The Silent Killer
Antibiotics are failing because bacteria are becoming increasingly resistant to them. The world is on the cusp of a “post-antibiotic era” as the alarming rate of resistant strain emergence far outpaces our ability to develop the drugs that are urgently needed to combat them. In the past half-century, only two completely new classes of antibiotics have reached the market. The last of these was in 1987, and since then many bacterial strains have adapted to become almost, and in some cases completely, impervious to our dwindling supply of drugs.
If we fail to address this problem quickly and comprehensively then antimicrobial resistance (AMR) will make providing quality universal healthcare coverage much more difficult, if not impossible. This is because antibiotics underpin modern medicine in all its finery. Without them, the practice of medicine itself will become much more hazardous: routine medical procedures such as minor surgeries, cancer treatments and even child birth will become life-threatening.
AMR is not only a threat to the future of medicine, it is already a significant global public health concern. Each year, antibiotic resistant infections cause more than 700,000 deaths, and this number is expected to rise dramatically over the coming years [ReAct, 2018]. Even more worryingly, pan-drug resistant superbugs have already emerged. For example, in a Nevada hospital in 2016, a patient infected with an imported superbug entered septic shock and died despite having received treatment comprising all 26 antibiotics in the US medicine cabinet. Unfortunately, stories like this, almost unthinkable in the past, will become more common as antimicrobial resistance continues to develop at breakneck speed. Life expectancy in the UK has recently fallen for the first time because the Office for National Statistics fears the “re-emergence of existing diseases and increases in antimicrobial resistance”. This contrasts with the period between 1945 and 1972, when the introduction of antibiotics to treat infections that were previously considered life-threatening contributed to an eight-year increase in average life expectancy.
The UK’s Chief Medical Officer, Professor Dame Sally Davies, has issued a stark warning:
“If we do not act now, any one of us could go into hospital for minor surgery in 20 years and die because of an ordinary infection that can’t be treated by antibiotics.”
She has also emphasized the importance of improving hygiene:
“We need to address the growing problem of drug-resistant infections as the global medicine cabinet is becoming increasingly bare … Preventing infections in the first place is key.”
AMR is a complex global problem which requires international cooperation. In 2015, the UN Secretary General Ban Ki-moon described AMR as a “fundamental threat to human health” at the first General Assembly meeting on antibiotic resistant bacteria. This was only the fourth time that the General Assembly has ever held a high-level meeting for a health issue, and all 193 Member States signed a declaration to combat the proliferation of AMR. This Accord routes the global response to superbugs along a similar path to the one used to combat climate change. Improving hospital hygiene and antibiotic stewardship programs to prevent the spread of such infections, and conserving existing antibiotics for when they are most acutely needed, are some of its key objectives.
This is because many of the bacteria that are causing the biggest resistance concerns are also responsible for increasingly difficult to treat hospital-acquired infections (HAIs). HAIs arise in healthcare facilities and are often transmitted within them due to failures in hygiene practices. The gathering storm that is forming from the combination of increasing HAI incidence and the rising prevalence of multi-drug resistant strains, has transformed a once commonly accepted need to improve hospital infection control programs into an urgent global healthcare priority. This places a great deal of responsibility upon the shoulders of the clinicians leading these programs. If they are adequately armed to rise to the challenge of AMR then the battle for the future of medicine can be won. In an age in which antibiotics are all too often rendered completely ineffective against antibiotic resistant superbugs, the old adage “Prevention is better than cure” has seldom rung truer.
AMR is spreading
More than 78% of the populations in some areas carry multidrug-resistant bacteria in their normal bacterial flora [Monira et al, 2017] and as the world is becoming more globalized and interconnected, the spread of AMR is accelerating.
AMR is expensive
By 2050, AMR will add $1.2 trillion per year in healthcare costs globally, and cause the world economy to decline by 3.8% [World Bank] – an economic impact much more devastating than the 2008/2009 global financial crisis.
By 2050, the world population will be up to 444 million persons lower than it would otherwise be if we fail to tackle AMR [Rand, 2018]. Every year, an estimated 214,000 newborns die from blood infections (sepsis) caused by antibiotic resistant bacteria [Sankar et al., 2016].
What are antibiotics and how do they work?
