scholarly journals Ochratoxigenic fungi and Ochratoxin A determination in dried grapes marketed in Tunisia

2020 ◽  
Vol 70 (1) ◽  
Author(s):  
Samir Chebil ◽  
Wafa Rjiba-Bahri ◽  
Souheib Oueslati ◽  
Hanen Ben Ismail ◽  
Anis Ben-Amar ◽  
...  
Keyword(s):  
Aspergillus Niger ◽  
Ochratoxin A ◽  
High Performance ◽  
Its Region ◽  
Production Test ◽  
Potential Health ◽  
Dried Grapes ◽  
Pcr Product ◽  
Polymerase Chain

Abstract Purpose With the present work, we aimed to assess the occurrence of ochratoxigenic fungi and Ochratoxin A (OTA) in dried grapes from Tunisia. Methods Dried grapes samples (n = 90) were investigated for the presence of ochratoxigenic fungi, which were further characterized at the species level through amplification of the internal transcribed spacer (ITS) region and polymerase chain reaction (PCR) product sequencing. Fungal isolates were tested for their ochratoxigenic potential by high-performance liquid chromatography with fluorescence detection (HPLC-FLD), as well as dried grapes samples after an immunoaffinity column (IAC) clean-up procedure. Results Black Aspergilli isolates were the dominant genre among the filamentous fungi found in dried grapes samples and were the only OTA-producing fungi encountered. Aspergillus niger aggregate were the most frequently found isolates reaching 70%, 80%, and 85% in dried grapes samples from regions of Kelibia, Sfax, and Rafraf, respectively, while covered 100% of the relevant mycobiota found in imported samples. Aspergillus carbonarius isolates were found only in Sfax’s and Kelibia’s samples, while uniseriate Aspergilli were found between 7 and 20% in dried grapes from Kelibia, Sfax, and the imported samples. The in vitro OTA production test showed that 88.9% of OTA-producing isolates belonged to A. carbonarius with OTA levels varying from 0.06 to 1.32 μg/g of Czapek Yeast Agar (CYA). The remaining OTA-producing fungi (11.1 %) belonged to A. niger aggregate group having a maximum OTA potential of 2.88 μg/g CYA, and no uniseriate Aspergilli isolate was able to produce OTA. All dried grapes samples were free of OTA presence. Conclusion According to the present study’s findings, no OTA contamination was recorded in the investigated samples from Tunisian market. Nevertheless, the presence of strong OTA producers A. carbonarius in samples originated from the two out of three studied Tunisian regions, as well the high incidences of Aspergillus niger aggregate group with an attested potential for OTA production in all samples, necessitates further research on Tunisian dried grapes. Additionally, a continuous analysis of staple food of the Mediterranean diet is imperative to insure the best quality for the consumers and prevent potential health problems.

In Vitro Removal of Ochratoxin A by Wine Lactic Acid Bacteria

2007 ◽  
Vol 70 (9) ◽  
pp. 2155-2160 ◽  
Author(s):  
VINCENZO DEL PRETE ◽  
HECTOR RODRIGUEZ ◽  
ALFONSO V. CARRASCOSA ◽  
BLANCA de las RIVAS ◽  
EMILIA GARCIA-MORUNO ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Ochratoxin A ◽  
High Performance ◽  
Culture Medium ◽  
Degradation Products ◽  
Cell Binding ◽  
A Cell ◽  
In Vitro Interaction

A study was carried out to determine the in vitro interaction between ochratoxin A (OTA) and wine lactic acid bacteria (LAB). Fifteen strains belonging to five relevant oenological LAB species were grown in liquid synthetic culture medium containing OTA. The portion of OTA removed during the bacterial growth was 8 to 28%. The OTA removed from the supernatants was partially recovered (31 to 57%) from the bacterial pellet. Cell-free extracts of three representative strains were produced by disrupting cells in a French pressure cell. The ability of crude cell-free extracts to degrade OTA was studied. OTA was not degraded by cell-free extracts of wine LAB strains, and no degradation products of OTA were detected in the high-performance liquid chromatograms of the methanol extract of the bacterial pellet. On the basis of these results, we conclude that OTA removal by wine LAB is a cell-binding phenomenon. The chemistry and the molecular basis of OTA binding to wine LAB remains unknown.

