Diagnosis and Management of Cryptococcal Disease in Resource-Limited Settings
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- Vanselow, M., Brandt, M.E. & Park, B.J. Curr Fungal Infect Rep (2012) 6: 35. doi:10.1007/s12281-011-0082-6
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Cryptococcal meningitis is one of the most important fungal infections in the developing world, where deaths related to this disease are numerous. In resource-limited settings, mortality is high in large part because of difficulties in the diagnosis and management of this infection. This paper outlines many of the realities in many resource-limited settings, and describes priorities for public health action and research.
KeywordsCryptococcusCryptococcosisCryptococcal meningitisDiagnosisManagementHIV/AIDSResource-limited settingsAmphotericinFluconazoleLateral flow assayCryptococcal antigenLatex agglutination
Cryptococcus infections are typically caused by C. neoformans and C. gattii, which are ubiquitous environmental fungi associated with soil enriched with bird droppings and with various trees, including eucalyptus and almond trees . Exposure to Cryptococcus occurs via inhalation of cryptococcal spores, which may result in a primary pulmonary infection or an asymptomatic infection. Later, primary symptomatic or asymptomatic infection may advance to disseminated disease . The most frequent manifestation of disseminated disease is meningitis, especially in immunocompromised hosts such as individuals with HIV/AIDS.
C. neoformans, specifically, is a major opportunistic pathogen worldwide that causes cryptococcal meningitis (CM) [3••]. CM is an important cause of morbidity and mortality, especially in HIV-infected patients in resource-limited settings. In developed countries, the widespread use of highly active antiretroviral therapy (HAART) has dramatically reduced the incidence of cryptococcal disease [4–8], but in countries with limited access to HAART or other healthcare resources, and where many patients are not yet receiving HAART, the incidence of cryptococcal infection and the resulting mortality remain high [9–15]. The global burden of CM is estimated at almost 1 million cases, with over 700,000 of those cases in Africa and 120,000 in South and Southeast Asia [3••]. The annual mortality from CM is estimated at about 625,000 deaths, with over 500,000 deaths per year in sub-Saharan Africa [3••].
Resources and capabilities at three infrastructure levels common in the developing world
Health-care setting (personnel)
Summary of resources and capabilities
No laboratory infrastructure
In the community or home (possibly healthcare worker, pharmacist, or family member)
No electricity or clean water available; no trained personnel; no laboratory space; cold storage not available; room temperature not controlled; venipuncture impossible; sputum impossible to process; rapid answer required to prescribe treatment before patient leaves; no physician oversight
Minimal laboratory infrastructure
Health clinics in Africa; rural health clinics in Latin America and Asia (nurse)
No reliable electricity and clean water; minimal trained personnel; no or minimal laboratory space; cold storage occasionally available; room temperature rarely controlled; venipuncture unlikely; sputum difficult to process; rapid answer required to prescribe treatment before patient leaves; no physician oversight
Moderate to advanced laboratory infrastructure
Urban health clinics in Asia and Latin America; hospitals in Africa, Latin America, and Asia (nurses, technicians, and physicians)
Dependable electricity and clean water available; trained personnel available; dedicated laboratory space; cold storage available; room temperature sometimes controlled; venipuncture routine; sputum acceptable (except for children); time to answer usually less crucial with hospitalized patients, but still important for clinical patients; physician oversight routine
Various modalities are available for the detection of Cryptococcus species, often depending on existing laboratory capacity, cost, and the availability of cerebrospinal fluid (CSF) for testing.
Microscopy and Culture
Microscopic examination of CSF using an India ink preparation, at a cost of less than $1 per test, is frequently performed in resource-limited settings. It is reasonably specific with adequate technical training, but it is less sensitive than antigen-based assays and is dependent on the fungal burden in the CSF [17, 18]. Culture of CSF or blood is considered the gold standard for cryptococcal infection diagnosis, but like the India ink stain, culture has limited sensitivity (56% sensitivity for culture; 72% for India ink) ; the likelihood of a positive result is higher in patients with a high fungal organism burden, no prior antifungal treatment, or both.
These tests require basic laboratory equipment (microscope, slides, cover slips, etc.), which may not be available in many areas of sub-Saharan Africa. For example, it was estimated that there was less than one working microscope per 100,000 people in Malawi in 2000, and none of the microscopes being used had ever received service or repair . Similar results were seen in Ghana, with fewer than two microscopes available per 100,000 people . Even if the equipment is available, it is not likely to be reliable. It is estimated that 40–70% of equipment in resource-limited settings is out of service . Lack of dependable electrical service puts even greater limitations on equipment use. For example, in Uganda, the majority of medical facilities in 50 of 54 districts were without electricity (http://apps.who.int/healthinfo/systems/datacatalog/index.php/catalog/9).