Antibiotics are a type of antimicrobial drug used to treat and prevent bacterial infections. Bacterial cells differ from our own, which has allowed us to take advantage of these differences by disrupting the biological mechanisms that are essential for their survival. Antibiotics are therefore able to prevent the growth of, or kill, bacterial cells while minimizing harm to our own bodies.
Bacterial species are immensely diverse but can be broadly categorised into two groups: gram-negative and gram-positive. Gram-negative bacteria have two cell membranes separated by a thin layer of peptidoglycan (cell wall); gram-positive bacteria do not have an outer cell membrane and instead have a thicker peptidoglycan cell wall surrounding them.
Narrow-spectrum antibiotics only show activity against a small range of bacterial species, whereas broad-spectrum antibiotics act on both gram-negative and gram-positive bacteria. This makes broad-spectrum antibiotics powerful and flexible drugs that can used to treat bacterial infections when the infecting bacterium has not yet been identified or remains unknown, or as a prophylactic measure following surgery. Unfortunately, the same characteristics that make broad-spectrum antibiotics so clinically useful are also their Achilles heel: they attack the normal flora of healthy bacteria that inhabit the body as well as disease-causing bacteria. Because they act non-specifically on a wide range of bacteria they create selection pressures for the consequential development and transmission of antibiotic resistance.
In 1928, biologist Alexander Fleming was sorting through some petri dishes that were growing bacteria and noted an unusual colony of mold that had grown on one of the plates. The unexpected mold colony was somehow preventing bacteria from growing around it in a clear ring. Fleming identified the mold as a rare strain of Penicillium notatum and had in fact discovered the first antibiotic, penicillin.Fleming’s discovery of penicillin heralded the arrival of the golden age of antibiotic discovery. These miracle drugs significantly reduced mortality rates worldwide and contributed to a longer and better quality of life.
However, Fleming foresaw the danger of resistance developing to these drugs
“The time may come when penicillin can be bought by anyone in the shops. Then there is the danger that the ignorant man may easily underdose himself and by exposing his microbes to non-lethal quantities of the drug make them resistant.” (December 11th, 1945).
Indeed, the global optimism surrounding antibiotic wonder drugs didn’t last. Infectious diseases are becoming ever more difficult to treat and cure, and the last few decades have shown that we have to continue to battle old diseases even as new infections emerge.
|Target||Family||Mode of Action||Examples and Uses|
|Cell Wall||Penicillins||Penicillins and cephalosporins are the major β-lactam antibiotics, which act by inhibiting the synthesis of bacterial cell walls.Penicillins and cephalosporins are the major β-lactam antibiotics, which act by inhibiting the synthesis of bacterial cell walls.Most bacteria have a cell wall built from peptidoglycan. Humans do not use peptidoglycan, making it an incredibly useful antibiotic target. Penicillins and cephalosporins interfere with the last step in peptidoglycan synthesis in bacteria by mimicking one of its constituents. The enzymes involved in this final reaction react with the β-lactam antibiotics due to their similar shape, inactivating the reaction and resulting in the formation of peptidoglycan chains that are not cross-linked and lack strength.||Penicillin G|
Pneumonia, strep throat, syphilis, necrotizing enterocolitis, diphtheria, gas gangrene, leptospirosis, cellulitis, and tetanus
Ear and bladder infections, pneumonia, gonorrhea, E. coli and Salmonella
|Cephalosporins||Weak points develop in the growing cell wall, which results in the cell rupturing. In addition to cell walls, there are more targets for the action of penicillins, collectively termed penicillin-binding proteins (PBP), that all bacteria possess.||Cephalexin|
Upper respiratory, ear, skin and urinary tract infections
|Protein Synthesis||Macrolides||Macrolides comprise a 12- to 16-membered ring. Macrolides inhibit protein synthesis by different mechanisms, depending on their structure. These include:– Blocking elongation of protein chains as they are created by binding to ribosomes and blocking their exit– Peptidyl tRNA dissociation (tRNAs are small molecules that guide the right amino acid to the growing peptide chain in the ribosome)- Blocking Peptidyl Transferase (the enzyme that joins amino acids in a protein chain)– Preventing Ribosomal Assembly||Erythromycin|
Respiratory tract infections, skin infections, chlamydia infections, pelvic inflammatory disease, and syphilis
