Characterization of Streptococcus bovis from the rumen of the dromedary camel and Rusa deer

Aims: Isolation and characterization of Streptococcus bovis from the dromedary camel and Rusa deer. Methods and Results: Bacteria were isolated from the rumen contents of four camels and two deer fed lucerne hay by culturing on the semi-selective medium MRS agar. Based on Gram morphology and RFLP analysis seven isolates, MPR1, MPR2, MPR3, MPR4, MPR5, RD09 and RD11 were selected and putatively identified as Streptococcus. The identity of these isolates was later confirmed by comparative DNA sequence analysis of the 16S rRNA gene with the homologous sequence from S. bovis strains, JB1, C14b1, NCFB2476, SbR1, SbR7 and Sb5, from cattle and sheep, and the Streptococcus equinus strain NCD01037T. The percentage similarity amongst all strains was >99%, confirming the identification of the camel isolates as S. bovis. The strains were further characterized by their ability to utilize a range of carbohydrates, the production of volatile fatty acids (VFA) and lactate and the determination of the doubling time in basal medium 10 supplemented with glucose. All the isolates produced L-lactate as a major fermentation end product, while four of five camel isolates produced VFA. The range of carbohydrates utilized by all the strains tested, including those from cattle and sheep were identical, except that all camel isolates and the deer isolate RD11 were additionally able to utilize arabinose. Conclusions: Streptococcus bovis was successfully isolated from the rumen of camels and deer, and shown by molecular and biochemical characterization to be almost identical to S. bovis isolates from cattle and sheep. Significance and Impact of the Study: Streptococcus bovis is considered a key lactic acid producing bacterium from the gastrointestinal tract of ruminants, and has been implicated as a causative agent of lactic acidosis. This study is the first report of the isolation and characterization of S. bovis from the dromedary camel and Rusa deer, and suggests a major contributive role of this bacterium to fermentative acidosis.

The genetic diversity of lactic acid producing bacteria in the equine gastrointestinal tract

Seventy-two lactic acid producing bacterial isolates (excluding streptococci) were cultured from the gastrointestinal tract of six horses. Two of the horses were orally dosed with raftilose to induce lactic acidosis and laminitis while the remaining four were maintained on a roughage diet. Near complete 16S rDNA was amplified by PCR from the genomic DNA of each isolate. Following RFLP analysis with the restriction enzymes MboI, HhaI and HinfI, the PCR products from the IS isolates that produced L- and/or D-lactate were subsequently cloned and sequenced. DNA sequence analysis indicated that the majority of the isolates were closely related to species within the genus Lactobacillus, including Lactobacillus salivarius, Lactobacillus mucosae and Lactobacillus delbrueckii. Four isolates were closely related to Mitsuokella jalaludinii. Lactic acid producing bacteria (LAB) from the equine gastrointestinal tract was dominated by representatives from the genus Lactobacillus, but also included D-lactate-producing bacteria closely related to M. jalaludinii. Identification and characterization of LAB from the equine gastrointestinal tract should contribute to our understanding and management of fermentative acidosis, ulceration of the stomach and laminitis. (c) 2005 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.

Inherent size constraints on prokaryote gene networks due to "accelerating" growth

Networks exhibiting accelerating growth have total link numbers growing faster than linearly with network size and either reach a limit or exhibit graduated transitions from nonstationary-to-stationary statistics and from random to scale-free to regular statistics as the network size grows. However, if for any reason the network cannot tolerate such gross structural changes then accelerating networks are constrained to have sizes below some critical value. This is of interest as the regulatory gene networks of single-celled prokaryotes are characterized by an accelerating quadratic growth and are size constrained to be less than about 10,000 genes encoded in DNA sequence of less than about 10 megabases. This paper presents a probabilistic accelerating network model for prokaryotic gene regulation which closely matches observed statistics by employing two classes of network nodes (regulatory and non-regulatory) and directed links whose inbound heads are exponentially distributed over all nodes and whose outbound tails are preferentially attached to regulatory nodes and described by a scale-free distribution. This model explains the observed quadratic growth in regulator number with gene number and predicts an upper prokaryote size limit closely approximating the observed value. (c) 2005 Elsevier GmbH. All rights reserved.

