eChapters

Introduction

Antibiotic-resistant bacteria are a major cause of health care-associated infections around the world, and resistance has also emerged in infections in the wider community. Infections caused by multi resistant organisms significantly increase morbidity, mortality, and health care costs. Molecular analyses have revealed that widespread multi resistance has commonly been achieved by the acquisition of preexisting determinants followed by amplification in response to selection. The capture, accumulation, and dissemination of resistance genes are largely due to the actions of mobile genetic elements (MGE), a term used to refer to elements that promote intracellular DNA mobility (e.g., from the chromosome to a plasmid or between plasmids) as well as those that enable intercellular DNA mobility.

Insertion sequences (IS) and transposons (Tn) are discrete DNA segments that areable to move themselves (and associated resistance genes) almost randomly to newlocations in the same or different DNA molecules within a single cell. Other elements, such as integrons (In), use site-specific recombination to move resistance genes between defined sites. As these types of MGE are often present in multiple copies indifferent locations in a genome, they can also facilitate homologous recombination(exchange of sequences between identical or related segments). Intercellular mechanisms of genetic exchange include conjugation/mobilization (mediated by plasmidsand integrative conjugative elements [ICE]), transduction (mediated by bacteriophages), and transformation (uptake of extracellular DNA). Interactions between thevarious types of MGE underpin the rapid evolution of diverse multi resistant pathogens in the face of antimicrobial chemotherapy. Primarily the most important and/ or topical elements in Gram-negative and Gram-positive bacterial species of particular concern clinically, namely, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. (the so-called ESKAPE group) (Rice LB., 2008) as well as Escherichia coli (giving the ESKAPEE group) (Llaca-Díaz J.M. et al.,2012); nonetheless, many of the MGE types described are of course broadly relevant to many other bacterial taxa.

The term transgenic animal refers to an animal in which there has been a deliberate modification of the genome - the material responsible for inherited characteristics - in contrast to spontaneous mutation (FELASA September 1992, revised February 1995). Foreign DNA is introduced into the animal, using recombinant DNA technology, and then must be transmitted through the germ line so that every cell, including germ cells, of the animal contain the same modified genetic material.  By simple means a transgenic animal is an animal in which foreign DNA has been incorporated into its original DNA. Transgenic animals are altered so that their DNA produces chemicals that normally they would not produce.

I. Definitions

Transpiration - evaporation of water from a plant surface

Evapotranspiration - evaporation of water from a plant surface and the soil (and abiotic surroundings.

Take-home-lesson: Plants loose a lot of water by transpiration. See text for some specific examples.

Transplantation refers to the implantation of a cell, tissue or organ from one site to another site in an individual or from one individual to another. It involves implanting of organs or grafting of tissues to repair defects and to stimulate regeneration.

Transplantation refers to the implantation of a tissue or organ from one site  to another site in an individual or from one individual to another.It involves implanting of organs or grafting of tissues to repair defects and to stimulate regeneration. Auto-transplantation or autoplasty of the skin, cartilage, bone, muscle, tendon, nerve and fatty tissue is used in plastic and cosmetic surgery. The replacement of a diseased organ by a transplant of a healthy tissue is called transplantation.

Kingdom: Protozoa
Phylum: Euglenozoa
Class: Kinetoplasta
Order: Trypanosomatida
Family: Trypanosomatidae
Genus: Trypanosoma
Members of the genus Trypanosoma are responsible for trypanosomosis in animals and human beings and are found in the blood vascular system and tissues all over the world. All the species of trypanosomes are transmitted biologically or mechanically through an arthropod vector except T. equiperdum, which is transmitted through coitus in equines.

A tumor or neoplasm is an abnormal growth of the cells to form a lump or mass of transformed cells. This growth occurs due to loss of regulatory control of cell division. The majority of cells and organs have a balance between cell renewal and cell death. 

