Antifungal Activity of Lactic Acid Bacteria, Isolated from (Mytilus Galloprovincialis Lam.) in the Bulgarian Black Sea Aquatory

Home , Aspergillus niger

https://doi.org/10.5272/jimab.2020261.2875 Journal of IMAB Journal of IMAB - Annual Proceeding (Scientific Papers). 2020 Jan-Mar;26(1) ISSN: 1312-773X https://www.journal-imab-bg.org Original article

ANTIFUNGAL ACTIVITY OF LACTIC ACID BACTERIA, ISOLATED FROM (MYTILUS GALLOPROVINCIALIS LAM.) IN THE BULGARIAN BLACK SEA AQUATORY

Sevginar Ibryamova1, Nina Arhangelova2, Teodora Koynova1, Dimitar Dimitrov1, Zheni Dimitrova1, Radoslav Ivanov1, Karamphil Kalchev1, Nescho Chipev1, Nikolay Natchev1, Tsveteslava Ignatova-Ivanova1* 1) Department of Biology, Shumen University, Shumen, Bulgaria 2) Department of Physics and Astronomy, Shumen University, Shumen, Bulgaria..

ABSTRACT: Currently, aquaculture has become an important economic In the present study we present the results from an- activity in many countries. In large-scale production fa- tifungal activity of Lactic Acid Bacteria, isolated from M. cilities, where aquatic animals are exposed to stressful con- galloprovincialis Lam. from the Bulgarian Black Sea ditions, problems related to diseases and deterioration of aquatory. The Black Sea mussel (M. galloprovincialis environmental conditions often result in serious economic Lam.) is in fact the only marine species grown as losses. During recent decades, prevention and control of aquaculture in the Bulgarian section of the Black Sea. diseases have led to a substantial increase in the use of The mussels have an exceptional nutritional value, mak- veterinary medicines. However, the utility of antimicro- ing them highly suitable for the human diet. The cultiva- bial agents as apreventive measure has been questioned, tion of sea products, in particular the Black Sea clam, is given extensive documentation of the evolution of anti- becoming an important part of the Western Black Sea in- microbial resistance among pathogenic bacteria [1]. From dustry. With increasing of the tourist pressure and the con- a social point of view, safe food forms a significant prob- struction activities on the Black Sea coast, maritime pol- lem for modern civilization. The cultivation of sea prod- lution increased in the last decades. The pollutants in- ucts, in particular the Black Sea clam, had becom an im- clude various pathogenic microorganisms - Escherichia portant part of the Western Black Sea industry. The mus- coli, Staphylococcus aureus, Salmonella thyphimurium, sels are flter-feeding organisms and can concentrate bacte- Hymeniacidon perlevis, incl. the cholera cause (Vibrioo ria in high number [2]. The microbiota found in shellfishes cholera). Such pathogenic microorganisms are often can be divided into autochtonous and allochtonous mi- present in the mussels and can cause the spread of vari- croorganisms, and reflects the microbial population of the ous diseases. In this study 2 isolates Lactic acid bacteria, water in which they grow [3]. On the other hand, it has from M. galloprovincialis Lam have been identified as been found that seafood is a source of lactic acid bacteria Lactobacillus sp. and characterized as cultures with prom- (LAB), which synthesize substances with antimicrobial ising antifungal activity. The antifungal activity of our activity against many pathogenic microorganisms [2, 4]. new isolates Lactobacillus sp. lineages, seems to be a Consuming foods containing high quantities of chemical promising advantage, suggesting their potential applica- conservants provoked the interest in the use of natural and tions in different food technologies. minimally processed foods, with a focus on the use of natu- rally produced antimicrobial agents - bioconversants such Keywords: Bivalvia, Mytilus galloprovincialis Lam, as bacteriocins [5]. Bacteriocin producing LABs can im- probiotic bacteria, pathogens, Black Sea, prove the aquatic environment of the shrimp and fish aquacultures. The development of bacteriocinogenic strains and INTRODUCTION the isolation of bacteriocins from seafood and products Worldwide, people obtain about 25% of their ani- may result in valuable strains that are used both as mal protein from fish and shellfish and consumer’s demand bioconversants and as food (and feed) additives in aqua- for fish continues to climb. The Black Sea mussel (M. culture animals [6]. galloprovincialis Lam.) is the only species of mussels Currently, the food industry depends on chemical grown as aquaculture in the Bulgarian Black Sea aquatory. preservatives to extend the shelf life and control the growth Mussels have an exceptional nutritional value making them of spoilage fungi. The consumer’s awareness about the suitable for the human diet. Mussel flesh is rich in sele- health hazards associated with chemicals has recently in- nium, calcium, iron, magnesium, phosphorous and vita- creased, and they are demanding for processed foods that mins (A, B1, B2, B6, B12 and C). Mussel farming has an are free of preservatives. There are many natural alterna- important economic impact in many European countries. tive preservatives produced by certain microorganisms,

