Leptospira borgpetersenii from free-living white-eared opossum (Didelphis albiventris): First isolation in Brazil

Leptospirosis is a zoonotic disease that occurs all over the world, caused by bacteria of the genus Leptospira. Marsupial and didelphidae families are considered susceptible to infection caused by a wide range of Leptospira serovars for which they serve as reservoirs. Thirty-three free-living white-eared opossums (Didelphis albiventris) were captured in Southern Brazil and bodily fluids were collected. From the urine samples it was possible to obtain an isolate identified as Leptospira borgpetersenii by rpoB gene sequencing and belonging to serovar Castellonis by Multilocus Variable-Number Tandem-Repeat Analysis. This is the first report of the isolation of Leptospira spp. from the white-eared opossum in Brazil. In addition, the new strain was also virulent in the hamster model of lethal leptospirosis. The microscopic agglutination test (MAT) was used for detecting the presence of antibodies against Leptospira spp. in white-eared opossum, human, cattle and canine sera using a panel of 59 Leptospira strains that included the new isolate. The inclusion of the new strain in the MAT battery increased the MAT sensitivity for canine sera. These findings suggest that the white-eared opossum is an important reservoir of pathogenic Leptospira spp.     

Polynucleobacter necessarius,a model for genome reduction in both free-living and symbiotic bacteria

We present the complete genomic sequence of the essential symbiont Polynucleobacter necessarius (Betaproteobacteria), which is a valuable case study for several reasons. First, it is hosted by a ciliated protist, Euplotes; bacterial symbionts of ciliates are still poorly known because of a lack of extensive molecular data. Second, the single species P. necessarius contains both symbiotic and free-living strains, allowing for a comparison between closely related organisms with different ecologies. Third, free-living P. necessarius strains are exceptional by themselves because of their small genome size, reduced metabolic flexibility, and high worldwide abundance in freshwater systems. We provide a comparative analysis of P. necessarius metabolism and explore the peculiar features of a genome reduction that occurred on an already streamlined genome. We compare this unusual system with current hypotheses for genome erosion in symbionts and free-living bacteria, propose modifications to the presently accepted model, and discuss the potential consequences of translesion DNA polymerase loss.Symbiosis, defined as a close relationship between organisms belonging to different species (1), is a ubiquitous, diverse, and important mechanism in ecology and evolution (e.g., refs. 2–4). In extreme cases, through the establishment of symbiotic relationships, quite unrelated lineages can functionally combine their genomes and generate advantageous emergent features or initiate parasite/host arms races. Ciliates, common unicellular protists of the phylum Ciliophora, are extraordinary receptacles for prokaryotic ecto- and endosymbionts (5, 6) that provide varied examples of biodiversity and ecological roles (6). Nevertheless, most of these symbionts are understudied, partially owing to the scarcity of available molecular data and the absence of sequenced genomes. Yet, thanks to their various biologies and the ease of sampling and cultivating their protist hosts, they are excellent potential models for symbioses between bacteria and heterotrophic eukaryotes. Until recently this field was dominated by studies on endosymbionts of invertebrates, especially insects (e.g., ref. 7), although unicellular systems like amoebas (e.g., refs. 8 and 9) have been shown to be suitable models.Polynucleobacter necessarius was first described as a cytoplasmic endosymbiont of the ciliate Euplotes aediculatus (10, 11). Further surveys detected its presence in a monophyletic group of fresh and brackish water Euplotes species (12, 13). All of the investigated strains of these species die soon after being cured of the endosymbiont (10, 12, 13). In the few cases in which P. necessarius is not present, a different and rarer bacterium apparently supplies the same function (12, 14). No attempt to grow symbiotic P. necessarius outside their hosts has yet been successful (15), strongly suggesting that the relationship is obligate for both partners, in contrast to most other known prokaryote/ciliate symbioses (6).Thus, the findings of many environmental 16S rRNA gene sequences similar to that of the symbiotic P. necessarius (16) but belonging to free-living freshwater bacteria came as a surprise. These free-living strains, which have been isolated and cultivated (17), are ubiquitous and abundant in the plankton of lentic environments (17, 18). They are smaller and do not show the most prominent morphological feature of the symbiotic form: the presence of multiple nucleoids, each containing one copy of the genome (10, 11). It is clear that free-living and endosymbiotic P. necessarius are not different life stages of the same organism (15). Nevertheless, these strikingly different bacteria, occupying separate ecological niches, exhibit >99% 16S rRNA gene sequence identity, and phylogenetic analyses fail to separate them into two distinct groups (15). Rather, several lines of evidence point to multiple, recent origins of symbiotic strains from the free-living bacterial pool (14, 15).Thus, the Euplotes–Polynucleobacter symbiosis provides a promising system for the study of changes promoting or caused by the shift to an intracellular lifestyle. The remarkably small (2.16 Mbp) genome of the free-living strain QLW-P1DMWA-1 has been sequenced and studied, especially for features that would explain the success of this lineage in freshwater systems worldwide (19, 20). Phylogenies based on the 16S rRNA gene (13, 14) and multiple-gene analyses (19, 21, 22) consistently cluster Polynucleobacter with bacteria of the family Burkholderiaceae (Betaproteobacteria), either in a basal position or as the sister group of Ralstonia and Cupriavidus.Here we provide the complete genomic sequence of a symbiotic P. necessarius harbored in the cytoplasm of E. aediculatus and present a comparative analysis of the two sequenced Polynucleobacter genomes, addressing the possible biological basis of the Euplotes–Polynucleobacter symbiosis. We also provide insights into the evolution of the unique two-step genome reduction in this bacterial species: the first step involving streamlining in a free-living ancestor and the second a more recent period of genome erosion confined to the symbiotic lineage.     

