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Rhizopus oligosporus – Overview

  • Rhizopus oligosporus, a fungus belonging to the family Mucoraceae, is a popular starter culture for the domestic and industrial production of tempeh.
  • As the mould grows, it generates white, fluffy mycelia that bind the beans together to form a “cake” of partially catabolized soybeans. It is believed that the organism was domesticated in Indonesia several centuries ago.
  • Multiple factors make R. oligosporus the preferred initial culture for tempeh production. It thrives in the typical warm temperatures (30–40 °C or 85–105 °F) of the Indonesian islands; it possesses strong lipolytic and proteolytic activity, which confers desirable properties on tempeh; and it produces metabolites that allow it to inhibit and outcompete other moulds and gram-positive bacteria, including the potentially harmful Aspergillus flavus and Staphylococcus aureus.
  • Currently, R. oligosporus is believed to be a domesticated form of Rhizopus microsporus, resulting in the synonym Rhizopus microsporus var. oligosporus.
  • R. microsporus produces several potentially toxic metabolites, including rhizoxin and rhizonins A and B, but it appears that domestication and mutation of the R. oligosporus genome have resulted in the loss of genetic material involved in toxin production.
  • The synonym is presently unrecognised in fungal taxonomy, so its taxonomic status is best described as a member of the R. microsporus species group.
  • Rhizopus oligosporus is a fungus belonging to the Zygomycetes class, which is one of two classes in the Zygomycota phylum.
  • Rhizopus oligosporus is a member of the group Rhizopus microsporus. This group consists of taxa with similar morphology that are linked to the production of undesirable metabolites, pathogenesis, and dietary fermentation.
  • Although other species of Rhizopus microsporus may produce potentially hazardous metabolites, Rhizopus oligosporus is not associated with the production of such substances. It is not found in nature and is commonly employed by humans.
  • Strains of Rhizopus oligosporus have a large diameter (up to 43 μm) and irregular spores with a wide volume range (typically 96–223 mm3).
  • Large, subglobose to globose spores are produced by Rhizopus oligosporus, with a high proportion of irregular spores (>10%). Rhizopus oligosporus has spores with nonparallel valleys and ridges, as well as granular plateaus.

Rhizopus oligosporus classification

Rhizopus oligosporus is a filamentous fungus belonging to the kingdom Fungi and the phylum Zygomycota (also known as the phylum Glomeromycota). It is further classified into the following taxonomic ranks:

  • Class: Zygomycetes: This class includes a diverse group of fungi characterized by the formation of zygospores during sexual reproduction. Rhizopus oligosporus belongs to this class.
  • Order: Mucorales: This order consists of various genera of fungi that share similar characteristics, including the formation of sporangia and asexual spores. Rhizopus oligosporus is classified within the Mucorales order.
  • Family: Mucoraceae: This family includes several genera of fungi, including Rhizopus. Rhizopus oligosporus is a member of the Mucoraceae family.
  • Genus: Rhizopus: This genus comprises numerous species of filamentous fungi, including Rhizopus oligosporus. These fungi are commonly found in soil, decaying organic matter, and various food fermentation processes.
  • Species: Rhizopus oligosporus: This is the specific species name for the fungus. “Oligosporus” refers to the fungus’s characteristic production of few (oligo) spores during asexual reproduction.

So, the complete taxonomy classification of Rhizopus oligosporus is as follows:

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  • Kingdom: Fungi
  • Phylum: Zygomycota (or Glomeromycota)
  • Class: Zygomycetes
  • Order: Mucorales
  • Family: Mucoraceae
  • Genus: Rhizopus
  • Species: oligosporus

Morphology of Rhizopus oligosporus

The morphology of Rhizopus oligosporus can be described as follows:

  1. Hyphae: Rhizopus oligosporus consists of long, branching hyphae, which are the thread-like filaments that make up the body of the fungus. The hyphae are typically colorless or pale and have a cottony or woolly appearance.
  2. Sporangiophores: Sporangiophores are specialized hyphae that bear sporangia, which are structures that produce spores. In Rhizopus oligosporus, sporangiophores are erect and rise above the substrate. They are often unbranched or sparingly branched.
  3. Sporangia: The sporangia of Rhizopus oligosporus are spherical to oval-shaped structures that develop at the tips of sporangiophores. They are usually black or dark brown in color and have a rough texture. Each sporangium contains numerous spores.
  4. Spores: Rhizopus oligosporus produces asexual spores, which are known as sporangiospores. These spores are typically spherical or oval-shaped and are released from the sporangia. They are usually dark-colored and can be seen within the sporangia or dispersed in the environment.
  5. Rhizoids: Rhizopus oligosporus may produce root-like structures called rhizoids. Rhizoids anchor the fungus to the substrate and aid in nutrient absorption.
Morphology of Rhizopus oligosporus
Morphology of R. oligosporus (Photo: Inger Ohlsson).

Habitat of Rhizopus oligosporus

Rhizopus oligosporus is a filamentous fungus that belongs to the class Zygomycetes. It is commonly found in the natural environment and has a wide habitat range. Here are the typical habitats where Rhizopus oligosporus can be found:

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  1. Soil: Rhizopus oligosporus thrives in soil environments and is commonly found in agricultural fields, gardens, and compost piles. It plays a crucial role in the decomposition of organic matter in the soil.
  2. Fermented Foods: This fungus is a key component in the production of several fermented foods, including tempeh. Tempeh is a traditional Indonesian food made by fermenting soybeans using Rhizopus oligosporus. The fungus grows on the beans, producing a mycelial network that binds them together and imparts a characteristic flavor.
  3. Rotting Plant Material: Rhizopus oligosporus is known to colonize decaying plant material, such as fallen fruits, vegetables, and plant residues. It contributes to the natural process of decomposition, breaking down complex organic compounds into simpler forms.
  4. Animal Waste: The fungus can also be found in animal waste, including manure and compost. It thrives in nutrient-rich environments, utilizing the organic matter present in the waste as a food source.
  5. Indoor Environments: In addition to its natural habitats, Rhizopus oligosporus can also be found in indoor environments where food fermentation occurs. If conditions are suitable, such as in improperly stored food or in humid environments, the fungus can grow and contribute to food spoilage.

Life Cycle of Rhizopus oligosporus

The life cycle of Rhizopus oligosporus, a filamentous fungus belonging to the class Zygomycetes, involves both sexual and asexual reproduction. Let’s explore the different stages of its life cycle:

  1. Spore Formation: The life cycle of Rhizopus oligosporus begins with the production of asexual spores called sporangiospores. These spores are formed within specialized structures called sporangia, which are multicellular, sac-like structures that develop at the tips of hyphae (filamentous structures of the fungus). Each sporangium contains numerous spores.
  2. Spore Dispersal and Germination: The mature sporangia rupture, releasing the sporangiospores into the surrounding environment. These spores are dispersed by air currents, water, or other means. When favorable conditions are present, such as moisture and a suitable substrate, the sporangiospores germinate.
  3. Hyphal Growth and Mycelium Formation: Upon germination, each sporangiospore gives rise to a single hypha. Hyphae are elongated, tubular structures that form the main body of the fungus. As the hyphae grow and branch, they form a network called a mycelium. The mycelium consists of a mass of interconnected hyphae and is responsible for nutrient uptake and fungal growth.
  4. Asexual Reproduction – Conidiospore Formation: Under certain conditions, such as nutrient limitation or environmental stress, Rhizopus oligosporus can undergo asexual reproduction through the production of conidiospores. Conidiospores are formed directly from the hyphae without the involvement of specialized structures like sporangia. These spores are smaller and more resistant than sporangiospores and can be easily dispersed.
  5. Sexual Reproduction – Gametangia Formation: Rhizopus oligosporus can also reproduce sexually. Under specific conditions, two different mycelia of opposite mating types (designated “+” and “-“) come into contact and fuse. The fusion of these compatible mycelia leads to the formation of specialized structures called gametangia.
  6. Gametangial Fusion and Zygospore Formation: Within the gametangia, the cytoplasm of the “+” and “-” mating types fuse, resulting in the formation of a diploid zygote. This process is known as plasmogamy. The zygote undergoes a series of developmental changes and enlarges to form a thick-walled structure called a zygospore.
  7. Zygospore Maturation and Germination: The zygospore is a dormant and resistant structure capable of surviving unfavorable conditions. When conditions become favorable again, the zygospore undergoes meiosis, resulting in the formation of haploid spores called zygospores. These spores can germinate and give rise to new hyphae, completing the life cycle of Rhizopus oligosporus.

