A Sensitive and Specific DNA Probe for the Oyster Pathogen Haplosporidium nelsoni

ABSTRACT. Haplosporidium nelsoni is a pathogen of the eastern oyster, Crassostrea virginica , along the middle Atlantic coast of the U.S. Genomic DNA was extracted from H. nelsoni plasmodia and small subunit (SSU) rDNA was amplified by PCR, cloned and sequenced. The sequence of H. nelsoni SSU rDNA was aligned with that of another haplosporidian, Minchinia teredinis , and with SSU rDNA data of C. virginica and various protists in GenBank. A 21-base oligonucleotide unique to H. nelsoni , designated MSX1347, was commercially synthesized and tested for sensitivity and specificity. In dot blot hybridizations the probe detected 100 pg of cloned H. nelsoni rDNA and the presence of H. nelsoni in 1 μg of genomic DNA from an infected oyster. It did not hybridize with 1 μg of genomic DNA from uninfected C. virginica or with cloned SSU rDNA of M. teredinis. The probe was further tested for specificity with in situ hybridizations on AFA-fixed, paraffin-embedded tissue sections. The probe hybridized well with H. nelsoni plasmodia and immature spores, but poorly with mature spores. The probe did not hybridize with oyster tissue, with other common oyster parasites such as P. marinus or Nematopsis sp., or with the haplosporidians Haplosporidium louisiana from mud crabs ( Panopeus spp.), Haplosporidium costale from C. virginica or M. teredinis from shipworms ( Teredo spp.).     

Ion Compartmentation in the Marine Fungus Dendryphiella salina in Response to Salinity: X-ray Microanalysis

X-ray microanalysis was performed on hyphae of the filamentousmarine fungus Dendryphiella salina growing at different salinitiesto give sodium, potassium and chloride concentrations in thecytoplasm, vacuole and cell wall. Sodium and chloride concentrationsincreased with salinity in all compartments. Cytoplasmic andvacuolar sodium and chloride concentration were broadly similar,and vacuolar contents represented, at most, 19% of the totalprotoplasmic content of an individual ion species. Potassiumconcentrations decreased to some extent with salinity, althoughconcentrations were not severely affected by competition withsodium uptake. Results are discussed with regard to the roleof ions in the overall osmotic adjustment in this species. Key words: Dendryphiella salina, marine fungus, salt-tolerance, x-ray microanalysis     

Fine Structure of Sarcocystis cruzi Schizonts

SYNOPSIS Schizogony of Sarcocystis cruzi Hasselmann (syn. S. fusiformis Railliet) takes place in vascular endothelial cells 26 to 33 days after cattle ingest sporocysts from dogs. Kidney cortex from a heavily infected, dexamethasone-treated bovine was fixed for electron microscopy to determine the method of schizogonie development. Schizogony takes place by endopolygeny characterized by marked enlargement of the parasite nucleus, formation of nuclear lobes, presence of numerous spindles with adjacent pairs of centrioles along the nucleus, and simultaneous formation of daughter merozoites in the cytoplasm adjacent to the spindle poles. Endopolygeny in S. cruzi differs from that in other Sporozoa in that merozoite anlagen are seen in the cytoplasm before any nuclei divide. The resultant merozoites continue development and, when mature, resemble other sporozoan zoites. Upon release from the host cell into capillaries, they travel to muscle tissue to continue the life cycle by forming sarcocysts.     

Combining surveys of river habitats and river birds to appraise riverine hydromorphology

1. Biological monitoring is vital to river conservation. Aside from providing census data, regular monitoring may detect population trends that reflect the degradation or remediation of riverine environments. Birds are major candidates for this purpose because of their connections to riverine food webs and river habitat features. However, much information on factors affecting river bird distributions is qualitative. 2. River bird populations have been surveyed annually in the U.K. since 1974. The value of this monitoring could be increased if links between population trends and changes in riverine landscapes or channel hydromorphology were better understood. We modelled the relationships between bird species’ distributions recorded by the British Trust for Ornithology's Waterways Breeding Bird Survey (WBBS) and hydromorphology recorded by the Environment Agency's River Habitat Survey (RHS). 3. Regression models were built for 20 bird species associated with the river channel and neighbouring floodplains. This provided one of the first illustrations in ecology of generalized estimating equations (GEEs) for extending generalized linear models to instances where individual observations are not statistically independent. GEEs could offer a range of benefits to freshwater biologists, as they are a simple approach to analysing hierarchical, correlated data while maintaining the ease of interpretation with a range of link functions and data types. 4. Upland, fast water species and reedbed specialists showed the closest links with RHS, followed by lowland, slow water species, where correlations to RHS varied widely in magnitude. The distributions of floodplain wading birds were poorly modelled by RHS, suggesting that floodplain land uses as quantified by RHS were poor predictors of distributions. Marked preferences between upland and lowland channels were evident for most species. Specific features were also important, including hydraulics, bank/channel vegetation, depositional features and anthropogenic structures. 5. Monitoring the abundance and distribution of a range of bird species, with a diversity of life histories, could be a valuable tool for assessing trends in riverine landscapes and whole river basins. RHS captures useful information about channel hydromorphology and vegetation, but needs supplementing with additional environmental information to describe (i) the wider riverine landscape (e.g. more detailed floodplain variables, proximity of other waterbodies) and (ii) water chemistry. Although further work is required, this study suggests that WBBS and RHS add mutual value in appraising the river environment. We advocate the further development of birds as indicators in riverine landscapes that could convey the importance of the ecological integrity, biological production and conservation value of river systems to a wider audience.     