Skin and respiratory tract infections. It can also be used in conjunction with other drugs to treat stomach ulcers caused by Helicobacter pylori
Skin, respiratory, ear infections and sexually-transmitted infections
|Tetracyclines||Tetracyclines consist of 4 fused cyclic rings and only vary in minor alterations of the chemical groups attached to this structure. Tetracyclines bind to part of the protein synthesis machinery within bacterial cells called ribosomes and interrupt the structure needed to commence protein synthesis.These bacteria therefore cannot synthesize the proteins necessary to survive, resulting in cell death.||Tetracycline|
Urinary tract, acne, gonorrhea, chlamydia infections
Urinary tract, acne, gonorrhea, chlamydia, periodontitis infections
|Aminoglycosides||Weak points develop in the growing cell wall, which results in the cell rupturing. In addition to cell walls, there are more targets for the action of penicillins, collectively termed penicillin-binding proteins (PBP), that all bacteria possess.||Gentamicin|
Used to treat severe or serious bacterial infections
Injections used to treat skin, heart, stomach, brain, spinal, respiratory and urinary tract infections. Also used to treat cystic fibrosis
|DNA Replication||Fluoroquinolones||Nearly all quinolone antibiotics used are fluoroquinolones, which contain a fluorine atom in their chemical structure. They are effective against both gram-negative and gram-positive bacteria.Fluoroquinolones share a bicyclic core structure and act by inhibiting enzymes involved in DNA replication. Without the synthesis of new DNA, bacterial cells cannot replicate and divide, causing death by bacteriostatic action.||Levofloxacin|
Skin, sinus, kidney, bladder or prostate infections. It can also be used to treat bacterial infections that cause bronchitis or pneumonia, or those who have been exposed to anthrax or plague
Bone and joint infections, intra-abdominal infections, certain type of infectious diarrhea, respiratory tract infections
Bronchitis, pneumonia, chlamydia, gonorrhea, skin, urinary tract and prostate infections
|Cellular Metabolism||Sulfonamides||Sulfonamide is the basis of several groups of drugs. Antibacterial sulfonamides are a group of antibiotic agents that contain the sulfonamide group.Antibacterial sulfonamides inhibit an enzyme involved in the biosynthesis of folate (vitamin B9). Sulfonamides do this by mimicking natural substrates and reacting with them. This forms a ‘pseudo-metabolite’ that is also reactive and antibacterial.||Co-trimoxazole|
Mixture of trimethoprim and sulfamethoxazole antibiotics, used to treat ear and urinary tract infections, bronchitis, traveller’s diarrhea, shigellosis and Pneumocystis jiroveci
Bladder, kidney and ear infections
|Cellular Membrane||Polymyxins||Polymyxins are a group or last-resort antibiotics, typically used in the treatment of multidrug-resistant gram-negative bacterial infections.Polymyxins bind to a molecule called lipopolysaccharide in the outer membrane of gram-negative bacteria and disrupt both the outer and inner membranes. Like detergents, these antibiotics are thought to displace membrane lipids and cause cell lysis.||Colistin|
Colistin is a decades-old drug that is rarely used in medicine due to its kidney toxicity. It remains one of the last-resort antibiotics for multidrug-resistant bacteria, such as Pseudomonas aeruginosa, Klebsiella pneumoniae, and Acinetobacter.
How does antibiotic resistance occur?
AMR is the ability of microorganisms to overcome the toxic effects of an antimicrobial agent. AMR is a natural phenomenon, occurring as a result of the extraordinary evolutionary capacity of bacteria to adapt and survive. Resistance often occurs as a result of genetic mutation following exposure to an antibiotic. Bacteria also have the ability to transfer and receive mobile genetic elements (MGEs) that can carry a range of resistance genes. This promotes both inter- and intraspecies transfer of resistance genes.
Due to the fluid genetic makeup of bacteria, MGEs can quickly contribute to multi-drug resistance in single bacterial species. Recognizing the increasing severity of resistance, the terms extensively drug resistant and pandrug-resistant (impervious to all antibiotics) have been introduced and are documented with increasing frequency.
Carbapenems are some of the most powerful antibiotics in clinical use. They are used as a last resort for many bacterial infections, such as E. coli. However, some bacteria now carry the NDM-1 gene which confers resistance to carbapenems. Bacteria carrying NDM-1 genes are some of the most dangerous superbugs in existence. First isolated in India in 2008, NDM-1 has since been detected in more than 70 countries worldwide. If NDM-1 is acquired by other bacterial species, it could lead to a health crisis in itself.
