Journal of Infection (I99I) 23, 303-306
CASE REPORT Mycobacterium
h a e m o p h i l u m i n f e c t i o n in a p a t i e n t w i t h AIDS
J. Holton,*~ P. Nye* and R. MillerT
Departments of * Microbiology and t Medicine, University College and Middlesex School of Medicine, London, U.K. Accepted for publication 2.2 April r99I Summary We report the isolation of a nutritionally fastidious mycobacterial species, Mycobacterium haemophilum, from the skin of a patient with AIDS. The diagnosis of the mycobacterial nature of the patient's lesions was complicated by extensive Kaposi's sarcoma. It is recommended that slowly growing fastidious mycobacteria be sought in any patients with unusual skin lesions.
Introduction Patients infected with the human immunodeficiency viruses are prone to a variety of infectious agents. Many are opportunist pathogens which gain an advantage because of the waning cellular immunity. Opportunist infections are believed to be responsible for almost 9o % HIV-related deaths. 1 Of the various pathogens responsible for opportunist infections a significant proportion are Mycobacterial species. 2 In African countries Mycobacterium tuberculosis is found in up to 3o % HIV-positive individuals, a In America and Europe, the predominant Mycobacterial species isolated is Mycobacterium avium-intracellulare, where up to 5o ~/o patients may be colonised or infected. 4 Other species may also cause disease and occasionally the mycobacterium may have special growth requirements or even be impossible to isolate in culture, 5 despite the use of a variety of media or injection into mice. Mycobacterium haemophilum is a slowly growing mycobacterium that was described in I978 by Sompolinsky 6 as requiring haemin for growth. Other strains have been isolated which require ferric ammonium citrate. 7 T h e taxonomic status of this organism is uncertain as only a few isolates have been examined. It does have biochemical similarities to M. avium-intracellulare and Mycobacterium xenopi, in being niacin-, nitrate- and urease-negative; nicotinamidase and pyrazinamidase-positive. Mycobacterium haemophilum grows at 25-30 °C (unlike M. xenopi) but not at 37-42 °C (unlike M. aviumintracellulare). Mycobacterium haemophilum does not hydrolyse Tween, does not produce catalase and is resistant to isoniazid. We report an infection caused by M. haemophilum in the skin of a patient with A I D S who also had extensive Kaposi's sarcoma. ~: Address correspondence to: Dr J. Holton, Department of Microbiology, University College and Middlesex School of Medicine, 67-73 Ridinghouse Street, London Wx, U.K. oi63-4453/9I/o6o3o3+o4 $o3.oo/o
© I99I The British Society for the Study of Infection
304
J. HOLTON
ET AL.
Case report
A 37-year-old Caucasian homosexual male presented in June I99o with a painful and swollen calf. He had been found to be H I V - I antibody positive in November I987 and had experienced an episode of Pneumocystis carinii pneumonia in February 1988. Since that time he had had recurrent peri-anal herpetic ulcers and was receiving long-term cotrimoxazole and acyclovir prophylaxis. He also had recurrent oropharyngeal candidiasis for which he was receiving fluconazole prophylaxis. He first noticed lesions of Kaposi's sarcoma on his trunk in October I989 and subsequently many new lesions appeared on his face, trunk and limbs. Those on the patient's foot and calf resembled the Kaposi's sarcoma lesions elsewhere on his body but were painful and exquisitely tender when touched. Examination revealed a large 8 x 9 cm tender lesion on the sole of the foot. In addition there were several raised and tender nodules along the medial edge of the gastrocnemius muscle. There was no associated inguinal lymphadenopathy. T h e patient was pyrexial 38"5 °C and investigations revealed a haemoglobin of 9"6 g/d1, WBC count of 4"8 x iog/1 (granulocytes = 93 %, lymphocytes = 5%, m o n o c y t e s - - 2 % ) , urea and electrolytes and liver function tests were normal. Cultures of blood and urine were negative for mycobacteria and other micro-organisms. T h e lesions were treated with radiotherapy, 16oo centigray in four fractions over 6 days to the sole of the foot and 60o centigray at one session to each of the lesions on the calf. This brought about a rapid reduction in pain and swelling of the foot and calf. Two to 4 weeks after irradiation the lesion on the sole of the foot ulcerated. At this time a skin biopsy was taken for culture and histology. Methods
