Partial Purification of Thai Isolate of Citrus Huanglongbing (Greening) Bacterium and Antiserum Production for Serological Diagnosis

We aimed to improve the purification of citrus Huanglongbing (greening) bacterium (HB), Candidatus Liberobacter asiaticum and to produce an antiserum against HB. Periwinkle plants Catharantus roseum L. graft-inoculated with HB were used to produce an antiserum. All young leaves of new shoots incubated at 20–25°C and 25–30°C, a few mature leaves incubated at 20–25°C, and all mature leaves incubated first at 25–30°C and later transferred to 20–25°C developed yellowing symptoms and were then used to prepare immunogen. The HB was partially purified from these leaves by an improved method that included a macerating enzyme treatment of the midribs of infected leaves and homogenization of infected phloem sieve tissues. An antiserum raised against partially purified HB reacted clearly at a dilution of 1/16 with HB-infected citrus extract prepared at a concentration of 40 times, but did not react with healthy or tristeza virus-infected citrus extract in microprecipitin tests. Received 23 August 2002/ Accepted in revised form 4 December 2002     

Application of DNA markers for discrimination between Japanese and Australian Wagyu beef

The objective of this study was to discriminate between original Japanese and Australian Wagyu beef, which is sold in the Singapore markets, using six previously developed DNA markers. To effectively evaluate the six markers for breed identification, the probability of identification as Australian Wagyu beef was calculated based on the estimated allele frequencies using 130 Australian Wagyu individuals. The combined use of six markers would allow the discrimination of Australian Wagyu beef with an estimated probability of 0.776. The probability to discriminate Australian Wagyu from Japanese Wagyu beef was sufficiently high. In addition, Australian Wagyu has maternal mitochondrial DNA of Bos indicus cattle with moderate high frequency of 0.377. The DNA marker system could also be used as a deterrent force against false sales, and contribute to the reduction and prevention of incorrect or falsified labeling of beef.     

Neuronal activation modulates enhancer activity of genes for excitatory synaptogenesis through de novo DNA methylation

Post-mitotic neurons do exhibit DNA methylation changes, contrary to the longstanding belief that the epigenetic pattern in terminally differentiated cells is essentially unchanged. While the mechanism and physiological significance of DNA demethylation in neurons have been extensively elucidated, the occurrence of de novo DNA methylation and its impacts have been much less investigated. In the present study, we showed that neuronal activation induces de novo DNA methylation at enhancer regions, which can repress target genes in primary cultured hippocampal neurons. The functional significance of this de novo DNA methylation was underpinned by the demonstration that inhibition of DNA methyltransferase (DNMT) activity decreased neuronal activity-induced excitatory synaptogenesis. Overexpression of WW and C2 domain-containing 1 (Wwc1), a representative target gene of de novo DNA methylation, could phenocopy this DNMT inhibition-induced decrease in synaptogenesis. We found that both DNMT1 and DNMT3a were required for neuronal activity-induced de novo DNA methylation of the Wwc1 enhancer. Taken together, we concluded that neuronal activity-induced de novo DNA methylation that affects gene expression has an impact on neuronal physiology that is comparable to that of DNA demethylation. Since the different requirements of DNMTs for germ cell and embryonic development are known, our findings also have considerable implications for future studies on epigenomics in the field of reproductive biology.     

Efficacy of leflunomide for treatment of refractory inflammatory colorectal polyps in 15 Miniature Dachshunds

Inflammatory colorectal polyp (ICRP), common in miniature dachshunds, presents with hematochezia, tenesmus and mucoid feces. Although an 80% response rate has been reported when treated with prednisolone and cyclosporine, effective treatment is needed for the remaining 20% of ICRP dogs. Leflunomide is an immunosuppressive drug reported as effective in several immune-mediated diseases. In the present study, we retrospectively evaluated the efficacy and adverse effects of leflunomide in 15 ICRP dogs that were refractory to treatment with prednisolone and cyclosporine. Treatment efficacy was assessed by endoscopy, clinical symptoms and rectal palpation. Adverse effects were determined by clinical symptoms and blood testing during follow-up. The leflunomide treatment response rate was 93.3%. The median dosage of leflunomide and the median response time were 3 mg/kg (range: 1.7–4.0 mg/kg) and 35 days (range: 20–119 days), respectively. Adverse effects observed included lethargy (3 dogs), anorexia (1 dog), respiratory symptoms (1 dog), leukocytopenia (2 dogs), thrombocytopenia (1 dog), anemia (1 dog) and liver enzyme elevation (8 dogs). Most of the adverse effects improved with symptomatic treatment and leflunomide discontinuation or dosage reduction. In conclusion, leflunomide treatment is effective in ICRP dogs refractory to treatment with prednisolone and cyclosporine. Because several adverse effects were observed, close monitoring is needed during leflunomide treatment follow-up.     

