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Microscopy and latex antigen negative cryptococcal meningitis
Case Reports
329
Microscopy and Latex Antigen Negative Cryptococcal Meningitis I. F. Laurenson .1, J. D. C. R o s s 2 and L. J. R. Milne 3 1Department of Medical Microbiology, University of Edinburgh Medical School, Edinburgh EH8 9AG, 2Departrnent of GU Medicine, Edinburgh Royal Infirmary, Edinburgh EH3 9YW, 3RegionaI Mycology Reference Laboratory, Western General Hospital, Edinburgh EH4 2XU, U.I(. A HIV-positive patient presented with cryptococcal meningitis that was not detected by cerebrospinal fluid (CSF) latex antigen and direct microscopy. The diagnosis was confirmed by culture of the CSF and subsequent urine culture, both of which yielded an apparently acapsular strain of Cryptococcus neoformans, After 19 months the patient relapsed and capsulated yeasts were observed on this occasion on direct microscopy of the CSF. The latex antigen test was strongly positive. Culture again yielded an apparently acapsular isolate. Retrospective culture of all isolates obtained from this patient in sterile CSF resulted in the formation of capsules. This was confirmed by the requirement of normal non heat inactivated serum for neutrophil-cryptococcus attachment to occur in vitro. Although antigen and direct microscopy are frequently relied upon to diagnose cryptococcal meningitis, a negative result does not exclude the condition.
Introduction In the U.K. cryptococcal meningitis caused by Cryptococcus neoformans occurs in at least 4% of patients with AIDS, while in the U.S.A. 6-10% of cases are affected. 1'2 In Edinburgh in 1995 there were four immunocompromised patients diagnosed with cryptococcosis, of whom three were infected with the human immunodeficiency virus. In 1996 there were no new cases diagnosed. Headache is a common presenting symptom, but there are often no focal findings. The initial cerebrospinal fluid (CSF) findings may be virtually normal. We report a patient in whom the initial diagnosis of cryptococcal meningitis was only made after culture of CSF deposit.
Case Report A 17-year-old Caucasian male was diagnosed as HIV-positive following routine screening during blood donation. He had previously been in a heterosexual relationship, but denied any other risk factors. On questioning at initial presentation he gave a 4-month history of mild intermittent headache associated with fever and night sweats. There was no history of travel, pigeon or antifungal exposure. Examination found him alert and orientated. His temperature was 39.8 °C, and he had oral thrush and axillary lymphadenopathy without clinical hepatosplenomegafly. There were no signs of meningitis. Negative tests included biochemistry and haematology profiles, repeated blood cultures, chest X-ray, induced sputum for Pneumocystis carinii, urinary bacterial culture and computerized tomography of the head. At presentation the CD4 lymphocyte count was 125/mm 3, being 6% of the total peripheral white blood cell count. At lumbar puncture the CSF protein was 474 mg/1 (reference range 100-400 mg/1), glucose 2.8 mmol/1 * Address all correspondence to: I. F. Laurenson. Accepted for publication 10 July 1997.
with no cells or organisms present, the pressure was not recorded. The concurrent serum glucose was 5.8 mmol/l. Gram staining of the CSF and cryptococcal antigen testing (IMMYImmuno-mycologics, Norman, OK, U.S.A.) of CSF and blood were negative, as was examination for other pathogens. After 3 days' culture of the spun CSF deposit on 4% malt extract agar in air at 30°C, four colonies of an apparently noncapsulated, relatively fluconazole resistant isolate (MIC 16big/ ml, confirmed at the PHLS Mycology Laboratory, Bristol) of Cryptococcus neoformans was cultured. A similar isolate was subsequently cultured from urine, which then gave a positive antigen test at a 1:4 titre. Serotyping at the PHLS Reference Mycology Laboratory, Leeds showed the CSF isolate to be serotype A. The patient responded rapidly to amphotericin B preparations given for 3 weeks followed by oral itraconazole, to which the isolate was sensitive in vitro. The latter was poorly tolerated and the patient continued on fluconazole 200 mg per day. He remained well while being prescribed fluconazole, antiretroviral therapy and pentamidine nebulizers until 19 months later. He then relapsed complaining of headache, vomiting and photophobia. At this stage he admitted that he had not been taking his fluconazole for the past 2-3 weeks. On this occasion CSF examination showed capsular cryptocoeci in India-ink staining but was again otherwise acellular. The CSF glucose was 2.4 mmol/1, protein 1144 mg/1 and serum glucose 7.9 mmol/1. Culture and latex antigen testing of CSF and peripheral blood were positive (antigen CSF titre 1:512), while the CD4 count was 0/mm 3, On this admission a CSF opening pressure of 38 cm was recorded. 'Abelcet' (liposomal amphotericin) was commenced and he was discharged after 3 weeks, improving on oral intaconazole suspension. The isolate was reported to be more resistant to flnconazole on disc testing than the original isolate. The original CSF isolate was grown in air at 37 °C and a neutrophil attachment assay showed that normal, non heatinactivated human serum was necessary for attachment to occur. This implies that opsonization by complement is required for attachment and therefore that some capsule is present.
