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Abstract

  1. Top of page
  2. Abstract
  3. Case Report
  4. Discussion
  5. Declaration of Interests
  6. References

We describe an imported case of histoplasmosis, whose serological profile was established by means of a protein-based microarray platform, the recently described mycoarray. Because of its peculiarities, such a novel tool greatly facilitates the rapid and multiparametric assessment of patients' serological status and lends itself to be employed as an aid in the diagnosis of primary mycoses, especially in nonendemic countries.

Histoplasmosis, caused by Histoplasma capsulatum, is a granulomatous infection occurring only in specific endemic areas.[1] The environmental reservoir is soil, contaminated by bird or bat stool. Infection occurs by inhalation of conidia during activities, implying contact with the contaminated soil. Pulmonary involvement is the most common clinical presentation: chest radiographs show pulmonary infiltrates and mediastinal lymphadenopathy. Rheumatologic and/or dermatologic manifestations occur in approximately 5% of patients.[2] Immunocompetent individuals usually present an acute pulmonary histoplasmosis which undergoes resolution without treatment.

Recently, a protein microarray-based assay, the mycoarray, has been described as a rapid, sensitive, and specific tool for the serodiagnosis of endemic mycoses, including histoplasmosis.[3] This approach has been proposed as especially useful in areas such as Europe, where endemic mycoses represent a diagnostic challenge, given their rare occurrence.[4]

Case Report

  1. Top of page
  2. Abstract
  3. Case Report
  4. Discussion
  5. Declaration of Interests
  6. References

We report the case of a 30-year-old Italian male presenting with chronic dry cough, sore throat, and fever, 15 days after return from Brazil. He had gone on a 1-month trip around the south-western part of the country, visiting caves and trekking through the forest close to the Iguazu River and falls in the region of Mato Grosso. At clinical examination, the patient was eupneic and his heart rate was 80 bpm. Chest and abdominal examination findings were normal. Empiric antibiotic therapy had already been started but without any clinical improvement. Chest X-ray showed evidence of a solid lesion of 1.5 cm diameter, localized in the upper lobe of the left lung. Computed tomography (CT) scan demonstrated multiple intrapulmonary nodular lesions and mediastinal adenopathies (Figure 1). After some weeks, the patient presented progressive malaise, joint pain, and a nodular, erythematous eruption limited to the extensor aspects of the lower legs, a typical feature of erythema nodosum. At the same time, symmetric polyarthralgia and signs of arthritis at the knees, ankles, and fingers developed, with partial loss of function. Erythema nodosum and polyarthralgia solved gradually with nonsteroidal antinflammatory drugs (NSAIDs). Laboratory parameters showed only a moderate increase of C-reactive protein levels (6.4 mg/dL—normal values inferior to 0.80 mg/dL). Erythrocyte sedimentation rate was 44 mm/hour. Conventional serological tests for typhus, paratyphus, brucellosis, leishmaniosis, rickettsioses, leptospirosis, toxoplasmosis, syphilis, and human immunodeficiency virus (HIV) infection were negative. QuantiFERON-TB test was negative. Complete rheumatic panel and immunological status, including quantitative immunoglobulins, complement and lymphocyte subpopulation, all tested negative. Blood cultures were also performed and no microbial growth was observed. An abdominal ultrasound examination showed some enlarged retroperitoneal lymph nodes. Based on these results, further investigations were found necessary.

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Figure 1. Image of the lung computed tomography (CT) scan. Multiple intrapulmonary nodular lesions and mediastinal adenopathy are evident.

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Bronchoscopy with multiple lung sampling revealed no pathological changes. Microbiological cultures of the bronchoalveolar lavage (BAL) fluids provided negative results. Real-time polymerase chain reaction for Mycobacterium tuberculosis DNA, as well as galactomannan test, returned negative results.

A positron emission tomography (18-FDG PET-CT-Discovery STE, GE Healthcare, Waukesha, WI, USA) scan with dual-time imaging helped to identify ipercaptant lesions to guide surgical sampling. It revealed that the F-18 FDG uptake was very low in pulmonary nodules with a standardized uptake value (SUV) of 2.5 and very high in mediastinal and hilar lymph nodes with SUVs of 20.3 and 23.2, respectively (Figure 1). A percutaneous mediastinal biopsy of an intra-thoracic lymphadenopathy was performed. Histological examination revealed areas of parenchymal necrosis and granulomas, with multinucleated giant cells suggesting an infective etiology (Figure 2). Few isolated, distorted, intra- and extracellular, single and budding yeast forms were evident, suggesting an infection by H capsulatum (Figure 3). However, the biopsy culture did not show any fungal growth.

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Figure 2. Histological analysis on a percutaneous mediastinal biopsy. A residual nodule presents a large zone of caseous necrosis, including some scattered epithelioid histiocytes and multinucleate giant cells with peripheral lymphoid aggregates (PAS staining, ×400 magnification).

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Figure 3. Microphotograph of the lesion. Isolated, distorted, intra- and extracellular, single and budding yeast forms are indicated by the arrows (hematoxylin–eosin staining, ×1000 magnification).

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Finally, the serological profile of the patient was established by the recently described mycoarray,[3] further expanded for assessing antibody reactivities against H capsulatum, Coccidioides immitis, Paracoccidioides brasiliensis, and Blastomyces dermatitidis (Figure 4A). High and specific reactivity was revealed only against the histoplasmin antigen (Figure 4B), both in terms of IgG (left panel) and IgM (right panel) antibodies. When quantitative analysis was performed, the histoplasmin spots returned IgG and IgM antibody masses of 25.54 and 4.72 pg, well above the cut-off values. No appreciable signals were detected in the spots from all the other antigens, thus allowing exclusion of a coinfection.

