Advances in the Understanding of the Pathogenesis of Opportinistic Fungal Diseases.

ZygomycosisGeorge L. Petrikkos

Professor of Internal Medicine and Infectious DiseaseNational and Kapodistrian University of Athens

Greece

Percentages of zygomycosis cases documented by culture since the 1940s, by decade

Roden et al CID 2005:41

60

36

35

22

21

16

12

9

6

5

2

2

2

1

1

Italy

Greece

Germany

Switzerland

France

Belgium

Austria

Spain

Russia

Norway

Finland

Czech Republic

Turkey

Netherlands

UK

G.Petrikkos, and the ECMM Zygomycosis Working Group, ICAAC, 2008.

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Zygomycosis refers to a group of uncommon but frequently fatal mycoses caused by fungi of the class Zygomycetes.

This class is subdivided into two orders,the Mucorales and the Entomophthorales.

This disease is also sometimes referred to as mucormycosis,or phycomycosis but the term zygomycosis is preferred.

The genera in the Order Mucorales cause most human infection.

These organisms are ubiquitous in nature, and can be found on decaying vegetation,fruit, bread and in the soil.

These fungi grow rapidly and release large numbers of spores that can become airborne.

All humans have ample exposure to these fungi during day-to-day activities.

The fact that mucormycosis is a rare human infection reflects the effectiveness of the intact human immune system.

This is further supported by the finding that almost all human infections due to a zygomycete occur in the presence of some underlying compromising condition.

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Healthy humans have a strong natural immunity against Zygomycetes.

Most commonly, the infection presents as an opportunistic disease in patients with underlying risk factors.

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The most important exception to this generalization is the disease caused by Entomophthorales, which commonly affects normal hosts.

Other exceptions are rare reports of zygomycosis caused by the agents of the order Mucorales in immunocompetentindividuals with no identifiable risk factors and in employees of both the malt and lumber industries.

Hematological Malignancy

54%Diabetes10%

Trauma10%

Other6%

Surgery6%

Cancer5%

Solid organ transplantation

5%

Burn3%

AIDS1%

Underlying disease Mortality (%)

Hematological malignancy 57.3

Diabetes 37.5

Trauma 12.5

Burn 20.0

Solid organ transplant 50.0

AIDS 0

Other 50.0

Surgery 50.0

Cancer 55.6

Total 47.8

The genera most commonly found in human infections are :

Rhizopus, Mucor, and Rhizomucor; Cunninghamella, Absidia,Saksenaea, and Apophysomyces are genera

that are less commonly implicated in infection .

Absidia sp.19%

Cunninghamella sp5%

Rhizopus sp33%

Apophysomyces sp1%

Mucor sp30%

Acrophialophora sp.0%

Rhizomucor12%

Zygomycetes enter the human host through inhalation, percutaneousinoculation or ingestion

Cutaneous infection following traumatic inoculation of a high load of sporangiosporesunder the skin is the most common form of zygomycosis in normal hosts.

Prolonged neutropenia Hyperglycemia and acidosis (DKA) Steroids Immunosuppressive therapy Burns, trauma (skin form) Excess iron Deferoxamine (not iron chelators in general) Voriconazole use?

Knowing the pathogenesis helps understand risk factors, manifestations and later on therapeutic implications

Intact mucosal and endothelial barriers serve as structural defenses against tissue invasion by Zygomycetes.

The mechanisms of attachment to and invasion of mucosal surfaces have not been fully clarified.

Spores may invade epithelium previously damaged by infection, cytotoxic chemotherapy or direct trauma.

Kontoyiannis DP, Lewis RE. Invasive zygomycosis: update on pathogenesis, clinical manifestations and management. Infect Dis Clin N Am 2006;20:581-

607.

Recent studies have suggested that zygomycetessporangiospores may have unique capability for tissue adherence and invasion of intact endothelial barriers.

A surprising finding is that viable spores are not essential for causing tissue damage and injection of heat-killed Rhizopus spp. can result in significant mortality in diabetic mice.

This observation suggests that some component of sporangiospores may be directly toxic to endothelial cells in the mucosal membranes.

Kontoyiannis DP, Lewis RE. Invasive zygomycosis: update on pathogenesis, clinical manifestations and management. Infect Dis Clin N Am 2006;20:581-607.

A greater understanding of the pathogenesis of the disease may lead to future therapies. For example, it is now clear that iron metabolism plays a central role in regulating mucormycosis infections and that deferoxamine predisposes patients to mucormycosis by inappropriately supplying the fungus with iron.

Brad Spellberg, John Edwards Jr.,1 and Ashraf IbrahimClinical Microbiology Reviews, July 2005, p. 556-569, Vol. 18, No. 3Brad Spellberg, John Edwards Jr.,1 and Ashraf IbrahimClinical Microbiology Reviews, July 2005, p. 556-569, Vol. 18, No. 3

To cause disease, the agents of mucormycosis must scavenge from the host sufficient iron for growth, must evade host phagocytic defense mechanisms, and must access vasculature to disseminate.

