Glutamine Emerges as a Hidden Nitrogen Lifeline for the Leptospirosis Pathogen

Leptospirosis is one of the most widespread bacterial zoonoses on the planet, yet it remains stubbornly difficult to treat in its severe forms. The disease is caused by spiral-shaped bacteria of the genus Leptospira, most notably Leptospira interrogans, which shed into the environment by infected animals and enter humans through contaminated water and soil. Now, a team of researchers led by Matthew H. Ward, Nathan Scherer, Leah P. Shriver, and Gary J. Patti has uncovered a metabolic vulnerability that had gone unnoticed for decades: the pathogen can draw nitrogen from the amino acid glutamine, a nutrient abundant in mammalian blood, to fuel its own proliferation. The discovery, published in PLOS Pathogens, challenges a long-standing assumption about how these bacteria feed themselves and opens a potential new avenue for drug development.

For most of the history of leptospiral research, scientists have grown these fastidious organisms in Ellinghausen-McCullough-Johnson-Harris medium, commonly abbreviated as EMJH. That formulation, while effective for keeping cultures alive in the laboratory, bears little resemblance to the chemical landscape a pathogen actually encounters inside a mammalian host. Nutrient concentrations in EMJH differ dramatically from those in blood plasma, which means that metabolic behaviors observed in conventional cultures may not reflect what happens during a genuine infection. The research team recognized that this mismatch could be masking important aspects of leptospiral biology, and they set out to build a more faithful experimental system.

Their solution was a culture medium they call supplemented Human Plasma-Like Medium, or sHPLM. As the name suggests, the formulation was designed to mimic the composition of human plasma far more closely than traditional media do, with physiologically relevant concentrations of amino acids, vitamins, salts, and other metabolites. The researchers established and validated the medium specifically for Leptospira culture, confirming that the organism could grow in it reliably. By moving the bacteria into an environment that better approximates the host, the team hoped to reveal metabolic dependencies that standard conditions would never expose. The gamble paid off almost immediately.

To interrogate what the bacteria were actually consuming, the group pioneered a set of metabolomics approaches tailored to leptospiral cultures. They developed a liquid chromatography coupled with mass spectrometry workflow capable of performing two complementary tasks: profiling the metabolites present in the medium itself, and tracing the fate of specific labeled nutrients as the bacteria metabolized them. Stable isotope tracing is a powerful technique in which a nutrient is supplied in a chemically identical but isotopically tagged form, allowing researchers to follow exactly where its atoms end up inside the cell. Applying this workflow to L. interrogans growing in sHPLM gave the team an unprecedented, real-time picture of the pathogen’s feeding habits.

The central finding was striking. Among the many nitrogen-containing compounds available in the plasma-like medium, glutamine stood out as a major nitrogen source for the bacteria. This was unexpected because ammonium has long been considered the sole nitrogen source for Leptospira species. Nitrogen is an indispensable element for every living cell, forming the backbone of amino acids, nucleotides, and countless other building blocks, so identifying an alternative and physiologically abundant nitrogen supply fundamentally changes how scientists should think about the pathogen’s nutritional strategy inside the host. Glutamine, one of the most plentiful amino acids in human blood, appears to be far more than passive background chemistry for these organisms.

To confirm that the observation was not an artifact of a single strain or condition, the researchers ran growth experiments across three pathogenic Leptospira strains as well as the nonpathogenic strain L. biflexa Patoc-1. When glutamine-derived nitrogen was the only nitrogen available, all of the tested strains proliferated robustly. That breadth matters. It indicates that the capacity to assimilate glutamine nitrogen is not a quirk of one laboratory isolate but a shared trait across the genus, present in both disease-causing and benign relatives. From an evolutionary standpoint, this suggests that glutamine utilization is a deeply rooted metabolic capability rather than a recent adaptation confined to virulent lineages.

The findings also carried immediate therapeutic implications. The team tested JHU-083, a small-molecule inhibitor designed to interfere with glutamine utilization, and found that it effectively impaired the proliferation of L. interrogans cultures. Because severe leptospirosis currently lacks consistently effective treatments, and because the bacterium’s unusual biology makes it resistant to many conventional antibiotic strategies, a metabolic inhibitor that targets nitrogen assimilation represents an intriguing conceptual blueprint. The result does not mean JHU-083 itself is ready for clinical use, but it demonstrates that blocking glutamine metabolism can starve the organism of a resource it clearly depends on under host-like conditions.

Perhaps the most surprising result came when the researchers added glutamine to the non-physiological EMJH medium. Even in that artificial environment, the amino acid rapidly induced a short-term boost in proliferation and increased biofilm formation in L. interrogans. Biofilms, the structured bacterial communities that adhere to surfaces and often display heightened tolerance to antimicrobial agents, are increasingly recognized as important in persistent infections. The fact that a single nutrient could trigger both faster growth and biofilm development hints that glutamine is not merely food but also a signal, a molecule the bacterium uses to sense its surroundings and reprogram its behavior accordingly.

RNA sequencing reinforced that interpretation. After exposing the bacteria to glutamine, the team observed transcriptional trends pointing toward increased biosynthetic activity, consistent with the organism ramping up its internal manufacturing to support the accelerated growth and biofilm phenotypes. In other words, the presence of glutamine appears to shift the pathogen’s gene expression in ways that prepare it for expansion. If similar signaling occurs during infection, glutamine availability in the host could influence not only how fast L. interrogans multiplies but also how it organizes itself and resists clearance, factors that directly affect disease severity and transmission.

Taken together, the study delivers a two-part message. First, glutamine provides a second nitrogen source for biosynthesis in multiple Leptospira strains, overturning the textbook view that ammonium is the only option. Second, the amino acid may act as a metabolite signal that alters L. interrogans physiology in ways that could shape infection. Both conclusions underscore the methodological lesson embedded in the work: studying pathogens under physiological conditions is not a luxury but a necessity, because nutrient environments that look adequate in the laboratory can conceal the very dependencies that matter most in the host. Nitrogen assimilation now stands as a candidate target for new therapies, and the sHPLM platform combined with isotope tracing offers a template for uncovering similar metabolic blind spots in other understudied pathogens.

Subject of Research: Glutamine utilization as a nitrogen source in the pathogenic bacterium Leptospira interrogans

Article Title: Glutamine from plasma-like medium fuels nitrogen metabolism and influences proliferation in Leptospira interrogans

Article References: Ward, M. H., Scherer, N., Shriver, L. P., & Patti, G. J. (2026). Glutamine from plasma-like medium fuels nitrogen metabolism and influences proliferation in Leptospira interrogans. PLOS Pathogens, 22(10), e1014534. https://doi.org/10.1371/journal.ppat.1014534

Image Credits: AI Generated

DOI: 10.1371/journal.ppat.1014534

Keywords: Leptospira interrogans, leptospirosis, glutamine, nitrogen metabolism, metabolomics, stable isotope tracing, Human Plasma-Like Medium, biofilm, JHU-083, PLOS Pathogens, bacterial physiology, antimicrobial targets