Molecular characterization and cancer risk associated with BRCA1 and BRCA2 splice site variants identified in multiple-case breast cancer families

Genetic screening of women from multiple-case breast cancer families and other research-based endeavors have identified an extensive collection of germline variations of BRCA1 and BRCA2 that can be classified as deleterious and have clinical relevance. For some variants, such as those in the conserved intronic splice site regions which are highly likely to alter splicing, it is not possible to classify them based on the identified DNA sequence variation alone. We studied 11 multiple-case breast cancer families carrying seven distinct splice site region genetic alterations in BRCA1 or BRCA2 (BRCA1, c.IVS6-2delA, c.IVS9-2A>C, c.IVS4-1G>T, c.IVS20+1G>A and BRCA2, c.IVS17-1G>C, c.IVS20+1G>A, c.IVS7-1G>A) and applied SpliceSiteFinder to predict possible changes in efficiency of splice donor and acceptor sites, characterized the transcripts, and estimated the average age-specific cumulative risk (penetrance) using a modified segregation analysis. SpliceSiteFinder predicted and we identified transcipts that illustrated that all variants caused exon skipping, and all but two led to frameshifts. The risks of breast cancer to age 70 yrs, averaged over all variants, over BRCA1 variants alone, and over BRCA2 variants alone, were 73% (95% confidence interval 47-93), 64% (95%CI 28-96) and 79% (95%CI 48-98) respectively (all P

hnRNP A2, a potential ssDNA/RNA molecular adapter at the telomere

The heterogeneous nuclear ribonucleoprotein (hnRNP) A2 is a multi-tasking protein that acts in the cytoplasm and nucleus. We have explored the possibility that this protein is associated with telomeres and participates in their maintenance. Rat brain hnRNP A2 was shown to have two nucleic acid binding sites. In the presence of heparin one site binds single-stranded oligodeoxyribonucleotides irrespective of sequence but not the corresponding oligoribonucleotides. Both the hnRNP A2-binding cis-acting element for the cytoplasmic RNA trafficking element, A2RE, and the ssDNA telomere repeat match a consensus sequence for binding to a second sequence-specific site identified by mutational analysis. hnRNP A2 protected the telomeric repeat sequence, but not the complementary sequence, against DNase digestion: the glycine-rich domain was found to be necessary, but not sufficient, for protection. The N-terminal RRM (RNA recognition motif) and tandem RRMs of hnRNP A2 also bind the single-stranded, template-containing segment of telomerase RNA. hnRNP A2 colocalizes with telomeric chromatin in the subset of PML bodies that are a hallmark of ALT cells, reinforcing the evidence for hnRNPs having a role in telomere maintenance. Our results support a model in which hnRNP A2 acts as a molecular adapter between single-stranded telomeric repeats, or telomerase RNA, and another segment of ssDNA.

Epigenetics of lung cancer

Epigenetics is the study of heritable changes in gene expression that occur without changes in DNA sequence. It has a role in determining when and where a gene is expressed during development. Perhaps the most well known epigenetic mechanism is DNA methylation whereby cytosines at position 5 in CpG dinucleotides are methylated. Histone modification is another form of epigenetic control, which is quite complex and diverse. Histones and DNA make up the nucleosome which is the structural unit of chromatin which are involved in packaging DNA. Apart from the crucial role epigenetics plays in embryonic development, transcription, chromatin structure, X chromosome inactivation and genomic imprinting, its role in an increasing number of human diseases is more and more recognized. These diseases include cancer, and lung cancer in particular has been increasingly studied for the potential biological role of epigenetic changes with the promise of better and novel diagnostic and therapeutic tools.

