eChapters

Objectives

To acquaint with the various methods of sterilization and disinfection used in laboratory animals.

Materials Required

Boiler, autoclave, hot air oven, chemical disinfection.

Sterilization

It is a process of the complete removal or destruction of living organisms including viruses, vegetative forms and spores. Sterilization is an absolute term. Nothing can be “partially sterile’’, either, it is or it is ‘not’. An article must be protected from recontamination after sterilization.

Introduction
Sterilization of culture media, containers, and instruments is crucial in bacteriological work to ensure the isolation and maintenance of pure cultures. Sterilization and sanitization are essential processes in a microbiology lab to prevent contamination and ensure accurate results. Sterilization refers to the complete elimination of all microorganisms, including bacteria, fungi, viruses, and spores, using methods like autoclaving, dry heat, filtration, and chemical agents. Sanitization, on the other hand, reduces microbial load to a safe level through cleaning and disinfection using alcohols, detergents, and UV light. These practices maintain aseptic conditions, protect lab personnel, and preserve sample integrity. Proper sterilization and sanitization are fundamental to microbiological research, clinical diagnostics, and industrial applications, ensuring reliable and reproducible experimental outcomes.
Preparations for Sterilization Before sterilization, cleaned and dried test tubes and flasks should be carefully plugged with clean, non-absorbent cotton. Pipettes should also be plugged at
the mouth end, and the plugs should remain in place during use to minimize oral contamination. Cotton plugs provide sufficient ventilation for bacterial growth while preventing external contamination.

Introduction

Microbes like all other living beings require suitable nutrients as well as favourable environment like pH, osmotic pressure, temperature, humidity etc. for growth. The substrate/food base which fulfills any of these requirements and supports the growth of an organism is called culture medium. Selection of a medium to grow a particular type of microorganism is extremely important, as the microbes vary in their nutritional requirements. Thus, different media are used for different groups of microbes. The following are the main types of media:

a) Natural/Empirical media: These are made up of natural substrates, such as herbaceous or woody stems, seeds, leaves, corn meal, wheat germ, and oatmeal etc.The exact chemical composition of this media is not known. e.g. Nutrient agar

b) Semi-synthetic media: 

Sterilization Techniques

Sterilization: All the materials, e.g., vessels, instruments, medium, explants, etc., used in culture work must be freed from microbes. This can be achieved by one of the following approaches:

Sterilization is a process of killing all microorganisms, including bacteria, fungi and their spores, and viruses from animate or inanimate surfaces with the use of either physical or chemical agents. Joseph Lister first gave antiseptic principle in 1867.

Sterilization is defined as the process by which an article, surface or medium is freed of all living microorganisms either in the vegetative or spore state.

Disinfection means the destruction or removal of all pathogenic organisms, or organisms capable of giving rise to infection. Chemical disinfectants which can be safely applied to skin or mucous membrane and are used to prevent infection by inhibiting the growth of bacteria are called antiseptics.

Five main methods are used for sterilization. 1. Red heat (flaming) 2. Dry heat (hot air) 3. Moist heat 4. Filtration 5. Radiation.

The killing effect of dry heat is due to protein denaturation, oxidative damage and toxic effect of elevated levels of electrolytes. The lethal effect of moist heat is due to the denaturation and coagulation of protein. In the case of spore, steam condenses on it, increasing its water content with ultimate hydrolysis and breakdown of the bacterial protein.

Sterilization is a thermal process aimed at destroying all microorganisms and microbial spores in milk, ensuring the product is microbiologically safe and shelf-stable at room temperature for extended periods. It is primarily used in the production of sterilized milk and evaporated milk, with typical conditions involving heating at 116°C for 15 minutes. This process kills all microorganisms and also inactivates all milk enzymes, including plasmin, which might otherwise contribute to spoilage during storage. However, enzymes produced by psychrotrophic bacteria should not be present in raw milk, as they are highly heat-stable and may remain active even after sterilization. Sterilized milk can be stored at room temperature for longer period. Sterilization kills all microorganisms and inactivates all microbial spores that may germinate under the storage conditions of the product in the market.
The Food Safety and Standards Authority of India (FSSAI) define “sterilization” in relation to milk and milk products as the process of heating the product in a sealed container continuously to at least 115°C for 15 minutes. This treatment ensures that the milk can be preserved at room temperature for a period of not less than 30 days.
In-bottle sterilised milk is a shelf-stable dairy product produced by sealing milk in glass bottles and subjecting it to high-temperature heat treatment. Typically, the milk is heated to temperatures ranging from 110°C to 121°C for 15 to 30 minutes. This process effectively destroys all vegetative microorganisms and bacterial spores, making the milk microbiologically safe and extending its shelf life up to 6–9 months without refrigeration. The heating is usually done in batch retorts or autoclaves after the milk has been bottled and sealed. Although the process ensures safety, it often leads to a slight cooked flavour and some nutritional losses, particularly in heat-sensitive vitamins such as vitamin B1 and vitamin C.

