Antibiotic resistance of bacteria: A global challenge

177RESONANCE February 2012

GENERAL ARTICLE

Antibiotic Resistance of Bacteria: AGlobal Challenge

Saswati Sengupta and Madhab K Chattopadhyay

KeywordsAntibiotic-resistance, bacteria,genetic basis, biochemical ba-sis, antibiotic paradox, multidrug-resistant.

Bacterial resistance to antibiotics poses a serious challenge tothe prospect of chemotherapy. Rational use of antibiotics ismost desirable but it cannot provide a permanent solution tothe problem. In this article the biochemical and genetic basisof antibiotic resistance in bacteria is discussed with examples.The non-clinical aspects of antibiotic-resistance are also dealtwith in brief.

1. The Wonder Drugs

Antibiotics constitute one of the most significant contributions ofmodern science. The discovery of these life-saving drugs trans-formed the health-care scene during the last century. A signifi-cant decline in the fatality rate of many diseases was noticed afterthe introduction of antibiotics into clinical practice. For example,20 to 85% death rate due to pneumonia in theUS during the 1930scame down to about 5% in the 1960s, 100% death due to chronicinfection of a heart valve was reduced to 5%, 20 to 90% fatalitydue to epidemic of spinal meningitis infection caused by thebacterium Neisseria meningitides, was decreased to almost 2%.Antibiotics help not only in curing the diseases but also in theirprevention. Penicillin, for instance, prevents throat infectioncaused by Streptococcus and thereby prevents recurrences ofrheumatic fever in susceptible individuals. Onset of a sexuallytransmitted disease may be prevented by the timely use of theproper antibiotic. By eliminating the causative organism of diph-theria from carriers (people harboring the organism withoutfalling sick) using an antibiotic, the chances of dissemination ofinfection could be effectively reduced.

2.What They Are

In 1942, Selman Waksman defined antibiotics as low molecular

1Saswati Sengupta is areserach associate at theIndian Institute ofChemical Technology(CSIR). Her areas ofinterest are therapeuticproteins and intellectual

property rights.

2Madhab K Chattopadhyayis a scientist at the Centrefor Cellular and MolecularBiology (CSIR). His areasof interest are stress

adaptation of bacteria andantibiotic resistance inmicroorganisms.

1 2

178 RESONANCE February 2012

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weight chemical substances produced by microorganisms, hav-ing growth-inhibitory effects on other microorganisms in highdilution. However in the present context, any chemical substanceeither of microbial, synthetic or semi-synthetic origin, used in theclinical treatment of infections, is called antibiotic. Thus strepto-mycin produced by fermentation is an antibiotic. The sulphadrugs and quinolones, produced by chemical synthesis, are anti-biotics. Ampicillin (semisynthetic penicillin), produced partly byfermentation and partly by chemical synthesis, is also an antibi-otic. The discussion in this article will be confined to the antibac-terial antibiotics.

The significance of antibiotics in nature remains a riddle. Theyare secondary metabolites and therefore are not indispensable forsustenance of life of the producer organism. The widely- believednotion about their role in preventing the growth of other organ-isms which thrive on the same source of food in natural environ-ments, is essentially anthropocentric and yet to be validated. Therole of antibiotics as signaling molecules in natural ecosystemshas been suggested by some scientists.

3. Historical Background

Though the use of microorganisms for the control of infectionswas in vogue even in ancient civilizations, the modern era ofantibiotics was ushered only at the beginning of the twentiethcentury. During the screening of hundreds of synthesized organicarsenical compounds for efficacy in the treatment of syphilis,salvarsan was discovered by Sahachiro Hata in 1909 in thelaboratory of Paul Ehrlich, the GermanNobel-laureate. Prontosil,the first sulfonamide drug, was developed by Gerhard Domagk in1932 for commercial use. The serendipitous discovery of penicil-lin by Alexander Fleming at St Mary’s Hospital (UK) in 1928 andthe subsequent isolation and purification of the compound byFlorey and Chain made a giant stride in the field of chemotherapy.Availability of such a compound with activities against a widerange of infectious organisms and (unlike sulfonamides) no lossof activity in the presence of biological materials such as pus,

Any chemicalsubstance either ofmicrobial, syntheticor semi-syntheticorigin, used in the

clinical treatment ofinfections, is called

antibiotic.

