SciELO - Scientific Electronic Library Online

 
vol.76 issue2Genetic diversity and population densities of endophytic Bacillus spp. in yam plantsMaintenance pruning in physalis commercial production author indexsubject indexarticles search
Home Pagealphabetic serial listing  

Services on Demand

Journal

Article

Indicators

Related links

Share


Bragantia

Print version ISSN 0006-8705On-line version ISSN 1678-4499

Bragantia vol.76 no.2 Campinas Apr./Jun. 2017  Epub May 15, 2017

http://dx.doi.org/10.1590/1678-4499.171 

Plant Breeding

SH1 leaf rust and bacterial halo blight coffee resistances are genetically independent

Lucas Mateus Rivero Rodrigues1  * 

Masako Toma Braghini1 

Oliveiro Guerreiro Filho1 

1Instituto Agronômico - Centro de Café Alcides Carvalho - Campinas (SP), Brazil.

ABSTRACT

Coffee resistance to Pseudomonas syringae pv. garcae has been associated to pleiotropic effect of SH1 allele, present in coffee plants resistant to certain races of Hemileia vastatrix, the causal agent of leaf rust, or genetic linkage between resistance alleles to both pathogens. To validate this hypothesis, 63 coffee plants in F2 generation were evaluated for resistance to 2 isolates of H. vastatrix carriers of alleles, respectively, v2, v5 (isolate I/2015) and v1; v2; v5 (isolate II/2015) with the objective to confirm presence of SH1 allele in resistant plants to isolate I/2015. The same coffee plants were evaluated for resistance to a mixture of P. syringae pv. garcae strains highly pathogenic to coffee. Results showed that, among F2 coffee allele SH1 carriers, resistant to isolate I/2015, resistant and susceptible plants to bacterial halo blight were found; the same segregation occurs between F2 homozygous for SH1 allele, susceptible to the same isolate (I/2015) of H. vastatrix. Results also indicate that there is no pleiotropic effect of gene or allele SH1 connection between genes conferring resistance to leaf rust caused by H. vastatrix and bacterial halo blight caused by P. syringae pv. garcae.

Key words Coffea arabica L.; resistant cultivars; Pseudomonas syringae pv. garcae; linkage; Hemileia vastatrix; pleiotropic effect

INTRODUCTION

Leaf rust is the most important disease of coffee plantations, and widespread in major Coffee arabica L. producing countries. Variability of the causal agent, fungus Hemileia vastatrix Berkeley & Broome, is quite wide, and there are currently 45 known races of this fungus in world (Várzea and Marques 2005). In Brazilian coffee plantations, 17 races have already been identified (Zambolim et al. 2005).

Races of H. vastatrix are characterized by variable combinations of resistance genes, in total of 9, denominated SH1 to SH9 (Várzea et al. 2005), with direct implications for pathogenicity and in determining amplitude and nature of their hosts.

Studies conducted by Moraes et al. (1975) with diverse Coffea spp. germplasm exhibiting resistance to different races of H. vastatrix resulted in the identification of sources of resistance to bacterial halo blight, caused by Pseudomonas syringae pv. garcae, in C. arabica exotic varieties of Ethiopian origin, known as Harar, Dilla and Alghe, S 12 Kaffa, and Geisha, all of which are carriers of SH1 allele.

Multiple resistance to leaf rust and bacterial halo blight presented in these exotic varieties, could be explained according to Carvalho (1988), by the genetic linkage between resistance alleles to both pathogens or pleiotropic effect of SH1 allele. Sera (2001) and Fazuoli et al. (2009) also attributed to SH1 allele, present in the Ethiopian varieties (Bettencourt and Carvalho 1968), evaluated by Moraes et al. (1975), the simultaneous resistance to some races of H. vastatrix and bacterial halo blight caused by P. syringae pv. garcae.

