Porcine circovirus type 2 infection before and during an outbreak of postweaning multisystemic wasting syndrome on a pig farm in the UK.

The presence of porcine circovirus type 2 (PCV-2) and other pathogens before and during an outbreak of postweaning multisystemic wasting syndrome (PWMS) in pigs is evaluated in this study. At the time of the outbreak on a large commercial pig farm in the UK, serum samples and data were collected in two independent on-going research projects, one in weaned pigs and the other in sows. Serum samples of growing pigs and sows were PCV-2-antibody and PCR positive before and during the PMWS outbreak. Upon sequencing, PCV-2 isolates collected before the outbreak were identified as PCV-2a, and isolates collected during the outbreak were identified as PCV-2b, suggesting a shift of PCV-2 genotypes present on the farm. Pigs in the weaner study were from sows originating from different breeders and an association of sow origin and PCV-2 serostatus in offspring was found. Further, pigs had higher odds to be PCV-2 antigen positive if the sow was PCV-2 antibody positive around farrowing, the sow was of higher parity, and were less likely to test antigen positive if the sow was sourced from a particular breeder. The findings of this study highlight the potential role of the immune status of the sow on the occurrence of PMWS.

Clinical and pathological responses of weaned pigs to atmospheric ammonia and dust.

Nine hundred and sixty weaned pigs were exposed for five weeks to controlled concentrations of atmospheric ammonia and dust in a single, multifactorial experiment. The treatments were a mean dust concentration of either 1.2, 2.7, 5.1 or 9.9 mg/m3 (inhalable fraction) and a mean ammonia concentration of either 0.6, 10.0, 18.8 or 37.0 ppm, concentrations representative of commercial conditions. The experiment was carried out over two years and the pigs were used in eight batches, each consisting of five lots of 24 pigs. Each treatment combination was replicated once, and an additional control lot (nominally 0 mg/m3 dust and 0 ppm ammonia) was included in each batch. The dust concentration was the same in the other four lots in each batch in which the four concentrations of ammonia were used; thus, the split-plot design was more sensitive to the effects of ammonia than dust. The groups of pigs were kept separately in five rooms in a purpose-built facility, and the pollutants were injected continuously into the air supply. Ammonia was supplied from a pressurised cylinder, and the endogenous dust in each room was supplemented by an artificial dust manufactured from feed, barley straw and faeces, mixed by weight in the proportions 5:1:4; its ingredients were oven-dried, milled and mixed, and then resuspended in the air supply. The health of the pigs was assessed in terms of general pathology, respiratory tract pathology, and the microbiology of the nasal cavity, trachea and lung. In each batch, postmortem examinations were carried out on 40 pigs after five weeks' exposure to the pollutants and on 30 pigs two weeks later to test for carryover and recovery--a total of 560 pigs. These examinations revealed minimal gross pathology and widespread minor pathological changes of little significance. The pigs' turbinate and lung scores were low and unaffected by exposure to pollutants. All the putative bacterial pathogens, with the exception of toxigenic Pasteurella multocida type D, were isolated from the respiratory tract of the pigs of both ages, but there were no differences between the effects of the different concentrations of pollutants.

Porcine dermatitis and nephropathy syndrome. clinical and pathological features of cases in the United Kingdom (1993-1998).

This paper documents the salient clinical and pathological features of porcine dermatitis and nephropathy syndrome (PDNS) in 96 pigs submitted from 55 units in the UK from 1993 to 1998. This series of cases pre-dated the emergence of post-weaning multisystemic wasting syndrome (PMWS) in the UK. The morbidity during outbreaks was 1% or less. Affected pigs ranged from 14 to 70 kg in weight and most died after a short clinical illness. Fifty-five pigs had multifocal or coalescing erythematous skin lesions, some progressing to dermal necrosis. Biochemistry showed raised serum urea, creatinine and gamma globulin levels accompanied by proteinuria. All cases showed bilateral renal enlargement with petechiae throughout the cortices. Microscopically these renal lesions ranged in chronology from acute necrotizing glomerulitis and vasculitis with multiple hyaline casts in renal tubules to chronic glomerular sclerosis with interstitial inflammation and fibrosis. Haemorrhagic dermatitis when present was associated with necrotizing vasculitis in the dermal vessels. Vasculitis was sometimes detected in other tissues including subcutis, lymph nodes, spleen, liver, joint synovial membrane, gastric and intestinal submucosa or serosa and meninges but its frequency and distribution varied considerably in individual pigs. Immunostaining showed deposits of IgG and IgM in damaged glomeruli, renal casts and skin lesions. The aetiology and pathogenesis of the condition remain unknown but the histopathological and immunological findings suggest a systemic immune-complex disorder resulting in vasculitis with particular predilection for kidney and skin.

Locating interaction sites on proteins: the crystal structure of thermolysin soaked in 2% to 100% isopropanol.

