Cryptosporidium: Cryptosporidium: Director, UK Cryptosporidium Reference Unit the global challenge in monit toring

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1 Cryptosporidium: still cryptic after all these years Dr Rachel Chalmers Cryptosporidium: Director, UK Cryptosporidium Reference Unit the global challenge in monit toring

2 urtesy of the Francis A. Countway Library of dicine, Boston, MA Cryptosporidium Ernest Tyzzer Established the genus Cryptosporidium = hidden spores 1907 A sporozoan found in the peptic glands of the common mouse 1910 An extracellular coccidium, Cryptosporidium muris (gen. et sp. no.) of the gastric glands of the common mouse 1912 Described the oocyst structure and endogenous stages of Cryptosporidium parvum in mice

3 Original illustrations of C. parvum Tyzzer, 1912; Slapeta, 2011

4 Living things Eukaryota Prokaryota Bacteria Archaea Charles Darwin, 1837 Cryptosporidium

5 Eukaryota Animalia Fungi Amaebozoa Plantae Chromalveolata Rhizaria Excavata Alveolata Apicomplexa Ciliophora Dinflagellata Conoidasida Aconoidasida Coccidiasina Gregarinasina Haemosporasina Piroplasmasina Eucoccidiorida Eimeriorina Cryptosporidiidae

6 00 years of taxonomic uncertainty Tyzzer, 1907 possession of an organ of attachment and of iodophilic granules, it resembles the gregarines morphology, lack of motion in the adult and in sexual dimorphism it resembles the coccidia Tyzzer, 1910 its habitat of life is similar...to the order Gregarinida it appears to be.a coccidian Placed in the then suborder Coccidia Cryptosporidium

7 rta and Thompson, 2006 distantly related lineage of icomplexan parasites that e not in fact coccidia but at do occupy many of the me ecological niches. Cryptosporidium

8 n imperfect Coccidian, an atypical Gregarine or a parate lineage? oes it matter? Explains some of the difficulties we face: insensitive to anti-coccidial drugs false-positive results in environmental samples when antibody-based methods are used: multi-level assurance that it s crypto Take off the Coccidian blinkers: screening new drugs and targets; use of more appropriate models improve our understanding of life cycle, pathogenesis and ecology PCR primers for accurate ID Cryptosporidium

9 ryptosporidium species supported by genetic tudies ecies in humans Species in animals (type host) C. parvum sensu lato (cattle) C. parvum sensu stricto (mice) C. hominis (humans) C. bovis (cattle) C. meleagridis (turkeys) C. serpentis (snakes) C. varanii (lizards) C. felis (cats) C. scophthalmi (turbot) C. canis (dogs) C. molnari (fish) C. suis (pigs) C. galli (birds) C. muris (mice) C. baileyi (chickens) C. andersoni (cattle) C. wrairi (guinea pigs) C. ubiquitum (sheep) C. fragile (toads) C. fayeri (red kangaroo) C. ryanae (cattle) C. cuniculus (rabbit) C. macropodum (grey kangaroo) C. xiaoi (sheep) Cryptosporidium

10 orldwide diversity of Cryptosporidium spp. in human infection Cryptosporidium spp. C. parvum C. hominis Other species and genotypes Red = caused an outbreak C. leagridis C. cuniculus = rabbit gt C. felis C. ubiquitum = cervine gt C. canis C. andersoni, C. muris, C. suis, C. fayeri skunk, horse, monkey chipmun pig genotyp Cryptosporidium

11 Cryptosporidium genotypes Have no taxonomic status Identified by DNA sequence differences Temporary name Biological details needed to define a new species: 1. Morphometric data (oocyst and developmental stages) 2. Natural or experimental host specificity 3. Detailed biological characterisation, including localisation and pathology associated with an individual host 4. Differential diagnosis 5. Genetic characteristics 6. Deposition of sequences and material 7. Compliance with ICZN species-naming rules. Cryptosporidium

12 Rabbit genotype: C. hominis in disguise or a separate species? Cryptosporidium

13 he morphology of the oocysts and iagnostic characteristics typical of other intestinal species of Cryptosporidium slightly larger oocysts mean 5.98 x 5.38 µm; length:width = 1.1 ) Differential interference contrast ) FITC ) DAPI (arrows indicate sporozoite nuclei) ) auramine phenol ) modified Ziehl Neelsen. Re-description of Cryptosporidium cuniculus etc. Robinson et al.., IJP 2010

14 ost range atural hosts: European rabbits (Oryctolagus cuniculus) humans. perimental infections established in: weanling rabbits immunosuppressed Mongolian gerbils immunosuppressed adult Porton mice but not in neonatal mice. Re-description of Cryptosporidium cuniculus etc. Robinson et al.., IJP 2010

15 hedding profiles in experimentally infected animals. Cryptosporidium shedding profiles in rabbits 100 Shedding Score Percentage days post infection C. hominis (blue bar) C. cuniculus (yellow bars) Re-description of Cryptosporidium cuniculus etc. Robinson et al.., IJP 2010

16 Infection and pathology Patterns of infection measured by oocyst shedding are significantly different compared with C. hominis, particularly in rabbits. Endogenous stages in the brush border of the epithelium of the small intestinal villi RABBIT IMM. GERBIL IMM. MOUSE clinical signs are absent caused disease in humans. Re-description of Cryptosporidium cuniculus etc. Robinson et al.., IJP 2010

17 Phylogenetics A close relationship with C. hominis inferred from molecular analyses.. Sequences contained limited, consistent polymorphisms: SSU rrna gene 4/787 bp (0.51%) HSP70 gene 1/403 bp (0.25%) Actin gene 1/833 bp (0.12%) 60 kda glycoprotein (GP60): 2 unique families, Va and Vb But only 0.07% of the entire genome has been studied and further investigation of the whole genome would be helpful. Re-description of Cryptosporidium cuniculus etc. Robinson et al.., IJP 2010

18 e concluded: Although genetically closely related, there are significant biological differences between C. cuniculus and C. hominis that support separate species status. This relies on the assumption that mating between these species would not normally occur: it is highly unlikely that C. cuniculus and C. hominis interbreed within the type host, the European rabbit. Re-description of Cryptosporidium cuniculus etc. Robinson et al.., IJP 2010

19 Are other unusual Cryptosporidium species or genotypes a public health risk? Other species and genotypes in human infection C. leagridis C. cuniculus = rabbit gt C. felis C. ubiquitum = cervine gt C. canis C. andersoni, C. muris, C. suis, C. fayeri skunk, horse, monkey chipmun pig genotyp Re-description of Cryptosporidium cuniculus etc. Robinson et al.., IJP 2010

20 Fig. 1. Bayesian belief network of risk assessment for risk of Cryptosporidium infections associated with very small private supplies. unter et al., Risk Analysis, Vol. 31, No. 2, 2011

21 uantitative Microbial Risk Assessment of Cryptosporidiosis from Very all Private Water Supplies. Median annual risk of infection for Cryptosporidium 25% In the poorer quality supplies the risk could be much higher Risk is substantially greater than for public water supplies and well above the risk considered tolerable. unter et al., Risk Analysis, Vol. 31, No. 2, 2011

22 Factors influencing likelihood of waterborne cryptosporidiosis Primary source: farmed animals; wild life; humans Catchment: weather; run off; barriers Water treatment: type; function; ingress Distribution: ingress; blending; dilution; bolus Cryptosporidium Consumer population: immunity; consumption Oocysts: Numbers; viability; strain (infectivity & pathogenicity)