Cultural Characteristics of Rhizopus oligosporus

The cultural characteristics of Rhizopus oligosporus refer to the observable features and behaviors of the fungus when grown under laboratory conditions. These characteristics can help in identifying and distinguishing Rhizopus oligosporus from other fungi. Here are the cultural characteristics typically associated with Rhizopus oligosporus:

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  1. Colony Appearance: When grown on solid culture media, Rhizopus oligosporus colonies exhibit distinct characteristics. The colony appears fast-growing and initially appears as a white or cream-colored cottony mass. As the colony matures, it may develop a grayish or brownish color due to the accumulation of spores. The colony is generally fluffy and can cover a significant portion of the culture medium.
  2. Mycelium: The mycelium of Rhizopus oligosporus is composed of branched hyphae that form a dense network. The hyphae are usually coenocytic, meaning they lack cross-walls (septa) and contain multiple nuclei within a continuous cytoplasmic mass.
  3. Sporangia Formation: Rhizopus oligosporus is known for its ability to produce sporangia, which are specialized structures containing sporangiospores. Sporangia are typically spherical to oval in shape and are formed at the tips of hyphae. They are often black or dark brown in color due to the pigmentation of the spores.
  4. Spore Dispersal: As the sporangia mature, they undergo a characteristic swelling and eventually rupture, releasing the sporangiospores. The release of spores can create a powdery appearance on the surface of the colony or culture medium.
  5. Growth Conditions: Rhizopus oligosporus prefers warm and humid conditions for optimal growth. It thrives in temperatures ranging from 20 to 37 degrees Celsius (68 to 99 degrees Fahrenheit) and requires moisture to support its growth and sporulation.
The form of Rhizopus oligosporus is commonly found in tempeh starter and serves to help the process of fermentation of soybeans into tempeh. (Source : alchetron.com)
The form of Rhizopus oligosporus is commonly found in tempeh starter and serves to help the process of fermentation of soybeans into tempeh. (Source : alchetron.com)
Sporangium colour of Rhizopus oligosporus isolates FB-01 and FB-06. Sporangium of FB-01 is brownish, whereas sporangium of FB-06 is blackish.
Sporangium colour of Rhizopus oligosporus isolates FB-01 and FB-06. Sporangium of FB-01 is brownish, whereas sporangium of FB-06 is blackish.
Rhizopus microsporus var. oligosporus: a Sporangiophores, columellae, and rhizoids (Lactophenol cotton blue, 9200); b Columellae (Lactophenol cotton blue, 9400); c Sporangiospores (Lactophenol cotton blue, 91,000)
Rhizopus microsporus var. oligosporus: a Sporangiophores, columellae, and rhizoids (Lactophenol cotton blue, 9200); b Columellae (Lactophenol cotton blue, 9400); c Sporangiospores (Lactophenol cotton blue, 91,000)
Development of Rhizopus oligosporus in PDA medium with Rice (A) and in PDA medium only (B).
Development of Rhizopus oligosporus in PDA medium with Rice (A) and in PDA medium only (B).

