Ingerslev, Hans-Christian

Jensen, Tim Kåre

Schou, Kirstine Klitgaard

Weiss Nielsen , Martin

Online Research Database In Technology

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Downloaded from orbit.dtu.dk on: Dec 18, 2017Genetic diversity of Treponemes in dairy herds and their surrounding environmentWeiss Nielsen , Martin ; Jensen, Tim Kåre; Ingerslev, Hans-Christian; Schou, Kirstine KlitgaardPublication date:2014Document VersionPublisher's PDF, also known as Version of recordLink back to DTU OrbitCitation (APA):Weiss Nielsen , M., Jensen, T. K., Ingerslev, H-C., & Schou, K. K. (2014). Genetic diversity of Treponemes indairy herds and their surrounding environment. Poster session presented at Gordon Research Conference onSpirochetes, Ventura, CA, United States.Gene$c	  diversity	  of	  Treponemes	  	  in	  dairy	  herds	  and	  their	  surrounding	  environment	  Martin W. Nielsen, Tim K. Jensen, Hans-Christian Ingerslev, Kirstine Klitgaard 	  Na$onal	  Veterinary	  Ins$tute,	  Sec$on	  for	  Bacteriology,	  Pathology	  and	  Parasitology,	  The	  Technical	  University	  of	  Denmark	  	  Introduc$on	  	  Conclusion	  	  The	  causa$ve	  agents	  of	   the	  highly	  contagious	   inﬂammatory	  skin	  disease	  digital	  derma$$s	  (DD),	  which	  has	  reached	  endemic	   levels	   in	  parts	  of	  the	  world,	  have	  been	  iden$ﬁed	  as	  mainly	  governed	  by	  Treponema	  species.	  Of	  the	   20	   diﬀerent	   phylotypes	   iden$ﬁed	   from	   DD	   biopsies,	   Treponema	  phagedenis-­‐like,	  T.	  medium/T.	  vincen3i-­‐like	  and	  T.	  den3cola/T.	  pedis-­‐like	  phylotypes	  appears	  to	  be	  highly	  associated	  with	  the	  disease	  progression	  [1,2].	  	  Speciﬁc	   niches	   of	   infec$on	   reservoirs	   and	   transmission	   routes,	   however	  s$ll	  remains	  to	  be	  clariﬁed	  in	  order	  to	  target	  and	  control	  the	  spread	  of	  the	  infec$on.	  	  To	   address	   the	   issue	   of	   infec$on	   reservoirs	   of	   DD,	   we	   surveyed	   for	  evidence	  of	  treponemes	  in	  the	  content	  of	  the	  bovine	  gastrointes$nal	  tract	  	  (feces),	  in	  DD	  lesions,	  and	  in	  the	  dairy	  farm	  environment	  (slurry).	  Methods	  Figure	  1.	  Healthy	  and	  digital	  derma$$s	  hoofs	  	  Severe	   lesions	   are	   easily	   iden$ﬁed	   (b,c)	   in	   comparison	   to	   a	   healthy	   hoof	   (a).	   Typical	   DD	   lesions	   characterized	   by	   irregular,	   raw	   raised	   areas	   that	   oRen	   develop	   kera$naceous	   hairlike	  projec$ons	  (b,	  c).	  	  ='Sample	  type'!$A$2:$J$21	  0	  10	  20	  30	  40	  50	  60	  70	  80	  90	  100	  Bacteria	  spp	   Treponema	  spp	   Treponema	  rumen	  spp	   T.	  medium-­‐like	   Treponema	  canine	  oral	  taxon	   Treponema	  sp.	  PT1	   Treponema	  sp.	  PT2	  Treponema	  sp.	  PT3	   Treponema	  sp.	  PT4	   T.	  den$cola-­‐like	   Treponema	  sp.	  PT8	   Treponema	  sp.	  9T-­‐42	   Treponema	  sp.	  PN-­‐20	   Treponema	  sp.	  PT15	  Treponema	  KO1_aai43a12	   Treponema	  SP2_g07_2	   Treponema	  pedis	   Treponema	  phagedenis	   Treponema	  refringens	  Percentage	  	  	  	  	  	  	  	  Field	  (feces)	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  Slurry	  samples	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  Feces	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  DD	  lesions	  	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	   	  	  	  Ulcera$ve	  mammary	  derma$$s	  	  	  	  	  	  	  	  	  Field	  (feces)	  	  	  	  	  	  	  	  	  	  	  	  	  	  Slurry	  samples	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  Feces	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  	  DD	  lesions	  	  	  	  	  	  	  	  	  0	  10	  20	  30	  40	  50	  60	  70	  80	  90	  100	  Average	  percentage	  	  ='Sample	  type'!