̽��ֱ�� of Cambridge - Lalita Ramakrishnan

Rare genetic disease may protect Ashkenazi Jews against TB

cjb250 — Mon, 06 Feb 2023 20:00:02 +0000

In research published today in Proceedings of the National Academy of Sciences (PNAS), Cambridge scientists – with colleagues in the Netherlands, Spain, and Pennsylvania, USA – show that the same biological mechanisms that underlie Gaucher disease are also effective at clearing TB infection.

̽��ֱ��discovery – made while studying TB susceptibility in zebrafish – suggests that genetic variants that increase the risk of Gaucher also help protect against TB, giving them a selective advantage – that is, making the variants more likely to be passed down from generation to generation.

In 2021, an estimated 10.6 million people worldwide fell ill with TB and 1.6 million people died from the disease. Most people manage to clear the infection themselves, however – only around one in 10 to 20 people will go on to develop the disease.

Professor Lalita Ramakrishnan and colleagues from the ̽��ֱ�� of Cambridge and the Medical Research Council Laboratory of Molecular Biology, Cambridge, are interested in what makes some people susceptible to TB while others appear to be protected. She uses zebrafish to model human disease as it is relatively easy to manipulate zebrafish’s genetics, and their immune systems share many similarities with those of humans.

During their research, her team had previously found that zebrafish with mutations that impaired the digestion of proteins by lysosomes became more susceptible to TB. Lysosomes are components of our cells that break down unwanted materials, including proteins and fats, using enzymes. When a mutation affects the production of these enzymes, it can lead to a build-up of toxic materials.

One type of cell that is vulnerable to this build-up is the macrophage, a type of immune cell that ‘eats’ toxic material, including bacteria and waste products. In lysosomal disorders, the macrophages become enlarged because of accumulation of undigested material in their lysosomes and move slowly, hampering their ability to fight infection.

Professor Ramakrishnan said: “Macrophages need to move quickly to attack invading bacteria and viruses. Their name means ‘big eater’, and this is exactly what they do. But with lysosomal disorders, they’re unable to break down the food they eat, which makes them bloated and sluggish, unable to perform their duties.”

However, when Ramakrishnan and colleagues modelled a lysosomal storage disease known as Gaucher disease, they found something very unexpected: TB resistance rather than susceptibility.

Gaucher disease is a rare disease, affecting around one in 40,000 to 60,000 births in the general population, but rates are significantly higher among Ashkenazi Jews – around one in 800 births. In most cases, the disease can be relatively mild – with symptoms including enlarged spleen and liver, and anaemia – and around two-thirds of people carrying two copies of the most common genetic variant are unaware they are carriers.

When the researchers genetically engineered zebrafish with genetic variants causing Gaucher disease that are common among Ashkenazi Jews, as anticipated their macrophages became enlarged and unable to break down the toxic materials, in this case an unusual type of fat (called sphingolipids) rather than protein. But when the team exposed the fish to TB, they discovered unexpectedly that the fish were resistant to infection, not susceptible.

̽��ֱ��reason for this resistance to infection was because of the fatty chemical that accumulates within the macrophages in Gaucher disease, called glucosylsphingosine. Glucosylsphingosine was found to act as a detergent-like microbicide that kills TB mycobacteria within minutes by disrupting their cell walls.

Professor Ramakrishnan added: “We’d unknowingly landed in a debate that’s been going on in human genetics for decades: are Ashkenazi Jews – who we know are at a much greater risk of Gaucher disease – somehow less likely to get TB infection? ̽��ֱ��answer appears to be yes.”

̽��ֱ��Ashkenazi Jewish diaspora has experienced centuries of persecution, often forced to live in ghettos and migrate from country to country. They would almost certainly have been exposed to TB, which spreads more widely among poorer living conditions and densely-populated urban areas.

Although this genetic mutation is associated with Gaucher disease, the fact that it makes people more resistant to TB would likely have outweighed the potential fitness cost of Gaucher disease. This would have increased the likelihood of affected individuals passing on their genes to future generations and therefore spread the mutation within the population. A similar phenomenon is seen among some individuals who carry genetic variants that protect them from malaria but, when more than one copy is present, cause harmful anaemia or even sickle cell disease.

Unlike the example of sickle cell anaemia, however, only individuals who carry two copies of the Gaucher genetic variant – one from each parent – are likely to be protected against TB. That’s because the one ‘healthy’ gene generates enough of the enzyme to clear the macrophages of their accumulating material – and hence gets rid of the antimicrobial substrate.

Professor Timothy Cox from the ̽��ֱ�� of Cambridge, a co-author on the paper, added: “Our discovery may provide clues to possible new treatments for TB. Drugs that mimic the effects of Gaucher disease – specifically the build-up of glucosylsphingosine – might offer antimicrobial effects against TB.”

Several such drugs have already been designed by Professor Hans Aerts from Leiden ̽��ֱ��, another co-author on the paper. Because these drugs would only need to be administered for a relatively short amount of time, any side-effects should be limited and temporary.

̽��ֱ��research was funded by Wellcome, Gates Cambridge and the National Institute for Health and Care Research Cambridge Biomedical Research Centre.