Sterols are derivatives of a complex ring system called cyclopentano perhydro phenanthrene. The term sterol means solid alcohol. Sterols, though alcohols, are solids with a M.P. of 100-120^ºC. The 27 carbon atoms impart hydrocarbon character, namely, solubility in non-polar solvents. Free sterols in plants are mono hydroxy  secondary alcohols varying in the constitution of methyl or ethyl group and in double bonds. Cholesterol, ergosterol, bile acids, sex harmones, cortical harmones and D-vitamins are some of the sterol derivatives.
   
The most widely distributed sterols in higher plants are the C₂₉ sterols, stigmasterol, β sitosterol and g sitosterol. Together, they are called steroids. In plants, steroids are present as glycosides. Cholesterol was first isolated from gall stones. The word cholesterol means “solid alcohol from bile”. Bile is a fluid which is formed and stored in the gall bladder. It aids in the digestion of fats in the digestive tract. Sterols form an important group among the steroids. Unsaturated steroids with most of the skeleton of cholestane containing a 3β-hydroxyl group and an aliphatic side chain of 8 or more carbon atoms attached to position 17 form the group of sterols.

1. Introduction

The global population is very cautious about gaining weight and diabetics. Calorie less and sugar free substances are gaining momentum in food sector. The miracle plant stevia is very fit in this scenario as it is both zero calorie and sugar free and one more additional benefit that it is completely natural. Stevia rebaudiana Bert. belonging to the family asteraceae has been used in many countries for thousands of years as a source of natural sweetener eg., in Paraguay and Brazil Central America, Thailand, Korea and China (Megeji et al., 2005, Sivaram and Mukundan, 2003; Jain et al., 2009). It is also known as sweet herb, sweet leaf, honey leaf, candy leaf, honey yerba (Carakostas et al., 2008) and meeti tulsi. It possesses an extensive root system and brittle stems producing small, elliptic leaves. In natural conditions S. rebaudiana grows in a form of a shrub and reaches even 1 m of height. Leaves are the major economic part. They are sessile and oppositely arranged, lanceolate to oblancoelate in shape, and are serrated above the middle. Trichome structures on the leaf surface are of two different sizes, one large and one small (Shaffert and Chetobar 1994). The white florets are, borne in small corymbs of 2–6 in number. Corymbs are arranged in panicles. Dr. Moises Santiago Bertoni discovered this plant in 1888 at Paraguay. In 1905, the plant was scientifically named as S. rebaudiana after a Paraguayan chemist Dr. Rebaudi. (Gupta et al., 2013). The active components of Stevia i.e., steviol glycosides (SGs) includes rebaudioside A (Reb A), rebaudioside C and stevioside (ST) that are 250-300 times sweeter than sugar (Chalapathi & Thimmegowda 1997). Stevia is a short-day plant and its flowering is induced when days are shorter. Long photoperiods stimulates leaf growth and steviol glycoside content. The plant can survive only mild winters of its native regions. It cannot survive in waterlogged areas. Stevia extracts can be used in a wide variety of food and beverage applications. Individual usages and use levels may vary depending on the country. They act synergistically when used in combination with other sweeteners. The higher market demand favors the cultivation and commercialization of this particular species.

15.1. Origin and distribution

• Also known as Cheeni tulsi or Mou tulsi.

• Originated from Seirra Amambay in North eastern Paraguay and adjacent areas in Bolivia and Brazil.

• Widely distributed in U SA and Brazil.

15.2. Economic importance

• Economic part - leaves

• Candy leaf

• Major constituents-Stevioside (3 to 10%), rebaudiside-A (13%), rebaudiside- B and C

Data mining is a powerful technology which helps in extracting future trends and behaviour from large databases. Thereby it helps decision makers to make knowledge-driven decisions. One common form of real-world data is time series. The suitable models are required to analyze time series data. This chapter provides a short introduction to stochastic system, uncertainty, basic statistics time series analysis and some stochastic models.

Stock or Gillyflower (Matthiola spp.) belongs to family Brassicaceae. It is a hardy annual, biennial and perennial plant, flowering in terminal clusters. Matthiola incana is ornamentally important and grown for various purposes i.e. cut flower, fragrance, pot culture, etc. This species develops a woody base in areas where it grows as a biennial or short-lived perennial, hence the name of stock. Generic name “Matthiola” was given in the honour of “Dr. Pietro Andrea Gregoria Matthiole” (1501-1577), personal physician of Emperor Maximilian of Austria. “Incana” means “hairy or gray-white” in reference to plant leaves. It exists as both single and double forms and is valued for fragrant flowers that can be used as fresh or dried cut flower. They are excellent pot plants for the greenhouse in winter and spring and provide good cut flowers. Flowers are well arranged on a long column and colours are variable from white to rose, crimson, purple, yellow, mauve, pink, etc. The highly fragrant flowers are used as a garnish, especially with sweet deserts. Many old names were given to Matthiola incana including stocks, sea stocks, wallflowers and wall or gillyflowers.

