Functional heterogeneity within rhodamine123(lo) Hoechst33342(lo/sp) primitive hemopoietic stem cells revealed by pyronin Y

Objective. The aim of this study was to determine the function of primitive hematopoietic stem cells (PHSC) at phases G(0) and G(1) of the cell cycle. Materials and Methods. A combination of supravital dyes rhodamine123 (Rh), Hoechst33342 (Ho), and pyronin (PY) was used to isolate the G(0) and G(1) subsets of PHSC. A competitive repopulation assay was used to evaluate their in vivo function. Results. We confirmed that the Rh(lo)Lin(-)Kit(+)Sca-1(+) PHSC were relatively quiescent when compared with the more mature Rh(hi)Lin(-)Kit(+)Sca-1 HSC and Rh(hi)Lin(-)Kit(+)Sca-1(-) progenitors. In addition, cells with Rh(lo)Lin(-)Kit(+)Sca-1(+), Rh(lo)Ho(lo)Lin(-)Sca-1(+), or Rh(lo)Ho(sp)Lin(-)Sca-1(+) phenotypes identified the same cell population. We further subfractionated the Rh(lo)Ho(lo/sp)Lin(-)Sca-1(+) PHSC using PY into PYlo and PYhi subsets. Limiting dilution analysis revealed that the frequency of long-term in vivo competitive repopulating units (CRU) of the (PYRhHolo/sp)-Rh-lo-Ho-lo PHSC was 1 in 10 cells, whereas there was at least a three-fold lower frequency in those isolated at the G(1) phase (PYhi) We found a dose-dependent PY-mediated cytotoxicity that at moderate concentration affected most of the murine hematopoietic compartment but spared the early HSC compartment. Conclusion. Our data confirm that the HSC compartment is hierarchically ordered on the basis of quiescence and further extend this concept to PY-mediated cytotoxicity. PY supravital dye can be used to reveal functional heterogeneity within the (RhHolosp)-Ho-lo PHSC population but is of limited use in dissecting the relatively more mature hematopoietic stem/progenitor cell population. (C) 2001 International Society for Experimental Hematology. Published by Elsevier Science Inc.

Habitat requirements for the conservation of arboreal marsupials in dry sclerophyll forests of Southeast Queensland, Australia

The habitat requirements of arboreal marsupials were investigated in the dry sclerophyll forests of southeast Queensland, Australia. Species richness and abundance of arboreal marsupials was correlated to the proportion of total stand basal area occupied by lemon-scented gum (Corymbia citriodora), the height of the tallest trees, and density of hollow-bearing trees. The first two factors suggested that the most productive forests were also the most suitable habitats for arboreal marsupials. Importantly, the number of hollow-bearing trees was a significant factor in determining species richness and abundance of arboreal marsupials in this study, with the maximum number of species reached at sites containing greater than or equal to4 hollow-bearing trees/ha, and maximum abundance occurring at sites with :6 hollow-bearingtrees/ha. The proportion of C. citriodora was significant for the presence of the common brushtail possum (Trichosurus vulpecula), greater glider (Petauroides volans), and the yellow-bellied glider (Petaurus australis), while understory Acacia sp. density was important for the presence of the sugar glider (Petaurus breviceps). The yellow-bellied glider was also affected by two other variables: the density of hollow-bearing trees >50 cm diameter at breast height (dbh), and the time since the last logging. Current Codes of Practice regulating the density of hollow-bearing trees and silvicultural practices in state-owned timber production forests appear to provide adequate protection for arboreal marsupials, but the recently introduced increase in timber extraction rates within state forests may be detrimental to the animals. Also, protective prescriptions do not apply to the privately owned and leasehold estates, which contain the majority of the dry sclerophyll forests in southeast Queensland.

Testicular myxosporidiasis in anurans, with a description of Myxobolus fallax n. sp.

