Cryptococcosis is caused by a Gram-positive yeast that has worldwide distribution with an increased incidence in temperate regions such as southern California and Australia. It affects numerous mammalian species, including dogs, cats, and humans. Cryptococcus neoformans is the species that primarily causes this disease in domestic animals. C. neoformans is a saprophytic, round to oval, yeast-like fungus measuring 3.5-7.0 microns in diameter. It also has a characteristic microscopic "soap-bubble" appearance in lesions due to the large heteropolysaccharide capsule that protects the organism from desiccation when invading tissues. Two variants, Cryptococcus neoformans var. neoformans and Cryptococcus neoformans var. gattii, and five serotypes (A, B, C, D, AD) have been identified.
Several sources of C. neoformans exist, including bird (especially pigeon) excreta which contain high amounts of nitrogen-containing compounds such as creatinine, soil and fruits. Cryptococcus sp. can survive in faeces for up to two years, unless viability is reduced by dry conditions or UV light. It does not spread via direct contact but instead is transmitted by inhalation of the aerosolised organism from the soil or feces. Cryptococcus neoformans var. gattii is primarily found in tropic and subtropic areas due to its very specific habitat of Eucalyptus trees.
Species which have been recorded as pathogenic in cattle include:
- Cryptococcus neoformans var grubii
- Cryptococcus neoformans var neoformans
- Cryptococcus neoformans var bacillisporus
Cryptococcal reproduction occurs by the formation of blastoconidia, which are buds connected by a narrow isthmus to the parent cell. Cryptococcus neoformans differs from other dimorphic fungi in that it is always found in the yeast phase in laboratory cultures and infected tissues. Inhalation of the aerosolised organism is the most probable source of infection; however, the exact mode of transmission is unknown. The organism can deposit in the upper or lower respiratory tract. However, the organism primarily remains in the upper airways since the encapsulated organisms (5-20 micrometers in diameter) are larger than the diameter of the terminal airways. This leads to a predominance of upper respiratory infections rather than pulmonary disease. Once established in the airways, the infection can spread hematogenously or by tissue invasion through the cribriform plate to the brain. Mammals with normal immune systems usually clear the cryptococcal infection prior to its spread. The majority of resistance is provided by cell-mediated immunity. Infections with Cryptococcus neoformans may indicate immunosuppression in some cattle.
The infection is thought to be acquired from the environment, with no reported cases of disease transmission from one affected animal to another. Thus, cryptococcosis is not a contagious or anthropozoonotic disease.
Cryptococcosis is most commonly caused by two species of the genus Cryptococcus; C. neoformans and C. bacillisporus. The ability of C. neoformans and C. bacillisporus to grow at 37oC may in part explain their pathogenicity, because other members of the genus grow poorly at this temperature. Elaboration of a polysaccharide capsule and the enzymes laccase and phospholipase by C. neoformans and C. bacillisporus are additional virulence factors that contribute to pathogenicity. C. neoformans and C. bacillisporus are dimorphic, basidiomycetous fungi. They exist in animal tissues as the yeast form (Cryptococcus spp), but are capable of transforming under special laboratory conditions into a filamentous form (Filobasidiella spp). Thus far, the filamentous phase has been demonstrated only under strictly controlled laboratory conditions, but this perfect state is likely to exist in certain natural environments. The importance of the perfect state is that spores resulting from sexual or asexual filamentous reproduction likely represent the infectious propagules that give rise to mammalian disease.
In animal tissues, Cryptococcus neoformans and C. bacillisporus exists as a round, yeast-like organism, with a variably-sized polysaccharide capsule as its distinguishing feature. The capsule provides protection from environmental insults (e.g., desiccation) and the phagocytic response of the host. In tissues, Cryptococcus reproduces by forming one or two daughter cells (buds) that are connected to the parent cell by a narrow isthmus. Buds may break off when small and thus the cell population varies in size.