Corticosterone concentrations in free-living spotted salamanders (Ambystoma maculatum)

A non-lethal technique for drawing repeated blood samples from spotted salamanders (Ambystoma maculatum) was used to examine sex, seasonal, and capture technique differences in the physiological stress response under natural, undisturbed conditions. Baseline and stress-induced (30 min of handling and restraint) corticosterone (CORT) concentrations were measured at night during the spring migrations into and out of the breeding pond, as well as during the fall migration to over-wintering sites. Females had significantly higher CORT concentrations than males during the spring migration toward the breeding pond, but this difference was not present when animals emerged from the pond post-breeding or during the fall migration. CORT concentrations did not vary seasonally, but during the acute stress response, CORT concentrations nearly doubled during the spring inbound migration, exhibited a nearly significant increase during the spring outbound migration, and did not change in the fall. Allowing animals to sit overnight in the buckets in which they were captured elicited CORT concentrations that tended to be higher (although not statistically significant) than when animals were sampled on the night of capture. The ability to sample blood from spotted salamanders using a non-lethal technique offers an opportunity to better understand both amphibian physiology and ecology.     

Tick-borne encephalitis (TBE) trends in epidemiology and current and future management

Tick-borne encephalitis (TBE) is considered an international health issue, as the number of risk areas and reported cases across Europe, Russia, and parts of Asia continues to increase. The incidence of TBE has fluctuated considerably from year to year in many countries, but in the past decade the number of TBE cases has significantly increased in the Baltic states, the Czech Republic, and Germany, in addition to occurring in countries previously considered to be free from TBE, such as Denmark (specifically the main island of Zealand), France, and Italy. A number of factors have been suggested to explain the increase in incidence, including climate change, and increased travel and outdoor pursuits, placing people in increased contact with infected ticks. There is no causal treatment available once infected, but TBE can be effectively prevented by vaccination, for which several vaccines are widely available. Three vaccination schedules are available for immunization against TBE, and the recommendations for TBE vaccination vary considerably across the countries in which TBE foci are found. However, plans are in place to raise awareness of TBE and to standardize the vaccination programme across Europe, with the aim of reducing the number of future cases of TBE.     

Tick-borne rickettsioses in Pune district, Maharashtra, India

An extensive study on tick-borne rickettsioses in the Pune district of Maharashtra revealed that Indian tick typhus exists as a zoonosis, which only occasionally causes disease in man. By sero-conversion in guinea pigs, presumptive isolates of Rickettsia conori and Coxiella burnetii were recovered from 4 of the 11 species of ticks examined. Boophilus microplus and Rhipicephalus haemaphysalis were found to be harbouring R. conori whereas C. burnetii was isolated from Haemaphysalis intermedia and Hyalomma hussaini in addition to the above mentioned 2 tick species. Complement fixation tests carried out on sera from various species of rodents and gerbils revealed the presence of antibodies against the R. conori antigen in the sera of Rattus blanfordi, R.r. rufescens and Suncus murinus. In the case of large mammals, similar antibodies were detected in the sera from dog, cow, horse and sheep. C. burnetii infection was found to exist in both the sylvan and domestic cycle, as evidenced from the involvement of ticks, large and small mammals and man in its natural history.     

