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Larone's Medically Important Fungi
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Table of Contents

List of Tables xv Preface to the Fifth Edition xvii Preface to the First Edition xix Acknowledgments xxi How to Use the Guide 1 Use of Reference Laboratories 3 Safety Precautions 7 Part I Direct Microscopic Examination of Clinical Specimens Introduction 11 Histological Terminology 13 Tissue Reactions to Fungal Infection 17 Stains 21 Table 1 Stains for direct microscopic observation of fungi and/or filamentous bacteria in tissue 22 Guide to Interpretation of Direct Microscopic Examination 23 Detailed Descriptions 31 Actinomycosis 33 Mycetoma, Actinomycotic or Eumycotic 34 Nocardiosis 36 Zygomycosis 37 Aspergillosis 38 Miscellaneous Hyalohyphomycoses 40 Dermatophytosis 42 Tinea versicolor 43 Tinea nigra 44 Phaeohyphomycosis 45 Chromoblastomycosis 46 Sporotrichosis 47 Histoplasmosis capsulati 48 Penicilliosis marneffei 50 Blastomycosis 52 Paracoccidioidomycosis 53 Candidiasis (Candidosis) 54 Trichosporonosis 56 Cryptococcosis 57 Pneumocystosis 59 Protothecosis 60 Coccidioidomycosis 61 Rhinosporidiosis 62 Adiaspiromycosis 64 Special References 65 Part II Identification of Fungi in Culture Guide to Identification of Fungi in Culture 69 Detailed Descriptions 101 Filamentous Bacteria 103 Introduction 105 Table 2 Differentiation of filamentous aerobic actinomycetes encountered in clinical specimens 107 Nocardia spp. 108 Table 3 Phenotypic characteristics of most common clinically encountered Nocardia spp. 110 Streptomyces spp. 111 Actinomadura spp. 112 Nocardiopsis dassonvillei 113 Yeasts and Yeastlike Organisms 115 Introduction 117 Candida albican 119 Table 4 Characteristics of the genera of clinically encountered yeasts and yeastlike organisms 120 Candida dubliniensis 121 Table 5 Characteristics of Candida spp. most commonly encountered in the clinical laboratory 122 Table 6 Characteristics that assist in differentiating Candida dubliniensis from Candida albicans 124 Candida tropicalis 125 Candida parapsilosis complex 126 Candida lusitaniae 127 Candida krusei 128 Table 7 Differentiating characteristics of Blastoschizomyces capitatus versus Candida krusei 129 Table 8 Differentiating characteristics of C. krusei, C. inconspicua, and C. norvegensis 129 Candida kefyr (formerly Candida pseudotropicalis) 130 Candida rugosa 131 Candida guilliermondii complex 132 Table 9 Differentiating characteristics of Candida guilliermondii versus Candida famata 133 Candida lipolytica 134 Candida zeylanoides 135 Candida glabrata 136 Cryptococcus neoformans 137 Cryptococcus gattii 138 Table 10 Characteristics of Cryptococcus spp. 139 Table 11 Characteristics of yeasts and yeastlike organisms other than Candida spp. and Cryptococcus spp. 140 Rhodotorula spp. 141 Sporobolomyces salmonicolor 142 Saccharomyces cerevisiae 143 Wickerhamomyces anomalus (formerly Pichia anomala and Hansenula anomala) (sexual state); Candida pelliculosa (asexual state) 145 Malassezia spp. 146 Malassezia pachydermatis 148 Ustilago sp. 149 Prototheca spp. 150 Trichosporon spp. 151 Table 12 Key characteristics of the most common clinically encountered Trichosporon spp. 152 Blastoschizomyces capitatus 153 Geotrichum candidum 154 Thermally Dimorphic Fungi 155 Introduction 157 Histoplasma capsulatum 158 Blastomyces dermatitidis 160 Paracoccidioides brasiliensis 162 Penicillium marneffei 164 Sporothrix schenckii complex 166 Table 13 Characteristics for differentiating species of the Sporothrix schenckii complex 168 Thermally Monomorphic Moulds 169 Zygomycetes 171 Introduction 173 Table 14 Differential characteristics of similar organisms in the class Zygomycetes 175 Table 15 Differential characteristics of the clinically encountered Rhizopus spp. 175 Rhizopus spp. 176 