Microorganisms: Infectious Disease and Ecology

Blog entries about bacteria, viruses, fungi and other microorganisms.

Synthetic Biology: Minimal Cell, Maximal Opportunity

Posted on by Ken Doyle

According to the National Human Genome Research Institute, synthetic biology is “a field of science that involves redesigning organisms for useful purposes by engineering them to have new abilities”. Synthetic biology has a broad range of applications, from manufacturing pharmaceuticals and other biologically active chemicals and biofuels, to accelerating the adoption of plant-based burgers (1).

At the heart of the synthetic biology revolution is the rapid technological advancement—and accompanying drop in costs—of DNA oligonucleotide synthesis. Typically, synthetic biology researchers use oligonucleotides as building blocks to assemble genes of interest that are then introduced into, and expressed by, a different organism. For example, to create the plant-based Impossible Burger, the soy leghemoglobin gene (normally found in the root nodules of leguminous plants) was synthesized and expressed in yeast cells (1). This component gives the burger its meaty flavor and appearance of “bleeding” when cooked.

An Impossible Burger served with fries on the side

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2023 Promega iGEM Grant Winners: Tackling Global Problems with Synthetic Biology Solutions

Posted on by Michele Arduengo

On June 15, 2023, we announced the winners of the 2023 Promega iGEM grant. Sixty-five teams submitted applications prior to the deadline with projects ranging from creating a biosensor to detect water pollution to solving limitations for CAR-T therapy in solid tumors. The teams are asking tough questions and providing thoughtful answers as they work to tackle global problems with synthetic biology solutions. Unfortunately, we could only award nine grants. Below are summaries of the problems this year’s Promega grant winners are addressing.

UCSC iGEM

An immature night heron against the green surface of Pinto Lake. 2023 Promega iGEM Grant Winner, UCSC iGEM seeks to mitigate these harmful aglal blooms.
A night heron hunts on Pinto Lake, California.

The UCSC iGEM team from the University of California–Santa Cruz is seeking a solution to mitigate the harmful algal blooms caused by Microcystis aeruginosa in Pinto Lake, which is located in the center of a disadvantaged community and is a water source for crop irrigation. By engineering an organism to produce microcystin degrading enzymes found in certain Sphingopyxis bacteria, the goal is to reduce microcystin toxin levels in the water. The project involves isolating the genes of interest, testing their efficacy in E. coli, evaluating enzyme production and product degradation, and ultimately transforming all three genes into a single organism. The approach of in-situ enzyme production offers a potential solution without introducing modified organisms into the environment, as the enzymes naturally degrade over time.

IISc-Bengaluru

Endometriosis is a condition that affects roughly 190 million (10%) women of reproductive age worldwide. Currently, there is no treatment for endometriosis except surgery and hormonal therapy, and both approaches have limitations. The IISc-Bengaluru team at the Indian Institute of Science, Bengaluru, India, received 2023 Promega iGEM grant support to investigate the inflammatory nature of endometriosis by targeting IL-8 (interleukin-8) a cytokine. Research by other groups has snow that targeting IL-8 can reduce endometriotic tissue. This team will be attempting to create an mRNA vaccine to introduce mRNA for antibody against IL-8 into affected tissue. The team is devising a new delivery mechanism using aptides to maximize the delivery of the vaccine to the affected tissues.

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Streamlining Disease Diagnostics to Protect Potato Crops

Posted on by AnnaKay Kruger
A potato farmer holds a handful of potatoes. Scientists are working to protect potato crops from disease.
The WSPCP works to provide seed potato growers with healthy planting stock

The mighty potato—the Midwest’s root vegetable of choice—is susceptible to a variety of diseases that, without proper safeguards, can spell doom for your favorite side dishes. Founded in 1913 and housed in the Department of Plant Pathology at the University of Wisconsin-Madison, the Wisconsin Seed Potato Certification Program (WSPCP) helps Wisconsin seed potato growers maintain healthy, profitable potato crops year-to-year through routine field inspections, a post-harvest grow-out and laboratory testing.

While WSPCP conducts visual inspections for various seed potato pathogens, their diagnostic laboratory testing is primarily focused on viruses such as Potato virus Y (PVY), which can cause yield reduction and tuber defects, along with select bacteria such as Dickeya and Pectobacterium species that cause symptoms like wilting, stem rot and tuber decay.

