Cryptosporidium update
Prof. Rachel Chalmers
Director, Cryptosporidium Reference Unit
Public Health Wales Microbiology
The challenge of Cryptosporidium
and
Singleton Hospital
swimming pools
Swansea
The significant public health threat
from cryptosporidiosis
• Cryptosporidiosis-associated diarrhoeal syndromes are
somewhat dependent on the population affected:
1. a self-limited diarrhoea most often seen in immune
competent individuals
2. a persistent diarrhoea which most commonly afflicts children
in developing countries and can be associated with
nutritional and growth effects
3. a chronic diarrhoea syndrome most often seen in immune
compromised individuals, who may experience other
complications.
Clear need to diagnose and prevent cryptosporidiosis in all populations, with
a need for treatment in some circumstances.
Cryptosporidium is one of the most
significant entero-pathogens
worldwide.
• The global enteric multicentre study (GEMS) (Kotloff et al.,
Lancet 2013):
 one of the leading causes of moderate to severe diarrhoea in
children under 5 years in all seven sites in sub-Saharan Africa
and South Asia,
 with an increased risk of death in toddlers aged 12–23 months,
regardless of HIV prevalence.
• threatens human potential (growth shortfalls, cognitive deficit)
and is involved in the ‘vicious cycle’ of infection and
malnutrition (Bartlet et al., PLOS Neglected Tropical Diseases
2013)
Can cryptosporidiosis still be referred to as
a mild self-limiting illness, even in a “low
risk” population?
• 427 lab confirmed immune-competent cases in the UK with
diarrhoea (Hunter et al., Emerg Infect Dis 2004):
 96% abdominal pain
 65% vomiting
 59% fever
Mean duration 12.7 days; 3 to 4 weeks not uncommon
Symptoms relapse and remit in ~30% cases
14% hospitalised 1 to 9 days (mean 3 days)
Long-term sequelae require further investigation
Acute symptoms
indicate the
pathogenesis:
 infection
predominantly
of the small
bowel
 with
malabsorption
 some
elements of
inflammation
 After clearance, the epithelium usually recovers............
Cryptosporidium putative virulence factors
(thanks to Maha Bouzid for pic; CMR 2013)
Post acute cryptosporidosis health
effects in “low risk” groups
• Case reports of reactive arthritis (Hay et al. 1987; Ozgul et al. 1999;
Shepherd et al. 1989; Cron et al. 1995)
• Suggested cause for relapse of Crohn’s and ulcerative colitis:
including a test for Cryptosporidium is indicated for IBD patients
with sudden exacerbation of digestive symptoms (Manthey et al.
1997)
• C. hominis: joint pain, eye pain, fatigue, headaches in 2 months
post infection (Hunter et al. 2004)
• Anecdotal reports of an association between Cryptosporidium
and IBS are supported by the observation of long-term
pathological changes in rat intestine triggered by C. parvum,
similar to those in IBS patients (Khaldi et al., 2009)
A journey of discovery
Living things
Eukaryota
Cells contain a nucelus and
organelles enclosed within
membranes
Bacteria
Prokaryota
Archaea
Cryptosporidium: a protozoan parasite
The oocyst stage is shed in faeces:
 Robust “shell”; transmissive stage
 Contains 4 sporozoites,
each capable of invading a host cell
 Reproduces by asexual cycling
and by sexual reproduction
 Autoinfection within the host
 Massive numbers of oocysts can be
shed in faeces and some hosts are
chronically infected; infectious dose
<10 oocysts
4-6µm or 5x7µm
depending on species:
currently 26
Sources of Cryptosporidium in human
infection
Two main species cause 96% human
C. parvum
cryptosporidiosis in UK:
C. parvum and C. hominis
C. hominis
Anthroponotic and zoonotic
cycles
Anthroponotic cycle
The problems with Cryptosporidium
• High potential for contamination/transmission from infected
hosts
• Multiple sources; multiple transmission routes
• Robust oocyst stage, survives in environment, waste water
treatment, water treatment ......
• Resistance to common disinfection (e.g. chlorine)
• Causes outbreaks
• Some patients highly vulnerable; some immune-compromised
patients susceptible to severe, life-threatening illness
• Long term sequelae
• Limited treatment options, regimes poorly defined, no licensed
treatment in EU
Human cryptosporidiosis in the UK
• 3000 to 6000 cases reported in England and Wales p.a.
