eChapters

Introduction
• A cell is the most basic structural and functional unit of life. These are of two types:
i) Prokaryotic cells– Prokaryotes are those cells which lack a membrane bound nucleus and other membrane bound cell organelles. These cells are primitive cells. Examples, bacterial cells, archaeal cells.
ii) Eukaryotic cells– Eukaryotes are those cells which have distinct membrane bound organelles. These are advanced cells. Example, animal cells, plant cells, fungi.
• A microorganisms (from the Greek: mikros, “small” and organism’s, “organism”) are those small living organisms which cannot be seen with naked eyes. These are of two types:
i) Single cell– Organism made up of a single cell.
ii) Multi-cellular– Organism made up of multiple cells.
Shape and Arrangement of Bacterial Cells
• Prokaryotes display following shapes
• Spherical cells called Cocci
• Cylinders called rods or Bacilli
• Coma shaped called Vibrio
 

Introduction
Ocular micrometer and stage micrometer are used to measure the size of microorganisms. The Ocular micrometer consists of a circular disk of glass which has graduations engraved on surface. A stage micrometer or objective micrometer has scribed lines on it that are exactly 0.01mm (10 micrometer) apart. Ocular micrometer is calibrated for the objective first. To calibrate the ocular micrometer for a given objective, the two scales are superimposed and determined how many of the ocular gradation coincide with one graduation on the stage micrometer scales. Thus, the distance between the graduations on the ocular micrometer can be determined and so the size of the organism can be determined.
Materials
1. Ocular micrometer
2. Stage micrometer
3. Microscope
4. Stained slides of yeast or bacteria

A. Construction, Use and Care of Light Microscope
Introduction
Microorganism are invisible to naked eyes, hence their detailed study requires a microscope. A microscope can be defined as an optical instrument consisting of a lens or a combination of lenses for getting enlarged or magnified images of minute objects. A simple microscope is essentially an ordinary magnifying lens held on an adjustable stand. Usually it gives magnification up to 20 times the size of the object. A compound microscope consists of two separate lens system – the objective and the eye piece. It gives much greater magnification (100-1500 times) than a simple microscope and is, therefore useful for viewing and examining very minute objects like protozoa, bacteria, yeast, mold and algae which cannot be seen with naked eyes.
The objective of this exercise is to familiarize the students with construction and the use of compound light microscope.
1. Construction of Microscope
A microscope contrite various parts in which principle parts are described below:
i) Frame work: All microscopes have a basic frame structure which includes the arm and base. All other parts are attached to this framework and the base support the microscope.
ii) Stage: It is a horizontal platform that supports the microscopic slide. It has a mechanical stage, which is used for holding and moving the slide around on the stage.

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.

1. Introduction
All glassware used for bacteriological work must be meticulously cleaned and sterilized. Used and soiled glassware should be thoroughly washed to remove dirt and solid residues before drying and undergoing sterilization.
2. Cleaning of New Glassware
Borosilicate glass (e.g., Pyrex) and factory-washed soda glass require only standard washing before use. However, new, unwashed soda glass needs additional treatment to neutralize alkali content.
• Place the items (e.g., test tubes, Petri dishes, and flasks) in a suitable container, cover with a 1% tri-sodium phosphate solution, and bring to a boil.
• Remove the items, rinse with tap water, and immerse them in a 1% hydrochloric acid solution to neutralize alkalinity.
• Rinse thoroughly with tap water, followed by distilled water. Allow the glassware to dry before sterilization.
3. Cleaning of Used Glassware and Other Apparatus
• Autoclave all glassware contaminated by microorganisms (121°C for 15–20 minutes). This liquefies solid media, facilitating removal.
• Rinse the empty glassware with tap water, then warm water (50–55°C).
• Soak in a detergent solution (e.g., 1% washing soda or tri-sodium phosphate solution) and scrub with a brush.
• Rinse thoroughly with tap water to remove detergent traces, followed by distilled water.

