567 Cacefo, V. et al.

BIOLOGICAL CONTROL OF Hemileia vastatrix Berk. & Broome WITH

Bacillus subtilis Cohn AND BIOCHEMICAL CHANGES IN THE COFFEE

Viviane Cacefo1, Fábio Fernando de Araújo2, Ana Cláudia Pacheco3

(Recebido: 04 de abril de 2016; aceito: 03 de junho de 2016)

ABSTRACT: Hemileia vastatrix stands out as one of the main phytosanitary problems of the coffee culture. The objective of
this study was to evaluate the efficiency of the biological control of Hemileia vastatrix and detect some biochemical changes in
plants of cultivars Icatu and Mundo Novo treated with foliar spray of Bacillus subtilis and azoxystrobin under field conditions.
After the applications, leaves were collected for visual assessment of H. vastatrix severity in accordance with diagrammatic
scales. For the biochemical evaluations related to plants defense, leaf tissue samples were collected for the analysis of proline,
total phenols, total protein and lipid peroxidation. B. subtilis acted effectively in control of H. vastatrix in both cultivars, but
with less efficiency than the azoxystrobin. Plants treated with B. subtilis presented greater proline, total phenols and total
proteins in their leaves on the application of azoxystrobin. The cultivar Mundo Novo was more responsive to treatments that Icatu.

Index terms: Coffea arabica, induced resistance, rhizobacterium.

CONTROLE BIOLÓGICO DA FERRUGEM COM Bacillus subtilis E ALTERAÇÕES

BIOQUÍMICAS NO CAFEEIRO
RESUMO: A ferrugem destaca-se como um dos principais problemas fitossanitários da cultura do café. O objetivo deste
trabalho foi avaliar a eficiência do controle biológico da ferrugem e detectar alterações bioquímicas nas plantas das cultivares
Icatu e Mundo Novo em função da pulverização foliar com Bacillus subtilis e azoxistrobina em condições de campo. Após as
aplicações foram coletadas folhas para avaliação visual da severidade da ferrugem de acordo com escalas diagramáticas.
Para a avaliação bioquímica relacionada com a defesa nas plantas, foram coletadas amostras de tecido foliar para realização
de análises de prolina, fenóis totais, proteínas totais e peroxidação de lipídeos. B. subtilis atuou de forma eficaz no controle
da ferrugem nas duas cultivares testadas, porém com desempenho inferior a azoxistrobina. O tratamento com B. subtilis
proporcionou maiores teores de prolina, fenóis totais e proteínas totais nas folhas em relação à aplicação de azoxistrobina,
com a cultivar Mundo Novo sendo mais responsiva aos tratamentos que a Icatu.

Termos de indexação: Coffea arabica, indução de resistência, rizobactéria.

1 INTRODUCTION pesticides (CARVALHO et al., 2012).

The cultivar Icatu has more rustic features,
The fungus Hemileia vastatrix Berk. & high size, canopy with larger diameter, vigorous
Broome (coffee rust) stands out as the main cause growth, average maturity late and moderate
of the coffee disease due to major damages. It resistance to H. vastatrix and drought, as well as
causes a foliar disease that initially cause chlorotic being widely used as a way of expressing drink.
spots on the underside of the leaf and in time Already the Mundo Novo presents high size, high
will spread throughout the limbo, forming larger vigor and longevity, with average maturity and
patches of yellow-orange coloration, leading to susceptibility to coffee rust, besides presenting
death of the affected tissue (MAIA et al., 2009). great quality drink (GUERREIRO FILHO;
Chemical control is the most used way to FAZUOLI; AGUIAR, 2006). Both cultivars have
control the disease through the use of protective excellent adaptation in the producing regions of
and systemic fungicides. Fungicides chemical Brazil coffee (CARVALHO et al., 2006).
group of strobilurins, which belongs azoxystrobin, Biological control has been gaining ground
have a biochemical mode of action to inhibition in order to minimize the use of fungicides, reduce
of mitochondrial respiration. This is by blocking environmental impacts and costs in production. It is
the electron transfer in the mitochondrion site, a form of control that aims to reduce the inoculum
interfering with breathing pathogen (PARREIRA; and the activities of pathogens through natural
NEVES; ZAMBOLIM, 2009). agents that activate the plant defense system. This
Genetic improvement has already released defense is based primarily on the production and/
in the market cultivars resistant to H. vastatrix, or accumulation of some molecular compounds
which have high productivity and quality of grain, that the plant develops when under stress (COSTA;
and this would lead to decreased application of ZAMBOLIM; RODRIGUES, 2007).

