scholarly journals Fast and Accurate Large-Scale Detection of β-Lactamase Genes Conferring Antibiotic Resistance

2015 ◽  
Vol 59 (10) ◽  
pp. 5967-5975 ◽  
Author(s):  
Jae Jin Lee ◽  
Jung Hun Lee ◽  
Dae Beom Kwon ◽  
Jeong Ho Jeon ◽  
Kwang Seung Park ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Pathogenic Bacteria ◽  
Large Scale ◽  
Diagnostic Methods ◽  
Control Measures ◽  
Clinical Isolates ◽  
Molecular Diagnostic ◽  
Resistant Bacteria ◽  
Specificity And Sensitivity ◽  
Almost All

ABSTRACTFast detection of β-lactamase (bla) genes allows improved surveillance studies and infection control measures, which can minimize the spread of antibiotic resistance. Although several molecular diagnostic methods have been developed to detect limitedblagene types, these methods have significant limitations, such as their failure to detect almost all clinically availableblagenes. We developed a fast and accurate molecular method to overcome these limitations using 62 primer pairs, which were designed through elaborate optimization processes. To verify the ability of this large-scalebladetection method (large-scaleblaFinder), assays were performed on previously reported bacterial control isolates/strains. To confirm the applicability of thelarge-scaleblaFinder, the assays were performed on unreported clinical isolates. With perfect specificity and sensitivity in 189 control isolates/strains and 403 clinical isolates, thelarge-scaleblaFinder detected almost all clinically availableblagenes. Notably, thelarge-scaleblaFinder detected 24 additional unreportedblagenes in the isolates/strains that were previously studied, suggesting that previous methods detecting only limited types ofblagenes can miss unexpectedblagenes existing in pathogenic bacteria, and our method has the ability to detect almost allblagenes existing in a clinical isolate. The ability oflarge-scaleblaFinder to detectblagenes on a large scale enables prompt application to the detection of almost allblagenes present in bacterial pathogens. The widespread use of thelarge-scaleblaFinder in the future will provide an important aid for monitoring the emergence and dissemination ofblagenes and minimizing the spread of resistant bacteria.

scholarly journals A large-scale comparison shows that genetic changes causing antibiotic resistance in experimentally evolved Pseudomonas aeruginosa predict those in naturally evolved bacteria

2019 ◽  
Author(s):  
Samuel J. T. Wardell ◽  
Attika Rehman ◽  
Lois W. Martin ◽  
Craig Winstanley ◽  
Wayne M. Patrick ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Experimental Evolution ◽  
Large Scale ◽  
Genetic Basis ◽  
Clinical Isolates ◽  
Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Health Crisis ◽  
Wide Range

AbstractPseudomonas aeruginosa is an opportunistic pathogen that causes a wide range of acute and chronic infections. An increasing number of isolates have acquired mutations that make them antibiotic resistant, making treatment more difficult. To identify resistance-associated mutations we experimentally evolved the antibiotic sensitive strain P. aeruginosa PAO1 to become resistant to three widely used anti-pseudomonal antibiotics, ciprofloxacin, meropenem and tobramycin. Mutants were able to tolerate up to 2048-fold higher concentrations of antibiotic than strain PAO1. Genome sequences were determined for thirteen mutants for each antibiotic. Each mutant had between 2 and 8 mutations. There were at least 8 genes mutated in more than one mutant per antibiotic, demonstrating the complexity of the genetic basis of resistance. Additionally, large deletions of up to 479kb arose in multiple meropenem resistant mutants. For all three antibiotics mutations arose in genes known to be associated with resistance, but also in genes not previously associated with resistance. To determine the clinical relevance of mutations uncovered in experimentally-evolved mutants we analysed the corresponding genes in 457 isolates of P. aeruginosa from patients with cystic fibrosis or bronchiectasis as well as 172 isolates from the general environment. Many of the genes identified through experimental evolution had changes predicted to be function-altering in clinical isolates but not in isolates from the general environment, showing that mutated genes in experimentally evolved bacteria can predict those that undergo mutation during infection. These findings expand understanding of the genetic basis of antibiotic resistance in P. aeruginosa as well as demonstrating the validity of experimental evolution in identifying clinically-relevant resistance-associated mutations.ImportanceThe rise in antibiotic resistant bacteria represents an impending global health crisis. As such, understanding the genetic mechanisms underpinning this resistance can be a crucial piece of the puzzle to combatting it. The importance of this research is that by experimentally evolving P. aeruginosa to three clinically relevant antibiotics, we have generated a catalogue of genes that can contribute to resistance in vitro. We show that many (but not all) of these genes are clinically relevant, by identifying variants in clinical isolates of P. aeruginosa. This research furthers our understanding of the genetics leading to resistance in P. aeruginosa and provides tangible evidence that these genes can play a role clinically, potentially leading to new druggable targets or inform therapies.

