scholarly journals A Small Regulatory RNA Controls Cell Wall Biosynthesis and Antibiotic Resistance

mBio ◽  
2018 ◽  
Vol 9 (6) ◽  
Author(s):  
Jessica Borgmann ◽  
Sina Schäkermann ◽  
Julia Elisabeth Bandow ◽  
Franz Narberhaus
Keyword(s):  
Antibiotic Resistance ◽  
Agrobacterium Tumefaciens ◽  
Antibiotic Resistance Gene ◽  
Tumor Formation ◽  
Regulatory Rna ◽  
Beta Lactamase ◽  
Microbial Composition ◽  
Ampicillin Resistance ◽  
Content Type ◽  
Small Regulatory Rna

ABSTRACT Small regulatory RNAs play an important role in the adaptation to changing conditions. Here, we describe a differentially expressed small regulatory RNA (sRNA) that affects various cellular processes in the plant pathogen Agrobacterium tumefaciens. Using a combination of bioinformatic predictions and comparative proteomics, we identified nine targets, most of which are positively regulated by the sRNA. According to these targets, we named the sRNA PmaR for peptidoglycan biosynthesis, motility, and ampicillin resistance regulator. Agrobacterium spp. are long known to be naturally resistant to high ampicillin concentrations, and we can now explain this phenotype by the positive PmaR-mediated regulation of the beta-lactamase gene ampC. Structure probing revealed a spoon-like structure of the sRNA, with a single-stranded loop that is engaged in target interaction in vivo and in vitro. Several riboregulators have been implicated in antibiotic resistance mechanisms, such as uptake and efflux transporters, but PmaR represents the first example of an sRNA that directly controls the expression of an antibiotic resistance gene. IMPORTANCE The alphaproteobacterium Agrobacterium tumefaciens is able to infect various eudicots causing crown gall tumor formation. Based on its unique ability of interkingdom gene transfer, Agrobacterium serves as a crucial biotechnological tool for genetic manipulation of plant cells. The presence of hundreds of putative sRNAs in this organism suggests a considerable impact of riboregulation on A. tumefaciens physiology. Here, we characterized the biological function of the sRNA PmaR that controls various processes crucial for growth, motility, and virulence. Among the genes directly targeted by PmaR is ampC coding for a beta-lactamase that confers ampicillin resistance, suggesting that the sRNA is crucial for fitness in the competitive microbial composition of the rhizosphere.

scholarly journals The FasX Small Regulatory RNA Negatively Regulates the Expression of Two Fibronectin-Binding Proteins in Group A Streptococcus

2015 ◽  
Vol 197 (23) ◽  
pp. 3720-3730 ◽  
Author(s):  
Jessica L. Danger ◽  
Nishanth Makthal ◽  
Muthiah Kumaraswami ◽  
Paul Sumby
Keyword(s):  
Cell Surface ◽  
Virulence Factors ◽  
Binding Proteins ◽  
Mrna Translation ◽  
Regulatory Rna ◽  
Collagen Binding ◽  
Content Type ◽  
Small Regulatory Rna ◽  
Group A ◽  
Fibronectin Binding

ABSTRACTThe group AStreptococcus(GAS;Streptococcus pyogenes) causes more than 700 million human infections each year. The success of this pathogen can be traced in part to the extensive arsenal of virulence factors that are available for expression in temporally and spatially specific manners. To modify the expression of these virulence factors, GAS use both protein- and RNA-based regulators, with the best-characterized RNA-based regulator being the small regulatory RNA (sRNA) FasX. FasX is a 205-nucleotide sRNA that contributes to GAS virulence by enhancing the expression of the thrombolytic secreted virulence factor streptokinase and by repressing the expression of the collagen-binding cell surface pili. Here, we have expanded the FasX regulon, showing that this sRNA also negatively regulates the expression of the adhesion- and internalization-promoting, fibronectin-binding proteins PrtF1 and PrtF2. FasX posttranscriptionally regulates the expression of PrtF1/2 through a mechanism that involves base pairing to theprtF1andprtF2mRNAs within their 5′ untranslated regions, overlapping the mRNA ribosome-binding sites. Thus, duplex formation between FasX and theprtF1andprtF2mRNAs blocks ribosome access, leading to an inhibition of mRNA translation. Given that FasX positively regulates the expression of the spreading factor streptokinase and negatively regulates the expression of the collagen-binding pili and of the fibronectin-binding PrtF1/2, our data are consistent with FasX functioning as a molecular switch that governs the transition of GAS between the colonization and dissemination stages of infection.IMPORTANCEMore than half a million deaths each year are a consequence of infections caused by GAS. Insights into how this pathogen regulates the production of proteins during infection may facilitate the development of novel therapeutic or preventative regimens aimed at inhibiting this activity. Here, we have expanded insight into the regulatory activity of the GAS small RNA FasX. In addition to identifying that FasX reduces the abundance of the cell surface-located fibronectin-binding proteins PrtF1/2, fibronectin is present in high abundance in human tissues, and we have determined the mechanism behind this regulation. Importantly, as FasX is the only mechanistically characterized regulatory RNA in GAS, it serves as a model RNA in this and related pathogens.

