Thermodynamics of transfer of indocarbocyanines from gel to fluid phases of phospholipid bilayers.

Application of the regular solution model to thermal transition data obtained by differential scanning calorimetry has allowed the determination of partition coefficients, Kp, and the thermodynamics of transfer of a series of indocarbocyanine solutes between the gel and fluid phases of phospholipid bilayers. The indocarbocyanines with alkyl chain lengths of 12 to 22 carbons were partitioned between dipalmitoyl- and distearoylphosphatidylcholine phases at the transition temperatures of the gel-liquid-crystal phase transition. The results indicate that as the alkyl chain length of the solute nears that of the acyl chains of the bilayer lipid, the free energy of transfer from gel to fluid is least negative, and the enthalpy of transfer is most positive. There is almost complete entropy-enthalpy compensation in the transfer process. Comparison of the partition coefficients with published values determined by a fluorescence method show good agreement when the differences in temperature of the measurements are accounted for.

Simulation of the 13C nuclear magnetic resonance spectra of trisaccharides using multiple linear regression analysis and neural networks.

Predictive models are developed for the 13C NMR chemical shifts of the carbon atoms comprising the central rings of 46 trisaccharide compounds. Thirty-nine trisaccharides are used as a training set for development of models using regression analysis and computational neural networks, and seven compounds are used as an external prediction set. The descriptors used in the models are developed directly from the molecular structures of the trisaccharides. Three different methods of descriptor selection are compared. The dependence of the models on the geometries of the trisaccharides is explored. The models developed with geometric descriptors are better than those developed without geometric descriptors, although the latter models are still of a comparable quality. Overall, the best model found is a neural network based on descriptors selected by multiple linear regression.

Differentiation of a porcine reproductive and respiratory syndrome virus vaccine strain from North American field strains by restriction fragment length polymorphism analysis of ORF 5.

The suitability of restriction fragment length polymorphism (RFLP) analysis for differentiating a porcine reproductive and respiratory syndrome virus (PRRSV) vaccine strain from other North American field strains was investigated. Open reading frame 5 nucleotide sequence data of the vaccine virus, its parent strain VR-2332, and 22 other strains of PRRSV included in this study indicated that 3 restriction enzyme gel patterns characterize the vaccine virus and the parent strain genotype. The combined 3 RFLP patterns differentiate the vaccine and parent virus from other PRRSV strains. This test will be a valuable tool in epidemiologic studies and will be useful in identifying individual strains in cases of multistrain PRRSV infections.

Identification and clinical assessment of suspected vaccine-related field strains of porcine reproductive and respiratory syndrome virus.

OBJECTIVE: To determine the origin and clinical relevance of selected strains of porcine reproductive and respiratory syndrome (PRRS) virus (PRRSV). ANIMALS: 38 pigs without antibodies for PRRSV. PROCEDURE: A seemingly uncommon restriction endonuclease digestion site in a commercially available vaccine strain of attenuated PRRSV was tested for its stability and prevalence under defined conditions. Selected field strains of PRRSV, with or without the restriction-site marker, were subsequently tested in pigs for virulence and for their ability to replicate competitively in pigs simultaneously given the vaccine. RESULTS: Under experimental conditions, the restriction-site marker was stable during long-term infection of pigs. It was not detected in any of the 25 field strains of PRRSV that were isolated before use of the vaccine or 21 of 25 field strains that were isolated after use of the vaccine but that, on the basis of previous testing, were believed unrelated to the vaccine strain. Conversely, it was detected in 24 of 25 field strains that were isolated after use of the vaccine and that, on the basis of previous testing, were believed to be direct-line descendants of the vaccine strain. Putative vaccine-related strains caused more pronounced pathologic changes than did the vaccine strain alone, and they predominated during replication in pigs also given the vaccine strain. CONCLUSIONS: In some swine herds, the vaccine strain may have persisted and mutated to a less attenuated form. CLINICAL RELEVANCE: The potential for persistence and mutation of specific strains of virus should be an important consideration when designing vaccination programs involving attenuated PRRSV.

Diagnostic implications of concurrent inoculation with attenuated and virulent strains of porcine reproductive and respiratory syndrome virus.