Decaying toxic wood as sodium supplement for herbivorous mammals in Gabon

African rainforest harbors herbivores at high density. However, because plants and soils typically lack in some essential minerals, rainforest is not always a suitable habitat for herbivores. How they fulfill the mineral requirements is therefore an important question to animal ecology and conservation. Although large marshes, called ‘bais’, are often mentioned as efficient mineral-resource, little information on other sodium resources has still been available. Our laboratory works and field surveys found that a peculiar item, decaying wood stumps of Anthostema aubryanum, played as a major sodium resource for herbivores in Moukalaba-Doudou National Park, Gabon. When A. aubryanum is alive, the sodium content of its bark is low and its latex is toxic. Sodium is accumulated in decaying stumps (mean=1,343 mg/kg dry matter). Eight herbivores visited stumps to ingest the dead wood. Fecal sample analysis revealed that western lowland gorillas, a species most-frequently using the stumps, consumed large amount of the dead wood as regular food. Our findings suggest that decaying A. aubryanum is critical sodium-resources and is a key species for herbivores in our study area. Importance of the A. aubryanum may be particularly large there, because it is a limited sodium-rich material that is available year round. Our study site is known as the site where the densities of several herbivores are among the highest at Central Africa. The relatively high herbivores density in our study site may partly depend on decaying A. aubryanum as sodium resources.     

Complex constitutive nature of Japanese upland rice, Oryza sativa L.

Two rice ecotypes, the so-called lowland and upland populations, which carry different isozyme genotypes mostly at a single locus, are cultivated in Japan. The aim of this study was to examine the origin and the mechanism for keeping these genetic differences. The upland population is cultivated in upland fields and carries a different allele for a particular isozyme gene, Pgd-1, which has never been found in the lowland population. RFLP markers showed a weak trend for genetic differentiation between the two ecotypes. On the other hand, morphological, and physiological traits showed marked differences between the two ecotypes. Furthermore, based on the genotypic difference, two Japonica subgroups are defined in the upland population. Subgroup I is the minor group and carries key lowland characters, including the genotype for PGD. Subgroup II carries different traits and the genotype for PGD of the alternative subgroup. As an allelic difference for Pgd-1 is known to occur between the two ecospecies, Tropical (Tr) and Temperate (Tm) Japonicas, upland cultivars can be classified by diagnostic characters which distinguish a variety into Tr or Tm type. The upland population consists of three types of cultivars, Tr-, Tm- and intermediate-type. In contrast, the lowland population consists of a uniform Tm type Japonicas. As Japanese upland cultivars still have an isozyme allele specific to the Tr type, the upland population has a rather complex constitution which is presumably now being introgressed by lowland genetic material, but still represents a major difference at some genetic levels. Upland rice carries several stress-resistant genes which would be useful for lowland rice breeding. The genetic difference would be efficient for tagging upland specific traits by upland specific genetic markers. This revised version was published online in July 2006 with corrections to the Cover Date.     

Long‐term impact of puerperal metritis on the profiles of peripheral blood leukocytes in peripartum dairy cows

To determine the effects of puerperal metritis on the immune response, changes in the differential peripheral blood leukocyte counts were analyzed during the peripartum period in cows with or without metritis. Multiparous Holstein cows were examined for uterine health disorders and classified into two groups: healthy (n = 11) or metritis (n = 5) cows. The lymphocyte and monocyte counts and the proportion of CD8⁺ lymphocytes were higher in cows with metritis compared to healthy cows. Moreover, the effects of puerperal metritis on the lymphocyte counts and CD4⁺/CD8⁺ ratio persisted weeks after the uterine inflammation had self‐resolved. Taken together, the findings of the present study indicate the possible long‐term alterations of systemic immune responses in cows with puerperal uterine inflammation. © 2015 Japanese Society of Animal Science     

Genetic diversity of African wild rice (Oryza longistaminata Chev. et Roehr) at the edge of its distribution

Ethiopia is at the edge of the distribution for African wild rice, Oryza longistaminata. Here, chloroplast (cp) and simple sequence repeat (SSR) markers were applied to wild rice accessions in Ethiopia to evaluate how they differ from control O. longistaminata, O. barthii and O. glaberrima accessions which originated from African countries. Based on the cp genomes of African wild rice species, maternally inherited cpINDEL markers were developed. The cp indels helped to elucidate 20 plastid types. African cultivated rice shared a particular plastid type with one of annual O. barthii. Parts of northern wild rice in Ethiopia shared Type 6 with control O. longistaminata. The north group shared another type with parts of the south group. The 16 SSR markers amplified a total of 155 alleles in 215 rice accessions, with mean allelic richness of 9.688 per locus, observed heterozygosity of 0.241, expected heterozygosity of 0.724, polymorphic information content of 0.700, and a significant genetic differentiation of 0.215. Both cpINDEL and nuclear markers analyses suggested that wild rice in Ethiopia belongs to O. longistaminata. However, they carry both a unique plastid type and different population structure from control O. longistaminata collected from other areas in Africa. We concluded that the edge of its distribution maintains unique variation. These populations are regarded as valuable genetic resources for future rice breeding.