The Biology of Fungi Impacting Human Health: A Tropical Asia-Pacific Perspective

By WA Shipton

Fungi have become increasingly significant determinants of human health and may cause as heavy a burden to health as viruses, bacteria and parasites. This outcome has occurred on account of the rise in diseases affecting the immune system and in the risk factors associated with advances in technologies used to treat various diseases and human conditions. These trends are no more evident than in tropical locations. This text emphasizes the biology of fungi impacting human health, with an emphasis on the Asia-Pacific region. The author draws on his own experience working in tropical Australia, Papua New Guinea and Thailand.

A range of information is presented on the natural relationships of fungi, which helps the reader to understand the interactions these microbes engage in with other living organisms including plants and microfauna. Highlighted are the abilities of fungi to survive in soil, on plants and animals and their capacity to adapt to changing conditions and evade attempts to control them. The successes and problems encountered in controlling fungi biologically are outlined, including the development of vaccines. Practical methods to limit the impact of mycotoxins produced by fungi are suggested, including moderating plant growth conditions and being aware of human nutritional status.

• Language: English
• Publisher: Partridge Publishing Singapore
• Release date: Dec 10, 2012
• ISBN: 9781482894790

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Copyright © 2014 by WA Shipton.

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CONTENTS

Preface

Acknowledgments

Illustrations

Tables

1 Introduction

2 Old Treats

3 Fungal Toxins, Cell Components And Health

4 New Challenges

5 Patterns Of Reproduction

6 Mechanisms Available For Adaptation

7 Microbial Ecology

8 Habitat Relationships

9 Virulence And Related Factors

10 Mycotoxin Risk Minimization

11 Fungal Control Through Host Modification And The Use Of Microbes

References

Dedicated to the memory of the microbiologists of the Asia-Pacific region

who have advanced their science according to the best traditions.

PREFACE

On a world-wide basis, fungal diseases of humans are becoming more prominent. The new threats arising from these microbes primarily are a result of an increase in the immunocompromised population. This is a consequence of the AIDS epidemic, the rise of life-style diseases, the advances in transplant technologies and in the treatment of neoplastic diseases. Such developments, together with the increasing popularity of tourist medicine in tropical locations, place an increased focus on developments in countries in these regions.

The Asia-Pacific region has and is still experiencing a heavy impact from the AIDS epidemic, China and India are in the midst of a diabetes epidemic, and the region is recording a boom in travel medicine. Infections from both primary and opportunistic fungal pathogens are on the increase in the area. These are difficult to treat, for fungi are of the same basic cellular organisation as found in animals, and the number of antifungal agents available to treat the diseases is limited. Old treats coming from mycotoxins continue to plague agricultural commodities produced and traded in tropical regions. The widespread adoption of international trade in food commodities, with its strict guidelines, continues to place rural populations at risk. These developments are taking place amidst increasing population pressures in the region, which impact on the risk of infection. Such population increases have increased the financial pressures on a portion of the population and made them less able to discriminate in food choices. On the other hand the increases have exposed selected populations to greater contact with non-domestic animals with the increased risk of contracting infections that previously were unknown or rare.

These considerations indicate that a wider knowledge of the biology and ecology of fungi will be valuable for students of microbiology, particularly since many general texts give sparse attention to this group of microbes. Even when some mention is made, very little is said about diseases in the Asia-Pacific region.

This text is meant for students with a sound knowledge of the principles of microbiology. Fungal perspectives are introduced on a range of topics with particular focus being given to the organisms rather than their pathogenesis.

ACKNOWLEDGMENTS

It is a pleasure to acknowledge those who have helped in the preparation of this book. First of all one needs inspiration to launch on such a project and I am indebted to Drs Siriporn Tantipoonwini and Kamolnan Taweeyanyongkul for providing this. Unwittingly Drs Andrew Greenhill and Jeffrey Warner contributed to the inspiration through their enthusiasm for tropical diseases and for helping to provide insights into the factors contributing to sago hemolytic disease while working on our Australian Centre for International Agricultural Research project in Papua New Guinea.

Others have provided valuable technical support, access to information and facilities, without which the project would have been impossible. To all these I say a heartfelt thank you. Special thanks are due to Ash McVeigh for completing the illustrations and to Elizabeth Lough and Michael Bucknell for designing the front cover. The cover picture of Aspergillus nidulans was supplied courtesy of Dennis Kunkel Microscopy, Incorporated.

Of course, day to day support for such a project comes from those who love and care for you. Through the long and sometimes tedious work involved this has been provided by my wife Jan. For her indulgence, I am particularly grateful.