1,2,3
Universidade do Oeste Paulista/UNOESTE - Rodovia Raposo Tavares, km 572 - Bairro Limoeiro, Presidente Prudente - SP
19.067-175 - [email protected], [email protected], [email protected]

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Biological control of Hemileia vastatrix ... 568

The Bacillus subtilis Cohn, is a biocontrol The severity of H. vastatrix in the coffee
agent widely used for acting antagonist manner, was evaluated after the four applications. The
releasing toxic substances that act and compete evaluations were conducted in eight reproductive
with pathogens. This species has been assessed as branches per plant, four branches on each side of
an antagonist of several pathogenic fungi under the row, in the middle third of the plants, and each
controlled conditions (FIGUEIREDO et al., 2010; branch was rated the third or fourth pair of leaves.
KUPPER; BELLOTTE; GOES, 2009). In addition, The quantification of the severity of H.
this microorganism promotes plant growth, proving vastatrix was made according to the diagrammatic
to be very versatile and efficient in disease control scale proposed by Cunha et al. (2001), establishing
and crop development (GRIGOLETTI JÚNIOR; scores from 0 to 5, with 0 (no disease), 1 (<3%
SANTOS; AUER, 2004). This is due to production severity), 2 (3 to 6% severity), 3 (6 to 12%
growth regulators and plant phytohormones in the severity), 4 (from 12 to 25% severity) and 5 (25
rhizosphere, as abscisic acid and indole acetic
to 50% severity).
acid (ARAÚJO; HENNING; HUNGRIA, 2005),
as well as solubilization of nutrients (MACHADO The collection of leaves for biochemical
et al., 2012). assessment was done concurrently with the
The objective of this study was to evaluate collection for severity assessment, 10 days after the
the efficiency of the biological control of H. last spraying. The proline concentration in leaves
vastatrix and detect some biochemical changes in analysis (µg mg-1) was performed according to the
plants of cultivars Icatu and Mundo Novo treated method described by Bates, Waldren and Teare
with foliar spray of B. subtilis and azoxystrobin (1973), in which 400 mg of fresh material was
under field conditions. taken from the middle third of the plants of each
treatment. The material was macerated in 10 mL
2 MATERIAL AND METHODS of 3% sulfosalicylic acid. Filtering material and
The experiment was carried out in the 2 mL of the supernatant obtained was performed
experimental area of Universidade do Oeste was removed and placed in a test tube with 2 mL of
Paulista of Presidente Prudente, SP, Brazil, 22° ninhydrin acid and 2 mL of glacial acetic acid. The
07’ 04” S and 51° 22’ 57” W, altitude 472 m, from samples were kept in boiling water bath for 1 hour
October 2013 to June 2014. On site is made coffee and after this period were cooled on ice. Then, the
cultivation for nine years, being the area with the reading was performed at the spectrophotometer
cultivars Mundo Novo and Icatu chosen for this at 520 nm in triplicate.
study presents infestation history of H. vastatrix. The determination of total phenols content
The experimental design was randomized blocks (µg mL-1) present in the leaves was carried out
with distribution in factorial arrangement of two using the Folin-Ciocalteu method, according to
cultivars, three treatments (bacteria, fungicide and the methodology proposed by Singleton, Orthofer
control) and four replicates, totaling 24 plots. Each and Lamuela-Raventós (1974), using gallic acid
replicate was represented by four coffee plants as standard, solution of methanol and sodium
properly identified with the name of the cultivar carbonate, in triplicate.
and treatment. For analysis of total protein, leaf samples
The control strategy was used to apply were weighed (1g) and macerated with liquid
a biologically input the base of bacteria of the nitrogen. Then put the extraction buffer and
species B. subtilis in liquid suspension, with centrifuged for 30 minutes (10.000 rpm and 4°C).
a concentration of 1,0 x 109 cells/ml and a With drew aliquots of 100 mL of the supernatant,
conventional chemical fungicide of the strobilurin from which was made the determination of protein
group and the active ingredient azoxystrobin content (mg mL-1), based on the reactant adsorption
(commercial product AMISTAR® WG). The Coomassie Brilliant Blue G-250 (BRADFORD,
control treatment received no spraying. 1976) using bovine serum albumin as standard.
Four monthly foliar sprays with products The lipid peroxidation was measured by
using backpack sprayer, constant pressure and determining the concentration of malondialdehyde
manual drive have been performed. Each spray (MDA), according to the methodology of Heath
was used dosage of 5 L of water with 50 mL of and Packer (1968), 100 mg of leaf tissue without the
bacteria and fungicide to 5 L of water to 1,25 g midrib were processed. The assay was performed in
of the product, the first spray being conducted in triplicate and the values expressed in nanomol of MDA
October 2013. by fresh weight of grass (nmol g MF-1).

