Structure based docking and molecular dynamic studies of plasmodial cysteine proteases against a South African natural compound and its analogs:

Identification of potential drug targets as well as development of novel antimalarial chemotherapies with unique mode of actions due to drug resistance by Plasmodium parasites are inevitable. Falcipains (falcipain-2 and falcipain-3) of Plasmodium falciparum, which catalyse the haemoglobin degradation process, are validated drug targets. Previous attempts to develop peptide based drugs against these enzymes have been futile due to the poor pharmacological profiles and susceptibility to degradation by host enzymes. This study aimed to identify potential non-peptide inhibitors against falcipains and their homologs from other Plasmodium species

Analysis of non-peptidic compounds as potential malarial inhibitors against Plasmodial cysteine proteases via integrated virtual screening workflow

Falcipain-2 (FP-2) and falcipain-3 (FP-3), haemoglobin-degrading enzymes in Plasmodium falciparum, are validated drug targets for the development of effective inhibitors against malaria. However, no commercial drug-targeting falcipains has been developed despite their central role in the life cycle of the parasites. In this work, in silico approaches are used to identify key structural elements that control the binding and selectivity of a diverse set of non-peptidic compounds onto FP-2, FP-3 and homologues from other Plasmodium species as well as human cathepsins. Hotspot residues and the underlying non-covalent interactions, important for the binding of ligands, are identified by interaction fingerprint analysis between the proteases and 2-cyanopyridine derivatives (best hits). It is observed that the size and chemical type of substituent groups within 2-cyanopyridine derivatives determine the strength of protein–ligand interactions. This research presents novel results that can further be exploited in the structure-based molecular-guided design of more potent antimalarial drugs.http://www.tandfonline.com/loi/tbsd20hb2017Forestry and Agricultural Biotechnology Institute (FABI)Genetic

Structure based docking and molecular dynamic studies of plasmodial cysteine proteases against a South African natural compound and its analogs

Identification of potential drug targets as well as development of novel antimalarial chemotherapies with unique mode of actions due to drug resistance by Plasmodium parasites are inevitable. Falcipains (falcipain-2 and falcipain-3) of Plasmodium falciparum, which catalyse the haemoglobin degradation process, are validated drug targets. Previous attempts to develop peptide based drugs against these enzymes have been futile due to the poor pharmacological profiles and susceptibility to degradation by host enzymes. This study aimed to identify potential non-peptide inhibitors against falcipains and their homologs from other Plasmodium species. Structure based virtual docking approach was used to screen a small non-peptidic library of natural compounds from South Africa against 11 proteins. A potential hit, 5α-Pregna-1,20-dien-3-one (5PGA), with inhibitory activity against plasmodial proteases and selectivity on human cathepsins was identified. A 3D similarity search on the ZINC database using 5PGA identified five potential hits based on their docking energies. The key interacting residues of proteins with compounds were identified via molecular dynamics and free binding energy calculations. Overall, this study provides a basis for further chemical design for more effective derivatives of these compounds. Interestingly, as these compounds have cholesterol-like nuclei, they and their derivatives might be well tolerated in humans.The National Institutes of Health Common Fund under grant number U41HG006941 to H3ABioNet; the National Research Foundation (NRF), South Africa [grant numbers 79765].http://www.nature.com/srepam2016Forestry and Agricultural Biotechnology Institute (FABI)Genetic

Analysis of non-peptidic compounds as potential malarial inhibitors against plasmodial cysteine proteases via integrated virtual screening workflow

Malaria is an infectious disease caused by a diverse group of erythrocytic protozoan parasites of the genus Plasmodium. It remains an exigent public health problem in the tropical areas of Africa, South America and parts of Asia and continues to take its toll in morbidity and mortality with half of the world’s population under a permanent risk of infection leading to more than half a million deaths annually (WHO, 2013). Five Plasmodium species, namely P. falciparum (Pf ), P. vivax (Pv), P. ovale (Po), P. malariae (Pm) and P. knowlesi (Pk), are known to infect humans with Pf responsible for more than 90% of the malarial fatalities reported in sub-Saharan Africa. The predominance of Pf is attributed to its adaptability (Ashley, McGready, Proux, & Nosten, 2006; Prugnolle et al., 2011). Although the high occurrence of the Duffy negative trait among African populations lowers the threat posed by Pv, it is the most frequent and widely causative agent of benign tertian malaria in other parts of the world (Mendis, Sina, Marchesini, & Carter, 2001). In addition to the listed human malarial parasite forms, several other Plasmodium species, which infect non-human laboratory models, have been identified and are of significant importance in understanding the parasite biology, the host–parasite interactions and in the drug development process (Langhorne et al., 2011)

