Disseminated visceral coccidiosis in a wild white-naped crane (Grus vipio)

Disseminated visceral coccidiosis (DVC) was unexpectedly recognized in a wild white-naped crane (Grus vipio) killed by phosphamidon insecticide. On gross pathologic examination, widely disseminated white nodules were found on the serosa of the pro-ventriculus, gizzard, and intestine, as well as on the surface and in the parenchyma of liver, spleen, and cardiac muscle. Microscopically, asexual stages of a coccidia were observed in some nodules. However, the species of coccidia could not be determined because no oocysts were found on fecal examination. This is believed to be the first reported case of DVC in a wild white-naped crane infected with Eimeria spp.     

Accurate analysis of prevalence of coccidiosis in individually identified wild cranes in inhabiting and migrating populations in Japan

Eimeria gruis and E. reichenowi cause coccidiosis, a major parasitic disease of cranes. By non-invasive molecular approaches, we investigated the prevalence and genetic characterization of pathogens in two Japanese crane habitats; one is Hokkaido inhabited by the endangered red-crowned crane, and the other is Izumi in Kyushu where populations that consist mainly of vulnerable hooded and white-naped cranes migrate in winter. The non-invasively collected faecal samples from each wintering population were first subjected to host genomic DNA-targeted analyses to determine the sample origin and avoid sample redundancy. Extremely high prevalence was observed in the Izumi populations (> 90%) compared with the Hokkaido population (18-30%) by examining 470 specimens by microscopy and PCR-based capillary electrophoresis (PCR-CE), using genetic markers in the second internal transcribed spacer (ITS2). Correspondence analysis of PCR-CE data revealed differences in community composition of coccidia between hooded and white-naped cranes. 18S rRNA and ITS2 sequences were determined from single oocysts excreted by red-crowned and hooded cranes. Phylogenetic analysis of 18S rRNA suggested that E. reichenowi was polyphyletic while E. gruis was monophyletic. Together with PCR-CE data, these results indicate different host specificity among the E. reichenowi type. Our data suggest that E. reichenowi comprises multiple species.     

Parasites of the laboratory woodchuck (Marmota monax)

The parasite burden of captive and colony-born woodchucks maintained in laboratory animal facilities was determined and compared to woodchucks recently trapped. Microfilariae of Ackertia marmotae in blood or dermis were most commonly observed in trapped woodchucks. There was a linear decrease in woodchucks positive for microfilariae after introduction in the laboratory colony, but even after 39 months, 53% were positive. Microfilaremia or microfilariderma were never observed in colony born woodchucks. Eggs or oocysts of Obeliscoides cuniculi, Citellina triradiata and Eimeria sp. were found by fecal flotation. The O. cuniculi population declined exponentially in captive woodchucks following colony entry, and this nematode was never found in colony-born woodchucks. However, low rates of infection with Citellina and Eimeria sp. were found in the colony-born group. Ixodes cookei, Oropsylla sp., and Androlaelaps sp. ectoparasites were observed commonly in recently trapped woodchucks, but were not present in any woodchucks maintained in the colony. Clinical disease associated with parasitic infection was not observed.     

Changes in parasite transmission stage excretion after pheasant release

The production of parasite transmission stages was investigated in the faeces of 77 farm-bred ring-necked pheasants (Phasianus colchicus). Coccidian oocysts (Eimeria sp.), and nematode eggs (Heterakis sp., and Capillaria-like eggs) were recovered before and after release but all birds were treated prior to release. Treatment with fenbendazole significantly reduced the abundance of transmission-stage excretion for all parasites, and reduced the prevalence in the case of Eimeria sp. and Heterakis sp. Nonetheless, a significant increase in the excretion abundance for all parasites and in the prevalence of Eimeria sp. and Heterakis sp. was found after release. Eggs of Ascaridia sp. were found only after releasing, suggesting infection ocurred in the wild. A negative relationship was found between the pheasant body condition and Heterakis excretion abundance and a higher abundance of Capillaria sp. eggs in female birds. No significant relationship was found between parasite excretion abundance and pheasant survival. Despite this, results suggest that an increase in the excretion of parasite transmission stages follows the release of captive pheasants into the wild. This can in part explain restocking failures, but also means that autochtonous free-living birds may become exposed to new and potentially harmful pathogens. To avoid these risks it is proposed that improved prophylactic measures should be taken.     

