Laffitte MCN, Leprohon P, Papadopoulou B and Ouellette M. Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.12688/f1000research.9218.1)
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1Centre de Recherche en Infectiologie du Centre de Recherche du CHU Québec, and Département de Microbiologie, Infectiologie et Immunologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada
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Abstract
Leishmania has a plastic genome, and drug pressure can select for gene copy number variation (CNV). CNVs can apply either to whole chromosomes, leading to aneuploidy, or to specific genomic regions. For the latter, the amplification of chromosomal regions occurs at the level of homologous direct or inverted repeated sequences leading to extrachromosomal circular or linear amplified DNAs. This ability of Leishmania to respond to drug pressure by CNVs has led to the development of genomic screens such as Cos-Seq, which has the potential of expediting the discovery of drug targets for novel promising drug candidates.
Keywords
Leishmania, Ploidy, Drug Resistance, Mode of action, Cos-Seq
Corresponding authors:
Barbara Papadopoulou, Marc Ouellette
Competing interests:
The authors declare that they have no competing interests.
Grant information:
BP is supported by grants from the Canadian Institutes of Health Research, the Natural Sciences and Engineering Research Council of Canada, and the Ministère du Developpement Économique de l’Innovation et de l’Exportation du Québec. MO is supported by grants from the Canadian Institutes of Health Research and holds a Canada Research Chair on Antimicrobial Resistance.
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Leishmania are dimorphic parasites living as extracellular promastigotes in the digestive tract of Phlebotomus or Lutzomyia sandflies and as intracellular amastigotes within phagocytic cells (mainly macrophages) of the vertebrate hosts. Leishmania species cause leishmaniasis, the second largest parasite killer; there are 1.3 million new cases annually, 12 million people are affected worldwide, and 350 million people are currently at risk1. The Leishmania genus includes several species, among which more than 20 are pathogenic to humans2. Leishmania can be divided into two subgenera: the Leishmania Leishmania subgenus responsible for visceral and cutaneous leishmaniasis and the Leishmania Viannia subgenus often associated with either cutaneous or muco-cutaneous forms of the disease2. Visceral leishmaniasis is mainly caused by L. donovani and L. infantum and is characterized by fever, hepatosplenomegaly, and pancytopenia3, making it the most severe and deadly form of the disease when compared with the self-healing but nonetheless debilitating skin lesions of cutaneous leishmaniasis4,5.
No effective human vaccine is currently available against Leishmania (a canine Leishmania vaccine was recently registered in Europe6), and control measures mainly involve chemotherapy7–9. Pentavalent antimony (Sb) has been the standard drug for 70 years and remains the mainstay in many endemic regions, apart from Northern India, where antimonial formulations have been rendered obsolete because of widespread parasite resistance. Other first-line therapies include the polyene antibiotic amphotericin B (AMB) for which a single dose was shown to be 95% effective against visceral leishmaniasis in India10. Liposomal AMB has become a standard treatment in many countries11 but requires administration by the intravenous route. Geographical differences in the response to liposomal AMB were reported, and visceral leishmaniasis cases in India were more responsive than those from East Africa or South America12. Clinical AMB resistance is scarce and parasites remained susceptible even after multiple rounds of treatment in the same patient13. The alkyl-lysophospholipid analogue miltefosine (MTF) is the only oral drug against Leishmania14,15. It has been successfully used for the treatment of visceral leishmaniasis since its registration in 2002 in India16. However, relapse rates are on the rise in India17,18 and Nepal19,20, making MTF resistance a likely problem. The aminoglycoside paromomycin (PMM) is also approved for the treatment of visceral leishmaniasis in India21,22. So far, the scarce use of PMM has limited the emergence of resistance, but geographical variations in PMM efficacy against visceral leishmaniasis were noted between East Africa (especially Sudan) and India23,24. Lastly, pentamidine (PTD) has been abandoned for the treatment of visceral leishmaniasis because of serious toxicities and is mainly restricted to patients with cutaneous leishmaniasis in South America25–27.
Despite six decades of use, the mode of action (MOA) of antimonials is not known. It has been shown to lead to the production of reactive oxygen species28–31, the depletion of trypanothione32, and apoptosis-like death33–36, but an exact MOA is still awaited. The same applies for MTF, AMB, PTD, and PMM, with the possible exception of AMB, which kills Leishmania by forming pores in ergosterol-containing membranes. New molecules with well-defined drug targets are clearly needed.
Leishmania and its genome
The Leishmania genome is around 32 Mb and displays over 8,300 coding genes37,38. Within the Leishmania genus, gene synteny is conserved for more than 99% of genes between L. major, L. infantum, and L. braziliensis, and only few species-specific genes were found38. Leishmania species have between 34 and 36 chromosomes ranging in size from 0.3 to 2.8 Mb37–41. One unique feature characterizing trypanosomatid parasites lies in their genome architecture, their protein-coding genes being organized as large polycistronic units42,43. In the absence of defined RNA polymerase II promoters, transcription of the long polycistronic units occurs in a bidirectional fashion from transcriptional start sites located at strand switch regions43,44. Processing into individual messenger RNAs (mRNAs) occurs by the addition through trans-splicing of a spliced leader RNA (39 nt) to the 5′ ends of each mRNA, coupled to 3′ end polyadenylation45,46. Because of its lack of transcriptional control, Leishmania uses several adaptive mechanisms to regulate gene expression when facing changing environmental conditions during its development. 3′ untranslated regions (3′ UTRs) were shown to be major players in monitoring mRNA stability and translation rates in this parasite47–53. To overcome stressful conditions like drug pressure, Leishmania also often relies on DNA copy number variations (CNVs) (aneuploidy, gene amplification, or gene deletion) for regulating the expression of drug targets, drug transporters, or other determinants of resistance. This is not restricted to Leishmania, however, and variations in gene dosage or chromosome copy numbers also influence drug susceptibility, adaptability, and proliferation in fungi and cancer cells54–57. In addition to CNVs, single-nucleotide polymorphisms (SNPs) in drug targets or in transporters can lead to drug resistance without the need for altering gene expression.
