Common invasive fungal diseases : an overview of invasive candidiasis , aspergillosis , cryptococcosis , and Pneumocystis pneumonia

Every year, Candida, Aspergillus, Cryptococcus and Pneumocystis infect an estimated two million individuals worldwide. Most are immunocompromised or critically ill. Candida is the most common fungal pathogen of the critically ill and of recipients of transplanted abdominal organs. In high-risk haemato-oncological patients, in contrast, the introduction of antifungal prophylaxis with fluconazole and later with mould-active posaconazole has led to a remarkable reduction of invasive candidiasis and is likely to have a similar effect on invasive aspergillosis. Invasive aspergillosis remains the dominant invasive fungal disease (IFD) of haemato-oncological patients and solidorgan transplant recipients and is increasingly found in individuals with exacerbated chronic obstructive pulmonary disease on corticosteroids. In the developed world, owing to antiretroviral therapy Pneumocystis pneumonia and cryptococcosis have become rare in patients with human immunodeficiency virus (HIV) and are mainly found in solid-organ transplant recipients or immunocompromised patients. In the developing world, cryptococcosis remains a common and highly lethal disease of HIV positive individuals. With invasive candidiasis and invasive aspergillosis, timely diagnosis is the principal challenge. The clinical presentation is nonspecific and current diagnostic tests lack sensitivity and specificity. The combination of several tests improves sensitivity, but not specificity. Standardised polymerase chain-reaction-based assays may be promising tools for more rapid and specific diagnosis of candidiasis and invasive aspergillosis. Nevertheless, initiation of treatment is often based solely on clinical suspicion. Empirical therapy, however, may lead to over-treatment of patients without IFD or it may miss its target in the case of resistance. Despite the success of antifungal prophylaxis in reducing the incidence of IFDs in haemato-oncological patients, there are a considerable number of breakthrough infections demonstrating not only fungal resistance but also the emergence of rare and often lethal fungal pathogens. Knowledge of the local epidemiology and antifungal resistance is therefore pivotal. Current trial-based guidelines leave major gaps in identifying those most at risk, who may benefit from prophylaxis. Ongoing searches for diseaseassociated genetic polymorphisms may contribute to the establishment of individual risk profiles and targeted prophylaxis.


Introduction
Out of more than 100'000 known fungal species, only about 300 cause disease in humans [1].Our body temperature may provide a protective thermal barrier against the majority of species that grow best at ambient temperature [2].The most common pathogens are Candida, Aspergillus, Cryptococcus, and Pneumocystis spp.-causing more than 90% of reported deaths due to fungal disease [3].The top ten fungal infections are responsible for at least as many deaths as tuberculosis or malaria [3][4][5].Yet, on a global scale, fungal diseases are neglected.This is reflected by the lack of initiatives by the World Health Organization and the paucity of national surveillance programmes [3].Although current trends show an overall increase of invasive fungal diseases (IFDs), their incidence is likely to be underestimated [6].IFDs are associated with high morbidity and mortality.Their diagnosis is challenging and their timely treatment often depends on a high level of clinical suspicion.Therefore, this review aims to give an overview of the current epidemiology, clinical presentation, diagnosis, and management of the four most common IFDs: invasive candidiasis, aspergillosis, cryptococcosis, and Pneumocystis pneumonia.

Definitions of invasive fungal disease
IFDs are proven by the presence of moulds or yeasts in a deep tissue biopsy or a culture obtained by a sterile procedure [7].Additional definitions for probable and possible IFDs based on host-specific, clinical, and mycological features, which were originally established for haemato-oncological research purposes, are now commonly applied in the clinical setting [8].However, these definitions lack pre-cision and universal applicability to all patients groupsnotably the critically ill -and do not include Pneumocystis pneumonia.

