scholarly journals A Population-level Strain Genotyping Method to Study Pathogen Strain Dynamics in Human Infections

2021 ◽  
Author(s):  
Sarah J Morgan ◽  
Samantha L Durfey ◽  
Sumedha Ravishankar ◽  
Peter Jorth ◽  
Wendy Ni ◽  
...  
Keyword(s):  
Relative Abundance ◽  
Bacterial Infections ◽  
Ex Vivo ◽  
Bacterial Species ◽  
Pcr Amplification ◽  
Population Level ◽  
Bacterial Cells ◽  
Chronic Infections ◽  
Single Strain ◽  
Sequencing Errors

A hallmark of chronic bacterial infections is the long-term persistence of one or more pathogen species at the compromised site. Repeated detection of the same bacterial species can suggest that a single strain or lineage is continually present. However, infection with multiple strains of a given species, strain acquisition and loss, and changes in strain relative abundance can occur. Detecting strain-level changes and their effects on disease is challenging as most methods require labor intensive isolate-by-isolate analyses, thus, only a few cells from large infecting populations can be examined. Here we present a population-level method for enumerating and measuring the relative abundance of strains called PopMLST. The method exploits PCR amplification of strain-identifying polymorphic loci, next-generation sequencing to measure allelic variants, and informatic methods to determine whether variants arise from sequencing errors or low abundance strains. These features enable PopMLST to simultaneously interrogate hundreds of bacterial cells that are either cultured en masse from patient samples, or are present in DNA directly extracted from clinical specimens without ex vivo culture. This method could be used to detect epidemic or super-infecting strains, facilitate understanding of strain dynamics during chronic infections, and enable studies that link strain changes to clinical outcomes.

scholarly journals Slow Genetic Divergence of Helicobacter pylori Strains during Long-Term Colonization

2005 ◽  
Vol 73 (8) ◽  
pp. 4818-4822 ◽  
Author(s):  
Annelie Lundin ◽  
Britta Björkholm ◽  
Ilya Kupershmidt ◽  
Magnus Unemo ◽  
Peter Nilsson ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Genetic Divergence ◽  
Bacterial Species ◽  
Pcr Amplification ◽  
Genetic Change ◽  
Genetic Changes ◽  
Single Strain ◽  
Divergent Gene ◽  
Asymptomatic Adult ◽  
H Pylori

ABSTRACT The genetic variability of Helicobacter pylori is known to be high compared to that of many other bacterial species. H. pylori is adapted to the human stomach, where it persists for decades, and adaptation to each host results in every individual harboring a distinctive bacterial population. Although clonal variants may exist within such a population, all isolates are generally genetically related and thus derived from a common ancestor. We sought to determine the rate of genetic change of H. pylori over 9 years in two asymptomatic adult patients. Arbitrary primed PCR confirmed the relatedness of individual subclones within a patient. Furthermore, sequencing of 10 loci (∼6,000 bp) in three subclones per time and patient revealed only two base pair changes among the subclones from patient I. All sequences were identical among the patient II subclones. However, PCR amplification of the highly divergent gene amiA revealed great variation in the size of the gene between the subclones within each patient. Thus, both patients harbored a single strain with clonal variants at both times. We also studied genetic changes in culture- and mouse-passaged strains, and under both conditions no genetic divergence was found. These results suggest that previous estimates of the rate of genetic change in H. pylori within an individual might be overestimates.

scholarly journals Potentiation of Tobramycin by Silver Nanoparticles against Pseudomonas aeruginosa Biofilms

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Marc B. Habash ◽  
Mara C. Goodyear ◽  
Amber J. Park ◽  
Matthew D. Surette ◽  
Emily C. Vis ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Silver Nanoparticles ◽  
Bacterial Infections ◽  
Pathogenic Bacteria ◽  
Antimicrobial Agents ◽  
Bacterial Species ◽  
Acute Infection ◽  
Public Health Problem ◽  
Chronic Infections ◽  
Content Type

