All visible bands were assigned a number based upon relative position to the DNA ladder. Each position was then assigned a “0” or a”1” to indicate absence or presence of the band, respectively. The 0/1 matrix was then used to generate the dendrograms. Similarities based on the Dice coefficient were calculated and UPGMA clustering was obtained using NTSYS software . The comparative cluster analysis of different strains integrates banding pattern data from two separate experiments. The data were combined in a single matrix. Although banding patterns differed there were some band positions in common in some strains across the two studies. The reagents used in the two studies were identical; however the PCR thermal cycler was different, a GeneAmp PCR System 2700 was used in this study. In a previous study 349 strains isolated from different native fermentations in China were screened by interdelta sequence typing. The number of strains tested in this analysis did not allow definitive comparisons of highly similar strains as each strain was only run once. However based on this preliminary analysis 54 different banding patterns were identified. These 54 strains were studied in more detail under conditions enabling conclusive comparisons of DNA band profile. Samples of the strains were run on gels multiple times and band positions calculated in comparison to the DNA ladder on each gel. Strains were run on different gels and yielded identical patterns on the replicate gels. This method generated stable replicate banding patterns for each isolate. The replicated interdelta sequence typing methodology enabled clear differentiation of forty-two distinct S. cerevisiae genotypes among the 54 S. Twenty-one strains of S. cerevisiae isolated from Xinjiang revealed eighteen different patterns . The analysis of thirty-three S. cerevisiae isolates from Ningxia revealed the existence of twenty-four distinct profiles in other words, 24 differentiated strains. The results also showed that the interdelta profile of genotype NX10 was identical to that of commercial yeast Lalvin RC212,maceta 5 litros which has been used in this region as a commercial starter strain.
In general, the Xinjiang and Ningxia regions evaluated in this study exhibited different S. cerevisiae populations. There were no identical strains between the regions. In addition, comparison with the forty-four interdelta genetic patterns found among the fifty-two Saccharomyces strains from Liu et al. revealed that no identical strains were present between the studies. The PCR amplification profiles obtained by interdelta sequence typing were used to obtain independent presence-absence matrices for S. cerevisiae isolates from Xinjiang and Ningxia, China. The dendrograms deduced by interdelta sequence typing are presented in Figures 3 and 4, respectively. The dendrograms demonstrated that native wine fermentations were conducted by a mixture of S. cerevisiae strains. In general a minimum of two interdelta sequence profiles could be detected during each of the spontaneous fermentations with the exception of the fermentation of Big-berry Thompson Seedless. Some strains showed highly similar but not identical banding patterns and are thought to represent genetically related strains. Six clusters of two isolates each, cluster XJ4 and XJ5, XJ12 and XJ15, XJ2 and XJ7 ; NX2 andNX3, NX29 and NX33, and NX9 and NX32 showed highly conserved interdelta sequence patterns indicating that they are likely genetically distinct derivatives of the same strain. Two clusters of four isolates, cluster NX1, NX4, NX5, and NX8; cluster NX17, NX30, NX31, and NX26 also displayed conserved interdelta sequencing patterns again suggesting a genetic relationship among the sets of strains. The S. cerevisiae isolates from Xinjiang that clustered together were from the same grape variety, but, in contrast, the isolates from Ningxia that clustered together were from more than two different grape varieties. NX10 from a native Riesling fermentation was indistinguishable from the commercial strain RC212. A difference was observed in the genetic relationships of S. cerevisiae among strains isolated from table and wine grape varieties planted in Xinjiang . Both table and wine grapes are used for wine production in Xinjiang. Four groups containing nine, eight, three, and one isolate, respectively, were distinguished with a Dice coefficient of 0.448. All nine isolates in group I and all the 3 isolates in group III were obtained from the table grape varieties Red Globe, Small-berry Thompson Seedless, Big-berry Thompson Seedless, and Mixed white . Group II had seven isolates from wine grape varieties of Merlot and Mixed red , and one isolate from the table grape variety Red Globe.