330
Case Reports
Further study of all isolates from this patient grown at 3 7 °C in CSF showed significant capsules on India-ink examination.
Discussion The capsule of the yeast-like fungus C. neoformans varies in size depending on growth conditions. 3 In experimental mice the capsule size does not seem to affect virulence, * although acapsular mutants are less virulent. 3 The capsule inhibits phagocytosis and may impair leucocyte migration, 6'7 as well as being immunosuppressive. 8 Recent work suggests that capsular glucoronoxylomannan (GXM) induces interleukin-lO (a potent downregulator of proinflammatory cytokines) secretion by h u m a n monocytes. 9 Release of tumor necrosis factor cz, interlenkin-l]3 and interleukin-8, proinflammatory cytokines, by h u m a n neutrophils has also been correlated to capsule size and quantity of GXM present by the same group of workers. 1° There is a report that capsules may enhance HIV replication. 1~ Acapsular strains activate both classical and alternative complement pathways, .2 while capsular strains only activate the latter. In one study, 13 9% of AIDS patients had culture-positive cryptococcal meningitis but negative CSF antigen detection tests, although serum was negative in only 1% of patients. Low levels of antigen, I4 presence of immune-complexes (in nonprotease treated serum samples), .5 high titres (prozone effect) or poorly/non-encapsulated strains may contribute to such negative antigen tests. In cases with extraneural cryptococcosis a m u c h lower percentage of samples are positive. These factors and the possibility of a false-negative antigen test may not be commonly appreciated. Urinary screening is not always routinely carried out and antigen testing not standardized, although the prostate in particular may act as a reservoir. Commercial kits to detect cryptococcal capsular antigen by latex agglutination and enzyme immunoassay are widely used, although there is some variation in performance. 16 The I m m y kit used here has a sensitivity and specificity in serum of 9 7% and 93%, and in CSF of 93% and 93%, respectively. ~6 This can be further improved without loss of sensitivity by raising the threshold for a positive result to 1:2 from the 1:1 indicated in the manufacturer's instructions. Pronase treatment of serum and CSF is used with this kit to limit non-specific interference. All four test kits in this study were compared with culture of CSF. 16 The sensitivity of antigen detection in the CSF was 9 3 - 1 0 0 % , and specificity ranged from 93-98%, while in serum sensitivity ranges were 8 3 - 9 7 % and specificity ranges 9 5 100%. It must therefore be emphasized to clinicians and microbiologists that cryptococcal meningitis cannot be excluded simply on the basis of negative (even repeatedly so) CSF and serum antigen tests or microscopy. In this case the isolate was apparently acapsular on culture. It seems that this isolate could produce capsular material as shown by its serotyping result, behaviour in the neutrophil assay and subsequent in vitro culture in CSF. It may be that capsule formation was stimulated by the host and that initial antigen tests were negative due to low numbers of cryptococci present. It is interesting to note that this patient was relatively well until 19 months after presentation and responded to further treatment. This is considerably longer than the median survival of 152 days in culture positive cases in the study reported by Chuck and Sande. 13 Improvement in antiretroviral therapy since that study was carried out may lead to a slower decline
in immunity. Collins and Bancroft have shown that encapsulation of C. neoformans impairs antigen-specific T-cell responses, w h e n compared with an acapsular mutant. 17 Therefore it may be that the lack of apparent capsule in this case also contributed to the longer time until relapse. Fluconazole resistance, though rarely documented in C. neoformans, m a y not correlate with clinical outcome. In this patient treatment was successful until compliance became a problem, although the new isolate is capsular and more resistant than the original, and the CD4 count was O/ram 3 at relapse. Reinfection with a different strain cannot be ruled out. This case illustrates the importance of culture at 2 5 - 3 0 °C on an appropriate medium such as 4% malt extract agar in making the diagnosis of cryptococcosis. Standard 48-h incubation, as commonly carried out for more c o m m o n bacterial pathogens, is insufficient. Culture may be positive even where other parameters such as CSF cell count are normal. If the diagnosis is still elusive, repeated CSF examination should be considered. Such procedures should be carried out, particularly in immunosuppressed patients, even w h e n antigen tests are negative.