ABSTRACT Increasing antibiotic resistance among pathogenic bacterial species is a serious public health problem and has prompted research examining the antibacterial effects of alternative compounds and novel treatment strategies. Compounding this problem is the ability of many pathogenic bacteria to form biofilms during chronic infections. Importantly, these communities are often recalcitrant to antibiotic treatments that show effectiveness against acute infection. The antimicrobial properties of silver have been known for decades, but recently silver and silver-containing compounds have seen renewed interest as antimicrobial agents for treating bacterial infections. The goal of this study was to assess the ability of citrate-capped silver nanoparticles (AgNPs) of various sizes, alone and in combination with the aminoglycoside antibiotic tobramycin, to inhibit established Pseudomonas aeruginosa biofilms. Our results demonstrate that smaller 10-nm and 20-nm AgNPs were more effective at synergistically potentiating the activity of tobramycin. Visualization of biofilms treated with combinations of 10-nm AgNPs and tobramycin reveals that the synergistic bactericidal effect may be caused by disrupting cellular membranes. Minimum biofilm eradication concentration (MBEC) assays using clinical P. aeruginosa isolates shows that small AgNPs are more effective than larger AgNPs at inhibiting biofilms, but that the synergy effect is likely a strain-dependent phenomenon. These data suggest that small AgNPs synergistically potentiate the activity of tobramycin against P. aeruginosa in vitro and may reveal a potential role for AgNP/antibiotic combinations in treating patients with chronic infections in a strain-specific manner.

scholarly journals Beyond the pan-genome: current perspectives on the functional and practical outcomes of the distributed genome hypothesis

2020 ◽  
Vol 48 (6) ◽  
pp. 2437-2455
Author(s):  
Jocelyn A. Hammond ◽  
Emma A. Gordon ◽  
Kayla M. Socarras ◽  
Joshua Chang Mell ◽  
Garth D. Ehrlich
Keyword(s):  
Bacterial Infections ◽  
Bacterial Species ◽  
Genotypic Diversity ◽  
Life Cycles ◽  
Chronic Infections ◽  
Novel Technologies ◽  
Symbiotic Relationships ◽  
Pan Genome ◽  
Potential Applications ◽  
Imaging Science

The principle of monoclonality with regard to bacterial infections was considered immutable prior to 30 years ago. This view, espoused by Koch for acute infections, has proven inadequate regarding chronic infections as persistence requires multiple forms of heterogeneity among the bacterial population. This understanding of bacterial plurality emerged from a synthesis of what-were-then novel technologies in molecular biology and imaging science. These technologies demonstrated that bacteria have complex life cycles, polymicrobial ecologies, and evolve in situ via the horizontal exchange of genic characters. Thus, there is an ongoing generation of diversity during infection that results in far more highly complex microbial communities than previously envisioned. This perspective is based on the fundamental tenet that the bacteria within an infecting population display genotypic diversity, including gene possession differences, which result from horizontal gene transfer mechanisms including transformation, conjugation, and transduction. This understanding is embodied in the concepts of the supragenome/pan-genome and the distributed genome hypothesis (DGH). These paradigms have fostered multiple researches in diverse areas of bacterial ecology including host–bacterial interactions covering the gamut of symbiotic relationships including mutualism, commensalism, and parasitism. With regard to the human host, within each of these symbiotic relationships all bacterial species possess attributes that contribute to colonization and persistence; those species/strains that are pathogenic also encode traits for invasion and metastases. Herein we provide an update on our understanding of bacterial plurality and discuss potential applications in diagnostics, therapeutics, and vaccinology based on perspectives provided by the DGH with regard to the evolution of pathogenicity.

scholarly journals Phage–Bacteria Interactions in Potential Applications of Bacteriophage vB_EfaS-271 against Enterococcus faecalis

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 318
Author(s):  
Gracja Topka-Bielecka ◽  
Bożena Nejman-Faleńczyk ◽  
Sylwia Bloch ◽  
Aleksandra Dydecka ◽  
Agnieszka Necel ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Mammalian Cells ◽  
Bacterial Infections ◽  
Phage Therapy ◽  
Bacterial Species ◽  
Resistant Bacteria ◽  
Bacterial Cells ◽  
Potential Applications ◽  
High Level