Brad Spellberg,et al Clinical Microbiology Reviews, July 2005, p. 556-569, Vol. 18, No. 3

A) In a normal host, primary defense mechanismsagainst mucormycosis include :

Sequestration of iron in serum by specialized iron-binding proteins (1), phagocytes including circulatingneutrophils (2a) and tissue macrophages (2b), andendothelial cells (3), which regulate vascular tone andpermeability.

Acting in concert, these mechanisms preventestablishment of infection in tissue and subsequentendovascular invasion.

Brad Spellberg, John Edwards Jr., and Ashraf IbrahimClinical Microbiology Reviews, July 2005, p. 556-569, Vol. 18, No. 3

B) In susceptible hosts, normal defense mechanisms break down.

For example, in diabetic ketoacidosis (DKA), the acidic pH of the serum causes dissociation of free iron from sequestering proteins (1).

This release of free iron allows rapid fungal growth. Defects in phagocytic defense mechanisms (2), for example,

deficiency in cell number (neutropenia) or functional defects caused by corticosteroids or the hyperglycemia and acidosis of diabetic ketoacidosis, allow proliferation of the fungus.

Finally, adherence to and damage of endothelial cells by the fungus (3) allows fungal angioinvasion and vessel thrombosis and subsequent tissue necrosis and dissemination of the fungal infection.

Brad Spellberg, John Edwards Jr., and Ashraf IbrahimClinical Microbiology Reviews, July 2005, p. 556-569, Vol. 18, No. 3

M. LIU, B. SPELLBERG, T. GEBREMARIAM, Y. FU, J. EDWARDS, JR., A. S. IBRAHIM; Habor-UCLA Med. Ctr., Torrance, CA. Identification of Rhizopus oryzae Ferrioxamine Receptor

Conclusions: Our results suggest that we have identified inducible receptors for ferrioxamine. Further studies to confirm the role of these receptors in R. oryzae iron uptake from ferrioxamine and pathogenesis of mucormycosis is warranted.

Drosophila melanogaster flies injected with a standardized amount of fungal spores from clinical Zygomycetes isolates

to study virulence & host defense mechanisms

Chamilos G et al. PNAS; 2008 vol. 105 no. 27: 9367–9372

Zygomycetes rapidly infect and kill wild-type flies Toll-deficient flies exhibited susceptibility

Constitutive overexpression of the antifungal peptide Drosomycin in transgenic flies partially restored resistance to zygomycosis

Chamilos G et al. PNAS; 2008 vol. 105 no. 27: 9367–9372

Classic enhancers of Zygomycetes virulence corticosteroids increased iron supply & iron availability - treatment with deferoxamine

dramatically Zygomycetes pathogenicity

Chamilos G et al. PNAS; 2008 vol. 105 no. 27: 9367–9372

iron starvation induced by the iron chelator deferasirox

significantly protected flies infected with Zygomycetes

Chamilos G et al. PNAS; 2008 vol. 105 no. 27: 9367–9372

Chamilos G et al. PNAS; 2008 vol. 105 no. 27: 9367–9372

In normal hosts, the traumatic inoculation ofa high load of sporangiospores under the skinleads to cutaneous zygomycosis.

Extension to the subcutaneous tissue or bone is common in patients who have delayed or ineffectively treated cutaneous zygomycosis

In immunocompromised patients the infection may disseminate.

Rhino-cerebral or cranio-facial (1/3-1/2 of the cases)

Pulmonary Cutaneous Gastro-intestinal: rare Disseminated>90% mortality Others: endocarditis, kidneys, etc..

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Predisposing conditions Clinical presentationImmunosuppression Respiratory tract infection,

disseminated infectionMetabolic Respiratory tract infection,

rhinocerebral infectionDeferoxamine therapy Disseminated infectionSkin and soft tissue breakdown Primary cutaneous and soft

tissue infectionIntravenous illicit drug use Endocarditis, cerebral infectionNeonatal prematurity Gastrointestinal infection,

disseminated infectionMalnourishment Gastrointestinal infection

Acquired by the inhalation of spores Infection starts in the nasal turbinates

then spreads to involve the orbits and the brain

Mucor is angioinvasive and can lead to infarction of infected tissues

It occurs most frequently amongneutropenic patients.

It presents with nonspecific symptoms such as fever, cough and dyspnea; hemoptysis may occur with vascular invasion.

Radiological presentation includes segmental consolidation that progresses to contiguous areas of the lung, with occasional cavitation.

Fatality 80%

It has been reported with minor trauma, insect bites, contaminated dressings and patches, wounds, and burns.The necrotic lesions progressively evolve from the epidermis into dermis and even muscle and bone.

Minor trauma by lemon tree thorn.

A patient who had a brother with soft tissue zygomycosis was taking care of him, changing the dressings. She had an abdominal scar due to previous cholecystectomy and she reported scratching it. She developed abdominal soft tissue zygomycosis.

