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AimsThe invasion success of alien plants is strongly affected by both biological and human-associated factors. Evaluation of the relative contribution of each factor is important not only for the further understanding of invasion processes but also for the better management of invasion risk, particularly in protected areas of high conservation priority. Here, we quantified the relative importance of species biological traits and association with a human activity, i.e., agriculture, in explaining the invasion success of alien plants across the entire region and in protected areas in Hokkaido, Japan.MethodsAs a quantitative measure of invasion success, the distribution extent of naturalized populations across the entire prefecture and in protected areas was calculated for 63 alien species with equal residence time based on species occurrence records at a spatial resolution of 5-km mesh grid units. For each species, we identified seven biological traits (seed mass, dispersal mode, maximum plant height, capability of vegetative reproduction, flowering start time and period, and life span) and two human-associated factors (introduction purpose and cultivation frequency for agricultural use). Cultivation frequency was determined based on the frequency of seed-sowing in pastures: 1. not sown, 2. accidentally sown as a seed contaminant, and 3. intentionally sown for commercial cultivation. The importance of biological traits and human-associated factors in explaining the distribution extent was determined using an information-theoretic approach.Important FindingsIn explaining the distribution extent across the entire prefecture, species biological traits and human-associated factors showed comparable importance; cultivation frequency exhibited the highest importance value closely followed by seed mass, maximum height, and flowering period. In contrast, when focusing on protected areas, human association was more important than biological traits, as indicated by the greatest importance of cultivationfrequency and much lower values for most biological traits. The results demonstrated that species biological traits and human association almost equally contributed to invasion success across the entire region, while invasions into protected areas were more attributable to human association than to biological traits. We highlight that the control of propagule pressure associated with artificial cultivation may be key to preventing further invasions into protected areas.

Miscanthus ×ogiformis is a hybrid between Miscanthus sacchariflorus and Miscanthus sinensis and its several synonyms indicate the insufficient understanding of the morphological variation of its awn on the spikelet, which is a key characteristic for identifying Miscanthus species. In the present study, all the Miscanthus specimens in 19 herbaria were examined in order to find new localities for Miscanthus ×ogiformis. Thirty-nine Miscanthus ×ogiformis specimens appeared to be naturally occurring hybrids, as their callus hair length, awn length, and callus-to-spikelet ratio differed from those found in the parental species. Therefore, new Miscanthus ×ogiformis individuals with morphological variations are likely to be discovered and these strains might be useful as new genetic resources for this biomass crop.

Increasing incidences of swallowing dysfunction, or dysphagia, a risk factor for aspiration pneumonia, are being reported in aging populations.To investigate the relationship between swallowing function and oral bacteria in independent, community-dwelling elderly.This study recruited 139 community-dwelling individuals aged ≥ 70 years with poor swallowing function. The presence of anaerobic (Prevotella spp. and Fusobacterium spp.) and aerobic bacteria was examined in the participants' oral cavity flora. Swallowing function was evaluated using a 30 mL water swallowing test. Multivariate logistic regression analyses were performed to examine the association between oral bacteria and swallowing function.Swallowing function was assessed as abnormal in 2.9% and as abnormal in 47.5% of the subjects. The colony-forming units (CFUs/ml) of Prevotella spp. were associated with the swallowing dysfunction (odds ratio [OR] 3.45, 95% confidence interval [CI] 1.49-8.11). Further, CFUs/ml of Fusobacterium spp. and aerobes did not correlate with the swallowing dysfunction but were related with the number of teeth (OR 2.71; 95% CI 1.28-5.74, and OR 0.40; 95% CI, 0.18-0.91, respectively) CONCLUSIONS: Swallowing dysfunction in community-dwelling elderly is associated with increased abundance of Prevotella spp., which indirectly may be an increased risk factor for aspiration pneumonia.

