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Direct, species-specific detection of Waitea circinata var. zeae
No full textThe species-specific detection of soil-borne plant pathogenic fungi is often challenging.
Many different detection procedures have been used, from direct microscopic examination,
incubation under wet conditions and isolation, to the application of serological and various
molecular methods. Most of these methodological approaches are not species-specific and have
to be combined in order to identify the causal agent. They are also time-consuming and require
experienced and trained personnel. The development of direct, rapid, easy-to-perform and
species-specific molecular tools would significantly speed up the work and expand the capacity of diagnostic laboratories. Waitea circinata Warcup & Talbot is a soil-borne pathogen that
comprises five varieties: circinata, oryzae, zeae, agrotis and prodigus, separated based on
differences in colony morphology and host range. W. circinata var. zeae is the teleomorph stage
of the anamorph Rhizoctonia zeae and is a pathogen that infects mainly monocotyledonous and
some dicotyledonous plants. Recently, this species was detected on new hosts for the first time
in the world, in Serbia in Futog and Rimski Sancevi, as the causal agent of root rot and decay of
cabbage and oilseed rape. As W. circinata var. zeae expands its host range and geographical
distribution, there is a need for a rapid species-specific diagnostic tool. In this study we have
developed conventional and real-time PCR protocols for direct detection from mycelium and
diseased plant tissue. To achieve this, we used mycelium and symptomatic tissue from oilseed
rape seedlings artificially inoculated with W. circinata var. zeae and different Rhizoctonia spp, as
well as with mycelia of of common soil-borne pathogens. We designed primer pair
zeaefor1/zeaerew1 that specifically amplified only target DNA region from isolates of W. circinata
var. zeae. Also, the newly designed primers proved to be specific as no positive reaction was
observed with DNA from the mycelium and plant tissue inoculated with isolates of 11 different
binucleate and multinucleate anastomosis groups of Rhizoctonia spp. (AG-A, AG-G, AG-F, AG-U,
AG-2-1, AG-2-2, AG-3, AG-4 HGI, AG-4 HGII, AG-4 HGIIl and AG-6), and neither with the mycelium
of common soil-borne pathogens (Fusarium spp, Alternaria spp., Macrophomina spp., Sclerotinia
spp., Phytophtora spp.). The conventional PCR protocol was less sensitive (detection limit of 102
than the real-time PCR protocol (10%), but both methods are useful tools for the direct detection
and identification of W. circinata var. zeae from mycelium and diseased oilseed rape seedlings, as
confirmed by sequencing (Acc. No. 0Q646649 and 0Q646548). Depending on the equipment
availability in the diagnostic laboratories, either a conventional or a real-time PCR detection
protocol will enable the detection, future research and control of W. circinata var. zeae
Beneficial microorganisms joining forces to fight green mold in mushrooms
No full textThe mushroom industry in Serbia is growing rapidly, producing fresh mushrooms of different quality for consumption,
processing or export. A lack of effective chemicals for disease control, and pathogen resistance to pesticides,
consequentially lead to unsuccessful control of the green mold agent, Trichoderma aggressivum. The principles
of sustainable use of pesticides require a change in the attitude of composters and mushroom growers towards
biological disease control. Altering the microbial comunities in compost and casing soil to stimulate the growth
of healthy button mushrooms will hopefully increase mushroom yield and green mold control, and provide a
practice-based understanding of the microbial community-pathogen relationships. The objective of the study was
to implement two indigenous beneficial microorganisms in the cultivation of button mushroom Agaricus bisporus:
Bacillus amyloliquefaciens, strain B-241, a bacterium with fungicidal activity against T. aggressivum, and Streptomyces
flavovirens, strain A06, an actinobacterium which stimulates mushroom yield. Interrelationship between
the two beneficial microorganisms was investigated in an experimental mushroom growing room after inoculation
with compost green mold, T. aggressivum f. europaeum, which was added one day after the spawned compost was
placed in boxes (106 conidia/mL per m2). The concentration of beneficial bacterial and actinobacterial suspension
was adjusted to 108 CFU/mL per m2 and applied at seven-day intervals. The synergy factor (SF) was calculated as
a ratio between the observed and expected impact on yield or effectiveness in suppression of disease symptoms.