ILLUSTRATIONS

Figure 1.1.   General features displayed by Ascomycetes and Basidiomycetes.

Figure 1.2.   Fruiting body types typically recognized in the Coelomycetes.

Figure 1.3.   Representation of blastic and thallic conidiogenesis.

Figure 5.1.   Conidium formation and discharge in Basidiobolus and Conidiobolus.

Figure 5.2.   Sexual reproductive structures found in Conidiobolus and Basidiobolus.

Figure 5.3.   Typical asexual and sexual spore forms found in the Mucorales.

Figure 5.4.   A generalized life cycle found in the Ascomycetes (Gymnoascaceae).

Figure 5.5.   Typical asexual structures found in the genera Aspergillus and Penicillium and an example of sexual reproductive structures.

Figure 5.6.   Diagrammatic illustration of the sequence of events following fusion between compatible yeast cells of Saccharomyces cerevisiae and illustrations of a mature zygote together with mating in Candida albicans resulting in the formation of a daughter cell.

Figure 5.7.   Suggested life cycle of Pneumocystis.

Figure 5.8.   Diagrammatic representation of monokaryotic fruiting and mating found in Cryptococcus neoformans.

Figure 5.9.   Diagrammatic representation of the Schizophyllum commune life cycle.

Figure 5.10.   Asexual and sexual structures found in Pythium insidiosum.

Figure 6.1.   Diagrammatic representation of nuclear behaviour during clamp connection formation.

Figure 6.2.   Diagrammatic representation of the formation of dikaryons initiating the sexual reproduction process in the Ascomycetes.

Figure 6.3.   Diagrammatic representation of the parasexual cycle (heterokaryosis, karyogamy and haploidization during mitosis) and mitotic crossing-over.

Figure 6.4.   General structure of transposable elements classes I and II.

Figure 6.5.   Representation of opposite and same sex mating cycles in Candida albicans.

Figure 6.6.   Representation of opposite and same sex mating in C. neoformans.

Figure 8.1.   Representation of the life cycle of Coccidioides.

Figure 8.2.   Asexual spores encountered in the dermatophytes.

TABLES

Table 1.1.   Examples of pathogenic fungi and yeasts causing disease of humans in the Asia-Pacific tropical region.

Table 1.2.   Higher level phylogenetic classification scheme constructed for the kingdom Fungi or Mycota.

Table 1.3.   Taxonomy of fungi previously placed in the phylum Zygomycotina.

Table 2.1.   Range of regulatory limits specified for aflatoxins in selected Asian countries.

Table 2.2.   Toxicoses in the Asia-Pacific tropics or associated with the presence of toxin contaminated foods originating there.

Table 3.1.   Evaluation of the carcinogenicity of various mycotoxins.

Table 3.2.   Range of regulatory limits adopted for mycotoxins in various products.

Table 3.3.   Important agricultural mycotoxins that influence animal and human health.

Table 4.1.   Host defence defects that predispose individuals to specific fungal infection.

Table 4.2.   Some records of fungal species encountered in AIDS patients in the tropics and associated regions in the Asia-Pacific region.

Table 4.3.   Some fungal infections noted in transplant recipients in tropical and associated regions in the Asia-Pacific region.

Table 4.4.   Fungal enzymes implicated in sensitization and asthma in occupational situations.

Table 5.1.   Some Ascomycete pathogens encountered in the class Sordariomycetes. The species nominated often represent species complexes.

Table 6.1.   Representative examples of reproductive success (+) following matings involving bipolar and tetrapolar systems.

Table 7.1.   Interactions observed between microbial populations.

Table 7.2.   Cardinal temperatures for fungal growth shown by various categories of organisms.

Table 7.3.   Some factors influencing mycotoxins production by fungi.

Table 9.1.   Some suggested virulence and virulence moderating factors in primary pathogens, commensals and opportunists.

Table 10.1.   Dietary protein (casein) effects on foci number of pre-neoplastic hepatic lesions in rats.

Table 10.2.   The hazards, appropriate control measures and identification of practicable CCPs associated with food safety in traditional sago production.

Table 11.1.   Reduction in aflatoxin levels in peanuts in three drought seasons following soil inoculation with a non-aflatoxigenic strain of A. flavus.

1

Introduction

Microrganisms are capable of causing disease in all categories of living organisms. They include algae, bacteria, fungi, helminths, protozoa and viruses. Among plant populations, fungi and viruses are the most significant disease-causing agents. Among the animal population it is the bacteria and viruses. Fungi, together with algae, helminths and protozoa are eukaryotic in cell organisation, which is quite different to that found in the bacteria (prokaryotes). The viruses on the other hand are non-living agents with no energy generating apparatus.

