Bacteriophages from the forestomachs of Australian marsupials.

Bacteriophages were observed in forestomach contents from three species of Australian macropodoid marsupials possessing a foregut fermentative digestion: the eastern grey kangaroo (Macropus giganteus), the eastern wallaroo (Macropus robustus robustus), and the rufous bettong (Aepyprymnus rufescens). Forty-six morphologically distinct phage types, representing the families Myoviridae, Siphoviridae, and Podoviridae, were identified. The range of forms varied between host species. The greatest diversity of phage types was found in forestomach contents of the wallaroo, and few phage types were recorded from the rufous bettongs. It is concluded that macropodoid marsupials, in common with their eutherian counterparts, possess diverse populations of bacteriophages in their fermentative forestomachs.     

Bacteriophages from the forestomachs of Australian marsupials.

Bacteriophages were observed in forestomach contents from three species of Australian macropodoid marsupials possessing a foregut fermentative digestion: the eastern grey kangaroo (Macropus giganteus), the eastern wallaroo (Macropus robustus robustus), and the rufous bettong (Aepyprymnus rufescens). Forty-six morphologically distinct phage types, representing the families Myoviridae, Siphoviridae, and Podoviridae, were identified. The range of forms varied between host species. The greatest diversity of phage types was found in forestomach contents of the wallaroo, and few phage types were recorded from the rufous bettongs. It is concluded that macropodoid marsupials, in common with their eutherian counterparts, possess diverse populations of bacteriophages in their fermentative forestomachs.     

Divergent neuropeptide evolutionary drifts between American and Australian marsupials

Present-day marsupials, which are supposed to have arisen from a single stem diverging from the placental stem some 130 million years ago, exist only in the American and Australian continents. Comparison of the homologous genes and their protein products, which evolved under different environmental conditions, may provide arguments for either selective or neutral evolution. In contrast to Australian Macropodidae, which have pecuIiar neurohypophysial peptides, namely mesotocin and two pressor peptides, lysine vasopressin and phenypressin, the South American oppossum,Didelpbis marsupialis, has oxytocin, lysine vasopressin, and arginine vasopressin. Because placental mammals have oxytocin and usually arginine vasopressin, and nonmammalian tetrapods have mesotocin and arginine vasotocin, it is assumed that (1) selective change of arginine vasotocin into arginine vasopressin occurred in mammalian ancestors and a subsequent gene duplication in the marsupial line gave rise to two pressor peptides with divergent neutral drifts in American and Australian groups, and (2) mesotocin of nonmammalian tetrapods has been preserved in Australian marsupials and reclaimed for milk-ejecting function whereas it has been converted into oxytocin in South American oppossums. The change of mesotocin into oxytocin seems neutral rather than selective.     

Phylogenetic analysis of Porphyromonas species isolated from the oral cavity of Australian marsupials

Porphyromonas species are frequently isolated from the oral cavity and are associated with periodontal disease in both animals and humans. Black, pigmented Porphyromonas spp. isolated from the gingival margins of selected wild and captive Australian marsupials with varying degrees of periodontal disease (brushtail possums, koalas and macropods) were compared phylogenetically to Porphyromonas strains from non-marsupials (bear, wolf, coyote, cats and dogs) and Porphyromonas gingivalis strains from humans using 16S rRNA gene sequence analysis. The results of the phylogenetic analysis identified three distinct groups of strains. A monophyletic P. gingivalis group (Group 1) contained only strains isolated from humans and a Porphyromonas gulae group (Group 2) was divided into three distinct subclades, each containing both marsupial and non-marsupial strains. Group 3, which contained only marsupial strains, including all six strains isolated from captive koalas, was genetically distinct from P. gulae and may constitute a new Porphyromonas species.     