Acknowledgements We are grateful to Dr D. W. Warnock, Bristol PHLS Mycology Reference Laboratory for performing the MIC and Dr V. Hopwood of the Leeds PHLS Reference Mycology Laboratory for serotyping the isolate.
References 1 Dismukes WE. Cryptococcal meningitis in patients with AIDS. ] Infect Dis 1988: 157: 624-628. 2 Knight FR, Macket3zie DW, Evans BG, Porter K, Barrett NI, White GC. Increasing incidence of cryptococcosis in the United Kingdom. J Infect 1993; 27: 185-191. 3 Vartivarian SE, Anaissie EJ, Cowart RE, Sprigg HA, Tingler iV[], lacobson ES. Regulation of cryptococcal capsular polysaccharide by iron. J Infect Dis 1993; 167: 186-190. 4 Dykstra MA, Friedman L, Murphy JW. Capsule size of Cryptoeoccus neoformans: control and relation to cirulence. Infection & Immunity 1977; 16: 129-135. 5 Kwon-Chung KJ, Rhodes JC. Encapsulation and melanin formation as indicators of virulence in Cryptococcus neoformans. Infection & Immunity 1986; 51: 218-223. 6 Kozel TR, Hermerath CA. Binding of cryptococcal polysaccharide to Cryptococeus neoformans. I@ction & Immunity 1984; 43: 879-886. 7 Drouhet E, Segretain G. Inhibition de la migration leucocytaire in vitro par un polyoside capsulaire de Torulopsis (Cryptococcus) neoformans. Annales de l'Institut Pasteur 1951; 81: 674-676. 8 Khakpour FR, Murphy JW. Characterization of a third-order suppressor T cell (Ts3) induced by cryptococcal antigen(s). Infection & Immunity 1987; 55: 1657-1662. 9 Vecciarelli A, Retini C, Pietrella D et al. Purified capsular polysaccharide of Cryptococcus neoformans induces interleukin-lO secretion by human monocytes. Infection & Immunity 1996; 64: 2846-2849. 10 Retini C, Vecchiarelli A, Monari C, Tascini C, Bistoni 13, Kozel TR. Capsular polysaccharide of Cryptococcus neoforrnans induces prointlammatory cytokine release by human neutrophils. Infection & Immunitg 1996; 64:: 2897-2903. 11 Pettoello-Mantovani M, Casadevall A, Kollmann TR, Rubenstein A, Goldstein H. Enhancement of HIV-1 infection by the capsular
Case Reports polysaccharide of Cryptococcus neoformans. Lancet 1992; 339: 2123. 12 Kozel TR, Wilson MA, Welch WH. Kinetic analysis of the amplification phase for activation and binding of C3 to encapsulated and nonencapsulated Cryptococcusneoformans.I@ction gJ Immunity 1992; 60: 3122-3127. 13 Chuck SL, Sande MA. Infections with Cryptococcus neoformans in the acquired immuno deficiency syndrome. N Eng ] Med 1989; 321: 794-799. 14 Currie BP, Freundlich LF, Soto MA, Casadevall A. False-negative cerebrospinal fluid cryptococcal latex agglutination tests for patients
331
with culture-positive cryptococcal meningitis. ] ClinMicrobio11993; 31: 2519-2522. 15 Sadamoto S, Ikeda R, Nishikawa A, Shinoda T. Evidence for interference by immune complexes in the serodiagnosis of cryptococcosis. Microbiol & Immunol 1993; 37: 129-133. 16 Tanner DC, Weinstein MP, Fedorciw B, Joho KL, Thorpe JJ, Reller LB. Comparison of commercial kits for detection of cryptococcal antigen. J Clin Microbiol 1994; 32: 1680-1684. 17 Collins HL. Bancroft GJ. Encapsulation of Cryptococcus neoformans impairs antigen-specific T-cell responses. Infection& Immunity 1991; 59: 3883-3888.