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Phage Therapy: Meeting the Challenge of Drug-Resistant Bacterial Infections

Posted on by Ken Doyle

Global pandemics, such as COVID-19, have taught us to abhor viruses. The emergence of new, highly infectious viruses is—rightfully so—a cause for concern. However, despite the average human body harboring 380 trillion viruses, most of them simply coexist with us and are harmless. When it comes to an ancient lineage of viruses within the realm Duplodnaviria, researchers are even using them as weapons in the battle against infectious diseases.

In 1915, Frederick William Twort, an English bacteriologist at the University of London, reported the discovery of an unusual “ultramicroscopic virus” (1). Twort was culturing vaccinia virus as part of an experiment to determine if he could prepare smallpox vaccines in vitro. These vaccines, made in calves, were typically contaminated with Staphylococcus bacteria. When Twort plated the vaccines, he found small, clear areas on the agar plates where the bacteria would not grow, and these clear areas were the source of his ultramicroscopic virus. Two years later, a French-Canadian microbiologist, Félix d’Hérelle, independently discovered a similar phenomenon when culturing Shigella bacteria from fecal samples of patients with bacillary dysentery. He called the new virus “un bactériophage obligatoire” (2). Shortly after his discovery, he found that bacteriophages (phages) could be used as powerful agents to treat a variety of bacterial infections, and the field of phage therapy was born (3).

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The Microbial Secrets that Lie within Yellowstone National Park Hot Springs

Posted on by Anna Bennett
picture of grand prismatic hot spring; steam rising up from orange and yellow hot springs pools
Grand Prismatic Spring, Yellowstone National Park; Photo Credit: Anna Bennett

Yellowstone National Park —located partially in Idaho, Montana and Wyoming—puts modern volcanic activity on full display. Near boiling, ominous pools of water in the form of geysers, mud pots, fumaroles (vents that release steam) and hot springs are all present and active in the park and visitors flock to the park to view a handful of thermal features every year during the peak summer visitor season. Coincidentally, this is when a large portion of scientific research also takes place at the park. Combining both the boardwalk paths that are open to all who visit the park and the expansive backcountry, Yellowstone is host to over 10,000 thermal features. These thermal features are fed by superheated water that travels through a complex groundwater system—think the pipes under your kitchen sink—where subsurface water collects gases and chemical compounds en route to the surface. As a result, near-boiling water that bubbles through to the surface is often rife with chemicals like sulfur, iron or magnesium. Early scientists thought of hot springs as uninhabitable, but as it turns out, these conditions are just the right environment for thermophilic (or “heat-loving”) bacteria to thrive.

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New Vaccine for Honeybees Could Take the Sting Out of Devastating American Foulbrood Disease

Posted on by Kelly Grooms

Our world is a complex, interdependent system, and invertebrate pollinators such as honeybees play a pivotal role in its survival. Threats to populations numbers of pollinators like honeybees can be equated to threats to the overall health and survival of the ecosystem in which they live. Of the over 20,000 known bee species, one—the western honeybee (Apis mellifera)—acts as the single most frequent pollinator for crops worldwide (1). Found on every continent except Antarctica, the western honeybee owes its status as a top pollinator to its widespread geographic distribution, generalist foraging behavior and competence as pollinators (1).

Deadly American Foulbrood Disease

Honeybees are the most economically valuable pollinators and are threatened by several pathogens (2). Perhaps the biggest threat to honeybee colony health and survival is the bacterial disease, American Foulbrood (AFB; (3). Caused by the spore-forming, Gram+ bacteria, Paenibacillus larvae, the highly contagious AFB disease affects the young brood of colonies.  When newly hatched larvae are fed spore-contaminated food, the spores germinate and replicate causing septicemia and death. P. larvae spores are incredibly resilient and can remain viable for decades (3). Each infected larva can produce over 1 billion new spores.  Thus, a colony can produce large numbers of spores with just a few cases of symptomatic brood (4).

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Growing Our Understanding of Rose Rosette Virus Through Reverse Genetics

Posted on by Kelly Grooms
Viral particle representing lethal Rose Rosette Emaravirus with red rose.

