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1.  Naturally selecting solutions 
Bioengineered  2012;4(5):266-278.
For decades, computer scientists have looked to nature for biologically inspired solutions to computational problems; ranging from robotic control to scheduling optimization. Paradoxically, as we move deeper into the post-genomics era, the reverse is occurring, as biologists and bioinformaticians look to computational techniques, to solve a variety of biological problems. One of the most common biologically inspired techniques are genetic algorithms (GAs), which take the Darwinian concept of natural selection as the driving force behind systems for solving real world problems, including those in the bioinformatics domain. Herein, we provide an overview of genetic algorithms and survey some of the most recent applications of this approach to bioinformatics based problems.
doi:10.4161/bioe.23041
PMCID: PMC3813526  PMID: 23222169
genetic algorithm; optimization; multiple sequence alignment; protein structure prediction
2.  The role of the Cronobacter sakazakii ProP C-terminal coiled coil domain in osmotolerance 
Gut Pathogens  2014;6(1):46.
Background
We investigate the role of the C-terminal coiled coil of the secondary proline porter ProP in contributing to Cronobacter sakazakii osmotolerance.
Findings
The extended C-terminal domain of ProP1 (encoded by ESA_02131) was spliced onto the truncated C-terminal end of ProP2 (encoded by ESA_01706); creating a chimeric protein (ProPc) which exhibits increased osmotolerance relative to the wild type.
Conclusions
It appears that the C-terminal coiled coil domain tunes ProP at low osmolality, whereas ProP transporters lacking the coiled coil domain are more active at a higher osmolality range.
doi:10.1186/s13099-014-0046-9
PMCID: PMC4272814  PMID: 25530808
3.  Combined metagenomic and phenomic approaches identify a novel salt tolerance gene from the human gut microbiome 
In the current study, a number of salt-tolerant clones previously isolated from a human gut metagenomic library were screened using Phenotype MicroArray (PM) technology to assess their functional capacity. PM's can be used to study gene function, pathogenicity, metabolic capacity and identify drug targets using a series of specialized microtitre plate assays, where each well of the microtitre plate contains a different set of conditions and tests a different phenotype. Cellular respiration is monitored colorimetrically by the reduction of a tetrazolium dye. One clone, SMG 9, was found to be positive for utilization/transport of L-carnitine (a well-characterized osmoprotectant) in the presence of 6% w/v sodium chloride (NaCl). Subsequent experiments revealed a significant growth advantage in minimal media containing NaCl and L-carnitine. Fosmid sequencing revealed putative candidate genes responsible for the phenotype. Subsequent cloning of two genes did not replicate the L-carnitine-associated phenotype, although one of the genes, a σ54-dependent transcriptional regulator, did confer salt tolerance to Escherichia coli when expressed in isolation. The original clone, SMG 9, was subsequently found to have lost the original observed phenotype upon further investigation. Nevertheless, this study demonstrates the usefulness of a phenomic approach to assign a functional role to metagenome-derived clones.
doi:10.3389/fmicb.2014.00189
PMCID: PMC4010731  PMID: 24808895
metagenomics; functional metagenomics; gut microbiome; microbiota; salt tolerance; BIOLOG; phenotype microarray; transcriptional regulator
4.  Draft Genome Sequence of Campylobacter ureolyticus Strain CIT007, the First Whole-Genome Sequence of a Clinical Isolate 
Genome Announcements  2014;2(2):e00262-14.
Herein, we present the draft genome sequence of Campylobacter ureolyticus. Strain CIT007 was isolated from a stool sample from an elderly female presenting with diarrheal illness and end-stage chronic renal disease.
doi:10.1128/genomeA.00262-14
PMCID: PMC3983301  PMID: 24723712
5.  Digitizing humanity 
Artificial DNA, PNA & XNA  2013;4(2):37-38.
The application of ex vivo synthetic DNA as a high capacity information storage medium is well documented. Herein, we consider the potential for synthetic DNA to be incorporated as part of the human genome; providing a definitive, accessible, in vivo database of patient history.
doi:10.4161/adna.25489
PMCID: PMC3771996  PMID: 23912716
synthetic DNA; diagnostics; information; storage; identification
6.  The synthetic biology future 
Bioengineered  2014;5(2):69-72.