Ecthyma Gangrenosum in a Patient with Hypogammaglobulinemia W, Ng .1, C. L, Tan 1, V, Y e o w 2, M. Yeo 3 and S. H. Teo ~ Departments of 1Paediatrics, 2Plastic Surgery and 3Pathology, Singapore General Hospital, Singapore Ecthyma gangrenosum is a characteristic skin lesion that is caused by Pseudomonas aeruginosa (P. aeruginosa) in the majority of cases. Systemic P. aeruginosa usually complicates debilitating conditions like leukaemia, burns and cystic fibrosis. We report a patient wth underlying hypogammaglobulinemia who developed ecthyma gangrenosum secondary to P. aeruginosa septicaemia, which was potentially life-threatening. Recognition of the characteristic skin lesions with prompt initiation of appropriate antibiotics and intravenous immunoglobulins were life-saving. A review of the English literature reports three other cases of ecthyma gangrenosum in patients with underlying hypogammaglobulinemia.
introduction
Case Report
Ecthyma gangrenosum is a characteristic skin lesion that was first described by Baker 1 in 189 7. In 19 71, Dorff2 described the skin lesion of ecthyma gangrenosum progressing through a characteristic pattern: oedema, erythema, haemorrhagic bullae and frank necrosis. The entire sequence may evolve within hours, with lesions of varying ages appearing simultaneously. Pseudomonas aeru~inosa (E aeruginosa) is the causative organism in the majority of cases. We report this case to highlight that ecthyma gangrenosum due to E aeruginosa can be life-threatening, especially in a patient with hypogammaglobulinemia. However, prompt recognition of the skin lesions with immediate institution of appropriate antibiotic therapy and intravenous immunoglobulins may be life-saving, as illustrated in our reported case. Although pseudomonads may produce disease in any individual, they most commonly cause disease in those with burns, cystic fibrosis, malignancies, underlying immunodeficiency disease and in recipients of immunosuppressive therapy. 3 A review of the English literature reports three other cases of ecthyma gangrenosum in patients with underlying hypogammaglobulinemia.4-6
A 7-year-old Chinese boy was admitted with a 4-day history of fever and skin lesions. At 2.5 years of age he was admitted for left otitis media, and a left cortical mastoidectomy operation was done. Investigations showed that he had underlying hypogammaglobulinemia (IgG 0.88 g/l, IgM 0.18g/1, IgA 0.04 g/l). He was readmitted at 6.5 years of age for Haemophilus influenzae chest infection, which responded to a course of antibiotics. His parents are non-consanguineous, and there is no family history of any underlying immunodeficiencies. On physical examination, the child was well nourished but appeared extremely ill. Axillary temperature was 39 °C, pulse 190 per min and respiration 70 per min. Blood pressure was 74/36 mmHg. A large necrotic ulcer covered with a black eschar, measuring 7 cm by 5 cm, was noted on his upper chest. (Fig. 1). There were multiple nodular erythematous lesions with a bluish to purple core seen over all four limbs, the largest measuring 3 cm by 4 cm in diameter. These lesions progressed from erythema to haemorrhagic bullae and necrosis. The liver was enlarged 7 cm below the right costal margin. The spleen was not enlarged. Heart sounds were normal and the lungs were clear. Intravenous ceftazidime and amikacin were started on admission. Prompt resuscitation using normal saline 2 0 m l / k g and fresh frozen plasma 2 0 m l / k g were given; in addition he required inotropic support using dopamine 15 gg/kg/min, dobutamine 15 gg/kg/min and adrenaline 0.15 gg/kg/min to maintain his blood pressure. Intravenous immunoglobnlin 900 mg/kg was given on admission and subsequently every 3 weeks. The laboratory findings showed a Hb of 11.3 g/1, WBC
* Address all correspondence to: W. Ng (Attention Ms R Jennings), Southwestern Medical School, Department of Paediatrics, Division of Infectious Diseases, 5323 Harry Hines Blvd, Dallas, Texas 75235-9063, USA. Accepted for publication 10 luly 1997.
