prevalence and molecular characterisation of eimeria species in ethiopian village chickens

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Prevalence and molecular characterisa2on of Eimeria species in Ethiopian village chickens L. Luu 1 , J. Be*ridge 1,2 , R. Christley 1 , K. Melese 3 , D. Blake 4 , T. Dessie 2 , P. Wigley 1 , T. Desta 2,5 , O. Hano*e ,5 , P. Kaiser 6 , Z. Terfa 2,7 , M. Collins 1 and S. Lynch 1,2 1 Ins&tute of Infec&on and Global Health, University of Liverpool, UK; 2 Interna&onal Livestock Research Ins&tute, Addis, Ababa, Ethiopia; 3 Debre Zeit Agricultural Research Centre, Ethiopian Ins&tute for Agriculture Research, Debre Zeit, Ethiopia; 4 Department of Pathology and Infec&ous Diseases, Royal Veterinary College, UK; 5 Centre for Gene&cs and Genomics, School of Biology, University of NoPngham, UK; 6 The Roslin Ins&tute and Royal (Dick) School of Veterinary Science, University of Edinburgh, UK; 7 Management School, University of Liverpool, UK Sampling Strategy Eimeria Protozoan parasites are found worldwide and in all types of chicken producOon system 7 species are known, but are difficult to differenOate based on appearance under the microscope Some species cause severe disease and death, whilst others are only mildly pathogenic, but may reduce growth rates References 1. Permin A, Hansen JW: Epidemiology, diagnosis and disease control of poultry parasites. Food and Agriculture OrganizaOon of the United NaOons: Rome; 1998. 2. Vrba V, Blake DP, Poplstein M: Quan2ta2ve real2me PCR assays for detec2on and quan2fica2on of all seven Eimeria species that infect the chicken. Vet Parasitol 2010, 174:183–190. 3. McDougald LR: Coccidiosis. In Diseases of poultry. 11th ediOon. Edited by Saif YM, Barnes J, Glisson JR, Fadly AM, McDougald LR, Swayne DE. Ames: Iowa State Press ; 2003. Roundworm egg Eimeria oocyst Eimeria oocysts X100 magnifica2on X200 magnifica2on Methods Faecal samples were collected from healthy adult chickens from two Ethiopian woredas in May (n=383) and October 2011 (n=384) Where possible, chickens sampled in May were resampled in October, giving 110 further samples McMasters counts were performed on each sample to quanOfy oocysts [1] A selecOon of samples from those collected in May from each woreda were speciated using real Ome PCR [2] Detec2on of Eimeria oocysts in faeces 427 (56%) of samples were posiOve for oocysts, but more samples were posiOve in October (65%) compared to May (47%) There was moderate clustering of posiOve birds within households, suggesOng common risk factors or exposure There was no difference in overall prevalence between the regions or villages, but different species were more prevalent in each region Birds posiOve in May were just as likely to be posiOve in October as randomlysampled birds, perhaps suggesOng infecOon with different species, or immunologically disOnct strains Region Mixed species infec2on (n = 23) Single species infec2on (n = 13) Species Horro Jarso Fisher’s exact pvalue (n = 22) (n = 25) E. acervulina 9 (40%) 10 (40%) 0.8 12 (52%) 1 (8%) E. bruneP 1 4 (18%) 5 (20%) 0.9 5 (22%) 0 (0%) E. maxima 5 (22%) 15 (60%) 0.02 2 13 (57%) 4 (31%) E. mi&s 0 (0%) 7 (28%) 0.01 2 2 (9%) 2 (15%) E. necatrix 1 4 (18%) 2 (8%) 0.4 4 (17%) 0 (0%) E. praecox 21 (95%) 9 (36%) <0.001 2 17 (74%) 6 (46%) E. tenella 1 3 (12%) 7 (28%) 0.3 3 (13%) 0 (0%) Frequency of PCR detec2on of Eimeria species in chicken faecal samples 1 Considered to be highly pathogenic [3]. 2 Significantly different (p <0.05). Conclusions Eimeria oocysts, including those with the potenOal to be highly pathogenic, can be frequently detected in faecal samples from village chickens in the absence of clinical signs of disease There is variaOon in the prevalence of some species between regions, perhaps suggesOng local risk factors associated with the birds, management or environment Further invesOgaOons are required to assess the role of Eimeria in clinical disease and subclinical effects on producOon in village chickens Research is also required as to whether intervenOons may have any posiOve impacts, or impact negaOvely by disrupOng exisOng endemic stability Weighing faecal samples in the Debre Zeit laboratory in prepara&on for parasite examina&on All seven species were detected, and highly pathogenic species were detected in 20/47 (43%) of samples Mixed infecOons were common (in 36 samples known to come from a single bird), and tended to occur with the most prevalent species in each region

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Poster prepared by L. Luu , J. Bettridge, R. Christley, K. Melese, D. Blake, Tadelle Dessie, P. Wigley, Takele Taye Desta, O. Hanotte, P. Kaiser, Zelalem Gutu Terfa, M. Collins and S. Lynch for the Annual Meeting of the Society of Veterinary Epidemiology and Preventive Medicine, Madrid, Spain, 20-22 March 2013.

