Sensory quality of coffee beverage produced thereof linked to the inhibition of molds growth and ochratoxin a removal from coffee cherries using lactobacillus plantarum strains

Guézéré Corinne Beugre, Adobi Christian Kadjo, Konan Mathurin Yao, Koumba Maî Koné, Isabelle Piro-Metayer, Charlie Poss, Noël Durand, Angélique Fontana, Tagro Simplice Guehi

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Guézéré Corinne Beugre, Adobi Christian Kadjo, Konan Mathurin Yao, Koumba Maî Koné, Isabelle Piro-Metayer, et al.. Sensory quality of coffee beverage produced thereof linked to the inhibition of molds growth and ochratoxin a removal from coffee cherries using lactobacillus plantarum strains. Current Journal of Applied Science and Technology, 2023, 42 (13), pp.10-20. 10.9734/CJAST/2023/v42i134112. hal-05180650

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# Sensory Quality of Coffee Beverage Produced Thereof Linked to the Inhibition of Molds Growth and Ochratoxin a Removal from Coffee Cherries Using Lactobacillus Plantarum Strains

Guézéré Corinne Beugre<sup>a</sup>, Adobi Christian Kadjo<sup>a</sup>, Konan Mathurin Yao<sup>b</sup>, Koumba Maï Kone<sup>c</sup>, Isabelle Piro-Metayer<sup>d,e</sup>, Charlie Poss<sup>d,e</sup>, Noël Durand<sup>d,e</sup>, Angélique Fontana<sup>d,e</sup> and Tagro Simplice Guehi<sup>a*</sup>

## Authors' contributions

This work was carried out in collaboration among all authors. Author GCB is the Ph'D student who conducted this research work for her thesis. Author ACK assisted author GCB during the collection of coffee cherry samples in producing regions. Author KMK as a specialist in statistics has supervised the statistical analysis. Authors KMY and IPM contributed to the microbial analysis. Author CP have conducted all molecular identification of microorganisms. Author ND have supervised the Ochratoxin A analysis. Author AF have facilitated the course and the scientific stay of author GCB at UMR Qualisud in Montpellier, France. She contributed also to the correction of the manuscript. Author TSG is a supervisor of author GCB, has contributed to the correction of the manuscript of this article. All authors read and approved the final manuscript.

<footnote><sup>a</sup> Laboratoire de Biotechnologie et Microbiologie Alimentaire, Unité de formation et de Recherche des Sciences et Technologies des Aliments, Université Nangui ABROGOUA, 02 Bp 801 Abidjan 02, Côte d'Ivoire.</footnote>

<footnote><sup>b</sup> Laboratoire de Biotechnologie et Valorisation des Agro-Ressources, Unité de Formation et de Recherche des Sciences Biologiques, Université Péléforo Gon COULIBALY de Korhogo, BP 1328 Korhogo, Côte d'Ivoire.</footnote>

<footnote><sup>c</sup> Institut National Polytechnique Félix Houphouët-Boigny, Bp 1093, Yamoussoukro, Côte d'Ivoire.</footnote>

<footnote><sup>d</sup> CIRAD, UMR Qualisud, TA B 96/16, 75 Av. JF Breton, 34398 Montpellier Cedex 5, France.</footnote>

<footnote><sup>e</sup> Qualisud, Univ Montpellier, CIRAD, Université d'Avignon, Université de la Réunion, Montpellier SupAgro, Montpellier, 1101, Avenue Agropolis 34090, Montpellier, France.</footnote>
## Article Information

DOI: 10.9734/CJAST/2023/v42i134112

**Open Peer Review History:**

This journal follows the Advanced Open Peer Review policy. Identity of the Reviewers, Editor(s) and additional Reviewers, peer review comments, different versions of the manuscript, comments of the editors, etc are available here: https://www.sdiarticle5.com/review-history/100341

**Original Research Article**

Received: 18/03/2023

Accepted: 23/05/2023

Published: 31/05/2023

## ABSTRACT

**Aims:** Mold contamination of foods especially by mycotoxin producing fungi is not only a global food quality concern for food manufacturers, but it also constitutes a high risk for human and animal health resulting in massive economic losses globally. This study investigated the effect of Lactic Acid Bacteria (LAB) on the growth of *Aspergillus carbonarius* strains and their production of ochratoxin A (OTA).