The fungus kingdom contains an estimated 1.5 million members and may, in fact, be much larger (Hawksworth 2001). Only a small proportion of these are capable of causing disease in animals, but when they do these fungi are often quite difficult to control on account of the similarity in cellular organization and functioning to the host cell.

Fungi have been prominent in previous times in spectacular outbreaks of ergotism (Claviceps purpurea), untimely, mass deaths among turkey poults (Aspergillus flavus), deaths in the human population during times of privation and war through eating contaminated grain (Fusarium species) and during peace as a consequence of predisposing individuals to various forms of carcinoma (Aspergillus species). Then there are the allergic diseases connected with various occupations and the morbidity associated with sick buildings. The focus on fungi as agents of disease of domestic animals and humans is becoming more pronounced as we enter the age of multiple drug resistance, immunological deficits in individuals due to other diseases and immunological-dampening medications. Then the epidemic of diabetes and fascination with transplant technology has ensured that fungi will be of increasing significance and be challenging in their own right.

While most fungi encountered in the new environments we have mentioned often are present in soils, on plant surfaces and in airspora, they are opportunistic by nature. However, there are also species capable of launching pathogenic attacks in their own right. These continue their significance and in some instances are increasing among specialized groups of people such as diabetics and the immunocompromised. Then there are the zoonotic diseases. We will mention these groups in turn commencing with the primary fungal pathogens.

Primary Fungal Pathogens

Primary pathogens are able to cause disease in immunocompetent individuals as well as in the immunocompromised. The portal of entry is commonly via the lungs or it may be through subcutaneous injury. These organisms are encountered most commonly in their endemic areas when aerosolized spores are inhaled following some environmental disturbance (e.g., dust storm, soil cultivation). The organisms encountered here are the dimorphic fungi (possess yeast and hyphal forms) such as Blastomyces, Coccidioides, Histoplasma, and Paracoccidioides (Reiss et al. 2012). The dimorphic fungi are responsible for systemic or superficial infection, which means that spread occurs to organs other than the lungs. The dermatophytes also come into the category of primary pathogens. However, they typically are limited to colonization of keratinized tissues (Hardy 2002). Cryptococcus gattii (Filobasidiella bacillispora) can be considered to be a primary pathogen too. It is typically found among immunocompetent individuals, although immunocompromised individuals commonly are infected. The change in view regarding the status of C. gattii has come particularly following the North America outbreaks of the hypervirulent molecular type VGII. It is able to exist in a range of natural habitats (Bahn et al. 2005, Baddley & Dismukes 2011, Chowdhary et al. 2011).

Zoonoses

Diseases of animals and humans may be connected. Among the microbial groups other than fungi, there are well known human disease outbreaks that have commenced in the animal population. These include severe acute respiratory syndrome (SARS) and human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS), West Nile fever and avian influenza. Previous culprits include Mycobacterium tuberculosis/M. bovis (Baker 2007).

Fungi also are associated with animals and may cross the species barrier to infect human beings. The best known example involves the dermatophytes, those fungi which live on the dermal layers causing diseases of the skin, nails and hair. These are primary pathogens. Microsporum canis is a classic example of a skin disease passing from dogs and cats to human beings. It is capable of passing from stray dogs and causing micro-epidemics in the right circumstances—as has been recorded recently in a day care centre in Brazil (Gürtler et al. 2005). M. canis is common in Southeast Asia too (Lim et al. 1992). Other dermatophytes have the capacity to move across the species barrier (Microsporum and Trichophyton species—Dvorak et al. 2008), although it must not be imagined that all are capable of doing so. Rarely, some largely human-associated dermatophyte infections can move to animals (Dvorak et al. 2008—the so-called anthropophilic group).

The fungi Basidiobolus ranarum, Cyrptococcus neoformans and Penicillium marneffei are associated with animals, but do not fit into the category of zoonoses, except with the possible exception of the latter. For example, Basidiobolus ranarum (syn. B. haptosporus) is carried by kangaroos and wallabies (Speare & Thomas, 1985) besides amphibians and reptiles (Zahari & Shipton 1988, Nelson et al. 2002), but is not associated with disease states in the animals. Specialized geographically defined subgroups of C. neoformans appear to be associated with disease of animals. However, it is not at all certain that cross species movement occurs involving these specialized subgroups (Bui et al. 2008). P. marneffei similarly is problematic. It is capable of causing infection/disease in both humans and selected animal species (Chariyalertsak et al. 1996, Gugnani et al. 2004). The later researchers, working in India, found some evidence, through microsatellite typing, that infection of humans with the animal type was a possibility. However, no genotype was shared by all bamboo rats (Cannomys badius) from all sites.