J of IMAB. 2020 Jan-Mar;26(1) https://www.journal-imab-bg.org 2875 however these microorganisms have to be non-toxic, easy 18 h) on MRS at 370C and in limitation of oxygen (tubes to grow and require simple media for cultivation [7]. Lac- or Petri dishes with the strains were incubated in plastic tic acid bacteria (LABs) represent a known potential source bags, which limited the oxygen content). for generating a variety of secondary metabolits such as X-Gal-based medium bacteriocines, organic acids and peptides [8]. In the past The modified MRS agar was used as the medium for decade the interest for antifungal LABs has increased and the incorporation of 5-bromo-4-chloro-3-indolyl β-D- different studies have showed that many LAB strains have galactopyranoside X-Gal (Boehringer Mannheim, the potential to impact the proliferation of fungi in vari- Montréal, Canada) [14]. ous food and feed materials [9]. LABs, due to their long API 50CHL System Assay Initial identiûcation of history of safe use in food and feed fermentations, [10] all the strains was performed by API 50CHL system have received both GRAS (Generally Recognised as Safe) (BioMerieux, Craponne, France), according to the manu- and QPS (Qualified Presumption of Safety) status in the facturer’s instructions. The fermentation proûles were read EU. Therefore, LABs have high potential as antifungal after incubation at 37C in anaerobic conditions, for 3 days. biocontrol agents in the future. The antifungal effect by Test Microrganisms Aspergillus niger, Penicillium the culture supernatant of various LAB strains against few claviforme, Saccharomyces cerevisae, Candida albicans fungi including food-deteriogens has been studied by vari- 8673 and Candida glabrata 72 were obtained from the ous authors [7]. Several compounds have been proposed National Bank for Industrial Microorganisms and Cell Cul- as responsible for the antifungal activity of LAB, like or- tures, Sofia, Bulgaria. All the isolates were checked for ganic acids, low molecular weight compounds, phenylace- purity and maintained in slants of Nutrient agar. tic and fatty acids, cyclic dipeptides, proteinaceous com- Assay for Antifungal Activity Antifungal assay was pounds and other miscellaneous compounds e.g. lactones performed by the well disc diffusion method using soft [11]. The fungistatic activity of pentocin TV35b against 0.8% agar. Agar medium was added to sterile Petri dishes Candida albicans and of two LAB strains against Fusarium seeded with 100 µL of each test yeast and fungi strains. (isolated from cereals) has been reported [12]. Due to their Wells of equal distance were dug on the seeded plates. inhibitory spectrum, these peptides are considered to be Before the assays, the strains LAB were twice pre-cultured powerful antimicrobials to target bacteria, fungi and para- in MRS broth, for 24h at 37°C. Exponential Lactobacillus sites [13]. The use of probiotics or beneficial bacteria, cultures in MRS broth were used as inoculum for the anti- which control pathogens through a variety of mechanisms, fungal tests. The plates were incubated at 37°C for 48 h. is increasingly viewed as an alternative to antibiotic treat- The antifungal activity was assayed by measuring the di- ment. ameter of the inhibition zone formed around the well [15]. In this study 2 isolates Lactic acid bacteria, from M. All Antifungal Activity of Lactic Acid Bacteria, Isolated galloprovincialis Lam have been identified as Lactoba- from M. galloprovincialis Lam 3 experiments were per- cillus sp. and characterized as cultures with promising an- formed in triplicates. tifungal activity. The antifungal activity of the new isolates Lactobacillus sp., seems to be possess potential for RESULTS AND DISCUSSION applications in different food technologies. The microscopic pictures were performed using stereomicroscope OPTIKA (Italy) with a camera Canon MATERIALS AND METHODS EOS 60D. Place and duration of the study: The study was con- The microorganisms were isolated from M. gallo- ducted at the Department of Biology, University of Shumen, provincialis Lam. (figure 1) in the Bulgarian Black Sea Bulgaria, from September 2018 to December 2018. aquatory. The microscopic pictures were performed using Collection of samples stereomicroscope OPTIKA (Italy) with a camera Canon The mussels were harvested from northern part of EOS 60D. the Bulgarian Black Sea aquatory. After collection, the samples (about 10 kg) were immediately refrigerated (4°C) Fig. 1. General view of a dissected M. galloprovin- and transported to the laboratory for the analyses (see cialis Lam. Fig.1). Microbiological analyses Three subsamples (each of about 1 kg of mussels) were used for the microbiological analyses. The mussels were scrubbed free of dirt, washed in hypochlorite solution (20 mg l-1), rinsed with sterile distilled water, and shucked with a sterile knife. Tissue liquor samples (about 100 g) were homogenized. Lactic acid bacteria (LAB) were isolated in media of MRS (de Mann Rogosa Sharpe, Biolife 272-20128, Milano, Italia). The strains were cultured overnight (16–

