Meet the Microbiologist

A very small proportion of people infected with HIV do not develop AIDS. Mark Connors talks about 2 patient populations that his lab studies, the elite controllers and the elite neutralizers, who control HIV infection with their respective T cell or B cell responses. Connors hopes his work on killer T cells and broadly neutralizing antibodies will help scientists develop better HIV therapies or an effective HIV vaccine. Links for This Episode: Mark Connors labsite at NIAID Immunity article: Identification of a CD4-binding-site antibody to HIV that evolved near-pan neutralization breadth. Immunity commentary: Class II-restricted CD8s: New lessons violate old paradigms. Science article: Trispecific broadly neutralizing HIV antibodies mediate potent SHIV protection in macaques. Imagining an HIV-Free Future (Smithsonian Worlds AIDS Day Event (Live Dec 4th at 6:45pm) HOM Tidbit: 12 Diseases that Changed Our World MTM Listener Survey

In the early 2000s, Andrew Read predicted that non-sterilizing vaccines would lead to more virulent disease. He was able to test his hypothesis with the real-world example of Marek’s disease, a disease of chickens. Read tells the story of his discovery, and talks about his work on myxoma virus. Take the MTM Listener Survey Subscribe (free) on Apple Podcasts, Google Podcasts, Android, RSS, or by email. Also available on the ASM Podcast Network app. Julie’s Biggest Takeaways: Every chicken on the market is vaccinated against Marek’s disease. Infection with Marek’s disease causes tumors on the bird and can lead to direct death, or condemnation of a flock requiring their culling. Birds are vaccinated with a live, attenuated virus, and there have been 3 vaccine iterations. The first used a related herpesvirus isolated from turkeys, while the second vaccine added a second virus strain. Each of these vaccines conferred protection for about 10 years, after which the disease began popping up again. The 3rd generation vaccine added yet another serotype - this additional strain is a mutant strain of the chicken-infecting serotype - and has been effectively protecting chickens since the 1990s. Chickens do not get sterilizing immunity from the Marek’s disease vaccine; they can be infected by the wild-type virus, but the vaccine prevents infected animals from having disease symptoms. These asymptomatically infected animals can still shed the virus. Contrast this to human immunity from many of our vaccines, such as measles or smallpox vaccines, in which our immune response stops the virus from entering our cells and therefore blocks virus replication. Vaccination inhibits strains with lower virulence more than strains with higher virulence. This fact, combined with asymptomatic infection, means that although the infected birds don’t show disease symptoms, they are more likely to be shedding more virulent (or ‘hot’) strains. This generates selection for these hot strains that wouldn’t normally be successful. Without vaccination, host strains kill the host too quickly to allow viral replication and transmission to occur; Vaccines allow these hot strains to propagate. Vaccine resistance is much more rare than antibiotic or antimicrobial resistance. This is due to a number of factors, including the diversity of microbial population being acted upon (small with initial infection, large when treated with antimicrobial drugs). Vaccines are much more evolution-proof for these reasons. Purposeful release of myxoma virus during the 1950s in Australia killed between 10 and 100 million animals, or 99.9% of the rabbit population. Frank Fenner followed the virus and surviving rabbit populations and discovered that myxoma viruses that were too virulent were less likely to be transmitted, because they killed the host too quickly. He also showed that the small surviving number of rabbits were more resistant to viral infection. The arms race between the two has generated a virus so immunosuppressive that Read’s group has found the currently circulating myxoma virus has changed the way it kills its host: the virus disables the rabbit immune system and allows the rabbit’s own microbiome to cause invasive bacterial disease.

