Antimicrobial drug use and its implications

The term antibiotic, meaning “substance against life," thus “Antibiotic resistance means life”.

Antibiotic therapy, if indiscriminately used, may turn out to be a medicinal flood that temporarily cleans and heals, but ultimately destroys life itself (Felix Marti-Ibanez, 1955)

Bhoj R SinghSection of Epidemiology, CADRAD, IVRI, Izatnagar

Bacteria

Most numerous denizens of agriculture, with populations ranging from 100 million to 3 billion in a gram of soil.

One bacterium is capable of producing 16 million more in just 24 hours.

Bacteria perform chemical transformations, including degradation, disease suppression, disease, and nutrient transformations.

Cloud formation in Troposphere (NASA’s Genesis and Rapid Intensification Processes experiment).

Generation of electricity: Using microbial fuel cell Bacillus stratosphericus generates twice as much electricity as other bacteria.

Bacteria

I always thought the most significant thing that we ever found on the whole goddamn Moon was that little bacteria who came back and lived and nobody ever said shit about it. — Pete Conrad 

On April 20, 1967, unmanned lunar lander Surveyor 3, left a camera on Moon. Two-and-a-half years later, on November 20, 1969, Apollo 12 astronauts Pete Conrad and Alan L. Bean recovered the camera. NASA scientists examined it back and find still alive Streptococcus mitis, survived for 31 months in the vacuum of the moon's atmosphere.

Zero gravity: “Generally speaking, bacteria tend to grow better in space,” Klaus (2004) said. “The first and most fundamental b

Bacteria found in rocks taken from the cliffs at Beer have survived a grueling year-and-a-half exposure to space conditions on the exterior of the ISS and returned home alive, becoming the longest-lived photosynthesizing microbes to survive in space.acterial response to spaceflight is a shortened lag phase.” The bacteria sent was: OU-20 resembling the cyanobacteria genus Gloeocapsa

Bacteria in our body

We are 90% bacteria and only 10% human(approximately 1014 versus 1013). Sears, 2005.

500 to 1000 species of bacteria live in the human gut and a roughly similar number on the skin. Science, 2008; Grice et al., 2009.

The human gut alone contains on average: 40,000 bacterial species 9 million unique bacterial genes and 100 trillion microbial cells and only 1% of them are characterized (Yang et al., 2009).

332,000 genetically distinct bacteria belonging to 4,742 different species were detected on hands of students using molecular techniques. Fierer et al., 2008.

The mass of microorganisms are estimated to account for 1-3% total body mass. 1.0 to 2.26 kilograms of live bacteria inside our bodies (Berg, 1996; Raymond, 2012).

Bacterial makeup of our body change little over time (National Human Genome Research Institute in Bethesda, Maryland).

Infections increase from overuse of antibiotics (Human Microbiome Project; HMP). They help in development of our immune system and how we fight off pathogens, they

influence our metabolism, our odor and even our behavior. Eisen, 2012.

How we get them?

Any two hands – even belonging to the same person – had only 13% of their bacterial species in common (Fierer et. al., 2008). It is due to the influence of sex, handedness, and washing habits but relatively unrelated with type of food intake.

We get them from: From mothers while come to this world. From Air, Food, Water, Environment and friends and family. How we can get healthy bacteria?

From healthy donors From excrements of the healthy persons Poop transplant (the best way to fight enteritis necroticans, Cl.

difficile)

Where bacteria are absent?

Where there is no life? NASA’s Clean room: 100 types of bacteria, about 45 percent of

which were previously unknown to scientists. Two miles below the surface of a Greenland glacier – “ultrasmall

bacteria” in a glacial core – a habitat which is low-temperature, high-pressure, reduced-oxygen, and nutrient-poor. The core was estimated to be 120,000 ye

Deepest layer of the earth's crust –the Earth's oceanic crust has revealed a new ecosystem living over a kilometer beneath our feet.

In tissue sites once deemed sterile – The Relman Lab at Stanford used real time PCR to target conserved regions of the bacterial 16S ribosomal DNA (rDNA). They concluded that there is a substantial and “normal” population of bacterial DNA sequences in the blood of even healthy individuals.

Why bacteria persist so diversely?

Need very few genes to persist –Encephalitozoon cuniculi (a pathogen of rabbits) has 2.9 Mbp, encoding approximately 2,000 densely packed genes. Genome Encephalitozoon intestinalis, at 2.3 Mbp, a severely compacted genome.