Reference
Fan, J et al. Gaucher Disease Protects Against Tuberculosis. PNAS; 6 Feb 2023; DOI: 10.1073/pnas.2217673120

Scientists may have solved the question of why Ashkenazi Jews are significantly more susceptible to a rare genetic disorder known as Gaucher disease – and the answer may help settle the debate about whether they are less susceptible to tuberculosis (TB).

We’d unknowingly landed in a debate that’s been going on in human genetics for decades: are Ashkenazi Jews somehow less likely to get TB infection? ̽��ֱ��answer appears to be yes.

Lalita Ramkrishnan

halbergman (Getty Images)

Grandfather Helping Little Boy to Wash His Hands at Passover Seder with Family - stock photo

̽��ֱ��text in this work is licensed under a Creative Commons Attribution 4.0 International License. Images, including our videos, are Copyright © ̽��ֱ�� of Cambridge and licensors/contributors as identified. All rights reserved. We make our image and video content available in a number of ways – as here, on our main website under its Terms and conditions, and on a range of channels including social media that permit your use and sharing of our content under their respective Terms.

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Cambridge scientists recognised by major European research organisation

cjb250 — Tue, 18 Jun 2019 11:42:19 +0000

Cambridge ̽��ֱ�� has the highest number of new members of any institution within Europe. Five ̽��ֱ�� of Cambridge researchers are among the 48 scientists from 17 countries elected:

Professor Sadaf Farooqi – Wellcome-Medical Research Council (MRC) Institute of Metabolic Science
Dr Fanni Gergely – Cancer Research UK Cambridge Institute
Professor Paul Lehner – Department of Medicine and the Cambridge Institute for Medical Research
Professor Lalita Ramakrishnan – Department of Medicine and the MRC Laboratory of Molecular Biology
Professor Nicole Soranzo – Department of Haematology and Wellcome Sanger Institute

In addition, Dr Garib Murshudov from the MRC Laboratory of Molecular Biology has also been elected.

EMBO is an organisation of more than 1800 leading researchers in Europe and around the world, whose mission is to promote excellence in the life sciences in Europe and beyond. ̽��ֱ��major goals of the organisation are to support talented researchers at all stages of their careers, stimulate the exchange of scientific information and help build a research environment where scientists can achieve their best work.

“EMBO Members are excellent scientists who conduct research at the forefront of all life science disciplines, ranging from computational models or analyses of single molecules and cellular mechanics to the study of higher-order systems in development, cognitive neuroscience and evolution,” says EMBO Director Maria Leptin.

“We’re very honoured to have been elected as members of EMBO,” says Professor Farooqi. “This is great recognition for the excellent science taking place across our city, particularly on the Cambridge Biomedical Campus. We are proud of the role we play in European science and look forward to continuing to work in partnership with colleagues across the continent.”

Researchers from the Cambridge Biomedical Campus have featured prominently in this year’s election to the prestigious European Molecular Biology Organisation (EMBO).

We are proud of the role we play in European science and look forward to continuing to work in partnership with colleagues across the continent

Sadaf Farooqi

CDC/ Melissa Brower

This illustration depicts a three-dimensional (3D) computer-generated image of a cluster of rod-shaped drug-resistant Mycobacterium tuberculosis bacteria, the pathogen responsible for causing the disease tuberculosis (TB).

Yes

Academy of Medical Sciences announces 2018 Fellowships

cjb250 — Thu, 10 May 2018 10:22:09 +0000

̽��ֱ��new Fellows have been elected for their outstanding contributions to biomedical and health science, leading research discoveries, and translating developments into benefits for patients and the wider society.

This year's elected Fellows have expertise that spans sleep research, infectious and tropical diseases, diabetes medicine, parasite biology and ultrasound research and technology among many other fields.

Professor Sir Robert Lechler PMedSci, President of the Academy of Medical Sciences said: “ ̽��ֱ��Academy simply could not tackle major health and policy challenges without our dynamic and diligent brain trust of Fellows. I extend my warmest congratulations to all who are joining us this year, each of whom has earnt this prestige by advancing their own field of biomedical science.

“Later this year the Academy will celebrate 20 years of supporting the translation of biomedical and health research into benefits for society. As we celebrate this special anniversary we stand at a crossroads of both enormous health challenges and great opportunity for medical sciences. So I am delighted to see the remarkable breadth and depth of the expertise within our 48 new Fellows. We look forward to these experts joining us in addressing the health challenges we face head on and exploiting opportunities to improve health in the UK and internationally.”

̽��ֱ��Cambridge researchers among the new Fellows are:

Professor Simon Baron-Cohen FBA, Autism Research Centre
Professor Simon Griffin, Department of Public Health and Primary Care
Professor James Huntington, Cambridge Institute for Medical Research
Professor Peter Hutchinson, Department of Clinical Neurosciences
Professor Jonathan Mant, Department of Public Health and Primary Care
Professor Lalita Ramakrishnan, Department of Medicine
Professor David Rowitch, Department of Paediatrics
Professor Nicole Soranzo, Department of Haematology

̽��ֱ��new Fellows will be formally admitted to the Academy at a ceremony on 27 June 2018.

Eight Cambridge academics are among 48 of the UK’s world leading researchers who have been elected to join the prestigious Fellowship of the Academy of Medical Sciences.

