Morphological Characterization of Phytopathogenic Fungi Isolated from Seeds of Barley Plants (Hordeum Vulgare) in Mexico

Table of Contents:

TERESA ROMERO CORTES1, VICTOR HUGO PEREZ ESPANA1,

PABLO ANTONIO LOPEZ PEREZ1, EDUARDO RANGEL CORTES1,

MARIO A. MORALES OVANDO2, MARIO RAMIREZ-LEPE3, and JAIME ALIOSCHA CUERVO -PARRA1

  • 1Escuela Superior de Apan-Universidad Autonoma del Estado de Hidalgo, Carretera Apan-Calpulalpan, Km 8, Chimalpa Tlalayote s/n, Colonia Chimalpa, 43900 Apan, Hidalgo, Mexico
  • 2Universidad de Cienciasу Artes de Chiapas, Sede Acapetahua, Calle central norte s/n entre 4aynorte, 30580. Acapetahua, Chiapas, Mexico
  • 3Unidad de Investigaciony Desarrollo en Alimentos, Institute Tecnologico de Veracruz, Av. Miguel Angel de Quevedo No. 2779, Colonia Formando Hogar, Veracruz, Ver, Mexico

'Corresponding author. E-mail: This email address is being protected from spam bots, you need Javascript enabled to view it

ABSTRACT

Worldwide, barley (Hordeum vulgare L.) is the fourth most important cereal after wheat, maize, and rice and it is used in the livestock, food, and brewing industry. The main producer of this cereal is the Russian Federation with a production of 18 million tons per year. In Mexico, the barley is planted in an approximate area of 283,386 ha; 83% of this area is located in the States of Hidalgo (44%), Mexico (11%), Puebla (14%), and Tlaxcala (14%). The cultivation conditions depend on rainfall as a unique source of moisture during the summer cycle. Unfortunately, the minimum tillage methods and monoculture of cereals have favored the increase of phytopathogens causing foliar spots, root rot, and fusariosis; which is particularly important because it affects yield and produces toxins in the grain that are harmful to human and animal health. Among the phytopathogenic fungi that cause losses in barley plantations worldwide, Puccinia striiformis can be mentioned that causes yellow or linear rust. Puccinia hordei that causes leaf rust or brown rust, Drechslera teres/ Pyrenophora teres that causes net stain, with losses in yield greater than 50%, to the fungi Altemaria spp., Fusarium spp., Epicoccwn sp., which cause the black tip, Fusarium spp., which cause fusariosis, F. graminearumIGibberela zeae that causes seedling blight, Bipolaris sorokiniana/CochlioboItts sativus that causes seedling blight or blurred spot, Ustilago nuda that causes loose smut, and U. hordei causing covered smut disease. The yellow rust and the red spot diseases cause the greatest damage to the barley plantations. The phytopathogenic fungi classification of barley worldwide has been based on the use of morphological characters and the relationship with then hosts. Therefore, performing the morphological characterization of fungal isolates from the barley plant in Mexico will allow us to have knowledge about the microorganisms that are present in our country and thus to control those that are harm fill to the plant. For that, the aim of this research was the isolate and morphological characterization of fungi associated with diseased barley plants in Mexico.

INTRODUCTION

Worldwide, barley (Hordeum vulgare) is the fourth most important cereal, followed by wheat, maize, and rice. Both the seed and the plant are used in the livestock, food, and brewing industry (Sanchez, 2011). The largest producer of this cereal is the European Union with a production of 859,500,000.00 tons, followed by Russia with a production of 17,000,000 metric tons per year. In the American continent, the main producer is Argentina with 3,400,000 tons per year (PMC, 2017).

In Mexico, the barley is planted in an approximate area of 336,000 ha; where 90% of this area is located among the states of Guanajuato, Hidalgo, Mexico, Puebla, and Tlaxcala (Zamora et al., 2008; SAGARPA, 2017). Based on the United States Department of Agriculture (USDA) estimated, annual production of barley for Mexico will be 735,000 tons, ranking No. 19 globally (PMC, 2017).

