Diversity in metabolite production by Fusarium langsethiae, Fusarium poae, and Fusarium sporotrichioides

The production of mycotoxins and other metabolites by 109 strains of Fusarium langsethiae, Fusarium poae, Fusarium sporotrichioides, and F. kyushuense was investigated independently in four laboratories by liquid or gas chromatography analyses of cultural extracts with UV diode array, electron capture, or mass spectrometric detection systems. From the compiled results, it was found that F. langsethiae consistently produced the trichothecenes diacetoxyscirpenol (DAS), T-2 toxin (T-2), HT-2 toxin (HT-2), and neosolaniol (NEO) and, to a lesser extent, some additional trichothecene derivatives. F. langsethiae also produced culmorins, chrysogine (CHRYS), aurofusarin (AUF), and enniatin (EN). F. sporotrichioides showed a metabolite profile similar to that of F. langsethiae, while F. poae had a different profile as 41 of 49 strains produced nivalenol (NIV) and other 8-keto trichothecenes, in addition to DAS and derivatives of this metabolite. Only a trace amount of NIV was detected from one strain of F. kyushuense. In summary, all the three core taxa of this joint study were found to produce trichothecenes. Fusarin C (F-C) was not detected from F. langsethiae, but it was produced by F. poae and F. sporotrichioides. Aurofusarin was only detected from a few strains of F. langsethiae, while nearly all strains of F. poae and F. sporotrichioides produced this compound. In contrast, chrysogine was not detected from F. poae, but was produced by the other two taxa. Production of enniatins was scattered among the three main taxa of this study, whereas beauvericin (BEA) was produced by many strains of F. poae and F. sporotrichioides. Only one odd strain of F. langsethiae (IBT 9959) produced beauvericin. However, the status of this strain is uncertain. By a polyphasic approach using species-specific metabolite profiles, the fruity odour of F. poae, and morphological observations, it was concluded that F. langsethiae, F. poae, and F. sporotrichioides should be regarded as three significant taxa at a species level.     

Phylogenetic analyses of the Fusarium poae, Fusarium sporotrichioides and Fusarium langsethiae species complex based on partial sequences of the translation elongation factor-1 alpha gene

Phylogenetic relationships between four Fusarium species were studied using parts of the nuclear translation elongation factor-1 alpha (EF-1alpha) gene as a phylogenetic marker. Sequences from 12 isolates of Fusarium poae, 10 isolates of Fusarium sporotrichioides and 12 isolates of Fusarium langsethiae yielded 4, 5 and 5 haplotypes, respectively. In addition, we included one isolate of Fusarium kyushuense. The aligned sequences were subjected to neighbor-joining (NJ), maximum parsimony and maximum likelihood (ML) analyses. The results from the different analyses were highly concordant. The EF-1alpha-based phylogenies support the classification of F. langsethiae as a separate taxon in the section Sporotrichiella of Fusarium, as the closest sister taxon to F. sporotrichioides, while F. kyushuense is the sister taxon to F. poae. This corresponds well with the ability of F. langsethiae and F. sporotrichioides to produce T-2 and HT-2 toxins. In contrast, morphological characters indicate a closer relationship between F. langsethiae and F. poae on the one hand, and between F. sporotrichioides and F. kyushuense on the other hand.     

IGS-RFLP analysis and development of molecular markers for identification of Fusarium poae, Fusarium langsethiae, Fusarium sporotrichioides and Fusarium kyushuense