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569 Cacefo, V. et al.

The data were submitted to analysis of Angonese et al. (2009) showed in a study
variance by F test by SISVAR statistical program, on the fungistatic effect of Bacillus spp., which
and when significant, the averages were compared isolates this bacterium or molecules released by
by Tukey test at 5% probability. them have potential to be used in the integrated
management of plant diseases, associated for
3 RESULTS AND DISCUSSION
example with the use of fungicides, allowing
According to Table 1, the results of the the rotation of the use of products with different
analysis of variance F were significant treatment modes of action to prevent pathogen resistance,
and cultivar in H. vastatrix. Regarding the ensuring greater security for the production
biochemical evaluations - proline, total phenols, system. This becomes possible due to the mode
total protein and MDA, all results were significant, of action antagonist of this microorganism and
including the interaction between treatment and the characteristic to stay alive for a long time
cultivar. on the plant, interfering with the attachment of
The results demonstrate that both the the fungus (LANNA FILHO; FERRO; PINHO,
biological and chemical control was effective to 2010), releasing toxic substances, such as iturin,
reduce the H. vastatrix in the coffee (Table 2). that act and compete with pathogens (ARAÚJO;
The cultivar Icatu showed greater resistance to the HENNING; HUNGRIA, 2005; FIGUEIREDO et
pathogen due to lower incidence of H. vastatrix. al., 2010).
In a study by Montes, Paulo and Fischer (2012), Tofoli et al. (2002), in a job with azoxystrobin
to evaluate the H. vastatrix in the coffee, disease in controlling of the H. vastatrix in the coffee under
severity was lower in Icatu and Obatã cultivars. induced wet conditions, showed the potential of
The cultivar Icatu is a hybrid obtained from the chemical group of strobilurin fungicides for
Coffea arabica L. and Coffea canephora Pierre the disease control under adverse conditions, in
ex A.Froehner, crossing, having inherited from addition to promoting lower levels of defoliation,
an important factor in plant maintenance and
C. canephora greater resistance to H. vastatrix
productivity of future harvests. This chemical
(BOTTCHER et al., 2012). Martins, Mendez and
group of fungicides include azoxystrobin inhibits
Alvarenga (2004), comparing agro-ecosystems
mitochondrial respiration by blocking the transfer
of Minas Gerais, Brazil, planted with resistant
of electrons in the mitochondria site, interfering
cultivars and susceptible to H. vastatrix, observed
with breathing pathogen (PARREIRA; NEVES;
that the cultivar Icatu not reached the level of ZAMBOLIM, 2009). Studying the control of the
damage of the disease, unlike other susceptible Puccinia triticina in the wheat with fungicides,
cultivars. According to Botelho et al. (2007), the Navarini and Balardin (2012) concluded that the
population of Icatu cultivars has a germplasm effective control of this disease is due to the action
of great importance in breeding programs for of strobilurins.
resistance to H. vastatrix. With respect to biochemical changes in the
The treatments performed in both cultivars coffee depending on the treatment, it was found that
presented significant amounts of H. vastatrix the leaf proline contents were larger in the Mundo
control and B. subtilis had underperformed the Novo cultivar compared to Icatu. Treatment with
control provided by the fungicide (Table 2). B. subtilis resulted in a 84% increase in proline
Treatment with fungicide showed 53% control in content in the cultivar Mundo Novo. On the other
the cultivar Icatu and 45% in the Mundo Novo, hand, for the cultivar Icatu, treatment with B.
while the bacteria provided with a control of 24% subtilis and fungicide did not differ but resulted in
and 17%, respectively. This result confirms what higher proline contents than the control (Table 3).
was reported by Dorighello (2013), that by using Praxedes, Ferreira and Gomes Filho (2009),
isolates of B. subtilis QST 713 and AP-3 in the in studies with bean under salt stress, found that
control of Phakopsora pachyrhizie in the soybean, stressed plants had higher proline accumulation in
noted a significant reduction in germination their leaves, suggesting a protective mechanism.
uredospores, and in field trials treatments with The accumulation of proline in plants occurs in
fungicides alone or associated with B. subtilis response to saline, water stress, UV radiation,
have excelled in reducing the severity of disease heavy metals, pathogenic and oxidative stress
and defoliation. (LIANG et al., 2013).

