | 7 | 5 | 1 | 1 | 0 | 0 | 0 |
| M. intracellulare subsp. chimaera | 6 | 2 | 2 | 2 | 0 | 0 | 0 |
| M. tuberculosis | 4 | 3 | 1 | 0 | 0 | 0 | 0 |
| M. spec. | 3 | 2 | 0 | 0 | 0 | 0 | 1 |
| M. intracellulare | 1 | 0 | 1 | 0 | 0 | 0 | 0 |
| M. intracellulare complex | 2 | 0 | 2 | 0 | 0 | 0 | 0 |
| M. chelonae/immunogenum | 2 | 2 | 0 | 0 | 0 | 0 | 0 |
| M. mucogenicum | 2 | 2 | 0 | 0 | 0 | 0 | 0 |
| M. intermedium | 1 | 1 | 0 | 0 | 0 | 0 | 0 |
| M. fortuitum | 1 | 1 | 0 | 0 | 0 | 0 | 0 |
| M. simiae complex | 1 | 1 | 0 | 0 | 0 | 0 | 0 |
| M. nebraskense | 1 | 1 | 0 | 0 | 0 | 0 | 0 |
| M. celatum | 1 | 1 | 0 | 0 | 0 | 0 | 0 |
| Total | 53 | 36 | 8 | 5 | 0 | 3 | 1 |
Bold values indicate the culture condition recovering the highest number of isolates of each species.
A total of 223 MGIT cultures incubated at 30°C were contaminated, whereas 169 cultures of MGIT incubated at 37°C were contaminated after the second oxalic decontamination procedure. For the solid media, 168, 170, 151, and 158 cultures were contaminated from LJ incubated at 36°C, LJ at 30°C, Stonebrink at 36°C, and Stonebrink at 30°C. Nocardia sp. isolates recovered from three cultures of MGIT incubated at 30°C, one culture of MGIT at 37°C, two cultures of LJ at 30°C, and four cultures of Stonebrink at 30°C were not included in the number of contaminated samples.
DISCUSSION
The inclusion of one MGIT tube, incubated at 30°C, to the conventional mycobacterial culture methodology increased the recovery of isolates in cystic fibrosis patients and tissue samples. The MGIT incubated at 30°C alone significantly recovered more mycobacterial isolates (170) compared with any other culture condition (P < 0.0001), and recovered more isolates than all other culture methods combined (166), although this was not statistically significant.
The standard incubation temperature for mycobacterial culture using MGIT liquid medium is 37°C, which is equivalent to the human body temperature. This is considered as the optimum growth temperature for M. tuberculosis. Studies have demonstrated that incorporating liquid media in addition to solid media incubated at 36°C enhances the speed and sensitivity of mycobacterial culture (4–7), and the use of a liquid medium along with solid media is standard for mycobacterial culture (2, 13). The recommendation to include two additional solid media incubated at 30°C for mycobacterial culture in CF patients and tissue samples is based on the frequent presence of clinically relevant mycobacteria that have a lower optimum growth temperature, such as M. abscessus complex for CF patients or M. marinum and M. ulcerans for tissue samples. An improved recovery of primarily rapidly growing mycobacteria in CF and non-CF-patients has been reported from solid media incubated at 30°C compared with 36°C (10, 11). However, there is a lack of studies analyzing the performance of detecting mycobacteria in CF patients and tissue samples from MGIT liquid culture incubated at both 37°C and 30°C. In our study, the conventional culture conditions without MGIT 30°C recovered mycobacteria in 10.1% of CF sputum samples (109 NTM positive of 1,077 sputum cultures). This falls within the range of studies reporting a positive culture rate between 5% and 14%, depending on the geography (14, 15). The NTM prevalence in CF patients from Germany is estimated to be approximately 8% (16).