A tumor is said to be an abnormal growth of cells to form a lump or mass of transformed cells. This growth occurs due to loss of regulatory control of cell division. The majority of cells and organs have a balance between cell renewal and cell death. A cell has a particular life span;when it dies a new cell is generated. In normal condition, this system is regulated through cell cycle and cell division but at the time of any abnormality this growth mechanism is totally disturbed and failed. These cells give rise to clone of the cells and eventually produce tumor.

A. Answer the following.

Q.1.     What are tumour specific transplantation antigens (TSTAs) or tumour specific antigens (TSAs) ?
Ans.     Some antigens present on the surface of malignant cells are unique to the cancerous cells and are not present on their normal counterparts. They are called tumour specific transplantation antigens (TSTAs) or tumour specific antigens (TSAs).

Q.2.     What are tumour associated antigens (TAAs) ?
Ans.    Some antigens present on the tumour cells that are not different qualitatively from those found on normal cells but are overexpressed and present in increased number on the cancer cells as products of cellular oncogenes. They are called tumour associated antigens (TAAs). Ex the high level of a growth factor receptor due

There is only one type of scientific research and that is to solve a real problem or to understand the universe and provide a solution –either with the experiments or by putting a hypothesis. However, we have classified the research in various types as per our comfort. But for the employed researchers, it may be a matter of concern if they are working on time bound research schemes as their services may be co-terminus with the project and they may face the problem of un-employment, after termination of the scheme. According to the scholars’ activities and type of research being carried out in the country, we can discuss the subject under following topics.

This chapter examines the major forms and mechanisms of immunity. It distinguishes innate immunity from adaptive immunity and explains specificity, diversity, immunological memory and self–non-self recognition. Natural and artificial forms of active and passive immunity are discussed through examples involving infection, vaccination, maternal antibodies and antiserum administration. The chapter introduces pattern-recognition receptors, pathogen-associated molecular patterns and Toll-like receptors involved in microbial recognition. It also covers inflammation, its cardinal signs, acute-phase proteins and the roles of neutrophils and other innate defence components. These objective questions provide a clear foundation for understanding how animals resist infections and develop long-lasting protective immune responses.

Antibiotics, often referred to as “magic bullets” in the battle against bacteria, stand out as one of the most remarkable medical breakthroughs of the 20th century. Their advent revolutionized the therapeutic landscape, consistently saving millions of lives from bacterial infections (WHO, 2020). Undoubtedly, antibiotics are a blessing to humanity, not merely for their medicinal applications but also for their diverse uses. They have long been employed in various contexts, including animal husbandry and production, serving as preventive measures in numerous underdeveloped and developing nations for decades (Majumder et al., 2020). A range of antimicrobial agents, including antibiotics, antifungals, antivirals, disinfectants, and food preservatives, function to either inhibit microbial growth or eradicate them altogether. Among these, antibiotics, specifically tailored to combat bacterial infections, are the most widely used class of antimicrobials. With their increasing and often improper use, microorganisms have developed antimicrobial resistance (AMR) (Murray et al., 2022).

Antimicrobial resistance is an inevitable evolutionary process observed in all microorganisms, and is driven by genetic mutations aimed at resisting the lethal pressures of selection. Bacteria, adapt to environmental selection pressures by developing resistance to antibacterial drugs, thereby diminishing the efficacy of these treatments (Murray et al., 2022). The widespread use of antibiotics, particularly in developing nations, further flares up the process, leading to significant repercussions such as increased rates of morbidity and mortality (WHO, 2020). The incidence and prevalence of antimicrobial resistant bacterial infections have reached alarming levels in the 21st century, posing a substantial threat to global public health akin to a silent pandemic. Antibiotic resistance knows no borders and can impact individuals of any age or gender, regardless of their location. In its present state, antimicrobial resistance (AMR) stands as one of the foremost threats, not only to global health but also to food security (Murray et al., 2022).

