Aspergillus niger- An Overview

Aspergillus niger is a common fungus that belongs to the genus Aspergillus, which consists of hundreds of species of molds that are widely distributed in nature. Aspergillus niger is characterized by its black spores that form on decaying plant matter, soil, food, and indoor environments. It is a filamentous, aerobic, and saprobic fungus that can grow on a variety of substrates and under different environmental conditions.

Aspergillus niger is an important microorganism for biotechnology and industrial applications. It is known for its ability to produce citric acid, which is used as a preservative and flavoring agent in soft drinks and other products. It can also produce other organic acids, enzymes, bioactive compounds, and biofuels from various biomass sources. Aspergillus niger is generally regarded as safe by the US Food and Drug Administration for use in food production.

However, Aspergillus niger can also cause some diseases in plants and animals. It can cause black mold in fruits and vegetables like grapes, apricots, onions, and peanuts, which can reduce their quality and shelf life. It can also cause opportunistic infections in humans and animals with compromised immune systems or damaged respiratory tracts. The most common infection caused by Aspergillus niger is otomycosis, which is a fungal ear infection that can cause pain, itching, and hearing loss.

In this article, we will provide an overview of Aspergillus niger, covering its history, habitat, morphology, cultural characteristics, life cycle, pathogenesis, laboratory diagnosis, treatment, prevention and control, and industrial uses. We hope that this article will help you to learn more about this fascinating fungus and its role in nature and society.

Aspergillus niger is the most common and the most studied species in the Aspergillus genus, which consists of common molds found throughout the environment. It is known for its extensive metabolic diversity, high production yield, secretion capability, and ability to conduct post-translational modifications.

The industrial use of A. niger began in 1917, when a food chemist named James Currie discovered that any strain of this mold would produce high concentrations of citric acid when grown in sugar medium. Citric acid is a tricarboxylic acid that is an intermediate of the Krebs cycle and has applications in food and beverage production as a preservative, flavor enhancer, and acidulant. Before Currie`s discovery, citric acid was extracted from citrus fruits, which was expensive and unreliable. Two years after Currie`s discovery, industrial-level production using A. niger began, and the biochemical fermentation industry started to flourish.

Other studies revealed that A. niger can also produce other useful acids, such as gluconic acid, oxalic acid, and malic acid, as well as various enzymes, such as glucoamylase, alpha-galactosidase, cellulases, lactase, invertase, pectinases, and proteases. These enzymes have applications in food processing, textile industry, paper industry, biofuel production, and pharmaceutical industry.

Based on morphological and biochemical research, scientists concluded that A. niger has different strains with different properties. In 2004, several species similar to A. niger were discovered by a group of researchers studying the characteristic production of ochratoxins by A. niger. Ochratoxins are mycotoxins that can cause kidney damage and cancer in humans and animals. These species include subgenus Circumdati, section Nigri which has 15-related black-spored species that are very similar to those of A. niger. They are A. tubingensis, A. foetidus, A. carbonarius, and A. awamori. In 2007, the strain of ATCC 16404 Aspergillus niger was reclassified as Aspergillus brasiliensis based on genomic sequencing.

A century later, citric acid production using A. niger is a multi-billion dollar industry, with A. niger additionally producing a diverse range of proteins, enzymes and secondary metabolites. A. niger is classified as Generally Recognized as Safe (GRAS) by the US Food and Drug Administration for use in food production, although the microbe is capable of producing toxins that affect human health.

Aspergillus niger is a widespread and adaptable fungus that can be found in various habitats around the world. Some of the common places where A. niger can grow are:

• Soil: A. niger is a soil inhabitant that can decompose organic matter and release nutrients into the environment. It can also colonize plant roots and form symbiotic or pathogenic relationships with them.
• Decaying vegetation: A. niger can grow on dead leaves, compost piles, and other plant debris, where it can produce enzymes and acids that break down cellulose and lignin. It can also cause black mold on fruits and vegetables, such as onions, grapes, apricots, and peanuts.
• Indoor environments: A. niger can be found in damp and dark places, such as basements, bathrooms, kitchens, and air ducts. It can also contaminate food products, such as cereals, nuts, dried fruits, and spices.
• Industrial settings: A. niger is widely used for producing enzymes, acids, and bioactive compounds that have applications in food, pharmaceutical, biotechnology, and biofuel industries. It can also be involved in bioremediation of pollutants and waste materials.

