scholarly journals Screening Methods for Isolation of Biocontrol Epiphytic Yeasts against Penicillium digitatum in Lemons

2021 ◽  
Vol 7 (3) ◽  
pp. 166
Author(s):  
Martina María Pereyra ◽  
Mariana Andrea Díaz ◽  
Fabricio Fabián Soliz-Santander ◽  
Anja Poehlein ◽  
Friedhelm Meinhardt ◽  
...  
Keyword(s):  
Action Spectrum ◽  
Penicillium Digitatum ◽  
Rrna Genes ◽  
Screening Methods ◽  
Dual Culture ◽  
Citrus Fruits ◽  
Postharvest Diseases ◽  
Yeast Strains

Worldwide, the green rot caused by Penicillium digitatum is one of the most aggressive postharvest diseases of lemons. Searching for sustainable alternatives to chemical fungicides, epiphytic yeasts as potential biocontrol agents were isolated from citrus fruits using a tailor-made selective medium. For disclosing their antagonistic potential against P. digitatum, obtained isolates were subjected to direct screening methods, both in vitro and in vivo. In the course of the primary in vitro screening that comprised dual culture assays, 43 yeast strains displaying antagonistic activities against the pathogen were selected. Subsequently, such strains were subjected to an in vivo screening that consisted of a microscale test, allowing the selection of six yeast strains for further analysis. In the final screening using macroscale in vivo tests, three strains (AcL2, AgL21, and AgL2) displaying the highest efficiencies to control P. digitatum were identified. The protection efficiencies in lemons were 80 (AcL2), 76.7 (AgL21), and 75% (AgL2). Based on sequence analysis of the PCR amplified D1/D2 domains of the 26S rRNA genes, they were identified as representatives of the species Clavispora lusitaniae. Interestingly, the strains exhibited a broad action spectrum among citrus fruits as they were also able to combat the green mold disease in grapefruit and two orange varieties. The direct screening methods applied in this study favored the recovery of efficient candidates for application as biological control agents to combat fungal infestations of citrus fruits.

Inactivation of Penicillium digitatum and Penicillium italicum under In Vitro and In Vivo Conditions by Using UV-C Light

2013 ◽  
Vol 76 (10) ◽  
pp. 1761-1766 ◽  
Author(s):  
GÜLTEN TİRYAKİ GÜNDÜZ ◽  
FIKRET PAZIR
Keyword(s):  
Potato Dextrose Agar ◽  
Penicillium Digitatum ◽  
Citrus Fruits ◽  
Penicillium Italicum ◽  
Inoculation Methods ◽  
Penetration Ability ◽  
Working Distance ◽  
Uv C

In this study, the effects of UV-C on two of the main wound pathogens of citrus fruits, Penicillium digitatum and Penicillium italicum, were investigated with different inoculation methods in vitro and on oranges. P. digitatum and P. italicum spores were inoculated onto the surface of potato dextrose agar or oranges using spread, spot, wound, and piercing inoculation methods. UV-C treatment for 1 min from a working distance of 8 cm reduced the numbers of P. italicum and P. digitatum by about 3.9 and 5.3 log units, respectively, following spread inoculation under in vitro conditions. Significant reductions were obtained after 1-min UV-C treatments of the tested fungi following inoculation using the spread and spot methods. With inoculation by the wound and piercing methods, the tested spores were not inactivated completely even after 10- and 20-min treatment times, respectively. The application of UV-C (7.92 kJ m−2) on oranges reduced the percentage of oranges infected at least threefold compared with the rate of infection in the untreated control samples. UV-C irradiation could effectively inactivate spores of P. italicum and P. digitatum inoculated by the spread plate and spot inoculation methods under in vitro and in vivo conditions. On the other hand, because of the low penetration ability of UV-C light, the tested fungi were not completely inactivated following inoculation with the wound and piercing methods. UV-C treatment has potential for use in surface decontamination of citrus fruits.

scholarly journals Phytotoxic tryptoquialanines produced in vivo by Penicillium digitatum are exported in extracellular vesicles

2020 ◽  
Author(s):  
Jonas Henrique Costa ◽  
Jaqueline Moraes Bazioli ◽  
Luidy Darllan Barbosa ◽  
Pedro Luis Theodoro dos Santos Júnior ◽  
Flavia C. G. Reis ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Extracellular Vesicles ◽  
Metabolic Response ◽  
Penicillium Digitatum ◽  
Citrus Fruits ◽  
Green Mold ◽  
Host Pathogen ◽  
Citrus Seeds

