CaseIsolation Report of Mycobacterium fortuitum from BACTEC 9240 Blood Culture System Isolation of Mycobacterium Fortuitum from BACTEC 9240 Blood Culture System: A Case Report Fang-Lan Yu1, Jau-Ching Lee1, Tsung-Han Wu2, Yi-Lin Liang1, Chia-Wen Lin1, Tzu-Ting Chen1, Giueng-Chueng Wang1 1Department of Laboratory Medicine 2Tuberculosis center Taipei Medical University-Wan Fang Hospital We describe an 89-year-old male patient who admitted due to pneumonia documented bacteremia by isolating Mycobacterium fortuitum (M.fortuitum) from BACTEC 9240 blood culture system in April 2009. During his admission, two sets of blood culture were reported as positive. Microscopic examination revealed atypical gram-positive bacilli and acid-fast organism. This organism was subsequently identified as M. fortuitum. Concerning that M. fortuitum could be cultured and iso- lated in this case, not only owing to it is a rapidly growing mycobacteria, but also the professional- ism that the experienced technicians have. Laboratory staff should have awareness of performing mycobacteria culture when encountering an atypical gram-positive pathogen accompanying with delayed aerobic culture result. Key words: Rapidly growing mycobacteria, Mycobacterium fortuitum, BACTEC 9240 Blood Culture System M. fortuitum is a gram-positive and acid-fast bacilli. It is also a saprophyte whose natural habitat includes Introduction soil, water and dust. Nowadays M. fortuitum is increas- ingly recognized as an opportunistic pathogen causing Runyon described the four groups of non-tuberculous disseminated infection [3-5]. Clinical presentation of M. mycobacteria (NTM), a grouping that encompasses all fortuitum includes mainly cutaneous and soft tissue in- mycobacteria outside of the Mycobacterium tuberculosis fections, localized posttraumatic wound infections, sur- complex, according to pigment production and rate of gical wound infections and keratitis [6-8]. In general, growth in 1959 [1]. These are Runyon Group I: the immunocompetent patients tend to experience limited photochromogens, Runyon Group II: the scotochro- infections associated with low mortality. mogens, Runyon Group III: the nonchromogens, which Bacteremia is a serious clinical condition which can are classified as slowly growing mycobacteria, and Runyon lead to death, and consequently a rapid and accurate de- Group IV: the rapid growers, which may be photochro- tection and identification of the pathogen plays the cru- mogenic, scotochromogenic, or more usually nonchro- cial role in effective treatment. More than one blood mogenic, are defined as visible growth on Löwen- sample are collected and examined to detect the micro- stein-Jensen slant medium (Becton Dickinson) within organism exists and, to identify its species, and to deter- seven days on subculture [2]. Mycobacterium fortuitum mine its drug susceptibility. (M.fortuitum) is a member of the rapidly growing It goes without saying that shortening the turn- Runyon Group IV NTM. Besides M. fortuitum, the around time of microbiological analyses is fairly sig- common organisms of Runyon group IV are Mycobacte- nificant and is closely related to declining patients’ mor- rium peregrinum, Mycobacterium senegalense, Myco- bidity and mortality [9-10]. In clinical laboratories, the bacterium abscessus and Mycobacterium chelonae [1]. BACTEC 9240 blood culture system (Becton Dickinson Received: September 21, 2009 Revised: March 8, 2010 Accepted: April 7, 2010 Address for correspondence: Giueng-Chueng Wang, Department of Laboratory Medicine, Taipei Medical University-Wan Fang Hospital, No. 111, Section 3, Hsing-Long Road, Taipei, Taiwan, R.O.C. TEL:0968-745941 E-mail: [email protected] 70 J Biomed Lab Sci 2010 Vol 22 No 2 Diagnostic Instrument Systems, Sparks, Md.) is one of following microbiological and biochemical reactions the automated, continuous-monitoring and widely ex- were applied. There was growth at 3 days on Löwen- ploited systems [10]. It uses noninvasive fluorescent stein-Jensen slant medium, individual colonies were patented technology to detect increases in CO2 produced smooth and buff colored in the dark and after exposure by microbial growth. Each bottle contains a fluorescent to light. The isolate was positive for arylsulfatase, ni- CO2 sensor which is monitored every ten minutes for trate, 5% NaCl and urease and negative for niacin and increases in fluorescent appearance. Computer algo- Tween 80 hydrolysis. rithms determine whether sustained linear increases or increasing rates of change in fluorescent indicate micro- bial growth. Commercial bottles are available for every variety of clinical use. For instance, BD BACTEC Plus Aerobic/F and Plus Anaerobic/F media are used in a qualitative procedure for the aerobic and anaerobic