Morphological and Molecular Diversity of Ginger (Zingiber officinale Roscoe) Pathogenic Fungi in Chilga District, North Gondar, Ethiopia

Ginger diseases caused by fungal pathogens have become one of the most serious problems causing reduced production around the world. It has also caused a major problem among farmers in different parts of Ethiopia resulting in a huge decline in rhizome yield. However, the exact causative agents of this disease have not been identified in the state. Although there are few studies related to pathogenic fungus identification, molecular level identification of fungal pathogen was not done in the area. Therefore, this study was undertaken to isolate and characterized the fungal causative agent of ginger disease from the diseased plant and the soil samples collected around the diseased plant from Chilga district, Gondar, Ethiopia. Samples from infected ginger plants and the soil around the infected plant were collected. Culturing and purification of isolates were made using Potato Dextrose Agar supplemented with antibacterial agent chloramphenicol. The morphological characterization was done by structural identification of the isolates under the microscope using lactophenol cotton blue stains. Isolated fungi were cultured and molecular identification was done using an internal transcribed spacer (ITS) of ribosomal DNA (rDNA). A total of 15 fungal morphotypes including 11 Aspergillus spp. (73.3%), 2 Penicillium spp. (13.3%), and single uncultured fungus clone S23 were isolated from the samples representing all the plant organs and the soil. Aspergillus spp. (73.3%) was the most common and seems to be the major causative agent. To the best of our knowledge, this is the first report of ginger pathogenic fungi in Ethiopia identified using ITS rDNA molecular techniques. This study will lay foundation for the development of management strategies for fungal diseases infecting ginger.

N6-Methyladenosine Regulatory Machinery is Involved In Viral Infection of Plum Pox Virus And Potato Virus Y In Nicotiana Benthamiana

Abstract N6-methyladenosine (m6A) is a post-transcriptional modification of biological mRNA and non-coding RNAs, which by regulating the mRNA stability and translation. It has been demonstrated that m6A methylation has a regulatory effect on human RNA virus replication. In this project, Plum pox virus (PPV) and Potato Y virus (PVY) were used to examine the m6A modification in Nicotiana benthamiana during natural infection. The results showed that the global level of m6A in both PVY and PPV infected plants were significantly decreased than non-infected plants. Particularly, the PPV and PVY infection could alter the m6A level of the host endogenous gene. This is suggesting that plant viruses may disrupt the balance of the m6A in plant. Meanwhile, we found that viral genome RNA can be targeted by m6A methylation. Two m6A-enrich regions in PPV genome RNA and four in PVY genome RNA were detected, which are located in the coding region of viruses. Based on the ALKB and METTL sequences in the transcriptome sequencing data of the virus-infected plant, we cloned 2 NbALKB genes and 2 NbMETTL genes in N. benthamiana . According to results of transient expression and VIGS assay, NbALKB gene appears slightly contributing PPV and PVY infection. NbMETTL gene showed certain inhibition effect in PPV infection, but not PVY. Therefore, our data suggested that m6A methylation in plant might be an anti-viral strategy in some plant viruses.

Mapping of Plant-Parasitic Nematode Using GIS Technology in Tainal Watershed, Sulaimaniyah Province, Iraq

The present experiment was conducted to evaluate soil samples which had been taken from depths ranging from 0-30 cm in greenhouses planted with different vegetables such as cucumbers, tomatoes, peppers and eggplants in 24 agricultural sites in 19 villages. GIS application was used to make maps showing the results of a field survey that was conducted. The experiment was conducted for an area of 12 km2 in the Tainal Plain, west of Sulaimaniyah Province. Nearly 30% of the soil of the greenhouses were heavily infected. Plant analysis showed that the characteristics of ash, fat and moisture were significantly affected as a result of infection with nematodes. The spread of root-knot nematode juveniles in the villages of Al-Mahmoudia, Kani Pengsharma and Kani Shaya was 706, 695 and 622 per 250 g of soil. In tissues of the studied plants, the highest nitrogen content was 68.2 mg in Ziyeka, 60.4 mg in Gawani and 59.7 mg in Ali Bzaw. Phosphorus concentrations were highest in Kani Shaya, Shwankara and Ziyeka (25.5, 25.3 and 23.1 mg, respectively). Warmizyar had 91.8 mg of potassium, Kani Shaya had 78.6 mg and Kani Big had 71.6 mg. The calcium concentrations in Ziyeka, Gawani and Mewk were 54.5, 48.4 and 47.7 mg, respectively.