329
Microscopy and Latex Antigen Negative Cryptococcal Meningitis I. F. Laurenson .1, J. D. C. R o s s 2 and L. J. R. Milne 3 1Department of Medical Microbiology, University of Edinburgh Medical School, Edinburgh EH8 9AG, 2Departrnent of GU Medicine, Edinburgh Royal Infirmary, Edinburgh EH3 9YW, 3RegionaI Mycology Reference Laboratory, Western General Hospital, Edinburgh EH4 2XU, U.I(. A HIV-positive patient presented with cryptococcal meningitis that was not detected by cerebrospinal fluid (CSF) latex antigen and direct microscopy. The diagnosis was confirmed by culture of the CSF and subsequent urine culture, both of which yielded an apparently acapsular strain of Cryptococcus neoformans, After 19 months the patient relapsed and capsulated yeasts were observed on this occasion on direct microscopy of the CSF. The latex antigen test was strongly positive. Culture again yielded an apparently acapsular isolate. Retrospective culture of all isolates obtained from this patient in sterile CSF resulted in the formation of capsules. This was confirmed by the requirement of normal non heat inactivated serum for neutrophil-cryptococcus attachment to occur in vitro. Although antigen and direct microscopy are frequently relied upon to diagnose cryptococcal meningitis, a negative result does not exclude the condition.
Introduction In the U.K. cryptococcal meningitis caused by Cryptococcus neoformans occurs in at least 4% of patients with AIDS, while in the U.S.A. 6-10% of cases are affected. 1'2 In Edinburgh in 1995 there were four immunocompromised patients diagnosed with cryptococcosis, of whom three were infected with the human immunodeficiency virus. In 1996 there were no new cases diagnosed. Headache is a common presenting symptom, but there are often no focal findings. The initial cerebrospinal fluid (CSF) findings may be virtually normal. We report a patient in whom the initial diagnosis of cryptococcal meningitis was only made after culture of CSF deposit.
Case Report A 17-year-old Caucasian male was diagnosed as HIV-positive following routine screening during blood donation. He had previously been in a heterosexual relationship, but denied any other risk factors. On questioning at initial presentation he gave a 4-month history of mild intermittent headache associated with fever and night sweats. There was no history of travel, pigeon or antifungal exposure. Examination found him alert and orientated. His temperature was 39.8 °C, and he had oral thrush and axillary lymphadenopathy without clinical hepatosplenomegafly. There were no signs of meningitis. Negative tests included biochemistry and haematology profiles, repeated blood cultures, chest X-ray, induced sputum for Pneumocystis carinii, urinary bacterial culture and computerized tomography of the head. At presentation the CD4 lymphocyte count was 125/mm 3, being 6% of the total peripheral white blood cell count. At lumbar puncture the CSF protein was 474 mg/1 (reference range 100-400 mg/1), glucose 2.8 mmol/1 * Address all correspondence to: I. F. Laurenson. Accepted for publication 10 July 1997.
with no cells or organisms present, the pressure was not recorded. The concurrent serum glucose was 5.8 mmol/l. Gram staining of the CSF and cryptococcal antigen testing (IMMYImmuno-mycologics, Norman, OK, U.S.A.) of CSF and blood were negative, as was examination for other pathogens. After 3 days' culture of the spun CSF deposit on 4% malt extract agar in air at 30°C, four colonies of an apparently noncapsulated, relatively fluconazole resistant isolate (MIC 16big/ ml, confirmed at the PHLS Mycology Laboratory, Bristol) of Cryptococcus neoformans was cultured. A similar isolate was subsequently cultured from urine, which then gave a positive antigen test at a 1:4 titre. Serotyping at the PHLS Reference Mycology Laboratory, Leeds showed the CSF isolate to be serotype A. The patient responded rapidly to amphotericin B preparations given for 3 weeks followed by oral itraconazole, to which the isolate was sensitive in vitro. The latter was poorly tolerated and the patient continued on fluconazole 200 mg per day. He remained well while being prescribed fluconazole, antiretroviral therapy and pentamidine nebulizers until 19 months later. He then relapsed complaining of headache, vomiting and photophobia. At this stage he admitted that he had not been taking his fluconazole for the past 2-3 weeks. On this occasion CSF examination showed capsular cryptocoeci in India-ink staining but was again otherwise acellular. The CSF glucose was 2.4 mmol/1, protein 1144 mg/1 and serum glucose 7.9 mmol/1. Culture and latex antigen testing of CSF and peripheral blood were positive (antigen CSF titre 1:512), while the CD4 count was 0/mm 3, On this admission a CSF opening pressure of 38 cm was recorded. 'Abelcet' (liposomal amphotericin) was commenced and he was discharged after 3 weeks, improving on oral intaconazole suspension. The isolate was reported to be more resistant to flnconazole on disc testing than the original isolate. The original CSF isolate was grown in air at 37 °C and a neutrophil attachment assay showed that normal, non heatinactivated human serum was necessary for attachment to occur. This implies that opsonization by complement is required for attachment and therefore that some capsule is present.