According to Bettencourt and Carvalho (1968), SH1 allele is widespread in major coffee growing areas of Ethiopia. Studies conducted by these authors, allowed SH1 allele identification in various selections as Barbuk Sudan, BE-2 Ghembi, BE-4 Ennarea, BE-5 Wush-Wush, BE-6 Moderalo, BE-7 Boggia, BE-8 Era, BE-14 Loulo, Dilla & Alghe, Geisha, Lejeune’s, S 6 Cioiccie, S 9 Arba Gougou, S 12 Kaffa, S 17 Yrgalem, U l Dalecho, occurring individually or in combination with other resistance genes, as SH4 (SH1 SH4); SH5 (SH1 SH5) and SH4 and SH5 (SH1 SH4 SH5) and conferring resistance to 16 of 24 H. vastatrix races known at the period in which the research was conducted.

With SH1 allele conferring resistance to both pathogens, the search for sources of coffee resistant plants to P. syringae pv. garcae will be facilitated and occur simultaneous with tests to evaluate resistance to H. vastatrix. Therefore, the present study aimed to investigate whether coffee simultaneous resistance to H. vastatrix and P. syringae pv. garcae is due to pleiotrophic effect of SH1 allele or genetic linkage between resistance alleles to both pathogens.

MATERIAL AND METHODS

F2 progenies of coffee, from H8089-2 and H8089-7, composed respectively of 34 and 29 plants, were evaluated to infection by bacterium P. syringae pv. garcae and fungus H. vastatrix. These plants were derived from crossing between Catuaí Vermelho IAC 24 cultivar and IAC 1137-5 of Geisha originally from Ethiopia, carrier of SH1 and SH5 alleles (Bettencourt and Carvalho 1968). The Geisha selection was introduced in the coffee collection of Agronomic Institute (IAC) in 1953, coming from the United States Department of Agriculture under registration PI 205.928.

Plants were inoculated with bacteria obtained from the IBSBF Culture Collection of the Biological Institute, Campinas, Brazil, strains IBSBF 75 and IBSBF 1197 of P. syringae pv. garcae, in mixture, with approximate concentration of 108 UFC∙mL−1 (600 nm, absorbance = 0.25), by abrasion technique, which constitutes of friction against ab axial surface of leaves with medium grit sandpaper disposed in circular area (Ø1.5 cm) previously soaked in bacterial suspension causing limb injuries with no tissue drilling. Analysis of disease severity was carried out by using a rating scale of 0 to 5 points adapted from Paradela et al. (1974), as follows: 0 = no anasarca or chlorosis symptoms or hypersensitivity reaction around injured tissues; 1 = initial of bacterial colonization around lesions, with up to 10% of inoculated area showing disease symptom; 2 = 11 – 25% of inoculated area showing disease symptoms and with or without yellow halo; 3 = 26 – 50% of inoculated area showing disease development, yellowish halo pronounced around lesions; 4 = 51 – 75% of inoculated area with necrosis, yellowish halo throughout inoculated area; 5 = 100% necrosis of inoculated area. Plants were evaluated weekly up to 42 days after inoculation.

To confirm the presence of dominant SH1 allele, 63 F2 coffee plants were inoculated with 2 isolates of H. vastatrix obtained on differentiating plants grown in Centro de Café Alcides Carvalho from IAC.

The isolates used in this research were I/2015 carrying alleles V2 and V5 and isolate II/2015 the carrier of the alleles of virulence V1; V2; and V5. These aspects were confirmed by inoculation of this isolates on known differentiating plants to H. vastatrix races.

Disks of coffee leaves were inoculated with a drop of uredospore suspension with approximate concentration of 1.5 × 105 uredospore∙mL−1 of isolates I/2015 and II/2015, and subsequently kept in humid chambers in accordance with the methodology proposed by Eskes and Toma-Braghini (1981).

Two parameters were used to determine rust reaction: disease severity, evaluated with a 0 to 9 points scale, 0 being assigned to resistant plants without symptoms and 9, plants with high incidence of large lesions and regular and intense sporulation; and also type of reaction of lesions, evaluated by 0 to 4 point scale, in which 0 was assigned to immune plants without injuries and, four points to high susceptible plants with intense sporulation (Eskes and Toma-Braghini 1981).

Possibility of pleiotropic effect of SH1 allele or any genetic connection between resistance alleles to the pathogens used in this was investigated through reactions of plants to infection by bacteria strains IBSBF 75 and IBSBF 1197 and isolate I/2015 of fungus, and concordance between evaluated methods through following parameters:

  • Accuracy of method (AM) = (a + d)/n, calculated from sum of simultaneously resistant plants (a) and simultaneously susceptible (d) to rust and bacterial halo blight divided by total number of plants (n).

