A New Era in Parasitology 3 Entamoeba coli cases detected - 1 co-infection of Entamoeba...
Transcript of A New Era in Parasitology 3 Entamoeba coli cases detected - 1 co-infection of Entamoeba...
A New Era in Parasitology
Jennifer O’ Leary
Dr Roy Sleator & Dr Brigid Lucey
Introduction
• Enteric parasitology encompasses the diagnosis of a wide range of species
• Cryptosporidiumspp & Giardia lamblia commonly encountered in Irish clinical microbiology labs
Cryptosporidium spp
• Obligate enteric protozoan parasite
• Faecal-oral transmission
• C. parvum & C. hominis –90%
• Cramps, fever, nausea, vomiting & diarrhoea.
• Ireland – highest infection rate in Europe
Giardia lamblia
• Flagellate protozoan parasite
• Broad spectrum of symptoms
• Chronic sequelae
• Contracted via faecal-oral route
• Travel associated infection
Current Diagnostic Techniques
• Brightfield & fluorescent staining for Cryptosporidium
• Wet-prep and special stains employed for other parasites
• Limits speciation
• Immuno-based methods also available
Molecular Diagnostics• Real time PCR
• Commercial kits currently limited to a small number of parasitic species
- Cryptosporidium spp- Giardia duodenalis
• Commercial kits and in-house assays (Hadfield et al., 2011; Manser et al., 2013)
• EntericBio® Gastro Panel 2 real time PCR kit, Clinical Microbiology Department, CUH
• Roche LightMix ® Modular assay
Microscopy vs Real Time PCR• GastroPanel II (1) Validation – 100% correlation
• 104 samples tested- 22 samples requested OCP: 17 tested, 5 lacked relevant
clinical data- 2 non-requested samples positive
• 6 positive samples detected by both PCR & microscopy- 5 cases of Giardia duodenalis- 1 case of Cryptosporidium parvum/hominis
• 4 positive samples detected by microscopy alone:- 3 Entamoeba coli cases detected - 1 co-infection of Entamoeba histolytica/dispar and Giardia duodenalis detected
• RIDA®GENE Parasitic stool panel II (2) & VIASURE (3) monoplex assays: 97.3 –100% correlation
Giardia duodenalis & Entamoeba spp
Microscopy vs Real Time PCRAdvantages Disadvantages
Microscopy Relatively low cost
Widely available
LOD: 1,000 oocysts /mL faeces
Time consuming
Skilled microscopist essential
Molecular
methods
LOD: 1 oocyst /mL of faeces
Exceptional sensitivity reported
Time efficient
Multiplex capabilities
Speciation
Expensive reagents and
instrumentation required
PCR inhibitors
Incorporation of Real Time PCR
Future Studies• Cryptosporidium research:- Speciating and genotyping
Cryptosporidium spp involved in clinically significant human infections
- Bioinformatics based approach to identify molecular bio-markers
- Relate findings to the clinical history and follow-up of patients
- Potential for diagnosis of a broader range of species – Continued collaboration with Serosep
• Giardia lamblia genotyping
Concluding Remarks
• Molecular diagnosis:
- Improved diagnostic capabilities
- Streamlined lab work flow
- Time efficiency
• May be equivalent when microscopy used by skilled microscopists
• Employed only when clinically indicated
• May impact on national notification figures
Acknowledgements
• I would like to express my gratitude to the staff of the Microbiology Department, Cork University Hospital, particularly Mr. Liam Blake and Dr. Dan Corcoran.