scholarly journals Assessing the Role of Potential Biomarkers in Antimony Susceptible and Resistant Clinical Isolates of L. donovani from India

2016 ◽  
Vol 4 (1) ◽  
pp. 1-13
Author(s):  
Mahendra Maharjan ◽  
Swati Mandal ◽  
Rentala Madhubala
Keyword(s):  
Protein A ◽  
Building Blocks ◽  
Underlying Mechanism ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Polyamine Biosynthesis ◽  
Glutathione Biosynthesis ◽  
Antimony Resistance ◽  
Rate Limiting

Failure of antimonial drugs, the mainstay therapy for leishmaniasis has become an escalating problem in the treatment of Indian leishmaniasis. Using 14 clinical isolates from both visceral (VL) and post-kala-azar dermal leismaniasis (PKDL) patients, we have examined the role of ATP-binding cassette transporter (ABC transporter) gene, multidrug resistant protein A (MRPA) and two building blocks of the major thiol, trypanothione namely, ornithine decarboxylase gene (ODC) (a rate limiting enzyme in the polyamine biosynthesis) and γ-glutamylcysteine synthetase (γ-GCS) (a rate limiting enzyme in glutathione biosynthesis) in antimony resistance. Amplification of these three genes was observed in some but not all clinical isolates. Increased expression of the three RNAs as determined by real-time PCR was observed in all SAG-R clinical isolates. Significant increase in cysteine and glutathione levels was observed in the resistant isolates. Our studies report the underlying mechanism of antimony resistance in the clinical isolates.

scholarly journals Decreased expression of femXAB genes and fnbp mediated biofilm pathways in OS-MRSA clinical isolates

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Umarani Brahma ◽  
Paresh Sharma ◽  
Shweta Murthy ◽  
Savitri Sharma ◽  
Shalini Chakraborty ◽  
...  
Keyword(s):  
Antibiotic Susceptibility ◽  
Biofilm Formation ◽  
Multidrug Resistant ◽  
Agglutination Test ◽  
Clinical Isolates ◽  
Ocular Infections ◽  
Biofilm Production ◽  
Fibronectin Binding ◽  
Sequence Types

Abstract Methicillin-Resistant Staphylococcus aureus (MRSA) is a significant threat to human health. Additionally, biofilm forming bacteria becomes more tolerant to antibiotics and act as bacterial reservoir leading to chronic infection. In this study, we characterised the antibiotic susceptibility, biofilm production and sequence types (ST) of 74 randomly selected clinical isolates of S. aureus causing ocular infections. Antibiotic susceptibility revealed 74% of the isolates as resistant against one or two antibiotics, followed by 16% multidrug-resistant isolates (MDR), and 10% sensitive. The isolates were characterized as MRSA (n = 15), Methicillin-sensitive S. aureus (MSSA, n = 48) and oxacillin susceptible mecA positive S. aureus (OS-MRSA, n = 11) based on oxacillin susceptibility, mecA gene PCR and PBP2a agglutination test. All OS-MRSA would have been misclassified as MSSA on the basis of susceptibility test. Therefore, both phenotypic and genotypic tests should be included to prevent strain misrepresentation. In addition, in-depth studies for understanding the emerging OS-MRSA phenotype is required. The role of fem XAB gene family has been earlier reported in OS-MRSA phenotype. Sequence analysis of the fem XAB genes revealed mutations in fem × (K3R, H11N, N18H and I51V) and fem B (L410F) genes. The fem XAB genes were also found down-regulated in OS-MRSA isolates in comparison to MRSA. In OS-MRSA isolates, biofilm formation is regulated by fibronectin binding proteins A & B. Molecular typing of the isolates revealed genetic diversity. All the isolates produced biofilm, however, MRSA isolates with strong biofilm phenotype represent a worrisome situation and may even result in treatment failure.

scholarly journals Role of Aminoglycoside-Modifying Enzymes (AMEs) in Resistance to Aminoglycosides among Clinical Isolates of Pseudomonas aeruginosa in the North of Iran

2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Leila Ahmadian ◽  
Zahra Norouzi Bazgir ◽  
Mohammad Ahanjan ◽  
Reza Valadan ◽  
Hamid Reza Goli
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Resistance Pattern ◽  
Dilution Method ◽  
The North ◽  
Agar Diffusion Test ◽  
North Of Iran ◽  
Encoding Genes

