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ANTIBIOTICS IN MANURE AND SOIL – A GRAVE
THREAT TO HUMAN AND ANIMAL HEALTH
PREAMBLE
Growth promoting properties of antibiotics in farm animals were first discovered in the late1940’s in chickens and pigs. Feeding of sub-therapeutic doses of antimicrobials to thefarm animals was readily adopted and it has now become an integral part of the farmanimal/fish production systems. In spite of tremendous beneficial effects in improvingfeed efficiency and live-stock productivity, antibiotics are now found loosing ground asadditives in animal feed, because of their effect on development of resistance in somedeadly bacteria in the animal gut and in the terrestrial environment. Today, there is agrowing concern over the use of antibiotics to promote growth of animals.
Use of antibiotics as growth promoter is loosely defined as administration of antibioticsto healthy animals at concentrations below 200 ppm in feed for more than 14 days. Thisfeed dose, in terms of animal body weight will be around 200 mg per 100 kg, which worksout to a concentration of 2 ppm in animals. This much dose is well distinguished fromtherapeutic and prophylactic antibiotic use, which is generally delivered at a higher minimumdose of about 20 ppm in animals and are generally administered in water.
Recommended inclusion levels in poultry and pig diets were 4 ppm for the narrow spectrumand 10 ppm for the broad-spectrum antibiotics in 1950’s. Since then these levels haveincreased 10 to 20 folds. In case of fish and fish products certain antibiotics are permittedup to 100 ppm.
The lower concentration (non-lethal dose for any bacteria) through animal feeds over longperiods results in a condition conducive for the development of resistance in bacteria. Thecontinued feeding of antibiotics in feed also introduces low levels of antibiotics in the soiland water through the animal excreta. This in turn produces antibiotic resistance in soilbacteria including pathogenic bacteria. Overuse of prescribed drugs in human and veterinarymedicine, their use as growth promoters in live-stock feeds has all been blamed as thecause of growing antibiotic resistance.
Some of the antibiotics approved for use as feed additives in livestock production andacqua culture are Beta-Lactams (Cefadroxil, Ceftiofur, Penicillin, G benzathine, G potassium,G procaine, G sodium, Amoxicillin, Ampicillin), Macrolides (Erythromycin, TilmicosinPhosphate, Tylosin) Spectinomycin, Chloramphenicol, Florfenicol (Aqua-flor, Nuflor),Nitrofurans (Furazolidone, Furaltadone, Nitrofurazone, Nitrofurantoine), Tetracycline,Oxytetracycline, Chlortetracycline, Quinolones (Enrofloxacin, Sarafloxacin, Oxolinic acid,Flumequine), Sulphonamides (Sulphonamides, Sulfachlorpyridazine, Sulfadimethoxine, National Academy of Agricultural Sciences Sulfamethazine, Sulfanilamide, Sulfaquinoxaline, Sulfathiazole) and Aminoglycosides(Aminoglycosides, Amikacin, Apramycin Dihydrostreptomycin, Gentamicin, Kanamycin,Neomycin, Streptomycin,) Some of the bacterial infections which could normally be treated by specific antibioticshave turned out to be untreatable. For example, methicillin was introduced in 1960 for thetreatment of Staphylococcus aureus infection and with in a few years, methicillin resistantStaphylococcus aureus (MRSA) strains were reported. Then in 1980, fluoroquinoloneswere introduced for treatment of MRSA, but a majority of Staphylococcus strains becameresistant to fluoroquinolones with in a year. A national news paper (The Telegraph,Kolkata, Monday, 11 December, 2006) reported that a bacteria displaying resistance tovirtually all antibiotics known to humans has surfaced in the country, sending ripples ofalarm among medical researchers. A nationwide surveillance was conducted and it steppedup efforts to stem its emergence. Scientists of Institute of Medical Sciences, Varanasidetected the “super bug" among strains of Staphylococcus aureus. These bacteria cancause life-threatening infections such as pneumonia and septicemia. Two strainsof Staphylococcus aureus were found resistant to Vancomycin, the drug of last resort inthe arsenal of conventional antibiotics.
Worldwide legislation to control use of antibiotics has eventually fuelled the reduction ofantibiotic use, at least their non-therapeutic use. Consumers in many advanced countriesare no longer keen to eat meat / livestock products from the animals raised on feedscontaining antibiotics and efforts are being made to ban their use in many countries.
World Health Organization of United Nations (WHO), American Medical Association, andthe American Public Health Association have urged a ban on use of antibiotics as growthpromoting additives (GPAs) due to increased antibiotic-resistant infections in humans.
Some of the warnings / actions against the use of antibiotics in animal feed are inTable 1.
Table 1. Warnings/Actions against the use of antibiotics as animal feed additives
Warnings / action
Alexander Flemming warns against misuse of penicillin as ‘microbes Antibiotic resistance widely recognised — vertical transmission.
Tuberculosis bacteria resistant to Streptomycin.
Certain strains of dysentery bacillus was found resistant to Chloromphenicol, Tetracycline, Streptomycin, and Sulphanilamides Warnings / action
Swann Committee recommends severe restrictions on antimicrobial supplementations in animal feeds.
Swann committee recommendations implemented in the UK and EU Swann committee recommendations relaxed: tolysin and spiramycin still permitted as growth promoters; vancomycin comes into use.
Swedish Agriculture Board considers potential risk of antibiotic resistance, but concludes it is negligible Swedish farmers ask for government ban on antimicrobials in animal feed because of health and consumer concerns.
Swedish ban on grounds of antibiotic resistance in animals and it’s Avoparcin and Vancomycin resistant Enterococci in pigs and poultry.
Swedish report concludes that risk of antibiotic resistance in humans WHO scientific meeting concludes that it is ‘essential to replace Danish government banned Virginiamycin due to Streptococcus EU bans five antimicrobials in animal feed as ‘precautionary’ measure, such as Avoparcin, Bacitracin Zn, Spiramycin, Virginiamycin and EU Scientific Steering Committee recommends phase-out of antimicrobials that may be used in human/animal therapy Pharmaceutical industry opposes EU bans and takes EU to the European Court; judgement expected end 2001 WHO recommends ban on antimicrobials as growth promoters if used in human therapy and in absence of risk-based evaluation.