Automated closed-vessel system for in vitro diagnostics based on polymerase chain reaction

1993 ◽  
Vol 39 (9) ◽  
pp. 1927-1933 ◽  
Author(s):  
J B Findlay ◽  
S M Atwood ◽  
L Bergmeyer ◽  
J Chemelli ◽  
K Christy ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Pcr Amplification ◽  
Automated System ◽  
Closed Vessel ◽  
Chain Reaction ◽  
Panel Testing ◽  
Pcr Product ◽  
Pcr Products ◽  
Polymerase Chain

Abstract An automated system for polymerase chain reaction (PCR) amplification and detection combats false-positive results caused by "PCR product carryover." The system uses a single vessel for both PCR amplification and the subsequent detection of PCR products, eliminating the need to handle PCR products in an open environment and risk product carryover. The sample and PCR reagents are introduced into one compartment within the vessel, and amplification occurs as they are thermally cycled. Other compartments contain the reagents for detection of PCR products. Pressure from a roller provides for sequential delivery of the contents of the compartments to a detection area. The PCR products are biotinylated at their 5' ends during amplification through the use of biotinylated primers. After delivery to the detection area, they are specifically captured by hybridization with immobilized oligonucleotide probes. Subsequent reaction with streptavidin-horseradish peroxidase conjugate forms a complex that catalyzes dye formation from dye precursor. Wash steps minimize nonspecific background. This format is amenable to multiplexing, permitting internal controls, speciation of bacteria, typing of viruses, and panel testing. An HIV assay performed with this system demonstrated 100% sensitivity and 95% specificity for 64 patients' samples relative to a conventional PCR assay based on 32P solution hybridization. Similarly, an automated closed-vessel assay of cytomegalovirus exhibited 97.5% sensitivity and 100% specificity.

scholarly journals Other Natural Hosts of Potato virus T

Plant Disease ◽  
2000 ◽  
Vol 84 (7) ◽  
pp. 736-738 ◽  
Author(s):  
C. Lizárraga ◽  
M. Querci ◽  
M. Santa Cruz ◽  
I. Bartolini ◽  
L. F. Salazar
Keyword(s):  
Potato Virus ◽  
Germplasm Bank ◽  
Chain Reaction ◽  
Oxalis Tuberosa ◽  
Pcr Product ◽  
Ullucus Tuberosus ◽  
Natural Hosts ◽  
Polymerase Chain ◽  
Andean Tuber Crops

Potato virus T (PVT), a member of the genus Trichovirus, was isolated from leaves of naturally infected ulluco (Ullucus tuberosus), oca (Oxalis tuberosa), and mashua (Tropaeolum tuberosum). These Andean tuber crops are often grown in small plots in association with potato (Solanum tuberosum) in the Peruvian highlands. PVT isolates from ulluco, oca, mashua, and potato infected virus-free ulluco, oca, and potato genotypes by mechanical inoculation. The incidence of PVT in mashua, oca, and ulluco accessions from the International Potato Center (CIP) in vitro germplasm bank was less than 10%. A polymerase chain reaction (PCR) product of approximately 330 bp was obtained from each of the four isolates using primers designed from the published PVT sequence. Restriction enzyme digestions of the PCR product did not demonstrate variability.

scholarly journals OPTIMALISASI PCR IKAN TONGKOL KRAI (Auxis thazard) DAN LISONG (Auxis rochei) PADA ANALISIS KERAGAMAN GENETIK

2019 ◽  
Vol 11 (2) ◽  
pp. 95
Author(s):  
Raymon Rahmanov Zedta ◽  
Bram Setyadji
Keyword(s):  
Genetic Diversity ◽  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Temperature Range ◽  
Pcr Polymerase Chain Reaction ◽  
Fisheries Resources ◽  
Pcr Product ◽  
Tuna Fishing ◽  
Polymerase Chain