Nucleotide sequence variations were investigated with respect to the geographical distribution patterns of Miscanthus sinensis populations that were sampled from 26 Japanese national parks and three populations of the Ryukyu Islands. Twelve homozygous sequences in the nuclear ribosomal DNA internal transcribed spacer region were detected. The populations of M. sinensis in mainland Japan mainly were composed of a monophyletic group with a symapomorphic character, whereas those in the Ryukyu Islands included a polyphyletic group. Only an internal transcribed spacer haplotype with a plesiomorphic character was found in both mainland Japan and the Ryukyu Islands. Thus, no clear geographical isolation was observed in this species. These facts might be caused by the ability of M. sinensis as a pioneer plant to have a high migration potential and high gene flow by outcrossing. On the basis of the results of this study (nuclear ribosomal DNA) and the previous study (chloroplast DNA), a phylogeographical history of M. sinensis was presumed where the ancestral and derived lineages were distributed in the southern and northern regions of Japan, respectively, without strict geographical isolation.

Miscanthus sinensis Andersson is a dominant perennial grass in East Asia, where it is a pioneer species in disturbed habitats. To obtain basic information about the genetic structure of this pioneer species, we investigated chloroplast DNA (cpDNA) variation in M. sinensis within and among native populations across Japan. We detected nine haplotypes in 636 individuals from 30 populations. Two distinct haplotypes had a wide geographical distribution and coexisted within most of the populations. The degree of genetic differentiation among the populations was relatively low. Two putative ancestral lineages were distributed in the Ryukyu Islands. Therefore, it is possible that lineages that originated in subtropical areas of mainland China migrated into the Ryukyu Islands via Taiwan. Another possible migration route is the Korean Peninsula. Two major haplotypes may have migrated from the Eurasian continent to the northern coast in Kyushu via the Korean Peninsula and then expanded throughout Japan. There was no clear genetic boundary among the populations north of Kyushu. The enhanced immigration ability and high rate of gene flow of Miscanthus may have modulated the genetic structure created during previous glacial periods. Alternatively, the shortage of unique haplotypes north of Kyushu may indicate the recent expansion of this species to northern Japan.

Experiments were carried out to investigate whether papillomas could be induced on the teat skin of heifers by intradermal injection with bovine papillomavirus type 9 (BPV-9). Three heifers (#1 and 2, two 0.5-year-old Holsteins; #3, a 1.5-year-old Japanese Black) were injected with BPV-9 and one heifer (#4, a 0.5-year-old Holstein) was mock-infected. Viral DNA load in the inocula was quantified by real-time polymerase chain reaction assay and adjusted to 1.56 × 10 12 copies per injection. Papillomas appeared at the injection sites in the BPV-9-injected heifers #1, 2 and 3 and grew over the 8 (#1 and 2) and 4 (#3) mo observation period, respectively. However, no papillomas were found in the mock-infected heifer #4. The experimentally induced papillomas were excised and examined. Histologically, the lesions were characterized by hyperplasia of the epidermis with hyperkeratosis and marked acanthosis and were morphologically similar to naturally occurring lesions. BPV-9 DNA and bovine papillomavirus capsid antigen were abundant in the lesions. Therefore, we conclude that BPV-9 is an etiological agent causing epithelial papillomas on the teat skin of heifers.

The aim of this study was to evaluate the hypothesis that cerebral hemoglobin (Hb) oxygenation is related to phosphorylation potential during primary and secondary cerebral energy failure in newborn infants who have experienced birth asphyxia. We subjected newborn piglets to severe transient cerebral hypoxic-ischemia followed by resuscitation and examined cerebral energy metabolism by 31P-magnetic resonance spectroscopy and evaluated changes in cerebral Hb oxygen saturation (ScO2) using full-spectrum near-infrared spectroscopy before, during, and up to 54 h after the hypoxic-ischemic insult. ScO2 was significantly decreased during the hypoxic-ischemic insult compared with baseline values. During secondary energy failure, piglets were separated based on the relationship between the ratio of phosphocreatine to inorganic phosphate and ScO2; those with a negative correlation were less injured than those with a positive correlation. These results indicate that changes in ScO2 as measured by near-infrared spectroscopy are related to phosphorylation potential during secondary energy failure in asphyxiated infants.