Can take and give genes rapidly. Can survive anywhere at any temperature

(even in boiling springs), eat anything, breath any air.

AMDR is Natural?

Call of the wild: antibiotic resistance genes in natural environments (Allen et al., 2010).

Antibiotic resistance genes are detected in ‘antibiotic-naïve’ strains isolated in pre-antibiotic era (before 1930).

24% strains of Murray collection (1917-1952) are resistant to ampicillin and tetracycline.

Bacteria that may grow on antibiotics as the sole carbon and nitrogen sources include: Pseudomonas fluorescens (on streptomycin and penicillin); Burkholderia cepacia (on Penicillin); Flavobacterium spp., Streptomyces venezuelae (on chloramphenicol), actinomycetes cultured from forest (on telithromycin)

Use of anti-microbials in Nature

Antibiotics have been produced for over 500 million years, dating back to the Cambrian period and the emergence of vertebrate fish. Antibiotic-like molecules, are likely to be even older than this.

The non-protein amino acids that are found as components of peptide antibiotics have been detected in meteorites and other primordial sources (Davies, 2009).

The fungus-growing ant system, in which ants carry an antibiotic-producing actinomycete (a Pseudonocardia sp.) on their cuticle and use this bacterium specifically for biocontrol of the fungal garden parasite, Escovopsis sp. (Currie et al., 1999; Cafaro and Currie, 2005)

Biocontrol of the causative agent of potato scab, Streptomyces scabies str. RB4, by the antibiotic-producing suppressive strain, Streptomyces diastatochromogenes str. (Neeno-Eckwall et al., 2001)

Some autoinducers used in quorum sensing have antibiotic activity (Kravchenko et al., 2008).

Bacteria as natural pesticides in the biological pest control (Bacillus thuringiensis) (Chattopadhyay et al., 2004)

Antimicrobials used by human

We hardly know about 1% of antibacterial molecules in nature. Most of these molecules cannot be detected, and the few that have been identified some of them are the most used antibiotics (Baltz, 2008).

Of the known antibiotics, < 1% of antimicrobial agents have medical value (Marino, 2007).

Antibiosis was first described in 1877 in bacteria when Louis Pasteur and Robert Koch observed that an airborne bacillus could inhibit the growth of Bacillus anthracis.

Salvarsan Discovered by Paul Ehrlich was produced in 1932 at the Bayer Laboratories and was used for treatment of syphilis.

Alexander Fleming: Discovered penicillin in 1928, could not purify. In 1939, gramicidin from B. brevis. was one of the first commercially

manufactured antibiotics (Sykes, 2001). Ernst Chain and Howard Florey successfully purified penicillin, and in

1941 tested on human subjects. The term antibiotic was coined by Selman Waksman in 1942.

Use of Antimicrobials for Animals

Antibiotics are used in food-producing animals for three major reasons (CDC, 2009). A. in high doses for short periods to treat sick animals. B. in high doses for short periods to prevent diseases (after

weaning, or during transport). This use “usually involves treating a whole herd or flock, which increases the likelihood of selecting for organisms that are resistant to the antibiotic.”

C. antibiotics are commonly given in the feed at low doses for long periods to promote the growth of cattle, poultry, and swine (How? No one knows exactly, 1-15% Improvement in production, poorer the hygiene better the effect).

A=17%, B+C=83% (About ten million Kg alone in USA, i.e., 70% of total antibiotic production): AHI, 2009, Union of Concerned Scientists, 2009.

A+B+C=87%, 40% of it is added in to feed (APHA, 2009)

Use of Antimicrobials----Conti--

By the mid-1990s the EU had authorised 9 antibiotics, plus the antibacterials carbadox and olaquindox, for use in animal feed as ‘growth promoters’ and preventive antibiotic use had become a routine aspect of intensive farming.

FDA authorized the use of 18 antibiotics for ‘growth promotion’, of which 8 were identical or chemically similar to drugs used in human medicine

By 1995 around 90% of poultry production units used feed containing antibiotics.

1999 in the US, 90% of the diets of recently weaned piglets, 70% of the diets of growing pigs and 50% of the diets of ‘finishing’ pigs contained some form of antibiotic.

DANMAP (Denmark) estimated that in 1997, 80% of total antibiotics produced were used in animals.

In 2001, the Union of Concerned Scientists in the US estimated that around 70% of all US antibiotic usage were in animals.

