86) Use of e.Coli for the biotechnological production

E. coli is frequently used as a model organism in microbiology studies. Cultivated strains (e.g. E. coli K12) are well-adapted to the laboratory environment, and, unlike wild type strains, have lost their ability to thrive in the intestine. Many lab strains lose their ability to form biofilms. These features protect wild type strains from antibodies and other chemical attacks, but require a large expenditure of energy and material resources.

Because of its long history of laboratory culture and ease of manipulation, E. coli also plays an important role in modern biological engineering and industrial microbiology. The work of Stanley Norman Cohen and Herbert Boyer in E. coli, using plasmids and restriction enzymes to create recombinant DNA, became a foundation of biotechnology.

E. coli is a very versatile host for the production of heterologous proteins, and various protein expression systems have been developed which allow the production of recombinant proteins in E. coli. Researchers can introduce genes into the microbes using plasmids which permit high level expression of protein, and such protein may be mass-produced in industrial fermentation processes. One of the first useful applications of recombinant DNA technology was the manipulation of E. coli to produce human insulin.

Many proteins previously thought difficult or impossible to be expressed in E. coli in folded form have also been successfully expressed in E. coli. For example, proteins with multiple disulphide bonds may be produced in the periplasmic space or in the cytoplasm of mutants rendered sufficiently oxidizing to allow disulphide-bonds to form,while proteins requiring post-translational modification such as glycosylation for stability or function have been expressed using the N-linked glycosylation system of Campylobacter jejuni engineered into E. coli.

Modified E. coli cells have been used in vaccine development, bioremediation, production of biofuels and production of immobilised enzymes.

87) Microbes in milk and dairy products

The fermentation is usually performed by lactic acid bacteria which ferment the lactose in milk and convert it to lactic acid leading to precipitation of the proteins.

There is a tremendous variety of fermented dairy products in many regions in the world. The properties of each product depend on the local strains used for thefermentation.

Many lactic acid bacteria have also been investigated for medicinal health benefits in the past few decades but so far the results are inconclusive.

Fermented milk or dairy products have been part of human diet since ancient times. Various fermented products are made by different strains. Lactic acidfermentation is performed most often by lactic acid bacteria. Due to their abundance in nature, including mucosal surfaces of the human body, and their use in fermented foods they are labeled as GRAS (generally recognized as safe). The main genera that belong to the lactic acid bacteria group are: Lactobacillus,Leuconostoc, Lactococcus, Pediococcus and Streptococcus. These bacteriaferment the carbohydrates in milk, the major one being lactose, to lactic acid and some other products. The acid precipitates the proteins in the milk and that is why fermented products are usually of thicker consistency than milk. The high acidity and low pH hinders the growth of other bacteria, including pathogens. Some lactic acid bacteria can produce agents with antimicrobial properties. Since milk is rich in many nutrients such as protein, calcium, phosphorus, and B vitamins dairy products are an excellent food.

Some of the most popular and widespread cultured dairy products are yogurt and cheese.

Records of yogurt preparation as food date back to centuries BCE. Classic yogurt is the result of the fermentation of two main bacterial species: Lactobacillus bulgaricus and Streptococcus thermophilus. Sometimes other lactic acid bacteriaare added as well. Yogurt is most often made of cow's milk although milk from sheep, goat, water buffalo, camels and yaks is used as well depending on the region of cultivation.

To make yogurt, the milk is first heated to 80ºC or boiled to kill any pathogenicbacteria and to denature the milk proteins to prevent the formation of curds. After it is cooled down to about 45ºC, the starter culture of the two species is mixed well with the milk and incubated at the same temperature for a few hours. In many countries, the traditional food is yogurt without any sweeteners which could be consumed plain or used to prepare a variety of dishes usually with vegetables (Figure 0). Yogurt has been traditionally consumed in Eastern cultures as a cold drink after mixing with water (e.g., lassi, ayran, doogh). After the industrialization of yogurt production in the twentieth century, yogurt with added sweetener and fruit or fruit jam has become popular in the Western world.

Cheese is another popular and ancient dairy product. It consists of milk proteins and fat together with lactic acid bacteria. It has longer shelf life than uncultured milk. Currently there are a few hundred varieties of cheese produced all over the world. Making cheese is similar to yogurt but after acidification usually with lactic acid bacteria (Lactococci, Lactobacilli, Streptococci), the solids are separated from the whey by coagulation with rennet and processed further to yield the final product. Depending on the type of cheese, the solids could go straight to packaging or other bacteria or mold could be added (e.g., Penicillium mold for blue cheese) for additional fermentation.

Other fermented and widely consumed cultured dairy products include kefir (lactic acid bacteria and yeasts are used for the fermentation), sour cream (fermented cream), cultured buttermilk (fermented cow's milk withStreptococcus lactis or Lactobacillus bulgaricus only).

Lactic acid bacteria have been researched for medicinal health benefits. In the early twentieth century, the Nobel laureate in medicine, Elie Metchnikoff, believed that the longevity of peasants in Bulgaria and the Russian steppes was due to their high consumption of milk-fermented products. He hypothesized that the lactic acidbacteria would inhabit the gut after consumption, create and acidic environment as they grow and multiply, and hence prevent the growth of proteolytic. After it was discovered that Lactobacillus bulgaricus can not live in the human gut, the idea was abandoned. Years later, strains of Lactobacillus acidophilus were found to thrive in the gut after implantation and the research started again. The term "probiotics" was introduced and defined as live microorganisms that provide beneficial effects for their host when administered in adequate concentration. Most of the researched species were isolated from different fermented dairy products. The research has been focused on curing or preventing a number of diseases like diarrhea, intestinal inflammations, urogenital infections, allergies, etc. Some species have been prepared and sold as nutritious supplements. However, so far there has not been enough evidence to establish a definite cause and effect relationship about any of the marketed products.