**Methodology:** Seven fresh coffee cherry and 9 dry coffee cherry samples were collected from Man, Daloa and Akoupé 3 main coffee producing regions in Côte d'Ivoire. LAB were isolated from fresh coffee cherries while mold strains were from both fresh and dry coffee cherry. The inhibitory effect against mold growth and the ability for OTA removal of selected LAB strains were tested successively in vitro and then during coffee cherry primary postharvest processing before evaluating their influence on sensory quality of beverage. OTA production ability of molds strains are studied using both solid (CYA) and liquid (CYB) Cazapeck Yeast medium.

**Results:** About 34 fungal isolates belonging to *Aspergillus* and *Penicillium* genus were studied for their OTA production using the agar plug technique and HPLC-FD. Five *A. carbonarius* strains were capable of OTA production between 15.9 and 83mg.kg⁻¹. Out of seven isolates of *Lactobacillus plantarum*, two were successful in inhibition of mycelial growth produced fungicidal activity; five were successful in retarding it produced fungistatical activity All of *L. plantarum* isolates exhibited OTA reduction ability at about 99 %. The inoculation of two highest anti-ochratoxigenic LAB to fermenting coffee cherries resulted in great inhibition of mold growth and OTA contents reduction varying from 63.2 to 82.2%. The addition of LAB to coffee cherries did not influence the sensory attributes of the beverages produced thereof.

**Conclusion:** This study highlighted that LAB are very promising biological candidates for reduction of mold contamination and removal OTA from coffee cherry during primary postharvest processing.

**Keywords:** Coffee sherry; biocontrol; lactic acid bacteria; mold growth; ochratoxin A; sensory quality.

## 1. INTRODUCTION

Coffee grows in over 85 countries through Latino America, Asia and Africa [1]. Various foodstuffs and beverages including coffee cherry and their products are commonly exposed to ochratoxin A (OTA) [2]. Coffee has been reported as a crop currently contaminated by OTA [3] at different stages: immature, mature and overripe cherries from trees, overripe cherries from the ground and beans during drying and storage on the farm [4]. OTA is a secondary metabolite produced mainly by molds belonging to *Penicillium* [5] and *Aspergillus* [6] genus. Unfortunately, removal of

OTA from foodstuffs is particularly difficult [7] because OTA has been reported to be resistant to acidity and high temperatures. The destruction of OTA was not complete when exposed 3 hours to high pressure steam sterilization of 121°C and even at 250°C [8]. Some studies highlighted that roasting decreased partially OTA content in coffee beans [9]. OTA is still considered as nephrotoxic, carcinogenic, embryotoxic and teratogenic metabolite [10]. Many countries have statutory limits for OTA, and concentrations need to be reduced to as low as technologically possible in foodstuffs. The most important measures which have been taken to control OTA
were preventive in order to avoid fungal growth and OTA production. However, these measures including chemical and physical methods are difficult to implement in all cases with the consequence of OTA remaining in crops [11]. Nowadays, biological control is more and more recommended as a prophylactic control on several foodstuffs [12]. Antagonist microorganisms or their products can inhibit or destroy undesired microorganisms in food and agricultural products, particularly mycotoxinogenic molds [13]. Among the microorganisms, lactic acid bacteria (LAB) have been considered to be promising natural biological antagonists for mycotoxigenic mold growth in various agricultural commodities [14]. The main mechanism involved in antimicrobial efficiency of LAB is the production of organic acids, antagonistic compounds and competition for nutrients [15]. LAB have been reported to have a reliable ability to inhibit mycelia growth of different species [16]. So they could be used as the best alternative for reduction of pre/post-harvest mold infections [17]. *A. carbonarius* is reported to be a greater OTA-producer in coffee cherry [18]. Although Côte d'Ivoire is one of the greatest coffee producing countries, coffee cherry sourced from this country are currently exposed to the high OTA level [19]. In addition, the ability of the LAB strains to inhibit the growth of *A. carbonarius* and to reduce OTA content in coffee cherry and the influence of the addition of these microorganisms on the sensory quality of coffee beverage produced thereof have not yet been studied. This work aimed to investigate the ability of LABs for reduction of mold growth and elimination of ochratoxin A in coffee cherries during the primary post-harvest processing and to evaluate the effect of inoculation of LAB to the raw coffee beans on the sensory quality of coffee beverage produced thereof.