scholarly journals Development of a DNA Microarray for Enterococcal Species, Virulence, and Antibiotic Resistance Gene Determinations among Isolates from Poultry

2011 ◽  
Vol 77 (8) ◽  
pp. 2625-2633 ◽  
Author(s):  
J. Champagne ◽  
M. S. Diarra ◽  
H. Rempel ◽  
E. Topp ◽  
C. W. Greer ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Dna Microarray ◽  
Virulence Genes ◽  
Antibiotic Resistance Gene ◽  
Antibiotic Resistance Genes ◽  
Content Type ◽  
Virulence Potential ◽  
Enterococcal Species ◽  
Species Specific ◽  
Simultaneous Identification

ABSTRACTA DNA microarray (Enteroarray) was designed with probes targeting four species-specific taxonomic identifiers to discriminate among 18 different enterococcal species, while other probes were designed to identify 18 virulence factors and 174 antibiotic resistance genes. In total, 262 genes were utilized for rapid species identification of enterococcal isolates, while characterizing their virulence potential through the simultaneous identification of endogenous antibiotic resistance and virulence genes. Enterococcal isolates from broiler chicken farms were initially identified by using the API 20 Strep system, and the results were compared to those obtained with the taxonomic genesatpA,recA,pheS, andddlrepresented on our microarray. Among the 171 isolates studied, five different enterococcal species were identified by using the API 20 Strep system:Enterococcus faecium,E. faecalis,E. durans,E. gallinarum, andE. avium. The Enteroarray detected the same species as API 20 Strep, as well as two more:E. casseliflavusandE. hirae. Species comparisons resulted in 15% (27 isolates) disagreement between the two methods among the five API 20 Strep identifiable species and 24% (42 isolates) disagreement when considering the seven Enteroarray identified species. The species specificity of key antibiotic and virulence genes identified by the Enteroarray were consistent with the literature adding further robustness to the redundant taxonomic probe data. Sequencing of thecpn60gene further confirmed the complete accuracy of the microarray results. The new Enteroarray should prove to be a useful tool to accurately genotype strains of enterococci and assess their virulence potential.

scholarly journals Plasmid with Colistin Resistance Genemcr-1in Extended-Spectrum-β-Lactamase-Producing Escherichia coli Strains Isolated from Pig Slurry in Estonia

2016 ◽  
Vol 60 (11) ◽  
pp. 6933-6936 ◽  
Author(s):  
Age Brauer ◽  
Kaidi Telling ◽  
Mailis Laht ◽  
Piret Kalmus ◽  
Irja Lutsar ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Resistance Gene ◽  
Antibiotic Resistance Gene ◽  
Dna Transfer ◽  
The Other ◽  
Pig Slurry ◽  
Colistin Resistance ◽  
Content Type ◽  
Slurry Sample

ABSTRACTA plasmid carrying the colistin resistance genemcr-1was isolated from a pig slurry sample in Estonia. The gene was present on a 33,311-bp plasmid of the IncX4 group.mcr-1is the only antibiotic resistance gene on the plasmid, with the other genes mainly coding for proteins involved in conjugative DNA transfer (taxA,taxB,taxC,trbM, and thepilXoperon). The plasmid pESTMCR was present in three phylogenetically very differentEscherichia colistrains, suggesting that it has high potential for horizontal transfer.

scholarly journals Housefly Larva Vermicomposting Efficiently Attenuates Antibiotic Resistance Genes in Swine Manure, with Concomitant Bacterial Population Changes