2876 https://www.journal-imab-bg.org J of IMAB. 2020 Jan-Mar;26(1) After 24 h of cultivated on MRS culture media microbial colonies were obtained colonies represented on fig. 2 and fig. 3. Figure 2 indicates that two strains of Lactobacillus spp. gave light blue colonies on modified MRS medium supplemented with 1 mM X-Gal.

Fig. 2. Pictures of the colonies of isolated species: A) colonies of Lactic acid bacteria on media MRS agar, B) colonies of Lactic acid bacteria on media MRS agar with X-Gal. The pictures were taken using stereomicroscope OPTIKA (Italy) and DinoEye, Eyepiece camera, USB, 1.3 megapixsel, up to 5 megapixel. Arrowheads indicate the LAB cells;

Figure 2 indicates that two strains of Lactobacillus cells. The cells gave negative probe for catalase and we spp. gave light blue colonies on modified MRS medium propose that these are representatives of the Lactobacillus supplemented with 1 mM X-Gal. sp. Following the classical microbial methods, we isolated The micromorphologic characteristic of the LAB, 2 strains of LAB from M. galloprovincialis Lam. The data which were isolated over MRS agar, demonstrated that the show that the isolates represent L. plantarum (fig. 3). cells possess elongated form and strain as Gram positive

Fig. 3. Light microscopic visualizations of Gram staining of Lactic acid bacteria. The picture were taken using microscope OPTIKA (Italy) at magnification 1000 by immersion and DinoEye, Eyepiece camera, USB, 1.3 megapixsel, up to 5 megapixel.