330
Case Reports
Further study of all isolates from this patient grown at 3 7 °C in CSF showed significant capsules on India-ink examination.
Discussion The capsule of the yeast-like fungus C. neoformans varies in size depending on growth conditions. 3 In experimental mice the capsule size does not seem to affect virulence, * although acapsular mutants are less virulent. 3 The capsule inhibits phagocytosis and may impair leucocyte migration, 6'7 as well as being immunosuppressive. 8 Recent work suggests that capsular glucoronoxylomannan (GXM) induces interleukin-lO (a potent downregulator of proinflammatory cytokines) secretion by h u m a n monocytes. 9 Release of tumor necrosis factor cz, interlenkin-l]3 and interleukin-8, proinflammatory cytokines, by h u m a n neutrophils has also been correlated to capsule size and quantity of GXM present by the same group of workers. 1° There is a report that capsules may enhance HIV replication. 1~ Acapsular strains activate both classical and alternative complement pathways, .2 while capsular strains only activate the latter. In one study, 13 9% of AIDS patients had culture-positive cryptococcal meningitis but negative CSF antigen detection tests, although serum was negative in only 1% of patients. Low levels of antigen, I4 presence of immune-complexes (in nonprotease treated serum samples), .5 high titres (prozone effect) or poorly/non-encapsulated strains may contribute to such negative antigen tests. In cases with extraneural cryptococcosis a m u c h lower percentage of samples are positive. These factors and the possibility of a false-negative antigen test may not be commonly appreciated. Urinary screening is not always routinely carried out and antigen testing not standardized, although the prostate in particular may act as a reservoir. Commercial kits to detect cryptococcal capsular antigen by latex agglutination and enzyme immunoassay are widely used, although there is some variation in performance. 16 The I m m y kit used here has a sensitivity and specificity in serum of 9 7% and 93%, and in CSF of 93% and 93%, respectively. ~6 This can be further improved without loss of sensitivity by raising the threshold for a positive result to 1:2 from the 1:1 indicated in the manufacturer's instructions. Pronase treatment of serum and CSF is used with this kit to limit non-specific interference. All four test kits in this study were compared with culture of CSF. 16 The sensitivity of antigen detection in the CSF was 9 3 - 1 0 0 % , and specificity ranged from 93-98%, while in serum sensitivity ranges were 8 3 - 9 7 % and specificity ranges 9 5 100%. It must therefore be emphasized to clinicians and microbiologists that cryptococcal meningitis cannot be excluded simply on the basis of negative (even repeatedly so) CSF and serum antigen tests or microscopy. In this case the isolate was apparently acapsular on culture. It seems that this isolate could produce capsular material as shown by its serotyping result, behaviour in the neutrophil assay and subsequent in vitro culture in CSF. It may be that capsule formation was stimulated by the host and that initial antigen tests were negative due to low numbers of cryptococci present. It is interesting to note that this patient was relatively well until 19 months after presentation and responded to further treatment. This is considerably longer than the median survival of 152 days in culture positive cases in the study reported by Chuck and Sande. 13 Improvement in antiretroviral therapy since that study was carried out may lead to a slower decline
in immunity. Collins and Bancroft have shown that encapsulation of C. neoformans impairs antigen-specific T-cell responses, w h e n compared with an acapsular mutant. 17 Therefore it may be that the lack of apparent capsule in this case also contributed to the longer time until relapse. Fluconazole resistance, though rarely documented in C. neoformans, m a y not correlate with clinical outcome. In this patient treatment was successful until compliance became a problem, although the new isolate is capsular and more resistant than the original, and the CD4 count was O/ram 3 at relapse. Reinfection with a different strain cannot be ruled out. This case illustrates the importance of culture at 2 5 - 3 0 °C on an appropriate medium such as 4% malt extract agar in making the diagnosis of cryptococcosis. Standard 48-h incubation, as commonly carried out for more c o m m o n bacterial pathogens, is insufficient. Culture may be positive even where other parameters such as CSF cell count are normal. If the diagnosis is still elusive, repeated CSF examination should be considered. Such procedures should be carried out, particularly in immunosuppressed patients, even w h e n antigen tests are negative.