Detection of parasitizing coccidia and determination of host crane species, sex and genotype by faecal DNA analysis

In Japan, the three main crane species are the endangered red-crowned crane (Grus japonensis) inhabiting Hokkaido, the northernmost island of Japan; the vulnerable hooded crane (Grus monacha); and the vulnerable white-naped crane (Grus vipio). Both the hooded and white-naped cranes migrate in winter to Izumi in Kyushu, the southern island of Japan. In this study, we investigated the cranes and their coccidian parasites, through a targeted molecular approach using faecal DNA to develop a noninvasive method for infectious disease research. To determine the origin of noninvasively collected faecal samples, host species were identified by sequencing a region of approximately 470 bp of the mitochondrial 16S ribosomal RNA gene in the faecal DNA. Furthermore, to avoid sample redundancy, individual determination was performed by fragment analysis using microsatellite and sex-linked markers. For microsatellite genotyping, previously reported markers and markers isolated in this study were examined, and seven loci for red-crowned cranes, eight for hooded cranes and six for white-naped cranes displayed polymorphisms. A low error rate was demonstrated by comparing microsatellite data generated from faecal DNA samples with that generated from feather DNA samples, indicating a high reliability. Polymerase chain reaction-based capillary electrophoresis (PCR-CE), employing genetic markers in the second internal transcribed spacer (ITS2) of nuclear ribosomal DNA, was employed to detect crane coccidia. The sensitivity of detection of PCR-CE using faecal DNA was inferior to that with traditional microscopy; however, our results suggest that PCR-CE can depict crane coccidia diversity with higher resolution and it is a useful tool to characterize community composition of coccidia in detail.     

Obtaining an Interspecific Hybrid of Cranes by Artificial Insemination with Frozen–thawed Semen

Studies of artificial insemination of cranes and cryoconservation of their semen have been carried out in the nursery of rare species at the Oka Biosphere Reserve for many years. The criterion of successful cryoconservation of the semen is the obtaining of fertilized eggs after artificial insemination by the thawed semen. An experiment is described on artificial insemination of females of the white-naped crane Grus vipio by the frozen–thawed semen of the Siberian white crane G. leucogeranus after one-year storage of semen in liquid nitrogen. As a result, an interspecific hybrid of cranes was obtained, which confirmed the possibility of producing a bank of cryoconserved crane semen. The use of the white-naped crane females was due to the absence of conspecific males and unavailability of Siberian white crane females. Problems of artificial insemination and cryoconservation of semen of rare crane species are discussed.     

Obtaining an Interspecific Hybrid of Cranes by Artificial Insemination with Frozen–thawed Semen

Studies of artificial insemination of cranes and cryoconservation of their semen have been carried out in the nursery of rare species at the Oka Biosphere Reserve for many years. The criterion of successful cryoconservation of the semen is the obtaining of fertilized eggs after artificial insemination by the thawed semen. An experiment is described on artificial insemination of females of the white-naped crane Grus vipio by the frozen–thawed semen of the Siberian white crane G. leucogeranusafter one-year storage of semen in liquid nitrogen. As a result, an interspecific hybrid of cranes was obtained, which confirmed the possibility of producing a bank of cryoconserved crane semen. The use of the white-naped crane females was due to the absence of conspecific males and unavailability of Siberian white crane females. Problems of artificial insemination and cryoconservation of semen of rare crane species are discussed.     