Corticosterone, brood size, and hatch order in free-living Florida scrub-jay (Aphelocoma coerulescens) nestlings

It is well known that variation in developmental conditions can have profound effects upon lifetime fitness. In altricial avian species, nestlings undergo a substantial portion of development in the nest after hatching, often in the presence of nest-mates. This can result in the formation of brood hierarchies based on age, size, and competitive ability. Measurement of baseline corticosterone (CORT) levels in developing birds may provide a means to assess whether individuals within a brood experience stress due to sibling competition or nutritional state. However, few studies have attempted to correlate corticosterone concentrations with brood hierarchies in free-living, developing birds. We investigated the degree to which corticosterone levels in nestling Florida scrub-jays (Aphelocoma coerulescens) were predicted by brood size, hatching order, and body mass. Nestling corticosterone levels were inversely correlated with body mass but did not differ among brood sizes. Within broods of two, second-hatched nestlings had lower CORT than first-hatched nestlings, although there was no effect of hatch order in broods of three or four nestlings. The results of this study suggest that conditions within the nest, particularly those associated with within-brood hierarchies, are manifested through differential body condition and nestling corticosterone secretion. The consequences of this variation in nestling corticosterone are unknown, but development of the adult phenotype, as well as life-long survival, may be impacted.     

Circulating prolactin in free-living California ground squirrels (Spermophilus beecheyi)

During a long-term field study of a free-living population of California ground squirrels (Spermophilus beecheyi), blood samples were drawn at regular intervals from marked animals via femoral venipuncture, and plasma prolactin (PRL) was measured by homologous radioimmunoassay. Marked fluctuations with season and reproductive condition occurred in circulating PRL levels in both males and females, with peak levels occurring during the spring breeding season. Peak PRL levels in females were reached 4 weeks after peak male levels, but mean peak values did not differ significantly between the sexes. Among females, juveniles had lower mean PRL levels than adults, and yearlings had lower mean levels throughout their initial reproductive episodes than did older females.     

Reproductive endocrinology of a free-living, opportunistically breeding passerine (white-winged crossbill, Loxia leucoptera)

The aim of the study was to investigate, in a free-living opportunistic breeder, the White-winged Crossbill (Loxia leucoptera), the degree to which annual changes in photoperiod and differences in the timing of breeding in different years correlate with the secretion of reproductive hormones. Seasonal changes in concentrations of luteinizing hormone (LH) and prolactin were compared in White-winged Crossbills in interior Alaska (64 degrees 50'N, 147 degrees 50'W) in plasma samples taken in May-December 1994 and July 1998-August 1999. The birds were in breeding condition in May-July 1994 and in February-April 1999. Additional observations were made on plasma testosterone (T) in males in 1998/1999. In 1998, in adult males, plasma LH and T were low in September-October (LH: <0.4 ng/ml; T: undetectable) and increased significantly in November (LH: 3.65 +/- 2.20 ng/ml) and December (T: 0.69 +/- 0.88 ng/ml). Thereafter, plasma T increased to 2.35 +/- 2.03 ng/ml in March/April and returned to basal levels by June/July. In 1998/1999, in females, plasma LH did not change seasonally. In males in 1994, plasma LH was higher in June (2.15 +/- 1.22 ng/ml) and July (2.86 +/- 0.69 ng/ml) than in the same period in 1999 (June: 1.20 +/- 1.82 ng/ml; July: 1.37 +/- 1.03 ng/ml), which is consistent with the later breeding season in 1994. In 1994 and 1998/1999 in both sexes, concentrations of plasma prolactin were elevated when day lengths exceeded about 12 h, being high during late spring to early fall (males: 40-130 ng/ml; females: 40-140 ng/ml) and low during winter (<15 ng/ml in both sexes). In 1994 and 1998, molt occurred during the fall, at a time when breeding did not occur, although conifer seeds were sufficiently abundant to support a large bird population. These data are consistent with the view that White-winged Crossbills have the potential to breed at all times of the year except when molting in the fall. It is likely that a photoinduced increase in prolactin secretion is a factor that induces molt. Crossbills may not breed when they are molting because of increased metabolic demands for feather formation and for the maintenance of basal metabolism at a time of year when ambient temperatures fall well below 0 degrees. Alternatively or in addition, the birds may become reproductively photorefractory.     

Corticosterone responses change seasonally in free-living house sparrows (Passer domesticus)

Avian corticosterone (CORT) concentrations vary seasonally for many species, but most studies examined species from extreme environments or from only a few times of the year. It is unclear how general a phenomenon this is. We examined both baseline and stress-induced CORT from house sparrows (Passer domesticus) in two habitats-Massachusetts, similar to their ancestral habitat; and New Mexico, a semi-arid desert where the house sparrow only thrives as an obligate human commensal. We captured both males and females during four times of the year-Spring (when the male cloacal protuberance indicated birds were in breeding condition), Fall, Winter, and in the late summer when birds were undergoing a prebasic molt. Birds were heaviest and had the longest wing chord lengths in the Fall at both sites and Massachusetts birds were approximately 10% heavier than New Mexico birds. House sparrows also showed a seasonal variation in the amount of fat stores, but the seasonal pattern differed. Massachusetts birds were fatter overall and showed the most fat during Fall and Winter, whereas New Mexico birds showed the most fat in the Spring. Both baseline and stress-induced total CORT did not differ between sexes or sites, with the exceptions of baseline CORT in the Fall and stress-induced CORT in the Spring being elevated in Massachusetts males. There was a distinct seasonal pattern at both sites, however, with total CORT being highest in the Spring and Winter and lowest during Fall and molt. This seasonal pattern was mirrored in corticosterone binding globulin (CBG) capacities, and when free CORT was estimated, the seasonal pattern disappeared. Stress-induced free CORT, however, was higher in Massachusetts males and females during the Fall and Winter, suggesting a potential differential response to stress associated with commensalism.     