ORIGIN AND DISTRIBUTION

Matthiola incana is native to the Iberian Peninsula, France, the Apennine Peninsula, the Balkan Peninsula, the British Isles, Turkey and Cyprus. According to Greuter et al. (1986), four species of Matthiola are known in Sicily and its surroundings: M. fruticulosa(L.) Maire, M. incana (L.) R. Br. & Ait., M. tricuspidata (L.) R. Br. and M. sinuata (L.)R. Br. Unlike its congeners, which only have a single taxon in this archipelago,M. incana consists of three infra-specific subdivisions (Lojacono-Pojero, 1888 and Greuter et al. 1986): ssp. incana (L.) R. Br., which is widespread in other insular and continental Mediterranean enclaves, and the narrow endemics ssp. pulchella (Conti) Greuter & Burdet (confined to the islet of Pantelleria) and ssp. rupestris (Raf.) Nyman, distributed in the surroundings of Palermo (Sicily) and in the Aeolids, the Egadian and Ustica, respectively (Raimondo et al. 1994).

Stock earlier known as Leucoium album or Viola alba was later renamed as Mattiola after Dr. Mattioli, an Italian physician and botanist. This was being grown by the ancient Greeks and was linked for its frangrance. In the wild state, stocks are found in the Mediterranean, Egypt, South Europe and in South Africa and two species, Mattiola incana and M. sinuate are among the rare British natives. Ruth et al. (2009) also reported that Matthiola is widespread in the Mediterranean and Irano-Turanian regions. Both single and double forms were grown in Europe from 10^th century onwards and became popular garden flowers. Apart from its countries of origin, Matthiola incana can be found in Australia, Austria, Bolivia, Bulgaria, China, Colombia, Ecuador, Finland, France, Germany, Greece, Ireland, Israel, Japan, Kazakhstan, Mexico, The Netherlands, New Zealand, Peru, Romania, Sweden, United Kingdom and the United States.

Stock, primarily represented by Matthiola species, is an important cool-season cut flower valued for attractive colours and fragrance. This chapter covers its origin, taxonomy, botanical description, genetics and commercially important series and cultivars. It explains climate, soil, propagation, nursery raising, planting, spacing, irrigation, nutrition and greenhouse production. Special attention is given to maintaining strong stems and quality flowers, as well as avoiding excessive nitrogen that can result in weak growth. Harvesting, postharvest management, ethylene control, vase-life extension and yield are discussed. The chapter provides practical information for producing commercially valuable stock flowers for domestic and specialized markets.

Stock or Gillyflower (Matthiola spp.) belongs to family Brassicaceae. It is a hardy annual, biennial and perennial plant, flowering in terminal clusters. Matthiola incana is ornamentally important and grown for various purposes i.e. cut flower, fragrance, pot culture, etc. This species develops a woody base in areas where it grows as a biennial or short-lived perennial, hence the name of stock. Generic name “Matthiola” was given in the honour of “Dr. Pietro Andrea Gregoria Matthiole” (1501-1577), personal physician of Emperor Maximilian of Austria. “Incana” means “hairy or gray-white” in reference to plant leaves. It exists as both single and double forms and is valued for fragrant flowers that can be used as fresh or dried cut flower. They are excellent pot plants for the greenhouse in winter and spring and provide good cut flowers. Flowers are well arranged on a long column and colours are variable from white to rose, crimson, purple, yellow, mauve, pink, etc. The highly fragrant flowers are used as a garnish, especially with sweet deserts. Many old names were given to Matthiola incana including stocks, sea stocks, wallflowers and wall or gillyflowers.

INTRODUCTION

Stock or Gilly flower is an annual or biennial plant flowering in terminal clusters. It exists as both single and double forms and is valued for fragrant flowers that can be used as fresh or dried cut flower. Stocks are a most valuable race of garden plants for they are easy to grow and can be in bloom during the greater part of the year. They are excellent pot plants for the greenhouse in winter and spring and provide good cut flowers. Flowers are well arranged on a long column and the colours are variable from white to rose, crimson, purple, yellow, mauve, pink, etc. Seeds of stock are aphrodisiac, bitter, diuretic, expectorant, stimulant and are used as tonic. An infusion has been used in the treatment of cancer and when mixed with wine, it has been used as an antidote to poisonous bites. The highly fragrant flowers are used as a garnish, especially with sweet deserts.

