Antifungal activities of basil (Ocimum basilicum L.) extract on Fusarium species

The basil extract composition was determined by the GC-MS method and 38 different components were identified. The major components of the basil extract were estragol (86.72%), trans-α-bergamotene (2.91%), eucalyptol (2.67%), trans-ocimene (1.04%), linalool (0.72%), methyl-eugenol (0.71%), etc. The antifungal potential of the basil extract was tested against Fusarium oxysporum, F. proliferatum, F. subglutinans, and F. verticillioides isolated from cakes, using the agar plate method. Extract concentrations of 0.35 and 0.70% (v/v) significantly inhibited the growth of F. proliferatum (33.37 and 44.30%, respectively) and F. subglutinans (24.74 and 29.27%, respectively) whereas other investigated Fusarium species exhibited much lower sensitivity. The basil extract completely inhibited the growth of investigated Fusarium spp. at the concentration of 1.50% (v/v). Higher concentrations (0.35 and 0.70% (v/v)) reduced growth of aerial mycelium in all tested species. Strong medium pigmentation in the case of F. proliferatum and F. verticillioides was observed. The microscopic examination of the samples confirmed the presence of hyphae deformations with a frequent occurrence of fragmentations, thickenings and diminished sporulation. In addition to the basic, sensory, role the extract of basil has in the food product, it exerted significant antifungal properties, depending on its concentration.Key words: Basil (Ocimum basilicum L.) extract, components, antifungal activity, Fusarium spp

Pathogenicity and diversity of vegetative compatibility of Fusarium verticillioides

Pathogenicity of 10 Fusarium verticillioides isolates, originated from grain of wheat (five isolates) and maize (five isolates), were studied under greenhouse conditions. Based on different parameters of the pathogenicity estimate (a scale for % of nonemerged plants, % of survived plants, plant vigour - the growth and dry weight of roots and epicotyls and disease severity) it was determined that all F. verticillioides isolates expressed a different degree of pathogenicity. According to % of nonemerged plants six three and one F. verticillioides isolates expressed low, moderate and high degree of pathogenicity, respectively. All F. verticillioides isolates reduced the plant survival rate and vigour, while the disease severity ranged from 2.0 to 3.54. Two types of nit mutants, nit1 and NitM, were obtained by the use of the method of vegetative compatibility. The frequency of nit1 mutants was greater (58.79%) than the frequency of NitM mutants (5.77%). A total of 10 vegetative compatibility groups (VCGs) of F. verticillioides were established in the complementation tests. These results point out to a high genetic diversity of F. verticillioides population

Plesni i mikotoksini u uskladištenom kukuruzu

In this study the presence of moulds and mycotoxins in samples of stored maize grains in the period from October 2011 to September 2012 was investigated. Mycological analyses of whole and broken grains showed the presence of species from the genera Alternaria, Aspergillus, Fusarium, Penicillium, Rhizopus and others. Among the Aspergillus and Fusarium genera as potentially toxigenic fungi, Aspergillus flavus was identified with the highest percentage on broken grains (20.38%) whereas F. verticilioides was the predominant species in the whole maize grains (34.04%). In addition, it was obtained that tested samples of stored maize grains were 100% positive with aflatoxin B1 (AFB1), zearalenone (ZON), deoxynivalenol (DON) and fumonisin B1 (FB1) with an average concentration of 1.39 μg kg-1, 71.79 μg kg-1, 128.17 μg kg-1, and 1610.83 μg kg-1, respectively. A significant positive correlation was found between the moisture content and the presence of Fusarium spp. on the broken grains (r = 0.44) and between the moisture content and the concentration of DON (r = 0.61). However, a significant negative correlation was found between moisture content and FB1 (r = -0.34), and between the concentration of ZON and DON mycotoxins (r = -0.58).U radu je ispitivano prisustvo plesni i mikotoksina u uzorcima zrna uskladištenog kukuruza u periodu od oktobra 2011. do septembra 2012. godine. Mikološkim analizama celog i slomljenog zrna kukuruza ustanovljeno je prisustvo vrsta iz rodova Alternaria, Aspergillus, Fusarium, Penicillium, Rhizopus i drugih. Od potencijalno toksigenih vrsta iz rodova Aspergillus i Fusarium, identifikovane su u najvećem procentu A. flavus na slomljenom (20,38%) i F. verticillioides na celom zrnu kukuruza (34,04%). Ispitivani uzorci uskladištenog kukuruza bili su 100% pozitivni sa aflatoksinom B1 (AFB1), zearalenonom (ZON), deoksinivalenolom (DON) i fumonizinom B1 (FB1) sa prosečnim koncentracijama 1,39 μg kg-1, 71,79 μg kg-1, 128,17 μg kg-1 i 1610,83 μg kg-1, respektivno. Statistički značajna pozitivna korelacija ustanovljena je između sadržaja vlage i prisustva Fusarium spp. na slomljenom zrnu kukuruza (r = 0,44), kao i između sadržaja vlage i koncentracije DON (r = 0,61). Statistički značajna negativna korelacija ustanovljena je između sadržaja vlage i FB1 (r = -0,34), kao i između koncentracija ZON i DON mikotoksina (r = -0,58)