Herein, I track the evolution of synthetic biology from its earliest incarnations more than 50 years ago, through the DIYbio revolution, to the next 50 years.
doi:10.4161/bioe.28317
PMCID: PMC4049910  PMID: 24561910
7.  Metagenomic Identification of a Novel Salt Tolerance Gene from the Human Gut Microbiome Which Encodes a Membrane Protein with Homology to a brp/blh-Family β-Carotene 15,15′-Monooxygenase 
PLoS ONE  2014;9(7):e103318.
The human gut microbiome consists of at least 3 million non-redundant genes, 150 times that of the core human genome. Herein, we report the identification and characterisation of a novel stress tolerance gene from the human gut metagenome. The locus, assigned brpA, encodes a membrane protein with homology to a brp/blh-family β-carotene monooxygenase. Cloning and heterologous expression of brpA in Escherichia coli confers a significant salt tolerance phenotype. Furthermore, when cultured in the presence of exogenous β-carotene, cell pellets adopt a red/orange pigmentation indicating the incorporation of carotenoids in the cell membrane.
doi:10.1371/journal.pone.0103318
PMCID: PMC4110020  PMID: 25058308
8.  Letter from the Editor: Bioengineered Bugs begins … 
Bioengineered Bugs  2010;1(1):1.
doi:10.4161/bbug.1.1.10496
PMCID: PMC3035153
9.  Bio-Bites! 
Bioengineered  2013;4(4):183-184.
doi:10.4161/bioe.25287
PMCID: PMC3728186
10.  Ferretting out the facts behind the H5N1 controversy 
Bioengineered Bugs  2012;3(3):139-143.
Recent recommendations by the National Science Advisory Board for Biosecurity (NSABB) to redact key methodological details of two studies involving mammal-to-mammal transmission of the H5N1 (H5) subtype influenza viruses, has led to a temporary moratorium on all research involving live H5N1 or H5 HA reassortant viruses shown to be transmissible in ferrets. Herein, I review the events which led to this impasse and comment on their impact.
doi:10.4161/bbug.19943
PMCID: PMC3370930  PMID: 22402712
11.  Synthetic DNA 
Bioengineered  2013;4(3):123-125.
With world wide data predicted to exceed 40 trillion gigabytes by 2020, big data storage is a very real and escalating problem. Herein, we discuss the utility of synthetic DNA as a robust and eco-friendly archival data storage solution of the future.
doi:10.4161/bioe.24296
PMCID: PMC3669150  PMID: 23514938
12.  A single point mutation in the listerial betL σA-dependent promoter leads to improved osmo- and chill-tolerance and a morphological shift at elevated osmolarity 
Bioengineered  2013;4(6):401-407.
Betaine uptake in Listeria monocytogenes is mediated by three independent transport systems, the simplest of which in genetic terms is the secondary transporter BetL. Using a random mutagenesis approach, based on the E. coli XL1 Red mutator strain, we identified a single point mutation in a putative promoter region upstream of the BetL coding region which leads to a significant increase in betL transcript levels under osmo- and chill-stress conditions and a concomitant increase in stress tolerance. Furthermore, the mutation appears to counter the heretofore unreported “twisted” cell morphology observed for L. monocytogenes grown at elevated osmolarities in tryptone soy broth.
doi:10.4161/bioe.24094
PMCID: PMC3937201  PMID: 23478432
Listeria; osmotolerance; chill-tolerance; salt stress; twisted cells
13.  Proteins 
Bioengineered Bugs  2012;3(2):80-85.
