Are all VTEC created Equal?
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1 PHL-HSE-Dublin Mid Leinster Are all VTEC created Equal? Anne Carroll
2 Escherichia coli Commensal Microrganism but some strains are cause of infections in humans Syndromes associated to E. coli infections: Urinary tract infections Sepsis/meningitis Enteric Infections-diarrhoea
3 Diarrheagenic E. coli Abbreviation Pathotypes Genes Action DAEC EAEC EIEC ETEC EPEC VTEC EHEC Diffusely-adherent afa Afimbrial adhesion Enteroaggregative aggr Regulator, plasmid encoded Enteroinvasive ipah invasive Enterotoxigenic esta eltb Heat stable/labile toxins Enteropathogenic eae, EAF Intimin attaching effacing, LEE. (chr) Adherence factor (Plasmid) Verotoxigenic vt1, vt2 vero./shiga toxins encoded on bacteriophages Enterohaemorrhagic vt1, vt2 eae ehly Haemorrhagic colitis aat anti-agg protein transporter aap (dispersin)protein aaic chr gene. VT1, a,c,d. VT2, a,b,c,d,e,f,g.
4 attaching and effacing (A/E) lesion
5 Human VTEC Infection Adults visiting GP abdominal pain 93% bloody diarrhoea 25% Severe diarrhoea 60% Fever atypical vomiting 11% Median duration of diarrhoea 9 days Source: Foods Standards Agency. A report of the study of infectious intestinal disease in England. London: The Stationery Office,
6 Severe Infection Hemolytic uremic syndrome (HUS) up to 10% Children, elderly, immunocompromised Usually develops post-diarrhoea Mortality Clinical signs Kidney failure, hemolytic anemia, Thrombocytopenia 6
7 VTEC in Ireland Total VTEC Cases. PCR positive culture negative and Isolates Total isolated strains O157 isolated strains only O26 isolated strains only Total non-o157/o26vtec Cases. PCR positive culture negative and Isolates non-o157/o26isolated strains only
8 Number of Specimens Analysed Number of tests
9 VTEC in Ireland 100% 90% 80% 70% 60% 50% 40% Other O26 O157 30% 20% 10% 0%
10 Serogroup VT1 VT1+VT2 VT2 Total O Unidentifiable O O O O O O O O O O O O O O O O O O O O O O169-O O O O O O O O O O2 4 4 O O O O O O O O O O O8 3 3 O O O9 1 1 O O O PCR only Grand Total :49 serogroups
11 Number of notifications % symptomatic among known VTEC Symptoms % % 80% % % 50% 40% 30% 20% 10% Unknown Asymptomatic Symptomatic % symptomatic among known year of notification 0% Number of VTEC notifications by symptom status, Ireland
12 Most common Questions Why do we investigate vtx1 only, international evidence suggests they are not virulent? Is it necessary to do so much screening in outbreaks of VTEC in creches? EntericBio is positive reference lab result is negative, what to do? What is the significance of PCR positive culture negative results?
13 Should vtx1 be investigated?
14 Virulence factors vtx1 subtypes (vtx1a, vtx1c, vtx1d) vtx2 subtypes (vtx2a, vtx2b, vtx2c,vtx2d, vtx2e, vtx2f and vtx2g) Investigate the prevalence of virulence genes eae, hlya, aggr, and aaic No one proven virulent genotype vtx2 associated with HUS
15 Assess if there is a correlation between virulence profile and clinical presentation HUS Bloody Diarrhoea Diarrhoea Asymptomatic
16 Methodology 176 confirmed VTEC isolates 4 cohort clinical groups 50 per group 1:1 outbreak/sporadic (where possible)).