A. niger is a thermotolerant and acidophilic fungus that can grow in a wide range of temperatures (6-47°C) and pH levels (1.5-9.8). It can also survive in extreme conditions, such as high radiation and low oxygen levels. A recent study showed that A. niger can adapt to space radiation and grow on the International Space Station.

Aspergillus niger is a filamentous fungus that forms multicellular structures called hyphae. Hyphae are long, thin, and branching tubes that grow and extend through the substrate. Hyphae can be divided into two types: vegetative hyphae and aerial hyphae. Vegetative hyphae are responsible for absorbing nutrients and water from the environment, while aerial hyphae are involved in reproduction and spore formation.

Some of the aerial hyphae develop into specialized structures called conidiophores, which are the main reproductive organs of A. niger. Conidiophores are upright stalks that bear spore-producing cells called conidia at their tips. Conidia are small, spherical, and dark brown or black in color. They are arranged in chains or columns on top of the conidiophores, forming a radiating structure called a conidial head.

The conidiophore consists of several parts: a basal foot cell, a smooth or slightly granular stalk, and a swollen vesicle at the apex. The vesicle is covered with short branches called metulae, which in turn bear smaller branches called phialides. The phialides are the actual spore-producing cells that extrude conidia from their tips. The conidiophore can reach up to 3 mm in length and has a thick wall that can be hyaline or brown.

The conidia of A. niger have a rough or warty surface and a diameter of 4-5 μm. They can have different shapes, such as globose, subglobose, elliptical, or with longitudinal striae. They are resistant to desiccation and can survive for long periods of time in the environment. They can also germinate and form new hyphae when they encounter suitable conditions for growth.

The morphology of A. niger can vary depending on the strain, the culture medium, and the environmental factors. Different strains can have different sizes, shapes, and colors of conidial heads and conidia. Different media can affect the growth rate, branching pattern, and pigmentation of hyphae and conidiophores. Different environmental factors such as temperature, pH, oxygen, light, and nutrients can also influence the morphology of A. niger by regulating its gene expression and metabolism.

The morphology of A. niger is important for its industrial applications, as it affects its productivity, quality, and stability of the products it produces. For example, the size and shape of the pellets formed by A. niger in submerged fermentation can affect the mass transfer, oxygen uptake, rheology, and citric acid yield of the process. Therefore, understanding and manipulating the morphology of A. niger is a key aspect of optimizing its biotechnological potential.

Aspergillus niger is a filamentous fungus that exhibits specific cultural characteristics when grown on various media. Here are some of the cultural characteristics of Aspergillus niger:

• Colony morphology: Aspergillus niger colonies appear as flat, velvety, and olive-black in color. The colony can grow up to 7 cm in diameter. The colony surface is covered with conidiophores that produce black conidia in radiate heads. The reverse of the colony is white to yellow. The colony may produce radial fissures in the agar due to its rapid growth.
• Microscopic morphology: Aspergillus niger produces smooth, hyaline, and septate hyphae that branch dichotomously. The aerial hyphae form conidiophores that are 400-3000 um long and swell at the apex to form a globose vesicle that is 30-75 um in diameter. The vesicle is covered with metulae and phialides that produce conidia. The conidia are dark brown, rough-walled, and spherical with a diameter of 4-5 um. The conidia are arranged in columns on top of the phialides.
• Growth conditions: Aspergillus niger is a strict aerobe that requires oxygen for growth. It can grow in a wide range of temperatures from 6 to 47°C, but its optimal temperature range is 35-37°C. It can also tolerate a wide range of pH from 1.5 to 9.8, but it prefers acidic conditions. It can grow on various substrates such as carbohydrates, proteins, fats, and organic acids.
• Media and tests: Aspergillus niger can be cultured on different media such as potato dextrose agar (PDA), malt extract agar (MEA), Czapek yeast agar (CYA), and Sabouraud dextrose agar (SDA). On PDA, it produces black colonies with yellow reverse after 7 days of incubation at 25°C or 37°C. On MEA, it produces slightly brown colonies with smooth-walled conidia after 7 days of incubation at 25°C or 37°C. On CYA, it produces black colonies with wooly smooth-walled conidia after 5 days of incubation at 25°C or 37°C. On SDA, it produces black colonies with rough-walled conidia after 5 days of incubation at 25°C or 37°C.

