5. Conclusions and future perspectives

Doster et al. used A. flavus strain AF36 as a BCA to control AFs in pistachio orchards for four consecutive seasons (2008–2011) and he could diminish AFs level by 20–45% [114]. In groundnuts, more trials in vitro [61, 66] and in the field [58–60] have been done. Zhou et al. 2015 found a positive correlation between AFs reduction rate and inoculum dose while Hulikunte Mallikarjunaiah et al. 2017 measured total AFs in rhizospheric and geocarpospheric soil and groundnut seeds after he treated them with two strains isolated from India. A significant reduction of mycotoxin concentration below the maximum permissible levels for ground nuts was obtained [61]. Field trials in Argentina were designed to control AFs in groundnut. However, the author reported a high level of AFs reduction, and the results were inconsistent

High levels of AFs and CPA control in maize field were achieved after challenging two strains of A. flavus with atoxigenic strains K49 and NRRL 21882 [65]. Mauro et al. could obtain similar results in vitro after screening for local atoxigenic strains from Italy [67]. In Nigeria, a successful maize field trial exhibited the promising use of two locally isolated strains, La3279 and La3303, in controlling AFB1 and AFB2 up to 99.9% [120]. When these two strains mixed with other two strains to make a mixture applied to the soil before flowering, a similar conclusion was obtained [55] with the advantage of persistence of the

Researchers have also tested different species of Trichoderma such as T. viride, T. harzianum and T. asperellum [38, 95, 115, 116]; bacteria [84, 121, 124]; yeast [36, 174]; and algae [118] as a potential alternative BCAs to control Aspergillus spp., although not all have looked into mycotoxins (Figure 2B). Production of two volatile compounds, dimethyl trisulfide and 2,4-bis(1,1 dimethylethyl)-phenol, by Shewanella algae strain YM8 showed a 100% inhibition on aflatoxin synthesis in maize and peanuts stored at different water activities [118]. Previously, B. subtilis RCB 90 in vitro was also reported to completely inhibit AFB1 [121]. The yeast, Candida parapsilosis IP1698 was also able to inhibit aflatoxin production (90–99%) at different pH and temperatures [174]. This was also in line with the same reduction percentage obtained but with Bacillus spp. P1 and Bacillus spp. P11 [40]. Aiyaz et al. tested in the field, four BCAs and all the formulations, by maize seeds treatment application, had a significant reduction in AFs level [95].

Hundreds of BCAs have been tested against different types and strains of mycotoxigenic fungi in vitro. However, not all of them were effective against mycotoxigenic fungi under field conditions. For instance, Johansson et al. selected 164 bacterial isolates out of 600 for a field experiment to control F. culmorum infection in wheat and three strains of Fluorescent pseudomo-

In general, the difference in BCAs performance from in vivo condition to field conditions might be related to the influence of other factors present in the field such as meteorological parameters, soil characteristics, nutrient availability, microbial community which may affect the efficacy of the screened BCAs. Other important parameters which are not present in in vivo studies include the way of delivery of the BCAs to the host (spray or direct inoculation), form of

nads and a species of Pantoea gave a high level of control and consistent results [159].

between the two seasons [58, 59].

72 Mycotoxins - Impact and Management Strategies

biocontrol effect during storage.

4. From lab bench to field trials

Despite the considerable amount of research that have been done to screen and select effective BCAs to control mycotoxigenic pathogens and their mycotoxins, still there are several pitfalls for using BCAs. For instance, the broad spectrum antagonistic activity of some BCAs such as Trichoderma spp., against several pathogenic fungi may also affect other beneficial organisms present in rhizosphere [178] and this may require more research for target specific BCAs. Even though implementation of a biological control strategy is strongly recommended to replace the use of synthetic pesticides, there are several concerns regarding the biological and environmental stability of BCAs. For example, the population of A. flavus including atoxigenic strains is highly diverse. This entails that there is a risk under certain environmental conditions that atoxigenic strains outcross with toxigenic A. flavus and thereafter produce mycotoxins [26, 62]. In addition, it is not guaranteed whether the atoxigenic strains can survive for a long time and what is the short term and long term effect on the soil microenvironment.

mycotoxins "one fits for all may not be the case here" [183, 184]. To tackle this problem, maybe a combination of multiple BCAs or with fungicides could be considered. Application dose should be deeply investigated to achieve the desirable control. As in previous research, it has been shown that a suboptimal or sublethal treatment with fungicides [185] may lead to induction of mycotoxins production by the pathogen as a stress response. Searching for new BCAs with novel modes of action can assist to effectively control mycotoxigenic plant pathogens. Recently, Enterobacter spp., a root-inhabiting bacterial endophyte, was reported to have a different mode of action than those previously described through formation of physicochemical barrier that blocks the invasion of F. graminearum. However it is unclear whether this mode of action can be applied to maize and wheat [186]. Finally, the sound implantation of preharvest strategies can help in saving crop loss but does not fully ensure the safety of food as the fungal attack can also happen during storage or during processing which necessitate a post-