Ikan tongkol lisong dan krai merupakan salah satu jenis tuna yang berperan nyata untuk usaha perikanan tangkap di Indonesia. Pengelolaan sumberdaya ikan tersebut harus selalu dapat dilakukan untuk menjaga tingkat pemanfaatannya supaya tidak lebih tangkap. Kajian keragaman genetik merupakan salah satu teknik dalam pengelolaan pemanfaatan sumberdaya perikanan dengan cara mengetahui tingkat keragaman genetik pada suatu struktur populasi. Kajian keragaman genetik ini diharapkan dapat menjadi basis kajian stok dan opsi dalam pengelolaan sumberdaya perikanan tongkol agar pemanfaatannya dapat dilakukan secara berkelanjutan. Awal mula analisis keragaman genetik dilakukan dengan memperbanyak DNA secara in vitro menggunakan teknik PCR (Polymerase Chain Reaction). Keberhasilan proses PCR dipengaruhi oleh beberapa faktor seperti suhu dan waktu penempelan oligonukleotida primer. Berdasarkan hal tersebut, penelitian ini bertujuan untuk mengetahui suhu dan waktu optimal pada primer Aro2-38. Sampel penelitian diperoleh dari hasil tangkapan pukat cincin yang didaratkan di PPN Palabuhanratu, Jawa Barat. Optimasi PCR menggunakan 12 suhu dan 2 waktu penempelan yang berbeda yaitu : 520C; 52,80C; 540C; 55,50C; 57,20C; 59,10C; 60,90C; 62,80C; 64,50C; 65,90C; 67,20C dan 680C, dan suhu penempelan 30 dan 15 detik. Hasil analisis menunjukkan bahwa produk PCR optimum (menghasilkan pita alel DNA) pada ikan tongkol krai berhasil waktu penempelan 30 detik dengan rentang suhu 52-540C. Sedangkan pada sampel ikan tongkol lisong, produk PCR yang optimum muncul pada waktu penempelan 15 dan 30 detik, dengan rentang suhu 52-60,90C.Frigate and bullet tuna constitute one of tuna species that plays a significant role in Indonesian fishing business. Management of fisheries resources must always be done to maintain the level of utilization so that it is not excessive. Genetic study is one of techniques in managing fisheries resource utilization by knowing the level of genetic diversity in a population structure. This genetic diversity study is expected to be the basis and option in the management of tuna fishing resources so that their utilization can be carried out sustainably. Genetic diversity analysis is start by multiplying fish DNA using PCR (Polymerase Chain Reaction) technique. The success of the PCR process is influenced by several factors such as temperature and time of primary oligonucleotide attachment. Based on this, this study aims to determine the optimal temperature and time in primers Aro2-38. The research sample was obtained from the catch of purse seine landed in PPN Palabuhanratu, West Java. PCR optimization uses 12 temperatures and 2 different annealing times: 520C; 52.80C; 54ÚC; 55,50C; 57.20C; 59.10C; 60.90C; 62.80C; 64,50C; 65,90C; 67.20C and 680C, and the annealing times are 30 and 15 seconds. The results of the analysis showed that the optimum PCR product (producing DNA allele bands) on the cretaceous tuna was successfully pasted for 30 seconds with a temperature range of 52-540C. Whereas in the sample of tuna lisong, the optimum PCR product appeared at the time of attachment of 15 and 30 seconds, with a temperature range of 52-60.90C.

scholarly journals Effect of a Debaryomyces hansenii and Lactobacillus buchneri Starter Culture on Aspergillus westerdijkiae Ochratoxin A Production and Growth during the Manufacture of Short Seasoned Dry-Cured Ham

2020 ◽  
Vol 8 (10) ◽  
pp. 1623
Author(s):  
Lucilla Iacumin ◽  
Martina Arnoldi ◽  
Giuseppe Comi
Keyword(s):  
Ochratoxin A ◽  
Starter Culture ◽  
Debaryomyces Hansenii ◽  
Aspergillus Ochraceus ◽  
In Situ Tests ◽  
Potential Health ◽  
Lactobacillus Buchneri ◽  
Dry Cured Ham ◽  
Aspergillus Westerdijkiae

Recently, specific dry-cured hams have started to be produced in San Daniele and Parma areas. The ingredients are similar to protected denomination of origin (PDO) produced in San Daniele or Parma areas, and include pork leg, coming from pigs bred in the Italian peninsula, salt and spices. However, these specific new products cannot be marked as a PDO, either San Daniele or Parma dry cured ham, because they are seasoned for 6 months, and the mark PDO is given only to products seasoned over 13 months. Consequently, these products are called short-seasoned dry-cured ham (SSDCH) and are not branded PDO. During their seasoning period, particularly from the first drying until the end of the seasoning period, many molds, including Eurotium spp. and Penicillium spp., can grow on the surface and work together with other molds and tissue enzymes to produce a unique aroma. Both of these strains typically predominate over other molds. However, molds producing ochratoxins, such as Aspergillus ochraceus and Penicillium nordicum, can simultaneously grow and produce ochratoxin A (OTA). Consequently, these dry-cured hams may represent a potential health risk for consumers. Recently, Aspergillus westerdijkiae has been isolated from SSDCHs, which could represent a potential problem for consumers. Therefore, the aim of this study was to inhibit A. westerdijkiae using Debaryomyces hansenii or Lactobacillus buchneri or a mix of both microorganisms. Six D. hansenii and six L. buchneri strains were tested in vitro for their ability to inhibit A. westerdijkiae. The strains D. hansenii (DIAL)1 and L. buchneri (Lb)4 demonstrated the highest inhibitory activity and were selected for in situ tests. The strains were inoculated or co-inoculated on fresh pork legs for SSDCH production with OTA-producing A. westerdijkiae prior to the first drying and seasoning. At the end of seasoning (six months), OTA was not detected in the SSDCH treated with both microorganisms and their combination. Because both strains did not adversely affect the SSDCH odor or flavor, the combination of these strains are proposed for use as starters to inhibit OTA-producing A. westerdijkiae.

