Team:IISER-Tirupati India/Inspiration


1.1 Antimicrobial Resistance (AMR): A global concern

The discovery of antibiotics in the 1940s revolutionized medical care and positively impacted human and animal health. Antimicrobials help treat and prevent serious infections. When microorganisms start to develop the ability to resist these antimicrobials and continue to proliferate, then AMR arises. Resistance development in microorganisms is an evolutionary process; however, it is accelerated by the selective pressure exerted by the misuse of antibiotics in both humans and animals, eventually favouring the selection and spread of resistant bacteria. As a result, the medicines become ineffective, and infections persist, increasing the risk of spread to others.
According to the United Nations, by the year 2050, AMR alone could cause 10 million deaths globally; thus, it currently poses the most serious health threat. This also means that progress in modern medicine, which relies on the availability of effective antibacterial drugs, is now at risk. Though there are a lot of information gaps on the extent of AMR, and on the types and number of infections caused by bacteria that have become resistant to antibacterial drugs, it is clear that AMR is a concern that needs immediate attention.
With the current COVID-19 pandemic situation, there is a huge potential that certain activities could drive antimicrobial resistance even further. Increase in the number of patients in hospitals increases the risk of healthcare-associated infections and the transmission of multidrug-resistant organisms which in turn lead to increased antimicrobial use [9]. Disruptions to health services during the pandemic cause interruptions to treatments, such as for tuberculosis and human immunodeficiency virus, which can also lead to selection for drug resistance. Similarly, disruption to vaccination services can increase the risk of infection, potentially leading to an overuse of antimicrobials. Another potential threat is the wide use of biocidal agents for environmental and personal disinfection, including in non-health-care settings. Low-level exposure to biocidal agents can select for drug-resistant strains and enhance the risk of cross-resistance to antibiotics [10], particularly those that treat Gram-negative bacteria [11].

1.2 AMR in India

AMR emergence in India is majorly driven by factors like the high burden of bacterial infections, poor sanitary, hygiene conditions, and the increasing proportion of intensive animal farming, and this is ramped by the unregulated access to antibiotics, including sale without prescription or with an invalid prescription. The use of antibiotics as growth promoters in animal husbandry is a common practice and this, in turn, affects human health, as antibiotic-resistant bacteria can be transmitted between humans and animals through contact, food products, and from the environment [7]. In 2010 it was estimated that India was the fifth-largest consumer of antibiotics in food animals (poultry, pigs, and cattle), after China, the United States, Brazil, and Germany based on livestock density [5].

Module 1

Fig 1: Number of formulation companies manufacturing various antibiotics for animal use. This shows sulfonamides are one amongst the highly produced antibiotics in India [6].

StudyYear(s) data collection and stateSpecimenOrganismFindings
Brower et al. 20172014 PunjabCloacal swab samples Broilers (n=270) Layers (n=260)Not applicableESBL producing strains (%) Broilers: 87% of cloacal swabs Layers: 42% of cloacal swabs
Shrivastav et al. 20162015 Madhya PradeshCecal swabsE. coli (n=400)ESBL producers (%) Broilers: 33.5%
Kar et al. 20152013-2014 OdishaFeeal sampleE. coli (n=170)ESBL producers (%) Poultry: 9.4%
Naik et al. 20152013-2014 ChattisgarhChicken meat samples (n=200)Salmonella species (n=14)Prevalence of Salmonella: 7% Resistance % Ciprofloxacin:o9 Erythromycin:100% Oxytetracyeline: 42.8%
Kaushik et al. 20142010-2013 BiharChicken meat samples (n=228)Salmonella species (n=54)Prevalence of Salmonella: 23.7% 100% resistance AmpicillinGentamicin Highly sensitive Ceftriaxone Azithromycin Moderately sensitive Ciprofloxacin Tetracyeline
Samanta et al. 2014Year not mentioned West BengalCloacal samples, eggs and environment samples of backyard poultry flocks (n=360)Salmonella species (n=22)Prevalence of Salmonella: 6.1% Resistance % Ciprofloxacin: 100% Gentamicin: 100% Tetracycline:1009 Ceftriaxone:
Singh et al. 2013Year not mentioned Uttar PradeshCloacal samples, eggs and environment samples (n=720)Salmonella species (n=26)Prevalence of Salmonella- 3.3% Resistance% Ampicillin: 0% Ciprofloxacin: 11.5% Gentamicin: 7.7% Tetracycline:23.1%

Table 1: Antibiotic resistance in poultry in various studies in India [6]

In the studies conducted involving farms, it was observed that there was high resistance to the commonly used antibiotics including sulfonamides, ciprofloxacin, β-lactams, and tetracycline [6]. However, the data available for India is still quite inadequate and there is a lot of information gap that needs to be bridged with the help of surveillance and studies. From the data available from other nations it was noted that agricultural waste contains high levels of antibiotics. This is because of high consumption and incomplete metabolism in the animal bodies [8].

