scholarly journals Application of kDNA Minicircle PCR-RFLP to Characterize Leishmania donovani Clinical Isolates Obtained from Post-Kala-Azar Dermal Leishmaniasis in Eastern Nepal

2019 ◽  
Vol 2019 ◽  
pp. 1-8
Author(s):  
Ojesh Pokhrel ◽  
Keshav Rai ◽  
Narayan Raj Bhattarai ◽  
Suman Rijal ◽  
Arpana Rijal ◽  
...  
Keyword(s):  
Leishmania Donovani ◽  
Clinical Isolates ◽  
Rdna Genes ◽  
Kala Azar ◽  
Pcr Rflp ◽  
Potential Reservoir ◽  
Rflp Assay ◽  
Epidemiological Characterization

Post-kala-azar dermal leishmaniasis (PKDL) is a skin manifestation of visceral leishmaniasis (VL) which develops after apparent cure in some patients. PKDL is considered as the potential reservoir for the VL infection. Molecular epidemiological characterization of L. donovani isolates obtained from VL and PKDL isolates is essentially required in order to understand the transmission dynamics of the VL infection. To date, genetic variation among the VL and PKDL L. donovani isolates was not fully elucidated. Therefore, 14 clinical isolates from VL and 4 clinical isolates from PKDL were speciated by hsp70 and rDNA genes. Further characterization of L. donovani by haspB PCR demonstrates two different genotypes. All PKDL isolates have the same genetic structure. kDNA PCR-RFLP assay revealed 18 different genotypes; however, structural analysis showed the two distinct kDNA genotype population (k = 2). The kDNA fingerprint patterns of parasites from hilly districts were clustered separately from low-land districts. Therefore, further study with a large number of samples is urgently required for systematic characterization of the clinical isolates to track the molecular epidemiology of the Leishmania donovani causing VL and the role of PKDL as a reservoir.

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.

First Detection of OXA-10 Extended-Spectrum Beta-Lactamases and the Occurrence of mexR and nfxB in Clinical Isolates of Pseudomonas aeruginosa from Nigeria

Chemotherapy ◽  
2015 ◽  
Vol 61 (2) ◽  
pp. 87-92 ◽  
Author(s):  
Bamidele T. Odumosu ◽  
Bola A. Adeniyi ◽  
Ram Chandra
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Efflux Pump ◽  
Clinical Isolates ◽  
Beta Lactamases ◽  
First Report ◽  
Extended Spectrum ◽  
Pcr Rflp ◽  
Extended Spectrum Beta Lactamases ◽  
Regulator Genes

Background: The characterization of β-lactamase production in Pseudomonasaeruginosa is rarely reported in Nigeria. The objective of this study was to investigate the occurrence and characterize the different β-lactamases as well as mechanisms of fluoroquinolones resistance among P. aeruginosa isolated from various clinical sources from Nigeria. Materials and Method: Isolates were investigated using PCR, RFLP and sequencing for the detection of various β-lactamases and efflux pump regulator genes. Result: The prevalence of OXA-10, AmpC, CTX-M and SHV in P. aeruginosa was 80, 70, 5 and 5%, respectively. The coexistence of blaOXA-10 with blaAmpC, blaSHV and blaCTX-M was reported in 40, 5 and 5% of isolates, respectively. The efflux pump regulator genes mexR and nfxB were both amplified in 45% of the OXA-10-positive isolates. Conclusion: This is the first report of the characterization of OXA-10 extended-spectrum β-lactamases and occurrence of mexR and nfxB efflux regulator genes in clinical isolates of P. aeruginosa in Nigeria.

scholarly journals Characterization of Biofilm Formation and the Role of BCR1 in Clinical Isolates of Candida parapsilosis

2013 ◽  
Vol 13 (4) ◽  
pp. 438-451 ◽  
Author(s):  
Srisuda Pannanusorn ◽  
Bernardo Ramírez-Zavala ◽  
Heinrich Lünsdorf ◽  
Birgitta Agerberth ◽  
Joachim Morschhäuser ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Candida Parapsilosis ◽  
Clinical Isolates ◽  
Wild Type ◽  
Content Type ◽  
Initial Adhesion ◽  
High Level ◽  
Increased Susceptibility