Construction of PCR primers to detect and distinguish Eimeria spp. in northern bobwhites and a survey of Eimeria on gamebird farms in the United States

Coccidiosis is an important disease in captive gamebirds, including northern bobwhites (Colinusvirginianus). Three Eimeria species, Eimeria lettyae, Eimeria dispersa, and Eimeria colini, have been described in bobwhites. Distinguishing the various Eimeria spp. is often problematic because of similarity in oocyst morphology and site of infection and thus requires live bird infections to distinguish between the coccidian species. To aid in identification and diagnosis, PCR specific primers were generated against the internal transcribed spacer region 1 (ITS-1) of the ribosomal RNA gene using sequences obtained from coccidian-positive samples collected from diagnostic cases or litter from captive bobwhites. Three distinct Eimeria spp. were detected. Species-specific primers were constructed and used to survey the prevalence of the species in 31 samples collected from 13 states. The primers survey results identified E. lettyae, E. dispersa, and Eimeria sp. in 20 (64.5%), 22 (71%), and 29 (93.5%) of the samples, respectively. Mixed infections were common: 13 (41.9%) samples had 3 Eimeria spp., 14 (45.2%) had 2 spp., and 4 (12.9%) samples had only 1 species. The species were widely distributed over the area sampled and were not associated with the age of the flock.     

Molecular survey of Apicomplexa in Podarcis wall lizards detects Hepatozoon, Sarcocystis, and Eimeria species

The occurrence of apicomplexan parasites in Podarcis sp. wall lizards from the Iberian Peninsula and Balearic islands was studied by amplification and sequencing of the 18S rRNA gene. Species from 3 genera, Hepatozoon , Sarcocystis , and Eimeria , were found. The phylogenetic analysis of the 18S rRNA gene provides unexpected insights into the evolutionary history of these parasites. All Hepatozoon spp. specimens were recovered as part of a clade already identified in lizards from North Africa. The Sarcocystis species, detected in Podarcis lilfordi from Cabrera Island in the Balearic Islands, appears related to Sarcocystis gallotiae , known only from endemic Gallotia sp. lizards from the Canary Islands. Based on the lack of snake predators on this island, this parasite presumably presents an atypical transmission cycle that uses the same host species as both intermediate and final host through cannibalism, like S. gallotiae . Eimeria sp. is reported for the first time from Podarcis spp. lizards. This study shows the power of detecting multiple different apicomplexan parasites through screening of tail tissue samples and blood drops that are often collected in reptiles for other purposes.     

Blood and intestinal parasites of squirrels (Rodentia: Sciuridae) in Amazonian Brazil

We report the result of an examination for blood and intestinal protozoa in 12 specimens of the red squirrel Sciurus spadiceus (Rodentia: Sciuridae) from Birroque, municipality of Placido de Castro, state of Acre, Brazil. No parasites were detected in thin, Giemsa-stained blood films of the animals, but culture of the blood of three in Difco B45 medium blood-agar slants gave rise to isolates of epimastigotes. Inoculation of one isolate into laboratory mice resulted in the appearance of Trypanosoma cruzi-like trypomastigotes in their peripheral blood, and the other two isolates gave rise to transient infections with a T. lewisi-like parasite in inoculated mice and hamsters. The failure of the latter parasite to develop in the triatomine bug Rhodnius robustus suggests that it is probably not T. rangeli. This appears to be the first record of a T. lewisi-like trypanosome in neotropical squirrels. Oocysts of an Eimeria sp., were detected in the faeces of 10 animals (83.3%). The parasite develops in the epithelial cells of the intestine, where it may cause severe damage and sometimes results in death of the animal. No oocysts were detected in bile.     

圈养丹顶鹤与白枕鹤杂交繁育

我园一只雄性丹顶鹤与一只雌性白枕鹤于2001年开始自然配对产蛋,3年共产51枚,经人工孵化育雏,目前存活15只。杂种鹤在第4周后进入快速增长阶段,在12周达到最高峰,16周后增长速度明显下降,18周后部分指标停止增长。外观上,出生至3月龄时,幼鹤整体颜色都呈黄褐色,以后变化基本一致。目前,2001及2002年存活杂种鹤颜色以白色为主,夹杂着灰色羽毛。     

Parasitological study of a population of Tiber River eels (Anguilla anguilla)

