Analysis of Genetic Structure and Genetic Diversity in Japanese Grey-Headed Lapwing Population Using mtDNA.

The grey-headed lapwing (Vanellus cinereus) is a wading species in East Asia. However, examples of regional population dynamics and genetic research are limited. To reconsider the natural history and current status of the grey-headed lapwing in Japan, we analyzed the genetic diversity of the Japanese grey-headed lapwing population. We collected 77 grey-headed lapwing samples from 12 locations across Japan during the breeding season and three individuals during the wintering season and extracted DNA; 496-bp sequences of the ND2, which form part of the mitochondrial DNA, were determined for genetic analysis of the population. Consequently, 10 haplotypes were detected in 80 individuals, and 67 individuals, 84% of the total, shared two haplotypes, namely Vc1 and Vc2. Furthermore, the results showed that the prevalence of Vc1 was higher mainly in northern Japan, while that of Vc2 was higher mainly in southern Japan. Genetic diversity analysis showed that the overall haplotype diversity in Japan was 0.617, which is not particularly low. The sequence of Vc1 was exactly the same as that of grey-headed lapwing in China. Our study revealed the genetic structure of the grey-headed lapwing, suggesting that as the grey-headed lapwing expanded its distribution area into southern Japan, many Vc2-positive individuals migrated southward, resulting in a higher detection rate of Vc2 in southern Japan.

Analysis of Genetic Structure and Genetic Diversity in Japanese Black Kite Population Using mtDNA.

The black kite (Milvus migrans) is widespread in the "Old World" and is a common raptor species in Japan. However, examples of regional population dynamics and genetic research are limited. To reconsider the natural history and current status of the black kite in Japan, we analyzed the genetic diversity of the Japanese black kite population. We collected 59 black kite samples from 22 locations across Japan and extracted DNA; 1585 bp sequences of the cytochrome b and control region, which form part of the mitochondrial DNA, were determined for genetic analysis of the population. Consequently, six haplotypes were detected in 59 individuals, 50 of which had the same major haplotype, namely, Mm1. Moreover, the genetic analysis indicated that the Japanese black kite population would fit the population expansion model. Phylogenetic analysis using sequences obtained in this study or from a DNA database indicated that the Japanese black kite population can be divided into two groups: (1) Mm1 and its close haplotypes and (2) Mm5. The sequence of Mm1 was exactly the same as that of black kite in Pakistan, India, and Korea, suggesting that this haplotype is generally widespread in East Eurasia, and that the ancestral haplotype of the Japanese population likely migrated from continental East Asia and expanded its distribution throughout Japan. In summary, we found that the black kite population in East Eurasia, including Japan, is composed of at least two lineages.

Satellite Tracking of Migration Routes of the Eastern Buzzard (Buteo japonicus) in Japan through Sakhalin.

We satellite-tracked the eastern buzzard (Buteo japonicus) wintering in Japan to delineate both northward and southward migration routes, destinations, and stopover behavior. Twenty-two buzzards were captured and fitted with functional platform transmitter terminals. For these buzzards that departed from the capture sites, we observed a total of 65 northward migrations during 2008-2016 and a total of 55 southward migrations during 2008-2015. In spring, the eastern buzzards migrated eastward along the Seto Inland Sea in the Chugoku region or further inland. In eastern Honshu, they followed two different routes. One was to Hokkaido via the Tsugaru Peninsula from central or northern central Honshu northward along the side of the Sea of Japan in northern Honshu. The other was to Hokkaido via the Shimokita Peninsula, mainly from the Pacific Ocean side of northern Honshu, moving eastward through central Honshu. Of the 17 birds tracked, 10 summered in Sakhalin, three in Hokkaido, three in northern Honshu, and one unknown. In autumn, the buzzards retraced their northward migration routes. Of the 14 birds that were tracked the entire southward migration, 13 (92%) returned to their respective capture sites. One juvenile wintered in an area different from the capture site. Our study contributes to a deeper understanding of the distribution of breeding and wintering grounds and the migration routes of B. japonicus. In addition, the information on migration obtained in this study can contribute toward appropriate environmental impact assessment for wind power facilities in Japan.

Genetic Structure in Japanese and Thai Populations of the Japanese Sparrowhawk Accipiter gularis.

The Japanese sparrowhawk Accipiter gularis is a small raptor that breeds in Northeast Asia. The species consists of the widespread and mostly migratory subspecies A. g. gularis that is common in East Asia, including Japan, and the resident and endangered subspecies A. g. iwasakii which inhabits the Ryukyu and Yaeyama Islands in Okinawa, southern Japan. Given the minimal knowledge about the migration of the species, in this study we sought to compare the genetic variation of the populations breeding in Japan with those migrating through Southeast Asia. We sequenced 761 bp of mitochondrial DNA Control Region from each of 21 A. gularis collected during the breeding season in Japan and from 20 individuals intercepted on migration in Thailand. We detected 26 haplotypes among the 41 individuals which differed significantly between Japan and Thailand. Migrants in Thailand were presumed to have originated from a wide area in Eastern Eurasia. The phylogenetic and network analyses demonstrated that the haplotypes of all A. g. gularis detected in Japan were genetically close. Moreover, the Okinawa haplotypes of A. g. iwasakii were clustered with moderate genetic variation. The information presented here can be used towards implementing future conservation actions.