J of IMAB. 2020 Jan-Mar;26(1) https://www.journal-imab-bg.org 2877 API Tests API tests showed that all the strains tested belong to L. plantarum (figure 4)

Fig. 4. API test

The effects of LAB on the pathogenics fungi were cans 8673 and Candida glabrata 72. The sensitivities of summarized in Table 1. Varying degrees of inhibition were the test organisms to infusions were indicated by the clear detected against the isolates of Aspergillus niger, Penicil- zone around the wells (Fig. 5). lium claviforme, Saccharomyces cerevisae, Candida albi-

Fig. 5. Zone of inhibition with LAB along without LAB as control after seven days. A) Strain L. plantarum Ts2 cultivated with P. claviforme; B) Strain L. plantarum Ts1 cultivated with A. niger; C) Strain L. plantarum Ts2 cultivated with C. albicans 8673; D) Strain L. plantarum Ts2 cultivated with S. cerevisae;

2878 https://www.journal-imab-bg.org J of IMAB. 2020 Jan-Mar;26(1) Table 1. Inhibition zones (mm), of lactobacilli iso- showed the highest inhibitory activity. Valerio et al. (2009) lated from M. galloprovincialis Lam in the Bulgarian Black [18] tested fermentation products of 17 LAB strains against Sea aquatory. Data are presented as average values ± standard Penicillium roqueforti and Aspergillus niger. Lactobacil- deviation in mm. lus citreum, Weissella cibaria, and L. rossiae inhibited (>98%) the growth of A. niger. L. plantarum was the most effective against P. roqueforti. Ahmad Rather et al. (2013) Zone of inhibition (mm) [19] screened 1400 bacteria isolated from different kimchi Target strains by strains LAB samples for their antifungal activity against A. niger by L. plantarum Ts1 L. plantarum Ts2 dual-culture agar plate assay. Luz C, et. al, (2017) [7] re- A. niger 33.81 ± 0.15 31.60 ± 0.15 ported, that the strain exhibiting the high antifungal ac- P. claviforme 26.43 ± 0.16 26.18 ± 0.17 tivity was L. plantarum YML007. This strain was further screened against various pathogens showing the highest S. cerevisae - 8.80 ± 0.09 inhibition against A. niger, followed by Aspergillus fla- C. albicans 8673 15.24 ± 0.19 13.35 ± 0.17 vus, Aspergillus oryzae and Fusarium oxysporum. The biopreservative activity of L. plantarum YML007 was The tested strains completely suppressed the growth evaluated using dried soybeans and no fungal growth was of against C. glabrata 72 (Table 1). observed in the beans treated with fivefold concentrated Strains of L. plantarum Ts2 completely suppressed cell-free supernatant of L. plantarum YML007. Gupta et the growth of S. cerevisae (Table 1 and Fig. 1D). While, al. (2014) [20] studied the antifungal activity of 88 L. in the sample with L. plantarum strains Ts1 and Ts2, a plantarum strains against A. niger, A. flavus, Fusarium retarded and weak growth of A. niger and P. claviforme culmorum, Penicillium roqueforti, Penicillium expansum, (fig. 1A and 1B) was observed. Penicillium chrysogenum and Cladosporium spp. Nine of The moulds produce compounds potentially toxics these strains that strongly inhibited at least three moulds to the consumers called mycotoxins which can cause ill- were further screened based on the antifungal properties of ness and death. Numerous incidences of mould and my- their cell free supernatant. None of the tested L. plantarum cotoxin contamination in food and feed happened due to strains was able to completely inhibit the growth of the fungi ubiquitous nature - their ability to colonize differ- analyzed filamentous fungi and the maximum antagonis- ent substrates and to the lack of effective control meas- tic effect was observed after 5 days of incubation in plates ures [16]. supplemented with CFS (12% v/v) from L. plantarum UFG However, the spore germination and the colony 108 and L. plantarum UFG 121, in which the growth of P. growth stared started only on the fifth day of the mould expansum and F. culmorum was reduced about 50 and 60%, lactobacilli co-cultivation, which also should be consid- respectively. ered as a good result. In the literature, most of the active Lavermicocca et al. [21] reported the production of antifungal strains in fermented milk products were related antifungal compounds by a sourdough L. plantarum strain. to the L. casei group [16]. Ryan et al. (2008) [17] investi- The same was observed for Lactobacillus coryneformis Si3 gated the antifungal activity of two strains of L. plantarum [22] as well as isolates of L. plantarum, L. coryneformis, fermented sourdoughs comparing their activity to that of a Lactobacillus salivarius, Lactobacillus sake, E. hirae, and sourdough fermented by L. sanfranciscensis LTH2581, as Enterococcus durans from various sources. Ström et al. [23] well as to that of a chemically acidified dough and a dough also found that L. plantarum MiLAB 393 produced anti- containing antibiotics. L. plantarum fermented sourdough fungal substances. Addis, et al. [24] reported that LAB in-