November 2011 in US, 8 times more antimicrobials are used for non-therapeutic purposes in the three major livestock sectors [i.e., chickens, pigs and cattle than are used in human medicine.

Use of Antimicrobials----Conti--

Fifty million pounds of antibiotics are produced in the U.S. each year. Twenty million pounds are given to animals, of which 80% (16 million pounds) is used on livestock merely to promote more rapid growth.. American Medical News, "FDA Pledges to Fight Overuse of Antibiotics in Animals", February 15, 1999.

FDA (2010) reported use of antibiotics totals for food-producing animals in 2009 were 13,068 tonnes of antibiotics for domestic use and a further 1,632 tonnes exported.

Who, 2011. In several parts of the world, more than 50% in tonnage of all antimicrobial production is used in food-producing animals. In addition.

Veterinarians in some countries earn at least 40% of their income from the sale of drugs, creating a strong disincentive to limit their use.

In Australia, 500,000kg antibiotics are used in animals and 300,000kg in humans per year (JETACAR 1999).

In 1992, over 120,000kg of avoparcin (10% active ingredient by weight) was used in animals in Australia (predominantly as a growth promoter), while only 68kg of vancomycin was used in people (JETACAR 1999).

From: Edquist and Pederson, 2000

Legalities of Antibiotic use in animals

Penicillin Act and the Therapeutic Substances (Prevention of Misuse) Acts in the UK, FDA in USA and similar acts in most of the developed countries restricted the use of antibiotics to therapeutic use on prescription by a doctor, veterinarian or dentist. (1947).

1960s scientists discovered that antibiotic resistance could be transferred from one bacterial species to another. In the UK, the Netherthorpe Committee in 1962, the Swann Committee in 1969 and the Lamming Committee in 1992 sounded the alarm.

In Denmark, The Feedingstuffs Act was passed by parliament in November 1985 and came into force in January 1986. Antibitotic consumtipn got reduced by 60% in 1996. Ban was full effective in 1998.

In 1997 the World Health Organization recommended that the use of any antibiotic for ‘growth promotion’ in animals should be terminated if that antibiotic is used for human medicine or if its use in animals increases resistance to other antibiotics used in human medicine.

Between 1999 and January 1st 2006 the EU banned the use of 8 antibiotics for ‘growth promotion’: virginiamycin, tylosin phosphate, bacitracin zinc, spiramycin, avilamycin, flavophospholipol, monensin and salinomycin, plus the drugs carbadox and olaquindox.

2000, WHO recommended total ban on feed antibiotics used as growth promoter.

In 2010, in USA, FDA limited the use of antibiotics for growth promotion.

Trends in Tylosin use in growth promotion and Erythromycin Resistance in Enterococcus faecalis isolated from pigs.

(Aarestrup et al al., 2001, WHO, 2003)

Antibiotic in feed had effect on production: A MythDenmark Experience, totally stopped antibiotics in feed in 1998

After ban, poultry production was unaffected other than a one percent increase in feed intake (there were no effects on weight gain or mortality). In finisher pigs there were also no important detrimental effects.

No detrimental effects on overall pork production which continued to rise. (WHO, 2003)

Effect of Antibiotic use on Salmon production (Source: Norvegian directorate of Fisheries)

Why AMDR is essential?

For survival of Bacteria. It is Natural.

If there was no antibiotic resistance, there would be no need for a microbiology lab!

Infections could be treated “syndromically”. It actually stimulate innovation in drug

discovery (Outterson, 2010).

Survival of Bacteria in Nature

The ancestors of modern bacteria appeared on Earth, about 4 billion years ago. For about 3 billion years, all organisms were microscopic, and bacteria and archaea were the dominant forms of life.

Bacteria are present in most habitats on Earth, growing in soil, acidic hot springs, radioactive waste, water, and deep in the Earth's crust, as well as in organic matter and the live bodies of plants and animals, providing outstanding examples of mutualism in the digestive tracts of humans, termites and cockroaches (Fredrickson et al., 2004).

There are typically 40 million bacterial cells in a gram of soil and a million bacterial cells in a millilitre of fresh water; in all, there are approximatelyfive nonillion (5×1030) bacteria on Earth (Whitman et al., 1998)].

Bacteria form a biomass that exceeds that of all plants and animals (Hogan, 2010). Bacteria are vital in: recycling nutrients, fixation of nitrogen. Provide the nutrients needed to sustain life by converting

dissolved compounds such as hydrogen sulphide and methane.