# 2. MATERIALS AND METHODS

## 2.1 Sampling of Coffee Cherries

Nine samples of 1-5 kg fresh coffee cherries (robusta *Coffea canephora*) were harvested directly from coffee trees and seven samples of dry cherries stocks were collected from Akoupé, Daloa, Man; 3 main coffee producing region of Côte d'Ivoire in January 2018. All coffee cherry samples were stored 5 hours at -20°C until further use.

## 2.2 Isolation and Identification of Microorganisms

LAB were isolated from 10 g of fresh coffee fruit per sample blended with 90 mL of sterilized peptone water diluted to 10⁻⁶ [20]. Molds were isolated by direct plating of five coffee cherries per sample on PDA medium, pH 3.5 by addition of tartric acid solution (0.1N) in order to inhibit the bacterial growth and then incubated (25°C, 3 days). The morphological characteristics of the mycelia and conidia were used [20] to identify *Aspergillus* and *Penicillium*. Bacterial DNA was extracted using thermal shock heating to 100°C for 10 min, then rapid cooling to -80°C for 10 min. The 16S rRNA genes of DNA presumptive LAB strain were amplified using specific primers as indicated by Sebastian et al. [21]. The genomic DNA of presumptive OTA producing mold was extracted as described previously by Atoui et al. [22]. The β-tubulin gene of DNA was amplified with specific primers as described previously [23].

## 2.3 Ochratoxin A (OTA) Production

Molds identified as belonging to *Aspergillus* section *Nigri* were investigated for OTA production. Conidia suspensions (10⁵ conidia.mL⁻¹) were prepared from sporulating fungal cultures [24]. This suspension (1mL) was spread on former Czapeck yeast Agar (CYA) medium added to 25 mL of Czapeck yeast broth (CYB). Both cultures were incubated at 25°C, 3 days but liquid cultures were done with shaking. OTA contents were evaluated from 4 agar plugs of about 5 mm diameter taken around of fungal [25] or from 5mL of CYA.OTA was extracted for 20 min using an ultrasonic bath and filtrated by appropriated method [26]. Detection of OTA was performed by HPLC (Shimadzu LC-10 ADVP, Japan) using fluorimetric detector (Shimadzu RF20A, Japan) according to the method previously described by Kedjebo et al. [6].

## 2.4 Inoculation of Antifungal LAB to Coffee Cherries

Fresh coffee cherries sample was divided into 14 fractions of 2.5 kg. The first fraction inoculated with 200 mL of sterile distilled water was considered as the negative control. Two hundred milliliters of LAB D12 (4×7 10⁸ UFC.g⁻¹) and LAB D13 (3×7 10⁸ UFC.g⁻¹) were applied individually to the coffee fruits of fractions 2 and 3. The coffee cherries of fractions 4 and 5 were inoculated with a mixture of 100 mL of conidial
suspension (4×10⁷ conidia.g⁻¹) of OTA producing mold strain (AcA41) and 100 mL of LAB D12 and LAB D13 cells suspension respectively. Fraction 5 was inoculated with a mixture of 100 mL of conidial suspension of OTA producing mold (AcA41) and 100 mL of LAB D13 cells suspension. Fraction 6 was inoculated with 200 mL of only conidial suspension of mold strain AcA41. Fraction 7 was not inoculated (paysant control). All experiments of inoculation were duplicated. The different inoculated coffee cherries were incubated for 16 hours (overnight) at ambient temperature and sun-dried on a plastic tarpaulin for 11 days.

