Identification of the porcine cytomegalovirus major capsid protein gene.

A major capsid protein (MCP) gene homologue of porcine cytomegalovirus (PCMV) was identified. Sequence analysis indicated that the PCMV MCP gene is 4,026 nucleotides in length encoding a protein of 1,341 amino acid residues. The predicted molecular weight of the PCMV MCP is 151,456 Da, equivalent to those of other herpesvirus MCP counterparts. Phylogenetic analysis using herpesviral MCP gene sequences confirmed that PCMV is a betaherpesvirus with higher homology with human herpesvirus-6 and -7 than human and mouse cytomegaloviruses. The serum of pig experimentally infected with PCMV did not react with bacterially expressed MCP, suggesting that the PCMV MCP may not be related to the humoral immune response in the course of PCMV infection. Also, we established polymerase chain reaction (PCR) protocols using primers corresponding to MCP gene sequences for detection of PCMV infection. The PCR protocol would be effective for the diagnosis of slow-growing PCMV infection, for which traditional methods involving virus-isolation are not useful.     

Natural Stable Isotopes for Determination of Gastrointestinal Transit Time in Fish

This study evaluated the application of stable isotopes of carbon as an alternative and more accurate method to determine gastrointestinal transit time (GTT) in fish by comparing it to the inert marker method. The stable isotope method detects alterations of the normal carbon flow in a biological system by analyzing naturally occurring isotopes of carbon, contrary to studies based on conventional techniques that apply external markers to the diet to determine GTT through visual observation of the color change in feces. Therefore, 320 pacu, Piaractus mesopotamicus juveniles were reared in 32 tanks under two different temperatures (25 and 29 C). The pacu juveniles received two different diets, one based on ingredients derived from C₃ photosynthetic cycle plants and the other based on C₄ plant ingredients, both containing titanium oxide (TiO₂) as a marker. After 40 d, the isotopic signature of the diets was changed, and the marker was replaced by chromic oxide (Cr₂O₃). In the isotopic technique, the feces were analyzed to determine the exchange in the isotopic ratio of carbon δ¹³C. Both methods found that GTT was faster (nearly 6 h) in fish at 29 C when using the C₄/C₃ feeding strategy and slower in fish at 25 C using the C₃/C₄ strategy (15 h by inert marker and 18 h by the isotopic method). In conclusion, GTT determination in pacu juveniles using the stable isotope technique exhibits the same accuracy obtained with the inert marker method at temperatures suitable (nearly 29 C) for the metabolism of these animals.     

Analysis of porcine cytomegalovirus DNA polymerase by consensus primer PCR

We used a consensus primer PCR method to amplify a region of herpesviral DNA-directed DNA polymerase gene using degenerate primers for initial characterization of the porcine cytomegalovirus (PCMV) genome. The sequence of the PCR product from PCMV DNA template and its alignment with other herpesvirus DNA polymerase counterparts showed that both conserved amino acid residues and conservative amino acid substitutions are in parallel. Phylogenetic analysis revealed that PCMV should be included in the clade comprising human herpesvirus 6 and 7, rather than human and mouse cytomegaloviruses, in Betaherpesvirus subfamily.     

Determination of organic acids by high-performance liquid chromatography with electrochemical detection during wine brewing

Voltammetric determination of acids by means of the electrochemical reduction of quinone was applied to high-performance liquid chromatography (HPLC) with electrochemical detection (ED) for determining organic acids in fruit wines. A two-channel HPLC-ED system was fabricated by use of an ion-exclusion column and an electrochemical detector with a glassy carbon working electrode. Aqueous solution of 0.1 mM HClO(4) and ethanol containing 2-methyl-1,4-naphthoquinone served as a mobile phase and reagent solution, respectively. Determination of acetic, citric, lactic, malic, succinic, and tartaric acids was made by measuring the peak areas of the flow signals due to the reduction current of quinone caused by the eluted acids. The peak area was found to be linearly related to the acid amount ranging from 0.1 to 40 nmol per 20 microL injection. The present method was characterized by reproducibility with the simple and rapid procedure without derivatization of analytes. The method was shown as an effective means for following acid contents in fruit juices during fermentation with wine yeast.     

Antigenic differences between H5N1 human influenza viruses isolated in 1997 and 2003

To assess whether the antigenic properties of H5 hemagglutinin (HA) change over time due to antigenic drift, we produced a panel of monoclonal antibodies (mAbs) against the HA of the index H5N1 human influenza A virus, A/Hong Kong/156/97. By immunizing mice with a plasmid expressing this HA and boosting the initial immunization with cell lysates transfected with the plasmid, a total of six hybridomas producing HA-specific mAbs were established: four to the HA1 subunit with hemadsorption-inhibiting activity and two to the HA2 subunit. None of the mAbs to HA1 could bind to the HA of a recent human isolate, A/Hong Kong/213/2003, indicating that there are substantial antigenic differences between the H5N1 human influenza virus isolated in 1997 and that isolated in 2003.     