Improving the quality of bacteriological studies for infections caused by carbapenem resistant Klebsiella pneumoniae using molecular diagnostic methods

2020 ◽  
pp. 6-17
Author(s):  
A. Chizhikova ◽  
E. Lisitsyna
Keyword(s):  
Klebsiella Pneumoniae ◽  
Pathogenic Bacteria ◽  
Test System ◽  
Ease Of Use ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Fast Analysis ◽  
Carbapenem Resistant ◽  
Specificity And Sensitivity

The resistance of gram-negative pathogenic bacteria to antibiotics of the β-lactam group — carbapenems — is associated with the ability of bacteria to produce carbapenemases. The relevant task is to determine carbapenem-resistant microorganisms to control the spread of resistant strains and conduct effective pharmacotherapy. The results of the development of molecular diagnostic methods, including use of polymerase chain reaction (PCR), for the detection of carbapenem resistant Klebsiella pneumoniae bacteria are presented. A multiplex PCR test system was developed for detecting carbapenem-resistant K. pneumoniae producing KPC and OXA 48-like carbapenemases. The test system is characterized by high specificity and sensitivity, ease of use, fast analysis time (up to 3 hours). Its introduction into clinical and diagnostic practice is promising in terms of improving the quality of bacteriological studies.

scholarly journals Determination of antibiotic resistance patterns of Vibrio parahaemolyticus from shrimp and shellfish in Selangor, Malaysia

2019 ◽  
Vol 2 (1) ◽  
Author(s):  
Vengadesh Letchumanan ◽  
Nurul-Syakima Ab Mutalib ◽  
Sunny Hei Wong ◽  
Kok-Gan Chan ◽  
Learn-Han Lee
Keyword(s):  
Antibiotic Resistance ◽  
Vibrio Parahaemolyticus ◽  
Bacterial Infections ◽  
Pathogenic Bacteria ◽  
Large Scale ◽  
Health Concern ◽  
Resistance Pattern ◽  
Resistant Bacteria ◽  
Resistance Patterns ◽  
Plasmid Profiling

High consumer demand for seafood has led to the need for large-scale, reliable supply through aquaculture farming. However, bacterial infections - which can spread rapidly among the dense farming area pose a major threat to this industry. The farmers therefore often resort to extensive use of antibiotics, both prophylactically and therapeutically, in order to protect their stocks. The extensive use of antibiotics in aquaculture has been postulated to represent a major contributing factor in the rising incidence of antimicrobial resistant pathogenic bacteria in seafood; which may then lead to the spread of antimicrobial resistant bacteria in the environment as well as posing a significant threat to human health. This study aimed to characterize antibiotic resistance of Vibrio parahaemolyticus from shrimp and shellfish in Selangor, Malaysia. The antibiotic susceptibility of 385 V. parahaemolyticus isolates was investigated against 14 antibiotics followed by plasmid profiling and plasmid curing to determine the antibiotic mediation. A large number of isolates showed resistance to ampicillin (85%), amikacin (66.8%), and kanamycin (50.1%). A notable resistance pattern was also observed to the third generation cephalosporins (cefotaxime 55.8% and ceftazidime 34%). Only 338 V. parahaemolyticus isolates had 1-7 different plasmids and could be categorized into 27 patterns based on the number and pattern of plasmid present. Interestingly, there was no correlation between the number of plasmids and antibiotic resistant patterns seen in the isolates. The antibiotic resistance was mediated by both chromosomal and plasmid mediation among the resistant isolates. In summary, our results demonstrate that incidence of pathogenic V. parahaemolyticus in seafood in Selangor remains in relatively assuring levels, however the identification of antibiotic resistance among the isolates does rises a public health concern and warrants for continuous surveillance.

scholarly journals Assessing the Molecular Targets and Mode of Action of Furanone C-30 on Pseudomonas aeruginosa Quorum Sensing

Molecules ◽  
2021 ◽  
Vol 26 (6) ◽  
pp. 1620
Author(s):  
Victor Markus ◽  
Karina Golberg ◽  
Kerem Teralı ◽  
Nazmi Ozer ◽  
Esti Kramarsky-Winter ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Quorum Sensing ◽  
Conformational Changes ◽  
Mode Of Action ◽  
Pathogenic Bacteria ◽  
Molecular Targets ◽  
Resistant Bacteria ◽  
Public Healthcare ◽  
C 30