Genetic Structure and Diversity of Two Populations of the Eastern Buzzard (Buteo japonicus japonicus and B. j. toyoshimai) in Japan.

The nominotypical subspecies of the Eastern buzzard (Buteo japonicus japonicus; BJJ) is a common raptor inhabiting East Asia and Japan. Another subspecies, B. j. toyoshimai (BJT), inhabits only the Bonin Islands of the Ogasawara Islands, where there are only an estimated 85 breeding pairs. Because of this low population size, this subspecies is classified as endangered (class IB) in Japan. The aims of the present study were to examine genetic differences between BJJ and BJT, determine the genetic structure of the Eastern Buzzard, and assess genetic diversity within each subspecies. We sequenced 1526 bp within the control region of the mtDNA of 10 BJJ individuals during the breeding season in four sites; similarly, we sequenced 23 BJJ individuals during winter in three sites. We detected 24 haplotypes among the 33 individuals. In a similar analysis performed with 12 BJT individuals, three haplotypes were detected. The phylogenetic analysis showed that BJJ and BJT have diverged into distinct clades, supporting the genetic differentiation between the subspecies. Network and mismatch distribution analyses indicated that BJJ may have experienced population expansion. In addition, comparisons with other raptors revealed a high degree of genetic diversity in the BJJ population. In contrast, the genetic diversity of the BJT population is lower than that in other raptors. Our results indicated that it is necessary to protect BJT to prevent the reduction in its genetic diversity.

Hindgut Bacterial Flora Analysis in Oriental Honey Buzzard (Pernis ptilorhynchus).

The intestinal microbiome is known to affect host health through various effects on nutrition and immunity. The oriental honey buzzard (OHB) is a raptor that feeds on bees and wasps. Due to its restricted diet, we reasoned that the OHB may have a unique microbiome. The aim of this study was to characterize the structure of the intestinal flora of oriental honey buzzards and to investigate the difference of intestinal bacterial flora between individuals in the wild and those reared in captivity. We investigated the intestinal microbiome of seven wild buzzards (Wild), one zoo-reared (Zoo), and one individual reared in captivity for one month (Rearing). Average operational taxonomic units in Wild and Rearing were 69.4 and 113, respectively. Diversity indices such as ACE, Chao 1, Shannon, and Alpha were significantly lower in the Wild than in the Rearing samples. These results suggest that the variety of Wild microbiome is remarkably low. At the phylum level, the composition of the microbiome was similar in all three groups, with firmicutes and bacteroidetes predominating. The third most abundant bacterium in Wild was Proteobacteria, whereas it was Actinobacteria in Rearing and unclassified bacteria in Zoo. Thus, microbiome composition is affected even with just one month of human rearing.

Migration of Tundra Swans (Cygnus columbianus) Wintering in Japan Using Satellite Tracking: Identification of the Eastern Palearctic Flyway.

Migration through the Eastern Palearctic (EP) flyway by tundra swans (Cygnus columbianus) has not been thoroughly documented. We satellite-tracked the migration of 16 tundra swans that winter in Japan. The objectives of this study were 1) to show the migration pattern of the EP flyway of tundra swans; 2) to compare this pattern with the migration pattern of whooper swans; and 3) to identify stopover sites that are important for these swans' conservation. Tundra swans were captured at Kutcharo Lake, Hokkaido, in 2009-2012 and satellite-tracked. A new method called the "MATCHED (Migratory Analytical Time Change Easy Detection) method" was developed. Based on median, the spring migration began on 18 April and ended on 27 May. Autumn migration began on 9 September and ended on 2 November. The median duration of the spring and autumn migrations were 48 and 50 days, respectively. The mean duration at one stopover site was 5.5 days and 6.8 days for the spring and autumn migrations, respectively. The number of stopover sites was 3.0 and 2.5 for the spring and autumn migrations, respectively. The mean travel distances for the spring and autumn migrations were 6471 and 6331 km, respectively. Seven migration routes passing Sakhalin, the Amur River, and/or Kamchatka were identified. There were 15, 32, and eight wintering, stopover, and breeding sites, respectively. The migration routes and staging areas of tundra swans partially overlap with those of whooper swans, whose migration patterns have been previously documented. The migration patterns of these two swan species that winter in Japan confirm the importance of the Amur River, Udyl' Lake, Shchastya Bay, Aniva Bay, zaliv Chayvo Lake, zal Piltun Lake, zaliv Baykal Lake, Kolyma River, Buyunda River, Sen-kyuyel' Lake, and northern coastal areas of the Sea of Okhotsk.

Spring migration routes of mallards (Anas platyrhynchos) that winter in Japan, determined from satellite telemetry.

