Limosilactobacillus reuteri as sustainable biological control agent against toxigenic Fusarium verticillioides.

Corn contamination with Fusarium verticillioides (Sacc.) Nirenberg is a worldwide problem that affects yield and grain quality resulting in severe economic losses and implications for food safety. Control of F. verticillioides is a challenge, but lactic acid bacteria (LAB) has high potential as a biological control agent. In this study, the antifungal effect of Limosilactobacillus reuteri (formerly Lactobacillus reuteri) LR-92 against F. verticillioides 97L was investigated. Cell-free supernatant (CFS) from L. reuteri showed concentration-dependent fungicidal and fungistatic activity against F. verticillioides 97L. The antifungal compounds from CFS showed heat stability and pH dependence, and antifungal activity was not affected by treatment with proteolytic enzymes. High-performance liquid chromatography analysis indicated that L. reuteri LR-92 produces lactic and acetic acids. After liquid-liquid extraction, electrospray ionization mass spectrometry analysis of the active ethyl acetate fraction containing antifungal compounds revealed the production of 3-phenyllactic acid, cyclo-(L-Pro-L-Leu), cyclo-(L-Pro-L-Phe), and cyclo-(L-Phe-trans-4-OH-L-Pro). L. reuteri LR-92 has potential as a biocontrol agent for F. verticillioides and contributes to food safety.

Lactic acid bacteria and Bacillus spp. as fungal biological control agents.

Fungal pathogens are one of the most important agents affecting crop production and food safety, and agrochemical application is one of the main approaches to reduce phytopathogenic fungi contamination in agricultural products. However, excessive and inadequate use can cause environmental damage, human and animal hazard, and increased phytopathogen resistance to fungicides. Biological control using lactic acid bacteria (LAB) and Bacillus spp. is an environmentally friendly strategy for phytopathogenic fungi management. Several molecules produced by these bacteria indeed affect fungal growth and viability in different plant crops. In this article, the activity spectra are reviewed along with the antifungal effect and antifungal compounds produced by LAB (e.g. organic acids, peptides, cyclic dipeptides, fatty acids, and volatile compounds) and Bacillus spp. (e.g. peptides, enzymes, and volatile compounds).

Transgenic versus conventional corn: fate of fumonisins during industrial dry milling.

The aim of this study was to compare the fate of fumonisins in transgenic and non-transgenic corn during industrial dry milling. For this purpose, whole corn samples and their fractions (germ, pericarp, endosperm, corn meal, and grits) were collected from one of the major Brazilian milling plants, totaling 480 samples. There was no significant difference (p > 0.05) between mean fumonisin (FB1 + FB2) levels in transgenic (1130 µg/kg) and non-transgenic (920 µg/kg) whole corn. However, in non-transgenic germ, endosperm and corn meal fraction fumonisin levels were higher (2940 µg/kg, 250 µg/kg and 190 µg/kg, respectively) than in transgenic fractions (2180 µg/kg, 130 µg/kg and 85.0 µg/kg, respectively). Furthermore, the highest percentages of fumonisins were distributed in the germ, corresponding to about 87 and 76% of the total fumonisins present in the whole corn from non-transgenic and transgenic hybrids, respectively. Concerning the endosperm from non-transgenic and transgenic corn, approximately, 23% and 13% of the total fumonisins were retained after the dry milling. Further processing in corn meal (300 to 420 µm particle size) and grits (590 to 1190 µm) decreased the percentages of remaining fumonisins to 4% and 2% (transgenic) and 10% and 3% (non-transgenic corn), respectively. These results suggested that fumonisin concentration was higher in outer and inner non-transgenic fractions when compared to transgenic ones and that the fate of fumonisins during the industrial dry milling could be affected by the transgenic status. However, it was not possible to conclude that the difference was exclusively due to this variable.

Safety of Corn and Corn-Based Products Intended for Human Consumption Concerning Fumonisins from a Brazilian Processing Plant.

Brazil is one of the world's largest corn producers and is a leader in exportation. Due to intense globalization, corn may be commercialized worldwide and the issue concerning the safety of corn-based products has become a topic of widespread international interest. Dietary exposure evaluation is a relevant criterion for mycotoxin risk assessment. Thus, human exposure to fumonisins were assessed for corn grain and its derivatives (endosperm, cornmeal, and grits; n = 320) sampled from one of the large-scale corn processing plants in Brazil. The total probable daily intake (PDI) for fumonisins in Brazil was 96.9 ng kg-1 body weight day-1, which corresponds to 5% of the provisional maximum tolerable daily intake (PMTDI) of 2000 ng kg-1 b.w. day-1 for fumonisins. In countries that import Brazilian corn, the total PDI is lower in European countries (from 35.7 to 177 ng kg-1 b.w. day-1) and higher in Angola (1553 ng kg-1 b.w. day-1). Taking into account that dietary exposure in populations in Brazil and importing countries was low, the corn-based products were safe for human consumption regarding fumonisins, even for regions with high corn consumption.

Development of Indirect Competitive Enzyme-Linked Immunosorbent Assay to Detect Fusarium verticillioides in Poultry Feed Samples.

Fumonisins are a group of toxic secondary metabolites that are produced by Fusarium verticillioides which are associated with poultry health hazard and great economic losses. The objective of the present study was to develop an immunological method to detect F. verticillioides in poultry feed samples. An indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) based on a polyclonal antibody against 67 kDa protein of the F. verticillioides 97K exoantigen was developed to detect this fungus. Antibody anti-67 kDa protein showed cross-reactivity against F. graminearum (2⁻7%) and F. sporotrichioides (10%), but no or low cross-reactivity against Aspergillus sp. and Penicillium sp. exoantigens. The detection limit for the 67 kDa protein of F. verticillioides was 29 ng/mL. Eighty-one poultry feed samples were analyzed for Fusarium sp. count, 67 kDa protein of F. verticillioides and fumonisin concentrations. Eighty of the 81 feed samples (98.6%) showed Fusarium sp. contamination (mean 6.2 x 104 CFU/g). Mean 67 kDa protein and fumonisin concentration in the poultry feed samples was 21.0 µg/g and 1.02 µg/g, respectively. The concentration of 67 kDa protein, as determined by ic-ELISA correlated positively (p < 0.05) with fumonisin levels (r = 0.76). These results suggest that this ic-ELISA has potential to detect F. verticillioides and predict fumonisin contamination in poultry feed samples.

Detection of Fusarium verticillioides by PCR-ELISA based on FUM21 gene.

Fusarium verticillioides is a primary corn pathogen and fumonisin producer which is associated with toxic effects in humans and animals. The traditional methods for detection of fungal contamination based on morphological characteristics are time-consuming and show low sensitivity and specificity. Therefore, the objective of this study was to develop a PCR-ELISA based on the FUM21 gene for F. verticillioides detection. The DNA of the F. verticillioides, Fusarium sp., Aspergillus sp. and Penicillium sp. isolates was analyzed by conventional PCR and PCR-ELISA to determine the specificity. The PCR-ELISA was specific to F. verticillioides isolates, showed a 2.5 pg detection limit and was 100-fold more sensitive than conventional PCR. In corn samples inoculated with F. verticillioides conidia, the detection limit of the PCR-ELISA was 1 × 104 conidia/g and was also 100-fold more sensitive than conventional PCR. Naturally contaminated corn samples were analyzed by PCR-ELISA based on the FUM21 gene and PCR-ELISA absorbance values correlated positively (p < 0.05) with Fusarium sp. counts (CFU/g). These results suggest that the PCR-ELISA developed in this study can be useful for F. verticillioides detection in corn samples.