Microbial Resistance Mechanism

Huge use of antibiotics in nosocomical environment has lead to the occurrence of multidrug resistance mechanism by microorganism against particular antibiotics (Munita et al., 2016; Zhao et al., 2020). Main operation includes chemical modification, efflux pump through passivation, modification of drug targeting gene and systemic elimination of antibiotics as many of them can form biofilm (Rajput et al., 2018) which can show extreme resistance. Chemical and enzymatic mechanism can include inactivation, activity loss, degradation and derivatizaton of antibiotics chemical group (Liu et al., 2019). Best idea may include production of enzymes or they can also expel antibiotic to efflux pump. Antibiotics drug efflux resistance can be accomplished by drug efflux pump playing an important role in microbial resistance. A different type of efflux pump system has been identified that include lipophilic and hydrophilic efflux system targeting the drug at different chemical properties (Wasaznik et al., 2009; Kumar et al., 2012). Another strategy adopting in resistance mechanism is modification of drug targeting gene. This mechanism makes the changes in drug to lose its target through drug targeting gene modification. Another important resistance mechanism is effectiveness of antibiotics by interfering with the target site. Therefore, bacteria has evolved different mechanism that include target site protection. A bactericidal role played by beta lactum antibiotics in inhibition of mucopeptide synthase and bacterial penicillin binding protein (PBP) in cell wall transpeptidation reaction that prevents the formation of complete call wall and hence dies (Sauvage et al., 2016). Bacteria also developed its own strategies to cope up with the environmental stress that can also include human body. In course of gaining the advantage bacteria needs to compete for nutrients and to keep away from the molecules produced by other competing organism (Zhao et al., 2019). Recently researchers have found the importance of biofilm resistance in bacteria. Bacterial biofilm resistance produce by complex and systematic drug resistance mechanism.

Globally, antimicrobial resistance (AMR) is an important public health and development threat (WHO, 2020). Being one of the top 10 global public health threats, the ever-increasing rates in the occurrence of drug-resistant bacterial infections have become a grave public health concern for the governments (Laxminarayan et al., 2013). Moreover, such infections would be difficult to treat and therefore can lead to worse clinical outcomes (Laxminarayan et al., 2013). Worldwide, each year about 700,000 people die from antimicrobial resistant infections and the mortality figures are projected to reach 10 million/ year by 2050 (O’Neill, 2016). The crude infectious disease mortality rate in India is more than 400 per 100,000 persons and at least 23,000 deaths in adults and 58,000 neonatal deaths per year are caused by AMR bacteria (Laxminarayan and Chaudhury, 2016). A 2017 estimate puts the cost of developing an antibiotic at around US$1.5 billion (Towse, et al. 2017). Meanwhile, industry analysts estimate that the average revenue generated from an antibiotic’s sale is roughly $46 million per year. “That’s tiny and nowhere near the.

Different basic as well as advanced laboratory equipment need to be used in a laboratory while conducting experiments. Many of them like pH meter, laminar air-flow hood, viscometer, thermometer, magnetic stirrer, microwave ovens, incubators, sand-bath, water-bath, oil-bath, etc. are the basic and simple equipment which require to be used routinely while conducting experiments in a laboratory. However, basic knowledge of their operation, handling procedure, when to be used, how to be used, and more importantly what not to be done with it, etc. are essential for keeping them in serviceable condition as well as for the accuracy/precision in their operation. In this chapter, we will discuss about some of the routinely used laboratory equipment along, while some other sophisticated equipment will be discussed in other chapters.

pH Meter

A pH meter measures exactly the H+ concentration of a solution; thus indicates acidity or alkalinity of a solution with greater precision. Actually, it measures the difference in electrical potential between a pH electrode and a reference electrode, and display the results converted into the corresponding pH value. Electrode is the key part (rod-like structure, usually made of glass) of a pH meter, having a bulb at the bottom containing a sensor. The pH meters range from simple and inexpensive pen-like devices (portable/pocket pH meter) to complex and expensive laboratory instruments with computer interface (Figure 5.1).