Copy number variations
During the last few decades, Leishmania parasites were considered to be essentially diploid but recent data have shown that aneuploidy seems to be the norm58–64. Within populations of Leishmania parasites, distinct aneuploidy patterns were shown to occur at the level of individual cells. This phenomenon was called mosaic aneuploidy and can translate into a seemingly average diploid population when the cumulative ploidy is derived from next-generation DNA sequencing data but in which few parasites actually share the same ploidy for individual chromosomes62,63,65. Interestingly, variations in the size and content of chromosomes have also been observed between different strains of the related trypanosomatid parasite Trypanosoma cruzi66,67. In the case of Leishmania, circumstantial links between the presence of supernumerary chromosomes or chromosomal losses and drug resistance have been observed58–60,64,68–71, suggesting that a particular group of genes on the variant chromosomes may possibly act together in establishing resistance, but this has yet to be demonstrated.
Aneuploidy is generally linked to developmental abnormalities as best exemplified by the trisomy 21 syndrome in humans. However, Leishmania uses aneuploidy as a lifestyle. This is raising a number of questions about aneuploidy generation, stability, transmission, and biological significance (reviewed elsewhere60,62,72). In the absence of transcription initiation control, increases (or decreases) in chromosome copy number may serve as a strategy for regulating expression under environmental cues. This can happen at the level of whole chromosomes, and indeed there was a good correlation between chromosome ploidy and the level of DNA and RNA expression59. Increasing the copy numbers of entire chromosomes may lead to the overexpression of toxic genes, but the Leishmania genome (32 Mb) is spread in 34 to 36 chromosomes, thus reducing the co-expression of many genes. However, as explained in detail below, Leishmania also has the ability to amplify (or delete) specific smaller regions of DNA as part of extrachromosomal elements by recombination/rearrangements at the level of homologous repeated sequences (RSs). RNA levels derived from these amplifications are correlated to DNA copy number.
The genome of Leishmania is populated with repeated DNA sequences. A recent study highlighted the entire set of RSs in different Leishmania species, and it was found that the whole Leishmania genome has the potential to be rearranged at the level of those RSs for generating extrachromosomal elements73. Indeed, almost 2,000 RSs are distributed over the genome of L. infantum and these potentially support the formation of more than 3,000 extrachromosomal DNA elements73. Short interspersed degenerate retroposons (SIDERs) account for up to 65% of all RSs. SIDERs are truncated versions (~0.55 kb) of formerly active retroposons that are predominantly located in 3′ UTRs and have been associated with post-transcriptional regulation at the levels of both mRNA stability and translation47,48,52,53,74,75. Because SIDERs are degenerated, they were found in different RS groups. Remarkably, SIDER elements would have dual roles: one functional by regulating gene expression and a second one structural, providing the backbone to facilitate gene rearrangements for changing copy number of chromosomal DNA regions.
Extrachromosomal DNA amplifications are frequently detected in Leishmania parasites challenged with drugs or other stressful conditions58,59,64,76–87. The episomes are amplified as either circular or linear extrachromosomal DNA and formed through rearrangements at the level of direct or inverted homologous RSs, respectively (Figure 1)73,80,88. Interestingly, between 60% and 80% of the predicted amplicons appear to be already present in the population in the absence of selection and these pre-existing stochastic gene amplifications were shown to foster the selection of adaptive traits in response to drug pressure73. Beneficial amplicons were shown to increase in abundance upon higher drug pressure and to decrease when the drug is removed, allowing parasites to respond to a changing environment73.
Figure 1. Potential mechanisms for gene amplification in Leishmania.
(a) The RAD51 recombinase mediates homologous recombination between direct repeated sequences (DRS) and leads to (i) extrachromosomal circular amplicon or (ii) intrachromosomal tandem duplication by unequal sister chromatid exchange or RAD51-mediated break-induced replication. Black arrows represent DRS. (b) The MRE11 nuclease processes DNA ends after single-strand break (SSB), double-strand break (DSB), or hairpin formation during replication and leads to extrachromosomal linear amplification. Black arrows represent inverted repeated sequences (IRS). The green segments represent the amplified DNA regions. dsDNA, double-stranded DNA; ssDNA, single-stranded DNA.
Since gene rearrangements through RSs are primary responses to drug pressure, a reasonable hypothesis was that identifying recombinase proteins involved in these rearrangements could lead to strategies to prevent the emergence of resistance. A first candidate was the RAD51 DNA repair protein, a key protein involved in homologous recombination (HR), a mechanism evolutionarily conserved in trypanosomatids89. Interestingly, the expression of RAD51 was induced in Leishmania by DNA double-strand breaks (DSBs)90,91. Inactivating RAD51 led to viable parasites unable to generate circular extrachromosomal elements but still capable of producing linear amplicons upon drug pressure73. Leishmania has three RAD51 paralogs (RAD51-3, RAD51-4, and RAD51-6) that were shown to work as a complex in promoting HR through their capacity to stimulate RAD51 activity92. Inactivation of RAD51-4 was also shown to prevent the formation of circular amplicons in L. infantum exposed to drugs, but not of linear amplicons92.
Linear amplicons are formed by the annealing of RSs found in an inverted orientation64,73,93. MRE11 is a DNA repair nuclease that interacts with RAD50 and NBS1 to form the MRN complex94,95 and is important for DSB repair by HR96,97 or for non-homologous end joining89 (Figure 1b). Inactivation of MRE11 impaired the ability of L. infantum to form linear amplicons upon drug selection at the level of inverted RSs, although the capacity to generate circular amplicons was similar to that of wild-type parasites93. Moreover, a fully functional MRE11 is important for linear amplification, as parasites expressing DNA-binding-proficient but nuclease-deficient MRE11 exclusively generated circular amplicons during drug selection93. Interestingly, inactivation of MRE11 alone or along with its partner RAD50 led to extensive chromosomal translocation in L. infantum98, showing that the MRE11/RAD50 complex is important for the maintenance of genome integrity in addition to its role in gene rearrangements. The number of enzymes involved in the formation of extrachromosomal elements suggests that targeting this pathway may not be a viable strategy for preventing the emergence of resistance, although this remains to be experimentally tested.
Single-nucleotide polymorphisms and small nucleotide insertions or deletions
Although CNVs are important contributors of drug resistance, SNPs and small nucleotide insertions or deletions (indels) can also contribute to resistance. This was proven with experimental drugs99,100 and was highlighted with MTF where amino acid substitutions or non-sense mutations were observed in the MTF transporter (MT)101 or in its Ros3 subunit102. This was further confirmed in additional mutants using whole genome sequencing103,104 or by deep sequencing of MT105. Mutations detected in the MT gene of L. infantum isolates serially collected from an MTF-treated patient who had multiple relapses were shown to correlate with resistance106, suggesting that MTF resistance could become a clinical reality in the near future.
