Global sequence variation in the histidine-rich proteins 2 and 3 of Plasmodium falciparum: implications for the performance of malaria rapid diagnostic tests

Background Accurate diagnosis is essential for prompt and appropriate treatment of malaria. While rapid diagnostic tests (RDTs) offer great potential to improve malaria diagnosis, the sensitivity of RDTs has been reported to be highly variable. One possible factor contributing to variable test performance is the diversity of parasite antigens. This is of particular concern for Plasmodium falciparum histidine-rich protein 2 (PfHRP2)-detecting RDTs since PfHRP2 has been reported to be highly variable in isolates of the Asia-Pacific region. Methods The pfhrp2 exon 2 fragment from 458 isolates of P. falciparum collected from 38 countries was amplified and sequenced. For a subset of 80 isolates, the exon 2 fragment of histidine-rich protein 3 (pfhrp3) was also amplified and sequenced. DNA sequence and statistical analysis of the variation observed in these genes was conducted. The potential impact of the pfhrp2 variation on RDT detection rates was examined by analysing the relationship between sequence characteristics of this gene and the results of the WHO product testing of malaria RDTs: Round 1 (2008), for 34 PfHRP2-detecting RDTs. Results Sequence analysis revealed extensive variations in the number and arrangement of various repeats encoded by the genes in parasite populations world-wide. However, no statistically robust correlation between gene structure and RDT detection rate for P. falciparum parasites at 200 parasites per microlitre was identified. Conclusions The results suggest that despite extreme sequence variation, diversity of PfHRP2 does not appear to be a major cause of RDT sensitivity variation.

The complete genome sequence of Escherichia coli EC958: a high quality reference sequence for the globally disseminated multidrug resistant E. coli O25b:H4-ST131 clone

Escherichia coli ST131 is now recognised as a leading contributor to urinary tract and bloodstream infections in both community and clinical settings. Here we present the complete, annotated genome of E. coli EC958, which was isolated from the urine of a patient presenting with a urinary tract infection in the Northwest region of England and represents the most well characterised ST131 strain. Sequencing was carried out using the Pacific Biosciences platform, which provided sufficient depth and read-length to produce a complete genome without the need for other technologies. The discovery of spurious contigs within the assembly that correspond to site-specific inversions in the tail fibre regions of prophages demonstrates the potential for this technology to reveal dynamic evolutionary mechanisms. E. coli EC958 belongs to the major subgroup of ST131 strains that produce the CTX-M-15 extended spectrum β-lactamase, are fluoroquinolone resistant and encode the fimH30 type 1 fimbrial adhesin. This subgroup includes the Indian strain NA114 and the North American strain JJ1886. A comparison of the genomes of EC958, JJ1886 and NA114 revealed that differences in the arrangement of genomic islands, prophages and other repetitive elements in the NA114 genome are not biologically relevant and are due to misassembly. The availability of a high quality uropathogenic E. coli ST131 genome provides a reference for understanding this multidrug resistant pathogen and will facilitate novel functional, comparative and clinical studies of the E. coli ST131 clonal lineage.

Autotransporters of Escherichia coli : a sequence-based characterization

Autotransporter (AT) proteins are found in all Escherichia coli pathotypes and are often associated with virulence. In this study we took advantage of the large number of available E. coli genome sequences to perform an in-depth bioinformatic analysis of AT-encoding genes. Twenty-eight E. coli genome sequences were probed using an iterative approach, which revealed a total of 215 AT-encoding sequences that represented three major groups of distinct domain architecture: (i) serine protease AT proteins, (ii) trimeric AT adhesins and (iii) AIDA-I-type AT proteins. A number of subgroups were identified within each broad category, and most subgroups contained at least one characterized AT protein; however, seven subgroups contained no previously described proteins. The AIDA-I-type AT proteins represented the largest and most diverse group, with up to 16 subgroups identified from sequence-based comparisons. Nine of the AIDA-I-type AT protein subgroups contained at least one protein that possessed functional properties associated with aggregation and/or biofilm formation, suggesting a high degree of redundancy for this phenotype. The Ag43, YfaL/EhaC, EhaB/UpaC and UpaG subgroups were found in nearly all E. coli strains. Among the remaining subgroups, there was a tendency for AT proteins to be associated with individual E. coli pathotypes, suggesting that they contribute to tissue tropism or symptoms specific to different disease outcomes.

