Shiga Toxin-Producing E. coli
and Ruminant Diets: A Match
Made in Heaven?
Todd Callaway
Agricultural Research Service/USDA...
Escherichia coli O157:H7 & friends
• N. America has a VERY safe food supply, but it is
still a focus of concern
• Enterhem...
phuman infection = panimal infection x pcarcass contamination
x pproduct abuse x peaten by susceptible
= 0.05 x 0.4 x 0.02...
Carcass
Contamination
Hide
Contamination
Fecal
Shedding/
Super-Shedder
Environment
Fecal
Microbiome
Host
Diet
Why should we worry about
EHEC in live cattle?
• Several points for improvement
– Food Safety (Reduced pathogen burden)
• ...
Bovine Gastrointestinal Tract, ‘Generic’ E. coli, and E. coli O157:H7
Total Bacteria consortium (per gram digesta) > 2000 ...
Bovine Colonization and E. coli O157:H7
Colonization appears to be primarily at RAJ
(rectoanal junction)
Infection in catt...
• Management/ecological niche factors
– Diet (including specific feeds and additives)
Effects
Distillers Grains
Orange pee...
Dietary factors in cattle
• Co-evolution of host (lack of toxin receptors)
• Fasting increases shedding of EHEC (Buchko) a...
0
2
4
6
8
10
-20 0 20 40 60 80 100
Rumen
Colon
E.coli
Grain (%)
Effect of Diet on intestinal coliform and E. coli populati...
Bovine Diet and E. coli O157:H7: Forage vs Grain Diets
Wells et al, unpublished
In a on-going study, animals fed
forage-ba...
DDGS impacts
• Distillers grains are increasingly included in
animal rations
– Significantly alter the gut microbiome (Pin...
Fecal Samples
High Moisture/Dry-rolled Corn
Based Diet
(0 or 40% WDGS)Fecal prevalence and percent of fecal
samples with e...
Finishing Phase (0 or 40% WDGS)
Hide Samples
Hide prevalence and percent of
hide samples with enumerable
E. coli O157:H7 w...
Finishing Phase
Prior to Shift
(0, 40 or 70% WDGS)
Fecal Samples
Bovine Diet and E. coli O157:H7: WDGS in Finishing Diets
...
Finishing Phase
Time after Shift
Fecal Samples
Reductions in WDGS in
the diet did not result in
reduction of E. coli
O157:...
The Bovine Fecal Microbiome: Corn vs WDGS
Finishing feedlot diets.
N = 4, 0 or 40% WDGS
Shifts in Prevotella and
Bacteroid...
The Bovine Fecal Microbiome: Diet and OTUs Association
A B
Heifer-10
Steer-10
Steer-09
Steer-09
Heifer-10
Steer-10
Princip...
Red: Top 10% Average
Enumerable E. coli O157:H7
Blue: Lowest 10% Average
Enumerable E. coli O157:H7
Community structure ma...
Specific Microbiome Taxa Association with E. coli O157:H7
30% Alfalfa Haylage
70% Corn Silage
66% Corn
26% Corn Silage
8% ...
The Bovine Fecal Microbiome and E. coli O157:H7
Propensity to shed E. coli O157:H7 appears to associate with
groups of OTU...
Tasco, tannins, and essential oils
• Produced from brown seaweed
• Reduced growth of
EHEC and Salmonella in
In vitro ferme...
Dried orange peel/pulp
• By-product, competitively prices into least-
cost ration formulation
– Used widely in dairy and f...
Sheep fed 20% orange
peel consumed less peel
than did sheep fed 10%
0
1
2
3
4
5
6 0
10
20
SalmonellaentericaTyphimurium
(l...
Dowd et al., 2006
Stress can affect microbial populationsStress can affect microbial populations
Bailey et al., 2011
Cecal...
Oral administration of
Norepinephrine
Pullinger et al., 2010
Stress is a complicated issue
It’s like pornography
Hard to r...