J of IMAB. 2020 Jan-Mar;26(1) https://www.journal-imab-bg.org 2879 hibited the growth at several yeasts with L. plantarum be- and disease resistance. In aquaculture, the range of ing active against S. cerevisiae and other yeast species. probiotics evaluated for use is considerably wider than in Durlu-Ozkaya, Karabiçak, Kayali, and Essen [25] found terrestrial agriculture. Several probiotics both as mono- LAB, such as L. plantarum, L. paracasei, subsp. paracasei species, as well multispecies supplements are commercially and L. lactis isolated from a Turkish traditional cheese to available for aquaculture practices [29]. Apart from the inhibit S. cerevisiae, several species of Candida, Torulop- nutritional and other health benefits, certain probiotics as sis glabrata and Rhodotorula rubra, isolates from the same water additives can also play a significant role in decom- cheese. position of organic matter, reduction of nitrogen and phos- The effects of LAB on the pathogenics fungi were phorus level, as well as control of ammonia, nitrite, and summarized in Table 1. Varying degrees of inhibition were hydrogen sulphide [30]. Numerous microbes have been detected against the isolates of Aspergillus niger, Penicil- identified as probiotics for aquaculture practices, many of lium claviforme, Saccharomyces cerevisae, Candida albi- which differ markedly in their mode of action. There are, cans 8673 and Candida glabrata 72. The sensitivities of however, some common mechanisms of action that have the test organisms to infusions were indicated by the clear been reported for the majority of probiotic strains. zone around the wells (fig. 5). Probiotics help in feed conversion efficiency and live The tested strains completely suppressed the growth weight gainand confer protection against pathogens by of against C. glabrata 72 (table 1). Strains of L. plantarum competitive exclusion for adhesion sites, production of Ts2 completely suppressed the growth of S. cerevisae (Ta- organic acids (formic acid, acetic acid, lactic acid), hydro- ble 1 and Fig. 1D). While, in the sample with L. plantarum gen peroxide and several other compounds such as antibi- strains Ts1 and Ts2, a retarded and weak growth of A. niger otics, bacteriocins, siderophores, lysozyme and also modu- and P. claviforme (fig. 1A and 1B) was observed. late physiological and immunological responses in fish. Probiotic applications, in biological control of sea- Perusal of available literatures indicates that several food associated pathogens can be an alternative solution, probiotics either individually or in combination can en- providing the consumer with a product of good quality hance both systemic as well as local immunity in fish [31]. owing to the use of nontoxic compounds. Most of the con- To date, data concerning isolated from mussels LAB sumed shellfish in Bulgaria is produced on aquaculture are lacking. In this context our investigation for first time farms. Because much of the shellûsh is grown and harvested provides results for the presence and determination of mus- as aquacultures, this offers an opportunity to monitor and sel inhabiting microbiota, which possess antimitotic ac- improve microbiological safety of the product both tivity and have a probiotic potential. These qualities suite preharvest and postharvest. Several processes, including these microorganisms as agents in the industry, produc- freezing, low-temperature pasteurization, high pressure tion and conservation of aqua cultures. processing and irradiation have been reported to be capable of reducing patogenics bacteria. The massive use of CONCLUSIONS antibiotics may lead to the emergence of resistant bacte- Based on our results, LAB could be used as a bio- ria, which can spread in the environment and jeopardize protective culture in M. galloprovincialis Lam. depura- human health [26]. Probiotics are among the most promis- tion to prevent pathogenic growth. Additional studies are ing alternatives to antibiotics and the application in needed to provide data concerning the role played by con- aquaculture is now widely accepted. It may be possible to taminated shellfish consumption in human diseases. The reduce the load of pathogens in seafood by improving water LAB colonization mechanism and the probiotic potential quality and also by introducing biocontrol bacteria capa- of LAB in M. galloprovincialis remains to be investigated. ble of excluding pathogens from shellûsh associated microbial communities [27]. The Black sea mussels are ûlter feeders, capable of ûltering up to 10 L of water per hour Acknowledgements [28]. This feeding behavior leads to the accumulation (at The contributors express their gratitude for the least transient) of human pathogens within shellûsh. The funding by fund scientific research Bulgaria programme consumption of raw bivalve mollusks is associated with grant KP-06-N21/7/ 16 Dec 2018, the project by Shumen high risk of food poisoning. University project RD-08-104/01.02.2019 and RD-08-45/ In aquaculture practices, probiotics are used for a 18.01.2019. Department of Biology and National Program quite long time, but in last few years probiotics became an Young Scientists and Postdoctoral Students. integral part of the culture practices for improving growth