During studies of amphibian sperm cryopreservation, a new species of myxosporidean parasite (Myxozoa, Myxosporae) was observed in the testes of the Australian dwarf green tree frog Litoria fallax (Peters). Myxosporidiasis was found to have no affect on L. fallax body condition or sperm numbers. Myxobolus spores from L. fallax are morphologically distinct from Myxobolus hylae spores (infecting the sympatric Litoria aurea Lesson) and the three previously named (exotic to Australia) Myxobolus species found in anurans. Myxobolus fallax n. sp. is characterised by: pseudocyst white, spherical to ovoid, 141 x74 to 438 x337 mum in diameter (mature); plasmodium with spores loosely arranged within interior. Spores ovoid 13.4 +/- 0.5 (12.6-14.6) mum length, 9.5 +/- 0.4 (8.3-10.6) mum width, 6.8 +/- 0.4 (6.5-7.6) mum depth, 1.4 +/- 0.1 (1.3-1.6) length/width; polar capsules broadly pyriform and equal in size 4.2 +/- 0.3 (3.3-4.7) mum length, 2.4 +/- 0.2 (2.1-2.8) mum width; filament coils 7-8, wound tightly and perpendicular to the longitudinal axis of the capsule; polar filament 34 +/- 7.0 (18-50) mum length; intercapsular appendix and sutural ridge folds absent; and iodinophilous vacuole and mucous envelope lacking. In addition to this new species, data from archival samples of M. hylae are provided which show two morphologically distinct spore types. Both appeared rarely in the same pseudocysts and we cautiously retain the single species.

Growth of subtropical ECM fungi with different nitrogen sources using a new floating culture technique

Eight species of ectomycorrhizal (ECM) fungi in the genera Amanita. Gymnoboletus, Lactarius, and Russula were isolated from subtropical plant communities in eastem Australia. Two species were isolated from each of rainforest, Nothofagus forest, Eucalyptus forest, and Eucalyptus dominated wallum (heath) forest. These communities differ strongly in their soluble soil nitrogen (N) composition. The ability of the fungi to use inorganic (nitrate, ammonium) and organic (amide, peptide, protein) nitrogen sources was determined. As the fungi did not grow in liquid culture, a 'floating culture' technique was devised that allows hyphal growth on a screen floating on liquid medium. With some exceptions, fungal biomass production in floating culture closely reflected fungal growth on solid media assessed by total colony glucosamine content. Most isolates grown in floating culture had similar glucosamine concentrations on all N sources, with isolate specific concentrations ranging from 6 to 12 mug glucosamine g(-1) DW. However, Russula spp. had up to 1.7-fold higher glucosamine concentrations when growing with glutamine or ammonium compared to nitrate, glutathione or protein. Floating cultures supplied with 0.5, 1.5. 4.5, or 10 mm N mostly produced greatest biomass with 4.5 mM N. In vitro nitrate reductase activity (NRA) ranged from very low (0.03 mumol NO2- g(-1) fw h(-1)) in Russula sp. (wallum) to high (2.16 mumol NO2- g(-1) fw h(-1)) in Gymnoboletus sp. (rainforest) and mirrored the fungi's ability to use nitrate as a N source. All Russula spp. (wallum, Nothofagus and Eucalyptus forests), Lactarills sp, (rainforest) and.4manita sp. (wallum) utilized ammonium and glutamine but had little ability to use other N sources. In contrast,Amanita species (Nothofagus and Eucalyptus forests) grew on all N sources but produced most biomass with ammonium and glutamine. Only Gymnoboletus sp. (rainforest) showed similar growth with nitrate and ammonium as N sources. Fungal N source use was not associated with taxonomic groups, but is discussed in the context of soil N sources in the different habitats.

Monostephanostomum georgianum n. sp (Digenea : Acanthocolpidae) from Arripis georgianus (Valenciennes) (Perciformes : Arripidae) off Kangaroo Island, South Australia, with comments on Monostephanostomum Kruse, 1979 and Stephanostomum Looss, 1899

Monostephanostomum georgianum n. sp. is described from Arripis georgianus off Kangaroo Island, South Australia. It differs from its congeners by the presence of a short second row of oral spines. M. manteri Kruse, 1979 is reported from A. georgianus off southern Western Australia and Kangaroo Island, South Australia and A. trutta off northern Tasmania. It is considered that the other two species, M. yamagutii Ramadan, 1984 and M. krusei Reimer, 1983, should probably be removed from this genus. Two new combinations are formed, M. gazzae (Shen, 1990) n. comb. (from Stephanostomum) and M. mesospinosum (Madhavi, 1976) n. comb. (from Stephanostomum). A key to the four recognised species of Monostephanostomum is given.

Eucryphia honeys of Tasmania

A total of 22 honey samples from the west of Tasmania were analysed. All of them were monofloral leatherwood honeys containing between 79.9% and 99.1% of Eucryphia sp. pollen. Furthermore, 27 different pollen types were identified, the most frequent of which were Eucalyptus, Leptospermum (Myrtaceae), Trifolium (Fabaceae), Rosaceae, Banksia (Proteaceae), Lotus (Fabaceae), Bursaria (Pittosporaceae) and Brassica (Brassicaceae), which appeared in a great number of honeys with different percentages. The pollen spectrum characteristics of Tasmania's monofloral leatherwood honeys differentiate them from Chilean honeys of the same floral origin.

