Potential of plant-derived antimicrobials for controlling zoonotic and food-borne diseases

Transcription

1 Potentil of plnt-derived ntimicrobils for controlling zoonotic nd food-borne diseses Kumr Venkitnrynn, DVM, MVSc, MS, Ph.D. Professor of Microbiology Grdute Progrms Chir Deprtment of Animl Science University of Connecticut Storrs, CT 06269, USA

2 Zoonotic Diseses About 75% of recently emerging infectious diseses ffecting humns re diseses of niml origin, nd pproximtely 60% of ll humn pthogens re zoonotic. Food-borne diseses Wide rnge of niml reservoirs Emergence of ntibiotic resistnce

3 Plnt-Derived Antimicrobils (PDAs) An lterntive pproch Phytophenolics Plnt defense mechnism Wide spectrum of biologicl effects Bcteril resistnce low Burt et l., 2004; Ohno et l., 2007; Wollenweber, 1988

4 Plnt-derived ntimicrobils (PDAs) Trns-cinnmldehyde(TC) Crvcrol(CR) Thymol(TH) Eugenol (EU) Cprylic cid (CA) Cinnmon oil Oregno oil Clove oil Coconut oil

5 Efficcy of PDAs for reducing egg-borne trnsmission of Slmonell Enteritidis

6 Slmonell Mjor food-borne pthogen worldwide Highest incidence rte for Slmonell infections 1 Totl estimted nnul cost 4 billion USD 2 1 CDC 2010, 2 Schrff, 2012

7 Slmonelloutbreks in the US Slmonell Pthogenic E. coli Listeri monocytogenes No reported outbreks

8 Foods ssocited with Slmonelloutbreks Source: CDC Ntionl Outbrek Reporting System,

9 SlmonellEpidemiology Chicken reservoir host SlmonellEnteritidis(SE) most common Slmonellfrom poultry 1 Consumption of rw or undercooked eggs 100 billion tble eggs re produced nnully 1 Mrcus et l., 2007; 2 AMI, 2009; 3 CDC, 2010

10 Slmonellchicken link Primry coloniztion site cecum Other sites crop, intestine, cloc Internl orgns liver, spleen, oviduct Trnsmission routes: Horizontl bird to bird Verticl Trnsovrin-bird to yolk (mcrophges involved in systemic spred) 1,2 1 Gstet l., 2007; 2 Gntoiset l., 2009

11 Controlling Slmonell Idel Intervention Strtegy Economiclly vible Prcticl for frmers to dopt No toxicity Orgnic frming Environmentlly friendly No bcteril resistnce development

12 Rtionle Supplementtion of PDAs Reduce cecl coloniztion Reduce oviduct coloniztion Reduce fecl contmintion of shelled eggs Reduce yolk nd membrne contmintion Control slmonellosis

13 Chllenge experiment in chickens 120 White Leghorn lyer chickens 25 & 40 weeks of ge Tretments 1% TC control (No SE, 1% TC) 1.5% TC control (No SE, 1.5% TC) Positive control (SE, No TC) 1% TC (SE, 1% TC) 1.5% TC (SE, 1.5% TC) SE-28, SE-21, SE-12, SE-31, SE-90 13

14 Protocol TRANS-CINNAMALDEHYDE FEEDING PHASE Weeks Dy 1 Grouping of birds Dy 14 Egg Collection strts Dily egg collection Weekly fecl swb Dy 70 End of tril Collection of tissues Dy 10 Chllenge with SE Dy 12 Chllenge check

15 RESULTS

16 Effect of TC on SE contmintion of eggs *, week 1 25 WEEKS 40 WEEKS b b b b b b b c *Tretments with different superscripts significntly differed from ech other t P< 0.05

17 Effect of TC on SE contmintion of eggs *, week 2 25 WEEKS 40 WEEKS b b b b c b b c *Tretments with different superscripts significntly differed from ech other t P< 0.05

18 Effect of TC on SE contmintion of eggs *, week 3 25 WEEKS 40 WEEKS b b b b c b b c *Tretments with different superscripts significntly differed from ech other t P< 0.05

19 Effect of TC on SE contmintion of eggs *, week 4 25 WEEKS 40 WEEKS b b b b c b c c *Tretments with different superscripts significntly differed from ech other t P< 0.05

20 Effect of TC on SE contmintion of eggs *, week 5 25 WEEKS 40 WEEKS b b b b c b c c *Tretments with different superscripts significntly differed from ech other t P< 0.05

21 Effect of TC on SE contmintion of eggs *, week 6 25 WEEKS 40 WEEKS b b b c c b c c *Tretments with different superscripts significntly differed from ech other t P< 0.05

22 Effect of TC on SE contmintion of eggs *, week 7 25 WEEKS 40 WEEKS b b b c c b b c *Tretments with different superscripts significntly differed from ech other t P< 0.05

23 Effect of TC on SE contmintion of eggs *, week 8 25 WEEKS 40 WEEKS b b b b b b b c *Tretments with different superscripts significntly differed from ech other t P< 0.05

24 Effect of TC on SE contmintion of eggs (N=2195) cumultive dt 25 WEEKS 40 WEEKS b b b b c b c c No chnge in egg production due to TC supplementtion *Tretments with different superscripts significntly differed from ech other t P< 0.05

25 *p < 0.05 Effect of TC on SE in Chicken Orgns* S E p o s it iv e s m p le s ( % ) LIVER CECUM OVIDUCT CONTROL 1% TC 1.5% TC S E p o s it iv e s m p le s ( % ) LIVER CECUM OVIDUCT CONTROL 1% TC 1.5% TC ORGANS ORGANS

26 Impct of reserch In-feed supplementtion of TC could be used to reduce egg-borne trnsmission of SE nd improve microbiologicl sfety of eggs.