For accurate analysis, both the India ink test and culture require experienced technicians to correctly prepare the examinations and identify the fungus, but properly trained technicians are not prevalent in many resource-limited settings. In Ghana, for example, of the 693 laboratory technicians working in 205 laboratories, 74% had no professional qualifications, with as little as 6 weeks to 1 year of training . Other studies in Africa showed that a large proportion of basic diagnostic test results were being read inaccurately, often leading to misdiagnoses [22, 23].
Cryptococcal Antigen-Based Tests
Latex Agglutination and Enzyme Immunoassay
Antigen tests (latex agglutination [LA] or enzyme immunoassay [EIA]) of CSF or serum have many advantages over culture and microscopy, but many limitations make them less feasible in resource-limited settings. They have high specificity and sensitivity (>90% in both) and require less time and labor than culture [24–26]. Results from LA are fairly quick (less than an hour), are less subject to interpretation than India ink, and require minimal technical training to perform. EIA, on the other hand, requires more advanced laboratory expertise and infrastructure, including a refrigerator, freezer, centrifuge, and microplate reader. Both LA and EIA are more expensive than microscopy and culture, with estimates for the full cost of testing (including supplies, labor, and laboratory overhead) ranging from $5.61 per test in South Africa to $16.43 in Uganda [27••]. With these limitations, cryptococcal antigen tests may be available only in larger reference and diagnostic laboratories. When specimen collection is far from these laboratories, a cold chain for transporting specimens is needed, making it less likely that specimens will arrive in adequate condition for testing [23, 28].
Lateral Flow Assay
A new cryptococcal antigen assay, called the lateral flow assay (LFA; Immuno-Mycologics, Norman, OK, USA), has recently been approved by the US Food and Drug Administration for use in serum. Similar to the LA and EIA, the LFA detects the presence of cryptococcal antigen by using immunochromatographic test strips coated with Cryptococcus-specific monoclonal antibodies and gold particles. The test strips are placed in a drop of diluted serum or CSF and incubated at room temperature for 10 minutes. Unlike LA, which requires a pronase treatment step, LFA requires no pretreatment of the specimen. A positive result occurs when two bands (control and test) develop in the test zone, whereas a negative result is indicated with the presence of a single band (control). The sensitivity and specificity of the LFA are similar to those of EIA and LA [29••, 30••].
World Health Organization ASSURED criteria: characteristics of ideal diagnostic tests for resource-limited settings
Affordable by those at risk of infection
Sensitive (few false negative results)
Specific (few false positive results)
User-friendly (simple to perform by persons with little training)
Rapid treatment at the first visit and Robust use without the need for refrigerated storage
Equipment-free (easily collected, noninvasive specimens)
Delivered to those who need it
Early initiation of treatment for CM is extremely important. Without appropriate antifungal treatment for HIV-infected patient populations with CM, mortality rates reach 100% within 2 weeks after arrival at a healthcare facility . Management is achieved through administering combination antifungal therapy by the principles of induction, consolidation, and maintenance therapy, along with reducing elevated intracranial pressure .
First-line pharmacotherapy for induction and consolidation therapy of CM includes the 2-week fungicidal regimen of amphotericin B (AmB) plus flucytosine, followed by maintenance therapy with fluconazole for a minimum of 8 more weeks. However, in many resource-limited settings, AmB is not available or is too expensive. Additionally, the use of AmB requires appropriate facilities for the administration of intravenous therapy, and because it can cause nephrotoxicity, facilities must be able to provide quick and reliable potassium and renal monitoring.
Such capacities, unfortunately, are relatively uncommon, particularly in rural areas. Of 100 medical sites assessed in Mexico, Vietnam, Ghana, and India, none of the 51 rural sites had electrolyte analysis available, and none of the small or large hospitals in Ghana and India had these capabilities . Even if facilities with these capacities do exist, there are other general barriers to accessing medical care, including lack of transportation, loss of income with hospitalization, and the cost of treatment . One or all of these limitations can affect the ability to administer AmB. Flucytosine (which is administered orally) is recommended as an adjunctive therapy to AmB , but although it is more easily administered, it is very expensive and currently is available in Africa only through clinical trials [34•]. Flucytosine should not be administered as a single therapy.