Fig 2: The graph above depicts the detection of antibiotics in poultry (“Poultry Litter”), swine (“Swine Manure”), and beef cattle (“Cattle Manure”) manure. Antibiotic class codes on the y-axis are as follows: MC macrolide, LM lincosamide, TM trimethoprim, TC tetracycline, SA sulfonamide, PP polypeptide, FQ fluoroquinolone, COC coccidiostat, BL beta-lactam [8].

From Table 2, fluoroquinolones, sulfonamides, and tetracyclines were found consistently in all the litters (poultry, swine, cattle) across different countries. All of these have been listed by WHO as critically important for human health. The relatively low concentrations of beta-lactams and polypeptides were due to their ability to get readily metabolized. This shows that the previously mentioned classes of antibiotics are of greater concern and there is a need to design and test treatment technologies that ensure degradation of antimicrobials in animal waste before use as fertilizers and soil amendments [8].

Table 2
Sample typeSource of SampleAntiblotic classMaximum Detected RangeRemarks
Poultry LitterChina, Egypt, and AustriaEnrofloxacin (fluoroquinolone)1421mg/kg. 31mg/kg and 8mg/kg 
Poultry litterChinaSulfonamides and sulfadiazine6mg/kg and 51mg/kg 
Poultry LitterUSATetracyclines66 mg/kgAlso, in Austria, China and Egypt tetracycline residues were detected
Swine LitterUSAsulfamethoxazole and trimethoprim400 ug/L and 2.5 ug/L 
Swine LitterUSAMacrolides (i.e., erythromycin and tylosin)0.001 to 10 mg/L 
Swine litterAustria China, Germany, Switzerland and the USAAll classes (fluoroquinolone sulfonamide and tetracycline)0.01 and 100 mg/kg (or mg/L).Tetracycline antibiotics and key metabolic products. have been widely reported in swine manure with detection frequencies as high as 73 % in Austria
Cattle LitterUSAAll classes (fluoroquinolone sulfonamide and tetracycline)The range observed for fluoroquinolones and tetracyclines was 0.1 to 100 mg/kg. but sulfonamide levels were lower. 
Cattle litterChinaOxytetracycline, enrofloxacin, ciprofloxacin and chlortetracycline59.59mg/kg. 46.70 mg/kg. 29.59 mg/kg and 27.59 mg/kg

1.3 Our Solution Coli Kaze

Based on all the above data, to address this issue of antibiotic pollution which is causing AMR, we designed a synthetic biology-based solution. Our genetically engineered bacteria Coli Kaze would reduce the excessive efflux of antibiotics that are present in animal waste. Following degradation, our bacteria would undergo self DNA degradation and ultimately die. We would then be left with excreta free of both the antibiotics as well our engineered bacteria and could then be processed and used as safe manure. For proof of concept, we will initially target sulfonamides present in the poultry waste. However, we plan to extend it to other antibiotics in the future. For the implementation of our project, we are planning to set up a treatment tank on farms. This tank will have 2 different chambers. The topmost chamber, that is, the pre-treatment tank is the place where we would grow our Coli Kaze bacteria in culture media to an extent that would have produced enough antibiotic degrading enzymes. We will then induce the kill switch in this tank itself so that our bacteria would die by cell lysis, that is, by breaking down, releasing all the antibiotic degrading enzymes into the second chamber.
The second chamber would be the digester tank. Here, the slurry of the excreta contaminated with antibiotics (Sulfonamides, in this case) would be incubated with the enzymes released from chamber 1 until the antibiotics are degraded.
After the above process, cell lysate and enzymes could be found in the supernatant. Also to keep the temperature stable and suitable for enzyme action, the tank would be partially buried underground. To know more about our project and its modules please visit the description page.


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