A recent Nipah virus outbreak in Kerala, India, was halted due to improved detection capabilities. G. Arunkumar tells the story of his involvement. Host: Julie Wolf  Take the MTM Listener Survey Subscribe (free) on Apple Podcasts, Google Podcasts, Android, RSS, or by email. Also available on the ASM Podcast Network app. Julie’s Biggest Takeaways: Because bats are the normal reservoir, Nipah virus outbreaks appear to be seasonal, with an increase in cases coinciding with the spring, when the bat reproduction season is. Once a person is infected through direct contact with the virus, the virus is transmitted person-to-person through respiratory droplets. Family clusters combined with the right incubation time acted as a clue that a Nipah virus outbreak had begun. Molecular tests improved virus detection during the 2018 Nipah outbreak because patients presented symptoms within a few days, which was too short for them to have developed antibodies. Molecular tests allowed identification of infected patients within days. Previous outbreaks have taken weeks to months, or even years, to identify the infectious virus. A single crossover event in the recent Nipah outbreak led to person-to-person transmission within the 22 additional individuals. Hospital infection control practices are important to reduce transmission to healthcare workers and hospital attendants. Featured Quotes: “Most of the Nipah outbreaks, you find a lot of hospital transmission from the infected patient to healthcare workers, the other patients in the ward as well as the patient attendants.” “The only virus that can cause encephalitis in a family cluster is Nipah. With other encephalitis viruses like herpes or Japanese encephalitis virus, you don’t see family clusters.” “Nipah virus is a level 4 pathogen, so the cultivation can be only done in a level 4 laboratory. But molecular tests allow you to test for it at a lower level laboratory, such as a BSL-3 lab, because you inactivate the virus. You are only focusing on RNA. The risk can be reduced.” “When you use serological diagnosis, the antibodies are detectable only after 8-10 days after onset of illness. Nipah is a very, very acute, serious fatal disease. Many people may die before they develop antibody. So we need to use a combination of real-time PCR and antibody.” “This is the first time in the history of Nipah that the diagnosis was done in country. All the previous diagnoses were done at CDC Atlanta.” Links for This Episode: Department of Virus Research at Manipal Academy of Higher Education Journal of Clinical Microbiology Review on Nipah virus ASM Global Impact Report HOM Tidbit: NPR piece interviewing K. B. Chua and others HOM Tidbit: Science article first describing Nipah virus

Robin Patel discusses her work on prosthetic joint infections and how metagenomics is changing infectious disease diagnostic procedures. Take the listener survey: asm.org/mtmpoll Julie’s Biggest Takeaways: The term antimicrobial resistance can mean many things. Although acquisition of genetic elements can lead to drug resistance, so can different growth lifestyles of bacteria; the same bacteria growing in liquid culture may be more susceptible to a drug than those bacteria growing on a biofilm. Lifestyle and genetics can intertwine, however, when bacteria growing as a biofilm exchange resistance genes through horizontal gene transfer. How do bacteria reach an implanted surface, such as on a prosthetic joint, to cause infection? It may rarely occur during surgery, if even a single bacterium reaches the joint surface despite the sterile conditions; alternatively, it could occur through hematogenous spread (through the blood) after the surgery is over. Most infections are believed to be seeded at the time of implantation. While scientists don’t perform teeny, tiny implants in animal models of infection, the materials are placed in animal bone to mimic as similar an immune response as possible. Targeted metagenomics and shotgun metagenomics are both being developed clinically. Targeted metagenomics looks at one specific gene found in a number of species, such as the 16S ribosomal RNA gene. Shotgun metagenomic looks at all DNA present, and requires a lot more cleaning up to eliminate human genomic material, which is the major sequence of any human-derived sample.  

To eliminate malaria, you have to stop transmission, and that’s what Carolina Barillas-Mury hopes to do. Her work on the interaction of the malaria parasite Plasmodium falciparum may lead to a transmission-blocking vaccine. She explains how, and discusses the co-evolution of malaria, mosquitos, and man.   Take the listener survey: asm.org/mtmpoll   Julie’s Biggest Takeaways: When born, babies carry antibodies from their mothers, which may protect them through passive immunity; additionally, babies are more easily protected from mosquito exposure by placing them under bed netting. As they grow, children become more active, and their passive immunity concurrently wanes. They may be exposed to mosquitoes carrying malaria parasites and their still-developing immune systems aren’t able to keep the parasites from replicating, leading to more severe disease, including cerebral malaria.   The Culicines and Anopholines are two major groups of mosquitoes that carry disease. The culicines have recently spread around the world, but the Anopholines species moved from Africa into South America one hundred million years ago, but malaria only moved into the New World a few hundred years ago with the slave trade. The relationship between the mosquitoes and malaria parasites has been evolving much longer in Africa than it has been with the specific population of mosquitoes in South America - one of the reasons why the disease is less devastating in South America.   The ‘invisibility gene,’ pfs47, is expressed in the banana-shaped ookinete and helps the malaria parasite to avoid detection by the mosquito immune system. The pfs47 malarial gene is adapted for the localized mosquito populations from the same region as the parasite; if an African mosquito is infected with a South American parasite, the parasite is more likely to be recognized and killed than if the African mosquito is infected with an African parasite.   The most immunogenic proteins in parasites may produce an immune response, but this immune response may not block infection. New vaccines are concentrating on where antibodies bind, to ensure there is a biological effect of the immune response, and this is why Barillas-Mury has used a modified Pfs47 protein to generate immune responses, rather than its native form. Links for this Episode: Carolina Barillas-Mury NIAID website NPJ Vaccines: Antibody Targeting of a Specific Region of Pfs47 Blocks Plasmodium falciparum Malaria Transmission. PLoS One: Molecular Analysis of Pfs47-Mediated Plasmodium Evasion of Mosquito Immunity. PNAS: Plasmodium Evasion of Mosquito Immunity and Global Malaria Transmission: The Lock-and-Key Theory. HOM Tidbit: History of the Discovery of the Malaria Parasites and their Vectors MTM Listener Survey  