The intergenic spacer (IGS) regions of the rDNA of several Fusarium spp. strains obtained from the collaborative researchers (Int. J. Food Microbiol. (2003)) were amplified by polymerase chain reaction (PCR), and an IGS-RFLP analysis was performed. Restriction digestion with AluI, MspI and PstI allowed differentiation between the related Fusarium poae and Fusarium kyushuense species. Fusarium langsethiae was also separated from Fusarium sporotrichioides (including var. minus) on the basis of the banding patterns after MspI digestion, while specific XhoI, AluI and MspI restriction patterns were found in the IGS amplicons of F. sporotrichioides var. minus. According to the phylogenetic analysis of IGS-RFLP patterns, F. langsethiae (except for one strain), F. sporotrichioides, F. poae and F. kyushuense strains formed four well-supported clades with high-bootstrap values. Based on the sequence differences in the IGS region, species-specific primers were designed for the F. langsethiae/F. sporotrichioides group and for F. poae. The specificity and sensitivity of the primers were tested on various Fusarium species and isolates, and on several other important fungal genera associated with cereals. The F. poae-specific primers, designed in this study, showed the same specificity as primers Fp82f/Fp82r developed previously. The two phylogenetic subgroups of F. langsethiae, found by IGS sequencing analysis, were separated on the basis of size differences of the amplification products with primers CNL12/PulvIGSr specific for the F. langsethiae/F. sporotrichioides group. RFLP analysis of the amplified IGS region is a useful molecular assay for characterisation and a phylogenetic study of several related Fusarium species-F. langsethiae, F. sporotrichioides, F. sporotrichioides var. minus, F. poae and F. kyushuense. The primers designed in this study were highly specific and allowed identification of F. poae and the F. langsethiae/F. sporotrichioides group.     

An integrated taxonomic study of Fusarium langsethiae, Fusarium poae and Fusarium sporotrichioides based on the use of composite datasets

An integrated systematic study was carried out to clarify the taxonomical position and relationship of Fusarium langsethiae to other taxa within the Fusarium section Sporotrichiella. Strains of this species were compared with strains of the closely related species Fusarium poae and Fusarium sporotrichioides using a composite dataset. This set consisted of DNA sequences derived from the ribosomal internal transcribed spacer (ITS) regions, partial sequences of the ribosomal intergenic spacer (IGS) region, the beta-tubulin and translation elongation factor-1 alpha (EF-1alpha) genes, AFLP fingerprints, chromatographic data on secondary metabolites and morphological data and growth characteristics. From these combined data, a consensus matrix was calculated by taking the mean of all pairwise distances between single isolates over all separate datasets. The consensus matrix was used as the basis for the construction of a UPGMA dendrogram and a multidimensional scaling, both of which revealed a clear separation of the three taxa. Partial IGS, EF-1alpha and beta-tubulin sequence-as well as chromatography-and AFLP-derived similarities turned out to be comparably consistent, while ITS sequence- and morphology-derived similarity matrices were rather divergent.     

Fusarium sibiricum sp. nov, a novel type A trichothecene-producing Fusarium from northern Asia closely related to F. sporotrichioides and F. langsethiae

Production of type A trichothecenes has been reported in the closely related species Fusarium langsethiae and F. sporotrichioides. Here, we characterized a collection of Fusarium isolates from Siberia and the Russian Far East (hereafter Asian isolates) that produce high levels of the type A trichothecene T-2 toxin and are similar in morphology to the type A trichothecene-producing F. langsethiae, and to F. poae which often produces the type B trichothecene nivalenol. The Asian isolates possess unique macroscopic and microscopic characters and have a unique TG repeat in the nuclear ribosomal intergenic spacer (IGS rDNA) region. In Asian isolates, the TRI1-TRI16 locus, which determines type A versus type B trichothecene production in other species, is more similar in organization and sequence to the TRI1-TRI16 locus in F. sporotrichioides and F. langsethiae than to that in F. poae. Phylogenetic analysis of the TRI1 and TRI16 gene coding regions indicates that the genes in the Asian isolates are more closely related to those of F. sporotrichioides than F. langsethiae. Phylogenetic analysis of the beta-tubulin, translation elongation factor, RNA polymerase II and phosphate permease gene sequences resolved the Asian isolates into a well-supported sister lineage to F. sporotrichioides, with F. langsethiae forming a sister lineage to F. sporotrichioides and the Asian isolates. The Asian isolates are conspecific with Norwegian isolate IBT 9959 based on morphological and molecular analyses. In addition, the European F. langsethiae isolates from Finland and Russia were resolved into two distinct subgroups based on analyses of translation elongation factor and IGS rDNA sequences. Nucleotide polymorphisms within the IGS rDNA were used to design PCR primers that successfully differentiated the Asian isolates from F. sporotrichioides and F. langsethiae. Based on these data, we formally propose that the Asian isolates together with Norwegian isolate IBT 9959 comprise a novel phylogenetic species, F. sibiricum, while the two subgroups of F. langsethiae only represent intraspecific groups.     

