1. Cryptosporidium monitoring of
Ireland's waters
Theo de Waal
UCD School of Veterinary Medicine Scoil Leighis Tréidliachta UCD
1

2. Outline
• Introduction
• What is Cryptosporidium
• How is it spread
• Cryptosporidium in humans
• Cryptosporidium in surface water
• National reference laboratory
• Survey of Irish water supplies
2

3. Introduction
• Cryptosporidium first described in 1907 Tyzzer
– C. muris
• Only associated with disease and death in 1955
in Turkeys
– C. meleagridis
• In early 1970’s first reported of its association
with diarrhoea in cattle
• In 1976 first two human case described
– 3-year-old child
– 39-year old immunosuppressed patient
• Today Cryptosporidium one of the most
commonly identified intestinal pathogens
3

4. What is Cryptosporidium?
• Small single cell eukaryotic
organism
• Found in GIT
• Oocyst  environment
– Small
• 4-8 µm in size
– Smooth, thick outer wall
– Contain infective sporozoites
4

5. Cryptosporidium life cycle
• Direct life cycle
• Sporulated oocyst ingested
• Infect microvillus border of
GIT – vertebrates
– 3 species : Gastric mucosa
– 1 specie: Respiratory system
• Complex development
– Asexual multiplication
– Sexual reproduction
• Autoinfection
5

6. How is it spread?
• Transmission: faecal-oral route
– Close contact
– Waterborne
– Foodborne
6
Fayer, R., 1997. Cryptosporidium and Cryptosporidiosis . CRC Press

7. Cryptosporidium oocyst survival
• Very resistant!
– Oocysts can remain viable in environment
& animal liquid waste ~ 1 year
– Resistant to environmental stressors
– Resistant to most chemical disinfectants
http://www.bio-uv.com/fr/site/Piscines-spas-collectifs/Prevention-Cryptosporidium/Prevention-contre-les-pathogenes-parasitaires_129_.html 7

8. CRYPTOSPORIDIUM IN
HUMANS
8

9. Cryptosporidium spp:
• More than 22 recognised Cryptosporidium species
– 39 Cryptosporidium genotypes
• Host specific - C. hominis to broad host range - C.
parvum and C. ubiquitum
• Only few considered infectious to humans
– Human cryptosporidiosis in Ireland1,2
• C. hominis (20%)
• C. parvum (80%)
1. Zintl, et al, 2009, The prevalence of Cryptosporidium species and subtypes in human faecal samples in Ireland. Epidemiol. Infect. 137, 270-277. 9
2. Graczyk, et al., 2007. Human enteropathogen load in activated sewage sludge and corresponding sewage sludge end products. Applied and Environmental
Microbiology 73 (6):2013-2015.

10. Cryptosporidium in Ireland: Human
• Crude incidence rate
Cryptosporidium1
– 6.9 – 13.4/100,000 annually
700 • Strong urban-rural divide
600 • Rural areas reported more
Number cases
500 cases
400 • Regional as high as
300 – 31.4/100,000 per year
200
100
0
Year
2004 2005 2006 2007
2008 2009 2010 2011
10
1Human cryptosporidiosis became a notifiable disease on January 1st 2004

11. Ireland: Seasonal distribution in humans
11
http://www.hpsc.ie/hpsc/A-Z/Gastroenteric/Cryptosporidiosis/Publications/EpidemiologyofCryptosporidiosisinIrelandAnnualReports/

12. CRYPTOSPORIDIUM IN
SURFACE WATER
12

13. Cryptosporidium waterborne outbreaks
• First waterborne outbreak
Braun Station, Texas
(1984)
• Largest epidemic –
Milwaukee, Wisconsin
(1993)
• To date outbreaks affecting
>106 individuals
documented Finnegan's Lake, County Galway.
13

14. Cryptosporidium waterborne outbreaks -
Ireland
Area Year Cases Source & Deficiency Species
Belfast 2000 & 2001 246 & 191 Septic tank seepage into reservoir C. bovine strain & C.
human strain
Mullingar 2002 26 Heavy rain, agricultural runoff into C. genotype 2 (=C.
lake. No filtration. parvum) in humans
Ennis 2003, 2005, 2008 ? Surface water into spring source. ?
No filtration
Carlow 2004 31 ? C. parvum, C. andersoni,
C. muris in water. C.
hominis in humans
Galway 2007 240 Agricultural runoff, sewage plant. C. hominis and C.
Inadequate filtration parvum in water and
humans
Clonmel 2007 ? Surface water contamination ?
following heavy rain
14

15. Drinking water in Ireland- vulnerable?
• Surface water (82% of drinking
water)
• Climate
• High rainfall
• Geology
– Shallow layer of soil and subsoil Groundwater vulnerability map
over karst limestone
– Heavy soils  either rapid
surface runoff or preferential
flow
• Unprotected catchments
• Septic tanks
• Livestock
• Inadequate treatment on some
supplies
Households with septic tanks
15

16. Cryptosporidium in Ireland: Environment
• Several Irish studies have detected
Cryptosporidium species in Irish river basins1,2
• 2005 EPA risk assessment - Irish public water
supply
– 8% high risk
– 13% very high risk
16
1. Graczyk, et al., 2004. Human waterborne parasites in zebra mussels (Dreissena polymorpha) from the Shannon River drainage area, Ireland. Para Research 93: 385-391.
2. Lucy, et al., 2008..Biomonitoring of surface and coastal water for Cryptosporidium, Giardia and human virulent microsporidia using molluscan shellfish. Para Research 103:1369-1375

17. Cryptosporidium in Irish Water
• Source of contamination and public health risk
– Very limited information
– No genotyping facilities in Ireland
– Few local authorities get samples genotyped in UK
• Survey of Cryptosporidium monitoring in
public water supplies
– 24.5 % supplies being monitored
• >83% high Crypto risk score
• Monitoring frequency low
17

