Aspergillus flavus is the main producer of aflatoxin B1, one of the most toxic contaminants of food and feed. With global warming, climate conditions have become favourable for aflatoxin contamination of agricultural products in several European countries, including Serbia. The infection of maize with A. flavus, and aflatoxin synthesis can be controlled and reduced by application of a biocontrol product based on non-toxigenic strains of A. flavus. Biological control relies on competition between atoxigenic and toxigenic strains. This is the most commonly used biological control mechanism of aflatoxin contamination in maize in countries where aflatoxins pose a significant threat. Mytoolbox Af01, a native atoxigenic A. flavus strain, was obtained from maize grown in Serbia and used to produce a biocontrol product that was applied in irrigated and non-irrigated Serbian fields during 2016 and 2017. The application of this biocontrol product reduced aflatoxin levels in maize kernels (51–83%). The biocontrol treatment had a highly significant effect of reducing total aflatoxin contamination by 73%. This study showed that aflatoxin contamination control in Serbian maize can be achieved through biological control methods using atoxigenic A. flavus strains.

Contamination of maize with aflatoxins and fumonisins is one of the major food safety concerns worldwide. Knowing the contamination in advance can help to reduce food safety risks and related health issues and economic losses. The current study aimed to develop forecasting models for the contamination of maize grown in Serbia with aflatoxins and fumonisins. An integrated modeling approach was used, linking mechanistic modeling with artificial intelligence, in particular Bayesian network (BN) modeling. Two of such combined models, i.e., the prediction model for aflatoxins (PREMA) and for fumonisins (PREFUM) in maize, were developed. Data used for developing PREMA were from 867 maize samples, collected in Serbia during the period from 2012 to 2018, of which 190 were also used for developing PREFUM. Both datasets were split randomly in a model training set and a model validation set. With corresponding geographical and meteorological data, the so-called risk indices for total aflatoxins and total fumonisins were calculated using existing mechanistic models. Subsequently, these risk indices were used as input variables for developing the BN models, together with the longitudes and latitudes of the sites at which the samples were collected and related weather data. PREMA and PREFUM were internally and externally validated, resulting in a prediction accuracy of PREMA of, respectively, 83 and 70%, and of PREFUM of 76% and 80%. The capability of PREMA and PREFUM for predicting aflatoxins and fumonisins contamination using data from the early maize growth stages only was explored as well, and promising results were obtained. The integrated approach combining two different modeling techniques, as developed in the current study, was able to overcome the obstacles of unbalanced data and deficiency of the datasets, which are often seen in historical observational data from the food safety domain. The models provide predictions for mycotoxin contamination at the field level; this information can assist stakeholders of the maize supply chain, including farmers, buyers/collectors, and food safety authorities, to take timely decisions for improved mycotoxin control. The developed models can be further validated by applying them into practice, and they can be extended to other European maize growing areas.

Biochar is considered one of the most promising long-term solutions for soil quality improvement, representing an ideal environment for microorganisms’ immobilization. Hence there is a possibility to design microbial products formulated using biochar as a solid carrier. The present study was aimed at development and characterization of Bacillus-loaded biochar to be applied as a soil amendment. The producing microorganism Bacillus sp. BioSol021 was evaluated in terms of plant growth promotion traits, indicating significant potential for production of hydrolytic enzymes, indole acetic acid (IAA) and surfactin and positive tests for ammonia and 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase production. Soybean biochar was characterised in terms of physicochemical properties to evaluate its suitability for agricultural applications. The experimental plan for Bacillus sp. BioSol021 immobilisation to biochar included variation of biochar concentration in cultivation broth and adhesion time, while the soil amendment effectiveness was evaluated during maize germination. The best results in terms of maize seed germination and seedling growth promotion were achieved by applying 5% of biochar during the 48 h immobilisation procedure. Germination percentage, root and shoot length and seed vigour index were significantly improved when using Bacillus-biochar soil amendment compared to separate treatments including biochar and Bacillus sp. BioSol021 cultivation broth. The results indicated the synergistic effect of producing microorganism and biochar on maize seed germination and seedling growth promotion, pointing out the promising potential of this proposed multi-beneficial solution for application in agricultural practices.

