Near-edge X-ray absorption spectra for metallic Cu and Mn.

The measurement of X-ray absorption fine structure of metals- both in the extended region (EXAFS) as well as in the near edge region (XANES)-has been widely discussed (see refs 1-6 for Cu and refs 7-9 for Mn). The recent availability of intense X-ray fluxes from storage rings has usually been exploited for EXAFS leaving the XANES often with poorer resolution than earlier work performed on conventional sources (for example, compare the near edge structure for copper in ref. 1 with refs 3 or 6). In addition, whilst the theory and analysis of EXAFS is relatively well-established2,10, a theory for the strong scattering regime near to the absorption edge has only recently been developed11. We report here the first high resolution XANES spectra for Cu and Mn which were performed at the SRS storage ring at Daresbury. Although both metals have close-packed structures consisting of atoms of similar size their local atomic structure is different in detail. Significant differences are found in their respective XANES reflecting the senstivity of this region of the X-ray absorption fine structure to the local atomic structure. Spectra for the two metals have been analysed using the new multiple scattering formalism. This is a real space calculation and unlike a conventional band structure approach it does not require structural periodicity but works from the local arrangement of atoms.

Changes in Fe site structure from fetal to adult hemoglobin probed by XANES.

Iron X-ray absorption near edge structure (XANES) spectra of human fetal (F) and adult (A) deoxyhemoglobin (deoxyHb) measured at the Frascati synchrotron radiation facility reveal the different geometrical structure of the Fe-porphyrin complexes in the two proteins. By this method, having determined for the first time the variation of atomic positions in fetal and adult hemoglobin in solution (close to the 'in vivo' situation), we give further insight into the structure-function relationship in hemoglobins.

Heterogeneity of the isolated subunits of the fetal and adult human hemoglobin in solution, detected by XANES spectroscopy.

Differences in the local structure of the heme in the isolated alpha-, beta- and gamma-chains of the adult and fetal human hemoglobin are detected by XANES (X-ray absorption near-edge structure) spectroscopy. The ligand bonding angle to the iron ion in the ligated forms and the displacement of the Fe respect to the porphyrin plane in the deoxy forms are found to be different for each chain.

XANES study of iron displacement in the haem of myoglobin.

The XANES (X-ray absorption near edge structure) spectra of deoxy human adult haemoglobin (HbA) and myoglobin (Mb) have been measured at the wiggler beam line of the Frascati synchrotron radiation facility. The XANES are interpreted by the multiple scattering cluster theory. The variations in the XANES between HbA and Mb are assigned to changes in the Fe-porphyrin geometry.

Studies on the replication of a bovine parvovirus.

Optimal replication of a bovine parvovirus type 1 was found to occur when parasynchronous bovine embryonic lung cells were infected during the S phase of the cell cycle, just prior to maximum DNA synthesis. Viral antigen was first detected in the cytoplasm by immunofluorescence at 8 h post-infection, reaching a maximum at this location by 16 h and then disappearing. In the nucleus, antigen was first detected at 12 h, concurrent with early inclusion body formation and first detection of intracellular virus production. Intranuclear antigen then increased rapidly to a maximum at 20 h, as the inclusions progressively matured, large amounts of virus were produced within the cell, with some release to the environment. From 24 h, the nuclear inclusions became increasingly shrunken and basophilic as virus migrated to the cytoplasm and was progressively released to the exterior concurrent with cell degeneration and fragmentation. The majority of virus remained cell associated, even at 28 h post-inoculation. Two morphological types of early and late stage intranuclear inclusions were produced by the virus, these appearing to be a distinct feature of bovine strains. In other aspects, the replication of bovine parvovirus appeared similar to that of other members of the genus.

Properties of an Australian isolate of bovine parvovirus type 1.

