Desiccation and persistence of Cronobacter spp. (E. sakazakii) in infant formula. 1 st Intl Conference on Cronobacter UCD January 2009
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1 Desiccation and persistence of Cronobacter spp. (E. sakazakii) in infant formula 1 st Intl Conference on Cronobacter UCD January 2009
2 Desiccation and persistence of Cronobacter spp. (E. sakazakii) in infant formula 1 st Intl Conference on Cronobacter UCD January 2009
3 Desiccation and persistence of Cronobacter spp. (E. sakazakii) in infant formula 1 st Intl Conference on Cronobacter UCD January 2009
4 Structure! " # $!! %
5 Characteristics of Cronobacter spp. &'()* + $ "
6
7 Low impact journal! Only 5 pages,--).'/!0 Six strains of Cr. sakazakii 3 from Franco and Jeff Enteral feeding tubes and baby bottles FAO/WHO 2004 One PAGE! JEMRA Risk Model FAO/WHO 2004 Key method reference: Druggan, Forsythe & Silley (1993)
8 Sources of Cronobacter spp.
9
10 Being right for the wrong reason!
11 Sources of Cronobacter spp. Plants Mixed salad vegetables Rice seed Herbs & spices Grain Cereal Rhizosphere Ingredients Dried flour or meal (corn, soy, wheat and rice) Dried sodium caseinate Starches Potato flour Products Powdered infant formula Follow-on formula Weaning foods Milk powder Dried foods Dried vegetables and spices Infant cereals Adult cereals Tofu Iced tea Other Household dust Hospital air Preparation equipment For more details see Friedemann 2007
12 Sources of Cronobacter spp. Yellow pigmentation Oxygen radical protection Capsule Attachment and desiccation resistance Efflux pumps Essential oil resistance
13 Desiccation persistence
14 Desiccation persistence,*.- 1 Caubilla-Barron & Forsythe 2007
15 Desiccation persistence Cronobacter spp. Strain Capsulation Source TSA VRBGA Difference Cr. sakazakii 1 No Clinical Cr. sakazakii 4 Clinical Cr. sakazakii 23 PIF Cr. muytjensii 3 Clinical Cr. sakazakii 2 Yes Clinical Cr. sakazakii 7 PIF Cr. sakazakii 10 Nuts Cr. sakazakii 13 Herb Cr. sakazakii 25 PIF Cr. turicensis 9 Baby food Caubilla-Barron & Forsythe 2007
16 Desiccation persistence 8 6 Viable count (log cfu/ml) 4 2 P antoea spp. C. freundii S. Enteritidis Time (months) Pantoea spp. C. freundii Salm. Enteritidis Caubilla-Barron & Forsythe 2007
17 Desiccation persistence 8 6 Viable count (log cfu/ml) Time (months) Strain number C. sakazakii 1 C. sakazakii 2 C. sakazakii 25 C. turicensis C. muytjensii Caubilla-Barron & Forsythe 2007
18 Desiccation persistence 8 Viable count (log cfu/ml) Biphasic decline Subpopulation more resistant? Time (months) Caubilla-Barron & Forsythe 2007
19 Capsule and biofilm formation Intrinsic (<1 cfu/g) Equipment (Reported) People (unknown) Reconstitution Ambient temperature storage Baby stomach Probability of ingestion and Probability of infection Biofilm formation (recontamination) Forsythe 2005
20 Capsule and biofilm formation Organism Source Capsule production a Polyvinylchloride (log 10 cfu/tube) Av Max Min Cr. sakazakii NCTC T Infant clinical Cr. sakazakii ATCC Unknown Cr. sakazakii 1 PIF Cr. sakazakii 2 PIF Cr. sakazakii 3 PIF Cr. sakazakii 4 PIF Cr. sakazakii 5 PIF Cr. sakazakii 6 PIF Cr. sakazakii 7 Unknown Cr. sakazakii 8 Unknown Cr. sakazakii 9 Environmental Poster P14
21 Thermal resistance Organism D 55 min Cr. sakazakii 3-49 Cr. malonaticus 6 Cr. malonaticus 3 Cr. turicensis 3 Cr. muytjensii 3 Cr. dublinensis 3 Salm. Senftenberg 36 E. hormaechei 2-54 Poster P25
22 D-values of unstressed and stressed Cronobacter spp. in reconstituted PIF Temperature Unstressed Stress treatment ( C) Desiccation Starvation Heat Cold ,--3
23 Mechanisms of resistance Compatible solutes Trehalose and maltose are both disaccharides of glucose Trehalose has two (11)-glycosidic linkages and no reducing sugar residue Maltose (14)-glycosidic linkage, plus reducing end sugar !!27! 7,78' Breeuwer et al. 2003
24 Mechanisms of resistance Compatible solutes Proteins peptides amino acids ecotine ecotine peptides proline betaine betaine trehalose sugars choline taurine carnitine choline taurine carnitine Breeuwer et al. 2003
25 Mechanisms of resistance "..
26 Mechanisms of resistance Proteins associated with desiccation and osmotic stress adaptation DNA protecting protein under starved conditions Dps Cold shock-like protein CspC OmpC OmpA Thermoregulated motility protein Super oxide dismutase Chaperonoin GroES ABC-type proline/glycine betaine transport systems Hypothetical protein Mfla_1164 Hypothetical protein Mfla_1165 Hypothetical protein Psyc_0523 Trehalose uptake Capsule production Putative gene in C. sakazakii BAA-894 Yes Yes Yes Yes Yes Yes Yes Yes NF NF Yes Yes* Yes* / :;<!!!! 7 9/ For more detail see Reidel & Lehner 2007
27 Summary Thank you Origin of Cronobacter Physiological resistance traits Cross-protection : heat and desiccation stress Desiccation during processing Induction of thermotolerance reconstitution temperature efficacy Capsulation - lower proportion of sublethally injured cells which revive and multiply and form biofilms!
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