Frequent calibration of a pH meter using the solutions of known pH (standard solution of pH 4.01, 7.00, 10.01, Figure 5.2), generally before each use, ensures the accuracy of the instrument. For very precise work the pH meter should be calibrated before each measurement. For normal use calibration should be performed at the beginning of each day. This is necessary because the glass electrode does not give a reproducible measurement over a longer period of time.

Introduction

To accomplish experimentations in laboratories, sometimes we need microscope to test/validate our ideas/findings. A microscope is needed to examine objects which are very small which cannot be visualised by naked eyes. However, microscopic study requires some expertise in operating the microscope, particularly the sophisticated microscopes, as well as to prepare (peeling, sections, cutting, fixation, staining, dehydration, mounting, etc.) the sample, so that it can be appropriately examined for the anticipated details.

According to the scientific requirements different types of microscopes are used in the laboratories for research purpose. In this chapter, our aim is to acquaint you with the basics of using and handling of microscopes, particularly a compound microscope that is commonly used by the graduate and post graduate students for basic research.

Microscopy is often designated as the scientific investigation that is employed to review and analyse minute structures and objects using a microscope. The term “microscope” has been derived from ancient Greek words “micros” meaning “small” and “skopeîn” meaning “to look at or see,”.

Microscope

A microscope is a laboratory instrument used to examine objects (biological specimens) which are too small to be seen by the naked eye. The microscope must accomplish three tasks:

• Magnification- to provide a magnified image of the specimen.

• Resolution- to separate the details in the image.

• Contrast- to render the details clearly visible to the eyes.

Although we will tnroduce to you the different types of microscopes used for different speciments, our main focus is to discuss about compound microscope, which is commonly used in the laboratories. Compound microscope was developed around the beginning of 17th century.

A variety of glassware like burettes, pipettes, measuring cylinders, flasks, separating funnel etc. are used for experimental purposes in a laboratory. All of these need to be used and handled correctly, otherwise the chances of experimental error and damage to the glassware increase significantly. Being made of glass, they need special care while washing and using in experiments because even a minor mistake in handling of glassware may result in breakage of the apparatus. With this precautionary note, let us discuss about some of the specific glassware used in the laboratories.

Burette

Burette is a long graduated glass tube with a stopcock at one end for delivering a known/desired volume of a liquid, especially in titration experiments. It contains a stopcock at the lower end and a tapered capillary tube as the stopcock’s outlet. The flow rate of liquid from the tub stopcock’s outlet can be controlled with the help of stopcock valve. Burette is used for measuring the volume of a solution/chemical added in a quantitative (titrimetric) estimation experiment.

Procedure of using a Burette

The burette reading must be noted before and after delivering the chemical/ solution from the burette. The difference in these two readings is the volume of the chemical/solution released out of the burette. The solution should be released slowly/drop-by-drop to avoid any experimental error/mistake. If the solution/chemical is allowed to pour out too fast, the wall of the burette will not drain properly and some liquid may remain sticking to the surface of the walls. Moreover, excessive amount of chemical may be released/dropped during the experiment, particularly during the titration experiment wherein we need to know the exact amount of acid/base needed to get neutralization point. Thus, any minor mistake may lead to get a faulty reading. The measuring capacity of a burette usually used in a laboratory is 50 mL.

Since mouth pipetting of liquid reagents in a laboratory is not advisable, different tools are needed for safe, easier, and precise dispensing of liquid reagents. This chapter is devoted to make you aware of the tools like vacupette and micropipette which can be used for the handling of liquid reagents in a safe and precise way.

Vacupette

Glass pipettes have been traditionally used by sucking liquid solvent/solution directly using mouth. For safety reasons, mouth pipetting is no longer permitted in a laboratory. Hence, to fill a pipette vacupette is used by placing it on the top of glass or plastic pipette. It acts as a device to create vacuum for filling reagent in the pipette as well as for releasing the reagent with mechanical regulation on the flow out of the liquid. Vacupette can be of different types.

