Masters Project Overview

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Masters Project Overview

• Genetic Stability of a genetically-engineered
  chimeric porcine circovirus (PCV) vaccine, PCV1-2

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Objective

• To determine the in vitro and in vivo genetic
  stability of the chimeric PCV1-2 vaccine

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Hypothesis

• The chimeric PCV1-2 live vaccine is genetically
  stable after serial passages in PK-15 cells and
  in pigs.

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Background of PCV1

• Discovered as a contaminant of porcine kidney
  PK-15 cells
• Determined to be non-pathogenic in swine

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Background of Postweaning multisystemic wasting
syndrome (PMWS)

• PMWS was first reported in weaning piglets in
  Canada in 1991
• Today, it effects every major swine producing
  country in the world
• Case fatalities can reach up to 50

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Background of PCV2

• First isolated from pigs infected with PMWS.
• Now known to be the primary causative agent of
  PMWS.

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PCV1 and PCV2

• Members of the family Circoviridae
• Single stranded, non-enveloped DNA viruses about
  1.76 kb in size
• Sequence analysis shows 75 nucleotide identity
  with similar genomic organization

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PCV1 and PCV2 continued
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Chimeric PCV1-2 vaccine

• Our lab has developed a vaccine against PCV2
• The capsid gene of PCV2 was cloned into the
  genetic backbone of the non-pathogentic PCV1
• The chimeric vaccine is attenuated in pigs and
  produces protective immunity against PCV2
  infection in pigs

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Materials and MethodsStudy 1 In vitro study

• Preparation of the PCV1-2 chimeric DNA clone
• Transfection of PK-15 cells with infectious
  PCV1-2 plasmid clone
• Serial passage of the PCV1-2 vaccine virus in
  PK-15 cells

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Materials and Methods cont

• Genetic characterization of the PCV1-2 vaccine
  virus during serial passage

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Materials and Methods contStudy 2 In vivo study

• Propagation of the PCV1-2 vaccine virus for in
  vivo study
• Serial passaging of PCV1-2 vaccine virus in SPF
  pigs

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Table 1. Serial passages of PCV1-2 vaccine in
pigs Passage 1
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Fig. 1. Serial passages of PCV1-2 virus in pigs
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Materials and Methods cont

• ELISA
• Sequence analysis and comparison

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Progress

• Procedures
• Primers
• Passages