scholarly journals Genomics of an endemic cystic fibrosis Burkholderia multivorans strain reveals low within-patient evolution but high between-patient diversity

2021 ◽  
Vol 17 (3) ◽  
pp. e1009418
Author(s):  
Cédric Lood ◽  
Charlotte Peeters ◽  
Quentin Lamy-Besnier ◽  
Jeroen Wagemans ◽  
Daniel De Vos ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Burkholderia Cepacia ◽  
Genomic Analysis ◽  
Genomic Diversity ◽  
Epidemiological Surveillance ◽  
Nucleotide Polymorphisms ◽  
Whole Genome Analysis ◽  
Structural Genomic ◽  
Burkholderia Multivorans ◽  
Sequencing Technologies

Burkholderia multivorans is a member of the Burkholderia cepacia complex (Bcc), notorious for its pathogenicity in persons with cystic fibrosis. Epidemiological surveillance suggests that patients predominantly acquire B. multivorans from environmental sources, with rare cases of patient-to-patient transmission. Here we report on the genomic analysis of thirteen isolates from an endemic B. multivorans strain infecting four cystic fibrosis patients treated in different pediatric cystic fibrosis centers in Belgium, with no evidence of cross-infection. All isolates share an identical sequence type (ST-742) but whole genome analysis shows that they exhibit peculiar patterns of genomic diversity between patients. By combining short and long reads sequencing technologies, we highlight key differences in terms of small nucleotide polymorphisms indicative of low rates of adaptive evolution within patient, and well-defined, hundred Kbps-long segments of high enrichment in mutations between patients. In addition, we observed large structural genomic variations amongst the isolates which revealed different plasmid contents, active roles for transposase IS3 and IS5 in the deactivation of genes, and mobile prophage elements. Our study shows limited within-patient B. multivorans evolution and high between-patient strain diversity, indicating that an environmental microdiverse reservoir must be present for this endemic strain, in which active diversification is taking place. Furthermore, our analysis also reveals a set of 30 parallel adaptations across multiple patients, indicating that the specific genomic background of a given strain may dictate the route of adaptation within the cystic fibrosis lung.

scholarly journals Mucoid morphotype variation of Burkholderia multivorans during chronic cystic fibrosis lung infection is correlated with changes in metabolism, motility, biofilm formation and virulence

Microbiology ◽  
2011 ◽  
Vol 157 (11) ◽  
pp. 3124-3137 ◽  
Author(s):  
Inês N. Silva ◽  
Ana S. Ferreira ◽  
Jörg D. Becker ◽  
James E. A. Zlosnik ◽  
David P. Speert ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Phenotypic Variation ◽  
Biofilm Formation ◽  
Burkholderia Cepacia ◽  
Galleria Mellonella ◽  
Acute Infection ◽  
Lung Infection ◽  
Infection Model ◽  
Burkholderia Multivorans ◽  
Expression Of Genes

Burkholderia cepacia complex (Bcc) bacteria are opportunistic pathogens infecting hosts such as cystic fibrosis (CF) patients. Long-term Bcc infection of CF patients’ airways has been associated with emergence of phenotypic variation. Here we studied two Burkholderia multivorans clonal isolates displaying different morphotypes from a chronically infected CF patient to evaluate trait development during lung infection. Expression profiling of mucoid D2095 and non-mucoid D2214 isolates revealed decreased expression of genes encoding products related to virulence-associated traits and metabolism in D2214. Furthermore, D2214 showed no exopolysaccharide production, lower motility and chemotaxis, and more biofilm formation, particularly under microaerophilic conditions, than the clonal mucoid isolate D2095. When Galleria mellonella was used as acute infection model, D2214 at a cell number of approximately 7×106 c.f.u. caused a higher survival rate than D2095, although 6 days post-infection most of the larvae were dead. Infection with the same number of cells by mucoid D2095 caused larval death by day 4. The decreased expression of genes involved in carbon and nitrogen metabolism may reflect lower metabolic needs of D2214 caused by lack of exopolysaccharide, but also by the attenuation of pathways not required for survival. As a result, D2214 showed higher survival than D2095 in minimal medium for 28 days under aerobic conditions. Overall, adaptation during Bcc chronic lung infections gave rise to genotypic and phenotypic variation among isolates, contributing to their fitness while maintaining their capacity for survival in this opportunistic human niche.

scholarly journals Genomic analysis unveils important aspects of population structure, virulence, and antimicrobial resistance inKlebsiella aerogenes