Unlike other dimorphic fungi, the yeast phase of Cryptococcus is found under routine laboratory conditions and in infected tissues. Historically, five serotypes (A, B, C, D, AD) have been identified on the basis of antigenic differences in capsular polysaccharide. Recent advances in the taxonomy of the genus Cryptococcus have led to a new nomenclature that was proposed at the 5th International Conference on Cryptococcus and cryptococcosis. C. neoformans and C. bacillisporus differ biochemically, genetically, ecologically and epidemiologically. C. neoformans has a world wide distribution, while C bacillisporus is largely restricted to tropical and subtropical climates. C. neoformans can be divided into two varieties based on serotyping, C. n var grubii and C. n var neoformans. Both varieties are strongly associated with disease in immunocompromised human patients, although the same may not be true for companion animals. C n var grubii is by far the most common isolate from cryptococcosis in people and animals worldwide, although C. bacillisporus is important in certain geographical regions such as Australia, Papua New Guinea, South East Asia and Central Africa. There is strong evidence that several of Australian eucalyptus trees provide a natural environmental niche for C. bacillisporus. Interestingly, koalas seem capable of amplifying the number of cryptococci in certain environments. The definitive environmental niche for C. neoformans has not been determined, although there is a strong 'historic' association with weathered bird (especially pigeon) guano and more recent evidence for growth in decaying plant matter in hollows of certain trees. The organism passes through the gut of pigeons, but systemic infection of pigeons is extremely rare. Perhaps the pigeon's high body temperature protects it from infection. Pigeon guano provide an alkaline, hyperosmolar environment that is rich in many nitrogen-containing compounds including creatinine that favour cryptococcal growth. Cryptococci may remain viable for at least two years in accumulations of pigeon guano protected from drying or sunlight; pigeon lofts provide such an environment.
Most basidiomycetes reproduce sexually in their natural environment, and the teleomorphs of C. neoformans (Filobasidiella neoformans) and C. bacillisporus (F. bacillisporus) can be induced to undergo sexual reproduction in the laboratory and produce dikaryotic hyphae, blastoconidia, basidia and basidiospores. The recent documentation of both α and a-mating types of C. bacillisporus in Eucalyptus trees suggests that this may occur in nature. However, recent work has suggested that C. neoformans may be evolving into an asexual fungus and that basidiospores may result from haploid (monokaryotic) fruiting as well as by sexual recombination. In either case, the notion that the basidiospore is the infectious propagule for Cryptococcus is attractive, as this stage is suited to dispersal by air currents and has physical properties that favour penetration into the respiratory system, thereby facilitating primary infection of mammalian hosts.
The exact mode of infection is unproven, but the most likely route is via inhalation of air-borne organisms. These may be basidiospores or yeast cells desiccated by environmental exposure. Shrunken, poorly capsulated cryptococci that are small enough for alveolar deposition have been isolated from pigeon guano and soil. Although human patients with cryptococcosis typically present with neurological signs referable to meningoencephalitis, there is strong circumstantial evidence that the infection starts in the lungs and subsequently spreads to the nervous system hematogenously via macrophages. Respiratory involvement usually does not result in clinical signs, although lesions can be detected in thoracic radiographs, thoracic computed tomography scans or at necropsy. The small particle size of infectious propagules is said to be the reason the lung is primary site of infection, as only very small particles are capable of penetrating deep into the lower respiratory tract.