~ 10 / 100 000 population
• Mostly C. parvum or C. hominis; <4% other species
• Watery diarrhoea, acute onset
• Incubation period up to 12 days; usually 5 to 7 days
• 1 to 3 weeks duration; 30% patients experience relapse
• 40% cases in under-5’s
• Immune-compromised patients severe, life-threatening illness
• Treatment regimes poorly defined
• Management by good hygienic practice and exclusion for 48h after first
normal stool
Ascertainment of cryptosporidiosis cases
E&W
n
Potentially: x16
IID studies: for every case reported
Report +ve there are 7 to 8 in the community
to surveillance (Adak et al., 2002; Tam et al., 2011)
Lab tests for
Cryptosporidium
Submits a specimen
Sick patient seeks
medical assistance
True number of cases = ?
PCR detected twice as many
infections (Amar et al., 2007)
Seasonal distribution of Cryptosporidium
species in human cases
400
number of reported cases
350
300
250
200
150
100
50
0
2 5 8 11 14 17 20 23 26 29 32 35 38 41 44 47 50
Year 2000, by week
HPA national surveillance data
CRU typing data
Cryptosporidium reports E&W:
Cases per week as a % of cases per year
changes over time
300
250
200
150
100
50
0
• Cryptosporidium
cases declined since
2001.
• Improved drinking
1989 to 2000
water quality.
• Reduction mostly in
2006 to 2011
spring (C. parvum).
• Autumn cases have
remained (mostly C.
hominis and no
disease reduction).
• Travel and pools are
01 04 07 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52
the likely sources.
Week of year
Data from CfI; thanks to Gordon Nichols
for the slide
Risk factors for Cryptosporidium species
Various species
• Drinking contaminated water: C. parvum, C. hominis, C. cuniculus
• Travel to less industrialised countries: Mainly C. hominis, C. meleagridis,
C. viatorum
• Contact with cats: C. felis
Mainly C. hominis
• Use of swimming pools and water based recreation
• Contact with another person with diarrhoea, especially a child
• Attendance at child care settings
• Changing nappies or toileting young children (even those with no diarrhoea
• Urban
C. parvum
• Contact with animals, especially young ruminants, or animal dung,
• Rural; proximity to muck spreading
Outbreaks of human cryptosporidiosis
Waterborne:
C. cuniculus
C. parvum
C. hominis
C. parvum
C. hominis
drinking water
recreational waters
Farm visits:
C. parvum
open farms, farm open days, residential
farms
Person to person:
all species
households, schools, nurseries, hospitals,
prisons
Environmental:
C. parvum
surface waters, campsites, outdoor centres
Food:
C. parvum
C. hominis
milk (pasteurisation problems), salad items
food handlers
2012: Cryptosporidium outbreak settings
Farm (5, C. parvum)
Enviro (1, C. parvum)
Nursery (1, C. hominis)
Food (1, C. parvum)
Pool (10, C. hominis)
Breakdown of pool settings:
6 at Leisure centres
3 at Holiday or caravan parks
1 at a Hydrotherapy pool in a hospital,
being used for baby and toddler
swimming lessons
Data from CfI
Requirement for better subtyping tools
Systematically connect human pathogen strains/cases with
 other human strains/cases
 animal strains
 food, water and environmental isolates
to identify pathways to exposure and disease, and investigate
outbreaks.
Two approaches have been utilised most
commonly
1. sequence analysis of a single gene encoding a polymorphic sporozoite
surface glycoprotein (gp60 gene)
2. multi-locus analysis of variable number tandem repeats (VNTR) within
a variety of loci
Estimate of zoonotic risk in sporadic cases
Proportion of indigenous
C. parvum cases reporting
contact with farmed animals
(35%)
x
proportion of these cases
with gp60 subtype found in
livestock dung (71%)
= 25%
Applied to national
surveillance data
Outbreak, England, Spring 2012
Over 300 excess cases of acute cryptosporidiosis
were identified between 14th May and 3rd June.
Cases were predominantly female adults, and had
no history of foreign travel.
Geographical clustering, mainly across four
regions of northern England.
Caused by C. parvum gp60 subtype IIaA15G2R1
Case control study found infection was strongly
associated with consumption of pre-cut,
mixed leaf, bagged salad.
Cryptosporidium cases in 2012 compared to the
average from the previous six years
350
Average 2006 to 2011
cases per week
300
Cases 2012
250
Mainly C. hominis
IbA10G2
C. parvum
IIaA15G2R1
foodborne
outbreak
200
150
100
50
0
1
5
9
13
17
21
25
29
Week of year
33
Data from CfI; thanks to Gordon Nichols
for the slide
37
41
45
49
A multi-locus approach would better address
diversity arising from potential re-assortment of
genes during the sexual phase of the life-cycle
Multi-locus markers have been investigated in isolates from:
• a variety of sampling frames
• in varying combinations
• using different assays/platforms and methods of analysis……
Single-locus variant eBURST network
generated from 7 loci of C. parvum in Italy
(Drumo et al., AEM 2008)
Number of
isolates
Global phylogeography of C. hominis:
single-locus variant eBURST network generated from 9 loci,
comparing UK and Ugandan isolates (Chalmers et al., EID 2008;
Tanrivderdi et al., AEM 2008)
UK
Uganda
Microsatellite typing of
Cryptosporidium parvum in
isolates from a waterborne
outbreak
(Hunter et al., JCM 2008)
April/May 2000
238 cryptosporidiosis cases,
though not all were thought to be
related to the primary outbreak.
Nearly all C. parvum P36 strains
were strongly clustered in the
area around Preston and Chorley,
the area most dependent on
water from the implicated supply.
Clustering was highly significant.
Strategy for developing a subtyping
scheme for epidemiology of C. parvum
and C. hominis
underway
MLFT scheme
based on VNTR
Exploring NGS
technologies
Literature review
Biological hurdles
Selection of
markers for MLFT
Bioinformatics
Technical
evaluation
New and improved
markers?
Validation of
scheme
New
assays/platforms
underway
Annual distribution of 82 outbreaks of cryptosporidiosis
linked to swimming pools reported to national
surveillance 1992-2012 (PHE eFOSS data).
Settings of outbreaks of cryptosporidiosis linked to
swimming pools reported to national surveillance
1992-2012 (PHE eFOSS data)
Setting
Leisure centre
Holiday centre
School
Hydrotherapy pool
Sports/health club
Hotel
Caravan park
Baby swimming facility
Shower
Combined spts club and
leisure centre
Number of repted
outbreaks
53
9
5
4
4
2
2
1
1
1
Swimming pool outbreak epidemiology
• Most outbreaks are in
summer/autumn
12
Number of outbreaks
• Mainly linked to learner,
toddler or leisure pools
14
10
8
6
4
2
• High child : adult case ratio
• Precise age groups depend
on risk group
• Association with head
immersion (a proxy for
water consumption)
0
Jan
Feb
Mar
Apr
May
Jun
Jul
Aug
Sept
Oct
Nov
Month
Monthly distribution of outbreaks of cryptosporidiosis linked to
swimming pools reported to national surveillance 1992-2012 (PHE
eFOSS data).
Dec
The swimming pool
Components of a
swimming pool:
1.
Tank
2.
Circulation system
3.
Filtration
4.
Dosing for treatment
chemicals.
A re-circulating system:
continuous, to keep
water clear of dirt,
debris and microorganisms.
Swimming pool treatment: disinfection
 no effective residual against Cryptosporidium
Pathogen
E. coli O157
Giardia
Cryptosporidium
Chlorine survival*
1mg/L, pH7.5, 25oC
< 1 min
45 min
10.6 days
 additional treatments e.g. UV, ozone, are progressive,
in the plant room
*Source http://www.cdc.gov/healthywater/swimming/pools/chlorine-disinfection-timetable.html
Swimming pool treatment: filtration
Swimming pool filtration was designed to provide a physically
clean, clear and safe environment, not specifically to remove
Cryptosporidium
•
Small size (oocysts 4-6µm)
•
Require low or medium rate filters with coagulation
•
As the filtration rate increases the log removal rate decreases
and requires multiple turnover cycles to remove the risk
Rachel Chalmers
Cryptosporidium Reference Unit
Public Health Wales Microbiology
Singleton Hospital
Swansea
01792 285341