Genotypic and phenotypic diversity differences of presumptive commensal and avian pathogenic E. coli

1. The objective of the experiment was to characterise the genotypic and phenotypic differences between presumptive commensal E. coli and avian pathogenic E. coli (APEC) of poultry. 2. DNA was extracted from 65 confirmed APEC E. coli from chicken, 100 presumptive commensal E. coli from healthy turkey and 35 from healthy chicken. Enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and virulence factors genotyping was performed to characterise genetic features. 3. Carbon source utilisation and antimicrobial susceptibility tests were performed to characterise phenotypic features of isolates. 4. The genetic divergence between E. coli strains tested by ERIC-PCR profiles and virulence associated genes showed a clear genetic separation between E. coli APEC and turkey E. coli strains. 5. The carbon utilisation profile of turkey isolates was different from chicken and APEC strains; whereas antimicrobial susceptibility was highest for turkey isolates (53%), and lowest for APEC strains (33.8%). 6. The study showed a significant negative correlation between utilisation of arabitol and adonitol with different virulence determinants tested, which suggests that the ability to utilise some uncommon carbon sources may be used to discriminate between presumptive commensal E. coli and APEC

Comparative genomics of Salmonella enterica serovars Derby and Mbandaka, two prevalent serovars associated with different livestock species in the UK

Background Despite the frequent isolation of Salmonella enterica sub. enterica serovars Derby and Mbandaka from livestock in the UK and USA little is known about the biological processes maintaining their prevalence. Statistics for Salmonella isolations from livestock production in the UK show that S. Derby is most commonly associated with pigs and turkeys and S. Mbandaka with cattle and chickens. Here we compare the first sequenced genomes of S. Derby and S. Mbandaka as a basis for further analysis of the potential host adaptations that contribute to their distinct host species distributions. Results Comparative functional genomics using the RAST annotation system showed that predominantly mechanisms that relate to metabolite utilisation, in vivo and ex vivo persistence and pathogenesis distinguish S. Derby from S. Mbandaka. Alignment of the genome nucleotide sequences of S. Derby D1 and D2 and S. Mbandaka M1 and M2 with Salmonella pathogenicity islands (SPI) identified unique complements of genes associated with host adaptation. We also describe a new genomic island with a putative role in pathogenesis, SPI-23. SPI-23 is present in several S. enterica serovars, including S. Agona, S. Dublin and S. Gallinarum, it is absent in its entirety from S. Mbandaka. Conclusions We discovered a new 37 Kb genomic island, SPI-23, in the chromosome sequence of S. Derby, encoding 42 ORFS, ten of which are putative TTSS effector proteins. We infer from full-genome synonymous SNP analysis that these two serovars diverged, between 182kya and 625kya coinciding with the divergence of domestic pigs. The differences between the genomes of these serovars suggest they have been exposed to different stresses including, phage, transposons and prolonged externalisation. The two serovars possess distinct complements of metabolic genes; many of which cluster into pathways for catabolism of carbon sources

A process for analysis of microarray comparative genomics hybridisation studies for bacterial genomes

<p>Abstract</p> <p>Background</p> <p>Microarray based comparative genomic hybridisation (CGH) experiments have been used to study numerous biological problems including understanding genome plasticity in pathogenic bacteria. Typically such experiments produce large data sets that are difficult for biologists to handle. Although there are some programmes available for interpretation of bacterial transcriptomics data and CGH microarray data for looking at genetic stability in oncogenes, there are none specifically to understand the mosaic nature of bacterial genomes. Consequently a bottle neck still persists in accurate processing and mathematical analysis of these data. To address this shortfall we have produced a simple and robust CGH microarray data analysis process that may be automated in the future to understand bacterial genomic diversity.</p> <p>Results</p> <p>The process involves five steps: cleaning, normalisation, estimating gene presence and absence or divergence, validation, and analysis of data from test against three reference strains simultaneously. Each stage of the process is described and we have compared a number of methods available for characterising bacterial genomic diversity, for calculating the cut-off between gene presence and absence or divergence, and shown that a simple dynamic approach using a kernel density estimator performed better than both established, as well as a more sophisticated mixture modelling technique. We have also shown that current methods commonly used for CGH microarray analysis in tumour and cancer cell lines are not appropriate for analysing our data.</p> <p>Conclusion</p> <p>After carrying out the analysis and validation for three sequenced <it>Escherichia coli </it>strains, CGH microarray data from 19 <it>E. coli </it>O157 pathogenic test strains were used to demonstrate the benefits of applying this simple and robust process to CGH microarray studies using bacterial genomes.</p