Seasonal variations in thyroid hormone levels in free-living echidnas (Tachyglossus aculeatus)

Hulbert and Augee (1982) have suggested that the thyroid has little effect on energy metabolism in the echidna. In order to investigate whether thyroid status changes during hibernation, when metabolism drops dramatically, we measured levels of thyroid hormones in 31 free-living echidnas at various times during the year. Unlike eutherian hibernators, in which thyroid hormone levels may rise to seasonally high values in late hibernation, total and free T(4) and T(3) were all significantly depressed throughout hibernation. TT(4) from nonhibernating echidnas was 11.8 +/- 0.9 ng/ml (n = 23), confirming previously published values, but fell to half this level (5.9 +/- 0.7 ng/ml, n = 8) during hibernation. By contrast to the low TT(4) values, nonhibernating FT(4), TT(3), and FT(3) values were similar to normal values for eutherian mammals. Differences in the seasonal pattern of variation in thyroid hormones between echidnas and hibernating eutherians may be due to differences in thyroid hormone transporting proteins.     

新疆阿勒泰山地蜱传脑炎疫源地调查

目的 调查新疆阿勒泰山地蜱传脑炎疫源地特征,分离鉴定蜱传脑炎病毒。方法 通过家畜体表捡法采集寄生蜱;利用间接免疫荧光法检测当地健康人群血清中蜱传脑炎病毒IgG抗体;通过将蜱研磨液接种实验小鼠进行病原动物分离;通过接种BHK-21细胞对蜱传脑炎病毒进行分离培养;利用RT-PCR方法对病毒E蛋白基因片段进行扩增和测序,通过序列分析明确病毒系统进化特征。结果 新疆阿勒泰山地白哈巴地区分布有2种蜱,森林革蜱为优势种(55.6%),其次为边缘革蜱(44.4%);当地人群蜱传脑炎IgG抗体阳性率5.31%(6/113);通过动物试验和细胞分离培养,从森林革蜱中分离出一株森林脑炎病毒;对病毒E蛋白基因序列的系统进化分析表明,蜱传脑炎病毒阿勒泰分离株属于远东亚型。结论 首次从病原学上证实新疆阿勒泰山地存在蜱传脑炎疫源地,媒介为森林革蜱,病毒流行株为远东亚型。     

Seasonal changes in hypothalamic-pituitary-adrenal axis sensitivity in free-living house sparrows (Passer domesticus)

Recent evidence indicates that house sparrows (Passer domesticus) seasonally regulate corticosterone responses to capture, handling, and restraint. Responses during molt and in the fall are lower than responses in the winter and while breeding. This study tested whether changes in either adrenal tissue responsiveness to adrenocorticotropin (ACTH) or pituitary responsiveness to corticotropin releasing factor (CRF) or arginine vasotocin (AVT) could provide the mechanism regulating these seasonal changes. House sparrows were captured at two sites (Massachusetts and New Mexico, USA) and during the above four seasons and injected with exogenous ACTH, CRF, and AVT. ACTH stimulated further corticosterone release in all birds except Massachusetts birds in the winter, suggesting that reduced adrenal sensitivity to ACTH cannot explain reduced corticosterone release during fall and molt. However, exogenous ACTH was less effective during molt at both sites, implying that adrenal sensitivity does change. Pituitary sensitivity also changed seasonally, but these pituitary changes did not match the seasonal changes in corticosterone release. CRF and AVT only succeeded in elevating corticosterone in the spring in Massachusetts birds and in the winter in New Mexico birds, whereas CRF alone also stimulated corticosterone release in New Mexico birds in the fall. Taken together, these data indicate that house sparrows can alter the amount of corticosterone released from adrenal tissue, the amount of ACTH released from the pituitary, and the amount of CRF and AVT released from the hypothalamus, but that none of these changes correlate with seasonal changes in corticosterone release. Consequently, seasonal modulation of corticosterone release in house sparrows appear to result from a complicated mix of adrenal, pituitary, and hypothalamic changes that also vary seasonally.