Specific detection of Fusarium langsethiae and related species by DGGE and ARMS-PCR of a beta-tubulin (tub1) gene fragment

Fusarium langsethiae was recently described as a new toxigenic Fusarium species, which morphologically resembles Fusarium poae, but exhibits a mycotoxin pattern related to Fusarium sporotrichioides. To develop tools for early and specific detection of F. langsethiae and distinguishing it from related species of section Sporotrichiella and Discolor (F. poae, F. sporotrichioides, Fusarium kyushuense, Fusarium robustum, Fusarium sambucinum and Fusarium tumidum) sequence variations in their beta-tubulin-encoding (tub1) gene were employed to design two PCR-based methods, denaturing gradient gel electrophoresis (DGGE) and amplification refractory mutation system (ARMS)-PCR. DGGE reliably separated all these strains, even from mixtures and in the presence of DNA from their natural hosts Zea mais, Triticum aestivum and Avena sativa. In addition, a tetraprimer ARMS-PCR, which employs two primer pairs to amplify, respectively, two different fragments of tub1 in a single PCR reaction resulted in rapid differentiation between F. langsethiae, F. sporotrichioides and F. poae according to the number of amplicons (four, two and one, respectively). These two methods will thus be worthwhile tools in the specific detection of F. langsethiae in infected crops.     

Accumulation of type A trichothecenes in maize, wheat and rice by Fusarium sporotrichioides isolates under diverse culture conditions

Toxigenic isolates of Fusarium sporotrichioides were tested for the production of type A trichothecenes (T-2 toxin, HT-2 toxin, diacetoxyscirpenol and neosolaniol) when grown on three substrates (maize, rice and wheat) under various conditions of temperature and water activity in the laboratory for 3 weeks. Trichothecenes were determined by high-performance liquid chromatography with fluorescence detection, after derivatisation with coumarin-3-carbonyl chloride. This is the first time this analytical method has been applied to an extensive study of trichothecene accumulation. With minor exceptions, greater trichothecene production occurred when samples were incubated at 20 degrees C and moistened with 35% water (water activity 0.990) although incubation conditions affected the substrates studied in different ways. No correlation between the different pairs of trichothecenes was found except for neosolaniol and diacetoxyscirpenol (r=0.56). Principal component analysis results show that the data points can be grouped in three rough clusters related to cereal type, which points out that the composition of these cereals can influence the production of type A trichothecenes.     

Effect of fenpropimorph, prochloraz and tebuconazole on growth and production of T-2 and HT-2 toxins by Fusarium langsethiae in oat-based medium