Microbiologists are constantly at risk of exposure to hazardous microorganisms that can cause diseases. These microorganisms may enter the body through inhalation, ingestion, cuts, abrasions, or the eyes. Therefore, handling cultures, slides, and live microorganisms requires utmost care. To ensure safety in the laboratory, several precautions must be followed:
1. Maintain cleanliness in microbiological labs at all times, taking extra care to prevent contamination from the air, equipment, personnel, or the environment.
2. Always wear a clean white lab coat or apron while working. After handling infectious microorganisms, the lab coat should be autoclaved.
3. No eating, drinking, or smoking is allowed in the laboratory.
4. Personal hygiene is essential in microbiological labs.
5. Wash hands with soap and water or a mild disinfectant solution (e.g., hypochlorite solution containing 50-100 ppm chlorine or 1-2% lysol solution) before starting work.
6. Disinfect the work surface by wiping down the lab bench with a disinfectant solution, such as 5% phenol or hypochlorite solution containing 200-300 ppm chlorine, both before and after work.
7. Keep the work area neat and organized at all times.
8. Use self-adhesive labels instead of moistening labels with your tongue.
9. Sterilize inoculating loops and needles by heating them in a Bunsen burner flame until red hot before and after use.

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.

Introduction

The epidemiological risk and hazards to global health security posed by AMR has been reiterating in several World Health Assembly (WHA) declarations and has been prioritized under the Global Health Security Agenda (GHSA). The Indian Ministry of Health & Family Welfare (MoHFW) has also identified AMR as one of the top priorities for collaborative work with WHO and the first National Action Plan on AMR has been implemented (2017-2022). India ranked fifth in antibiotic consumption in food producing animals (ruminants, pigs, and poultry) in 2010. Even though there has been a gradual reduction of 3.6% in antibiotic consumption during 2011-2019 (Koya et al., 2022), with mounting incomes and shifting in dietary patterns in favour of the consumption of animal protein, especially for poultry, the veterinary antibiotic utilization is projected to further rise by 312%, elevating India to the fourth rank by 2030 (Van Boeckel et al., 2015). FAO has already reported some of the peak resistance rates among common commensal and environmental bacteria against antibiotics like penicillin, cephalosporins, erythromycin and co-trimazoles in India (INFAAR). Resistance to the broad spectrum fluoroquinolones and third generation cephalosporin antibiotics is already more than 70% in Escherichia coli, Klebsiella pneumoniae, and, Acinetobacter baumannii and more than 50% in Pseudomonas aeruginosa. Even the microbial resistance to carbapenems, the last-resort antibiotics, is extremely high among gram-negative microorganisms (Sumanth et al., 2016). To overcome these incidences, it is impulsive to understand the microbe and drug interaction in host system.

Introduction

The study of infectious diseases, alongside other clinical and natural strengths, is going through fast change introduced on by the approach of reasonable next generation sequencing (NGS) advancements (Koser et al., 2012).These technologies become adopted as routine methods easily, and laboratory techniques are transformed throughout the process. The amount of genomic bacterial data generated is huge. As of this writing, for example, over 200,000 Salmonella genomes alone are in the public domain with hundreds being added weekly. A whole genomic DNA sequence characterizes the maximum practicable level of structural detail on the individuating traits of an microbial organism or population. It can therefore be used to provide more accurate microbial genome recognition, evolutionary relations and a detailed catalogue of epidemiologically important characteristics. This has a significant role to play for outbreak research, diagnosis and treatment of infectious diseases, microbiology and epidemiology (Allard et al., 2019). Besides, DNA sequences are also a universal dataset which can theoretically be used to determine any biological function. This means that antimicrobial resistant (AMR) can be detected and to track the development and spread of AMR bacteria in a hospital or the community.

Introduction

Until the 20thcentury, bacterial infections were the number one cause of human death. In the 1940s, antibiotics became the lifelines for treating serious infections that saved millions of lives. Despite the advantages derived from the use of antibiotics, resistance to these drugs emerged. Unfortunately, resistance to most antibiotics emerged in time-course even for the most powerful antibiotics developed.Resistance to antibiotics is a challenge to the scientific community in keeping up the pace of antibiotic discovery.World Economic Forum’s Global Risks Reporthas listed antibiotic resistance as one of the greatest threats (WEF, 2020).United Nations (UN) has warned that drug-resistant diseases could account for 10 million deaths each year by 2050. By 2030, antimicrobial resistance (AMR) could likelylead up to 24 million people into extreme poverty(IACG,2019). Currently, at least 7,00,000 people die each year due to drug-resistant diseases (IACG,2019). The emergence and widespread of antibiotic-resistant organisms is a global challenge and needs a multi-dimensional approach to address the fundamental issues to minimize drug-resistance.The Antibiotic Resistance Genes Database (ARDB) contains a repository of around 23,000 antibiotic resistance gene or protein sequences conferring resistance against over 240 antibiotics (ARDB). Computational Biology (Bioinformatics) has become one of the important tools in studying antimicrobial resistance. It employs a range of computation techniques to make predictions about the biological phenomenon.

