Pilobolus crystallinus

Pilobolus crystallinus
Scientific classification
Kingdom: Fungi Phylum: Zygomycota Class: Zygomycetes Order: Mucorales Family: Pilobolaceae Genus: Pilobolus Species: P. crystallinus
Binomial name
Pilobolus crystallinus var. crystallinus (F.H. Wigg.)Tode(1784)

Description and significance

The species Pilobolus crystallinus is a type of spore forming fungus that falls into the order of a mucor that grows in the feces of herbivorous animals and decays it. Two other common names for this fungus are the "Dung Cannon" or the "Hat Thrower". It has predominantly been found growing in China. It has also been found growing in eastern and midwestern United States and the Grand Canyon National Park. This fungus begins its life cycle in the form of a sporangium that has been discharged onto some grass. Herbivorous animals which are animals that eat grass such as cows, horses, deer’s, etc. comes along and eat the grass with the sporangium sitting on it. The sporangium passes through the digestive tract of the animal without causing any harm to the animal and without germinating. It is excreted outside the host in the feces of the animal where it begins germination growing as mycelium. They grow 2-4 centimeters tall under surfaces where oxygen concentration is low because oxygen prevents radial growth of it hyphae. Radial expansion of the hyphae occurs when the partial pressure of oxygen is less that 30 mm Hg. The hyphae expand beneath the surface when the partial pressure of oxygen was greater than 30 mm Hg and growth cease at atmospheric conditions of 760 mm Hg. The pilobolus decays the feces of the animal using the nitrogen, water etc. found in the feces as its source of food. Once the nutrients in the feces are depleted the pilobolus shoots it sporangium away from the feces into a new set of grass where it will sit waiting again to be eaten by herbivorous animals so it can repeat the lifecycle. McVickar in 1942 and later Ootaki et al. in 1993 categorized the development of pilobolus crystallinus into six stages. In the first stage there is elongation of the sporangiophore at the apex without rotation. The second stage a sporangiophore develops into a sporangium. In the third stage no growth occurs and in the fourth stage a subsporangial vesice starts expanding under the sporangium. Stage five spore maturation occurs and the hypha region below the subsporangial vesicle elongates and in the last stage, stage six the subsporangial vesicle bursts open and the sporangium are releases into the air.

This time lapse video of pilobolus crystallinus spore development and dispersal was documented by Dave Kalb and Kent Loeffler from Cornell University

Genome structure

There is not much sequencing of the genome that has been done for this fungus. There has been partial sequencing of the large subunit ribosomal RNA gene, partial sequencing of the 18S and 28S ribosomal RNA gene. The 5.8S ribosomal RNA gene has been completely sequenced. Putative blue-light photoreceptor was used to study mRNA to isolate and characterization of MADA homolog in pilobolus crystallinus.

Cell structure and metabolism

Scanning and transmission electron microscope has been used in studying the structure of pilobolus crystallinus. The stalk which is a structure that resembles a plant stem is transparent and it rises above the feces. The stalk is always oriented towards the sunlight which increases the chances that the sporangium will land on a new set of grass once they have been dispersed. The subsporangial vesicle is a balloon like structure found at the end of the stalk. This vesicle fills with fluid and builds up a pressure called turgor pressure. Turgor pressure is the force that keeps the pilobolus upright and once the pressure builds up to 7 ATM or greater and the volume reaches 0.08 cubic millimeters the pilobolus is able shoot its sporangium to a fresh set of grass up to 2 meters away which is 6 feet reaching speed of 0-45 mph during the first millimeter of travel. The subsporangial vesicle also serves the function as a lens focusing light towards the base of the vesicle through the carotenoid pigments. On top of the subsporangial vesicle there is one black sporangium which is the structure that produces and contains the spores. The spores within the sporangium are called sporangiospores and the sporangiospore is an asexual fruiting structure found in the pilobolus species of fungus. There are two different sizes of calcium oxalate crystals, some large and some small covering the surface of the sporangium. These crystals are hydrophobic so when the spores of the pilobolus are shot into the air the hydrophobic part is repelled by the dew on the grass causing the sporangium to flip over so that they land on their bottom which has a sticky mucus like substance. The large crystals surround the spines with a central pore which serves as protection. The vesicle contains a mucus like substance which enables the spores to stick to the grass it lands on. Pilobolus crystallinus travels through the digestive systems of animals and begins it growth in their feces. It reproduces asexually through the hyphae called the sporangiophore. The mycelium of pilobolus crystallinus spreads throughout the feces of the animal and its function is to absorb the nutrients found in the dung of the animal.