Opportunistic Fungi

Opportunistic pathogens are organisms that are able to infect individuals who are compromised in some fashion. They normally do not invade healthy individuals (Reiss et al. 2012). Fungi are found in most habitats including bodies of water. The majority of fungi encountered are able to grow on dead organic matter where they complete their life cycle. In order to survive, the majority of these organisms produce spores and resting bodies that are able to tide them over until a fresh substrate arrives. They commonly are dispersed by rain splash and wind, with assistance from animals, birds, invertebrates and other agencies including humans. Spores are produced often in large numbers and when these are wind dispersed they are widely available. The airspora encountered in the outside environment differs from that indoors, but common elements exist and large numbers of fungal propagules may be available. Some propagules originate from growth on substrates in the building itself (Gregory 1973, Ingold & Hudson 1993, Andersen et al. 2011).

The spores commonly are carried by air currents and lodge on substrates using a number of strategies or are inhaled. If the exposed surfaces contain nutrients and the abiotic conditions are suitable, particularly if moisture and temperature regimes are favourable, then spores germinate and colonization may occur. If the spores are deposited on wound tissues for example, then these unprotected surfaces may be colonized, as in burn victims. Similarly, if the surface of the skin is punctured, the wound may be contaminated by soil and debris and carry with it active hyphae or dormant spores or other survival structures. Fungal spores or vegetative cells may arrive at the suitable site via human activity, as on the hands of surgeons and nurses, surgical equipment, and such like or may originate from the patient’s own body. Inhaled spores may give rise to infection under suitable predisposing conditions, such as metabolic acidosis, diabetes, leukopenia (low white blood cell count) and hyperglycemia (Romani 2002, Reiss et al. 2012).

A number of fungi are carried on or in the human body as harmless organisms, such as Candida albicans. If the biological balance existing between these organisms and the multitude of other microbes is upset, such as by antibiotic treatments, illness, pregnancy and other events, the organism may be able to increase its population levels dramatically and cause disease (Timoney et al. 1988, Reiss et al. 2012).

The number of fungi associated with diseases in humans is increasing. There are some limits to the numbers that might be involved as systemic pathogens for they must be able to grow at 37 °C. However, the involvement of opportunistic fungi associated with superficial infections are not so limited as surface temperatures of the human body are lower than the core body temperature.

Geographic Region Highlighted

In selecting the tropical Asian and Oceania region as the focus of this study, I am concentrating on the general region of the world where information gathering has been less intense and the details are often hidden in more general texts. It also is representative of the region where most of my scientific career has been spent—tropical Australia, Papua New Guinea and Thailand.

The tropics comprise the zone bordering the equator stretching from the Tropic of Capricorn (southern hemisphere or latitude approximately 23.3 degrees south) to the Tropic of Cancer (northern hemisphere or latitude approximately 23.3 north). The Tropic of Capricorn passes through Australia (northern third commencing at Rockhampton in Queensland). In the northern hemisphere the Tropic of Cancer passes through India (southern half—line passes close to Ahmadabad—and West Bengal), Bangladesh (coastal delta with the line passing south of Dhaka), Myanmar (southern two thirds), the three southern provinces of China (Yunnan, Guangxi and Guangdong), and the southern half of Taiwan (commences at Chianyi city). It has not always be possible to determine whether a report from a country relates precisely to the tropical or subtropical region due to the lack of detail provided; this means that some reports from India and Taiwan may be grouped somewhat incorrectly. In reality, then, the study area embraced is the tropics and associated areas. Of course all readers will recognize that microbes do not recognize latitudinal lines, although some do occupy geographic zones of preference (cf. Walker 1996).

Tropical Asia consists of segments of South and East Asia and Southeast Asia. South Asia has at its core India, Bangladesh, Myanmar and Sri Lanka in the tropical zone. The People’s Republic of China (three southern mainland provinces and Hong Kong) and Taiwan are part of East Asia. They fit into the tropics as do the Southeast Asian countries—Laos, Cambodia, Peninsular Malaysia, Singapore, Thailand and Vietnam. The tropics include Brunei, Indonesia, Sabah, Sarawak, Philippines and the smaller Pacific Islands (those just south of Necker Island, Hawaii to the equator and south of it to and including New Caledonia, French Polynesia, and Tonga)—the oceanic region separated from continental Asia is sometimes referred to as Oceania. In this account the term tropical Oceania is taken to incorporate the tropical zone between Asia and the Americas.