Quorum sensing (QS), a sophisticated system of bacterial communication that depends on population density, is employed by many pathogenic bacteria to regulate virulence. In view of the current reality of antibiotic resistance, it is expected that interfering with QS can address bacterial pathogenicity without stimulating the incidence of resistance. Thus, harnessing QS inhibitors has been considered a promising approach to overriding bacterial infections and combating antibiotic resistance that has become a major threat to public healthcare around the globe. Pseudomonas aeruginosa is one of the most frequent multidrug-resistant bacteria that utilize QS to control virulence. Many natural compounds, including furanones, have demonstrated strong inhibitory effects on several pathogens via blocking or attenuating QS. While the natural furanones show no activity against P. aeruginosa, furanone C-30, a brominated derivative of natural furanone compounds, has been reported to be a potent inhibitor of the QS system of the notorious opportunistic pathogen. In the present study, we assess the molecular targets and mode of action of furanone C-30 on P. aeruginosa QS system. Our results suggest that furanone C-30 binds to LasR at the ligand-binding site but fails to establish interactions with the residues crucial for the protein’s productive conformational changes and folding, thus rendering the protein dysfunctional. We also show that furanone C-30 inhibits RhlR, independent of LasR, suggesting a complex mechanism for the agent beyond what is known to date.

scholarly journals Bacteria and Their Antibiotic Resistance Profiles in Ambient Air in Accra, Ghana, February 2020: A Cross-Sectional Study

2021 ◽  
Vol 6 (3) ◽  
pp. 110
Author(s):  
Godfred Saviour Kudjo Azaglo ◽  
Mohammed Khogali ◽  
Katrina Hann ◽  
John Alexis Pwamang ◽  
Emmanuel Appoh ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Pathogenic Bacteria ◽  
Ambient Air ◽  
Cross Sectional Study ◽  
Bacillus Species ◽  
Resistant Bacteria ◽  
Cross Sectional ◽  
Antibiotic Resistant ◽  
Bacterial Counts ◽  
Inappropriate Use

Inappropriate use of antibiotics has led to the presence of antibiotic-resistant bacteria in ambient air. There is no published information about the presence and resistance profiles of bacteria in ambient air in Ghana. We evaluated the presence and antibiotic resistance profiles of selected bacterial, environmental and meteorological characteristics and airborne bacterial counts in 12 active air quality monitoring sites (seven roadside, two industrial and three residential) in Accra in February 2020. Roadside sites had the highest median temperature, relative humidity, wind speed and PM10 concentrations, and median airborne bacterial counts in roadside sites (115,000 CFU/m3) were higher compared with industrial (35,150 CFU/m3) and residential sites (1210 CFU/m3). Bacillus species were isolated in all samples and none were antibiotic resistant. There were, however, Pseudomonas aeruginosa, Escherichia coli, Pseudomonas species, non-hemolytic Streptococci, Coliforms and Staphylococci species, of which six (50%) showed mono-resistance or multidrug resistance to four antibiotics (penicillin, ampicillin, ciprofloxacin and ceftriaxone). There was a positive correlation between PM10 concentrations and airborne bacterial counts (rs = 0.72), but no correlations were found between PM10 concentrations and the pathogenic bacteria nor their antibiotic resistance. We call for the expansion of surveillance of ambient air to other cities of Ghana to obtain nationally representative information.

scholarly journals Antimicrobial Susceptibility of Lactic Acid Bacteria Strains of Potential Use as Feed Additives - The Basic Safety and Usefulness Criterion

2021 ◽  
Vol 8 ◽  
Author(s):  
Ilona Stefańska ◽  
Ewelina Kwiecień ◽  
Katarzyna Jóźwiak-Piasecka ◽  
Monika Garbowska ◽  
Marian Binek ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Resistance Genes ◽  
Pathogenic Bacteria ◽  
Feed Additives ◽  
Health Concern ◽  
Resistant Bacteria ◽  
Safety Criterion ◽  
Phenotypic Resistance

