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Mycobacterium malmoense in Italy
Tubercle and Lung Disease (1995) 76, 171-I 72 © 1995 Pearson Professional Ltd
Tubercleand Lung Disease
Mycobacterium malmoense in Italy C. Piersimoni*, L. Felici t, V. Penati*, C. Lacchini*
*Department of Clinical Microbiology, General Hospital 'Umberto I°-Torrette', Ancona, Italy, tDepartment of Pediatrics, University of Ancona, Ancona, Italy, ~'Villa Marelli', Institute for Chest and Lung Disease, Milan, Italy S U M M A R Y. A case of mycobacterial lymphadenitis due to Mycobacterium malmoense was recently diagnosed in a 5-year-old girl. The organism was isolated from pus and tissue fragments obtained by surgical excision of the affected nodes. This is the first documented case of h u m a n infection due to this organism in Italy.
R E S U Mfi~. Un cas de lymphad~nite mycobact6rienne due ~ Mycobacterium malmoense a r~cemment 6t~ diagnostiqu~ chez une fillette fig~e de 5 ans. Le germe a ~t~ isol~ du pus et des fragments tissulaires obtenus par excision chirurgicale des ganglions concern~s. C'est le premier cas document~ d'infection humaine due cet organisme en Italie. R E S U M E N. Se diagnostic6 recientemente un caso de linfoadenitis micobacteriana debida a Mycobacterium malmoense en una nifia de 5 afios de edad. El microorganismo fue aislado del pus y de fragmentos de tejido obtenidos quirfirgicamente de los ganglios afectados. Es el primer caso documentado de infecci6n humana debida a este germen en Italia.
Infection caused by non-tuberculous mycobacteria (NTM) is on the increase in many developed countries. 1,2 Mycobacterium malmoense was first described as a new pathogenic mycobacterial species in 1977. 3 Since then an increasing number of cases have been reported from the UK, 4-6 North America, 7,8 Sweden, 9 Finland, I° Switzerland, H France 12 and A u s t r i a . 13 To our knowledge no cases of disease due to M. malmoense have been documented in Italy. In this paper we report the isolation and discuss details of the clinical outcome and identification of M. malmoense from a 5-year-old child with mycobacterial lymphadenitis.
2 X 3.5 cm oval masses, one near the angle of the jaw and the other in the preauricular area. Routine laboratory findings and a standard chest radiograph were normal. Skin tests with atypical mycobacterial antigens (Statens Seruminstitut, Copenhagen, DK), including PPD-A (M. avium), PPD-G (M. scofuIaceum), PPD-Y (M. kansasii), PPD-F (M. fortuitum) and a standard tuberculin antigen (PPD-S), resulted in the following reactions (mm of induration): PPD-S, 6 mm; PPD-A, 17 mm; PPD-G, 15 mm; PPD-Y, 9 mm; PPD-F, 10 mm. Chemotherapy with rifampicin and isoniazid was started, followed a few days later by partial excision and di'ainage of the submandibular nodes. Histological examination of the tissue fragments obtained by surgical excision revealed granulomas containing central necrosis and Langhans' giant cells. Tissue fragments and pus were homogenized in sterile saline in tissue grinders and the suspension decontaminated with NaOH (final concentration 2%), centrifuged and neutralized according to standard procedures. 14 The sediment was inoculated onto two L-J slants and into a Middlebrook 12B medium bottle (Becton-Dickinson, Sparks, USA). Both media grew mycobacteria: acid-fast bacilli were observed in liquid 12B medium after 9 days and thin colonies appeared on L-J medium after 26 days. Subcultures on L-J medium were slow-growing and remained colourless after exposure to light.
CASE R E P O R T In March 1991, a 5-year-old girl was referred to the surgical department of our hospital with right neck adenitis and fever. The child was living in a country area and had not received any prior BCG vaccination. Similarly her parents did not report any history of tuberculosis or other mycobacterial infections. Physical examination was unremarkable except for two firm, Correspondence to: C. Piersimoni MD, Department of Clinical Microbiology,General Hospital 'UmbertoI°-Torrette', via Conca, 60020 Ancona,Italy. 171
172 Tubercle and Lung Disease In biochemical tests TM the mycobacterium gave negative results for niacin production, nitrate reductase and 3-day arylsulfatase. No growth was observed on L-J medium containing 5% sodium chloride and on MacConkey agar without crystal violet. The mycobacterium showed catalase activity (< 45 m m of foam) at 37°C and retained slight catalase activity after heating at 68°C for 20 min. The urease test was positive and the organism was able to hydrolyze Tween 80 with a markedly positive reaction within 2 days. Lipid analysis performed on this isolate by P. A. Jenkins, Mycobacterium Reference Unit (PHLS), confirmed that the culture was M. malmoense. Antimicrobial susceptibility tests were performed using the Bactec radiometric system. 15 Broth determined MICs showed that the strain was susceptible to ethambutol (4 mcg/mi), rifabutin (0.05 mcg/ml), clarithromycin ( l m c g / m l ) and sparfloxacin (0.5mcg/ml); it was moderately susceptible to ciprofloxacin (4 mcg/ml) and rifampicin (1 mcg/mi) and resistant to isoniazid (> 2.0 gg/ ml), streptomycin (8 gg/ml) and PAS (> 2.0 gg/ml). The patient was treated with antituberculosis therapy (initially isoniazid plus rifampicin and subsequently rifampicin and ethambutol) for 6 months after surgery. She developed a draining sinus at the site of excision, followed by healing 4 months later. Progressive reduction of the preauricolar node was observed during chemotherapy. At present the patient feels well and the affected nodes appear to have completely healed.