̽��ֱ��text in this work is licensed under a Creative Commons Attribution 4.0 International License. For image use please see separate credits above.

Yes

̽��ֱ��Royal Society announces 2018 Fellows

cjb250 — Wed, 09 May 2018 14:26:31 +0000

̽��ֱ��50 newly-elected Fellows announced today join a list of scientists, engineers and technologists from the UK and Commonwealth. Past Fellows and Foreign Members have included Newton, Darwin and Einstein.

̽��ֱ��Cambridge academics announced today as Royal Society Fellows are:

Alexander Dawid, Emeritus Professor of Statistics, Department of Pure Mathematics and Mathematical Statistics
Gregory Hannon, Royal Society Wolfson Research Professor of Molecular Cancer Biology and Director, Cancer Research UK Cambridge Institute
Judy Hirst, Deputy Director, MRC Mitochondrial Biology Unit
Lalita Ramakrishnan, Professor of Immunology and Infectious Diseases and Head of Molecular Immunity Unit, Department of Medicine

Sir Venki Ramakrishnan, President of the Royal Society, says: “Our Fellows are key to the Royal Society’s fundamental purpose of using science for the benefit of humanity. For their outstanding contributions to research and innovation, both now and in the future, it gives me great pleasure to welcome the world’s best scientists into the ranks of the Royal Society.”

̽��ֱ��new Fellows will be formally admitted to the Society at the Admissions Day ceremony in July, when they will sign the Charter Book and the Obligation of the Fellows of the Royal Society.

View the full list of new Fellows and Foreign Members

Four Cambridge academics are among the new Fellows announced today by the Royal Society and chosen for their outstanding contributions to science.

For their outstanding contributions to research and innovation, both now and in the future, it gives me great pleasure to welcome the world’s best scientists into the ranks of the Royal Society

Sir Venki Ramakrishnan

Royal Society

Professor Simon Tavare elected as a Foreign Member of US National Academy of Sciences

Professor Simon Tavare, former Director of the Cancer Research UK Cambridge Institute, has been elected as a Foreign Member of the US National Academy of Sciences.

Professor Tavare works at the interface of the mathematical sciences and the biological and medical sciences. He pioneered probabilistic and statistical aspects of coalescent theory, full likelihood-based methods for sequence variation data, methods for ancestral inference, evolutionary approaches to cancer, and approximate Bayesian computation for inference in complex stochastic processes.

̽��ֱ��text in this work is licensed under a Creative Commons Attribution 4.0 International License. For image use please see separate credits above.

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Leprosy turns the immune system against itself, study finds

cjb250 — Wed, 23 Aug 2017 08:33:57 +0000

Leprosy is an infectious disease that affects the skin and peripheral nerves and is caused by Mycobacterium leprae and, less commonly, Mycobacterium lepromatosis. According to the World Health Organization, there has been a dramatic decrease in the global disease burden in the past few decades: from 5.2 million people with leprosy in 1985 to 176,176 at the end of 2015.

Despite the disease having been known about for thousands of years – many people will have first heard about it through references in the Bible – very little is understood about its biology. This is in part because the bacteria are difficult to grow in culture and there are no good animal models: M. leprae can grow in the footpads of mice, but do not cause nerve damage; the disease causes nerve damage in armadillos, but these animals are rarely used in research.

Now, an international team led by researchers at the ̽��ֱ�� of Cambridge, UK, and the ̽��ֱ�� of Washington, the ̽��ֱ�� of California Los Angeles and Harvard ̽��ֱ��, USA, have used a new animal model, the zebrafish, to show for the first time how M. leprae damage nerves by infiltrating the very cells that are meant to protect us. Zebrafish are already used to study another species of mycobacteria, to help understand tuberculosis (TB).

Scientists have previously shown that the nerve damage in leprosy is caused by a stripping away of the protective insulation, the myelin sheath, that protects nerve fibres, but it was thought that this process occurred because the bacteria got inside Schwann cells, specialist cells that produce myelin.

In new research published today in the journal Cell, researchers used zebrafish that had been genetically modified so that their myelin is fluorescent green; young zebrafish are themselves transparent, and so the researchers could more easily observe what was happening to the nerve cells. When they injected bacteria close to the nerve cells of the zebrafish, they observed that the bacteria settled on the nerve, developing donut-like ‘bubbles’ of myelin that had dissociated from the myelin sheath.

When they examined these bubbles more closely, they found that they were caused by M. leprae bacteria inside of macrophages – literally ‘big eaters’, immune cells that consume and destroy foreign bodies and unwanted material within the body. But, as is also often the case with TB, the M. leprae was consumed by the macrophages but not destroyed.

“These ‘Pac-Man’-like immune cells swallow the leprosy bacteria, but are not always able to destroy them,” explains Professor Lalita Ramakrishnan from the Department of Medicine at the ̽��ֱ�� of Cambridge, whose lab is within the Medical Research Council’s Laboratory of Molecular Biology. “Instead, the macrophages – which should be moving up and down the nerve fibre repairing damage – slow down and settle in place, destroying the myelin sheath.”

Professor Ramakrishnan working with Dr Cressida Madigan, Professor Alvaro Sagasti, and other colleagues confirmed that this was the case by knocking out the macrophages and showing that when the bacteria sit directly on the nerves, they do not damage the myelin sheath.