454-Pyrosequencing: A Molecular Battiscope for Freshwater Viral Ecology

Viruses, the most abundant biological entities on the planet, are capable of infecting organisms from all three branches of life, although the majority infect bacteria where the greatest degree of cellular diversity lies. However, the characterization and assessment of viral diversity in natural environments is only beginning to become a possibility. Through the development of a novel technique for the harvest of viral DNA and the application of 454 pyrosequencing, a snapshot of the diversity of the DNA viruses harvested from a standing pond on a cattle farm has been obtained. A high abundance of viral genotypes (785) were present within the virome. The absolute numbers of lambdoid and Shiga toxin (Stx) encoding phages detected suggested that the depth of sequencing had enabled recovery of only ca. 8% of the total virus population, numbers that agreed within less than an order of magnitude with predictions made by rarefaction analysis. The most abundant viral genotypes in the pond were bacteriophages (93.7%). The predominant viral genotypes infecting higher life forms found in association with the farm were pathogens that cause disease in cattle and humans, e.g. members of the Herpesviridae. The techniques and analysis described here provide a fresh approach to the monitoring of viral populations in the aquatic environment, with the potential to become integral to the development of risk analysis tools for monitoring the dissemination of viral agents of animal, plant and human diseases

Collimation and asymmetry of the hot blast wave from the recurrent nova V745 Scorpii

The recurrent symbiotic nova V745 Sco exploded on 2014 February 6 and was observed on February 22 and 23 by the Chandra X-ray Observatory Transmission Grating Spectrometers. By that time the supersoft source phase had already ended and Chandra spectra are consistent with emission from a hot, shock-heated circumstellar medium with temperatures exceeding 10^7K. X-ray line profiles are more sharply peaked than expected for a spherically-symmetric blast wave, with a full width at zero intensity of approximately 2400 km/s, a full width at half maximum of 1200 +/- 30 km/s and an average net blueshift of 165 +/- 10 km/s. The red wings of lines are increasingly absorbed toward longer wavelengths by material within the remnant. We conclude that the blast wave was sculpted by an aspherical circumstellar medium in which an equatorial density enhancement plays a role, as in earlier symbiotic nova explosions. Expansion of the dominant X-ray emitting material is aligned close to the plane of the sky and most consistent with an orbit seen close to face-on. Comparison of an analytical blast wave model with the X-ray spectra, Swift observations and near-infrared line widths indicates the explosion energy was approximately 10^43 erg, and confirms an ejected mass of approximately 10^-7 Msun. The total mass lost is an order of magnitude lower than the accreted mass required to have initiated the explosion, indicating the white dwarf is gaining mass and is a supernova Type 1a progenitor candidate.Comment: To appear in the Astrophysical Journa

Curing vector for IncI1 plasmids and its use to provide evidence for a metabolic burden of IncI1 CTX-M-1 plasmid pIFM3791 on Klebsiella pneumoniae

Using a sequence based approach we previously identified an IncI1 CTX-M-1 plasmid,pIFM3791, on a single pig farm in the UK that was harboured by K. pneumoniae,Escherichia coli and Salmonella enterica serotype 4,5,12,i:-. To test the hypothesis that the plasmid had spread rapidly into these differing host bacteria we wished to assess whether the plasmid conferred a fitness advantage. To do this an IncI1 curing vector was constructed and used to displace the IncI1 CTX-M-1 plasmids from K. pneumoniae strain B3791 and several other unrelated IncI1 harbouring strains indicating the potential wider application of the curing plasmid. The IncI1 CTX-M-1 plasmid was re-introduced by conjugation into the cured K. pneumoniae strain and also a naturally IncI1 plasmid free S. enterica serotype 4,5,12,i:-, S348/1. Original, cured and complemented strains were tested for metabolic competence using BiologTM technology and in competitive growth, association to mammalian cells and biofilm formation experiments. The plasmid-cured K. pneumoniae strain grew more rapidly than either the original plasmid-carrying strain or plasmid-complemented strains in competition experiments. Additionally, the plasmid-cured strain was significantly better at respiring with L-sorbose as a carbon source and putrescine, γ-amino-n-butyric acid,L-alanine, L-proline as a nitrogen sources. By contrast, no differences in phenotype were found when comparing plasmid harbouring and plasmid free S. enterica S348/11. In conclusion, the IncI1 curing vector successfully displaced multiple IncI plasmids. The IncI1 CTX-M1 plasmid conferred a growth disadvantage upon K. pneumoniae, possibly by imposing a metabolic burden the mechanism of which remains to be determined

Identification of differentially temperature regulated virulence factors of Campylobacter jejuni and tested in Galleria mellonella