STAR : Predicting recombination sites from amino acid sequence

Background Designing novel proteins with site-directed recombination has enormous prospects. By locating effective recombination sites for swapping sequence parts, the probability that hybrid sequences have the desired properties is increased dramatically. The prohibitive requirements for applying current tools led us to investigate machine learning to assist in finding useful recombination sites from amino acid sequence alone. Results We present STAR, Site Targeted Amino acid Recombination predictor, which produces a score indicating the structural disruption caused by recombination, for each position in an amino acid sequence. Example predictions contrasted with those of alternative tools, illustrate STAR'S utility to assist in determining useful recombination sites. Overall, the correlation coefficient between the output of the experimentally validated protein design algorithm SCHEMA and the prediction of STAR is very high (0.89). Conclusion STAR allows the user to explore useful recombination sites in amino acid sequences with unknown structure and unknown evolutionary origin. The predictor service is available from http://pprowler.itee.uq.edu.au/star.

A transcriptome resource for the koala (Phascolarctos cinereus) : insights into koala retrovirus transcription and sequence diversity

Background The koala, Phascolarctos cinereus, is a biologically unique and evolutionarily distinct Australian arboreal marsupial. The goal of this study was to sequence the transcriptome from several tissues of two geographically separate koalas, and to create the first comprehensive catalog of annotated transcripts for this species, enabling detailed analysis of the unique attributes of this threatened native marsupial, including infection by the koala retrovirus. Results RNA-Seq data was generated from a range of tissues from one male and one female koala and assembled de novo into transcripts using Velvet-Oases. Transcript abundance in each tissue was estimated. Transcripts were searched for likely protein-coding regions and a non-redundant set of 117,563 putative protein sequences was produced. In similarity searches there were 84,907 (72%) sequences that aligned to at least one sequence in the NCBI nr protein database. The best alignments were to sequences from other marsupials. After applying a reciprocal best hit requirement of koala sequences to those from tammar wallaby, Tasmanian devil and the gray short-tailed opossum, we estimate that our transcriptome dataset represents approximately 15,000 koala genes. The marsupial alignment information was used to look for potential gene duplications and we report evidence for copy number expansion of the alpha amylase gene, and of an aldehyde reductase gene. Koala retrovirus (KoRV) transcripts were detected in the transcriptomes. These were analysed in detail and the structure of the spliced envelope gene transcript was determined. There was appreciable sequence diversity within KoRV, with 233 sites in the KoRV genome showing small insertions/deletions or single nucleotide polymorphisms. Both koalas had sequences from the KoRV-A subtype, but the male koala transcriptome has, in addition, sequences more closely related to the KoRV-B subtype. This is the first report of a KoRV-B-like sequence in a wild population. Conclusions This transcriptomic dataset is a useful resource for molecular genetic studies of the koala, for evolutionary genetic studies of marsupials, for validation and annotation of the koala genome sequence, and for investigation of koala retrovirus. Annotated transcripts can be browsed and queried at http://koalagenome.org

Phylogenetics of small horseshoe bats from east Asia based on mitochondrial DNA sequence variation

We undertook analyses of mitochondrial DNA gene sequences and echolocation calls to resolve phylogenetic relationships among the related bat taxa Rhinolophus pusillus (sampled across China), R. monoceros (Taiwan), R. cornutus (main islands of Japan), and R. c. pumilus (Okinawa, Japan), Phylogenetic trees and genetic divergence analyses were constructed by combining new complete mitochondrial cytochrome-b gene sequences and partial mitochondrial control region sequences with published sequences. Our work showed that these 4 taxa formed monophyletic groups in the phylogenetic tree. However, low levels of sequence divergence among the taxa, together with similarities in body size and overlapping echolocation call frequencies, point to a lack of taxonomic distinctiveness. We therefore suggest that these taxa are better considered as geographical subspecies rather than distinct species, although this should not diminish the conservation importance of these island populations, which are important evolutionarily significant units. Based on our findings, we suggest that the similarities in body size and echolocation call frequency in these rhinolophids result from their recent common ancestry, whereas similarities in body size and call frequency with R. hipposideros of Europe are the result of convergent evolution.