Globalization means that we are now simultaneously exposing ourselves to previously hard-to-reach human, animal and bacterial populations, triggering an explosion in the global spread of infectious diseases and bacterial resistance.
Bacterial resistance mechanisms
1. Enzymatic inhibition
This is the most common mechanism of resistance employed by bacteria. Enzymatic inhibition occurs when bacteria develop enzymes to chemically modify the structure of antibacterial compounds and render them ineffective. β-lactamases are an enzyme family that poses a major problem in the treatment of gram-negative bacteria and can confer resistance against penicillin antibiotics.
NDM-1 (New Delhi metallo-beta-lactamase 1) is an enzyme that makes bacteria resistant to a broad range of beta-lactam antibiotics, including the carbapenem family, which are a cornerstone for the treatment of antibiotic resistant bacterial infections. The most common bacteria that make this enzyme are gram-negatives such as Escherichia coli and Klebsiella pneumoniae, but the gene for NDM-1 can easily spread from one strain of bacteria to another.
2. PBP modifications
Penicillin-binding proteins (PBPs) are key proteins involved in the construction of bacterial cell walls. These proteins are essential to the assembly of peptidoglycan, the major constituent of cell walls. β-lactam antibiotics mimic a part of peptidoglycan to fit into the active site of PBPs and form a tight complex that deactivates them.
The mecA gene is an example of a Penicillin-binding protein that provides bacteria with resistance to antibiotics such as methicillin, penicillin and other penicillin-like antibiotics. The gene encodes a protein called Penicillin Binding Protein 2A that does not allow the ring-like structure of penicillin antibiotics to bind to the enzymes that help form the cell wall of bacteria, allowing the bacteria to replicate as normal.
3. Porin mutations
The main constituent in the outer bilayer of gram-negative bacteria is lipopolysaccharide, a highly hydrophobic (water-repelling) lipid that makes the passage of hydrophilic (water-attracting) compounds difficult. Porins are proteins that span the outer-membrane of gram-negative bacteria to allow the allow the passage of compounds into and out of the cell. These porins can mutate to become selective against antibiotics or can be expressed in lower concentrations to reduce uptake to below minimum inhibitory concentration levels of antibiotics.
The OmpF porin confers general antibiotic resistance to a range of gram-negative bacteria by preventing the influx of antibiotics. The gene is widespread and is found in E. coli, Enterobacter and Salmonella, with the ability to prevent entry of β-lactam, carbapenem and chloramphenicol antibiotics into bacterial cells.
4. Cell wall modifications
Most antibiotics target the ribosome, the protein synthesis machinery of bacterial cells, necessary for replication. Antibiotics that target this machinery are potent inhibitors of bacterial growth. However, over decades of clinical use pathogens have developed resistance to inhibitors of protein synthesis. Bacteria often modify the amino acid sequences of antibiotic targets through point mutations that result in rapid and easy resistance, with minimized impact on the microbe’s fitness.
A well-characterized example of antibiotic resistance by these mechanisms are oxazolidinones. These drugs are synthetic bacteriostatic antibiotics with broad gram-positive activity that exert their mechanism by interaction with bacterial ribosomes and blocking synthesis. The most commonly characterized mechanisms of linezolid resistance include mutations in genes encoding the ribosomal proteins L3 and L4, which prevent linezolid and tedizolid from binding and disrupting protein synthesis.
5. Efflux pumps
Bacteria use membrane-spanning proteins, known as efflux pumps, to actively eliminate substances from the cell. Efflux pumps can be used to eliminate specific antimicrobial compounds and higher levels of generic efflux pumps can be expressed to ensure antibiotics are below their minimum inhibitory concentration. There are a wide range of efflux pumps that can simultaneously confer multi-drug resistance to a variety of antibiotics.
Tetracycline resistance is a classic example of efflux-mediated resistance, where tet genes encode efflux pumps that actively extrude tetracyclines from the cell. Currently, more than 20 different tet genes have been described, most of which are harbored in mobile genetic elements. The majority of these genes are found in gram-negative bacteria, such as E. coli.