T h e biopsy material was homogenised, decontaminated with 5 % oxalic acid for 20 min and neutralised with phosphate buffer pH 6"8. It was inoculated on to Lowenstein-Jensen (L J) medium and LJ medium containing I5 mg/1 of ferric ammonium citrate (FAC) and incubated at 3o ° and 35 °C. T h e isolate was subcultured on to chocolate agar and Columbia agar base (CBA) upon which an X factor (haemin) disc was placed. T h e y were incubated at 30 ° and 35 °C. Results
A Ziehl-Nielsen stain of the biopsy revealed numerous acid-fast bacilli and the histology was reported as typical tuberculoid granuloma. After 8 weeks, growth occurred on the FAC medium incubated at 3o °C but not at 35 °C. There was no growth on the unsupplemented LJ medium. T h e growth was smooth and buff-coloured and a Ziehl-Nielsen stain of the colonies demonstrated strongly acid fast bacilli and cord formation, the morphology of the organism resembling 34. tuberculosis. Upon subculture, growth occurred after 2 weeks at 3o °C but not at 35 °C on the chocolate agar and the CBA medium in the region of the X disk. T h e
M. h a e m o p h i l u m infection in A I D S
305
growth was dry and rugose, quite dissimilar from the growth obtained on F A C m e d i u m on primary isolation. T h e organism was identified as M. haemophilum by the Mycobacterial Reference Laboratory, Cardiff. It was reported to be resistant to streptomycin, isoniazid, ethambutol, ciprofloxacin, gentamicin, augmentin, ceftazidime and sulphamethoxazole, and sensitive to rifampicin, erythromycin, imipenem, minocycline, doxycycline and amikacin. T h e patient was started on treatment with minocycline and rifampicin but suffered vestibular side-effects from the minocycline and was changed to erythromycin and rifampicin. T h e ulcerated lesion began to heal after several weeks. T h e nodules slowly resolved and no more have developed. T h e patient remains apyrexial and well. Discussion
In ~978 Sompolinsky isolated a Mycobacterial species from the skin of an i m m u n o c o m p r o m i s e d patient which required haemin for growth and which he called M. haemophilum. Subsequently Dawson demonstrated the identity of Sompolinsky's isolate to previous isolates from the skin of other i m m u n o compromised p a t i e n t s / T h e species of Mycobacterium isolated from the skin of the patient reported here only grew u p o n Lowenstein-Jensen m e d i u m with ferric a m m o n i u m citrate as a supplement. Its growth was also demonstrated to be dependent u p o n haemin. T h e cultural characteristics of this isolate on F A C and CBA media were similar to previous isolates of M. haemophilum, 8 as was its temperature-dependence. T h e isolate was identified as M. haemophilum by the Mycobacterial Reference Laboratory. T h e presentation in this patient was complicated by extensive Kaposi's sarcoma u p o n the trunk and limbs. T h e skin lesion on the foot was a flat area, with sporotrichoid spread up the calf in the form of painful nodules. T h e histology of a tuberculoid granuloma was consistent with previously reported cases. T h e site of the lesion is consistent with the infection occurring after an inoculation injury to the foot, although the patient did not remember any such event. After radiotherapy the inflammation subsided, presumably due to the death of cells mediating the inflammatory response. This is consistent with the recognised effects of radiation on the inflammatory process. T h e subsequent ulceration of the irradiated lesion is likely to have been a further effect of the radiation u p o n the local cellular i m m u n e defence mechanisms allowing mycobacterial proliferation and ulceration. Once the mycobacterial nature of the lesion was recognised, the patient was started on appropriate chemotherapy and the lesions slowly regressed. T h e r e have been a n u m b e r of reports recently of slowly growing on or nutritionally exacting organisms being isolated from patients with AIDS.9' 10 A high index of suspicion for such organisms must be maintained in these patients. We would r e c o m m e n d that, particularly with skin biopsies, there should be prolonged culture of the specimen on a range of media, including LJ m e d i u m supplemented with ferric a m m o n i u m citrate, in order to maximise the chance of isolating nutritionally fastidious mycobacteria.