The results indicated strong synergism in promoting A. bisporus productivity in both uninoculated (SF=1.56) and
inoculated plots (SF=1.62) after combined application of the two beneficial microorganisms. Regarding efficacy
in preventing green mold symptoms, the beneficial bacterium and actinobacterium displayed an additive effect
(SF=1.05). Further investigatation is expected to reveal adequate simultaneous application procedure of the beneficial
organisms in order to maximize biopesticidal/stimulatory effects, and reduce environmental pollution26-27 February 2024, Las Vegas, US
First Report of Collar and Root Rot of Lettuce Caused by Plectosphaerella cucumerina in Serbia
No full textIn March 2021, unusual plant stunting and collar and wet root rot of
lettuce (Lactuca sativa L.) during the rosette stage were observed in two
commercial fields in Serbia (44°589N, 20°329E; 44°459N, 20°439E). Disease
incidence in the fields ( 0.9 ha each) was approximately 15 and 20%,
respectively. Initial above-ground symptoms were yellowing and wilting of
leaves, while below-ground symptoms were collar and wet root rot and
necrotic lesions. Eventually, whole plants wilted, collapsed, and died. A total
of 35 symptomatic plants were collected from the fields and diseased tissues
were cut into small pieces, surface sterilized, and plated on potato dextrose
agar. Isolation resulted in 20 morphologically uniform monoconidial isolates.
The isolates formed white to creamy colonies, which gradually became
salmon pink, slimy, or moist in appearance, with sparse aerial mycelia.
Numerous hyphal coils with conidiophores and hyaline, smooth-surfaced,
ellipsoid to ovoid, septate or aseptate conidia were formed (4.5 to 10.1 ×
1.2 to 3.7 μm [n = 100]). To confirm the species identity, the internal
transcribed spacer (ITS) region and the D1/D2 region of a selected representative
isolate 13-3-c were amplified and sequenced by using the primer
pairs ITS1/ITS4 (White et al. 1990) and N1/N2 (O’Donnell and Gray 1995),
respectively. The sequences were deposited in GenBank (ITS, OR880564;
D1/D2, OR880567). Sequence analysis revealed 100% nucleotide identity
with Plectosphaerella cucumerina isolates from different countries
deposited in NCBI GenBank, including the isolates MH860704 (Vu et al.
2019) (ITS region) and KY662256 (Su et al. 2017) (D1/D2 region).
Neighbor-joining analysis was conducted based on combined ITS and D1/
D2 regions, and the tree was constructed with the substitution models
(1,000 bootstrap). Combined phylogeny confirmed that the sequences shared
a common clade with P. cucumerina. Hence, morphological, microscopic,
and molecular characterization confirmed the pathogen as P. cucumerina
(Carlucci et al. 2012; Palm et al. 1995). In a pathogenicity assay, 10 isolates
were tested. Five 30-day-old lettuce plants (cv. Majska Kraljica) per isolate
were root-dipped in the conidial suspensions (1 × 105 conidia/ml). The
10 inoculated plants were transplanted in l-liter pots containing the sterile
substrate (Floragard, Germany). Plants treated with sterile distilled water
were used as controls. Plants were maintained in a greenhouse at 25 to 28°C
under a 12-h photoperiod (Cai et al. 2021). Four weeks after inoculation,
stunting, chlorosis, and wilting of plants were observed, while collars and
roots exhibited typical decaying symptoms. No symptoms were observed on
the control plants. The pathogen was reisolated from symptomatic tissue as
previously described. Koch’s postulates were completed by confirming the
identity of reisolates based on the morphological features. To our knowledge,
this is the first report of P. cucumerina on lettuce or any other crops in
Serbia. P. cucumerina is already known as a pathogen of lettuce and other
hosts grown in many countries worldwide, as well as in some European
countries (Belgium, England, Italy, Netherland, and Switzerland) (Zhang
et al. 2019). This emerging pathogen may cause significant economic losses
in lettuce production in Serbia and in the entire Balkan region. Our results
may help to develop effective management strategies based on accurate and
timely identification and regular pathogen monitoring
Suzbijanje šampinjonske mušice i prouzrokovača zelene plesni šampinjona primenom 'Neem Cake' biopesticida
No full textPrevetnivna dezinfekcija pokrivke toksičnim formalinom i tretiranje pokrivke hemijskim
pesticidima je uobičajen postupak u zaštiti od prouzrokovača bolesti i štetočina. Nov
tehničko-tehnološki proces biološke zaštite šampinjona primenom 'Neem Cake'
biopesticida 2,5 % u sastavu pokrivke doprinosi rešavanju problema pojave zelene plesni T.