Screening of mycotoxins in animal feed from the region of Vojvodina

This paper shows the results of screening of mycotoxins in animal feed originating from the region of Vojvodina. Permanent screening is needed on all levels of production and storage, as well as the use of known methods to reduce mould contamination or toxin content in feedstuffs and feed. A total of 56 representative samples were collected from feed companies from the region of Vojvodina. Samples were collected during February 2009. The collected samples included 41 samples of feedstuffs (soybean, soybean meal, soybean grits, soybean cake, maize, sunflower meal, barley, wheat feed flour, rapeseed meal, dehydrated sugar beet pulps, alfalfa meal, yeast, dried whey, fish meal, meat-bone meal) and 15 samples of complete feedingstuffs. The amounts of aflatoxins, ochratoxin A, zearalenone, fumonisin and deoxynivalenol were determined. Screening method for the analysis was done using Neogen Veratox® testing kits. The test itself is a competitive direct enzyme-linked immunosorbent assay (CD-ELISA). Mycotoxins were present in 71.4% of the samples, but the values determined were below the maximum allowed limits for both Serbian and EC reference values. Zearalenone was found with the highest incidence (57.1% of samples), followed by ochratoxin A (37.5%), fumonisin (33.9%), deoxynivalenol (14.3%) and aflatoxins (3.6%)

Prisustvo deoksinivalenola u ozimoj pšenici tretiranoj s fungicidima

Natural occurrence of Fusarium spp. and concentrations of mycotoxin deoxynivalenol (DON) in the grain of the winter wheat moderately susceptible to Fusarium head blight (FHB) has been studied. Grain samples were collected from wheat crops intended mainly for human consumption. All wheat crops were treated with fungicides before (a.i. flutriafol - formulated as Fluoco, applied in dose of 0.5 l ha-1) and during the flowering phase of growing (a.i. thiophanate-methyl + epoxiconazole formulated as Eskorta plus and a.i. thiophanate-methyl formulated as Funomil, applied in doses of 0.75 and 0.5 l ha-1, respectivily). Among of Fusarium species only F. graminearum, as a well known producer of DON, was identified. This fungus was identified in 15 of 19 samples (78.9%) with incidence in positive samples of 2 to 28% (average, 14.0%). Presence of DON was established in 13 of a total 19 investigated wheat grain samples (68.4%). In positive samples DON was detected in concentrations from 69 to 918 μg kg-1 (average, 478 μg kg-1). DON showed a significant and positive correlation at P≥0.05 with grain moisture content (r = 0.52*). Between the frequency of F. graminearum and concentration of DON and between the frequency of F. graminearum and grain moisture content, positive correlation was determined, but without statistical significance (r = 0.44 and r = 0.29, respectively).U radu je proučavana prirodna pojava Fusarium spp. i koncentracija mikotoksina deoksinivalenola (DON) u zrnu ozime pšenice srednje osetljive prema fuzariozi klasa (FHB). Uzorci zrna su prikupljeni sa proizvodnih useva pšenice namenjene uglavnom za ljudsku upotrebu. Svi usevi pšenice bili su tretirani sa fungicidima pre (a.m. flutriafol - formulisana kao preparat Fluoco, primenjen u dozi 0,5 l ha-1) i tokom cvetanja biljaka (a.m. tiofanat-metil + epoksikonazol formulisana kao preparat Eskorta plus i a.m. tiofanat-metil formulisana kao Funomil, primenjeni u dozi 0.75 i 0.5 l ha-1, respektivno). Među Fusarium vrstama jedino je identifikovana F. graminearum, koja je poznati producent DON. Ova gljiva je bila identifikovana u 15 od 19 uzoraka (78.9%) sa incidencom od 2 do 28% (prosek 14.0%) u pozitivnim uzorcima. Prisustvo DON je utvrđeno u 13 od ukupno 19 proučavanih uzoraka pšenice (68.4%). U pozitivnim uzorcima DON je detektovan u koncentracijama od 69 do 918 μg kg-1 (prosek 478 μg kg-1). DON je pokazao značajnu i pozitivnu korelaciju pri P≥0.05 sa sadržajem vlage zrna (r = 0.52*). Između učestalosti F. graminearum i koncentracije DON i učestalosti F. graminearum i sadržaja vlage zrna utvrđena je, takođe, pozitivna korelacija ali statistički nije značajna (r = 0.44 i r = 0.29, respektivno)

Učestalost toksigenih vrsta Fusarium i fuzariotoksina u zrnu pšenice u Srbiji

A total of 88 and 40 wheat samples collected immediately prior to harvest in 2005 and 2006, respectively, under different agroecological conditions, were studied in respect to the occurrence of Fusarium spp. and the production of fusariotoxins. The greatest number of samples was infected with species of the genera Fusarium (81.8 and 65.0%), and Alternaria (36.3 and 17.5%) with the intensity ranging from 9.4 to 84.0% in 2005 and from 23.4 to 80.6% in 2006. Out of 13 identified species belonging to the genus Fusarium, F. graminearum had the highest frequency (35.2 and 12.5%) and the intensity up to 67.2%, and 21.9%, in 2005 and 2006, respectively, followed by F. poae but only in 2005 (20.4%), and F. proliferatum in 2006 (19.7%). The natural occurrence of mycotoxins in positive samples varied from 37 to 331 ppb for zearalenone and from 31 to 125 ppb for diacetoxyscirpenol (DAS) and T-2 toxin. The concentration of mycotoxins amounted, on average, to 133.4, 61.0 and 45.7 ppb for zearelenone, DAS and T-2 toxin, respectively.Osamdeset osam i 40 uzoraka pšenice, prikupljenih neposredno pred žetvu u 2005. i 2006. godini u različitim agroekološkim uslovima u Srbiji proučavani su radi praćenja pojave Fusarium spp. i stvaranja fuzariotoksina. Najveći broj uzoraka bio je zaražen vrstama roda Fusarium (81,8 i 65,0%), a zatim sa Alternaria spp. (36,3 i 17,5%) sa intenzitetom 9,4-84,0% u 2005. i 23,4-80,6% u 2006. godini. Od ukupno 13 identifikovanih vrsta iz roda Fusarium najučestalija je bila F. graminearum (35,2 i 12,5%) sa intenzitetom do 67,2% u 2005. i do 21,9% u 2006. godini, a zatim F. poae, ali samo u 2005. godini (20,4%), i F. proliferatum u 2006. godini (19.7%). Prirodna pojava mikotoksina u pozitivnim uzorcima je varirala od 37 do 331 ppb za zearalenon i od 31 do 125 ppb za diacetoksiscirpenol (DAS) i T-2 toksin. U proseku, koncentracija mikotoksina je bila 133,4 ppb za zearelonon, 61,0 ppb za DAS i 45,7 ppb za T-2 toksin

Utjecaj eteričnog ulja bijeloga pelina (Artemisia absinthium) na antioksidativni status tovnih pilića invadiranih oocistama Eimeria spp.

The aim of this study was to investigate the effects of Artemisia absinthium essential oil (AAEO) on the enzymatic activity of superoxide-dismutase (SOD), glutathione-peroxidase (GSHPx), glutathione-reductase (GR), peroxidase (POD) and xanthine-oxidase (XOD) and the content of lipid peroxides (LPx) and glutathione (GSH) in broilers infected with an oocyst mixture of Eimeria tenella, Eimeria mitis and Eimeria necatrix, compared to coccidicide salinomycine. The investigation was carried out on 240 Arbour acres broilers of both sexes. Broilers were distributed completely randomly into four treatment groups: treatment group A was uninfected and untreated; treatment group B was infected and kept untreated; treatment group C preventively received coccidicide salinomycine in a dose of 60 mg/kg of feed and inoculated with an oocyst mixture on the 21st day-of-age; treatment group D received AAEO in their feed in a dose of 3 g/kg and were infected with the oocyst mixture on the 21st day of age. During the study, bloody diarrhoea was observed from the 3rd to 9th day after the challenge. After six days of infection, the most intensive bloody diarrhoea was noticed in the un-medicated treatment group. In order to evaluate the effects of essential oil on poultry coccidiosis induced by Eimeria spp., oocysts per gram of faeces (OPG) were also investigated in all treatment groups. During the experiment, the oocyst output and mortality rate were significantly lower (P<0.05) in the AAEO treatment group (D2) in comparison to the positive control (B), while significant excretion of oocysts was noticed in the faeces of non-treated broilers infected with Eimeria spp. The broilers treated with salinomycin (C2) showed complete reduction of oocysts in their faeces at 30 days of age. The results obtained in this study indicate changes in the content and the activity of the non-enzymatic and enzymatic antioxidative protective systems in blood hemolysates of infected chickens. The positive preventive effects of AAEO, applied in a concentration of 3g/kg of feed, were high on the antioxidative system of erythrocytes. On the basis of the obtained results, it was concluded that AAEO was effective in lowering the intensity of bloody diarrhoea, as well in reducing the oocyst output of the preventively treated and infected broilers; hence it may be used as a prophylactic feed additive. Moreover, AAEO showed an important role in the activation of antioxidative protection systems in infected broilers, which is of great interest since free radicals and lipid peroxides, formed as a result of lower food intake and exhaustion of the organism induced by diarrhoea, could cause cellular membrane damage.U radu je istražen utjecaj eteričnog ulja bijeloga pelina (Artemisia absinthium) (AAEO) na enzimsku aktivnost superoksid dismutaze (SOD), glutation peroksidaze (GSHPx), glutation reduktaze (GR), peroksidaze (POD) i ksantin-oksidaze (XOD), kao i sadržaj lipidnih peroksida (LPx) i glutationa (GSH) u tovnim pilićima inficiranim smjesom oocista Eimeria tenella, Eimeria mitis i Eimeria necatrix u odnosu na kokcidiostatik salinomicin. Istraživanje in vivo provedeno je na 240 pilića Arbor acres, oba spola. Pilići su bili podijeljeni u četiri skupine. Skupina A je bila neinficirana i netretirana (negativna kontrola). Skupina B je bila inficirana i netretirana (pozitivna kontrola). Skupina C je preventivno dobivala kokcidiostatik salinomicin u količini od 60 mg/kg hrane te je bila inokulirana kokcidijama 21. dana starosti. Skupina D je u hrani dobivala AAEO u količini 3 g/kg te je 21. dana starosti bila inficirana mješavinom oocisti Eimeria sp. Tijekom istraživanja krvava dijareja javljala se od 3. do 9. dana nakon infekcije. Šest dana nakon inficiranja, najintezivnija krvava dijareja zapažena je u netretiranih pilića. U cilju procjene djelovanja eteričnog ulja bijeloga pelina na kokcidiozu kod peradi izazvanu Eimeria oocistama, broj oocista po gramu fecesa (OPG) također je bio istražen u svim skupinama. Tijekom pokusa utvrđeno je da je broj oocista i razina smrtnosti bila znatno niža (P<0,05) u AAEO skupini (D2) u usporedbi s pozitivnom kontrolom (B), dok je značajno izlučivanje oocista uočeno u fecesu pilića koji nisu bili tretirani. Pilići koji su dobivali salinomicin (C2) pokazali su potpunu redukciju oocista u fecesu 30. dana starosti. Rezultati dobiveni u ovoj studiji pokazali su promjene u sadržaju i aktivnosti neenzimskih i enzimskih sustava zaštite u krvi inficiranih tovnih pilića. Zapaženi su pozitivni učinci preventivne primjene AAEO u koncentraciji 3 g/kg hrane na antioksidativni sustav eritrocita. Na osnovi dobivenih rezultata zaključeno je da je eterično ulje Artemisia absinthium vrlo učinkovito u smanjenju jačine krvavog proljeva kao i u smanjenju broja oocista u pilića koji su preventivno tretirani s AAEO. Stoga se može zaključiti da eterično ulje bijeloga pelina može biti davano kao profilaktički dodatak hrani za životinje. Također, AAEO ima pozitivan utjecaj na aktiviranje antioksidativnog sustava zaštite u krvi pilića, što je vrlo značajno s obzirom na to da slobodni radikali i lipidni peroksidi, koji nastaju kao rezultat manjeg unosa hrane i iscrpljenosti organizma uzrokovanog dijarejom, mogu izazvati oštećenje stanične membrane

Toksigeni potencijal izolata Fusarium poae poreklom sa pšenice

Eleven isolates of F. poae, originated from wheat grain at 9 locations mainly in Vojvodina, were encompassed by the present study. The greatest number of samples was collected in 2005, in which the climatic conditions favoured a more intensive occurrence of Fusarium ear blight of wheat. In order to determine toxicological potential of this species, cultures of the selected isolates were grown in liquid media (GPY and SPY) on a rotary shaker (180 revolutions min-1), at room temperature (21-26°C) for three days. Crude toxins were isolated from liquid culture filtrates of isolates by the use of ethyl acetate, while quantification of mycotoxins was done by the thin layer chromatography method. A liquid culture of the isolate GZ-LES (F. graminearum) was used as a control for the evaluation of the zearalenone biosynthesis potential. On the other hand, the liquid culture of the isolate KF-38/1 (F. sporotrichioides) was used as a control for both type-A trichothecenes (T-2 toxin and diacetoxyscirpenol - DAS). The obtained results show that F. poae, in contrast to F. graminearum, has no potential for the zearalenone biosynthesis. The presence of DAS was determined only in one isolate of F. poae (MRIZP-666), and in the control isolate of F. sporotrichioides (KF-38/1/R), that were grown in the GPY liquid medium. The T-2 toxin was detected in the isolate MRIZP-666, grown in both media, and in the isolates MRIZP-37 and MRIZP-860, cultured in the GPY and SPY liquid medium, respectively. The control culture KF-38/1/R (F. sporotrichioides) produced the T-2 toxin at the concentration of 4,000 μg L-1. According to the gained information, it can be concluded that the potential of F. poae for the type-A trichothecene biosynthesis was low, as the concentration of DAS or T-2 toxin did not exceed 80 μg L-1 or 240 μg L-1, respectively.U ovom radu je u in vitro uslovima proučena sposobnost izolata F. poae za biosintezu jedne grupe fuzariotoksina - trihotecena tipa A (T-2 toksin i diacetoksiscirpenol - DAS), kao i zearalenona (ZEA). Toksikološki profil ove vrste je kod nas nedovoljno ispitan s obzirom na njenu zastupljenost i toksigena svojstva prema literaturnim podacima. Proučavanjima je bilo obuhvaćeno 11 izolata F. poae, poreklom sa pšenice iz 9 lokaliteta, uglavnom sa područja Vojvodine. Najveći broj uzoraka prikupljen je 2005. godine, kada su klimatski uslovi pogodovali intenzivnijoj pojavi fuzarioza klasa ove poljoprivredne kulture. Za određivanje toksikološkog potencijala F. poae kulture odabranih izolata su gajene u tečnim podlogama (GPK i SPK) tokom 3 dana na sobnoj temperaturi (21-26°C) i na rotacionoj tresilici (180 obrtaja min-1). Sirovi toksini su izolovani iz filtrata tečnih kultura ispitanih izolata pomoću etil acetata, dok je kvantifikacija mikotoksina izvršena metodom tankoslojne hromatografije. Tečna kultura izolata GZ-LES (F. graminearum) je korišćena kao kontrolna kultura za utvrđivanje potencijala za biosintezu zearalenona, a KF-38/1/R (F. sporotrichioides) za oba trihotecena tipa A (T-2 toksin i DAS). Dobijeni rezultati pokazuju da F. poae, za razliku od F. graminearum, ne poseduje potencijal za biosintezu zearalenona. Prisustvo DAS-a je utvrđeno samo kod jedne kulture F. poae (MRIZP-666) i kontrolnog izolata F. sporotrichioides (KF-38/1/R) koji su gajeni u tečnom GPK medijumu. T-2 toksin je detektovan kod izolata MRIZP-666 pri gajenju u obema podlogama, kao i izolata MRIZP-37 u GPK, odnosno MRIZP-860 u SPK medijumu. Kontrolna kultura KF-38/1/R (F. sporotrichioides) proizvodila je T-2 toksin u koncentraciji od 4000 μg L-1. Na osnovu iznetih podataka može se zaključiti da je potencijal F. poae za biosintezu trihotecena tipa A bio nizak u datim uslovima s obzirom da koncentracija DAS-a nije prelazila 80 μg L-1, odnosno T-2 toksina 240 μg L-1. Imajući u vidu prikazane rezultate, smatramo da je za dobijanje konačnog odgovora na pitanje o toksikološkom profilu izolata F. poae u Srbiji neophodno preduzeti dodatna ispitivanja, ne samo sa novim izolatima iz godina koje slede, nego i u drugim uslovima kultivisanja, u prvom redu na sterilnom prirodnom supstratu kao što su pšenica i kukuruz

Radio-frequency electrical characterization of viable-cell suspensions

Suspended viable cells significantly influence the relative complex permittivity (dielectric constant) of the suspension. This influence depends on the concentration, viability, size, and even the cell shape. Hence, by measuring the permittivity, various fermentation processes can be monitored (e.g., in wine making and preparation of milk products). Thereby, the complex impedance (or admittance) between electrodes immersed into the suspension is measured and the permittivity is obtained by a suitable de-embedding procedure. There exist various data in the literature on this influence, but they do not provide sufficient information for wide implementation of the technique. The aim of the paper was to experimentally investigate various aspects of the measurement procedures and establish practical guidelines and limitations. Measurements were performed using an LCR meter, a classical vector network analyzer, and a nanoVNA analyzer, in the frequency range from 10 Hz to 100 MHz. Various suspension containers and systems of electrodes were designed and manufactured. Bipolar systems of electrodes were found to be more suitable for higher frequencies than tetrapolar ones. Open electrode systems were found to be more prone to the influence of the environment than shielded systems (e.g., coaxial). Good results were obtained for simple yeast suspensions. However, the presence of other cells and dispersed solids (e.g., in pressed grape juice) was found to completely overshadow the yeast cells. Electrochemical effects on the electrodes were found to be insignificant above around 200 kHz. Bubbles on electrodes, delamination, high concentrations of ions, and even temperature variations significantly hindered measurements in many cases

Presence of deoxynivalenol in winter wheat treated with fungicides

Natural occurrence of Fusarium spp. and concentrations of mycotoxin deoxynivalenol (DON) in the grain of the winter wheat moderately susceptible to Fusarium head blight (FHB) has been studied. Grain samples were collected from wheat crops intended mainly for human consumption. All wheat crops were treated with fungicides before (a.i. flutriafol - formulated as Fluoco, applied in dose of 0.5 l ha-1) and during the flowering phase of growing (a.i. thiophanate-methyl + epoxiconazole formulated as Eskorta plus and a.i. thiophanate-methyl formulated as Funomil, applied in doses of 0.75 and 0.5 l ha-1, respectivily). Among of Fusarium species only F. graminearum, as a well known producer of DON, was identified. This fungus was identified in 15 of 19 samples (78.9%) with incidence in positive samples of 2 to 28% (average, 14.0%). Presence of DON was established in 13 of a total 19 investigated wheat grain samples (68.4%). In positive samples DON was detected in concentrations from 69 to 918 μg kg-1 (average, 478 μg kg-1). DON showed a significant and positive correlation at P≥0.05 with grain moisture content (r = 0.52*). Between the frequency of F. graminearum and concentration of DON and between the frequency of F. graminearum and grain moisture content, positive correlation was determined, but without statistical significance (r = 0.44 and r = 0.29, respectively)