DISCUSSION The isolation of mycobacteria from a sterile body site indicates infection and there is no reason to suspect contamination? To our knowledge this report is the first documented case of disease due to M. malmoense in Italy. In our report lipid analysis was used in addition to standard criteria to identify the strain. Since this method is not presently available in Italy, M. malmoense may have been misidentified in the past or even missed completely. When N T M lymphadenitis occurs, medical therapy with the presently available drugs is not usually helpful; the most reliable treatment is surgical excision of the involved nodes. 2 Nevertheless in this case chemotherapy, administered according to in vitro susceptibility
data, promoted regression of the enlarged node in the preauricular area and enhanced healing at the site of surgical drainage. Although more extensive study is necessary before recommendations on effective treatment of M. malmoense disease can be made, ~6 it seems reasonable that in vitro susceptibility tests may be useful to indicate the drug therapy of choice and to predict the clinical outcome.
References 1. SaubolleM A. Nontuberculousmycobacteriaas agents of human disease in the United States. Clin MicrobiolNewsl 1989; 11: 113-117. 2. WolinskyE. Mycobacterialdisease other than tuberculosis. Clin Infect Dis 1992; 15: 1-12. 3. Scr6derK H, Juhlin I. Mycobacterium malmoense sp. nov. Int J Sistem Bacteriol 1977; 27: 241-246. 4. Banks J, Jenkins P A, Smith A P. Pulmonaryinfectionwith Mycobacterium malmoense - a review of treatment and response. Tubercle 1985; 66: 197-203. 5. Whit M P, Bangash H, Goel K M, Jenkins P A. Non-tuberculous mycobacteriallymphadenitis.Arch Dis Child 1986; 61: 368-371. 6. France A J, McLeod D T, Calder M A, Seaton A. Mycobacterium malmoense infectionsin Scotland: an increasing problem. Thorax 1987; 42: 593-595. 7. Warren N G, Body B A, Silcox V A, Manhews J H. Pulmonary disease due to Mycobacterium malmoense. J Clin Microbiol 1984; 20: 245-247. 8. Alberts W M, ChandlerK W, SolomonD A, GoldmanA L. Pulmonarydisease caused by Mycobacterium malmoense. Am Rev Respir Dis 1987; 135: 1375-1378. 9. Hoffner S E, HenriquesB, Petrini B, K~illeniusG. Mycobacterium malmoense: an easily missed pathogen. J Clin MicrobioI 1991; 29: 2673-2674. 10. Katila M L, Brander E, Vijanen T. Difficultywith Mycobacterium malmoense. Lancet 1989; ii: 510-511. 11. Zangg M, SalfingerM, OpravilM, Ltithy R. Extrapulmonaryand disseminatedinfectionsdue to Mycobacterium malmoense: case report and review. Clin Infect Dis 1993; 16: 40-49. 12. BurucoaC, BourgoinA, Castets Met al. Pulmonaryinfection due to Mycobacterium malmoense in an immunocompromised patient. Clin MicrobiolNewsl 1993; 15: 86-87. 13. AllerbergerF, Fille M, Dierich M. Letter to the editor. Clin Microbiol Newsl 1993; 15: 191-192. 14. Roberts G D, KonemanE W, Kim Y K. Mycobacterium.In: Balows A, HanslerW J Jr, HerrmannK L, IsembergH D, Shadomy H J, eds. Manual of clinical microbiology,5th ed. WashingtonDC: AmericanSociety for Microbiology, 1991: 304-339. 15. Hoffner S E, Hjelm U, K~illeniusG. Susceptibilityof Mycobacterium malmoense to antibacterialdrugs maddrug combinations.AntimicrobAgents Chemother 1993; 37: 1285-1288. 16. Wallace R J Jr, O'Brien R, GlassrothJ, Raleigh J, Dutt A. Diagnosis and treatment of disease caused by nontuberculous mycobacteria. Am Rev Respir Dis 1990; 142: 940-953.