ABSTRACTPenicillium digitatum is the most aggressive pathogen of citrus fruits. Tryptoquialanines are major indole alkaloids produced by P. digitatum. It is unknown if tryptoquialanines are involved in the damage of citrus fruits caused by P. digitatum. To investigate the pathogenic roles of tryptoquialanines, we initially asked if tryptoquialanines could affect the germination of Citrus sinensis seeds. Exposure of the citrus seeds to tryptoquialanine A resulted in a complete inhibition of germination and an altered metabolic response. Since this phytotoxic effect requires the extracellular export of tryptoquialanine A, we investigated the mechanisms of extracellular delivery of this alkaloid in P. digitatum. We detected extracellular vesicles (EVs) released by P. digitatum both in culture and during infection of citrus fruits. Compositional analysis of EVs produced during infection revealed the presence of a complex cargo, which included tryptoquialanines and the mycotoxin fungisporin. The EVs also presented phytotoxicity activity in vitro, and caused damage to the tissues of citrus seeds. Through molecular networking, it was observed that the metabolites present in the P. digitatum EVs are produced in all of its possible hosts. Our results reveal a novel phytopathogenic role of P. digitatum EVs and tryptoquialanine A, implying that this alkaloid is exported in EVs during plant infection.IMPORTANCEDuring the post-harvest period, citrus fruits can be affected by phytopathogens such as Penicillium digitatum, which causes the green mold disease and is responsible for up to 90 % of the total citrus losses. Chemical fungicides are widely used to prevent the green mold disease, leading to concerns about environmental and health risks. To develop safer alternatives to control phytopathogens, it is necessary to understand the molecular basis of infection during the host-pathogen interaction. In the P. digitatum model, the virulence strategies are poorly known. Here, we describe the production of phytotoxic extracellular vesicles (EVs) by P. digitatum during the infection of citrus fruits. We also characterized the secondary metabolites in the cargo of EVs and found in this set of molecules an inhibitor of seed germination. Since EVs and secondary metabolites have been related to virulence mechanisms in other host-pathogen interactions, our data are important for the comprehension of how P. digitatum causes damage to its primary hosts.

An Attempt at Biological Control of Blossom Blight of Rose Caused by Botrytis cinerea Using some Local Trichoderma spp. Strains

2020 ◽  
Vol 55 (1) ◽  
pp. 27-34
Author(s):  
G. Zadehdabagh ◽  
K. Karimi ◽  
M. Rezabaigi ◽  
F. Ajamgard
Keyword(s):  
Botrytis Cinerea ◽  
Mycelial Growth ◽  
Limiting Factor ◽  
Dual Culture ◽  
Trichoderma Spp ◽  
Khuzestan Province ◽  
Inhibitory Potential ◽  
Cut Rose

The northern of Khuzestan province in Iran is mainly considered as one of the major areas of miniature rose production. Blossom blight caused by Botrytis cinerea has recently become a serious limiting factor in rose production in pre and post-harvest. In current study, an attempt was made to evaluate the inhibitory potential of some local Trichoderma spp. strains against B. cinerea under in vitro and in vivo conditions. The in vitro results showed that all Trichoderma spp. strains were significantly able to reduce the mycelial growth of the pathogen in dual culture, volatile and non-volatile compounds tests compared with control, with superiority of T. atroviride Tsafi than others. Under in vivo condition, the selected strain of T. atroviride Tsafi had much better performance than T. harzianum IRAN 523C in reduction of disease severity compared with the untreated control. Overall, the findings of this study showed that the application of Trichoderma-based biocontrol agents such as T. atroviride Tsafi can be effective to protect cut rose flowers against blossom blight.