An Australian bovine parvovirus isolate (BPV 267) was found to haemagglutinate human, guinea-pig, rat and dog erythrocytes, out of a range of 16 species of erythrocyte tested. The haemagglutinating activity was generally found to be both pH and temperature dependent. The virus was found to replicate best in intestinal epithelium, macrophage and lung cells, out of 9 bovine cell types tested. Highest yields of virus were obtained by the use of selected cell strains at low-passage levels which were maintained near neutral pH under conditions of high rates of cell growth. Studies of the rates of thermal inactivation with time showed the virus to be relatively stable at 37 degrees C, 56 degrees C and 70 degrees C, the incorporation of serum proteins, 1 M MgCl2 and 2 M NaCl in the medium having no influence on stability at 56 degrees C. The virus was resistant to the action of CHCl3, ether and 1% trypsin, and its replication was inhibited by BUDR, this effect being reversed by thymidine. Actinomycin D was found to inhibit virus replication, but only when applied during the first 8 h post-infection. Density gradient studies showed infective virus to have a density of 1.41 g cm-3; haemagglutinating non-infective virus with defective morphology having a density of 1.31 g cm-3. In addition, a proportion of morphologically-complete haemagglutinating, but non-infective virus particles was found at a density of 1.36 g cm-3. The virus proved to have a mean diameter of 22 nm.

An enzyme-linked immunosorbent assay (ELISA) for antibodies to bovine viral diarrhea virus.

A single dilution enzyme-linked immunosorbent assay (ELISA) was developed to detect antibodies to bovine viral diarrhea (BVD) virus in cattle sera. Viral antigen (NADL strain) was grown in a pig kidney cell line (PK15), and after removal of nuclear debris, was purified by ultracentrifugation through a potassium tartrate cushion. Antigen grown in embryonic bovine tracheal epithelial cells was also satisfactory. The test used a high salt buffer to minimize nonspecific reactivity, polyethylene glycol to enhance the reaction, and Protein G as the labelling agent. Comparative testing with the virus neutralization test (VNT) showed the ELISA results to have a high level of correlation with the VNT titers (r = 0.83). In vaccinated animals the ELISA detected antibodies earlier than the VNT. All animals sampled from a BVD-free herd were negative for BVD antibody. The single dilution test showed close agreement (r = 0.84) with ELISA values obtained using a serial dilution technique, and also proved to have a high level of reproducibility. The test procedures were relatively easy to carry out, and were economic in their use of materials.

Survival of Aujeszky's disease virus in frozen pig meat.

The survival of Aujeszky's disease virus was studied in muscle, lymph node and bone marrow frozen at -18 degrees C, following infusion of a large dose of the virus into the hindquarter of a freshly killed pig. Previous attempts to induce an adequate viraemia for such studies, using intranasal and intravenous routes of inoculation of large doses of virus in live pigs, were unsuccessful. In frozen meat and marrow, the virus showed a biphasic inactivation curve with time, similar to that seen with cell-cultured virus. Most virus was rapidly inactivated initially but a small population of more stable virus persisted for a considerable period of time. In contrast, virus in lymph node showed a uniform inactivaton rate, like that of the more stable componet only. Virus was not detectable in any of the tissues after 35 days of storage at -18 degrees C.

Pathological and virological studies of experimental parvoviral enteritis in calves.

Calves held in isolation showed a progressive decline in maternally derived antibody titres to bovine parvovirus but low concentrations of inhibitors resistant to heat and kaolin treatment persisted as the animals matured. These inhibitors had both haemagglutination inhibition and plaque neutralising activity and were considered to be of non-specific origin. Following oral challenge with bovine parvovirus, calves developed mild to moderate diarrhoea, with lymphopenia and viraemia. Sequential virological and immunofluorescent studies showed that the virus initially infected tonsils and intestinal tract, subsequently spreading to systemic lymphoid tissues. Histological and scanning electron microscopic examinations revealed moderate small intestinal villus atrophy and fusion due to crypt damage, together with lymphoid necrosis predominantly associated with the intestinal tract and thymus. Although the disease was not very severe, this may have been because the low parasite burden in the animals reduced mitotic activity in susceptible tissues.

Exacerbation of experimental parvoviral enteritis in calves by coccidia and weaning stress.

The disease induced in calves following oral dosing with bovine parvovirus (BPV) was found to be substantially exacerbated when the animals had subclinical coccidiosis. Low levels of passive serum antibody appeared to have little influence on the response to BPV. Imposition of weaning stress on coccidia-infected calves which had apparently recovered from prior infection with BPV, was found to induce severe diarrhoea with recrudescence of BPV excretion in faeces. This was in marked contrast to the relatively mild diarrhoea which was found following weaning of control animals and animals infected with either agent alone. It was concluded that BPV activity and damage in the intestinal tract was probably enhanced as a consequence of the extra mitotic activity induced in the region by coccidial infection and the local effects of weaning. On the basis of these and previous findings in the field, it is suggested that BPV may play a significant role in the aetiology of post-weaning diarrhoea in calves.