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Biological control of Hemileia vastatrix ... 570

TABLE 1 - F values analysis of variance.

F.V. H. vatatrix Proline Total phenols Total protein MDA

Treatment 52,05** 167,46** 929,71** 669,63** 20,62**
Cultivar 8,78** 787,42** 785,02** 800,23** 30,36**
Treat.XCult. 0,21 154,91** 543,79** 864,96** 41,43**
C.V. 12,36 6,80 1,25 8,06 6,41
* 5% probability ** 1% probability.

TABLE 2 - Effect of the application B. subtilis and azoxystrobin in controlling the H. vastatrix in two coffee
cultivars. Values established by grading scale.

Treatments Icatu Mundo Novo

Control 2,95 a 3,20 a
Fungicide 1,40 c 1,77 c
B. subtilis 2,22 b 2,66 b
Mean 2,19 B 2,54 A
Values followed by the same letter, in the columns and the mean line, have no significant value by Tukey test at
the 5% level of probability.

TABLE 3 - Forms of control and cultivars in coffee infested with H. vastatrix for proline content (µg mg-1) in the leaves.

Treatments Icatu Mundo Novo

Control 6,72 b 16,67 b
Fungicide 10,89 a 14,49 c
B. subtilis 9,57 a 30,73 a
Mean 9,06 B 20,63 A
Values followed by the same letter, in the columns and the mean line, have no significant value by Tukey test at
the 5% level of probability.

Generally, the function of the proline is by inoculation with rhizobacteria in sugarcane,

associated with osmotic adjustment and its action and the inoculated plants and under water stress
in regulating water stress. However, proline also increased 2,2 times the concentration of proline in
serves to balance the cellular oxidative stress in the the leaves when compared to inoculated without
chelation of metals and removal of reactive oxygen presence of stress. This information can be related
species (ROS) (MATYSIK; ALIA BHALU; to the data from this study, in which to cultivate the
MOHANTY, 2002; SZABADOS; SAVOURE, Mundo Novo, where there was a higher incidence
2010). Thus, understanding the mechanisms by of the H. vastatrix and consequently greater
which proline increases the response to biotic
stress, showing higher concentration of proline in
and abiotic stresses is of great importance for
treatment with B. subtilis.
agricultural research and genetical enhancement.
However, the Mundo Novo cultivar, despite Regarding the total phenol content in leaves,
having a higher concentration of proline in the the highest values were found in the cultivar
leaves, not found to be more resistant to H. vastatrix Mundo Novo. However, treatments with bacteria
(Table 2), indicating by this that other mechanisms and fungicides did not differ, but both cultivars
may be related to this resistance. Rampazzo (2013) responded to treatments applied in increasing
noted that the proline accumulation was affected phenolic content with the control (Table 4).

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571 Cacefo, V. et al.

TABLE 4 - Forms of control and cultivars in coffee infested with H. vastatrix for total phenols content (µg mL-1)
in the leaves.

