Vegetative and floral characteristics of interspecific Brassica chimeras produced by in vitro grafting

Summary Interspecific Brassica chimeras (Brassica campestris+B. oleracea) were synthesized according to an in vitro graftculture method, and propagated by tissue culture of axillary buds or chimeric explants. A total of 127 regenerants obtained were investigated. The mericlinal chimera type was more easily produced than the other periclinal and revertant types. No sectorial chimera was produced. The flowering habit and inflorescence type of the chimeras were found to be controlled by the constitution of three tissue layers, but the petal shape and color were controlled only by the two outer tissue layers. Pollen viability of the chimeras were generally lower and more variable in parts than those of the parental types.     

A method for detecting complex vertebral malformation in Holstein calves using polymerase chain reaction-primer introduced restriction analysis.

Complex vertebral malformation (CVM), a hereditary lethal disease in Holstein calves, is characterized by complex anomalies of the vertebral column and limbs in an aborted fetus and in prematurely born, stillborn, and neonatal calves. The mode of inheritance of CVM is autosomal recessive, and CVM is caused by a point mutation from G to T at nucleotide position 559 of the bovine solute carrier family 35 member 3 (SLC35A3) gene. Although an allele-specific polymerase chain reaction (AS-PCR) is a useful method for diagnosis of CVM, the AS-PCR requires selected DNA polymerases and strictly controlled reaction conditions to obtain reliable results. Therefore, an alternative screening method for the CVM gene would be useful. Polymerase chain reaction-primer introduced restriction analysis (PCR-PIRA) is a method that can be used for detecting a single nucleotide mutation in any gene without a restriction site around the mutation site. In this study, primers were designed to introduce PstI or EcoT22 sites into PCR products from the wild-type and CVM alleles, respectively. The wild-type allele, a heterozygote, and a homozygote of the CVM allele could be discriminated by restriction fragment length polymorphism analysis. Specific introduction of restriction sites into PCR products depending on the change in a single nucleotide of template was shown using a variety of DNA polymerases and PCR machines. Therefore, the PCR-PIRA technique using primers designed in this study might provide a more useful method for extensive screening of CVM.     

Phylogenetic analysis and PCR detection of Clostridium chauvoei, Clostridium haemolyticum, Clostridium novyi types A and B, and Clostridium septicum based on the flagellin gene

The flagellin genes (fliC) of Clostridium chauvoei, Clostridium haemolyticum, Clostridium novyi types A and B, and Clostridium septicum were analysed by PCR amplification and DNA sequencing. The five Clostridium species have at least two copies of the flagellin gene (fliC) arranged in tandem on the chromosome. The deduced N- and C-terminal aminoacid sequences of the flagellin proteins (FliCs) of these clostridia are well conserved but their central region aminoacid sequences are not. Phylogenic analysis based on the N-terminal aminoacid sequence of the FliC protein revealed that these clostridia, which belong to Clostridium 16S rDNA phylogenic cluster I (), are more closely related to Bacillus subtilis than to Clostridium difficile, which belongs to the cluster XI. Moreover, a multiplex polymerase reaction (PCR) system based on the fliC sequence was developed to rapidly identify C. chauvoei, C. haemolyticum, C. novyi types A and B, and C. septicum. PCR of each Clostridium amplified a species-specific band. The multiplex PCR system may be useful for rapid identification of pathogenic clostridia.     

ENDOSCOPIC ULTRASONOGRAPHY OF THE PANCREAS IN THE DOG

Endoscopic ultrasonography was done in 12 normal adult dogs to investigate its efficacy in visualization of the pancreas. The endoscopic ultrasonographic device used in the present study had a curved-array ultrasound transducer mounted in front of the objective lens. The tip of the ultrasonic endoscope was inserted into the stomach, and all examinations of the pancreas were performed from within the stomach. Endoscopic ultrasonography provided good images of most parts of the pancreas except for the ends of each lobe. Useful information about the pancreatic parenchyma, including pancreatic lobular structure, pancreatic duct, and vessels of the pancreas was obtained by endoscopic ultrasonography. Blood flow within vessels was detected using color Doppler and pulsed-wave Doppler examination. These results suggest that endoscopic ultrasonography is available as an effective diagnostic modality in small animal practice.     