Genome-wide surveys of genetic variations in L. donovani isolates from the Indian subcontinent supported the notion that resistance to antimonials emerged on several distinct occasions58,107. Isolates could also be clustered on the basis of their genetic structure and haplotypes, with some groups being enriched for non-responsive strains58,107,108. Interestingly, a particular group of highly resistant isolates that clustered together were found to share genomic features associated with resistance107. Among these were a higher copy number for the H-locus, coding for the well-characterized ABC transporter MRPA109, and a homozygous two-base-pair insertion in the aquaglyceroporin 1 (AQP1) gene involved in Sb uptake and whose inactivation or downregulation is strongly correlated with resistance71,107,110–117. The potential for these genomic variants in predicting treatment outcome is exciting, given the lack of molecular markers for Sb resistance, but will need to be thoroughly evaluated by using larger and geographically diversified sets of well-defined isolates.
Exploiting copy number variations for understanding drug mode of action and resistance mechanisms
Target-based assays and phenotypic whole-cell-based assays are the cornerstones of drug discovery. The current trend for anti-parasitic agents is for whole cell assays. A drawback of phenotype-based assays is the lack of knowledge about the targets of hit compounds. Although the molecules could be brought to the clinic without further knowledge about their MOA, a clear understanding of the molecular targets will facilitate the improvement of a candidate drug through lead optimization. Characterization of drug-resistant mutants, which often revealed mutations or CNVs in drug targets or in proteins responsible for drug transport, is one strategy to pinpoint drug targets. However, it is salient to point out that this strategy has not yet led to targets against the current anti-leishmanials, although amplification of gene targets was observed in mutants made resistant to experimental drugs64,99. Since CNVs are often associated with resistance, forward genetic tools can experimentally mimic this. One such gain-of-function screen was based on functional cloning where Leishmania cosmid libraries were electroporated into Leishmania and these transfectants were selected for a specific phenotype118. Selection is possible because of the high copy number (and gene expression) of the cosmids. This screen was successfully applied while selecting for drug resistance or susceptibility101,114,119–123. This technique selects for cosmids conferring dominant phenotypes (leaving out less enriched cosmids) and is not easily amenable to high-throughput screening. The sensitivity of cosmid-based functional screening was enhanced by its recent coupling to next-generation sequencing in an approach termed Cos-Seq124. The proportion of parasites with cosmids providing a selective advantage is expected to rise with increasing drug pressure, and these can be tracked and quantified at each drug increment by Illumina sequencing124,125. Thus, the dynamics of cosmid enrichment can be followed over the entire course of selection instead of being monitored only at endpoint. Published or ongoing Cos-Seq screens using experimental drugs with known targets (for example, methotrexate, terbinafine, and 5-fluorouracil) confirmed the recovery of the relevant target genes by Cos-Seq124. Interestingly, Cos-Seq supported the hypothesis that the current anti-leishmanials (MF, AMB, Sb, PMM, and PTD) may not act via specific major protein targets124. Indeed, although an unprecedented number of resistance genes (known and novel) were isolated using Cos-Seq, none emerged as a clear target candidate and it is conceivable that these antiquated drugs are broadly cytotoxic by disrupting multiple minor targets. Whether some of the genes are genuine drug targets remains to be established, and non-protein targets represent another possibility as these would not be detected by Cos-Seq.
The advent of high-content screening for intracellular L. donovani amastigotes126 is also key in the search for novel molecules having favourable anti-leishmanial properties directly on the intracellular stage of the parasite. This allowed the discovery of 192 new leads against visceral leishmaniasis from an initial set of 1.8 million compounds from GlaxoSmithKline127. An MOA could be hypothesized for 80 of the lead compounds using prior proprietary knowledge and bioinformatics analyses of TriTryp genomes, which revealed an over-representation of putative kinase inhibitors127. Cos-Seq was initially carried out with the insect form of the parasite but this could easily be adapted to intracellular parasites and this technique could be used to find the targets of these promising novel molecules or of other drugs repurposed against Leishmania128.
The Cos-Seq technique does not allow the isolation of loss-of-function mutations such as those found in the aquaglyceroporin AQP1 or in the MT transporter genes (see above). These require high-throughput dominant negative screening approaches, like inducible RNA interference target sequencing (RIT-Seq), which proved instrumental in elucidating mechanisms of drug uptake in trypanosomes129. Although RNA interference is absent from the L. Leishmania subgenus, it is active in species of the L. Viannia subgenus130. The lack of inducible expression in Leishmania was also a limitation of this technique, but two recent reports have shown the feasibility of inducible expression in Leishmania131,132. Thus, it is theoretically possible to develop a technology similar to RIT-Seq in L. Viannia parasites. An alternative approach to RIT-Seq would be to rely on RNA-guided nuclease systems using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) enzymes, as these have proven very efficient for achieving targeted genomic modifications in a wide range of genomes133–135. In trypanosomatid parasites, the CRISPR/Cas9 system derived from Streptococcus pyogenes has been used for disrupting genes in L. major136, L. donovani137, and T. cruzi138 and in principle could be used for generating whole-genome Cas9-mediated gene deletion libraries.
Concluding remarks
The toolkit for drug target discovery and resistance mechanism elucidation for Leishmania is expanding. With new promising drug candidates in the pipeline and further technological developments, it should now be possible to find new targets which should further help in the control of this important neglected tropical disease.
The authors declare that they have no competing interests.
Grant information
BP is supported by grants from the Canadian Institutes of Health Research, the Natural Sciences and Engineering Research Council of Canada, and the Ministère du Developpement Économique de l’Innovation et de l’Exportation du Québec. MO is supported by grants from the Canadian Institutes of Health Research and holds a Canada Research Chair on Antimicrobial Resistance.
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
2.
Bañuls AL, Hide M, Prugnolle F:
Leishmania and the leishmaniases: a parasite genetic update and advances in taxonomy, epidemiology and pathogenicity in humans.
Adv Parasitol.
2007; 64: 1–109. PubMed Abstract
| Publisher Full Text
3.
Guerin PJ, Olliaro P, Sundar S, et al.:
Visceral leishmaniasis: current status of control, diagnosis, and treatment, and a proposed research and development agenda.
Lancet Infect Dis.