Enset Production, Its Challenges and Controlling Methods in South Omo Zone, Southern Ethiopia

Enset (Ensete ventricosum (Welw.) Cheesman) is economically a useful crop that provides forage for animals, construction materials, fuel and traditionally it is used as a medicine. The production and productivity of enset is mainly affected by many diseases and pests which causes serious devastating cultivar diversity and finally a yield lose of the crop. Moreover, its production sustainability is also threatened by many factors like wild animal pests, enset root pests, high population pressure and the shift of the producers to cash-oriented crop production. The assessment on enset production, its challenges and controlling methods was carried out in Debub Ari district of South Omo Zone, in southern Ethiopia from November 2021 to January 2021 in four randomly selected representative Kebeles. Simple random sampling method were employed to select a representative sample of the study (n = 138). Qualitative and quantitative data were collected through structured questionnaire, focus group discussions, individual interview and personal observations. The collected data were analyzed by using appropriate simple descriptive statistics. The land under enset production is declining from time to time, as well as its production also declines due to social, bio-physical and environmental factors such as land fragmentation, enset bacterial welt, insects and pests and environmental variability factors. To overcome these factors farmers practicing many controlling methods such as by burying the infected plants, restricting the movement of infected plant materials, sterilizing production tools and raising awareness about how to protect disease like enset bacterial welt and use of aluminum phosphate for pests like mole rate.

Ustilago maydis Secreted Endo-Xylanases Are Involved in Fungal Filamentation and Proliferation on and Inside Plants

Plant pathogenic fungi must be able to degrade host cell walls in order to penetrate and invade plant tissues. Among the plant cell wall degrading enzymes (PCWDEs) produced, xylanases are of special interest since its degradation target, xylan, is one of the main structural polysaccharides in plant cell walls. In the biotrophic fungus Ustilago maydis, attempts to characterize PCWDEs required for virulence have been unsuccessful, most likely due to functional redundancy. In previous high-throughput screening, we found one xylanase to be important for U. maydis infection. Here, we characterize the entire U. maydis endo-xylanase family, comprising two enzymes from the glycoside hydrolase (GH) 10 family, Xyn1 and Xyn2, one from GH11, Xyn11A, and one from GH43, Xyn3. We show that all endo-xylanases except Xyn3 are secreted and involved in infection in a non-redundant manner, suggesting different roles for each xylanase in this process. Taking a closer look inside the plant during the pathogenic process, we observed that all secreted xylanases were necessary for fungal proliferation. Finally, we found that at least Xyn11A accumulated in the apoplast of the infected plant after three days, highlighting the role of these enzymes as important secreted proteins during fungal proliferation inside plant tissues.

Molecular Variability and Host Distribution of ‘Candidatus Phytoplasma solani’ Strains from Different Geographic Origins

The knowledge of phytoplasma genetic variability is a tool to study their epidemiology and to implement an effective monitoring and management of their associated diseases. ‘Candidatus Phytoplasma solani’ is associated with “bois noir” disease in grapevines, and yellowing and decline symptoms in many plant species, causing serious damages during the epidemic outbreaks. The epidemiology of the diseases associated with this phytoplasma is complex and related to numerous factors, such as interactions of the host plant and insect vectors and spreading through infected plant propagation material. The genetic variability of ‘Ca. P. solani’ strains in different host species and in different geographic areas during the last two decades was studied by RFLP analyses coupled with sequencing on vmp1, stamp, and tuf genes. A total of 119 strains were examined, 25 molecular variants were identified, and the variability of the studied genes was linked to both geographic distribution and year of infection. The crucial question in ‘Ca. P. solani’ epidemiology is to trace back the epidemic cycle of the infections. This study presents some relevant features about differential strain distribution useful for disease monitoring and forecasting, illustrating and comparing the phytoplasma molecular variants identified in various regions, host species, and time periods.