The role ofantibiotics as

signalingmoleculesin natural

ecosystems hasbeen suggested bysome scientists.


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provided a distinct advantage to the physicians in controllingbacterial infections. Discovery of streptomycin, the antitubercu-lar antibiotic, by Albert Schatz, a graduate student in the labora-tory of Selman Waksman (associated with the discovery of aseries of antibiotics in the 1940s and 1950s) at Rutgers University(USA) in 1943, led to the idea that all the infections could becontrolled whenever required. Major pharmaceutical companiesstarted diverting substantial portion of their funds, earmarked forresearch on anti-infectives, to the development of other drugs.

4. Emergence of Antibiotic-Resistant Bacteria

The euphoria did not last long. Within four years following theintroduction of penicillin during the Second World War, occur-rence of resistant strains was reported. According to an estimateby the The Centers for Disease Control and Prevention (USA),13,300 patients died of antibiotic-resistant bacterial infection inthe US during 1992. An incredible 150% increase in the occur-rence of drug-resistant Pneumococci was noted between 1987and 1994. A 20-fold increase in the frequency of hospital-ac-quired Enterococci, resistant to vancomycin, was seen between1989 and 1993. The frequency of methicillin-resistant Staphylo-coccus rose from 2% in 1975 to 32% in 1992. By this time,resistance to virtually all the therapeutically useful antibioticshad been evidenced. Emergence of methicillin-resistant Staphy-lococcus aureus (MRSA) and vancomycin-resistant Enterococci(VRE) have raised serious concern all over the world since thesetwo antibiotics were believed to be invincible when they werereleased in the market. A couple of years back, some Gram-negative bacteria (Escherichia coli, Klebsiella pneumoniae)werefound to produce an enzyme, which confers resistance to virtuallyall the commonly used antibiotics including carbapenems, one ofthe last resorts in the clinical management of infections caused bymultidrug-resistant organisms. The organisms were believed tobe originated from India. That is why the enzymewas named NewDelhi Metallo -lactamase (NDM-1). Some of the other mostimportant types of multiple drug-resistant organisms includeextended-spectrum beta-lactamase producers (which are resis-

Within four yearsfollowing theintroduction ofpenicillin during theSecondWorld War,occurrence ofresistant strains wasreported.


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tant to cephalosporins and monobactams) and penicillin-resistantStreptococcus pneumoniae. Studies on resistance in soil bacteriahave been shown that these organisms serve as a reservoir formultiple antibiotic resistances. A recent analysis of 264 soilisolates obtained from different natural habitats in and aroundHyderabad has identified 5 isolates resistant to as many as 10antibiotics. During the recent past, soil metagenomics also re-vealed several aminoglycoside resistances in nonculturable bac-teria. Notwithstanding the availability of so many antimicrobialagents, infectious diseases still remain the second leading causeof death worldwide. Eventually, the widespread occurrence ofantibiotic-resistant bacteria has added a new dimension to theemerging threat of bioterrorism.

5. Genetic and Biochemical Basis of Antibiotic Resistance

5.1 Genetic Basis:

5.1.1 Spontaneous Mutation and Gene Transfer: Antibiotic-tolerance in bacteria emerges as a result of error in DNA replica-tion, a phenomenon known as spontaneous mutation having afrequency of one in 107 cells. During the epidemic of Shigellainfections in Japan during 1950s, it was observed that bacteriacould transfer copies of antibiotic resistance genes to susceptiblebacteria thus making the latter antibiotic-tolerant. The mecha-nism predominantly used for this purpose is conjugation1, apowerful gene-mobilizing mechanism. The other known mecha-nisms of gene transfer among bacteria, e.g., transformation2 andtransduction3 also play some role in dissemination of antibioticresistance. Resistance thus acquired is transferred to the progenycells by a process called vertical gene transfer. Another process,called horizontal gene transfer4, enables bacteria to transfer cop-ies of antibiotic resistant genes even to the distantly relatedspecies in their neighborhood and thus contributes significantlyin dissemination of resistance. Different types of such resistantgenes, when accumulated in a single cell, result in multi-resistantphenotype.