Acknowledgements We are grateful to Dr D. W. Warnock, Bristol PHLS Mycology Reference Laboratory for performing the MIC and Dr V. Hopwood of the Leeds PHLS Reference Mycology Laboratory for serotyping the isolate.
References 1 Dismukes WE. Cryptococcal meningitis in patients with AIDS. ] Infect Dis 1988: 157: 624-628. 2 Knight FR, Macket3zie DW, Evans BG, Porter K, Barrett NI, White GC. Increasing incidence of cryptococcosis in the United Kingdom. J Infect 1993; 27: 185-191. 3 Vartivarian SE, Anaissie EJ, Cowart RE, Sprigg HA, Tingler iV[], lacobson ES. Regulation of cryptococcal capsular polysaccharide by iron. J Infect Dis 1993; 167: 186-190. 4 Dykstra MA, Friedman L, Murphy JW. Capsule size of Cryptoeoccus neoformans: control and relation to cirulence. Infection & Immunity 1977; 16: 129-135. 5 Kwon-Chung KJ, Rhodes JC. Encapsulation and melanin formation as indicators of virulence in Cryptococcus neoformans. Infection & Immunity 1986; 51: 218-223. 6 Kozel TR, Hermerath CA. Binding of cryptococcal polysaccharide to Cryptococeus neoformans. I@ction & Immunity 1984; 43: 879-886. 7 Drouhet E, Segretain G. Inhibition de la migration leucocytaire in vitro par un polyoside capsulaire de Torulopsis (Cryptococcus) neoformans. Annales de l'Institut Pasteur 1951; 81: 674-676. 8 Khakpour FR, Murphy JW. Characterization of a third-order suppressor T cell (Ts3) induced by cryptococcal antigen(s). Infection & Immunity 1987; 55: 1657-1662. 9 Vecciarelli A, Retini C, Pietrella D et al. Purified capsular polysaccharide of Cryptococcus neoformans induces interleukin-lO secretion by human monocytes. Infection & Immunity 1996; 64: 2846-2849. 10 Retini C, Vecchiarelli A, Monari C, Tascini C, Bistoni 13, Kozel TR. Capsular polysaccharide of Cryptococcus neoforrnans induces prointlammatory cytokine release by human neutrophils. Infection & Immunitg 1996; 64:: 2897-2903. 11 Pettoello-Mantovani M, Casadevall A, Kollmann TR, Rubenstein A, Goldstein H. Enhancement of HIV-1 infection by the capsular
Case Reports polysaccharide of Cryptococcus neoformans. Lancet 1992; 339: 2123. 12 Kozel TR, Wilson MA, Welch WH. Kinetic analysis of the amplification phase for activation and binding of C3 to encapsulated and nonencapsulated Cryptococcusneoformans.I@ction gJ Immunity 1992; 60: 3122-3127. 13 Chuck SL, Sande MA. Infections with Cryptococcus neoformans in the acquired immuno deficiency syndrome. N Eng ] Med 1989; 321: 794-799. 14 Currie BP, Freundlich LF, Soto MA, Casadevall A. False-negative cerebrospinal fluid cryptococcal latex agglutination tests for patients
331
with culture-positive cryptococcal meningitis. ] ClinMicrobio11993; 31: 2519-2522. 15 Sadamoto S, Ikeda R, Nishikawa A, Shinoda T. Evidence for interference by immune complexes in the serodiagnosis of cryptococcosis. Microbiol & Immunol 1993; 37: 129-133. 16 Tanner DC, Weinstein MP, Fedorciw B, Joho KL, Thorpe JJ, Reller LB. Comparison of commercial kits for detection of cryptococcal antigen. J Clin Microbiol 1994; 32: 1680-1684. 17 Collins HL. Bancroft GJ. Encapsulation of Cryptococcus neoformans impairs antigen-specific T-cell responses. Infection& Immunity 1991; 59: 3883-3888.