Roses, the universal symbol of love and affection, are one of the most popular ornamental flowering shrubs used by landscapers and home gardeners and account for almost half of the billion-dollar ornamental plant market. The growing prevalence of rose rosette disease poses a significant threat to these industries.  This lethal disease is caused by the Rose rosette emaravirus (RRV) and transmitted by the tiny eriophyid mite, Phyllocoptes fructiphilus. Infection by RRV results in prolific growth of clustered and bunched plant shoots (witches’ broom), malformed flowers and leaves, malformed shoots and enlarged stems and abundant leaf growth and thorniness. This excessive growth depletes the plant’s energy, eventually causing death.

Emerging and Devastating Plant Viruses of the Genus Emaravirus

RRV is a single-stranded, segmented, negative-sense RNA virus belonging to the genus Emaravirus, a relatively new genus that was established in 2012. These emerging viruses can be devastating to trees, herbaceous woody plants and vines. At Texas A&M University, Dr. Jeanmarie Verchot’s lab is working to better characterize and understand these new viruses. In addition to threatening roses, these viruses cause damage to important agriculture crops such as wheat and pigeon peas. They also endanger sensitive ecosystems when they infect plants specialized to a particular habitat, as is the case with Palo verde broom virus infection of palo verde trees of the Sonoran Desert (1).

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A Virus Makes a Comeback: Emerging Poliovirus Transmission in the West

Posted on by Michele Arduengo

In 1921, at age 39, Franklin D. Roosevelt, the man who would later be elected the 32nd president of the United States, was diagnosed with polio (poliomyelitis). His symptoms included fever, gastrointestinal issues, numbness, and leg and facial paralysis. The disease left him paralyzed from the waist down, relying on a wheelchair and leg braces to walk.

The paralysis from poliovirus infection affected the involuntary muscles that allow breathing, and iron lungs were used to keep patients breathing until they cleared the infection.
The paralysis from poliovirus infection affected the involuntary muscles that allow breathing, and iron lungs were used to keep patients breathing until they cleared the infection.

At the height of the polio epidemic in 1952, more than 3,000 people died of polio in the United States, and 20,000 more people suffered paralysis. Pictures of the era show children in special hospital wards, inside ominous-looking iron lungs, while “recovered” children played on the grounds of hospitals wearing leg braces.

In 1938, Roosevelt founded the March of Dimes, which funded the development of the Salk polio vaccine. Two years after the introduction of the Salk vaccine in 1955, polio cases in the US dropped by 90%. In fact, sustained polio transmission has been absent from the US for nearly 40 years; according to the CDC, the last case of wild poliovirus in the US occurred in 1979.

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PROTAC Virus Vaccines: A New Approach to Vaccine Development

Posted on by Michele Arduengo

Vaccine research and development is a major area of focus for life scientists across the globe. Clinical trials have shown that vaccines that target tumors show promise for cancer treatment. Additionally, the emergence of new zoonotic diseases has revealed a need to develop vaccines quickly as the world becomes more global and human populations interact more often with each other and wild habitats. Importantly, these vaccines need to be suitable for distribution in a variety of settings, including those that do not have easy access to refrigeration.

Influenza Virus. Si et al used influenza as a model to engineer and test PROTAC Virus vaccines

There are many ways to classify the different types of vaccines that are currently available. The National Institute of Allergy and Infectious Diseases in the United States, categorizes vaccines as: whole pathogen vaccines, subunit vaccines, and nucleic acid vaccines—based on how the antigen that stimulates the immune response is delivered to the host.

Whole-pathogen vaccines, which include many of vaccines used in clinical settings, use the entire pathogen (organism that causes the disease) that has been either weakened or killed to elicit a protective immune response. Killed vaccines are what their name implies: the pathogen has been killed so that it cannot cause disease, but enough of its structure remains to generate antibody response. Often, the immune response generated with killed vaccines is not as robust as that generated with other kinds of vaccines. 

Weakened or attenuated vaccines use whole pathogens that have been weakened in the laboratory through long-term culture or other means. Our modern MMR (measles, mumps and rubella) vaccine is an example of an attenuated vaccine. These vaccines tend to generate strong, long-lasting immune responses, but have increased risk for immunocompromised individuals.