̽��ֱ��team further demonstrated how this damage occurs. A molecule known as PGL-1 that sits on the surface of M. leprae ‘reprograms’ the macrophage, causing it to overproduce a potentially destructive form of the chemical nitric oxide that damages mitochondria, the ‘batteries’ that power nerves.

“ ̽��ֱ��leprosy bacteria are, essentially, hijacking an important repair mechanism and causing it to go awry,” says Professor Ramakrishnan. “It then starts spewing out toxic chemicals. Not only does it stop repairing damage, but it creates more damage itself.”

“We know that the immune system can lead to nerve damage – and in particular to the myelin sheath – in other diseases, such as multiple sclerosis and Guillain–Barré syndrome,” says Dr Cressida Madigan from the ̽��ֱ�� of California, Los Angeles. “Our study appears to place leprosy in the same category of these diseases.”

̽��ֱ��researchers say it is too early to say whether this study will lead to new treatments. There are several drugs being tested that inhibit the production of nitric oxide, but, says Professor Ramakrishnan, the key may be to catch the disease at an early enough stage to prevent damage to the nerve cells.

“We need to be thinking about degeneration versus regeneration,” she says. “At the moment, leprosy can be treated by a combination of drugs. While these succeed in killing the bacteria, once the nerve damage has been done, it is currently irreversible. We would like to understand how to change that. In other words, are we able to prevent damage to nerve cells in the first place and can we additionally focus on repairing damaged nerve cells?”

̽��ֱ��research was funded by the National Institutes of Health, the Wellcome Trust, and the AP Giannini Foundation.

Reference
Madigan, CA et al. A Macrophage Response To Mycobacterium leprae Phenolic Glycolipid Initiates Nerve Damage In Leprosy. Cell; 24 Aug 2017; DOI: 10.1016/j.cell.2017.07.030

Leprosy hijacks our immune system, turning an important repair mechanism into one that causes potentially irreparable damage to our nerve cells, according to new research that uses zebrafish to study the disease. As such, the disease may share common characteristics with conditions such as multiple sclerosis.

̽��ֱ��leprosy bacteria are, essentially, hijacking an important repair mechanism and causing it to go awry

Lalita Ramakrishnan

Wellcome Library, London

Hand showing leprosy

̽��ֱ��text in this work is licensed under a Creative Commons Attribution 4.0 International License. For image use please see separate credits above.

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‘Clogged-up’ immune cells help explain smoking risk for TB

cjb250 — Thu, 24 Mar 2016 16:00:00 +0000

TB is an infectious disease caused by Mycobacterium tuberculosis that primarily infects the lungs, but can also infect other organs. It is transmitted from person to person through the air. ̽��ֱ��disease can cause breathlessness, wasting, and eventual death. While treatments do exist, the drug regimen is one of the longest for any curable disease: a patient will typically need to take medication for six months.

For people exposed to TB, the biggest risk factor for infection is exposure to smoke, including active and passive cigarette smoking and smoke from burning fuels. This risk is even greater than co-infection with HIV. However, until now it has not been clear why smoke should increase this risk.

When TB enters the body, the first line of defence it encounters is a specialist immune cell known as a macrophage (Greek for ‘big eater’). This cell engulfs the bacterium and tries to break it down. In many cases, the macrophage is successful and kills the bacterium, preventing TB infection, but in some cases TB manages not just to avoid destruction, but to use macrophages as ‘taxi cabs’ and get deep into the host, spreading the infection. TB’s next step is to cause infected macrophages to form tightly-organised clusters known as tubercles, or granulomas. Once again here, the macrophages and bacteria fight a battle – if the macrophages lose, the bacteria use their advantage to spread from cell to cell within this structure.

An international team of researchers, led by the ̽��ֱ�� of Cambridge, and the ̽��ֱ�� of Washington, Seattle, studying genetic variants that increase susceptibility to TB in zebrafish – a ‘see-through’ animal model for studying the disease – identified a variant linked to ‘lysosomal deficiency disorders’. ̽��ֱ��lysosome is a key component of macrophages responsible for destroying bacteria. This particular variant caused a deficiency in an enzyme known as cathepsin, which acts within the lysosome like scissors to ‘chop up’ bacteria; however, this would not necessarily explain why the macrophages could not destroy the bacteria, as many additional enzymes could take cathepsin’s place.

̽��ֱ��key, the researchers found, lay in a second property of the macrophage: housekeeping. As well as destroying bacteria, the macrophage also recycles unwanted material from within the body for reuse, and these lysosomal deficiency disorders were preventing this essential operation.

Professor Lalita Ramakrishnan from the Department of Medicine at the ̽��ֱ�� of Cambridge, who led the research, explains: “Macrophages act a bit like vacuum cleaners, hoovering up debris and unwanted material within the body, including the billions of cells that die each day as part of natural turnover. But the defective macrophages are unable to recycle this debris and get clogged up, growing bigger and fatter and less able to move around and clear up other material.

“This can become a problem in TB because once the TB granuloma forms, the host’s best bet is to send in more macrophages at a slow steady pace to help the already infected macrophages.”