1 A process of gene transfer inbacteria facilitated by direct up-take of exogenous genetic ma-terial (such as plasmid DNA)from a related strain in the sur-roundings. Transformation takesplace naturally in some speciesand can also be attained by arti-ficial means. This technique isbeing used very frequently in thefield of biotechnology for thepurpose of cloning.

2 Another process of bacterialgene transfer mediated by bac-teriophages which carry DNAfrom one cell to another. Thisprocess is known to occur inseveral bacterial species suchas Salmonella, Escherichia, Mi-crococcus etc.

3 The process which involvesthe transfer of genetic materialfrom one bacterial cell to an-other by direct cell-to-cell con-tact through a tube-like struc-ture known as pilus.

4 The movement of genetic ma-terial between unrelated speciesof bacteria. This can happenthrough direct incorporation offree DNA by bacterial cells. Thisdirect form of gene transfer, forinstance in the soil or in thedigestive tract of animals, is themost common mode for thetransfer of genetic material. Itfosters rapid dissemination ofantibiotic resistance in the bac-terial community. Horizontalgene transfer can happen eitherthrough transformation, trans-duction or conjugation.


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Mobile genetic elements, viz., transposons5 and integrons6 canalso transfer antibiotic resistance genes en block to susceptiblebacteria. Resistance genes mostly reside on bacterial plasmids.However, in some cases they are also found to be located onchromosomes and in a few cases organized in operons. It wasbelieved earlier that antibiotic-resistant strains were less compe-tent to survive in the environment and are slow-growers com-pared to their antibiotic-sensitive counterparts, but subsequentstudies have established that such disadvantages might be coun-terbalanced by additional compensatory mutations.

5.2 Biochemical Basis

5.2.1 Drug Inactivation by Microbial Enzymes: Among severalmechanisms involved in the development of antibiotic resistance,drug modification plays a significant role in rendering manytherapeutically useful drugs useless. For example -lactamase,an enzyme elaborated by many Gram-positive and some Gram-negative bacteria, converts penicillin into penicilloic acid, whichis therapeutically inactive. The production of the enzyme isinducible in Gram-positive bacteria whereas it is constitutive inGram-negative bacteria. This reaction also forms the basis ofchemical assay of the antibiotic since penicilloic acid can betitrated with iodine. The enzyme also provides a therapeutictarget in the management of infections caused by penicillin-resistant bacteria. Likewise, chloramphenicol is converted to thetherapeutically inactivecompound 1, 3-Diacetoxychloramphenicolby chloramphenicol acetyl transferase (CAT), produced by someresistant bacteria. N-acetylation of kanamycin by a bacterialenzyme leads to tolerance of the organism to the antibiotic.

5.2.2 Modification of Target Site: The other mechanism ofantibiotic-resistance e.g., modification of the target is best exem-plified by streptomycin and erythromycin resistance of bacteria.Both of these antibiotics act by ribosomal binding thereby inhib-iting bacterial protein synthesis. Modification of the S12 proteinof the 30S subunit of the ribosome makes the ribosome insensi-tive to streptomycin. Mutations affecting protein L4 or L12 of the

5 Small segments of DNA whichcan hop from one DNA locationto another. It is found as part ofa bacterium’s nucleoid (conju-gative transposons) or in plas-mids. It contains a number ofgenes, coding for antibiotic re-sistance or other traits, flankedat both ends by insertion se-quences called transposase,which catalyzes the cutting andresealing of the DNA duringtransposition. Thus it is able tocut itself out of a bacterial nucle-oid or a plasmid and insert intoanother location leading to thetransmission of antibiotic resis-tance among a population ofbacteria.

6 A mobile genetic element,which contains sitespecific re-combination system capable ofcapturing and mobilising genecassettes. It is composed of anintI gene encoding an integrase,a recombination site attI, and apromoter (5’ to the cassette) thatmediates expression of the re-sistant determinant. The 3’ endof the cassette has a variableregion termed as 59bp element,which contains the recognitionsequence of the integrase. Theintegrase is able to integrate orexcise genes-cassettes (usu-ally antibiotic-resistance genes),by site-specific recombination.For dissemination, the integronmust be able to insert itself intoa plasmid or transposon. To dateintegrons are found in Gram-negative bacteria.