Epidemiological studies of parvovirus infections in calves on endemically infected properties.

Bovine parvovirus serology and virus excretion were monitored in calves located on three endemically infected North Queensland properties. Maternally derived serum antibody to bovine parvovirus was found to have a half-life of 19 days. On all three properties, calves developed intestinal bovine parvovirus infection with seroconversion soon after weaning. This occurred more promptly where the environment was subject to heavier bovine parvovirus contamination due to management practices. The concurrent presence of moderate levels of residual serum antibody had only minor influence on the onset of the infection. On one beef cattle property, onset of intestinal bovine parvovirus infection was associated with an outbreak of post-weaning diarrhoea. Anthelmintic treatment trials indicated that this syndrome was unrelated to helminth burdens, though coccidiosis appeared responsible for occasional subsequent cases of dysentery. It was considered that bovine parvovirus may have significantly contributed to the development of the diarrhoea syndrome, in conjunction with substantial weaning stresses.

An adenovirus infection of the kidney of Franklin's ground squirrels (Spermophilus franklini) in Saskatchewan, Canada.

During routine pathological studies of Franklin's ground squirrels (Spermophilus franklini) collected during a predator control program, basophilic intranuclear inclusions were found in the collecting tubule epithelium of the renal papillae in seven of 13 squirrels. This was associated with marked karyomegaly in affected cells. An inflammatory response was not seen in the adjacent tissues. Electron microscopic examination of affected cells demonstrated that the enlarged nuclei contained numerous virus-like particles. Autoculture and serial passage of renal medullary cells resulted in the isolation of virus particles producing intranuclear inclusions and cytopathic effect. The virus possessed properties typical of adenoviruses, but showed no evidence of hemagglutinating activity with a range of species of erythrocytes tested under several temperature conditions. The isolates were relatively host-cell specific; they failed to grow in hamster and rabbit kidney cell lines and in ground squirrel kidney cortical cells.

Experimentally induced severe canine parvoviral enteritis.

Severe enteritis was produced in recently weaned, 8-week old pups 3 to 9 days after being given parvovirus by mouth. The most severe manifestations of disease were observed 7 days after infection, when one pup died. Viraemia was detected on days 4 and 5 and a severe leucopenia was present 6 to 8 days after infection. Antibody was demonstrated in serum 4 days after infection, high titres being present 3 days later. Sequential pathological studies revealed necrosis of Peyer's patches on day 3. Between days 5 and 7 typical lesions of the disease became widespread with necrosis of tonsil and thymus being prominent. By the fifth day after infection viral inclusion bodies were numerous. Virus isolation from tissues was greatest at this stage and had diminished by the seventh day. Although tissue repair was well advanced by the tenth day thymic necrosis remained prominent and villous atrophy was still present on day 13. Based on these findings a possible pathogenesis is discussed.

Epidemic of blindness in kangaroos--evidence of a viral aetiology.

OBJECTIVE: To determine the cause of an epidemic of blindness in kangaroos. DESIGN AND PROCEDURES: Laboratory examinations were made of eyes and brains of a large number of kangaroos using serological, virological, histopathological, electron microscopical, immunohistochemical methods, and PCR with cDNA sequencing. In addition, potential insect viral vectors identified during the disease outbreak were examined for specific viral genomic sequences. SAMPLE POPULATION: For histopathological analysis, 55 apparently blind and 18 apparently normal wild kangaroos and wallabies were obtained from New South Wales, Victoria, South Australia, and Western Australia. A total of 437 wild kangaroos and wallabies (including 23 animals with apparent blindness) were examined serologically. RESULTS: Orbiviruses of the Wallal and Warrego serogroups were isolated from kangaroos affected with blindness in a major epidemic in south-eastern Australia in 1994 and 1995 and extending to Western Australia in 1995/96. Histopathological examinations showed severe degeneration and inflammation in the eyes, and mild inflammation in the brains. In affected retinas, Wallal virus antigen was detected by immunohistochemical analysis and orbiviruses were seen in electron microscopy. There was serological variation in the newly isolated Wallal virus from archival Wallal virus that had been isolated in northern Australia. There were also variations of up to 20% in genotype sequence from the reference archival virus. Polymerase chain reactions showed that Wallal virus was present during the epidemic in three species of midges, Culicoides austropalpalis, C dycei and C marksi. Wallal virus nucleic acid was also detected by PCR in a paraffin-embedded retina taken from a blind kangaroo in 1975. CONCLUSION: Wallal virus and perhaps also Warrego virus are the cause of the outbreak of blindness in kangaroos. Other viruses may also be involved, but the evidence in this paper indicates a variant of Wallal virus, an orbivirus transmitted by midges, has the strongest aetiological association, and immunohistochemical analysis implicates it as the most damaging factor in the affected eyes.