Mucor spp. 177 Rhizomucor spp. 178 Lichtheimia corymbifera complex (formerly Absidia corymbifera) 179 Apophysomyces elegans 181 Saksenaea vasiformis 183 Cokeromyces recurvatus 184 Cunninghamella bertholletiae 185 Syncephalastrum racemosum 186 Basidiobolus sp. 187 Conidiobolus coronatus 188 Dematiaceous Fungi 189 Introduction 191 Fonsecaea pedrosoi 193 Fonsecaea compacta 195 Rhinocladiella spp. 196 Phialophora verrucosa 197 Table 16 Characteristics of Phialophora, Pleurostomophora, Phaeoacremonium, Acremonium, Phialemonium, and Lecythophora 198 Pleurostomophora richardsiae (formerly Phialophora richardsiae) 199 Phaeoacremonium parasiticum (formerly Phialophora parasitica) 200 Phialemonium spp. 201 Cladosporium spp. 203 Table 17 Characteristics of Cladosporium and Cladophialophora spp. 204 Cladophialophora carrionii 205 Cladophialophora bantiana 206 Cladophialophora boppii (formerly Taeniolella boppii) 207 Pseudallescheria boydii (sexual state) / Scedosporium apiospermum (asexual state) complex 208 Table 18 Differentiating phenotypic characteristics of the clinically encountered members of the Pseudallescheria boydii complex and Scedosporium prolificans 210 Scedosporium prolificans (formerly Scedosporium inflatum) 211 Ochroconis gallopava (formerly Dactylaria constricta var.gallopava) 212 Table 19 Differentiation of the clinically encountered Ochroconis species 213 Table 20 Characteristics of some of the "black yeasts" 213 Exophiala jeanselmei complex 214 Exophiala dermatitidis (Wangiella dermatitidis) 215 Hortaea werneckii (Phaeoannellomyces werneckii) 216 Madurella mycetomatis 217 Madurella grisea 218 Piedraia hortae 219 Aureobasidium pullulans 220 Table 21 Differential characteristics of Aureobasidium pullulans versus Hormonema dematioides 222 Hormonema dematioides 223 Neoscytalidium dimidiatum (formerly Scytalidium dimidiatum) 224 Botrytis sp. 226 Stachybotrys chartarum (S. alternans, S. atra) 227 Graphium eumorphum 228 Curvularia spp. 229 Bipolaris spp. 230 Table 22 Characteristics of Bipolaris, Drechslera, and Exserohilum spp. 231 Exserohilum spp. 232 Helminthosporium sp 233 Alternaria sp 234 Ulocladium sp. 235 Stemphylium sp. 236 Pithomyces sp. 237 Epicoccum sp. 238 Nigrospora sp. 239 Chaetomium sp. 240 Phoma spp. 241 Dermatophytes 243 Introduction 245 Microsporum audouinii 247 Microsporum canis var. canis 248 Microsporum canis var. distortum 249 Microsporum cookei 250 Microsporum gypseum complex 251 Microsporum gallinae 252 Microsporum nanum 253 Microsporum vanbreuseghemii 254 Microsporum ferrugineum 255 Trichophyton mentagrophytes 256 Table 23 Differentiation of similar conidia-producing Trichophyton spp 257 Trichophyton rubrum 258 Trichophyton tonsurans 259 Trichophyton terrestre 260 Trichophyton megninii 261 Trichophyton soudanense 262 Table 24 Growth patterns of Trichophyton species on nutritional test media 263 Trichophyton schoenleinii 264 Trichophyton verrucosum 265 Trichophyton violaceum 266 Trichophyton ajelloi 267 Epidermophyton floccosum 268 Hyaline Hyphomycetes 269 Introduction 271 Coccidioides spp. 272 Table 25 Differential characteristics of fungi in which arthroconidia predominate 274 Malbranchea spp. 275 Geomyces pannorum 276 Arthrographis kalrae 277 Hormographiella aspergillata 278 Emmonsia spp. 279 The Genus Aspergillus 281 Aspergillus fumigatus 283 Aspergillus niger 284 Aspergillus flavus 285 Table 26 Identification of the most common species of Aspergillus 286 Aspergillus versicolor 288 Aspergillus calidoustus 289 Aspergillus nidulans (asexual state); Emericella nidulans (sexual state) 290 Aspergillus glaucus (asexual state); Eurotium herbariorum (sexual state) 291 Aspergillus terreus 292 Aspergillus clavatus 293 Penicillium spp. 294 Paecilomyces spp. 295 Scopulariopsis spp. 297 Table 27 Differential characteristics of Paecilomyces variotii versus