The crop is produced under rainfed or seasonal conditions in the summer cycle, depending on rainfall as a single source of irrigation (SAGARPA,

  • 2009). In the Hidalgo state, barley cultivation occupies the second socioeconomic importance after maize, in terms of area sown, production volumes obtained and the number of producers (Gomez et ah, 1997). However, this crop is affected by several phytopathogenic fungi responsible for diseases such as rust, foliar spots, root rot, and fusariosis. The latter is of great importance as it affects yield and contaminated grain produces toxins that are harmful to animal and human health (Gilchrist-Saavedra, 2000).
  • 15.1.1 USES OF BARLEY PLANT

In Mexico, barley is a crop of great economic and social importance in the high valleys of the country, because farmers prefer it because their vegetative cycle is short, has resistance to drought, low temperatures, and saline soils. Of which, the states of Guanajuato and Hidalgo are the two main producers of barley grains in Mexico, with 372,167 and 268,595 tons (SAGARPA, 2017).

At present, this cereal is produced in almost all the world, destining it mainly to two types of market: like food for cattle and for the malt production. Particularly in Mexico, approximately 70% of the barley produced is used by the malting industry and the remaining 30% corresponds to varieties that are mainly used for cattle feed.

The consolidation in the domestic and export markets of the two large breweries in Mexico and the good positioning of their products in the world markets has led to the development integrated of a malt production industry in Mexico. This industry has, in turn, developed its own barley grain traders, which enter into contracts with agricultural producers for the production of malt varieties demanded by industry (Espinosa et al., 2003). However, the numerous diseases present in this crop are a strong limitation of production, especially due to the high cost of fungicides and their application (Gilchrist- Saavedra, 2000).

15.1.1.1 PHYTOPATHOGENIC FUNGI AFFECTING BARLEY CULTIVATION

Barley plantations are affected by phytopathogenic fungi that cause losses worldwide, may be mentioned the following: Puccinia striiformis causes “yellow or stripe rust” (Gilchrist-Saavedra et al., 2005), Puccinia liordei causes “leaf rust” or “brown rust” (Sanchez, 2011), Drechslera teres causes “the net blotch,” causing losses of up to 50% (Gilchrist-Saavedra, 2000),

Altemaria spp., Fusarium spp., and Epicoccitm sp., cause “the black tip” (Carmona et al., 2011), Fusarium spp., they cause “fusariosis” (Zuniga et al., 2010), F. graminearum/Gibberella zeae causes “seedling blight,” Bipolaris sorokiuiaua/Cochliobolus sativus causes “seedling blight” or “blurred spot,” Ustilago nuda causes “flying coal” (Carmona et al., 2011), and Ustilago hordei causes “covered smut of barley” (Zuniga et al., 2010). Of these diseases, which cause greater damage to the barley plantations are the yellow rust and the red spot (Carmona et al., 2011; Sanchez, 2011). In general, the crop is frequently affected by foliar stains and the different species of pathogenic fungi of the genus Fusarium which has been favored by the cultural methods used for the tillage and the retention of the harvest residues, which has favored the increase of the inoculum. The infected grain is the primary inoculum of the vast majority of diseases (Gilchrist-Saavedra et al., 2005).

METHODOLOGY

Thirteen samples of 500 g of seed per lot from 13 different localities, varieties, and years of cultivation in the upper valleys area were analyzed for the states of Guanajuato, Hidalgo, and Puebla. The general data for the samples studied are specified in Table 15.1 and Figure 15.1.

TABLE 15.1 Origin of Barley Grains Samples.

