by Daniel Keays

 

Pathogenic Shapeshifter

Ground-to-air attack,

Disruption liberation.

Fungus among us.

 

            In 1891, at the very infancy of understanding the nature of infectious diseases, a medical student in Argentina made a remarkable discovery. A patient named Domingo Ezcurra, a soldier in the military stationed in Buenos Aires, Argentina, presented at clinic with a peculiar-looking purple lesion on his cheek. He also had lesions on his nose, arm, and trunk. First believed to be skin cancer, a biopsy was taken and examined after staining. With careful observation under the microscope, young doctor Alejandro Posadas observed some curious-looking round structures. To the best of his knowledge at the time, the organisms he noticed appeared to be a type of parasite, perhaps one related to the group known as the Coccidia, a large group of parasites that infect a wide range of mammals. The best-known among the human species of Coccidia is Toxoplasma gondii. What he actually discovered, though, was the first example of a fungus that infects humans rather commonly in some regions of the world, Coccidioides immitis. 

            This wasn’t just any fungus. We’re all familiar with fungi that grow as fuzzy white, green, or some other color creature on an orange or slice of bread. That type of fungus is well-characterized and well-studied and usually is of no consequence to human health. But this pathogenic fungus first observed by Dr. Posada is different. It changes its shape and nature depending on where it resides. In the soil, it looks much like other fungi, with mycelial strands forming a matrix of growth. When it enters the body, though, it becomes something very different, with rounded bodies containing multiple spores. Because of this double way of existing, the term dimorphic is applied: di meaning “two,” morphic meaning “shapes.” 

            Since it resembled the Coccidia group of parasites, the genus name Coccidioides, or coccidia-like, was applied. In the U.S., much of the early work on the fungus was carried on in the San Joaquin Valley of California. The fungus is resident in the soil in that area, and in the 1930s many immigrants from the Midwest came to the valley to work in agriculture. Being new to the area, they encountered the fungus for the first time, and some became ill. Scientists studying the infection in the San Joaquin Valley were able to provide a great deal of information about the disease.

              Because some cases are very serious, the Latin term immitis, meaning “not small” or “harsh,” was assigned. A similar fungus, the one discovered by Dr. Posadas, is found in parts of South America. It is assigned the name Coccidioides posadas, named for its discoverer.

            The official disease name for the ailment, coccidioidomycosis, is pretty hard to both pronounce and spell, so the more familiar term “valley fever” is usually used. Workers in the healthcare industry often use the term “cocci.”

 

            Coccidioides lives in alkaline, arid soil. This fits the general description of the soil found in the Sonoran Desert, a broad area of the southwestern United States and Baja California, Mexico. It is also found in parts of Central and South America. That sounds simple enough, but in reality, the fungus is very difficult to find just by looking at dirt. A broad swath of land that appears homogeneous might contain very little fungus, which is confined to one small, unique area. It fares better in soils that have elevated levels of minerals such as sodium, sulfates, and magnesium. The organism does not exist on the top of the soil but down about 6-20 inches. How it gets down there is a matter of conjecture, perhaps by rain or hooking a ride on some desert rodent or reptile. There is good evidence to show that the fungus increases in amount in years of elevated rainfall. In endemic areas, this would be during the winter months. It grows primarily as mycelia in the soil during the wet season but forms infectious arthrospores when it dries out in late spring and summer. It contains enzymes that allow it to decompose organic material, so it probably lives where some animal has died or has left some of its food. If it weren’t for the fact that it can cause significant human disease, the organism would be of no real importance. But it does indeed cause illness, sometimes very serious, and much has been learned about the fungus’ biology and lifestyle. 

            Since it lives half a foot or more under the soil, it can infect only when the ground has been disrupted, either by natural means like a windstorm, or by extensive digging, such as a construction project. In the soil, the organism exists as mycelia, or strands of long, branching cells. In this regard, it is like many run-of-the-mill fungi. Where it differs significantly is the way it produces spores. Many fungi we are familiar with produce spores on a stalk that project up from the mycelia. A breeze or some other physical event spreads the spores. Think of the fungus you see on an orange, giving off a small cloud of spores when you pick it up. Coccidioides lives under the soil, so this type of spore production wouldn’t work for it. Instead, it makes what is known as arthrospores, a hardening of the hyphae at regular intervals. The prefix “arthro” comes from the Greek, meaning a joint, or combination of two adjoining structures, in this case, two sections of mycelia. The arthrospores are tough and can remain in the ground until the soil is disrupted. Then they fly around like any other fungal spore, blindly trying to find a new place to set up house. If it is on the ground of the desert, it doesn’t much matter, but if the arthrospores find their way into the trachea and lungs of a human, trouble begins.