An overwhelming array of structural variants has evolved from a comparatively small number of protein structural domains; which has in turn facilitated an expanse of functional derivatives. Herein, I review the primary mechanisms which have contributed to the vastness of our existing, and expanding, protein repertoires. Protein function prediction strategies, both sequence and structure based, are also discussed and their associated strengths and weaknesses assessed.
doi:10.4161/bbug.18303
PMCID: PMC3357337  PMID: 22095055
circular permutation; combination; divergence; evolution; gene duplication; homology-based transfer; ontologies; protein domains; protein function; sequence and structure motifs
14.  Synthetic ribosomes 
Bioengineered  2013;4(2):63-64.
doi:10.4161/bioe.23640
PMCID: PMC3609622  PMID: 23324614
15.  Analysis of the role of the Cronobacter sakazakii ProP homologues in osmotolerance 
Gut Pathogens  2014;6:15.
Bacteria respond to elevated osmolality by the accumulation of a range of low molecular weight molecules, known as compatible solutes (owing to their compatibility with the cells' normal physiology at high internal concentrations). The neonatal pathogen Cronobacter sakazakii is uniquely osmotolerant, surviving in powdered infant formula (PIF) which typically has a water activity (aw) of 0.2 – inhospitable to most micro-organisms. Mortality rates of up to 80% in infected infants have been recorded making C. sakazakii a serious cause for concern. In silico analysis of the C. sakazakii BAA-894 genome revealed seven copies of the osmolyte uptake system ProP. Herein, we test the physiological role of each of these homologues following heterologous expression against an osmosensitive Escherichia coli host.
doi:10.1186/1757-4749-6-15
PMCID: PMC4047261  PMID: 24910715
Osmolytes; Proline; Osmotolerance; Stress; Cronobacter
16.  Digital biology 
Bioengineered  2012;3(6):311-312.
doi:10.4161/bioe.22367
PMCID: PMC3489704  PMID: 23099453
17.  Draft Genome Sequence of Campylobacter corcagiensis Strain CIT045T, a Representative of a Novel Campylobacter Species Isolated from Lion-Tailed Macaques (Macaca silenus) 
Genome Announcements  2014;2(2):e00248-14.
Campylobacter corcagiensis CIT045T (=CCUG 64942T, LMG 27932T), a new member of the Campylobacter genus, has recently been isolated from lion-tailed macaques in Cork, Ireland. To further characterize this new species and its potential pathogenicity, the genome sequence of C. corcagiensis was determined and is presented here.
doi:10.1128/genomeA.00248-14
PMCID: PMC3990743  PMID: 24744327
18.  Detection and molecular analysis of Campylobacter ureolyticus in domestic animals 
Gut Pathogens  2014;6:9.
Previous studies showed the presence of Campylobacter ureolyticus in a large proportion of diarrhoeal samples from patients in Ireland. This emerging gastrointestinal pathogen was the second most common Campylobacter species detected in patients presenting with gastroenteritis, surpassed only by C. jejuni. However, the source of C. ureolyticus infections in humans remains unknown. The aim of this study was to investigate the presence of C. ureolyticus in a range of domestic animals. Over a period of 6 months, 164 samples collected from various domestic animals were tested using molecular method based on detection of the C. ureolyticus specific hsp60 gene. These included canine faeces (n = 44), feline faeces (n = 31) and porcine faeces (n = 89). C. ureolyticus was detected in 32% (10/31) of feline faeces, 9% (4/44) of canine faeces and 18% (16/89) of porcine faeces. Random Amplified Polymorphic DNA (RAPD) analysis of C. ureolyticus isolates showed that an isolate from a cat is genetically similar to a strain isolated from a patient presenting with gastroenteritis.
This study reports the first detection and isolation of this organism in domestic animals in Ireland, with a potential source for human infection. Together with the previously reported detection of C. ureolyticus in bovine samples, it is likely that this emerging pathogen has a zoonotic potential.
doi:10.1186/1757-4749-6-9
PMCID: PMC3996856  PMID: 24739468
Campylobacter ureolyticus; Emerging pathogen; Reservoirs; Domestic animals; Zoonosis
19.  Mining the human gut microbiome for novel stress resistance genes 
Gut Microbes  2012;3(4):394-397.