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50S ribosomal subunit render it resistant to erythromycin. An-other example is the modification of the -subunit of RNA-polymerase of Mycobacterium leading to the failure of the anti-tubercular drug rifampicin to bind the subunit and inactivate theenzyme.

5.2.3 Reduction in Permeability to Antibiotics: In some cases,emergence of mutants with reduced permeability of the cellmembrane to antibiotics compared to that of the wild-type strain,leads to tolerance to the antibiotic. For example Neisseria gonor-rhoea, the causative organism of gonorrhea (one of the mostprevalent sexually transmitted disease), can gain antibiotic resis-tance by acquiring a mutation in the gene encoding the membraneprotein ‘porin’, thus inhibiting the transport of the antibioticspenicillin and tetracycline into the cell and rendering the cellsimmune to the effect of drugs.

5.2.4 Exclusion of Antibiotics from the Cell: Among the mecha-nisms involved in the resistance of bacteria to tetracycline, en-ergy mediated efflux is a powerful strategy, which does not allowthe drug to accumulate in sufficient concentration to exert itsinhibitory effect. It is mediated by a trans-membrane exportprotein that functions as an elctroneutral antiport system. Theprotein catalyzes the exchange of a tetracycline-divalent metalcation complex for a proton.

5.2.5 Overproduction of a Target Metabolite: In some cases, themolecule, which is competitively antagonized by the antibiotic, isoverproduced. For example, sulphonamides act by competitivelyinhibiting the enzyme dihydropteroate synthetase, which plays acrucial role in the synthesis of folate. Sulfonamides (or sulfadrugs) are structural analogs of p-aminobenzoic acid (PABA), thesubstrate of this enzyme. The indispensable role of folate in thesynthesis of nucleic acids viz., DNA and RNA is well-known. Insome PABA-overproducing mutants of Staphylococcus aureus,sulfonamide molecules are outnumbered by the substrate andtherefore the activity of the enzyme is not inhibited even in thepresence of the drug.


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5.2.6 Gene Deletion: Gene deletion is also known to contribute tothe mechanism of antibiotic resistance. Deletion of the katG genein some strains of Mycobaterium tuberculosis is associated withthe tolerance of the organism to the anti-tubercular drugisonicotinic acid hydrazide (INH). It is postulated that INH isactually a prodrug [15], which is converted into the active formbycatalase/peroxidase, encoded by katG. Thus, absence of the katGgene makes the organism immune to the effect of the drug.

6. Adaptive Resistance

In the laboratory, a bacterial population can adapt to tolerate anantibiotic following exposure to gradually increasing concentra-tion of the drug. While resistance due to mutation occurs in lowfrequency, resistance due to adaptation imparts tolerance to thewhole population at a time. It is usually caused by a change in thelipid composition of the cell wall ultimately leading to reducedpermeability to the antibiotic. The maintenance of the resistancephenotype requires the presence of an antibiotic in the growthmedium unlike mutational resistance, which does not require thepresence of the selection pressure. However, with the evidenceavailable so far, adaptive resistance does not appear to play anysignificant role in therapeutics.

A special type of non-heritable resistance to chloramphenicol andampicillin was also found to be induced in Escherichia coli whenit was incubated in the presence of repellants like sodium acetate,acetyl salicylate, salicylate and sodium benzoate. The inducers inturn made the cells tolerant to ampicillin and nalidixic acid.Induction of tolerance to structuarally unrelated antibiotics by avariety of substances including some pharmaceuticals (acetylsalicylate) is commonly known as phenotypic antibiotic resis-tance and it appears to have some significance in therapeutics.