306
j. H O L T O N E T AL.
References I. Masur H. Problems in the management of opportunistic infections in patients infected with human immunodeficiency virus. J Infect Dis I99o; I6X : 858-864. 2. Clumeck N, Hermans P, DeWits, S. Current problems in the management of A I D S patients, Eur J Clin Microbiol Infect Dis r988; 7: 2-IO. 3. Harries AD. Tuberculosis and human immunodeficiency virus infection in developing countries. Lancet I99O; 335: 387-390. 4- Guthertz LS, Damsker B, Bottone EJ, Ford EG, Midura T F , Janda MJ. Mycobaeterium avium and Mycobacterium intercellulare infections in patients with and without A I D S . J Infect Dis I989; I6o: IO37-IO4I. 5. Hirschel B, Chang H R , Mach N e t al. Fatal infection with a novel unidentified mycobacterium in a man with A I D S . N EnglJ Med I99o; 3z3: I o 9 - I I 3 . 6. Sompolinsky D, Lagziel A, Neveh D, Yonkilevitz L. Myeobacterium haemophilum sp.nov. a new pathogen from humans. Int J Syst Bacteriol I978; z8: 67-75. 7- Dawson D J, Jennis F. Mycobacteria with a growth requirement for ferric ammonium citrate identified as Mycobacterium haemophilum. J Clin Microbiol I98O; I I : 19o--I92. 8. Moulsdale M T , Harper J, Thatcher GN. Infection by Mycobacterium haemophilum, a metabolically fastidious acid fast bacillus. Tubercle I983 ; 64:24-36. 9. Holton J, Miller R, Furst V, Malnick H. Isolation of Protomonas extorquens (the Red Phantom) from a patient with A I D S . J Infect I99O; zx : 87-93. io. Slater L N , Welch D F , Hensel D, Goody DW. A newly recognized fastidious Gramnegative pathogen as a cause of fever and bacteremia. N EnglJ Med I99O; 323: I587-I592.
CASE REPORT Mycobacterium
h a e m o p h i l u m i n f e c t i o n in a p a t i e n t w i t h AIDS
J. Holton,*~ P. Nye* and R. MillerT
Departments of * Microbiology and t Medicine, University College and Middlesex School of Medicine, London, U.K. Accepted for publication 2.2 April r99I Summary We report the isolation of a nutritionally fastidious mycobacterial species, Mycobacterium haemophilum, from the skin of a patient with AIDS. The diagnosis of the mycobacterial nature of the patient's lesions was complicated by extensive Kaposi's sarcoma. It is recommended that slowly growing fastidious mycobacteria be sought in any patients with unusual skin lesions.
Introduction Patients infected with the human immunodeficiency viruses are prone to a variety of infectious agents. Many are opportunist pathogens which gain an advantage because of the waning cellular immunity. Opportunist infections are believed to be responsible for almost 9o % HIV-related deaths. 1 Of the various pathogens responsible for opportunist infections a significant proportion are Mycobacterial species. 2 In African countries Mycobacterium tuberculosis is found in up to 3o % HIV-positive individuals, a In America and Europe, the predominant Mycobacterial species isolated is Mycobacterium avium-intracellulare, where up to 5o ~/o patients may be colonised or infected. 4 Other species may also cause disease and occasionally the mycobacterium may have special growth requirements or even be impossible to isolate in culture, 5 despite the use of a variety of media or injection into mice. Mycobacterium haemophilum is a slowly growing mycobacterium that was described in I978 by Sompolinsky 6 as requiring haemin for growth. Other strains have been isolated which require ferric ammonium citrate. 7 T h e taxonomic status of this organism is uncertain as only a few isolates have been examined. It does have biochemical similarities to M. avium-intracellulare and Mycobacterium xenopi, in being niacin-, nitrate- and urease-negative; nicotinamidase and pyrazinamidase-positive. Mycobacterium haemophilum grows at 25-30 °C (unlike M. xenopi) but not at 37-42 °C (unlike M. aviumintracellulare). Mycobacterium haemophilum does not hydrolyse Tween, does not produce catalase and is resistant to isoniazid. We report an infection caused by M. haemophilum in the skin of a patient with A I D S who also had extensive Kaposi's sarcoma. ~: Address correspondence to: Dr J. Holton, Department of Microbiology, University College and Middlesex School of Medicine, 67-73 Ridinghouse Street, London Wx, U.K. oi63-4453/9I/o6o3o3+o4 $o3.oo/o
© I99I The British Society for the Study of Infection
304
J. HOLTON
ET AL.