Conclusions
• E. coli O157:H7 (and other EHEC?) co evolved
with cattle, so combatting it is neither easy, nor
straightforw...
9th
International Symposium on
Shiga Toxin (Verocytotoxin)
Producing Escherichia coli
Infections
Boston, Massachusetts
Tod...
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Shiga Toxin-Producing E. coli and Ruminant Diets: A Match Made in Heaven?

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Presented at 2013 Arkansas Association for Food Protection annual conference.

Todd Callaway
Agricultural Research Service/USDA
Food and Feed Safety Research Unit
College Station, TX

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Shiga Toxin-Producing E. coli and Ruminant Diets: A Match Made in Heaven?

  1. 1. Shiga Toxin-Producing E. coli and Ruminant Diets: A Match Made in Heaven? Todd Callaway Agricultural Research Service/USDA Food and Feed Safety Research Unit College Station, TX
  2. 2. Escherichia coli O157:H7 & friends • N. America has a VERY safe food supply, but it is still a focus of concern • Enterhemorrhagic E. coli (EHEC) – Bloody diarrhea, HUS, children – O157:H7, O26, O45, O103, O111, O121 and O145 are adulterants (FSIS) in beef • O157 evolved with cattle, lives in intestinal tract, adhering to intestinal mucosa – Recto anal junction, lymphoid tissue – 50,000+ years of evolution • Depending on season 40 % of cattle may have O157 (non-O157 under investigation) – May be higher during summer, near 80% – “Supershedders” >105 CFU/g Source: ASM
  3. 3. phuman infection = panimal infection x pcarcass contamination x pproduct abuse x peaten by susceptible = 0.05 x 0.4 x 0.02 x 0.2 = 0.00008 or 24,800 cases/year in a population of 310 million Thus reductions in pathogens before harvest can yield large returns in human health and reduced recalls Focus on food safety “From Farm to Fork” Processors say “below 103 cells/g, and we can take care of it” Preharvest prevalence directly affects human illnesses
  4. 4. Carcass Contamination Hide Contamination Fecal Shedding/ Super-Shedder Environment Fecal Microbiome Host Diet
  5. 5. Why should we worry about EHEC in live cattle? • Several points for improvement – Food Safety (Reduced pathogen burden) • Cross contamination of carcasses and foods • Increased in-plant effectiveness – Water Issues (indirect) • irrigation and run-off – Spinach outbreak; Walkerton, ON – Agricultural Fair/Open Farms (Petting zoo)/Farm worker safety (Keen et al., 2003) • Dust from cattle and O157:H7 outbreak
  6. 6. Bovine Gastrointestinal Tract, ‘Generic’ E. coli, and E. coli O157:H7 Total Bacteria consortium (per gram digesta) > 2000 species 1010 106 1010 1011 Total ‘generic’ E. coli (per gram digesta) 104 (104 ) (107-8 ) 107 E. coli O157:H7, when present (per gram digesta) <102 <102 <102 <106 Rumen, reticulum, omasum, and abomasum (fermentation reservoir) Small Intestine Large Intestine Colon Cecum
  7. 7. Bovine Colonization and E. coli O157:H7 Colonization appears to be primarily at RAJ (rectoanal junction) Infection in cattle is asymptomatic (lack of toxin receptors) Seasonality and variable Super-shedders (>104 CFU E. coli O157:H7 per gram feces) are a minor proportion Need >20 animals to sustain levels of E. coli O157:H7 in the pen (recirculation) Wells et al., 2008
  8. 8. • Management/ecological niche factors – Diet (including specific feeds and additives) Effects Distillers Grains Orange peel and pulp can reduce (essential oils) Forage feeding can alter microbiome and EHEC; forage quality – Microbial Environment/stress Cross-talk between host and microbes Norepinephrine role for “microbial organ” Microbiome and nutrient transporters and EHEC What factors affect EHEC in cattle GIT?