2019 ◽  
Author(s):  
Hemanoel Passarelli-Araujo ◽  
Jussara K. Palmeiro ◽  
Kanhu C. Moharana ◽  
Francisnei Pedrosa-Silva ◽  
Libera M. Dalla-Costa ◽  
...  
Keyword(s):  
Population Structure ◽  
Genomic Analysis ◽  
Genomic Diversity ◽  
Nucleotide Polymorphisms ◽  
Effective Population ◽  
Single Nucleotide ◽  
Deleterious Alleles ◽  
Large Effective Population Size ◽  
Healthcare Associated ◽  
Antibiotic Efflux

ABSTRACTKlebsiella aerogenesis an important pathogen in healthcare-associated infections. Nevertheless, in comparison to other clinically important pathogens,K. aerogenespopulation structure, genetic diversity, and pathogenicity remain poorly understood. Here, we elucidateK. aerogenesclonal complexes (CCs) and genomic features associated with resistance and virulence. We present a detailed description of the population structure ofK. aerogenesbased on 97 publicly available genomes by using both, multilocus sequence typing and single nucleotide polymorphisms extracted from core genome. We also assessed virulence and resistance profiles using VFDB and CARD, respectively. We show thatK. aerogeneshas an open pangenome and a large effective population size, which account for its high genomic diversity and support that negative selection prevents fixation of most deleterious alleles. The population is structured in at least ten CCs, including two novel ones identified here, CC9 and CC10. The repertoires of resistance genes comprise a high number of antibiotic efflux proteins as well as narrow and extended spectrum β-lactamases. Regarding the population structure, we identified two clusters based on virulence profile due to the presence of the toxin-encodingclboperon and the siderophore production genes,irpandybt.Notably, CC3 comprises the majority ofK. aerogenesisolates associated with hospital outbreaks, emphasizing the importance of its constant monitoring. Collectively, our results can be useful in the development of new therapeutic and surveillance strategies worldwide.

scholarly journals Long-Term Evolution of Burkholderia multivorans during a Chronic Cystic Fibrosis Infection Reveals Shifting Forces of Selection

mSystems ◽  
2016 ◽  
Vol 1 (3) ◽  
Author(s):  
Inês N. Silva ◽  
Pedro M. Santos ◽  
Mário R. Santos ◽  
James E. A. Zlosnik ◽  
David P. Speert ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Lung Function ◽  
Antimicrobial Resistance ◽  
Opportunistic Pathogen ◽  
Content Type ◽  
Burkholderia Multivorans ◽  
Strong Selection ◽  
Sequencing Technologies ◽  
Bacterial Genes

ABSTRACT Bacteria may become genetically and phenotypically diverse during long-term colonization of cystic fibrosis (CF) patient lungs, yet our understanding of within-host evolutionary processes during these infections is lacking. Here we combined current genome sequencing technologies and detailed phenotypic profiling of the opportunistic pathogen Burkholderia multivorans using sequential isolates sampled from a CF patient over 20 years. The evolutionary history of these isolates highlighted bacterial genes and pathways that were likely subject to strong selection within the host and were associated with altered phenotypes, such as biofilm production, motility, and antimicrobial resistance. Importantly, multiple lineages coexisted for years or even decades within the infection, and the period of diversification within the dominant lineage was associated with deterioration of the patient’s lung function. Identifying traits under strong selection during chronic infection not only sheds new light onto Burkholderia evolution but also sets the stage for tailored therapeutics targeting the prevailing lineages associated with disease progression. Burkholderia multivorans is an opportunistic pathogen capable of causing severe disease in patients with cystic fibrosis (CF). Patients may be chronically infected for years, during which the bacterial population evolves in response to unknown forces. Here we analyze the genomic and functional evolution of a B. multivorans infection that was sequentially sampled from a CF patient over 20 years. The population diversified into at least four primary, coexisting clades with distinct evolutionary dynamics. The average substitution rate was only 2.4 mutations/year, but notably, some lineages evolved more slowly, whereas one diversified more rapidly by mostly nonsynonymous mutations. Ten loci, mostly involved in gene expression regulation and lipid metabolism, acquired three or more independent mutations and define likely targets of selection. Further, a broad range of phenotypes changed in association with the evolved mutations; they included antimicrobial resistance, biofilm regulation, and the presentation of lipopolysaccharide O-antigen repeats, which was directly caused by evolved mutations. Additionally, early isolates acquired mutations in genes involved in cyclic di-GMP (c-di-GMP) metabolism that associated with increased c-di-GMP intracellular levels. Accordingly, these isolates showed lower motility and increased biofilm formation and adhesion to CFBE41o− epithelial cells than the initial isolate, and each of these phenotypes is an important trait for bacterial persistence. The timing of the emergence of this clade of more adherent genotypes correlated with the period of greatest decline in the patient’s lung function. All together, our observations suggest that selection on B. multivorans populations during long-term colonization of CF patient lungs either directly or indirectly targets adherence, metabolism, and changes in the cell envelope related to adaptation to the biofilm lifestyle. IMPORTANCE Bacteria may become genetically and phenotypically diverse during long-term colonization of cystic fibrosis (CF) patient lungs, yet our understanding of within-host evolutionary processes during these infections is lacking. Here we combined current genome sequencing technologies and detailed phenotypic profiling of the opportunistic pathogen Burkholderia multivorans using sequential isolates sampled from a CF patient over 20 years. The evolutionary history of these isolates highlighted bacterial genes and pathways that were likely subject to strong selection within the host and were associated with altered phenotypes, such as biofilm production, motility, and antimicrobial resistance. Importantly, multiple lineages coexisted for years or even decades within the infection, and the period of diversification within the dominant lineage was associated with deterioration of the patient’s lung function. Identifying traits under strong selection during chronic infection not only sheds new light onto Burkholderia evolution but also sets the stage for tailored therapeutics targeting the prevailing lineages associated with disease progression.