P. lilacinus 299 Table 28 Differential characteristics of Scopulariopsis brevicaulis versus S. brumptii 299 Gliocladium sp. 300 Trichoderma sp. 301 Beauveria bassiana 302 Verticillium sp. 303 Acremonium (formerly Cephalosporium) spp. 304 Fusarium spp. 305 Lecythophora spp. 307 Trichothecium roseum 308 Chrysosporium spp. 309 Table 29 Differential characteristics of Chrysosporium versus Sporotrichum 311 Sporotrichum pruinosum 312 Sepedonium sp. 313 Chrysonilia sitophila (formerly Monilia sitophila) 314 Part III Basics of Molecular Methods for Fungal Identification Introduction 317 Molecular Terminology 318 Overview of Classic Molecular Identification Methods 322 Fungal Targets 322 Selected Current Molecular Methodologies 323 Amplification and Non-Sequencing-Based Identification Methods 323 PCR (Polymerase Chain Reaction) 323 Nested PCR 324 Real-time PCR 324 Melting curve analysis 324 Fluorescence resonance energy transfer (FRET) 325 TaqMan 5' nuclease 325 Molecular beacons 325 Microarray 326 Repetitive-element PCR (rep-PCR) 327 Sequencing-Based Identification Methods 327 Sanger sequencing 327 Pyrosequencing 328 DNA barcoding 328 Applications of DNA Sequencing 329 Accurate method identification 329 Phylogenetic analysis 330 Organism typing 332 Commercial Platforms and Recently Developed Techniques 332 AccuProbe test 332 PNA FISH 332 Luminex xMAP 333 MALDI-TOF 333 Selected References for Further Information 334 Part IV Laboratory Technique Laboratory Procedures 339 Collection and Preparation of Specimens 341 Methods for Direct Microscopic Examination of Specimens 344 Primary Isolation 346 Table 30 Media for primary isolation of fungi 348 Table 31 Inhibitory mould agar versus Sabouraud dextrose agar as a primary medium for isolation of fungi 349 Macroscopic Examination of Cultures 349 Microscopic Examination of Growth 350 Procedure for Identification of Yeasts 352 Direct Identification of Yeasts from Blood Culture (by PNA FISH) 354 Isolation of Yeast When Mixed with Bacteria 355 Germ Tube Test for the Presumptive Identification of Candida albicans 356 Rapid Enzyme Tests for the Presumptive Identification of Candida albicans 356 Caffeic Acid Disk Test 357 Olive Oil Disks for Culturing Malassezia species 357 Conversion of Thermally Dimorphic Fungi in Culture 358 Method of Inducing Sporulation of Apophysomyces elegans and Saksenaea vasiformis 358 In Vitro Hair Perforation Test (for Differentiation of Trichophyton mentagrophytes and Trichophyton rubrum) 359 Germ Tube Test for Differentiation of Some Dematiaceous Fungi 359 Temperature Tolerance Testing 360 Maintenance of Stock Fungal Cultures 360 Controlling Mites 361 Staining Methods 363 Acid-Fast Modified Kinyoun Stain for Nocardia spp. 365 Acid-Fast Stain for Ascospores 366 Ascospore Stain 366 Calcofluor White Stain 366 Giemsa Stain 367 Gomori Methenamine Silver (GMS) Stain 368 Gram Stain (Hucker Modification) 370 Lactophenol Cotton Blue 371 Lactophenol Cotton Blue with Polyvinyl Alcohol (PVA) (Huber's PVA Mounting Medium, Modified) 371 Rehydration of Paraffin-Embedded Tissue (Deparaffination) 372 Media 373 Introduction 375 Acetamide Agar 375 Arylsulfatase Broth 376 Ascospore Media 376 Assimilation Media (for Yeasts) 377 Birdseed Agar (Niger Seed Agar; Staib Agar) 381 Brain Heart Infusion (BHI) Agar 382 Buffered Charcoal Yeast Extract (BCYE) Agar 382 Canavanine Glycine Bromothymol Blue (CGB) Agar 383 Casein Agar 384 CHROMagar Candida Medium 384 ChromID Candida Medium 385 Citrate Agar 386 Cornmeal Agar 386 Dermatophyte Test Medium (DTM) 387 Dixon Agar (Modified) 388 Esculin Agar 388 Fermentation Broths for Yeasts 389 Gelatin Medium 389 Inhibitory Mould Agar (IMA) 391 Leeming-Notman Agar (Modified) 391 Loeffler Medium 392 Lysozyme Medium 392 Middlebrook Agar Opacity Test for Nocardia farcinica 393 Mycosel Agar 393 Nitrate Broth 394 Polished Rice, or Rice Grain, Medium 394 Potato Dextrose Agar and Potato Flake Agar 395 Rapid Assimilation of Trehalose (RAT) Broth 395 Rapid Sporulation Medium (RSM) 397 SABHI Agar 397 Sabouraud Dextrose Agar (SDA) 398 Sabouraud Dextrose Agar with 15% NaCl 399 Sabouraud Dextrose Broth 399 Starch Hydrolysis Agar 399 Trichophyton Agars 400 Tyrosine, Xanthine, or Hypoxanthine Agar 401 Urea Agar 401 Water Agar 402 Yeast Extract-Phosphate Agar with Ammonia 402 Color Plates 405 Glossary 435 Bibliography 447 Selected Websites 465 Index 469