Engineering an Influenza A PROTAC Virus Vaccine

A recent paper by Si et al published in Nature Biotechnology describes a new type of live-attenuated whole pathogen vaccine: the PROTAC virus. PROTAC viruses are prevented from replicating by targeting critical viral proteins for degradation using the host cell protein degradation pathway. The vaccine is live-attenuated by the host cells that degrade critical proteins.

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Detecting SARS-CoV-2 In Wastewater: The New Frontier in Pandemic Surveillance

Posted on by AnnaKay Kruger

Tracking the spread of COVID-19 has been a tremendous challenge throughout the pandemic, but doing so is a key step toward containing the virus. Many communities have relied on patient testing and contact tracing, with limited success. In search of better methods, some countries have made inroads in a different form of disease surveillance: wastewater-based epidemiology (WBE). This approach involves testing wastewater for the presence of pathogens, primarily through DNA and RNA analysis, and has proved to be an accurate and highly effective way to keep tabs on the prevalence and progression of COVID-19 at the population level.

Switzerland is among those countries that have implemented WBE in their efforts to stay ahead of the pandemic. Since WBE first emerged in 2020 as a promising tool, several Swiss laboratories undertook wastewater testing, and protocols were established early.

“At the beginning, the methods to actually detect coronavirus in wastewater were rather laborious and complicated, and involved a lot of resources,” said Dr. Claudia Bagutti, microbiologist and molecular biologist in the State Laboratory of Basel-City, Switzerland.

Bagutti heads a small team performing applied biosafety research. In 2020, her lab was tasked with developing an assay for detecting COVID-19 in wastewater. However, the available methods were prohibitively complex and resource intensive.

In the meantime, researchers at Promega recognized that Promega products and methodologies could potentially be applied to WBE and set to work developing simpler and more efficient method for wastewater analysis. In the spring of 2021, Bagutti’s team decided to try adopting this method.

“Promega had a very nice method which was less laborious and much easier to handle, and that’s why we gave it a try,” said Bagutti.

In the ensuing study, Bagutti and her team analyzed effluent from the catchment area of one municipal wastewater plant in Switzerland. They examined the total wastewater output of around 270,000 people. Viral RNA was extracted using Promega’s Maxwell® RSC Environ Wastewater TNA Kit. The number of RNA copies present, representing the overall concentration of COVID-19 in each sample, was determined via quantitative reverse transcriptase (RT-qPCR) using the GoTaq® Enviro Wastewater SARS-CoV-2 Systems, also from Promega. The viral RNA was subsequently sequenced with next generation sequencing, and the results correlated quite well with the COVID-19 cases in the catchment area. Remarkably, this study detected the Omicron variant in a wastewater sample one day prior to the first reported case identified through patient testing.

“We observed a similar spread to most other western countries with respect to the time of the first discovery of these variants,” said Bagutti. “We were also able to demonstrate the presence [of Omicron] in the wastewater before it came up in a sample of a COVID-19 patient test, which of course shows the usefulness of wastewater monitoring for the prediction of new variants and infection dynamics.”

WBE is especially promising in that it provides population-level data independent of patient testing. Health departments can be alerted to the presence of COVID-19 earlier than would otherwise be possible with traditional testing and can take precautions to contain the spread. In creating a more user-friendly method for wastewater analysis, Promega has opened the door for more laboratories to conduct WBE, which could provide communities around the world with the information they need to preempt the progression of COVID-19.

“The Promega method is very straightforward to handle,” said Bagutti. “It only takes a small volume of wastewater, which makes it handy. It’s less time-consuming compared to the methods which were in the literature at the beginning of the pandemic, and it just works very well. We also did experience great support from Promega.”

At this point, much of the wastewater analysis performed in Switzerland is done with the Promega method, including in federal, state or private labs. The swift advance of WBE in Switzerland speaks to the colossal effort put forth both by Promega researchers in developing the necessary products and methodologies, as well by those labs that have made use of Promega’s products to monitor COVID-19 in wastewater.

“It’s really been a success story for us, from the beginning,” said Bagutti.

Learn more about Promega’s work with wastewater-based epidemiology.