Genome-informed loop-mediated isothermal amplification assay for specific detection of Pectobacterium parmentieri in infected potato tissues and soil

Pectobacterium parmentieri (formerly Pectobacterium wasabiae), which causes soft rot disease in potatoes, is a newly established species of pectinolytic bacteria within the family Pectobacteriaceae. Despite serious damage caused to the potato industry worldwide, no field-deployable diagnostic tests are available to detect the pathogen in plant samples. In this study, we aimed to develop a reliable, rapid, field-deployable loop-mediated isothermal amplification (LAMP) assay for the specific detection of P. parmentieri. Specific LAMP primers targeting the petF1 gene region, found in P. parmentieri but no other Pectobacterium spp., were designed and validated in silico and in vitro using extensive inclusivity (15 strains of P. parmentieri) and exclusivity (94 strains including all other species in the genus Pectobacterium and host DNA) panels. No false positives or negatives were detected when the assay was tested directly with bacterial colonies, and with infected plant and soil samples. Sensitivity (analytical) assays using serially diluted bacterial cell lysate and purified genomic DNA established the detection limit at 10 CFU/mL and 100 fg (18–20 genome copies), respectively, even in the presence of host crude DNA. Consistent results obtained by multiple users/operators and field tests suggest the assay’s applicability to routine diagnostics, seed certification programs, biosecurity, and epidemiological studies.

Characterization of bronze leaf disease in western Canadian aspen and poplar trees

Aspen and poplar trees are important horticultural plants grown in Canada for aesthetic, commercial woodlot and windbreak applications. Bronze leaf is a destructive disease in <i>Populus </i>spp. and is caused by the fungal pathogen <i>Apioplagiostoma populi </i>Barr. This pathogen is often difficult to isolate and confirm from infected plant tissues and has been mainly identified by disease symptoms and morphological characteristics of <i>A. populi</i> when fruiting bodies form on infected leaves or branches. Affected leaves and branches typically become necrotic and bronze in colour. Air-borne spores and nursery shipments containing infected plants play an important role in the efficient movement of the pathogen. In this study, bronze leaf disease samples from symptomatic trees in Canada were examined microscopically for <i>A. populi</i> perithecia and asci. Pathogen-specific genomic sequences were identified for the development of sensitive stringent diagnostics that indicated branches and petioles were the most effective tissues for detecting <i>A. populi</i>. Leaf samples from symptomatic trees were collected in Canada and examined for perithecia to microscopically characterize <i>A. populi</i> asci and ascospores. Disease associated DNA sequences of the internal transcribed spacer (ITS) 5.8S region of the nuclear ribosomal were isolated from perithecia and symptomatic tree samples. Morphological and molecular biological data from this study characterized the relationship and epidemiology of <i>A. populi</i> and enabled the development of rapid diagnostic methods that restrict the extent of further losses in amenity and commercial plantings of aspen and poplar.

An Isoform of the Eukaryotic Translation Elongation Factor 1A (eEF1a) Acts as a Pro-Viral Factor Required for Tomato Spotted Wilt Virus Disease in Nicotiana benthamiana

The tripartite genome of the negative-stranded RNA virus Tomato spotted wilt orthotospovirus (TSWV) is assembled, together with two viral proteins, the nucleocapsid protein and the RNA-dependent RNA polymerase, into infectious ribonucleoprotein complexes (RNPs). These two viral proteins are, together, essential for viral replication and transcription, yet our knowledge on the host factors supporting these two processes remains limited. To fill this knowledge gap, the protein composition of viral RNPs collected from TSWV-infected Nicotiana benthamiana plants, and of those collected from a reconstituted TSWV replicon system in the yeast Saccharomyces cerevisiae, was analysed. RNPs obtained from infected plant material were enriched for plant proteins implicated in (i) sugar and phosphate transport and (ii) responses to cellular stress. In contrast, the yeast-derived viral RNPs primarily contained proteins implicated in RNA processing and ribosome biogenesis. The latter suggests that, in yeast, the translational machinery is recruited to these viral RNPs. To examine whether one of these cellular proteins is important for a TSWV infection, the corresponding N. benthamiana genes were targeted for virus-induced gene silencing, and these plants were subsequently challenged with TSWV. This approach revealed four host factors that are important for systemic spread of TSWV and disease symptom development.