Transpecific microsatellites for hard pines M. Shepherd, M. Cross, T. Maguire, M. Dieters, C. Williams, R. Henry

Microsatellites are difficult to recover from large plant genomes so cross-specific utilisation is an important source of markers. Fifty micro satellites were tested for cross-specific amplification and polymorphism to two New World hard pine species, slash pine (Pinus elliottii var. elliottii) and Caribbean pine (R caribaea var. hondurensis). Twenty-nine (58%) markers amplified in both hard pine species, and 23 of these 29 were polymorphic. Soft pine (subgenus Strobus) microsatellite markers did amplify, but none were polymorphic. Pinus elliottii var. elliottii and R caribaea var. hondurensis showed mutational changes in the flanking regions and the repeat motif that were informative for Pinus spp. phylogenetic relationships. Most allele length variation could be attributed to variability in repeat unit number. There was no evidence for ascertainment bias.

A comparative ultrastructural study of spermatozoa of the teiid lizards Cnemidophorus gularis gularis, Cnemidophorus ocellifer, and Kentropyx altamazonica (Reptilia, Squamata, Teiidae)

The ultrastructure of the spermatozoa of Cnemidophorus gularis gularis, Cnemidophorus ocellifer, and Kentropyx altamazonica is described for the first time. Mature spermatozoa of Cnemidophorus spp. and K. altamazonica differ in the occurrence of a perforatorial base plate, the enlargement of axonemal fibers 3 and 8, and shape of mitochondria. The comparisons of the ultrastructure sperm of Cnemidophorus spp. and K. altamazonica with Ameiva ameiva [J. Morphol. (2002) in press] suggest that Ameiva and Cnemidophorus are more similar to each other than either is to Kentropyx. Statistical analyses reveal that sperm of all three species studied are significantly different in the following dimensions: head, acrosome, distal centriole length, and nuclear shoulders width. There was no variable statistically different between the Cnemidophorus spp. only. The length of the tail, midpiece, entire sperm, and nuclear rostrum are significantly different between K. altamazonica and Cnemidophorus spp. Our results indicate that sperm ultrastructure presents intra and intergeneric variability. (C) 2002 Elsevier Science Ltd. All rights reserved.

Of mites and bees: A review of mite-bee associations in Australia and a revision of Raymentia Womersley (Acari : Mesostigmata : Laelapidae), with the description of two new species of mites from Lasioglossum (Parasphecodes) spp. (Hymenoptera : Halictidae)

Social bees have a diverse fauna of symbiotic mesostigmatic mites, including highly pathogenic parasites of the honeybee, but there are few reports of Mesostigmata phoretic on or inhabiting the nests of solitary or communal, ground-nesting bees. In south-eastern Australia, however, native bees in the family Halictidae carry what appears to be a substantial radiation of host-specific mesostigmatans in the family Laelapidae. Herein, we redescribe the obscure genus Raymentia , associated with Lasioglossum (Parasphecodes ) spp. bees (Halictidae) and describe two new species, R. eickwortiana from L. lacthium (Smith) and R. walkeriana from L. atronitens (Cockerell). The type species, R. anomala Womersley, is associated with L. altichum (Smith). In addition, we review the mites known to be associated with Australian bees, provide a key to differentiate them, and describe and illustrate acarinaria of the Halictinae. We also report on the first occurrences in Australia of the genera Trochometridium Cross (Heterostigmata: Trochometridiidae), from L. eremaean Walker (Halictidae), and Cheletophyes Oudemans (Prostigmata: Cheyletidae) from Xylocopa Latreille (Xylocopinae), and on the previously unknown association between a Neocypholaelaps Vitzthum (Mesostigmata: Ameroseiidae) and Lipotriches tomentifera (Friese) (Halictidae).

Do female rainbowfish (Melanotaenia spp.) prefer to shoal with familiar individuals under predation pressure?

