Molecular approaches for mycotoxin risk reduction

Mycotoxin contamination in food is a serious concern for human and animal health. Today, we do not know how to detoxify food materials that are contaminated with mycotoxins in ways that retain their edibility. Therefore, avoiding mycotoxins from entering the food chain is an important approach. This needs early and easy identification, detection, and quantification of mycotoxinproducing fungi. The conventional methods for the identification, taxonomy, detection, and quantification of toxigenic fungi are challenging because they require a high level of expertise and a set of sophisticated equipment. The aim of current study was to use molecular-based approaches (as they are practical, rapid, and more reliable) to identify, classify, detect, and quantify mycotoxin-producing fungi including aflatoxin-producing Aspergillus and fumonisin- and trichothecene (TC)-producing Fusarium species. The results obtained from this study suggested that there are 2 main populations of F. graminearum in Europe. The population of the 3-acetyl-deoxynivalenol (3ADON) chemotype is dominant in northern Europe and it has probably recently been spreading from Finland to north-western Russia, while the population of the 15ADON chemotype is dominant in the central and southern Europe and it has been spreading to the Denmark and Norway. The results also suggested that the homogenization of the oat flour by milling with a 1 mm sieve is important for the reproducibility of deoxynivalenol (DON) and F. graminearum DNA levels, which is evident from a higher correlation between the DON and F. graminearum DNA levels in oat grain samples that were sieved after milling. In the thesis, the F. langsethiae isolate obtained from Iranian wheat was reidentified as F. sibiricum, and the identification was confirmed by IGS sequencing. This work reports the first record of F. sibiricum from Iran, outside northern Asia and Norway, and the first isolation of F. langsethiae (a European pathogen) from western Siberia. Large variations in the DON content and the amounts of F. graminearum DNA, and poor coefficient of determination (R2 ) between these were detected in oat grain when the RIDA®QUICK SCAN kit was used for DON content estimation. This study confirmed that the coefficient of determination was usually less when DNA or DON levels were estimated from oat flour that was not ground with 0.8 mm or 1 mm sieves. DON levels obtained with the Rida®Quick method were usually higher than those obtained with accredited GC-MS method in the Finnish oat, barley, and wheat samples. The homogenization of the oat flour by sieving is therefore likely to be connected to the variations in the DON detection. Also, it was suggested that the amounts of DON close to the legislative limits should be reconfirmed with accredited quantitative analyses. In addition, isolation, identification, detection, and quantification of Fusarium and Aspergillus isolates from Egypt and the Philippines maize, wheat, and soil samples were implemented in this thesis. A. parasiticus isolates, which produced higher amounts of aflatoxins, were only found in the Philippines.Elintarvikkeiden sisältämät mykotoksiinit ovat vakava huolenaihe ihmisten ja eläinten terveydelle. Nykyään emme tiedä, miten mykotoksiineja sisältäviä elintarvikkeita voi puhdistaa niin, että niiden käyttökelpoisuus säilyy. Siksi on tärkeää välttää mykotoksiinien pääsyä elintarvike-ketjuun. Tämä vaatii mykotoksiineja tuottavien sienten varhaista tunnistamista, detektointia ja kvantifiointia helposti käyttöönotettavilla menetelmillä. Perinteiset menetelmät tunnistusta, taksonomiaa, detektointia ja toksigeenisten sienten kvantifiointia varten ovat hyvin haastavia, koska ne vaativat huippuluokan osaamista ja laitteita. Työn tarkoituksena oli käyttää molekyylibiologisia menetelmiä, koska ne ovat käytännöllisiä, nopeita ja luotettavia tunnistamaan, luokittelemaan, havaitsemaan ja kvantifioimaan mykotoksiinia tuottavia sieniä, mukaan lukien aflatoksiinia tuottavia Aspergillus-homeita sekä fumonisiinia ja trikotekeeneitä tuottavia Fusarium-punahomeita. Väitöskirjatyön tulosten perusteella esitetään, että Euroopassa on kaksi F. graminearum-populaatiota. 3ADON-kemotyyppi on vallalla Pohjois-Euroopassa ja se on ilmeisesti levinnyt hiljattain Suomesta Luoteis-Venäjälle, kun taas 15ADON-kemotyyppi on vallalla Keski- ja EteläEuroopassa, ja se on hiljattain levinnyt Tanskaan ja Norjaan. Lisäksi esitetään, että kauranäytteiden homogenointi jauhamalla 1 mm:n seulalla näyttää olevan tärkeä DON- ja F. graminearum-DNApitoisuusmittausten toistettavuudelle, mikä on osoitettu kauran korkeammalla korrelaatiolla DON ja F. graminearum-DNA-pitoisuuksien välillä. F. langsethiae -kanta, joka eristettiin Iranissa vehnästä, tunnistettiin myöhemmin F. sibiricumiksi ja tunnistus varmistettiin IGS-sekvensoinnilla. Työssä raportoidaan tästä ensimmäisestä iranilaisesta F. sibiricum-kannasta, joka oli alun perin määritetty F. langsethiae-lajiin, sekä ensimmäisestä Euroopan ulkopuolelta löydetystä F. langsethiae-kannasta, joka löytyi Länsi-Siperiasta. Aikaisemmin on usein havaittu, että kauranjyvän F. graminearum-DNA- ja DONpitoisuuksien välillä on melko huono korrelaatio. Nyt saadut tulokset vahvistivat, että korrelaatio F. graminearum-DNA ja DON-pitoisuuden välillä oli pienempi, kun ne uutettiin kauroista, jota ei oltu jauhettu käyttäen 0,8 mm: n tai 1 mm:n seuloja. Rida®Quick-pikamääritysmenetelmällä mitatut DON-pitoisuudet, olivat tavallisesti korkeammat kuin akreditoitujen GC-MS-menetelmällä mitatut DONpitoisuudet suomalaisessa kaura-, ohra- ja vehnänäytteessä. Kaurajauhojen homogenointi seulan avulla on siten todennäköisesti yhteydessä Fusarium-DNA ja DON-määrien vaihteluihin. Pikamääritysmenetelmillä havaitut lähellä suurinta sallittua määrää olevat DON-pitoisuudet olisikin vahvistettava akreditoiduilla kvantitatiivisilla kromatografisilla analyyseillä. Ensimmäisessä julkaisussa Egyptistä ja Filippiineiltä peräisin olevista maissi-, vehnä- ja maaperänäytteistä eristettiin Fusarium–isolaatteja. Lisäksi Fusariumpunahomeita tunnistettiin ja fumonisiinia tuottavien Fusarium-punahomeiden DNA:n määrää mitattiin Egyptistä ja Filippiineiltä peräisin olevista maissi- ja vehnänäytteistä. A. parasiticus-isolaatteja, jotka tuottivat suurimmat mitatut aflatoksiinimäärät, löydettiin vain Filippiineiltä

