Utility of mitochondrial DNA sequences from four gene regions for systematic studies of Australian freshwater crayfish of the Genus Cherax (Decapoda: Parastacidae)

One of the most important requirements for systematic and phylogenetic studies is the identification of gene regions with the appropriate level of variation for the question of interest. Molecular phylogenetic and systematic studies of freshwater crayfish have made use of DNA sequences mainly from ribosomal genes, especially the 16S rRNA gene region. Thus, little information is available on other potentially useful mitochondrial gene regions for systematic studies in these animals. In this study, we look at nucleotide variation and phylogenetic relations within and between four species of freshwater crayfish of the genus Cherax from the southwest of Western Australia using four fragments amplified from the 16S rRNA, 12S rRNA, Cytochrome Oxidase I (COI), and Cytochrome b (Cyt b) gene regions. Samples of Engaeus strictifrons, Euastacus bispinosus, and Geocharax falcata were also sequenced for comparative purposes. The size of the fragments varied from 358 bp to 600 bp. Across all samples, the four fragments showed significant phylogenetic signal and showed similar proportions of variable sites (28.81–37.33%). Average divergence within species for the mitochondrial gene regions varied from 1.18% to 4.91%, with the 16S rRNA being the least variable and Cyt b the most variable. Average divergence between species ranged 7.63–15.53%, with 16S rRNA being the least variable and COI the most variable. At the generic level, average divergence ranged 17.21–23.82%. Phylogenetic analyses of the 16S rRNA, 12S rRNA, and COI regions generated four clades consistent with the presence of four species previously identified on the basis of allozyme and morphological studies. The relationships among samples were largely congruent across the data set, although some relationships remained unresolved. Not all samples could be amplified using the Cyt b primers, and some of those that were showed quite anomalous relationships, suggesting that one or more Cyt b pseudogenes were being amplified.

Complete mitochondrial DNA sequences of the decapod crustaceans pseudocarcinus gigas (Menippidae) and macrobrachium rosenbergii (Palaemonidae)

The complete mitochondrial DNA sequence was determined for the Australian giant crab Pseudocarcinns gigas (Crustacea: Decapoda: Menippidae) and the giant freshwater shrimp Macrobrachium rosenbergii (Crustacea: Decapoda: Palaemonidae). The Pse gigas and Mrosenbergii mitochondrial genomes are circular molecules, 15,515 and 15,772 bp in length, respectively, and have the same gene composition as found in other metazoans. The gene arrangement of M. rosenbergii corresponds with that of the presumed ancestral arthropod gene order, represented by Limulus polyphemus, except for the position of the tRNA^Leu(UUR) gene. The Pse. gigas gene arrangement corresponds exactly with that reported for another brachyuran, Portunus trituberculatus, and differs from the M. rosenbergii gene order by only the position of the tRNA^His gene. Given the relative positions of intergenic nonoding nucleotides, the “duplication/random loss” model appears to be the most plausible mechanism for the translocation of this gene. These data represent the first caridean and only the second brachyuran complete mtDNA sequences, and a source of information that will facilitate surveys of intraspecific variation within these commercially important decapod species.

Does behavior reflect phylogeny in swiftlets (Aves: Apodidae)? A test using cytochrome b mitochondrial DNA sequences

Swiftlets are small insectivorous birds, many of which nest in caves and are known to echolocate. Due to a lack of distinguishing morphological characters, the taxonomy of swiftlets is primarily based on the presence or absence of echolocating ability, together with nest characters. To test the reliability of these behavioral characters, we constructed an independent phylogeny using cytochrome b mitochondrial DNA sequences from swiftlets and their relatives. This phylogeny is broadly consistent with the higher classification of swifts but does not support the monophyly of swiftlets. Echolocating swiftlets (Aerodramus) and the nonecholocating "giant swiftlet" (Hydrochous gigas) group together, but the remaining nonecholocating swiftlets belonging to Collocalia are not sister taxa to these swiftlets. While echolocation may be a synapomorphy of Aerodramus (perhaps secondarily lost in Hydrochous), no character of Aerodramus nests showed a statistically significant fit to the molecular phylogeny, indicating that nest characters are not phylogenetically reliable in this group.