Characterization of Kudoa monodactyli n. sp (Myxosporea : Multivalvulida) from the muscle of Monodactylus argenteus (Teleostei : Monodactylidae) from Moreton Bay, Queensland, Australia

Kudoa monodactyli n. sp. is described from the somatic musculature of Monodactylus argenteus from several localities in southern Queensland, Australia. This is the first record of a myxozoan parasite from the family Monodactylidae. The spores typically have five polar capsules, making this species similar to the four other five-valved Kudoa species (K. neurophila, K. muscularis, K. shulmani, K. cutanea) that have been described to date. However, morphometric measurements particularly of spore length and width make the species from M. argenteus distinct from the other species. Comparison of the small subunit ribosomal DNA sequence of this species with its congeners for which sequence data are available, provides further evidence of novelty. Kudoa monodactyli n. sp. displays 38 (of 1,554) nucleotide differences compared with rDNA sequence of Kudoa neurophila, which on phylogenetic analysis places these species in clades exclusive of each other. Phylogenetic analyses also provide evidence that the number of valves per spore in this genus is an imperfect indicator of relatedness.

Extraordinary number of gene rearrangements in the mitochondrial genomes of lice (Phthiraptera : Insecta)

The arrangement of genes in the mitochondrial (mt) genomes of most insects is the same, or near-identical, to that inferred to be ancestral for insects. We sequenced the entire mt genome of the small pigeon louse, Campanulotes bidentatus compar, and part of the mt genomes of nine other species of lice. These species were from six families and the three main suborders of the order Phthiraptera. There was no variation in gene arrangement among species within a family but there was much variation in gene arrangement among the three suborders of lice. There has been an extraordinary number of gene rearrangements in the mitochondrial genomes of lice!

Molecular discrimination of taeniid cestodes :

DNA approaches are now being used routinely for accurate identification of Echinococcus and Taenia species, subspecies and strains, and in molecular epidemiological surveys of echinococcosis/taeniasis in different geographical settings and host assemblages. The publication of the complete sequences of the mitochondrial (int) genomes of E. granulosus, E. multilocularis, T solium and Asian Taenia, and the availability of mtDNA sequences for a number of other taeniid genotypes, has provided additional genetic information that can be used for more in depth phylogenetic and taxonomic studies of these parasites. This very rich sequence information has provided a solid molecular basis, along with a range of different biological, epidemiological, biochemical and other molecular-genetic criteria, for revising the taxonomy of the genus Echinococcus and for estimating the evolutionary time of divergence of the various taxa. Furthermore, the accumulating genetic data has allowed the development of PCR-based tests for unambiguous identification of Echinococcus eggs in the faeces of definitive hosts and in the environment. Molecular phylogenies derived from mtDNA sequence comparisons of geographically distributed samples of T solium provide molecular evidence for two genotypes, one being restricted to Asia, with the other occurring in Africa and America. Whether the two genetic forms of T solium differ in important phenotypic characteristics remains to be determined. As well, minor DNA sequence differences have been reported between isolates of T saginata and Asian Taenia. There has been considerable discussion over a number of years regarding the taxonomic position of Asian Taenia and whether it should be regarded as a genotype, strain, subspecies or sister species of T saginata. The available molecular genetic data do not support independent species status for Asian Taenia and T saginata. What is in agreement is that both taxa are closely related to each other but distantly related to T solium. This is important in public health terms as it predicts that cysticercosis in humans attributable to Asian Taenia does not occur, because cysticercosis is unknown in T saginata. (C) 2005 Elsevier Ireland Ltd. All rights reserved.

