Menu

Auto Tag: Virulence

July 7, 2019

Genomics-driven discovery of the gliovirin biosynthesis gene cluster in the plant beneficial fungus Trichoderma virens

Gliovirin is a strong anti-oomycete and a candidate anticancer compound. It is produced by “P” strains of the plant disease biocontrol fungus Trichoderma virens and is involved in biological control of certain plant pathogens. Even though the compound is known for more than three decades, neither the genes involved nor the biosynthetic pathway are known. We have sequenced the whole genome of a gliovirin producing strain of T. virens and discovered a novel gene cluster comprising of 22 genes. Disruption of the non-ribosomal peptide synthetase eliminated biosynthesis of gliovirin. The gene cluster is very similar to a hitherto un-described gene cluster of Aspergillus udagawae, a human pathogen. Our findings open-up the possibility of strain improvement of T. virens for improved biocontrol of plant diseases through enhanced production of gliovirin. Research also can now be initiated on the role of this gene cluster in pathogenicity of the human pathogen A. udagawae.

Read more
July 7, 2019

Expanding landscapes of the diversified mcr-1-bearing plasmid reservoirs.

Polymyxin is a cationic polypeptide antibiotic that can disrupt bacterial cell membrane by interacting with its lipopolysaccharide molecules and is used as a last resort drug against lethal infections by the carbapenem-resistant superbugs (like NDM-1). However, global discovery of the MCR-1 colistin resistance dramatically challenges the newly renewed interest in colistin for clinical use.The mcr-1-harboring plasmids were acquired from swine and human Escherichia coli isolated in China, from 2015 to 2016, and subjected to Illumina PacBio RSII and Hi-Seq2000 for full genome sequencing. PCR was applied to close the gap of the assembled contigs. Ori-Finder was employed to predict the replication origin (oriC) in plasmids. The phenotype of MCR-1-producing isolates was evaluated on the LBA plates with various level of colistin. Genetic deletion was used to test the requirement of the initial “ATG” codon for the MCR-1 function.Here, we report full genomes of over 10 mcr-1-harboring plasmids with diversified replication incompatibilities. A novel hybrid IncI2/IncFIB plasmid pGD17-2 was discovered and characterized from a swine isolate with colistin resistance. Intriguingly, co-occurrence of two unique mcr-1-bearing plasmids (pGD65-3, IncI2, and pGD65-5, IncX4) was detected in a single isolate GD65, which might accelerate dissemination of the mcr-1 under environmental selection pressure. Genetic analyses of these plasmids mapped mobile elements in the context of antibiotic resistance and determined two insertion sequences (ISEcp1 and ISApl1) that are responsible for the mobilization of mcr-1. Gene deletion also proved that the first ATG codon is redundant in the mcr-1 gene.Collectively, our results extend landscapes of the diversified mcr-1-bearing plasmid reservoirs.

Read more
July 7, 2019

Rapid and consistent evolution of colistin resistance in XDR Pseudomonas aeruginosa during morbidostat culture.

Colistin is a last resort antibiotic commonly used against multidrug-resistant strains of Pseudomonas aeruginosa To investigate the potential for in-situ evolution of resistance against colistin and to map the molecular targets of colistin resistance, we exposed two P. aeruginosa isolates to colistin using a continuous culture device known as morbidostat. As a result, colistin resistance reproducibly increased 10-fold within ten days, and 100-fold within 20 days, along with highly stereotypic, yet strain specific mutation patterns. The majority of mutations hit the pmrAB two component signaling system and genes involved in lipopolysaccharide (LPS) synthesis, including lpxC, pmrE, and migA We tracked the frequencies of all arising mutations by whole genome deep sequencing every 3-4 days to provide a detailed picture of the dynamics of resistance evolution, including competition and displacement among multiple resistant sub-populations. In seven out of 18 cultures, we observed mutations in mutS along with a mutator phenotype that seemed to facilitate resistance evolution. Copyright © 2017 American Society for Microbiology.

Read more
July 7, 2019

Comparative genomic and phylogenetic analysis of a toxigenic clinical isolate of Corynebacterium diphtheriae strain B-D-16-78 from Malaysia.