Extensible motif and aberrant nucleotide extraction in genomic sequences

The project focusses on the discovery of conserved DNA sequences in bacterial genomes and comparative analysis of bacterial genomes to elicit evolutionary trends. The outcomes have produced novel techniques for modelling motifs in DNA and the characterisation of evolutionary processes in medically significant bacterial pathogens.

Macrocyclic Zn complexes for DNA detection

Researchers from Monash University have developed an electrocatalytic method based on a charge transport through DNA films, which allows detection of complementary over non-complementary and mismatched DNA sequences in fully hybridized duplexes.

EST-PAC HPC - a web portal for high-throughput EST annotation and protein sequence prediction

Expressed Sequence Tags (ESTs) are short DNA sequences generated by sequencing the transcribed cDNAs coming from a gene expression. They can provide significant functional, structural and evolutionary information and thus are a primary resource for gene discovery. EST annotation basically refers to the analysis of unknown ESTs that can be performed by database similarity search for possible identities and database search for functional prediction of translation products. Such kind of annotation typically consists of a series of repetitive tasks which should be automated, and be customizable and amenable to using distributed computing resources. Furthermore, processing of EST data should be done efficiently using a high performance computing platform. In this paper, we describe an EST annotator, EST-PACHPC, which has been developed for harnessing HPC resources potentially from Grid and Cloud systems for high throughput EST annotations. The performance analysis of EST-PACHPC has shown that it provides substantial performance gain in EST annotation.

Effects of alterations of primer-binding site sequences on human immunodeficiency virus type 1 replication

The human immunodeficiency virus type 1 genomic RNA primer-binding site (PBS) sequence comprises 18 nucleotides which are complementary to those at the 3' end of the replication initiation primer tRNA(3Lys). To investigate the role of the PBS in viral replication, we either deleted the original wild-type PBS (complementary to tRNA(3Lys) or replaced it with DNA sequences complementary to either tRNA(1,2Lys) or tRNA(Phe). Transfection of COS cells with such molecular constructs yielded similar levels of viral progeny that were indistinguishable with regard to viral proteins and tRNA content. Virus particles derived from PBS-deleted molecular clones were noninfectious for MT-4, Jurkat, and CEM-T4 cells. However, infectious viruses were derived from constructs in which the PBS had been altered to sequences complementary to either tRNA(1,2Lys) or tRNA(Phe), although mutated forms showed significant lags in replication efficiency in comparison with wild types. Molecular analysis of reverse-transcribed DNA in cells infected by the mutated viruses indicated that both tRNA(1,2Lys) and tRNA(Phe) could function as primers for reverse transcription during the early stages of infection. Sequencing of full-length proviral DNA, obtained 6 days after infection, revealed the mutated PBS, indicating that a complete cycle of reverse transcription had occurred. During subsequent rounds of infection, reversion of the mutated PBS to wild-type sequences was observed, accompanied by increased production of viral gene products. Reversion to wild-type PBS sequences was confirmed both by specific PCR analysis, using distinct primer pairs, and by direct sequencing of amplified segments. We also performed endogenous in vitro reverse transcription experiments in which synthesis of minus-strand strong-stop viral DNA was primed from a synthetic RNA template containing a PBS complementary to various tRNA isoacceptors. These results showed that tRNA(3Lys) was a much more efficient primer of such reactions than either tRNA(1,2Lys) or tRNA(Phe).