Segmenting eukaryotic genomes with the generalized Gibbs sampler

Eukaryotic genomes display segmental patterns of variation in various properties, including GC content and degree of evolutionary conservation. DNA segmentation algorithms are aimed at identifying statistically significant boundaries between such segments. Such algorithms may provide a means of discovering new classes of functional elements in eukaryotic genomes. This paper presents a model and an algorithm for Bayesian DNA segmentation and considers the feasibility of using it to segment whole eukaryotic genomes. The algorithm is tested on a range of simulated and real DNA sequences, and the following conclusions are drawn. Firstly, the algorithm correctly identifies non-segmented sequence, and can thus be used to reject the null hypothesis of uniformity in the property of interest. Secondly, estimates of the number and locations of change-points produced by the algorithm are robust to variations in algorithm parameters and initial starting conditions and correspond to real features in the data. Thirdly, the algorithm is successfully used to segment human chromosome 1 according to GC content, thus demonstrating the feasibility of Bayesian segmentation of eukaryotic genomes. The software described in this paper is available from the author's website (www.uq.edu.au/similar to uqjkeith/) or upon request to the author.

How lifetimes shape epigenotype within and across generations

Despite our detailed characterization of the human genome at the level of the primary DNA sequence, we are still far from understanding the molecular events underlying phenotypic variation. Epigenetic modifications to the DNA sequence and associated chromatin are known to regulate gene expression and, as such, are a significant contributor to phenotype. Studies of inbred mice and monozygotic twins show that variation in the epigenotype can be seen even between genetically identical individuals and that this, in some cases at least, is associated with phenotypic differences. Moreover, recent evidence suggests that the epigenome can be influenced by the environment and these changes can last a lifetime. However, we also know that epigenetic states in real-time are in continual flux and, as a result, the epigenome exhibits instability both within and across generations. We still do not understand the rules governing the establishment and maintenance of the epigenotype at any particular locus. The underlying DNA sequence itself and the sequence at unlinked loci (modifier loci) are certainly involved. Recent support for the existence of transgenerational epigenetic inheritance in mammals suggests that the epigenetic state of the locus in the previous generation may also play a role. Over the next decade, many of these processes will be better understood, heralding a greater capacity for us to correlate measurable molecular marks with phenotype and providing the opportunity for improved diagnosis and presymptomatic healthcare.

Phylogenetic analysis to define feline immunodeficiency virus subtypes in 31 domestic cats in South Africa

Feline immunodeficiency virus (FIV), a lentivirus, is an important pathogen of domestic cats around the world and has many similarities to human immunodeficiency virus (HIV). A characteristic of these lentiviruses is their extensive genetic diversity which has been an obstacle in the development of successful vaccines. Of the FIV genes, the envelope gene is the most variable and sequence differences in a portion of this gene have been used to define 5 FIV subtypes (A, B, C, D and E). In this study, the proviral DNA sequence of the V3-V5 region of the envelope gene was determined in blood samples from 31 FIV positive cats from 4 different regions of South Africa. Phylogenetic analysis demonstrated the presence of both subtypes A and C, with subtype A predominating. These findings contribute to the understanding of the genetic diversity of FIV

The SEF14 fimbrial antigen of Salmonella enterica serovar Enteritidis is encoded within a pathogenicity islet

The DNA sequence of the chromosomal gene cluster encoding the SEF14 fimbriae of Salmonella enterica serovar Enteritidis was determined. Five contiguous open reading frames, sefABCDE, were identified. The sefE gene shared significant homology with araC-like positive regulators. Serovar-associated virulence plasmid (SAP) genes orf7,8,9 and pefI were identified immediately adjacent to the sef operon. The pefI gene encoded a putative regulator of the Plasmid-encoded fimbrial antigen (PEF) expression. The entire sef--pef region, flanked by two IS-like elements, was inserted adjacent to leuX that encoded a transfer RNA molecule. The organisation of this region was suggestive of a classic pathogenicity islet. Southern hybridisation confirmed two copies of the SAP derived orf7,8,9 and pefI region in S. Enteritidis, one in the chromosome and one on the SAP. Of other group D Salmonella, only S. Blegdam and S. Moscow harboured both chromosomal and plasmid copies of pefI--orf9 region although polymorphism was evident.