Introduction

It is recognized the incidence of phytopathogens and susceptibilities in the different variants of pepper, and in different production systems (greenhouse, biosphere, open sky, etc), so that diseases cause large losses in quality and yield, depending on favorable environmental conditions for its presence and crop management practices (^{Guigon-López and González-González, 2001}). Among the major pathogens that attack the crop pepper are: Phytophthora capsici, Xanthomonas campestri sp. Vesicatoria, Alternaria spp., A. alternata, Oidiopsis taurica, Leveillula taurica, Fusarium spp., Rhizoctonia solani, Pythium spp., Sclerotinia sclerotium, Sclerotium rolfsii, various nematodes (Meloidogyne spp. and Heterodera spp.) and viral complexes tomato, cucumber, snuff, potatoes, peppers and others (^{Guigon-López and González-González, 2001}; ^{Velázquez et al., 2002}; ^{Velásquez-Valle and Amador-Ramírez, 2007}; ^{Chew et al., 2008}).

The wilt of pepper or drier is one of the main diseases that cause losses in production from 26% to 90%, not only in Mexico but to worldwide. It is caused by a complex phytopathological of Fusarium spp., Phytophthora capsici, Verticillium spp., Macrophomina spp., Rhizoctonia solani, Pythium spp. and Sclerotium rolfsii, acting alone or in combination (^{Ristaino and Johnston, 1999}; ^{Velásquez et al., 2000} and 2001; ^{González-Pérez et al., 2004}; ^{Rico-Guerrero et al., 2004}; ^{Vásquez et al., 2009}; ^{Montero-Tavera et al., 2013}). As a result of the incidence of this disease, the acreage has declined or has moved into new areas (^{González-Pérez et al., 2004}). The complex etiology or pathological complex makes it difficult to establish a specific epidemiological pattern of disease development, preventing establish a specific management program coupled with high genetic variability in pathogens involved (^{Montero-Tavera et al., 2013}).

The symptoms of wilting of pepper or drier are varied but essentially reflected in leaf wilting without color change, chlorosis and then premature leaf fall, leaf ripening, early and irregular ripening of fruits, root rot or stem base rot and plant death (^{Avelar and Marban, 1989}; ^{Velásquez-Valle et al., 2001}; Vásquez et al., 2002). However, it has improved pathogens identification techniques based on DNA extraction and PCR amplification (^{Rico-Guerrero et al., 2004}).

Among the main management strategies of the pepper wilting, the use of rhizospheric fungi antagonistic to Fusarium spp, Rhizoctonia solani and Phytophthora capsici (^{Guillen-Cruz et al., 2006}; ^{Jiang et al., 2006}; ^{Robles-Yerena et al., 2010}; ^{Ramos-Sandoval et al., 2010}); rhizobacteria antagonistic to P. capsici (^{Sang et al., 2008}; ^{Sang and Kim, 2012}); combination of biofumigaciones and antagonistic microorganisms (^{Wang et al., 2014}); inoculations with rhizospheric mycorrhizal fungi (^{Ozgonen and Erilic, 2007}); combination of use of composts and grafting in resistant rootstocks (^{Gilardi et al., 2013}); crop rotation (^{Lamour and Hausbeck, 2003}), use of resistant germplasm to main complex Fusarium-Rhizoctonia-Phytophthora-Pythium (^{Koc et al., 2011}; ^{Babu et al., 2011}); even when there is no total resistance to the complex; and fungicide application (^{Chew et al., 2008}). However, they begin to identify expression profiles of specific DNA sequences (transcriptome) related phenotypic resistance to P. capsici from the creole collection Morelos-334 (^{Walker and Bosland, 1999}; ^{Richins et al., 2010}). Also in native Mexican pepper varieties they have been identified sources of resistance to P. capsici (^{Moran-Bañuelos et al., 2010}).

In general, it is necessary to continue documenting the presence and magnitude of damage caused by pepper wilt, the regional situation and associated pathogens, with the purpose of making decisions about management programs and duly justified. The regional diagnostics help detect changes and prevalence of the disease, relations associated pathogens, pathogen-host interactions and aspects of culture system that favor or stop the development of the disease, among other aspects (^{Velásquez-Valle et al., 2001}a; ^{Guigón-López and González-González, 2001}; ^{González-Pérez et al., 2004}; ^{Chew et al., 2008}; ^{Vásquez et al., 2009}). In this context, the objective of the study was to perform a rapid diagnosis of the symptom and pathogens associated with chill wilt in the Central Valleys of Oaxaca, Mexico.