The spread of resistance to antibiotics is a major health concern worldwide due to the increasing rate of isolation of multidrug resistant pathogens hampering the treatment of infections. The food chain has been recognized as one of the key routes of antibiotic resistant bacteria transmission between animals and humans. Considering that lactic acid bacteria (LAB) could act as a reservoir of transferable antibiotic resistance genes, LAB strains intended to be used as feed additives should be monitored for their safety. Sixty-five LAB strains which might be potentially used as probiotic feed additives or silage inoculants, were assessed for susceptibility to eight clinically relevant antimicrobials by a minimum inhibitory concentration determination. Among antimicrobial resistant strains, a prevalence of selected genes associated with the acquired resistance was investigated. Nineteen LAB strains displayed phenotypic resistance to one antibiotic, and 15 strains were resistant to more than one of the tested antibiotics. The resistance to aminoglycosides and tetracyclines were the most prevalent and were found in 37 and 26% of the studied strains, respectively. Phenotypic resistance to other antimicrobials was found in single strains. Determinants related to resistance phenotypes were detected in 15 strains as follows, the aph(3″)-IIIa gene in 9 strains, the lnu(A) gene in three strains, the str(A)-str(B), erm(B), msr(C), and tet(M) genes in two strains and the tet(K) gene in one strain. The nucleotide sequences of the detected genes revealed homology to the sequences of the transmissible resistance genes found in lactic acid bacteria as well as pathogenic bacteria. Our study highlights that LAB may be a reservoir of antimicrobial resistance determinants, thus, the first and key step in considering the usefulness of LAB strains as feed additives should be an assessment of their antibiotic resistance. This safety criterion should always precede more complex studies, such as an assessment of adaptability of a strain or its beneficial effect on a host. These results would help in the selection of the best LAB strains for use as feed additives. Importantly, presented data can be useful for revising the current microbiological cut-off values within the genus Lactobacillus and Pediococcus.

Comparing antibiotic resistance in commensal and pathogenic bacteria isolated from wild-caught South Carolina shrimps vs. farm-raised imported shrimps

2007 ◽  
Vol 53 (7) ◽  
pp. 919-924 ◽  
Author(s):  
Kavitha Boinapally ◽  
Xiuping Jiang
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
South Carolina ◽  
Antimicrobial Resistance ◽  
Pathogenic Bacteria ◽  
Vibrio Vulnificus ◽  
Resistant Bacteria ◽  
Microbiological Analysis ◽  
Salmonella Spp ◽  
Plate Counts

The objective of this study was to assess and differentiate wild-caught South Carolina (SC) shrimps from imported shrimps on the basis of microbiological analysis. Seven wild-caught SC shrimp and 13 farm-raised imported shrimp samples were analyzed. Total plate counts from wild-caught shrimp samples ranged from 4.3 to 7.0 log10 CFU/g, whereas counts from imported shrimp samples ranged from 3.2 to 5.7 log10 CFU/g. There was no difference (P > 0.05) between total bacterial counts of wild-caught SC shrimp and farm-raised imported shrimp. However, the percentages of bacteria with reduced susceptibility towards ceftriaxone and tetracycline were higher (P < 0.05) for farm-raised shrimp than for wild-caught samples. Salmonella spp. detected only in one farm-raised sample was resistant to ampicillin, ceftriaxone, gentamicin, streptomycin, and trimethoprim. Vibrio vulnificus was detected in both wild-caught and farm-raised shrimp samples; however, only the isolate from farm-raised shrimp was resistant to nalidixic acid and trimethoprim. Escherichia coli detected in one wild-caught sample was resistant to ampicillin. Both Listeria spp. and Salmonella spp. were absent with wild-caught SC samples. Therefore, the presence of more ceftriaxone- and tetracycline-resistant bacteria and the observed antimicrobial resistance phenotypes of isolates from the imported shrimp may reflect the possible use of antibiotics in raising shrimp in those countries.

A novel molecular diagnostic method for the gut content analysis of Philaenus DNA

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Isabel Rodrigues ◽  
Vítor Ramos ◽  
Jacinto Benhadi-Marín ◽  
Aránzazu Moreno ◽  
Alberto Fereres ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Control Strategies ◽  
Control Measure ◽  
Coi Gene ◽  
Molecular Diagnostic ◽  
Generalist Predators ◽  
Specificity And Sensitivity ◽  
Vector Borne ◽  
Polymerase Chain ◽  
Primer Sets