## 2.5 Determination of OTA Contents of Inoculated Coffee Cherries

Dried coffee cherries (1 kg) of each fraction were weighted and then dehusked. Green coffee beans (100 g) per fraction were frozen at -80 °C for 2 hours for grinding. Ground coffee (10 g) were collected from each sample and added to 100 mL of specific solvent (methanol + 3% sodium bicarbonate solution, 50+50, v/v). The different suspensions obtained were mechanically shaken (300 rpm, 30 min) and then centrifuged (6000 rpm, 10 min and 25°C). Each extract (25 mL) was purified for extraction of OTA with an immuno-affinity column (Ochraprep®, R-Biopharm, France). Final different eluates were added to 1 mL of the OTA mobile phase (purified water + methanol + glacial acetic acid, 30:69:1, v/v/v) for the OTA quantification using previous HPLC-FLD method [6].

## 2.6 Analysis of Beverage Sensory Quality

Three green coffee fractions containing OTA amount below 8 µg.kg⁻¹ were sampled for sensory analysis. The coffee beverage was prepared by brewing 50 g of roasted coffee in 1 L of water for 5 min as previously described by Sanchez and Chambers [27]. The beverages were prepared using 50 g of roasted coffee beans, which ground in 1000 mL of filtered water (pH 7). The cup quality of the coffee beans samples was assessed twice by 8 expert tasters using 7 sensory criteria: aroma (intensity and quality), acidity, sourness, body, astringency, bitterness and global quality [28]. A hedonic assessment was carried out when the beverage temperature reached 55 °C. Scoring was on a scale of 0 to 10, where a score of 0 corresponded to the total absence of the criterion in the coffee [29].

## 2.7 Statistical Analyses

Statistica software (XLSTAT, USA 2022) was used to perform all statistical analyses. Data were expressed as mean ± standard deviation. Following ANOVA, the sensory and volatile compound values were compared by Tukey test (p< 0.05) [28]. For the sensory analysis of the coffee drinks, the results were analysed with the XLSTAT 2022.1.2.1274 software (Fisher LSD test at the 5%). Statistical differences with a probability of less than 0.001 (p < 0.001) are considered significant and those with a probability of more than 0.001 (p > 0.001) are not significant.

# 3. RESULTS

## 3.1 Isolation and Identification of Detected Microorganisms

Sixteen LAB isolates presented common morphological and biochemical characteristics of LAB (results not showed). Molecular identification showed that eleven isolates (75 %) were *Lactobacillus plantarum*, 2 isolates were *Weissella paramesenteroides* (12.5 %), 1 isolate was *W. confusa* (6.3 %) and 1 unidentified isolate (Table 1). Mycological study revealed that 34 molds strains dominated by isolates belonging to genus *Aspergillus* were found in coffee cherries samples. Twenty-three mold strains (67.6 %) belonged to *Aspergillus* section *Nigri* while 4 isolates (11.8 %) were *Aspergillus* section *Fumigati*, 4 isolates were *Rhizopussp.* (11.8 %) and 2 isolates were *Penicilliumsp.* strains (5.9 %).

## 3.2 OTA Production Ability of *Aspergillus* Section *Nigri* Isolates

Mycotoxin analysis revealed that all 5 mold isolates produced OTA greater than other isolated mold strains belonged to *Aspergillus* section *Nigri*. These mold isolates were identified as *A. carbonarius* strains. They produced OTA quantities ranging from 15.9 to 83.0mg.kg⁻¹ of CYA medium and from 4.9 to 75.8 ng.mL⁻¹ in CYB medium (Table 2).