Fusarium langsethiae has been isolated from infected cereals in central and northern Europe where it has been identified in the last decade as the main species involved in the occurrence of high levels of T-2 and HT-2 toxins, mainly in oats. The efficacy of three fungicides (prochloraz, tebuconazole, fenpropimorph) for controlling growth of two strains of F. langsethiae isolated from oats was examined at 0.96 and 0.98 a_w at 15, 20 and 25 °C on oat-based media. The concentrations necessary for 50 and 90% growth inhibition (ED₅₀ and ED₉₀ values) were determined. The effect on the trichothecene type A mycotoxins T-2 and HT-2 was also determined. Without fungicides both strains grew faster at 0.98 than at 0.96 a_w and the influence of temperature on growth rates was 25 > 20 > 15 °C. Prochloraz and tebuconazole were more effective than fenpropimorph against F. langsethiae. Strain, temperature and type of fungicide significantly influenced the ED₅₀ and ED₉₀ values for growth. The concentration ranges under different environmental conditions were: prochloraz (0.03-0.1 and 0.3-1.5), tebuconazole (0.06-0.9 and 1.3-8.2), and fenpropimorph (22-59 and 125-215 mg l⁻¹). Production of T-2 and HT-2 toxins was influenced by temperature, a_w, type of fungicide and dose. Levels of T-2 were usually higher than those of HT-2 under the same conditions. The biosynthesis of T-2 toxin increased after 10 day incubation, but was reduced with decreasing temperature and increasing fungicide dose. At 0.98 a_w T-2 levels increased in cultures containing fenpropimorph while at 0.96 a_w the toxin concentrations increased in response to the other two fungicides. Low doses of prochloraz or tebuconazole enhanced toxin production when compared with untreated cultures for strain 2004-59 at 0.96 a_w and 20-25 °C. HT-2 was hardly detectable in the treatments with prochloraz or tebuconazole at 0.98 a_w. This is the first study on the effect of these anti-fungal compounds on control of growth of F. langsethiae and on production of T-2 and HT-2 toxins in oat-based media.     

Phylogenetic relationships among ammonia-oxidizing bacteria as revealed by gene sequences of glyceraldehyde 3-phosphate dehydrogenase and phosphoglycerate kinase

The three previously recognized genera of 'Nitrosolobus', Nitrosospira and 'Nitrosovibrio' were combined into one genus, Nitrosospira, on the basis of 16S rDNA sequence similarities. However, this classification has been controversial for some time, since the marked differences in their shapes suggest that they are not closely related. In this study, the phylogenetic analyses of the three groups using two genotypical markers, glyceraldehyde-3-phosphate dehydrogenase (GAP, gap), and 3-phosphoglycerate kinase (PGK, pgk), were performed. In the phylogenetic tree inferred from gap and pgk, the three genera appeared as clearly separated clusters. This is the first study of markers that are able to reveal the precise phylogenetic relationship among 'Nitrosolobus', Nitrosospira and 'Nitrosovibrio'.     

Formation of higher alcohols by wine yeasts,and relationship to taste thresholds

The amounts of n-propanol, iso-butanol and iso-amyl plus active amyl alcohol produced during fermentation of grape juice were found to vary considerably according to the yeast used. The yeasts studied included 11 wine yeasts and one brewery yeast, all belonging to the genus Saccharomyces, and 4 yeasts belonging to genera which sometimes cause wine spoilage. Experiments were carried out in the laboratory and confirmed on pilot-plant scale. The average production of n-propanol in juices from 4 grape varieties varied from 13 to 106 ppm depending on the wine yeast used. The corresponding variations in the production of iso-butanol and isoamyl plus active amyl alcohol were 9 to 37 ppm and 115 to 262 ppm respectively,. Juices from different varieties' of grapes (Vitis vinifera) differed somewhat in the amounts of higher alcohols formed, irrespective of the yeast strain used. An increase in temperature of fermentation from 15° to 25° with four yeasts produced an average of 24% more iso-amyl plus active alcohol and 39% more iso-butanol, and 17% less n-propanol. A yeast/temperature interaction occurred. An increase in pH from 3.0 to 4.2 with four yeasts produced 28% more iso-amyl plus active amyl alcohol, 85% more iso-butanol, and 11% more n-propanol. A yeast /pH interaction occurred. Spoilage yeasts examined also formed higher alcohols and the amounts were not related to sugar consumed during the fermentation. Taste thresholds of the higher alcohols, with the exception of active amyl alcohol, were measured in dry white wine with 7 tasters. Iso-amyl alcohol thresholds ranged from 100 to 900 ppm (mean 300 ppm) and iso-butanol and n-propanol both exceeded 500 ppm. The average threshold of iso-amyl alcohol in distilled water was 4 ppm. For certain tasters strains of yeast can produce sufficient differences in iso-amyl alcohol to be detectable organoleptically, but differences in iso-butanol and n-propanol are not large enough to be detectable.     

