Abd-El-Kareem et al. Egyptian Journal of Biological Pest Control (2021) 31:25 Egyptian Journal of https://doi.org/10.1186/s41938-021-00371-z Biological Pest Control RESEARCH Open Access Application of Bacillus pumilus isolates for management of black rot disease in strawberry Farid Abd-El-Kareem, Ibrahim E. Elshahawy and Mahfouz M. M. Abd-Elgawad* Abstract Background: Black root rot of strawberry plants caused by Rhizoctonia solani, Fusarium solani, and Pythium sp. is a serious disease in Egypt. Biocontrol agents have frequently proved to possess paramount and safe tools against many diseases. The impact of soil treatments with 3 Bacillus pumilus isolates on black root rot disease of strawberry plants caused by R. solani, F., and Pythium sp. under laboratory and field conditions was examined herein on the commonly used ‘Festival’ strawberry cultivar. To increase the bacterial adhesion and distribution on the roots, each seedling was dipped in bacterial cell suspension at 1 × 108 colony-forming units/ml of each separate bacterial isolate for 30 min then mixed with 5% Arabic gum. Results: The tested B. pumilus isolates significantly reduced the growth area of these 3 fungi. The two bacterial isolates Nos. 2 and 3 reduced the growth area by more than 85.2, 83.6, and 89.0% for R. solani, F. solani, and Pythium sp., respectively. Likewise, the 3 bacterial isolates significantly (P ≤ 0.05) inhibited the disease under field conditions. Isolates Nos. 2 and 3 suppressed the disease incidence by 64.4 and 68.9% and disease severity by 65.3 and 67.3%, respectively. The fungicide Actamyl had effect similar to that of the 2 isolates. B. pumilus isolates significantly enhanced growth parameters and yields of strawberry plants; isolates Nos. 2 and 3 raised the yield by 66.7 and 73.3%, respectively. Conclusions: Bacillus pumilus isolates could effectively manage the black rot disease in strawberry herein. Due to the significant impact of the root rot disease on strawberry yield, B. pumilus should be further tested to manage the disease on strawberry on large scale in Egypt. Keywords: Black root rot disease, Strawberry, Biological control, Bacillus pumilus, Egypt Background the most serious and damaging fungi on strawberry are An increasing interest in strawberry (Fragaria × ananassa Rhizoctonia solani and Fusarium solani together or in Duchesne) cultivation, as a high-value crop in Egypt, ex- addition to Pythium sp. (Abdel-Sattar et al. 2008; Abd- panded its growing areas to various governorates with El-Kareem et al. 2019). Many other pathogenic fungi different soil capacities. Nevertheless, soil-borne plant were reported on strawberry in Egypt and worldwide. pathogens are found in most of the strawberry-planted Main symptoms of black root rot disease include gradual soils. They can cause too serious diseases to maintain blackening and decay of the plant root system with con- profitable strawberry yield (Abd-Elgawad 2019). Black sequent suppression in vigor and yield of the strawberry root rot of strawberry is one such a disease that is plant (Abdel-Sattar et al. 2008). The disease is more ag- caused by one or even more fungal pathogens. In Egypt, gravated in plants that suffer other stresses such as bad soil drainage and other invading pathogens; e.g., plant- * Correspondence: [email protected] parasitic nematodes (PPNs). Plant Pathology Department, National Research Centre, El-Behooth St., Dokki, Giza 12622, Egypt © The Author(s). 2021 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. Abd-El-Kareem et al. Egyptian Journal of Biological Pest Control (2021) 31:25 Page 2 of 5 Methyl bromide (MB) alone or in combination with Dextrose Agar (PDA) medium. The same technique of other pesticides has been effectively utilized as a pre- Abd-El-Kareem et al. (2019), but using other factors for plant fumigant to suppress many soil-borne pathogens, disease management, was followed where five 9-cm weeds, and PPNs in Egyptian strawberry fields (Abd- diameter Petri plates containing PDA medium were rep- Elgawad 2019). Nevertheless, this fumigant has recently licated per each treatment (fungal species) and the un- been banned worldwide for almost all cultivated crops treated check. The inhibition in mycelial growth of the including strawberry due to its hazardous residues (Nol- pathogenic fungi was then estimated via the formula ing 2016). Generally, growing dissatisfaction with the ap- stated by Pandey et al. (2000) as follows: plication of MB and other chemical pesticides has R =(C – T/C) × 100, where R = mycelial growth