PATHOGENESIS

3.1 Overview

Cryptococcosis occurs sporadically in a broad range of Australian native mammal species (Table 25.3). There is a predominance of respiratory and neurological disease among these cases, with pneumonia most common.

Fig.

CNS involvement), but less commonly as the only organ system reportedly involved (14% of koala cryptococcosis cases). Although cryptococcosis is commonly observed and documented in managed koalas, case clusters in free-ranging populations are being more commonly rec­ognised (39% of cases were in free-ranging koalas). There is little evidence to suggest a sex bias of koala cases, with 47% of cases in males and 53% in females. The role of potential immune suppression or association with koala retrovirus variants in the pathogenesis of cryptococcosis remains unclear but is the subject of investigation. Measures of immune function in koalas have been difficult to develop, although there has been steady progress (Maher et al. 2014; Maher and Higgins 2016). The predominance of C. gattii species complex in koala cases is at odds with a strong retroviral association in the pathogenesis of cryptococcosis, but in itself does not exclude the possibility. In humans, there is a strong association between overt immune compromise, par­ticularly retroviral disease, and cryptococcosis caused by C. neoformans species complex.

For cryptococcosis to occur, contact with the organ­ism is necessary for colonisation to occur. Once colonisa­tion is established, subclinical or clinical cryptococcosis may develop (Fig. 25.1). Progression may be rapid or slow, but is not inevitable, and colonisation and even subclini- cal disease may spontaneously resolve over time. Sponta­neous resolution is a common occurrence in young koalas soon after independence.

3.2 Virulence factors

The most important virulence factors of the C. neofor­mans and C. gattii species complexes are: the formation of a thick polysaccharide capsule to avoid and suppress the host immune response (Plate 25.1); capacity to grow at mammalian body temperature; production of melanin by laccase to resist host free radicals; production of urease and phospholipase B to facilitate tissue invasion; produc­tion of superoxide dismutase and trehalose to evade the host immune response; and development of polyploid ‘titan cells’ to further resist phagocytosis (Ma and May 2009).

Table 25.3. Summary of all reported cases Ofcryptococcosis in Australian mammals

25 - Cryptococcosis 377

Table 25.3. (continued)

378 CurrentTherapyin MedicineofAustraIian Mammals

FR = free-ranging; LN = lymph node; LRT = lower respiratory tract; M = managed; UGT = urogenital tract; URT = upper respiratory tract; U = unknown.

¹Rose 1999; ²Ladds 2009; ³Gaynor etal. 1990; ⁴HoIz 2008; ⁵ARWH 2018 (case no. 3361.1); ⁶ARWH 2018 (case nos 5402.1, 5423.1); ⁷ARWH 2018 (case no. 9359.1); ⁸MaIiketa/. 2011; ⁹ARWH 2018 (case no. 8769.1); ¹⁰Lynch 2008; ¹¹ARWH 2018 (case no. 5616.2); ¹²Johnson 2008; ¹³ARWH 2018 (case no. 5515.1); ¹⁴Backhouse and Bolliger 1960; ¹⁵Backhouse and Bolliger 1961; ¹⁶BoIIiger and Finckh 1962; ¹⁷Bowater etal. 2022; ¹⁸Gardiner etal. 1964; ¹⁹CanfieId etal. 1986; ²⁰CanfieId 1987; ²¹Spencer etal. 1993; ²²MaIiketa/. 1997; ²³Woods and Blyde 1997; ²⁴Krockenberger etα∕. 2003; ²⁵Bercier eta∕. 2012; ²⁶Wynne eta∕. 2012; ²⁷Kido eta∕. 2012; ²⁸Martinez-Nevado etα∕. 2017; ²⁹ARWH 2018 (case nos 333.1, 358.1,4583.1, 5465.1, 5941.1, 7282.1,

10183.1.10721.1) ; ³⁰Reece and Hartley 1994; ³¹ARWH 2018 (case no. 7227.1); ³²ARWH 2018 (case no. 7126.1); ³³Booth 1994; ³⁴ARWH 2018 (case no. 6908.1); ³⁵Krockenberger etα∕. 2005; ³⁶ARWH 2018 (case no. 8578.1); ³⁷Vaughan etα∕. 2007; ³⁸ARWH 2018 (case no. 4665.1); ³⁹ARWH 2018 (case nos 1875.1,6877.1); ⁴⁰VogeInest and Portas 2008; ⁴¹ARWH 2018 (case nos 8539.1, 8684.1); ⁴²ARWH 2018 (case nos 11168.4,11168.5); ⁴³CanfieId and Hartley 1992; ⁴⁴ARWH 2018 (case nos 1809.1, 4170.1,

4873.1.6267.1.11262.1.11480.1.1 ; ⁴⁵ARWH 2018 (case no. 5743.1); ⁴⁶GaIes etα∕. 1985; ⁴⁷ARWH 2018 (case nos 6327.1, 7295.1, 10876.1,11396.1,11778.1); ⁴⁸Payne etα∕. 2005; ⁴⁹ARWH 2018 (case nos 5445.1, 5647.1, 7041.1, 9228.1, 9337.1,10722.1,