Treatments Icatu Mundo Novo

Control 6,60 b 9,24 b
Fungicide 9,38 a 9,53 a
B. subtilis 9,53 a 9,71 a
Mean 8,50 B 9,49 A
Values followed by the same letter, in the columns and the mean line, have no significant value by Tukey test at
the 5% level of probability.

In a study of resistant soybean cultivars protein content was not a useful parameter for the
and susceptible to bacterial blight (Pseudomonas discrimination of resistant or susceptible cultivars.
savastanoi pv glycinea), Sbalcheiro (2010) held No differences were found in the leaf
the application of Bacillus spp. and found increase content of malondialdehyde (MDA) between
in phenol content in plants. Mello, Frighetto and any of the treatments in Mundo Novo, which
Valarini (2009) observed an increase in the total on average had lower MDA values compared to
phenol content in soybean leaves exposed to
Icatu. This cultivar, in turn, exhibited higher MDA
P. pachyrhizie in response to treatment with an
isolated endophytic bacteria. Phenolic compounds content when treated with fungicide whereas
are secondary metabolites produced by plants treatment with B. subtilis significantly reduced
that are not directly related to development of the this level compared to control (Table 6).
same, but with resistance to pests and diseases. The MDA is produced by peroxidation
In this defense mechanism of plants deviated of lipids of the cell membrane when the plant is
carbohydrates of the major metabolic pathway for suffering from some kind of stress (HENDGES
the secondary producing the phenols, which are et al., 2015). The MDA content is often used
protective substances, fungitoxic, antibacterial as a marker of the degree of oxidative damage
and antiviral, and inhibit the germination of fungal (OLIVEIRA; GOMES-FILHO; ENÉAS-FILHO,
spores, such as the isoflavones (SALGADO, 2010). According to Langaro et al. (2014), when
2004). the plant is treated with herbicide, the protective
Leaf contents of total protein in treatments ability of the carotenoid to act against oxidative
with fungicide and B. subtilis in the cultivar Icatu
were lower than the control (Table 5). This effect stress occurs and decreases the lipid peroxidation.
was also verified by Kuhn (2007) in the induction The reduction of the MDA from plants treated with
of resistance in beans by treatment with Bacillus B. subtilis shows the ability of this biocontrol agent
cereus, where plants had a lower protein content has to maintain the integrity of cell membranes,
in the leaves. which requires further study.
In contrast, treatment with B. subtilis was B. subtilis has been shown to be an important
that provided the greatest protein content in the option in biological control, especially of foliar
cultivar Mundo Novo. Silva et al. (2009) found that diseases. This control can be effected by direct
the inoculation of bean seeds with isolated from inhibition of the pathogen through the production
Pseudomonas DFs842 was responsible for the of inhibitory substances, and also by physiological
induction of resistance attributed to the significant changes in the metabolism of plants, resulting in
increase in concentration of total soluble proteins. activation of the defense mechanism of the same.
The increase in total protein in the treatments of
The results of this experiment can be
B. subtilis is also related to the increase in proline
found in this study. The proteins are part of the inferred that the variety Mundo Novo showed
plasma membrane and out as regulators osmosis, higher production of defense substances in
and if there is intense stress proteins break their response to the treatment used, especially with B.
plants to generate electricity or other amino acids subtilis, demonstrating with it greater potential for
accumulate to defend (ABADE et al., 2014). In plant protection when using induced resistance.
the assessment of biochemical descriptors in Bottcher et al. (2012) also found that this cultivar
cotton cultivars focusing on increasing disease had better performance in the production of
resistance, Silva et al. (2009) stated that the total defense substances compared to Icatu.

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Biological control of Hemileia vastatrix ... 572

TABLE 5 - Forms of control and cultivars in coffee infested with H. vastatrix for total protein content (mg mL-1)
in the leaves.

Treatments Icatu Mundo Novo

Control 0,07 a 0,04 c
Fungicide 0,03 b 0,07 b
B. subtilis 0,04 b 0,32 a
Mean 0,05 B 0,14 A
Values followed by the same letter, in the columns and the mean line, have no significant value by Tukey test at
the 5% level of probability.

TABLE 6 - Forms of control and cultivars in coffee infested with H. vastatrix for content of MDA (nmol g-1) in
the leaves.