REFERENCES: 1. Balcazar JL, de Blas I, Ruiz- 2. Šoliæ M, Krstuloviæ N, Joziæ S, 3. Vernocchi P, Maffei M, Lanciotti Zarzuela I, Cunningham D, Vendrell D, Curaæ D. The rate of concentration of R, Suzzi G, Gardini F. Characterization Muzquiz JL. The role of probiotics in faecal coliforms in shellfish under dif- of Mediterranean mussels (Mytilus aquaculture. Vet Microbiol. 2006 May ferent environmental conditions. Envi- galloprovincialis) harvested in Adri- 31;114(3-4):173-86. [PubMed] ronment International. 1999 Dec; atic Sea (Italy). Food Control. 2007 [Crossref] 25(8):991-1000. [Crossref] Dec;18(12):1575-1583. [Crossref]

2880 https://www.journal-imab-bg.org J of IMAB. 2020 Jan-Mar;26(1) 4. Khouadja S, Haddaji N, Hanchi [Crossref] 21. Gupta R, Srivastava S. Antifun- M, Bakhrouf A. Selection of lactic 13. Laitila A, Alakomi HL, Raaska gal effect of antimicrobial peptides acid bacteria as candidate probiotics L, Mattila-Sandholm T, Haikara A. An- (AMPs LR14) derived from Lactobacil- for Vibrio parahaemolyticus depura- tifungal activities of two Lactobacil- lus plantarum strain LR/14 and their tion in pacific oysters (Crassostrea lus plantarum strains against Fusarium applications in prevention of grain gigas). Aquaculture Research. 2017 moulds in vitro and in malting of bar- spoilage. Food Microbiol. 2014 Sep; Apr;48(4):1885-1894 [Crossref] ley. J Appl Microbiol. 2002; 93(4):566- 42:1-7. [PubMed] [Crossref] 5. Françoise L. Occurrence and role 76. [PubMed] [Crossref] 22. Lavermicocca P, Valerio F, of lactic acid bacteria in seafood prod- 14. Mason AJ, Moussaoui W, Evidente A, Lazzaroni S, Corsetti A, ucts. Food Microbiol. 2010 Sep;27(6): Abdelrahman T, Boukhari A, Bertani P, Gobbetti M. Purification and charac- 698-709. [Crossref] Marquette A, et al. Structural determi- terization of novel antifungal com- 6. Falanga A, Lombardi L, Franci G, nants of antimicrobial and antiplas- pounds from the sourdough Lactoba- Vitiello M, Iovene MR, Morelli G, et modial activity and selectivity in his- cillus plantarum strain 21B. Appl al. Marine Antimicrobial Peptides: Na- tidine-rich amphipathic cationic pep- Environ Microbiol. 2000 Sep;66(9): ture Provides Templates for the Design tides. J Biol Chem. 2009 Jan 2;284(1): 4084-90. [PubMed] [Crossref] of Novel Compounds against Patho- 119-33. [PubMed] [Crossref] 23. Saenz de Rodriguez MA, Diaz- genic Bacteria. Int J Mol Sci. 2016 15. Nwamaioha NO, Ibrahim SA. A Rosales P, Chabrillon M, Smidt H, Arijo May 21;17(5) pii: E785. [PubMed] selective medium for the enumeration S, Leon-Rubio JM. Effect of dietary ad- [Crossref] and differentiation of Lactobacillus ministration of probiotics on growth 7. Luz C, Saladino F, Luciano FB, delbrueckii ssp. bulgaricus. J Dairy and intestine functionally of juvenile Mañes J, Meca G. In vitro antifungal Sci. 2018 Jun;101(6):4953-4961. Senegalese sole (Solea senegalensis, activity of bioactive peptides pro- [PubMed] [Crossref] Kaup 1858). Aquac Nutr. 2009 Apr; duced by Lactobacillus plantarum 16. Muhialdin BJ, Hassan Z, Saari 15(2):177-85. [Crossref] against Aspergillus parasiticus and N. Chapter 6: Lactic Acid Bacteria in 24. Ström K, Sjögren J, Broberg A, Penicillium expansum. LWT - Food Biopreservation and the Enhancement Schnürer J. Lactobacillus plantarum Science and Technology. 