How do researchers study a new pathogen? Stanley Perlman talks about how virus researchers studied SARS and MERS after they emerged, what they learned, and why there are no more cases of SARS. He also discusses his work on a coronavirus model of multiple sclerosis.   We want to hear from you! Please take our listener survey.   Julie’s Biggest Takeaways: Coronaviruses have the largest RNA genomes, with up to 40 kB of sequence, but why their genomes is so big is unclear - their genomes don’t seem to code for more genes than viruses with smaller genomes. Before the SARS coronavirus outbreak in 2002, few severe human infectious coronaviruses were known, but the several coronaviruses had been identified that cause serious disease in animals such as pigs, cats, and cows. Where did SARS go? SARS coronavirus had to cross into people and mutate for better infectivity, and when infecting people, it caused a lower respiratory disease. Quarantining SARS patients is extremely effective because the symptoms coincide with infectivity, and spread of SARS was quenched by strict use of quarantine. Quarantine is less effective for diseases like influenza or measles, because patients are contagious before showing symptoms. Because of its low person-to-person transmission, there’s very small possibility of major outbreaks from large gatherings such as the Hajj. MERS acts more like an opportunistic infection, and its transmission among people has been mostly among immunocompromised or otherwise sick people in the hospital. By the time patients present with multiple sclerosis, it may be 20 years after an inciting event that triggers the disease. By using a murine coronavirus inciting event for neuron demyelinization in mice, the role of the immune system in this process can be interrogated. Scientists may not understand the exact cause of MS in people, but this model helps them to understand how different immune cells contribute to disease.

You may know that beer is fermented, but did you know making chocolate requires a fermentation step? Kevin Verstrepen discusses how his lab optimizes flavor profiles of the yeast used in this fermentation step, and explains how yeast was domesticated before microorganisms had been discovered. Take the MTM listener survey, we want to hear from you. Thanks! Julie’s Biggest Takeaways: Microbes are used to ferment foods, but they do more than just add ethanol or carbon dioxide: their metabolic byproducts add flavors and aromas that are an essential part of the fermented food. In cocoa bean fermentation, the yeast that are part of the initial fermentative microbial population control the development of the subsequent microbial populations and the quality of the final product. How the volatile flavor compounds generated during fermentation survive the roasting step remains unclear. Heat can destroy these labile compounds, but Kevin thinks the compounds were able to survive roasting because they become embedded in lipids (fat) of the cocoa beans. Similar compounds produced during bread rising are destroyed during baking, possibly because there is less fat to protect these molecules. Mixing data science and beer: a computer scientist in the Verstrepen lab analyzed the flavor profiles of several hundred beers, which were also analyzed by a trained tasting panel. The goal is to link the chemistry to the aroma, which requires complex algorithms due to the integration of hundreds of flavor molecules. A spontaneous hybridization between Saccharomyces cerevisiae, the normal fermentative yeast, and S. eubayanus, a cold-tolerant yeast, resulting in a hybrid that can ferment at colder temperatures, as is required for brewing lager beers. There are 2 lineages that are used by most breweries, and while different characteristics have evolved over time, the genetic bottleneck limits characteristic diversity. The Verstrepen lab made several crosses between these two species and selected for hybrids that generated those with desirable characteristics. Molecular means can determine the offspring that are most likely to confer desired characteristics, but the commercial yeasts are not specifically genetically manipulated to this end. Domesticated yeast have different characteristics than their wild counterparts. Domesticated yeasts have lost the ability to use certain sugars, but have gained abilities associated with their use; beer yeasts use maltose at much higher rates, for example. When the origins are traced using molecular methods, it goes back to medieval times. How to domesticate an organism that hasn’t been identified? Brewers have long transferred sediment from one batch of beer into new batches, which is how selection for human-desired characteristics began. Wine yeasts, which are not passaged but are likely inoculated from the same vineyard annually, show less domestication than the beer yeast.