Introduction

Resistance to antibiotics in the bacterial world is a genetically determined phenomenon that was first observed within a few years after the introduction of antibiotics for therapeutic purposes. In Nature, ARGs are believed to have originated about three billion years ago as an intrinsic defense mechanism in antibiotic-producing bacteria (Perry et al., 2016; Iskandar et al., 2022), and subsequently, evolved by complex and quantum jump trajectories to mobilize, diversify and interconnect all ecosystems in the micro-biosphere. Bacterial genomes exhibit plasticity and continue to evolve under the selection pressure of antibiotics and other stressors (Patel, 2016). The ARGs in circulation at present are thus a mixture of ancient and modern counterparts. Recently, the interplay between bacterial metabolism and antibiotic resistance has also been revealed (Zampieri, 2021).

The use of antimicrobials in health management, livestock, and food industry is driving the emergence of resistant microbes at a rapid pace (Wall et al., 2016; Adegoke et al., 2016; Hu et al., 2017). As a result, AR pathogens have spread globally to cause loss of lives, production, productivity, and negatively impact economic growth. Recently, AR pathogens in a large study have been found directly responsible for millions of deaths, and the figures are likely to increase in the coming decades (Murray et al., 2022). The pace of the emergence of AR can however be slowed down by improving human interventions. The global and local AR combat plans implemented during the past few years have enhanced the mass awareness of the pandemic problem (WHO, 2015; FAO, 2016; NAP, GOI, 2017; WHO, 2022; WOAH, 2022). These Action Plans on AR containment encourage avoiding of indiscriminate use of antibiotics for the treatment of sick humans and animals, recommend the ban on the use of antibiotics as growth promoters in food animals, promote AR surveillance and monitoring, and support the ‘One-Health’ strategy and application of the biosecurity principles for containment of the resistant microbes.

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.

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).

The response of any living cell to a xenobiotic (including antibacterial agent) is significantly affected by the prevailing local environmental conditions. In case of bacterial infection, it may include the presence of other microbes, no matter commensal or pathogenic, or biochemical alterations at that particular site of bacterial infection in host. All these host and bacterial factors play a critical role in the evolutionary responses of bacteria but usually go unconsidered while determining the microbial sensitivity to a particular antibiotic in a clinical setting. These also include the direct and indirect interspecies interactions that can affect the responses of individual bacterial species and surviving microbial communities to a particular antibiotic therapy. These interactions may significantly alter the qualitative and quantitative responses, potential phenotypic changes and the final trajectory of evolution of antimicrobial resistance. Due acknowledgement and plausible interpretation to these interactions may help to reduce the upsurge in AMR and its associated morbidity and fatality and pave path to more proficient clinical success.

The factors participating in drug- microbe interaction can be classified into two broad groups: host factors and microbial factors. The environmental milieu of the interaction is chiefly regulated by the gut microbiota with stressors causing deflections. The gastrointestinal bacteria amend host mental and physiological cadence via microbial metabolites such as butyrate, polyphenolic antioxidants, vitamins, and amines, affecting the overall ready perception and active rebuke to a stimulus. Lifestyle stressors such as fluctuating sleep and erratic eating patterns (western diet) disturb the host circadian system along with the gut microbiome. The overall impact is impaired detoxification (decreased conjugation of bile acids or increased production of hydrogen sulphide), substrate oxidation and energy regulation (reduced butyrate production) in the host affecting the overall health and productivity.

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.