7. Non-Specific Resistance

Apart from harboring antibiotic resistance genes in mobile ge-netic elements such as plasmids and transposons, many bacterialspecies possess an intrinsic mechanism for resistance to multiple


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structurally unrelated compounds. The chromosomal multipleantibiotic resistance (mar) locus of some members of the Entero-bacteriaceae confers resistance to several antibiotics, householddisinfectants, organic solvents and other toxic chemicals. In Ecoli, multiple antibiotic resistance is known to be conferred bydifferent sets of genes. The mar phenotype is induced followingexposure to a variety of chemicals with aromatic rings such assalicylates, non-antibiotic antimicrobials (triclosan) and also su-peroxide radicals.

Bacteria can also survive antibiotics by the formation of biofilms,a multilayer conglomeration of diverse-species of bacteria, em-bedded in a self-produced exopolysaccharide matrix and spe-cially attached to surfaces including objects such as prostheses orcatheters. Antibiotic-resistance of bacteria in many infections(e.g., gingivitis, cystic fibrosis) is attributed to the biofilmsformed by them. The precise mechanism of this resistance is stillnot very clear. Bacteria detached from the biofilms are found tobe antibiotic-sensitive. Slow diffusion of antibiotics and a slowrate of growth and metabolism are believed to contribute signifi-cantly to antibiotic-tolerance in biofilms. Further, biofilms couldbe an ideal niche for horizontal gene mobilization, allowingresistance genes to be transferred and expressed rapidly. Conju-gation promotes formation of biofilms in E. coli. The ability ofbacteria to respond to quorum sensing [16] is a pre-requisite forthe formation of biofilm. It is not surprising therefore that somequorum sensing inhibitors (baicalin hydrate, cinnamaldehyde,hamamelitannin) have recently been found to enhance the effi-cacy of some antibiotics against pathogens.

8.1 The Antibiotic Paradox

In 1992, Professor Stuart B Levy, Director of the Center forAdaptation Genetics and Drug Resistance at Tufts UniversitySchool of Medicine (USA), proposed the theory of AntibioticParadox. It states that the widespread use of antibiotics itself ispromoting the emergence of resistant strains. The basis of thistheory lies in the fact that our body (as well as our surroundings)

Bacteria can alsosurviveantibioticsby the formation of

biofilms, amultilayerconglomeration ofdiverse-species of

bacteria, embeddedin a self-producedexopolysaccharide

matrix.


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is populated by millions of bacteria most of which are susceptibleto antibiotics. The frequency of occurrence of antibiotic-resistantbacteria is usually negligible in these naturally occurring popula-tions. Their growth is restrained by the susceptible bacteria,which outnumber them. Nowadays we resort to indiscriminateuse of antibiotics on trifle illness (a pill for every ill) and even incases like common cold for which the antibacterial antibiotics areof no use. These antimicrobials annihilate or suppress the friendlybacteria and thus provide opportunity to the antibiotic-resistantbacteria to grow and predominate in the population. We alsooften dump unused antibiotic formulations on the soil. Thusantibiotics help to select resistant bacteria in the soil by killing orsuppressing the susceptible bacteria.

Evidences available in the literature speak volumes in support ofthe theory. Following the Second World War, outbreaks of dys-entery caused by sulfonamide-resistant Shigella sp. was a majorconcern in Japan. The situation was improved by the introductionof a number of newly discovered antibiotics viz., streptomycin,and chloramphenicol, into the clinical practice in the year 1950.However by 1956, again a strain of Shigella flexneri, resistant tosulfonamides and all these three new antibiotics, was detected. Inthe year 1969, the frequency of occurrence of the multidrug-resistant strains was found to be 69% among the 10462 strainstested. Fluroquinolones were used only in 1.4% of the casesduring 1983–85. The fluoroquilone-resistance was not detectedin E coli tested during the period, 1983 to 1990. With a sharp risein the use of this drug in clinical practice during 1991 to 1993,28% of theE. coli strains testedwere found to be fluoroquinolone-resistant. A couple of years back, a team of investigators led byRanadhir Chakraborty, at the Department of Biotechnology in theNorth Bengal University, demonstrated a correlation between theprevalence of resistance to specific antibiotics in some riverisolates, obtained from north Bengal, and over the counter sale ofthe same antibiotics in and around the town of Shiliguri.

Antibiotics help toselect resistantbacteria in the soilby killing orsuppressing thesusceptiblebacteria.