2017 Aug; of the Functional Quality of Bread. In MiLAB 393 produces the antifungal 81:128-135. [Crossref] Book: Lactic Acid Bacteria - R & D for cyclic dipeptides cyclo(L-Phe-L-Pro) 8. Cizeikiene D, Juodeikiene G, Food, Health and Livestock Purposes. and cyclo(L-Phe-trans-4-OH-L-Pro) Paskevicius A, Bartkiene E. Antimicro- INTECH. 2015. pp:155-172. and 3-phenyllactic acid. Appl Environ bial activity of lactic acid bacteria 17. Hassan YI, Zhou T, Bullerman Microbiol. 2002 Sep;68(9):4322-7. against pathogenic and spoilage mi- LB. Sourdough lactic acid bacteria as [PubMed] [Crossref] croorganism isolated from food and antifungal and mycotoxin-controlling 25. Addis E, Fleet GH, Cox JM, their control in wheat bread. Food agents. Food Sci Tech Int. 2015 Jan; Kolak D, Leung T. The growth, prop- Control. 2013 Jun; 31(2):539-545. 22(1):79-90. [Crossref] erties and interactions of yeasts and [Crossref] 18. Ryan LA, Dal Bello F, Arendt bacteria associated with the maturation 9. Mauch A, Dal Bello F, Coffey A, EK. The use of sourdough fermented of Camembert and blue-veined chee- Arendt EK. The use of Lactobacillus by antifungal LAB to reduce the ses. Int J Food Microbiol. 2001 Sep; brevis PS1 to in vitro inhibit the out- amount of calcium propionate in 69(1-2):25-36. [PubMed] [Crossref] growth of Fusarium culmorum and bread. Int J Food Microbiol. 2008 Jul 26. Durlu-Ozkaya F, Karabicak N, other common Fusarium species found 31;125(3):274-8. [PubMed] [Crossref] Kayali R, Essen B. Inhibition of yeasts on barley. Int J Food Microbiol. 2010 19. Valerio F, Favilla M, De Bellis isolated from traditional Turkish Jun 30;141(1-2):116-21. [PubMed] P, Sisto A, de Candia S, Lavermicocca cheeses by Lactobacillus spp. Int J [Crossref] P. Antifungal activity of strains of lac- Dairy Technol. 2005 May;58(2):111- 10. Stoianova LG, Ustiugova EA, tic acid bacteria isolated from a semo- 114. [Crossref] Netrusov AI. [Antibacterial metabolites lina ecosystem against Penicillium 27. Antimicrobial resistance. WHO. of lactic acid bacteria: their diversity roqueforti, Aspergillus niger and 15 February 2018. [Internet] and properties]. [in Russian] Prikl Endomyces fibuliger contaminating 28. Khouadja S, Haddaji N, Hanchi Biokhim Mikrobiol. 2012 May- bakery products. Syst Appl Microbiol. M, Bakhrouf A. Selection of lactic acid Jun;48(3):259-75. [PubMed] [Crossref] 2009 Sep;32(6):438-48. [PubMed] bacteria as candidate probiotics for Vi- 11. Hugenholtz J. Traditional bio- [Crossref] brio parahaemolyticus depuration in technology for new foods and bever- 20. Ahmad Rather I, Seo BJ, Rejish pacific oysters (Crassostrea gigas). ages. Curr Opin Biotechnol. 2013 Kumar VJ, Choi UH, Choi KH, Lim JH, Aquaculture Research. 2017 Apr; Apr;24(2):155-9. [PubMed] [Crossref] et al. Isolation and characterization of 48(4):1885-1894. [Crossref] 12. Peyer LC, Axel C, Lynch KM, a proteinaceous antifungal compound 29. Catarino AI, Macchia V, Sand- Zannini E, Jacob F, Arendt EK. Inhibi- from Lactobacillus plantarum erson WG, Thompson RC, Henry TB. tion of Fusarium culmorum by car- YML007 and its application as a food Low levels of microplastics (MP) in boxylic acids released from lactic acid preservative. Lett Appl Microbiol. wild mussels indicate that MP inges- bacteria in a barley malt substrate. 2013 Jul;57(1):69-76. [PubMed] tion by humans is minimal compared Food Control. 2016 Nov;69:227-236. [Crossref] to exposure via household fibres fall-