Intron derived size polymorphism in the mitochondrial genomes of closely related chrysoporthe species

In this study, the complete mitochondrial (mt) genomes of Chrysoporthe austroafricana (190,834 bp), C. cubensis (89,084 bp) and C. deuterocubensis (124,412 bp) were determined. Additionally, the mitochondrial genome of another member of the Cryphonectriaceae, namely Cryphonectria parasitica (158,902 bp), was retrieved and annotated for comparative purposes. These genomes showed high levels of synteny, especially in regions including genes involved in oxidative phosphorylation and electron transfer, unique open reading frames (uORFs), ribosomal RNAs (rRNAs) and transfer RNAs (tRNAs), as well as intron positions. Comparative analyses revealed signatures of duplication events, intron number and length variation, and varying intronic ORFs which highlighted the genetic diversity of mt genomes among the Cryphonectriaceae. These mt genomes showed remarkable size polymorphism. The size polymorphism in the mt genomes of these closely related Chrysoporthe species was attributed to the varying number and length of introns, coding sequences and to a lesser extent, intergenic sequences. Compared to publicly available fungal mt genomes, the C. austroafricana mt genome is the second largest in the Ascomycetes thus far.S1 Fig. Midrooted phylogenetic tree of rnpb genes. Rnpb genes were mannualy retrieved from annotated fungal mt genomes publicly available in GenBank. Phylogenetic analysis was performed using maximum likelihood method implemented in RAxML with HKY model of selection. Branch support was calculated using 1000 bootstrap replicates. Green boxes depict atp6 gene, red; small sub-unit of ribosomal RNA (rns) and dark red; small sub-unit of ribosomal RNA (rnl). (PDF)S2 Fig. Maximum likelihood phylogeny of Rps3. Phylogenetic analysis of orf540, orf551 and orf551 from C. austroafricana, C. cubensis and C. deuterocubensis which show sequence similarity to C. parasitica S5 ribosomal protein/maturase fusion protein. Sequences used in this phylogeny were retrieved from GenBank using BLAST. The LG+G model of substitution was used. Branch support for was calculated using the bootstrap method with 1000 replicates. (PDF)S3 Fig. RPKM values for 14 OXPHOS genes. The graph shows the average expression for genes that are involved in oxidative phosphorylation and electron transport and the rnpb gene of Chrysoporthe austroafricana grown in complete and minimal media. (PDF)S4 Fig. Physical maps of the mt genomes of C. austroafricana, C. cubensis, C. deuterocubensis and C. parasitica showing locations of all intronic and intergenic ORFs. Intronic ORFs are depicted by yellow arrowed boxes on introns of genes where found and are labelled according to the gene and intron position. Intergenic ORFs are also depicted by yellow arrowed boxes and are labelled with “IN” prefix. (PDF)S1 Table. Accession number and species names for sequences used for rps3 gene phylogeny. (PDF)S2 Table. Codon usage analysis. Comparison of codon usage and tRNAs for the 14 genes involved in oxidative phosphorylation and electron transport in the mitochondrial genomes of Chrysoporthe austroafricana, C. cubensis, C. deuterocubensis and Cryphonectria parasitica. (PDF)S3 Table. BLAST analysis of all identified introns against mt genome sequences deposited in NCBI GenBank. The best hits for each query (intron) is shown. Default BLAST parameters were used. The prefix CA, CC, CD and CP is added to the intron names for clarity. (PDF)S4 Table. BLAST analysis of identified intron encoded and free standing HEG protein sequences. Results from all possible comparisons were calculated. Only Blast hits with a threshold of over 50% alignment coverage, 50% sequence identity and E-value of 0.00005 are shown. (PDF)S5 Table. BLAST analysis of intronic and free standing HEG protein sequences against mt genomes deposited in NCBI GenBank database. HEG type were annotated using the NCBI Conserved Domain Database (CDD). (PDF)The Genomics Research Institute (GRI) University of Pretoria, the University of Pretoria Research Development Programme, the DST/NRF Centre of Excellence in Tree Health Biotechnology (FABI, University of Pretoria), and the National Research Foundation (NRF) (grant number 87332).http://www.plosone.orgam2016Forestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

Intron derived size polymorphism in the mitochondrial genomes of closely related chrysoporthe species