With the rapid advances in sequencing technologies in recent years, the human genome is now considered incomplete without the complementing microbiome, which outnumbers human genes by a factor of one hundred. The human microbiome, and more specifically the gut microbiome, has received considerable attention and research efforts over the past decade. Many studies have identified and quantified “who is there?,” while others have determined some of their functional capacity, or “what are they doing?” In a recent study, we identified novel salt-tolerance loci from the human gut microbiome using combined functional metagenomic and bioinformatics based approaches. Herein, we discuss the identified loci, their role in salt-tolerance and their importance in the context of the gut environment. We also consider the utility and power of functional metagenomics for mining such environments for novel genes and proteins, as well as the implications and possible applications for future research.
doi:10.4161/gmic.20984
PMCID: PMC3463497  PMID: 22688726
functional metagenomics; human gut microbiome; salt tolerance; meta-biotechnology
20.  What’s in a name...? 
Bioengineered  2012;3(4):191.
doi:10.4161/bioe.21230
PMCID: PMC3476870  PMID: 22743681
21.  Acinetobacter baumannii 
Virulence  2012;3(3):243-250.
Acinetobacter baumannii is an opportunistic bacterial pathogen primarily associated with hospital-acquired infections. The recent increase in incidence, largely associated with infected combat troops returning from conflict zones, coupled with a dramatic increase in the incidence of multidrug-resistant (MDR) strains, has significantly raised the profile of this emerging opportunistic pathogen. Herein, we provide an overview of the pathogen, discuss some of the major factors that have led to its clinical prominence and outline some of the novel therapeutic strategies currently in development.
doi:10.4161/viru.19700
PMCID: PMC3442836  PMID: 22546906
Acinetobacter baumannii; antibiotic; infection; pathogen; stress; virulence
22.  Improved Detection of Bacterial Pathogens in Patients Presenting with Gastroenteritis by Use of the EntericBio Real-Time Gastro Panel I Assay 
Journal of Clinical Microbiology  2013;51(8):2679-2685.
In this study, we evaluated the use of EntericBio real-time Gastro Panel I (Serosep, Limerick, Ireland) for routine use in a clinical microbiology laboratory for simultaneous detection of Campylobacter jejuni, coli, and lari, Shiga toxin-producing Escherichia coli (STEC), Salmonella spp., and Shigella spp. in feces. This system differs from its predecessor (the EntericBio Panel II system, Serosep) in that it allows real-time detection of pathogens directly from feces, without pre-enrichment. It also specifically detects Campylobacter jejuni, coli, and lari rather than all Campylobacter species, as is the case with the previous system. A total of 528 samples from patients presenting with acute gastroenteritis were screened prospectively with this assay, and results were compared with those of the current method, which combines screening the samples with a molecular assay (the EntericBio Panel II assay) and retrospective culture of the specimens in which the target was detected. Discrepancy analysis was conducted using culture and molecular methods. The real-time assay produced 84 positive results, specifically, Campylobacter spp. (n = 44); Stx1 and/or Stx2 (n = 35); Shigella spp. (n = 3); and Salmonella spp. (n = 6). Of these, 4 samples represented coinfections with Campylobacter spp. and STEC. The real-time assay showed an increased detection rate for pathogens, apart from Salmonella spp. Four Campylobacter-positive and 6 Stx-positive results remained unconfirmed by any other method used. The isolation rates for PCR-positive samples were as follows: Campylobacter spp., 80%; STEC, 45.7%; Salmonella spp., 100%; and Shigella spp., 66.7%. The sensitivity, specificity, positive predictive value, negative predictive value, and efficiency were 100%, 97.8%, 88.1%, 100%, and 98.1%, respectively.