  • False positive rate (FPR) = b/(b + d), calculated by dividing the number of resistant plants to leaf rust, but susceptible to bacterial halo blight (b) by total of susceptible plants to bacterial halo blight (b + d).

  • False negative rate (FNR) = c/(a + c), calculated by division of number of susceptible plants to leaf rust, but resistant to bacterial halo blight (c) by total of resistant plants to bacterial halo blight (a + c).

  • Total rate error (TRE) = (b + c)/n, calculated from sum of resistant plants to leaf rust, but susceptible to bacterial halo blight (b) and number of susceptible plants to leaf rust, but resistant to bacterial halo blight (c) and divided by total number of plants (n).

  • McNemar chi-square with Yates continuity correction: (χ2McNemar ) = (|b − c| − 0.5)2/b + c, used to estimate significance between 2 assessments.

  • P value = quantitative measure of significance robustness of χ2McNemar test calculated.

  • Yule association coefficient (Q): measure degree of association between determined classes on evaluated plants for resistance to leaf rust and bacterial halo blight.

RESULTS AND DISCUSSION

The results related to resistance expression in progenies F2 from H8089-2 and H8089-7 to infection by P. syringae pv. garcae and isolates I/2015 and II/2015 of H. vastatrix are shown in Table 1.

Table 1 Reaction to infection by Pseudomonas syringae pv. garcae and isolates I/2015 and II/2015 of Hemileia vastatrix on F2 progeny from H8089-2 and H8089-7, hybrids between Catuaí Vermelho IAC 24 cultivar and Geisha IAC 1137-5 selection. 

Parameter H8089-2 H8089-7 Total
Plants (n°) 34 29 63
PHV I I/20151 (R:S) 6:28 10:19 16:47
PHV I I/20152 (R:S) 0:34 0:29 0:63
PPSG3 (R:S) 0:34 15:14 15:48

1Proportion of resistant and susceptible plants to isolate I/2015 of H. vastatrix;

2Proportion of resistant and susceptible plants to isolate II/2015 of H. vastatrix;

3Proportion of resistant and susceptible plants to P. syringae pv. garcae. P = Proportion; R = Resistant; S = Susceptible.

According to the obtained results, F2 plants are segregating for resistance to isolate I/2015 of H. vastatrix, but they proved to be susceptible to isolate II/2015. On the other hand, 34 plants of H8089-2 were susceptible to bacterial halo blight while 29 progenies of H8089-7 segregate in approximate 1R:1S ratio (Table 1).

From the total progeny analyzed regarding response to infection with isolate I/2015 of H. vastatrix, 16 plants presented resistance reaction and 47 were susceptible. On the other hand, regarding reaction to infection by P. syringae pv. garcae, 15 plants were identified with resistance reaction while remaining 48 showed to be susceptible (Table 1).

These results show heterozygous nature of F1 matrices (SH1 sH1) and, as demonstrated by Bettencourt and Carvalho (1968), genitors, Catuaí Vermelho IAC 24 cultivar and Geisha IAC 1137-5 selection were respectively homozygous for sH1 and SH1 alleles. Furthermore, according to these results, it was possible to confirm the presence of dominant allele SH1 in a third of plants, six coffee progenies of H8089-2 and ten coffee plants of H8089-7. In the gene-to-gene theory (Flor 1971), these coffee plants, carriers of at least one dominant allele SH1, had broken down resistance when inoculated with isolate II/2015 that carried v1 allele.

In the hypothesis of SH1 allele present pleiotropic effect or occurrence of genetic linkage between alleles that confer resistance to H. vastatrix and P. syringae pv. garcae as reported by Carvalho (1988), coffee plants carrying SH1 allele should be resistant to both pathogens.

Thus, F2 progenies, segregating, when inoculated with isolate I/2015 of H. vastatrix, should have the same reaction when inoculated with bacterium P. syringae pv. garcae, that is, resistant and susceptible plants to isolate I/2015 of H. vastatrix should be, respectively, resistant and susceptible to P. syringae pv. garcae.