18. Monitoring
Reasons given for routine monitoring Reasons why supplies are not routinely
40
monitored
50
30 40
responses (%)
responses (%)
30
20
20
10
10
0 0
18

19. Development of a National Reference
Facility for Cryptosporidium: 2010
• National survey of Irish public
water supplies
• Adopt best practice procedures
– Laboratory accreditation
• Pilot study of water supplies
• Significance of emerging
waterborne contaminants
• Strategies for service delivery
beyond project

20. National Reference Facility for
Cryptosporidium
• Detection of Cryptosporidium in water
– Based on USEPA 1622
• Filtration
• Immuno-magnetic separation
• Fluorescent antibody
• INAB Accreditation – ISO17025
– April 2012
20

21. Cryptosporidium Reference Laboratory
 Genotyping
• Source of contamination
• Public health risk
• Catchment protection
FITC stained
• Water safety plan development for supply Cryptosporidium oocysts
• Frontline help in source contamination events
or outbreak investigations without need for
samples to be sent overseas
• Nurture and provide local knowledge and
expertise
21

22. Pilot Scheme - 2011
• Detection and identification of Cryptosporidium
species in supplies on RAL
• 5 supplies selected
• “Type” supplies established
– Groundwater under influence of surface water
– Pristine upland lake
– Spring/Borehole supply
22

23. Results to date
Pilot scheme details No samples %
Samples submitted 152
Positive USEPA 1622 74 48.6
Genotyped 46 62.2
23

24. Type supply:
Groundwater under influence of surface water
• No barrier for Cryptosporidium
• On boil water notice
• Previously had one sample genotyped in Scotland
– very mixed results – up to 7 different species implicated
• No clear idea of source of contamination/ public health
risk
25

25. Type Supply :
Groundwater under influence of surface water
Date No of Oocysts/10 L Genotype Possible source Public
oocysts Health
detected Risk
March 15 0.19 C. andersoni Uncertain
6 0.3
April 64 0.45 C. parvum High
May 3 0.01 ND
June 2 <0.01 C. parvum High
July 3 0.01 ND
August 4 0.01 C. muris No risk
September 24 0.11 C. andersoni Uncertain
October 55 0.52 C. parvum/ High
C.ubiquitum
November 24 0.12 C. bovis/C.ubiquitum Uncertain
26

26. Type Supply: Upland Lake
• Town supply - source water upland lake
• No barrier for Cryptosporidium
• Cryptosporidium detected in 2007 during intense
monitoring period
– C. parvum detected once in raw water
– C. ubiquitum also detected once
• EPA audit conducted in 2009
27

27. Type supply: Upland lake monitoring results
Date No of Oocysts/10 L Genotype Possible Public Health
oocysts source Risk
detected
March 3 0.02 ND
April 0 <0.01 N/A
May 16 0.14 C. ubiquitum Uncertain
June 52 0.33 C.ubiquitum/ C. Uncertain
xiaoi
August 1 <0.01 N/A
September 2 <0.01 no amplification
October 2 0.01 C. envir genotype ??? No known risk
November 3 0.01 C. envir genotype ??? No known risk
28

28. Type supply: Spring/Borehole supply
• Spring & Borehole supply
• Spring supply located downstream of lake
– concern over influence of lake over spring
• No barrier for Cryptosporidium
• July 2011 - 5 oocyts detected (0.02/10 L)
29

29. Type supply: Spring/Borehole supply monitoring
results
Date No of Oocysts/10 L Genotype Possible Public
oocysts source Health Risk
detected
August 264 2.4 C. ubiquitum Uncertain
30

30. SUMMARY
31

31. Cryptosporidium spp. and genotypes in
Irish Drinking Water Supplies
Cryptosporidium spp. detected No. Samples Possible source Public Health Risk *
C. andersoni 14 adult cattle/yearlings Uncertain risk
C. ubiquitum 11 deer/sheep Uncertain risk
C. parvum 4 preweaned calves/human High risk
C. bovis 4 weaned calves No known risk
C. environmental genotype 3 unknown No known risk
C. ryanae 1 weaned calves No known risk
C. muris 1 mouse Uncertain risk
C. andersoni / C. bovis mixed 1 calves/yearlings/adult cattle Uncertain risk
C. ubiquitum / C. xaoi mixed 1 wildlife/sheep Uncertain risk
C. andersoni / C. canis mixed 1 cattle/dog Uncertain risk
C. andersoni / C. muris mixed 1 cattle/mouse Uncertain risk
C. parvum / C. ubiquitum mixed 1 wildlife/cattle/sheep High risk
C. bovis / C.ubiquitum mixed 3 deer/sheep /cattle Uncertain risk
Total 46
As described in the UK Environment agency Microbiology of Drinking Water (2009)- Blue Book
High risk: Known human pathogen and causative agent of outbreaks
Uncertain risk: Isolated from sporadic human cases but pathogenicity uncertain
No known risk: No human isolates reported
32

32. Summary & Conclusions
• Drinking water in Ireland particularly vulnerable to
Cryptosporidium contamination
• Risk of recreational waters?
• Humans incidence
– Predominant spring peak
• C. parvum, C. hominis
• Cryptosporidium reference facility established in
Ireland
– INAB Accreditation – ISO17025
33

33. Acknowledgements
– Carolyn Read
– Jenny Pender
– Annetta Zintl
– Marzieh Mirhashemi
– Frances Lucy
– Declan Feeney
– Hui-Wen Cheng
34

34. Original illustrations and photographs of Cryptosporidium parvum - Tyzzer, 1912
35