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Histopathology of Alternaria Infected Chilli Seeds

A.S. Basak
Department of Botany, University of Chittagong

G.A. Fakir
Khulna University, Khulna, Bangladesh

M.A.U. Mridha
Department of Botany, University of Chittagong

Abstract/ Executive Summary:

Histopathological studies were carried out with chilli seeds infected by Alternaria capsici-annui by blotter method and whole mount cleared preparations following microtome section. The seed parts, such as seed coat, endosperms and embryo were found associated with these pathogens. Seed coat infected by Alternaria spp. showed profuse thick dark-brown mycelium. In case of severely infected seeds, mycelium invaded the endosperms and they completely disintegrated the embryonic tissue.

Keywords: Fungi, Chilli seeds, Alternaria

Location: Chittagong District

Start Date:

End Date:

Program Area: Pest Management

Commodity: Chilli

Objectives:

To determine the site of infection of these fungi in chilli seeds.

Methodology:

Collection of samples: Fruit samples having black or Alternaria rot symptoms produced by A. capsici-annui and A. tenuis were collected from fields of Chittagong district. All the samples were properly sundried and stored in an air-cooled room at 18 ± 2°C until the seeds were analysed for the presence of selected fungi. Seeds from the fruits of infected samples were separated just before use.

Blotter method: The presence of Alternaria spp. with seeds was determined by blotter method as described by ISTA. Seeds' from the infected fruits were graded at five ratings of disease severity. These were 1 = 1-10% fruit area infected, 2 = 11-25% fruit area infected, 3 = 26-50% fruit area infected, 4 = 51-75% fruit area infected and 5 = 76-100% fruit area infected. Two hundred seeds from each grade were selected randomly. The seeds were washed for four to five times with distilled water and soaked individually in sterile water, one seed per test tube, for 16 h before dissecting different parts such as seed coat, endosperms and embryo. Each part was surface sterilised by 1: 1000 HgCl₂ solution and placed directly on water-soaked blotter in such a way that the seed coat lies in outer periphery, endosperms in the middle and embryo in the centre of the Petri dish. In each Petri dish, dissected parts of 10 seeds were plated. After 7 days incubation at room temperature (18 ± 2°C), seed parts were examined for fungal infection. Correlation analysis was done to find out the relationship between the rating scales of disease severity of fruit rot and the percentage recovery of fungi from Alternaria infected seeds.

Histopathological studies were carried out by wholemount cleared preparations. In this method, selected seeds were boiled in water for about 45 min and' then cleaned in concentrated HCI by boiling for 10-20 min. The seeds were then washed thoroughly with water and fixed in 70% ethanol for 48 h. Each seed was handled separately. The fixed seed was dehydrated through a tertiary butyl alcohol series, infiltrated and embedded in paraffin. The materials were softened before sectioning by paring the paraffin from the blocks on one side upto the material and then immersing in a 1% aqueous solution of sodium lauryl sulphate for 24 h. The blocks were washed in water and transferred to a mixture of glycerine and acetic acid (1:1) for four weeks. Serial microtome sections were cut at 25 flm thickness, stained with saffranin and fast-green and mounted in caedax solution (two parts caedax + one part xylene). The sections were examined under a microscope for the presence of fungal propagules in seed tissue.

Source of Information: Bangladesh Journal of microbiology,Vol-11.No:1, June, (ISSN 1011-9981), Page:37-41,

Article Link (Online Journal):

Funding Source:

1. Government

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The pathogens were present in all the three seed parts. Seed coat infected by Alternaria spp. showed profuse thick dark brown to black branched mycelia which was found to invade the endosperm after breaking the structural barrier of cuticle. It was found that Alternaria spp. were deep-seated pathogens. Moreover, the outer epidermal layer of chilli seeds is hard with a thick layer made of cutin . If the pathogen once breaks this layer.

Knowledge Management: Journal