The Villains: Some Significant Pathogens

At the outset, it must be stated that pathogenicity is rare among the fungi. Most live a secret life of usefulness. The fungi involved as human and animal pathogens may be classified as primary pathogens or as opportunistic. Among the latter is a group of rarely encountered organisms. The following Table highlights this information with the emphasis on human diseases, but it is not meant to be exhaustive for the countries in the zone under consideration. By convention the yeasts often are separated from the fungi. The latter are organisms with a predominant filamentous or hyphal habit, although readers will recognise that both fungi and yeasts are eukaryotic and are found in the kingdom Fungi. I have included one water mould in our discussions. This organism, a Pythium, previously enjoyed a place with the fungi, but now it is classified in a different kingdom (Stramenopila). It is included at the risk of losing information about this significant pathogen.

One of the confusing aspects of working with fungi is that some organisms carry two names. This has come about since many fungi reproduce by both asexual methods (anamorphic—no union of nuclei involved) and sexual means (teleomorphic—union of two nuclei involved). To illustrate, if the anamorphic form of a fungus is discovered first, it is named according to its perceived morphologically similar relatives and placed in a form-genus or artificial genus. Later if the teleomorphic form is discovered its relationships became clearer and it is named accordingly (some genera are known solely by their sexual structures). The International Code of Nomenclature dictates that the whole fungus should be named after that of the teleomorph where it is known (Kirk et al. 2008). However, all this will change in 2013 when a fungus will be known by one name. Generally the whole fungus (holomorph) will take the teleomorph name, but under some circumstances it can take the anamorph name (Hawksworth 2011). In this account, where both asexual and sexual reproduction is known and an organism has two names, we will give both anamorph and teleomorph names at the first mention of the fungus in a chapter and then will refer to it by its holomorph name when this is reasonably well established. The time schedule for name changes is in the immediate future, but it will take some time for names to become settled.

About one fifth of fungal species classified among the two great phyla Ascomycota and Basidiomycota (sub-kingdom Dikarya) are known by their asexual structures, but this is not to infer that they lack sexual reproduction even though it may not yet have been found (Shenoy et al. 2007, Hyde et al. 2011, Taylor 2011). With the advent of DNA technology, the relationships among fungi have been revolutionized and in the near future they will become clearer. As this happens, some species will be moved to other genera. These changes will make information gathering more complex at one level and easier at another.

Some organisms are known by their asexual fruiting bodies alone. These have been placed into an artificial group known as the Deuteromycetes or Anamorphic fungi. This is a classification of convenience. Within this grouping, various subgroupings are made to help scientists in their identification. These groupings do not suggest genetic relationships. Indeed, a morphological species may be separated into a number of phylogenetic species using DNA technology, which makes statements about the ecology and disease-eliciting potential of a number of fungi problematic (Shenoy et al. 2007).

Before we enter into a brief discussion of taxonomy, I have listed some fungi and fungal-like organisms that have been shown to be pathogenic somewhere in the geographical area of emphasis (Table 1.1). These have been organized mainly under descriptive headings and some under distinctive classification groupings. We will deal with classification issues more fully in the next section.

Table 1.1. Examples of pathogenic fungi and yeasts causing disease of humans in the Asia-Pacific tropical region.

The lists constructed under the various headings are not meant to be exhaustive.

The teleomorph name is given in brackets where it is known.

The taxonomic group given in this table is according to long held tradition, but this will be updated as we move through the chapter.

Many more fungi than listed under each grouping have been found to be pathogenic (Ellis et al. 2007, Ellis 2011). Other fungi are being added to the list continually; although it must be stated that Koch’s postulates have not always been satisfied for such candidates. In Table 1.1 the ascomycete now known as Pneumocystis jirevecii (previously P. carinii) is highly significant among the immunocompromised. Closely related species are found in many mammalian species (Cushion et al. 2004). Other fungi mentioned as human pathogens also are pathogenic to plants (e.g., Colletotrichum dematium, Lasiodiplodia theobromae, Nattrassia mangiferae) and still others infect both mankind and a variety of animals. The vast majority are saprophytic in nature and are not dependent on an animal or plant host for completion of their life cycle. Some have intimate relationships with plants, animals and specific organic substrates. We will mention some of these associations in later chapters.