  9. 9. Dietary factors in cattle • Co-evolution of host (lack of toxin receptors) • Fasting increases shedding of EHEC (Buchko) and Salmonella (Grau) – Lack of VFA which are toxic to E. coli/Salmonella • Barley increased EHEC shedding (Buchko) • Steam flaked corn increased EHEC relative to dry rolled corn (Fox) • Monensin generally has no effect on EHEC (Edrington, McAllister, Van Baale) – Increased dosage of 44 mg/kg did reduce (Paddock)
  10. 10. 0 2 4 6 8 10 -20 0 20 40 60 80 100 Rumen Colon E.coli Grain (%) Effect of Diet on intestinal coliform and E. coli populations (Adapted from Diez Gonzalez et al., 1998) -4 -2 0 2 4 6 8 10 12 Time (d) Abrupt shift from 90% grain to 100% alfalfa hay E. coli (Log10 cells/g) 9 8 6 4 7 5 3
  11. 11. Bovine Diet and E. coli O157:H7: Forage vs Grain Diets Wells et al, unpublished In a on-going study, animals fed forage-based growing rations over 2 study years had higher fecal prevalence for E. coli O157:H7 than animals on corn- based finishing rations. Long term feeding of forage-type diets may increase E. coli O157:H7. May be related to survival in manure and soils (Durso; Berry) Forage quality appears to play a role in some form
  12. 12. DDGS impacts • Distillers grains are increasingly included in animal rations – Significantly alter the gut microbiome (Pinchak) • Increase fecal shedding of E. coli O157:H7 in cattle (Wells, 2009; Jacob, 2008) • It appears that there is an effect of DG compared to corn on O157 populations, but magnitude is fairly small overall ~1.5 log10, but affects incidence – Unknown mechanism – Batch variability/QC issues
  13. 13. Fecal Samples High Moisture/Dry-rolled Corn Based Diet (0 or 40% WDGS)Fecal prevalence and percent of fecal samples with enumerable E. coli O157:H7 were significantly different for the pens. Fecal counts for E. coli O157:H7 were not significantly different at any Bovine Diet and E. coli O157:H7: WDGS in Finishing Diets
  14. 14. Finishing Phase (0 or 40% WDGS) Hide Samples Hide prevalence and percent of hide samples with enumerable E. coli O157:H7 were different for the pens. Hide counts for E. coli O157:H7 were significantly different from 40 CFU/100cm2 to 480 CFU/100cm2 . Bovine Diet and E. coli O157:H7: WDGS in Finishing Diets Wells et al, 2008
  15. 15. Finishing Phase Prior to Shift (0, 40 or 70% WDGS) Fecal Samples Bovine Diet and E. coli O157:H7: WDGS in Finishing Diets Wells et al, 2010 After Shift
  16. 16. Finishing Phase Time after Shift Fecal Samples Reductions in WDGS in the diet did not result in reduction of E. coli O157:H7 in feces by day 28, but did by day 56. Bovine Diet and E. coli O157:H7: Shift WDGS from Finishing Diets Wells et al, 2010
  17. 17. The Bovine Fecal Microbiome: Corn vs WDGS Finishing feedlot diets. N = 4, 0 or 40% WDGS Shifts in Prevotella and Bacteroides. WDGS diet resulted in more Sporacetigenium and Anaerovorax E. coli O157:H7 shedding not detected Durso et al, 2010
  18. 18. The Bovine Fecal Microbiome: Diet and OTUs Association A B Heifer-10 Steer-10 Steer-09 Steer-09 Heifer-10 Steer-10 Principle Coordinate Analysis Weighted UniFrac Principle Coordinate Analysis Unweighted UniFrac Diet drives the fecal microbial composition in cattle Heifer-10 30% Alfalfa Haylage 70% Corn Silage Steer-09 66% Corn 26% Corn Silage 8% Supplement Steer-10 83% Corn 12% Corn Silage 5% Supplement Kim et al, submitted JAS