National Academy of Agricultural Sciences Warnings / action
U.S. Food and Drug Administration banned the non-therapeutic use of Eenrofloxacin for growth promotion in food animals on the grounds that its use has contributed to fluoroquinolone-resistance in human European Commission authorized use of Flavophospholipol, Monensis Na, Salinomycin Na and Avilamycin in poultry, beef cattle, pigs, rabbits and calves diets, as these are not used in human medicine. The act is Animal Drug user Fee Act (ADUFA), Strategies to address antimicrobial drug resistance (STAAR) and Preservation of Antibiotics for Medical Treatment Act (PAMTA) US congress passed a legislation regarding AGP’s Purpose of this policy paper is to invoke general awareness about the indiscriminate useof antibiotics in agriculture and its impact on human health and terrestrial environment.
A brief account of information that emerged during the deliberations is given below: Use of Antibiotics in Animal Feed for Growth Promotion
The overall outcome of use of antimicrobial growth promoters (AGPs) is the availabilityof more nutrients for growth and production of livestock and poultry. Improvement ofgrowth rate and feed conversion ratio (feed : gain) has been reported as 16% and 9% inpiglets, 9% and 5.5% in growing pigs, 3-10% and 3-5% in broiler chickens, 2% and 1%in layers and 7-10% in veal calves. Antimicrobials for therapeutic purposes can bepurchased only on prescription of a registered medical practitioner, however, as growthpromoters; these are freely accessible and sold over the counter. Effects of uses ofAGP’s on broad issues are depicted in Table 2.
Table 2. Effects of AGP in relation to some broad issues of animal production.
Broad issue
Positive effect
Negative effect
1. Development of antimicrobial resistance 2. Masks sub-clinical disease and infection 1. Camouflages stress associated with sub- Broad issue
Positive effect
Negative effect
Stimulates, increases and intensifies animal Lowers labour demand 1. Hampers the development of animal- even for pathogens of farm animals.
1. Threat of infection by antibiotic resistant Exact mechanism as to how AGP’s promote growth is not entirely clear. It is widelyassumed that AGP’s act mainly through their effect on intestinal flora. With less than 10% of intestinal micro-flora identified, there has been little chance of fully understandingAGP’s mode of action. It is postulated that AGP’s allow the animal to express theirnatural potential for growth through their direct influence on gut bacteria. AGP’s arebeneficial to the host as these reduce the total number of intestinal microorganisms.
Secondly, these may also create a more favourable balance between beneficial and non-beneficial ones. Thirdly, there will be a proportional sparing of nutrients, which are utilizedby the animal for absorption and weight gain.
In general, the antibiotic feed will be beneficial to the animal because of inhibition of subclinical infections, reduced gut motility, reduced mucin secretion, reduced toxin (eg.
ammonia and biogenic amine from proteins) production, bile salts modification, thinningintestinal wall, increase in digestive enzyme output, improved digestibility, increased National Academy of Agricultural Sciences uptake of nutrients along the alimentary canal, reduced opportunity for harmful bacteriato establish in the gut, activation of the intestinal immune system and reduction inmicrobial use of nutrients, thus sparing the nutrients for the host. The ultimate impactsof antimicrobials as growth promoters in farm animal/aquaculture production systems areincreased growth rate of animals/fish/shell fish, better feed conversion, improved eggproduction in laying hens, increased litter size in sows, early weaning of piglets, increasedmilk yield in dairy cows, economized animal production systems, reducedincidence of disease in aquaculture, high stocking rates and some protection againstcertain diseases.
Antibiotics in Soil Environment and Food Chain
A major portion i.e.30- 80 percent of antibiotic dose fed to the animals as growth promotersmay be excreted as waste because of poor absorption. When antibiotic-laden manure isused to fertilize crop lands, antibiotics in the manure may get into the soil and eventuallyend up in streams, lakes or rivers. Antibiotics enter the environment by two ways, (i)directly when using the drugs i.e. the unabsorbed as waste and (ii) subsequent excretionof absorbed antibiotic residues and their metabolites through urine and feaces of theanimal. The dominating pathways of environmental release of antibiotics in the terrestrialcompartments are through application of FYM in arable soil and in fish farms. An unknownpart of food-pellets containing the medical compound may not be eaten by the animalsand hence will reach the sediment directly without any change. Antibiotics after consumptionmay be excreted partly as unchanged compounds or as metabolites, which finally reachthe sediment. No information is available on the fate of veterinary medicinal products with antibioticsduring storage of manure/cow dung slurry. Two types (hydrophilic and hydrophobic) ofsubstances are available in the manure /slurry. The antibiotic residues present in the dungdepending on its chemical properties, either undergo degradation or leach to the soil. Ingrazing animals drugs released via the urine immediately reach the soil and if watersoluble, leaching down to ground water or adjacent water systems is a rapid process.
Most drugs excreted through the urine are water soluble, whereas drugs excreted viafeaces, in general, are less soluble.
Antibiotics, which kill disease causing bacteria, especially broad spectrum antibiotics,could work havoc on natural microbial communities in the soil. Antibiotics might disruptessential biological activity in the soil. One major consequence of this phenomenon is theemergence of antibiotic-resistant bacteria that could infect humans, livestock, fish &shellfish and wildlife. After consumption of antibiotic laced feed to the animals, unusedantibiotics enter the environment. In fish and shellfish farming, antibiotics are given asfeed additives and approximately 70 to 80 % are directly released into the aquaticenvironment. Antibiotic residues with significant antibacterial activity have been reportedin the sediments from fish and prawn hatcheries and farms in our country.
Tetracyclines (oxytetracycline and chlortetracycline), tylosin, sulfamethazine, amprolium,monensin, virginiamycin, penicillin, and nicarbazine are the most common antibioticspresent in swine, beef, and poultry/turkey manures. The concentration of these antibioticsvaries from traces to as high as 216 (mg L–1) of manure slurry. These antibiotics generallyremain stable during manure storage and end up in agricultural fields on manure applications.