Image: Left - normal macrophages (green); Right - dysfunctional macrophages whose lysosomes (red) are clogged with cell debris. Credit: Steven Levitte

“When these distended macrophages can’t move into the TB granuloma,” adds co-author Steven Levitte from the ̽��ֱ�� of Washington, “the infected macrophages that are already in there burst, leaving a ‘soup’ in which the bacteria can grow and spread further, making the infection worse.”

̽��ֱ��researchers looked at whether the effect seen in the lysosomal deficiency disorders, where the clogged-up macrophage could no longer perform its work, would also be observed if the lysosome became clogged up with non-biological material. By ‘infecting’ the zebrafish with microscopic plastic beads, they were able to replicate this effect.

“We saw that accumulation of material inside of macrophages by many different means, both genetic and acquired, led the same result: macrophages that could not respond to infection,” explains co-author Russell Berg.

This discovery then led the team to see whether the same phenomenon occurred in humans. Working with Professor Joe Keane and his colleagues from Trinity College Dublin, the researchers were able to show that the macrophages of smokers were similarly clogged up with smoke particles, helping explain why people exposed to smoke were at a greater risk of TB infection.

“Macrophages are our best shot at getting rid of TB, so if they are slowed down by smoke particles, their ability to fight infection is going to be greatly reduced,” says Professor Keane. “We know that exposure to cigarette smoke or smoke from burning wood and coal, for example, are major risk factors for developing TB, and our finding helps explain why this is the case. ̽��ֱ��good news is that stopping smoking reduces the risk – it allows the impaired macrophages to die away and be replaced by new, agile cells.”

Image: Smoke-clogged macrophages of cigarette smokers are unable to move to engulf infecting TB bacteria, which may explain why cigarette smokers are more susceptible to tuberculosis. Credit: Kevin Takaki and drawn by Paul Margiotta

̽��ֱ��research was supported by the National Institutes of Health, the Wellcome Trust, the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre (BRC), the Health Research Board of Ireland and ̽��ֱ��Royal City of Dublin Hospital Trust.

Also contributing to this research were Professor David Tobin from Duke ̽��ֱ��, Dr Cecilia Moens from the Fred Hutchinson Cancer Research Institute, Drs C.J. Cambier and J. Cameron from ̽��ֱ�� of Washington, Dr Kevin Takaki from ̽��ֱ�� of Cambridge and Drs Seonadh O’Leary and Mary O’Sullivan from Trinity College Dublin.

Reference
Berg, RD, Levitte, S et al. Lysosomal Disorders Drive Susceptibility to Tuberculosis by Compromising Macrophage Migration. Cell; 24 Mar 2016; 10.1016/j.cell.2016.02.034

Smoking increases an individual’s risk of developing tuberculosis (TB) – and makes the infection worse – because it causes vital immune cells to become clogged up, slowing their movement and impeding their ability to fight infection, according to new research published in the journal Cell.

Macrophages act a bit like vacuum cleaners, hoovering up debris and unwanted material within the body, including the billions of cells that die each day as part of natural turnover

Lalita Ramakrishnan

ZEISS Microscopy

Macrophage engulfing Tuberculosis bacteria

̽��ֱ��text in this work is licensed under a Creative Commons Attribution 4.0 International License. For image use please see separate credits above.

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Attribution-Noncommerical

Even without lungs, zebrafish help us study TB

amb206 — Wed, 25 Nov 2015 09:25:17 +0000

Scroll to the end of the article to listen to the podcast.

Professor Lalita Ramakrishnan shares her workspace at the Laboratory of Molecular Biology (LMB) with thousands of tiny stripy fish. Zebrafish have long been a favourite in domestic aquariums: they are strikingly pretty and constantly on the move. ̽��ֱ��zebrafish at the LMB, each one no bigger than your little finger, are helping Ramakrishnan and her colleagues to find novel ways of preventing and treating tuberculosis (TB). We asked her about her work.

Why are zebrafish such good models for scientists?

Around 40 years ago scientists began to realise that zebrafish, as vertebrates, could tell us a lot about human development and human diseases. This discovery represented a real breakthrough in terms of what could be achieved using zebrafish in laboratories.

There are two key reasons why zebrafish, in particular, are so valuable. Firstly, when the new fish hatches as a tiny larva, it is optically transparent for the first two weeks of its development. This transparency means that, using powerful imaging technology, we are able to observe in real time the development of the organism as it grows to maturity. In our laboratory, we exploit the optical transparency to directly look at how the tuberculosis bacteria cause disease.

̽��ֱ��second reason why zebrafish is such a good model is that a single mating can produce hundreds of eggs – and female zebrafish are capable of producing a new batch of eggs each week. So we have access to large numbers of animals for the work. On top of all this, zebrafish are relatively straight-forward to keep and easy to breed.

We can also create zebrafish with different mutations and we can then assess the impact of host genes on the course of disease. This kind of fundamental work enables us to identify, by a process of deduction and elimination, what genes do – which is essential to developing new medical interventions.

But surely zebrafish and humans have little in common – we’re not fish!

Humans and fish are much more alike than people might suppose – even though we diverged from our last common ancestor at least 300 million years ago. Most of the genes found in fish are also found in humans – and most of the genes that cause disease in fish also cause disease in humans. ̽��ֱ��human immune system, which fights off disease, is a lot like the immune system of fish.