Invasive candidiasis
Candida is the most common cause of IFDs in the developed world [9].As a normal commensal of humans, this yeast may be found in the oral cavity, the gastrointestinal tract, the female genital tract or on the skin [10].An estimated 24-70% of healthy people above one year of age are colonised by Candida.Presumably, everyone is temporarily colonised at least once during their lifetime [11,12].Invasive candidiasis originates from the patient's own flora: it is introduced into the bloodstream or deep tissue following iatrogenic breaks of the skin or mucosal barrier.Since its introduction in the 1990s, antifungal prophylaxis with fluconazole has led to a remarkable reduction of candidaemia in haemato-oncological patients [13][14][15][16].Invasive candidiasis is predominantly a disease of the critically ill, hospitalised patient.

Epidemiology
The incidence rate of invasive candidiasis shows a large geographical variation.In Europe, invasive candidiasis makes up 2-3% of all nosocomial infections -four times less than in the USA [9,[17][18][19][20].Despite an increased use of antifungals, the incidence of candidaemia is on the rise.Between 2000 and 2010 its incidence rate in Swiss hospitals has doubled from 0.49 to 1.01/10 000 patient days [21].A similar trend was observed in other European surveillance studies, with an average incidence rate increasing to approximately 0.59/10 000 population and 11/10 000 admissions [22][23][24][25].Incidence rates in intensive care unit (ICU) patients are 5-10 times higher than in patients from medical or surgical wards [20,22,26].The average 30-day mortality rate for candidaemia is 43% [27,28].This is substantially higher than for any other blood stream infection [29].The main risk factor and principal portal of entry for candidaemia is an intravenous (IV) catheter [30].Candida biofilms formed on IV catheters are also an important source of continued infection [31].Other critical risk factors for candidaemia are the use of broad-spectrum antibiotics, total parenteral nutrition, dialysis, and chemotherapy.Notably, all of these factors are associated with critical illness [32][33][34].In the USA TRANSNET study on solidorgan transplant recipients, more than 50% of all fungal infections were due to Candida with incidence rates increasing from 1.4% to 2.1% between 2001 and 2006.In particular, recipients of abdominal organs (liver, pancreas, small bowel) were likely to become infected [35].The principal risk factors for invasive intra-abdominal candidiasis are complicated abdominal surgery and pancreatitis, both often leading to ICU admission.Its mortality rate in recent studies was between 27-38% [36,37].

Different Candida species
Worldwide, C. albicans remains the dominant Candida species.Between 1991 and 2010, Funginos, the "Fungal infection network of Switzerland", reported a stable distri-bution of blood-stream isolates of 65% C. albicans, 15% C. glabrata, 6% C. tropicalis, 5% C. parapsilosis and 2% C. krusei [21].In contrast to the USA, in Switzerland and Northern Europe no shift to fluconazole-resistant C. krusei and C. glabrata has been observed [27,38].In some areas (Southern Europe, the Americas and Asia) C. parapsilosis is the second most common isolate after C. albicans.The identification of Candida isolates to the species level is important because of species-specific antifungal drug resistance patterns, which have major impact on the choice of best treatment.More than 98% of C. albicans isolates are susceptible to fluconazole [39], whereas C. krusei is constitutiively resistant to fluconazole, but susceptible to newer-generation azoles.Because of its wide array of resistance mechanisms, C. glabrata poses the greatest treatment challenge [23,27]: azole therapy is usually advised against because of the capacity of C. glabrata to develop or to extend resistance under treatment, and some studies also report rapidly occurring and/or increased resistance to echinocandins [40][41][42].Previous azole exposure increases the risk for infection with azole-resistant Candida isolates [43][44][45][46].