Treatments Icatu Mundo Novo

Control 3,19 b 2,49 a
Fungicide 3,54 a 2,52 a
B. subtilis 2,21 c 2,73 a
Mean 2,98 A 2,58 B
Values followed by the same letter, in the columns and the mean line, have no significant value by Tukey test at
the 5% level of probability.

The use of biological control with B. ARAÚJO, F. F.; HENNING, A. A.; HUNGRIA, M.
subtilis has shown promise for its introduction Phytohormones and antibiotics produced by Bacillus
as a tactic of diseases in coffee, because beyond subtilis and their effects on seed pathogenic fungi
the control of diseases such as H. vastatrix, this and on soybean root development. World Journal of
microorganism can stimulate the plant to trigger Microbiology & Biotechnology, Dordrecht, v. 21, p.
defense mechanisms important for increased
1639-1645, 2005.
resistance. This can mitigate the damaging effects
of different pathogens culture during their cycle.
BATES, L. S.; WALDREN, R. P.; TEARE, I. D.
4 CONCLUSIONS Rapid determination of free proline for water-stress
studies. Plant and Soil, Netherlands, v. 39, n. 2, p. 205-
B. subtilis acted effectively in control of H.
vastatrix in both cultivars, but with less efficiency 207, 1973.
than the azoxystrobin. Plants treated with B.
subtilis presented greater proline, total phenols BOTELHO, C. E. et al. Avaliação de progênies de
and total proteins in their leaves on the application café obtidas por cruzamentos das cultivares de Icatu e
of azoxystrobin, with the cultivar Mundo Novo Catimor. Coffee Science, Lavras, v. 2, n. 1, p. 10-19,
being more responsive to treatments that Icatu. 2007.
5 REFERENCES
BOTTCHER, A. et al. Antioxidative responses of cell
ABADE, M. T. R. et al. Determinação dos suspension cultures of two Coffea arabica varieties to
teores de proteínas solúveis totais em plantas de low aluminum levels at pH 5.8. Hoehnea, São Paulo,
girassol (Helianthus annuus L.). In: Workshop v. 39, n. 1, p. 1-10, 2012.
Agroenergia, 8., 2014, Ribeirão Preto. Anais...
Ribeirão Preto, 2014. 1 CD-ROM. BRADFORD, M.  M. A  rapid and sensitive method
for the quantification of microgram quantities
ANGONESE, M. T. et al. Efeito fungistático de Bacillus of protein utilizing the principles of protein-dye
spp. sobre fungos fitopatogênicos. Revista Brasileira binding.  Analytical Biochemistry, New York, v.  72,
de Agroecologia, Cruz Alta, v. 4, n. 2, p. 97-100, 2009. n. 1/2, p. 248-254, 1976.

Coffee Science, Lavras, v. 11, n. 4, p. 567 - 574 out./dez. 2016

573 Cacefo, V. et al.

CARVALHO, A. M. et al. Desempenho agronômico de Revista Brasileira de Energias Renováveis, Palotina,

cultivares de café resistentes à ferrugem no Estado de v. 4, n. 2, p. 52-63, 2015.
Minas Gerais, Brasil. Bragantia, Campinas, v. 71, n. 4,
p. 481-487, 2012. KUHN, O. J. Indução de resistência em feijoeiro
(Phaseolus vulgaris) por acibenzolar-Smetil e
CARVALHO, G. R. et al. Comportamento de Bacillus cereus: aspectos fisiológicos, bioquímicos e
progênies de cafeeiro cultivar mundo novo. Ciência e parâmetros de crescimento e produção. 2007. 139 p.
Agrotecnologia, Lavras, v. 30, n. 5, p. 853-860, set./ Tese (Doutorado Fitopatologia) - Escola Superior de
out. 2006. Agricultura “Luiz de Queiroz”, Piracicaba, 2007.