            Arthrospores are rectangular shaped, about five microns long. On the outside of them is a hardened coat to protect them from the environment. It doesn’t take many arthrospores to set up an infection. Experiments in mice have shown that a single spore is all it takes. Generally, the more spores inhaled, the sooner symptoms will be displayed. Once they enter the lungs, the coat is dissolved away.

             The remaining structure rounds up and is known as an endospore. The endospore begins a meticulous and intricate phase when liberated inside the body. It sets up a series of walls, dividing the growing structure into many compartments. Inside each little compartment is a new endospore.  When allowed to grow unchecked, the single initial endospore makes a rather large, multi-compartmented structure known as a spherule. Spherules are round, about 30 microns in diameter, and contain about one hundred endospores. Each endospore can create a new spherule, which is what happens. When a spherule gets large enough it ripens, bursts, and releases its many endospores, each capable of initiating another spherule. Clearly, this is not good for the infected person. It doesn’t take long for many spherules to infest the lung. They don’t invade the lung cells but grow in the spaces between the cells of lung tissue. This crowding eventually restricts the infected person’s air space, and shortness of breath ensues. It usually takes 7 to 21 days for symptoms to appear after exposure.

 

            In some cases, endospores and spherules escape the lungs. If uncontrolled by the immune system, the endospores can use the bloodstream or lymphatic channels to traverse the body and end up in remote locations. The skin, especially the area around the nose and lips, is the organ most commonly affected. Coccidioidomycosis can also occur in joints, bones, and the intestinal tract. Meningitis is nearly always fatal if not aggressively treated.

            Of course, the immune system cells recognize the danger of the invader and are mobilized. Arthrospores and endospores are small enough to be gobbled up by neutrophils and macrophages, and they bear on their surface a protein that alerts the phagocytes they are foreign material. The fungal protein is called Spherule Outer Wall glycoprotein, or SOWgp. Known as a pathogen-associated molecular pattern, or PAMP, the surface material gives the arthrospores away, and they wouldn’t last long if SOWgp is left there. To mask itself, in the early stages of its growth, the organism produces an enzyme called metalloproteinase (Mep-1) to digest the tell-tale SOW and get rid of it. If the fungus successfully removes the PAMP it stands a better chance of evading the phagocytic cells designed to attack it.

            Spherules are, of course, much larger than endospores, so neutrophils can’t handle them.  That leaves it up to CD4 T-helper cells. The immune attack on spherules is similar to that conducted against tuberculosis: wall it off with the formation of a granuloma. The infection can be contained as long as the T-cells are in good working order. If they aren’t, it can spell trouble.

            

            Our macrophages and dendritic cells express a group of signaling molecules called Toll-like receptors. Each is designed to react when they encounter a molecule unique to a microbe, sending signals to unleash the molecules of the immune response. In addition to the TLR group, we also have a group of microbe-detecting and signaling molecules called the C-type lectin receptors. An important member of this group is a protein called Dectin-1. Dectin-1 has a specific receptor for a portion of a fungal cell wall, glucans. When they encounter this fungal PAMP, they spring into action, giving a chemical signal to the host genome to unleash appropriate cytokines and other defensive molecules. Two very important immune responses initiated after Dectin-1 becomes active and sends its chemical signals are the Th₁₇ subset of CD4 lymphocytes, and interferon-gamma.

            The T-helper lymphocytes, which bear the chemical marker CD4, are extremely important in aiding the body’s fight against invading microbes. There are several sub-types of the CD4 T-helper lymphocytes. The one known as the Th₁₇ variety is activated by the activity of Dectrin-1 after encountering an invading fungus. Among the T-helper Th₁₇’s most important roles are the chemical signals they send to enhance the creation of neutrophils in the bone marrow, and the neutrophils’ attraction to and concentration in an infected body site. Neutrophils are very effective attackers of Coccidioides endospores, and the more of them at the infected site, the better. The CD4 Th₁₇ lymphocytes, stimulated by Dectin-1, are vital in this endeavor.

