eChapters

Anthocyanins are members of the flavonoid group of phytochemicals, a group predominant in teas, honey, wines, fruits, vegetables, nuts, olive oil, cocoa and cereals. The primary players in this group include the anthocyanins (eg, cyanidin, pelargonidin, petunidin), flavonols (quercetin, kaempferol), flavones (luteolin, apigenin), flavanones (myricetin, naringin, hesperetin, naringenin), flavan-3-ols (catechin, epicatechin, gallocatechin), and although sometimes classified separately, the isoflavones (genistein, daidzein) (Skibola and Smith, 2000).

ABSTRACT

A plant grows in a highly competitive environment. It is continually being threatened by other plants encroaching upon the space from which it draws its sustenance, by microorganisms, by insects, avian, reptilian or mammalian herbivores. In order to survive, each plant must draw upon a complex of defenses, which may be physical, such as Spines or Leathery Leaves or Chemical. These defenses are passive, coming into action only when the plant is attacked which repel insects or the herbivores that attempt to feed on them. Plants protect themselves from insect attack by providing a multitude of repellent chemicals including toxic and bitter alkaloids and tannins. Non-specialist insects and other animals that attempt to feed on the plant may be repelled, injured through severe gastrointestinal malfunction or even killed. The cabbage, spinach, mustard etc. are well known for their mustard pungent flavor that deters most animals from consuming it. Even most medicinal plants like Cynanchum collialatum of the family Asclepediaceae have tannins or alkaloids like “cardenolides” that repel most animals and insects. All these plants have a basic enzyme “Myrosinase” which react with these specific tannins to produce the toxic metabolite, which repels insects and animals. But caterpillars of few insects are known to tolerate these toxic alkaloids (ex: caterpillar of Striped Tiger Butterfly-Danaus genutia) and in turn sequester the toxic alkaloids into their tissues making themselves non-palatable to their enemies. We hypothesize the analysis of possible roles of toxic alkaloids isolated from these butterflies as bio-insecticides

Introduction
According to the U.S. Food and Drug Administration (FDA), Good Laboratory Practices (GLP) is a quality system that ensures the nonclinical safety studies are appropriately designed, conducted, and documented to generate reliable data. The laboratory techniques are defined as the processes and practices that are recommended for equipment use in the laboratory, while laboratory methods are based on established scientific principles involving biology, chemistry, and physics, and encompass all aspects of the clinical laboratory from testing the amount of cholesterol in blood to analysing DNA to growing microscopic organisms that may be causing an infection. One of the most significant parts of laboratory techniques and laboratory methods is the proper validation of the equipments (old or new equipments) as well as all the testing procedures/ methods (established or newly developed methods). Analytical method validation is a critical process to ensure the quality and safety of products. The objective of the validation of an analytical method is to demonstrate that it is suitable for its intended purpose; the results are authentic and universally acceptable. An analytical method is suitably well considered when the results generated through different method are consistent, reproducible, and thus, reliable.
The validation is required when
• A new method has been developed
• Revision of established method
• Use of established methods in different laboratories and different analysts, etc.
• Comparison of methods required
• When quality control indicates method changes

? Lantana camara (lantana, wild sage, bunch berry) one of the ten most toxic weeds in the world. Brought to India in early part of the 19th century as an ornamental plant.

? Very hardy shrub grows very luxuriantly, capable of prospering in adverse soil.

? It has no. of adverse interactions with the biosphere.

? Causes hepatotoxicity, photosensitization in grazing livestock and growth inhibition of the neighboring vegetation due to allelopathic action which is rapidly changing the ecological balance.

? Reports of poisoning have been received form India, Australia, Cuba, Mexico, Kenya, Brazil and New Zealand.

? Very pungent plant. Cattle eat only in scarcity of other feed/fodder.

? Bos taurus more susceptible than Bos indicus.

LD 50- It is defined as the concentration/dose that kills 50 % of the experimental population.

Acute toxic studies have been conducted in order to calculate LD50. There are several methods to calculate LD50, some are as follows:

Methods to Determine LD 50

• Karber’s method.