Background Campylobacter jejuni is the major cause of bacterial gastroenteritis in man, while it is generally regarded as a commensal of the avian gut. Consumption and handling of contaminated poultry meat products are a major risk factor for human infection. The body temperature in man 37°C and chickens 42°C differ markedly, and differential gene regulation and protein expression at different temperatures may in part explain the behaviour in the two hosts. Results We performed proteomics analyses with C. jejuni cells grown at 37°C and 42°C. Q tof analysis was carried out after samples were digested with the Fasp method and peptides were fractionated by strong anion exchanges. Differentially regulated proteins were identified by Mascot and Scaffold analyses. QQQ analysis confirmed that a total of 33 proteins were differentially regulated between 37°C and 42°C. Several upregulated proteins were selected for their corresponding gene knock-out mutants to be tested for their virulence in the Galleria mellonella model. To correlate with other tissue/animal models, the GADH mutant was selected for its reduced ability to colonize chickens. At 37°C, the mutants of Omp50 and GroEL significantly increased virulence; while at 42°C, the mutants of YceI, Omp50, and GADH reduced virulence against Galleria mellonella compared with the wild type strains. Conclusion The results of current and previous studies indicate that GADH is a virulent factor in G. mellonella and a colonization factor in chickens. The workflow of this study may prove a new way to identify stress related virulent factors. The implications of these findings are discussed for pathogenesis in the model and other hosts

A Structured Approach to Modifying Successful Heuristics

In some cases, heuristics may be transferred easily between different optimisation problems. This is the case if these problems are equivalent or dual (e.g., maximum clique and maximum independent set) or have similar objective functions. However, the link between problems can further be defined by the constraints that define them. This refining can be achieved by organising constraints into families and translating between them using gadgets. If two problems are in the same constraint family, the gadgets tell us how to map from one problem to another and which constraints are modified. This helps better understand a problem through its constraints and how best to use domain specific heuristics. In this position paper, we argue that this allows us to understand how to map between heuristics developed for one problem to heuristics for another problem, giving an example of how this might be achieved

Antimicrobial activities of ellagitannins against Clostridiales perfringens, Escherichia coli, Lactobacillus plantarum and Staphylococcus aureus

In this study, we tested the growth inhibition effect of 22 individual ellagitannins and of pentagalloylglucose on four bacterial species, i.e., Clostridiales perfringens, Escherichia coli, Lactobacillus plantarum and Staphylococcus aureus. All tested compounds showed antimicrobial effects against S. aureus, and almost all against E. coli and C. perfringens. For L. plantarum, no or very weak growth inhibition was detected. The level of inhibition was the greatest for S. aureus and the weakest for C. perfringens. For S. aureus, the molecular size or flexibility of ellagitannins did not show a clear relationship with their antimicrobial activity, even though rugosins E and D and pentagalloylglucose with four or five free galloyl groups had a stronger growth inhibition effect than the other ellagitannins with glucopyranose cores but with less free galloyl groups. Additionally, our results with S. aureus showed that the oligomeric linkage of ellagitannin might have an effect on its antimicrobial activity. For E. coli, the molecular size, but not the molecular flexibility, of ellagitannins seemed to be an important factor. For C. perfringens, both the molecular size and the flexibility of ellagitannin were important factors. In previous studies, corilagin was used as a model for ellagitannins, but our results showed that other ellagitannins are much more efficacious; therefore, the antimicrobial effects of ellagitannins could be more significant than previously thought

Escherichia coli isolates from extraintestinal organs of livestock animals harbour diverse virulence genes and belong to multiple genetic lineages

Escherichia coli, the most common cause of bacteraemia in humans in the UK, can also cause serious diseases in animals. However the population structure, virulence and antimicrobial resistance genes of those from extraintestinal organs of livestock animals are poorly characterised. The aims of this study were to investigate the diversity of these isolates from livestock animals and to understand if there was any correlation between the virulence and antimicrobial resistance genes and the genetic backbone of the bacteria and if these isolates were similar to those isolated from humans. Here 39 E. coli isolates from liver (n=31), spleen (n=5) and blood (n=3) of cattle (n=34), sheep (n=3), chicken (n=1) and pig (n=1) were assigned to 19 serogroups with O8 being the most common (n=7), followed by O101, O20 (both n=3) and O153 (n=2). They belong to 29 multi-locus sequence types, 20 clonal complexes with ST23 (n=7), ST10 (n=6), ST117 and ST155 (both n=3) being most common and were distributed among phylogenetic group A (n=16), B1 (n=12), B2 (n=2) and D (n=9). The pattern of a subset of putative virulence genes was different in almost all isolates. No correlation between serogroups, animal hosts, MLST types, virulence and antimicrobial resistance genes was identified. The distributions of clonal complexes and virulence genes were similar to other extraintestinal or commensal E. coli from humans and other animals, suggesting a zoonotic potential. The diverse and various combinations of virulence genes implied that the infections were caused by different mechanisms and infection control will be challenging