2002; 2(8): 494–501. PubMed Abstract
| Publisher Full Text
11.
Bern C, Adler-Moore J, Berenguer J, et al.:
Liposomal amphotericin B for the treatment of visceral leishmaniasis.
Clin Infect Dis.
2006; 43(7): 917–24. PubMed Abstract
| Publisher Full Text
12.
Berman JD, Badaro R, Thakur CP, et al.:
Efficacy and safety of liposomal amphotericin B (AmBisome) for visceral leishmaniasis in endemic developing countries.
Bull World Health Organ.
1998; 76(1): 25–32. PubMed Abstract
| Free Full Text
13.
Lachaud L, Bourgeois N, Plourde M, et al.:
Parasite susceptibility to amphotericin B in failures of treatment for visceral leishmaniasis in patients coinfected with HIV type 1 and Leishmania infantum.
Clin Infect Dis.
2009; 48(2): e16–22. PubMed Abstract
| Publisher Full Text
14.
Croft SL, Neal RA, Pendergast W, et al.:
The activity of alkyl phosphorylcholines and related derivatives against Leishmania donovani.
Biochem Pharmacol.
1987; 36(16): 2633–6. PubMed Abstract
| Publisher Full Text
15.
Jha TK, Sundar S, Thakur CP, et al.:
Miltefosine, an oral agent, for the treatment of Indian visceral leishmaniasis.
N Engl J Med.
1999; 341(24): 1795–800. PubMed Abstract
| Publisher Full Text
16.
Sundar S, Jha TK, Thakur CP, et al.:
Oral miltefosine for Indian visceral leishmaniasis.
N Engl J Med.
2002; 347(22): 1739–46. PubMed Abstract
| Publisher Full Text
17.
Burza S, Nabi E, Mahajan R, et al.:
One-year follow-up of immunocompetent male patients treated with miltefosine for primary visceral leishmaniasis in Bihar, India.
Clin Infect Dis.
2013; 57(9): 1363–4. PubMed Abstract
| Publisher Full Text
| Faculty Opinions Recommendation
19.
Pandey BD, Pandey K, Kaneko O, et al.:
Relapse of visceral leishmaniasis after miltefosine treatment in a Nepalese patient.
Am J Trop Med Hyg.
2009; 80(4): 580–2. PubMed Abstract
20.
Rijal S, Ostyn B, Uranw S, et al.:
Increasing failure of miltefosine in the treatment of Kala-azar in Nepal and the potential role of parasite drug resistance, reinfection, or noncompliance.
Clin Infect Dis.
2013; 56(11): 1530–8. PubMed Abstract
| Publisher Full Text
| Faculty Opinions Recommendation
21.
Jha TK, Olliaro P, Thakur CP, et al.:
Randomised controlled trial of aminosidine (paromomycin) v sodium stibogluconate for treating visceral leishmaniasis in North Bihar, India.
BMJ.
1998; 316(7139): 1200–5. PubMed Abstract
| Publisher Full Text
| Free Full Text
22.
Sundar S, Jha TK, Thakur CP, et al.:
Injectable paromomycin for Visceral leishmaniasis in India.
N Engl J Med.
2007; 356(25): 2571–81. PubMed Abstract
| Publisher Full Text
23.
Hailu A, Musa A, Wasunna M, et al.:
Geographical variation in the response of visceral leishmaniasis to paromomycin in East Africa: a multicentre, open-label, randomized trial.
PLoS Negl Trop Dis.
2010; 4(10): e709. PubMed Abstract
| Publisher Full Text
| Free Full Text
24.
Musa AM, Younis B, Fadlalla A, et al.:
Paromomycin for the treatment of visceral leishmaniasis in Sudan: a randomized, open-label, dose-finding study.
PLoS Negl Trop Dis.
2010; 4(10): e855. PubMed Abstract
| Publisher Full Text
| Free Full Text
25.
Lai A Fat EJ, Vrede MA, Soetosenojo RM, et al.:
Pentamidine, the drug of choice for the treatment of cutaneous leishmaniasis in Surinam.
Int J Dermatol.
2002; 41(11): 796–800. PubMed Abstract
| Publisher Full Text
26.
Roussel M, Nacher M, Frémont G, et al.:
Comparison between one and two injections of pentamidine isethionate, at 7 mg/kg in each injection, in the treatment of cutaneous leishmaniasis in French Guiana.
Ann Trop Med Parasitol.
2006; 100(4): 307–14. PubMed Abstract
| Publisher Full Text
27.
Soto J, Buffet P, Grogl M, et al.:
Successful treatment of Colombian cutaneous leishmaniasis with four injections of pentamidine.
Am J Trop Med Hyg.
1994; 50(1): 107–11. PubMed Abstract
28.
Mandal G, Wyllie S, Singh N, et al.:
Increased levels of thiols protect antimony unresponsive Leishmania donovani field isolates against reactive oxygen species generated by trivalent antimony.
Parasitology.
2007; 134(Pt 12): 1679–87. PubMed Abstract
| Publisher Full Text
| Free Full Text
29.
Mehta A, Shaha C:
Mechanism of metalloid-induced death in Leishmania spp.: role of iron, reactive oxygen species, Ca2+, and glutathione.
Free Radic Biol Med.
2006; 40(10): 1857–68. PubMed Abstract
| Publisher Full Text
30.
Mookerjee Basu J, Mookerjee A, Sen P, et al.:
Sodium antimony gluconate induces generation of reactive oxygen species and nitric oxide via phosphoinositide 3-kinase and mitogen-activated protein kinase activation in Leishmania donovani-infected macrophages.
Antimicrob Agents Chemother.
2006; 50(5): 1788–97. PubMed Abstract
| Publisher Full Text
| Free Full Text
31.
Moreira W, Leprohon P, Ouellette M:
Tolerance to drug-induced cell death favours the acquisition of multidrug resistance in Leishmania.
Cell Death Dis.
2011; 2: e201. PubMed Abstract
| Publisher Full Text
| Free Full Text
32.
Wyllie S, Cunningham ML, Fairlamb AH:
Dual action of antimonial drugs on thiol redox metabolism in the human pathogen Leishmania donovani.
J Biol Chem.
2004; 279(38): 39925–32. PubMed Abstract
| Publisher Full Text
33.
Lee N, Bertholet S, Debrabant A, et al.:
Programmed cell death in the unicellular protozoan parasite Leishmania.