Restricting the area of interest to the tropics reduces the consideration of some disease states as these have not been recorded there. For example, north India is an endemic region for paranasal sinus infections among young farmers caused by a wide variety of fungi (Chakrabarti & Sharma 2000); we will not consider these infections in any detail as they occur most intensely outside the tropics. A wide range of fungi may also be involved in corneal ulceration over the geographic region of interest—such occurrences have multiple causes (e.g., Srinivasan et al. 1997; Gopinathan et al. 2002). It is mostly beyond the scope of this study to give consideration of all the fungi thought to be involved in ulceration. One example is mentioned in the Table 1.1—corneal ulceration caused by Colletotrichum dematium—in order to give another example of a fungus from the form-group known as the Coelomycetes (Mendiratta et al. 2005).

Diseases of reasonably restricted geographical distribution generally will not be considered here where they occur outside the Asia-Pacific tropics. Examples in this category are coccidioidomycosis (Coccidioides immitis/posadasii complex—American southwest, Mexico, South America), paracoccidiodomycosis (Paracoccidioides brasiliensis—Brazil, Venezuela, Colombia) and lacaziosis (Lacazia loboi—Mexico to Argentina). By leaving these organisms out of our discussions, we are not inferring that they will never be encountered in the Asia-Pacific tropical region or become adapted to life there. For example, in Asia, paracoccidioidomycosis has been reported from Japan and several authentic cases of coccidioidomycosis have been reported from India and one from Australia. However, so far all have been imported cases that came from an endemic area (Ericsson et al. 2002, Verghese et al. 2002, Onda et al. 2011, Ong et al. 2012).

Following the recent determination that the agent of rhinosporidiosis (Rhinosporidium seeberi) is not a fungus but a mesomycetozoan, its common occurrence in Sri Lanka will not be commented upon further (Mendoza et al. 2001). We might mention that this organism can be confused with Coccidioides.

In other sections of the book it will be convenient to use a number of terms to refer to disease states caused by fungi. Some of these will be introduced here. The diseases associated with typically aseptate fungi (those without regular cross walls) reproducing by means of a variety of asexual spores (many are capable of zygospore formation under special conditions) are referred to as mucormycoses rather than zygomycoses. The group of fungi with aseptate or sepatate mycelium reproducing by means of forcibly discharged true conidia are responsible for entomophthoromycoses (they also are capable of forming zygospores). The term zygomycoses has been abandoned by fungal taxonomists (practitioners will take longer to adapt) based on molecular phylogenetic information that divides fungi previously located in the phylum Zygomycotina into various phyla (Ibrahim et al. 2011).

The superficial cutaneous mycoses are encountered commonly. The fungi associated with this group of mycotic diseases invade the skin (keratinized portion), nails and hair. In the literature many names are used to describe these diseases such as ringworm and athlete’s foot or they are described according to location on the body where the condition occurs—tinea capitis (scalp), tinea barbae (beard), tinea pedis (feet) or tinea corporis (body) are some examples. The fungi involved are found in three genera—Epidermophyton, Microsporum and Trichophyton. These are known as the dermatophytes. Moving on to the subcutaneous mycoses, these are infections that occur beneath the skin. Fungi gain access through puncture wounds in the skin that implant either spores or hyphal fragments. A number of species are involved but one well known fungus is Sporothrix schenckii. This organism occurs widely in vegetation and soil and enters through wounds in the skin caused by thorns or splinters. A small nodule develops at the entry site that develops into an ulcer and the number of lesions may increase along the route of the lymph vessels.

The fungus S. schenckii can also be referred to as an opportunistic organism. Such agents are usually harmless in the environment and enter the host when trauma occurs or debilitation is experienced. We will comment on the latter states in a future chapter, but at this point we mention that the use of broad spectrum antibiotics or corticosteroids and risk factors such as diabetes, chronic disease, immunosuppression and alcoholism may be sufficient to predispose an individual to serious fungal invasion. As an additional example, we might mention C. albicans. This yeast is a normal, harmless organism found on and in the human body. It lives in balance (homeostasis) with the other microbes. The use of broad spectrum antibiotics that destroy large numbers of bacteria allows it to colonize in an unusual manner; other factors may also predispose it to become pathogenic. It may cause inflammation of the mouth and throat for example. This type of candidiasis can be referred to as mucocutaneous mycosis. We must not imagine that the yeast will not cause infection at other body sites under conditions where the normal biota is disrupted—indeed vulvovaginal candidiasis is commonly encountered and involves infection of the vulva and vagina.