cul- ture respectively. Generally speaking, when blood cul- ture bottles were positive, removed them from the BACTEC instrument, the contents were gently mixed, direct Gram staining of the blood culture fluid, and some of the fluid was inoculated onto a combination of agar plates, suited for culturing aerobic, anaerobic, and fas- tidious microorganisms. In this case report, subsequently Fig. 1. Mycobacterium fortuitum (Gram stain 1000x) revealed mycobacteria culture should be considerated whenever gram-positive rods suspicion of mycobacterial infection. Finally, identifica- tion of the microorganism and determination of its sus- ceptibility pattern. Case Report We describe a case of an 89-year-old male patient with poor-controlled diabetes who was bedridden and under- went a long-term antibiotics and steroid therapy docu- mented M. fortuitum bacteremia in April 2009. This pa- tient had a prolonged hospitalization for two years be- cause of repeated episodes of pulmonary edema and nosocomial pneumonia. Two sets of blood culture were Fig. 2. Mycobacterium fortuitum ( Ziehl-Neelsen stain1000x) collected appropriately, incubated in BACTEC 9240 showed acid-fast bacilli blood culture system and were positive after 48 hours of incubation. Gram stain of these positive blood culture specimens revealed Gram-positive rods (Figure 1), which were preferred spore forming bacilli than cocci. Due to no microbial growth on aerobic agar plates in the first two days, we tried to perform Ziehl-Neelsen stain and unexpectedly found the acid-fast bacilli (Figure 2). For further identification, mycobacteria culture was fol- lowed by submitting the positive culture broth to Löwen- stein-Jensen slant medium. The individual colonies dis- played smooth and buff-color appearance both on aero- bic agar plates (Figure 3) and Löwenstein-Jensen slant medium (Figure 4) in the following two and three days. The microorganism was subsequently identified as M. Fig. 3. Smooth and buff-color appearance of Mycobacterium fortuitum by traditional biochemical method [11]. The fortuitum colonies grew on aerobic sheet blood agar plates J Biomed Lab Sci 2010 Vol 22 No 2 71 Isolation of Mycobacterium fortuitum from BACTEC 9240 Blood Culture System In view of the predilection groups of mycobacteria infection, it is essential to administer appropriate medi- cal treatment, and therefore the susceptibility testing for mycobacteria is extremely recommended. References 1. Brown-Elliott, B. A., and R. J. Wallace, Jr.: Clinical and taxonomic status of pathogenic nonpigmented or late-pigmenting rapidly growing mycobacteria. Clin. Mi- crobiol. Rev 2002; 15: 716–746. Fig. 4. The individual colonies of Mycobacterium fortuitum were 2. Wallace, R. J., Jr., B. A. Brown-Elliott, and C. Turenne: smooth and buff colored on Löwenstein-Jensen medium Clinical and laboratory features of Mycobacterium por- cinum. J. Clin. Microbiol 2004; 42: 5689–5697. 3. Corrado Serra, Giovanni Loi, Barbara Saddi, Marisa Pautasso, and Aldo Manzin: Unusual Clinical Presenta- Discussion tion of Mycobacterium fortuitum Infection in an Im- munocompetent Woman. J. Clin. Microbiol 2007; 45: 1663–1665. This is the first case of M. fortuitum isolation from 4. Toı¨di Ade´kambi, Andre´as Stein, Joseph Carvajal, positive blood culture bottle in the laboratory of Taipei Didier Raoult, and Michel Drancourt: Description of My- Medical University- Wan Fang Hospital. According to cobacterium conceptionense sp. nov., a Mycobacterium the manufacturer’s instructions, samples of blood culture fortuitum Group Organism Isolated from a Posttraumatic bottle are going to be asserted negative if BACTEC Osteitis Inflammation. J. Clin. Microbiol 2006; 44: 9240 does not detect the appearance of fluorescence in 1268–1273. 5. M P A Lessing, M M Walker: Fatal pulmonary infection culture bottles within six days. M. fortuitum, a member due to Mycobacterium fortuitum. J Clin Pathol 1993; 46: of rapidly growing Runyon IV NTM, has the property to 271-272. grow on standard mycobacterial media within seven 6. Hsin-Hung Wu, Rong-Luh Lin, Chao-Hsien Lee, days and this property may contribute to the primary Ming-Jen Peng, Chien-Liang Wu: Mycobacterium For- positive culture in blood sample culture system in this tuitum Bacteremia and Pulmonary Disease in a Hemo- case. dialysis Patient. Thorac Med 2007; 22: 332-337. This finding has highlighted that M. fortuitum is not 7. Zainal Muttakin A R, Tan A M: Mycobacterium fortuitum only a well-known acid-fast bacillus, but can also forms catheter-related sepsis in acute leukaemia. Singapore Med J 2006; 47: 543-545. diphtheroids, Gram positive rods appearance, under 8. Samuel C M Huang, H Kaz Soong, Jen-Shiang Chang, Gram stain microscopy [1]. Consequently, clinical tech- Yu-Sung Liang: Non-tuberculous