re- recently attracted much attention towards biocontrol of duction (%) of the pathogen, C = radial growth of the plant diseases as a safe and alternative strategy for the pathogen in control plates (cm), and T = radial growth frequently risky chemical control (Noling 2016, Abd- of the pathogen in dual culture plate (cm). Elgawad 2020). Hence, new safe and effective solutions to control black root rot of strawberry are desperately Field experiments needed (Abd-El-Kareem et al. 2019). Biocontrol agents Field experiments were carried out under field condi- included several Bacillus spp., which could not only sup- tions during 2018/19 and 2019/20 growing seasons at press the causal pathogens but also increase the yields of Eldeer village, Toukh Centre, Qalyubia Governorate, the treated plant species. One such rod-shaped Egypt, where soil is light loamy textured with natural in- endospore-forming, Gram-positive, aerobic bacteria of festation. Three plots (each of 1.2 × 5 m) were utilized as the genus Bacillus is B. pumilus. As others related to the replicates for each treatment in addition to the untreated same genus, B. pumilus can withstand a range of variable check plots. Each replicate contained 100 strawberry environmental stresses via its spores and consequently transplants. All strawberry transplants received the same adapt easily to diverse habitats. Also, it possesses the dis- production practices of fertilizers and irrigation regime. tinguished toxin gene cesB and acids that can adhere to The strawberry cultivar Festival was dipped in bacterial diverse surfaces such as their host cells (Potekhina et al. cell suspension at concentration of 1 × 108 colony- 2011). Moreover, each of two tested B. pumilus strains forming units (CFU)/ml of each separate bacterial isolate has demonstrated distinguishable toxins between them for 30 min then mixed with 5% Arabic gum to enhance (Hoult and Tuxford 1991). Mahmoud et al. (2006) ap- adhesive capacity and perfect distribution of the bio- praised Pseudomonas fluorescens and several Bacillus agent on the outer parts of the treated roots just before species as they showed a remarkable efficacy against sev- transplanting. Seedlings were soaked in just water for eral pathogenic fungi such as F. solani and R. solani on the same period to act as untreated check. The experi- peanut roots. mental layout was complete randomized block of the The objective of the present study was to evaluate the plots. effect of soil treatments with three Bacillus pumilus iso- lates on yield parameters of strawberry plants infected Assessment of disease incidence and severity by black root rot disease and on the incidence and sever- Percentages of disease incidence were evaluated 100 days ity of the disease. after transplanting as follows: Number of infected plants Methods Disease incidence% ¼ Black root rot pathogens and biocontrol agents Total number of plants  Local pathogenic isolates of R. solani, F. solani, and 100 Pythium sp. the causal agents of black root rot disease of Disease severity (DS) was recorded at the end of the B. pumilus strawberry plants and 3 isolates of as antag- experiments (5 months after transplanting) based on a Bacillus pumilus B pumilus onistic bacteria i.e., (1), . (2), 0–5 scale according to Morocko (2006) as follows: and B. pumilus (3) were supplied by the Plant Pathology Σ ðÞDisease grade  number of plants in each grade Department, National Research Centre, Giza, Egypt. Disease severity% ¼  100 Total number of plants  highest disease grade Test of the isolates against the root rot pathogens The total strawberry yield (ton/feddan) for each treat- The 3 isolates of B. pumilus (1, 2, and 3) were tested ment was recorded. against the black root rot pathogens F. solani, R. solani, and Pythium sp. via the dual culture technique using the Determination of plant growth parameters and yield method described by Estrella et al. (2007). Each bacterial Effect of the tested Bacillus pumilus isolates on fresh isolate was cultured (by streaking) at 1 cm from the edge and dry weights of the strawberry plants under field con- of a Petri plate containing freshly sterilized Potato ditions was recorded. Also, accumulated yield of Abd-El-Kareem et al. Egyptian Journal of Biological Pest Control (2021) 31:25 Page 3 of 5 Table 1 Effect of three Bacillus pumilus isolates on growth area of strawberry black root rot disease under laboratory conditions Treatment F. solani R. solani Pythium sp. Growth area (cm2) Reduction % Growth area (cm2) Reduction % Growth area (cm2) Reduction % B.