Soil and water contamination from manure fertilization has been frequently reported. It isa matter of great concern that residual concentrations of antibiotics in soils can easilyreach levels similar to pesticides. Such a contamination by veterinary antibiotics exposeshumans and animals to a constant threat of unknown consequences due to the presenceof low concentrations of antibiotics in the environment. This threatens the human andanimal health by diminishing the success of antibiotic treatment. Evidences show thatantibiotic resistant genes from microorganisms in the environment can transfer directly tohumans. Leaching and runoff of antibiotics from manure-fertilized lands is threatening thequality of drinking water. Effects of long-term exposure to low concentrations of antibioticsare not yet clear, but the potential danger resulting from veterinary antibiotic contaminationto human and animal health cannot be neglected.
Persistence of Antibiotics in Soil
There are very few studies on reaction of antibiotics in soil. Photolysis, hydrolysis, bio-degradation and binding on to soil particles through adsorption process are some of thereactions of these antibiotics and their products that can take place and influence theirpersistence in soil. They may form complexes with soluble organic materials and becomemore mobile and contaminate even groundwater while still in its parent form.
Antibiotics like ciprofloxacin, ofloxacin, and virginiamycin degrade very slowly and maypersist in soil in its original form up to 30-80 days while bambermycin, tylosin, anderythromycin completely degrade in a period of one month at temperatures ranging from20-300C. Persistence of an antibiotic in the terrestrial environment is the key factordetermining its environmental impact. Most of the antibiotic residues in manure generallyremain stable during manure storage until its application to agriculture fields.
So far, there have been very few studies on the impact of the antibiotics added to thesoil through manure, sludge and waste waters on the environment and perhaps none inthis country. We need to collect precise data on antibiotic use in animal farming, aquacultureand agriculture and the potential reservoir for residual antibiotics in the terrestrial environmentin the country. Besides, research work is needed to understand kinetics of biodegradationand potencies of degraded products of various antibiotics in different soils, manures andwaste water. This would help us to better understand the eco-toxicological impacts ofvarious antibiotic residues in the environment.
Effects of some soil properties on the persistence of anti biotics in soil are brieflydescribed.
National Academy of Agricultural Sciences Effect of soil pH
Soil pH plays an important role in ionization of most of the antibiotics. Besides molecularstructure, molar mass, physico-chemical properties and their dissociation constant (pKa)value of antibiotics determine as to how they will ionize in soil due to pH variation. Theanti-microbial properties of the antibiotics in soil are determined by their active functionalgroups.
Most of the antibiotics of tetracycline group are amphoteric in nature and are stable underacid conditions. These compounds can form chelates with divalent metal ions.
Sulfonamides, on the other hand, have two pKa values, they behave as weak acids andform salts both under acid and basic conditions. Amino-glycosides being polar compoundsmove easily with the percolating water and can contaminate the ground water resources.
But they are photodegradable therefore, easily decompose when subjected to sun-light.
Penicillin belongs to ß-lactam class of antibiotics. It is stable under a wide range of pHvalues from strong acid to strong alkali conditions. This speaks of its long persistencein soil as parent molecule and is a potential health hazard. Fluoroquinolones, also resistbreak down through hydrolysis and therefore are highly stable in soil.
Adsorption reactions
Antibiotics in soil can be retained on the mineral and organic colloids. In this form, theyare less liable to degradation forces and less potent towards its targets. Antibiotics thatcan ionize in soil to furnish positive charges can be retained on colloidal soil surfacethrough the adsorption process. Binding strength of antibiotics with soil is determined bythe negative charge in the soil and positive charge developed in antibiotic molecule. Theextent of such binding can be quantified by taking the ratio of antibiotic concentrationadsorbed in soil to the same in water in equilibrium with soil. This is also called distributioncoefficient (Kd). Antibiotics with higher Kd value are strongly bound with the soil and areless mobile. Compounds with less Kd value are less strongly bound and more mobile inthe soil. The later group of antibiotics can be easily transported to contaminate the groundas well as surface waters. Strongly bound antibiotics can however, be transported mainlyto surface waters with the sediments during run off losses of soil. Some of the antibioticcompounds form complexes with soluble organic matter in the soil. This increases theirmobility and they easily find their way to contaminate ground waters. Under ordinaryconditions, they are strongly bound to soil solids and thus highly immobile. In highlydeveloped dairy farming countries like Germany, sulfamethoxazole concentration as highas 40 ng L-1 has been reported in almost 10% of the ground water samples tested.
Sulfonamides have little adsorption tendency and do not form immobile complexes in soil.
Therefore, sulfonamides are strong contaminators of ground and surface waters whiletetracycline is likely to contaminate mainly the surface water bodies.
The tenacity with which the antibiotics are held on the soil solid surfaces is also determinedby pH, clay and soil organic matter contents. Amphoteric antibiotics like tetracyclinewhich is most widely used in animal feed and fish feed, may exist as anions and cationsdepending on pH of the medium. Cationic antibiotics bind to the soil particles through ionicinteraction, while acidic and amphoteric antibiotics may bind to the soil through non-ionicinteraction.
Effect of soil texture
Soil type, whether loam, silt loam or sandy loam matters in the persistence of antibioticsin soil. For example, it has been found that ciprofloxacin was mineralized to CO less than 1% in all the three soils in 80 days of incubation. Strong binding of this antibiotic wassited as the reason for its slow degradation. Half-life of ceftiofur was more than 49 daysin sand and only 22 days in clay loam. Half-life of oxytetracycline in marine sedimentsat a depth of 5 to 7 cm was more than 300 days as compared to 87 to 173 days forvirginiamycin in sandy soil. This shows that antibiotic persistence in soil is determinedby not only the soil type but also soil depth. Antibiotics can persist for longer periods ifthey are lodged in sub-surface soil layers and deep in waters away from sunlight andaeration.