Biological Control of Mycotoxigenic Fungi and Their Toxins: An Update for the Pre-Harvest Approach

http://dx.doi.org/10.5772/intechopen.76342

75

This work was supported by the EU project Horizon 2020-MYCOKEY "Integrated and inno-

The authors have mentioned some trade names of certain BCAs for the scientific purpose only

\*Address all correspondence to: mohamed.fathi@ugent.be; geert.haesaert@ugent.be

1 Laboratory of Applied Mycology and Phenomics, Department of Plants and Crops, Faculty

2 Centre of Excellence in Mycotoxicology and Public Health, Department of Bioanalysis,

, Sarah De Saeger<sup>2</sup>

, Kris Audenaert<sup>1</sup> and

vative key actions for mycotoxin management in the food".

harvest control.

Acknowledgements

Conflict of interest

Other declarations

Author details

Geert Haesaert<sup>1</sup>

The authors declare no conflict of interest.

Mohamed F. Abdallah1,2\*, Maarten Ameye1

\*

and this does not reflect any recommendation for use.

of Bioscience Engineering, Ghent University, Ghent, Belgium

Faculty of Pharmaceutical Sciences, Ghent University, Ghent, Belgium

Care should be taken that besides successful control of plant pathogens, and BCAs themselves do not produce toxic substances. For instance, C. rosea secretes gliotoxin which is toxic metabolite to human. Also, it was reported that some Trichoderma strains harbor trichothecenes (Tri) genes that translate into proteins similar to Fusarium Tri proteins [179, 180]. This entails that Trichoderma spp. share the production of trichothecenes toxins (such as T-2 toxin) with Fusarium spp. In addition, gliotoxin and viridian produced by T. harzianum, T. viride and T. virens showed their phytotoxic effect by reducing seed germination rate in wheat and human toxicity [28]. Therefore, spreading such a microorganism into the environment may impose an extra burden to food safety and public health. Additionally, from the economical point of view, it is necessary to estimate the total cost of application and the need for seasonal reapplication of the BCAs, so it does not exceed costs of current practices.

Controlling mycotoxins is an important aspect in the management of mycotoxigenic pathogens, which adds an extra challenge to find an effective biocontrol agent to control the fungal growth and toxin production simultaneously. It is very well known that one fungal pathogen can produce simultaneously several unrelated mycotoxins, as an example F. graminearum produces DON and ZEN which both have two different biosynthetic pathways. The scientific research has mostly been focusing to control one type of mycotoxin. Consequently, it will be more valuable to select a single biocontrol agent able to simultaneously suppress the production of both toxins. It is crucial that the selected BCAs are tolerant to mycotoxins [169] which will guarantee the long term efficiency in the field.

Some mycotoxins can be modified by the plant through alteration of their chemical structure "i.e. conjugation to a glucose moiety and hence called plant metabolites of mycotoxins or modified or masked mycotoxins" [181]. For example, DON is transformed to deoxynivalenol-3-glucoside (DON3G) in the plant as a part of the plant defense mechanism. These masked forms of mycotoxins can be hydrolyzed back into their parent forms "DON" inside human and animal body. Therefore, it is of paramount importance to take into account the effect of biocontrol agents on the production of (masked) mycotoxins and to deeply investigate whether the efficacy of the selected BCAs is due to an actual reduction of mycotoxin content based on a direct inhibition of their production by the pathogen or due to enhancing the plant immunity which may increase the plant ability to form more DON3G as in this case the total mycotoxin content in the plant will remain unchanged. Furthermore, the underlying mechanism between the parent mycotoxin, host and BCAs remains obscure and should be further investigated. In addition, other categories of mycotoxins, however they pose health risks, are underexplored such as enniatins, emerging mycotoxins produced by Fusarium spp., [14, 182] have not been tested with BCAs and this necessitates the need for further investigation.

Different BCAs with different modes of action, formulation, treatments, application time were tested showing that it may be difficult to have a single BCA able to diminish all the regulated mycotoxins "one fits for all may not be the case here" [183, 184]. To tackle this problem, maybe a combination of multiple BCAs or with fungicides could be considered. Application dose should be deeply investigated to achieve the desirable control. As in previous research, it has been shown that a suboptimal or sublethal treatment with fungicides [185] may lead to induction of mycotoxins production by the pathogen as a stress response. Searching for new BCAs with novel modes of action can assist to effectively control mycotoxigenic plant pathogens. Recently, Enterobacter spp., a root-inhabiting bacterial endophyte, was reported to have a different mode of action than those previously described through formation of physicochemical barrier that blocks the invasion of F. graminearum. However it is unclear whether this mode of action can be applied to maize and wheat [186]. Finally, the sound implantation of preharvest strategies can help in saving crop loss but does not fully ensure the safety of food as the fungal attack can also happen during storage or during processing which necessitate a postharvest control.