Aspergillus niger reproduces asexually by forming conidial spores. The life cycle starts with the dispersion of the conidia onto a platform with favorable conditions of at least 25-40°C. The conidia then germinate forming a vegetative cell.

The cells develop into hyphal mycelium which branches dichotomously forming aerial hyphae. The aerial hyphae then grow to form conidiophores which swell at the apex forming the vesicle part of the conidiophore.

From the vesicles, form the primary sterigmata known as the phialides. The sterigmata form the secondary sterigmata that start to produce the conidial spores. The spores are arranged in columns (several rows) on top of the phialides.

The conidia are dark brown, rough and have a diameter of 4-5um. They can be easily dispersed by air currents or water splashes. The conidia can survive for long periods in dry or moist conditions and can germinate when they encounter a suitable substrate.

The life cycle of Aspergillus niger is schematically shown below:

Aspergillus niger is commonly regarded as a pathogenic allergen that can cause lung infections in individuals with weak immune systems. It can also cause otomycosis (fungal ear infection), keratitis (corneal infection), and cutaneous infections. However, compared with other species of Aspergillus, such as A. fumigatus and A. flavus, A. niger is less virulent and less frequently isolated from clinical specimens.

The pathogenesis of A. niger involves the following steps:

• Inhalation or inoculation of conidia (spores) into the host tissues.
• Germination of conidia into hyphae (filaments) that penetrate the epithelial barriers and invade the blood vessels.
• Production of secondary metabolites, such as ochratoxin A and fumonisin B2, that have toxic effects on the host cells and organs.
• Evasion of host immune responses by modifying the cell wall components, such as melanin and galactosaminogalactan, that interfere with phagocytosis and complement activation.
• Formation of fungal balls (aspergillomas) or granulomas in the lungs or sinuses that cause tissue damage and bleeding.

The severity and outcome of A. niger infection depend on several factors, such as the dose and route of exposure, the underlying health status and immune function of the host, and the availability and efficacy of antifungal treatment.

The diagnosis of Aspergillus niger infection depends on the site and type of infection. The following methods can be used to detect and identify Aspergillus niger from clinical specimens:

• Microscopic examination: A direct smear of the specimen (such as sputum, sinus drainage, bronchial washing, or ear discharge) can be stained with potassium hydroxide (KOH), lactophenol cotton blue (LPCB), or calcofluor white and examined under a microscope for the presence of dark brown, rough-walled conidia and hyaline conidiophores . The conidia are typically arranged in radial columns on top of the phialides.
• Cultural examination: The specimen can be inoculated on various media, such as potato dextrose agar (PDA), malt extract agar (MEA), or Czapek yeast agar (CYA), and incubated at 25°C and 37°C for 5 to 7 days. The colonies of Aspergillus niger are initially white to yellow, but soon turn black due to conidial production. The reverse is pale yellow and the growth may produce radial fissures in the agar .
• Serological tests: The detection of Aspergillus-specific IgE antibodies in serum can be useful for establishing a diagnosis of allergy to Aspergillus niger. A fluorescence enzyme immunoassay (FEIA) can measure the level of IgE antibodies and provide a class interpretation from 0 (negative) to 6 (strongly positive). The detection of galactomannan antigen in serum or bronchoalveolar lavage (BAL) fluid can also provide a presumptive diagnosis of invasive aspergillosis caused by Aspergillus niger or other Aspergillus species. Galactomannan is a polysaccharide component of the fungal cell wall that is released during active growth. A positive result indicates a high likelihood of invasive aspergillosis, while a negative result does not exclude the diagnosis .
• Molecular tests: Polymerase chain reaction (PCR) assays can amplify and detect Aspergillus-specific DNA sequences from various specimens, such as blood, BAL fluid, or tissue biopsies. PCR can provide a rapid and sensitive diagnosis of invasive aspergillosis, as well as differentiate between different Aspergillus species. However, PCR assays are not standardized or widely available, and their interpretation may be affected by contamination or false-negative results .