aggressivum i šampinjonske mušice L. ingenua u proizvodnji šampinjona. Proizvodnjom
šampinjona bez ostataka sintetskih hemijskih pesticida doprinosi se očuvanju životne
sredine, neciljanih organizama, zdravlja ljudi i povećava konkurentnost proizvođača u
regionu
Oxalic acid production and aggressiveness of Sclerotinia spp. isolates
No full textSclerotinia sclerotiorum and Sclerotinia minor are two necrotrophic fungal species that cause
white mould disease in many crops. Oxalic acid, secreted by Sclerotinia species, is recognized to
be a major virulence factor involved in the induction of cell death in host plants during infection
establishment. The aim of this study was to determine relationship between oxalic acid produc-
tion in S. sclerotiorum and S. minor isolates and their aggressiveness toward lettuce as a model
plant. Ten isolates of S. sclerotiorum and ten isolates of S. minor, derived from diseased lettuce
plants, were tested simultaneously for their ability to produce oxalic acid and to cause necrosis
in an inoculation experiment. Oxalic acid production was measured spectrophotometrically in a
liquid medium, while the inoculation test was conducted on lettuce leaf discs. As a control, sterile
liquid medium and PDA (Potato Dextrose Agar) plugs were used, respectively. Higher production
of oxalic acid was recorded in S. minor isolates, ranging from 19 to 96.3 μg oxalic acid mg−1 dry
mycelium weight, while S. sclerotiorum isolates produced significantly lower amount of oxalic acid
(21.9 to 29.9 μg oxalic acid mg−1 dry wt). All Sclerotinia spp. isolates were able to establish infec-
tion in inoculated leaf disks. On average, the diameter of necrosis was slightly higher for S. minor
isolates (11.7 mm), although the difference between species was not statistically significant. Cor-
relation between aggressiveness and oxalic acid production among Sclerotinia spp. isolates was
not recorded, suggesting the contribution of some other factors to the necrosis establishment
and spreading
Unapređenje strategije suzbijanja štetnih glodara: I - Izbor odgovarajućeg konzervansa za održivost baze mamka u nepovoljnim uslovima sredine
No full textPreservation of the appearance, structure and most importantly the attractiveness of baits
targeting harmful rodents over extended periods of their exposure is one of the main tasks
in the process of bait formulation. The impact of several preservatives, different methods of
homogenization of main bait components, and base-paraffin ratios on the extension of longevity
of bait base was examined under controlled laboratory conditions. Baits based on ground maize
grain were exposed to unfavourable environmental conditions, such as high temperature (30 °C-35 °C)
and air humidity (90%-95%). The exposure period was 9 days, and potential mold development
was monitored over the same period of time. Sodium benzoate was found deficient in providing
a satisfactory effect on bait longevity (7 days) and
sorbic acid, which provided the longest extension in bait sustainability (>9 days). Preservative
application can significantly extend bait longevity under unfavourable environmental conditions.