Antibiotic resistant bacteria
The grave threat of spreading multidrug and pandrug-resistance has led the World Health Organization to develop a growing list of bacteria that are most concerning to global human health. These organisms are prioritized into three categories:
Carbapenem-resistant Pseudomonas aeruginosa,
Vancomycin-resistant Staphylococcus aureus,
Methicillin-resistant, vancomycin-intermediate and resistant Helicobacter pylori,
Clarithromycin-resistant Campylobacter spp.,
Fluoroquinolone-resistant Neisseria gonorrhoeae
Penicillin-non-susceptible Haemophilus influenzae,
Ampicillin-resistant Shigella spp.
‘Medium’ priority organisms already cause severe bacterial infections worldwide. Haemophilus influenzae, for example, can cause a multitude of different infections in the human body, including meningitis, which kills 1 in 20 children it infects, and those that survive may suffer long-term problems, such as hearing loss, seizures and learning disabilities.
Antibiotic overuse in healthcare and agriculture
Humans are playing a crucial role in the development and spread of antibiotic resistant microorganisms. Consistent antibiotic overuse and inappropriate prescriptions, coupled with high density and turnover of patients, has made hospitals a key battleground in the fight against resistance. A 2016 CDC report estimates that one in every three antibiotic prescriptions are unnecessary [CDC, 2016], or 20,000 per day in the NHS by General Practitioners in the UK [GPOnline, 2018]. This is further exacerbated by the fact that when bacteria develop resistance to one drug, such as penicillin, the changes in the bacteria’s biology often make it impervious to a range of other antibiotics too.
The overconsumption of antibiotics, both in humans and in agriculture, is a global concern. Like humans, animals are susceptible to infectious diseases. To combat this, antibiotics are used therapeutically to treat individual animals with clinical diseases. However, antibiotics are often administered to livestock in feed to prophylactically prevent infections and marginally improve growth. As a result of this, some animals receive a sub-therapeutic dose, creating the perfect conditions for the emergence of antibiotic resistant bacteria, where susceptible bacteria are killed off while allowing selected bacteria to survive.
Bacteria live in a plethora of environments where they are regularly exposed to sub-lethal doses of antibiotics, and resistant populations can emerge in a matter of hours. In a mixed culture of bacteria, there might be thousands of bacteria that are susceptible to antibiotics, but if just one antibiotic resistant bacterial cell has the competitive advantage of a resistance gene then it will rapidly proliferate.
What is so alarming about this approach to agriculture is that many of the antibiotics used to treat animals are identical to human medicines. Unsurprisingly, there is clear correlation between antibiotic resistance in humans and the widespread use of non-therapeutic antibiotics in animals [O’Neill, 2015]. Antibiotic resistant bacteria are transmitted to humans through direct contact with animals, animal manure, consumption of undercooked meat and contact with uncooked meat.
Hospital acquired infections: Prevention is better than cure
Many of the bacteria that are causing the biggest resistance concerns among the scientific and medical community are also responsible for hospital-acquired infections (HAIs), or nosocomial infections (infections that are contracted in hospital or healthcare facilities). HAIs are the most frequent adverse events in the delivery of care worldwide and are a leading cause of death in the developed world [WHO]. It is estimated that in the US and UK alone, HAIs contribute to approximately 100,000 and 20,000 deaths each year, respectively.
Surfaces are rapidly contaminated with harmful microbes, and environmental contamination is often the root cause of infections spreading from person to person in the hospital environment. For example, up to 60% of surfaces in the patient care zone are contaminated with pathogens known to cause HAIs. These pathogens persist for months on surfaces, facilitating cross-contamination and subsequently infection transmission between healthcare professionals (HCPs) and patients.
Many HAIs are preventable and result from failures in hygiene that cause the needless suffering of patients. Preventing the occurrence and spread of hospital acquired infections obviously saves lives, but perhaps more importantly, reducing infection incidence also leads to fewer antibiotic treatments, which in turn decreases the likelihood of the development of antibiotic resistant strains. Conserving antibiotics in this manner buys medicine time as it awaits the discovery of the new antibiotics that are urgently needed to replenish a presently diminishing drug arsenal.