Cell Death Differ.
2002; 9(1): 53–64. PubMed Abstract
| Publisher Full Text
34.
Sereno D, Holzmuller P, Mangot I, et al.:
Antimonial-mediated DNA fragmentation in Leishmania infantum amastigotes.
Antimicrob Agents Chemother.
2001; 45(7): 2064–9. PubMed Abstract
| Publisher Full Text
| Free Full Text
35.
Sudhandiran G, Shaha C:
Antimonial-induced increase in intracellular Ca2+ through non-selective cation channels in the host and the parasite is responsible for apoptosis of intracellular Leishmania donovani amastigotes.
J Biol Chem.
2003; 278(27): 25120–32. PubMed Abstract
| Publisher Full Text
36.
Vergnes B, Gourbal B, Girard I, et al.:
A proteomics screen implicates HSP83 and a small kinetoplastid calpain-related protein in drug resistance in Leishmania donovani clinical field isolates by modulating drug-induced programmed cell death.
Mol Cell Proteomics.
2007; 6(1): 88–101. PubMed Abstract
| Publisher Full Text
39.
Britto C, Ravel C, Bastien P, et al.:
Conserved linkage groups associated with large-scale chromosomal rearrangements between Old World and New World Leishmania genomes.
Gene.
1998; 222(1): 107–17. PubMed Abstract
| Publisher Full Text
40.
Raymond F, Boisvert S, Roy G, et al.:
Genome sequencing of the lizard parasite Leishmania tarentolae reveals loss of genes associated to the intracellular stage of human pathogenic species.
Nucleic Acids Res.
2012; 40(3): 1131–47. PubMed Abstract
| Publisher Full Text
| Free Full Text
41.
Wincker P, Ravel C, Blaineau C, et al.:
The Leishmania genome comprises 36 chromosomes conserved across widely divergent human pathogenic species.
Nucleic Acids Res.
1996; 24(9): 1688–94. PubMed Abstract
| Publisher Full Text
| Free Full Text
42.
El-Sayed NM, Myler PJ, Blandin G, et al.:
Comparative genomics of trypanosomatid parasitic protozoa.
Science.
2005; 309(5733): 404–9. PubMed Abstract
| Publisher Full Text
43.
Martinez-Calvillo S, Yan S, Nguyen D, et al.:
Transcription of Leishmania major Friedlin chromosome 1 initiates in both directions within a single region.
Mol Cell.
2003; 11(5): 1291–9. PubMed Abstract
| Publisher Full Text
44.
Martinez-Calvillo S, Nguyen D, Stuart K, et al.:
Transcription initiation and termination on Leishmania major chromosome 3.
Eukaryotic Cell.
2004; 3(2): 506–17. PubMed Abstract
| Publisher Full Text
| Free Full Text
45.
Haile S, Papadopoulou B:
Developmental regulation of gene expression in trypanosomatid parasitic protozoa.
Curr Opin Microbiol.
2007; 10(6): 569–77. PubMed Abstract
| Publisher Full Text
46.
Matthews KR, Tschudi C, Ullu E:
A common pyrimidine-rich motif governs trans-splicing and polyadenylation of tubulin polycistronic pre-mRNA in trypanosomes.
Genes Dev.
1994; 8(4): 491–501. PubMed Abstract
| Publisher Full Text
47.
Boucher N, Wu Y, Dumas C, et al.:
A common mechanism of stage-regulated gene expression in Leishmania mediated by a conserved 3'-untranslated region element.
J Biol Chem.
2002; 277(22): 19511–20. PubMed Abstract
| Publisher Full Text
49.
Dupe A, Dumas C, Papadopoulou B:
An Alba-domain protein contributes to the stage-regulated stability of amastin transcripts in Leishmania.
Mol Microbiol.
2014; 91(3): 548–61. PubMed Abstract
| Publisher Full Text
50.
Haile S, Dupe A, Papadopoulou B:
Deadenylation-independent stage-specific mRNA degradation in Leishmania.
Nucleic Acids Res.
2008; 36(5): 1634–44. PubMed Abstract
| Publisher Full Text
| Free Full Text
51.
McNicoll F, Muller M, Cloutier S, et al.:
Distinct 3'-untranslated region elements regulate stage-specific mRNA accumulation and translation in Leishmania.
J Biol Chem.
2005; 280(42): 35238–46. PubMed Abstract
| Publisher Full Text
52.
Muller M, Padmanabhan PK, Papadopoulou B:
Selective inactivation of SIDER2 retroposon-mediated mRNA decay contributes to stage- and species-specific gene expression in Leishmania.
Mol Microbiol.
2010; 77(2): 471–91. PubMed Abstract
| Publisher Full Text
53.
Muller M, Padmanabhan PK, Rochette A, et al.:
Rapid decay of unstable Leishmania mRNAs bearing a conserved retroposon signature 3'-UTR motif is initiated by a site-specific endonucleolytic cleavage without prior deadenylation.
Nucleic Acids Res.
2010; 38(17): 5867–83. PubMed Abstract
| Publisher Full Text
| Free Full Text
54.
Gordon DJ, Resio B, Pellman D:
Causes and consequences of aneuploidy in cancer.
Nat Rev Genet.
2012; 13(3): 189–203. PubMed Abstract
| Publisher Full Text
56.
Rutledge SD, Cimini D:
Consequences of aneuploidy in sickness and in health.
Curr Opin Cell Biol.
2016; 40: 41–6. PubMed Abstract
| Publisher Full Text
59.
Leprohon P, Legare D, Raymond F, et al.:
Gene expression modulation is associated with gene amplification, supernumerary chromosomes and chromosome loss in antimony-resistant Leishmania infantum.
Nucleic Acids Res.
2009; 37(5): 1387–99. PubMed Abstract
| Publisher Full Text
| Free Full Text
60.
Mannaert A, Downing T, Imamura H, et al.:
Adaptive mechanisms in pathogens: universal aneuploidy in Leishmania.
Trends Parasitol.
2012; 28(9): 370–6. PubMed Abstract
| Publisher Full Text
62.
Sterkers Y, Lachaud L, Bourgeois N, et al.:
Novel insights into genome plasticity in Eukaryotes: mosaic aneuploidy in Leishmania.Mol Microbiol.
2012; 86(1): 15–23. PubMed Abstract
| Publisher Full Text
63.