AbstractPhilaenus spumarius is a vector of Xylella fastidiosa, one of the most dangerous plants pathogenic bacteria worldwide. There is currently no control measure against this pathogen. Thus, the development of vector control strategies, like generalist predators, such as spiders, could be essential to limit the spread of this vector-borne pathogen. In this study, a polymerase chain reaction (PCR)-based approach was developed to principally detect DNA of P. spumarius in the spider’s gut. Accordingly, 20 primer pairs, targeting the mitochondrial cytochrome oxidase I (COI) and cytochrome b (cytB) genes, were tested for specificity, sensitivity, and efficiency in detecting P. spumarius DNA. Overall, two primer sets, targeting COI gene (COI_Ph71F/COI_Ph941R) and the cytB gene (cytB_Ph85F/cytB_Ph635R), showed the highest specificity and sensitivity, being able to amplify 870 pb and 550 bp fragments, respectively, with P. spumarius DNA concentrations 100-fold lower than that of the DNA of non-target species. Among these two primer sets, the cytB_Ph85F/cytB_Ph635R was able to detect P. spumarius in the spider Xysticus acerbus, reaching 50% detection success 82 h after feeding. The feasibility of this primer set to detect predation of P. spumarius by spiders was confirmed in the field, where 20% of the collected spiders presented positive amplifications.

Antimicrobial Resistance of Bacterial Flora Associated with Bovine Products in South Africa

1999 ◽  
Vol 62 (6) ◽  
pp. 615-618 ◽  
Author(s):  
THUREYAH MANIE ◽  
VOLKER S. BRÖZEL ◽  
WALTER J. VEITH ◽  
PIETER A. GOUWS
Keyword(s):  
South Africa ◽  
Antibiotic Resistance ◽  
Pathogenic Bacteria ◽  
Bacterial Flora ◽  
Resistant Bacteria ◽  
Antibiotic Resistant ◽  
Selective Pressures ◽  
Unpasteurized Milk ◽  
Plate Counts ◽  
Resistance To Penicillin

The administration of subtherapeutic doses of antibiotics to livestock introduces selective pressures that may lead to the emergence and dissemination of resistant bacteria. This study determined the antibiotic-resistance spectra of the microbial flora found on freshly slaughtered and retail beef and in unpasteurized and pasteurized packaged milk. Staphylococci, Enterobacteriaceae, and isolates from total aerobic plate counts were tested for resistance to vancomycin, streptomycin, methicillin, tetracycline, and gentamicin using the disc diffusion susceptibility test and resistance to penicillin was determined by using oxacillin. A larger proportion of resistance to most antibiotics, except for vancomycin, was displayed by isolates from abattoir samples. The incidence of multiple antibiotic resistance (MAR) pathogenic bacteria is also higher in the abattoir. Resistance genes lost because of lack of selective pressure or resistant flora being replaced by more sensitive flora during processing is the reason for the lower incidence of MAR pathogenic bacteria among retail samples. These resistant bacteria can be transferred to humans through the consumption of rare or raw beef and unpasteurized milk, thus rendering the resultant food-related infections difficult to treat. The present findings clearly demonstrate that antibiotic-resistant bacteria in beef and milk pose a serious problem in South Africa.

scholarly journals Evaluation on the Use of β-Lactamase and Aminoglycoside Modifying Enzyme Gene Sequences as Markers for the Early Detection of Antibiotic Resistance Profile ofPseudomonas aeruginosa

2004 ◽  
Vol 20 (6) ◽  
pp. 317-323 ◽  
Author(s):  
Victor A. Doss ◽  
S. Parvathi ◽  
B. Appala Raju ◽  
N. Abitha Devi
Keyword(s):  
Antibiotic Resistance ◽  
Dna Sequences ◽  
Antibiotic Resistance Genes ◽  
Diagnostic Methods ◽  
Clinical Isolates ◽  
Resistance Profile ◽  
Antibiotic Resistance Profile ◽  
Polymerase Chain ◽  
Labeled Probe ◽  
Hospital Acquired

Pseudomonas aeruginosais one of the major causes of infections including the hospital acquired (Nosocomial) infections. Detection of them and their antibiotic resistance profile by conventional method takes about three days. Recently, DNA based diagnostic methods are being used for the identification of the pathogens. Hence we have tested a rapid and sensitive method using DNA sequences as markers for detecting the presence of three genes coding for the enzymes that inactivate the two most commonly used Anti-pseudomonadal drugs such as β-lactam antibiotics (Penicillin, and its derivatives) and Aminoglycosides such as Gentamicin, Tobramycin, Amikacin, Streptomycin. The internal region of these genes were used for designing and synthesizing primers and these primers were used in Polymerase Chain Reaction (PCR) to screen for the presence of these genes in the clinical isolates and to label them non-radioactively with Biotin. They in turn were used to detect the presence of the antibiotic resistance genes in the clinical isolates by hybridization. The specificity (ratio of positive results obtained in both methods and the sensitivity (the minimum amount of sample DNA and the labeled probe required for the tests) were evaluated.

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