Clinical presentation
Signs and symptoms of invasive candidiasis are nonspecific.Candidaemia is the most common manifestation, deepseated infections with or without concomitant candidaemia are rarer.Intra-abdominal candidiasis may occur after complicated abdominal surgery or with necrotising pancreatitis; simultaneous candidaemia is detected in only 10% of cases [47][48][49].Candidaemia may go unnoticed when it produces a single febrile peak among many -typically in the critically ill patient.It may induce sepsis or ultimately septic shock, both indistinguishable from bacterial infection [43].Signs of intra-abdominal candidiasis include persistent fever and clinical deterioration despite continued antibiotic treatment.Deep-seated organ infections are more common in solid organ transplant recipients, whereas candidaemia occurs more often in haematological or critically ill patients [50].Candida pneumonia is rare [11,51].Candida chorioretinitis, found in approximately 8-16% of candidaemic patients, may be an easily detectable sign of recent or ongoing candidaemia.It is usually asymptomatic unless progression to vitritis and vision-threatening endophthalmitis produces blurred vision or floating black spots [52,53].Progression beyond chorioretinitis is very rare in patients treated for candidaemia but may occur up to several months later if candidaemia was missed

Diagnosis
The diagnosis of candidaemia is often difficult.Because of the lack of disease-specific signs and symptoms, clinical suspicion drives diagnostic testing [56].Despite their low sensitivity of only 50-70%, blood cultures are still the gold standard to diagnose candidaemia.Intermittent shedding of organisms from deep-seated foci of infection, low pathogen counts per volume of blood or nonviable organisms reduce the sensitivity of modern blood culture methods [56].Frequent sampling is recommended [57][58][59].The diagnosis of deep-seated candidiasis is based on positive cultures of sterilely collected tissue samples, which are often difficult to obtain [56].Molecular tests such as the assay for (1-3)-β-D-glucan, a cell wall component of various medically important fungi (except Cryptococcus and mucormycetes), may detect candidaemia and intra-abdominal candidiasis.However, the low sensitivity (65-75%) and specificity (80-85%) limit the usefulness of the test, which is not commonly offered by European microbiology laboratories [57,60,61].A commercial, whole-blood, multiplex polymerase chainreaction (PCR) assay that detects the five clinically most important Candida species, Aspergillus fumigatus as well as several bacteria had a sensitivity of 94% in candidaemic patients [62].The T2 magnetic resonance assay, an automated, PCR-based method, appears to be a promising new test for the rapid diagnosis of candidaemia [63].MALDI-TOF (matrix-assisted laser desorption/ionisation time of flight) has been shown to accelerate correct identification of Candida species [64].To search for endophthalmitis, a dilated fundoscopy is recommended for all candidaemic patients with any visual symptom or those unable to report symptoms (i.e.sedated patients) [52].The detection of Candida in a respiratory or urinary sample is almost always due to colonisation [11,65].Symptomatic candiduria in a neutropenic patient, however, may indicate a Candida cystitis that merits therapy.Persistence of candiduria after catheter removal may be due to colonised foreign bodies in the urinary tract (i.e.kidney or bladder stones) and requires imaging studies.

Treatment
Indications and agents for antifungal prophylaxis are listed in table 1. Antifungal therapy (table 2) is indicated in all patients with evidence of Candida in their blood culture regardless of the presence of clinical symptoms [66].As a result of the nonspecific clinical presentation as well as the poor sensitivity of diagnostic tests, treatment is mostly initiated on the basis of clinical suspicion.Following the latest European Society of Clinical Microbiology and Infectious Diseases (ESCMID) guidelines all non-neutropenic candidaemia patients -regardless of the underlying diseaseshould primarily receive an echinocandin or alternatively liposomal amphotericin B [66][67][68].For neutropenic patients (mostly after haematopoietic stem cell transplantation [HSCT]) echinocandins and amphotericin B are both first choice [69].For susceptible organisms, fluconazole may be a reasonable choice for continuation therapy in clinically improved patients.For all non-albicans Candida spp.treatment is adapted in accordance with resistance testing.They should initially be treated either with an echinocandin or amphotericin B [66].Treatment for candidaemia should be continued for 14 days after the first negative blood culture [70].All patients with candidaemia should have daily blood cultures until clearance of the blood stream infection is documented.In a suspected catheter-associated infection, the invasive device should be removed as soon as possible [66,69].The 2016 update of the Infectious Diseases Society of America (IDSA) guideline includes suggested treatment durations for some deep-seated infections [71].Combination therapy is generally not recommended.Chorioretinitis is usually sufficiently treated with the systemic antifungal therapy for candidemia [54,55].Candida endophthalmitis extending into the vitreous body commonly warrants surgical intervention as well as intraocular antifungals.