Structural analysis of enniatin H,I, and MK1688 and beauvericin by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and their production by Fusarium oxysporum KFCC 11363P

The molecular structures of enniatins H, I, and MK1688 and beauvericin were investigated by liquid chromatography-tandem mass spectrometry (LC-MS/MS). MS fragmentation occurred by loss of -CO after opening of the cyclic molecule to carbonyl carbon, and cleavage of the peptide and ester bonds in the molecular structure. Fusarium oxysporum KFCC 11363P was tested for its ability to produce beauvericin and enniatins H, I, and MK1688 on five cereal substrates: rice, barley, maize, wheat, and Indian millet kernels. Furthermore, optimal conditions for the production of the four mycotoxins by the Fusarium isolate were examined on maize at four temperatures (15, 20, 25, and 30°C) and at three moisture contents (10, 20, and 40%). Large amounts of beauvericin and enniatin H were present in maize cultures at 25°C (232.4 and 196.4 µg g^?1, respectively). Enniatins I and MK1688 were maximally formed at 20°C (221.5 and 180.2 µg g^?1, respectively). The optimal moisture contents for the production of enniatins H (196.4 µg g^?1) and MK1688 (165.6 µg g^?1), were 40%.     

Physicochemical characteristics,proximate analysis and mineral composition of ostrich meat as influenced by muscle

The influence of muscle on the physicochemical characteristics, proximate analysis, and mineral composition of meat from 10 ostriches (10–12 months old), slaughtered according to commercial abattoir procedures, were evaluated. Muscle had no influence (p > 0.05) on L*-values (32.5), a*-values (11.9), water-holding capacity (11.9%), final pH (pH₂₄) values (6.07), and ash contents (1.12 g/100 g edible meat). However, intramuscular lipid contents varied (p < 0.05) from 0.88 (M. fibularis longus) to 1.44 (M. flexor cruris lateralis) g/100 g edible meat, at a mean value of 1.16 g/100 g edible meat for 10 different muscles. Sodium (34.7 mg/100 g edible meat) and iron (3.14 mg/100 g edible meat) contents, both influenced (p < 0.05) by muscle, possessed substantially lower and higher values, respectively, than values reported for beef and chicken.     

Further data on the production of beauvericin, enniatins and fusaproliferin and toxicity to Artemia salina by Fusarium species of Gibberella fujikuroi species complex

The knowledge of toxigenic profiles of fungal plant pathogens is of extreme importance for evaluating the potential toxicity of infected plant products. Ninety-six fungal isolates belonging to 28 species in the Gibberella fujikuroi complex were studied for the production of beauvericin, enniatins and fusaproliferin in rice cultures. Toxin production ranged from 5 to 3000 microg/g for beauvericin, 2 to 131 microg/g for enniatins, and 4 to 440 microg/g for fusaproliferin. Beauvericin was the most common metabolite produced by 16 species followed by fusaproliferin with 11 species and enniatins with 4 species. The production of beauvericin by F. bulbicola, F. denticulatum, F. lactis, F. phyllophilum, F. pseudocircinatum, and F. succisae and fusaproliferin by F. antophilum, F. begoniae, F. bulbicola, F. circinatum, F. concentricum, F. succisae, and F. udum is reported here for the first time. Brine shrimp larvae were most sensitive to culture extracts of F. acutatum (up to 94+/-3%), F. concentricum (up to 99+/-1%), F. denticuatum (up to 100%) and F. sacchari (up to 100%). Toxicity towards brine shrimp was significantly correlated with the beauvericin content of the fungal extracts with few exceptions. These data indicate that beauvericin and fusaproliferin are common metabolites of species of the G. fujikuroi complex and pose a risk for a possible toxin accumulation in their respective host plant products. However, data from the brine shrimp bioassay showed that further toxic metabolites within this complex need to be characterized.     