Petrikkos GL, Skiada A, Sambatakou H, et al. Mucormycosis: Ten year experience in a tertiary-care center in Greece. Eur J Clin Microbiol Infect Dis 2003;22:753–756

Cutaneous zygomycosis in a patient with kidney transplantation

Needlestick exposures have been implicated in zygomycotic infections, occurring at the site of medicine injection, catheter insertion sites, injection sites for illicit drug use and tattooing.

Insect bites or stings have also been implicated.

Adam RD, Hunter G, DiTomasso J, Comerci G Jr.Mucormycosis: emerging prominence of cutaneous infections. Clin Infect Dis 1994; 19:67–76.

Burrell SR, et al. Apophysomyces elegansinfection associated with cactus spine injury in an immunocompetent pediatric patient. Pediatr Infect Dis J 1998; 17: 663–4.

Blair JE et al. Locally invasive cutaneousApophysomyces elegans infection acquired from snapdragon patch test. Mayo Clin Proc 2002; 77: 717–20.

Nitroglycerin patch. Petrikkos et al. Eur J Clin Microbiol Infect Dis 2003;22:753–756

«...His left index finger was swollen and tender, with multiple puncture wounds on the distal phalanx. There was loss of sensation and power in the left arm...»

«C. bertholletiae was isolated from the pus swab of the patient’s finger»

Intravenous intake of illicit drugs contaminated with Zygomycetes sporangiospores can produce disseminated and cerebral zygomycosis even in immunocompetent hosts.

Elasticized adhesive dressings Wooden tongue depressors Intravenous catheters

Mead JH, Lupton GP, Dillavou CL, et al. Cutaneous Rhizopus infection. Occurrence as a postoperative complication associated with an elasticized adhesive dressing. JAMA

1979;242(3):272–4. Mitchell SJ, Gray J, Morgan ME, et al. Nosocomial infection with Rhizopus microsporus in preterm

infants: association with wooden tongue depressors. Lancet 1996;348(9025):441–3.

…Although the CVC was changed high fever persisted and discharge continued from the large abdominal wound. Repeated tampons in different sections and wound secretion smears were positive for A. corymbifera. Fluconazole was stopped and liposomal amphotericin (Ambisome; 5 mg/Kg i.v.) given for over 3 months….

Zygomycosis of the scalp after a car accident

A volcanic cataclysm of major proportions, thefourth largest in terms of total casualties in thehistory of mankind, wiped out the town of Armero,Colombia, in 1985 resulting in over 23,000 deathsand 4,500 wounded. Among the hundreds ofsurvivors who were transferred to hospitals in thecapital city of Bogota, there was as overwhelmingnumber who developed necrotizing fasciitis.

8 developed necrotizing fasciitis due to Rhizopus arrhizus.

Only two survived.

Apophysomyces elegans

Invasive Fungal Disease (IFD) Following Remote Inoculation in Transplant Recipients

E. M. PETSCHNIGG, F. M. MARTY , S. P. HAMMOND , V. T. HO , C. MAGEE , D. A. MILNER , J. H. ANTIN , L. R. BADEN .

Two patients developed zygomycosis 2 and 10 months after their initial injuries.

One patient sustained a penetrating injury while snowboarding and the other required surgical packing of a thigh abscess.

Four and 23 days after transplantation respectively, both patients developed progressive necrotic skin lesions; angioinvasive hyphae consistent with Zygomycetes were seen on biopsy.

E. M. PETSCHNIGG et all

Conclusions: Fungal organisms may remain latent after

inoculation if plant debris or other foreign bodies are introduced during trauma.

Subsequently, when significant immunosuppression is utilized, IFD may develop.

IFD resulting from remote inoculation is a distinct syndrome that should be considered in immunocompromised patients presenting with soft tissue abnormalities in areas of prior injury.

E. M. PETSCHNIGG et all

Usually affects premature neonates and patients with severe malnutrition

May involve the stomach, ileum, and colon

Clinical picture mimics intra-abdominal abscess. The diagnosis is often made at autopsy

Clinical manifestations▪May involve virtually any organ▪ Can spread from lungs, sinus, or rarely form the gastrointestinal tract▪Most commonly seen in people who are immunosuppressed, IV drug users, or hematological malignancies

.Mucormycosis is an increasingly common infection in immunocompromised patients. The central role of iron in the organism’s pathogenesis has only recently been appreciated.The interaction between the Mucorales and endothelial cells is also beginning to be understood.Both of these pathogenetic features of disease may be amenable to novel therapeutic intervention in the future.

Iron overload Neutropenia Solid organ transplantation HIV Hypoglycemia

Zygomycosis is found only in immunosuppressed patients

Diabetes mellitus is a risk factor for zygomycosis only if it is accompanied by diabetic ketoacidosis.

Zygomycetes are the only fungi which can invade blood vessel walls.

Zygomycosis can be transmitted by inhalation of spores

Zygomycosis can be transmitted by blood transfusion.