doi:10.1128/JCM.00809-13
PMCID: PMC3719627  PMID: 23761157
23.  Enhanced expression of codon optimized Mycobacterium avium subsp. paratuberculosis antigens in Lactobacillus salivarius 
It is well documented that open reading frames containing high GC content show poor expression in A+T rich hosts. Specifically, G+C-rich codon usage is a limiting factor in heterologous expression of Mycobacterium avium subsp. paratuberculosis (MAP) proteins using Lactobacillus salivarius. However, re-engineering opening reading frames through synonymous substitutions can offset codon bias and greatly enhance MAP protein production in this host. In this report, we demonstrate that codon-usage manipulation of MAP2121c can enhance the heterologous expression of the major membrane protein (MMP), analogous to the form in which it is produced natively by MAP bacilli. When heterologously over-expressed, antigenic determinants were preserved in synthetic MMP proteins as shown by monoclonal antibody mediated ELISA. Moreover, MMP is a membrane protein in MAP, which is also targeted to the cellular surface of recombinant L. salivarius at levels comparable to MAP. Additionally, we previously engineered MAP3733c (encoding MptD) and show herein that MptD displays the tendency to associate with the cytoplasmic membrane boundary under confocal microscopy and the intracellularly accumulated protein selectively adheres to the MptD-specific bacteriophage fMptD. This work demonstrates there is potential for L. salivarius as a viable antigen delivery vehicle for MAP, which may provide an effective mucosal vaccine against Johne's disease.
doi:10.3389/fcimb.2014.00120
PMCID: PMC4154528  PMID: 25237653
MAP antigens; MptD; MMP; codon optimization; expression host; paratuberculosis; MAP vaccine; Johne's disease
24.  Functional Environmental Screening of a Metagenomic Library Identifies stlA; A Unique Salt Tolerance Locus from the Human Gut Microbiome  
PLoS ONE  2013;8(12):e82985.
Functional environmental screening of metagenomic libraries is a powerful means to identify and assign function to novel genes and their encoded proteins without any prior sequence knowledge. In the current study we describe the identification and subsequent analysis of a salt-tolerant clone from a human gut metagenomic library. Following transposon mutagenesis we identified an unknown gene (stlA, for “salt tolerance locus A”) with no current known homologues in the databases. Subsequent cloning and expression in Escherichia coli MKH13 revealed that stlA confers a salt tolerance phenotype in its surrogate host. Furthermore, a detailed in silico analysis was also conducted to gain additional information on the properties of the encoded StlA protein. The stlA gene is rare when searched against human metagenome datasets such as MetaHit and the Human Microbiome Project and represents a novel and unique salt tolerance determinant which appears to be found exclusively in the human gut environment.
doi:10.1371/journal.pone.0082985
PMCID: PMC3861447  PMID: 24349412
25.  Functional metagenomics reveals novel salt tolerance loci from the human gut microbiome 
The ISME Journal  2012;6(10):1916-1925.
Metagenomics is a powerful tool that allows for the culture-independent analysis of complex microbial communities. One of the most complex and dense microbial ecosystems known is that of the human distal colon, with cell densities reaching up to 1012 per gram of faeces. With the majority of species as yet uncultured, there are an enormous number of novel genes awaiting discovery. In the current study, we conducted a functional screen of a metagenomic library of the human gut microbiota for potential salt-tolerant clones. Using transposon mutagenesis, three genes were identified from a single clone exhibiting high levels of identity to a species from the genus Collinsella (closest relative being Collinsella aerofaciens) (COLAER_01955, COLAER_01957 and COLAER_01981), a high G+C, Gram-positive member of the Actinobacteria commonly found in the human gut. The encoded proteins exhibit a strong similarity to GalE, MurB and MazG. Furthermore, pyrosequencing and bioinformatic analysis of two additional fosmid clones revealed the presence of an additional galE and mazG gene, with the highest level of genetic identity to Akkermansia muciniphila and Eggerthella sp. YY7918, respectively. Cloning and heterologous expression of the genes in the osmosensitive strain, Escherichia coli MKH13, resulted in increased salt tolerance of the transformed cells. It is hoped that the identification of atypical salt tolerance genes will help to further elucidate novel salt tolerance mechanisms, and will assist our increased understanding how resident bacteria cope with the osmolarity of the gastrointestinal tract.
doi:10.1038/ismej.2012.38
PMCID: PMC3446828  PMID: 22534607
Akkermansia ;  Collinsella ;  Eggerthella ; human gut microbiome; metagenomics; salt tolerance

Results 1-25 (53)