## 3.3 Inhibition of *A. carbonarius* Growth by Cells of LAB Strain

The results of assay showed that 10 LAB strains exhibited antifungal activities. Among them, 3 *L. plantarum* coded M24, D20 and D23 had low rate inhibition against mold growth less than 20 %. Four *L. plantarum* coded D13, D31, D32 and D10
showed antifungal activity ranged between 20 to 40 %. Two *L. plantarum* (D24 and D12) and one *Weissella confusa* (M21) showed high rates inhibition over than 70 % against mold growth. However, 2 *Weissella paramesenteroides* (M31 and M33) and four *L. plantarum* (A11, M20, A10 and A12) had no inhibitor effect on *A. carbonarius* AcD64 growth (Table 3).

## 3.4 Effect of Antifungal LAB Strains on OTA of Postharvest Processed Coffee Cherries

The results about the effect of LAB strains D12 and D13 addition to coffee cherries showed that OTA contents were reduced from 6.46 (control) to 1.15 and to 2.38 µg.kg⁻¹ in green coffee beans respectively. The OTA reduction rates were 82.2 and 63.2 % for LAB strains D12 and D13 respectively. In addition, inoculation of OTA producing *A. carbonarius* AcA41 promoted the production of OTA content reached about 20 µg.kg⁻¹. However, LAB strain D12 and LAB strain D13 reduced OTA content three times and stimulated OTA production from 19.95 to 21.6 µg.kg⁻¹ in green coffee beans respectively when they were individually co-inoculated with *A. carbonarius* AcA41 (Fig. 1).

Changes in average OTA content measured in coffee cherries inoculated by antifungal LAB strains D12 and D13 co-inoculated with *A. carbonarius* AcA41 during 11 days on the farm. Data points are mean values of two replicates ± SE.Data with different letters are significantly different (One-way ANOVA, Tukey Test, p-value < 0.05).

## 3.5 Sensory Attributes of Coffee Beverage Linked to the Inoculation of OTA Reducer LAB Strains

Fig. 2 presents the sensory attributes of the coffee beverages made from the detoxified coffee beans samples in OTA. The results showed that the coffee beverage from the coffee cherries (farmer's control) recorded most intense coffee flavour with the score of 6.37. Both the beverages made from coffee cherries inoculated with LAB strains D12 or D13 recorded the score about 5. However, no significant difference (p < 0.05) was observed at the 5% level about the attributes such as "acidity", "bitterness", "astringency", "body in the cup", "sourness" and "overall quality" between all analyzed coffee beverages.