Ecology

Pilobolus crystallinus is a corprophilous or dung loving fungi. It is a heterotrophic organism meaning it is not able to produce its own food and depends on the nutrients in the feces of herbivorous animals as its food source. The feces of herbivorous animals contains many nutrients which include undigested plant remains, large quantities of mostly dead bacteria, water soluble vitamins, growth factors, carbohydrates and mineral ions such as corpogens It has a high nitrogen content, high water content, broken down red blood cells and bile pigment. Pilobolus crystallinus produces exoenzymes to digest its food outside its body and then it absorbs the nutrients and although it does not utilize all these nutrients in the animal feces it is dependent on the nitrogen source found in the dung of animals as it food source. It also needs the water found in the feces of animals in order to disperse its spores. Pilobolus uses the corpogen in the feces for its growth and reproduction. Pilobolus crystallinus has some unique features to enhance its chances for survival. They have the ability to disperse their spores up to six feet away to a new food source since herbivorous animals do not eat near their feces. It has phototropic response which means the entire sporangium grows towards sunlight and the sporangia are usually dispersed around mid-morning when the sun is at its highest position in the sky so the sporangia is shot at a 45 degree angle maximizing the distance that the sporangia will travel. This determines the direction of the spores when they are dispersed and since grass grows in sunlight, this increases their chances that they will land on grass.Light rays that falls within the range of blue light of 450nm stimulates the phototrophic response in pilobolus crystallinus.The dark pigmentation of the spores protects them from being damaged by ultraviolet light. The spores vesicle has a sticky mucus that allows the spores to stick to vegetation. The spores does not germinate within the digestive tract of the herbivorous animals so they are able to with stand the high temperature and digestive enzymes of the gut. Nematodes has a symbiotic relationship with pilobolus crystallinus because they assist the fungi with decomposition of the animal feces but when the nutrients become depleted the nematodes larva usually sits on top of the fungi where it gets launched towards a new food source.

Pathology

Pilobolus crystallinus is not a parasitic organism and there have been no reported cases of this fungus causing any diseases in animals or plants. In the book, Diseases Of Greenhouse Crops And There Control by Jacob J. Taubenhaus there was one reported case by Clinton of mechanical spotting on rose plants that was caused by pilobolus crystallinus. It was found that two rose beds that were mulched with cow manure resulted in roses that had black spots on the rose foliage and blossoms. The black spots were a result of the spores of the pilobolus getting stuck to these parts of the flowers when it was discharged. The problem did not incur great losses to the farmers and the problem resolved once all the spores were discharge. Herbivorous animals such as deer’s, cattle, elk, horses’ etc. can become infected by parasitic organisms called lungworms. The larva of the lungworm is excreted in the feces of these animals. The larva can position themselves on the sporangiophores of philobolus crystillinus and it is launched with the sporangium where they now have the ability to infect a new host.

Current Research

Observations on relationships between infective juveniles of bovine lungworm, Dictyocaulus viviparus (Nematoda: Strongylida) and the fungi, Pilobolus kleinii and P. crystallinus (Zygomycotina: Zygomycetes)

Robinson in 1962 filmed the lungworm larvae called dictyocaulus viviparous which is parasite to herbivorous animals as it climbs on to the sporangia of pilobolus crystallinus to be dispersed with the sporangia. It was found that about 50 larvae climbs onto the sporangia of the pilobolus to be discharged. These nematodes like pilobolus crystallinus has phototropic responses. Dictyocaulus viviparous is at risk for desiccation or drying out during the process of spore dispersal. However, the dried sporangia of pilobolus crystallinus forms a sheath around the lungworm larvae keeping the larvae enclosed and protecting them from water loss and from being washed away by the rain. It was also found that the walls of the sporangia of pilobolus crystallinus protects the dictyocaulus from ultraviolet rays damage giving them better chances for survival.

Effects of microbeam light on growth and phototropism of Pilobolus crystallinus sporangiophores

This study done by Kubo and Mihara used blue-light beam to analyze the light beam effects on the growth of the sporangiophore. They used a blue-light beam 50 micrometer in diameter and focused it 100-150 micrometer from the apex and found that 15 minutes later it promoted growth of the sporangiophore after a 1-2 minutes lag period and then the growth rate declined significantly. The team then took a 10 micrometer light beam and focused it continuously along one side of the sporangiophore and they found that sporangiophore bends towards the shaded side. The implication of this result is that the side with the light shining on it grew faster that the shaded side and that the lens has an effect on phototropism in the sporangiophore. This implication was tested using a mutant without carotenoid. The results confirmed that the lens did play a role in phototropism because these mutants were 1.5-2.0 times more sensitive with unilateral bending towards the blue light than the wild type.

Photo-induction of sporangia in Pilobolus Crystallinus

Richard Ellis studied how light affect sporangia production by using three different wavelength of monochromatic light with different photon fluence and varying the length of exposure. He found that cultured plate that were grown for 9 days in the dark resulted in growth of 1200 tropocyst and40-90 long sporangiophores but no sporangia were seen. Cultured plated that were grown exposed to continuous blue light yielded 50-90 short sporangiophores and sporangia production on sporangiophore. Cultures grown at the fluence rates of 1.7 and 0.17 micromoles/m2 sec and wavelength of 420 nm and 450 nm both stimulated sporangiophore and sporangia growth but wavelength at 450 nm at the lowest fluence rate of 0.017 and 0.001 micromoles/m2 sec was the most effective at stimulating growth. Cultures grown in light wavelength of 480 nm was the least effective at promoting growth. These wavelengths were chosen for this study because they are the points in the blue light spectra where the largest variations can be seen thus far from publishes data for plants and fungi.