Tubercleand Lung Disease
Mycobacterium malmoense in Italy C. Piersimoni*, L. Felici t, V. Penati*, C. Lacchini*
*Department of Clinical Microbiology, General Hospital 'Umberto I°-Torrette', Ancona, Italy, tDepartment of Pediatrics, University of Ancona, Ancona, Italy, ~'Villa Marelli', Institute for Chest and Lung Disease, Milan, Italy S U M M A R Y. A case of mycobacterial lymphadenitis due to Mycobacterium malmoense was recently diagnosed in a 5-year-old girl. The organism was isolated from pus and tissue fragments obtained by surgical excision of the affected nodes. This is the first documented case of h u m a n infection due to this organism in Italy.
R E S U Mfi~. Un cas de lymphad~nite mycobact6rienne due ~ Mycobacterium malmoense a r~cemment 6t~ diagnostiqu~ chez une fillette fig~e de 5 ans. Le germe a ~t~ isol~ du pus et des fragments tissulaires obtenus par excision chirurgicale des ganglions concern~s. C'est le premier cas document~ d'infection humaine due cet organisme en Italie. R E S U M E N. Se diagnostic6 recientemente un caso de linfoadenitis micobacteriana debida a Mycobacterium malmoense en una nifia de 5 afios de edad. El microorganismo fue aislado del pus y de fragmentos de tejido obtenidos quirfirgicamente de los ganglios afectados. Es el primer caso documentado de infecci6n humana debida a este germen en Italia.
Infection caused by non-tuberculous mycobacteria (NTM) is on the increase in many developed countries. 1,2 Mycobacterium malmoense was first described as a new pathogenic mycobacterial species in 1977. 3 Since then an increasing number of cases have been reported from the UK, 4-6 North America, 7,8 Sweden, 9 Finland, I° Switzerland, H France 12 and A u s t r i a . 13 To our knowledge no cases of disease due to M. malmoense have been documented in Italy. In this paper we report the isolation and discuss details of the clinical outcome and identification of M. malmoense from a 5-year-old child with mycobacterial lymphadenitis.
2 X 3.5 cm oval masses, one near the angle of the jaw and the other in the preauricular area. Routine laboratory findings and a standard chest radiograph were normal. Skin tests with atypical mycobacterial antigens (Statens Seruminstitut, Copenhagen, DK), including PPD-A (M. avium), PPD-G (M. scofuIaceum), PPD-Y (M. kansasii), PPD-F (M. fortuitum) and a standard tuberculin antigen (PPD-S), resulted in the following reactions (mm of induration): PPD-S, 6 mm; PPD-A, 17 mm; PPD-G, 15 mm; PPD-Y, 9 mm; PPD-F, 10 mm. Chemotherapy with rifampicin and isoniazid was started, followed a few days later by partial excision and di'ainage of the submandibular nodes. Histological examination of the tissue fragments obtained by surgical excision revealed granulomas containing central necrosis and Langhans' giant cells. Tissue fragments and pus were homogenized in sterile saline in tissue grinders and the suspension decontaminated with NaOH (final concentration 2%), centrifuged and neutralized according to standard procedures. 14 The sediment was inoculated onto two L-J slants and into a Middlebrook 12B medium bottle (Becton-Dickinson, Sparks, USA). Both media grew mycobacteria: acid-fast bacilli were observed in liquid 12B medium after 9 days and thin colonies appeared on L-J medium after 26 days. Subcultures on L-J medium were slow-growing and remained colourless after exposure to light.
CASE R E P O R T In March 1991, a 5-year-old girl was referred to the surgical department of our hospital with right neck adenitis and fever. The child was living in a country area and had not received any prior BCG vaccination. Similarly her parents did not report any history of tuberculosis or other mycobacterial infections. Physical examination was unremarkable except for two firm, Correspondence to: C. Piersimoni MD, Department of Clinical Microbiology,General Hospital 'UmbertoI°-Torrette', via Conca, 60020 Ancona,Italy. 171
172 Tubercle and Lung Disease In biochemical tests TM the mycobacterium gave negative results for niacin production, nitrate reductase and 3-day arylsulfatase. No growth was observed on L-J medium containing 5% sodium chloride and on MacConkey agar without crystal violet. The mycobacterium showed catalase activity (< 45 m m of foam) at 37°C and retained slight catalase activity after heating at 68°C for 20 min. The urease test was positive and the organism was able to hydrolyze Tween 80 with a markedly positive reaction within 2 days. Lipid analysis performed on this isolate by P. A. Jenkins, Mycobacterium Reference Unit (PHLS), confirmed that the culture was M. malmoense. Antimicrobial susceptibility tests were performed using the Bactec radiometric system. 15 Broth determined MICs showed that the strain was susceptible to ethambutol (4 mcg/mi), rifabutin (0.05 mcg/ml), clarithromycin ( l m c g / m l ) and sparfloxacin (0.5mcg/ml); it was moderately susceptible to ciprofloxacin (4 mcg/ml) and rifampicin (1 mcg/mi) and resistant to isoniazid (> 2.0 gg/ ml), streptomycin (8 gg/ml) and PAS (> 2.0 gg/ml). The patient was treated with antituberculosis therapy (initially isoniazid plus rifampicin and subsequently rifampicin and ethambutol) for 6 months after surgery. She developed a draining sinus at the site of excision, followed by healing 4 months later. Progressive reduction of the preauricolar node was observed during chemotherapy. At present the patient feels well and the affected nodes appear to have completely healed.