COSTA, M. J. N.; ZAMBOLIM, L.; RODRIGUES, KUPPER, K. C.; BELLOTTE, J. A. M.;  GOES,
F. A. Avaliação de produtos alternativos no controle A.  Controle alternativo de Colletotrichum
da ferrugem do cafeeiro. Fitopatologia Brasileira, acutatum agente causal da queda prematura dos
Brasília, v. 32, p. 150-152, 2007. frutos cítricos. Revista Brasileira de Fruticultura,
Jaboticabal, v. 31, n. 4, p. 1004-1015, 2009.
CUNHA, R. L. et al. Desenvolvimento e validação
de uma escala diagramática para avaliar a severidade LANGARO, A. C. et al. Alterações fisiológicas
da ferrugem (Hemileia vastatrix) do cafeeiro. In: na cultura do tomateiro devido à deriva simulada
SIMPÓSIO DE PESQUISA DOS CAFÉS DO de herbicidas. Revista Brasileira de Herbicidas,
BRASIL, 2., 2001, Vitória. Resumos Expandidos... Londrina, v. 13, n. 1, p. 40-46, jan./abr. 2014.
Brasília: Embrapa Café, 2001. v. 3, p. 1101-1108.
LANNA FILHO, R.; FERRO, H. M.; PINHO, R. S.
DORIGHELLO, D. V. Controle da ferrugem asiática C. Controle biológico mediado por Bacillus subtilis.
da soja (Phakopsora packyrhizi) com óleo de café e Revista Trópica - Ciências Agrárias e Biológicas,
Bacillus spp. 2013. 45 p. Dissertação (Mestrado em Lavras, v. 4, n. 2, p. 12-20, 2010.
Agronomia) - Faculdade de Ciências Agronômicas,
Universidade Estadual Paulista, Botucatu, 2013. LIANG, X. et al. Proline mechanisms of stress survival.
Antioxidants and Redox Signaling, New York, v. 19,
FIGUEIREDO, J. E. F. et al. Atividade antagonista in n. 9, p. 998-1011, 2013.
vitro de Bacillus subtilis contra fungos fitopatogênicos
do milho e sorgo. In: CONGRESSO NACIONAL DE MACHADO, V. et al. Bactérias como agentes de
MILHO E SORGO, 28., 2010, Goiânia. Resumos controle biológico de fitonematóides. Oecologia
Expandidos... Sete Lagoas: Embrapa Milho e Australis, Rio de Janeiro, v. 12, n. 2, p. 165-182, 2012.
Sorgo, 2010. p. 667-673.
MAIA, T. A. et al. Estrutura genética da população
GRIGOLETTI JÚNIOR, A.; SANTOS, A. F. dos; de Hemileia vastatrix com base no mercado AFLP.
AUER, C. Perspectivas do uso do controle biológico In: SIMPÓSIO DE PESQUISA DOS CAFÉS DO
contra doenças florestais. Floresta, Curitiba, v. 30, n. BRASIL, 6., 2009, Vitória. Resumos Expandidos...
1/2, p. 155-165, 2004. Brasília: Embrapa Café, 2009. 1 CD-ROM.

GUERREIRO FILHO, O.; FAZUOLI, L. C.; AGUIAR, MARTINS, M.; MENDEZ, A. N. G.; ALVARENGA, M.
A. T. Cultivares de Coffea arabica selecionadas I. N. Incidência de pragas e doenças em agrossistemas
pelo IAC: características botânicas, tecnológicas, de café orgânico de agricultores familiares em Poço
agronômicas e descritores mínimos. 2006. Disponível Fundo-MG. Ciência Agrotecnologia, Lavras, v. 28, n.
em: <http://www.infobibos.com/Artigos/2006_2/ 6, p. 1306-1313, nov./dez. 2004.
Cultivares_cafe/Index.htm>. Acesso em: 1 out. 2014.
MATYSIK, J.; ALIA BHALU, B.; MOHANTY, P.
HEATH, R. L.; PACKER, L. Photoperoxidation in Molecular mechanisms of quenching of reactive
isolated chloroplast: I., kinetics an stoichiometry of oxygen species by proline under stress in plants.
fatty acid peroxidation. Archives of Biochemistry and Current Science, Bangalore, v. 82, p. 525-532, 2002.
Biophysics, New York, v. 125, n. 1, p. 189-198, 1968.
MELLO, U. Q.; FRIGHETTO, R. T. S.; VALARINI,
HENDGES, F. B. et al. Avaliação enzimática e P. J. Teor de fenólicos totais e crescimento radicular
fisiológica de plântulas de milho submetidas à seca. em soja (Glycine max (L.) Merr.) inoculada com