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Page 1: Prevalence and molecular characterisation of Eimeria species in Ethiopian village chickens

Prevalence  and  molecular  characterisa2on  of  Eimeria  species  in  Ethiopian  village  chickens  

L.  Luu1  ,  J.  Be*ridge1,2,  R.  Christley1,  K.  Melese3,    D.  Blake4,  T.  Dessie2,  P.  Wigley1  ,  T.  Desta2,5  ,  O.  Hano*e,5,  P.  Kaiser6,  Z.  Terfa2,7,    M.  Collins1  and  S.  Lynch1,2    

 1  Ins&tute  of  Infec&on  and  Global  Health,  University  of  Liverpool,  UK;  2  Interna&onal  Livestock  Research  Ins&tute,  Addis,  Ababa,  Ethiopia;  3  Debre  Zeit  Agricultural  Research  Centre,  Ethiopian  Ins&tute  for  Agriculture  Research,  Debre  Zeit,  Ethiopia;  4  Department  of  Pathology  and  Infec&ous  Diseases,  Royal  Veterinary  College,  UK;  5  Centre  for  Gene&cs  and  Genomics,  School  of  Biology,  University  of  NoPngham,  UK;  6  The  Roslin  Ins&tute  and  Royal  

(Dick)  School  of  Veterinary  Science,  University  of  Edinburgh,  UK;  7  Management  School,  University  of  Liverpool,  UK        

 

Sampling  Strategy    

           

Eimeria    •  Protozoan  parasites  are  found  worldwide  and    in  all  types  of  chicken  producOon  system  •  7  species  are  known,  but  are  difficult  to  differenOate  based    on  appearance  under  the  microscope  •  Some  species  cause  severe  disease  and  death,  whilst  others    are  only  mildly  pathogenic,  but  may  reduce  growth  rates  

References  1.  Permin  A,  Hansen  JW:  Epidemiology,  diagnosis  and  disease  control  of  poultry  parasites.  Food  and  Agriculture  OrganizaOon  of  the  United  NaOons:  Rome;  1998.  2.  Vrba  V,  Blake  DP,  Poplstein  M:  Quan2ta2ve  real-­‐2me  PCR  assays  for  detec2on  and  quan2fica2on  of  all  seven  Eimeria  species  that  infect  the  chicken.  Vet  Parasitol  2010,  174:183–190.  3.  McDougald  LR:  Coccidiosis.  In  Diseases  of  poultry.  11th  ediOon.  Edited  by  Saif  YM,  Barnes  J,  Glisson  JR,  Fadly  AM,  McDougald  LR,  Swayne  DE.  Ames:  Iowa  State  Press  ;  2003.    

Roundworm  egg  

Eimeria  oocyst  

Eimeria  oocysts  

X100  magnifica2on   X200  magnifica2on  

Methods    •  Faecal  samples  were  collected  from  healthy  adult  chickens  from  two  Ethiopian  woredas  in  May  (n=383)  and  October  2011  (n=384)    • Where  possible,  chickens    sampled  in  May  were  resampled  in  October,  giving  110  further  samples  •  McMasters  counts  were  performed  on  each  sample  to  quanOfy  oocysts    [1]  •  A  selecOon  of  samples  from  those  collected  in  May  from  each  woreda  were  speciated  using    real  Ome  PCR  [2]  

Detec2on  of  Eimeria  oocysts  in  faeces                        •  427  (56%)  of  samples  were  posiOve  for  oocysts,  but  more  samples  were  posiOve  in  October  (65%)  compared  to  May  (47%)  •  There  was  moderate  clustering  of  posiOve  birds  within  households,  suggesOng  common  risk  factors  or  exposure  •  There  was  no  difference  in  overall  prevalence  between  the  regions  or  villages,  but  different  species  were  more  prevalent  in  each  region  •  Birds  posiOve  in  May  were  just  as  likely  to  be  posiOve  in  October  as  randomly-­‐sampled  birds,  perhaps  suggesOng  infecOon  with  different  species,  or  immunologically  disOnct  strains  

Region    Mixed  species  infec2on    (n  =  23)  

 Single  species  infec2on    (n  =  13)  

Species   Horro   Jarso   Fisher’s  exact    p-­‐value  

(n  =  22)   (n  =  25)  

E.  acervulina   9  (40%)   10  (40%)   0.8   12  (52%)   1  (8%)  

E.  bruneP  1   4  (18%)   5  (20%)   0.9   5  (22%)   0  (0%)  

E.  maxima   5  (22%)   15  (60%)   0.022   13  (57%)   4  (31%)  

E.  mi&s   0  (0%)   7  (28%)   0.012   2  (9%)   2  (15%)  

E.  necatrix  1   4  (18%)   2  (8%)   0.4   4  (17%)   0  (0%)  

E.  praecox   21  (95%)   9  (36%)   <0.0012   17  (74%)   6  (46%)  

E.  tenella  1   3  (12%)   7  (28%)   0.3   3  (13%)   0  (0%)  

Frequency  of  PCR  detec2on  of  Eimeria  species  in  chicken  faecal  samples  

1Considered  to  be  highly  pathogenic  [3].  2Significantly  different  (p  <0.05).  

Conclusions    

•  Eimeria  oocysts,  including  those  with  the  potenOal  to  be  highly  pathogenic,  can  be  frequently  detected  in  faecal  samples    from  village  chickens  in  the  absence  of  clinical  signs  of  disease  •  There  is  variaOon  in  the  prevalence  of  some  species  between  regions,  perhaps  suggesOng  local  risk  factors  associated  with  the  birds,  management  or  environment  •  Further  invesOgaOons  are  required  to  assess  the  role  of  Eimeria  in  clinical  disease  and  subclinical  effects  on  producOon  in  village  chickens  •  Research  is  also  required  as  to  whether  intervenOons  may  have  any  posiOve  impacts,  or  impact  negaOvely  by  disrupOng  exisOng  endemic  stability  

Weighing  faecal  samples  in  the  Debre  Zeit  laboratory  in  prepara&on  for    parasite  examina&on  

•  All  seven  species  were  detected,  and  highly  pathogenic  species  were  detected  in  20/47  (43%)  of  samples  •  Mixed  infecOons  were  common  (in  36  samples  known  to  come  from  a  single  bird),  and  tended  to  occur  with  the  most  prevalent  species  in  each  region