COLLEGE OF MEDICINE, NURSING & HEALTH SCIENCES

MLS 602 : Medical Microbiology LECTURE 6 : Superficial, Cutaneous and Subcutaneous Mycoses

Ms. Edwina Razak [email protected].

3/11/2020 MLS 602 EDWINA RAZAK 1 Expected Student Outcomes oList superficial, cutaneous and subcutaneous Mycoses o Describe caustic agents of superficial, cutaneous and subcutaneous mycoses o-Discuss the laboratory diagnosis of mycoses of skin, , and nails. o Explain how skin scrapings are collected and transported to the Microbiology laboratory for testing.

3/11/2020 MLS 602 EDWINA RAZAK 2 Overview of Integumentary system

3/11/2020 MLS 602 EDWINA RAZAK 3 Introduction Superficial and cutaneous • Fungal infections involving hair, skin or nails • No direct invasion of the deeper tissues • –agents of ringworm and athlete’s foot • Infect keratinized tissues

Subcutaneous ◦ Confined to subcutaneous tissue without dissemination to distant sites ◦ Slow growing

3/11/2020 MLS 602 EDWINA RAZAK 4 Superficial  outer most layers of the skin and hair There are two types  Infection of skin: Caused by furfur (pityriasis versicolor) and Exophiala werne ckii ()  Infection of hair: Caused by (black ) and beigelii ().

3/11/2020 MLS 602 EDWINA RAZAK 5 Pityriasis Versicolor ()  chronic, usually asymptomatic, involvement of the stratum corneum  discoloration or depigmentation of the skin.  areas involved are mainly the chest, abdomen, upper limbs and back  Etiological agent: A lipophilic, -like . Spaghetti and meatball like appearance in skin scrapping

3/11/2020 MLS 602 EDWINA RAZAK 6 3/11/2020 MLS 602 EDWINA RAZAK 7 Exophiala werne ckii  disease is Tinea nigra Black macules Black and 2 cell oval yeast in skin scrapping particularly of the palms On Sabouraud agar the fungus develops as grey, yeast-like colonies, which gradually become, more mycelial and darker coloured with age.

3/11/2020 MLS 602 EDWINA RAZAK 8 Piedra  infection of the hair firm, irregular nodules along the hair shaft. Two types: 1. black piedra caused by Piedraia hortae oBlack nodule on hair shaft o club shaped asci, each with eight ascospores. 2. white piedra caused by . o Cream coloured nodule on hair shaft o Axillary, pubic, beard, and scalp hair may be infected.

3/11/2020 MLS 602 EDWINA RAZAK 9 Cutaneous mycoses: . Fungi invade the keratinized layers of skin, hair and nails and the diseases are limited in these layers. .The fungi causing these diseases are termed dermatophytes .The diseases are referred to as tinea. .filamentous fungi that infect only superficial keratinized tissues Classification A. : Trichophyton species infect hair, skin or nails. B. Microsporum: Microsporum species infect only hair and skin. C. Epidermophyton: Epidermophyton attacks the skin and nails but not the hair.

3/11/2020 MLS 602 EDWINA RAZAK 10 common cutaneous mycoses

3/11/2020 MLS 602 EDWINA RAZAK 11 3/11/2020 MLS 602 EDWINA RAZAK 12 Subcutaneous mycoses:

The infections involve the dermis, subcutaneous tissue, muscle and fascia. mycetoma, chromomycosis, and rhinosporidiosis.

3/11/2020 MLS 602 EDWINA RAZAK 13 sporotrichosis

3/11/2020 MLS 602 EDWINA RAZAK 14 sporotrichosis .Frequently encountered in gardeners, forest workers etc. .Yeast form is mostly cigar shaped .Contains melanin that protects it from macrophages

3/11/2020 MLS 602 EDWINA RAZAK 15 3/11/2020 MLS 602 EDWINA RAZAK 16 chromomycosis

3/11/2020 MLS 602 EDWINA RAZAK 17 What is Mycetoma

3/11/2020 MLS 602 EDWINA RAZAK 18 Mycetoma

3/11/2020 MLS 602 EDWINA RAZAK 19 Rhinosporidiosis. chronic granulomatous disease characterized by the development of large polyps or wart-like lesions in the nose, conjunctiva and occasionally in ears, larynx, bronchus, penile urethra, vagina, rectum and skin. The causative fungus . polypoid growth and histologic examination is best way of diagnosis Sporangia containing endospores

3/11/2020 MLS 602 EDWINA RAZAK 20 Subcutaneous  a painless subcutaneous nodule develops which enlarges to involve a whole limb or large areas of the body. Basiodiobolus haptosporus, a saprophytic phycomycete.