## 4. DISCUSSION

Sixteen LAB and 34 fungal strains were isolated from coffee cherries collected from different areas of Côte d'Ivoire. *Lactobacillus* species dominated the bacterial microbiota with 75% of isolates. LAB isolates were lower than those found in Taiwanese coffee cherries [30]. These differences could be ascribed to the impact of various factors including climatic factors, altitude where the coffee farm located, genotype of coffee and post-harvest storage dry coffee cherries. Also, the aerobic conditions as well as the low moisture content could cause the low contamination level of coffee cherries by LAB [31]. Some *Lactobacillus plantarum* strains were frequently reported to be predominant species among LAB microbiota found in coffee cherries source from Taiwan. The fungal strains isolated from our tested coffee cherry belonged mainly to *Aspergillus*, *Penicillium* and *Rhyzopus* genera. Our results are similar to those found in coffee beans [30]. The presence of these fungi could be due to the conditions promoting spoilage of coffee cherries before harvesting and to the fungal contamination during primary postharvest processing [32]. Pre-harvest fungal invasions were mainly due to the interactions between coffee plants and other organisms such as insects. While post-harvest fungal invasions were caused by nutrient availability, temperature, humidity and biotic factors [33]. Proliferation of mold in coffee cherry could be also due to bad post-harvest practices [34]. Our results are different from those found by Vale et al. [31]. indicated that *Aspergillus*, *Penicillium*, *Fusarium*, *Cladosporium* as main genera and by Martinez et al. [35] concluded that *Aspergillus*, *Penicillium*, *Fusarium* and *Rhizopus* genera were dominated. However, *Aspergillus*, *Penicillium*, *Fusarium* and *Cladosporium* were the most common fungal genera contaminated Ivoirian coffee cherries [19,34]. Our results showed that molds species belonged to *Aspergillus* section *Nigri* were predominant. These results are in agreement with those found by Lu et al. [32] and highlighted some risks for OTA production. Most of our *L. plantarum* isolates (9) were exhibited ability varying from 18 to 78% for the reduction of fungal growth. So, *L. plantarum* had a greater negative impact on the fungal growth than other LAB species. Our observations are in agreement with those made by Møller et al. [36]. Moreover, Shehata et al. [37]. have concluded that this may be due to the production of metabolites as well as toxicity of the compounds. Furthermore,
Møller et al. [36] evoked bacteriocin-forming ability of the LAB strains to explain the inhibition or reduction of molds growth. Dong et al. [38] reported that the antifungal activity of LAB is expressed either directly through competition of live bacterial cells for growth nutrients. The interactions between LAB and *Aspergillus carbonarius* can cause over-fermentation of coffee cherries, induce undesirable flavours and produce OTA [39]. *A. carbonarius* strains found in coffee cherries were able to produce OTA at concentration between 15.9 and 83.0mg.kg⁻¹ of solid medium and between 4.9 and 75.8 ng.mL⁻¹ in liquid medium respectively. Our isolated *A. carbonarius* strains produced OTA in contrary to Martins et al. [40] who have observed that all *A. carbonarius* strains did not produce OTA. Changes in OTA production abilities of *A. carbonarius* could be explained by many factors such as the strain, the nutrients concentration, the availability space for the mycelial growth and the physico-chemical conditions such as temperature and pH [41]. Moreover, some microbial strains were reported to be able to utilize the OTA as a source of carbon in the case of lack of organic nutrients [42]. A total reduction of OTA production of tested *A. carbonarius* strains by the cells of all tested *L. plantarum* strains varied from round 92 to about 99.9 %. Our study hihglighted that many *L. plantarum* species exhibited OTA removal ability as previously obtained by Luz et al. [43]. The individual inoculation of two previously greater inhibitors of mold growth as *L.plantarum* D12 and D13 to coffee cherries showed high OTA

reduction in dry green coffee beans of 82.2 and 63.2% respectively. This OTA reduction could be due to the ability of *L. plantarum* for adherence to the surface of coffee cherries [44]. The high OTA levels detected in inoculated coffee cherries with OTA producing *A. carbonarius* strains showed that production of OTA could be due to the previous contamination of coffee cherries by mycotoxigenic fungi [45]. However, the OTA-producing fungal contaminants of *Arabica* and *Robusta* coffee in Phillipines are *A. niger* and *A. ochraceus* [46]. Other study reported that *A. niger* and *A. carbonarius* are the main OTA producing molds found in coffee cherries [47]. OTA was reported to be produced during both preharvest [22] and post-harvest processing of coffee cherries [48]. The inoculation of antifungal LAB strains to coffee cherries reduced strongly the OTA contents. These results could be explained by the production of various compounds with antifungal effects which could damage fungal hyphae and conidia by LAB [49]. Furthermore, the low OTA levels found in inoculated coffee beans confirm that the tested LAB have strong OTA removal ability as showed by Del Prete et al. [50]. The results could be due the ability of some bacterial metabolites produced at high concentrations to lyse of OTA-producing molds cells [44] and to disrupt their functionality [51]. The results showed also that *L. plantarum* D12 reduced OTA at 39.6% in coffee beans. This could be explained by the higher ability to adsorb and/or sequester OTA [52] or use it as a carbon by cells of LAB source for the their growth [43].