Degradation of ochratoxin A by supernatant and ochratoxinase of Aspergillus niger W-35 isolated from cereals

2020 ◽  
Vol 13 (2) ◽  
pp. 287-298
Author(s):  
M. Zhao ◽  
X.Y. Wang ◽  
S.H. Xu ◽  
G.Q. Yuan ◽  
X.J. Shi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Food Safety ◽  
Aspergillus Niger ◽  
Ochratoxin A ◽  
Promising Strategy ◽  
Ochratoxin Α ◽  
Food And Feed ◽  
Commercial Feeds ◽  
Penicillium Spp

Ochratoxin A (OTA) is a mycotoxin produced by Aspergillus spp. and Penicillium spp. and poses a threat to food safety. Biodegradation may be a promising strategy for reducing the OTA contamination in the future. In this study, Aspergillus niger strain W-35 was isolated from cereals and studied for its ability to degrade OTA. Results showed that the supernatant of W-35 could degrade OTA both in vitro and in commercial feeds after incubation at 37 °C for 12 h by 78.0 and 37.0%, respectively. Ochratoxin α (OTα) was assayed as a degradation product by HPLC-FLD. Furthermore, an enzyme specific for OTA degradation (ochratoxinase, OTase) obtained from W-35 was successfully expressed in Escherichia coli BL21, and degraded OTA at a rate of 85.1% for 12 h. These results indicated that this OTA degradation is enzymatic and that the responsible enzyme is extracellular OTase. Reliable degradation of OTA has the potential for wide-ranging applications in the food and feed industries.

In vitro effect of some fungicides used in cultivation of Capsicum spp. on growth and ochratoxin A production by Aspergillus species

2013 ◽  
Vol 6 (2) ◽  
pp. 159-165
Author(s):  
L. Santos ◽  
S. Marín ◽  
V. Sanchis ◽  
A.J. Ramos
Keyword(s):  
Ochratoxin A ◽  
High Performance ◽  
Fruit Production ◽  
Aspergillus Species ◽  
Aspergillus Ochraceus ◽  
Fungal Load ◽  
Capsicum Spp ◽  
Vitro Effect ◽  
Yeast Extract Sucrose

The present study aimed to assess the effect of some pre-harvest fungicides commonly used in Capsicum fruit production on growth and ochratoxin A production of three Aspergillus species found in Capsicum powder. Aspergillus tubingensis, Aspergillus ochraceus and Aspergillus westerdijkiae, previously isolated from paprika and chilli, were inoculated on yeast extract sucrose agar and paprika extract agar supplemented with different fungicides at their recommended dosage rates, and incubated at 20 and 30 °C during 7 days. Radial growth was measured after 3, 5 and 7 days and ochratoxin A production was determined by high-performance liquid chromatography with fluorescence detection on day 7. Dodine 40% and mancozeb 80% were the most effective fungicides in inhibiting growth and ochratoxin A production, regardless of the fungal strain tested or temperature conditions. Whereas the application of fungicides could be very attractive in reducing the mycotoxigenic fungal load, it can also stimulate ochratoxin A production in some cases.

Cloning and expression ofSpodoptera lituranucleopolyhedrovirusgp41gene inEscherichia coliand preparation of antibody

2005 ◽  
Vol 2 (2) ◽  
pp. 113-117
Author(s):  
Pan Li-Jing ◽  
Li Zhao-Fei ◽  
Yin Chong ◽  
Lv Lei ◽  
Pang Yi
Keyword(s):  
Fusion Protein ◽  
Prokaryotic Expression ◽  
Recombinant Plasmid ◽  
Nitrilotriacetic Acid ◽  
Cloning And Expression ◽  
Reading Frame ◽  
Pcr Product ◽  
Prokaryotic Expression Vector ◽  
Polymerase Chain