• Miller & Tainter method

• Lorke method

• Fixed dose method

• Acute Toxic Class method

• Up & down procedure

Karber’s Method

This method involves the administration of different doses of test substance to various groups, which has five animals each. The first group of animals is administered with the vehicle in which the test substance was dissolved or diluted

Introduction
The emergence and spread of antimicrobial resistance has changed the landscape of infectious disease treatment, creating an urgent need for new diagnostic and therapeutic approaches4’7. Traditional phenotypic antimicrobial susceptibility testing, while remaining the gold standard, requires 24-72 hours for results and provides limited insights into resistance mechanisms. Limited turnaround time is often a constraint that requires empirical use of antibiotics, potentially driving the propagation of other resistance emergence as well as suboptimal patient outcomes. Thanks to mass spectrometry, clinical microbiology has greatly benefitted through the rapid detection of pathogens. However, the application is far broader than species identification. The integration of multiple omics fields like proteomics, metabolomics, genomics, and transcriptomics through mass spectrometry provides unprecedented means to unravel, predict, and combat antimicrobial resistance (AMR) on a molecular level. The use of a multiomics approach is demonstrated to show that AMR is a complex phenotype that arises from the intersection between genetic mutations, variations in gene expression, metabolic differences, and changes in protein structures that all act in unison to affect bacterial survival in the presence of antibiotics. The conversion of raw mass spectrometry outputs into clinical insights amenable to decision-making is dependent upon bioinformatics. Through the use of sophisticated computational methodologies, such as machine learning and network analysis, bioinformatics provides the means to incorporate heterogeneous omics datasets. Through this approach, resistance-related biomarkers can be identified, predictions for therapeutic efficacy can be made, and new targets for intervention can be identified. The intersection between high-resolution mass spectrometry, broad-based omic profiling, and strictbioinformatics provides a powerful platform for addressing the intractable challenge of antimicrobial resistance.

Introduction

In the 1997 WHO guidance document: “WHO/VSQ/97.01: (A WHO guide to good manufacturing practice (GMP) requirements. Part 1: Standard operating procedures and master formulae)” some basic explanations and instructions were given for preparing various documents required by Good Manufacturing Practice guidelines from WHO and from several regulatory authorities.

GMP guidelines include the requirements for documents (individual), documentation (the systems and formats for documents), and documenting (recording) of production and control activities. Most GMP guidelines provide the same or very similar information as the principles of Good Manufacturing Practice are now international in scope.

In this guidance document, the requirement for master manufacturing instructions and the requirements as given in different GMP documents, different names for these documents and various forms that they can take will be described. This is to guide vaccine manufacturers who are applying for pre-qualification or re-qualification of their product(s) in the preparation or improvement of current documents for manufacturing operations.

Introduction

Measurement of pH of a solution and preparation of different types/strength of buffers are common practices in a laboratory, particularly in the Biochemistry and Molecular Biology laboratories. Therefore, in this chapter you will be acquainted with the procedures of measuring pH of a solution as well as preparation of some of the commonly used buffers in a laboratory.

Potential or Power of Hydrogen Ion (pH)

Measurement of pH of a solution is measuring the molar concentration of hydrogen ion [H+] in the solution, which is an indication of the acidity or basicity of the solution. The symbol pH stands for “power of hydrogen” and numerical value for pH is just the negative logarithm of the molar concentration of hydrogen ions [H+]. The ionic product of water (Kw ) is the basis for the pH scale. The total hydrogen ion concentration from all the sources and it can be experimentally measured.

The pH scale designates the concentration of H+ (and thus of OH−) in an aqueous solution, which ranges between 1.0 M H+ and 1.0 M OH−. The term pH is defined by the expression

Thus, the ionic product of water makes it possible to calculate the concentration of H+, with given concentration of OH−, and vice versa. The value of 7 for pH of a neutral solution is not an arbitrarily chosen figure; it is derived from the absolute value of the ionic product of water. A solution having pH greater than 7 is basic, and the concentration of OH− is greater than that of H+. A solution having a pH less than 7 is acidic.