Inferring phylogenies of evolving sequences without multiple sequence alignment

Alignment-free methods, in which shared properties of sub-sequences (e.g. identity or match length) are extracted and used to compute a distance matrix, have recently been explored for phylogenetic inference. However, the scalability and robustness of these methods to key evolutionary processes remain to be investigated. Here, using simulated sequence sets of various sizes in both nucleotides and amino acids, we systematically assess the accuracy of phylogenetic inference using an alignment-free approach, based on D2 statistics, under different evolutionary scenarios. We find that compared to a multiple sequence alignment approach, D2 methods are more robust against among-site rate heterogeneity, compositional biases, genetic rearrangements and insertions/deletions, but are more sensitive to recent sequence divergence and sequence truncation. Across diverse empirical datasets, the alignment-free methods perform well for sequences sharing low divergence, at greater computation speed. Our findings provide strong evidence for the scalability and the potential use of alignment-free methods in large-scale phylogenomics.

Variation in the OC locus of Acinetobacter baumannii genomes predicts extensive structural diversity in the Lipooligosaccharide

Lipooligosaccharide (LOS) is a complex surface structure that is linked to many pathogenic properties of Acinetobacter baumannii. In A. baumannii, the genes responsible for the synthesis of the outer core (OC) component of the LOS are located between ilvE and aspS. The content of the OC locus is usually variable within a species, and examination of 6 complete and 227 draft A. baumannii genome sequences available in GenBank non-redundant and Whole Genome Shotgun databases revealed nine distinct new types, OCL4-OCL12, in addition to the three known ones. The twelve gene clusters fell into two distinct groups, designated Group A and Group B, based on similarities in the genes present. OCL6 (Group B) was unique in that it included genes for the synthesis of L-Rhamnosep. Genetic exchange of the different configurations between strains has occurred as some OC forms were found in several different sequence types (STs). OCL1 (Group A) was the most widely distributed being present in 18 STs, and OCL6 was found in 16 STs. Variation within clones was also observed, with more than one OC locus type found in the two globally disseminated clones, GC1 and GC2, that include the majority of multiply antibiotic resistant isolates. OCL1 was the most abundant gene cluster in both GC1 and GC2 genomes but GC1 isolates also carried OCL2, OCL3 or OCL5, and OCL3 was also present in GC2. As replacement of the OC locus in the major global clones indicates the presence of sub-lineages, a PCR typing scheme was developed to rapidly distinguish Group A and Group B types, and to distinguish the specific forms found in GC1 and GC2 isolates.

Genome Sequence of Acinetobacter baumannii Strain A1, an Early Example of Antibiotic-Resistant Global Clone 1

Acinetobacter baumannii isolate A1 was recovered in the United Kingdom in 1982 and belongs to global clone 1 (GC1). Here, we present its complete 3.91-Mbp genome sequence, generated via a combination of short-read sequencing (Illumina), long-read sequencing (PacBio), and manual finishing.

The sequence: Inside the race for the human genome

The human genome project was a grand scientific enterprise which attracted both hyperbole and ridicule alike. The project was lauded as “the moon shot of the life sciences”, the “holy grail of man”, “the code of codes”, and “the book of life”. Such rhetoric has also received scorn. President George Bush senior managed to deflate the pretensions of the project with the accidental slip that it was the “human gnome initiative”. In The Sequence, Kevin Davies seeks to go beyond such metaphors, and provide a candid and honest account of the race of the human genome project. The author is indebted to the authoritative book The Gene Wars, which considered the early struggles over the human genome project. Robert Cook-Deegan observes that there was initially much debate over whether there should be a Human Genome Project at all: The debate became one of “big” science versus “small” science. The reliance on systematic technology development and goal-directed gene-mapping efforts presaged a new style for biology, one that elicited excitement from those attracted to whiz-bang technologies but drew gasps of revulsion from those who aspired to cultivate biology on a more modest scale and with decentralized organisation. The battle was, among other things, over whose vision would control the budget and which scientific aesthetic would prevail.