Maize is one of the leading export products in the Republic of Serbia. As a country where economic development depends on agriculture, maize production plays a critical role as a crop of strategic importance. Potential aflatoxin contamination of maize poses a risk to food and feed safety and tremendous economic losses. No aflatoxin contamination of maize samples harvested in 2019 and 2020 in different localities in the Republic of Serbia was detected by the Enzyme-Linked Immunosorbent Assay (ELISA) test and High-Performance Liquid Chromatography (HPLC) method. On the other hand, the Cluster Amplification Patterns (CAP) analyses of the isolated Aspergillus flavus strains from 2019 maize samples confirmed the presence of key biosynthesis genes responsible for aflatoxin production. Artificial inoculation and subsequent HPLC analysis of the inoculated maize samples confirmed the high capacity of the A. flavus strains for aflatoxin production, pointing to a high risk of contamination under favorable conditions. Prevention of aflatoxin contamination is primarily based on A. flavus control, where biocontrol agents play a significant role as sustainable disease management tools. In this study, antagonistic activity screening of the novel strains belonging to the Bacillus genus indicated superior suppression of A. flavus strains by two Bacillus strains isolated from the rhizosphere of Phaseolus vulgaris.

Pectobacterium is a diverse genus which comprises of multiple destructive bacterial species which cause soft rot/blackleg/wilt disease complex in a wide variety of crops by employing high levels of virulence factors. During the 2018, 2019 and 2020 potato growing seasons, numerous outbreaks of bacterial wilt, stem blackleg and tuber soft rot were recorded, and symptomatic plant samples from ten localities in the Province of Vojvodina (Serbia) were collected and analysed. Bacterial soft-rot pathogens were detected in 63 samples using genus and species-specific primers. Through 16S rRNA Sanger sequencing of 19 representative isolates, the identity of P. brasiliense (73.7%), P. punjabense (15.8%), and P. carotovorum (10.5%) species were revealed. To further validate the identification, genotypic profiling of Pectobacterium strains using rep-PCR (ERIC, BOX, REP) was conducted for 25 selected isolates and the phylogenetic assessment based on four selected housekeeping genes (gyrA, recA, rpoA, and rpoS). Physiological and biochemical properties were analysed using basic microbiological tests and VITEK^® 2 GN card, and pathogenicity was confirmed on cv. VR808 and cv. Desiree potato tubers and plants. This study confirmed the distinctiveness of the newly described P. punjabense in Serbia as well as the high diversity of Pectobacterium brasiliense and Pectobacterium carotovorum species in Serbia.

Aflatoxin contamination is a global risk and a concerning problem threatening food safety. The biotechnological answer lies in the production of biocontrol agents that are effective against aflatoxins producers. In addition to their biocontrol effect, microbial-based products are recognized as efficient biosolutions for plant nutrition and growth promotion. The present study addresses the characterization of the representative of Phaseolus vulgaris rhizosphere microbiome, Bacillus sp. BioSol021, regarding plant growth promotion traits, including the activity of protease, cellulase, xylanase, and pectinase with the enzymatic activity index values 1.06, 2.04, 2.41, and 3.51, respectively. The potential for the wider commercialization of this kind of product is determined by the possibility of developing a scalable bioprocess solution suitable for technology transfer to an industrial scale. Therefore, the study addresses one of the most challenging steps in bioprocess development, including the production scale-up from the Erlenmeyer flask to the laboratory bioreactor. The results indicated the influence of the key bioprocess parameters on the dual mechanism of action of biocontrol effects against the aflatoxigenic Aspergillus flavus, as well on maize seed germination activity, pointing out the positive impact of high aeration intensity and agitation rate, resulting in inhibition zone diameters of 60 mm, a root length 96 mm, and a shoot length 27 mm.