Materials and methods

Results and discussion

As for the description of the pepper production system, it was found that the age of the producers who cultivated pepper went from 40 to 60 years of age (65.1%) and there were farmers who had from 10 to 30 years planting the crop. However, most of them sow half a hectare or less (88.9%) and regionally, water pepper is the most cultivated type (92.1%). Although there were plantations of habanero, jalapeño, tabiche, poblano and tusta. At the time of the field visit, the crops had 90 to 120 days of being transplanted and were in flowering and production, when the presence of wilting was most noticeable. They always fertilize either organically or inorganic-organic combinations, and usually perform chemical controls of diseases or pests.

The seed comes from its own previous cultivation or exchange with neighbors for both irrigation and temporary crops. Farmers point out that they always have problems with the cultivation, whether of a sanitary type (pests or diseases) or high costs of inputs and low sales in the regional markets that are the main destination of production. The main crop is the water pepper, and is practically sown all year round in the Central Valleys. However, when wilt was present the damage was evaluated from mild to moderate (90.5%) of sites visited; that is, the disease began to appear and the base of the stem had an advanced degree of necrosis. Combinations of root rot, weevil attack, or presence of viral infections were sometimes present. The results show that the management of the pepper crop is variable.

The most water pepper plant almost all year, this is verified with statistical significant differences (p< 0.01) between classes of responses. The results corroborate previous signs of ^{Velasco et al. (1998)} and ^{López (2007)}, who stated that the production of water pepper in small areas, besides generating income for the producers, stimulates the local economy because, if the parcels are irrigated, they can grow pepper throughout year. On the other hand, the exchange of seed without health care can help to spread disease in the region, this coincides with the field observations of ^{López (1989)}, noting that the rapid increase in area sown between 1970 and 1985, brought with it the appearance and development of problems and diseases, because there are no improved varieties or seed health checks.

In correspondence analysis, it was determined that the age of the farmer, area planted, type of pepper planted, planting season and transplant, phytosanitary problems, fertilizer use and seed origin, were determinant in the description of the pepper production system of 63 farmers in the Central Valleys of Oaxaca. The fourth main dimension explained 82.4% of total variability (Table 1).

Table 1 Values and eigenvectors of the correspondence analysis based on farmers’ responses to the management of the pepper crop in Central Valleys of Oaxaca. 

In the graphical representation of the correspondence analysis, the first two main dimensions (Figure 1) showed that the pepper production systems follow a pattern determined by the area sown, planting and transplanting, type of pepper and use of fertilizers. Thus, some plots in the Coatlán-Sola of Vega subregion follow a particular pattern because they are essentially planted with tusta pepper, and in the case of certain plantations established in the Tlacolula-Zimatlán subregion, mainly of water pepper, based on the planting season and transplant and use of agrochemicals (30.7%). It was also possible to find a pattern of differences of small, medium and large sown areas of 0.25 or less than 1 ha. In the case of farmers with an area of less than 0.25, they correspond to the Coatlán-Sola of Vega, Ocotlán-Ejutla and Etla subregions, with the largest area corresponding to the Etla subregion.

Figure 1 Description of management of pepper production units in three subregions of Central Valleys of Oaxaca, based on the first two main dimensions of correspondence analysis.  

The 79.4% of the area sown corresponds to irrigation and irrigation-time, which means that when irrigation rolled flooding in the plot, coinciding with the rainy season and flooding. Thus, the water dispersed sporangia and motile zoospores then find a favorable environment to disperse released, and the waterlogged soil becomes focus of infection and means of survival of the pathogen, as indicated earlier reports to P. capsici (^{Erwin and Ribeiro, 1996}; ^{Ávila-Quezada et al., 2005} and ^{Silva-Rojas et al., 2009}).

As for incidence of pepper wilt caused by phytopathological complex it was observed in the entire region of the Central Valleys explored four diseases were presented: Rhizoctonia, Fusarium, Phytophthora and Pythium. The subdivision into four subregions made it possible to observe a relationship between pathogens and subregions. For example, for the regions of Coatlan-Sola of Vega and Ejutla-Ocotlán there was a higher incidence of Rhizoctonia and Fusarium, but also Zimatlán-Tlacolula Phytophthora and Pythium also presented. The presence of the four pathogens also became evident in the Etla region. It is thought that wilt damages occur more frequently if there is a prevalence of two or more pathogens in the soil, and specifically the regions of Etla and Zimatlán-Tlacolula presented with a higher incidence of the disease. This results are consistent with previous reports of ^{Vásquez et al. (2009)} in water pepper (Table 2).