DISCUSSION The isolation of mycobacteria from a sterile body site indicates infection and there is no reason to suspect contamination? To our knowledge this report is the first documented case of disease due to M. malmoense in Italy. In our report lipid analysis was used in addition to standard criteria to identify the strain. Since this method is not presently available in Italy, M. malmoense may have been misidentified in the past or even missed completely. When N T M lymphadenitis occurs, medical therapy with the presently available drugs is not usually helpful; the most reliable treatment is surgical excision of the involved nodes. 2 Nevertheless in this case chemotherapy, administered according to in vitro susceptibility
data, promoted regression of the enlarged node in the preauricular area and enhanced healing at the site of surgical drainage. Although more extensive study is necessary before recommendations on effective treatment of M. malmoense disease can be made, ~6 it seems reasonable that in vitro susceptibility tests may be useful to indicate the drug therapy of choice and to predict the clinical outcome.
References 1. SaubolleM A. Nontuberculousmycobacteriaas agents of human disease in the United States. Clin MicrobiolNewsl 1989; 11: 113-117. 2. WolinskyE. Mycobacterialdisease other than tuberculosis. Clin Infect Dis 1992; 15: 1-12. 3. Scr6derK H, Juhlin I. Mycobacterium malmoense sp. nov. Int J Sistem Bacteriol 1977; 27: 241-246. 4. Banks J, Jenkins P A, Smith A P. Pulmonaryinfectionwith Mycobacterium malmoense - a review of treatment and response. Tubercle 1985; 66: 197-203. 5. Whit M P, Bangash H, Goel K M, Jenkins P A. Non-tuberculous mycobacteriallymphadenitis.Arch Dis Child 1986; 61: 368-371. 6. France A J, McLeod D T, Calder M A, Seaton A. Mycobacterium malmoense infectionsin Scotland: an increasing problem. Thorax 1987; 42: 593-595. 7. Warren N G, Body B A, Silcox V A, Manhews J H. Pulmonary disease due to Mycobacterium malmoense. J Clin Microbiol 1984; 20: 245-247. 8. Alberts W M, ChandlerK W, SolomonD A, GoldmanA L. Pulmonarydisease caused by Mycobacterium malmoense. Am Rev Respir Dis 1987; 135: 1375-1378. 9. Hoffner S E, HenriquesB, Petrini B, K~illeniusG. Mycobacterium malmoense: an easily missed pathogen. J Clin MicrobioI 1991; 29: 2673-2674. 10. Katila M L, Brander E, Vijanen T. Difficultywith Mycobacterium malmoense. Lancet 1989; ii: 510-511. 11. Zangg M, SalfingerM, OpravilM, Ltithy R. Extrapulmonaryand disseminatedinfectionsdue to Mycobacterium malmoense: case report and review. Clin Infect Dis 1993; 16: 40-49. 12. BurucoaC, BourgoinA, Castets Met al. Pulmonaryinfection due to Mycobacterium malmoense in an immunocompromised patient. Clin MicrobiolNewsl 1993; 15: 86-87. 13. AllerbergerF, Fille M, Dierich M. Letter to the editor. Clin Microbiol Newsl 1993; 15: 191-192. 14. Roberts G D, KonemanE W, Kim Y K. Mycobacterium.In: Balows A, HanslerW J Jr, HerrmannK L, IsembergH D, Shadomy H J, eds. Manual of clinical microbiology,5th ed. WashingtonDC: AmericanSociety for Microbiology, 1991: 304-339. 15. Hoffner S E, Hjelm U, K~illeniusG. Susceptibilityof Mycobacterium malmoense to antibacterialdrugs maddrug combinations.AntimicrobAgents Chemother 1993; 37: 1285-1288. 16. Wallace R J Jr, O'Brien R, GlassrothJ, Raleigh J, Dutt A. Diagnosis and treatment of disease caused by nontuberculous mycobacteria. Am Rev Respir Dis 1990; 142: 940-953.