Effect of type of clay minerals
Depending on the reaction of antibiotics with the clay minerals, antibiotics can be dividedinto 4 groups: Streptomycin; dihydrostreptomycin; neomycin and kanamycin.
In soils dominated by montmorillonite or illite or kaolinite, clay mineral reacts with the firsttwo groups (strongly basic and amphoteric) of antibiotics to form complexes. But acidicand neutral antibiotics are adsorbed only in soil that dominantly contains montmorillonitetype of clay mineral; still the tenacity of adsorption is relatively weak. On an average, theamount of antibiotics adsorbed by clays varies from 9 mg g-1 for kaolinite clays andstrongly basic antibiotics to more than 300 mg g-1 in case of montmorillonite clays andamphoteric antibiotics. Strongly basic antibiotics are so strongly held on the clay surfaceof montmorillonite, vermiculite or illite minerals, that they are virtually un-releasable asassessed from bioassay studies. In kaolinite, however, where it is not held that strongly,there is some release in case of streptomycin and dihydrostreptomycin. But in case of National Academy of Agricultural Sciences amphoteric antibiotics, there is easy release from all kinds of clay minerals. Streptomycinthat is commonly used as growth promoter in swine is adsorbed strongly on the soilparticles, which is high in clay and low in sand fractions. Wide variation in tenacity of theiradsorption on soil exchange sites is apparent from a vide range of observed sorptiondistribution coefficient Kd from 0.2 to as high as 6000. Clay adsorption is the main reasonfor weakly adsorbed antibiotic compounds, such, as metronidazole and olaquindox to bemore mobile and can leach with percolating water. However, strongly adsorbedoxytetracycline and tylosin percolate the least in the leachate. In general, affinity of manyof the commonly used antibiotics as growth promoters is quite high to soil particles. Thisindicates that most of the mobility of these antibiotics in terrestrial environment is probablydue to run-off losses of antibiotic-laden sediments to surface waters from fields whereantibiotic laden manures are applied.
Effect of soil temperature
Most of the degradation process of antibiotics in soil is mediated by soil micro-organisms.
Therefore persistence of these compounds in the soil is affected by all those factors thataffect the activity of microbes. Soil temperature is an important factor in this respect. Asthe temperature decreases from the normal range of 25-300C, persistence of antibioticsincreases. At 300C, 44% of chlortetracycline and 23% of bacitracin remained in the soilafter 30days of their application. However, when temperature decreased to 200C, 88% ofchlortetracycline, 33% of bacitracin, 25% of erythrocin remained in soil. At 40C, almostall chlortetracycline, erythrocin and bambermycin persisted in soil. It is very likely thatunder north-western Indian conditions, antibiotics finding their way to the fields withmanures during kharif season, a rapid decomposition may eliminate it from the soil dueto prevailing moderate to high temperature. During rabi season, when atmospherictemperature is low, significant portions of antibiotics applied through manures may remainintact in their original parent form.
Eco-toxicological Impacts
The soil environment may be impacted by the antibiotics in the following ways: Alter the composition and diversity of indigenous soil microbial communities whichare of fundamental importance for ecosystem, Change function in nutrient cycling, especially of nitrifying bacteria.
Inhibit decomposition of organic matter.
Develop resistance, (even cross and multiple), in organisms in the soil environment.
Antibiotics in soil are well known to inhibit microbial growth. How these affect soil faunaand flora, enzymatic activity and nutrient cycling needs greater emphasis for investigation.
These impacts could be direct, such as, antibiotic toxicity to soil microbes and indirecteffect, such as, reduced nutrient availability due to changed microbiological activity andreduced rate of organic matter decomposition in soil. Decomposition of organic matterdepends on various microbial processes, which in turn depend on type and population ofmicroorganisms in soil and also by allowing only the antibiotic-resistant microbes toflourish that may affect decomposition differently. Oxytetracycline or chlortetracycline fedanimals has been found to result in the manure/feaces which when applied to the soilresult in more evolution of CO . It has also been reported that feaces or dung from the animals fed with antibiotics contained higher proportion of easily decomposable/degradablecarbon compounds. Ionophore antibiotics, such as, monensin favour growth of Gram-negative bacteria in the gut.
Impact on Non-target Microorganisms
Antibiotics like streptomycin-laden manure decrease bacterial count in soil up to 50-75%over several months depending upon the nature of bacterial population. Streptomycin hasbeen specifically found to adversely affect the nitrifying bacteria. Gram negative bacterialike Nitrosomonas spp are responsible for nitrification in soil. Therefore, broad spectrumantibiotics like tetracyclines, aminoglycosides, and sulfonamides in manure and soil areexpected to inhibit the nitrification process. Narrow spectrum antibiotics such as sefdiazine,oxolinic acid, and tylosin, on the other hand, stimulate the nitrification process. Veterinaryantibiotics may also inhibit SO reduction as well. Build up of tylosin in soil can cause shifting of bacterial communities from Gram positive to a Gram negative.
Emergence of Antibiotic Resistance
Widespread use of antibiotics and their subsequent release into the environment may leadto the selection of antibiotic resistant bacteria. There has been a frequent observation onshortening of the time between introduction of a new antibiotic and development ofresistance of the targeted microbial species. If spontaneous mutations were the onlycause of antibiotic resistance, it would have been restricted to only a few bacterialspecies amongst the hundreds of billion in one antibiotic-treated host and would not bethe epidemic problem as it is to-day. A higher degree of vancomycin resistance (60%) invarious enterococci isolates from the broiler droppings has been reported. Similarly, a highlevel of resistance among Gram-positive and Gram-negative isolates from various meatproducts have been observed for penicillin, erythromycin, sulfamethoxine, tetracycline,ceftibiotics, and gentamycin. Therefore, animal manures containing such antibiotics cancause elevation of resistance in soil bacteria. Development of resistance is quite rapidas within 3 weeks of antibiotic feeding, more than 70% of feacal bacteria become resistantto penicillin and tetracycline. This indicates that antibiotic feeding provides an environmentfor selection of resistant strains and may encourage the transfer of genetic informationfrom even unrelated bacterial species. Every time one or the other antibiotic drug becomes National Academy of Agricultural Sciences ineffective because of the emergence of resistance in the targeted bacteria. Discovery ofa new drug is very expensive costing more than one $ billion and may take 10-15 yearsbefore getting its regulatory approval. It is thus very important for us to look for safergrowth promoters and use antibiotics to a bare minimum. Antibiotics used for managinghuman infections should be totally banned for use as growth promoters.