Case report
A 37-year-old Caucasian homosexual male presented in June I99o with a painful and swollen calf. He had been found to be H I V - I antibody positive in November I987 and had experienced an episode of Pneumocystis carinii pneumonia in February 1988. Since that time he had had recurrent peri-anal herpetic ulcers and was receiving long-term cotrimoxazole and acyclovir prophylaxis. He also had recurrent oropharyngeal candidiasis for which he was receiving fluconazole prophylaxis. He first noticed lesions of Kaposi's sarcoma on his trunk in October I989 and subsequently many new lesions appeared on his face, trunk and limbs. Those on the patient's foot and calf resembled the Kaposi's sarcoma lesions elsewhere on his body but were painful and exquisitely tender when touched. Examination revealed a large 8 x 9 cm tender lesion on the sole of the foot. In addition there were several raised and tender nodules along the medial edge of the gastrocnemius muscle. There was no associated inguinal lymphadenopathy. T h e patient was pyrexial 38"5 °C and investigations revealed a haemoglobin of 9"6 g/d1, WBC count of 4"8 x iog/1 (granulocytes = 93 %, lymphocytes = 5%, m o n o c y t e s - - 2 % ) , urea and electrolytes and liver function tests were normal. Cultures of blood and urine were negative for mycobacteria and other micro-organisms. T h e lesions were treated with radiotherapy, 16oo centigray in four fractions over 6 days to the sole of the foot and 60o centigray at one session to each of the lesions on the calf. This brought about a rapid reduction in pain and swelling of the foot and calf. Two to 4 weeks after irradiation the lesion on the sole of the foot ulcerated. At this time a skin biopsy was taken for culture and histology. Methods
T h e biopsy material was homogenised, decontaminated with 5 % oxalic acid for 20 min and neutralised with phosphate buffer pH 6"8. It was inoculated on to Lowenstein-Jensen (L J) medium and LJ medium containing I5 mg/1 of ferric ammonium citrate (FAC) and incubated at 3o ° and 35 °C. T h e isolate was subcultured on to chocolate agar and Columbia agar base (CBA) upon which an X factor (haemin) disc was placed. T h e y were incubated at 30 ° and 35 °C. Results
A Ziehl-Nielsen stain of the biopsy revealed numerous acid-fast bacilli and the histology was reported as typical tuberculoid granuloma. After 8 weeks, growth occurred on the FAC medium incubated at 3o °C but not at 35 °C. There was no growth on the unsupplemented LJ medium. T h e growth was smooth and buff-coloured and a Ziehl-Nielsen stain of the colonies demonstrated strongly acid fast bacilli and cord formation, the morphology of the organism resembling 34. tuberculosis. Upon subculture, growth occurred after 2 weeks at 3o °C but not at 35 °C on the chocolate agar and the CBA medium in the region of the X disk. T h e
M. h a e m o p h i l u m infection in A I D S
305
growth was dry and rugose, quite dissimilar from the growth obtained on F A C m e d i u m on primary isolation. T h e organism was identified as M. haemophilum by the Mycobacterial Reference Laboratory, Cardiff. It was reported to be resistant to streptomycin, isoniazid, ethambutol, ciprofloxacin, gentamicin, augmentin, ceftazidime and sulphamethoxazole, and sensitive to rifampicin, erythromycin, imipenem, minocycline, doxycycline and amikacin. T h e patient was started on treatment with minocycline and rifampicin but suffered vestibular side-effects from the minocycline and was changed to erythromycin and rifampicin. T h e ulcerated lesion began to heal after several weeks. T h e nodules slowly resolved and no more have developed. T h e patient remains apyrexial and well. Discussion