Q1. Resistance in tetracycline is due to mutational changes in

1. Tet A gene
2. Kat G
3. Par C par E
4. Inh A

Q2. Single step mutation can be seen in

1. Erythromycin
2. Tetracycline
3. Chloramphenicol
4. Rifampicin

Q3. Multistep mutation is observed in

1. Streptomycin
2. Rifampicin
3. Both A and C
4. Tetracycline

Introduction
The advent of antibiotics, beginning with penicillin in 1941, transformed medicine by rendering bacterial infections treatable [1]. However, antimicrobial resistance (AMR) has emerged as a profound threat, enabling bacteria to survive in the presence of drugs designed to inhibit or kill them and these are rooted in ancient microbial ecosystems evidenced by resistome genes in 30,000-year-old permafrost [2]. AMR has been exacerbated by human practices: approximately 50% of antibiotic prescriptions are inappropriate, 70% of global antibiotics fuel agriculture, and environmental pollution creates resistance hotspots [1]. By 2025, AMR directly causes 1.3 million deaths annually, with economic losses projected at $100 trillion by 2050 [1]. Priority pathogens, including carbapenem-resistant Enterobacteriaceae (CRE), methicillin-resistant Staphylococcus aureus (MRSA), and colistin-resistant strains via mcr-1, challenge last-resort therapies, particularly in low-resource settings [3]. This chapter delivers a technically extraordinary exploration of AMR, structured into genetic, biochemical, and evolutionary mechanisms, each elaborated with molecular precision and step-by-step clarity for learners. Genetic alterations provide the DNA-level foundation, biochemical pathways execute molecular defenses, and evolutionary forces drive adaptation and dissemination. It ensures scientific rigor and broad appeal for students, researchers, clinicians, and policymakers.
Historical Context and Global Burden
AMR emerged shortly after antibiotics’ introduction, with penicillin-resistant S. aureus identified by 1942 due to ß-lactamase production [1]. The 1960s

1. Introduction

During the last decade, considerable attention has been focused on the involvement of oxygen free radical (OFR) in various diseases. Molecular oxygen, while providing an efficient energy from ingested food, results in free radical and peroxide by-products, which have high intrinsic toxicity. Numerous physiological processes in living organisms occasionally produce free radicals, reactive nitrogen species (RNS) and reactive oxygen species (ROS) as by-products. Their broad ranges of effects in biological systems have drawn the attention of many experimental works. ROS have been implicated in the pathophysiology of several clinical disorders, including ischemia, reperfusion injury, myocardial infarction, rheumatoid arthritis, neurodegenerative, atherosclerosis, acute hypertension, hemorrhagic shock and diabetes mellitus (Hemnani and Parihar, 1998).

Oxidative stress occurs in a cell or tissue when the concentration of reactive oxygen species (ROS) generated exceeds the antioxidant capability of the cell (Simic and Jovanovic, 1986). Although humans and other organisms possess antioxidant defense and repair systems to protect against oxidative damage, these systems are insufficient under certain conditions to prevent the damage totally (Chang and Miles, 1982). Hence, the oxygen free radicals mediated damage to protein, DNA and other biomolecules accumulates during the lifetime of organisms. Many synthetic antioxidant components exhibited toxic or mutagenic effects that have directed the attention to the naturally occurring antioxidants in recent years. A multitude of natural antioxidants have already been isolated from different plant materials such as oilseeds, cereal crops, vegetables, fruits, leaves, roots, spices and herbs (Ramarathnam et al., 1995).

ABSTRACT

Our country is well known for developing into rich and ancient traditional system of medicines based on plants. For centuries, these ancient cultures and professional practicing on traditional system viz. Ayurveda, Unani and Siddha could remain in house hold and pass through generations to generation. India is being a developing country, of late uses bioactive natural products (secondary metabolites) using in vitro culture technology of the medicinal plant species. Recently, commercial utilization of these compounds using modern techniques is in good progress. The developed western affluent world has to now depend on these compounds mostly in drug and pharmaceutical industry in recent years. Orchids as medicinal herbs, details on plant medicinal potentialities, traditional medicinal systems viz. Ayurveda, Unani and Siddha have been described.