10760.1) ; ⁵⁰ARWH 2018 (case nos 2382.1,4111.1,4899.1, 5211.1, 8161.1, 9725.1,10133.1, 11784.1,12002.1)

3.3 Colonisation

Colonisation of the nasal passages by the C. gattii species complex is known to occur commonly in koalas, espe­cially in zoo animals, and can occur in the absence of the development of clinical or subclinical disease. In a recent study investigating 181 free-ranging koalas from the Liv­erpool Plains region of NSW, 6.6% were found to have nasal colonisation with Cryptococcus spp. (predomi­nantly C. gattii VGI) (Schmertmann et al. 2019c). Coloni­sation in koalas can persist indefinitely, resolve spontaneously or progress to subclinical or clinical dis­ease (Connolly et al. 1999; Krockenberger et al. 2002b). A positive culture for Cryptococcus spp. from a nasal swab in the absence of sinonasal signs or a positive cryptococ- cal serological test is insufficient evidence of disease and should be considered colonisation until proven other­wise. This phenomenon has not been recognised in other Australian mammals to date. Nasal colonisation of east­ern ring-tailed (Pseudocheirus peregrinus) and common brush-tailed possums (Trichosurus vulpecula) in the Sydney region by Cryptococcus spp. is either extremely rare or not commonly detectable. An unspeciated Cryp­tococcus spp., C. neoformans var. grubii (serotype A) and C. magnus were detected from free-ranging quokkas (Setonix brachyurus) on Rottnest Is. and mainland WA using culture and molecular methods (Martinez-Perez et al. 2020). Quokkas from which these organisms were detected were clinically healthy suggesting colonisation rather than infection.

3.4 Subclinical disease

Subclinical disease is a relatively common outcome after colonisation. This is particularly so in koalas (especially zoo but increasingly in free-ranging koalas) (Krocken- berger et al. 2002a), but is also seen in humans (Gold­man et al. 2001) and is likely the case in other species. Subclinical cryptococcosis can be defined as cryptococ- cal capsular antigenaemia in the absence of clinical signs or identifiable lesions. It is presumed that this reflects early, limited invasion of the respiratory mucosa by Cryptococcus spp., which can (i) resolve with an effective host immune response, (ii) persist as a constrained local focus or (iii) progress to clinical disease, possibly after stress or malnutrition. The authors would generally con­sider treating any koala with a persistent LCAT of 1:8 or higher, although individual case details may influence the decision to initiate therapy. A titre of ~1:64 is consid­ered the limit of positivity seen with subclinical disease; at this point, a focus of infection will have developed (e.g. a focal sinonasal or pulmonary granuloma).

Although few other Australian mammals have been assessed for subclinical cryptococcosis, a small study of eastern ring-tailed and common brush-tailed possums in the Sydney region found no evidence for subclinical dis­ease in free-ranging animals.

3.5 Clinical cryptococcosis

The most common primary site for cryptococcal lesions in the koala is the respiratory tract, consistent with inha­lation as the typical route of infection. Clinical crypto­coccosis classically presents as sinonasal disease in the koala, although lower respiratory tract lesions and pneu­monia are also common (Krockenberger et al. 2003). Both upper and lower respiratory tract disease can lead to haematogenous dissemination, particularly to the CNS. Therefore, characteristic presenting signs include dysp­noea, coughing, sneezing, stertor, nasal discharge, epistaxis, facial distortion, neurological abnormalities (e.g. blindness, nystagmus, seizures), inappetence and lethargy (Blanshard and Bodley 2008). Skin lesions can also be seen as a primary focal lesion (presumably from direct inoculation) or subsequent to haematogenous dis­semination from a primary respiratory focus (Krocken- berger et al. 2003). Disseminated lesions can be found in many tissues, but with the exception of the CNS, these lesions are generally milder and unlikely to be fatal. In one koala, the presenting sign was an extensive ulcerated lesion involving the distal hindlimb, also involving the underlying distal tibia (Schmertmann et al. in press). This highlights the broad range of clinical presentations that occur with cryptococcosis in koalas. The majority of koala cases where the causative agent was definitively identified were attributable to the C. gattii species com­plex (Krockenberger et al. 2005), with a single case caused by the C. neoformans species complex in a zoo koala in Spain (Martinez-Nevado et al. 2017). Generally, crypto­coccosis is caused by C gattii VGI, although in south­western WA infections can involve C gattii VGII, including cases where koalas have been infected in WA and developed disease following translocation elsewhere within Australia.

Subclinical and clinical cryptococcosis in zoo koalas presents a significant management concern in some insti­tutions and serological monitoring is generally recom­mended as a component of a standard health check at least once or twice a year.

In other Australian native mammals, sinonasal crypto­coccosis appears to be less common (Johnson 2008; Vogel- nest and Woods 2008; Ladds 2009). Instead, the primary presenting signs and lesions are related to pneumonia and/ or meningitis, with upper respiratory tract disease only reported in an eastern barred bandicoot (Perameles gunnii) and a long-nosed potoroo (Potorous tridactylus) (Table 25.3). Both the C. gattii and C. neoformans species complexes have been implicated in cases of cryptococcosis in non-koala Australian native mammals, with the C. gattii species complex responsible for approximately twice as many cases as the C. neoformans species complex.

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