Extended bait functionality is important for rodent pest control procedures in habitats where
unfavourable environmental conditions prevail, such as sewers and water supply grids (collectors
and pumping stations), public areas, housing facilities (boiler rooms, moist sellers, etc.).Očuvanje izgleda, strukture i ponajviše atraktivnosti mamka za ciljane štetne vrste glodara
u dužem vremenskom periodu izlaganja jedan je od glavnih zadataka u procesu njihovog
formulisanja. U laboratorijskim, kontrolisanim uslovima ispitivan je uticaj različitih konzervanasa
na produženje perioda održivosti osnove mamka, način homogenizacije osnovnih komponenti
mamka kao i odnos baze i parafina na period održivosti mamka. Mamci na bazi mlevenog zrna
kukuruza izlagani su nepovoljnim uslovima sredine kao što su visoka temperatura (30° C-35° C)
i vlažnost vazduha (90%-95%). Period izlaganja bio je 9 dana, koliko je trajao i period praćenja
razvoja prouzrokovača plesni. Utvrđeno je da natrijum benzoat nema zadovoljavajući uticaj
na održivost mamka (7 dana) i sorbinske kiseline, koja
je uticala na najdužu održivost mamka (>9 dana). Primenom konzervanasa značajno se može
produžiti period održivosti mamka u nepovoljnim uslovima sredine. Produžena upotrebna
vrednost mamka, značajna je u praksi u postupku suzbijanja štetnih glodara na staništima
gde preovladavaju nepovoljni uslovi sredine, kao što su kanalizacioni i vodovodni sistemi
(kolektori, crpne stanice), javne površine, stambeni objekti (kotlarnice, vlažni podrumi i sl)
Pathogenicity of entomopathogenic nematodes to Phthorimaea operculella (Zeller) (Lepidoptera, Gelechiidae)
No full textEntomopathogenic nematodes (EPNs) are the most effective biological agents among
beneficial nematodes and, as they have no ecotoxicological side effects, are increasingly used
against various insect pests. The potato tuber moth, Phthorimaea operculella (Zeller)
(Lepidoptera, Gelechiidae), can be a serious pest of potato in the field and even more so in
warehouses during storage of potato tubers. In this work we present the results of the laboratory
test of virulence of ten populations of three EPN species to Phthorimaea operculella larvae at the
last instar stage. Two experiments were performed. In the first experiment, we tested the
virulence of commercial and Serbian and Italian native populations of Steinernema feltiae
(Filipjev), S. carpocapsae (Weiser) and Heterorhabditis bacteriophora Poinar. The bioassay was
performed in Petri dishes on filter paper with fresh infective juveniles (lJ) at four concentrations,
each in five replicates, repeating the entire experiment twice with each EPN population. The
second experiment was conducted by dipping P. operculella infested potato tubers in
suspensions of 1000 and 2000 IJ/ml to test the ability of two native populations of S. feltiae to
control the potato moth larvae in the tubers. The results showed high virulence of all EPN strains
against the larvae of P. operculella with 100% mortality at the highest concentration of 500 IJ/ml
with most populations tested. Immersion of tubers for five seconds in a suspension of 1000 J/ml
of both S. feltiae populations used, reduced the moth population by more than 50 percent. A
longer immersion time of 20 seconds in 2000 IJ/ml nematode suspension increased the mortality
of potato tuber moth larvae to 86 and 92 percent with populations TG4 and P9, respectively.
These results demonstrate the high virulence of EPN to P. opercullela and the potential of S.
feltiaeto be used for potato tuber moth control by dipping potato tubers before storage
The effect of aqueous extracts of hemlock (Conium maculatum L.) on seed germination of ragweed (Ambrosia artemisiifolia L.) and lettuce (Lactuca sativa L.)