Comparison of the transition states for folding of two Ig-like proteins from different superfamilies

In the fold approach proteins with a similar fold but different sequences are compared in order to investigate the relationship between native state structure and folding behaviour. Here we compare the properties of the transition states for folding of TI I27, the 27th immunoglobulin domain from human cardiac titin, and that of TNfn3, the third fibronectin type III domain from human tenascin. Experimental phi-values were used as restraints in molecular dynamics simulations to determine the structures that make up the transition state ensembles (TSEs) for folding of the two proteins. The restrained simulations that we present allow a detailed structural comparison of the two TSEs to be made. Further calculations show explicitly that for both proteins the formation of the interactions involving the residues in the folding nucleus is sufficient for the establishment of the topology of the Ig-like fold. We found that, although the folding nuclei of the two proteins are similar, the packing of the folding nucleus of TI I27 is much tighter than that of TNfn3, reflecting the higher experimental phi-values and beta(T) (Tanford Beta) of TI I27. These results suggest that the folding nucleus can be significantly deformed to accommodate extensive sequence variation while conserving the same folding mechanism.     

Mammalian genome evolution: new clues from comparisons of eutherians, marsupials and monotremes.

1. Comparisons of chromosomes and gene maps of different mammals are yielding a big picture of the evolution of mammalian genome form and function. It has been particularly instructive to compare gene arrangements on the sex chromosomes between the three major groups of mammals. Eutheria (so-called placental mammals). Metatheria (marsupials) and Prototheria (monotremes), which diverged 150 and 170 Myr BP respectively. 2. A region amounting to 3% of the haploid genome is located on the X chromosome in all three groups, implying that this region must have been part of the original X in a common ancestor. This region comprises the long arm of the human X. 3. A region represented by the short arm of the human X is common to the X in all eutherians, but is autosomal in marsupials and monotremes; thus it was not a part of the original X, and must have been acquired by the X early in the eutherian radiation. 4. This recently acquired region was probably translocated to a pseudoautosomal region shared by the eutherian X and Y. Thus it was originally paired and exempt from X chromosome inactivation; stepwise deletion of this region from the Y and recruitment of the newly unpaired region of the X into the inactivation system could account for some of the peculiarities of this region of the human X. 5. The sex-determining gene TDF must lie on the Y in all mammals in which the Y is male determining. The autosomal location of the candidate gene ZFY in marsupials and monotremes eliminates it from consideration. The recently described candidate gene SRY has yet to pass the "marsupial test".     

Observations on the vasculature of the reproductive tract in some Australian marsupials

The origin, distribution and structure of the blood vessels of the female reproductive tract and the testis of the brush possum (Trichosurus vulpecula) were studied using latex and silicone rubber casting and histological techniques. Latex casts of the vessels of the female tract were also studied in five macropod species – Macropus giganteus, M. eugenii, M. agilis, Megaleia rufa and Thylogale billardierii, and in the common wombat (Vombatus ursinus). The female reproductive tract in the brush possum was supplied and drained by four major sets of paired vessels – ovarian, cranial urogenital, caudal urogenital, and internal pudendal arteries and veins. These vessels formed substantial anastomoses with one another on each side of the midline, and also across-the-midline anastomoses. The proximal part of the ovarian artery ran in close apposition to the ovarian vein, which received one or more large uterine branches. In its distal protion the ovarian artery gave rise to a leash of small, tortuous ovarian branches, which wound around and between the plexiform ovarian veins. The testicular arteries and veins in this species also ran in close apposition to one another. Both arteries and veins branched into many smaller, mildly tortuous, parallel vessels in the spermatic cord, which reunited before entering the testis. The blood vessels of the reproductive tract in all of the macropod species studied, and in the common wombat, were basically similar to those of the brush possum. The intimate structural relationships between ovarian arteries and veins, and their ovarian branches, in these marsupials are suggestive of specializations for counter-current exchange between venous and arterial blood. However, in contrast to those of the testicular vessels where heat exchange is a demonstrated function, their physiological significance remains unknown.     