In this study, we report the comparative genomics and phylogenetic analysis of Corynebacterium diphtheriae strain B-D-16-78 that was isolated from a clinical specimen in 2016. The complete genome of C. diphtheriae strain B-D-16-78 was sequenced using PacBio Single Molecule, Real-Time sequencing technology and consists of a 2,474,151-bp circular chromosome with an average GC content of 53.56%. The core genome of C. diphtheriae was also deduced from a total of 74 strains with complete or draft genome sequences and the core genome-based phylogenetic analysis revealed close genetic relationship among strains that shared the same MLST allelic profile. In the context of CRISPR-Cas system, which confers adaptive immunity against re-invading DNA, 73 out of 86 spacer sequences were found to be unique to Malaysian strains which harboured only type-II-C and/or type-I-E-a systems. A total of 48 tox genes which code for the diphtheria toxin were retrieved from the 74 genomes and with the exception of one truncated gene, only nucleotide substitutions were detected when compared to the tox gene sequence of PW8. More than half were synonymous substitution and only two were nonsynonymous substitutions whereby H24Y was predicted to have a damaging effect on the protein function whilst T262V was predicted to be tolerated. Both toxigenic and non-toxigenic toxin-gene bearing strains have been isolated in Malaysia but the repeated isolation of toxigenic strains with the same MLST profile suggests the possibility of some of these strains may be circulating in the population. Hence, efforts to increase herd immunity should be continued and supported by an effective monitoring and surveillance system to track, manage and control outbreak of cases. Copyright © 2017 Elsevier B.V. All rights reserved.

Read more
July 7, 2019

Phenotypic and genomic comparison of Mycobacterium aurum and surrogate model species to Mycobacterium tuberculosis: implications for drug discovery.

Tuberculosis (TB) is caused by Mycobacterium tuberculosis and represents one of the major challenges facing drug discovery initiatives worldwide. The considerable rise in bacterial drug resistance in recent years has led to the need of new drugs and drug regimens. Model systems are regularly used to speed-up the drug discovery process and circumvent biosafety issues associated with manipulating M. tuberculosis. These include the use of strains such as Mycobacterium smegmatis and Mycobacterium marinum that can be handled in biosafety level 2 facilities, making high-throughput screening feasible. However, each of these model species have their own limitations.We report and describe the first complete genome sequence of Mycobacterium aurum ATCC23366, an environmental mycobacterium that can also grow in the gut of humans and animals as part of the microbiota. This species shows a comparable resistance profile to that of M. tuberculosis for several anti-TB drugs. The aims of this study were to (i) determine the drug resistance profile of a recently proposed model species, Mycobacterium aurum, strain ATCC23366, for anti-TB drug discovery as well as Mycobacterium smegmatis and Mycobacterium marinum (ii) sequence and annotate the complete genome sequence of this species obtained using Pacific Bioscience technology (iii) perform comparative genomics analyses of the various surrogate strains with M. tuberculosis (iv) discuss how the choice of the surrogate model used for drug screening can affect the drug discovery process.We describe the complete genome sequence of M. aurum, a surrogate model for anti-tuberculosis drug discovery. Most of the genes already reported to be associated with drug resistance are shared between all the surrogate strains and M. tuberculosis. We consider that M. aurum might be used in high-throughput screening for tuberculosis drug discovery. We also highly recommend the use of different model species during the drug discovery screening process.

Read more
July 7, 2019

Comparative genomic analysis of Acinetobacter strains isolated from murine colonic crypts.

A restricted set of aerobic bacteria dominated by the Acinetobacter genus was identified in murine intestinal colonic crypts. The vicinity of such bacteria with intestinal stem cells could indicate that they protect the crypt against cytotoxic and genotoxic signals. Genome analyses of these bacteria were performed to better appreciate their biodegradative capacities.Two taxonomically different clusters of Acinetobacter were isolated from murine proximal colonic crypts, one was identified as A. modestus and the other as A. radioresistens. Their identification was performed through biochemical parameters and housekeeping gene sequencing. After selection of one strain of each cluster (A. modestus CM11G and A. radioresistens CM38.2), comparative genomic analysis was performed on whole-genome sequencing data. The antibiotic resistance pattern of these two strains is different, in line with the many genes involved in resistance to heavy metals identified in both genomes. Moreover whereas the operon benABCDE involved in benzoate metabolism is encoded by the two genomes, the operon antABC encoding the anthranilate dioxygenase, and the phenol hydroxylase gene cluster are absent in the A. modestus genomic sequence, indicating that the two strains have different capacities to metabolize xenobiotics. A common feature of the two strains is the presence of a type IV pili system, and the presence of genes encoding proteins pertaining to secretion systems such as Type I and Type II secretion systems.Our comparative genomic analysis revealed that different Acinetobacter isolated from the same biological niche, even if they share a large majority of genes, possess unique features that could play a specific role in the protection of the intestinal crypt.