Aspergillus niger is generally considered as a non-pathogenic aspergillus, infecting only immunocompromised individuals and occasionally acting as a wound pathogen. However, in rare cases, it can cause opportunistic invasive aspergillosis, otomycosis (fungal ear infection), and other localized infections. The treatment of these infections depends on the type, severity, and location of the infection, as well as the patient`s overall health and immune status.

Invasive aspergillosis

Invasive aspergillosis is a serious and potentially life-threatening infection that occurs when Aspergillus spores invade the lungs, sinuses, brain, or other organs. It is most common in patients with hematologic malignancies, hematopoietic stem cell transplant recipients, solid organ transplant recipients, and those receiving high-dose corticosteroids or other immunosuppressive agents.

The treatment of invasive aspergillosis requires prompt initiation of antifungal therapy, preferably guided by culture and susceptibility testing. The first-line treatment for invasive aspergillosis is voriconazole, a broad-spectrum triazole antifungal agent that inhibits the synthesis of ergosterol, a component of the fungal cell membrane. Voriconazole has good penetration into various tissues and fluids, including cerebrospinal fluid, and has shown superior efficacy and survival rates compared with amphotericin B.

Other drugs that can be used to treat invasive aspergillosis include itraconazole , another triazole antifungal agent; amphotericin B , a polyene antifungal agent that binds to ergosterol and disrupts the fungal cell membrane; caspofungin, a echinocandin antifungal agent that inhibits the synthesis of beta-glucan, a component of the fungal cell wall; micafungin, another echinocandin antifungal agent; and posaconazole, a newer triazole antifungal agent with activity against some Aspergillus species that are resistant to other azoles.

Prolonged treatment is usually required for invasive aspergillosis, ranging from 6 to 12 weeks or longer depending on the response and resolution of the infection. When possible, immunosuppressive therapy, such as systemic corticosteroids, should be discontinued or reduced to minimize the risk of relapse or progression of the infection .

Otomycosis

Otomycosis is a fungal infection of the external ear canal that can be caused by Aspergillus niger or other fungi. It is characterized by ear pain, itching, discharge, temporary hearing loss, and inflammation of the ear canal and tympanic membrane. It is more common in tropical and subtropical regions, and in people who frequently expose their ears to water or trauma.

The treatment of otomycosis involves cleaning and debridement of the ear canal to remove fungal debris and secretions, followed by topical antifungal therapy. The most commonly used topical antifungal agents are clotrimazole, a imidazole antifungal agent that inhibits ergosterol synthesis; miconazole, another imidazole antifungal agent; and acetic acid, a weak acid that lowers the pH of the ear canal and inhibits fungal growth. These agents can be applied as drops or creams several times a day for 7 to 14 days or until symptoms resolve.

Oral antifungal therapy is usually not required for otomycosis unless there is evidence of invasive or systemic infection. However, some cases may benefit from oral itraconazole , especially if there is resistance or intolerance to topical agents. Oral itraconazole can be given at a dose of 200 mg once or twice daily for 7 to 14 days.

In addition to antifungal therapy, anti-inflammatory agents such as acetaminophen can be used to relieve pain and inflammation associated with otomycosis. Patients should also avoid getting water in their ears while swimming or showering, dry their ears after exposure to moisture, avoid putting cotton swabs or other objects inside their ears, avoid scratching or injuring their ears, and use acetic acid ear drops after getting water in their ears to prevent recurrence of the infection .

Other localized infections

Aspergillus niger can also cause other localized infections, such as skin, nail, eye, or sinus infections. These infections are usually mild and self-limiting, and can be treated with topical or oral antifungal agents depending on the site and extent of the infection. The choice of antifungal agent should be based on culture and susceptibility testing, if available, or on the empirical activity of the agent against Aspergillus species. The duration of treatment should be tailored to the response and resolution of the infection.