Shoaling with familiar individuals may have many benefits including enhanced escape responses or increased foraging efficiency. This study describes the results of two complimentary experiments. The first utilised a simple binary choice experiment to determine if rainbowfish (Melanotaenia spp.) preferred to shoal with familiar individuals or with strangers. The second experiment used a free range situation where familiar and unfamiliar individuals were free to intermingle and were then exposed to a predator threat. Like many other small species of fish, rainbowfish were capable of identifying and distinguishing between individuals and choose to preferentially associate with familiar individuals as opposed to strangers. Contrary to expectations. however. rainbowrish did not significantly increase their preference for familiar individuals in the presence of a stationary predator model. Griffiths [J Fish Biol (1997) 51:489-4951 conducted similar studies under semi-natural conditions examining, the shoaling preferences of European minnows and showed similar results. Both the current study and that of Griffiths were conducted using predator wary populations of fish. It is suggested that, in predator sympatric populations, the benefits of shoaling with familiar individuals are such that it always pays to stay close to familiar individuals even when the probability If predator attack is remote.

Evolution of the secondary structure of the rRNA internal transcribed spacer 2 (ITS2) in hard ticks (Ixodidae, Arthropoda)

ITS2 sequences are used extensively in molecular taxonomy and population genetics of arthropods and other animals yet little is known about the molecular evolution of ITS2. We studied the secondary structure of ITS2 in species from each of the six main lineages of hard ticks (family Ixodidae). The ITS2 of these ticks varied in length from 679 bp in Ixodes scapularis to 1547 bp in Aponomma concolor. Nucleotide content varied also: the ITS2 of ticks from the Prostriata lineage (Ixodes spp.) had 46-49% GC whereas ITS2 sequences of ticks from the Metastriata lineage (all other hard ticks) had 61-62% GC. Despite variation in nucleotide sequence, the secondary structure of the ITS2 of all of these ticks apparently has five domains. Stems 1, 3, 4 and 5 of this secondary structure were obvious in all of the species studied. However, stem 2 was not always obvious despite the fact that it is flanked by highly conserved sequence motifs in the adjacent stems, stems 1 and 3. The ITS2 of hard ticks has apparently evolved mostly by increases and decreases in length of the nucleotide sequences, which caused increases, and decreases in the length of stems of the secondary structure. This is most obvious when stems of the secondary structures of the Prostriata (Ixodes spp.) are compared to those of the Metastriata (all other hard ticks). Increases in the size of the ITS2 may have been caused by replication slippage which generated large repeats, like those seen in Haemaphysalis humerosa and species from the Rhipicepalinae lineage, and the small repeats found in species from the other lineages of ticks.

Neuro-angiostrongylosis in wild Black and Grey-headed flying foxes (Pteropus spp)

Objective To identify nematodes seen in histological sections of brains of flying foxes (fruit bats) and describe the associated clinical disease and pathology. Proceedures Gross and histological examination of brains from 86 free-living flying foxes with neurological disease was done as part of an ongoing surveillance program for Australian bat lyssavirus. Worms were recovered, or if seen in histological sections, extracted by maceration of half the brain and identified by microscopic examination. Histological archives were also reviewed. Results There was histological evidence of angiostrongylosis in 16 of 86 recently submitted flying foxes with neurological disease and in one archival case from 1992. In 10 flying foxes, worms were definitively identified as Angiostrongylus cantonensis fifth-stage larvae. A worm fragment and third stage larvae were identified as Angiostrongylus sp, presumably A cantonensis, in a further three cases. The clinical picture was dominated by paresis, particularly of the hind-limbs, and depression, with flying foxes surviving up to 22 days in the care of wildlife volunteers. Brains containing fifth-stage larvae showed a moderate to severe eosinophilic and granulomatous meningoencephalitis (n = 14), whereas there was virtually no inflammation of the brains of bats which died when infected with only smaller, third-stage larvae (n = 3). There was no histological evidence of pulmonary involvement. Conclusion This is the first report of the recovery and identification of A cantonensis from free-living Australian wildlife. While anglostrongylosis is a common cause of paresis in flying foxes, the initial clinical course cannot be differentiated from Australian bat lyssavirus infection, and wildlife carers should be urged not to attempt to rehabilitate flying foxes with neurological disease.