In ~978 Sompolinsky isolated a Mycobacterial species from the skin of an i m m u n o c o m p r o m i s e d patient which required haemin for growth and which he called M. haemophilum. Subsequently Dawson demonstrated the identity of Sompolinsky's isolate to previous isolates from the skin of other i m m u n o compromised p a t i e n t s / T h e species of Mycobacterium isolated from the skin of the patient reported here only grew u p o n Lowenstein-Jensen m e d i u m with ferric a m m o n i u m citrate as a supplement. Its growth was also demonstrated to be dependent u p o n haemin. T h e cultural characteristics of this isolate on F A C and CBA media were similar to previous isolates of M. haemophilum, 8 as was its temperature-dependence. T h e isolate was identified as M. haemophilum by the Mycobacterial Reference Laboratory. T h e presentation in this patient was complicated by extensive Kaposi's sarcoma u p o n the trunk and limbs. T h e skin lesion on the foot was a flat area, with sporotrichoid spread up the calf in the form of painful nodules. T h e histology of a tuberculoid granuloma was consistent with previously reported cases. T h e site of the lesion is consistent with the infection occurring after an inoculation injury to the foot, although the patient did not remember any such event. After radiotherapy the inflammation subsided, presumably due to the death of cells mediating the inflammatory response. This is consistent with the recognised effects of radiation on the inflammatory process. T h e subsequent ulceration of the irradiated lesion is likely to have been a further effect of the radiation u p o n the local cellular i m m u n e defence mechanisms allowing mycobacterial proliferation and ulceration. Once the mycobacterial nature of the lesion was recognised, the patient was started on appropriate chemotherapy and the lesions slowly regressed. T h e r e have been a n u m b e r of reports recently of slowly growing on or nutritionally exacting organisms being isolated from patients with AIDS.9' 10 A high index of suspicion for such organisms must be maintained in these patients. We would r e c o m m e n d that, particularly with skin biopsies, there should be prolonged culture of the specimen on a range of media, including LJ m e d i u m supplemented with ferric a m m o n i u m citrate, in order to maximise the chance of isolating nutritionally fastidious mycobacteria.
306
j. H O L T O N E T AL.
References I. Masur H. Problems in the management of opportunistic infections in patients infected with human immunodeficiency virus. J Infect Dis I99o; I6X : 858-864. 2. Clumeck N, Hermans P, DeWits, S. Current problems in the management of A I D S patients, Eur J Clin Microbiol Infect Dis r988; 7: 2-IO. 3. Harries AD. Tuberculosis and human immunodeficiency virus infection in developing countries. Lancet I99O; 335: 387-390. 4- Guthertz LS, Damsker B, Bottone EJ, Ford EG, Midura T F , Janda MJ. Mycobaeterium avium and Mycobacterium intercellulare infections in patients with and without A I D S . J Infect Dis I989; I6o: IO37-IO4I. 5. Hirschel B, Chang H R , Mach N e t al. Fatal infection with a novel unidentified mycobacterium in a man with A I D S . N EnglJ Med I99o; 3z3: I o 9 - I I 3 . 6. Sompolinsky D, Lagziel A, Neveh D, Yonkilevitz L. Myeobacterium haemophilum sp.nov. a new pathogen from humans. Int J Syst Bacteriol I978; z8: 67-75. 7- Dawson D J, Jennis F. Mycobacteria with a growth requirement for ferric ammonium citrate identified as Mycobacterium haemophilum. J Clin Microbiol I98O; I I : 19o--I92. 8. Moulsdale M T , Harper J, Thatcher GN. Infection by Mycobacterium haemophilum, a metabolically fastidious acid fast bacillus. Tubercle I983 ; 64:24-36. 9. Holton J, Miller R, Furst V, Malnick H. Isolation of Protomonas extorquens (the Red Phantom) from a patient with A I D S . J Infect I99O; zx : 87-93. io. Slater L N , Welch D F , Hensel D, Goody DW. A newly recognized fastidious Gramnegative pathogen as a cause of fever and bacteremia. N EnglJ Med I99O; 323: I587-I592.