My research is focused on tuberculosis in humans – a disease that affects millions of people worldwide. Without treatment, TB can be life threatening. We tend to associate human TB with the lungs, and of course fish don’t have lungs. TB does affect the lungs but it can affect almost all our organs. In humans, some 40% of TB infection is not in the lungs but elsewhere in the body - brain, bone, kidney, intestine, reproductive organs.

Fish are affected by a close relative of the human TB bacterium. If we can work out how TB works in fish, and how to prevent it and treat it in fish, then we’re a step closer to solving a major health problem in humans.

What is the life of a laboratory zebrafish like?

Our fish live in tanks that are kept pristine by a unit that cleans and circulates the water. We grow the food they need in the lab – it’s a kind of brine shrimp. Putting this live food into the tanks allows the fish to hunt for their food, creating a more natural environment for them. Zebrafish are sociable creatures so we keep them in groups. All our fish are on a programme of 16 hours of daylight and eight hours of night. This routine mimics, as much as possible, the natural environment in the regions of the world where they live. We make sure that they are as healthy and stress-free as possible. Happy fish are healthy fish – and the other way round!

You can identify the males from the females by the roundness of the female’s belly. When we want a new batch of eggs, we put a male and a female in a tank overnight, with the two fish separated by a transparent divider. When daylight comes, the two fish become excited and we take out the divider and they mate. When the eggs are laid, they fall through a fine grill that enables us to take them out of the tank.

All these procedures are done as carefully as possible so as not to harm the fish or eggs.

How do you use the zebrafish eggs?

In my laboratory, we’re studying TB so we need to infect some of the fish eggs, one by one, with bacteria so that we can observe what happens. This procedure is carried out under a microscope using a very fine needle that is hollow, enabling tiny amounts of bacteria to be delivered into the egg. Because zebrafish eggs are so tiny, it takes a while to learn how to do this. It requires good hand-eye coordination and a steady hand – but everyone learns to do it with time and practice.

Once the eggs are infected we put them into small dishes where we can observe them. Because the eggs and the larvae are transparent, we can observe the process by which the bacteria enters the cells – and we can watch what happens as the bacteria and immune system face off. By using fluorescence, we can colour the host (the organism affected by the disease) and the bacteria so that it’s easier to track what’s happening on a cellular level. We can, for example, observe how exactly bacteria invade and spread.

How is your work with fish helping to develop better ways of tackling TB?

At the moment, TB in the human population is treated with a long course of strong antibiotics – it often takes as long as six months to get rid of it. Strains of drug-resistant TB have developed, partly because people do not finish the courses of drugs prescribed to them.

̽��ֱ��work that my colleagues and I are doing suggests that there could be another, and perhaps more effective, approach to tackling TB. Rather than only targeting the bacteria, which are so clever in their invasive strategies, it might be better to additionally target the host and help the immune system to fight it off. We might do this by boosting or tweaking the immune system.

We now need to put to the test our ideas for helping the immune system by trying out a list of available drugs – and, in the initial stages of the research, we will be using zebrafish as models.

What’s the future for zebrafish as a model organism in research?

̽��ֱ��world of research using zebrafish is wonderfully collaborative and fast-moving. Our main partner is the Sanger Institute which is just a few miles from the LMB. We collaborate closely with the scientists there on tools and techniques – including producing the mutants in order to identify genetic pathways.

Zebrafish are still relatively new in terms of their contribution to research – but it’s difficult to overstate how important they are. Every research organism has its limitations, of course. However, there’s much, much more we can learn from zebrafish that will benefit humans in the future.

This is the last article in the Cambridge Animal Alphabet series. If you have missed the others, you can catch up on Medium here.

Inset images: A two day old transgenic zebrafish embryo (Wikimedia Commons / IchaJaroslav); Zebrafish embryos (Wikimedia Commons / Adam Amsterdam, Massachusetts Institute of Technology).

̽��ֱ��Cambridge Animal Alphabet series celebrates Cambridge's connections with animals through literature, art, science and society. Here, Z is for Zebrafish as we talk to eminent immunologist Professor Lalita Ramakrishnan about her research into new ways of treating tuberculosis.

If we can work out how TB works in fish, and how to prevent it and treat it in fish, then we’re a step closer to solving a major health problem in humans

Lalita Ramakrishnan

Wikimedia Commons / Pogrebnoj-Alexandroff

Danio rerio (Zebrafish)

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Yes

A whole host of options

cjb250 — Fri, 09 Oct 2015 08:30:04 +0000

Professor Lalita Ramakrishnan is, it’s fair to say, a world authority on the biology of TB. She studies the disease – one which most people will know of as a disease of the lungs – using what at first sight seems an unusual model: the zebrafish.

“What most people don’t realise is that about 40% of human TB occurs outside the lungs,” explains Ramakrishnan. “It can infect the brain, bone, heart, reproductive organs, skin, even the ear. In fact, TB infection is a basic biology question, and this is the same in zebrafish as it is in humans.”

TB is caused by Mycobacterium tuberculosis, which is generally transmitted from person to person through the air. It has been around since at least the Neolithic period, but its prevalence in 19th-century literature led it to be considered something of a ‘romantic’ disease. ̽��ֱ��truth is a long way from this portrayal. ̽��ֱ��disease can cause breathlessness, wasting and eventual death. And while treatments do exist, the drug regimen is one of the longest for any curable disease: a patient will typically need to take medication for six months.