How can the humble zebrafish teach us about the human microbiome? John Rawls discusses the benefits of using animal models Take the MTM Listener Survey  Julie’s Biggest Takeaways:   Zebrafish and other model animals provide opportunities to understand host-microbe interactions. Zebrafish are particularly useful for imaging studies, due to their translucent skin and the ease of in vivo microscopy. This allows zebrafish to be used to in studies of spatial architecture or longitudinal studies (imaging the same fish specimen over time) in ways that other model organisms can’t be.   Zebrafish get their first microbes from their mother, just like mammals! The chorion, a protective coating that surrounds the zebrafish embryo, is seeded with microbes from passing through the cloaca of the female zebrafish. Surface-sterilizing this chorion allows researchers to generate germ-free animals that are very useful for microbiome studies.   A gut epithelial transcription factor is regulated by a signal from the gut microbiota, and this signaling interaction is conserved among all vertebrates. The transcription factor itself, HNF4, is found in both complex and simple animals, like the sea sponge, and may serve a long-conserved function in regulating interactions between animals and their microbiota.   Enteroendocrine cells release hormones based on specific chemical cues, but they can also interact with the nervous system. This makes them an important part of the gut-brain system, and the power of in vivo imaging has made zebrafish a great model for better understanding their function. Specific members of the microbiome specifically stimulate these EECs, sending signals up the vagus nerve to the brain.   Featured Quotes:   “We know that the zebrafish functionality of its intestine is very similar to what one encounters in the mouse or human intestine and we and others have been able to translate our findings from zebrafish studies into human biology.”   On genomic studies that have found similar transcription profiles in zebrafish, stickleback fish, mice, and humans: “This suggested that there is a core transcriptome that gut epithelial cell use in different vertebrate species that haven’t shared an ancestor in 420 million years!”   Comparing fish and mouse: “Genes regulated by microbiota in these respective hosts display a lot of overlap. Many of the same signaling pathways and metabolic processes are affected by microbiotas in different hosts in similar ways.”   “There’s been a lot of interesting research documenting the role of the intestinal microbiome in promoting harvest of dietary nutrients we consume. Much of that literature has been focused on the events that occur in the distal intestine, in the colon, where recalcitrant carbohydrates and proteins that make it that far, many of which we are unable to digest, are made available to the colonic microbiome, members of which are able to digest and degrade them to things such as short chain fatty acids, which we can consume.”   “Eventually, we’ll have some strong candidates in terms of specific bacterial strains or communities or factors or pharmacologic agents that could be used to affect dietary fat absorption or metabolism. We’re still a long ways away from that.”   “One of the fascinating things about developmental biology is that the only way you get a viable animal is if the different tissues and the different cells within the body are coordinating amongst themselves for energy, for nutrients, for oxygen, et cetera. As you’re building an animal and as you’re sustaining an animal, the different tissues have to cooperate. When that doesn’t happen, when tissues or cells become selfish or don’t play by the rules, you get things like cancer and other diseases as well...when I began learning about the field of microbiome science and some of the work that was coming out from that field, it sounded to me like the microbiome was going to be a really important part of that. Not only can we think of the microbiome as a ‘microbial organ,’ as it is sometimes called, and therefore worthy of consideration within the context of developmental biology, but also the influence of the microbiome on any one tissue is going to modify its need and its ability to cooperate within the integrated system.”   Links for this Episode:   John Rawls’ lab website More amazing zebrafish images from the Rawls lab Duke University Microbiome Center Genome Research article on HNF4 regulation Cell Host and Microbe article on microbial influence on fatty acid absorption  