Culture media used for the growth of Rhizopus oligosporus

To support the growth of Rhizopus oligosporus in the laboratory, various culture media can be used. The choice of culture media depends on the specific objectives of the study or experiment. Here are some commonly used culture media for cultivating Rhizopus oligosporus:

  1. Sabouraud Dextrose Agar (SDA): SDA is a general-purpose medium that supports the growth of a wide range of fungi, including Rhizopus oligosporus. It contains dextrose (glucose) as the carbon source and peptone as a nitrogen source. SDA has a low pH, which helps inhibit the growth of bacteria and other contaminants, allowing the fungus to thrive.
  2. Potato Dextrose Agar (PDA): PDA is another widely used medium for fungal cultivation. It consists of mashed potatoes and dextrose, providing a rich source of nutrients for the growth of Rhizopus oligosporus. PDA supports rapid mycelial growth and sporulation of the fungus.
  3. Malt Extract Agar (MEA): MEA is a nutrient-rich medium that contains malt extract and agar. It provides a favorable environment for the growth of Rhizopus oligosporus, promoting mycelial growth and sporulation. MEA is commonly used when a higher biomass of the fungus or abundant spore production is desired.
  4. Czapek-Dox Agar: Czapek-Dox agar is a selective medium often used for the cultivation of filamentous fungi. It contains various ingredients such as sodium nitrate, sucrose, potassium chloride, and magnesium sulfate. Czapek-Dox agar provides essential nutrients for Rhizopus oligosporus growth and allows for the observation of specific cultural characteristics.
  5. Synthetic Media: Synthetic media are chemically defined and can be customized to provide specific nutrient compositions. Synthetic media allow for precise control over the nutrient availability and can be tailored to study specific aspects of Rhizopus oligosporus growth and metabolism.

It’s important to sterilize the culture media before use to prevent contamination by other microorganisms. The appropriate medium selection should consider the specific research objectives and growth requirements of Rhizopus oligosporus, ensuring optimal growth and development of the fungus in the laboratory setting.

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Pathogenesis of Rhizopus oligosporus

Rhizopus oligosporus is a filamentous fungus belonging to the order Mucorales. It is commonly used in the fermentation process for producing traditional fermented foods such as tempeh and sufu. While Rhizopus oligosporus is generally considered non-pathogenic, it can cause infections, particularly in individuals with compromised immune systems.

The pathogenesis of Rhizopus oligosporus involves several key factors:

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  1. Opportunistic Pathogen: Rhizopus oligosporus is an opportunistic pathogen, meaning it primarily affects individuals with weakened immune systems. Conditions such as uncontrolled diabetes, immunosuppressive therapies, organ transplantation, or prolonged corticosteroid use can increase the risk of infection.
  2. Inhalation or Contamination: The primary route of infection is inhalation of spores or exposure to contaminated materials. Rhizopus oligosporus spores are ubiquitous in the environment and can be found in soil, decaying vegetation, and various food substrates.
  3. Angioinvasion: Once the spores enter the respiratory tract or come into contact with a compromised area of the body, they can germinate and invade surrounding tissues. Rhizopus oligosporus is capable of angioinvasion, meaning it can invade blood vessels, leading to thrombosis and tissue necrosis.
  4. Rapid Growth and Sporulation: Rhizopus oligosporus has a fast growth rate, aided by its ability to utilize a wide range of carbon sources. The fungus forms mycelia, which are branching filaments that penetrate and invade tissues. As the mycelia mature, they produce asexual spores called sporangiospores, which can be released into the surrounding environment.
  5. Immune Response: The immune response to Rhizopus oligosporus infection is complex. Neutrophils and macrophages are important in early defense against the fungus, but in immunocompromised individuals, the response may be inadequate. The fungus can also evade immune detection by producing factors that inhibit phagocytosis or by altering its cell surface composition.
  6. Clinical Manifestations: Infections caused by Rhizopus oligosporus are known as mucormycosis or zygomycosis. The clinical manifestations depend on the site of infection. Rhinocerebral mucormycosis, the most common form, involves the nasal sinuses and can spread to the brain. Other forms include pulmonary mucormycosis, cutaneous mucormycosis, gastrointestinal mucormycosis, and disseminated mucormycosis.