Solution Structures of the cis- and trans-Pro30 Isomers of a Novel 38-Residue Toxin from the Venom of Hadronyche Infensa sp. that Contains a Cystine-Knot Motif within Its Four Disulfide Bonds

The primary sequence and three-dimensional structure of a novel peptide toxin isolated from the Australian funnel-web spider Hadronyche infensa sp. is reported. ACTX-HI:OB4219 contains 38 amino acids, including eight-cysteine residues that form four disulfide bonds. The connectivities of these disulfide bonds were previously unknown but have been unambiguously determined in this study. Three of these disulfide bonds are arranged in an inhibitor cystine-knot (ICK) motif, which is observed in a range of other disulfide-rich peptide toxins. The motif incorporates an embedded ring in the structure formed by two of the disulfides and their connecting backbone segments penetrated by a third disulfide bond. Using NMR spectroscopy, we determined that despite the isolation of a single native homologous product by RP-HPLC, ACTX-HI:OB4219 possesses two equally populated conformers in solution. These two conformers were determined to arise from cis/trans isomerization of the bond preceding Pro30. Full assignment of the NMR spectra for both conformers allowed for the calculation of their structures, revealing, the presence of a triple-stranded antiparallel sheet consistent with the inhibitor cystine-knot (ICK) motif.

Dry matter production and allocation in Eucalyptus cloeziana and Eucalyptus argophloia seedlings in response to soil water deficits

Effects of soil water availability on seedling growth, dry matter production and allocation were determined for Gympie ( humid coastal) and Hungry Hills ( dry inland) provenances of Eucalyptus cloeziana F. Muell. and for E. argophloia Blakely ( dry inland) species. Seven-month-old seedlings were subjected to well-watered (100% field capacity, FC), moderate (70% FC) and severe (50% FC) soil water regimes in a glasshouse environment for 14 wk. There were significant differences in seedling growth, biomass production and allocation patterns between species. E. argophloia produced twice as much biomass at 100% FC, and more than three times as much at 70% and 50% FC than did either E. cloeziana provenance. Although the humid provenance of E. cloeziana had a greater leaf area at 100% FC conditions than did the dry provenance, total biomass production did not differ significantly. Both E. cloeziana provenances were highly sensitive to water deficits. E. argophloia allocated 10% more biomass to roots than did E. cloeziana. Allometric analyses indicated that relative biomass allocation patterns were significantly affected by genotype but not by soil water availability. These results have implications for taxon selection for cultivation in humid and subhumid regions.

Leaf water relations of Eucalyptus cloeziana and Eucalyptus argophloia in response to water deficit

Leaf water relations responses to limited water supply were determined in 7-month-old plants of a dry inland provenance of Eucalyptus argophloia Blakely and in a humid coastal provenance (Gympie) and a dry inland provenance (Hungry Hills) of Eucalyptus cloeziana F. Muell. Each provenance of E. cloeziana exhibited a lower relative water content at the turgor loss point, a lower apoplastic water content, a smaller ratio of dry mass to turgid mass and a lower bulk modulus of elasticity than the single provenance of E. argophloia. Osmotic potential at full turgor and water potential at the turgor loss point were significantly lower in E. argophloia and the inland provenance of E. cloeziana than in the coastal provenance of E. cloeziana. There was limited osmotic adjustment in response to soil drying in E. cloeziana, but not in E. argophloia. Between-species differences in water relations parameters were larger than those between the E. cloeziana provenances. Both E. cloeziana provenances maintained turgor under moderate water stress through a combination of osmotic and elastic adjustments. Eucalyptus argophloia had more rigid cell walls and reached lower water potentials with less reduction in relative water content than either of the E. cloeziana provenances, thereby enabling it to extract water from dryer soils.