Ramakrishnan is involved in a new trial due to start soon that might allow doctors to reduce the length of this treatment. She is cautiously optimistic that it can be reduced to four months; if successful, however, it may eventually lead to treatments more on a par with standard antibiotic treatments of a couple of weeks.

̽��ֱ��trial builds on work in zebrafish carried out by Ramakrishnan and colleagues at the ̽��ֱ�� of Washington, Seattle, before she moved to the Department of Medicine in Cambridge in September 2014. These small fish, which grow to the length of a little finger, helped her and collaborator Professor Paul Edelstein from the ̽��ֱ�� of Pennsylvania (currently on sabbatical in Cambridge) to make an important discovery that could explain why it takes a six-month course of antibiotics to rid the body of the disease (rather than seven to ten days that most infections take) and yet in the lab can easily be killed.

Within our bodies, we have a host of specialist immune cells that fight infection. One of these is the macrophage (Greek for ‘big eater’). This cell engulfs the TB bacterium and tries to break it down. This, together with powerful antibiotics, should make eliminating TB from the body a cinch. Ramakrishnan’s breakthrough was to show why this wasn’t the case: once inside the macrophages, TB switches on pumps, known as ‘efflux pumps’. Anything that we throw at it, it just pumps back out again.

“Once we’d identified the pumps, we started to look for drugs that are out there in the market and tested a few of them,” she explains. “We found that verapamil, an old drug, made the bacteria susceptible to two of the antibiotics we use to fight TB.”

̽��ֱ��trial of verapamil, which is commonly used to treat high blood pressure, is due to start soon at the National Institute for Research in Tuberculosis (NIRT) in Chennai, India.

Ramakrishnan is one of a number of brilliant minds working as part of a collaboration between the NIRT and the ̽��ֱ�� of Cambridge to apply the very latest in scientific thinking and technology to the problem of TB.

An expansion of this collaboration has now become possible through the recent award of a £2 million joint grant from the UK Medical Research Council (MRC) and the Department of Biotechnology (DBT) in India, which will enable the exchange of British and Indian researchers. For Professor Sharon Peacock, the UK lead on the proposal, this means an opportunity to train a new cohort of early-career researchers in an environment where they will have access to outstanding scientific facilities and training, at the same time as becoming familiar with the clinical face and consequences of TB for people in India.

“India is home to a large pool of talented young people with the potential to help fight back against this deadly disease,” says Peacock. “Developing a close collaboration between Cambridge and Chennai involving two-way traffic of scientists and ideas is an exciting opportunity to start to tap into this.”

There are few places more suitable for the proposed work than India. According to the World Health Organization, India is home to almost one in four of all worldwide cases of TB, with over two million newly diagnosed cases in 2014.

Not only that, but it is one of the countries that has seen an increase in the number of cases of drug resistance to TB – including ‘multi-drug’-resistant, and even more worrying, ‘extremely’ drug-resistant strains of TB against which none of our first- and second-line drug treatments work. In part, this increase reflects improved access to diagnostic services, but the situation highlights why new approaches to tackling the disease are urgently needed, says Professor Soumya Swaminathan, Director of NIRT and the India lead in the collaboration.

“So far, the treatment of TB has focused almost exclusively on using drugs to try to kill the bacteria directly, but there’s increasing evidence that there may be benefits to targeting the host. TB is very clever and it manipulates the host immune system to its own advantage, so if we could use drugs to help the immune system, then we may be able to make it more effective.”

This is the approach that Professors Ken Smith and Andres Floto from the Department of Medicine at Cambridge, also part of the collaboration, are taking. Smith is looking at the role that specialist immune cells known as T cells play in the persistence of multi-drug-resistant strains of TB. His group has evidence that around two thirds of the population have T cells which have a tendency to become ‘exhausted’ when activated.

“It might be that exhausted T cells can’t fight multi-drug-resistant TB effectively, in which case we need to find a way to overcome this exhaustion and spur the T cells on to rid the body of the disease,” says Smith.

For Floto, the key may lie in the role played by the macrophages and their otherwise voracious appetites. As their Greek name suggests, macrophages ‘eat’ unwanted material (surprisingly similar in action to Pac-Man), effectively chewing it up, breaking it down and spitting it out again.

This process, known as autophagy (‘self-eating’), is a repair mechanism for clearing damaged bits of cells and recycling them for future use, but also works as a defence mechanism against some invading bacteria. So why, when it engulfs TB, does the bacterium manage to avoid being digested?

“Autophagy is partially inhibited by TB itself, but we found that if you overstimulate this mechanism – like flooring the accelerator of a car – you can overcome the bacteria,” explains Floto. “Clearly this will be applicable to normal TB, but we already have drugs that are effective against this. We want to know if this would work against multi-drug-resistant strains.”

Floto and colleagues already have a list of potential drugs that can stimulate autophagy, drugs that have already been licensed and are in use to treat other conditions, such as carbamazepine, which is used to treat epileptic seizures. These drugs are safe to use: the question is, will they work against TB?