Evolution Of Special Ruled Surfaces Via The Evolution Of Their Directrices In Euclidean 3-Space E3

In this paper, evolutions of ruled surfaces that are generated by the normal and binormal vector fields of space curve (normal and binormal surfaces) are presented. These evolutions of the ruled surfaces depend on the evolutions of their directrices. Geometric visualization of these ruled surfaces are presented. In addition, the conditions which make these surfaces of types inextensible, developable and minimal are obtained

The Idea of a Living Constitution in Jurisprudence and Constitutional Judiciary

تطل فكرة الدستور الحي برأسها تحت ظلال مبدأ أعلوية الدستور، هذا المبدأ الذي أريد به منذ البدء اسباغ الحماية على الوثيقة التي تؤسس لكيان الدولة وروحها، وتحصينها ضد تجاوزات السلطات الحاكمة، وحماية إرادة الشعب التي ترجمت في نصوص الدستور واتخذت من مباني ومعاني كلمات الدستور، مستقراً لها، غير ان هذا المبدأ لايمكن ان يقف عائقا امام حركة التطور الدستورية، فمهما بلغت اعلوية الدستور مقاماً مصوناً ضمن النظام القانوني في الدولة فإنها لا يمكن ان تؤدي باي حال من الاحوال الى جعل الوثيقة الدستورية وثيقة جامدة بالمطلق لان الدستور هو قانون في الاصل والقانون يستمد حياته من المجتمع، والمجتمع حركة متغيرة، بالتالي لابد لهذا القانون من المواكبة مع المستجدات ومحاولة استيعابها وتنظيمها بالشكل الأمثل، بحيث يصبح الدستور نابضا بالحياة لكي يتمكن من أداء الوظيفة التي وجد لأجلها الا وهي تنظيم الحياة في مجتمع ما تميز بشكل دولة. وتأسيساً على ما سبق بيانه، يتعرض البحث لفكرة الدستور الحي من خلال تناولها في ضوء اجتهادات الفقه واحكام وقرارات القضاء الدستوري.The idea of a living constitution is under the shadow of the constitutional supremacy, This principle, which was originally set to protect the document that establishes the state's entity and spirit and to protect the will of the people that has been translated into the constitution, but the principle of constitutional supremacy cannot transform the constitution into a dead rigid document, no matter how high the Constitution is, the latter is a law in its essence, the latter is a law in essence and the law derives its life and effectiveness from society. Society is a moving, innovative and evolving phenomenon. Therefore, this law must keep abreast of developments and try to assimilate them and organize them in an optimal manner, so that the constitution becomes viable so that it can perform the function that it has found for it, namely, organizing life in a society characterized as a state. Based on the above, the research explores the idea of a living constitution by addressing it in the light of jurisprudence and the rulings and decisions of the constitutional judiciary