Extrinsic versus intrinsic factors in the decline and extinction of Australian marsupials

Recent attempts to explain the susceptibility of vertebrates to declines worldwide have largely focused on intrinsic factors such as body size, reproductive potential, ecological specialization, geographical range and phylogenetic longevity. Here, we use a database of 145 Australian marsupial species to test the effects of both intrinsic and extrinsic factors in a multivariate comparative approach. We model five intrinsic (body size, habitat specialization, diet, reproductive rate and range size) and four extrinsic (climate and range overlap with introduced foxes, sheep and rabbits) factors. We use quantitative measures of geographical range contraction as indices of decline. We also develop a new modelling approach of phylogenetically independent contrasts combined with imputation of missing values to deal simultaneously with phylogenetic structuring and missing data. One extrinsic variable-geographical range overlap with sheep-was the only consistent predictor of declines. Habitat specialization was independently but less consistently associated with declines. This suggests that extrinsic factors largely determine interspecific variation in extinction risk among Australian marsupials, and that the intrinsic factors that are consistently associated with extinction risk in other vertebrates are less important in this group. We conclude that recent anthropogenic changes have been profound enough to affect species on a continent-wide scale, regardless of their intrinsic biology.     

Thermogenic capabilities of the opossum Monodelphis domestica when warm and cold acclimated: similarities between American and Australian marsupials

1. Monodelphis domestica is a small marsupial mammal from South America. Its thermogenic abilities in the cold were determined when the opossums were both warm (WA) and cold (CA) acclimated. Maximum heat production of M. domestica was obtained at low temperatures in helium-oxygen. 2. Basal metabolic rate (BMR) in the WA animals was 3.2 W/kg and mean body temperature was 32.6 degrees C at 30 degrees C. These values were lower than those generally reported for marsupials. Nevertheless, these M. domestica showed considerable metabolic expansibility in response to cold. Sustained (summit) metabolism was 8-9 times BMR, while peak metabolism was 11-13 times BMR. These maximum values were equal to, or above, those expected in small placentals. 3. Cold acclimation altered the thermal responses of M. domestica, particularly in warm TaS. However, summit metabolism was not significantly increased; nor did M. domestica show a significant thermogenic response to noradrenaline, which in many small placentals elicits non-shivering thermogenesis. The thermoregulatory responses of this American marsupial were, in most aspects, similar to those of Australian marsupials. This suggests that the considerable thermoregulatory abilities of marsupials are of some antiquity.     

Genomic organization and cloning of novel genes encoding toxin-like peptides of three superfamilies from the spider Orinithoctonus huwena

The bird spider Ornithoctonus huwena is one of the most venomous spiders in China. Its venom is a mixture of various compounds with diverse bioactivities. Ninety proteins and 47 peptides have been identified, and 67 cDNA sequences encoding different toxin precursors have been cloned. However, the genomic DNA of them is seldom reported. To characterize the genomic DNA structure of huwentoxins, the genomic DNA encoding toxins of three superfamilies were cloned by using sequence specific or partially degenerate primers based on their cDNA sequences. An unexpected finding was that the intron was lacking in the genomic sequences of three superfamilies. The genomic DNA information has predictive value for better understanding the relationship of spider toxin evolution. In addition, we have cloned and analyzed 19 novel genes encoding toxin-like precursors by using the genomic DNA of the spider O. huwena.     

Luminescence from the yeast Candida utilis and comparisons across three genera

Weak luminescence was detected from oxygenated liquid cultures of the yeast Candida utilis during two stages of its growth cycle. The first period of emission occurred during the exponential phase of growth and comprised an ultraviolet band (270-390 nm; ca 19 photons s^{? 1} cm^?2 of culture surface) and a visible band (450-620 nm; ca 68 photons s^{? 1} cm^{? 2}). The second period of emission occurred late in the stationary phase of growth and was comprised almost entirely of a visible region band (450-620 nm; 6.8 × 10² photons s^{? 1} cm^{? 2}). No luminescence was observed when the yeast was grown anaerobically. These observations are compared with those previously obtained for two other yeasts, Saccharomyces cerevisiae and Schizosaccharomyces pombe. The ratios of the intensities of blue/red emissions in the stationary phase luminescences correlated with the ratio of the saturated/unsaturated lipid content for the three yeasts. This result provided further support for the claim that the stationary phase luminescence arises from the reactions associated with lipid peroxidation. A number of previously suggested sources of the exponential phase luminescence are discussed and rejected. Oxidative side reactions accompanying protein synthesis remain a possible source of that emission.     