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8.2 Other Examples

The use of antibiotics in poultries and animal farms for clinicalmanagement of the infections of the livestock and also as agrowth promoter is well-documented. Substantial portion of theantibiotics applied to the animals is excreted into the soil, leadingto the selection of antibiotic-resistant strains. Washed by rain andcarried to the ponds and rivers, these antibiotics enter the body offishes, which serve as a reservoir of resistant bacteria. Emergenceof resistant strains in fishes is also favored by the antibiotics,used for the control of infections in fish farming. Isolation ofantibiotic-resistant bacteria from fishes and other marine organ-isms has been reported from several laboratories across the globe.

9. Prudent Use Not a Permanent Solution

It may appear from Prof Levy’s theory that the problem ofantibiotic-resistance could be bypassed by avoiding the use ofantibiotics. In the UK a joint committee on the use of antibioticsin Animal Husbandry and Veterinary Medicine was set up in1968 under the Chairmanship of Prof. M M Swan. Antibioticswere routinely used during that time in the country for thepurpose of promoting growth of animals. The committee recom-mended a stricture on the use of therapeutically useful antibioticsfor this purpose. Use of tetracycline as a growth promoter wasprohibited in March 1971. However, during the 16 months ofprohibition the incidence of pigs carrying tetracycline-resistantorganisms was not found to decrease. Following the fall ofsocialism in Eastern Europe, and also because of commercialblockade inflicted on the country, Cubans had less access toantibiotics during the 1990s. After 10 years, it was found that thedegree of antibiotic-resistance in the harmless bacteria isolatedfrom the mouth of Cubans was almost equal to that of theircounterparts from Mexico, where antibiotics is available withoutprescription and also used in agriculture. Widespread resistanceto a number of therapeutically useful antibiotics was observed bythe investigators at the Centre for Cellular andMolecular Biology


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(Hyderabad, India) among bacterial isolates, obtained from dif-ferent types of samples (soil, ice, cyanobacterial mat), collectedfrom Antarctica, where exposure of the organisms to antibioticsis highly improbable. During an exchange program conducted atthe Leibniz Institute of Freshwater Ecology and Inland Fisheries(Stechlin, Germany), it was found that incidence of chloram-phenicol-resistance (19.69%) was not substantially lower thanthat of erythromycin-resistance (24.24%) in various types oflake-isolates, sampled from some least populated areas of northGermany. It is well-known that use of chloramphenicol for thera-peutic purpose was banned in the Western countries quite sometime back while erythromycin is still being used. Many otherreports of this sort reveal that prudent use of antibiotics mighthelp slow down the emergence of resistant strains but the strategycannot ensure complete reversal and disappearance of resistance.

10. Co-occurrence of Antibiotic Resistance with Toleranceto Other Antimicrobials

Antibiotic resistance genes are often located on plasmids, whichalso carry genes conferring resistance to many other chemicalsincluding heavy metals and disinfectants. Substances like heavymetals and disinfectants are persistently present in the environ-ment since they are part and parcel of the industrial civilization.Consequently, they select resistant strains among the naturallyoccurring population of bacteria and in the process, cross-selectantibiotic-resistant strains. In some cases, bacteria are found toharbor plasmids conferring resistance to a number of antibioticsand heavy metals. It is also known that in the absence of antibiot-ics, resistance plasmids and their bacterial hosts co-evolve insuch a way that, after several generations, they grow better than astrain that lacks the plasmid. The metabolic burden needed tomaintain such a big plasmid does not seem to pose any challengeto the organisms and they are found to carry it through genera-tions without any selection pressure. That is why it is said thatantibiotic resistance genes are easy to get but difficult to lose.