Phage therapy is one of main alternative option for antibiotic treatment of bacterial infections, particularly in the era of appearance of pathogenic strains revealing resistance to most or even all known antibiotics. Enterococcus faecalis is one of such pathogens causing serious human infections. In the light of high level of biodiversity of bacteriophages and specificity of phages to bacterial species or even strains, development of effective phage therapy depend, between others, on identification and characterization of a large collection of these viruses, including understanding of their interactions with host bacterial cells. Recently, isolation of molecular characterization of bacteriophage vB_EfaS-271, infecting E. faecalis strains have been reported. In this report, phage–host interactions are reported, including ability of vB_EfaS-271 to infect bacteria forming biofilms, efficiency of eliminating bacterial cells from cultures depending on multiplicity of infection (m.o.i.), toxicity of purified phage particles to mammalian cells, and efficiency of appearance of phage-resistant bacteria. The presented results indicate that vB_EfaS-271 can significantly decrease number of viable E. faecalis cells in biofilms and in liquid cultures and reveals no considerable toxicity to mammalian cells. Efficiency of formation of phage-resistant bacteria was dependent on m.o.i. and was higher when the virion-cell ratio was as high as 10 than at low (between 0.01 and 0.0001) m.o.i. values. We conclude that vB_EfaS-271 may be considered as a candidate for its further use in phage therapy.

scholarly journals Robust, linear correlations between growth rates and β-lactam–mediated lysis rates

2018 ◽  
Vol 115 (16) ◽  
pp. 4069-4074 ◽  
Author(s):  
Anna J. Lee ◽  
Shangying Wang ◽  
Hannah R. Meredith ◽  
Bihan Zhuang ◽  
Zhuojun Dai ◽  
...  
Keyword(s):  
Growth Rate ◽  
Single Cells ◽  
Bacterial Species ◽  
Population Level ◽  
Instantaneous Growth Rate ◽  
Bacterial Cells ◽  
Bacterial Growth Rate ◽  
Lysis Rate ◽  
Dynamic Signature ◽  
Linear Correlations

It is widely acknowledged that faster-growing bacteria are killed faster by β-lactam antibiotics. This notion serves as the foundation for the concept of bacterial persistence: dormant bacterial cells that do not grow are phenotypically tolerant against β-lactam treatment. Such correlation has often been invoked in the mathematical modeling of bacterial responses to antibiotics. Due to the lack of thorough quantification, however, it is unclear whether and to what extent the bacterial growth rate can predict the lysis rate upon β-lactam treatment under diverse conditions. Enabled by experimental automation, here we measured >1,000 growth/killing curves for eight combinations of antibiotics and bacterial species and strains, including clinical isolates of bacterial pathogens. We found that the lysis rate of a bacterial population linearly depends on the instantaneous growth rate of the population, regardless of how the latter is modulated. We further demonstrate that this predictive power at the population level can be explained by accounting for bacterial responses to the antibiotic treatment by single cells. This linear dependence of the lysis rate on the growth rate represents a dynamic signature associated with each bacterium–antibiotic pair and serves as the quantitative foundation for designing combination antibiotic therapy and predicting the population-structure change in a population with mixed phenotypes.

scholarly journals CLIQ-BID: A method to quantify bacteria-induced damage to eukaryotic cells by automated live-imaging of bright nuclei

2017 ◽  
Author(s):  
Yann Wallez ◽  
Stéphanie Bouillot ◽  
Emmanuelle Soleilhac ◽  
Philippe Huber ◽  
Ina Attrée ◽  
...  
Keyword(s):  
High Throughput ◽  
Bacterial Infections ◽  
Pathogenic Bacteria ◽  
Ex Vivo ◽  
Morphological Changes ◽  
Cell Damage ◽  
Bacterial Species ◽  
Eukaryotic Cell ◽  
Live Imaging ◽  
Quantitative Detection