We evaluated the applicability of the Australian Weed Risk Assessment (AWRA) system in Japan. Native weeds (n = 117) and introduced plants (n = 142), whose weed status was classified by 20 plant experts, were assessed using a slightly modified version of the AWRA system designed to fit Japanese conditions. A receiver operating characteristic (ROC) curve for the system, when classifying two-thirds of the 259 taxa as weeds or non-weeds, was plotted and the area under the ROC curve was calculated. The area was 0.88 and significantly greater than 0.5. Thus, the validity of the system to classify plants was proven. The best cut-off level for the WRA score using Youden’s index was 10. When taxa whose AWRA scores were greater than 10 were regarded as weeds, the sensitivity and specificity were 0.88 and 0.78, respectively. These values were verified with the remaining one-third of the taxa. From these findings, the modified AWRA system was considered to be effective for use in Japan. However, further studies are required to set the best cut-off level in terms of maximising the benefits gained from using the system. A second screening test associated with the cut-off level also needs to be developed.

Noroviruses (NVs) cause many cases of oyster- or clam-associated gastroenteritis in various countries. We collected 191 samples from Japanese oysters intended for raw consumption that had been harvested from the sea in two different areas between December 2001 and February 2002. To detect, quantitate, and phylogenetically analyze the NV genome in purified concentrates from the stomachs and digestive diverticula of these oysters, we amplified the NV capsid gene by reverse transcription-PCR. Phylogenetic analysis was performed by using the neighbor-joining method. We detected the NV genome in 17 of 191 oysters (9%). Phylogenetic analysis indicated genogroup I (Norwalk virus type) in 3 of the 17 oysters and genogroup II (Snow Mountain virus type) in the other 14. Both genogroups showed wide genetic diversity. To quantify the NV capsid gene in these oysters, we performed real-time PCR using genogroup-specific probes. More than 10 2 copies of the NV genome were detected in 11 of 17 oysters. The results suggested that about 10% of Japanese oysters intended for raw consumption harbored NVs, and more than 50% of those oysters in which NVs were detected had a large amount.

A study was conducted to determine the length of exposure to heat necessary to kill water-imbibing weed seeds at 55 and 60°C. The examination covered ten upland weed species: Solanum carolinense L., S. americanum Mill., Abutilon theophrasti Medic., Phytolacca americana L., Amaranthus spinosus L., A. patulus Bertoloni, Persicaria lapathifolia (L.) S. F. Gray, Panicum dichotomiflorum Michx, Echinochloa crus-galli (L.) Beauv. var. crus-galli and Digitaria ciliaris (Retz.) Koeler. All seeds of each species were killed by exposure to heat at 55°C for 72hr and at 60°C for 24hr except for those of Abutilon theophrasti (dormant seed rate 80%) which were killed by exposure for 120 and 30hr at the respective temeratures.These weed species fell into four groups, categorized by a combination of short-duration heat-tolerance (SDHT) and long-duration heat-tolerance (LDHT). SDHT was determined by the viability of weed seeds when treated at 60°C for 3hr, with low and high subcategories of tolerance. LDHT was detemined by the duration required to kill all the seeds at 1% level of significance when treated at 60°C, into three subcategories (low, medium and high tolerance). Persicaria lapathifolia, E. crus-galli, and D. ciliaris were low for both SDHT and LDHT (Group I), and Amaranthus spinosus and Panicum dichotomiflorum were low-SDHT and medium-LDHT (Group II). Solanum species, Phytolacca americana, and Amaranthus patulus were high-SDHT and medium-LDHT (Group III), while Abutilon theophrasti was low-SDHT and high-LDHT (Group IV). The viability of seeds at short-duration heat treatment for Group I, II and IV was low, but the durations required to kill all seeds were different among the groups. Group III consisted of species for which the viability of seeds at short-duration heat treatment was high, and the fatal duration was medium.For Abutilon teophrasti which required the longest duration of heat exposure to kill all seeds at each temperature, 95% confidence intervals of LD90 values were calculated using the probit method, and were 42 to 58 and 10 to 17hr at 55 and 60°C, respectively.