Ecthyma Gangrenosum in a Patient with Hypogammaglobulinemia W, Ng .1, C. L, Tan 1, V, Y e o w 2, M. Yeo 3 and S. H. Teo ~ Departments of 1Paediatrics, 2Plastic Surgery and 3Pathology, Singapore General Hospital, Singapore Ecthyma gangrenosum is a characteristic skin lesion that is caused by Pseudomonas aeruginosa (P. aeruginosa) in the majority of cases. Systemic P. aeruginosa usually complicates debilitating conditions like leukaemia, burns and cystic fibrosis. We report a patient wth underlying hypogammaglobulinemia who developed ecthyma gangrenosum secondary to P. aeruginosa septicaemia, which was potentially life-threatening. Recognition of the characteristic skin lesions with prompt initiation of appropriate antibiotics and intravenous immunoglobulins were life-saving. A review of the English literature reports three other cases of ecthyma gangrenosum in patients with underlying hypogammaglobulinemia.
introduction
Case Report
Ecthyma gangrenosum is a characteristic skin lesion that was first described by Baker 1 in 189 7. In 19 71, Dorff2 described the skin lesion of ecthyma gangrenosum progressing through a characteristic pattern: oedema, erythema, haemorrhagic bullae and frank necrosis. The entire sequence may evolve within hours, with lesions of varying ages appearing simultaneously. Pseudomonas aeru~inosa (E aeruginosa) is the causative organism in the majority of cases. We report this case to highlight that ecthyma gangrenosum due to E aeruginosa can be life-threatening, especially in a patient with hypogammaglobulinemia. However, prompt recognition of the skin lesions with immediate institution of appropriate antibiotic therapy and intravenous immunoglobulins may be life-saving, as illustrated in our reported case. Although pseudomonads may produce disease in any individual, they most commonly cause disease in those with burns, cystic fibrosis, malignancies, underlying immunodeficiency disease and in recipients of immunosuppressive therapy. 3 A review of the English literature reports three other cases of ecthyma gangrenosum in patients with underlying hypogammaglobulinemia.4-6
A 7-year-old Chinese boy was admitted with a 4-day history of fever and skin lesions. At 2.5 years of age he was admitted for left otitis media, and a left cortical mastoidectomy operation was done. Investigations showed that he had underlying hypogammaglobulinemia (IgG 0.88 g/l, IgM 0.18g/1, IgA 0.04 g/l). He was readmitted at 6.5 years of age for Haemophilus influenzae chest infection, which responded to a course of antibiotics. His parents are non-consanguineous, and there is no family history of any underlying immunodeficiencies. On physical examination, the child was well nourished but appeared extremely ill. Axillary temperature was 39 °C, pulse 190 per min and respiration 70 per min. Blood pressure was 74/36 mmHg. A large necrotic ulcer covered with a black eschar, measuring 7 cm by 5 cm, was noted on his upper chest. (Fig. 1). There were multiple nodular erythematous lesions with a bluish to purple core seen over all four limbs, the largest measuring 3 cm by 4 cm in diameter. These lesions progressed from erythema to haemorrhagic bullae and necrosis. The liver was enlarged 7 cm below the right costal margin. The spleen was not enlarged. Heart sounds were normal and the lungs were clear. Intravenous ceftazidime and amikacin were started on admission. Prompt resuscitation using normal saline 2 0 m l / k g and fresh frozen plasma 2 0 m l / k g were given; in addition he required inotropic support using dopamine 15 gg/kg/min, dobutamine 15 gg/kg/min and adrenaline 0.15 gg/kg/min to maintain his blood pressure. Intravenous immunoglobnlin 900 mg/kg was given on admission and subsequently every 3 weeks. The laboratory findings showed a Hb of 11.3 g/1, WBC
* Address all correspondence to: W. Ng (Attention Ms R Jennings), Southwestern Medical School, Department of Paediatrics, Division of Infectious Diseases, 5323 Harry Hines Blvd, Dallas, Texas 75235-9063, USA. Accepted for publication 10 luly 1997.