Feed additives like antibiotics, hormones are successfully employed in chicken production along with basic minerals, vitamins, and nutrients. Because of worries about antibiotic residues in animal tissues and the resulting development of bacterial resistance, the use of antibiotics as growth promoters (AGP) in cattle was outlawed in 2005. Given the rising concerns about antibiotic use and its involvement in antibiotic resistance, the use of plant-based chemicals in animal feed has become essential. Phytogenic feed additives, which are made from plants and their secondary metabolites, are becoming more and more popular as a result of the search for sustainable farming methods.
These natural growth promoters are seen as alternatives to synthetic chemicals, which are often associated with adverse environmental and health impacts. The usage of plant additions, also known as phytobiotics, have existed since the beginning of time, when people and farm animals were treated and prevented with herbs. The unique qualities of herbs and plant additions were effectively utilised by the ancient Greek, Egyptian, Roman, and Chinese civilisations. Phytogenic feed additives offer multiple benefits for livestock, including enhancing growth, improving immunity, supporting digestion, and promoting
overall animal health. Water or alcohol extracts derived from plant additions, whether fresh or dried, fermented, or freeze-dried, can be used in poultry practices.

The therapeutic potential of plants and its parts has been explored since ancient times and has been used for the well being of humans as well as animals. In the era of emergence of multi drug resistant bacteria modern research aims to find out alternative therapeutic approaches preferably from herbal source to combat the infection. Medicinal plants owing to their antibacterial, antioxidant, anti inflammatory capabilities have been used against many bacterial infections in livestock and companion animals. Ethnoveterinary practices have been documented against bacterial diseases in animals like mastitis, colibacillosis, Salmonellosis, pyoderma, otitis externa, gastrointestinal infection, pneumonia and many others, some of which are enlisted below.
Bovine mastitis, predominantly a bacterial disease continues to threaten the dairy sector as it inflicts great economic losses to the dairy farmers. The disease caused by more than 200 species of bacteria is characterized by inflammation of the mammary gland with physical and chemical changes in the milk and pathological changes in the glandular tissue. There is a significant reduction in the milk yield and many times it’s accompanied with abnormalities in the udder, milk and the cow. In addition to this, milk from affected animals may harbor the organisms which are potentially pathogenic to humans. Use of antibiotics is a common practice to cure mastitis.

The control of external parasites and the pertinent disease vectors using chemicals is riddled with various issues and hence there is a huge demand for alternative approaches. Natural products, being a prominent resource for poor farmers, can majorly serve as a cheaper, environment friendly and sustainable alternative. Further, ethno-veterinary and traditional knowledge has provided a diverse array of plants, which need to be validated for their repellent, insecticidal and acaricidal activities to claim their effect against external parasites. Besides, phytochemical study of these therapeutic plants has progressively revealed the existence of a variety of bioactive components in these plant derived extracts, catering to their efficacy against external parasites.
External Parasites in Domestic Animals
External parasites, also known as ectoparasites are organisms that live on the surface of domestic animals, feeding on their blood or skin. Fleas, ticks, mites, lice and flies are among the most common external parasites. These parasites can cause significant health issues in animals including discomfort, skin irritation and even severe diseases. Infestations can lead to conditions such as dermatitis, anemia and secondary infections. Moreover, many external parasites are vectors for serious diseases that can affect both animals and humans. Hence, effective management of these parasites is essential to ensure
the health and well-being of domestic animals.

Introduction
Helminths are a group of internal parasites of animals and human beings that usually infest gastrointestinal tract that feed from the host which provides protection to the parasite. The infestation cause loss of blood and nutrients from the host, making it weak, and vulnerable to other infections. There can be injury to other organs due to the migration of different stages of the parasite, formation of cysts or secretion of toxins that can cause damage.
The heminths belong to two phyla including nemathelminthes which are round worms and Platyhelminthes that has flukes, and tapeworms. Nematodes are round worms who have a resistant cuticle and most of them have separate sexes with direct or indirect life cycle. The major nematodes include Ascarids, Strongyles, strongyloides, Trichostrongyles, Filarids, Trichuris etc. Tape worms have a flat body with a head or scolex, neck and strobili having proglottids. They are hermaphrodites and the proglottids contain reproductive organs. They require more than one host for completion of life cycle. The flukes are ususally leaf like , unsegmented and have suckers that help them to attach to the internal organs of the host. They also have a life cycle in more than one host.