The new conquistadors: Patent law and expressed sequence tags

It's akin to the old Spanish, English and Portuguese explorers. They would take their boats until they found some edge of land, then they would go up and plant the flag of their king or queen. They didn't know what they'd discovered; how big it is, where it goes to - but they would claim it anyway. David Korn of the Association of American Medical Colleges This article analyses recent litigation over patent law and expressed sequence tags (ESTs). In the case of In re Fisher, the United States Court of Appeals for the Federal Circuit engaged in judicial consideration of the revised utility guidelines of the United States Patent and Trademark Office (USPTO). In this matter, the agricultural biotechnology company Monsanto sought to patent ESTs in maize plants. A patent examiner and the Board of Patent Appeals and Interferences had doubted whether the patent application was useful. Monsanto appealed against the rulings of the USPTO. A number of amicus curiae intervened in the matter in support of the USPTO - including Genentech, Affymetrix, Dow AgroSciences, Eli Lilly, the National Academy of Sciences, and the Association of American Medical Colleges. The majority of the Court of Appeals for the Federal Circuit supported the position of the USPTO, and rejected the patent application on the grounds of utility. The split decision highlighted institutional tensions over the appropriate thresholds for patent criteria - such as novelty, non-obviousness, and utility. The litigation raised larger questions about the definition of research tools, the incremental nature of scientific progress, and the role of patent law in innovation policy. The decision of In re Fisher will have significant ramifications for gene patents, in the wake of the human genome project. Arguably, the USPTO utility guidelines need to be reinforced by a tougher application of the standards of novelty and non-obviousness in respect of gene patents.

Investigating the population structure of Queensland invasive Streptococcus pneumoniae isolates in children: Using a modified multi-locus variable number of tandem repeat analysis and a novel minimum SNPs capsular typing method

This research investigated the use of DNA fingerprinting to characterise the bacteria Streptococcus pneumoniae or pneumococcus, and hence gain insight into the development of new vaccines or antibiotics. Different bacterial DNA fingerprinting methods were studied, and a novel method was developed and validated, which characterises different cell coatings that pneumococci produce. This method was used to study the epidemiology of pneumococci in Queensland before and after the introduction of the current pneumococcal vaccine. This study demonstrated that pneumococcal disease is highly prevalent in children under four years, that the bacteria can `switch' its cell coating to evade the vaccine, and that some DNA fingerprinting methods are more discriminatory than others. This has an impact on understanding which strains are more prone to cause invasive disease. Evidence of the excellent research findings have been published in high impact internationally refereed journals.

Complex Symbolic Sequence Encodings for Predictive Monitoring of Business Processes

This paper addresses the problem of predicting the outcome of an ongoing case of a business process based on event logs. In this setting, the outcome of a case may refer for example to the achievement of a performance objective or the fulfillment of a compliance rule upon completion of the case. Given a log consisting of traces of completed cases, given a trace of an ongoing case, and given two or more possible out- comes (e.g., a positive and a negative outcome), the paper addresses the problem of determining the most likely outcome for the case in question. Previous approaches to this problem are largely based on simple symbolic sequence classification, meaning that they extract features from traces seen as sequences of event labels, and use these features to construct a classifier for runtime prediction. In doing so, these approaches ignore the data payload associated to each event. This paper approaches the problem from a different angle by treating traces as complex symbolic sequences, that is, sequences of events each carrying a data payload. In this context, the paper outlines different feature encodings of complex symbolic sequences and compares their predictive accuracy on real-life business process event logs.

Genome sequence of marine bacterium idiomarina sp. strain 28-8, isolated from Korean ark shells

Idiomarina sp. strain 28-8 is an aerobic, Gram-negative, flagellar bacterium isolated from the bodies of ark shells (Scapharca broughtonii) collected from underwater sediments in Gangjin Bay, South Korea. Here, we present the draft genome sequence of Idiomarina sp. 28-8 (2,971,606 bp, with a G+C content of 46.9%), containing 2,795 putative coding sequences.