Survival of Bacteria in Nature

Most bacteria have not been characterized, and only about half of the phyla of bacteria have species that can be grown in the laboratory (Rappé and Giovannoni, 2003)

Eukaryotes resulted from ancient bacteria entering into endosymbiotic associations with the ancestors of eukaryotic cells, which were themselves possibly related to the Archaea.

Due to their small size, commensal bacteria are ubiquitous and grow on animals and plants exactly as they will grow on any other surface.

They degrade a variety of organic compounds thus used in waste processing and bioremediation of industrial wastes. Bacteria capable of digesting the hydrocarbons in petroleum are often used to clean up oil spills.

Bacteria a highly social organisms with an uncanny ability to protect themselves, as a group (Wassenaar, 2011).

Secrets of survival

Altruism or selflessness is the principle or practice of concern for the welfare of others and Bacteria practice it. A traditional virtue in many cultures.

Lee et al (2010) observed that most individual bacteria within an antibiotic-resistant population can be significantly more sensitive to the antibiotic than the global population, then how they survive in presence of antibiotic.

In a 2010 Collins et al., periodically analyzed the levels of drug resistance in an E. coli the colony, they saw something unexpected: although the entire population was thriving in the presence of the drug, only a few individual bacteria were actually resistant. Further analysis revealed that the resistant mutants were secreting a molecule called indole that thwarts their own growth but helps the rest of the population to survive by activating drug-export pumps on the bacterial cell membranes.

Tanouchi et al (2012) while studying programmed death in bacterial cells they reported an altruistic trait, whereby some cells trigger the cell death program and release stress-relieving substances that increase the chances of survival of other cells within the population.

Why Survival of Bacteria Important for life?

The bacteria is: First and the last in the food-web. Life forms perform as their microbes evolved.

Ruminant digest fibrous food, Rumen and caecum: the fermenters Termites digest lignin

Over 1,000 bacterial species in the normal human gut flora of the intestines and contribute to: gut immunity, synthesize vitamins such as folic acid, vitamin K and biotin convert sugars to lactic acid fermenting complex indigestible carbohydrates inhibit the growth of potentially pathogenic bacteria (usually

through competitive exclusion) Life is not possible without bacteria (L. Pasteur, 1885). Nencki (1986) disagreed with Pasteur's thought, gnotobiotics. Gnotobiotics in not natural. Life without bacteria if even possible is worse (Wassenaar, 2011). Bacteria: The Benign, the Bad, and the Beautiful. (Wassenaar,

2011).

Life within life

There are approximately ten times as many bacterial cells in the human flora as there are human cells in the body, with large numbers of bacteria on the skin and as gut flora (Sears, 2005).

In human body, the Living mass of enteric flora accounts for 1 kg to 2 kg of body weight.

It represents a combined genetic microbiome exceeding the human genome by one hundred-fold.

Magnitude and complexity of the alimentary ecosystem is ill understood.

It seems surprising that the rediscovery of the importance of the gut flora is only a latter-day occurrence.

It exhibits a collective metabolic activity that eclipses that of the liver. The gut flora is tantamount to a hidden organ.

The vast majority of the bacteria in the body are rendered harmless by the protective effects of the immune system.

A few are beneficial and still fewer are pathogenic and cause infectious diseases (WHO, 2002).

Lesson from germfree life (Shanahan, 2004)

Life without bacteria is associated with reductions in: mucosal cell turnover, digestive enzyme activity, cytokine

production, lymphoid tissue, lamina propria cellularity, vascularity, muscle wall thickness and motility.

Absence of bacteria is associated with an increase in enterochromaffin cell area, larger GIT. caloric intake to sustain a normal body weight.

Thus, the normal host–flora relationship is not one in which the host is in nutritional deficit because of the presence of bacterial guests within the gut.

Important metabolic effects of the flora include: the production of short chain fatty acids—a major energy source

for colonic epithelia—from dietary fermentable carbohydrates. Breakdown dietary carcinogens. synthesis of biotin, folate, and K vitamins.

Understanding of the activity of the intestinal microbiome, may offer strong potential for therapeutic exploitation in health and disease.

Lesson from germfree life

The fertility of germfree animals is very low, but the reasons for this are not known (Levenson et al., 1959).

Antibodies can be produced by the germfree animal, but seemingly at a slower rate (Miyakawa et al., 1957).

The presence of a commensal flora is necessary for fine tuning of T cell repertoires and TH1/TH2 cytokine profiles (Rook and Stanford, 1998).

