الفهرس 
Folate and its importance in Human DietOnly 14 pages are availabe for public view
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Abstract
This thesis took place from 2012-2017, in collaboration between the
Genetics department, Faculty of Agriculture, the Genetic Engineering and
Biotechnology Research Center, Ainshams University, Cairo, Egypt, and
Food safety and Biotechnology Laboratories, the Regional Center for
Food and Feed, Agriculture Research Center, Ministry of Agriculture,
Cairo, Egypt
The research work was mainly done on two probiotics, Streptococcus
thermophilus and Enterococcus faecium. Specifically the capabilities of
these bacterium to synthesize folate was measured by bioassay. Results
showed that Streptococcus thermophilus has superior capability. The
average folate production by the three isolated strains of Streptococcus
thermophilus was 19 folds higher than the best folate producer of
Enterococcus faecium strain. Through these experiments, two bioassay
methods were tested, one used Enterococcus hirae as the tester strain, and
the other method Lactobacillus casei strain was used as the tester strain
and it was lypholized in a kit. The kit showed better coefficient of
determination indicating it would be better in the analysis of folate. Also,
there were no correlation between the ability of the strain to grow in the
media and its ability to produce folate, where Enterococcus faecium had a
higher growth rate than Streptococcus thermophilus but yet showed less
folate production.
The hypothesis of the idea that Streptococcus thermophilus produce more
folate than Enterococcus faecium because it contain the genetic makeup
required for folate production was confirmed since, the required seven
genes for folate production and metabolism was isolated from
Streptococcus thermophilus and sequenced, and one gene responsible for
folate metabolism was also isolated..
The practical work of the thesis was as follows: 1- Lactic acid bacterial strains were isolated from yoghurt and
milk using M17 broth medium and M17 agar medium.
2- Folate producers strains were selected using two folate
deprived media, the folic acid medium and the chemically
defined medium.
3- The bacteria that were able to grow on folate deprived
media were identified and characterized by PCR using 16s
rRNA gene specific primers. from the selected strains six
of them were Enterococcus faecium and three of them,
where Streptococcus thermophilus. The results were
confirmed by sequencing the obtained bands.
4- Quantitative analysis of folate was done on Enterococcus
faecium strains by the bioassay using the Enterococcus
hirae as the tester strain.
5- Quantitative analysis of folate was done on Streptococcus
thermophilus by the bioassay, but using a Vitafast kit, that
contain lypholized Lactobacillus casei as the tester strain.
6- The bioassay measurement of folate done by bioassay
using the Vitafast r-biopharm kit was better than the
bioassay using the Enterococcus hirae. The R2 of the first
method was 0.9, while the latter was 0.7.
7- The results of Enterococcus faecium folate quantitative
measurement showed that there was significant difference
among the strains. where strains produced from 2.4 μg/l to
6.5 μg/l folate. 8- The folate production was not correlated with the rate of
growth. where some strains that had less growth rate
produced more folate than strains that had higher growth
rate.
9- The results of Streptococcus thermophilus quantitative
measurement showed no significant difference between its
strains, where they produced folate in the range of 156 -
162 μg/l.
10- The Streptococcus thermophilus produced folate 27 folds
more than that of Enterococcus faecium.
11- In this study, eight primers were designed to isolate eight
genes responsible for folate metabolism in Streptococcus
thermophilus, each primer was produced specific band.
12- The isolated eight genes were sequenced and their role was
identified by insilico analysis. The eight genes were
identified as follows: FolA. FolB/Q, FolC1, FolC2, FolD,
FolE, FolP, and FolK.
The results revealed that Streptococcus thermophilus are very
challenging probiotic bacteria to be used in folate production. The
biosynthetic pathway of this organism still need to be studied extensively,
and the relationship between the gene expression and folate production,
needs to be addressed.