2015 ◽  
Vol 81 (22) ◽  
pp. 7668-7679 ◽  
Author(s):  
Hang Wang ◽  
Hongyi Li ◽  
Jack A. Gilbert ◽  
Haibo Li ◽  
Longhua Wu ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Waste Management ◽  
Resistance Genes ◽  
Management Practice ◽  
Antibiotic Resistance Gene ◽  
Tetracycline Resistance ◽  
Feed Additives ◽  
Content Type ◽  
Genes Encoding ◽  
Waste Management Practice

ABSTRACTManure from swine treated with antimicrobials as feed additives is a major source for the expansion of the antibiotic resistance gene (ARG) reservoir in the environment. Vermicomposting via housefly larvae (Musca domestica) can be efficiently used to treat manure and regenerate biofertilizer, but few studies have investigated its effect on ARG attenuation. Here, we tracked the abundances of 9 ARGs and the composition and structure of the bacterial communities in manure samples across 6 days of full-scale manure vermicomposting. On day 6, the abundances of genes encoding tetracycline resistance [tet(M),tet(O),tet(Q), andtet(W)] were reduced (P< 0.05), while those of genes encoding sulfonamide resistance (sul1andsul2) were increased (P< 0.05) when normalized to 16S rRNA. The abundances of tetracycline resistance genes were correlated (P< 0.05) with the changing concentrations of tetracyclines in the manure. The overall diversity and richness of the bacteria significantly decreased during vermicomposting, accompanied by a 100 times increase in the relative abundance ofFlavobacteriaceaespp. Variations in the abundances of ARGs were correlated with the changing microbial community structure and the relative abundances of the familyRuminococcaceae, classBacilli, or phylumProteobacteria. Vermicomposting, as a waste management practice, can reduce the overall abundance of ARGs. More research is warranted to assess the use of this waste management practice as a measure to attenuate the dissemination of antimicrobial residues and ARGs from livestock production before vermicompost can be safely used as biofertilizer in agroecosystems.

scholarly journals A Small Regulatory RNA Generated from the malK 5′ Untranslated Region Targets Gluconeogenesis in Vibrio Species

mSphere ◽  
2021 ◽  
Author(s):  
Xing Luo ◽  
Marick Esberard ◽  
Philippe Bouloc ◽  
Annick Jacq
Keyword(s):  
Untranslated Region ◽  
Vibrio Species ◽  
Economic Consequences ◽  
Regulatory Rna ◽  
Content Type ◽  
Small Regulatory Rna ◽  
Pacific Oysters ◽  
Summer Mortality

Juvenile pacific oysters have been subject in recent years to summer mortality episodes with deep economic consequences. The pathogen Vibrio tasmaniensis has been associated with such mortality events.

scholarly journals Metagenomic Analysis Reveals the Shared and Distinct Features of Bacterial Communities and Resistomes in Corn Silage from Different Climate Zones in China

2021 ◽  
Author(s):  
Dongmei Xu ◽  
Hongyan Han ◽  
Chao Wang ◽  
Yixin Zhang ◽  
Fuhou Li ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
High Risk ◽  
Relative Abundance ◽  
Bacterial Communities ◽  
Climatic Factors ◽  
Antibiotic Resistance Gene ◽  
Corn Silage ◽  
Pollution Level ◽  
Microbial Composition ◽  
Climate Zones

Abstract Background: The emergence and spread of antibiotic resistance are a significant threat to global health. Silage is a major forage feed for ruminants, and its safety is an important guarantee that high-quality ruminant products will remain available to humans. However, little attention has been given to the silage resistome. To define the antibiotic resistome and its potential risk to silage from different climate zones and in response to the ensiling process, this study used metagenomics to investigate bacterial communities and the type and amount of antibiotic resistance gene (ARG) in corn silage harvested from six climate zones (Cfa, BWk, Dwc, Dwa, BSk, and Aw based on Köppen-Geiger climate classification) in China. Results: The composition and succession of silage bacterial communities varied significantly between different climate zones. Lactobacillus was the predominant bacteria during corn ensiling. A total of 134 ARGs were observed in corn silage, with the dominant classes being beta-lactamase and multidrug resistance and the primary mechanisms being efflux pump, inactivation, and target protection. Differences in the resistome were mainly attributed to disparities in microbial composition, which was indirectly affected by climatic factors and fermentation pH. ARG abundance was lower in 90-day silages than 5-day silages except in Hainan silage. The diversity and relative abundance (0.65-0.4% based on total gene number) of ARGs was lower in silage microbiota from Tibet than other climate zones. The dominant ARGs were tetM, oqxB, lmrD, lnuA, ermB, and tetS, and Enterobacter, Klebsiella, Staphylococcus, Lactobacillus and Lactococcus were the primary ARG hosts. Eleven high-risk ARGs were chosen to evaluate the pollution level of silages harvested from different climate zones. The highest relative abundance of high-risk ARGs belonging to Lactobacillus occurred in corn silages from Cfa, Dwa and BWk climate zones. Conclusions: The ensiling process decreased ARG abundance. While resistome contamination of silage from Tibet was relatively low, ARGs with high risk were abundant in silages from Cfa, Dwa and BWk climate zones.