Exposure to cadmium from foods, estimated by analysis and calculation--comparison of the methods

Cadmium intake of 40 middle-aged Finnish men was determined both by calculation based on computer files and by analysis of duplicate portions. A significant difference was observed between intake estimates of the two methods. The average calculated cadmium intake was 15.8 micrograms/day compared to the 8.2 micrograms/day obtained by analysis. Factors causing the difference between the methods and affecting the comparability are: (i) expression of concentrations below the detection limit; (ii) the calculation is based on the analysis of foods being only washed and peeled whereas duplicate portions are normally processed; (iii) when duplicate portions are prepared using non-representative lots or brands of food, their cadmium content differs from average values used in calculation; (iv) the level of cadmium content in duplicate portion samples is very low affecting the accuracy of the results. The analytical level of cadmium during the analysis of duplicate portions was somewhat lower as compared to the analysis on which the food composition file and the calculation is based. It seems that the complicating factors identified in the present study potentially affect the reliability and comparability of studies of trace element intakes. Therefore one has to be careful when comparing intake estimates derived independently or by different methods.     

Response surface models to predict potato tuber infection by Fusarium sambucinum from duration of wetness and temperature,and dry rot lesion expansion from storage time and temperature

Sucrose-negative Yersinia enterocolitica isolates of bioserotype 4/O:3 have been recovered for the first time. They were found in 2% of the tonsils of clinically healthy fattening pigs. These sucrose-negative Y. enterocolitica isolates could not be differentiated from Y. kristensenii isolates using API 20E; thus, they were identified using PCR and sequencing. Using pulsed-field gel electrophoresis (PFGE). NotI profiles of sucrose-negative Y. enterocolitica 4/O:3 isolates showed a high similarity to sucrose-positive Y. enterocolitica 4/O:3 isolates. This study demonstrated that sucrose-negative Y. enterocolitica 4/O:3 isolates of porcine origin can harbour virulence genes; plasmid-encoded virulence markers were found in 8 out of 11 isolates and all isolates contained chromosomal-encoded virulence markers. Thus, the pathogenicity of sucrose-negative Yersinia isolates should always be assessed.     

In vitro induction of apoptosis, necrosis and genotoxicity by cosmetic preservatives: application of flow cytometry as a complementary analysis by NRU

Preservatives are used in cosmetics to prevent microbial contamination; however, some preservatives are not free of allergenic and cytotoxic potential. Allergenicity and cytotoxicity potential values are major aspects of preservative safety, which determine limitations and maximum concentration dose in a cosmetic product. The purpose of this study was to investigate and compare the in vitro apoptosis, necrosis and genotoxicity-inducing potential of five different types of preservatives: Phenoxyethanol (PE), Propylparaben (PP), Methylparaben (MP), Benzyl Alcohol (BA) and Ethylhexyl Glycerine (EG). In vitro experiments were carried out on human dermal fibroblasts by a quantitative flow cytometry method, using specific cell markers (Annexin V, Propidium Iodide and H2AX). We compared the resulting cell viability by means of neutral red uptake (NRU) and established the IC(50) . Our results showed that PE, PP, MP and BA have similar cytotoxic mechanisms (high apoptosis and necrosis levels only at the test concentration of 1%), whereas EG showed only an apoptosis pathway. For genotoxicity, both parabens yielded the highest values. Results obtained by flow cytometry for necrosis were comparable to those produced by NRU; however, NRU does not distinguish apoptosis from necrosis. We propose that flow cytometry is a more sophisticated methodology for understanding the cytotoxic mechanisms of cosmetic preservatives and can be used to complement the NRU.     