Some examples of antifungal agents that can be used for localized Aspergillus niger infections are:

• For skin infections: clotrimazole cream, miconazole cream, ketoconazole cream, or terbinafine cream applied once or twice daily for 2 to 4 weeks.
• For nail infections: itraconazole capsules taken at a dose of 200 mg once daily for 12 weeks, or terbinafine tablets taken at a dose of 250 mg once daily for 12 weeks.
• For eye infections: natamycin eye drops applied every hour for the first day, then every 2 hours for the next 2 days, then four times daily for up to 21 days; or voriconazole eye drops applied every hour for the first day, then every 2 hours for the next 2 days, then four times daily for up to 21 days.
• For sinus infections: itraconazole nasal spray administered at a dose of 10 mg per nostril twice daily for 14 days; or voriconazole nasal spray administered at a dose of 40 mg per nostril twice daily for 14 days.

Aspergillus niger is a common and ubiquitous fungus that can cause various infections and diseases in plants, animals, and humans. To prevent and control the spread of this fungus, some measures can be taken depending on the situation and the risk factors involved.

Removal of Aspergillus niger spores and growth using chemical and antifungal treatment

One way to prevent and control Aspergillus niger is to remove its spores and growth from the environment using chemical and antifungal agents. Some examples of these agents are :

• 70% ethanol or isopropyl alcohol for about 10 minutes, which is effective in penetrating the spore’s cell wall and its hyphae, and killing them.
• Phenols which kill Aspergillus niger spores within 20 minutes. They can be added to scrub soaps, mouthwashes, and surface disinfectants.
• Bleach containing hypochlorite which inhibits the growth of spores.

These agents can be applied to surfaces, materials, or objects that are contaminated or suspected to be contaminated with Aspergillus niger spores or growth.

Protection from the environment

Another way to prevent and control Aspergillus niger is to protect oneself from exposure to the fungus in the environment. This is especially important for people who have weakened immune systems or lung diseases, who are at a higher risk of developing invasive aspergillosis. Some steps to protect oneself from the environment are :

• Avoiding areas with a lot of dust like construction or excavation sites. If this is not possible, wearing an N95 respirator (a type of face mask) while in these areas.
• Avoiding activities that involve close contact with soil or dust, such as yard work or gardening. If this is not possible, wearing shoes, long pants, and a long-sleeved shirt when doing these activities. Wearing gloves when handling materials such as soil, moss, or manure.
• Cleaning any cuts or skin abrasions well with soap and water, especially if they have been exposed to soil or dust. This can reduce the chances of developing a skin infection.
• Using acetic acid ear drops after getting water in the ears. This can prevent otomycosis, a type of fungal ear infection caused by Aspergillus niger.

Antifungal medication

A third way to prevent and control Aspergillus niger is to use antifungal medication. This is mainly for people who are at high risk for developing invasive aspergillosis, such as those who have had a stem cell transplant or an organ transplant, are getting chemotherapy for cancer, or are taking high doses of corticosteroids. Antifungal medication can be prescribed by a healthcare provider to prevent aspergillosis or to treat it if it occurs . Some examples of antifungal drugs that are used for aspergillosis are:

• Itraconazole, which can be taken orally or intravenously. It is effective against most species of Aspergillus, but may have side effects such as liver toxicity, nausea, headache, and rash.
• Amphotericin B, which can be given intravenously. It is effective against most species of Aspergillus, but may have side effects such as kidney damage, fever, chills, and low blood pressure.
• Voriconazole, which can be taken orally or intravenously. It is effective against most species of Aspergillus, but may have side effects such as liver toxicity, visual disturbances, rash, and hallucinations.

The choice of antifungal drug depends on several factors, such as the type and severity of infection, the species of Aspergillus involved, the patient`s medical history and condition, and the drug`s availability and cost.