Aspergillosis
Aspergillosis is the most common mould infection in humans, accounting for >85% of invasive mould disease [72].Aspergillus is found in soil, decaying vegetation, food, air, and the water supply [73].Its ubiquitous spores reach the respiratory tract by inhalation.In immunocompromised hosts this results primarily in invasive pulmonary aspergillosis (IPA).Disseminated disease, the end-stage complication of IPA, involves predominantly the brain and kidneys.

Epidemiology
Worldwide there are 200 000 estimated annual cases of invasive aspergillosis (IA) [3].At present, approximately 50% of all IA are found in patients with haematological malignancy, mostly acute myeloid leukaemia (AML), acute lymphoblastic leukaemia (ALL), and recipients of allogeneic HSCT [15,74].Prolonged severe neutropenia (>10 days; <500 cell/mm 3 ) as a result of chemotherapy is still the single most important risk factor for IA.The introduction of mould-active prophylaxis with posaconazole after 2007 [13,14] is likely to result in a reduction of IA incidence in haemato-oncological patients (AML, HSCT): a French prospective single-centre study described not only  a reduction in IA incidence in AML patients with posaconazole prophylaxis (7.3%) versus those without prophylaxis (15.5%) but also a decrease in mortality at day 100: 3.6% versus 10.6%, respectively [75].An Austrian singlecentre study found unchanged rates of IFDs but no IA in AML, ALL and HSCT patients after introducing posaconazole prophylaxis.This indicates a remarkable decrease of IA, but also a high rate of breakthrough infections [76].
In keeping with this finding, the continued use of fluconazole for prophylaxis in almost 50% of HSCT recipients in a recent Swiss multicentre study resulted in almost 60% IAs among all proven and probable IFDs [77].Mortality rates of haematological patients with IA in large clinical treatment trials reached 29% at 3 months [14,78].
In SOT recipients Aspergillus causes one out of five IFDs; it is most commonly found in lung and heart-lung recipients.The 5-year cumulative incidence in all SOT recipients of the TRANSNET study remained stable at 0.7%, with a median time to diagnosis of 184 days [35,79].The dominant risk factor in SOT recipients is not a depletion of phagocytic neutrophils but their functional impairment by immunosuppressant drugs.ICU patients represent the second largest at-risk population for IA.Reported incidence rates are variable but generally high (6.1 to 57/1 000 ICU admissions) [80,81].Underlying malignancy in the ICU population is rare.Instead, patients often present with exacerbations of chronic obstructive pulmonary disease, which are treated with high-dose corticosteroids.Severe alcoholic liver cirrhosis is another more recently reported risk factor in ICU patients [82,83].Crit-ically ill patients with IA have remarkably high mortality rates ranging from 46 to 80% [84,85].Only patients with cerebral involvement, regardless of the underlying disease, reach higher rates of 90% at 4 months [86].
IA has also been associated with concurrent viral respiratory infections due to H1N1 influenza, adenovirus [87], and cytomegalovirus.Particularly in transplant recipients, infection with cytomegalovirus exerts a powerful immunosuppressive effect [88,89].In addition, genetic factors such as toll-like receptor-4 (TLR4) haplotypes [90] and pentraxin 3 (PTX3) deficiency [91,92] have been associated with an increased risk of IA.Ddespite a large number of patients at-risk, only a minority eventually develop IA.