Molecular phylogeny of the Cheilodactylidae and Latridae (Perciformes: Cirrhitoidea) with notes on taxonomy and biogeography

Phylogenetic relationships among cheilodactylid and latrid fishes were estimated from cytochrome oxidase I and cytochrome b mitochondrial DNA sequences. Two South African cheilodactylids, Cheilodactylus fasciatus and Cheilodactylus pixi, were divergent from the remaining members of their genus and family, and the monophyly of these groups was rejected based on parametric bootstrap analysis. As C. fasciatus is the nominal species for the genus and family, widespread taxonomic reassignment is implicated for the remaining 12 and 17 members of these groups, respectively. As these 17 cheilodactylids are not genetically or morphologically distinct from the latrids, it is proposed that the Latridae should be expanded to encompass them. The inferred relationships among those Cheilodactylus requiring generic reassignment were largely unresolved, and hence few recommendations can be made regarding their placement. Divergence time estimates indicate that chance oceanic dispersal subsequent to Gondwanan fragmentation best explains the Southern Hemisphere radiation of cheilodactylids.

Molecular phylogeny and zoogeography of the freshwater crayfish genus Cherax Erichson (Decapoda: Parastacidae) in Australia

The evolutionary history and biogeography of freshwater-dependent taxa in Australia is of intrinsic interest given the present-day aridity of this continent. Cherax is the most widespread and one of the most species-rich of Australia's nine freshwater crayfish genera. The phylogenetic relationships amongst 19 of the 23 Australian Cherax were established from mitochondrial DNA sequences representing the 12S rRNA and 16S rRNA gene regions. The relationships among species support an initial east–west separation, followed by a north–south divergence in eastern Australia. Molecular clock estimations suggest that these divergences date back to the Miocene. The phylogenetic relationships support endemic speciation within geographical regions and indicate that long-distance dispersal has not led to recent speciation as previously hypothesized. This new evolutionary scenario is consistent with the climatic history of Australia and the evolutionary history of other similarly distributed freshwater-dependent organisms in Australia.

Non-quantized minimum free energy in untranslated region exons

In an attempt to improve automated gene prediction in the untranslated region of a gene, we completed an in-depth analysis of the minimum free energy for 8,689 sub-genetic DNA sequences. We expanded Zhang's classification model and classified each sub-genetic sequence into one of 27 possible motifs. We calculated the minimum free energy for each motif to explore statistical features that correlate to biologically relevant sub-genetic sequences. If biologically relevant sub-genetic sequences fall into distinct free energy quanta it may be possible to characterize a motif based on its minimum free energy. Proper characterization of motifs can lead to greater understanding in automated genefinding, gene variability and the role DNA structure plays in gene network regulation.

Our analysis determined: (1) the average free energy value for exons, introns and other biologically relevant sub-genetic sequences, (2) that these subsequences do not exist in distinct energy quanta, (3) that introns exist however in a tightly coupled average minimum free energy quantum compared to all other biologically relevant sub-genetic sequence types, (4) that single exon genes demonstrate a higher stability than exons which span the entire coding sequence as part of a multi-exon gene and (5) that all motif types contain a free energy global minimum at approximately nucleotide position 1,000 before reaching a plateau. These results should be relevant to the biochemist and bioinformatician seeking to understand the relationship between sub-genetic sequences and the information behind them.

Molecular Genetic Evidence for a New Species of Bream of the Genus Acanthopagrus Peters (Perciformes: Sparidae)

Fishes of the genus Acanthopagrus are found throughout the coastal waters of Asia and Australia with several species being of commercial significance. In this study, genetic comparisons are made between widely disjunct populations of Acanthopagrus australis (Günther) from Australian and Taiwanese waters and among samples of A. butcheri (Munro), A. berda, (Forskal), A schlegeli (Day) and A. latus (Houttuyn) using mitochondrial DNA sequences obtained from the control region. The mean interspecific pairwise sequence divergence for all species is 17%, while the divergence between A. australis from Australia and that of Taiwan is slightly larger at 18%. These values are considerably higher than those found for intraspecific control region comparisons in some fish species. Phylogenetic analyses indicate that A. australis from Australia is more closely related to the Australian species A. butcheri than to A. australis from Taiwan. These findings suggest that the northern and southern hemisphere forms of A. australis are not monophyletic, with the former possibly representing a new undescribed species of Acanthopagrus.