Isolation of polymorphic microsatellite markers in the sub‐Saharan tree,Acacia (Senegalia) mellifera (Fabaceae: Mimosoideae)

We isolated 11 polymorphic microsatellite markers from Acacia mellifera, a savannah woodland tree in sub‐Saharan Africa and southern Arabia. The loci were screened for polymorphism using 48 Kenyan individuals. Allelic diversity ranged from three to 19 per locus and the polymorphic information content varied from 0.287 to 0.893. These loci will be useful in studies of genetic structure, gene flow and breeding systems.     

Evaluating large‐scale effects of Bacillus thuringiensis var. israelensis on non‐biting midges (Chironomidae) in a eutrophic urban lake

1. There is continued interest in controlling non‐biting midges (chironomids) in some freshwaters because of the potential nuisance caused by emergent adults. One option is to apply Bacillus thuringiensis var. israelensis (Bti), primarily used against mosquitoes and blackflies, to control benthic chironomid larvae. Chironomids are also at risk of collateral effects where Bti is used on other targets, and such instances may increase if climate change alters the incidence of dipteran‐borne diseases. However, most research on Bti effects on chironomids is available from mesocosms or ponds and might not scale‐up to larger waterbodies. 2. We present results on the effects of Bti on larval chironomids from eight experimental treatments, over 3 years, on a newly created eutrophic, urban lake of 200 ha, Cardiff Bay. 3. The first two experimental years provided limited evidence of Bti effects, with chironomid densities reduced by up to 14%. Increased scale of application and altered experimental design in the third year revealed reductions in chironomid larval densities of around 35% following Bti treatment, with suppression lasting several months. 4. These large‐scale Bti experiments – among the largest ever undertaken on chironomids – complement previous smaller‐scale experiments in illustrating how field conditions and application methods influence Bti effects on chironomid densities. Specifically, the work reveals how near‐neutral buoyancy formulations of Bti can reduce chironomid numbers in large lakes exceeding 3 m depth, but only where treatment methods avoid over‐dispersion. We advocate further evaluation to assess whether chironomids can be suppressed over longer periods using whole‐lake application without long‐term ecological implications or excessive cost. We also suggest further consideration be given to non‐buoyant Bti formulations for use in deeper lakes.     

Y chromosome microsatellite markers identified from the tammar wallaby (Macropus eugenii) and their amplification in three other macropod species

Microsatellites were identified from three fully sequenced Y chromosome‐specific bacterial artificial chromosome clones from the tammar wallaby, Macropus eugenii. Ten microsatellites were genotyped in male tammar wallabies. Four loci were polymorphic with between two and six alleles per locus. Eleven different haplotypes were identified from 22 male tammar wallabies. No amplifications were obtained from female samples. Each microsatellite was also shown to amplify reliably in at least one other macropod species. These markers may therefore prove useful as some of the first male‐specific genetic markers for marsupials, with potential application to studies of male‐biased dispersal and mating systems.     

Increased plumage darkness of female Shiny Cowbirds Molothrus bonariensis in the subtropics: an adaptation to bacterial degradation?

The Shiny Cowbird Molothrus bonariensis is a sexually dichromatic species, in which males have blackish‐blue iridescence and females are dull brown. However, in some subtropical parts of its distribution, females show a plumage polymorphism that ranges from dull brown to dark brown and even black. Plumage melanization has been shown to protect feathers from bacterial degradation, decreasing the effects of harmful bacterial activity and thus plumage damage. In this study, we assessed whether bacterial feather‐degrading activity is acting as the selective force to increase darkness in the plumage of the female Shiny Cowbirds in Argentina. We compared the degradation of female Shiny Cowbird feathers belonging to different colour morphs when exposed to bacterial strains isolated from subtropical and temperate zones of its distribution, as well as to Bacillus licheniformis. We did not find differences in susceptibility to bacterial degradation between brown feathers and darker feathers. These results suggest that female plumage polymorphism in Shiny Cowbirds has not arisen as a defence against bacterial feather‐degrading activity.     

Using DNA Barcodes to Identify Bird Species Involved in Birdstrikes

Abstract: We determined effectiveness of using mitochondrial DNA barcodes (cytochrome c oxidase subunit 1 [CO1]) to identify bird-aircraft collision (birdstrike) cases that lacked sufficient feather evidence for morphological diagnosis. From September through December 2006, 821 samples from birdstrike events occurring in the United States were submitted for DNA analysis. We successfully amplified a CO1 DNA barcode product from 554 (67.5%) of the samples; 267 (32.5%) did not contain viable DNA and depended on morphological methods (microscopy) for Order or Family level identification. We deemed 19 cases inconclusive either because the DNA barcode recovered from the sample did not meet our 98% match criteria when compared to the Barcode of Life Database (BoLD) or because the DNA barcode matched to a set of ≥ 2 closely related species with overlapping barcodes, preventing complete species identification. Age of the sample (≤6 months) did not affect DNA viability, but initial condition of the sample and the collection method was critical to DNA identification success. The DNA barcoding approach has great potential in aiding in identification of birds (and wildlife) for airfield management practices, particularly in regions of the world that lack the vast research collections and individual expertise for morphologic identifications.