scholarly journals Characterization of the AmpC β-Lactamase fromBurkholderia multivorans

2018 ◽  
Vol 62 (10) ◽  
Author(s):  
Scott A. Becka ◽  
Elise T. Zeiser ◽  
Melissa D. Barnes ◽  
Magdalena A. Taracila ◽  
Kevin Nguyen ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Active Site ◽  
Burkholderia Cepacia ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Enzyme Complex ◽  
Active Site Residues ◽  
Content Type ◽  
Burkholderia Multivorans

ABSTRACTBurkholderia multivoransis a member of theBurkholderia cepaciacomplex, a group of >20 related species of nosocomial pathogens that commonly infect individuals suffering from cystic fibrosis. β-Lactam antibiotics are recommended as therapy for infections due toB.multivorans, which possesses two β-lactamase genes,blapenAandblaAmpC. PenA is a carbapenemase with a substrate profile similar to that of theKlebsiella pneumoniaecarbapenemase (KPC); in addition, expression of PenA is inducible by β-lactams inB.multivorans. Here, we characterize AmpC fromB.multivoransATCC 17616. AmpC possesses only 38 to 46% protein identity with non-BurkholderiaAmpC proteins (e.g., PDC-1 and CMY-2). Among 49 clinical isolates ofB.multivorans, we identified 27 different AmpC variants. Some variants possessed single amino acid substitutions within critical active-site motifs (Ω loop and R2 loop). Purified AmpC1 demonstrated minimal measurable catalytic activity toward β-lactams (i.e., nitrocefin and cephalothin). Moreover, avibactam was a poor inhibitor of AmpC1 (Kiapp> 600 μM), and acyl-enzyme complex formation with AmpC1 was slow, likely due to lack of productive interactions with active-site residues. Interestingly, immunoblotting using a polyclonal anti-AmpC antibody revealed that protein expression of AmpC1 was inducible inB.multivoransATCC 17616 after growth in subinhibitory concentrations of imipenem (1 μg/ml). AmpC is a unique inducible class C cephalosporinase that may play an ancillary role inB.multivoranscompared to PenA, which is the dominant β-lactamase inB.multivoransATCC 17616.

scholarly journals A histone-like nucleoid structuring protein regulates several virulence traits in Burkholderia multivorans

2021 ◽  
Author(s):  
Sara C. Gomes ◽  
Mirela R. Ferreira ◽  
Andreia F. Tavares ◽  
Inês N. Silva ◽  
Jörg D. Becker ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cystic Fibrosis ◽  
Burkholderia Cepacia ◽  
Gc Content ◽  
Mobile Element ◽  
Surface Hydrophobicity ◽  
Upstream Region ◽  
Cell Physiology ◽  
Burkholderia Multivorans ◽  
Virulence Traits