Two-level scheduling scheme for integrated 4G-WLAN network

In this paper, a novel scheduling scheme for the Fourth Generation (4G)-Wireless Local Area Network (WLAN) network is proposed to ensure that end to end traffic transaction is provisioned seamlessly. The scheduling scheme is divided into two stages; in stage one, traffic is separated into Actual Time Traffic (ATT) and Non-Actual-Time Traffic (NATT), while in stage two, complex queuing strategy is performed. In stage one, Class-Based Queuing (CBQ) and Deficit Round Robin (DRR) are used for NATT and ATT applications, respectively to separate and forward traffic themselves according to source requirements. Whereas in the stage, two Control Priority Queuing (CPQ) is used to assign each class the appropriate priority level. Evaluation of the performance of the integrated network was done according to several metrics such as end-to-end delay, jitter, packet loss, and network’s throughput. Results demonstrate major improvements for AT services with minor degradation on NAT applications after implementing the new scheduling scheme

A Polyphasic Approach to Compare the Genomic Profiles of Aflatoxigenic and Non-Aflatoxigenic Isolates of Aspergillus Section Flavi

Aflatoxins (AF) are highly toxic compounds produced by Aspergillus section Flavi. Theyspoil food crops and present a serious global health hazard to humans and livestock. The aim ofthis study was to examine the phylogenetic relationships among aflatoxigenic and non-aflatoxigenicAspergillus isolates. A polyphasic approach combining phylogenetic, sequence, and toxin analyses wasapplied to 40 Aspergillus section Flavi isolates collected from eight countries around the world (USA,Philippines, Egypt, India, Australia, Indonesia, China, and Uganda). This allows one to pinpoint thekey genomic features that distinguish AF producing and non-producing isolates. Based on molecularidentification, 32 (80%) were identified as A. flavus, three (7.5%) as A. parasiticus, three (7.5%) asA. nomius and one (2.5%) as A. tamarii. Toxin analysis showed that 22 (55%) Aspergillus isolateswere aflatoxigenic. The majority of the toxic isolates (62.5%) originated from Egypt. The highestaflatoxin production potential was observed in an A. nomius isolate which is originally isolatedfrom the Philippines. DNA-based molecular markers such as random amplified polymorphic DNA(RAPD) and inter-simple sequence repeats (ISSR) were used to evaluate the genetic diversity andphylogenetic relationships among these 40 Aspergillus isolates, which were originally selected from80 isolates. The percentage of polymorphic bands in three RAPD and three ISSR primers was 81.9%and 79.37%, respectively. Analysis of molecular variance showed significant diversity within thepopulations, 92% for RAPD and 85% for ISSR primers. The average of Polymorphism InformationContent (PIC), Marker Index (MI), Nei’s gene diversity (H) and Shannon’s diversity index (I) in ISSRmarkers are higher than those in RAPD markers. Based on banding patterns and gene diversitiesvalues, we observed that the ISSR-PCR provides clearer data and is more successful in geneticdiversity analyses than RAPD-PCR. Dendrograms generated from UPGMA (Unweighted Pair GroupMethod with Arithmetic Mean) cluster analyses for RAPD and ISSR markers were related to thegeographic origin.</p

Different grain grinding methods affect detection of Fusarium graminearum DNA and mycotoxins

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Comparison of biomass and deoxynivalenol production of northern European and southern European Fusarium graminearum isolates in the infection of wheat and oat grains

The 3ADON chemotype of Fusarium graminearum predominates in northern Europe, whereas the 15ADON chemotype is predominant in central and southern Europe. Therefore, it has been suggested that there are two F. graminearum populations in Europe, which may have been specialized to different host plants. The aim of the present work was to test this hypothesis by comparing southern European isolates (15ADON chemotype) from southern Russia and northern European isolates (3ADON chemotype) from Finland in the infection of grains in wheat cultivar Wellamo and oat cultivar Venla. F. graminearum biomass levels were measured by TaqMan (2018) and SYBR Green (2019) qPCR, while DON levels were measured by chromatographic methods. Most of the qPCR and DON results are supporting the hypothesis that in F. graminearum the 15ADON isolates from southern Russia are more specialized to wheat than the 3ADON isolates from Finland. In oat, there were not as clear differences between the 15ADON and 3ADON isolates, but in 2018 higher F. graminearum DNA levels and in 2019 higher DON and F. graminearum DNA levels were found in oat samples inoculated with 3ADON isolates. Our results are in line with literature according to which F. graminearum DNA and DON levels are also highest in oat in northern Europe, while in southern Europe they are highest in wheat and maize