Sterkers Y, Lachaud L, Crobu L, et al.:
FISH analysis reveals aneuploidy and continual generation of chromosomal mosaicism in Leishmania major.
Cell Microbiol.
2011; 13(2): 274–83. PubMed Abstract
| Publisher Full Text
64.
Ubeda JM, Légaré D, Raymond F, et al.:
Modulation of gene expression in drug resistant Leishmania is associated with gene amplification, gene deletion and chromosome aneuploidy.
Genome Biol.
2008; 9(7): R115. PubMed Abstract
| Publisher Full Text
| Free Full Text
66.
Minning TA, Weatherly DB, Flibotte S, et al.:
Widespread, focal copy number variations (CNV) and whole chromosome aneuploidies in Trypanosoma cruzi strains revealed by array comparative genomic hybridization.
BMC Genomics.
2011; 12: 139. PubMed Abstract
| Publisher Full Text
| Free Full Text
68.
Brotherton MC, Bourassa S, Leprohon P, et al.:
Proteomic and genomic analyses of antimony resistant Leishmania infantum mutant.
PLoS One.
2013; 8(11): e81899. PubMed Abstract
| Publisher Full Text
| Free Full Text
69.
do Monte-Neto RL, Coelho AC, Raymond F, et al.:
Gene expression profiling and molecular characterization of antimony resistance in Leishmania amazonensis.PLoS Negl Trop Dis.
2011; 5(5): e1167. PubMed Abstract
| Publisher Full Text
| Free Full Text
70.
Kumar P, Lodge R, Raymond F, et al.:
Gene expression modulation and the molecular mechanisms involved in Nelfinavir resistance in Leishmania donovani axenic amastigotes.
Mol Microbiol.
2013; 89(3): 565–82. PubMed Abstract
| Publisher Full Text
71.
Mukherjee A, Boisvert S, Monte-Neto RL, et al.:
Telomeric gene deletion and intrachromosomal amplification in antimony-resistant Leishmania.
Mol Microbiol.
2013; 88(1): 189–202. PubMed Abstract
| Publisher Full Text
73.
Ubeda JM, Raymond F, Mukherjee A, et al.:
Genome-wide stochastic adaptive DNA amplification at direct and inverted DNA repeats in the parasite Leishmania.
PLoS Biol.
2014; 12(5): e1001868. PubMed Abstract
| Publisher Full Text
| Free Full Text
75.
Smith M, Bringaud F, Papadopoulou B:
Organization and evolution of two SIDER retroposon subfamilies and their impact on the Leishmania genome.
BMC Genomics.
2009; 10: 240. PubMed Abstract
| Publisher Full Text
| Free Full Text
77.
Beverley SM, Coderre JA, Santi DV, et al.:
Unstable DNA amplifications in methotrexate-resistant Leishmania consist of extrachromosomal circles which relocalize during stabilization.
Cell.
1984; 38(2): 431–9. PubMed Abstract
| Publisher Full Text
78.
Garvey EP, Santi DV:
Stable amplified DNA in drug-resistant Leishmania exists as extrachromosomal circles.
Science.
1986; 233(4763): 535–40. PubMed Abstract
| Publisher Full Text
79.
Grondin K, Haimeur A, Mukhopadhyay R, et al.:
Co-amplification of the gamma-glutamylcysteine synthetase gene gsh1 and of the ABC transporter gene pgpA in arsenite-resistant Leishmania tarentolae.
EMBO J.
1997; 16(11): 3057–65. PubMed Abstract
| Publisher Full Text
| Free Full Text
80.
Grondin K, Papadopoulou B, Ouellette M:
Homologous recombination between direct repeat sequences yields P-glycoprotein containing amplicons in arsenite resistant Leishmania.Nucleic Acids Res.
1993; 21(8): 1895–901. PubMed Abstract
| Publisher Full Text
| Free Full Text
81.
Haimeur A, Brochu C, Genest P, et al.:
Amplification of the ABC transporter gene PGPA and increased trypanothione levels in potassium antimonyl tartrate (SbIII) resistant Leishmania tarentolae.
Mol Biochem Parasitol.
2000; 108(1): 131–5. PubMed Abstract
| Publisher Full Text
82.
Kundig C, Leblanc E, Papadopoulou B, et al.:
Role of the locus and of the resistance gene on gene amplification frequency in methotrexate resistant Leishmania tarentolae.
Nucleic Acids Res.
1999; 27(18): 3653–9. PubMed Abstract
| Publisher Full Text
| Free Full Text
83.
Mittal MK, Rai S, Ashutosh, et al.:
Characterization of natural antimony resistance in Leishmania donovani isolates.
Am J Trop Med Hyg.
2007; 76(4): 681–8. PubMed Abstract
84.
Mukherjee A, Padmanabhan PK, Singh S, et al.:
Role of ABC transporter MRPA, gamma-glutamylcysteine synthetase and ornithine decarboxylase in natural antimony-resistant isolates of Leishmania donovani.
J Antimicrob Chemother.
2007; 59(2): 204–11. PubMed Abstract
| Publisher Full Text
85.
Ouellette M, Hettema E, Wüst D, et al.:
Direct and inverted DNA repeats associated with P-glycoprotein gene amplification in drug resistant Leishmania.
EMBO J.
1991; 10(4): 1009–16. PubMed Abstract
| Free Full Text
86.
Papadopoulou B, Roy G, Ouellette M:
Frequent amplification of a short chain dehydrogenase gene as part of circular and linear amplicons in methotrexate resistant Leishmania.
Nucleic Acids Res.
1993; 21(18): 4305–12. PubMed Abstract
| Free Full Text
87.
White TC, Fase-Fowler F, van Luenen H, et al.:
The H circles of Leishmania tarentolae are a unique amplifiable system of oligomeric DNAs associated with drug resistance.
J Biol Chem.
1988; 263(32): 16977–83. PubMed Abstract
88.
Navarro M, Liu J, Muthui D, et al.:
Inverted repeat structure and homologous sequences in the LD1 amplicons of Leishmania spp.
Mol Biochem Parasitol.
1994; 68(1): 69–80. PubMed Abstract
| Publisher Full Text
89.
Genois MM, Paquet ER, Laffitte MC, et al.:
DNA repair pathways in trypanosomatids: from DNA repair to drug resistance.
Microbiol Mol Biol Rev.
2014; 78(1): 40–73. PubMed Abstract
| Publisher Full Text
| Free Full Text
90.