About the Author

Davise H. Larone is well known as the originator of the book that many readers have come to rely upon for assistance in the accurate identification of fungi from patient specimens, a key step in treating mycotic infections. Dr. Larone has now been joined by Thomas J. Walsh and Randall T. Hayden to update this gold standard reference while retaining the format that has made this guide so popular for more than 40 years.

Reviews

This well-established manual has really withstood the test of time, evolving through the last four decades. It first came out in 1976, and the edition prior to this one was published by the American Society of Microbiology in 2011. This record demonstrates that it fulfills a real need amongst medical mycologists; indeed, it is referred to in the Preface as an esteemed, beloved, and time-honored book (p. xvii). This is perhaps in no small measure because the authors are all hospital or medical college based and at the sharp end of diagnosis of conditions due to fungi. They take a pragmatic approach and have endeavored to provide a manual that provides as much as possible to make this a one-stop-shop for clinicians and laboratory technicians taking them as far as they can and then with information on how to proceed with rare or atypical fungi. Indeed, guidance on the use of reference laboratories and how to safely package and transport material appears right at the start of the book, followed by sections of safety procedures to be followed and taxonomy and nomenclature. I was pleased to see the issue of the need to be aware of cryptic species being flagged up, and that the one-name-one-fungus decision had been embraced, albeit with the common misunderstanding that this was effective from January 2013 rather than the actual date of July 2011... While the book does perhaps have something of a North American slant, for example not including species such as Neotestudina rosatii, it is by far the best book on clinical mycology I am aware of tailored for the hospital laboratory, providing a bridge between more superficial texts and the monographic approach of the Atlas of Clinical Fungi. Davise Larone should be very pleased to see the work she started so long ago going from strength to strength, and continuing to fulfill a real need, as she commences her 80th year. - David Hawksworth, excerpted from the full review in IMA Fungus Vol. 9, No. 2, Book News" Recognizing the increase in fungal infections of humans and the decrease of formal training in mycology for clinical laboratory personnel, the bench-side guide and teaching aid describes the macroscopic and microscopic morphologies of cultured fungi as pertaining to those on Sabouraud dextrose agar. Black and white images depict different types of filamentous bacteria, yeasts, thermally dimorphic fungi, and thermally monomorphic molds. A section on laboratory technique details lab procedures, staining methods, and media preparation. The sixth edition adds an essay on taxonomy and nomenclature, identifies emerging pathogens, and updates technological advances in next-generation sequencing, real-time PCR, melt curve analysis, and T2 magnetic resonance. - PROTOVIEW (NO INDIVIDUAL REVIEWER NOTED)

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