No full textThe invasive weed species hemlock (Conium maculatum L.), fam. Apiaceae, is one of the most
toxic plants that is naturally widespread in Europe, Asia and Africa and has been introduced to
other parts of the world. It is also a common biennial weed in Serbia. Its bioactive compounds,
mainly alkaloids such as coniine, γ-coniceine and conhydrine, have phytotoxic properties that can
be utilized for bioherbicidal purposes. We investigated the potential phytotoxic effects of aqueous
extracts of hemlock leaves and stems on the germination of an invasive weed species – ragweed
(Ambrosia artemisiifolia L.), and on the germination of a commonly grown leafy vegetable – lettuce
(Lactuca sativa L.). The extract was obtained by immersing crushed dry leaves and stems in distilled
water (1:10, weight (g):volume (mL)) for 24 hours, and then filtered and diluted to 50%, 20%, 10%
and 5% concentrations. The hemlock leaf extract (20% and 50%) significantly inhibited the germi-
nation of ragweed (43% and 99%), while it inhibited the germination of lettuce only at the 50% di-
lution, where the inhibition was 100% compared to the control. In addition, the stem extract (5%,
10%, 20% and 50%) significantly inhibited ragweed germination (from 23% to 89%), while lettuce
germination did not change significantly, except at the 50% dilution, where it was inhibited by 9%.
The ability of the aqueous extract of hemlock leaves and stems to selectively inhibit ragweed seed
germination, while lettuce seed germination was not altered, could be useful for integrated weed
management strategies. Further research is essential to determine the appropriate concentration
of hemlock extract to maximize herbicide efficacy while minimizing negative effects on non-tar-
get plants and other organisms
Effects of leaf, stem and inflorescence water extracts of hemlock (Conium maculatum L.) on seed germination and seedling growth of chickweed (Stellaria media (L.) Vill.)
No full textChickweed (Stellaria media (L.) Vill.), which belongs to the Caryophyllaceae family, is one of the
most widespread weed species. It is native to Europe, including Serbia, and rapidly infests wheat,
barley, sugar beet and other fields, causing significant yield and quality losses. It has been ob-
served that seedlings and mature plants growing near hemlock (Conium maculatum L.), fam. Api-
aceae, exhibit stunted growth, reduced biomass and poor root development. Piperidine alkaloids,
coniine and γ-coniceine, are the major bioactive compounds in hemlock, whose occurrence in
different parts of the plant may vary depending on the age of the plant and environmental condi-
tions. In this study, we investigated the potential phytotoxic effects of leaf, stem and inflorescence
water extracts of hemlock (5%, 10%, 20% and 50%) on chickweed seed germination. In addition,
we studied the effect of hemlock leaf water extracts (5%, 10%, 20% and 50%) on the early devel-
opment of chickweed seedlings by measuring some growth parameters. Our work showed that
the water extract of inflorescence had the strongest inhibitory effect on seed germination, as the
highest concentration applied (50%) completely inhibited seed germination. The water extract of
leaf (50%), also, significantly inhibited seed germination (90%), while the water stem extract did
not affect seed germination compared to the control. In addition, the root growth of chickweed
seedlings was more inhibited by the water extract of hemlock leaf than shoots. Further research
is needed to fully understand the distribution of bioactive compounds in the different parts of
the plant. The development of standardized extraction and safety protocols will be crucial for the
further practical application of potential hemlock-based bioherbicides, which would represent a
more environmentally friendly alternative to synthetic herbicides
Description of fundatrix and sexual morphs of the Asian apricot aphid, Myzus mumecola (Matsumura, 1917) (Hemiptera: Aphididae)
No full textFundatrix and sexual morphs of the Asian apricot aphid, Myzus (Myzus) mumecola (Matsumura, 1917) are described from
Serbia with original, clarified slide mounts, and illustrations. The morphological identification of sexual morphs was
corroborated through COI barcode sequence analysis. The subgeneric position of this species is discussed and the transfer
to subgenus Nectarosiphon is proposed