Introduction

Antimicrobials, viz. antibiotics, antivirals, antifungals and anthelmintics are substances widely used to prevent and treat infections in humans, fish & prawn, livestock animals, and crops. In a situation where microorganisms such as bacteria, viruses, parasites or fungi become resistant to antimicrobial treatments to which they were previously susceptible, it is apprehended that antimicrobial resistance in the microbes has evolved. In fact, antibiotic resistant bacteria evade the effects of antibiotics and multiply in their own way leading to severe health consequences in human and animals. The menace is such that antimicrobial resistance (AMR) hinders treatment modalities targeted for infectious diseases. The hazards of antimicrobial resistance is frequently observed in various pathogenic and non-pathogenic organisms, viz. Salmonella spp., Campylobacter spp., Yersinia sp., E. coli serotype O157:H7, enterococci, Pasteurella sp., Actinobacillus sp. with varying severity (McEwen and Fedorka-Cray, 2002). Microorganisms that develop resistance to commonly used antimicrobials are referred to as superbugs. Globally it is a matter of great concern that it spreads rapidly and threatening the ability to treat common infectious diseases, resulting in prolonged illness, disability and death. Without any effective antimicrobial agent, treatment of infectious diseases, many medical procedures like cancer therapy, diabetes management, organ transplantation, and major surgery will not be possible. The microbes showing resistance to multiple antimicrobials are commonly known as multidrug resistance (MDR) organisms (Magiorakos et al., 2012).

Introduction

Today, antimicrobial resistance (AMR) has become a global issue, contributing to rise in mortality and cost of treatment. Antimicrobial resistance (AMR) is the resistance of a microorganism against an antimicrobial agent to which it was originally sensitive (WHO, 2013). Dispersion of antibiotics is done at large scale, in India even sometimes without any proper prescription. Animal Husbandry is not untouched and antibiotics are used to a great extent, which needs enforcement of strict government rules and regulations for their use in f ield conditions (PHFI, 2011).

Extended-spectrum β-lactamases (ESBLs) are enzymes that hydrolyse most of the β-lactam antibiotics and thus mediate resistance to penicillins, 3rd and 4th generation cephalosporins (Saravanan et al., 2018). Beta-lactamase inhibitors, inhibit these ESBLs such as clavulanic acid, sulbactam and tazobactam (Trott, 2013). The genes encoding for these enzymes are commonly found both on the chromosomes and plasmids among Enterobacteriaceae family (Kotsoana et al., 2019).

Among Enterobacteriacae, E. coli and Klebsiella species are main environmental pathogens associated with various illnesses such as acute bovine mastitis (Koovapra, 2015), pathogenic E. coli with diarrhoea and extra intestinal infections in humans and animals (Levine et al., 1987), K. pneumoniae with respiratory and urinary tract as well as bloodstream infections (Daehre et al., 2018). Resistance to extended-spectrum β-lactams occurs mainly in members of Enterobacteriaceae family through plasmids encoded ESBL genes. (Sharma et al., 2010).

Introduction

The antibiotics are low molecular weight natural, semi-synthetic, or synthetic substance that kills or slows the growth of germs while causing little or no harm to the host is considered an antimicrobial (Walsh, 2000). Antibiotics are the type of medicine produced by some microorganisms that kill or slow the growth of other microorganisms. It includes any of a large group of chemical substances, as penicillin or streptomycin, produced by various microorganisms and fungi, having the capacity in dilute solutions to inhibit the growth of or to destroy bacteria and other microorganisms, used chiefly in the treatment of infectious diseases (De Vetten et al., 2003). However today, a large number of bacteria are resistant to various antibiotics. Antibiotic resistance occurs when a bacterium becomes able to survive and multiply despite the presence of an antibiotic that would normally control or kill it. In other words, the bacteria become “resistant” and continue to grow even at therapeutic levels of the antibiotic. While antibiotic resistance is a natural phenomenon, it is accelerated by the selective pressure exerted when an antibiotic is used. Bacteria that are naturally resistant to the antibiotic have a better chance of survival compared to those that are susceptible. As the susceptible bacteria are killed or inhibited, resistant strains are favored and continue to thrive (Holme et al., 2006). Some resistance arises without human influence, as bacteria can naturally produce antibiotics against other microorganisms, resulting in a low-level, natural selection for antibiotic resistance. However, the current surge in antibiotic resistant bacteria is largely attributed to the overuse and misuse of antibiotics. One of the main issues facing public health is antimicrobial resistance, particularly in developing nations where access to and use of medications are easier than in developed nations, leading to a disproportionately higher rate of inappropriate antibiotic use and higher resistance levels (Ganguly et al., 2011). Antimicrobial resistance will make it more difficult to control infections in the community and make health care services less effective (Bhatia et al., 2010). It is well known that India has one of the highest rates of infectious disease burdens in the world, and that the country’s high rates of malnutrition and inadequate sanitation make these circumstances worse (http://www. resistancestrategies.org/wpcontent/uploads/2011/03/Indiareport).