Synthesis of Novel 3(N,N-dialkylamino)alkyl/phenyl Substituted Thieno [2,3-d]pyrimidinones as H1-Anti-Histaminic and Antimicrobial Agents

2019 ◽  
Vol 15 (1) ◽  
pp. 63-70
Author(s):  
Shiv Dev Singh ◽  
Arvind Kumar ◽  
Firoz Babar ◽  
Neetu Sachan ◽  
Arun Kumar Sharma
Keyword(s):  
Antimicrobial Activity ◽  
Potassium Hydroxide ◽  
Antimicrobial Agents ◽  
Pyrimidine Ring ◽  
Antimicrobial Activities ◽  
Screening Methods ◽  
Chlorpheniramine Maleate ◽  
In Vivo Screening

Background: Thienopyrimidines are the bioisoster of quinazoline and unlike quinazoline exist in three isomeric forms corresponding to the three possible types annulation of thiophene to the pyrimidine ring viz thieno[2,3-d] pyrimidine, thieno[3,2-d] pyrimidine and thieno[3,4-d]pyrimidine. Heterocyclic containing the thienopyrimidinone moiety exhibits various pronounced activities such as anti-hypertensive, analgesic and anti-inflammatory, antiviral, platelet aggregation inhibitory, antiprotozoal bronchodilatory, phosphodiesterase inhibitory, antihistaminic, antipsychotic and antimicrobial activity. Objective: Synthesis of novel 3(N,N-dialkylamino)alkyl/phenyl substituted thieno[2,3-d]pyrimidinones as H1-anti-histaminic and antimicrobial agents. Methods: A series of 3-[(N,N-dialkylamino)alkyl/phenyl]-2-(1H)thioxo-5,6,7,8-tetrahydrobenzo(b) thieno(2,3-d)pyrimidine-4(3H)-ones[4a-d], their oxo analogous [5a-d] and 3-[(N,N-dialkylamino)alkyl]- 2-chlorophenyl-5,6,7,8-tetrahydrobenzo(b)thieno(2,3-d)pyrimidine- 4 (3H)-ones[6a-d]derivative were synthesized from 2-amino-4,5,6,7-tetrahydrobenzo(b)thiophene-3-carboxylic acid by nucleophilic substitution of different N,N-dialkyl alkylene/phenylene diamines on activated 3-acylchloride moiety followed by cyclocondensation with carbon disulfide and ethanolic potassium hydroxide to get [4a-d] and in second reaction by condensation with 4-chlorobenzoyl chloride to get [6a-d] by single pot novel innovative route. The oxo analogous [5a-d] were prepared by treating derivatives [4a-d] with potassium permagnate in ethanolic KOH. The synthesized compound were evaluated for H1-antihistaminic and antimicrobial activities. Results: All synthesized compounds exhibited significant H1-antihistaminic activity by in vitro and in vivo screening methods and data were verified analytically and statistically. The compound 4a, 4b, 5a and 5b showed significant H1-antihistaminiic activity than the reference standard chlorpheniramine maleate. The compound 6d, 6c, 5c and 4c exhibited significant antimicrobial activity.

scholarly journals In Vitro Probitotic Evaluation of Saccharomyces boulardii with Antimicrobial Spectrum in a Caenorhabditis elegans Model

Foods ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1428
Author(s):  
Ramachandran Chelliah ◽  
Eun-Ji Kim ◽  
Eric Banan-Mwine Daliri ◽  
Usha Antony ◽  
Deog-Hwan Oh
Keyword(s):  
Pancreatic Enzyme ◽  
Saccharomyces Boulardii ◽  
Antimicrobial Spectrum ◽  
Antimicrobial Efficiency ◽  
Heat Stable ◽  
Yeast Strains ◽  
Probiotic Yeast ◽  
Probiotic Strains

In the present study, we screened for potential probiotic yeast that could survive under extreme frozen conditions. The antimicrobial and heat-stable properties of the isolated yeast strains Saccharomyces boulardii (S. boulardii) (KT000032, KT000033, KT000034, KT000035, KT000036, and KT000037) was analyzed and compared with commercial probiotic strains. The results revealed that the tested S. boulardii KT000032 strain showed higher resistance to gastric enzymes (bile salts, pepsin, and pancreatic enzyme) at low pH, with broad antibiotic resistance. In addition, the strain also showed efficient auto-aggregation and co-aggregation abilities and efficient hydrophobicity in the in-vitro and in-vivo C. elegens gut model. Further, the KT000032 strain showed higher antimicrobial efficiency against 13 different enteropathogens and exhibited commensal relationships with five commercial probiotic strains. Besides, the bioactive compounds produced in the cell-free supernatant of probiotic yeast showed thermo-tolerance (95 °C for two hours). Furthermore, the thermo-stable property of the strains will facilitate their incorporation into ready-to-eat food products under extreme food processing conditions.