Many bacterial species multiply rapidly enough to double their numbers every 20–30minutes. Their ability to adapt to changes in the environment and survive unfavourableconditions often results in the development of mutations that protect them. In addition,another factor contributing to their adaptability is that individual cells do not rely on theirown genetic resources alone. Bacterial capacity to adapt to external changes using thesemechanisms is called resistance development and this allows the resistant organisms toproliferate in the prevailing conditions. Resistance takes two forms (i) inherent or intrinsicresistance and (ii) acquired resistance. In intrinsic resistance, the species is not normallysusceptible to a particular drug. This may be due to the inability of the antibacterial agentto enter the bacteria to reach its target site. In acquired resistance, species is normallysusceptible to a particular drug but certain strains express drug resistance, which maybe mediated through a number of mechanisms. When resistance develops, the antibioticis no longer capable of curing or treating the disease caused by the infective agent. Alow level of resistance may be detected by a slight increase in the minimal inhibitoryconcentration (MIC), which is not necessarily of clinical significance. A higher degree ofresistance is characterized by an MIC that exceeds, sometimes by several orders ofmagnitude, the concentrations of drug safely attainable in the patient’s tissues.
Transfer of Resistance
Resistant genes are flowing freely between animal and human bacteria through the foodchain, which makes the situation more alarming. Of great concern is the possibility thatresistance generated on the farm could lead to a loss of effectiveness of key antibioticsin the management of human diseases. To assess the likelihood of the risk of resistancetransfer, two risks have to be considered independently : (i) risk associated with thetransmission of resistant bacteria from aquaculture environments to humans, i.e. theincrease of resistance in human bacterial pathogens as a direct consequence of the useof antibiotics in aquaculture, and (ii) risk associated with the introduction in the humanenvironment of non pathogenic bacteria containing antimicrobial resistant genes and thesubsequent transfer of such genes to human pathogens.
Different mechanisms exist in bacteria that make them resistant to a specific antibioticof a common chemical group. Antibiotic resistance is conferred by variations in thegenetic makeup of bacteria. The genes that confer antibiotic resistance are carried on thebacterial chromosome or on separate plasmids that can be transferred between different bacteria. In addition, genes that confer resistance tend to group together such thatresistance to multiple antibiotics is transferred by a single plasmid. These properties ofantibiotic resistant genes complicate the control of antibiotic resistance and contribute tothe concern over the use of antibiotics in livestock and public health. Antibiotics canpromote the establishment of an antibiotic resistant population of bacteria by killing thesusceptible bacteria and leaving behind the resistant bacteria. They do not induce thegenetic changes but select bacteria that already have the genetic changes responsible forresistance. Indiscriminate or inappropriate use of antibiotics can promote the selection ofantibiotic resistant bacteria. It is extremely important that antibiotics are used appropriatelyand according to the manufacturer’s recommendations unless otherwise specified by theanimal nutritionist/veterinarian.
Originally it was thought that resistance trait would be confined to the mutant clone andspread of resistance is confined to that clone only (Vertical Transmission). Later on,another type of resistance through mutation in existing genes (Horizontal transmission)was observed and this resistance could also be developed through the uptake of existinggenes. In this case, the resistance trait through mobile genetic elements can also spreadto other bacterial clones, to other bacterial species and even to other genera. Bacteriawhen exposed to antimicrobials develop strategies for their survival. Widespread use ofantimicrobials in human disease management undoubtedly is of more importance for theemerging antimicrobial resistance problems in humans. Continuous use of antimicrobialsin feed is one of the major sources of overuse and misuse of antimicrobials in animalproduction.
Resistance in Food-borne Pathogens
If there is development of resistance in this way into the food-borne pathogens, then itreally becomes a problem, because these infections can become difficult to treat withtraditional antibiotics, thus threatening human and animal life. The reports of severalincidences of infection by multidrug-resistant Salmonella typhimurium DT 104 is the resultof such a possibility. Around 20% of 120 isolates of E.coli from animal food were resistantto multi-antibiotic drugs. Similarly, a wide range of tetracycline-resistant genes of E.colihave been observed as isolated from human and animal sources. Gram-negativeenterobacteria and Gram-positive bacteria are the major source of antibiotic-resistantintegrons in animal litters. More than 40% of bacteria collected from surface waters wereresistant to one or more antibiotics in USA. Similarly, 54% of the coli form isolates ofKorean river were resistant to at least one antibiotic. In Greece, 20% of the Salmonellasamples isolated from surface waters were resistant to antibiotics.
Until recently, research on antibiotic use has been mainly directed toward their beneficialand adverse effects on the end user, humans and animals.
National Academy of Agricultural Sciences Allergic or Toxic Reactions
There have been relatively few studies on the effect of antibiotics on plants raised onmanure-amended soils. Consumers may unknowingly be ingesting some of these antibioticsby consuming vegetables grown on manure-applied lands. Some adverse effects ofconsuming antibiotics in vegetables and fruits are discussed. Some antibiotics wheningested by humans, especially children, cause serious allergies or toxicity. There maybe some interaction effects from simultaneous ingestion of two different antibiotics. It hasbeen shown that some of the macrolide antibiotics present in animal feed have interactedwith monensin resulting in its toxicity leading to death of affected cattle. Antibioticspresent in plant materials ingested by humans may provide resistance to human pathogensthus resulting in illnesses that may be difficult to cure with presently available antibiotics.