Multiple-solvent crystal structure determination (MSCS) allows the position and orientation of bound solvent fragments to be identified by determining the structure of protein crystals soaked in organic solvents. We have extended this technique by the determination of high-resolution crystal structures of thermolysin (TLN), generated from crystals soaked in 2% to 100% isopropanol. The procedure causes only minor changes to the conformation of the protein, and an increasing number of isopropanol interaction sites could be identified as the solvent concentration is increased. Isopropanol occupies all four of the main subsites in the active site, although this was only observed at very high concentrations of isopropanol for three of the four subsites. Analysis of the isopropanol positions shows little correlation with interaction energy computed using a molecular mechanics force field, but the experimentally determined positions of isopropanol are consistent with the structures of known protein-ligand complexes of TLN.

Ligand-induced conformational change in penicillin acylase.

The enzyme penicillin acylase (penicillin amidohydrolase EC 3.5.1. 11) catalyses the cleavage of the amide bond in the benzylpenicillin (penicillin G) side-chain to produce phenylacetic acid and 6-aminopenicillanic acid (6-APA). The enzyme is of great pharmaceutical importance, as the product 6-APA is the starting point for the synthesis of many semi-synthetic penicillin antibiotics. Studies have shown that the enzyme is specific for hydrolysis of phenylacetamide derivatives, but is more tolerant of features in the rest of the substrate. It is this property that has led to many other applications for the enzyme, and greater knowledge of the enzyme's structure and specificity could facilitate engineering of the enzyme, enhancing its potential for chemical and industrial applications. An extensive study of the binding of a series of phenylacetic acid derivatives has been carried out. A measure of the relative degree of inhibition of the enzyme by each of the compounds has been obtained using a competitive inhibition assay, and the structures of a number of these complexes have been determined by X-ray crystallography. The structures reveal a clear rationale for the observed kinetic results, but show also that some of the ligands cause a conformational change within the binding pocket. This change can generally be understood in terms of the size and orientation of the ligand within the active site.The results reveal that ligand binding in penicillin acylase is facilitated by certain amino acid residues that can adopt two distinct, energetically favourable positions in order to accommodate a variety of compounds within the active site. The structures of these complexes provide evidence for conformational changes in the substrate-binding region that may act as a switch in the mechanism of autocatalytic processing of this enzyme.

Prevention of transmission of sheep pulmonary adenomatosis by embryo transfer.

Two-hundred-and-fifteen embryos recovered from 76 donor ewes from flocks endemically infected with sheep pulmonary adenomatosis (SPA) and mated with uninfected rams were transferred to 131 uninfected recipients under strict sanitary conditions using International Embryo Transfer Society protocols. The recipients and their progeny were kept in a closed, isolated SPA-free flock. Thirty-eight of 51 progeny from SPA-positive donors and 55 of 74 progeny from donors in which no lesions of SPA were detected survived for at least five years after birth. In a similar study 11 embryos from four uninfected donors mated to an SPA-infected ram were transferred to seven recipients, and four of five progeny born to four recipients survived for at least five years. No evidence of SPA was found in the recipients or their progeny by embryo transfer in either study. On the basis of clinical and pathological criteria, it is concluded that embryo transfer can be used to provide an effective barrier against the transmission of SPA from donors from infected flocks, whether or not the parents show clinical signs of the disease.

Neuropathology of scrapie: a study of the distribution patterns of brain lesions in 222 cases of natural scrapie in sheep, 1982-1991.

Detailed neuropathological findings in 222 cases of naturally occurring scrapie from Great Britain are described. The material consisted of formalin-fixed brain from eight breeds of sheep submitted between 1982 and 1991. Paraffin-embedded histological sections were made from several specified brain areas, including the medulla oblongata, cerebellum, pons, mesencephalon, diencephalon, septal area, basal ganglia and frontal cortex. Sections were examined by conventional and polarised light microscopy and the type and distribution of the lesions were recorded. Histologically, the lesions included vacuolation of neuronal perikarya and grey matter neuropil, neuronal degeneration (especially "dark' neurons) and loss, a reactive glial (predominantly astrocytic) response and amyloidosis. Vacuolar lesions were present in the cerebral cortex of 37 per cent of cases, centred around the superior frontal gyrus. Vacuolar lesions were detected in the neocortex for as long as sections have been taken from the superior frontal gyrus and are thus probably not a new feature of the disease. The distribution of vacuolation in the grey matter neuropil could be classified into seven patterns. Data from individual breeds of sheep showed that in some breeds there were significant differences in the age at which animals with different patterns of vacuolation died from scrapie.

A new crystal form of penicillin acylase from Escherichia coli.

A new crystal form of penicillin acylase (penicillin amidohydrolase, E.C. 3.5.1.11) from Escherichia coli W (ATCC 11105) is reported. The crystals were grown using a combination of hanging-drop and streak-seeding methods. The crystals are in the monoclinic space group P2(1) with cell dimensions a = 51.52, b = 131.95, c = 64.43 A, beta = 106.12 degrees. There is one heterodimer in the asymmetric unit (V(m) = 2.45 A(3) Da(-1)) and the solvent content is 49%. Preliminary data have been collected to d(min) = 2.7 A using a MAR Research image plate and a rotating-anode X-ray source. Subsequent experiments show diffraction beyond 1.3 A at a synchrotron radiation source.