<table><caption>Table 1. LAB isolates from fresh coffee fruits identified using molecular technique</caption><thead><tr><th>Codes of isolates</th><th>Bacterial species</th><th>% Similarity</th><th>Query coverage</th><th>Accession number</th></tr></thead><tbody><tr><td>D12</td><td rowspan="10">Lactobacillus plantarum</td><td rowspan="10">100</td><td rowspan="10">100</td><td>MT322914</td></tr><tr><td>D13</td><td>CP017066</td></tr><tr><td>D20</td><td>MN636335</td></tr><tr><td>D23</td><td>MN602940</td></tr><tr><td>A11</td><td>CP046262</td></tr><tr><td>A12</td><td>LC512751</td></tr><tr><td>D24</td><td>CP021929</td></tr><tr><td>M20</td><td>99.5</td><td>99.5</td><td>CP046262</td></tr><tr><td>M24</td><td>99.6</td><td>100</td><td>EF439684</td></tr><tr><td>D31</td><td>99.6</td><td>100</td><td>MN602939</td></tr><tr><td>A10</td><td>99.6</td><td>99.6</td><td>MN700260</td></tr><tr><td>D10</td><td>99.6</td><td>99.3</td><td>MN700260</td></tr><tr><td>M31</td><td rowspan="2">Weissella paramesenteroides</td><td rowspan="2">100</td><td rowspan="2">100</td><td>MH845061</td></tr><tr><td>M33</td><td>CP017066</td></tr><tr><td>M21</td><td>W. confusa</td><td>100</td><td>100</td><td>LC506181</td></tr><tr><td>D32</td><td>Non identified LAB</td><td>-</td><td>-</td><td>-</td></tr></tbody></table>
<table><caption>Table 2. OTA amount produced by *A. carbonarius* isolates from dry coffee cherries on different culture media. Data points are mean values of two replicates ± SE</caption><thead><tr><th rowspan="2">Code of A.<br>carbonarius isolates</th><th rowspan="2">Similarity<br>(%)</th><th rowspan="2">Query<br>coverage (%)</th><th rowspan="2">Accession<br>number</th><th colspan="2">OTA quantity</th></tr><tr><th>PDA (mg.kg<sup>-1</sup>)</th><th>CYB (ng.mL<sup>-1</sup>)</th></tr></thead><tbody><tr><td>AcA41</td><td>100</td><td>100</td><td>GU296700</td><td>83.1 ± 0,9<sup>a</sup></td><td>25.5 ± 0,8<sup>b</sup></td></tr><tr><td>AcA42</td><td>100</td><td>100</td><td>KC520549</td><td>78.8 ± 7,6<sup>a</sup></td><td>26.0 ± 2,7<sup>b</sup></td></tr><tr><td>AcD61</td><td>99,6</td><td>100</td><td>KP259287</td><td>15.9 ± 0,5<sup>c</sup></td><td>4.9 ± 0,2<sup>c</sup></td></tr><tr><td>AcD63</td><td>100</td><td>97</td><td>KC520550</td><td>54.0 ± 3,6<sup>b</sup></td><td>75.8 ± 1,7<sup>a</sup></td></tr><tr><td>AcD64</td><td>100</td><td>98</td><td>MG701891</td><td>17.4 ± 1,2<sup>c</sup></td><td>6.4 ± 0,1<sup>c</sup></td></tr></tbody></table>

In a column, the values of OTA quantity assigned to the same alphabetical letter indicated no significant difference at $p \le 0.05$

<table><caption>Table 3. Comparative antifungal activity of 16 LAB strains against *Aspergillus carbonarius* growth PDA medium for 48h at 30°C</caption><thead><tr><th>LAB strains</th><th>Reduction in growth of A. carbonarius growth (%)</th></tr></thead><tbody><tr><td>L. plantarum (D12)</td><td>76.4 ± 2.6<sup>a</sup></td></tr><tr><td>L. plantarum (D13)</td><td>2.6 ± 1.0<sup>bc</sup></td></tr><tr><td>L. plantarum (D24)</td><td>78.7 ± 3.5<sup>a</sup></td></tr><tr><td>Weissella paramesenteroides (M33)</td><td>0.0</td></tr><tr><td>L. plantarum (M20)</td><td>0.0</td></tr><tr><td>L. plantarum (M24)</td><td>16.9 ± 0.6<sup>c</sup></td></tr><tr><td>W. confusa (M21)</td><td>76.4 ± 2.0<sup>a</sup></td></tr><tr><td>L. plantarum (D31)</td><td>29.6 ± 0.6<sup>b</sup></td></tr><tr><td>L. plantarum (D23)</td><td>17.6 ± 0.6<sup>c</sup></td></tr><tr><td>L. plantarum (A10)</td><td>0.0</td></tr><tr><td>L. plantarum (D10)</td><td>28.5 ± 0.6<sup>b</sup></td></tr><tr><td>L. plantarum (D20)</td><td>17.6 ± 0.6<sup>c</sup></td></tr><tr><td>W. paramesenteroides (M31)</td><td>0.0</td></tr><tr><td>L. plantarum (A11)</td><td>0.0</td></tr><tr><td>L. plantarum (A12)</td><td>0.0</td></tr><tr><td>Not identified (D32)</td><td>29.1 ± 1.0<sup>b</sup></td></tr></tbody></table>