Hydrolysis of the sarcoplasmic, myofibrillar and connective tissue proteins of lean beef by alcalase and its relationship to whole meat hydrolysis

The enzymic hydrolysis and solubilisation of the three meat protein fractions have been investigated in order to elucidate some aspects of the hydrolysis of whole beef protein. Much of the water-soluble sarcoplasmic protein fraction was insolubilised on heating for 25 min at 60°C. While most of this insoluble protein was easily resolubilised enzymically, some of it remained insoluble after 5 hours' hydrolysis. Solubilisation of connective tissue by Alcalase after a 3 h reaction increased markedly from 23% at 55°C to 99% at 60°C. This explains the high optimum temperature for solubilisation of whole beef by Alcalase. A significant portion of myofibrillar tissue remained insoluble after 3 hours' reaction. indicating that the majority of the insoluble solids remaining after whole meat hydrolysis at 60°C derived from myofibrillar tissue. There was good agreement between an experimental reaction progress curve for whole beef hydrolysis and one estimated from progress curves for hydrolysis of the three meat protein fractions.     

Uptake by perennial ryegrass of iodide, elemental iodine and iodate added to soil as influenced by various amendments.

Perennial ryegrass was grown in pots in a sandy loam soil, into which iodide, elemental iodine or iodate had been incorporated at a rate of 20 mg I/kg. The uptake of iodine into the herbage was much greater from iodate than from the other two forms. Replacement of 5 % of the soil by well-decomposed farmyard manure reduced uptake from all three forms of iodine more than ten-fold. Similar replacement by chalk reduced uptake from iodide but increased uptake from iodate. The recovery of added iodine in three successive harvests of ryegrass ranged from 0.03% for elemental iodine in combination with soil + FYM to 2.16% for iodate in combination with soil + chalk.     

High-resolution genotyping of Listeria monocytogenes by fluorescent amplified fragment length polymorphism analysis compared to pulsed-field gel electrophoresis, random amplified polymorphic DNA analysis, ribotyping, and PCR-restriction fragment length polymorphism analysis

The purpose of this study was to evaluate fluorescent amplified fragment length polymorphism (AFLP) analysis for the inter- and intraspecies differentiation of a collection of 96 strains of Listeria monocytogenes and 10 non-L. monocytogenes strains representing six other Listeria species of different origin. The AFLP technique was compared with three other molecular typing methods--ribotyping, random amplified polymorphic DNA analysis (RAPD), and pulsed-field gel electrophoresis (PFGE)--in terms of discriminatory ability. PCR-restriction fragment length polymorphism was included for virulence gene allele characterization. The 96 L. monocytogenes strains were divided into two major clusters by AFLP fingerprinting at a similarity level of 82% in concordance with the results of PFGE, RAPD, and ribotyping. One main cluster consisted of all of the 24 L. monocytogenes hly allele 1 strains, while another main cluster consisted of all of the 72 L. monocytogenes hly allele 2 strains. This indicates the existence of two distinct phylogenetic divisions. Isolates of the remaining Listeria species were not included in the clusters. AFLP, PFGE, and RAPD typing were highly discriminatory methods, with discrimination (D) indices of 0.974, 0.969, and 0.954, respectively, whereas ribotyping had a lower D index of 0.874. AFLP, PFGE, and RAPD typing showed some level of agreement in terms of strain grouping and differentiation. However, all three methods subdivided types of strains grouped by the other methods. Isolates with identical DNA profiles were distributed across the spectrum of origin. It was not possible to associate certain types with specific food sectors or clinical cases, which is indicative of the spread of L. monocytogenes clones across species. Overall, AFLP fingerprinting was suitable for the high-resolution genotyping of L. monocytogenes and had an equally high or higher differentiation power compared to PFGE or RAPD typing.     

Consumer perception of boar meat as affected by labelling information,malodorous compounds and sensitivity to androstenone

This study aimed to assess the influence of two label conditions on the acceptance of boar meat. A central location test was conducted with 145 consumers each assessing 4 pieces of pork loin.