Read more
July 7, 2019

Complete genome sequences of three Xanthomonas citri strains from Texas.

The complete genome sequences of three Xanthomonas citri strains isolated from lime trees in Texas were found to belong to the A(w) group. All carried nearly identical large plasmids with similarity to those of a citrus canker strain from India and to xanthomonads from Africa and Colombia. All three strains harbored unusual pthA homologs. Copyright © 2017 Munoz Bodnar et al.

Read more
July 7, 2019

Genomic insights into the virulence and salt tolerance of Staphylococcus equorum.

To shed light on the genetic background behind the virulence and salt tolerance of Staphylococcus equorum, we performed comparative genome analysis of six S. equorum strains. Data on four previously published genome sequences were obtained from the NCBI database, while those on strain KM1031 displaying resistance to multiple antibiotics and strain C2014 causing haemolysis were determined in this study. Examination of the pan-genome of five of the six S. equorum strains showed that the conserved core genome retained the genes for general physiological processes and survival of the species. In this comparative genomic analysis, the factors that distinguish the strains from each other, including acquired genomic factors in mobile elements, were identified. Additionally, the high salt tolerance of strains enabling growth at a NaCl concentration of 25% (w/v) was attributed to the genes encoding potassium voltage-gated channels. Among the six strains, KS1039 does not possess any of the functional virulence determinants expressed in the other strains.

Read more
July 7, 2019

Plasmid composition in Aeromonas salmonicida subsp. salmonicida 01-B526 unravels unsuspected type three secretion system loss patterns.

Aeromonas salmonicida subsp. salmonicida is a ubiquitous psychrophilic waterborne bacterium and a fish pathogen. The numerous mobile elements, especially insertion sequences (IS), in its genome promote rearrangements that impact its phenotype. One of the main virulence factors of this bacterium, its type three secretion system (TTSS), is affected by these rearrangements. In Aeromonas salmonicida subsp. salmonicida most of the TTSS genes are encoded in a single locus on a large plasmid called pAsa5, and may be lost when the bacterium is cultivated at a higher temperature (25 °C), producing non-virulent mutants. In a previous study, pAsa5-rearranged strains that lacked the TTSS locus on pAsa5 were produced using parental strains, including 01-B526. Some of the generated deletions were explained by homologous recombination between ISs found on pAsa5, whereas the others remained unresolved. To investigate those rearrangements, short- and long-read high-throughput sequencing technologies were used on the A. salmonicida subsp. salmonicida 01-B526 whole genome.Whole genome sequencing of the 01-B526 strain revealed that its pAsa5 has an additional IS copy, an ISAS5, compared to the reference strain (A449) sequence, which allowed for a previously unknown rearrangement to occur. It also appeared that 01-B526 bears a second large plasmid, named pAsa9, which shares 40 kbp of highly similar sequences with pAsa5. Following these discoveries, previously unexplained deletions were elucidated by genotyping. Furthermore, in one of the derived strains a fusion of pAsa5 and pAsa9, involving the newly discovered ISAS5 copy, was observed.The loss of TTSS and hence virulence is explained by one consistent mechanism: IS-driven homologous recombination. The similarities between pAsa9 and pAsa5 also provide another example of genetic diversity driven by ISs.

Read more
July 7, 2019

Evidence for contemporary switching of the O-antigen gene cluster between Shiga toxin-producing Escherichia coli strains colonizing cattle.