Values with the same alphabetical letter do not differ significantly at the 5% level

<figure><img src="image_3.png" /><figcaption>Fig. 1. Microbial inoculation tests</figcaption></figure>
<figure><img src="image_4.png" /><figcaption>**Fig. 2.** Effect of the *L. plantarum* strains addition to coffee cherries for detoxification in OTA on the sensory attributes of beverages produced</figcaption></figure>

In our study, there was no significant difference between beverages derived from detoxified coffee cherries and those made from non-inoculated coffee in terms of attributes including "acidity", "bitterness", "astringency", "body in the cup", "sourness" and "overall quality". The similarity about sensory attributes could be ascribed to the fact that the field tests were not conducted in controlled atmosphere, the microbial interactions that favored the dominance of endoegnous microorganisms and to the influence of uncontrolled microorganisms [34]. The low intensity scored by beverages from inoculated coffee cherries could be due to the lower amount of VOCs produced by LAB in inoculated green as compared to the naturally fermented green coffee [53]. Furthermore, the results showed also that the beverage prepared from the farmer control sample had a more intense coffee aroma. Main aroma compounds such as esters, higher alcohols, aldehydes and ketones, formed during roasting from organic compounds precursors would therefore be responsible for the intensity of the coffee odour of the beverage from the farmer control samples [54]. Finally, the inoculation of *L. plantarum* LAB D12 and LAB D13 certainly had an impact on the production of VOCs involved in beverages sensory qualities. According to Pereira et al. [54], the volatile organic compounds (VOCs) produced by LAB included ester, alcohol, alkane, acid, hydrocarbon, ether and nitrogen-containing. Bertrand et al. [28] concluded that these VOCs

seemed to be associated with a decrease in many aromatic quality attributes. Consequently, we can hypothesise that some VOCs play the inhibitory effetscs against the mycelial growth as like the *Bacillus* strains [55]. So, inoculated LABs did not negatively influence the sensory qualities of the coffee beverages.

**5. CONCLUSION**

LAB cells were efficient against mycelial growth of *A. carbonarius*. All antifungal *L. plantarum* strains exhibited OTA removal ability by adsorption.The findings of this study are very relevant, especially considering the critical toxic effect of OTA as well as the increasing OTA occurrence worldwide. The purpose of screening for LAB with the ability to reduce OTA production in green coffee was clearly demonstrated. The addition of antifungal and anti-mycotoxigenic LAB strains to coffee cherry did not generally influence the sensory attributes of beverage produced thereof except the intensity of odour. This study highlighted that tested *L. plantarum* strains are very promising biological candidates for various fermented foods safety such as coffee cherries cocoa beans and wine.

**ACKNOWLEDGEMENTS**

BEUGRE Guézéré Corinne's research is partially supported by a grant for PhD Student (N° SCAC_SC2 / BE-005 / Année 2019-2020 PGF-
RK\JNK) from French Embassy at Côte d'Ivoire awarded by "Service de Coopération et d'Action Culturelle (SCAC)." The authors thank also a lot hte family ANON, the family LOGBO, coffee farmers living at Akoupé and Daloa, coffee producing regions.

## COMPETING INTERESTS

Authors have declared that no competing interests exist.

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