1. Rubber Bulb Vacupette

Rubber bulb vacupette is often used to transfer (pump up and discharge) liquid using a pipette when a larger quantity of chemicals is to be transferred. Small/tiny rubber bulb is attached to glass/plastic pipette for drawing little amount of reagents. Small vacupette is used to precisely discharge smaller volume/ drops.

Handling of Vacupette

• Insert the top of a pipette in the bottom of the rubber bulb vacupette (Figure 6.1).

• Remove air from the pipette by squeezing valve “A” at the top of the vacupette while simultaneously squeezing the rubber bulb. The amount of air released depends on the size of the rubber bulb.

Introduction
The discovery of antibiotics and their subsequent use in combating infectious diseaseswas amedical breakthroughin the 20th century, as it completely changed the treatment approach to infections in general and bacterial infections in particular. In 1928, Alexander Fleming discovered Penicillin that started the started the so-called “golden age of antibiotic”. However, later in course of time the underuse, overuse and misuse of antibiotics in medical and veterinary practice led toemergence of drug-resistant pathogens resulting in phenomenon called antimicrobial resistance (AMR). In last few decades AMR has evolved silently and now threatening the human and animal health and has acquired the potential to be a next pandemic. AMR develops in a microorganism (e.g., bacteria, viruses, fungi, parasites) when it accumulates genetic changes over time and fails to respond to antibiotic(s) leading to severe illness and mortality due to infection. Although, AMR is an evolutionary event occurring naturally in microbes, but inappropriate use of antibiotics in humans and animal medicine act as key driver to its spread. Global antibiotic consumption increased between 2016 and 2023, with total consumption rising by 16.3% to 34.3 billion defined daily doses (DDDs) in reporting countries and a projected 20.9% increase when extrapolated to nonreporting countries. World Health Organization (WHO) and other authorities have called for coordinated action to address antimicrobial resistance, a threat looming large on horizon. In 2021, there were an estimated 1.14 million deaths attributed directly to Antimicrobial Resistance (AMR), with globally another 4.71 million deaths associated with AMR infections. This dataindicates a significant global health burden, with prophecy that AMR deaths could increase dangerously by 2050 if current trends continue. Global Action Plan (GAP) of the WHO highlights the importance of “One Health approach” calling for the joint action in human health, food production, animals and environmental sectors, to achieve better results. In the year 2015, the WHO initiated the Global Antimicrobial Resistance and Use Surveillance System (GLASS)to collect, analyze and share AMR data received from all countries. The Global Action Plan on AMR lists five strategic objectives to contain AMR viz., 1. optimizing the use of antimicrobials, 2. preventing infections (including the use of vaccines), 3. strengthening surveillance and research, 4. improving awareness and understanding of AMR, and 5. investing in new medical products. Vaccines stimulate the body’s immune system to recognize and attack specific pathogens and thus can prevent AMR through several ways viz., 1. reduce the incidence of infections leading to reduction in morbidity and mortality, 2. vaccines can prevent secondary bacterial infections e.g. Streptococcus pneumoniae after an initial infection with influenza, 3. produce herd immunity, 4. Prevention of infections resulting in decrease in use of antibiotics that reduces development of AMR. Immunotherapeutics can both combat and be complicated by Antimicrobial Resistance (AMR). Other immunotherapies antibodies, and immune-boosting agents can directly fight AMR by preventing infections and reducing the need for antibiotics.

This chapter examines the principles, preparation and application of vaccines and biological products. It compares live attenuated, inactivated, toxoid, subunit, recombinant, vector, DNA and mRNA vaccines. Vaccine development phases, immunogenicity, safety, efficacy and post-marketing surveillance are introduced. The chapter discusses adjuvants, including alum, Freund’s adjuvants, liposomes, ISCOMs and delivery systems. Important veterinary vaccines for brucellosis, foot-and-mouth disease, anthrax, Newcastle disease, infectious bronchitis, goat pox, sheep pox and peste des petits ruminants are covered. Questions also address bacterins, toxoids, attenuation, booster requirements and reversion to virulence, providing practical knowledge of immunoprophylaxis in animal-health management.