Many phylogeographic studies of various tree species have been conducted to elucidate the locations of refugia and the colonization patterns during the Pleistocene. However, only a few large-scale phylogeographic studies have been conducted on herbaceous plants, especially scarce on herbs that are adapted to disturbance. Artemisia indica is a fast-growing perennial herb found in open habitats. To examine the basic information on the genetic structure of this species, we investigated the chloroplast DNA variation within and among populations across Japan. We detected 26 haplotypes in 604 individuals from 28 Japanese populations. The haplotype A1 had wide geographic distribution, and its close relatives were locally present. Some putative ancestral lineages were found mainly in the Kyushu region. This may be because several lineages migrated from the Eurasian continent to the northern coast in Kyushu via the Korean peninsula during the Pleistocene, and the A1 haplotype expanded northward, whereas others remained in southern areas. Phylogenetic distant haplotypes were present mainly in the Kanto region. Because the geographic distribution pattern of these haplotypes in this region is believed to be unnatural, these haplotypes may be derived from commercial sources for re-vegetation during the last few decades.

A case of intranuclear coccidiosis in an 11-month-old female Japanese Black calf is described. Because the calf exhibited diarrhea and failure to grow and then became moribund, euthanasia was performed. Histologically, the small intestinal villi were highly atrophic, and the epithelial cells frequently had meronts or merozoites within the nuclei. A few intranuclear gametocytes were also seen. The presence of Eimeria alabamensis-specific DNA in the lesions was demonstrated by polymerase chain reaction and nucleotide sequence analyses. Electron microscopy disclosed that most parasites were within nuclear inclusions encircled by the two membranes derived from the nuclear envelope, and that intracytoplasmic meronts and merozoites were in direct contact with the cytosol. Apparently, mature merozoites could be free without destruction of the host cells, and this may be associated with the relatively low pathogenicity of this coccidium.

Norovirus (NV) is a causative agent of acute gastroenteritis in humans, and shellfishes including oysters act as major vehicles of the virus. To investigate the genetic characteristics of NVs, we collected 1,512 oysters for raw consumption between October 2002 and March 2005 from two distinct areas (area A: the Sanriku Sea area; area B: the Setouchi Sea area). We detected the capsid gene and subjected it to phylogenetic analysis. By further quantification of the copy number of the genome by using real-time PCR, the NV capcid gene was detected in approximately 5% of the oysters, and they showed wide diversity. Two percent of the oysters from area B showed relatively large number of NVs, i.e., over 100 copies of capsid gene/oyster, whereas this was not observed in area A. Most of the detected NVs from oysters and humans were genetically related when the capsid region was compared. These results suggested that NVs obtained from humans and those obtained from oysters showed a potential relationship to each other and that some populations of Japanese oysters accumulated a relatively large number of NVs.

The complete nucleotide sequence of a cDNA clone representing the M5 RNA segment of epizootic hemorrhagic disease virus Japan serotype 2 (EHDV-2), Ibaraki virus, was determined. The M5 segment is 1641 base pairs long with the single open reading frame which predicts a polypeptide of 527 amino acids. The comparison of the amino acid sequence of the VP5 with those of EHDV-1, bluetongue virus serotype 10, and African horse sickness virus serotype 4 revealed that the protein shared 67%, 57% and 42% homologies, respectively. In addition, the VP5 protein was expressed in insect cells by recombinant baculovirus, which could be recognized by the mouse anti-EHDV-2 sera at a position of the expected 59 kDa on immunoblot analysis.

Human sapovirus was detected in 4 of 57 clam packages by reverse transcription–PCR and sequence analysis. This represents the first finding of sapovirus contamination in food. Closely matching sequences have been detected in stool specimens from patients with gastroenteritis in Japan, which indicates a possible food-to-human transmission link.