Genois MM, Mukherjee A, Ubeda JM, et al.:
Interactions between BRCA2 and RAD51 for promoting homologous recombination in Leishmania infantum.
Nucleic Acids Res.
2012; 40(14): 6570–84. PubMed Abstract
| Publisher Full Text
| Free Full Text
91.
McKean PG, Keen JK, Smith DF, et al.:
Identification and characterisation of a RAD51 gene from Leishmania major.
Mol Biochem Parasitol.
2001; 115(2): 209–16. PubMed Abstract
| Publisher Full Text
92.
Genois MM, Plourde M, Éthier C, et al.:
Roles of Rad51 paralogs for promoting homologous recombination in Leishmania infantum.
Nucleic Acids Res.
2015; 43(5): 2701–15. PubMed Abstract
| Publisher Full Text
| Free Full Text
93.
Laffitte MC, Genois MM, Mukherjee A, et al.:
Formation of linear amplicons with inverted duplications in Leishmania requires the MRE11 nuclease.
PLoS Genet.
2014; 10(12): e1004805. PubMed Abstract
| Publisher Full Text
| Free Full Text
98.
Laffitte MC, Leprohon P, Hainse M, et al.:
Chromosomal Translocations in the Parasite Leishmania by a MRE11/RAD50-Independent Microhomology-Mediated End Joining Mechanism.
PLoS Genet.
2016; 12(6): e1006117. PubMed Abstract
| Publisher Full Text
| Free Full Text
99.
Ritt JF, Raymond F, Leprohon P, et al.:
Gene amplification and point mutations in pyrimidine metabolic genes in 5-fluorouracil resistant Leishmania infantum.
PLoS Negl Trop Dis.
2013; 7(11): e2564. PubMed Abstract
| Publisher Full Text
| Free Full Text
100.
Vasudevan G, Ullman B, Landfear SM:
Point mutations in a nucleoside transporter gene from Leishmania donovani confer drug resistance and alter substrate selectivity.
Proc Natl Acad Sci USA.
2001; 98(11): 6092–7. PubMed Abstract
| Publisher Full Text
| Free Full Text
101.
Pérez-Victoria FJ, Gamarro F, Ouellette M, et al.:
Functional cloning of the miltefosine transporter. A novel P-type phospholipid translocase from Leishmania involved in drug resistance.
J Biol Chem.
2003; 278(50): 49965–71. PubMed Abstract
| Publisher Full Text
102.
Pérez-Victoria FJ, Sánchez-Cañete MP, Castanys S, et al.:
Phospholipid translocation and miltefosine potency require both L. donovani miltefosine transporter and the new protein LdRos3 in Leishmania parasites.
J Biol Chem.
2006; 281(33): 23766–75. PubMed Abstract
| Publisher Full Text
103.
Coelho AC, Boisvert S, Mukherjee A, et al.:
Multiple mutations in heterogeneous miltefosine-resistant Leishmania major population as determined by whole genome sequencing.
PLoS Negl Trop Dis.
2012; 6(2): e1512. PubMed Abstract
| Publisher Full Text
| Free Full Text
104.
Mondelaers A, Sanchez-Cañete MP, Hendrickx S, et al.:
Genomic and Molecular Characterization of Miltefosine Resistance in Leishmania infantum Strains with Either Natural or Acquired Resistance through Experimental Selection of Intracellular Amastigotes.
PLoS One.
2016; 11(4): e0154101. PubMed Abstract
| Publisher Full Text
| Free Full Text
| Faculty Opinions Recommendation
105.
Laffitte MN, Leprohon P, Légaré D, et al.:
Deep-sequencing revealing mutation dynamics in the miltefosine transporter gene in Leishmania infantum selected for miltefosine resistance.
Parasitol Res.
2016; 1–5. PubMed Abstract
| Publisher Full Text
106.
Cojean S, Houzé S, Haouchine D, et al.:
Leishmania resistance to miltefosine associated with genetic marker.
Emerging Infect Dis.
2012; 18(4): 704–6. PubMed Abstract
| Publisher Full Text
| Free Full Text
108.
Vanaerschot M, Decuypere S, Downing T, et al.:
Genetic markers for SSG resistance in Leishmania donovani and SSG treatment failure in visceral leishmaniasis patients of the Indian subcontinent.
J Infect Dis.
2012; 206(5): 752–5. PubMed Abstract
| Publisher Full Text
| Free Full Text
109.
Légaré D, Richard D, Mukhopadhyay R, et al.:
The Leishmania ATP-binding cassette protein PGPA is an intracellular metal-thiol transporter ATPase.
J Biol Chem.
2001; 276(28): 26301–7. PubMed Abstract
| Publisher Full Text
110.
Decuypere S, Rijal S, Yardley V, et al.:
Gene expression analysis of the mechanism of natural Sb(V) resistance in Leishmania donovani isolates from Nepal.
Antimicrob Agents Chemother.
2005; 49(11): 4616–21. PubMed Abstract
| Publisher Full Text
| Free Full Text
111.
Gourbal B, Sonuc N, Bhattacharjee H, et al.:
Drug uptake and modulation of drug resistance in Leishmania by an aquaglyceroporin.
J Biol Chem.
2004; 279(30): 31010–7. PubMed Abstract
| Publisher Full Text
112.
Mandal G, Mandal S, Sharma M, et al.:
Species-specific antimonial sensitivity in Leishmania is driven by post-transcriptional regulation of AQP1.
PLoS Negl Trop Dis.
2015; 9(2): e0003500. PubMed Abstract
| Publisher Full Text
| Free Full Text
113.
Mandal S, Maharjan M, Singh S, et al.:
Assessing aquaglyceroporin gene status and expression profile in antimony-susceptible and -resistant clinical isolates of Leishmania donovani from India.
J Antimicrob Chemother.
2010; 65(3): 496–507. PubMed Abstract
| Publisher Full Text
114.
Marquis N, Gourbal B, Rosen BP, et al.:
Modulation in aquaglyceroporin AQP1 gene transcript levels in drug-resistant Leishmania.Mol Microbiol.
2005; 57(6): 1690–9. PubMed Abstract
| Publisher Full Text
115.
Monte-Neto R, Laffitte MC, Leprohon P, et al.:
Intrachromosomal amplification, locus deletion and point mutation in the aquaglyceroporin AQP1 gene in antimony resistant Leishmania (Viannia) guyanensis.