Testing for early infection

A fourth way to prevent and control Aspergillus niger is to test for early infection. This is also for people who are at high risk for developing invasive aspergillosis. Testing for early infection can help detect Aspergillus in the blood or other body fluids before symptoms appear. This can allow for prompt diagnosis and treatment of aspergillosis. Some tests that can be used for early detection of Aspergillus are:

• Galactomannan assay, which measures a component of the Aspergillus cell wall in the blood or other body fluids. It is a sensitive and specific test, but may give false positive results in some cases, such as when the patient is taking certain antibiotics or has other fungal infections.
• Beta-D-glucan assay, which measures a component of the fungal cell wall in the blood or other body fluids. It is a sensitive test, but not very specific, as it can detect other fungi besides Aspergillus. It may also give false positive results in some cases, such as when the patient has bacterial infections or is receiving certain blood products.
• Polymerase chain reaction (PCR), which detects the DNA of Aspergillus in the blood or other body fluids. It is a sensitive and specific test, but may be affected by the quality and quantity of the sample and the presence of inhibitors.

The choice of test depends on several factors, such as the availability and cost of the test, the patient`s clinical condition, and the likelihood of infection. The tests may also be used in combination to increase their accuracy and reliability.

Aspergillus niger is a mold that has many applications in various industries, especially in the production of acids, enzymes and bioactive compounds. Some of the industrial uses of A. niger are:

• Citric acid production: A. niger is the main microorganism used for the industrial fermentation of citric acid, a compound that is widely used as a food preservative, flavor enhancer, and acidulant in beverages, confectionery, and pharmaceuticals. A. niger can produce high yields of citric acid from various substrates, such as molasses, starch, and sucrose, under acidic and aerobic conditions.
• Gluconic acid production: A. niger can also produce gluconic acid, another organic acid that has applications in food, pharmaceutical, textile, and metal industries. Gluconic acid can be used as a sequestrant, a chelating agent, a pH regulator, and a cleaning agent. A. niger can convert glucose to gluconic acid by oxidation using glucose oxidase enzyme.
• Enzyme production: A. niger is a prolific producer of various enzymes that have industrial and biotechnological significance. Some of the enzymes produced by A. niger are:
  • Glycoside hydrolases: These are enzymes that can break down complex carbohydrates into simple sugars. A. niger can produce glycoside hydrolases such as cellulases, hemicellulases, amylases, pectinases, and xylanases, which can be used for biofuel production, animal feed improvement, pulp and paper processing, and textile bleaching.
  • Proteases: These are enzymes that can hydrolyze proteins into peptides and amino acids. A. niger can produce proteases such as aspergillopepsin, which can be used for cheese making, meat tenderization, leather processing, and detergent formulation.
  • Lipases: These are enzymes that can catalyze the hydrolysis of fats and oils into glycerol and fatty acids. A. niger can produce lipases that can be used for biodiesel production, oil extraction, flavor enhancement, and detergent formulation.
• Bioactive compound production: A. niger can also produce various bioactive compounds that have pharmacological and nutraceutical properties. Some of the bioactive compounds produced by A. niger are:
  • Ochratoxin A: This is a mycotoxin that has nephrotoxic, hepatotoxic, immunosuppressive, and carcinogenic effects in animals and humans. A. niger can produce ochratoxin A in contaminated food products such as cereals, coffee beans, dried fruits, and nuts. Ochratoxin A can be detected and quantified by thin-layer chromatography or other analytical methods.
  • Fructooligosaccharides: These are prebiotic compounds that can stimulate the growth of beneficial bacteria in the gut and improve the health of the host. A. niger can produce fructooligosaccharides from sucrose by transfructosylation using fructosyltransferase enzyme. Fructooligosaccharides can be used as dietary supplements or functional food ingredients to enhance the immune system, lower cholesterol levels, and prevent intestinal infections.
  • Naphthoquinones: These are aromatic compounds that have antimicrobial, antifungal, antiviral, anti-inflammatory, and anticancer activities. A. niger can produce naphthoquinones such as shikonin and alkannin from plant precursors such as shikonin lactone and alkannin lactone by biotransformation using cytochrome P450 enzymes. Naphthoquinones can be used as natural dyes or therapeutic agents for various diseases.

These are some of the industrial uses of Aspergillus niger that demonstrate its versatility and potential as a microbial cell factory for valuable products.

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