Table 2 Observed frequency of pathogens identified in the samples (n= 937) isolates in pepper plants with wilt symptoms, in the Central Valleys of Oaxaca.  

^*= diferencias significativas a nivel p≤ 0.05; ^**= diferencias significativas a nivel p≤ 0.01 (prueba de χ²).

Diagnostic results on the association of phytopathological complex Fusarium Rhizoctonia-Fusarium-Phytophthora-Pythium, was observed and identified the presence of two or more of them and manifesting symptoms of wilting, which coincide with the work of ^{Durán-Ortiz et al. (2001)} of wilt in the States of Zacatecas and Aguascalientes. In Camargo, Delicias and South of Chihuahua was determined to Phytophthora capsici as the causal agent of wilting pepper (^{Silva-Rojas et al., 2009} and ^{Guigon-López and González-González, 2001}). ^{Vásquez et al. (2009)} study found that Phytophthora and Rhizoctonia are the causative agents of wilting for three communities in the state of Oaxaca. Other studies indicate that pathogens were more frequently associated with Rhizoctonia spp. and Fusarium spp., in Aguascalientes and Zacatecas (Rico-Guerrero, 2002).

When pathogenicity test was performed at 17 and 52 isolations of Phytophthora and Rhizoctonia, respectively; in both cases, a number of non-pathogenic isolates were detected rather than pathogenic. Of these, three in Phytophthora and seven in Rhizoctonia were determined. Even though in these pathogenicity tests there were isolates that independently did not reproduce the symptoms, it is not a reflection of loss of pathogenicity because in the field two or more pathogens regularly converge in the same site to reproduce the symptoms of wilt. It should also be noted that inoculate were only made in water pepper plants at 45 days of age, the first growth phase (Table 3).

Table 3 Results of tests of pathogenicity of 17 isolates of Phytophthora and 52 in Rhizoctonia, and their relation to the subregion of origin in Central Valleys of Oaxaca.  

A inoculations 60 days, symptoms of Rhizoctonia plants were chlorosis, after necrosis the stem base with detachment of epidermis and finally death of the plant. In inoculations with Phytophthora, symptoms appeared 55 days of inoculation; among the most notable were loss of leaf turgor or wilting, defoliation, slight yellowing, necrosis at the base of the stem and plant death. Pathogenicity was quantified as the number of dead plants with symptoms of the disease.

Conclusions

The agro-systems of pepper production in Central Valleys of Oaxaca are diverse in infrastructure used, area planted, crop management, pests and diseases. In this sense, water pepper is highlighted as the most frequent and largest area planted throughout the year. Producers usually obtain their own seed, use chemical fertilizers, regularly present pest incidences such as fruit weevil and root rot associated with wilt symptoms, and 84.1% of them use agrochemicals for their control. In the correspondence analysis, it was determined that the age of the farmer, area planted, type of pepper planted, planting season and transplant, phytosanitary problems, fertilizer use and seed origin were the variables with the highest descriptive value of the production system of Pepper from 63 farmers in Central Valleys of Oaxaca.

The different symptoms were observed and listed as pepper wilting in several production sites in the Central Valleys and related pathogens were Rhizoctonia and Fusarium symptoms, and two oomycetes: Phytophthora and Pythium. A significant relationship is also determined (X², p< 0.05) between subregions visited and frequency of distribution of pathogens: in the regions of Coatlán-Sola of Vega and Ejutla-Ocotlán had higher incidence of Rhizoctonia and Fusarium, but Zimatlán-Tlacolula and Etla also presented Phytophthora and Pythium. The results show the presence of two or more pathogens in the same site, and consequently higher incidence in the field of the disease.

In the pathogenicity tests one or more symptoms of wilt pepper reproduced by inoculating 3 and 7 pathogenic isolates of Phytophthora and Rhizoctonia respectively; in plants of 45 days old water pepper. These pathogenic isolates are feasible to be used in tests of tolerances or resistance of pepper germplasm.