Polyclonal activation of B cells occurs in lymphoid organs from porcine reproductive and respiratory syndrome virus (PRRSV)-infected pigs

Porcine reproductive and respiratory syndrome virus (PRRSV) induces a persistent viral infection associated with an inefficient humoral immune response. A study of lymphoid B cells and specific humoral immune response was performed in blood and several lymphoid organs collected from PRRSV experimentally-infected pigs. Groups of specific pathogen-free (SPF) pigs were infected with the LHVA-93-3 isolate of PRRSV, and blood, tonsils, spleen and mediastinal lymph nodes (MLN) were collected at various times postinfection (p.i.) (3-60 days). Lymphoid cells were isolated, immunolabeled for cytofluorometric determination of B cell percentages, used for counting specific anti-PRRSV antibody secreting B cells by an ELISPOT assay, or cultured for metabolic activity. The presence of anti-PRRSV antibodies in the serum of infected pigs was determined using a commercial ELISA assay. Virus detection was performed in all tissues, including lungs, by virus isolation and RT-PCR. The results show that percentages of B cells increased in tonsils as soon as 3 days until 17 days p.i. in PRRSV-infected pigs while they increased in spleen at 3 days p.i. only, due to an increase of larger Ig(high)-producing B cells. Metabolic activity of lymphoid cells from blood and spleen increased at 3 days p.i. only while lymphoid cells from tonsils and MLN transiently decreased at that time and increased thereafter up to 60 days p.i. Anti-PRRSV antibody-secreting B cells occurred in tonsils after 10 days p.i. and strongly increased up to 60 days p.i. However, specific anti-PRRSV-secreting B cells were detected in blood and spleen after 17 days p.i and in MLN only after 45 days p.i. Specific antibodies were detectable in serum at 10 days p.i., reached the maximum level at 45 days and remained high up to 60 days p.i. Infectious virus was detected in lungs and MLN as soon as 3 days p.i., and remained detectable up to 45 days p.i. in tonsils of one pig while viral RNA was detected in most organs up to 60 days p.i. In vitro experiments revealed that inactivated virus induced a stimulation of lymphoid cells isolated from PRRSV-infected pigs while it was cytotoxic for lymphoid cells from control pigs. Taken together, these results indicate that viral infection induced simultaneously a polyclonal activation of B cells, mainly in tonsils, and an exaggerated and prolonged specific humoral immune response due to persistent viral infection in lymphoid organs.     

Pathogenic specialization in Melampsora epitea var. epitea on Salix

Eighteen isolates of the rust fungus Melampsora from different locations in England were tested for pathogenicity to a large range of willow clones ( Salix spp.) in experiments involving inoculation of leaf discs. Seventeen of the isolates were of leaf-infecting M. epitea var. epitea , 16 of which represented forms which alternated on Larix and one of which alternated on Ribes . The remaining isolate was of uncertain identity. Two experiments were carried out. In the first experiment, 24 willow clones consisting of 20 Salix species and interspecific hybrids were inoculated with eight isolates from clones of S. viminalis . In the other experiment, 77 clones from 57 species or hybrids were inoculated with 10 isolates from several Salix spp. The M. epitea var. epitea isolates from S. viminalis clones were all similarly pathogenic, whilst most of the other isolates expressed distinct host specificity. Eight distinct pathotypes were recognized within M. epitea var. epitea . All except one of these alternated on Larix and could be assigned to three formae speciales which had been reported previously in Europe: four pathotypes to f.sp. larici-epitea typica , two to f.sp. larici-retusae , and one to f.sp. larici-daphnoides . The Ribes -alternating pathotype of M. epitea var. epitea infected only S. purpurea . Nine differential willow hosts are proposed as reference clones to distinguish between the larch-alternating pathotypes, the Ribes -alternating rust and the 'stem-infecting' form. The extent of pathogenic variation encountered amongst sexually reproducing rusts suggests that more pathotypes probably exist and will arise in future in response to selection given by long-term clonal willow plantings.     

Contamination of soil with Toxocara eggs in urban (Prague) and rural areas in the Czech Republic

Contamination of soil with feline and canine ascarid eggs in public parks, backyards and sand pits in Prague, Czech Republic was investigated in this work. Soil samples from shelters and rural areas were also collected. The comparison of soil from different areas (urban, rural, backyards and shelters) exhibited significant difference (chi(2)=32.16, d.f.=3 and p<0.0001). The highest rate of contamination (45%) was found in backyards inhabited by feral cats. The eggs of Toxocara spp. were found in 20.4% of parks, 10% of shelters and 5% of rural samples. Mean egg density per sample from Prague parks was 6.2 eggs/100g of soil. In 126 composite samples from children's and pits, the prevalence of Toxocara eggs was 11.90%. The number of eggs in positive samples varied from 2 to 22 (per 100g). A high proportion (46.9%) of eggs was fully embryonated. There was no difference between the sand pits with or without formal exclusion of dogs (chi(2)=0.6, d.f.=1 and p<0.0001).     

Effects of exogenous amylase on the in vitro digestion kinetics of whole-crop maize silages made from flint or dent grain type at different phenological stages grown in tropical condition

The effect of exogenous amylase on the in vitro rumen digestion kinetics of whole-crop maize silage made from dent (RB9004) or flint grain type (RB9308) was evaluated at different phenological stages: soft dough (SOD), early dent (EAD), ½ milkline (½M) and ¾ milkline (¾M). Forage was harvested from 70 to 110 days after sowing. Two rumen-cannulated cows receiving or not exogenous amylase (0.7 g/kg dry matter—DM, provided to achieve 396 kilo Novo units of amylase activity/kg of TMR DM) were used as donor of ruminal fluid. The in vitro gas production kinetics was evaluated according to a dual-pool logistic model. The chemical composition and gas production kinetics were affected by the hybrid and phenological stages. The flint hybrid had lower range for chemical analysis among physiological stages. Harvesting at ½M and ¾M improved DM content, bromatological composition and silage quality parameters compared to dent or flint types. Amylase (i) increased methane (CH₄) production and in vitro dry matter digestibility (IVDMD) in ½M stage, (ii) improved digestion kinetics by reducing lag time and increasing total gas production and fermentation rates of non-fibrous carbohydrates (NFC) and fibrous carbohydrates (FC), and (iii) increased extent and fermentation rate of NFC and increased fermentation rate of FC fraction in whole-crop maize silages produced from dent or flint types in all phenological stages. Harvesting between ½M and ¾M is the best phenological stage to improve chemical composition and silage quality parameters. Exogenous amylase showed improvements on fibre digestion of silages at ½M and ¾M phenological stages in both grain types of corn.     

Preparation and evaluation of antigens used in serological tests for caprine syncytial retrovirus antibody in sheep and goat sera

Methods used to prepare antigens from caprine syncytial retrovirus (CSR) for use in the agarose gel immunodiffusion test (AGID) or an indirect enzyme-linked immunosorbent assay (ELISA) are described. Caprine and ovine sera were tested for antibody to CSR using the AGID test and ELISA incorporating a caprine system (CSR antigen and rabbit anti-goat IgG) or an ovine system (maedi-visna virus antigen and rabbit anti-sheep IgG). Good correlation was achieved in the results of the 3 tests when sera were devoid of antibody or were strongly positive. Variations in the results on weakly positive sera were considered to be more a matter of interpretation than due to basic differences in the reagents employed.