In recent years, the prevalence of resistance to aminoglycosides among clinical isolates of Pseudomonas aeruginosa is increasing. The aim of this study was to investigate the role of aminoglycoside-modifying enzymes (AMEs) in resistance to aminoglycosides in clinical isolates of P. aeruginosa. The clinical isolates were collected from different hospitals. Disk agar diffusion test was used to determine the antimicrobial resistance pattern of the clinical isolates, and the minimum inhibitory concentration of aminoglycosides was detected by microbroth dilution method. The PCR was performed for discovery of aminoglycoside-modifying enzyme-encoding genes. Among 100 screened isolates, 43 (43%) isolates were resistant to at least one tested aminoglycosides. However, 13 (13%) isolates were resistant to all tested aminoglycosides and 37 isolates were detected as multidrug resistant (MDR). The resistance rates of P. aeruginosa isolates against tested antibiotics were as follows: ciprofloxacin (41%), piperacillin-tazobactam (12%), cefepime (32%), piperacillin (26%), and imipenem (31%). However, according to the MIC method, 13%, 32%, 33%, and 37% of the isolates were resistant to amikacin, gentamicin, tobramycin, and netilmicin, respectively. The PCR results showed that AAC(6 ′ )-Ib was the most commonly (26/43, 60.4%) identified AME-encoding gene followed by AAC(6 ′ )-IIa (41.86%), APH(3 ′ )-IIb (34.8%), ANT(3 ″ )-Ia (18.6), ANT(2 ″ )-Ia (13.95%), and APH(3 ″ )-Ib (2.32%). However, APH(3 ′ )-Ib was not found in any of the studied isolates. The high prevalence of AME-encoding genes among aminoglycoside-resistant P. aeruginosa isolates in this area indicated the important role of AMEs in resistance to these antibiotics similar to most studies worldwide. Due to the transmission possibility of these genes between the Gram-negative bacteria, we need to control the prescription of aminoglycosides in hospitals.

scholarly journals Role of the ABC Transporter MRPA (PGPA) in Antimony Resistance in Leishmania infantum Axenic and Intracellular Amastigotes

2005 ◽  
Vol 49 (5) ◽  
pp. 1988-1993 ◽  
Author(s):  
Karima El Fadili ◽  
Nadine Messier ◽  
Philippe Leprohon ◽  
Gaétan Roy ◽  
Chantal Guimond ◽  
...  
Keyword(s):  
Abc Transporter ◽  
Leishmania Infantum ◽  
Dna Microarrays ◽  
Specific Inhibitor ◽  
Buthionine Sulfoximine ◽  
Protozoan Parasite ◽  
Sodium Stibogluconate ◽  
Glutathione Biosynthesis ◽  
Antimony Resistance

ABSTRACT Antimonial compounds are the mainstay for the treatment of infections with the protozoan parasite Leishmania. We present our studies on Leishmania infantum amastigote parasites selected for resistance to potassium antimonyl tartrate [Sb(III)]. Inside macrophages, the Sb(III)-selected cells are cross-resistant to sodium stibogluconate (Pentostam), the main drug used against Leishmania. Putative alterations in the level of expression of more than 40 genes were compared between susceptible and resistant axenic amastigotes using customized DNA microarrays. The expression of three genes coding for the ABC transporter MRPA (PGPA), S-adenosylhomocysteine hydrolase, and folylpolyglutamate synthase was found to be consistently increased. The levels of cysteine were found to be increased in the mutant. Transfection of the MRPA gene was shown to confer sodium stibogluconate resistance in intracellular parasites. This MRPA-mediated resistance could be reverted by using the glutathione biosynthesis-specific inhibitor buthionine sulfoximine. These results highlight for the first time the role of MRPA in antimony resistance in the amastigote stage of the parasite and suggest a strategy for reversing resistance.

scholarly journals In Vitro Activity of Tetracycline Analogs against Multidrug-resistant and Extensive drug resistance Clinical Isolates of Mycobacterium tuberculosis

2019 ◽  
Author(s):  
Qi Ouyang ◽  
Dachuan Lin ◽  
Guofang Deng ◽  
Zhihua Wen ◽  
Houming Liu ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Peak Plasma ◽  
Peak Plasma Concentration ◽  
Underlying Mechanism ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Ammonium Bromide ◽  
Resistant Tuberculosis ◽  
Mdr Tb