Systemic mycoses are the most worrying group encountered. Many of these fungi are capable of existing in two morphological forms—hyphal and yeast commonly. These latter pathogens are known as dimorphic fungi. They are capable of causing infections deep in the body with no region or organ being out of bounds. For example, our fungus S. schenckii can become systemic in the immunocompromised. Most of the fungi that are systemic enter the body through inhalation of spores. One organism encountered in the tropics that causes systemic disease is Histoplasma. It grows preferentially on bat and bird droppings and inhalation of infective conidia or spores can lead to disease.

Fungal Classification

The classification of fungi is based on perceived phylogenetic relationships emphasizing the essential characteristics of the organism. The data base may change as additional information is gained. There has been a revolution in taxonomic relationships among fungi brought about by the molecular analysis of fungal genomes, particularly rRNA genes. These insights are likely to influence the perception of relationships among the fungal groups for some time to come. Until recently fungi were classified by convention according to the characteristics of their sexual phase or, when this was lacking, features of asexual spore production were used. In practical everyday operations, this is still done to a considerable degree, although molecular methods show some promise of displacing this practice in some instances. The hypothesized phylogenetic relationships are used to construct a framework for the classification system and to establish a phylogeny (perceived evolutionary history). In establishing this framework, there is considerable variation adopted in the names applied by various authors. The scheme presented below (Table 1.2) was suggested recently by a large group of experts (Geiser et al. 2006, Hibbett et al. 2007, Oehl et al. 2011). However, we might expect the fine details to be questioned as more data are considered (e.g., Liu et al. 2009, Lee et al. 2010) and for change to continue to occur over time.

The majority of fungal species are dikaryotic, which means they possess a stage in the life cycle when paired nuclei are present in cells. These have been placed in the sub-kingdom Dikarya. The Ascomycota (popularly known as Ascomycetes) is the largest phylum in this sub-kingdom and is characterised by the production of ascospores held in specialized structures known as an ascus (pl. asci)—Figure 1.1. The asci may be held in specialized fruiting bodies with various construction characteristics—cleistothecia, perithecia, apothecia (James et al. 2006).

figure%201%2c1.jpg

Figure 1.1. General features displayed by Ascomycetes and Basidiomycetes.

(a) Basidia and basidiospores. (b) Asci and ascospores.

The phylum Basidiomycota (popularly known as the Basidiomycetes) is the second group of dikaryotic fungi (sub-kingdom Dikarya). They are characterised mostly by the production of basidiospores on the exterior of structures known as basidia (Fig. 1.1). The majority of phylum members possess typical mushroom-forming fruiting bodies (subphylum Agaricomycotina—James et al. 2006). Relatively few members of this phylum have been associated with disease in humans. However, one significant group of pathogenic yeasts is found in the class Tremellomycetes. This group has been placed in the genus Filobasidiella, which contains both opportunistic and primary pathogens. They are becoming increasingly significant among both the immunocompetent and immunocompromised population. They possess no fruiting body as such, but well-developed basidia form under ideal conditions. Another group of basidiomycetous yeasts (Malassezia), which are commonly encountered in the tropics, have been placed provisionally in the Class Exobasidiomycetes. This group consists primarily of plant pathogenic fungi. However, Malassezia represent a group of lipophilic yeasts that are commensals on the skin surface of many animals, including human beings. Under special circumstances they become pathogenic.

Based on molecular similarities among the fungi, the phyla Ascomycota, Basidiomycota and Glomeromycota are considered to be monophylic. The phylum Chyrtidiomycota consists of true fungi that possess flagellated spores (James et al. 2006). Most molecular phylogenies based on 18S DNA place Basidiobolus among the Chytridiomycota, but when additional information is used it can be placed close to the order Entomophthorales, where it has been traditionally positioned (James et al. 2006).

The group of organisms formerly considered to cause zygomycoses has been separated into various families. The two groups of diseases relevant to this new classification are the mucormycoses and entomophthoromycoses (Kwon-Chung 2012). The mucormycoses are the most frequently encountered group of diseases. The most commonly isolated species belong in the family Mucoraceae. The genera Lichtheimia (syn. Absidia), Apophysomyces, Mucor, Rhizomucor, Rhizopus are found in the family Mucoraceae. Other genera associated with infection are as follows: Cunninghamella (family Cunninghamellaceae), Saksenaea (family Saksenaceae), Cokeromyces (family Thamnidiaceae), Mortierella (family Mortierellaceae) and Syncephalastrum (family Syncephalastraceae)—refer to Table 1.3. Among the Entomopthoromycoses two fungal families are represented—Ancylistaceae (Conidiobolus) and Basidiobolaceae (Basidiobolus). The Mucoraceae group of fungi has been isolated to an increasing degree from transplant patients. The members often display characteristic morphological features when cultured, but clinically they are indistinguishable, which makes identification necessary. Their identity ideally must be pursued to the species level in order to aid in understanding infection and in designing satisfactory treatment protocols—Ibrahim et al. 2011.