Antimicrobial Resistance abbreviated as AMR is defined as the process by which microorganisms like bacteria, fungi, viruses, and parasites change over a period and become resistant to a drug to which they were earlier susceptible to. This makes the medicines ineffective for treatment overtime, thus making infections difficult to treat thereby increasing the risk of spread of diseases caused by them and severe illness which may lead to death. This in turn necessitates the need for development of newer antibiotics. However, there has been a sharp decline in the development and manufacture of new antibiotics in the recent years resulting in development of only few antibiotics in the past two decades. The increase in emergence of antibiotic resistance concurrent with decline in development of newer antibiotics poses a serious threat to health of both humans and animals. Every year about 700,000 deaths have been reported to occur due to resistant infections and this rate is likely to increase further with estimated infections due to resistant pathogens expected to reach over 50 million globally by 2050 (O’Neill, 2016). The worrisome trends of increasing antimicrobial resistance across the globe poses a serious threat to both animal and human health as AMR not only increases the morbidity and mortality rates but also increases the economic burden on health care (Zhu et al., 2022). This presents an extremely grave situation as the supply of new antibiotics is inadequate to cope with the increase of AMR pathogens. The unnecessary use of antibiotics globally especially in the livestock sector as growth promoter further helps in selective enrichment of AMR pathogens which further increases health risks. It is well known that resistant pathogenic and commensal bacteria can persist and spread within and between premises despite zero antimicrobial drug use. This may be explained due to maintenance, replication, and further dissemination of resistance genes by horizontal transfer of genes via natural ecosystem and can occur either by conjugation, transformation and transduction; and these mechanisms can rapidly spread AMR genes among microbial community in natural environment and human microbiome, where these resistant bacterial species may be selected due to presence of antimicrobial drugs (Zhu et al., 2022). The physical transfer of bacteria via movement of animals, workers, and equipment and ineffective cleaning and disinfection may also be responsible for increasing the AMR pathogens in the natural environments.

AMR has been prevalent in almost all European countries from past decades. A 2021-2022 report published by the European Food Safety Authority (EFSA) and the European Centre for Disease Prevention and Control (ECDC) on AMR in zoonotic and indicator bacteria from humans, animals and food origin reported vast differences in antimicrobials resistance levels among the European countries, with comparatively higher AMR percentages in the southern and eastern parts of Europe (ECDC, 2022, 2023). The resistance levels ranged from very high for tetracyclines, sulfonamides and ampicillin, moderate for ciprofloxacin to low levels for third-generation cephalosporins (cefotaxime and ceftazidime) in human cases infected with Salmonella spp. in 2022 (ECDC, 2023). The trends are dynamic and over a decade of 2013 2022, the human isolates from 12 and 15 countries reported a statistically significant reduction in tetracycline and ampicillin resistance, particularly for S. typhimurium (ECDC, 2023). The ciprofloxacin and cefotaxime combination, a highest priority critically important antimicrobials (CIA), had very low and rare resistance levels for Salmonella isolates of humans and animal and their derived meat.

Livestock species wise variation was also observed in the resistance pattern in imported meat, being higher in poultry meat isolates compared to pig or cattle meat for Escherichia coli (ECDC, 2023). High to extremely high levels of f luoroquinolone resistance from Campylobacter jejuni and C. coli of both human and animal origin isolates exist, due to which the fluoroquinolones can no more be used as a treatment option for human Campylobacter infections (ECDC 2023). The combined occurrence of erythromycin resistance and ciprofloxacin in Campylobacter spp is considered of high public health importance, and the report mentioned that the combined resistance for these antimicrobials was higher in C. coli compared to C. jejuni from food-producing animals and humans. However, while the ciprofloxacin resistance showed an increase in over 13 study countries, seven countries reported reduced erythromycin resistance for C. jejuni. Overall, resistance to conventional antimicrobials was frequent in Campylobacter and Salmonella from humans and animals. Increasing trend in resistance to one of two CIA for treatment in humans has been observed in poultry-originated Salmonella serovars and Campylobacter while combined resistance was is low in few Salmonella serovars and C. coli isolates from humans and animals in some countries (ECDC, 2023).