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11. Search for New Antibiotics

An antibiotic research reported by the London School of Eco-nomics and Political Science (LSE) showed that bacterial andparasitic diseases are the second leading cause of death world-wide. Because of the emergence of drug-resistant ‘superbugs’,(like MRSA and VRE), traditional antibiotics and its derivativesare becoming nonfunctional. The whole world is thus confrontedwith a looming drug crisis because of frequent emergence ofresistant strain and development of new antibiotics is desperatelyneeded. The search for new antibiotics during the past fewdecades was mostly based on the chemical modification of theexisting drugs but in the post genomic era, molecular genetics isbeing used to identify essential new targets with the help of highthroughput screening and combinatorial chemistry. Structuralbiology is being applied to rapidly explore and optimize theinteractions between lead compounds and their biological targets.Genomic tools are helping us to select for antibacterial targets andunderstand bacterial resistance. On the other hand, the availabil-ity of a plethora of natural products and combinatorial syntheticsmall molecule libraries, provides ample opportunities to dig intothese resources in search of new chemotherapeutic agents. Anumber of approaches to the discovery of new antibacterials areon the anvil. Some of them are based on the forward chemicalgenetic approach while the others are based on the reversechemical genetic strategy (Box 1).

12. Doomsday Ahead?

In the past few decades, because of the emergence of resistantstrains, therapeutic benefit of some of the potent antibiotics wascompletely lost. Over nearly 20 years, from the early 1980s to thelate 1990s, not a single truly new antibiotic was introduced intoclinical use. Even now, barely a trickle has reached the marketsince 1999. Only five new antibiotics were approved from 2003to 2007, compared to the 16 approved during 1983 to 1987. Thesituation for the treatment of Gram-negative infections is evenbleaker. The impermeable nature of the Gram-negative envelope


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and presence of multiple efflux pumps in combination with otherresistance mechanisms contribute to the difficulty of this task. Ifthe emergence of antibiotic resistance continues to follow thepresent trend, it is reasonable to assume that resistance to the newantibiotics will emerge sooner or later. In fact, reduced suscepti-bility to linezolid and daptomycin has already been encounteredin the clinical setting. Despite prudent usage, the life span of anew antibiotic is cut short by the emerging resistance. So it isevident that a continuous supply of structurally novel antibacte-rial agents with multiple mode of action is needed to combat theproblem of drug resistance. The optimism associated with antimi-crobial peptides because of their broad-spectrum activity, ab-sence of cross-resistance with the existing antibiotics and lowprobability of developing resistance is hindered by their poorbioavailability and high cost involved in their manufacture. Phy-sicians are often taking resort to multiple antibiotic therapies,which are likely to be more effective than using a single

Box 1. Chemical Genetics

Chemical genetics is a chemistry-based approach, which involves diverse small-molecule compounds toelucidate biological pathways in a manner analogous to the mutagenesis strategies used in classicalgenetics. Screening small-molecule libraries to induce a phenotype of interest represents the forwardchemical genetic approach, whereas the reverse approach involves small molecules targeting a singleprotein. The prerequisite for this analysis is a collection of potential compounds (natural products,combinatorial libraries, inventories and chemi-informatics) and high throughput functional assays (invitro binding assays, cellular assays, yeast, bacteria, animal models, bioinformatics).

In forward genetics, smallmolecules are used to at-tain a specific phenotypeand try to find the gene thatcontrols it; in reverse ge-netics, small molecules areused to manipulate genesor proteins to characterizetheir role by identifying theresulting phenotype.

If the emergence ofantibiotic resistancecontinues to follow thepresent trend, it isreasonable to assumethat resistance to thenew antibiotics willemerge sooner orlater.


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antibiotic. Scientists are also looking for alternative strategiese.g., enhancing the susceptibility of bacteria to antibiotics byusing plant-derived antimicrobials (viz., thymol, carvacrol,cinnamaldehyde, allyl isothiocyanate). Let us hope for a betterfuture while being aware of the precarious situation that isprevailing at present.