“We’ve already shown that carbamazepine stimulates autophagy in cells to kill TB – even multi-drug-resistant TB. We now want to refine it and test it in mice and in fish, alongside a shortlist of around 30 other potential drugs,” he adds.

TB evolves through ‘polymorphisms’ – spontaneous changes in the letters of its DNA to create variants. Because the drug regimen to fight the disease lasts so long, many patients do not take the full course of their medicines. If the TB is allowed to relapse, it can evolve drug resistance.

These patterns of resistance can be detected using genome sequencing – reading the DNA of the bacteria. Peacock believes this technique may be able to help doctors more easily diagnose drug resistance in patients.

“TB is very slow to grow in the laboratory, which means that testing an organism to confirm which antibiotics it is susceptible or resistant to can take several weeks, especially in the case of more resistant strains,” she says. “There is increasing evidence that antibiotic resistance can be predicted from the genome sequence of the organism, and we want to establish and evaluate this technology in India, where it is needed.”

This sequencing data could also then help inform the search for new drugs, explains Professor Sir Tom Blundell from the Department of Biochemistry. He is no stranger to TB: his grandfather died from the disease shortly after the war – though, as Blundell points out, this strain of TB is far less common now, as the organism has evolved in different communities throughout the world.

“We can take the polymorphisms and ask questions such as ‘What does this mean for the use of current drugs?’” says Blundell. “ ̽��ֱ��nature of the polymorphisms in the TB genome sequence of an infected individual can give us information on where that person was infected and what are the drugs that might be most effective. We can then begin to look at new targets for particular polymorphisms.”

Blundell plans to take the information gathered through the Chennai partnership and feed it into his drug discovery work. He takes a structural approach to solving the problem: look at the shape of the polymorphism and its protein products and try to find small molecules that can attach to and manipulate them. In essence, it’s akin to picking a lock by analysing the shape of its mechanism and trying to identify a key that could turn it, thus opening the door.

Yet even if the Chennai venture is successful, and research from the partnership leads to a revolution in how we understand and treat TB, the team recognise that this is unlikely to be enough to eradicate the disease for good.

“TB is as much a public health issue as one of infectious diseases,” says Ramakrishnan, pointing to Europe, where even before the introduction of antibiotics, the disease was already on the decline. “We need better nutrition, better air, less smoking, reductions in diabetes.”

Swaminathan agrees. “TB is very much associated with poverty and all the risk factors that go with it,” she says. “When people are living in very crowded conditions, when they’re malnourished, TB is going to continue to spread. This is happening in the slums of Mumbai, for example, where we’re seeing a mini-epidemic of multi-drug-resistant TB. Unless we see a rapid improvement in the living standards of people we’re not going to see a very major effect. There’s only so much we can do biomedically.”

Inset image: Macrophage engulfing Tuberculosis pathogen (ZEISS Microscopy).

Almost one in four of the world’s cases of tuberculosis (TB) are in India and the disease is constantly adapting itself to outwit our medicines. Could the answer lie in targeting not the bacteria but its host, the patient?

What most people don’t realise is that about 40% of human TB occurs outside the lungs ... It can infect the brain, bone, heart, reproductive organs, skin, even the ear

Lalita Ramakrishnan

Calcutta Rescue

Picture to educate people in villages that have no medical service about the spread of TB

̽��ֱ��Next Generation

If there’s one thing on the side of science v. TB, it’s the wealth of talent available in India.

Professor Sir Tom Blundell is quick to praise the Indian postdocs that come to work in his lab. “They tend to be naturally very inquisitive and interactive, with very enquiring minds,” he says.

This is something with which Professor Ashok Venkitaraman, Director of the Medical Research Council (MRC) Cancer Unit at Cambridge, wholeheartedly agrees. He has helped establish the Center for Chemical Biology and Therapeutics (CCBT) in Bangalore in part, he says, because “the number of really bright, well-trained young scientists in India is huge. ̽��ֱ��level of enthusiasm and commitment is something I find quite exceptional.”

̽��ֱ��CCBT is an inter-institutional centre that links the Institute for Stem Cell Biology and Regenerative Medicine and the National Center for Biological Sciences, both of which are world-class Indian research institutes studying fundamental biology. However, argues Venkitaraman, India needs the capacity to translate fundamental research to clinical application.

It is to help bridge this gap that the CCBT was established, with funding from the Department of Biotechnology (DBT) in India, recently supplemented by a £2 million joint award from the UK MRC and the DBT. ̽��ֱ��idea is to find innovative ways to discover ‘next-generation’ medicines against human diseases, by coupling biological research that reveals novel drug targets with approaches in chemistry and structural biology that create potential drug candidates.

Although Venkitaraman’s interest is in cancer, he predicts the work of the CCBT will be “disease agnostic”, because similar types of novel drug targets have been implicated in infectious diseases, cancer and even developmental defects.

“We desperately need to develop new medicines not just for currently problematic diseases like cancer and TB, but also for the new challenges that are being thrown at us all the time – antibiotic resistance, new infections, metabolic syndromes and diseases of ageing, for example. Nowhere is this need more critical than in emerging nations like India where the spectrum of disease is distinct from countries like the UK.”

̽��ֱ��text in this work is licensed under a Creative Commons Attribution 4.0 International License. For image use please see separate credits above.

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