scholarly journals In vitro and in vivo Assay for Assessment of the Biological Control Potential of Streptomyces sp. KT

2017 ◽  
Vol 7 (1) ◽  
pp. 10
Author(s):  
Tatsuya Ohike ◽  
Minori Maeda ◽  
Tetsuya Matsukawa ◽  
Masahiro Okanami ◽  
Shin’ichiro Kajiyama ◽  
...  
Keyword(s):  
Biological Control ◽  
Suppressive Effect ◽  
Phytopathogenic Fungi ◽  
Dual Culture ◽  
Biological Control Agents ◽  
Bipolaris Oryzae ◽  
Biological Control Potential ◽  
Dual Culture Assay

Rhizoctonia solani is fungal plant pathogen that infects many different host plants. Recently, biological control agents that are friendly to the environment and ecosystems have attracted much attention as an alternative to the use of chemical fungicide which have been used worldwide to control soil borne pathogens including R. solani. In this study, 53 strains of actinomycetes isolated from environmental soils, and antifungal activities of them were assessed by the dual culture assay. Strain KT showed strong inhibitory activities against 8 phytopathogenic fungi. A great suppressive effect on R. solani growth was observed in the inoculation test of plants using cucumber and chin-geng-sai. In addition, infection of Bipolaris oryzae also could be suppressed in the detached leaf assay using oats. As a result of genetic analysis, it was shown that KT was a species closely related to Streptomyces lavenduligriseus NRRL B-3173T. However, as far as we know, there is no report for biological control agents using S. lavenduligriseus. This study suggests that the strain KT may useful as biological control agents to suppress various crop diseases.

scholarly journals Therapeutic Potential of CUDC-907 (Fimepinostat) for Hepatocarcinoma Treatment Revealed by Tumor Spheroids-based Drug Screening

2020 ◽  
Author(s):  
Wei Liao ◽  
Wanren Yang ◽  
Yue Zhang ◽  
Fanhong Zeng ◽  
Jiecheng Xu ◽  
...  
Keyword(s):  
Cell Lines ◽  
Drug Screening ◽  
Screening Method ◽  
Screening Methods ◽  
Spheroid Culture ◽  
3D Print ◽  
Culture Plate ◽  
Hcc Cells

Abstract Background: Cancer is the second leading cause of death globally. However, most of the new anti-cancer agents screened by traditional drug screening methods fail in the clinic because of lack of efficacy. One of the reasons for this dilemma is that the two-dimensional (2D) culture cancer cell lines could not represent the in vivo cancer cells well. Fortunately, the development of a three-dimensional (3D) culture technique helps in this problem. Methods: The high-throughput spheroid culture plate was fabricated by using 3D print technique and agarose. 4 hepatocarcinoma (HCC) cell lines were 3D cultured to screen 19 small molecular agents based on the spheroid culture plate. 3D cultured primary HCC cells and tumor-bearing mice model were established to verify the candidate anti-hepatocarcinoma agent. Cell function experiments and western blotting were conducted to explore the anti-hepatocarcinoma mechanism of the candidate agent. Results: Based on the previous study, we established an in vitro 3D drug screening method by using our invented spheroid culture device and found that CUDC-907 can serve as a potent anti-hepatocarcinoma agent. The study data show that CUDC-907 (fimepinostat), a novel dual acting inhibitor of phosphoinositide 3-kinase (PI3K) and histone deacetylase (HDAC), has potent inhibitory effects on HCC cell lines and primary HCC cells in vitro, Animal studies have shown that CUDC-907 can also suppress HCC cells in vivo. Furthermore, we investigated the antitumor mechanism of CUDC-907 in HCC cells. We found that it inhibits the PI3K/AKT/mTOR pathway and downregulates the expression of c-Myc, leading to the suppression of HCC cells. Conclusion: Our results suggest that CUDC-907 can be a candidate anti-HCC drug, and the 3D in vitro drug screening method based on our novel spheroid culture device is promising for drug screening.

scholarly journals Evaluation of in-vitro methods to select effective streptomycetes against toxigenic fusaria