Wild birds, in particular waterfowl, are common reservoirs of low pathogenic avian influenza viruses, and infected individuals could spread the viruses during migrations. We used satellite telemetry to track the spring migration of the mallard ducks ( Anas platyrhynchos ) that winter in Japan. We studied their migration routes, distribution of stopover and breeding sites, and timing of migration movements. We tracked 23 mallards from four different wintering sites. Nine of the 23 mallards reached presumable breeding sites, where migration terminated. The migration routes of the birds greatly differed not only among the wintering sites but also within the same wintering site, although the general feature of the routes was shared among birds within the same wintering site. The mallards used several stopover sites, and they typically stayed for a long period (about one to four weeks) at a site between migration intervals of two to three days. Stopover sites were located in northeast Japan, the eastern coastline of South Korea and North Korea, and the interior of Far Eastern Russia. Mallards from three different wintering sites used a stopover area near the middle part of the Ussuri river in Russia. The terminal sites, which were presumably also breeding sites, were distributed widely over northeast Asia and Far Eastern Russia. These results suggest that mallards that winter in Japan originate from breeding areas widely distributed across eastern Asia. Mallards could potentially transmit avian influenza viruses between Japan and a broad region of northeastern Asia.

Centrifugal inputs modulate taste aversion learning associated parabrachial neuronal activities.

Our previous studies have demonstrated that gustatory neurons in the parabrachial nucleus (PBN) show altered responses after the acquisition of conditioned taste aversion (CTA) to NaCl. The present study was conducted 1) to examine centrifugal influences on the altered gustatory activity of CTA-trained rats, and 2) to evaluate the role of amiloride-sensitive (ASN) and -insensitive NaCl (AIN) best units in coding the taste of NaCl. Animals were separated into 2 groups: a CTA group that had acquired taste aversion to 0.1 M NaCl and a control group that underwent pseudoconditioning before the recording experiment. Single-neuron activity, in 2 separate series of experiments, was extracellularly recorded in anesthetized rats. In the stimulation studies, the effects of electrical stimulation of the gustatory cortex (GC) or the central nucleus of amygdala (CeA) were examined on firing of PBN taste units. CeA stimulation produced excitatory effect in significantly more neurons in the CTA group (n = 8) than in the control group (n = 1). Furthermore, ASN-best units in the CTA group showed larger responses to NaCl than similar units in the control group. In the decerebration experiment, there was no statistical difference among the taste responses between the 2 groups in any best-stimulus category. These results suggest that CTA conditioning uses an effective central amygdaloid input to modulate activity of gustatory neurons in the PBN. Data also substantiate that amiloride-sensitive components of NaCl-best neurons play a critical role in the recognition of distinctive taste of NaCl.

Synergistic responses of the chorda tympani to mixtures of umami and sweet substances in rats.

It has been known that umami substances such as monosodium L-glutamate (MSG) and 5'-inosine monophosphate (IMP) elicit a unique taste called 'umami' in humans. One of the characteristics of the umami taste is synergism: the synergistic enhancement of the magnitude of response produced by the addition of 5'-ribonucleotides to MSG. In addition to this well-documented synergism, we report here for the first time on another type of synergism between a glutamate receptor agonist, L-AP4, and sweet substances, by analyzing the chorda tympani responses in rats. The results are as follows: (i) when L-AP4 was mixed with one of the sweet substances, such as sucrose, glucose, fructose and maltose, large synergistic responses were observed. (ii) These synergistic responses, except to L-AP4 + sucrose, were not suppressed by sweet taste suppressants, gurmarin and pronase E. (iii) These synergistic responses were not suppressed by either metabotropic or ionotropic glutamate receptor antagonists. (iv) Fibers that responded well to the binary mixtures of L-AP4 and sweet substances also responded well to NaCl and HCl, but very weakly to sucrose. These findings are different from the characteristics of synergism between glutamate and IMP. The multiple transduction mechanisms for the umami taste in rat taste cells are discussed.

Parabrachial unit activities after the acquisition of conditioned taste aversion to a non-preferred HCl solution in rats.

Abstract In a behavioral experiment, rats reliably acquired a taste aversion to non-preferred 0.01 M HCl that had been previously paired with intraperitoneal injection of 0.15 M LiCl. These rats showed aversions to other acidic solutions such as malic acid and tartaric acid. In a neurophysiological experiment, the neuronal activities of the parabrachial nucleus (PBN) were recorded after the acquisition of conditioned taste aversion (CTA) to 0.01 M HCl in urethane-anesthetized rats. Neuronal responses to the conditioned stimulus (CS) did not change on the whole but decreased in the dorsal region to the brachium conjunctivum. The proportion of HCl-best to NaCl-best units was lower in the CTA group than in controls. The spontaneous firing rate was lower in the CTA group than in controls. Correlation coefficients between the HCl CS and normally preferred tastes (sucrose and NaCl) were more negative and those between HCl and quinine were more positive in the CTA group than in the controls. These results may be explained by the notion that gustatory responses of PBN neurons are concerned with alterations in taste hedonics after the acquisition of conditioned taste aversions.