13. Conclusion

The foregoing discussion on antibiotic-resistance in bacteria isfocused on the clinical impact of the phenomenon. But there aremany other aspects which are overshadowed by its devastatingeffects on the prospect of chemotherapy. The history of antibiot-ics as therapeutic agents is less than 100 years old while antibioticbiosynthetic pathways and antibiotic-resistance genes are be-lieved to have evolved thousands of years ago. So in nature bothantibiotic biosynthetic pathways and mechanisms involved inantibiotic resistance must have some other significance, whichwarrants extensive investigations. The multiplicity and non-speci-ficity of efflux pumps and occurrence of resistance-conferringgenes in non-pathogenic bacteria hint at some other role ofantibiotics in evolution. It is also believed that bacteria senseantibiotics as an environmental stress. Hence there might be somecorrelation between antibiotic-resistance and stress tolerance ofbacteria. Antibiotic-resistance has been detected in many bacte-ria isolated from extreme environments. Enhanced rate of hori-zontal gene transfer was observed by some investigators in somefood-borne bacteria treated with sublethal levels of stress factors.Therefore, it is obvious that the clinical aspect of antibiotic-resistance is only the tip of the iceberg and most of the aspects inthe study in antibiotic-resistance of bacteria remain unexploredtill now. We hope that thorough investigations will provide moreinsights in the years to come.

Suggested Reading

[1] C F Amabile-Cuevas, New antibiotics and new resistance, AmericanScientist, Vol.91, pp.138–149, 2003.


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[2] R I Aminov, The role of antibiotics and antibiotic resistance in nature,Environmental Microbiology, Vol.11, pp.2970–2988, 2009.

[3] HWBoucher, GH Talbot, J S Bradley, J E Edwards, DGilbert,L B Rice,M Scheld, B Spellberg and J Bartlett, Bad bugs, no drugs: No ESKAPE!An update from the Infectious Diseases Society of America, Clin. Infect.Dis., Vol.48, pp.1–12, 2009.

[4] G Brackman, P Cos, L Maes, H J Nelis and T Coenye, Quorum sensinginhibitors increase the susceptibility of bacterial biofilms to antibioticsin vitro and in vivo, Antimicrob. Agents Chemother., Vol.55, pp.2655–2661, 2009.

[5] M K Chattopadhyay and H-P Grossart, Antibiotic resistance, intrac-table and here’s why, British Medical Journal, Vol.341, p.c6848, 2010.

[6] R Jayraman, Antibiotic resistance: An overview of mechanisms and aparadigm shift, Curr. Sci., Vol.96, pp.1475–1484, 2009.

[7] P Keith, Uninhibited antibiotic target discovery via chemical genetics,Nature Biotechnol., Vol.22, pp.1528–1529, 2004.

[8] S B Levy, The antibiotic paradox: How the miracle drugs are destroyingtheir miracle, Plenum Press, New York, USA, 1992.

[9] M N Alekshun and S B Levy, The escherichia coli mar locus – antibioticresistance and more, ASM News, Vol.70, pp.451–456, 2004.

[10] K Palaniappan and R AHolley, Use of natural antimicrobials to increaseantibiotic susceptibility of drug resistant bacteria, Int. J. FoodMicrobiol.,Vol.140, pp.164–168, 2010.

[11] J L Rosner, Nonheritable resistance to chloramphenicol and otherantibiotics induced by salicylates and other chemotactic repellents inEscherichia coli K-12, Proc. Natl. Acad. Sci., USA. Vol.82, pp.8771–8774, 1985.

[12] G H Talbot, What is in the pipeline for Gram-negative pathogens?Expert Rev, Anti Infect. Ther., Vol.6, pp.39–49, 2008.

[13] S K Vooturi and S M Firestine, Synthetic membrane-targeted antibiot-ics, Curr. Med. Chem., Vol.17, pp.2292–2300, 2010.

[14] Saurabh Dhawan and Tomas J Ryan, The bacterium that got infected bya cow!, Resonance, Vol.12, No.1, pp.49–59, 2007.

[15] H Surya Prakash Rao, Capping drugs: Development of prodrugs,Resonance, Vol.8, No.2, pp.19–27, 2003.

[16] Avantika Lal, Quorum sensing: How bacteria talk to each other, Reso-

nance, Vol.14, No.9, pp.866–871, 2009.

Address for CorrespondenceSaswati Sengupta

Indian Institute of ChemicalTechnology (CSIR)

Hyderabad 500 007, India.Email:

[email protected]

Madhab K ChattopadhyayCentre for Cellular andMolecular Biology (CSIR)Hyderabad 500 007, IndiaEmail: [email protected]

Antibiotic resistance of bacteria: A global challenge

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