Clinical presentation
Like patients with invasive candidiasis, none of the at-risk groups show specific clinical symptoms.In neutropenic patients fever unresponsive to broad-spectrum antibiotics is often an early sign that should prompt further examination.Rarely, a cough or chest pain with or without haemoptysis -both signs of pulmonary infarction due to mould-induced vascular obstruction -are reported [93].On the other hand, the critically ill ICU patient with IA is mostly mechanically ventilated and may present with deteriorating lung function and refractory fever [86].Progression to disseminated disease is often an underdiagnosed complication, particularly in patients with severe or advanced underlying disease [94].Seizures or other focal neurological signs may be a late manifestation of cerebral dissemination [93].Primary extrapulmonary organ manifestation is rare [11].

Diagnosis
The diagnosis of IA is challenging and requires a high level of suspicion.The gold standard for proving invasive fungal infection is histopathology or culture of a tissue sample, which is rarely available on time because of the risks involved in performing biopsies during pancytopenia [8].Detection of galactomannan antigen (GM), a constituent of the hyphal cell wall, in blood is currently the most widely available albeit only moderately accurate noninvasive test for invasive aspergillosis.It has a sensitivity of approximately 71% and a specificity of approximately 89% in highrisk haematological patients [95].In corticosteroid-treated nonhaematology patients, GM detection in bronchoalveolar lavage (BAL) fluid has higher sensitivity for IPA than in blood [96].Caution is warranted in patients on mouldactive prophylaxis as the sensitivity of the GM assay is reduced [97], making it difficult to identify breakthrough infection in haematological patients [98,99].Repeat serological tests of (1-3)-β-D-glucan may be used as adjunct in the diagnosis of aspergillosis, although as a cell wall constituent shared between Aspergillus spp., Candida spp., and Pneumocystis jirovecii (but absent from Cryptococcus and mucormycetes), it is nonspecific and a high rate of falsepositive results may render its interpretation difficult [100,101].Blood cultures are virtually always negative in IA [3] and otherwise represent mostly contamination [8].PCR may be a useful tool to diagnose aspergillosis.A recent review reports mean sensitivity and specificity of a single PCR test (plasma and serum) of Imaging studies often provide the first evidence of IA.In neutropenic patients with fever not responding to antibiotic treatment, a high-resolution computed tomography (CT) scan of the thorax may reveal sharply demarcated nodes or dense lesions surrounded by a ground-glass halo compatible with, but not proof of, a fungal aetiology (fig.1a).In clinical practice, it may be difficult to dismiss the possibility of a fungal aetiology based on the radiomorphology of a lesion [104,105].Whereas in neutropenic patients centrally cavitating nodules (crescent sign) are a late sign of invasive fungal infection appearing with the recovery from neutropenia, cavitating pulmonary lesions may be an early sign in, for example, rheumatology patients treated with corticosteroids (fig. 1b).In contrast, in patients on steroidtreatment due to exacerbated chronic lung disease, the CT may only show non-specific pulmonary infiltrates [106].

Treatment
Mould-active prophylaxis is recommended for the duration of the neutropenia associated with remission-induction chemotherapy in high-risk patients with AML or myelodysplastic syndrome.Indications for prophylaxis are given in table 1. Starting treatment early is important in order to reduce mortality.First-line treatment for IA is voriconazole (table 2).Compared with amphotericin deoxycholate it showed a survival benefit in haematological patients (71% vs 58%).Furthermore, voriconazole has a more favourable adverse effect profile and, notably, no nephrotoxicity [78].However, because of extensive drugdrug interactions and large inter-and intra-individual variation in serum levels, therapeutic drug monitoring is advisable when using voriconazole.An equivalent alternative may be liposomal amphotericin B, though no comparative studies have been conducted.Although most Aspergillus isolates exhibit good susceptibility to mould-active azoles as well as amphotericin B, studies from the Netherlands, Belgium and Germany have reported an increasing number of azole-resistant clinical Aspergillus isolates [107][108][109].
It is recommended to continue treatment for invasive aspergillosis until all clinical signs and symptoms have resolved.
In haematological patients combination therapy with anidulafungin and voriconazole showed no significant survival advantage over monotherapy with voriconazole [110].Nevertheless, concomitant treatment with two antifungal drugs of different classes may occasionally be used to extend the spectrum of activity when the pathogen is unknown.Suspected coinfection with different fungal species may be another reason for combining antifungal agents.
Other measures such as reduction of iatrogenic immunosuppression should be considered [111].