ABSTRACTPathogenic bacteria induce eukaryotic cell damage which range from discrete modifications of signalling pathways, to morphological alterations and even to cell death. Accurate quantitative detection of these events is necessary for studying host-pathogen interactions and for developing strategies to protect host organisms from bacterial infections. Investigation of morphological changes is cumbersome and not adapted to high-throughput and kinetics measurements. Here, we describe a simple and cost-effective method based on automated analysis of live cells with stained nuclei, which allows real-time quantification of bacteria-induced eukaryotic cell damage at single-cell resolution. We demonstrate that this automated high-throughput microscopy approach permits screening of libraries composed of interference-RNA, bacterial strains, antibodies and chemical compounds in ex vivo infection settings. The use of fluorescently-labelled bacteria enables the concomitant detection of changes in bacterial growth. Using this method named CLIQ-BID (Cell Live Imaging Quantification of Bacteria Induced Damage), we were able to distinguish the virulence profiles of different pathogenic bacterial species and clinical strains.

scholarly journals The relationship between biofilm formation, genes of virulence and iron metabolism in Escherichia coli

2018 ◽  
Author(s):  
Lívia Handrová ◽  
Anna Čuvalová ◽  
Vladimír Kmeť
Keyword(s):  
Escherichia Coli ◽  
Iron Metabolism ◽  
Biofilm Formation ◽  
Bacterial Infections ◽  
Opportunistic Infections ◽  
Bacterial Species ◽  
Pcr Amplification ◽  
Bacterial Strains ◽  
E Coli ◽  
Wide Range

Escherichia coli is known as one of the bacterial species with the widest adaptability to variety of niches either within organisms or outside in environment. Most strains of E. coli are of low virulence and associated with opportunistic infections, whereas others are highly virulent. The success of E. coli in colonising such a wide range of hosts and environments is basically due to a noticeable ductility in exploiting the available resources. It is becoming increasingly clear that biofilms have an enormous impact on medicine because since 65% of animal and human bacterial infections involve biofilms. In present study, we isolated strains of E. coli from animals. 19 interesting isolates were selected and tested by PCR amplification to virulence – iutA, cvaC, iss, tsh, papC, kps, iha and iron metabolism genes – sitA, feoB, irp2, fyuA, iroN, ireA. The ability of biofilm formation was assessed in a quantitative assay using a microtiter-plate test. Bacterial strains were grown on BHI. We divided isolates of E. coli into four classes: very weak (63.0%), weak (10.5%), moderate (10.5%) and strong (16.0%) biofilm producers. Representation genes of virulence were highly in isolates from very weak biofilm producers – from 7 genes were 6 highly; only papC (P fimbrial adhesin) was low. Genes of iron metabolism were different. Genes – sitA, fyuA, ireA in strong isolates producing biofilm and feoB, irp2, iroN in weak producers were most represented. The results show possible relation between presence virulence factor and low biofilm formation.

scholarly journals Enantiomeric glycosylated cationic block co-beta-peptides eradicate Staphylococcus aureus biofilms and antibiotic-tolerant persisters

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Kaixi Zhang ◽  
Yu Du ◽  
Zhangyong Si ◽  
Yang Liu ◽  
Michelle E. Turvey ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Bacterial Infections ◽  
Ex Vivo ◽  
High Yield ◽  
Bacterial Cells ◽  
One Pot ◽  
Beta Peptide ◽  
Repeated Dosing ◽  
Multiple Drugs

Abstract The treatment of bacterial infections is hindered by the presence of biofilms and metabolically inactive persisters. Here, we report the synthesis of an enantiomeric block co-beta-peptide, poly(amido-D-glucose)-block-poly(beta-L-lysine), with high yield and purity by one-shot one-pot anionic-ring opening (co)polymerization. The co-beta-peptide is bactericidal against methicillin-resistant Staphylococcus aureus (MRSA), including replicating, biofilm and persister bacterial cells, and also disperses biofilm biomass. It is active towards community-acquired and hospital-associated MRSA strains which are resistant to multiple drugs including vancomycin and daptomycin. Its antibacterial activity is superior to that of vancomycin in MRSA mouse and human ex vivo skin infection models, with no acute in vivo toxicity in repeated dosing in mice at above therapeutic levels. The copolymer displays bacteria-activated surfactant-like properties, resulting from contact with the bacterial envelope. Our results indicate that this class of non-toxic molecule, effective against different bacterial sub-populations, has promising potential for the treatment of S. aureus infections.