OBJECTIVE: To determine the predominant strain of progeny virus in samples obtained from cell cultures and pigs exposed simultaneously to attenuated and virulent strains of porcine reproductive and respiratory syndrome virus (PRRSV). SAMPLE POPULATION: Cell cultures and twenty 4-week-old pigs. PROCEDURE: Cell cultures and pigs were simultaneously exposed to various relative concentrations of an attenuated, cell-culture-adapted vaccine strain and a virulent field strain of PRRSV. Progeny virus obtained at selected intervals thereafter was tested to determine strain identity by use of restriction fragment length polymorphism (RFLP) analysis. RESULTS: Progeny virus from infected cell cultures comprised the attenuated strain, alone or in combination with the virulent strain, except when cultures had been exposed to a large excess (> 100,000-fold) of the virulent strain. Progeny virus from infected pigs comprised only the virulent strain regardless of the relative concentrations of the 2 strains to which the pigs had been exposed. CONCLUSIONS: During concurrent replication in cell cultures, the attenuated strain quickly predominated. Conversely, during concurrent replication in pigs, the virulent strain quickly predominated. CLINICAL RELEVANCE: It is unlikely that only an attenuated strain of PRRSV would be identified by RFLP testing of samples obtained from pigs concurrently infected with a virulent strain of PRRSV. Nevertheless, the ability of a cell-culture-adapted attenuated strain of PRRSV to predominate during cell culture passage (the first step in the current RFLP testing procedure) indicated that, if possible, samples should be obtained from pigs that do not have a history of direct or indirect exposure to attenuated-virus vaccine.

Emerging infections: a tribute to the one medicine, one health concept.

Events in the last decade have taught us that we are now, more than ever, vulnerable to fatal zoonotic diseases such as those caused by haemorrhagic fever viruses, influenza, rabies and BSE/vCJD. Future research activities should focus on solutions to these problems arising at the interface between animals and humans. A 4-fold classification of emerging zoonoses was proposed: Type 1: from wild animals to humans (Hanta); Type 1 plus: from wild animals to humans with further human-to-human transmission (AIDS); Type 2: from wild animals to domestic animals to humans (Avian flu) and Type 2 plus: from wild animals to domestic animals to humans, with further human-to-human transmission (Severe Acute Respiratory Syndrome, SARS). The resulting holistic approach to emerging infections links microbiology, veterinary medicine, human medicine, ecology, public health and epidemiology. As emerging 'new' respiratory viruses are identified in many wild and domestic animals, issues of interspecies transmission have become of increasing concern. The development of safe and effective human and veterinary vaccines is a priority. For example, the spread of different influenza viruses has stimulated influenza vaccine development, just as the spread of Ebola and Marburg viruses has led to new approaches to filovirus vaccines. Interdisciplinary collaboration has become essential because of the convergence of human disease, animal disease and a common approach to biosecurity. High containment pathogens pose a significant threat to public health systems, as well as a major research challenge, because of limited experience in case management, lack of appropriate resources in affected areas and a limited number of animal research facilities in developed countries. Animal models that mimic certain diseases are key elements for understanding the underlying mechanisms of disease pathogenesis, as well as for the development and efficacy testing of therapeutics and vaccines. An updated veterinary curriculum is essential to empower future graduates to work in an international environment, applying international standards for disease surveillance, veterinary public health, food safety and animal welfare.

Molecular characterization of a caprine adenovirus.

Restriction endonuclease site maps were constructed for the genome of a caprine adenovirus (GAdV), strain NC90-7261, which was isolated in 1990 from a 3-year-old goat with encephalitis. Genomic GAdV DNA was digested with seven restriction endonucleases (RE). Genomic DNA libraries of GAdV were constructed by cloning BamHI and HindIII restriction fragments into a plasmid vector. Using cloned GAdV genomic fragments as probes in Southern blot hybridizations, an RE site map was constructed. The position of several clones was confirmed by limited nucleotide sequencing and the location of several RE sites was determined by single or double RE digestions of cloned fragments. The size of the GAdV genome was determined to be 28.2 kbp. The restriction pattern described in this report is different from that of other adenoviruses. Although the genomic organization of this GAdV is likely to be similar to that of other adenoviruses, the overall level of sequence similarity is low.

Genomic characterization of virulent, attenuated, and revertant passages of a North American porcine reproductive and respiratory syndrome virus strain.

Pigs were exposed to three passages of the NADC-8 strain of porcine reproductive and respiratory syndrome virus (PRRSV) to investigate the relationship between genotypic and phenotypic properties. Differences were found in the virulence of the three passages called virulent, intermediate, and avirulent. Avirulent virus was derived by attenuation of virulent virus in cell culture and intermediate virus was derived by passage of avirulent virus in a pig. Nucleotide sequence differences between virulent and avirulent virus consisted of 50 nucleotide changes and a three-nucleotide deletion, and between avirulent and intermediate virus consisted of 8 nucleotide changes resulting in six amino acid changes. Three of these amino acid changes were direct reversions to virulent virus. Genetic changes, especially those seemingly associated with attenuation followed by some degree of reversion to virulence, in ORF1a, ORF1b, and ORF 6 regions of the genome may be involved in the control of PRRSV replication and virulence.