Abstract Background Multidrug-resistant tuberculosis (MDR-TB) has become a big threaten to global health . The current strategy for treatment of MDR-TB and extensive drug resistant tuberculosis (XDR-TB) is with low efficacy and high side effect. While new drug is fundamental for cure MDR-TB, repurposing the Food and Drug Administration (FDA)-approved drugs represents an alternative soluation with less cost.Methods The activity of 8 tetracycline-class antibiotics against mycobacterium tuberculosis ( M.tb ) were determined by Minimum Inhibitory Concentration (MIC) in vitro. A transposon M.smeg libraries was generated by using the Harm phage and then used to isolate the conditional growth mutants in doxycycline containing plate. 11 mutants were isolated and genomic DNAs were extracted using the cetyltrimethyl ammonium bromide (CTAB) method and analyzed by whole genome sequencing.Results We found that three of eight drugs efficiently inhibited mycobacteria growth under the peak plasma concentration in the human body. Further tests showed these three tetracycline analogs (demeclocycline, doxycycline and methacycline) had antimicrobial activity against seven clinical isolates, including MDR and XDR strains. Among them, Doxycycline had the lowest MICs in all mycobacteria strains tested in this study. By using a transposon library, we identify the insertion of transposon in two genes, porin and MshA, associate with the resistant to doxycycline.Conclusions Our findings show that tetracycline analogs such as doxycycline, has bactericidal activity against not only drug sensitive M.tb , but also clinical MDR and XDR strains, provided proof of concept to repurpose doxycycline to fight MDR-TB and XDR-TB. Further investigations are warranted to clarify the underlying mechanism and optimize the strategy in combination with other anti-TB drugs.

scholarly journals Mutations in an Aquaglyceroporin as a Proven Marker of Antimony Clinical Resistance in the Parasite Leishmania donovani

2020 ◽  
Author(s):  
Jade-Eva Potvin ◽  
Philippe Leprohon ◽  
Marine Queffeulou ◽  
Shyam Sundar ◽  
Marc Ouellette
Keyword(s):  
Leishmania Donovani ◽  
Indian Subcontinent ◽  
Copy Number Variations ◽  
Clinical Isolates ◽  
Gene Copy ◽  
Specific Gene ◽  
Single Nucleotide Variants ◽  
Clinical Resistance ◽  
Antimony Resistance

Abstract Background Antimonial drugs have long been the mainstay to treat visceral leishmaniasis. Their use has been discontinued in the Indian subcontinent because of drug resistance, but they are still clinically useful elsewhere. The goal of this study was to find markers of antimony resistance in Leishmania donovani clinical isolates and validate experimentally their role in resistance. Methods The genomes of sensitive and antimony-resistant clinical isolates were sequenced. The role of a specific gene in contributing to resistance was studied by CRISPR-Cas9–mediated gene editing and intracellular drug sensitivity assays. Results Both gene copy number variations and single nucleotide variants were associated with antimony resistance. A homozygous insertion of 2 nucleotides was found in the gene coding for the aquaglyceroporin AQP1 in both resistant isolates. Restoring the wild-type AQP1 open reading frame re-sensitized the 2 independent resistant isolates to antimonials. Alternatively, editing the genome of a sensitive isolate by incorporating the 2-nucleotide insertion in its AQP1 gene led to antimony-resistant parasites. Conclusions Through genomic analysis and CRISPR-Cas9–mediated genome editing we have proven the role of the AQP1 mutations in antimony clinical resistance in L. donovani.

scholarly journals Screening antibiotics using an Hoechst 33342 dye-accumulation assay to detect efflux activity in Acinetobacter baumannii clinical isolates

2018 ◽  
Vol 11 (4) ◽  
pp. 371-378 ◽  
Author(s):  
In-Sun Choi ◽  
Choon-Mee Kim ◽  
Sook-Jin Jang
Keyword(s):  
Multidrug Resistance ◽  
Acinetobacter Baumannii ◽  
Efflux Pump ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Drug Efflux ◽  
Hoechst 33342 ◽  
Efflux Pump Inhibitor ◽  
Significant Difference