Burkholderia cepacia complex bacteria comprises opportunistic pathogens causing chronic respiratory infections in cystic fibrosis (CF) patients. These microorganisms produce an exopolysaccharide named cepacian, which is considered a virulence determinant. To find genes implicated in the regulation of cepacian biosynthesis, we characterized an evolved nonmucoid variant (17616nmv) derived from the ancestor, Burkholderia multivorans ATCC 17616, after prolonged stationary phase. Lack of cepacian biosynthesis was correlated with downregulation of the expression of bce genes implicated in its biosynthesis. Furthermore, genome sequencing of the variant identified the transposition of the mobile element IS406 upstream the coding sequence of an hns-like gene (Bmul_0158) encoding a histone-like nucleoid structuring protein, a known global transcriptional repressor. This IS element upregulated the expression of Bmul_0158 by 4-fold. Transcriptome analysis identified the global effects of this mutation on gene expression, with major changes in genes implicated in motility, pili synthesis, type VI secretion, and chromosome associated functions. Concomitant with these differences, the nonmucoid variant displays reduced adherence to a CF lung bronchial cell line, reduced surface hydrophobicity, forms smaller cellular aggregates, but has an increase in swimming and swarming motilities. Finally, analysis of the GC content of the upstream region of differentially expressed genes led to the identification of various genomic regions, possibly acquired by horizontal gene transfer, which were transcriptionally repressed by the increased expression of Bmul_0158 gene in the 17616nmv strain. Taken together, the results revealed a significant role for this H-NS protein in the regulation of B. multivorans persistence- and virulence-associated genes. IMPORTANCE Members of the histone-like nucleoid-structuring (H-NS) family of proteins, present in many bacteria, are important global regulators of gene expression. Many of the regulated genes were acquired horizontally and include pathogenicity islands and prophages, among others. Additionally, H-NS can play a structural role by bridging and compacting DNA, presenting a crucial role in cell physiology. Several virulence phenotypes have been frequently identified in several bacteria as dependent on H-NS activity. Here, we describe an H-NS-like protein of the opportunistic pathogen Burkholderia multivorans, a species commonly infecting the respiratory tract of cystic fibrosis patients. Our results indicate that this protein is involved in regulating virulence traits such as exopolysaccharide biosynthesis, adhesion to biotic surfaces, cellular aggregation, and motility. Furthermore, this H-NS-like protein, is one out of eight orthologs present in B. multivorans ATCC 17616 genome, posing relevant questions to be investigated on how these proteins coordinate the expression of virulence traits.

scholarly journals Origin and dynamics of admixture in Brazilians and its effect on the pattern of deleterious mutations

2015 ◽  
Vol 112 (28) ◽  
pp. 8696-8701 ◽  
Author(s):  
Fernanda S. G. Kehdy ◽  
Mateus H. Gouveia ◽  
Moara Machado ◽  
Wagner C. S. Magalhães ◽  
Andrea R. Horimoto ◽  
...  
Keyword(s):  
Native American ◽  
Latin American ◽  
Middle Eastern ◽  
Classical Model ◽  
African Ancestry ◽  
Genomic Analysis ◽  
Genomic Diversity ◽  
European Ancestry ◽  
Eastern Region ◽  
Whole Genome Analysis

While South Americans are underrepresented in human genomic diversity studies, Brazil has been a classical model for population genetics studies on admixture. We present the results of the EPIGEN Brazil Initiative, the most comprehensive up-to-date genomic analysis of any Latin-American population. A population-based genome-wide analysis of 6,487 individuals was performed in the context of worldwide genomic diversity to elucidate how ancestry, kinship, and inbreeding interact in three populations with different histories from the Northeast (African ancestry: 50%), Southeast, and South (both with European ancestry >70%) of Brazil. We showed that ancestry-positive assortative mating permeated Brazilian history. We traced European ancestry in the Southeast/South to a wider European/Middle Eastern region with respect to the Northeast, where ancestry seems restricted to Iberia. By developing an approximate Bayesian computation framework, we infer more recent European immigration to the Southeast/South than to the Northeast. Also, the observed low Native-American ancestry (6–8%) was mostly introduced in different regions of Brazil soon after the European Conquest. We broadened our understanding of the African diaspora, the major destination of which was Brazil, by revealing that Brazilians display two within-Africa ancestry components: one associated with non-Bantu/western Africans (more evident in the Northeast and African Americans) and one associated with Bantu/eastern Africans (more present in the Southeast/South). Furthermore, the whole-genome analysis of 30 individuals (42-fold deep coverage) shows that continental admixture rather than local post-Columbian history is the main and complex determinant of the individual amount of deleterious genotypes.

scholarly journals Loss and Gain in the Evolution of theSalmonella entericaSerovar Gallinarum Biovar Pullorum Genome

mSphere ◽  
2019 ◽  
Vol 4 (2) ◽  
Author(s):  
Yachen Hu ◽  
Zhenyu Wang ◽  
Bin Qiang ◽  
Yaohui Xu ◽  
Xiang Chen ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genomic Analysis ◽  
Developed Countries ◽  
Epidemiological Surveillance ◽  
Recent Common Ancestor ◽  
Whole Genome ◽  
Nucleotide Polymorphisms ◽  
Content Type ◽  
Most Recent Common Ancestor