Sequence analysis of a salt tolerant metagenomic clone

Metagenome represent an unlimited resource for discovery of novel genes. Here we report, sequence analysis of a salt tolerant metagenomic clone (6B4) from a pond water metagenomic library. Clone 6B4 had an insert of 2254 bp with G+C composition of 64.06%. DNA sequence from 6B4 showed homology to DNA sequences from proteobacteria indicating origin of 6B4 metagenomic insert from a yet uncharacterized proteobacteria. Two encoded proteins from clone 6B4 showed match with ATP-dependent Clp protease adaptor protein (ClpS) and phasin, while two truncated encoded proteins showed match with poly-3-hydroxybutyrate synthase and permease. Clp complex is known to play a role in stress tolerance. Expression of ClpS from metagenomic clone is proposed to be responsible for salt tolerance of the metagenomic clone 6B4.

Can phylogeny predict chemical diversity and potential medicinal activity of plants? A case study of amaryllidaceae

Background During evolution, plants and other organisms have developed a diversity of chemical defences, leading to the evolution of various groups of specialized metabolites selected for their endogenous biological function. A correlation between phylogeny and biosynthetic pathways could offer a predictive approach enabling more efficient selection of plants for the development of traditional medicine and lead discovery. However, this relationship has rarely been rigorously tested and the potential predictive power is consequently unknown.
Results We produced a phylogenetic hypothesis for the medicinally important plant subfamily Amaryllidoideae (Amaryllidaceae) based on parsimony and Bayesian analysis of nuclear, plastid, and mitochondrial DNA sequences of over 100 species. We tested if alkaloid diversity and activity in bioassays related to the central nervous system are significantly correlated with phylogeny and found evidence for a significant phylogenetic signal in these traits, although the effect is not strong.
Conclusions Several genera are non-monophyletic emphasizing the importance of using phylogeny for interpretation of character distribution. Alkaloid diversity and in vitro inhibition of acetylcholinesterase (AChE) and binding to the serotonin reuptake transporter (SERT) are significantly correlated with phylogeny. This has implications for the use of phylogenies to interpret chemical evolution and biosynthetic pathways, to select candidate taxa for lead discovery, and to make recommendations for policies regarding traditional use and conservation priorities.

Deploying aptameric sensing technology for rapid pandemic monitoring

The genome of virulent strains may possess the ability to mutate by means of antigenic shift and/or antigenic drift as well as being resistant to antibiotics with time. The outbreak and spread of these virulent diseases including avian influenza (H1N1), severe acute respiratory syndrome (SARS-Corona virus), cholera (Vibrio cholera), tuberculosis (Mycobacterium tuberculosis), Ebola hemorrhagic fever (Ebola Virus) and AIDS (HIV-1) necessitate urgent attention to develop diagnostic protocols and assays for rapid detection and screening. Rapid and accurate detection of first cases with certainty will contribute significantly in preventing disease transmission and escalation to pandemic levels. As a result, there is a need to develop technologies that can meet the heavy demand of an all-embedded, inexpensive, specific and fast biosensing for the detection and screening of pathogens in active or latent forms to offer quick diagnosis and early treatments in order to avoid disease aggravation and unnecessary late treatment costs. Nucleic acid aptamers are short, single-stranded RNA or DNA sequences that can selectively bind to specific cellular and biomolecular targets. Aptamers, as new-age bioaffinity probes, have the necessary biophysical characteristics for improved pathogen detection. This article seeks to review global pandemic situations in relation to advances in pathogen detection systems. It particularly discusses aptameric biosensing and establishes application opportunities for effective pandemic monitoring. Insights into the application of continuous polymeric supports as the synthetic base for aptamer coupling to provide the needed convective mass transport for rapid screening is also presented.