Photosynthetic light and temperature responses of Eucalyptus cloeziana and Eucalyptus argophloia

Acclimation of gas exchange to temperature and light was determined in 18-month-old plants of humid coastal (Gympie) and dry inland ( Hungry Hills) provenances of Eucalyptus cloeziana F. Muell., and in those of a dry inland provenance of Eucalyptus argophloia Blakely. Plants were acclimated at day/night temperatures of 18/13, 23/18, 28/23 and 33/ 28 degreesC in controlled-temperature glasshouses for 4 months. Light and temperature response curves were measured at the beginning and end of the acclimation period. There were no significant differences in the shape and quantum-yield parameters among provenances at 23, 28 and 33 degreesC day temperatures. Quantum yield [mumol CO2 mumol(- 1) photosynthetic photon flux density (PPFD)] ranged from 0.04 to 0.06 and the light response shape parameter ranged from 0.53 to 0.78. Similarly, no consistent trends in the rate of dark respiration for plants of each provenance were identified at the four growth temperatures. Average values of dark respiration for the plants of the three provenances ranged from 0.61 to 1.86 mumol m(-2) s(-1). The optimum temperatures for net photosynthesis increased from 23 to 32 degreesC for the humid- and from 25 to 33 degreesC for the dry-provenance E. cloeziana and from 21 to 33 degreesC for E. argophloia as daytime temperature of the growth environment increased from 18 to 33 degreesC. These results have implications in predicting survival and productivity of E. cloeziana and E. argophloia in areas outside their natural distribution.

Dactylomyza gibsoni n.g., n. sp (Digenea: Opecoelidae) from Schuettea woodwardi (Waite) (Monodactylidae) from off Western Australia

An opecoelid digenean, Dactylomyza gibsoni n. g., n. sp. is described and figured from Schuettea woodwardi (Waite), a monodactylid from off the coast of Western Australia. The new genus conforms to the concept of the opecoelid subfamily Opecoelinae. The resemblance of the new genus to three other opecoelid genera, Pseudopecoeloides Yamaguti, 1940, Opecoeloides Odhner, 1928 and Poracanthium Dollfus, 1948, is discussed. Dactylomyza n. g. is distinguished from these morphologically similar worms on the basis of its median genital pore, ventral sucker appendages, uroproct and the absence of an accessory sucker. Pseudopecoeloides equesi Manter, 1947 is transferred to the new genus as Dactylomyza equesi (Manter, 1947) n. comb.

Bartoliella pritchardae n.g., n.sp. (Digenea: Opecoelidae) in Epinephelides armatus (Serranidae) from temperate marine waters of Western Australia

An opecoelid, Bartoliella pritchardae n. g., n. sp., is described from the intestine of Epinephelides armatus from Western Australia. The new genus has been assigned to the subfamily Opecoelininae Gibson & Bray, 1984, bringing the number of genera in the subfamily to two. Although the new genus is similar to Opecoelina Manter, 1934 morphologically, the pedunculate ventral sucker and complete absence of a cirrus-sac necessitate the erection of a new genus. A formal re-definition of the subfamily is given, based on the diagnoses of the genera Opecoelina and Bartoliella n. g.

Allopodocotyle skoliorchis n. sp. (Opecoelidae: Plagiorporinae) from Parequula melbournensis (Castelnau) (Gerreidae), a temperate marine fish in Australian waters

A new species of Allopodocotyle Pritchard, 1966 is described from the intestine and pyloric caeca of Parequula melbournensis (Gerreidae) caught from the waters off South and Western Australia. The new species is distinguished from other species by its larger eggs, broader form, pre-bifurcal genital pore and a number of other measurable features that are discussed. Of the species that share morphological similarities with Allopodocotyle skoliorchis n. sp., it is the only species known from a gerreid; all the other species are from serranids.

Pseudotrypanosoma elphinstonae sp n., a trichomonad symbiotic in Schedorhinotermes (Isoptera : Rhinotermitidae)

A new species of Pseudotrypanosoma, P. elphinstonae sp. n., is described which is symbiotic within the hindguts of the rhinotermitid termites Schedorhinotermes secundus and S. intermedius. P. elphinstonae possesses most of the features of the genus: 4 anterior flagella, prominent costa and recurrent flagellum forming an undulating membrane and simple bean-shaped parabasal body. The mastigont complex is of similar composition and arrangement to other trichomonads but the pelto-axostylar complex is greatly simplified being composed of a single layer of microtubules which do not over lap and cannot be distinguished into separate structures. The undulating membrane is much smaller than in related species; the costa is smaller and simpler and there are no microtubular bundles connecting it to the recurrent flagellum. Comparison of the ultrastructure of P. elphinstonae sp. n. with that of P. giganteum demonstrated that P. elphinstonae sp. n. in addition to being much smaller in size had a correspondingly simper ultrastructural organisation lacking several organelles which characterise the latter species.