In Table 2, it is presented the comparative data analysis of F2 progenies reaction with regarding to infections by P. syringae pv. garcae and by isolate I/2015 of H. vastatrix.

Table 2 Total number of F2 coffee plants of 2 hybrids H8089 resistant and susceptible to Pseudomonas syringae pv. garcae and isolate I/2015 of Hemileia vastatrix. 

H. vastatrix isolate
I/2015
P. syringae pv. garcae Total
Resistant Susceptible
Resistant 6a 10b 16a + b
Susceptible 9c 38d 47c + d
Total 15a + c 48b + d 63n

From 63 F2 coffee plants evaluated, 44 revealed simultaneously resistance (6) and susceptibility (38) to the isolate I/2015 of H. vastarix and to bacterial halo blight. Nineteen plants showed an opposite reaction to biotic agents, 10 of them were resistant to leaf rust and susceptible to bacterial halo blight, and 9 were resistant to P. syringae pv. garcae and susceptible to H. vastarix (Table 2). The results of statistical analysis performed with experimental data are described in Table 3.

Table 3 Estimation of parameters related to coffee F2 progeny of 2 hybrids H8089 response to infection by the bacterium Pseudomonas syringae pv. garcae and fungus Hemileia vastatrix. 

Parameters H8089-2 H8089-7 Total
Accuracy of the method (%) 82.35 55.17 69.84
False positive rate (%) 17.64 13.79 15.87
False negative rate (%) 0 31.03 14.29
Total rate error (%) 17.64 44.82 30.16
χ2 McNemar 4.17* 1.23ns 0.56ns
P-value 0.04 0.27 0.45
Yule association coefficient - 0.25 0.43
Standard deviation of Yule association coefficient - 0.37 0.32
Confidence interval of 95% of Yule association coefficient - 0.47 - 0.98 0.40 - 1.06

*Significant at 5% probability;

nsNon-significant; -Non-calculated values, since false negative rate is 0.

Accuracy of method (AM) calculated from comparative data analysis of coffee reaction to infection by P. syringae pv. garcae and isolate I/2015 of H. vastatrix denote that plants simultaneously resistant or susceptible to both pathogens was 69.84% considering total number of F2plants in the same group the total rate error (TRE) was equal to 30.16%.

The χ2McNemar test was significant in progenies from H8089-2 (4.17; p = 0.04), not significant in progenies from H8089-7 (1.23; p = 0.27) and also not significant in total group of plants (0.56; p = 0.45). However, significant value of χ2McNemar test calculated on H8089-2 progeny reveals inconsistency between the 2 classifications; the number of resistant plants to isolate I/2015 of H. vastatrix (6) is not the same number of resistant plants to P. syringae pv. garcae (0).

On the other hand, even though χ2McNemar value was not significant, in the analysis of all plants evaluated, resistance to biotic agents was not proven to be simultaneous, since 10 plants resistant to the isolate I/2015 of H. vastatrix carrying SH1 allele proved to be susceptible to bacterial halo blight and 9 plants resistant to bacterial halo blight have shown susceptibility to the isolate I/2015 of H. vastatrix, and false positive rate, in same group of plants, was equal to 15.87% and the false negative in second was 14.29%; the total error rate was 30.16%.

Yule association coefficient (Q) was equal to 0.25 for H8089-7 progeny, 0.40 in total group of plants, and has not been calculated for H8089-2 progeny, since false negative rate, i.e. of susceptible plants to isolate I/2015 of H. vastatrix, but resistant to P. syringae pv. garcae, was equal to 0. The magnitude of Yule coefficient, which ranges between 0 and ±1, observed individually in progeny H8089-7, and on F2 population as a whole (H8089-2 and H8089-7), confirms the lack of association between resistant and susceptible classes determined in the evaluation ratings of plants resistance to leaf rust and bacterial halo blight.

CONCLUSION

In accordance with the analyzed data, we can conclude that SH1 gene has no pleiotropic effect or genetic linkage between genes which confer resistance to isolates homozygous for V1 allele of leaf rust caused by H. vastatrix and to bacterial halo blight caused by P. syringae pv. garcae.