A pentaplex real-time PCR assay for rapid identification of major beta-lactamase genes KPC, NDM, CTX, CMY, and OXA-48 directly from bacteria in blood

2021 ◽  
Vol 70 (12) ◽  
Author(s):  
Taalin R. Hoj ◽  
Bradley McNeely ◽  
Kylie Webber ◽  
Evelyn Welling ◽  
William G. Pitt ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Klebsiella Pneumoniae ◽  
Real Time ◽  
Real Time Pcr ◽  
Type Species ◽  
New Delhi ◽  
Beta Lactamase ◽  
Content Type ◽  
Link Type ◽  
Carbapenem Resistant

Introduction. Antibiotic resistance, particularly in cases of sepsis, has emerged as a growing global public health concern and economic burden. Current methods of blood culture and antimicrobial susceptibility testing of agents involved in sepsis can take as long as 3–5 days. It is vital to rapidly identify which antimicrobials can be used to effectively treat sepsis cases on an individual basis. Here, we present a pentaplex, real-time PCR-based assay that can quickly identify the most common beta-lactamase genes ( Klebsiella pneumoniae carbapenemase (KPC); New Delhi metallo-beta-lactamase (NDM); cefotaximase-Munich (CTX-M); cephamycin AmpC beta-lactamases (CMY); and Oxacillinase-48 (OXA-48)) from pathogens derived directly from the blood of patients presenting with bacterial septicemia. Aim. To develop an assay which can rapidly identify the most common beta-lactamase genes in Carbapenem-resistant Enterobacteriaceae bacteria (CREs) from the United States. Hypothesis/Gap Statement. Septicemia caused by carbapenem-resistant bacteria has a death rate of 40–60 %. Rapid diagnosis of antibiotic susceptibility directly from bacteria in blood by identification of beta-lactamase genes will greatly improve survival rates. In this work, we develop an assay capable of concurrently identifying the five most common beta-lactamase and carbapenemase genes. Methodology. Primers and probes were created which can identify all subtypes of Klebsiella pneumoniae carbapenemase (KPC); New Delhi metallo-beta-lactamase (NDM); cefotaximase-Munich (CTX); cephamycin AmpC beta-lactamase (CMY); and oxacillinase-48 (OXA-48). The assay was validated using 13 isolates containing various PCR targets from the Centre for Disease Control Antimicrobial Resistance Isolate Bank Enterobacterales Carbapenemase Diversity Panel. Blood obtained from volunteers was spiked with CREs and bacteria were separated, lysed, and subjected to analysis via the pentaplex assay. Results. This pentaplex assay successfully identified beta-lactamase genes derived from bacteria separated from blood at concentrations of 4–8 c.f.u. ml−1. Conclusion. This assay will improve patient outcomes by supplying physicians with critical drug resistance information within 2 h of septicemia onset, allowing them to prescribe effective antimicrobials corresponding to the resistance gene(s) present in the pathogen. In addition, information supplied by this assay will lessen the inappropriate use of broad-spectrum antimicrobials and prevent the evolution of further antibiotic resistance.

scholarly journals Expression of a Peptidoglycan Hydrolase from Lytic Bacteriophages Atu_ph02 and Atu_ph03 Triggers Lysis of Agrobacterium tumefaciens

2017 ◽  
Vol 83 (23) ◽  
Author(s):  
Hedieh Attai ◽  
Jeanette Rimbey ◽  
George P. Smith ◽  
Pamela J. B. Brown
Keyword(s):  
Cell Division ◽  
Agrobacterium Tumefaciens ◽  
Plant Pathogens ◽  
Transmembrane Domain ◽  
Cell Lysis ◽  
Tumor Formation ◽  
Plant Diseases ◽  
Bacterial Cells ◽  
Content Type ◽  
Endogenous Expression