Development and evaluation of a species diagnostic polymerase chain reaction-restriction fragment-length polymorphism procedure for cryptic members of the Culex sitiens (Diptera : Culicidae) subgroup in Australia and the southwest Pacific

Members of the Culex sitiens subgroup are important vectors of arboviruses, including Japanese encephalitis virus, Murray Valley encephalitis virus and Ross River virus. Of the eight described species, Cx. annulirostris Skuse, Cx. sitiens Wiedemann, and Cx. palpalis Taylor appear to be the most abundant and widespread throughout northern Australia and Papua New Guinea (PNG). Recent investigations using allozymes have shown this subgroup to contain cryptic species that possess overlapping adult morphology. We report the development of a polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP) procedure that reliably separates these three species. This procedure utilizes the sequence variation in the ribosomal DNA ITS1 and demonstrates species-specific PCR-RFLP profiles from both colony and field collected material. Assessment of the consistency of this procedure was undertaken on mosquitoes sampled from a wide geographic area including Australia, PNG, and the Solomon Islands. Overlapping adult morphology was observed for Cx. annulirostris and Cx. palpalis in both northern Queensland and PNG and for all three species at one site in northwest Queensland.

Mitochondrial genomes of parasitic flatworms

Complete or near-complete mitochondrial genomes are now available for 11 species or strains of parasitic flatworms belonging to the Trematoda and the Cestoda. The organization of these genomes is not strikingly different from those of other eumetazoans, although one gene (atp8) commonly found in other phyla is absent from flatworms. The gene order in most flatworms has similarities to those seen in higher protostomes such as annelids. However, the gene order has been drastically altered in Schistosoma mansoni, which obscures this possible relationship. Among the sequenced taxa, base composition varies considerably, creating potential difficulties for phylogeny reconstruction. Long non-coding regions are present in all taxa, but these vary in length from only a few hundred to similar to10 000 nucleotides. Among Schistosoma spp., the long non-coding regions are rich in repeats and length variation among individuals is known. Data from mitochondrial genomes are valuable for studies on species identification, phylogenies and biogeography.

Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs

Only a small proportion of the mouse genome is transcribed into mature messenger RNA transcripts. There is an international collaborative effort to identify all full-length mRNA transcripts from the mouse, and to ensure that each is represented in a physical collection of clones. Here we report the manual annotation of 60,770 full-length mouse complementary DNA sequences. These are clustered into 33,409 'transcriptional units', contributing 90.1% of a newly established mouse transcriptome database. Of these transcriptional units, 4,258 are new protein-coding and 11,665 are new non-coding messages, indicating that non-coding RNA is a major component of the transcriptome. 41% of all transcriptional units showed evidence of alternative splicing. In protein-coding transcripts, 79% of splice variations altered the protein product. Whole-transcriptome analyses resulted in the identification of 2,431 sense-antisense pairs. The present work, completely supported by physical clones, provides the most comprehensive survey of a mammalian transcriptome so far, and is a valuable resource for functional genomics.

On the intrinsic capacity for increase of Australian flying-foxes (Pteropus spp., Megachiroptera)

In the reproductive biology of organisms, a continuum exists from "highly reproductive species" at one end to "survivor species" at the other end. Among other factors, the position of a species along this continuum affects its sensitivity to human exploitation and its vulnerability to extinction. Flying foxes are long-lived, seasonal breeders, with a rigid, well-defined breeding season that is largely or wholly genetically determined. Unlike opportunistic, highly reproductive species, such as rabbits or mice, female flying foxes are unable to produce viable young before their second or third year of life, and are then capable of producing just one young per year. Such a breeding strategy will be successful only if flying-foxes are long-lived and suffer naturally low mortality rates. In this paper, we assess the vulnerability of flying foxes to extinction, using basic parameters of reproduction observed in the wild, and in captive breeding colonies of P. poliocephalus, P. alecto and P. scapulatus, and survival rates that are likely to apply to Australian conditions. Our models show explicitly that flying-fox populations have a very low capacity for increase, even under the most ideal conditions. The implications of our models are discussed in reference to the long-term management and conservation needs of Australian flying foxes. We conclude that current death-rates of flying-foxes in NSW and Queensland fruit orchards are putting state populations at serious risk.