Epidemiology
With more than one million cases and about 650 000 deaths annually, cryptococcosis is the most prevalent fatal fungal disease worldwide [3,112].The most common disease manifestation is cryptococcal meningitis presenting as an opportunistic infection principally in the HIV positive with CD4 cell counts below 100 cells/μl or the iatrogenically immunosuppressed [113].Cryptococcal meningitis is rare in Europe but in sub-Saharan Africa annual incidence rates range from 100/100 000 in high HIV prevalence settings to 4 000/100 000 among the HIV positive [114,115].The dominant species in this yeast genus is Cryptococcus neoformans, a ubiquitous environmental pathogen.Following inhalation, cryptococcal yeasts are deposited in the lung where they may cause life-long latent infection [116].In a USA study, 70% of 120 children older than five years were seropositive for C. neoformans, indicating oligosymptomatic infection early in life.Only in the immunosuppressed patient, usually after adolescence, infection may reactivate [117].Globally, HIV remains the leading risk factor for CM, although the widespread use of antiretroviral therapy has led to a steady decline in HIV-associated CM [112,113].In the developed world CM is now seen in approximately 2.8% of all SOT recipients, largely kidney transplant recipients.Infection occurs mostly >18 months after transplantation [118,119].Rarely, cryptococcosis is seen in apparently immunocompetent hosts.A USA study found that, in non-HIV and non-SOT patients, time to diagnosis was prolonged and outcome was worse compared with HIV positive individuals and SOT recipients [118].Apparently immunocompetent patients are more likely to have pulmonary disease and less likely to develop CM [120].Disseminated disease can reach any organ, but the lungs and the skin are predominately affected [11].

Clinical presentation
Symptoms of CM are nonspecific.The clinical picture ranges from minimal to severe symptoms with headache, malaise, fever, visual disturbance, nausea and vomiting.
In contrast to bacterial meningitis, meningism is rare [11,113].Disease onset is variable, but tends to be more insidious in the immunocompetent than in those with advanced HIV disease.Intracranial pressure is commonly raised and seizures may occur in advanced disease.Pulmonary disease produces nonspecific symptoms such as cough, fever and malaise.Skin lesions, which predominately occur in immunocompromised patients, may present as pustules, papules, ulcers, cellulitis, superficial granulomas, or abscesses [121,122].

Diagnosis
Diagnosis of CM is established by positive fungal culture or detection of cryptococcal antigen (CrAg) in cerebrospinal fluid.The CrAg test has a sensitivity and specificity of >90% in both cerebrospinal fluid and serum [123].In developing countries, a point-of-care urine dipstick test has shown promising results [124].The diagnosis of pulmonary disease is based on a positive sputum culture in conjunction with compatible imaging findings.A positive serum CrAg in pulmonary disease may represent disseminated disease [125].Extracranial disease manifestation warrants lumbar puncture to rule out possibly asymptomatic meningitis [126,127].