Antibiotic stewardship program
An antibiotic stewardship program is a system-wide approach in healthcare facilities to promote and monitor the use of antimicrobials to preserve their effectiveness [NICE, 2015]. These coordinated programs act to promote the appropriate use of antimicrobials, improve patient outcomes, reduce antimicrobial resistance, and decrease the spread of infections caused by multidrug-resistant organisms. Antibiotic stewardship programs are gaining global momentum as the issue of antibiotic resistance becomes a worldwide challenge. Effective programs improve the quality of patient care and have been shown to be significantly more cost effective, with reductions in antimicrobial use saving an average $200,000-900,000 per antibiotic stewardship program, per year [Kon & Rai, 2016].
An antibiotic stewardship program may feature any of the following elements:
- Prospective auditing with feedback
48-hour antibiotic time-out
- Physician feedback
- Dosing optimization
- Rapid diagnostics with antibiotic stewardship intervention
- Order sets
- Decision support for antibiotic time-out
- Formulary restriction
- Institution-wide conferences
Monitor for adverse events
Intravenous to oral conversion
Electronic order forms
Drug-bug mismatch decisions support
Monitor antimicrobial utilization
Ongoing education is a crucial element of an antibiotic stewardship program, such as educational conferences, written guidelines and teaching seminars. Continued education by infection preventionists is vital in keeping up-to-date with emergent pathogens, new guidelines and developing technologies. As antibiotic stewardship programs move away from cost-cutting strategies and towards quality and patient safety, adoption of innovative technologies is becoming ever-more important in ensuring best practice.
The Antibiotic Pipeline
A completely new class of antibiotic hasn’t been discovered since 1987, even though pathogenic bacteria are becoming increasingly resistant to our dwindling stock. Over the last decade, there has been an alarming increase in the number of resistant infections that can no longer be cured, and reports of superbug-related deaths have become common.
The costs to develop a new antibiotic drug are no less expensive compared to development of drugs for other therapeutic areas, yet the commercial potential and return on investment for companies developing new antibiotics are significantly lower than drugs to treat chronic conditions such as diabetes or heart disease. This has created an unwillingness for pharmaceutical companies to invest in much-needed research and development of new antibiotic classes. The problem for drugmakers is that new antibiotics are usually held in reserve and are not used unless they’re needed because patients develop resistance to an older medicine. Even the most expensive antibiotics, at around $1,000 a day, are cheap compared with a cancer medicine that will be given for months instead of a few days or weeks.
The other major issue in antibiotic discovery is target selection. Much of the potency of effective antibiotic treatments is due to the exclusivity of the bacterial mechanisms that they target. Targeting the membranes or distinct cellular machinery of bacteria ensures minimal damage to our own cells, but as resistance has developed and we have exhausted these exclusive targets over time, newer antibiotics are more harmful to our bodies and less effective at fighting bacteria. For example, the FDA is warning against fluoroquinolone antibiotics, which may cause sudden, serious, and potentially permanent nerve damage called peripheral neuropathy [FDA, 2016].
As a society, the need to cooperatively fight the immense threat of AMR is evident. We face a collective struggle, and while the odds are now stacked against us, taking immediate measures to combat this threat may allow us to safeguard healthcare for future generations.
A global response to the chronic shortfall in antibiotic innovation is urgently needed to combat antimicrobial resistance. At Hygenica, we are totally committed to combating this global threat. Our mission is to ensure that our innovative antimicrobial technology is appropriately adopted in clinical practice, to protect patients throughout the world from contracting unnecessary HAIs and reduce the development of AMR.
We are advancing an expanding range of products which are highly effective at killing a broad spectrum of harmful pathogens. When integrated within hospital-wide infection control programs, our disruptive antimicrobial technology helps to prevent the transmission of HAIs which emanate from cross-contamination at high-frequency touchpoint surfaces.
- Bowater: http://pubs.rsc.org/en/content/ebook/978-1-78262-167-6
Kon & Rai: https://www.elsevier.com/books/antibiotic-resistance/kon/978-0-12-803642-6
Monira et al.: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5399343/
O’Neill: https://amr-review.org/sites/default/files/Antimicrobials in agriculture
Sankar et al.: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4848744/
WHO (2016): http://www.who.int/bulletin/volumes/94/9/16-020916.pdf
World Bank: http://documents.worldbank.org/curated/en/323311493396993758/pdf/11467 9-REVISED-v2-Drug-Resistant-Infections-Final-Report.pdf