Molecular and antigenic similarities of the fimbrial major components between Porphyromonas gulae and P. gingivalis

Porphyromonas gulae is black-pigmented anaerobic bacteria associated with canine periodontitis. There is little information available about the specific identify and relative occurrence of pigmented anaerobes in companion animals. Our aim was to clarify the factor involved in the adherence and colonization of the organism in the oral cavity. Fimbrial protein was purified from P. gulae ATCC 51700. The molecular mass of this protein was approximately 41kDa as estimated by SDS-PAGE. An antibody against 41-kDa fimbrial protein from P. gingivalis ATCC 33277 reacted with fimbrillin of P. gulae ATCC 51700. Immunogold electron microscopy revealed that the anti-41kDa fimbrial serum bound to fimbria on the cell surface of P. gulae ATCC 51700. Thus, fimbrial protein of P. gulae ATCC 51700 had the same size and antigenicity as 41-kDa fimbriae of P. gingivalis ATCC 33277. The nucleotide sequence of the fimA gene from P. gulae ATCC 51700 showed 94% homology with that of P. gingivalis ATCC 33277. Moreover, the deduced amino acid sequences have 96.8% identity. P. gulae has adherent ability to gingival epithelial cells. The properties of P. gulae fimbriae are similar to those of P. gingivalis fimbriae. We suggest that the surface structure of P. gulae may play a role in the colonization of this organism in periodontal pockets in companion animals.     

Digestible protein requirements and muscle growth in juvenile tambaqui (Colossoma macropomum)

Several investigations have been carried out to improve the productivity of tambaqui, an economically important fish species in Brazil and other Latin American countries. This study determined the digestible protein (DP) requirements in juvenile tambaqui by assessing their productive performance and nutritional efficiency. It also evaluated the effects of different dietary DP levels on the morphology and cellularity of skeletal fast muscle fibres. The 1750 tambaqui tested (6.53 ± 0.43 g body weight, 7.58 ± 0.18 cm length) were randomly distributed into 35 tanks. Fish were fed one of the seven isocaloric diets, which contained 140, 170, 200, 230, 260, 290 or 320 g/kg DP. The DP requirement, calculated by segmented (broken line) regression of weight gain data, was 290 g/kg. An increase in diet DP to 290 g/kg significantly improved final weight, weight gain, feed intake, specific growth rate and crude protein gain, and changed fibre diameter in deep muscle. Muscle fibres were randomly distributed into a mosaic pattern, characterized by fibres with different diameters. Treatments with 290 and 320 g/kg DP increased the frequency of large‐diameter fibres (>50 µm), indicating hypertrophic growth of skeletal muscle during the juvenile phase, which occurred in conjunction with hyperplasia.     

Effect of dietary carbohydrate to lipid ratios on growth,digestive enzyme and blood metabolites of juvenile Brazilian sardines,Sardinella brasiliensis (Steindachner, 1879)

The limited availability of live bait for capturing skipjack tuna, Katsuwonus pelamis, is a bottleneck to increasing tuna production in many parts of the world. Therefore, a nutrition trial was performed to contribute to the production of the Brazilian sardine, Sardinella brasiliensis, for use as live bait. This study determined the best dietary carbohydrate to lipid ratio (CHO:L) for juvenile Brazilian sardines based on growth performance, feed utilisation, body composition, blood metabolites and digestive enzyme activity. Six isoenergetic and isonitrogenous diets were formulated with increased CHO:L ratios (2.05, 3.41, 4.15, 5.11, 5.80 and 6.72). Each diet was randomly assigned to triplicate groups of 100 fish with mean initial body weight of 2.97 ± 0.51 g, which were fed four times a day to apparent satiation. Survival was not affected by differences in diet, however, a low CHO:L ratio stimulated growth. Juveniles fed with a rich‐carbohydrate diet inhibit feed intake and protein intake. Body lipid increased as dietary lipid increased and was inversely correlated to body moisture. The diets did not affect the juvenile's blood metabolites. Alkaline and acid protease activities were not significantly different, but lipase and amylase responded positively to the dietary lipids and carbohydrates. Using segmented regression, the optimum CHO:L ratio for maximum weight gain of juvenile Brazilian sardines was estimated to be 3.41, which contain approximately 300 g kg^?1 carbohydrate and 88 g kg^?1 lipid.