Shiga toxin-producing Escherichia coli (STEC) comprise a group of zoonotic enteric pathogens with ruminants, especially cattle, as the main reservoir. O-antigens are instrumental for host colonization and bacterial niche adaptation. They are highly immunogenic and, therefore, targeted by the adaptive immune system. The O-antigen is one of the most diverse bacterial cell constituents and variation not only exists between different bacterial species, but also between individual isolates/strains within a single species. We recently identified STEC persistently infecting cattle and belonging to the different serotypes O156:H25 (n = 21) and O182:H25 (n = 15) that were of the MLST sequence types ST300 or ST688. These STs differ by a single nucleotide in purA only. Fitness-, virulence-associated genome regions, and CRISPR/CAS (clustered regularly interspaced short palindromic repeats/CRISPR associated sequence) arrays of these STEC O156:H25 and O182:H25 isolates were highly similar, and identical genomic integration sites for the stx converting bacteriophages and the core LEE, identical Shiga toxin converting bacteriophage genes for stx1a, identical complete LEE loci, and identical sets of chemotaxis and flagellar genes were identified. In contrast to this genomic similarity, the nucleotide sequences of the O-antigen gene cluster (O-AGC) regions between galF and gnd and very few flanking genes differed fundamentally and were specific for the respective serotype. Sporadic aEPEC O156:H8 isolates (n = 5) were isolated in temporal and spatial proximity. While the O-AGC and the corresponding 5′ and 3′ flanking regions of these aEPEC isolates were identical to the respective region in the STEC O156:H25 isolates, the core genome, the virulence associated genome regions and the CRISPR/CAS elements differed profoundly. Our cumulative epidemiological and molecular data suggests a recent switch of the O-AGC between isolates with O156:H8 strains having served as DNA donors. Such O-antigen switches can affect the evaluation of a strain’s pathogenic and virulence potential, suggesting that NGS methods might lead to a more reliable risk assessment.

Read more
July 7, 2019

Comparative genomic analysis reveals genetic features related to the virulence of Bacillus cereus FORC_013.

Bacillus cereus is well known as a gastrointestinal pathogen that causes food-borne illness. In the present study, we sequenced the complete genome of B. cereus FORC_013 isolated from fried eel in South Korea. To extend our understanding of the genomic characteristics of FORC_013, we conducted a comparative analysis with the published genomes of other B. cereus strains.We fully assembled the single circular chromosome (5,418,913 bp) and one plasmid (259,749 bp); 5511 open reading frames (ORFs) and 283 ORFs were predicted for the chromosome and plasmid, respectively. Moreover, we detected that the enterotoxin (NHE, HBL, CytK) induces food-borne illness with diarrheal symptom, and that the pleiotropic regulator, along with other virulence factors, plays a role in surviving and biofilm formation. Through comparative analysis using the complete genome sequence of B. cereus FORC_013, we identified both positively selected genes related to virulence regulation and 224 strain-specific genes of FORC_013.Through genome analysis of B. cereus FORC_013, we identified multiple virulence factors that may contribute to pathogenicity. These results will provide insight into further studies regarding B. cereus pathogenesis mechanism at the genomic level.

Read more
July 7, 2019

Evidence for the evolutionary steps leading to mecA-mediated ß-lactam resistance in staphylococci.

The epidemiologically most important mechanism of antibiotic resistance in Staphylococcus aureus is associated with mecA-an acquired gene encoding an extra penicillin-binding protein (PBP2a) with low affinity to virtually all ß-lactams. The introduction of mecA into the S. aureus chromosome has led to the emergence of methicillin-resistant S. aureus (MRSA) pandemics, responsible for high rates of mortality worldwide. Nonetheless, little is known regarding the origin and evolution of mecA. Different mecA homologues have been identified in species belonging to the Staphylococcus sciuri group representing the most primitive staphylococci. In this study we aimed to identify evolutionary steps linking these mecA precursors to the ß-lactam resistance gene mecA and the resistance phenotype. We sequenced genomes of 106 S. sciuri, S. vitulinus and S. fleurettii strains and determined their oxacillin susceptibility profiles. Single-nucleotide polymorphism (SNP) analysis of the core genome was performed to assess the genetic relatedness of the isolates. Phylogenetic analysis of the mecA gene homologues and promoters was achieved through nucleotide/amino acid sequence alignments and mutation rates were estimated using a Bayesian analysis. Furthermore, the predicted structure of mecA homologue-encoded PBPs of oxacillin-susceptible and -resistant strains were compared. We showed for the first time that oxacillin resistance in the S. sciuri group has emerged multiple times and by a variety of different mechanisms. Development of resistance occurred through several steps including structural diversification of the non-binding domain of native PBPs; changes in the promoters of mecA homologues; acquisition of SCCmec and adaptation of the bacterial genetic background. Moreover, our results suggest that it was exposure to ß-lactams in human-created environments that has driven evolution of native PBPs towards a resistance determinant. The evolution of ß-lactam resistance in staphylococci highlights the numerous resources available to bacteria to adapt to the selective pressure of antibiotics.

Read more
July 7, 2019

Reclassification of the specialized metabolite producer Pseudomonas mesoacidophila ATCC 31433 as a member of the Burkholderia cepacia complex.