Lot number

Variety

Origin place

Harvest date

Ml

Esmeralda

Apan, Hidalgo

2010

М2

Forrajera

Mijapa. Hidalgo

2011

М3

Forrajera

Tepeapulco. Hidalgo

2013

M4

Esmeralda

Apan, Hidalgo

2012

M5

Adabella

Tepepatlaxco, Hidalgo

2014

Мб

Esmeralda

San Felipe, Hidalgo

2009

M7

Esmeralda

Leon, Guanajuato

2013

M8

Esmeralda

Libres, Puebla

2008

M9

Gaviota

Leon, Guanajuato

2013

M10

Adabella

Almoloya. Hidalgo

2013

Mil

Josefa

Apan, Hidalgo

2015

M12

Josefa

Apan, Hidalgo

2016

M13

Josefa

Apan, Hidalgo

2017

Morphological characteristics of barley seed samples. Ml (a), М2 (b), М3 (с), M4 (d), M5 (e), M6 (f), M7 (g), M8 (h), M9 (i). M10 (j), МП (к), M12 (1), M13 (m)

FIGURE 15.1 Morphological characteristics of barley seed samples. Ml (a), М2 (b), М3 (с), M4 (d), M5 (e), M6 (f), M7 (g), M8 (h), M9 (i). M10 (j), МП (к), M12 (1), M13 (m).

15.2.1 METHODS OF CULTURING AND ISOLATING FUNGI

Barley seeds (10 g) were disinfected with 10% sodium hypochlorite solution (NaOCl) for one minute. Subsequently, the seeds were dried at room temperature and potato dextrose agar (PDA) was used for the development of fungal species. Samples were incubated for 7 days at 25°C. Subsequently, they were analyzed with a microscope stereoscopic and in those where fungal colonies were observed, 5 mm diameter discs were cut with the aid of a punch and placed in other Petri dishes with PDA medium for their growth, isolation and subsequent morphological characterization. The isolated strains were incubated for 5 days at 25°C or until all of the culture medium was filled.

15.2.2 MAINTENANCE AND PROPAGATION OF STRAINS

For maintenance and propagation of the strains, a spore wash was carried out with a sterile solution of 0.01% Tween 20 (Arevalo et al., 2005) at a concentration of 1 x 106 spores/mL for each isolated phytopathogenic fungus. The spore concentration was performed by the direct total count by the Thoma chamber method. One part of this spore suspension was transferred to another Petri dish, homogeneously distributed, incubated at 25°C for 7 days and stored refrigerated at 4°C under sterile conditions until use. In order to maintain the viable culture, the re-plantings were earned out every 30 days (Alvarez, 2007). After each microorganism was grown in the Petri dish, with PDA medium, 5 mL of a 0.01% Tween 20 solution was added and the spore suspension was recovered in sterile vials, to which were added 5 mL of a 40% glycerol solution and finally, the suspensions of spores were preserved in freezing at -86°C until use.

15.2.3 MICROORGANISMS USED

The microorganisms used in this study were strains of phytopathogenic fungi isolated from samples of barley plant seeds with symptoms of disease caused by fungi, which are listed below: Drechslera teres, Bipolaris soro- kiniana, Fusarium gromineorum, Fusarium oustroomericanum, Fusarium pseudograminearum, Rhizopus oryzae, Puccinia triticina, Puccinia stri- iformis, Puccinia recondita, Puccinia hordei, Alternaria alternata, Peni- cillium digitatum, PeniciUium citrinum, and Penicillium oxalicum. The fungal strains were isolated from seeds with characteristics of disease as yellow rust and brown rust from the upper valleys area of the states of Guanajuato, Hidalgo, and Puebla.

15.2A MORPHOLOGICAL CHARACTERIZATION

Morphological descriptions were based on comparisons with descriptions of other authors (Ireta and Gilchrist, 1994; Crous et al., 1995; Scudamore and Hetmanski, 1995; Maenetje and Dutton, 2007; Manamgoda et al., 2014; Lawrence et al., 2016), augmented by new observations as noted. PDA medium was the standard growth medium used for the growth rate and the morphological study of the reproductive structures of all fungi. Microscopic observations and measurements of the reproductive structures were made with an optical microscope (ZEIGEN, Model ZB-7100). All strains were stored as conidia and hyphae at -86°C until processed.

 
Source
< Prev   CONTENTS   Source   Next >