AbstractGP41, a major glycoprotein, identified in the occlusion-derived virions (ODV) of baculoviruses, is required for the egress of nucleocapsids from the nucleus in the pathway of budded virion (BV) synthesis. Using the polymerase chain reaction (PCR), the open reading frame (ORF) ofSpodoptera lituranucleopolyhedrovirus (SpltMNPV)gp41gene was obtained from SpltMNPV genomic DNA. The PCR product was cloned into pMD18-T vector to get the recombinant plasmid (pT-gp41). Thegp41gene was recombinedin vitrowith prokaryotic expression vector pQE30 and transformed intoEscherichia coliM15 [pREP4]. The M15 [pREP4] strain, containinggp41recombinant plasmid, expressed a 37.9 kDa 6×His-tag fusion protein after induction with 1 mmol/l isopropylthio-β-d-galactoside (IPTG). The fusion protein was purified with a nickel-nitrilotriacetic acid (Ni–NTA) resin column and used as the immunogen to raise GP41-specific antibody. Western blotting analysis indicated that the antibody was suitable to be used for further analysis of GP41 protein.

Growth and ochratoxin A production by Aspergillus niger group strains in coffee beans in relation to environmental factors

2010 ◽  
Vol 3 (1) ◽  
pp. 59-65 ◽  
Author(s):  
A. Astoreca ◽  
C. Barberis ◽  
C. Magnoli ◽  
A. Dalcero
Keyword(s):  
Growth Rate ◽  
Aspergillus Niger ◽  
Ochratoxin A ◽  
Mycelial Growth ◽  
High Performance ◽  
Lag Phase ◽  
Distilled Water ◽  
Coffee Beans ◽  
Growth Assessment ◽  
Producer Strains

The effect of water activity (aW), temperature and their interactions on lag phase, mycelial growth rate and ochratoxin A (OTA) production at 7, 14 and 21 days of incubation of two OTA-producer strains belonging to Aspergillus niger group on irradiated coffee beans was determined. Irradiated coffee beans were re-hydrated to 0.910-0.995 of aW with sterile distilled water. The temperatures assayed were 15, 25 and 30 °C. Growth assessment was measured every day during the incubation period to calculate the growth rate. OTA production was examined at 7, 14 and 21 days by high-performance liquid chromatography. Optimal aW for growth was 0.995 at 25 °C for RCC4 and RCC20 strains, being 1.10 and 2.36 mm/day, respectively. OTA concentration varied considerably depending on aW, temperature and incubation time assayed. Maximum OTA production was obtained at 0.973 and 0.995 aW at 30 °C for both strains. The results of the present work indicate that knowledge of the optimal and marginal conditions of black Aspergillus growth and OTA production allow methods to be established for preventing the development of these fungal and mycotoxin production on coffee beans. The data obtained provide useful information for predicting the risk factors for OTA contamination on coffee beans.

Fate of Ochratoxin A during Cooking of Naturally Contaminated Polished Rice

2005 ◽  
Vol 68 (10) ◽  
pp. 2107-2111 ◽  
Author(s):  
JE WON PARK ◽  
SOO-HYUN CHUNG ◽  
CHAN LEE ◽  
YOUNG-BAE KIM
Keyword(s):  
Ochratoxin A ◽  
High Performance ◽  
Glioma Cells ◽  
In Vitro Cytotoxicity ◽  
C6 Glioma Cells ◽  
Cytotoxic Potential ◽  
Cooked Rice ◽  
Polished Rice ◽  
Low Levels

Ochratoxin A (OTA), a mycotoxin widespread in cereals, occurs in polished rice that is consumed as cooked rice after washing and steaming. Cooking decreases OTA levels in food to varying extents, but little is known about how cooking changes the biological activity of this mycotoxin. We therefore evaluated the fate of OTA during rice cooking to determine the OTA residues and cytotoxic potential in vitro. Water-washed rice, ordinary cooked rice, and pressure-cooked rice were prepared from three polished rice lots naturally contaminated with OTA. Residual OTA in each sample was analyzed by high-performance liquid chromatography (HPLC), whereas in vitro cytotoxicity of OTA to C6 glioma cells, susceptible to low levels (nanograms per milliliter) of OTA, was used to confirm the chemical analysis. OTA concentration, as determined by HPLC analysis, in the cooked rice by both types of cookers was significantly lower than (59 to 75%) in the raw polished rice and water-washed rice. The cytotoxicity of the OTA that remained in the pressure-cooked rice from three lots was markedly decreased (approximately 20%, P < 0.05) when compared with other samples in respective lots. This confirms that cooking lowers OTA residues. Although washing polished rice with water had little effect on OTA levels, pressure steaming appeared to be the critical cooking step not only to reduce OTA residues in polished rice before reaching the consumer as the dietary staple of cooked rice, but also to diminish cytotoxicity of OTA.

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