J of IMAB. 2020 Jan-Mar;26(1) https://www.journal-imab-bg.org 2881 out during a meal. Environ Pollut. International Symposium on Micro- probiotic, Lactobacillus acidophilus, 2018 Jun;237:675-684. [PubMed] bial Ecology. Bell CR, Brylinsky M, on the growth performance, haematol- [Crossref] Johnson-Green P (eds). Atlantic ogy parameters and immunoglobulin 30. Moriarty DJW. Disease control Canada Society for Microbial Ecol- concentration in African Catfish in shrimp aquaculture with probiotic ogy, Halifax, Canada, 1999. [Internet] (Clarias gariepinus, Burchell 1822) bacteria. In: Microbial Biosystems: 31. Al-Dohail MA, Hashim R, fingerling. Aquac Res. 2009 Sep; New Frontiers. Proceedings of the 8th Aliyu-Paiko M. Effects of the 40(14):1642-52. [Crossref]

Please cite this article as: Ibryamova S, Arhangelova N, Koynova T, Dimitrov D, Dimitrova Z, Ivanov R, Kalchev K, Chipev N, Natchev N, Ignatova-Ivanova T. Antifungal Activity of Lactic Acid Bacteria, Isolated from (Mytilus galloprovincialis Lam.) in The Bulgarian Black Sea aquatory. J of IMAB. 2020 Jan-Mar;26(1):2875-2882. DOI: https://doi.org/10.5272/jimab.2020261.2875

Received: 17/10/2019; Published online: 22/01/2020

Address for correspondence: Tsveteslava Ignatova-Ivanova Department of Biology, Shumen University “Konstantin Preslavski” 115, Universitetska Str., Shumen, Bulgaria E-mail: [email protected] 2882 https://www.journal-imab-bg.org J of IMAB. 2020 Jan-Mar;26(1)