Sverdrup’s contribution could not take into account of the western boundary and so could not explain about the circulation what happened along the west coast of the ocean. Thus his circulation, as shown in Fig.8.1c in the chapter 8, is incomplete. This problem was solved by an American Oceanographer, Henry Stommel, in 1948.

The stone fruit crops namely peach, Prunus persica, plum, Prunus domestica, apricot, Prunus armeniaca, almond, Prunus dulcis and walnut, Juglans regia are grown in wide range of climatic conditions ranging from subtropical to temperate regions. Peach and plum are also cultivated in mid and low hills in addition to temperate regions. Peach is the major stone fruit crop followed by plum, apricot and cherry. Walnut and almond on the other hand are major nut crops. A number of insect pests (~80) infest stone and nut fruits in various stages of the tree growth and development and cause direct and indirect loss out. Only few pests are of economic concern, the details of which are given hereunder

Peach leaf curling aphid, Brachycaudus helichrysi (Kalt.), Aphididae: Hemiptera

It is a polyphagous pest reported infesting 175 plant species of 115 genera of 49 families. It is a regular and key pest of stone fruits viz. peach, nectarine, plum, apricot and almond and

32 – 33. Agronomic practices for fodder production: Feeding green forages to animal is economic and better for animal health and production. To get green forage throughout the year, it is necessary to sow the crop at proper time and also to harvest it at a proper stage. According to season, following agronomical practices may be followed:
(a) Rabi crops: Berseem, Lucerne, Oat, Mustard, Sugarcane tops
(b) Zaid crops: Chari, Maize, Sorghum, Cow pea, Guar.
(c) Kharif crops: Maize, Sorghum, Cow pea, Sudan grass, Napier, Para grass.
Berseem and Lucerne (legumes) are best forages for milk production. Berseem is called as king and Lucerne as queen of the fodder crops. Maize, Sorghum, Oat (non – legumes) at flowering stage are good forages. Hybrid Napier, Guinea grass, Para grass and other green grasses are palatable, but poorer in nutritive value than legumes and many non – leguminous forages. Straws and
stovers are poor quality roughages. Average voluntary intake of legumes is 2.0 kg; Non legumes and other grasses have 2.25 kg; while of straws, stovers is only 1.5 kg Dry Matter / 100 kg body weight.

1. Introduction

 Mushrooms have been part of man’s diet for centuries (Rajarathnam and Shashirekha, 2003). Mushrooms grow on wide range of agro-residues, biodegrade and biotransform them into edible fruiting bodies (Rajarathnam and Zakia Bano, 1988a; 1988b; 1990; Zakia Bano and Rajarathnam, 1988; Zakia Bano et al., 1993; Rajarathnam et al., 1998). Their post-harvest life is very ephemeral at room temperature (Rajarathnam et al., 1983) due to delicate flavor (Rajarathnam et al., 1990) and high moisture content, with a range of chemical constituents (Zakia Bano and Rajarathnam, 1986; 1982; 1988; Rajarathnam and Zakia Bano, 1990; Rajarathnam et al., 1983; 1992). Since, they contain large quantities of water and therefore suffer considerable weight loss during transportation and storage, causes serious economic losses, due to reductions in weight and quality (Gray, 1970). When the supply of water, organic substances and minerals are cut off at harvest, the fresh mushroom enters a deterioration or perishable phase. Cap opening and stem elongation are the usual symptoms of senescence constituting visible evidence of deterioration. At room temperature, the shelf-life of mushrooms cannot be for more than 24 h (Rajarathnam et al., 1983). During this period, there are considerable changes in color, texture and taste. In addition, water is continuously being lost as a result of transpiration and respiration. Additional loss occurs from surface cracks and bruises that develop during growth and handling (Salunkhe et al, 1991). Fresh mushrooms are white or light buff, with no dark marks on either the cap or stem. The veil is closed and the gills are not visible. The upper surface of the caps should be strongly convex and stems should be plump rather than elongated. Mushrooms are grown on compost, usually in mushroom houses where the temperature and humidity can be controlled. Eventually after harvest, the mushrooms can never be able to withstand any increase in temperature and decrease in relative humidity. Mushrooms are commercially preferred at the immature stage characterized by closed caps, invisible lamellae, round stipe and light color. Agaricus development is classified into various stages (Table 1). Thus, the concept of physiology of fresh mushrooms being very complicated, bears a great impact on the delicate problem of extending the storage life. The various means and methods studied and adopted by various researchers to aim at extended storage life of fresh produce, underlined by its physical, chemical and biochemical properties and also development of value added products are critically evaluated.