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Figure 4. Scheme and photos of the array. Panel A, design of the arrays employed for the IgG assay (left) and for the IgM assay (right). Panel B, pictures of the arrays processed with the patient's serum for the determination of the IgG (left) and IgM (right) levels. The low density arrays included the fungal antigens printed in four replicates [histoplasmin (H), coccidioidin (CF), C immitis “TP” (C TP), P brasiliensis (Pb), and B dermatitidis (Bd) antigens], the human IgG or IgM antibody serial dilutions (printed in duplicates), as internal calibration curves for the IgG and IgM assays, and the negative (bovine serum albumin), positive (Alexa 555-Invitrogen Corporation, Carlsbad, CA, USA), and carry-over (blanks) controls. The assays have been performed and data analyzed, as previously detailed.[3]

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Within 2 weeks of admission to the hospital, all signs and symptoms resolved and serial CT scans showed a continuous regression of the pulmonary lesion without any specific treatment, in accordance with the clinical Infectious Diseases Society of America 2007 guidelines.[5] Four months later, when assessing the convalescent serum, the mycoarray returned undetectable IgM levels against histoplasmin, while the IgG were reduced (4.67 pg), yet remaining above the cut-off. No reactivity was observed against the other antigens included in the array.

According to the European Organization for Research and Treatment of Cancer (EORTC) 2008 criteria for the diagnosis of endemic mycoses and based on the anamnesis of the patient, on the histological appearance of the granulomatous inflammation and of small intracellular budding yeast-like particles, this clinical case could be classified as an imported histoplasmosis.[6]

Discussion

  1. Top of page
  2. Abstract
  3. Case Report
  4. Discussion
  5. Declaration of Interests
  6. References

Histoplasmosis incidence is gradually rising in nonendemic countries, owing to increased frequency of both migratory movements and work- or tourism-related travels to and from endemic areas.[7-9] In spite of this, the diagnosis of histoplasmosis is a challenge for clinicians, especially in nonendemic countries, because clinical and radiographic features may be similar to other pulmonary infections or neoplastic diseases.

Histoplasmosis, among travelers, shows a broad spectrum of clinical manifestations. Acute histoplasmosis is the most frequent presentation in immunocompetent patients, presenting with febrile respiratory illness, associated with diffuse reticulonodular infiltrates and mediastinal lymphadenopathy in chest radiographs. Our patient presented a mild acute pulmonary infection associated with rheumatologic manifestations. Histoplasmosis usually heals without antifungal treatment; however, treatment is recommended if symptoms become severe or do not improve after 1 month.[7] It may be associated with arthralgia or arthritis with erythema nodosum, which represent systemic inflammatory responses to the primary pulmonary infection. In this case, similar to others described in literature, signs and symptoms resolved with NSAIDs.[10]

Laboratory diagnosis, based on fungal isolation in culture, represents the gold standard providing direct evidence of infection by H capsulatum; yet, cultures are often negative in patients with mild forms of histoplasmosis; moreover, fungal growth is slow and additional time is needed to achieve definitive species identification.[11] Bronchoscopy, with or without lung sampling, and trans-bronchial needle aspiration may be performed. Also, antigen detection in urine and serum samples can contribute to diagnosis. In particular, the Aspergillus galactomannan enzyme immunoassay can be positive in patients with disseminated histoplasmosis, especially in those who have high concentrations of circulating Histoplasma antigen, likely because of cross-reactive antigens present in the cell wall of both fungi. Yet, in this patient, the galactomannan assay resulted negative, similar to other examinations of the lung.

If a specific diagnosis is not achieved, surgical biopsy of the lung or mediastinal lymph nodes is performed. Histopathologic examination, using fungi-specific staining, allows a rapid diagnosis: morphologic findings include granulomas and diffuse mononuclear cell infiltrates often engulfed with yeast cells. Yet, the sensitivity of this analysis remains below 50%.[11] In our study, histopathology revealed yeast-like forms closely resembling Histoplasma.

Notwithstanding the several drawbacks of serology, ie, delay in antibody production (2–6 weeks after exposure), lack of discrimination between active and previous infection, and hyporesponsiveness in immunosuppressed patients,[11] specific anti-Histoplasma antibodies may be detected in the serum of about 90% of patients with histoplasmosis.[11] The recently described mycoarray[3] allows antibody profiling against a panel of endemic fungal antigens in a rapid, miniaturized, and multiparametric way, thus providing a novel aid in serodiagnosis. Although not clinically validated yet, it may be particularly useful in areas such as Italy, where endemic mycoses are rare events, thus difficult to be promptly identified. As described above, the mycoarray has allowed the rapid detection of antibodies against Histoplasma in a normo-immunoreactive young man, recently returned from Brazil. At first determination both IgG and IgM have been detected at levels well above the cut-off values, while their reduction (IgG) and disappearance (IgM) occurred with time. In contrast, no serum reactivity has been observed against all the other antigens. Overall, even though the absence of positive culture did not allow isolation and identification of the species responsible for the infection, our data provide a serological profile compatible with an imported case of histoplasmosis, ending with a spontaneous resolution.

Declaration of Interests

  1. Top of page
  2. Abstract
  3. Case Report
  4. Discussion
  5. Declaration of Interests
  6. References

The authors state they have no conflicts of interest to declare.

References

  1. Top of page
  2. Abstract
  3. Case Report
  4. Discussion
  5. Declaration of Interests
  6. References