Morphological variability of Fusarium oxysporum f.sp.capsici isolates infecting pepper (Capsicum annuum L.) landraces in West Gojjam Zone, Ethiopia

The study was conducted in Jabi Tehinan, Burie Zuria, Dembecha Zuria and Womberma districts of West Gojjam Zone, Amhara Regional State, Ethiopia to characterize the morphological variability of Fusarium oxysporum f.sp.capsici isolates collected from the main pepper producing areas of Amhara region, West Gojjam, Ethiopia.  These areas are known for high pepper production and incidence of Fusarium wilt. The study areas are situated in the Northwest direction of Addis Ababa at 385 km distance and 171 km South direction from the capital city of Amhara Region, Bahir Dar (Motbainor et al., 2016). The sampling areas are located between the geographical coordinates of 10°24’49” to 10°41’53” north latitude and 36°51’20” to 37°20’53” east longitude, with an altitude range of 1799 to 2115 m a.s.l. The districts have an average annual rainfall of 1168 mm and the mean annual temperature is 24.5°C.  The sample collection districts were purposively selected based on their pepper production levels and expansion potential. From each district, four farmer associations were selected and from each farmer association, three representative farmers’ fields were selected purposively based on their scale of production and incidence of disease. In each affected field, plant samples were collected randomly, from root, stem and leaf parts (Abebe, A. & Abera, 2019).  The sample plant parts were collected by looking into typical wilting symptoms including leaf yellowing, dropping, and partial or complete plant wilting (Akbar et al., 2018). Based on field size, 3–5 samples were collected per field (Altinok et al., 2019). Totally from all districts, 196 samples were collected and kept separately in labeled paper bags. The collected samples were placed in an icebox and taken to Injibara University, College of Natural and Computational Sciences, Microbiology and Parasitology Laboratory for identification. The samples were preserved at 4°C until isolation in the laboratory (Akbar et al., 2018; Endriyas et al., 2020). The sample collection was performed between September and October 2021 The collected symptomatic leaves, stems, and roots were thoroughly washed with running tap water. The washed parts were cut into small pieces (5 mm long) (Endriyas et al., 2020) and surface sterilized by soaking in 70% ethanol and 0.5% sodium hypochlorite solution (NaOCl) for 30 seconds and 1 minute, respectively (Oljira & Berta, 2020). The surface sterilized samples were rinsed for 1 minute in three successive changes of sterile distilled water and dried on filter paper (Altinok et al., 2019; Biri & Gomathinayagam, 2021). Then, the samples were placed on potato dextrose agar (PDA) plates (9 cm diameter) supplemented with chloramphenicol (0.25g/l) to restrict bacterial growth (Demissie et al., 2021; Kebede et al., 2020). All plates were labeled, sealed with parafilm and incubated at 25°C for 7 days to allow the development of mycelial growth of Fusarium (Getaneh et al., 2021; Hafizi et al., 2013). When the mycelia emerged, the colonies were sub-cultured onto a fresh sterile PDA to obtain a pure culture. Sub culturing was made till a distinct/pure culture are obtained and a pure culture of each isolate was stored on PDA slants at 4°C for further work (Hami et al., 2021). For the macroscopic data, colony color, pigmentation, margin, radial growth, shape, and mycelial growth pattern were recorded from the 7th day of pure culture following standard manuals (Campbell & Johnson, 2013; Leslie & Summerell, 2008; Nelson et al., 1983). Microscopic data were obtained using a compound microscope (400x) (Biri & Gomathinayagam, 2021) by preparing specimens from the 21 days grown mycelium.. Generally, morphological identification was performed according to the cultural characteristics and microscopic examination using the standard manuals used by different authors (Campbell & Johnson, 2013; Leslie & Summerell, 2008; Nelson et al., 1983).  The raw data is indicated in the tables below.