Treatment
The current treatment guidelines for CM are based on the results of a randomised clinical trial showing superiority of flucytosine over fluconazole in combination with amphotericin B deoxycholate for induction therapy (table 2) [127,128].Clinically, control of intracranial pressure is pivotal: if the initial intracranial pressure exceeds 250 mm, it should be decreased by 50% or at least to <200 mm.Repeated daily drainage may be necessary [127].For pulmonary disease, recommended treatment regimens vary with disease severity: severe disease is treated as CM [129], moderate disease responds to oral fluconazole [11].
In CM patients with HIV, initiation of antiretroviral therapy needs to be delayed for, generally, 5 weeks in order to avoid early excess mortality [130].In the early phase of antiretroviral therapy a potentially lethal immune-reconstitution inflammatory syndrome occurs in approximately 14-30% of individuals successfully treated for cryptococcosis [131].

Epidemiology
Traditionally considered a protozoan, Pneumocystis has been shown to belong to the family of fungi.An important reservoir of this opportunistic organism is children, most of whom are infected early in life and may transmit the organism from person to person via the airborne route [11,132].It is currently unclear whether clinical disease results from recent reinfection or from reactivation of latent infection in the naturally colonised host [133].It is estimated that approximately 400 000 individuals are affected by Pneumocystis jirovecii pneumonia (PCP) every year -as many as by C. albicans [3].In the Western world, PCP used to be the most common opportunistic infection in HIV-positive individuals [134].With the introduction of antiretroviral therapy, the incidence of PCP has significantly decreased.Today, the principal risk group comprises patients who are iatrogenically immunocompromised because of either malignancy, transplantation or rheumatological disease.In a recent European study PCP was the second most common invasive fungal infection in a mixed HIV-positive and -negative population, with an incidence rate of 1.5/100 000 and a fatality rate of 9.5% [135].Counterintuitively, mortality rates in HIV-negative patients were higher (30-60%) than in HIV-positive individuals (10-20%) [135][136][137].

Clinical presentation
The clinical course depends on the underlying disease.Generally, in HIV-positive patients the onset of disease is insidious and prolonged, whereas in non-HIV patients dis-ease progression tends to be more fulminant -possibly explaining the increased mortality in this group.Clinical symptoms are nonspecific including low-grade fever, dry cough and progressive dyspnoea.Lung auscultation is commonly normal [138].

Diagnosis
The diagnosis is confirmed by the detection of the fungal organisms obtained via BAL or induced sputum using fluorescence microscopy in conjunction with a radiological image [139,140].Radiological signs include bilateral perihilar interstitial infiltrates [141].In the early disease stage a chest x-ray may be normal.High-resolution CT is more sensitive, revealing perihilar ground-glass attenuation or cystic lesions [142].In patients who cannot tolerate bronchoscopy, the serum (1-3)-β-D-glucan is a good adjunct marker with a test sensitivity of 96% and specificity of 87% [143].The diagnostic yield of any specimen from HIV patients is higher than from HIV-negative patients [144].
Several trials have shown good sensitivities and specificities of real-time PCR [145,146].However, so far there is no standardised method and the additional problem of distinguishing between (frequent) colonisation and infection is unresolved.

Treatment
Prophylaxis should be given to all those at risk (table 1).
Owing to lack of ergosterol in the plasma membrane, Pneumocystis jirovecii is insensitive to polyenes and azoles [11].Current treatment regimens (table 2) are primarily based on trimethoprim/sulfamethoxazole.In HIV patients with an initial PO 2 <70 mm Hg, the additional administration of corticosteroids has resulted in improved survival [147].The treatment duration is only well established in HIV-positive patients and lasts for 3 weeks, while non-HIV patients are commonly given a 14-day course.In the case of intolerance to trimethoprim/sulfamethoxazole, clindamycin plus primaquine, atovaquone or trimethoprim/ dapsone are alternative regimens [148].

1
Figure 1 Computed tomography images of invasive pulmonary aspergillosis.a) Neutropenic patient, early stages: sharply demarcated nodes and dense lesions surrounded by a ground-glass halo.b) Rheumatology patient on corticosteroid treatment, early stages: centrally cavitating dense lesions.

Figures (large format) Figure 1
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