AbstractBackgroundUnderstanding the contribution of efflux pumps to the resistance of antibiotics is useful when considering strategies for antimicrobial therapy.ObjectivesTo assess the role of efflux activity on the resistance of antibiotics commonly used in hospitals.MethodsWe analyzed the efflux activity of 120 clinical isolates of Acinetobacter baumannii using an Hoechst 33342 (H33342) dye-accumulation assay. We compared the indicators for efflux activity of susceptible and non-susceptible groups of each of 16 tested antibiotics. To determine the role of efflux activity on resistance to an antibiotic, we used 3 criteria based on the results of the H33342-accumulation assay.ResultsThe evaluation suggests that efflux activity contributed to resistance to the following 11 antibiotics: cefepime, cefotaxime, ceftazidime, ciprofloxacin, gentamicin, imipenem, meropenem, piperacillin, piperacillin/tazobactam, ticarcillin/ clavulanic acid, and tigecycline. However, ampicillin/sulbactam, minocycline, and trimethoprim/sulfamethoxazole did not meet the criteria, suggesting resistance may not be mediated by efflux activity. A significant difference in efflux activity was observed between bacteria belonging to the multidrug-resistant Acinetobacter baumannii (MDRAB) group and those belonging to the non-MDRAB group.ConclusionsEfflux activity may contribute to multidrug resistance and particularly resistance to numerous antibiotics used in hospitals. These antibiotics would be good candidates for combination therapeutic regimens consisting of an antibiotic and an efflux pump inhibitor as an adjuvant to combat drug efflux.

Differential gene expression analysis in antimony-unresponsive Indian kala azar (visceral leishmaniasis) clinical isolates by DNA microarray

Parasitology ◽  
2007 ◽  
Vol 134 (6) ◽  
pp. 777-787 ◽  
Author(s):  
N. SINGH ◽  
R. ALMEIDA ◽  
H. KOTHARI ◽  
P. KUMAR ◽  
G. MANDAL ◽  
...  
Keyword(s):  
Leishmania Donovani ◽  
Leishmania Major ◽  
Clinical Isolates ◽  
Resistant Isolate ◽  
Closely Related Species ◽  
Kala Azar ◽  
Antimony Resistance ◽  
Sensitive Isolate ◽  
Differential Gene

SUMMARYIn this study, cDNA microarray analysis of a closely related species,Leishmania major, was used as a screening tool to compare antimonial-resistant and susceptible clinical isolates ofLeishmania donovaniin order to to identify candidate genes on the basis of antimony resistance. Clinically confirmed resistant isolate 39 and sensitive isolate 2001 were used in this study. Many differentially regulated genes were identified whose expression levels differ in sodium antimony gluconate (SAG)-treated patients. Interestingly, genes on the array, showing changes in expression of over 2-fold revealed the identity of ABC transporters, which are known determinants of drug resistance in laboratory mutants. The functionality of the transporters was validated by flow cytometry which, being biologically informative, provides direct clues to gene function. The results suggest that isolate 39 could have developed resistance by an increased multidrug resistance protein (MRP)-like pump. This study provides preliminary clues to the role of a thiol-dependent efflux system in antimonial resistant clinical isolates ofLeishmania donovani.

scholarly journals Role of OmpA in the Multidrug Resistance Phenotype of Acinetobacter baumannii

2013 ◽  
Vol 58 (3) ◽  
pp. 1806-1808 ◽  
Author(s):  
Younes Smani ◽  
Anna Fàbrega ◽  
Ignasi Roca ◽  
Viviana Sánchez-Encinales ◽  
Jordi Vila ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Acinetobacter Baumannii ◽  
Protein A ◽  
Multidrug Resistant ◽  
Nosocomial Pathogen ◽  
Content Type ◽  
Resistance Phenotype ◽  
Resistant Strains ◽  
First Time

ABSTRACTAcinetobacter baumanniihas emerged as a nosocomial pathogen with an increased prevalence of multidrug-resistant strains. The role of the outer membrane protein A (OmpA) in antimicrobial resistance remains poorly understood. In this report, disruption of theompAgene led to decreased MICs of chloramphenicol, aztreonam, and nalidixic acid. We have characterized, for the first time, the contribution of OmpA in the antimicrobial resistance phenotype ofA. baumannii.

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