In this study, the complete mitochondrial (mt) genomes of Chrysoporthe austroafricana (190,834 bp), C. cubensis (89,084 bp) and C. deuterocubensis (124,412 bp) were determined. Additionally, the mitochondrial genome of another member of the Cryphonectriaceae, namely Cryphonectria parasitica (158,902 bp), was retrieved and annotated for comparative purposes. These genomes showed high levels of synteny, especially in regions including genes involved in oxidative phosphorylation and electron transfer, unique open reading frames (uORFs), ribosomal RNAs (rRNAs) and transfer RNAs (tRNAs), as well as intron positions. Comparative analyses revealed signatures of duplication events, intron number and length variation, and varying intronic ORFs which highlighted the genetic diversity of mt genomes among the Cryphonectriaceae. These mt genomes showed remarkable size polymorphism. The size polymorphism in the mt genomes of these closely related Chrysoporthe species was attributed to the varying number and length of introns, coding sequences and to a lesser extent, intergenic sequences. Compared to publicly available fungal mt genomes, the C. austroafricana mt genome is the second largest in the Ascomycetes thus far.S1 Fig. Midrooted phylogenetic tree of rnpb genes. Rnpb genes were mannualy retrieved from annotated fungal mt genomes publicly available in GenBank. Phylogenetic analysis was performed using maximum likelihood method implemented in RAxML with HKY model of selection. Branch support was calculated using 1000 bootstrap replicates. Green boxes depict atp6 gene, red; small sub-unit of ribosomal RNA (rns) and dark red; small sub-unit of ribosomal RNA (rnl). (PDF)S2 Fig. Maximum likelihood phylogeny of Rps3. Phylogenetic analysis of orf540, orf551 and orf551 from C. austroafricana, C. cubensis and C. deuterocubensis which show sequence similarity to C. parasitica S5 ribosomal protein/maturase fusion protein. Sequences used in this phylogeny were retrieved from GenBank using BLAST. The LG+G model of substitution was used. Branch support for was calculated using the bootstrap method with 1000 replicates. (PDF)S3 Fig. RPKM values for 14 OXPHOS genes. The graph shows the average expression for genes that are involved in oxidative phosphorylation and electron transport and the rnpb gene of Chrysoporthe austroafricana grown in complete and minimal media. (PDF)S4 Fig. Physical maps of the mt genomes of C. austroafricana, C. cubensis, C. deuterocubensis and C. parasitica showing locations of all intronic and intergenic ORFs. Intronic ORFs are depicted by yellow arrowed boxes on introns of genes where found and are labelled according to the gene and intron position. Intergenic ORFs are also depicted by yellow arrowed boxes and are labelled with “IN” prefix. (PDF)S1 Table. Accession number and species names for sequences used for rps3 gene phylogeny. (PDF)S2 Table. Codon usage analysis. Comparison of codon usage and tRNAs for the 14 genes involved in oxidative phosphorylation and electron transport in the mitochondrial genomes of Chrysoporthe austroafricana, C. cubensis, C. deuterocubensis and Cryphonectria parasitica. (PDF)S3 Table. BLAST analysis of all identified introns against mt genome sequences deposited in NCBI GenBank. The best hits for each query (intron) is shown. Default BLAST parameters were used. The prefix CA, CC, CD and CP is added to the intron names for clarity. (PDF)S4 Table. BLAST analysis of identified intron encoded and free standing HEG protein sequences. Results from all possible comparisons were calculated. Only Blast hits with a threshold of over 50% alignment coverage, 50% sequence identity and E-value of 0.00005 are shown. (PDF)S5 Table. BLAST analysis of intronic and free standing HEG protein sequences against mt genomes deposited in NCBI GenBank database. HEG type were annotated using the NCBI Conserved Domain Database (CDD). (PDF)The Genomics Research Institute (GRI) University of Pretoria, the University of Pretoria Research Development Programme, the DST/NRF Centre of Excellence in Tree Health Biotechnology (FABI, University of Pretoria), and the National Research Foundation (NRF) (grant number 87332).http://www.plosone.orgam2016Forestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

Genetic variability in populations of Chrysoporthe cubensis and Chr. puriensis in Brazil