Habitat-predator association and avoidance in rainbowfish (Melanotaenia spp.)

The ability to recall the location of a predator and later avoid it was tested in nine populations of rainbowfish (Melanotaenia spp.), representing three species from a variety of environments. Following the introduction of a model predator into a particular microhabitat, the model was removed, the arena rotated and the distribution of the fish recorded again. In this manner it could be determined what cues the fish relied on in order to recall the previous location of the predator model. Fish from all populations but one (Dirran Creek) were capable of avoiding the predator by remembering either the location and/or the microhabitat in which the predator was recently observed. Reliance on different types of visual cues appears to vary between populations but the reason for this variation remains elusive. Of the ecological variables tested (flow variability, predator density and habitat complexity), only the level of predation appeared to be correlated with the orientation technique employed by each population. There was no effect of species identity, which suggests that the habitat that each population occupies plays a strong role in the development of both predator avoidance responses and the cues used to track predators in the wild.

Diseases of tunas, Thunnus spp.

Much is known about those aspects of tuna health which can be studied in wild populations, e.g. helminth parasites. However, because aquaculture of these species is in its infancy, knowledge of microbial, nutritional and environmental diseases is limited. This review is an attempt to bring together the available information on those diseases of Thunnus spp. which cause significant morbidity, mortality or economic loss. In doing so it has become clear that much more research needs to be undertaken on the physiology of the species (southern, northern and Pacific bluefin tuna) currently used in aquaculture in order for the pathogenesis of some conditions to be properly understood. Attempts at hatchery culture of Pacific bluefin tuna has indicated that Thunnus spp. will be problematic to hatch and propagate.

DNA vaccine coding for the full-length infectious Kunjin virus RNA protects mice against the New York strain of West Nile virus

A plasmid DNA directing transcription of the infectious full-length RNA genome of Kunjin (KUN) virus in vivo from a mammalian expression promoter was used to vaccinate mice intramuscularly. The KUN viral cDNA encoded in the plasmid contained the mutation in the NS1 protein (Pro-250 to Leu) previously shown to attenuate KUN virus in weanling mice. KUN virus was isolated from the blood of immunized mice 3-4 days after DNA inoculation, demonstrating that infectious RNA was being transcribed in vivo; however, no symptoms of virus-induced disease were observed. By 19 days postimmunization, neutralizing antibody was detected in the serum of immunized animals. On challenge with lethal doses of the virulent New York strain of West Nile (WN) or wild-type KUN virus intracerebrally or intraperitoneally, mice immunized with as little as 0.1-1 mug of KUN plasmid DNA were solidly protected against disease. This finding correlated with neutralization data in vitro showing that serum from KUN DNA-immunized mice neutralized KUN and WN,viruses with similar efficiencies. The results demonstrate that delivery of an attenuated but replicating KUN virus via a plasmid DNA vector may provide an effective vaccination strategy against virulent strains of WN virus.

Description of Paeon asymboli n. sp. (Copepoda : Sphyriidae), parasitic on Asymbolus spp. (catsharks) and a new host record for P-australis Kabata, 1993.

Paeon asymboli n. sp. ( Copepoda: Sphyriidae) is described and illustrated from two female specimens taken from the gills of a grey spotted catshark Asymbolus analis ( Ogilby) and an orange spotted catshark A. rubiginosus ( Last, Gomon & Gledhill) ( Scyliorhinidae) from off southeastern Queensland, Australia. The key features for identification are: a pear- shaped trunk, longer than it is wide, along with a cephalothorax characterised by two large ventral papillae, projecting laterally and supporting a number of secondary lobes; a single mid- line, subconical papilla located antero- dorsal to the ventral papillae; an anterior surface bearing two prominent stalked papillae; and an absence of posterolateral lobular processes. P. australis Kabata, 1993 is recorded for the first time from the eastern shovelnose ray Aptychotrema rostrata ( Shaw & Nodder) ( Rhinobatidae).