It has been shown that resistance of gut bacteria to antibiotics increased when fed withincreasing concentrations of penicillin in contaminated waste milk. Small amounts oftetracycline can act as a catalyst in triggering the horizontal gene transfer betweendifferent bacteria. Thus increasing resistance may be of concern both for human andanimal health if antibiotics are present in food crops. Our knowledge regarding the implicationsof manure-based antibiotics on the terrestrial environment and human health is limited.
There is an urgent need to study: Effect of cooking of food obtained from antibiotic fed animals / fish / poultry Fate of different antibiotics present in manure, Antibiotics and their degradation products taken up by plants grown on antibiotic-laden manure amended soils, Whether or not antibiotics or their degradation products are still bioactive to impartantibiotic resistance to gut and soil bacteria or cause adverse immunological reactionsin humans.
Alternatives to Antibiotic Growth Promoters (AGPs)/Veterinary nutraceuticals
Use of antimicrobial agents as feed additives is a complex issue with implications forhuman and animal health, animal welfare, food safety, environmental aspects, developmentof production systems, feeding practices and management of the animals. A completeban on the use of AGP’s will necessitate exploring the alternatives that can improve feedefficiency and general health status and enhance the immunity to fight against disease.
Some of the alternative products have proved useful. Examples of such materials arezinc oxide, copper sulphate, plasma proteins, egg yolk antibodies, organic acids, probiotics,prebiotics, enzymes, bioactive peptides, botanicals (herbs/spices), nutraceuticals, essentialoils and fermented liquid feeds. Several foods besides containing nutrients also contain certain compounds that enhance the production by providing either the nutritional balance,improving the metabolism or preventing the disease. Moreover at the same time there isincreased interest over the food safety, environmental contamination and the generalhealth risks that have made natural the norm, promoting the trend towards alternativestrategies to manage and feed the animals and birds without reliance on antibiotics. Suchtypes of foods are labeled as adaptogens, dietetics, nutracines, nutraceuticals ormultifunctional additives.
Nutraceuticals are a combination of nutrients and pharmaceuticals. Nutraceuticals mustimprove the performance effectively and economically, with little therapeutic use, withoutcausing cross resistance to other antibiotics at actual use level, without involving transferabledrug resistance and causing any deleterious disturbance to the normal gut flora andcreating environmental pollution. Moreover these must be non toxic to the animals andits handlers. Some of the nutraceuticals are briefly discussed : Herbs/Botanicals
Vegetative parts of the plants (leaves, bark, fruit, roots, seed and their extract) are calledherbs. The herbs contain a variety of chemical compounds that are used as bodyrestoratives. These chemical compounds are active in altering the physiological andbiochemical processes in the body. Herbs and spices have compounds with antibacterialeffects. For example garlic contains allicin and ajoene which exhibit broad spectrum antimicrobial properties and are effective in reducing cholesterol of liver, breast and thighmuscle. Another example is of Yucca Schidiger, which improves growth and feed efficiency.
Botanicals / herbs help in improving the performance by reducing the stress associatedwith handling, transport and poor health by providing nutrients and or active principles,which act as anti stress agents. These improve egg production in birds by ensuringnormal gut functioning and improving digestion by activating digestive secretions andimprove growth rate and animal production by increasing feed efficiency.
These properties of various herbs are due to the active secondary metabolites that belongto class of isoprene derivatives, flavonoides and glucosinolates. Interaction betweendifferent active components within and between extract may have either cumulative orantagonistic effects. Use of herbs in poultry and pig feeds is now gaining momentum asit claims to have no side effect, and is safe and eco-friendly. Botanicals have proved tobe equally beneficial as antibiotic growth promoters. Some botanical extracts have bothpositive and negative effect on the gut micro flora. This nature of botanicals can be usefulin the stabilization of gut environment. Reports indicated the reduced northern fowl miteinfestation with the topical application of garlic in laying hens. The desired activity ofherbs may vary due to variability of composition of plant secondary metabolites,environmental conditions, harvesting time, stage of maturity, method of extraction andconservation, anti nutritional factor and nature of diet in which it is supplemented.
National Academy of Agricultural Sciences Prebiotics
Prebiotics are short chain non-digestible oligosaccharides with 2-10 units of monosaccharideused as feed ingredients. Prebiotics are commonly found in soybean and rapeseed meal.
Legumes, cereals and yeast cell walls contain a-galactooligosaccharides (GOS),fructooligosaccharides (FOS) and mannanoligosaccharides (MOS), respectively. Lactobacilli,Bifidobacteria and Eubacteria selectively ferment some prebiotics. Prebiotics modify thegut microbial population balance by promoting the growth of beneficial flora in the intestines,thereby providing a healthier intestinal environment. These are not easily digestible andprovide competitive advantage to favourable bacteria, inhibiting the colonization of harmfulmicrobes by lowering intestinal pH and promoting the beneficial ones. Through a varietyof mechanisms prebiotics are thought to increase resistance to infection. Prebioticsenhance the physical barrier (modulation of paracellular permeability, mucosal trophicaction), improve functional barrier (mucosal immunity) and have a competitive adhesionto epithelial receptors. They increase SCFA production along the gastro-intestinal tract,and induce a shift to a more saccharolytic (carbohydrate fermenting) flora.
Galacto-oligosaccherides (GOS), Mannanoligosaccharides (MOS), Fructo- oligosaccharides(FOS) are frequently used in poultry diets. Harmful bacteria attach themselves to both theFOS and MOS and are excreted. Fructo- oligosaccharides (FOS), a derivative of inulin,inhibit the growth of pathogenic microorganism such as clostridia and salmonella. Increasein egg production and feed efficiency of layer with the use of dietary oligofructose andinulin has been widely reported. Oligosaccharides stimulate the secretion of cytokine andenhance the immune system of the pig to resist pathogenic bacterial challenges.