DATA AVAILABILITY STATEMENT: The data that support the findings will be available in NCBI Nucleotide at https://www.re3data.org/search?query=ncbi. The Genbank numbers are given in the text of the manuscript. The data is under embargo until a manuscript is accepted for publication. It will then be openly available.Chrysoporthe puriensis, a sibling species of the well-known Eucalyptus canker pathogen Chr. cubensis, has recently been described from Brazil. Both species are thought to be native to South America, but previous population genetic analyses were conducted prior to the ready availability of robust markers such as microsatellites to test this hypothesis. The objective of this investigation was to analyse the structure and genetic variability of Chr. cubensis and Chr. puriensis populations from non-native Myrtaceae and native Melastomataceae hosts in Brazil, using microsatellite markers developed for this purpose. The fungal isolates were obtained from Eucalyptus species, Corymbia citriodora and Tibouchina species in different regions of the country. Isolates were separated into sub-populations based on host families (Melastomataceae and Myrtaceae) and on region of origin. There was high genetic variability in all sub-populations with the highest levels detected within, rather than among sub-populations. Gene and genotypic diversities were higher for the isolates from the Melastomataceae than the Myrtaceae isolates. High levels of gene flow were found between sub-populations based on host and geographic distribution of the sub-populations. The presence of genetically diverse Chr. cubensis and Chr. puriensis populations on native hosts in Brazil supports a Latin American centre of origin for the two pathogens. Both undergo sexual and asexual reproduction and have a high potential for gene flow.The Tree Protection Co-operative Programme (TPCP) and the DST/NRF Centre of Excellence in Tree Health Biotechnology (South Africa). The FAPEMIG (Fundação de Amparo à Pesquisa do Estado de Minas Gerais), CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior), and CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) for providing scholarships.https://link.springer.com/journal/13313hj2023BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

Chrysoporthe puriensis sp. nov. from Tibouchina spp. in Brazil : an emerging threat to Eucalyptus

The discovery of Cryphonectriaceae and more specifically species related to the Eucalyptus canker pathogen Chrysoporthe cubensis on shrubs and trees in the Melastomataceae, has deepened our understanding of relevant, and potentially globally threatening tree pathogens. Recent isolations of Cryphonectriaceae associated with cankers on Tibouchina spp. in Brazil gave rise to an apparently undescribed species of Chrysoporthe associated with stem and branch cankers that lead to tree death. Cultures of this fungus were subjected to phylogenetic studies based on sequences for the ITS and β-tubulin gene regions. These analyses revealed a novel taxon that is described here as Chrysoporthe puriensis sp. nov., having both sexual and asexual states. Pathogenicity tests on two species of Tibouchina (T. granulosa, T. heteromalla) and hybrids of Eucalyptus grandis x E. urophylla showed that Chr. puriensis can infect and cause disease on all of these trees. It is clearly not only damaging on native Tibouchina spp. where environmental conditions are conducive to disease development, but also potentially threatening to non-native Eucalyptus spp., which form the basis of a major plantation forest industry.http://link.springer.com/journal/13313hj2022BiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

Intra-species genomic variation in the pine pathogen Fusarium circinatum

Fusarium circinatum is an important global pathogen of pine trees. Genome plasticity has been observed in different isolates of the fungus, but no genome comparisons are available. To address this gap, we sequenced and assembled to chromosome level five isolates of F. circinatum. These genomes were analysed together with previously published genomes of F. circinatum isolates, FSP34 and KS17. Multi-sample variant calling identified a total of 461,683 micro variants (SNPs and small indels) and a total of 1828 macro structural variants of which 1717 were copy number variants and 111 were inversions. The variant density was higher on the sub-telomeric regions of chromosomes. Variant annotation revealed that genes involved in transcription, transport, metabolism and transmembrane proteins were overrepresented in gene sets that were affected by high impact variants. A core genome representing genomic elements that were conserved in all the isolates and a non-redundant pangenome representing all genomic elements is presented. Whole genome alignments showed that an average of 93% of the genomic elements were present in all isolates. The results of this study reveal that some genomic elements are not conserved within the isolates and some variants are high impact. The described genome-scale variations will help to inform novel disease management strategies against the pathogen.DATA AVAILABILTY STATEMENT : The Whole Genome Shotgun project for Fusarium circinatum CMWF1803 has been deposited at DDBJ/ENA/GenBank under the accession JAEHFH000000000. The version described in this paper is version JAEHFH010000000. The Whole Genome Shotgun project for Fusarium circinatum CMWF560 has been deposited at DDBJ/ENA/GenBank under the accession JAEHFI000000000. The version described in this paper is version JAEHFI010000000. The Whole Genome Shotgun project for Fusarium circinatum CMWF567 has been deposited at DDBJ/ENA/GenBank under the accession JADZLS000000000. The version described in this paper is version JADZLS010000000. The Whole Genome Shotgun project for Fusarium circinatum UG27 has been deposited at DDBJ/ENA/ GenBank under the accession JAELVK000000000. The version described in this paper is version JAELVK010000000. The Whole Genome Shotgun project for Fusarium circinatum UG10 has been deposited at DDBJ/ENA/GenBank under the accession JAGJRQ000000000. The version described in this paper is version JAGJRQ010000000.The South African Department of Science and Innovation’s South African Research Chair Initiative and the DSI-NRF Centre of Excellence in Plant Health Biotechnology at the Forestry and Agricultural Biotechnology Institute (FABI), University of Pretoria.http://www.mdpi.com/journal/jofBiochemistryForestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog