Bakteriofagi kao baktericidi u zaštiti bilja

Control of plant pathogenic bacteria is a serious problem in production of many agricultural crops. High multiplication rate, adaptability and life inside plant tissue make bacteria unsuitable and inaccessible for most of control measures. Consequently, the list of bactericides available for plant protection is very short. Lately, biological control measures have been intensively studied as a potential solution of the problem. Investigation of bacteriophages, viruses that attack bacteria, is a fast-expanding area of research in plant protection. Several experiments have shown that they can be used as a very efficient tool for control of plant pathogenic bacteria. The fact that they are widespread natural bacterial enemies, simple for cultivation and management, host-specific, suitable for integration with other control practices, human and environment friendly, provide a great advantage for the application of phages over other bactericides.Zaštita bilja od patogenih bakterija predstavlja značajan problem u poljoprivrednoj proizvodnji. Visoka stopa umnožavanja, prilagodljivost i život u unutrašnjosti biljnih tkiva čine bakterije veoma nepogodnim i nedostupnim za suzbijanje. Zahvaljujući tome, lista baktericida, koji se mogu primeniti u zaštiti bilja, veoma je kratka. Poslednjih godina, rešenje problema sve više se traži u primeni bioloških mera zaštite. Bakteriofagi, virusi koji parazitiraju bakterije, sve su češće predmet istraživanja fitopatologa. Eksperimentalno je pokazano da se mogu iskoristiti kao veoma efikasno sredstvo za kontrolu štetnih bakterija. Činjenice da su opšte rasprostranjeni prirodni neprijatelji bakterija, jednostavni za gajenje i održavanje, specifični prema domaćinu, pogodni za integraciju sa drugim merama zaštite, bezopasni po čoveka i druge činioce biosfere, daju im značajnu prednost nad drugim sredstvima baktericidnog dejstva

Uticaj etarskih ulja timijana, cimeta i karanfilića na porast micelije Colletotrichum acutatum

Effects of the volatile phase of thyme, cinnamon and clove essential oils on Colletotrichum acutatum were investigated. Mycelial disc was placed in the center of the Petri dish (V=66 ml) containing PDA. Different volumes of either non- or ethanol-diluted essential oils were placed on the inner side of the dish cover to obtain final concentrations of 153, 107, 76, 46, 15, 14, 12, 11, 7.6, 3.82, 1.53, 0.153 and 0.0153 μl/L of air. The dishes were sealed with Parafilm and incubated in up-side-down position. After 7 days of incubation, mycelial growth was recorded by measuring the colony diameter. If no mycelial growth was recorded, the disc was transferred to a new PDA plate in order to evaluate whether the activity was either fungistatic or fungicidal. Mean growth values were obtained and then converted to inhibition percentage of mycelial growth compared with the control treatment. All the tested essential oils inhibited mycelial growth of C. acutatum in the dose dependent manner. Mycelial growth was totally inhibited by thyme oil in the concentration of 76 μl/L of air. The same results were obtained by cinnamon and clove oil in the concentration of 107 μl/L of air. Thyme and cinnamon oil had fungicidal effect in concentrations of 107 and 153 μl/L respectively. The results obtained provide evidence on the antifungal in vitro effect of the tested essential oils as potential means for the control of C. acutatum.Proučavan je efekat isparljive faze etarskih ulja timijana, cimeta i karanfilića na prouzrokovača antraknoze jagode Colletotrichum acutatum u uslovima in vitro. Fragmenti micelije gljive, prečnika 1 cm, zasejani su na KDA podlogu u petri-kutije (V = 66 ml). Različite koncentracije etarskih ulja (153, 107, 76, 46, 15, 14, 12, 11, 7,6, 3,82, 1,53, 0,153 i 0,0153 μl/l vazduha) dobijene su nanošenjem određene količine ulja, koncentrovanih ili razblaženih u etanolu, na središnji deo unutrašnje strane poklopca. Petri-kutije su zatvorene parafilmom i postavljene u obrnuti položaj, a porast micelije je meren posle 7 dana inkubacije. U kulturama u kojima nije došlo do porasta micelije fungicidno ili fungitoksično dejstvo određeno je prebacivanjem fragmenata na KDA podloge bez prisustva ulja. Efekat etarskih ulja predstavljen je procentom inhibicije porasta micelije, poređenjem sa kontrolom. Proučavana etarska ulja timijana, cimeta i karanfilića su svojom gasovitom fazom inhibirala porast micelije C. acutatum. Stepen inhibicije zavisio je od primenjene koncentracije. Etarsko ulje timijana je potpuno inhibiralo porast micelije pri koncentraciji 76 μl/l vazduha, a cimeta i karanfilića pri koncentraciji 107 μl/l vazduha. Fungicidno dejstvo ispoljilo je etarsko ulje timijana u koncentraciji 107 μl/l vazduha i cimeta u koncentraciji 153 μl/l vazduha. Rezultati istraživanja in vitro pokazali su antifungalno dejstvo testiranih etarskih ulja, što govori o potencijalu korišćenja etarskih ulja u kontroli C. acutatum

Pectobacterium carotovorum subsp. carotovorum – prouzrokovač vlažne truleži biljaka kale u Srbiji i Crnoj Gori

Bacterial strains were isolated from above- and underground parts of diseased calla plants originating from different localities in Serbia and one locality in Montenegro. They were characterized by studying their pathogenic, cultural, biochemical and physiological characteristics. All investigated strains caused soft rot of calla leaf stalks, potato slices and aloe leaves, and induced hypersensitive reaction on tobacco. Bacteriological properties of the strains indicated that symptoms on calla plants were caused by Gram-negative, nonfluorescent, oxidase negative, catalase positive and facultatively anaerobic bacterium belonging to the genus Pectobacterium. The investigated strains grew at 37ºC and in 5% NaCl, utilised lactose and trechalose, and produced neither indol nor lecitinase. These results, as well as the characteristic growth on Logan’s differential medium indicated that soft rot of tuber and stem base of calla plants was caused by Pectobacterium carotovorum subsp. carotovorum. This is the first report of this pathogen affecting calla plants in Serbia.Proučene su patogene, odgajivačke i biohemijsko-fiziološke odlike sojeva izolovanih iz nadzemnih i podzemnih delova obolelih biljaka kale gajene u različitim lokalitetima u Srbiji i jednom lokalitetu u Crnoj Gori. Svi proučavani sojevi ispoljili su izraženu pektolitičku aktivnost prouzrokujući vlažnu trulež lisnih drški kale, kriški krompira, liski aloje ili sanseverije, a prouzrokovali su i hipersenzitivnu reakciju duvana. Proučavanjem bakterioloških karakteristika utvrđeno je da promene na biljkama kale prouzrokuju Gram-negativni, nefluorescentni, oksidaza-negativni, katalaza-pozitivni i fakultativno-anaerobni sojevi bakterije, koja prema navedenim karakteristikama pripada rodu Pectobacterium. Proučavani sojevi se razvijaju pri 37ºC i u prisustvu 5% NaCl, razlažu laktozu i trehalozu, ne stvaraju indol i lecitinazu. Navedeni rezultati, kao i karakterističan razvoj na Loganovoj diferencijalnoj podlozi ukazuju da je vlažnu trulež korena i prizemnog dela biljaka kale prouzrokovala bakterija Pectobacterium carotovorum subsp. carotovorum. Ovo je prvo saopštenje o pojavi P. c. ssp. carotovorum kao patogena kale u Srbiji

Diferencijacija fitopatogenih vrsta roda Agrobacterium

Due to the difficulties in differentiation of phytopathogenic Agrobacterium spp. and lack of a standardized protocol, we carried out selection and evaluation of suitable methods based on the bacterial physiological, genetic and pathogenic properties. Strains of Agrobacterium tumefaciens, A. rhizogenes and A. vitis were differentiated using standard bacteriological and molecular methods. The biochemical and physiological tests confirmed authenticity of the strains. Two duplex PCR methods were conducted with four different primer pairs. In all strains, presence of plasmid virD2 and virC pathogenicity genes was detected. Chromosomal pehA gene was determined in A. vitis strain. Pathogenicity was confirmed on carrot slices and young plants of tomato and sunflower. Strains of A. tumefaciens and A. vitis were pathogenic on all test plants, while strain of A. rhizogenes induced characteristic symptoms only on carrot slices. The tests used in this study provided reliable discrimination between the three species and confirmed their identity as tumorigenic (Ti) Agrobacterium tumefaciens and A. vitis, and rhizogenic (Ri) A. rhizogenes.Usled poteškoća u razlikovanju vrsta roda Agrobacterium i nedostatka standardizovanog protokola izvršena je procena i odabir pogodnih metoda u cilju njihove diferencijacije na osnovu fizioloških, genetskih i patogenih odlika. Diferencirani su sojevi Agrobacterium tumefaciens, A. rhizogenes i A. vitis primenom standardnih bakterioloških i molekularnih metoda. Primenom diferencijalnih testova sojevi su ispoljili očekivane biohemijsko-fiziološke karakteristike. Izvedene su dve „duplex“ PCR metode sa 4 različita tipa prajmera. Kod proučavanih sojeva detektovano je prisustvo virD2 i virC gena patogenosti, koji se nalaze na plazmidnoj DNK bakterije. Prisustvo hromozomskog gena pehA, odgovornog za sintezu enzima poligalakturonaze, utvrđeno je kod soja A. vitis. Patogenost je proverena na kriškama mrkve i mladim biljkama paradajza i suncokreta. Sojevi A. tumefaciens i A. vitis bili su tumorogeni na svim test biljkama, dok je soj A. rhizogenes ispoljio patogenost jedino na kriškama mrkve. Na osnovu dobijenih rezultata, proučavani sojevi diferencirani su kao tumorogeni (Ti) Agrobacterium tumefaciens i A. vitis, i kao rizogeni (Ri) A. rhizogenes

The effect of corn grain micronization on diet digestibility and blood biochemical parameters in weaned Holstein calves

Aim of study: To evaluate corn grain micronization for calves fed a grower diet. Area of study: Padinska Skela – Belgrade, Serbia. Material and methods: Thirty weaned Holstein dairy calves (65–74 days of age) were randomly assigned to one of two treatments with growers containing micronized (MCG) or untreated corn grain (UCG). The experimental period lasted for 60 days. Main results: The values of total tract apparent digestibility of dry matter (DM), organic matter (OM), crude protein (CP), and nonfiber carbohydrates (NFC) were higher for calves fed MCG versus those within the UCG treatment by 3.9% (p<0.05), 7.0% (p<0.01), 7.1% (p<0.01) and 7.5% (p<0.05), respectively, for the days 25–30 of the experimental period. In addition, the values of digestibility of OM, CP, and NFC were higher by 4.9% (p<0.05), 5.7% (p<0.05), and 6.0% (p<0.05), respectively, for the days 55–60 of the experimental period. The density of metabolizable energy, net energy for maintenance and gain in consumed dietary DM was higher (p<0.001) by 4.7, 5.5, and 7.2%, respectively for calves fed on the grower containing micronized corn grain (MCG), during the first digestibility period, and by 3.0, 3.6, and 4.6%, respectively, during the second digestibility period. Energy intake was lower (p<0.05) during the second digestibility period, for calves fed a diet with micronized corn. Blood urea N was affected (p<0.001) by dietary treatments. Lower values (10.2%) were observed for calves fed the grower containing MCG. Research highlights: The micronization of corn grain is a useful tool for optimizing weaned calf production due to the improvement in the digestibility and energy content of the ration. © 2023 CSIC

61 Analiza masnih kiselina sojeva Erwinia amylovora iz Srbije i Crne Gore

Automated method of fatty acid analysis was used to identify and study heterogeneity of 41 Erwinia amylovora strains, originating from 8 plant species grown in 13 locations in Serbia and one in Montenegro. All strains contained 14:0 3OH fatty acid, characteristic for the “amylovora” group. According to fatty acid composition 39 strains were identified as E. amylovora as the first choice from the database. Due to their specific fatty acid composition, two strains were identified as E. amylovora, but as a second choice. Fatty acid analysis also showed that E. amylovora population from Serbia could be differentiated in three groups, designated in this study as α, β and γ. All strains originating from central or south Serbia, as well as four strains from north Serbia clustered into group α. Group β and γ contained only strains isolated in northern Serbia (Vojvodina). The results show that E. amylovora population in this area is heterogeneous and indicate pathogen introduction from different directions. Fatty acid analysis enabled identification at species level, as well as new insights of heterogeneity of E. amylovora population.Automatizovana metoda analize masnih kiselina primenjena je za identifikaciju i proučavanje heterogenosti Erwinia amylovora. Kao materijal za analizu prikupljen je 41 soj E. amylovora izolovan iz 8 različitih vrsta domaćina gajenih u 13 lokaliteta u Srbiji i jednom lokalitetu u Crnoj Gori. Rezultati ukazuju da svi proučavani sojevi poseduju 14:0 3OH masnu kiselinu, koja je karakteristična za „amylovora“ grupu. Na osnovu sastava masnih kiselina 39 sojeva je identifikovano kao E. amylovora, kao prvi izbor iz baze podataka. Dva soja su identifikovana kao E. amylovora, ali tek kao drugi izbor iz baze podataka, što je najverovatnije posledica specifičnosti u sastavu njihovih masnih kiselina. Rezultati analize masnih kiselina takođe pokazuju da populacija E. amylovora poreklom iz Srbije nije homogena i da među sojevima postoje tri grupe ili profila, koji su u ovom radu obeleženi sa α, β i γ. Svi sojevi koji su izolovani na prostoru centralne ili južne Srbije pripadaju grupi α, kao i četiri soja izolovana na području Vojvodine. Grupama β i γ pripadaju samo sojevi izolovani na području Vojvodine. Dobijeni rezultati predstavljaju dokaz heterogenosti populacije E. amylovora na ovim prostorima i ukazuju na mogućnost prodora patogena u naše područje iz različitih pravaca. Analiza masnih kiselina omogućila je ne samo identifikaciju do nivoa vrste, već i nova saznanja o heterogenosti populacije E. amylovora na ovim prostorima

Karakterizacija sojeva Xanthomonas euvesicatoria, patogena paprike u Srbiji

During spring and summer of 2008, 101 bacterial strains was isolated from the diseased pepper leaves collected from different pepper growing areas in the Republic of Serbia. The aim of this research was to characterize the isolated strains and determine their taxonomic position according to the most recent nomenclature. Pathogenic, biochemical and physiological characteristics of isolated bacteria were tested using standard bacteriological tests. The pathogen races were determined according to the reaction of differential varieties of Early Calwonder (ECW), their isogenic lines (ECW-10R, ECW-20R, ECW-30R) and Capsicum pubescens. The sensitivity of strains to bactericides was studied in vitro by culturing bacteria on sucrose pepton agar (SPA) plates, amended with filter-sterilized aqueous solution of streptomycin and kasugamycin (50, 100, 200 ppm) or copper-sulphate (100, 200 ppm). Based on pathogenic, biochemical and physiological characteristics, the investigated strains belonged to Xanthomonas euvesicatoria. The reaction of pepper differential varieties indicated that these strains belonged to pepper races P1, P3, P7, P8. Streptomycin resistant strains were not detected, but 6 strains were resistant to kasugamycin (50 ppm) and 13 strains to copper-sulphate (200 ppm), indicating bacterial resistance development.Tokom 2008. godine prikupljeni su uzorci obolelog lišća paprike sa simptomima bakteriozne pegavosti iz različitih lokaliteta Republike Srbije. Izolacijom iz zaraženih listova dobijen je 101 soj bakterija. Cilj ovog rada je proučavanje odlika i identifikacija izolovanih sojeva bakterije u skladu sa najnovijom nomenklaturom. Patogene i biohemijsko-fiziološke karakteristike sojeva proučene su korišćenjem standardnih bakterioloških testova. Određivanje fizioloških rasa bakterijske populacije izvršeno je na osnovu reakcije diferencijalnih sorti paprike Early Calwonder (ECW), njenih izogenih linija (ECW-10, ECW-20 ECW-30) i reakcijom sorte Capsicum pubescens. Proučena je osetljivost sojeva u in vitro uslovima na streptomicin, kasugamicin i bakar-sulfat. Rezultati proučavanja biohemijsko-fizioloških odlika sojeva ukazuju da naši sojevi pripadaju genetičkoj grupi „A“, odnosno vrsti Xanthomonas euvesicatoria. Proučavani sojevi, takođe, predstavljaju heterogenu populaciju u kojoj su zastupljene četiri fiziološke rase bakterije X. euvesicatoria (P1, P3, P7, P8). Sojevi rezistentni na streptomicin nisu detektovani ovim istraživanjima. Otpornost na 50 ppm kasugamicina utvrđena je kod 6 sojeva, a 13 sojeva je bilo otporno na 200 ppm bakar-sulfata. Prikazani rezultati ukazuju na opasnost od razvoja rezistentnosti bakterija na ova jedinjenja

Volcanic impacts on peatland microbial communities: A tephropalaeoecological hypothesis-test

Volcanic eruptions affect peatlands around the world, depositing volcanic ash (tephra) and a variety of chemicals including compounds of sulphur. These volcanic impacts may be important for many reasons, in particular sulphur deposition has been shown to suppress peatland methane flux, potentially reinforcing climatic cooling. Experiments have shown that sulphur deposition also forces changes in testate amoeba communities, potentially relating to the reduced methane flux. Large volcanic eruptions in regions with extensive peatlands are relatively rare so it is difficult to assess the extent to which volcanic eruptions affect peatland microbial communities; palaeoecological analyses across tephra layers provide a means to resolve this uncertainty. In this study, testate amoebae were analysed across multiple monoliths from a peatland in southern Alaska containing two tephras, probably representing the 1883 eruption of Augustine Volcano and a 20th Century eruption of Redoubt Volcano. Results showed relatively distinct and often statistically significant changes in testate amoeba community coincident with tephra layers which largely matched the response found in experimental studies of sulphur deposition. The results suggest volcanic impacts on peatland microbial communities which might relate to changes in methane flux

Differentiation of Pseudomonas syringae Pathovars Originating from Stone Fruits

Due to an overlapping host range, similar symptomatology and many common characteristics,Pseudomonas syringae pathovars originating from stone fruits can easily be misidentified.In order to select tests for rapid and efficient differentiation of P. s. pvs. syringae,morsprunorum and persicae, we studied the suitability and differentiating potential ofsome standard bacteriological and molecular methods. Differentiation of the strains wasperformed using LOPAT, GATTa and ice nucleation tests, nutrient sucrose broth growthand utilization of various carbon sources. PCR method enabled the detection of toxin-producinggenes: syrB and syrD in P. s. pv. syringae, and cfl gene in P. s. pv. morsprunorum race1. Syringomycin production by pv. syringae was confirmed in bioassay using Geotrichumcandidum, Saccharomyces cerevisiae and Rhodotorula pilimanae as indicator organisms.Pathogenicity test on lemon and immature nectarine fruits, as well as on string bean pods,showed different intensity of reaction of the inoculated material which could separate pv.syringae from the other two pathovars. PCR-based repetitive sequences, Rep-PCR withREP, ERIC and BOX primers revealed different genetic profiles within P. syringae pathovars

Complete genome and plasmid sequence data of three Xanthomonas arboricola pv. corylina strains, the bacterium responsible for bacterial blight of hazelnut

Xanthomonas arboricola pv. corylina is the causal agent of bacterial blight of hazelnut. The bacterium is listed as A2 quarantine pathogen in Europe since 1978 and on the Regulated Non-Quarantine Pest (RNQP) list since 2020. Three strains from various geographic regions and isolated at different times were sequenced using a hybrid approach with short- and long-read technologies to generate closed genome and plasmid sequences in order to better understand the biology of this pathogen