Probiotics
The live microbial food supplement i.e. probiotics or Direct Fed Microbials (DFM’s), whichwhen fed, improve the intestinal microbial balance of the host. Lactobacilli, Streptococci,Bifido bacteria, Bacillus, Bacteroides, Pediococcus, leuconostoc, Propionibacterium, andsome yeast (Saccharomyces cerevesiae) and fungi (Asperzillus oryzae) are commonlyused DFM’s. B Subtilis and B licheniformis are commonly used in nursery pig rations asthey are spore forming and are able to resist the environmental conditions of hightemperature and moisture occurring during the pelleting process. Probiotics improve thesurvival with better growth, better-feed conversion and inhibition of diarrhea in piglets.
Probiotics can be administered through drinking water and by mixing in the feed. Probioticsshould be given once or twice daily, after which the bacteria should establish itself in thealimentary canal and replace disease-promoting microorganisms. These must be addedto the feed on a daily basis. Use of Probiotics bacterial cultures have greater effect duringthe early stages of growth, when, the gut is sterile especially in pigs and when thealimentary flora are unstable. Probiotics improve health and growth by modifying intestinalmicrobial balance by competitive exclusion thereby increasing uptake of nutrients due toimproved gut permeability.
Some bacterial cultures when fed in single or multiple (few doses) to newly hatched birds,quickly establish as intestinal flora and prevent colonization of pathogenic bacteria. Forexample, Lactobacilli acidophilus produce lactocidin that has antibacterial effects onEscherchia coli. Some preparations are proven effective in protecting chicks from Salmonellainfections and improving weight gain and feed efficiency in chicks and broilers. Probioticsappear to have a more pronounced effect on farms where housing and hygiene are notoptimal. Supplementation of probiotics containing Lactobaccilus acidophilus, Streptococcusfaecium and yeast @ 0.025% in the diets of broilers were found to be beneficial in earlygrowth stage. In broilers supplementation of yeast culture at 0.1 % level increased thebody weight and performance due to quantitative and qualitative alteration in the digestivetract flora with better nutrient utilization.
Dietary supplementation of probiotics has better growth performance with improvement infeed efficiency and low mortality during finishing. In layers, improvement in egg productionand feed efficiency has also been reported. In pigs, the intestinal microflora is capableof resisting the establishment of certain intestinal pathogens.
Organic acids/acidifiers
Organic acids (C-1 to C-7) are widely distributed in nature as normal constituents of plantsor animal tissues. Organic acids posses both the antibacterial and anti mould activities.
They have long been used as preservative to prevent spoilage by checking microbialgrowth and maintain proper gut health. These are very efficacious when their use isadapted to the physiology and anatomy of birds.
Generally two types of acidifiers are used in the feed industry; (i) Feed acidifiers and(ii) Gut acidifiers. Feed acidifiers lower the pH of the feed and inhibit the growth ofpathogenic micro flora. This inhibition reduces the micro flora competing for the hostnutrients and prevents the occurrence of diseases and this results in better growth andperformance. On the other hand gut acidifiers acidify the intestinal tract and modulate theintestine bacterial population in a positive and natural way. Maintenance of healthy gutfor proper productivity is of utmost importance. Amongst various options available topoultry and pig feed industry, short chain fatty acids have shown tremendous promise inmaintaining gut health through their varied modes of action. Antimicrobial activity oforganic acids is related to reduction in pH. Acidifiers maintain an optimum pH in stomach,stimulate feed consumption, improve growth rates, improve feed conversion ratio, inhibitthe growth and colonization of pathogenic bacteria, prevent damage to epithelial cells ofintestines, and reduce microbial competition with host for nutrients. In poultry diets,organic acids are mainly used in order to sanitize the feed to avoid the problems relatedwith salmonella.
State of the organic acids whether un-dissociated or dissociated is extremely importantto define their capacity to inhibit the growth of bacteria. As a general rule, more than tento twenty times the level of dissociated acids to reach the same inhibition of bacteria are National Academy of Agricultural Sciences required as compared to un-dissociated acids. At a pH below 3.0-3.5, almost all organicacids are very efficacious in controlling bacterial growth. The key basic principle on themode of action of organic acids on bacteria is that non-dissociated (non-ionized) organicacids can penetrate the cell wall and disrupt the normal physiology of certain bacteria (E.
coli, Salmonella spp., C. perfringens, Listeria monocytogenes, and Campylobacter spp). Antioxidants
Auto oxidation of nutrients in the body results in the production of free radicals, whichdamage the cellular tissue and cause many disorders. To prevent auto oxidation,antioxidants are frequently used. Nutritional antioxidants are very helpful in reducingphysiological stress both at an organ and cellular level. Feed antioxidants protect nutrientsduring storage, help the absorption of the oxidation sensible substances in the GIT,reduce aging by keeping the membrane intact, while the level of these enzymes decrease.
ß-Carotene, vitamins A, E and C and its calcium and sodium salts, ethoxyquin, lecithin,butylated hydroxytoulene (BHT), propyl gallate, chelated metal ions are commonly usedantioxidants in poultry diets. Beneficial effects of antioxidants are due to their scavengingnature for free radicals; maintain the potency of dietary vitamins and stimulating bird’simmune- responsiveness to infections. Antioxidant defence system includes the enzymessuperoxide dismutase, catalase, and glutathione per oxidase. Many studies have shownthat supplementation of Vitamin A, C & E can attenuate the side effects of extremeenvironmental stress.
Non starch polysaccharides (NSP, cellulose, glucans and xylans etc.) of the cereal grains(wheat, rye, oats) possess antinutritive activity which leads to the formation of viscousgel in the gut that interferes with proper absorption of nutrients and also produces stickydroppings in poultry. Similarly phytic acid and its salts as phytates present in the feedstuffsalso bind minerals, carbohydrates, proteins and form insoluble complexes. These makethe nutrients especially minerals like phosphorus unavailable to the monogastrics and areexcreted in feaces. Supplementation of exogenous enzymes in the diets decreases gutviscosity and improves the availability of nutrients from feed, lowers the feed cost andhelps in reducing the environmental pollution by minimizing the waste excretion. Exogenousenzymes in the diets of young animals complement the endogenous enzymes. Use ofamylase, arabinase, cellulase, glucanase, hemicellulase, pectinase, xylanase, acid andalkali protease, lipases, esterases, phytase and tannase in poultry and pig feed industryhas become a routine. Enzymes in pig and poultry feeds are added to supplement theendogenous enzymes especially to young birds and pigs.