ABSTRACTSalmonella entericasubspeciesentericaserovar Gallinarum biovar Pullorum (S. Pullorum) is the etiological agent of pullorum disease, causing white diarrhea with high mortality in chickens. There are many unsolved issues surrounding the epidemiology ofS. Pullorum, including its origin and transmission history as well as the discordance between its phenotypic heterogeneity and genetic monomorphism. In this paper, we report the results of whole-genome sequencing of a panel of 97S. Pullorum strains isolated between 1962 and 2014 from four countries across three continents. We utilized 6,795 core genome single nucleotide polymorphisms (SNPs) to reconstruct a phylogenetic tree within a spatiotemporal Bayesian framework, estimating that the most recent common ancestor ofS. Pullorum emerged in ∼914 CE (95% confidence interval [95%CI], 565 to 1273 CE). The extantS. Pullorum strains can be divided into four distinct lineages, each of which is significantly associated with geographical distribution. The intercontinental transmissions of lineages III and IV can be traced to the mid-19th century and are probably related to the “Hen Fever” prevalent at that time. Further genomic analysis indicated that the loss or pseudogenization of functional genes involved in metabolism and virulence inS. Pullorum has been ongoing since before and after divergence from the ancestor. In contrast, multiple prophages and plasmids have been acquired byS. Pullorum, and these have endowed it with new characteristics, especially the multidrug resistance conferred by two large plasmids in lineage I. The results of this study provide insight into the evolution ofS. Pullorum and prove the efficiency of whole-genome sequencing in epidemiological surveillance of pullorum disease.IMPORTANCEPullorum disease, an acute poultry septicemia caused bySalmonellaGallinarum biovar Pullorum, is fatal for young chickens and is a heavy burden on poultry industry. The pathogen is rare in most developed countries but still extremely difficult to eliminate in China. Efficient epidemiological surveillance necessitates clarifying the origin of the isolates from different regions and their phylogenic relationships. Genomic epidemiological analysis of 97S. Pullorum strains was carried out to reconstruct the phylogeny and transmission history ofS. Pullorum. Further analysis demonstrated that functional gene loss and acquisition occurred simultaneously throughout the evolution ofS. Pullorum, both of which reflected adaptation to the changing environment. The result of our study will be helpful in surveillance and prevention of pullorum disease.

scholarly journals Discrimination of Burkholderia multivorans and Burkholderia vietnamiensis fromBurkholderia cepacia Genomovars I, III, and IV by PCR

1999 ◽  
Vol 37 (5) ◽  
pp. 1335-1339 ◽  
Author(s):  
Adolf Bauernfeind ◽  
Ines Schneider ◽  
Renate Jungwirth ◽  
Carsten Roller
Keyword(s):  
Cystic Fibrosis ◽  
Burkholderia Cepacia ◽  
General Sense ◽  
Burkholderia Multivorans ◽  
Burkholderia Vietnamiensis ◽  
Definition Of ◽  
Species Specific ◽  
Optimized Conditions ◽  
Ribosomal Dnas ◽  
Sense Primer

We present a PCR procedure for identification of Burkholderia cepacia, Burkholderia multivorans, andBurkholderia vietnamiensis. 16S and 23S ribosomal DNAs (rDNAs) of B. multivorans and B. vietnamiensiswere sequenced and aligned with published sequences for definition of species-specific 18-mer oligonucleotide primers. Specific antisense 16S rDNA primers (for B. cepacia, 5′-AGC ACT CCC RCC TCT CAG-3′; for B. multivorans, 5′-AGC ACT CCC GAA TCT CTT-3′) and 23S rDNA primers (for B. vietnamiensis, 5′-TCC TAC CAT GCG TGC AA-3′) were paired with a general sense primer of 16S rDNAs (5′-AGR GTT YGA TYM TGG CTC AG-3′) or with a sense primer of 23S rDNA (5′-CCT TTG GGT CAT CCT GGA-3′). PCR with these primers under optimized conditions is appropriate to specifically and rapidly identify B. multivorans, B. vietnamiensis, and B. cepacia (genomovars I, III, and IV are not discriminated). In comparison with the polyphasic taxonomic analyses presently necessary for species and genomovar identification within the B. cepacia complex, our procedure is more rapid and easier to perform and may contribute to clarifying the clinical significance of individual members of the complex in cystic fibrosis.

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