Characterisation of experimental infections of domestic pigs with genotype 2.1 and 3.3 isolates of classical swine fever virus

Abstract

Characterisation of experimental infections of domestic pigs with genotype 2.1 and 3.3 isolates of classical swine fever virus.

Assessment of the Phenotype and Functionality of Porcine CD8 T Cell Responses following Vaccination with Live Attenuated Classical Swine Fever Virus (CSFV) and Virulent CSFV Challenge

“…Clinical, hematological, and virological methods. Animals were inspected twice daily (a.m. and p.m.), and 9 parameters relevant for indication of CSF (liveliness, body tension, body shape, breathing, walking, skin, eye/conjunctiva, appetite/leftovers at feedings, and defecation) were examined and scored as 0 (normal), 1 (slightly altered), 2 (distinct clinical sign), or 3 (CSF symptoms) (29). A total clinical score for each animal was assigned twice daily, and temperatures were monitored by rectal thermometer readings and recorded once daily.…”

“…For experiment 1, temperature and clinical score monitoring commenced on day Ϫ11 postchallenge and continued until the termination of the experiment (day 5 postchallenge), and for experiment 2, temperature and clinical score were monitored for 7 days before and after each of the CSFV Brescia inoculations. Peripheral blood leukocytes were enumerated every 3 or 7 days during experiment 1 or 2, respectively, using flow cytometry (29). In brief, 50 l EDTA blood was incubated with 5 l of anti-porcine CD45-fluorescein isothiocyanate (FITC) monoclonal antibody (MAb) (0.1 mg/ml, K252-1E4; AbD Serotec, Oxford, United Kingdom) for 10 min at room temperature (RT) in the dark (details of all the antibodies used in this work are reported in Table 2).…”

“…Total RNA was extracted from EDTA blood using the QIAamp viral RNA mini-extraction kit, according to the manufacturer's protocols (Qiagen, Crawley, United Kingdom). CSFV viral copy numbers were assessed by quantitative reverse transcription-PCR (RT-PCR), using the One-Step Superscript III Platinum kit (Invitrogen) (29). Primers CSF100-F and CSF192-R were used at a concentration of 600 nM, and the 6-carboxyfluorescein (FAM)-labeled CSF probe (30) was used at 200 nM.…”

“…Primers CSF100-F and CSF192-R were used at a concentration of 600 nM, and the 6-carboxyfluorescein (FAM)-labeled CSF probe (30) was used at 200 nM. The viral genome copy number was determined by comparison to a 10-fold serial dilution of a standard, which consisted of RNA transcribed from plasmid pCRXLV234-6, containing the 5= untranscribed region (UTR) of CSFV strain Alfort 187 (29).…”

Factors affecting the infectivity of tissues from pigs with classical swine fever: Thermal inactivation rates and oral infectious dose

“…Viral loads and rates of CSFV inactivation were determined in muscle (longissimus dorsi or bicep fermoris), lymph node (mandibular, Ileocecal, retropharyngeal, ventral superficial cervical), fat and serum samples which had been stored at À80 8C after harvest from animals experimentally infected with CSFV CBR/93, UK2000/7.1 or Brescia, for other purposes, as described below or previously (Everett et al, 2010;Graham et al, 2012). Material from pigs exhibiting the acute form of CSF, between 7 and 21 days post infection (dpi), were used for the study.…”