The addition of MGIT incubated at 30°C increased the recovery of mycobacteria from 10.1% (109 of 1,077) to 12.8% (138 of 1,077) of sputum cultures from CF patients. Species with doubtful and low clinical significance, including M. gordonae, as well as those with high clinical significance, such as M. abscessus, were among those grown in higher numbers in MGIT incubated at 30°C. In our study, we found that incubating MGIT cultures at 30°C resulted in the recovery of a higher number of mycobacterial isolates, particularly those that thrive at lower temperatures, such as M. gordonae, M. abscessus, M. marinum, M. chelonae, M. mucogenicum, and M. nebraskense. Interestingly, the number of isolates recovered from MGIT incubated at 30°C was also not lower for most mycobacterial species considered to have a growth optimum between 35°C and 37°C, including M. tuberculosis than for MGIT incubated at 37°C. Of the four M. tuberculosis isolates obtained from four patients that were cultured exclusively from one culture condition, three were grown from MGIT cultured at 30°C, and only one was grown from MGIT cultured at 37°C. None of the 30 M. tuberculosis isolates was cultured exclusively from any of the other culture conditions. Two tissue samples of the MTB positive cultures grown exclusively from MGIT incubated at 30°C were positive for acid-fast bacilli and had a positive MTB complex PCR from the primary specimen. The other two tissue samples, one grown exclusively from MGIT at 30°C and one from MGIT at 37°C, were negative for acid-fast bacilli and had a negative MTB complex PCR.
The inoculation volume of 500 µL of decontaminated sample into MGIT is higher than the volume used for inoculating solid media with 100 µL. This difference in volume may contribute to a higher sensitivity of liquid media compared with solid media. However, the volume is equal for the liquid cultures of both temperature conditions and may not account for the difference in their sensitivity. MGIT at 30°C was incubated for 8 weeks, which was 2 weeks longer than MGIT at 37°C, and was read in parallel with the solid media. The higher recovery rate observed in MGIT incubated at 30°C compared with MGIT at 37°C may be partially attributed to this factor. For decades, it is well known that the culture from caseous tissues and calcified lesions of TB patients is sterile in a high percentage of cases, especially when the lesions are not communicating with a bronchus, in contrast to those communicating with a bronchus (17, 18). However, a percentage of 44% of those tissues closed off from bronchi produced positive cultures when the incubation time was prolonged to 3 to 10 months (17). When growing, the tuberculous mycobacterial isolates exhibited normal morphology, growth in culture, and were able to infect animals (17, 18). These results indicate that the local microenvironments are highly diverse and may impact the replication rate in vivo as well as the growth requirements ex vivo (19).
The higher percentage of contaminated samples of our study than aimed in the guidelines is due to the high number of samples from CF patients, which accounted for 75% of the samples included in the study, and may also exhibit contamination of conventional mycobacterial cultures >20% (20). A second decontamination step using oxalic acid was exclusively performed from MGIT incubated at 37°C but not from MGIT incubated at 30°C. The lacking decontamination step from MGIT incubated at 30°C may significantly contribute to the lower rate of contaminated samples from liquid media incubated at 37°C compared with MGIT incubated at 30°C.
The results of this study indicate that the detection of mycobacteria from CF patients and tissue samples can be enhanced by incorporating an MGIT culture incubated at 30°C for 8 weeks into the conventional culture conditions. Additionally, the findings indicate that Löwenstein Jensen incubated at 36°C can be omitted from these cohorts, as no mycobacterial isolate from 1,549 specimens from CF patients and tissues was exclusively cultured from it. The study was limited by its inclusion of patients from a single university hospital. Because NTM species isolated from clinical specimens vary between different countries and regions of the world (21), the improvement in mycobacterial culture observed in our study may not be generalizable to other geographic regions. Multi-center studies would be beneficial to confirm the efficacy of MGIT incubated at 30°C for 8 weeks in improving the yield of mycobacterial cultures in CF and tissue specimens, which could influence recommendations for mycobacterial culture in diagnostic guidelines. In addition, multi-center studies may be beneficial to evaluate the potential for replacing other media with MGIT incubated at 30°C. It seems reasonable to conclude that the incorporation of MGIT incubated at 30°C for enhancing mycobacterial culture is likely to be most effective when applied to the specimens included in our study. This is because the specimens more likely contain mycobacteria with lower optimal growth temperatures than would be the case if the method were to be applied to all specimens. Nevertheless, it would be beneficial to ascertain whether the 30°C incubation of MGIT is exclusive to CF and tissue specimens, or if it could potentially enhance the recovery of mycobacteria from other patients and specimens that are not enriched in mycobacteria with lower optimal growth temperatures.
ACKNOWLEDGMENTS
We thank Nermin Kurt, Cornelia Aschenbroich, Bianca Böke, Gabriela Inting, and Janine Lau for technical assistance.
Contributor Information
J. Kehrmann, Email: jan.kehrmann@uk-essen.de.
Christine Y. Turenne, University of Manitoba, Winnipeg, Manitoba, Canada
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