Asexually reproducing members of the kingdom Fungi are placed in the artificial phylum Deuteromycota or Fungi Imperfecti. These anamorphic fungi are frequently encountered and must be identified. Various morphological groups have been recognised in order to facilitate identification. These groupings have been given names as though they represented classes, although we emphasize that they represent classes of convenience only.

Table 1.2. Higher level phylogenetic classification scheme constructed for the kingdom Fungi or Mycota

(Geiser et al. 2006, Hibbett et al. 2007, Oehl et al. 2011).

Table 1.3. Taxonomy of fungi previously placed in the phylum Zygomycotina

(Ribes et al. 2000, Schüβler et al. 2001, Hibbert et al. 2007, Ibrahim et al. 2011).

The accepted terminology used is as follows: Hyphomycetes are mycelial fungi that bear conidia on separate or aggregated hyphae (conidiogenous cells), but they do not form spores inside discrete fruiting structures. The Agonomycetes are mycelial forms that are sterile, but may produce chlamydospores, sclerotia or other vegetative structures. The Coelomycetes form conida in loosely formed to highly organized fruiting bodies, such as acervuli and pycnidia. In these structures the conidia arise from special cells termed the conidiogenous cells present in considerable numbers—Figure 1.2. The term hyaline refers to essentially colourless mycelia while the dematiaceous fungi are pigmented (Kirk et al. 2008).

figure%201%2c2.jpg

Figure 1.2. Fruiting body types typically recognized in the Coelomycetes.

(a) Pycnidium. (b) Acervulus. Redrawn from de Hoog et al. (2000)

and used with kind permission of the American Society for Microbiology-Books.

The emphasis adopted with the identification of anamorphic fungi is on the method of spore production and release from the spore bearing structure (conidiophores). Conidiophores are modified hyphae. They may be branched and can have inflated apices that support a number of conidiogenous cells. However, the apex of the conidiophore becomes the conidiogenous cell with some species. The conidia may be produced singly or in chains. When chains are produced, the youngest or most recently formed conidium may be either at the apex of the chain (acropetal) or the base (basipetal). The conidia may be single celled or possess additional cells and the outer surface may be smooth or ornamented. Conidial development occurs through blastic or thallic means. In blastic conidiogenesis, the developing conidium can be recognised as a discrete structure before a cross wall is formed. In thallic conidiogenesis, a cross wall is laid down before a conidium is recognisable (Fig. 1.3). More detailed terminology is used to describe other features of conidium formation (Kirk et al. 2008, Moore et al. 2011).

figure%201%2c3.jpg

Figure 1.3. Representation of blastic and thallic conidiogenesis.

In thallic conidiogenesis the conidium initial does not enlarge. The cell becomes a conidium

by the laying down of septa. In blastic conidiogenesis the conidium initial enlarges before a

septum is laid down. (a) Examples of thallic conidiogenesis (left to right) are Coccidioides immitis and Geotrichum candidum. (b) Example of holoblastic conidiogenesis where all the cell wall

layers are involved in its formation. (c) Enteroblastic conidiogenesis involving a phialide where

the conidium is generated from materials made at the apex of the phialide.

The conidium cell wall is independent of the conidiogenous cell wall.

We will consider one organism that is not found among the true fungi but rather is placed in the kingdom Stramenopila. The water mould Pythium insidiosum is a significant human pathogen in the area of emphasis. This fungal-like organism has not yet found its place in a comprehensive phylogenetic scheme. However currently it is placed in the family Pythiales in the kingdom Stramenipila because it shows some features found among the algae (Lévesque 2011). It possesses sexual spores termed oospores, which is a characteristic of the oomycete group (pylum Oomycota) of organisms (Schardl & Craven 2003). Identification of Pythium species commonly is based on micro-morphological and growth characteristics on various media, which predictably are unreliable. DNA macro-array analysis has been developed to overcome some of these difficulties (Tambong et al. 2006). There are geographic variants or genetically distinct populations of P. insidiosum—i.e., North, Central & South America, Asia and Australia, Thailand and the USA. However, at the present time, the different populations have not been subdivided into different species (Schurko et al. 2003).

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Old Treats

Microorganisms have helped shape the future of the human race. Any discussion of diseases of mankind cannot be separated from a consideration of domestic and wild animal diseases. Almost all categories of microbes have been