Pseudomonas mesoacidophila ATCC 31433 is a Gram-negative bacterium, first isolated from Japanese soil samples, that produces the monobactam isosulfazecin and the ß-lactam-potentiating bulgecins. To characterize the biosynthetic potential of P. mesoacidophila ATCC 31433, its complete genome was determined using single-molecule real-time DNA sequence analysis. The 7.8-Mb genome comprised four replicons, three chromosomal (each encoding rRNA) and one plasmid. Phylogenetic analysis demonstrated that P. mesoacidophila ATCC 31433 was misclassified at the time of its deposition and is a member of the Burkholderia cepacia complex, most closely related to Burkholderia ubonensis The sequenced genome shows considerable additional biosynthetic potential; known gene clusters for malleilactone, ornibactin, isosulfazecin, alkylhydroxyquinoline, and pyrrolnitrin biosynthesis and several uncharacterized biosynthetic gene clusters for polyketides, nonribosomal peptides, and other metabolites were identified. Furthermore, P. mesoacidophila ATCC 31433 harbors many genes associated with environmental resilience and antibiotic resistance and was resistant to a range of antibiotics and metal ions. In summary, this bioactive strain should be designated B. cepacia complex strain ATCC 31433, pending further detailed taxonomic characterization.IMPORTANCE This work reports the complete genome sequence of Pseudomonas mesoacidophila ATCC 31433, a known producer of bioactive compounds. Large numbers of both known and novel biosynthetic gene clusters were identified, indicating that P. mesoacidophila ATCC 31433 is an untapped resource for discovery of novel bioactive compounds. Phylogenetic analysis demonstrated that P. mesoacidophila ATCC 31433 is in fact a member of the Burkholderia cepacia complex, most closely related to the species Burkholderia ubonensis Further investigation of the classification and biosynthetic potential of P. mesoacidophila ATCC 31433 is warranted. Copyright © 2017 Loveridge et al.

Read more
July 7, 2019

Synergistic interaction between phage therapy and antibiotics clears Pseudomonas aeruginosa infection in endocarditis and reduces virulence.

Increasing antibiotic resistance warrants therapeutic alternatives. Here we investigated the efficacy of bacteriophage-therapy (phage) alone or combined with antibiotics against experimental endocarditis (EE) due to Pseudomonas aeruginosa, an archetype of difficult-to-treat infection.In vitro fibrin clots and rats with aortic EE were treated with an antipseudomonas phage cocktail alone or combined with ciprofloxacin. Phage pharmacology, therapeutic efficacy, and resistance were determined.In vitro, single-dose phage therapy killed 7 log colony-forming units (CFUs)/g of fibrin clots in 6 hours. Phage-resistant mutants regrew after 24 hours but were prevented by combination with ciprofloxacin (2.5 × minimum inhibitory concentration). In vivo, single-dose phage therapy killed 2.5 log CFUs/g of vegetations in 6 hours (P < .001 vs untreated controls) and was comparable with ciprofloxacin monotherapy. Moreover, phage/ciprofloxacin combinations were highly synergistic, killing >6 log CFUs/g of vegetations in 6 hours and successfully treating 64% (n = 7/11) of rats. Phage-resistant mutants emerged in vitro but not in vivo, most likely because resistant mutations affected bacterial surface determinants important for infectivity (eg, the pilT and galU genes involved in pilus motility and LPS formation).Single-dose phage therapy was active against P. aeruginosa EE and highly synergistic with ciprofloxacin. Phage-resistant mutants had impaired infectivity. Phage-therapy alone or combined with antibiotics merits further clinical consideration.

Read more
July 7, 2019

Whole-genome comparative analysis of Salmonella enterica serovar Newport strains reveals lineage-specific divergence.

Salmonella enterica subsp. enterica serovar Newport has been associated with various foodborne outbreaks in humans and animals. Phylogenetically, serovar Newport is one of several Salmonella serovars that are polyphyletic. To understand more about the polyphyletic nature of this serovar, six food, environment, and human isolates from different Newport lineages were selected for genome comparison analyses. Whole genome comparisons demonstrated that heterogeneity mostly occurred in the prophage regions. Lineage-specific characteristics were also present in the Salmonella pathogenicity islands and fimbrial operons. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution 2017. This work is written by US Government employees and is in the public domain in the US.

Read more

Subscribe for blog updates:

Talk with an expert

If you have a question, need to check the status of an order, or are interested in purchasing an instrument, we're here to help.