Search for resistance sources to bacterial halo blight for use in breeding programs aiming to develop cultivars with simultaneous resistance to both biotic agents should not be restricted to Coffea germplasm carrying SH1 gene with resistance to races or isolates of H. vastatrix.

ACKNOWLEDGEMENTS

The authors are grateful to the National Council for Scientific and Technological Development (CNPq) for financial support (Project 479.589/2013-50) and research scholarship to OGF; Coordination for the Improvement of Higher Education Personnel (CAPES) (to LMRR) and Coffee Research Consortium (to MTB), as well as to Dr. Suzete A. Lanza Destéfano, Curator of IBSBF Culture Collection, for providing bacterial strains used in this study, and Dr. Júlio Rodrigues Neto, Dr. Flávia Rodrigues Alves Patrício, Msc. Irene Maria Gatti de Almeida and Dr. Luis Otávio Saggion Beriam for their helpful suggestions.

REFERENCES

Bettencourt, A. J. and Carvalho, A. (1968). Melhoramento visando à resistência do cafeeiro à ferrugem. Bragantia, 27, 35-68. http://dx.doi.org/10.1590/S0006-87051968000100004. [ Links ]

Carvalho, A. (1988). Principles and practice of coffee plant breeding for productivity and quality factors. Coffea arabica. In R. J. Clark and R. Macrae (Eds.), Coffee: agronomy (p. 129-165). London: Elsevier Applied Science. [ Links ]

Eskes, A. B. and Toma-Braghini, M. (1981). Assessment methods for resistance to coffee leaf rust (Hemileia vastatrix Berk. & Br.). FAO Plant Protection Bulletin, 29, 56-66. [ Links ]

Fazuoli, L. C., Braghini, M. T., Silvarolla, M. B., Mistro, J. C. and Patrício, F. R. A. (2009). Melhoramento do cafeeiro visando à resistência a doenças. Proceedings of the 9th Curso de Atualização em Café. Campinas; Brazil. (Documentos IAC 91). [ Links ]

Flor, H. H. (1971). Current status of the gene-for-gene concept. Annual Review of Phytopathology, 9, 275-296. http://dx.doi.org/10.1146/annurev.py.09.090171.001423. [ Links ]

Moraes, S. A., Sugimori, M. H., Tomazello Filho, M. and Carvalho, P. C. T. (1975). Resistência de cafeeiros a Pseudomonas garcae. Summa Phytopathologica, 1, 105-110. [ Links ]

Paradela Filho, O., Ribeiro, I. J. A. and Sugimori, M. H. (1974). Comportamento diferencial em progênies de cafeeiros de dois isolados do agente causador da mancha aureolada do cafeeiro. Fitopatología, 9, 64. [ Links ]

Sera, T. (2001). Coffee genetic breeding at IAPAR. Crop Breeding and Applied Biotechnology, 1, 179-190. [ Links ]

Várzea, V. M. P. and Marques, D. V. (2005). Population variability of Hemileia vastatrix vs. coffee durable resistance. In L. Zambolim, E. M. Zambolim and V. M. P. Várzea (Eds.), Durable resistance to coffee leaf rust (p. 53-74). Viçosa: UFV. [ Links ]

Várzea, V. M. P., Rodrigues Junior, C. J., Silva, M. C. M. L., Gouveia, M., Marques, D. V., Guerra-Guimarães, L. and Ribeiro, A. (2005). Resistência do cafeeiro a Hemileia vastatrix. In L. Zambolim (Ed.), O estado da arte de tecnologias na produção de café (p. 297-320). Viçosa: UFV. [ Links ]

Zambolim, L., Zambolim, E. M., Vale, F. X. R., Pereira, A. A., Sakyama, N. S. and Caixeta, E. T. (2005). Physiological races of Hemileia vastatrix Berk. et Br. in Brazil — Physiological variability, current situation and future prospects. In L. Zambolim, E. M. Zambolim and V. M. P. Várzea (Eds.), Durable resistance to coffee leaf rust (p. 75-98). Viçosa: UFV. [ Links ]

Received: May 09, 2016; Accepted: August 25, 2016

Creative Commons License This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.