Correct answer 3: Infection by Histoplasma spp.

Histoplasmosis, caused by Histoplasma spp., is highly endemic in the Americas.^1,2 In Colombia, histoplasmosis is not a disease requiring mandatory notification, but it is known that HIV infection, especially in the AIDS stage, is present in 70% of patients diagnosed with progressive disseminated histoplasmosis (PDH).^3,4 Risk factors for infection include handling soil infected with the fungus, as in the case of this patient, a farmer.^5,6,7

In the clinical presentation, it is important to note the fever, constitutional and respiratory symptoms, accompanied by skin lesions consistent with disseminated histoplasmosis, which are similar to the presentation described in more than 38% of patients with AIDS.^{4, 5} The risk of histoplasmosis is greatest in patients with HIV infection, especially those with CD4 counts ≤200 cells/mm³.^6,8,9

On the other hand, the disease produced by Talaromyces marneffei is also frequent in patients with AIDS, but it is endemic in Southeast Asia, with limited geographic distribution. In its clinical presentation it is very similar to PDH, including skin lesions in 60-70% of patients.¹⁰ In disseminated tuberculosis, skin involvement only occurs in 0.4-2.8% of patients with a predominance of respiratory symptoms.^11,12  In cryptococcosis occurring in immunocompromised patients, the predominant presentation is meningeal involvement, less frequently skin involvement. When skin lesions do occur, they often appear as with an ulcerated nodule in the center, and generally, few lesions.¹³

Correct answer 4: All of the above, depending on the local facilities

The results/reports of the direct skin examination and bone marrow are shown in Figures 3A, B, and C.

All the procedures indicated in the above question should be carried out if possible, in order to investigate the changes seen on the chest X-ray, the pancytopenia reported by the laboratory tests, and the skin lesions.

Common reasons for needing bronchoscopy are a persistent cough, infection, or something unusual seen on a chest X-ray, principally when interstitial infiltrates are reported. Bronchoscopy can also be used to obtain samples of mucus or tissue or to remove foreign bodies or other blockages from the airways or lungs. It is a minimally invasive procedure to diagnose problems with the lungs.

Wright, Giemsa, and Grocott´s methenamine silver (GMS) stains can be used to detect yeast cells of Histoplasma spp. in tissues, blood, or bone marrow smears. These cells can also be seen in tissue sections stained with Periodic Acid Schiff (PAS) but usually not with hematoxylin and eosin (H&E).⁶

Figure 3

A. Bone marrow aspirate smear stained with Giemsa showing numerous yeast-like cells inside macrophages (arrows) with morphology suggestive of Histoplasma spp. Magnification 100X.

B. Skin scraping stained with Grocott´s methenamine silver stain (GMS) showing numerous blastoconidia suggestive of Histoplasma spp. Magnification 100X.

C. Bone-marrow aspirate smear stained with GMS showing numerous yeast-like cells suggestive of Histoplasma spp. Magnification 40X. Images courtesy of Beatriz L. Gómez, PhD.

Correct answer 1. Yes

Detection of the small (2- to 4-μm), oval, budding yeasts allows a presumptive diagnosis of histoplasmosis. Organisms can be found within macrophages or free in the tissues. It is unusual to find yeast cells on cytological examination of sputum or other respiratory tract fluids. The thick-walled, narrow-based budding yeast cells causing African histoplasmosis, approximately 10 to 15 μm in diameter, are about fourfold larger than those of classical Histoplasma spp. in tissue sections and might occasionally be confused with Blastomyces spp.⁶

Histoplasma spp. can be isolated from samples such as sputum, bronchoalveolar lavage fluid specimens, and biopsies. Culture of clinical samples is considered the gold standard; the analytical performance of the assay can vary according to the clinical presentation. In disseminated disease, useful specimens for culture include blood, urine, lymph node, and bone marrow samples, and sensitivity varies from 74% to 95% depending on the sample available. Culture does not provide a rapid result since the fungus can take up to 4 to 6 weeks to grow.^6,9

Biopsy specimens of oral, cutaneous, and gastrointestinal lesions, adrenal glands, or liver and spleen can also provide a diagnosis. Sensitivity varies according to the clinical presentation, but in disseminated disease, sensitivity is around 87%.⁶

In cases of cutaneous tuberculosis, the skin lesions present with few or no detectable Mycobacterium bacilli.¹²

Correct answer 2: Histoplasma antigen

The landscape of histoplasmosis diagnosis has changed since the introduction of antigen detection tests. Detection of Histoplasma antigen in urine specimens by a commercial Histoplasma enzyme immunoassay (EIA) has good diagnostic performance and has been extensively validated in two cohorts of people living with HIV from Guatemala and Colombia, with a sensitivity of 86% and a specificity of 94% (cutoff > 0.84 ng/mL)¹⁴  in this study as well as in many other recently published studies.¹⁵ Some cross reactivity with other dimorphic fungi, such as Paracoccidioides spp.,¹⁴ and Emergomyces africanus, has been reported.¹⁶ The test is relatively easy to perform and samples are inexpensive to obtain—and the results can be obtained on the same day, with some formats in a few hours. It is also useful in the follow-up of therapy and post-therapy.¹⁴ The performance of Histoplasma antigen tests is superior to all other diagnostic tests, with a sensitivity and specificity over 90%. Detection of Histoplasma antigen has been shown to diagnose more cases when compared to conventional methods and have an impact on patients’ survival.¹⁷

The detection of antibodies using conventional serologic methods is less sensitive when performed in immunocompromised individuals. Antibodies may be negative due to the immunocompromised status of the patient; also, antibodies may take 6-8 weeks to appear and be detected. ^6,18,19

Molecular testing may be a desirable alternative for correct diagnosis, however, a lack of commercially available kits and standardized methods and the limited number of validation studies, limit their broad use in clinical microbiology laboratories. Investigators have yet to determine the optimal DNA extraction method, gene target and primers, or amplification methodology.^6,18,19

Correct answer 2: No

Itraconazole and rifampicin should not be administered simultaneously, since itraconazole is a potent inhibitor of the CYP3A4 enzyme system, which induces a large number of drug interactions. Rifampicin, being an inducer of this enzyme system, decreases the therapeutic effect of itraconazole, leading to therapeutic failure.²⁴

According to the guidelines proposed by PAHO and WHO for diagnosing and managing disseminated histoplasmosis among people living with HIV, there are several recommendations, ideally maintaining anti-tuberculosis treatment with rifampicin.

• Extend induction therapy with amphotericin B to 1 month and perform the recommended transition to itraconazole in the maintenance phase.⁸
• If the patient presents intolerance to the treatment with amphotericin B for a month or it is not possible to give this treatment for a month, in this case amphotericin B can be replaced by another azole different from itraconazole, with similar MIC (minimum inhibitory concentration) for Histoplasma spp.^6,8,25 Do not use fluconazole because of high rates of resistance.²⁵
• Replace rifampicin by rifabutin, which leads to less propensity for drug interactions.⁸
• In cases of administering simultaneously rifampicin and itraconazole, therapeutic drug monitoring should be carried out to ensure adequate itraconazole concentrations in the blood.⁸ 

When it is not possible to administer liposomal amphotericin B for prolonged periods of time and rifabutin is not available, rifampicin can be replaced by a fluoroquinolone such as moxifloxacin or levofloxacin. However, this choice prolongs the anti-TB treatment due to lack of rifampicin.²⁶ Although the literature supporting this approach is scarce,²⁶ it is an alternative that is being used in the treatment of patients with tuberculosis resistant to rifampicin.²³