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6905 ◽  
Author(s):  
Elena Maria Colombo ◽  
Cristina Pizzatti ◽  
Andrea Kunova ◽  
Claudio Gardana ◽  
Marco Saracchi ◽  
...  
Keyword(s):  
Plant Pathogens ◽  
Culture Media ◽  
Screening Methods ◽  
Dual Culture ◽  
In Planta ◽  
In Vitro Method ◽  
In Vitro Methods ◽  
Strain Diversity ◽  
Fungal Plant Pathogens

Biocontrol microorganisms are emerging as an effective alternative to pesticides. Ideally, biocontrol agents (BCAs) for the control of fungal plant pathogens should be selected by an in vitro method that is high-throughput and is predictive of in planta efficacy, possibly considering environmental factors, and the natural diversity of the pathogen. The purpose of our study was (1) to assess the effects ofFusariumstrain diversity (N= 5) and culture media (N= 6) on the identification of biological control activity ofStreptomycesstrains (N= 20) againstFusariumpathogens of wheat in vitro and (2) to verify the ability of our in vitro screening methods to simulate the activity in planta. Our results indicate that culture media,Fusariumstrain diversity, and their interactions affect the results of an in vitro selection by dual culture assay. The results obtained on the wheat-based culture media resulted in the highest correlation score (r= 0.5) with the in planta root rot (RR) inhibition, suggesting that this in vitro method was the best predictor of in planta performance of streptomycetes against Fusarium RR of wheat assessed as extension of the necrosis on the root. Contrarily, none of the in vitro plate assays using the media tested could appropriately predict the activity of the streptomycetes against Fusarium foot rot symptoms estimated as the necrosis at the crown level. Considering overall data of correlation, the activity in planta cannot be effectively predicted by dual culture plate studies, therefore improved in vitro methods are needed to better mimic the activity of biocontrol strains in natural conditions. This work contributes to setting up laboratory standards for preliminary screening assays ofStreptomycesBCAs against fungal pathogens.

scholarly journals Suppression of Stem-End Rot on Avocado Fruit Using Trichoderma spp. in the Central Highlands of Kenya

2021 ◽  
Vol 2021 ◽  
pp. 1-6
Author(s):  
E. K. Wanjiku ◽  
J. W. Waceke ◽  
J. N. Mbaka
Keyword(s):  
Growth Inhibition ◽  
Fungal Pathogen ◽  
Mycelial Growth ◽  
Culture Technique ◽  
Global Market ◽  
Dual Culture ◽  
Postharvest Diseases ◽  
Trichoderma Spp ◽  
Lasiodiplodia Theobromae

Demand for organic avocado fruits, together with stringent food safety standards in the global market, has made producers to use alternative, safe, and consumer-friendly strategies of controlling the postharvest fungal disease of avocado fruits. This study assessed the in vitro efficacy of Trichoderma spp. (T. atroviride, T. virens, T. asperellum, and T. harzianum) against isolated avocado stem-end rot (SER) fungal pathogens (Lasiodiplodia theobromae, Neofusicoccum parvum, Nectria pseudotrichia, and Fusarium solani) using a dual culture technique. The Trichoderma spp. were also evaluated singly on postharvest “Hass” avocado fruits. Spore suspension at 5 × 104 conidial/ml of the Trichoderma spp. was applied on the avocado fruits at three time points, twenty-four hours before the fungal pathogen (preinoculation), at the same time as the fungal pathogen (concurrent inoculation), and 24 hours after the fungal pathogen (postinoculation). In the in vitro study, T. atroviride showed the highest mycelial growth inhibition against N. parvum (48%), N. pseudotrichia (55%), and F. solani (32.95%), while T. harzianum had the highest mycelial growth inhibition against L. theobromae. Trichoderma asperellum was the least effective in inhibiting the mycelial growth of all the pathogens. Similarly, T. virens showed the highest mycelial growth inhibition against N. pseudotrichia at 45% inhibition. On postharvest “Hass” fruits, T. atroviride showed the highest efficacy against N. parvum, N. pseudotrichia, and F. solani in all the applications. Trichoderma virens and T. harzianum were most effective against all the pathogens during postinoculation, while Lasiodiplodia theobromae was best controlled by T. virens, T. harzianum, and T. asperellum during postinoculation. Both T. atroviride and T. harzianum present a potential alternative to synthetic fungicides against postharvest diseases of avocado fruits, and further tests under field conditions to be done to validate their efficacy. The possibility of using Trichoderma spp. in the management of SER on avocado fruits at a commercial level should also be explored.

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