When infection ensues, clinical signs of rostral nasal cavity disease such as sneezing, epistaxis and nasal discharge are conspicuous, and sometimes granulomatous protuberances can be seen at the nares. In some cases, destruction of adjacent facial bones facilitates spread of infection to contiguous regions, such as the bridge and side of the nose, the planum nasale or hard palate. When facial distortion develops, the clinical presentation is strongly suggestive of either fungal rhinosinusitis or nasal neoplasia. On the other hand, when infection begins in the caudal portion of the nasal cavity, signs of mycotic rhinitis may be subtle or absent, although it is possible to confirm the sinonasal region as the primary site of infection using cytology, culture, endoscopy or cross-sectional imaging. In some cases, infection spreads through the cribriform plate into the olfactory bulbs and olfactory tract, giving rise to meningoencephalitis. In these cases, the anatomical proximity of the optic nerves frequently results in concurrent cryptococcal optic neuritis, and secondary retinitis. Clinically, this is manifest as widely dilated pupils that respond poorly to light, swelling of the optic disc and focal retinal haemorrhage. There may also be anterior uveitis. The invasion of the meninges results in an extensive leptomeningitis throughout the CNS. This is often accompanied by a ventriculitis. Unilateral strabismus or nystagmus occur as the infection extends caudally along the brain stem. In other cases, caudal nasal cavity involvement gives rise to a mass lesion which occludes one or both choanae, resulting in nasopharyngeal signs viz. stertor, snoring, dyspnea or open mouth breathing. In occasional cases the infection spreads to the middle ear via the auditory tube. A focal granuloma may occur in the brain and clinically mimic a neoplasm.
Cutaneous involvement, if multifocal, reflects hematogenous dissemination from the primary site of infection, as do lesions in bone (e.g., digits) or periarticular soft tissues. In ferrets, and the exceptional cat, localized cutaneous cryptococcosis can develop following penetrating injury of the skin. In some cats, infection spreads to the mandibular lymph nodes, presumably via the lymphatics from the nasal cavity. Rarely, mandibular lymphadenomegaly can be massive and require surgical debridement. Occasionally salivary gland infection has been documented, although how organisms reach this site is a mystery.
There is a tendency for animals in rural environments to be infected with C. bacillisporus, presumably due to increased exposure to eucalyptus material. The development of granulomatous intracranial or pulmonary mass lesions (cryptococcomas) are strongly associated with immunocompetence.
The four primary systems affected are the respiratory, central nervous, ocular, and cutaneous systems.
Isolation of yeasts from bovine milk is not uncommon. Candida spp' are the yeasts most frequently isolated from milk or the mammary gland, but Cryptococcus spp have also been commonly isolated and do cause mastitis in cattle. The subclinical infections may resolve spontaneously or become chronic. The severity of infection is dependent on the number of organisms introduced into the gland.
With cryptococcal meningitis, affected cattle present with multifocal neurological deficits manifested by hypermetria, ataxia, depression, circling, impaired vision, head pressing, low head carriage, wide-based stance, and falling to the side or backwards.
Diagnosis is based on presenting clinical signs and confirmation of infection by Cryptococcus is confirmed by laboratory testing of infected tissue. Serum biochemical test results are usually within reference intervals.
Definitive diagnosis can also be determined via examination of tissue biopsies and fungal cultures. Histologically, the organism can be stained with hematoxylin and eosin, periodic acid-Schiff, Gomori's methenamine silver, Masson-Fontana, or Mayer’s mucicarmine techniques; the latter staining technique is considered definitive. C. neoformans has a thick capsule, thin cell wall, budding, and lack of endospores which differentiate it from Blastomycoides spp and Coccidioides spp. Cryptococcus neoformans can be cultured from exudates, cerebrospinal fluid, urine, joint fluid, and tissue samples on Sabouraud’s agar with antibiotics at 25-37°C. MR imaging may reveal thickened meninges or focal granulomas.
In cattle, Amphotericin B, ketoconazole, itraconazole and fluconazole are effective but expensive and are reserved for valuable cattle.
Severe affected cattle are usually culled as spontaneous recovery from cryptococcal mastitis is rare.
Outcomes of treatment of cryptococcosis are quite varied. Drug therapy is long-term (average of 8.5 months) and relapses occur frequently. Since Cryptococcus neoformans is ubiquitous, the best means of prevention is to decrease contact with areas containing a high concentration of organisms (pigeon droppings, damp buildings or barns).
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