Coffee Science, Lavras, v. 11, n. 4, p. 567 - 574, out./dez. 2016

Biological control of Hemileia vastatrix ... 574

as bactérias endofítica e epifítica e sua exposição à relações hídricas. 2013. 54 p. Dissertação (Mestrado
ferrugem. In: CONGRESSO INTERINSTITUCIONAL em Agricultura Tropical e Subtropical) - Instituto
DE INICIAÇÃO CIENTÍFICA, 3., 2009, Campinas. Agronômico de Campinas, Campinas, 2013.
Anais... Campinas: ITAL; Jaguariúna: Embrapa
Meio Ambiente, 2009. Poster 0902019. SALGADO, P. R. Fenóis totais no cafeeiro em
razão das fases de frutificação e clima. 2004. 60
MONTES, S. M. N. M.; PAULO, E. M.; FISCHER, p. Dissertação (Mestrado em Agronomia) - Escola
I. H. Incidência da ferrugem alaranjada em cafeeiros Superior de Agricultura “Luiz de Queiroz”, Piracicaba, 2004.
nas condições climáticas de Presidente Prudente, SP.
Coffee Science, Lavras, v. 7, n. 3, p. 208-214, 2012. SBALCHEIRO, C. C. Uso de Bacillus sp. e
Acibenzolar-S-Metil como indutores de resistência
NAVARINI, L.; BALARDIN, R. S. Doenças foliares e o ao crestamento bacteriano em soja: (Pseudomonas
controle por fungicidas na produtividade e qualidade savastanoi pv. glycinea). 2010. 195 p. Tese (Doutorado
de grãos de trigo.  Summa Phytopathologica, São em Agronomia) - Universidade de Passo Fundo, Passo
Paulo, v. 38, n. 4, p. 294-299, 2012. Fundo, 2010.

OLIVEIRA, A. B. de; GOMES-FILHO, E.; ENÉAS- SILVA, E. G. et al. Alterações metabólicas em plantas
FILHO, J. O problema da salinidade na agricultura e as de feijão originadas de sementes microbiolizadas
adaptações das plantas ao estresse salino. Enciclopédia por Pseudomonas sp. e inoculadas com Xanthomonas
Biosfera, Centro Científico Conhecer, Goiânia, v. 6, n. axonopodis pv. phaseoli. Summa Phytopathologica,
11, p. 1-16, 2010. São Paulo, v. 35, n. 2, p. 98-104, 2009.

PARREIRA, D. F.; NEVES, W. S.; ZAMBOLIM, SINGLETON, V. L.; ORTHOFER, R.; LAMUELA-
L. Resistência de fungos a fungicidas inibidores de RAVENTÓS, R. M. Analysis of total phenols and other
quinona. Revista Trópica - Ciências Agrárias e oxidation substrates and antioxidants by means of
Biológicas, São Luís, v. 3, n. 2, p. 24-34, 2009. Folin- Ciocaulteu reagent. Methods in Enzymology,
Netherlands, v. 29, p. 152-178, 1974.
PRAXEDES, S. C.; FERREIRA, T. M.; GOMES
FILHO, E. Acúmulo de prolina e aminoácidos em SZABADOS, L.; SAVOURE, A. Proline: a
cultivares de feijão caupi com tolerância diferencial à multifunctional amino acid. Trends in Plant Science,
salinidade. Caatinga, Mossoró, v. 22, n. 3, p. 211-214, London, v. 15, p. 89-97, 2010.
2009.
TOFOLI, J. G. et al. Desempenho de azoxystrobin no
RAMPAZZO, P. E. Interação entre rizobactérias controle da ferrugem do cafeeiro sob condições de
e cana-de-açúcar sob diferentes condições de chuva induzida. Arquivos do Instituto Biológico, São
umidade do substrato: crescimento, fotossíntese e Paulo, v. 69, n. 1, p. 93-96, 2002.

Coffee Science, Lavras, v. 11, n. 4, p. 567 - 574 out./dez. 2016