PLoS Negl Trop Dis.
2015; 9: e0003476. PubMed Abstract
| Publisher Full Text
| Free Full Text
116.
Plourde M, Ubeda JM, Mandal G, et al.:
Generation of an aquaglyceroporin AQP1 null mutant in Leishmania major.
Mol Biochem Parasitol.
2015; 201(2): 108–11. PubMed Abstract
| Publisher Full Text
117.
Uzcategui NL, Zhou Y, Figarella K, et al.:
Alteration in glycerol and metalloid permeability by a single mutation in the extracellular C-loop of Leishmania major aquaglyceroporin LmAQP1.
Mol Microbiol.
2008; 70(6): 1477–86. PubMed Abstract
| Publisher Full Text
| Free Full Text
118.
Ryan KA, Garraway LA, Descoteaux A, et al.:
Isolation of virulence genes directing surface glycosyl-phosphatidylinositol synthesis by functional complementation of Leishmania.
Proc Natl Acad Sci USA.
1993; 90(18): 8609–13. PubMed Abstract
| Publisher Full Text
| Free Full Text
119.
Carter NS, Drew ME, Sanchez M, et al.:
Cloning of a novel inosine-guanosine transporter gene from Leishmania donovani by functional rescue of a transport-deficient mutant.
J Biol Chem.
2000; 275(27): 20935–41. PubMed Abstract
| Publisher Full Text
120.
Coelho AC, Beverley SM, Cotrim PC:
Functional genetic identification of PRP1, an ABC transporter superfamily member conferring pentamidine resistance in Leishmania major.
Mol Biochem Parasitol.
2003; 130(2): 83–90. PubMed Abstract
| Publisher Full Text
121.
Cotrim PC, Garrity LK, Beverley SM:
Isolation of genes mediating resistance to inhibitors of nucleoside and ergosterol metabolism in Leishmania by overexpression/selection.
J Biol Chem.
1999; 274(53): 37723–30. PubMed Abstract
| Publisher Full Text
122.
Kundig C, Haimeur A, Legare D, et al.:
Increased transport of pteridines compensates for mutations in the high affinity folate transporter and contributes to methotrexate resistance in the protozoan parasite Leishmania tarentolae.
EMBO J.
1999; 18(9): 2342–51. PubMed Abstract
| Publisher Full Text
| Free Full Text
123.
Vasudevan G, Carter NS, Drew ME, et al.:
Cloning of Leishmania nucleoside transporter genes by rescue of a transport-deficient mutant.
Proc Natl Acad Sci U S A.
1998; 95(17): 9873–8. PubMed Abstract
| Publisher Full Text
| Free Full Text
124.
Gazanion E, Fernandez-Prada C, Papadopoulou B, et al.:
Cos-Seq for high-throughput identification of drug target and resistance mechanisms in the protozoan parasite Leishmania.
Proc Natl Acad Sci U S A.
2016; 113(21): E3012–21. PubMed Abstract
| Publisher Full Text
| Free Full Text
131.
Duncan SM, Myburgh E, Philipon C, et al.:
Conditional gene deletion with DiCre demonstrates an essential role for CRK3 in Leishmania mexicana cell cycle regulation.
Mol Microbiol.
2016; 100(6): 931–44. PubMed Abstract
| Publisher Full Text
| Free Full Text
134.
Selle K, Barrangou R:
Harnessing CRISPR-Cas systems for bacterial genome editing.
Trends Microbiol.
2015; 23(4): 225–32. PubMed Abstract
| Publisher Full Text
135.
Wright AV, Nuñez JK, Doudna JA:
Biology and Applications of CRISPR Systems: Harnessing Nature's Toolbox for Genome Engineering.
Cell.
2016; 164(1–2): 29–44. PubMed Abstract
| Publisher Full Text
1
Centre de Recherche en Infectiologie du Centre de Recherche du CHU Québec, and Département de Microbiologie, Infectiologie et Immunologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada
BP is supported by grants from the Canadian Institutes of Health Research, the Natural Sciences and Engineering Research Council of Canada, and the Ministère du Developpement Économique de l’Innovation et de l’Exportation du Québec. MO is supported by grants from the Canadian Institutes of Health Research and holds a Canada Research Chair on Antimicrobial Resistance.
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Laffitte MCN, Leprohon P, Papadopoulou B and Ouellette M. Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved] F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.12688/f1000research.9218.1)
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Spaeth G. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16438)
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Reviewer Report20 Sep 2016
Gerald Spaeth, Unité de Parasitologie moléculaire et Signalisation, Department of Parasites and Insect Vectors, Institut Pasteur, INSERM U1201, Paris, France
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Spaeth G. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16438)
Sterkers Y. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16439)
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Sterkers Y. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16439)
McCulloch R. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16440)
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Reviewer Report20 Sep 2016
Richard McCulloch, The Wellcome Trust Centre for Molecular Parasitology, Institute of Infection, Immunity and Inflammation, University of Glasgow, Glasgow, UK
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Faculty Reviews are commissioned and written by members of the prestigious Faculty Opinions Faculty, and are edited as a service to our readers. In order to make these reviews as comprehensive and accessible as possible, we seek the reviewers’ input before publication. The reviewers’ names and any additional comments they may have are published alongside the review, as is usual on F1000Research.
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McCulloch R. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16440)
Clos J. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16441)
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Faculty Reviews are commissioned and written by members of the prestigious Faculty Opinions Faculty, and are edited as a service to our readers. In order to make these reviews as comprehensive and accessible as possible, we seek the reviewers’ input before publication. The reviewers’ names and any additional comments they may have are published alongside the review, as is usual on F1000Research.
I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
Clos J. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16441)
Gamarro F. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16442)
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Competing Interests: No competing interests were disclosed.
Faculty Reviews are commissioned and written by members of the prestigious Faculty Opinions Faculty, and are edited as a service to our readers. In order to make these reviews as comprehensive and accessible as possible, we seek the reviewers’ input before publication. The reviewers’ names and any additional comments they may have are published alongside the review, as is usual on F1000Research.
I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
Gamarro F. Reviewer Report For: Plasticity of the Leishmania genome leading to gene copy number variations and drug resistance [version 1; peer review: 5 approved]. F1000Research 2016, 5(F1000 Faculty Rev):2350 (https://doi.org/10.5256/f1000research.9921.r16442)
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