Treatment of Rhizopus oligosporus infections involves a multi-disciplinary approach, including antifungal therapy, surgical debridement, and management of the underlying immunocompromising condition. Amphotericin B is the primary antifungal agent used, but other drugs such as posaconazole and isavuconazole may be considered. Prompt diagnosis and aggressive treatment are crucial for improving outcomes in mucormycosis cases.

Infections caused by Rhizopus oligosporus

  • Mucormycosis: This severe infection can affect the epidermis, sinuses, lungs, and brain. People with compromised immune systems, such as those with diabetes, cancer, or organ transplants, are most susceptible.
  • Rhizopus dermatitis: Rhizopus dermatitis is a skin infection that can produce patches that are red, itchy, and scaly. It is most prevalent in individuals with a skin break, such as a laceration or scrape.
  • Rhizopus otomycosis: This ear infection can produce pain, inflammation, and discharge. It is most prevalent among those who use hearing aides or have had ear surgery.
  • Rhizopus keratitis: Rhizopus keratitis is an infection of the cornea that can result in pain, irritation, and vision impairment. It is most prevalent among contact lens wearers.

Diagnosis of Rhizopus oligosporus infection

  1. Clinical evaluation: A thorough medical history and physical examination are important initial steps in diagnosing any infection. The doctor will inquire about symptoms such as fever, cough, difficulty breathing, and any recent exposure to the fungus.
  2. Microscopic examination: A sample from the infected site, such as tissue, skin, or sputum, may be collected and examined under a microscope. The presence of characteristic fungal hyphae and sporangia (spore-containing structures) can suggest the presence of Rhizopus oligosporus. Special stains, such as lactophenol cotton blue or potassium hydroxide (KOH), can be used to enhance the visibility of the fungal structures.
  3. Fungal culture: The collected sample can be cultured in the laboratory on appropriate growth media to encourage the growth of Rhizopus oligosporus. Fungal cultures typically require several days to weeks for visible growth. Once the fungus has grown, its identification can be confirmed through morphological characteristics and specialized tests.
  4. Molecular techniques: Polymerase chain reaction (PCR) assays can be used to detect the presence of specific fungal DNA or genetic markers associated with Rhizopus oligosporus. These tests provide rapid and accurate identification of the fungus.
  5. Imaging studies: Depending on the site of infection, imaging techniques such as X-rays, computed tomography (CT) scans, or magnetic resonance imaging (MRI) may be performed to visualize the affected area. These studies can help assess the extent and severity of the infection and aid in treatment planning.
    • Chest X-ray: This test can be used to look for lung infection.
    • CT scan: This test can be used to look for infection in the sinuses, brain, and other organs.
    • MRI: This test can be used to look for infection in the brain and other organs.

Treatment of Rhizopus oligosporus

Antifungal medications that may be used to treat Rhizopus oligosporus infection include:

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  • Amphotericin B
  • Liposomal amphotericin B
  • Voriconazole
  • Posaconazole
  • Caspofungin
  • Isavuconazole

Prevention of Rhizopus oligosporus

  • Maintain clean and dry epidermis. This will prevent the growth of the fungus on your epidermis.
  • Avoid contact with ill individuals. This will prevent you from becoming unwell due to the fungus.
  • Frequent hand washing with detergent and water is recommended. This will assist in eliminating the fungus from your hands.
  • If the air in your residence is dry, use a humidifier. This will help to keep the air moist, which can inhibit the growth of fungi.
  • Get regular exams from your physician. This will aid in the early detection of infections, when they are simpler to treat.

Industrial Uses of Rhizopus oligosporus

Rhizopus oligosporus has several industrial uses due to its ability to produce various enzymes and metabolites. Here are some of the industrial applications of Rhizopus oligosporus:

  1. Tempeh production: Rhizopus oligosporus is primarily used in the production of tempeh, a traditional Indonesian food product made from fermented soybeans. The fungus helps break down complex carbohydrates and proteins in the soybeans, improving their nutritional value and flavor.
  2. Enzyme production: Rhizopus oligosporus is capable of producing a range of enzymes, including amylases, proteases, lipases, and cellulases. These enzymes have industrial applications in various sectors, such as food, textile, and detergent industries. They can be used for starch hydrolysis, protein degradation, lipid modification, and cellulose degradation.
  3. Production of organic acids: Rhizopus oligosporus can produce organic acids such as citric acid and fumaric acid. Citric acid is widely used in the food and beverage industry as an acidity regulator, flavor enhancer, and preservative. Fumaric acid finds applications in the food, pharmaceutical, and chemical industries.
  4. Bioremediation: Some strains of Rhizopus oligosporus have been investigated for their potential in bioremediation processes. They can degrade organic pollutants, such as petroleum hydrocarbons and agricultural wastes, aiding in the cleanup of contaminated environments.
  5. Production of bioactive compounds: Rhizopus oligosporus has been studied for its ability to produce bioactive compounds with potential pharmaceutical applications. These include antimicrobial peptides, immunosuppressive compounds, and antioxidants. Research is ongoing to explore their potential therapeutic uses.
  6. Biofuels: Rhizopus oligosporus can be used to produce bioethanol, a type of biofuel. Bioethanol is a renewable fuel that can be derived from plant matter. Rhizopus oligosporus is capable of fermenting carbohydrates and starches into bioethanol.

FAQ

What is Rhizopus oligosporus?

Rhizopus oligosporus is a filamentous fungus that belongs to the Zygomycota phylum. It is widely known for its role in the fermentation process of tempeh, a traditional Indonesian food product made from soybeans.

What are the main characteristics of Rhizopus oligosporus?

Rhizopus oligosporus is a fast-growing fungus with a cottony mycelium that forms sporangia. It produces few (oligo) spores during asexual reproduction, which are used to inoculate substrates for fermentation.

What is the role of Rhizopus oligosporus in tempeh production?

Rhizopus oligosporus plays a crucial role in tempeh production. It breaks down complex carbohydrates and proteins present in soybeans through enzymatic activity, resulting in the fermentation and transformation of the soybeans into tempeh.

How does Rhizopus oligosporus contribute to the fermentation process in tempeh production?

Rhizopus oligosporus produces enzymes, such as amylases and proteases, that degrade starches and proteins present in soybeans. This enzymatic activity leads to the breakdown of complex nutrients into more digestible forms and contributes to the unique texture, flavor, and nutritional profile of tempeh.

Can Rhizopus oligosporus be harmful to human health?

Rhizopus oligosporus is generally safe for human consumption when used in controlled fermentation processes, such as tempeh production. However, it is important to ensure proper hygiene and quality control measures during fermentation to avoid contamination and ensure the safety of the final product.

What are the optimal growth conditions for Rhizopus oligosporus?

Rhizopus oligosporus thrives in slightly acidic conditions with a pH range of 4.0 to 6.0. It grows best at temperatures between 30°C and 37°C (86-98.6°F) and requires a relative humidity of 80-90% for optimal growth and fermentation.

How is Rhizopus oligosporus cultivated in industrial settings?

Rhizopus oligosporus is typically cultivated by inoculating cooked and partially sterilized soybeans or other legumes with a starter culture containing the fungus. The inoculated substrate is then tightly packed and allowed to ferment under controlled conditions.

Can Rhizopus oligosporus be used in other food fermentation processes besides tempeh production?

While Rhizopus oligosporus is most commonly associated with tempeh production, it can potentially be used in other food fermentation processes. However, its usage may vary depending on the specific requirements and characteristics of the food product being fermented.

Does Rhizopus oligosporus have any industrial applications outside of the food industry?

Yes, Rhizopus oligosporus has some industrial applications beyond the food industry. It can be used for the production of enzymes, such as amylases, proteases, lipases, and cellulases, which find applications in various sectors, including the food, textile, and detergent industries.

Is Rhizopus oligosporus found naturally in the environment, or is it a cultivated species?

Rhizopus oligosporus is found naturally in the environment, particularly in soil and decaying organic matter. However, it is also cultivated and used in controlled fermentation processes, such as tempeh production, to ensure consistent and reliable results.

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