A study was conducted on the parasitofauna of Anguilla anguilla caught in the section of the Tiber river south of Rome (Italy). The sample examined consisted of 417 specimens that were classified by stage (elver, young yellow eel, yellow eel and silver eel) and season of sampling. The following species were identified: Trypanosoma granulosum, Eimeria anguillae, Myxidium giardi, Sphaerospora reichenowi, Myxobolus sp., Trichodina anguilli, Ichthyohodo sp., Gyrodactylus anguillae, Bothriocephalus claviceps, Proteocephalus macrocephalus, Raphidascaris acus, Acanthocephalus clavula. Each parasite species was considered in its prevalence according to stage and season. For some parasite the pathogenic effect was also examined in a histological study of serial sections. The results of the statistical analysis of the single parasitic infection as related to different seasons pointed to marked seasonal trends only in some species (Trypanosoma granulosum and Gyrodactylus anguillae). The intensity of infection assessed for acanthocephalan infection only was found to affect neither weight nor condition index (weight/length3). No difference in weight or condition index was found to be ascribable to the presence or absence of a species. An attempt to find a rule-of-thumb algorithm to gauge the overall effect to parasitic diseases failed to give sufficiently significant results.     

Trematode infections in the small intestine of Egretta alba modesta in Kangwon-do]

We surveyed 12 migrating large egrets, Egretta alba modesta, for their infection status with intestinal trematodes, from June to September, 1990. All of them were infected with one or more flukes. Total 5 species were identified. Their infection rates and total numbers of worms were Metagonimus sp. 67% and 1,412, Centrocestus armatus 67% and 2,306, Echinochasmus japonicus 50% and 99, Strigea falconis 83% and 635, and a diplostomid fluke 33% and 36. The number of worms in a bird was in the range from 6 to 896. This is the first record that the egret is a reservoir host of Metagonimus sp., C. armatus, E. japonicus, and S. falconis. S. falconis is a new parasite fauna in Korea. Migrating birds must play an important role in the transmission and spreading of these intestinal trematodes in Korea.     

Effect of heterospecifics on foraging of endangered red-crowned and white-naped cranes in the Korean Demilitarized Zone

We determined how the presence of heterospecific individuals in the vicinity of a focal individual affects the behavior of two critically endangered species of cranes on their wintering grounds at Cheolwon in the Korean Demilitarized Zone. The red-crowned crane, Grus japonensis, is larger than the white-naped crane, Grus vipio, and it dominates the white-naped crane in aggressive interactions. We showed that the dominant species increases foraging activity in the presence of the subordinate species presumably because of scrounging of food from the subordinate. Because interspecific interactions may affect avian endangerments these behavioral findings should be taken into account when managing winter refuges for the two endangered crane species.     

滦河上游——白枕鹤西部种群的重要停歇地

白枕鹤(Grus vipio)为国家II级重点保护野生动物, 被IUCN列为易危(VU)物种。白枕鹤西部种群繁殖于中蒙俄交界处的达乌尔地区, 数量呈下降趋势。我们于2017-2018年在蒙古国东部给白枕鹤西部种群的50只个体佩戴了GPS-GSM跟踪设备。截至2019年5月, 获得春季和秋季迁徙路径各48条。分析结果显示: 春季91.67%和秋季72.91%的跟踪个体在滦河上游(河北省沽源-内蒙古正蓝旗-多伦区域)停歇, 春季停留时间36.16 ± 15.00天、秋季20.26 ± 11.08天, 分别占春季和秋季迁徙时间的75%和67%, 确定了这一区域是西部种群迁徙途中最重要的停歇地。迁徙路线栖息地选择模型结果显示, 白枕鹤常在距离湖泊较近(< 210 km)、海拔1,200-1,500 m, 且坡度小(< 1°)的区域停歇。而滦河上游和整条迁徙路线停歇位点比较的模型结果显示, 滦河上游停歇地的海拔1,200-1,500 m与整条迁徙路线栖息地选择模型的结果一致; 此外这个区域离河流更近(< 70 km), 不仅有湿地和水体的栖息环境, 还有草地和农田可供觅食和栖息。保护空缺分析发现滦河上游现有四处保护地, 但在保护地内的迁徙停歇点不超过总位点的1.63%。综上, 我们建议将滦河上游整体纳入保护地体系进行管理, 为这一受胁物种及其栖息地管理和保护提供可靠保障。     

Molecular characterization of the 18S rDNA gene of an avian Hepatozoon reveals that it is closely related to Lankesterella