Phytase improves the availability of phytate phosphorus as well as other organic nutrients,performance and mineral retention. Similarly supplementation glycosidase has been foundto increase the energy utilization in birds, diminishing digestive disturbance in weaner pigs. Impact of improvement is more in young pigs. Use of ß glucanase and xylanaseare beneficial with high fiber grains like wheat, barley and their by-products. a - galactosidaseis used to breakdown the galactose units in raffinose and stachyose found in soybean.
The efficacy of enzyme supplementation depends upon types of diet, animals, chemicallinkage in the substrate that needs to be cleaved etc.
Recommendations
There is an urgent need to gather precise information on the use of antibiotics inthe animal husbandry in the country and its potential reservoir in soil and water.
There is need for constant monitoring by compound livestock feed manufacturersassociation (CLFMA), ICAR, Drug controlling agencies for production and distributionof antibiotics for non-therapeutic uses.
Withdrawal of antibiotics all of a sudden from feed will lead to significant reductionin the production performance of live stock, poultry and aquaculture. This can raiseissues of food security. A workable strategy in this context will be to classify theavailable antibiotics into two categories.
Category A: Chloramphenicol, nitrofurans including: furaltadone, furazolidone,furylfuramide, nifuratel, nifuroxime, nifurprazine, nitrofurantoin, nitrofurazone,neomycin, nalidixic acid, sulphamethoxazole, chlorpromazine, colchicines, dapsone,dimetridazole, metronidazole, ronidazole, ipronidazole, other nitroimidazoles,clenbuterol, sulfonamide drugs (except approved sulfadimethoxine,sulfabromomethazine and sulfaethoxypyridazine) and fluroquinolones which are widelyused as therapeutic antibiotics in controlling human and animal bacterial infections,should be reserved for exclusive use in human and animal disease managementonly.
Category B: All other antibiotics including tetracyclines can be listed in the secondcategory of antibiotics, which can be permitted as antimicrobial growth promoters(AGPs). The use of these antibiotics as AGPs should be based on a clear packageof practices indicating maximum permissible concentration in feed, withdrawal timeas well as method of disposal of solid and liquid based including metabolic waste.
Alternative nutritional strategies should be developed for enhanced digestion andabsorption of the ingested feed stuffs. The challenge for live-stock productivity isto find suitable, reliable and cost effective management routines and feed additivesfor a sustainable and successful production.
Once suitable alternatives to AGP’s are developed, use of antibiotics as AGPs inanimal husbandry can be completely banned.
Health and hygiene should be the key to success without AGP’s. Cleaning anddisinfection routines should also be reviewed and upgraded. A failure to replaceAGP’s will result in an increase in adverse intestine-bacteria interactions.
National Academy of Agricultural Sciences Executive Council 2010
President:
Members:
Immediate Past President:
Vice President:
Secretaries:
Foreign Secretaries:
Dr N.S.L. Srivastava (Vallabha Vidyanagar) Editors:
Treasurer
CONVENER
Dr N.S. Pasricha, Former Director, Potash Research Institute of India,
Gurgaon
CO-CONVENER :
Dr S.S. Sikka, Senior Nutritionist Department of Animal Nutrition,
Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
Dr Rajendra Prasad
Dr P.S. Pathak
CITATION
NAAS 2010. “Antibiotics in Manure and Soil – A Grave Threat to Human andAnimal Health”. Policy Paper No.43,National Academy of Agricultural Sciences, New Delhi pp 20.
PARTICIPANTS
Abrol, Y.P., Former Professor Plant Physiology, IARI, New Delhi.
Aggarwal, P.K., Secretary NAAS, National Professor, Division of Environmental Sciences, IARI. NewDelhi.
Bhatia, S.K. Professor Animal Nutrition (Retd.), HAU, Hisar.
Handa, S.K., Former Project Co-coordinator, AICRI on Pesticide Residues, Indian Agricultural ResearchInstitute, New Delhi.
Jaswinder Singh, District Extension Specialist, Department of Veterinary and Animal HusbandryExtension, Guru Angad Dev Veterinary and Animal Sciences University (GADVASU), Ludhiana.
Kulshrestha, Gita, Former Head, Division of Agricultural Chemicals, IARI, New Delhi Kundu S.S., Principal scientist, National Dairy Research Institute Karnal.
Mandal, A.B., Principal Scientist, Nutrition & Feed Technology Division, CARI, Izat Nagar.
Mukundan, M.K., Professor, Central Institute of Fisheries Technology, Cochin.
Parmar, B.S., Former Joint Director Research IARI, New Delhi Parminder Singh, Associate Prof. Extension (Nutrition), Department of Veterinary and Animal HusbandryExtension, Guru Angad Dev Veterinary and Animal Sciences University (GADVASU), Ludhiana.
Parthasarathy, M., Professor Animal Nutrition, Department of Animal Nutrition, College of VeterinaryScience, Sri Venkateswara Veterinary University, Tirupati.
Pasricha, N.S., Former Director, Potash Research Institute of India, Gurgaon, Haryana Pattanaik, A. K., Senior Scientist, CAS in Animal Nutrition, IVRI, Izat Nagar.
Sachdev, M.S., Principal Scientist, Nuclear Research Laboratory, IARI, New Delhi.
Sikka. S.S., Senior Nutritionist, Department of Animal Nutrition, Guru Angad Dev Veterinary andAnimal Sciences University (GADVASU), Ludhiana.
Singh, Shashi Bala., Principal Scientist, Division of Agricultural Chemicals, IARI, New Delhi.
Tyagi J.S., Senior Scientist, Division of Physiology and Reproduction, Central Avian ResearchInstitute, Izatnagar.
Published by Shri H.C. Pathak, Executive Secretary on behalf of NATIONAL ACADEMY OF AGRICULTURAL SCIENCES
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