27 Vrunkumr Bhttrm PhD Student Attenution of Vibrio cholere infection using plnt molecules Deprtment of Animl Science University of Connecticut PI: Dr. Kumr Venkitnrynn

28 Vibrio cholere (VC) Custive gent of humn choler Toxin-medited wtery dirrhe Life thretening dehydrtion nd electrolyte imblnce Extreme cses led to kidney filure nd deth SerogroupsO1 nd O139 Globlly - excess of 300,000 cses; 7500 deths

29 Choler: Route of Trnsmission Contminted Soil Feces Food Humn Unsnitized Wter

30 Choler toxin CTB Choler toxin B subunit CTA1 nd CTA2 Choler toxin A subunit

31 Tretment/Control Orl Rehydrtion Therpy Fluids supplemented with electrolytes nd slts Antibiotics First ntibiotic resistnt strin 1970 (Kitok et l., 2011) Multiple drug resistnt Vibrio cholere(ds et l., 2008; Akochere., 2013) Vccine Orl vccine Not fully effective (WHO Guidelines, 2012) Sfe nd Effective lterntive strtegy needed

32 Effect of CR, TH nd EG on choler toxin production Strins of Vibrio cholereused in this study VC 11623; BAA nd BAA 2163

33 Protocol for quntifiction of choler toxin V. cholere with or without PDAS Culture superntnt collected fter 24 hours Dilution Purified choler toxin B subunit Seril dilution Fleming et l., 1988 Choler toxin custom kit(kpl protein detector kit) Std curve, & Quntifiction by colorimetry

34 Effect of CR, TH nd EG on choler toxin production (VC 11623)

35 Effect of CR, TH nd EG on choler toxin production (VC 569b)

36 Effect of CR, TH nd EG on choler toxin production (VC 2163)

37 Summry nd Future Studies PDAs could be potentilly dded in orl rehydrting solution to control V. choler infection in humns Vlidte the in vitro results in mmmlin in vivo model Chrcterize nd delinete the mechnism of ction CR, TH, nd EG

38 Controlling fltoxinsin chicken feed using crvcrol nd trnscinnmldehyde s feed dditives

39 Afltoxins(AF) in feed Fungl toxins in feed ingredients Aspergillus spp.- A. flvus, A. prsiticus Routes of contmintion of grins: Pre-hrvest, post-hrvest, nd trnsporttion Processing of feed ingredients Formulted feed fter processing

40 Why study AF? Afltoxin-contminted feed Consumption by Chickens Afltoxin residues in chicken products Crcinogenic nd heptotoxic effect Impct on Humn helth Economic losses to producers Impct on Chicken performnce Bboset l., 2013; Herzllhet l., 2013

41 Afltoxicosisinchickens Poor body weight gin Acute fltoxicosis: 50%mortlitywithin48hours Chronic fltoxicosis: Decrese egg production Decrese htchbility Liver necrosis Ftty liver syndrome Thrsher, 2012; Hmilton nd Grlich, 1971

42 Mterils nd Methods A. flvus NRRL 3357 or A. prsiticus NRRL 4123 or NRRL 2999 (5 log 10 CFU/g) 200 g chicken feed CR 0%, 0.4%, 0.8%, 1.0% TC 0%, 0.4%, 0.8%, 1.0% Incubted t 25 o C for 3 months 0, 1, 2, 3, 4, 8, 12 weeks Afltoxin concentrtion Frg et l., 1989; Rzzghi-Abyneh et l., 2008

43 Effect of CR on A. flvusfltoxin production in chicken feed *Tretments differed significntly from the control (n = 6) (p<0.05)

44 Effect of CR on A. prsiticus fltoxinproductioninchickenfeed * All tretments differed significntly from the control (n = 6) (p<0.05)

45 Effect of TCon A. flvusfltoxin production inchickenfeed *Tretments differed significntly from the control (n = 6) (p<0.05)

46 Effect of TCon A. prsiticus fltoxin production in chicken feed *Tretments differed significntly from the control (n = 6) (p<0.05)

47 Protective effect of CR nd TC on decresing fltoxin-induced cytotoxicity in heptocytes

48 Heptotoxic effect of fltoxins Afltoxin consumption by chicken Acute Afltoxicosis Chronic Afltoxicosis Liver hemorrhge Ftty liver Liver necrosis Doerret l., 1983

49 Mterils nd Methods Humn Heptocytes (ATCC HepG2) 24 hours incubtion to form monolyer in 96-wells plte 3 ppm or 10 ppm fltoxin Control 0 % CR 0.05% & 0.1% Incubte for 18 hours t 37 o C TC 0.05% & 0.1% Cytotoxicity % using LDH ssy Reddy et l., 2006; Zhou et l., 2006

50 Efficcy of CR in reducing fltoxin-induced cytotoxicity in heptocytes 10 ppm 3 ppm * All tretments differed significntly from the control (p < 0.05)

51 Efficcy of TC in reducing fltoxininduced cytotoxicity in heptocytes 10 ppm 3 ppm 3 * All tretments differed significntly from the control (p < 0.05)

52 Summry CR nd TC reduced fltoxins in chicken feed CR nd TC did not chnge feed composition CR nd TC significntly decresed fltoxin-induced cytotoxicity in heptocytes (p < 0.05)

53 Concluding remrks Determine the stbility of CRndTC in chicken feed during mnufcturing process Field trils in chicken frms

54 THANK YOU! QUESTIONS?