ABSTRACT To provide food security, innovative approaches to preventing plant disease are currently being explored. Here, we demonstrate that lytic bacteriophages and phage lysis proteins are effective at triggering lysis of the phytopathogen Agrobacterium tumefaciens. Phages Atu_ph02 and Atu_ph03 were isolated from wastewater and induced lysis of C58-derived strains of A. tumefaciens. The coinoculation of A. tumefaciens with phages on potato discs limited tumor formation. The genomes of Atu_ph02 and Atu_ph03 are nearly identical and are ∼42% identical to those of T7 supercluster phages. In silico attempts to find a canonical lysis cassette were unsuccessful; however, we found a putative phage peptidoglycan hydrolase (PPH), which contains a C-terminal transmembrane domain. Remarkably, the endogenous expression of pph in the absence of additional phage genes causes a block in cell division and subsequent lysis of A. tumefaciens cells. When the presumed active site of the N-acetylmuramidase domain carries an inactivating mutation, PPH expression causes extensive cell branching due to a block in cell division but does not trigger rapid cell lysis. In contrast, the mutation of positively charged residues at the extreme C terminus of PPH causes more rapid cell lysis. Together, these results suggest that PPH causes a block in cell division and triggers cell lysis through two distinct activities. Finally, the potent killing activity of this single lysis protein can be modulated, suggesting that it could be engineered to be an effective enzybiotic. IMPORTANCE The characterization of bacteriophages such as Atu_ph02 and Atu_ph03, which infect plant pathogens such as Agrobacterium tumefaciens, may be the basis of new biocontrol strategies. First, cocktails of diverse bacteriophages could be used as a preventative measure to limit plant diseases caused by bacteria; a bacterial pathogen is unlikely to simultaneously develop resistances to multiple bacteriophage species. The specificity of bacteriophage treatment for the host is an asset in complex communities, such as in orchards where it would be detrimental to harm the symbiotic bacteria in the environment. Second, bacteriophages are potential sources of enzymes that efficiently lyse bacterial cells. These phage proteins may have a broad specificity, but since proteins do not replicate as phages do, their effect is highly localized, providing an alternative to traditional antibiotic treatments. Thus, studies of lytic bacteriophages that infect A. tumefaciens may provide insights for designing preventative strategies against bacterial pathogens.

scholarly journals ARG-ANNOT, a New Bioinformatic Tool To Discover Antibiotic Resistance Genes in Bacterial Genomes

2013 ◽  
Vol 58 (1) ◽  
pp. 212-220 ◽  
Author(s):  
Sushim Kumar Gupta ◽  
Babu Roshan Padmanabhan ◽  
Seydina M. Diene ◽  
Rafael Lopez-Rojas ◽  
Marie Kempf ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Gene Sequence ◽  
Antibiotic Resistance Gene ◽  
Antibiotic Resistance Genes ◽  
Genetic Determinants ◽  
Bacterial Genomes ◽  
Content Type ◽  
A Genome ◽  
Bioinformatic Tool

ABSTRACTARG-ANNOT (Antibiotic Resistance Gene-ANNOTation) is a new bioinformatic tool that was created to detect existing and putative new antibiotic resistance (AR) genes in bacterial genomes. ARG-ANNOT uses a local BLAST program in Bio-Edit software that allows the user to analyze sequences without a Web interface. All AR genetic determinants were collected from published works and online resources; nucleotide and protein sequences were retrieved from the NCBI GenBank database. After building a database that includes 1,689 antibiotic resistance genes, the software was tested in a blind manner using 100 random sequences selected from the database to verify that the sensitivity and specificity were at 100% even when partial sequences were queried. Notably, BLAST analysis results obtained using thermtFgene sequence (a new aminoglycoside-modifying enzyme gene sequence that is not included in the database) as a query revealed that the tool was able to link this sequence to short sequences (17 to 40 bp) found in other genes of thermtfamily with significant E values. Finally, the analysis of 178Acinetobacter baumanniiand 20Staphylococcus aureusgenomes allowed the detection of a significantly higher number of AR genes than the Resfinder gene analyzer and 11 point mutations in target genes known to be associated with AR. The average time for the analysis of a genome was 3.35 ± 0.13 min. We have created a concise database for BLAST using a Bio-Edit interface that can detect AR genetic determinants in bacterial genomes and can rapidly and easily discover putative new AR genetic determinants.

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