Gemma Reguera discusses her studies of Geobacter pili, which transfers electrons to iron oxide and other minerals, and can be used for new biotech applications. Host: Julie Wolf  Subscribe (free) on Apple Podcasts, Google Podcasts, Android, RSS, or by email. Also available on the ASM Podcast Network app. Julie’s Biggest Takeaways: Geobacter sulferreducans, a bacterium that “breathes” rust, is the lab representative of the genus Geobacter that dump electrons onto rust. These specialized microbes use minerals like manganese oxide and iron oxide (also known as rust) for respiration in both terrestrial and aquatic sediments. Although many species are strict anaerobes, a few species can grow under microaerophilic conditions, in which the bacteria will respire the oxygen to eliminate its toxic effects on the cell. Iron oxide respiration relies on the Geobacter pili, a simple structure composed of a single peptide repeat. The pili concentrate on one side of the bacterial cell, where they connect the cell with the iron oxide to release the electrons that have been accumulating. The pili immediately depolymerize and retract, shedding the mineral before returning into the cell. Mass-producing pilin subunits in E. coli took a bit of trouble shooting, but now Reguera and her colleagues can make them on a much larger scale, which bodes well for expanding tests into electronic applications. Commercialization grants address the “valley of death,” the chasm between the technologies developed at the bench and the scale of production necessary for industrialization. Geobacter can bind and reduce many minerals using their pili, including uranium and other toxic heavy metals like lead and cobalt. Using Geobacter pili in agricultural soils or aquaculture waters may help remove these contaminants and improve the health of these ecosystems. Featured Quotes: “I remember when I started as a microbiology student, I think I underappreciated the role that electrons and the movement of electrons play in microbiology.” “There is absolutely not a single process in living organisms that is not energized by the movement of electrons.” “The Earth didn’t have oxygen for the first 2 billion years, if not longer - and there was life! On Earth! Those early organisms were really great at finding minerals, metals, just about anything other than oxygen to dump their electrons, continue to grow, and to colonize the Earth.” “When you start comparing the structure and the amino acid composition of this subunit to any other known bacterial pilins, you really see 2 remarkable changes: one of them is the pilin of Geobacter is very small. the second is that little stick has aromatic amino acids. When the sticks come together to make the filament, they cluster very close to each other and create like a staircase for the electrons to move fast. It’s like a magic combination in which you have the right structural reduction and the right amino acids to really fit like a puzzle to create paths for electrons.” “What has always motivated me is learning something new.” Links for This Episode: Gemma Reguera lab website Gemma Reguera interview on “People Behind the Science” HOM: Thirty-Second Annual Meeting of the Society of American Bacteriologists HOM: Barney Cohen: An Appreciation (Bacteriological Reviews memorial)

Why do Legionnaire’s Disease outbreaks occur mostly in the summer? What is the connection of the Flint change in water source and Legionella outbreaks in the area? Michele Swanson discusses her work on Legionella pneumophila and her path from busy undergraduate to ASM President. Julie’s Biggest Takeaways: Legionella pneumophila is a waterborne microbe that lives in fresh water and can colonize water systems of the built environment. Colonization of cooling systems, like those used in air conditioning systems, can lead to contaminated water droplets that can cause disease. Legionella are very adaptable to different environment, but scientists don’t have great models to determine the exact preferences of the bacterium. After Flint switched water sources from lake to the Flint river, a chemical that prevents corrosion was omitted from the water treatment. This led to lead in the water, which was detected in pediatric patients. An increase of legionella cases in the two years also occurred, and the question was whether the outbreak was related to the shift in water chemistry. Michele joined a team of water engineers, epidemiologists and sociologists to answer this question, and the team found an association between low chlorine levels and high risk of legionella disease. Across the globe, more than 80% of disease is associated with L. pneumophila serogroup 1. The serogroup is based on the bacterial lipopolysaccharide (LPS) structure, which in this strain is very hydrophobic and may allow this serogroup to withstand a higher degree of desiccation than other strains. A urine-based diagnostic test works well, but only to detect serogroup 1. The strain isolated from patients of the Flint outbreak were serogroup 6, as were Legionella isolated from the homes of Flint residents. Featured Quotes: “Amoeba are very good at digesting bacteria, eating them for food, but Legionella, because it’s been under this severe selective pressure of the amoeba, they’ve evolved tools to allow them not only to survive within the amoeba but to replicate within the vacuole of the amoeba.” “We now have equipment that throws water into the air and gives [Legionella] a new opportunity to be ingested by a macrophage. It can then deploy the same tricks it uses to grow inside amoeba to grow inside the macrophage.” “[Human infection] is a tragedy for the patient, but also for the microbe...humans are a dead-end for the bacterium.” “I was really delighted to be recruited to work with this interdisciplinary team on a public health crisis here in my home state. It has opened my eyes to a much more complex pathway and I just feel really privileged in this stage in my career to be able to turn my attention to these larger public health issues.” “People want to hear encouragement; we have a tendency to compare ourselves to those who are 5-10 years ahead of us. Encouragement really is valuable.” Links for this Episode: Michele Swanson at the University of Michigan mBio: Prevalence of infection-competent Legionella pneumophila within premise plumbing within southeast Michigan PNAS: Assessment of the Legionnaire’s disease outbreak in Flint, Michigan Microbial Sciences blog post: Examining Flint: New research highlights lack of Legionella public policy ASM membership