The germfree animal, generally is highly susceptible to pathogenic (and at times to ordinarily nonpathogenic) bacteria (Levenson et al., 1959).

About 70% germfree animals never reach the adulthood. Almost 30 per cent of germfree animals have spontaneous deaths of due to volvulus of the enlarged caecum. Forty per cent dies due to asphyxia (Wardcet al., 1958).

Bad Effects of Antibiotic use

Change in our food (antibiotic load) may alter the microbial education and fine tuning of the immune system.

With increase in use of antibiotics the following disorders have shown an increasing trend (Bach, 2002; Ekbom, 2003; Anthony Gucciardi , 2011):

Asthma, Autoimmune disorders and allergies (Penicillins, Carbapenems, cefalosporins and Sulpha drugs),

Mutagenicity and carcinogenicity (Sulphamethazine, Oxytetracycline, Furazolidone) Nephrotoxicity (Aminoglycosides, Penicillins, sulphonamides) Bone-marrow toxicity (Chloramphenicol, Tetracyclines, Linezolid, Sulphonamides) Hepatotoxicity (Macroloides, Telithromycin, Penicillins) Brain and Nerve damage (Carbapenems, penicillins, polypeptides, quinolones) Hearing impairment –Ototoxicity (Aminoglycosisdes) Visual disturbances (Telithromycin) Foetotoxic (Tetracyclines) Tendinosis (Quinolones) Photosensitivity (Sulphonamides, tetracyclines) Change in Taste (metallic by metronidazole, bitter by tinidazole) Multiple sclerosis Soaring obesity rates around the globe Increased cases of mental illness Insulin-dependent diabetes mellitus. Autoimmune disorders Obesity

Autoimmune disorder: About 75 percent of humans host a microorganism called Bacteroides fragilis which helps hold the immune system in check by producing a protein that restrains T-cells. When T-cells get too high, it can lead to inflammatory and autoimmune diseases.

Obesity: Human stomach contains bacteria called Helicobacter pylori. This bacterium regulates the amount of stomach acid that the stomach produces. Acidity inturn stimulate ghrelin, secretion which signals to the brain that the stomach is empty and the body needs to eat. When after eating acidity is low ‘leptin’ is secreted and brain sense that now it is to stop eating. Eradication of Helicobacter pylori Increases Ghrelin mRNA expression in the Gastric Mucosa and decrease in ‘leptin’ mRNA expression. Lack of ghrelin and leptin regulation after Helicobacter pylori eradication (after antibiotic use) may lead to over-eating and obesity (Roper et al., 2008).

Unwanted effects of Antibiotics(Blaser, 2011)

Reproductive disorders: Dr. Wassenaar (2011) explains how the antibiotic use alters intestinal bacterial microflora of a fruit fly which in-turn drives mating preference. If this applies to humans!

Mating preference was achieved by dividing a population of Drosophila melanogaster and rearing one part on a molasses medium and the other on a starch medium. When the isolated populations were mixed, “molasses flies” preferred to mate with other molasses flies and “starch flies” preferred to mate with other starch flies. The mating preference appeared after only one generation and was maintained for at least 37 generations. Antibiotic treatment abolished mating preference, suggesting that the fly microbiota was responsible for the phenomenon (Sharon et al., 2010).

Gastro-intestinal problems IBD (Krohn’s disease) Peptic ulcers Antibiotic Associated Pseudo-Membranous Colitis (AAPMC)

due to Clostridium difficile. Antibiotic-associated diarrhea (although all antibiotics may

be associated with AAD, cephalosporins, extended coverage penicillins, and clindamycin are the main)

Superinfection (or Suprainfection): Candida albicans vaginal infections Suppression of Normal Microbiota

Estimates of Adverse effects of Antibiotic chemotherapy

It is estimated that 10% to 15% of all hospitalized patients treated with antibiotics will develop AAD.

Most important, twice as many will become asymptomatic carriers. Risk factors include: compromised immune status advanced age, abdominal surgery Comorbidity types and length of use of antibiotics, and the length of hospitalization (infection rates for C.

difficile are reported to be around 10% after 2 weeks of hospitalization but may reach 50% after 4 or more weeks

Conclusion

Many of the emerging health problems are associated with change/ destruction of our microbiota.

Bacteria are necessary for healthy life on earth thus the antibiotic resistance.

Antibiotic is to destroy the life while ADR is for life.

Most life on earth perform due to life within (bacteria).