Occurrence of a Novel Strain of Moroccan Watermelon Mosaic Virus Infecting Pumpkins in Kenya.

The Potyvirus Moroccan watermelon mosaic virus (MWMV) naturally infects and severely threatens production of cucurbits and papaya. In this study, we identified and characterized MWMV isolated from pumpkin (Cucurbita moschata) intercropped with MWMV-infected papaya plants through next-generation sequencing (NGS) and Sanger sequencing approaches. Complete MWMV genome sequences were obtained from two pumpkin samples through NGS and validated using Sanger sequencing. The isolates shared 83.4 to 83.7% nucleotide (nt) and 92.3 to 95.1% amino acid (aa) sequence identities in the coat protein and 79.5 to 79.9% nt and 89.2 to 89.7% aa identities in the polyprotein with papaya isolates of MWMV. Phylogenetic analysis using complete polyprotein nt sequences revealed the clustering of both pumpkin isolates of MWMV with corresponding sequences of cucurbit isolates of the virus from other parts of Africa and the Mediterranean regions, distinct from a clade formed by papaya isolates. Through sap inoculation, a pumpkin isolate of MWMV was pathogenic on zucchini (Cucurbita pepo), watermelon (Citrullus lanatus), and cucumber (Cucumis sativus) but not on papaya. Conversely, the papaya isolate of MWMV was nonpathogenic on pumpkin, watermelon, and cucumber, but it infected zucchini. The results suggest the occurrence of two strains of MWMV in Kenya having different biological characteristics associated with the host specificity.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Metagenomic Analysis of Plant Viruses Associated With Papaya Ringspot Disease in Carica papaya L. in Kenya.

Carica papaya L. is an important fruit crop grown by small- and large-scale farmers in Kenya for local and export markets. However, its production is constrained by papaya ringspot disease (PRSD). The disease is believed to be caused by papaya ringspot virus (PRSV). Previous attempts to detect PRSV in papaya plants showing PRSD symptoms, using enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase-polymerase chain reaction (RT-PCR) procedures with primers specific to PRSV, have not yielded conclusive results. Therefore, the nature of viruses responsible for PRSD was elucidated in papaya leaves collected from 22 counties through Illumina MiSeq next-generation sequencing (NGS) and validated by RT-PCR and Sanger sequencing. Viruses were detected in 38 out of the 48 leaf samples sequenced. Sequence analysis revealed the presence of four viruses: a Potyvirus named Moroccan watermelon mosaic virus (MWMV) and three viruses belonging to the genus Carlavirus. The Carlaviruses include cowpea mild mottle virus (CpMMV) and two putative Carlaviruses-closely related but distinct from cucumber vein-clearing virus (CuVCV) with amino acid and nucleotide sequence identities of 75.7-78.1 and 63.6-67.6%, respectively, in the coat protein genes. In reference to typical symptoms observed in the infected plants, the two putative Carlaviruses were named papaya mottle-associated virus (PaMV) and papaya mild mottle-associated virus (PaMMV). Surprisingly, and in contrast to previous studies conducted in other parts of world, PRSV was not detected. The majority of the viruses were detected as single viral infections, while a few were found to be infecting alongside another virus (for example, MWMV and PaMV). Furthermore, the NGS and RT-PCR analysis identified MWMV as being strongly associated with ringspot symptoms in infected papaya fruits. This study has provided the first complete genome sequences of these viruses isolated from papaya in Kenya, together with primers for their detection-thus proving to be an important step towards the design of long-term, sustainable disease management strategies.

Comparing sequence and structure of falcipains and human homologs at prodomain and catalytic active site for malarial peptide based inhibitor design.

BACKGROUND: Falcipains are major cysteine proteases of Plasmodium falciparum involved in haemoglobin degradation and remain attractive anti-malarial drug targets. Several inhibitors against these proteases have been identified, yet none of them has been approved for malaria treatment. Other Plasmodium species also possess highly homologous proteins to falcipains. For selective therapeutic targeting, identification of sequence and structure differences with homologous human cathepsins is necessary. The substrate processing activity of these proteins is tightly controlled via a prodomain segment occluding the active site which is chopped under low pH conditions exposing the catalytic site. Current work characterizes these proteases to identify residues mediating the prodomain regulatory function for the design of peptide based anti-malarial inhibitors. METHODS: Sequence and structure variations between prodomain regions of plasmodial proteins and human cathepsins were determined using in silico approaches. Additionally, evolutionary clustering of these proteins was evaluated using phylogenetic analysis. High quality partial zymogen protein structures were modelled using homology modelling and residue interaction analysis performed between the prodomain segment and mature domain to identify key interacting residues between these two domains. The resulting information was used to determine short peptide sequences which could mimic the inherent regulatory function of the prodomain regions. Through flexible docking, the binding affinity of proposed peptides on the proteins studied was evaluated. RESULTS: Sequence, evolutionary and motif analyses showed important differences between plasmodial and human proteins. Residue interaction analysis identified important residues crucial for maintaining prodomain integrity across the different proteins as well as the pro-segment responsible for inhibitory mechanism. Binding affinity of suggested peptides was highly dependent on their residue composition and length. CONCLUSIONS: Despite the conserved structural and catalytic mechanism between human cathepsins and plasmodial proteases, current work revealed significant differences between the two protein groups which may provide valuable information for selective anti-malarial inhibitor development. Part of this study aimed to design peptide inhibitors based on endogenous inhibitory portions of protease prodomains as a novel aspect. Even though peptide inhibitors may not be practical solutions to malaria at this stage, the approach followed and results offer a promising means to find new malarial inhibitors.

Metagenomic analysis of viruses associated with maize lethal necrosis in Kenya.

BACKGROUND: Maize lethal necrosis is caused by a synergistic co-infection of Maize chlorotic mottle virus (MCMV) and a specific member of the Potyviridae, such as Sugarcane mosaic virus (SCMV), Wheat streak mosaic virus (WSMV) or Johnson grass mosaic virus (JGMV). Typical maize lethal necrosis symptoms include severe yellowing and leaf drying from the edges. In Kenya, we detected plants showing typical and atypical symptoms. Both groups of plants often tested negative for SCMV by ELISA. METHODS: We used next-generation sequencing to identify viruses associated to maize lethal necrosis in Kenya through a metagenomics analysis. Symptomatic and asymptomatic leaf samples were collected from maize and sorghum representing sixteen counties. RESULTS: Complete and partial genomes were assembled for MCMV, SCMV, Maize streak virus (MSV) and Maize yellow dwarf virus-RMV (MYDV-RMV). These four viruses (MCMV, SCMV, MSV and MYDV-RMV) were found together in 30 of 68 samples. A geographic analysis showed that these viruses are widely distributed in Kenya. Phylogenetic analyses of nucleotide sequences showed that MCMV, MYDV-RMV and MSV are similar to isolates from East Africa and other parts of the world. Single nucleotide polymorphism, nucleotide and polyprotein sequence alignments identified three genetically distinct groups of SCMV in Kenya. Variation mapped to sequences at the border of NIb and the coat protein. Partial genome sequences were obtained for other four potyviruses and one polerovirus. CONCLUSION: Our results uncover the complexity of the maize lethal necrosis epidemic in Kenya. MCMV, SCMV, MSV and MYDV-RMV are widely distributed and infect both maize and sorghum. SCMV population in Kenya is diverse and consists of numerous strains that are genetically different to isolates from other parts of the world. Several potyviruses, and possibly poleroviruses, are also involved.