17 % Serogroup distribution for symptom groups in this study 100% 90% Percentage (%) 80% 70% 60% 50% 40% 30% 20% 10% Ungroupable O111 O55 O1O3 O145 O26 O157 0% HUS BD D A Symptom Group
18 Results: Virulence Profiles 32 individual profiles identified (P1-P32) 12/32 profiles expressed by 4 samples vtx2e, vtx2f and vtx2g = negative aggr and aaic = negative (EAEC)
19 Most Common Virulence profiles (n=12/32) P Virulence profile Number of cases HUS BD D A 1 vtx1a + vtx1c + eae + hlya (6.5%) 1 (2%) 9 (19.6%) 8 (16.3%) 7 vtx2a + eae + hlya (12.9%) 3 (6%) - 3 (6.1%) 10 vtx2a + eae (6.5%) 9 (18%) - 1 (2%) 11 vtx2a + vtx2c + eae + hlya (6.5%) (%) 6 (12.2%) 12 vtx2a + vtx2c + eae (16.1%) 7 (14%) 2 (4.3%) - 15 vtx2a + vtx2d + eae (12.9%) 8 (16.%) 1 (2.2%) - 16 vtx2a + vtx2d + eae+ hlya 25 1 (3.2%) 1 (2.%) 13(28.3%) 10 (20.4%) 18 vtx1a + vtx1c + vtx2a + eae (9.7%) 1 (2.%) vtx1a + vtx1c+ vtx2a + vtx2c + eae (3.2%) 9 (18%) vtx1a + vtx1c + vtx2a + vtx2d + eae+ hlya (3.2%) - 8 (17.4%) 2 (4.1%) 24 vtx1a + vtx1c + vtx2c + eae + hlya (4.3%) 2 (4.1%) 29 vtx1a + vtx1c + vtx2a + eae + hlya (8.2%)
20 Potentially more severe disease P Virulence profile Number of cases HUS BD D A 1 vtx1a + vtx1c + eae + hlya (6.5%) 1 (2%) 9 (19.6%) 8 (16.3%) 7 vtx2a + eae + hlya (12.9%) 3 (6%) - 3 (6.1%) 10 vtx2a + eae (6.5%) 9 (18%) - 1 (2%) 11 vtx2a + vtx2c + eae + hlya (6.5%) (%) 6 (12.2%) 12 vtx2a + vtx2c + eae (16.1%) 7 (14%) 2 (4.3%) - 15 vtx2a + vtx2d + eae (12.9%) 8 (16.%) 1 (2.2%) - 16 vtx2a + vtx2d + eae+ hlya 25 1 (3.2%) 1 (2.%) 13(28.3%) 10 (20.4%) 18 vtx1a + vtx1c + vtx2a + eae (9.7%) 1 (2.%) vtx1a + vtx1c+ vtx2a + vtx2c + eae (3.2%) 9 (18%) vtx1a + vtx1c + vtx2a + vtx2d + eae+ hlya (3.2%) - 8 (17.4%) 2 (4.1%) 24 vtx1a + vtx1c + vtx2c + eae + hlya (4.3%) 2 (4.1%) 29 vtx1a + vtx1c + vtx2a + eae + hlya (8.2%)
21 VT1 P Virulence profile Number of cases HUS BD D A 1 vtx1a + vtx1c + eae + hlya (6.5%) 1 (2%) 9 (19.6%) 8 (16.3%) 7 vtx2a + eae + hlya (12.9%) 3 (6%) - 3 (6.1%) 10 vtx2a + eae (6.5%) 9 (18%) - 1 (2%) 11 vtx2a + vtx2c + eae + hlya (6.5%) (%) 6 (12.2%) 12 vtx2a + vtx2c + eae (16.1%) 7 (14%) 2 (4.3%) - 15 vtx2a + vtx2d + eae (12.9%) 8 (16.%) 1 (2.2%) - 16 vtx2a + vtx2d + eae+ hlya 25 1 (3.2%) 1 (2.%) 13(28.3%) 10 (20.4%) 18 vtx1a + vtx1c + vtx2a + eae (9.7%) 1 (2.%) vtx1a + vtx1c+ vtx2a + vtx2c + eae (3.2%) 9 (18%) vtx1a + vtx1c + vtx2a + vtx2d + eae+ hlya (3.2%) - 8 (17.4%) 2 (4.1%) 24 vtx1a + vtx1c + vtx2c + eae + hlya (4.3%) 2 (4.1%) 29 vtx1a + vtx1c + vtx2a + eae + hlya (8.2%)
22 Potentially less severe disease P Virulence profile Number of cases HUS BD D A 1 vtx1a + vtx1c + eae + hlya (6.5%) 1 (2%) 9 (19.6%) 8 (16.3%) 7 vtx2a + eae + hlya (12.9%) 3 (6%) - 3 (6.1%) 10 vtx2a + eae (6.5%) 9 (18%) - 1 (2%) 11 vtx2a + vtx2c + eae + hlya (6.5%) (%) 6 (12.2%) 12 vtx2a + vtx2c + eae (16.1%) 7 (14%) 2 (4.3%) - 15 vtx2a + vtx2d + eae (12.9%) 8 (16.%) 1 (2.2%) - 16 vtx2a + vtx2d + eae+ hlya 25 1 (3.2%) 1 (2.%) 13(28.3%) 10 (20.4%) 18 vtx1a + vtx1c + vtx2a + eae (9.7%) 1 (2.%) vtx1a + vtx1c+ vtx2a + vtx2c + eae (3.2%) 9 (18%) vtx1a + vtx1c + vtx2a + vtx2d + eae+ hlya (3.2%) - 8 (17.4%) 2 (4.1%) 24 vtx1a + vtx1c + vtx2c + eae + hlya (4.3%) 2 (4.1%) 29 vtx1a + vtx1c + vtx2a + eae + hlya (8.2%)
23 HUS genotypes Serogroup 1a 1a2a 1a2a2c 1a2a2d 2a 2a2b2c 2a2c 2a2c2d 2a2d Total O O O O O O Total
24 2017, 2 more HUS caused by O26 vtx1
25 2017, 2 more HUS caused by O26 vtx1 Yes we need to investigate vtx1 only VTEC
26 Is it necessary to do so much screening in outbreaks of VTEC in creches?
27 Number of notifications Rate per 100,000 population Age group (years) 0 Female - Cases Male - Cases Female - Rate Male - Rates
28 Number of outbreaks * Person-to-person Foodborne +/- P-P Other Year of notification Waterborne +/- P-P Animal contact/environmental +/- P-P Unknown/not specified
29
30 VTEC Risk assessment 1. Confirm the diagnosis in the index case 2. Transmission: Determine the likelihood of spread from index case to close contacts Determine the possibility of spread beyond immediate contacts (for example, is the index case in a risk category?) 3. Source: Determine the likely mechanism of exposure Determine if there could be a potential continuing source of infection 4. Assess what initial levels of control (if any) are required to be put in place.
31 CCF investigation An outbreak should be declared if: children/staff report symptoms of HUS in the previous two weeks, an children/staff report even a single episode of bloody diarrhoea within the previous week children/staff report diarrhoea in the week before the index became ill, should submit a stool sample for analysis and should not attend the CCF. children/staff develops diarrhoea in the week following the last attendance of the index at the CCF, should provide a single stool sample for testing and not attend the CCF If, after testing, any other children/staff are VTEC positive (or subsequently report symptoms of HUS/bloody diarrhoea) an outbreak should be declared. If any of the results return a toxin producing E. coli that is of a different serogroup or VT type than that of the index, a risk assessment should be performed to determine the potential impact of this finding.
32 CCF outbreak All staff and children are to be excluded All children regardless of age and all staff should submit two stool samples taken at least 24h apart to establish microbiological clearance Parents should be instructed not to allow their children to attend other childcare facilities while awaiting microbiological clearance Children and staff should be allowed back on an individual basis once they are asymptomatic for 48 hours and have two negative stool samples, taken at least 24 hours apart (these 2 negative samples must be taken after symptoms have subsided if symptomatic)
33 CCF outbreak Microbiological Clearance: Only children and staff who have met the conditions for microbiological clearance will be allowed to return to the CCF Hygiene Review: Food/kitchen hygiene Hand washing facilities Nappy changing facilities Toilet hygiene Waste disposal Drinking/washing water hygiene Pet/external environment hygiene Internal environmental hygiene (toys, door handles and other touch surfaces, etc) Deep Cleaning: Sufficient Personnel:
34 In Ireland Screen stools at local/regional lab by PCR Send positive stool or isolate to ref lab Or Send all stools to ref lab for screening
35
36 EntericBio is positive reference lab result is negative? What is the significance of PCR positive culture negative results?
37 Both proven sensitive and specific methods Different portions tested Different gene targets Very low numbers i.e. high cp
38 Clinical Relevance PCR Pos culture negative Enteric samples not sterile Presence of free vtx toxins Patient symptomatic? Risk group? Likely infection source
39 Clinical Relevance Interpretation of result is key Implications of false negative and positive result Lab and clinical need to be taken together Difficult for scientists!!!
40 Thank you Questions?
41
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