ABSTRACT In Candida parapsilosis , biofilm formation is considered to be a major virulence factor. Previously, we determined the ability of 33 clinical isolates causing bloodstream infection to form biofilms and identified three distinct groups of biofilm-forming strains (negative, low, and high). Here, we establish two different biofilm structures among strains forming large amounts of biofilm in which strains with complex spider-like structures formed robust biofilms on different surface materials with increased resistance to fluconazole. Surprisingly, the transcription factor Bcr1, required for biofilm formation in Candida albicans and C. parapsilosis , has an essential role only in strains with low capacity for biofilm formation. Although BCR1 leads to the formation of more and longer pseudohyphae, it was not required for initial adhesion and formation of mature biofilms in strains with a high level of biofilm formation. Furthermore, an additional phenotype affected by BCR1 was the switch in colony morphology from rough to crepe, but only in strains forming high levels of biofilm. All bcr1 Δ/Δ mutants showed increased proteolytic activity and increased susceptibility to the antimicrobial peptides protamine and RP-1 compared to corresponding wild-type and complemented strains. Taken together, our results demonstrate that biofilm formation in clinical isolates of C. parapsilosis is both dependent and independent of BCR1 , but even in strains which showed a BCR1 -independent biofilm phenotype, BCR1 has alternative physiological functions.

scholarly journals Genetic analysis of Cryptosporidium from 2414 humans with diarrhoea in England between 1985 and 2000

2006 ◽  
Vol 55 (6) ◽  
pp. 703-707 ◽  
Author(s):  
F. Leoni ◽  
C. Amar ◽  
G. Nichols ◽  
S. Pedraza-Díaz ◽  
J. McLauchlin
Keyword(s):  
Genetic Analysis ◽  
Cryptosporidium Parvum ◽  
18S Rdna ◽  
Oocyst Wall ◽  
Cryptosporidium Hominis ◽  
Rdna Genes ◽  
Cryptosporidium Andersoni ◽  
Faecal Samples ◽  
Pcr Rflp

The characterization of Cryptosporidium using DNA extracted from whole faecal samples collected from 2414 humans with diarrhoea in England between 1985 and 2000 where cryptosporidial oocysts were detected using conventional methods is described. Characterization was achieved by PCR/RFLP and DNA sequencing of fragments of the Cryptosporidium oocyst wall protein and the 18S rDNA genes. Cryptosporidium parvum was detected in 56.1 % of cases, Cryptosporidium hominis in 41.7 % and a mixture of C. parvum and C. hominis in 0.9 %. In the remainder of cases, Cryptosporidium meleagridis (0.9 %), Cryptosporidium felis (0.2 %), Cryptosporidium andersoni (0.1 %), Cryptosporidium canis (0.04 %), Cryptosporidium suis (0.04 %) and the Cryptosporidium cervine type (0.04 %) were detected.

scholarly journals Characterization of the recent clinical isolates of Indian Kala-azar patients by RAPD-PCR method

2011 ◽  
Vol 35 (2) ◽  
pp. 116-122 ◽  
Author(s):  
Supriya Khanra ◽  
Subir K. Bandopadhyay ◽  
Priyanka Chakraborty ◽  
Sanchita Datta ◽  
Dinesh Mondal ◽  
...  
Keyword(s):  
Clinical Isolates ◽  
Kala Azar ◽  
Rapd Pcr ◽  
Pcr Method

scholarly journals Accelerated Control of Visceral Leishmania donovani Infection in Interleukin-6-Deficient Mice

2008 ◽  
Vol 76 (9) ◽  
pp. 4088-4091 ◽  
Author(s):  
Henry W. Murray
Keyword(s):  
Visceral Leishmaniasis ◽  
Interleukin 6 ◽  
Leishmania Donovani ◽  
Wild Type ◽  
Therapeutic Blockade ◽  
Th1 Cell ◽  
Kala Azar ◽  
Type Response ◽  
Deficient Mice

ABSTRACT In patients with visceral leishmaniasis, increased levels of circulating interleukin-6 (IL-6) regularly accompany fully expressed, progressive infections (kala-azar). To experimentally test the role of IL-6, responses to an intracellular Leishmania donovani infection in the livers of IL-6−/− and wild-type mice were compared. IL-6−/− mice showed an enhanced control of the infection and earlier, rapid parasite killing along with additional evidence of a stimulated antileishmanial Th1-cell-type response: increased levels of circulating gamma interferon, accelerated granuloma assembly, and heightened responsiveness to chemotherapy. In this model of visceral leishmaniasis, IL-6 appears to act in a suppressive, macrophage-deactivating fashion, which identifies it as a potential target for therapeutic blockade.

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.

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