In contrast, fluconazole is widely available, thanks to the Pfizer Diflucan® Partnership Program (diflucanpartnership.org), which since 2000 has distributed free fluconazole to many parts of Africa for the treatment of CM and esophageal candidiasis. Fluconazole is currently the only antifungal treatment option in many areas. Fluconazole is orally administered, is well tolerated, and requires minimal laboratory and clinical monitoring. Monotherapy with high doses of fluconazole is an alternative treatment regimen to AmB and flucytosine .
Management of Elevated Intracranial Pressure
The elevated intracranial pressure that frequently occurs in CM is associated with high mortality, but this is often difficult to manage in resource-limited settings. Where advanced imaging is available, the results of a CT scan or MRI are recommended prior to diagnostic lumbar puncture in patients with focal neurologic signs or altered mental status . If scans are not available, lumbar puncture without CT or MRI is acceptable because contraindications to lumbar punctures (such as CNS mass lesions and cryptococcomas) are rare, and the potential benefits of lumbar puncture outweigh its risks . Subsequently, frequent lumbar punctures during therapy are often required to relieve intracranial pressure.
In developing countries, many patients do not receive this important treatment component because equipment or trained personnel are lacking. Rural clinics are often staffed only by nurses, with no regular doctors available . Patients who are unable to receive needed lumbar punctures need to be transferred to facilities that can appropriately manage elevated intracranial pressures. However, transfer to a referral center may add levels of complexity and cost to the management of the patient, requiring additional time (resulting in treatment delays), adequate roadways, appropriate transportation, and communication between the healthcare facilities. All of these are common barriers in resource-limited settings .
Early Diagnosis and Treatment
Early diagnosis, followed by early therapy for cryptococcal disease, may be the most practical strategy in resource-limited settings and has great potential to reduce mortality from this disease if instituted on a large scale. This strategy involves screening HIV-infected persons with CD4 counts below a certain threshold (usually 100 cells/μL) with a cryptococcal antigen-based test (LA, EIA, or LFA). It has been shown that cryptococcal antigen can be detected in serum prior to the onset of symptoms, often several weeks before the onset of symptoms [13, 35–38]. Treatment at this point, before the development of overt meningitis, may be accomplished using oral fluconazole alone, thereby avoiding the resource-intensive management of patients requiring AmB and therapeutic lumbar punctures.
Limited data suggest that treating these patients with fluconazole will prevent clinical disease [26, 27••, 39••]. In one recent study in Uganda, integrating cryptococcal antigen screening with early fluconazole treatment was shown to prevent CM and death [27••]. The study found that among the group of patients who were positive for cryptococcal antigen when starting antiretroviral therapy, 71% of those promptly treated with fluconazole were alive at 30 months, compared with 0% of those who were not treated.
Cryptococcal disease is one of the major causes of HIV infection-related mortality in some developing countries, so it remains an important public health issue, but major obstacles in providing adequate treatment include the lack of adequate laboratory infrastructure and medications. In addition to improving access to antiretroviral therapy, public health officials can reduce mortality from this infection by focusing efforts on improving access to diagnostic tests, improving access to antifungal medications, and implementing large-scale strategies to screen for cryptococcal disease.
One of the most promising developments in the public health battle to reduce the burden of cryptococcal disease is the development of the LFA. The ability to use this test with minimal laboratory capacity and as a point-of-care test in resource-limited settings should vastly improve the ability to diagnose cryptococcal disease. Further experience is needed with the use of this test in these settings, as well as with its utility in detecting antigen in other fluids such as plasma, urine, or whole blood.
Improving access to first-line antifungal medications (AmB and flucytosine) may also help in limiting morbidity and mortality. The current cost and distribution of both AmB and flucytosine hinder their use in low-resource settings. However, public health officials may improve access to these medications by initially targeting referral centers, which are most likely to have the infrastructure required to monitor the administration of AmB. Public health education in rural clinics can also help healthcare workers recognize the signs and symptoms of CM for early referral to these sites.
Although the ultimate goal is to have universal HAART coverage, early diagnosis of cryptococcal disease holds enormous potential as a large-scale public health strategy to combat this infection. Early diagnosis is particularly promising in resource-limited settings, where the existing realities make the management of CM so difficult. A public health strategy using reflexive screening for cryptococcal antigen among persons with CD4 counts below 100 cells/μL is in the planning stages in South Africa. Further programmatic research into the optimal implementation of such a screening strategy is needed.
The findings and conclusions in this presentation/report are those of the author(s) and do not necessarily represent the official position of the Centers for Disease Control and Prevention.
No potential conflicts of interest relevant to this article were reported.