As a part of intensive study of blood parasite infections in a population of the passerine bird blue tit (Cyanistes caeruleus, Paridae), we detected a parasite species that, based on its morphological similarity, was tentatively identified as Hepatozoon parus, the only species of this parasite genus described from birds of this family. However, morphological measurements show that H. parus is slightly larger than the parasite detected in our population. A molecular characterization of the parasite species was conducted by amplification of the 18S rDNA gene, using primers that were reported previously to amplify in Hepatozoon sp. of water pythons. Additional primers were developed based on the new sequence obtained. The 1,484-bp fragment amplified reveals that the parasite from our bird population is more closely related to Lankesterella minima than to Hepatozoon species from other vertebrates according to analysis using the BLAST comparison method (93% identity). In addition, phylogenetic analyses using parsimony and Kimura procedures unequivocally related the parasite species detected by PCR with L. minima. The bootstrap values obtained were 97% and 100%, respectively. These results imply that this parasite is a species of a lankesterellid instead of Hepatozoon.     

Embryo vaccination against Eimeria tenella and E. acervulina infections using recombinant proteins and cytokine adjuvants

Avian coccidiosis is an intestinal disease caused by protozoa of the genus Eimeria. To investigate the potential of recombinant protein vaccines to control coccidiosis, we cloned 2 Eimeria sp. genes (EtMIC2 and 3-1E), expressed and purified their encoded proteins, and determined the efficacy of in ovo immunization to protect against Eimeria infections. Immunogen-specific serum antibody titers, parasite fecal shedding, and body weight gains were measured as parameters of disease. When administered alone, the recombinant EtMIC2 gene product induced significantly higher antibody responses, lower oocyst fecal shedding, and increased weight gains compared with nonvaccinated controls following infection with E. tenella. Combined embryo immunization with the EtMIC2 protein plus chicken cytokine or chemokine genes demonstrated that all 3 parameters of vaccination were improved compared with those of EtMIC2 alone. In particular, covaccination with EtMIC2 plus interleukin (IL)-8, IL-16, transforming growth factor-beta4, or lymphotactin significantly decreased oocyst shedding and improved weight gains beyond those achieved by EtMIC2 alone. Finally, individual vaccination with either EtMIC2 or 3-1E stimulated protection against infection by the heterologous parasite E. acervulina. Taken together, these results indicate that in ovo vaccination with the EtMIC2 protein plus cytokine/chemokine genes may be an effective method to control coccidiosis.     

Ecology,not distance,explains community composition in parasites of sky-island Audubon’s Warblers

Haemosporidian parasites of birds are ubiquitous in terrestrial ecosystems, but their coevolutionary dynamics remain poorly understood. If species turnover in parasites occurs at a finer scale than turnover in hosts, widespread hosts would encounter diverse parasites, potentially diversifying as a result. Previous studies have shown that some wide-ranging hosts encounter varied haemosporidian communities throughout their range, and vice-versa. More surveys are needed to elucidate mechanisms that underpin spatial patterns of diversity in this complex multi-host multi-parasite system. We sought to understand how and why a community of avian haemosporidian parasites varies in abundance and composition across elevational transects in eight sky islands in southwestern North America. We tested whether bird community composition, environment, or geographic distance explain haemosporidian parasite species turnover in a widespread host that harbors a diverse haemosporidian community, the Audubon’s Warbler (Setophaga auduboni). We tested predictors of infection using generalized linear models, and predictors of bird and parasite community dissimilarity using generalized dissimilarity modeling. Predictors of infection differed by parasite genus: Parahaemoproteus was predicted by elevation and climate, Leucocytozoon varied idiosyncratically among mountains, and Plasmodium was unpredictable, but rare. Parasite turnover was nearly three-fold higher than bird turnover and was predicted by elevation, climate, and bird community composition, but not geographic distance. Haemosporidian communities vary strikingly at fine spatial scales (hundreds of kilometers), across which the bird community varies only subtly. The finer scale of turnover among parasites implies that their ranges may be smaller than those of their hosts. Avian host species should encounter different parasite species in different parts of their ranges, resulting in spatially varying selection on host immune systems. The fact that parasite turnover was predicted by bird turnover, even when considering environmental characteristics, implies that host species or their phylogenetic history plays a role in determining which parasite species will be present in a community.