            Macrophages wandering through lung tissue are of two main types, designated M1 and M2. The M1 types are warriors designed to kill invading microbes, mainly by destructive enzymes.  The M2 macrophages are tasked with tissue repair. Both are, of course, important. A naïve macrophage gets its signals from cytokines: interferon-gamma stimulates them to turn into M1, and interleukin-4 directs the development of the M2 type. Without the stimulated M1 macrophages, the body’s fight against the invading fungus is significantly disadvantaged. Absent the production of interferon-gamma and the attendant recruitment of active M1 macrophages, coccidioidal endospore and spherule production can advance quite rapidly. Dectin-1 and its chemical signals help direct this important mission.

 

            For most people, inhaling Coccidioides arthrospores does not result in a serious infection. Neutrophils engulf and digest many of them, and M1 macrophages, activated by interferon-gamma, handle the rest. Neutrophils have many destructive enzymes and any time they are active in an area there is the possibility of local tissue damage, but the M2 macrophages entering the area are very good at cleaning that up. Depending on how many arthrospores are inhaled, the efficiency of the fungus’ ability to hide its outer protein, and the number of neutrophils and macrophages in the area, the disease in most people ranges from asymptomatic to a relatively mild flu-like disorder that resolves in a week. In some, it can persist for several weeks or even months, but most cases resolve uneventfully. Many who are infected don’t know Coccidioides infected them because they are not tested for it; it just seems like a common viral infection.

            In some individuals, though, infection by Coccidioides is devastating. The fungus progresses at an alarming rate, with the cycle of spherule production, endospore release, and new spherule production progressing at a furious pace. Damage to lung tissue is great, and some of the endospores can escape the lungs, enter the bloodstream or lymphatic channels, and begin to reproduce in other areas of the body. Great systemic damage can ensue.

            The attack by neutrophils and macrophages against arthrospores and endospores is part of the innate immune response. The adaptive immune system must be engaged if the infection overwhelms the innate system. The principal mechanism of the adaptive immune response against dimorphic fungi is the activity of CD4 lymphocytes, working in concert with macrophages. Principal among the CD4 cells involved in fighting off Coccidioides are the Th₁ and Th₁₇subgroups of cells. Working in concert, they manage the overall coordination of the defense, usually with a good outcome.  But in some people, and it is often difficult to predict just whom, the marshaling of the proper CD4 lymphocyte mix is impaired, and the full thrust of the immune system defense is diminished. Certainly, individuals who have human immunodeficiency virus (HIV), and therefore markedly reduced numbers of CD4 lymphocytes, are in great danger of uncontrolled Coccidioides infection. That applies as well to someone receiving immunosuppressive therapy. But some people not known to have a suppressed immune system sometimes also develop an overwhelming infection, requiring aggressive, often toxic, anti-fungal therapy. The exact nature of some people's immune system failure to meet the fungal challenge is often elusive. It is probably due to a genetic anomaly giving an imbalance to their adaptive immune response. Members of some racial groups, such as Africans and Filipinos, are at many times greater risk for disseminated infections than whites. Also, pregnant women, especially those in the third trimester, are at higher risk for disseminated infection.

            The number of cases of coccidioidomycosis in the southwestern United States, primarily California and Arizona, has been growing in the last couple of decades. Just how much it has grown is subject to speculation. Certainly, population growth in endemic areas directly influences the total number of cases. Better diagnostic testing helps, especially with minimally symptomatic patients. But some serologic tests are subject to false negatives and false positives and may give misleading information, so the answer remains elusive.

            Unfortunately, anti-fungal therapy does not kill all invading organisms. It is more likened to bacteriostatic agents against bacteria, which slow down growth without eliminating all organisms. By reducing the number of bugs, we depend on the immune system to clear the remainder. But in disseminated disease that usually doesn’t work, and therapy must be maintained for years.

 

            Laboratory workers have known for a long time that Coccidioides immitis is a highly infectious organism. It can grow very well on artificial laboratory culture media designed to grow routine bacteria, giving a fuzzy little non-descript creature looking all the world like a contaminant. It’s a noteworthy hazard of the occupation. But because it is so easy to grow on lab media and so few arthrospores are needed to start an infection, the fungus has been classified as a potential bioweapons threat. It wouldn’t be able to kill millions, but it can sicken many, and its widespread use would certainly sow panic. 

            There is currently no vaccine for coccidioidomycosis. Treatment with anti-fungal medications has not been shown to be effective for mild cases, but for serious cases, drugs such as amphotericin and fluconazole can manage the disease, but unfortunately, they usually don’t eliminate it. 

 

PHIL

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