Enzyme-linked immunosorbent assays for detecting antibody to Rickettsia australis in sera of various animal species

New endemic areas of spotted fever-like rickettsial disease have been found in south-eastern Australia (Gippsland, Victoria and Flinders Island, Tasmania). The rickettsia responsible is currently unknown although it may be Rickettsia australis. To investigate serological evidence of rickettsial exposure in various wild animal species, a competitive ELISA was developed which detected antibodies to R. australis. It was based on inhibition of an indirect ELISA detecting antibody to R. australis in guinea pig sera. Pre- and post-infection sera from 2 dogs, 2 rabbits, 5 mice and 6 rats, experimentally infected with R. australis, were tested by competitive ELISA. The results showed that all pre-infection sera were negative and all post-infection sera positive for antibody to R. australis. To test the utility of the competitive ELISA for detecting natural rickettsial infection in non-laboratory animals, 51 dog sera, negative for rickettsial antibody by immunofluorescence (IF) and 20 IF positive dog sera (collected from various locations on the east coast of Australia) were tested. Compared to the IF test the competitive ELISA was 90% sensitive and 96% specific. This new test has potential for detecting antibody to R. australis in the sera of different wild animal species.     

Zinc bioavailability in feed-grade sources of zinc

Chick bioassays were used to assess bioavailability of zinc (Zn) from inorganic Zn sources. A soy isolate-dextrose diet containing 13 mg Zn/kg diet was supplemented with feed-grade sources of ZnSO4.H2O (ZnSO4) or ZnO and fed for 2 wk after a 7-d Zn-depletion protest period. Bioavailability of Zn in ZnO relative to ZnSO4 (set at 100%) was determined by multiple regression slope-ratio methodology, using both growth and tibia Zn accumulation in chicks fed graded levels of ZnO and ZnSO4. Linear responses for gain and tibia Zn occurred at dietary Zn levels (ZnSO4.7H2O) between 13 mg/kg (basal) and 33 mg/kg (gain) or 53 mg/kg (total tibia Zn). Therefore, two bioavailability assays were conducted using supplemental Zn levels of 0, 7.5 and 15 mg/kg from each Zn source. When weight gain was regressed on supplemental Zn intake, bioavailability of Zn in ZnO was only 61.2% (P less than .01) that of ZnSO4. When total tibia Zn was regressed on supplemental Zn intake, bioavailability of Zn compared with ZnSO4 (set at 100.0%) was 44.1% (P less than .001) for ZnO. With chicks fed soy-based diets, bioavailability of Zn from ZnO was less than that of ZnSO4.     

Preliminary evidence of growth factor(s) from chicken thrombocytes--growth effects on chicken embryo fibroblasts culture

The thrombocyte extracts (TE) and thrombocyte-secretion products (TSP) prepared from chicken peripheral blood were examined for the growth activity of chicken embryo fibroblasts (CEF). When the cells were incubated with TE and TSP, the enhancement of cell spreading was observed within one hour after seeding, but the control culture showed little spreading. Three to six days after cell seeding, the increase of the cell densities in the culture supplemented with TE and TSP was noticed in comparison with the culture without thrombocyte materials. The results strongly indicate the possibility for the presence of thrombocyte-derived growth factor(s) to CEF.     

The purification and characterisation of cervine IgM and IgG

A procedure is described for the isolation of immunoglobulin G (IgG) and immunoglobulin M (IgM) from hyperimmune cervine serum. Hybrids of red deer (Cervus elaphus) and wapiti (Cervus canadensis) were immunised with keyhole limpet hemocyanin (KLH). An immunoglobulin-containing fraction was precipitated from the hyperimmune serum using ammonium sulphate. The antigen-specific immunoglobulins were purified by KLH-conjugated sepharose affinity chromatography and further separated into IgM and IgG by gel-filtration chromatography. Purified immunoglobulin was analysed by polyacrylamide gel electrophoresis and isoelectric focusing. The molecular weights and isoelectric points of the composite chains of cervine IgG and IgM are presented.     

Mycoplasmas from donkeys and horses in the Sudan

Seventeen isolates (4.27%) were recovered from 398 samples. Twelve isolates (4%) were obtained from 300 donkey nasal swabs, three (4.3%) and two (6.89%) isolates were recovered from 69 horse nasal swabs and 29 mare uterine washings, respectively. Nine isolates were lost during storage at -20 degrees C and the remaining eight were identified as mycoplasmas and their biological, biochemical and serological reactions were investigated. The isolates could be divided into two groups on the basis of glucose fermentation and arginine hydrolysis. The first group neither fermented glucose nor hydrolysed arginine. Organisms in the second group hydrolysed arginine only.     

The relation between pathogenicity and plaque size of Getah virus Kanagawa strain in suckling mice

The relation among biological properties, particularly pathogenicity for suckling mice, and plaque size was studied in four virus strains: Getah virus strain Kanagawa; two strains obtained by plaque cloning of the Kanagawa strain, Getah Kanagawa SP (G-K-SP) strain which forms small plaques (SP) only and strain G-K-LP which forms large plaques (LP) only; and strain Haruna which forms SP only. There were no marked differences among the four strains in serological properties, growth curves and sensitivity to pH, trypsin and temperature. Strain G-K-LP showed higher pathogenicity for suckling mice than strain G-K-SP. However, the pathogenicity of strain Haruna, which forms SP only, was as high as that of strain G-K-LP. Some of the clones in SP of strain Kanagawa kill all mice in 5 to 6 days after inoculation while the others required 9 to 11 days or longer before causing death. The present study showed that the pathogenicity of Getah viruses shortly after being isolated from the field, such as the Kanagawa strain, is different between large and small plaques, and even among small plaques, at least in suckling mice, and that the pathogenicity has no relation to plaque size.     

Major outer membrane proteins of Brucella spp.: past,present and future

The major outer membrane proteins (OMPs) of Brucella spp. were initially identified in the early 1980s and characterised as potential immunogenic and protective antigens. They were classified according to their apparent molecular mass as 36–38 kDa OMPs or group 2 porin proteins and 31–34 and 25–27 kDa OMPs which belong to the group 3 proteins. The genes encoding the group 2 porin proteins were identified in the late 1980s and consist of two genes, omp2a and omp2b, which are closely linked in the Brucella genome, and which share a great degree of identity (>85%). In the 1990s, two genes were identified coding for the group 3 proteins and were named omp25 and omp31. The predicted amino acid sequences of omp25 and omp31 share 34% identity. The recent release of the genome sequence of B. melitensis 16 M has revealed the presence of five additional gene products homologous to Omp25 and Omp31. The use of recombinant protein technology and monoclonal antibodies (MAbs) has shown that the major OMPs appear to be of little relevance as antigens in smooth (S) B. abortus or B. melitensis infections i.e. low or no protective activity in the mouse model of infection and low or no immunogenicity during host infection. However, group 3 proteins, in particular Omp31, appear as immunodominant antigen in the course of rough (R) B. ovis infection in rams and as important protective antigen in the B. ovis mouse model of infection. The major OMP genes display diversity and specific markers have been identified for Brucella species, biovars, and strains, including the recent marine mammal Brucella isolates for which new species names have been proposed. Recently, Omp25 has been shown to be involved in virulence of B. melitensis, B. abortus and B. ovis. Mutants lacking Omp25 are indeed attenuated in animal models of infection, and moreover provide levels of protection similar or better than currently used attenuated vaccine strain B. melitensis Rev.1. Therefore, these mutant strains appear interesting vaccine candidates for the future. The other group 3 proteins identified in the genome merit also further investigation related to the development of new vaccines.