Group IV only included one genotype, XJ3, also from Red Globe . Thus the majority of isolates from the four different table grape fermentations were genetically similar in banding pattern and clustered together. In addition, a difference was observed in the genetic relationships of S. cerevisiae among strains isolated from red and white wine grape varieties planted in Ningxia . Five groups are indicated in Figure 4, when the Dice coefficient is 0.674. Groups I, II, and III consisted of isolates from more than three grapes varieties, while group V only included isolates from Pinot Noir. Group IV included only 1 isolate, NX20, from Cabernet Sauvignon. Most of the S. cerevisiae isolates included in group I were isolated from red grape varieties , while isolates in group III were mainly from white grape varieties . All the isolates in group II were from the red grape varieties of Cabernet Sauvignon, Merlot, and Cinsault . Genetic relatedness was also evaluated by constructing a dendrogram compiled from all interdelta sequence patterns in this study and Liu et al. . Differences were observed in the genetic relationships between the Chinese strains and the strains from the UC Davis collection. The similarity of strains was correlated with their geographical regions of origin: Ningxia strains were closer to the Xinjiang strains, while a clear separation between the indigenous Chinese and the UC Davis strains was observed. The dendrogram produced by interdelta sequence typing revealed five groups containing fifty-one, four, forty-eight, three, and one isolate each when the Dice coefficient is 0.418 . The largest group, group I, contained fifty isolates from China and one from California . There is a difference observed in the genetic relatedness between Xinjiang and Ningxia indigenous S. cerevisiae strains. Group I could be further divided into five subgroups, I-1 to I-5, when the Dice coefficient is 0.506. Most of the S. cerevisiae isolates included in subgroups I-1 and I-5 as well as all three in I-4 were isolated from Xinjiang, while isolates in I-3 and I-2 were mainly from Ningxia . Group II displayed a high similarity with XJ19 , UCD587 , UCD2515 and UCD2516 . Forty-eight out of fifty-two Saccharomyces isolates from the UC Davis collection were clustered in group III. XJ2, XJ7, and XJ20 were clustered in group IV. Group V consisted only of XJ3. Clusters IV and V containing XJ2, XJ7, XJ20, and XJ3 had a low degree of similarity with other Chinese indigenous genotypes and UC Davis collection strains . In addition,cultivo de la frambuesa this dendrogram showed that identical isolates were found only in the same geographic regions with the exception of UCD522 and UCD514 . Understanding the genetic diversity of S. cerevisiae strains from different geographical origins can make an important contribution towards delineating the genetic distance of these strains as well as providing genetic material for further strain development. The genetic diversity of indigenous S. cerevisiae was investigated during the spontaneous fermentations of grape must in Xinjiang and Ningxia, China. Eighteen distinct interdelta profiles were found in Xinjiang, and twenty-four in Ningxia. Forty-two different S. cerevisiae strains were distinguished out of a total of three hundred and forty nine isolates analyzed. All forty-two of these isolates were unique when compared to a set of strains from other major international wine producing regions . Lavin RC212, showing the same interdelta sequence patterns as genotype NX10 isolated from Ningxia was detected during the spontaneous fermentations in this study. Similar to other studies, commercial yeasts were detected in fermentations without inoculation . In this study, the detection of Lavin RC212 colonizing a spontaneous fermentation could be explained by the winery practice of dumping grape skins on the road for drying. Valero et al. analyzed the dissemination and survival of commercial wine yeast in the vineyards near wineries and they suggested that the dispersion of commercial strains is mainly mediated by water run-off and derived from macerated grape skin at dumping sites. Drying the grape skins on the roads for further processing is a normal practice at the Imperial Horse Winery, Qing Tongxia, Ningxia, China, where the spontaneous fermentations were conducted.
It is understandable that this practice could have contributed to the dissemination and survival of Lavin RC212 in the vineyards and wineries, and its occurrence in spontaneous fermentations. In this study, comparison between eighteen and twenty-four different S. cerevisiae patterns by interdelta sequence typing revealed that yeast strains from Xinjiang and Ningxia did not share the same interdelta profiles. The same observations made in the Western Cape, South Africa showed that different S. cerevisiae strains were present at different regions in the different climate zones. In addition, the differences could be attributed to the fact that the grape varieties studied were different in these two regions: table and wine grape varieties in Xinjiang vs. wine grape varieties in Ningxia. This result agrees with a previous study that demonstrated the impact of grape variety on yeast diversity . Further, this study suggests that these two wine making regions are biologically isolated from each other. Geographic location and ecological niches are both thought to play a significant role in Saccharomyces strain diversity . In comparison with strains isolated from other wine making regions, the Ningxia and Xinjiang strains showed a high degree of similarity. This suggests that the indigenous Chinese strains are distinct from European and new world lineages. Nearly identical strains were only found in the wine samples collected in the same viticulture region with the exception of UCD522 and UCD514 . These results are in agreement with previous studies on geographically close regions and widely distant geographic regions . According to Ezeronye and Legras , who studied the genetics of S. cerevisiae strains isolated from palm wine in eastern Nigeria, geographic and/or ecological isolation results in a specific population of S. cerevisiae. These analyses have led to the conclusion that geographic location plays a significant role in genetic divergence. Strain XJ19 displayed a high similarity with UCD587 , UCD2515 and UCD2516 . The relatedness of these four strains may imply a common origin or a commonality of evolutionary forces in the wild. Interestingly as a group the Chinese wine strains appear to show greater overall diversity as compared to the wine isolates from the rest of the world. This is consistent with the narrowness of the Wine European lineage previously described as well as the observations of overall greater diversity of natural S. cerevisiae isolates from China as compared to other regions . The greater natural diversity of isolates of S. cerevisiae from China many represent an untapped genetic reservoir for strain improvement and breeding programs. Knowledge about indigenous yeast strains can also help preserve and employ the most representative strains from a wine region .When making purchasing decisions about products, consumers traditionally include factors such as price, quality, and availability of substitutes.On the one hand, if a product with safety concerns is removed from the market and the remaining products experience additional safety checks, consumers may perceive the market as being at least as safe as before. On the other hand, if the removal of the unsafe product provides negative information about closely related products or the industry as a whole, consumers may respond by decreasing demand, even in the absence of safety concerns about the remaining products. The empirical question is whether a recall of an unsafe product can have a direct impact on consumer purchases and preferences, even if the remaining products are safe. From a safety perspective, the question is relevant if firm incentives to invest in risk reduction and regulatory compliance in existing regulation depend, to some degree, on consumer responses to recalls.In early July 2010, the Center for Disease Control and Prevention identified a nationwide, four-fold abnormal increase in the number of reported Salmonella infections. A month later, on August 13, 2010, a first egg farm, located in Iowa, conducted a nationwide voluntary recall of around 228 million eggs. By August 18, 2010, the same farm expanded its recall to around 152 million additional eggs. Within 48 hours, on August 20, 2010, a second egg farm, also located in Iowa, conducted another nationwide voluntary recall of around 170 million eggs. In total, from August 13 to August 20, 2010, more than 500 million eggs were recalled, in what would be the largest egg recall in U.S. history .