Mitochondrial DNA polymorphism within and among species of Capillaria sensu lato from Australian marsupials and rodents

The nucleotide variation in a mitochondrial DNA (mtDNA) fragment within and among species of Capillaria sensu lato from Australian marsupials and rodents was analyzed using a mutation scanning/sequencing approach. The fragment of the cytochrome c oxidase subunit I (COI) was amplified by PCR from parasite DNA, and analysed by single-strand conformation polymorphism (SSCP) and sequencing. There was no significant variation in SSCP profiles within a morphospecies from a particular host species, but significant variation existed among morphospecies originating from different host species. The same morphospecies was found to occur in 1-3 tissue habitats within one host individual or within different individuals of a particular species of host from the same or different geographical areas, and morphospecies appeared to be relatively host specific at the generic level. The results indicated that the species of Capillaria sensu lato examined, although highly variable in their host and tissue specificity, may exhibit the greatest degree of specificity at the level of host genus.     

Widespread occurrence in Australian marsupials of neutralizing antibodies to a herpesvirus from a parma wallaby.

Serum neutralizing antibodies to a parma wallaby herpesvirus (PWHV) have been detected in a wide range of marsupials from different locations across Australia, including several islands. A study of 242 animals (mostly macropods) sampled in the wild showed that 23% had antibodies; a significantly higher frequency (41%) of 116 animals in captivity had antibodies, which were generally at higher levels than those of animals in the wild. Antibodies to PWHV were also detected among parma wallabies from a colony on Kawau Island, New Zealand. The highest antibody levels were found in a group of captive tammar wallabies during recurrent outbreaks of clinical infection. It is suggested that PWHV has evolved along with a marsupial host, and that the high antibody levels among captive animals reflects ease of virus transmission due to crowding, or to conditions of stress leading to expression of latent virus.     

Diversity of color vision: not all Australian marsupials are trichromatic

Color vision in marsupials has recently emerged as a particularly interesting case among mammals. It appears that there are both dichromats and trichromats among closely related species. In contrast to primates, marsupials seem to have evolved a different type of trichromacy that is not linked to the X-chromosome. Based on microspectrophotometry and retinal whole-mount immunohistochemistry, four trichromatic marsupial species have been described: quokka, quenda, honey possum, and fat-tailed dunnart. It has, however, been impossible to identify the photopigment of the third cone type, and genetically, all evidence so far suggests that all marsupials are dichromatic. The tammar wallaby is the only Australian marsupial to date for which there is no evidence of a third cone type. To clarify whether the wallaby is indeed a dichromat or trichromatic like other Australian marsupials, we analyzed the number of cone types in the "dichromatic" wallaby and the "trichromatic" dunnart. Employing identical immunohistochemical protocols, we confirmed that the wallaby has only two cone types, whereas 20-25% of cones remained unlabeled by S- and LM-opsin antibodies in the dunnart retina. In addition, we found no evidence to support the hypothesis that the rod photopigment (rod opsin) is expressed in cones which would have explained the absence of a third cone opsin gene. Our study is the first comprehensive and quantitative account of color vision in Australian marsupials where we now know that an unexpected diversity of different color vision systems appears to have evolved.     

Comparative DNA analysis of three South American marsupials.

Published information on marsupials DNA is limited to a group of species belonging to only one genus. No previous reports have been written on South American species. In this paper we characterize the DNA of three out of the four marsupials found in Uruguay. Analytical and preparative ultracentrifugations in neutral CsCl gradients, including four intercalating agents and in Cs2SO4 gradients in presence of increasing amounts of Hg++ ion did not allow us to separate any satellite fraction. The buoyant density of the unique peak measured in CsCl gradients was in every case 1.697 g/cc with a G-C content of 37.7%. Digestion of total DNA with 11 restriction endonucleases produced a different pattern of bands for the three species, although some possible homologies could be established. Hybridization with 32P-rRNA of Southern blots of the gels containing digested DNAs demonstrated that the repeated sequences evidenced do not correspond to the ribosomal cistrons.