Distribution of blastodermal cells transferred to chick embryos for chimera production using windowed eggs

1. To improve the production of chimeras, the distribution of donor blastodermal cells after transferring into recipient embryos was examined morphologically.

2. Donor blastodermal cells were distributed near the site of injection in the epiblast and in the subgerminal cavity and yolk. Some filled the hole made by the micropipette and were distributed outside the epiblast. Many were buried in yolk. In some cases, more donor blastodermal cells were located in the yolk than in the subgerminal cavity and some were located 800 μm below the under‐surface of the epiblast.

3. It is recommended that injection should be as shallow as possible to increase the proportion of chimeras produced, and that some means is needed to prevent blastodermal cells from escaping from the hole produced by injection.     

Virological and serological studies of porcine respiratory coronavirus infection on a Japanese farm

We detected transmissible gastroenteritis virus (TGEV) antibodies in pig farms in Tochigi prefecture, although the farms had no past record of TGEV vaccination or TGE. Among the farms, Farm A showed a high antibody incidence. We could not confirm if either TGEV or porcine respiratory coronavirus (PRCV) induced the antibodies, since conventional tests failed to discriminate PRCV from TGEV. Therefore, we conducted virological and serological examinations of this farm for 4 years to establish the etiology - TGEV or PRCV. Although no TGEV was detected, PRCVs were isolated from the nasal samples of pigs. Using a commercial ELISA kit, it was found that the antibodies detected in pigs of all the raising stages and sows were raised against PRCV but not TGEV. The phylogenetic analysis of the nucleotide sequences of the isolates showed that they were closely related to each other, and formed a separate cluster apart from the U.S.A. and European strains. In Cesarean-derived, colostrums-deprived piglets inoculated with a PRCV isolate, no clinical signs were seen, and the viruses were mainly isolated from the nasal samples. Moreover, viral genes were detected from the nasal sample of the contact pig. The result suggested that PRCV infection was located in the nasal cavity of pigs, and horizontal transmission easily occurs. From these results, PRCVs with different origins from the exotic PRCVs might be prevalent in pig farms in Japan.     

Parentage analysis of tea cultivars in Japan based on simple sequence repeat markers

Tea cultivars have been bred by individual selection of landraces and by crossbreeding, but the validation of the parentage is limited. In this study, we performed parentage analysis of 79 tea cultivars in Japan based on SSR markers to confirm or identify the parent-offspring relationships among them. The effectiveness of nine SSR markers for parentage analysis was validated by comparing them to the existing cleaved amplified polymorphic sequence markers. The former markers were detectable more alleles than the latter. Simulation of parentage analysis of the tea cultivars predicted biparental origins for 12 cultivars (‘Houshun’, ‘Mie ryokuhou no. 1’, ‘Surugawase’, ‘Tenmyo’, ‘Yamanoibuki’, ‘Harumidori’, ‘Koushun’, ‘Minekaori’, ‘Okumusashi’, ‘Saemidori’, ‘Sofu’, and ‘Toyoka’), in the first five of which candidate parents of yet-to-be-defined pedigree were newly identified. Comparisons of a total of 41 SSR genotypes confirmed the newly-identified parentages of ‘Asahi’ for ‘Tenmyo’, ‘Rokurou’ for ‘Houshun’, ‘Surugawase’, and ‘Yamanoibuki’, and ‘Yamatomidori’ for ‘Mie ryokuhou no. 1’. The maternity of seven cultivars out of the 12 was also confirmed with chloroplast DNA sequences. Uniparental origins were confirmed for 25 cultivars. This parentage analysis has improved our knowledge of tea pedigrees and will aid in the development of new cultivars.     

Inhibition of growth in juvenile asari clams Ruditapes philippinarum fed Ulva spp. marine silage

Fisheries Science - The suitability of fermented products of Ulva fronds as diets for juvenile asari clams was tested. Ulva fronds were transformed by fermentation into a single-cell product (Ulva...     

Salinization intensifies the effects of elevated temperatures on Channa striata,a common tropical freshwater aquaculture fish in the Mekong Delta,Vietnam

Fisheries Science - Increasing water temperatures and salinity intrusion due to climate change are serious challenges for freshwater aquaculture. In this study, we assessed the combined effects of...