Porcine reproductive and respiratory syndrome (PRRS): a review, with emphasis on pathological, virological and diagnostic aspects.

Despite early attempts to control the spread of the disease, porcine reproductive and respiratory syndrome (PRRS) has now become endemic in many countries including Britain. The occurrence of subclinical herd infections, the prolonged circulation of virus within herds and probable aerogenic virus spread all mitigated against the success of control measures. The origin of the disease is unknown but the causative agent has been shown to be an arterivirus with shared features to lactate dehydrogenase virus of mice. There is evidence of extreme genetic and antigenic variability between American and European isolates. PRRS virus has a predilection for alveolar macrophages and does not grow in most cell lines. In infected pigs, viraemia can persist for many weeks in the face of circulating antibodies and little is known about the mechanisms by which immunity to infection develops. A wide spectrum of disease has been reported from the field, accompanied in some cases by heavy economic losses. Reproductive and perinatal losses were most prominent when the disease first appeared. In the endemic phase, PRRS may be more significant as a contributory factor to a post-weaning respiratory syndrome of young pigs of 3-8 weeks. On-farm techniques have been developed to reduce the recycling of PRRS virus from older infected nursery pigs to the younger newly weaned pig. Vaccines are now marketed for the control of PRRS, but are not licensed for use in Britain. Improvements in knowledge of virion composition and antigenic stability and in the nature of the immune response of the pig should result in genetically engineered subunit vaccines becoming available. Diagnosis of PRRS is still difficult as many animals do not show clinical signs and may only be detected by serology and often only when other respiratory diseases are being investigated. Now that the infection is widespread, serological testing must be properly targeted and interpreted to give meaningful results about virus circulation. An increasing arsenal of diagnostic methods are becoming available to detect virus in both fresh and fixed specimens. The pathogenic mechanisms of PRRS remain poorly defined and more work is needed to reveal the nature of the interaction between PRRS virus and other factors in disease.

Porcine reproductive and respiratory syndrome: clinical disease, pathology and immunosuppression.

Porcine reproductive and respiratory syndrome (PRRS) was first known as blue-eared pig disease in the United Kingdom and the causative agent as 'Lelystad virus'. The disease is characterised by very variable clinical signs, including reproductive failure and respiratory disease. The respiratory syndrome is often associated with severe infection with secondary bacterial agents including Pasteurella multocida, Haemophilus parasuis and Streptococcus suis. However, some seropositive herds show no clinical signs of disease. The secondary infections may be facilitated by the destruction of circulating lymphocytes, by the destruction of the mucociliary clearance system and, most importantly, by a large reduction in the numbers of alveolar macrophages. The clinical syndrome observed in a herd may therefore depend in part upon the other diseases present.

Thrombocytopenic purpura in lambs given bovine colostrum.

Newborn lambs which had been given colostrum from a single cow developed thrombocytopenia and seven died. The clinical signs included purpura, haemoglobinuria, melaena, dullness and lethargy. The lambs died at one to two days old. Post mortem examination revealed widespread haemorrhages throughout the body. Lambs given small amounts of colostrum developed a transient thrombocytopenia but did not die.

Congenital infection of pigs with ruminant-type pestiviruses.

Congenital infections of pigs were induced with two ruminant-type pestiviruses isolated from pigs. One of the viruses was bovine viral diarrhoea virus-like and the other border disease virus-like. Both produced symptoms similar to those observed with low virulence strains of classical swine fever virus. A striking effect of persistent virus infection in post-natal life was stunting in viraemic animals. It was also shown that a congenitally infected pig shed virus for 2.5 years and in sufficient quantity to infect other pigs, even by indirect contact. Unlike ruminants, congenitally infected pigs sometimes had persistent viraemia but eventually eliminated the virus. Clearance of virus from the blood was related to the appearance of neutralizing antibodies. However, clearance from the tissues sometimes took as much as 5 months longer than from the blood.

Effect of sequential porcine reproductive and respiratory syndrome and swine influenza on the growth and performance of finishing pigs.

Sows and gilts on a breeding and finishing unit seroconverted to porcine reproductive and respiratory syndrome (PRRS) virus between February and March 1992. The signs of the disease progressed through the breeding herd to the piglets, weaners, growers and finally the finishing pigs. Forty-eight finishing pigs, on a nutritional experiment in which the individual pigs' food intake was recorded daily, showed signs of PRRS and later of swine influenza virus infection. Blood samples collected at slaughter from 47 of the pigs were seropositive to PRRS virus but seronegative to swine influenza virus. However, an immunocytochemical examination of a representative sample of the lungs of the pigs showed that swine influenza virus was present, despite the negative serological results. The infection of the finishing pigs with PRRS virus was associated with recurrent periods of inappetence and a decline in growth rate and performance. The total financial loss from the sequential infection with the two viruses, in a herd without high health status, was estimated at approximately 7 pounds per pig.