Viral Diseases in Domestic Animals
Viral diseases significantly impact the health and productivity of domestic animals, including livestock and companion animals, leading to substantial economic and emotional costs. In livestock, diseases like Foot-and-Mouth Disease (FMD) and Peste des Petits Ruminants (PPR) cause severe production losses in terms of milk, meat, and wool. Similarly, in poultry, Newcastle disease and Avian Influenza outbreaks result in high mortality and disruptions to egg production. Among companion animals, viruses such as Canine Distemper and Feline Leukemia Virus (FeLV) lead to serious morbidity and reduced quality of life. Current control measures rely heavily on vaccination and biosecurity practices, but the lack of effective antiviral treatments, coupled with the emergence of drug-resistant strains, underscores the need
for novel therapeutic approaches. Medicinal plants with antiviral properties have garnered significant interest as sustainable and holistic solutions. Plantderived bioactive compounds offer a dual advantage of modulating immune responses and directly targeting viral pathogens, making them a promising area of research for the management of viral diseases in both livestock and pet animals.

Herbal galactagogues are medicinal plants used to stimulate, sustain, and boost milk production. Several plants known for their galactagogue activity are being used traditionally in many cultures to stimulate milk production in animals. However, there is a knowledge gap regarding the safety and efficacy of these medicinal plants. Moreover, the mechanism of action of the herbal constituents present in these plants is not yet elucidated. Although, many herbal galactagogues have been reported as safe in domestic animals and can be used as food supplements in the dairy industry. Nowadays, the use of plantderived supplements has increased in the dairy sector promoting a new trend of organic dairy farming. Traditional remedies being used for increasing milk production in animals often incorporate herbal treatments, mainly comprising
of the following important herbs [1-8]:

Introduction
Arthritis, a condition characterized by inflammation of the joints, is prevalent among livestock species, particularly in older animals or those subjected to intensive farming conditions. It results in pain, reduced mobility, and diminished productivity. Herbal plants with anti-inflammatory and analgesic qualities are being investigated as a result of the increased interest in natural therapies, even if treatments like non-steroidal anti-inflammatory medications (NSAIDs) and corticosteroids have been utilized to manage arthritis in livestock. This chapter addresses the use of herbal plants to treat arthritis in cattle, including
a summary of the best plant-based remedies, their modes of action, and the scientific data that backs them up.
Pathophysiology of Arthritis in Livestock
Arthritis in livestock is primarily caused by joint degeneration, trauma, infection, or autoimmune disorders. The inflammation of the synovial membrane leads to the discharge of cytokines that cause inflammation, which degrade cartilage and cause pain. The condition significantly impacts the animal’s ability to move, affecting their overall well-being and productivity. In farmed animals, arthritis can lead to decreased milk production, reduced growth rates, and compromised meat quality, thus representing a significant economic loss.

Cardiovascular System Disorders in Domestic Animals
Cardiovascular system disorders in domestic animals include a range ofM conditions that affect the heart (pericardium, myocardium, endocardium), veins, arteries and lymphatic vessels. Infectious disorders like Heartworms (Dirofilaria immitis, vegetative endocarditis in pigs, equine viral arteritis). Patency of the foramen ovale, interventricular foramena, patent ductus arteriosus, persistence of the right aortic arch, coarctation of the aorta, transposition of the aorta, congenital aneurysm, pulmonic stenosis, subaortic stenosis, ectopia cardis, acardia, and diplocardia are examples of congenital and developmental disorders that can result in these conditions. Traumatic disorders like Traumatic reticulopericarditis and cardiac failure. Degenerative diseases like fatty and hyaline degeneration. Inflammatory disorders like pericarditis,
myocarditis, endocarditis and phlebitis may also lead to cardiovascular system disorders. Some tumours associated with the cardiovascular system like rhabdomyosarcoma, fibroma and myxofibroma also cause cardiovascular system disorders.