scholarly journals Ɛ34 Phage Tailspike Protein is Resistant to Trypsin and Inhibits and Salmonella Biofilm Formation

2021 ◽  
Author(s):  
Joseph A. Ayariga ◽  
Logan Gildea ◽  
Robert Villafane
Keyword(s):  
Biofilm Formation ◽  
Bacterial Infections ◽  
Ex Vivo ◽  
Digestive Enzyme ◽  
Food Poisoning ◽  
Developed Countries ◽  
Cell Culture Supernatant ◽  
Chronic Infections ◽  
The Us ◽  
Tailspike Protein

Salmonella can cause acute and chronic infections in humans. Salmonella species are known to cause food poisoning and other diseases in developing countries. Their role in the pathogenesis of these diseases has received increased international attention. Despite numerous advances in sanitation, they still can infect humans and cause outbreaks in developed countries. For example, Salmonella causes about 1.2 million illnesses in the US each year with over 450 deaths. Additionally, Salmonella outbreaks cause significant losses to chicken producers globally. The Salmonella species is also prone to acquiring resistance to various classes of antibiotics. Hence, the need for a paradigm shift from antibiotics to bacteriophages to manage, control and treat bacterial infections. The ɛ34 phage belongs to Podoviruses and categorized into the P22-like phages. The P22-like phages include ɛ34, ES18, P22, ST104, and ST64T. In this work, we investigated the antibacterial property of ɛ34 phage tailspike protein against Salmonella newington (S. newington). We demonstrate here that, the phage’s tailspike protein enzymatic property as a LPS hydrolase synergizes with Vero Cell culture supernatant in killing S. newington. Using decellularized cartilage scaffold as an ex vivo tissue model, the ɛ34 TSP protected the scaffold from S. newington biofilm formation. Computational analysis of the ɛ34 TSP interaction with membrane proteins of S. newington demonstrated a higher probability (0.7318) of binding to ompA of S. newington, and when docked to ompA extracellular component, it produced a high free energy of -11.3kcal/mol. We also demonstrate the resistance/sensitivity of the tailspike to the digestive enzyme trypsin. The data obtained in this work indicates that the trypsin resistant tailspike protein of Ɛ34 phage can be formulated as a novel antibacterial agent against S. newington.

Small Is Mighty—Chemical Communication Systems in Pseudomonas aeruginosa

2019 ◽  
Vol 73 (1) ◽  
pp. 559-578 ◽  
Author(s):  
Stephen Dela Ahator ◽  
LianHui Zhang
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Chemical Communication ◽  
Communication Systems ◽  
Bacterial Infections ◽  
Environmental Changes ◽  
Therapeutic Interventions ◽  
Research Progress ◽  
Host Immune System ◽  
Bacterial Cells ◽  
Chronic Infections

Pseudomonas aeruginosa is an opportunistic pathogen that causes a variety of acute and chronic infections. Usually a commensal on the host body, P. aeruginosa is capable of transforming into a virulent pathogen upon sensing favorable changes in the host immune system or stress cues. P. aeruginosa infections are hard to eradicate, because this pathogen has developed strong resistance to most conventional antibiotics; in addition, in chronic infections it commonly forms a biofilm matrix, which provides bacterial cells a protected environment to withstand various stresses including antibiotics. Given its importance as a human pathogen and its notorious antimicrobial tolerance, P. aeruginosa has been the subject of intensive investigations internationally. Research progress over the last two decades has unveiled a range of chemical communication systems in this pathogen. These diversified chemical communication systems endow P. aeruginosa a superb ability and remarkable flexibility to coordinate and modulate accordingly the transcriptional expression of various sets of genes associated with virulence and other physiologic activities in response to environmental changes. A fair understanding of the chemical signaling mechanisms with which P. aeruginosa governs virulence gene expression may hold the key to developing alternative therapeutic interventions that control and prevent bacterial infections.

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