  19. 19. Red: Top 10% Average Enumerable E. coli O157:H7 Blue: Lowest 10% Average Enumerable E. coli O157:H7 Community structure may not be driving shedding Principle Coordinate Analysis Weighted UniFrac Heifers 30% Alfalfa Haylage 70% Corn Silage Steers 66% Corn 26% Corn Silage 8% Supplement Steers 83% Corn 12% Corn Silage 5% Supplement Microbiome OTU Composition and Shedding of E. coli O157:H7 Kim et al, in preparation
  20. 20. Specific Microbiome Taxa Association with E. coli O157:H7 30% Alfalfa Haylage 70% Corn Silage 66% Corn 26% Corn Silage 8% Supplement 83% Corn 12% Corn Silage 5% Supplement No obvious single taxa group driving shedding No community association for heifers. Weak separation for steers diets Community structure not driving supershedding Few OTUs associated supershedding for any of the three diets Kim et al, in preparation
  21. 21. The Bovine Fecal Microbiome and E. coli O157:H7 Propensity to shed E. coli O157:H7 appears to associate with groups of OTUs within diet for animals fed finishing diets that are corn-based. In contrast, there appeared to be little association between shedding E. coli O157:H7 and any single OTU or groups of OTUs for animals fed the corn silage/haylage diet. Shedding high levels of E. coli O157:H7 appears to have low association with any OTU or groups of OTUs for any diet.
  22. 22. Tasco, tannins, and essential oils • Produced from brown seaweed • Reduced growth of EHEC and Salmonella in In vitro fermentations • Improves meat quality and shelf life • Polyphenolics (tannins) can reduce EHEC – Naturally found in plants (Min; Lacombe) • Essential oils can be bactericidal (spices) – Limonene, terpenes, thymol, oregano (Varel)
  23. 23. Dried orange peel/pulp • By-product, competitively prices into least- cost ration formulation – Used widely in dairy and feedlot rations in Florida and Southern California • Orange oils (e.g., limonene) improve animal health and productivity (esp. in swine) • Source of pectin and fermentable fiber to ruminal microbial population – 9% CP, TDN 82%, DE 1.64, NEm 0.88, and NEg 0.59 Mcal/lb (CSU)
  24. 24. Sheep fed 20% orange peel consumed less peel than did sheep fed 10% 0 1 2 3 4 5 6 0 10 20 SalmonellaentericaTyphimurium (log 10 CFU/ml) Rumen Cecum Rectum 0 5 10 0 1 2 3 4 E.coliO157:H7CFUlog 10 /gdigesta Rumen Cecum Rectum a b b ba aa b b 5 and 10% OP reduced E. coli O157:H7 in rumen, cecum and rectum Final pH , TVFA, and A:P ratio were not affected Doesn’t work with Campylobacter
  25. 25. Dowd et al., 2006 Stress can affect microbial populationsStress can affect microbial populations Bailey et al., 2011 Cecal community structure changes following social disruption (SDR) Inverse relationship bet. cytokines (IL-6) and bacterial populations
  26. 26. Oral administration of Norepinephrine Pullinger et al., 2010 Stress is a complicated issue It’s like pornography Hard to replicate Host/bacterial crosstalk (Lyte) “Feed Me” signals Microbial organ as a driver of homeostasis and disease
  27. 27. Conclusions • E. coli O157:H7 (and other EHEC?) co evolved with cattle, so combatting it is neither easy, nor straightforward • Diet impacts O157:H7 populations – Forage quality, WDGS vs corn, dry rolled vs steam-flake • Diet changes the microbiome population • Diet-driven microbiota changes are not linked to O157:H7 shedding shifts – Generic E. coli yes… • What is driving the microbiome/EHEC change? – Future avenues for exploring this linkage are evolving rapidly
  28. 28. 9th International Symposium on Shiga Toxin (Verocytotoxin) Producing Escherichia coli Infections Boston, Massachusetts Todd Callaway Jeff LeJeune 31 May- 3 June, 2015

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