$A$2:$J$21	  References:	  1.Klitgaard	  K,	  Boye	  M,	  Capion	  N,	  Jensen	  TK:	  Evidence	  of	  mul$ple	  Treponema	  phylotypes	  involved	  in	  bovine	  digital	  derma$$s	  as	  shown	  by	  16S	  rRNA	  gene	  analysis	  and	  ﬂuorescence	  in	  situ	  hybridiza$on.	  J	  Clin	  Microbiol	  2008,	  46(9):3012-­‐3020.	  2.Evans	  NJ,	  TimoRe	  D,	  Isherwood	  DR,	  Brown	  JM,	  Williams	  JM,	  Sherlock	  K,	  Lehane	  MJ,	  Murray	  RD,	  Birtles	  RJ,	  Hart	  CA	  et	  al:	  Host	  and	  environmental	  reservoirs	  of	  infec$on	  for	  bovine	  digital	  derma$$s	  treponemes.	  Vet	  Microbiol	  2012,	  156(1-­‐2):102-­‐109.	  ='Sample	  type'!$A$2:$J$21	  Acknowledgements:	  Anastasia	  Isbrand	  is	  acknowledged	  for	  her	  technical	  support	  in	  the	  laboratory.	  Furthermore	  we	  would	  	  like	  to	  thank	  The	  Danish	  Camle	  Federa$on	  for	  ﬁnancial	  support	  of	  the	  project.	  Figure	  2.	  Phylogene$c	  composi$on	  within	  the	  samples	  (a)	  Phylogene$c	  composi$on	  of	  individual	  environmental	  and	  lesion	  samples,	  *	   denotes	   the	   ulcera$ve	   mammary	   derma$$s.	   (b)	   Representa$on	   of	   the	  average	   phylogene$c	   contribu$on	   in	   each	   sample	   category.	  Data	   represent	  34	  samples	  of	  feces	  (ﬁeld,	  slurry	  and	  individual	  cows)	  and	  9	  scrapings	  (8	  DD	  lesions	  and	  1	  ulcera$ve	  mammary	  lesion*)	  Samples	  from	  poten$al	  reservoirs	  were	  collected	  and	  subsequently	  DNA	  was	  puriﬁed	  by	  QIAamp®DNA	  stool	  mini	  kit.	   	  The	  hyper-­‐variable	  regions	  V3	  and	  V4	  of	  16S	  rRNA	  were	  PCR	  ampliﬁed	  as	  described	   in	   [1],	   and	   subsequently	   sequenced	  on	  a	  MiSeq,	   Illumina.	   Sequences	  were	  analyzed	  by	  BION-­‐meta	   (hmps://app.box.com/BION)	  	  We	   surveyed	   for	   the	   presence	   of	   DD	   Treponema	   in	   environmental	  samples	   and	   individual	   scrapings	   from	   (DD-­‐	   and	  mammary)	   lesions.	  We	  were	   not	   able	   to	   iden$fy	   DD	   Treponema	   in	   the	   environmental	   samples	  (ﬁeld,	  slurry	  and	  individual	  feces	  samples).	  	  The	   phylogene$c	   composi$ons	   of	   the	   environmental	   samples	   were	  remarkably	   alike	   and	   signiﬁcantly	   diﬀerent	   from	   the	   DD-­‐associated	  Treponema,	  ﬁgure	  2.	  	  The	  DD	  lesions	  	  and	  the	  ulcera$ve	  mammary	  derma$$s	  sample	  proved	  to	  be	  much	  more	  diverse	  and	  dominated	  by	  species	  previously	  iden$ﬁed	  in	  DD	  biopsies	  [1,2].	  	  The	  ques$on	   remains	   if	   this	   is	   a	   sensi$vity	   issue	  –	  or	   if	   the	   reservoir	   of	  these	   bacteria	   is	   actually	   to	   be	   found	   elsewhere	   in	   the	   dairy	   herd	  environment.	  	  	  *	  (a)	   (b)	   (c)	  (a)	  (b)	  

Genetic diversity of Treponemes in dairy herds and their surrounding environment

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Genetic diversity of Treponemes in dairy herds and their surrounding environment

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