Prevalence of antibodies to infectious bovine rhinotracheitis, parainfluenza 3, bovine respiratory syncytial, and bovine viral diarrhea viruses in cattle in Saskatchewan and Alberta.

A total of 1745 healthy cattle from 295 farms in Saskatchewan and Alberta was tested by ELISA for antibodies to four viruses. Antibodies to infectious bovine rhinotracheitis (IBR) virus were found in 37.8% of sera (59.5% of properties), to parainfluenza 3 (PI3) virus in 93.9% of sera (99.7% of properties), to bovine respiratory syncytial (BRS) virus in 78.5% of sera (86.6% of properties), and to bovine viral diarrhea (BVD) virus in 40.6% of sera (66.7% of properties)The prevalence of PI3 viral antibodies among Saskatchewan cattle was not affected by district of origin, breed, sex, age, or vaccination practices, though BRS viral antibodies appeared less frequent in young, male, and unvaccinated animals. Antibodies to IBR and BVD viruses were less prevalent in the Prince Albert/Tisdale districts and in young, male, and unvaccinated animals, but were more common in Holstein cattle. Antibodies to IBR virus appeared less frequent in Herefords. Antibodies were more prevalent in cattle which had been vaccinated against IBR, BRS, and BVD virus infections.The relatively small number of cattle sampled from Alberta had a similar prevalence of antibodies to PI3 and BRS viruses to that seen in cattle in Saskatchewan, though IBR and BVD prevalence rates were lower.

Epizootic infection of a minimal disease Swine herd with a herpesvirus.

A totally confined, farrow-to-finish, closed, minimal disease herd of pigs experienced high death and reproductive losses during a disease outbreak lasting five weeks, and during the ensuing three months. The losses were caused by infection of pigs of all ages with a herpesvirus, with devastating effects on the lungs of immature pigs and on the reproductive performance of the sows.

Five field trials on the efficacy of a bovine respiratory syncytial virus vaccine.

Five field trials evaluated whether immunization of beef cattle prior to weaning, at weaning, or immediately upon arrival at the feedlot with a commercial bovine respiratory syncytial virus (BRSV) vaccine would reduce subsequent treatment for respiratory disease.Bovine respiratory syncytial virus vaccination was associated with a significant (p<0.05) reduction in treatment rate in one of three groups of calves immunized prior to weaning (-12%) and in calves immunized upon arrival at the feedlot (-4%).There was no significant (p>0.05) effect of the BRSV vaccine on treatment rate in calves immunized at weaning, in calves immunized upon arrival at the Saskatoon bull test station, or in yearlings immunized upon arrival at the feedlot.Although the trend in these field trials was to a sparing effect of the BRSV vaccine, the small reduction in treatment rate may not justify the cost of the vaccination program.

Viruses and virus-like particles detected during examination of feces from calves and piglets with diarrhea.

A total of 763 fecal or intestinal samples from diarrheic calves and piglets were examined for viral content by immunofluorescence, electron microscopy or cell culture. Routine fluorescent antibody and cultural tests detected rotavirus (n=126), coronavirus (n=80) and bovine viral diarrhea virus (n=13). Electron microscopy detected rotaviruses (n=24) and coronaviruses (n=17) not identified by standard fluorescent antibody tests. Other viruses detected by electron microscopy included Breda virus-like particles (n=49), astroviruses (n=1), caliciviruses (n=1), rhabdoviruses (n=1), parvoviruses (n=2), enteroviruses (n=3), togavirus-like particles (n=2), and "chained" particles (n=5). Mixtures of several of the viruses were detected in a number of fecal samples.The survey emphasized the value of electron microscopy as a broad-spectrum diagnostic tool.