3/11/2020 MLS 602 EDWINA RAZAK 21 LABORATORY DIAGNOSTIC TESTS

3/11/2020 MLS 602 EDWINA RAZAK 22 3/11/2020 MLS 602 EDWINA RAZAK 23 3/11/2020 MLS 602 EDWINA RAZAK 24 3/11/2020 MLS 602 EDWINA RAZAK 25 20% Potassium hydroxide (KOH) .The potassium hydroxide is a clearing agent which makes the fungi more visible in tissue body fluids or body secretion. . It softens, digests and clears the tissues (e.g., keratin present in skin) surrounding the fungi so that the hyphae and conidia (spores) of fungi can be seen under microscope. .KOH is a strong alkali

3/11/2020 MLS 602 EDWINA RAZAK 26 Procedure of KOH Preparation Place a drop of KOH solution on a slide. Transfer the specimen (small pieces) to the drop of KOH, and cover with glass. Place the slide in a petri dish, or other container with a lid, together with a damp piece of filter paper or cotton wool to prevent the preparation from drying out. Note: To assist clearing, should not be more than 5 mm long, and skin scales, crusts and nail snips should not be more than 2 mm across. As soon as the specimen has cleared, examine it microscopically using the 10X and 40X objectives with the condenser iris diaphragm closed sufficiently to give a good contrast. If too intense a light source is used the contrast will not be adequate and the unstained fungi will not be seen.

3/11/2020 MLS 602 EDWINA RAZAK 27 Disadvantages of KOH preparation method Experience required since background artifacts are often confusing. Clearing of some specimens may require an extended time Procedure to make 100 ml of KOH 20% solution: 1.Weigh 20 g potassium hydroxide (KOH) pellets. 2.Transfer the chemical to a screw-cap bottle. 3.Add 50 ml distilled water, and mix until the chemical is completely dissolved, add remaining distilled water and make the volume 100 ml. 4.Label the bottle and mark it corrosive. Store it at room temperature. The reagent is stable for up to 2 years

3/11/2020 MLS 602 EDWINA RAZAK 28 Modification in KOH Preparation method Use of dimethylsulphoxide-KOH reagent: The addition of dimethylsulphoxide (DMSO) to KOH enables specimens to be examined immediately or after only a few minutes. KOH with blue-black fountain pen ink added: The ink is not specific for fungi as it stains cells and other components in the skin. The addition of ink is recommended when Malassezia furfur is suspected .

3/11/2020 MLS 602 EDWINA RAZAK 29 Lactophenol Cotton Blue Stain

3/11/2020 MLS 602 EDWINA RAZAK 30 Culture media • The medium commonly employed is Sabouraud Dextrose Agar (SDA) [pH – 5.4] • The media may supplemented with antibiotics, such as gentamicin and chloramphenicol to minimize bacterial contamination and cycloheximide to inhibit saprophytic fungi. • Potato dextrose agar, Brain heart infusion (BHI) agar and Czapek Dox agar

3/11/2020 MLS 602 EDWINA RAZAK 31 3/11/2020 MLS 602 EDWINA RAZAK 32 3/11/2020 MLS 602 EDWINA RAZAK 33 Summary  Superficial Mycoses .Malassezia furfur (pityriasis versicolor) .Exophiala werne ckii (tinea nigra) .Piedraia hortae (black piedra) . Trichosporon beigelii (white piedra). Cutaneous Mycoses .Tinea/dermatophytes A.Trichophyton: hair, skin or nails. B. Microsporum: only hair and skin. C. Epidermophyton- Skin, nails and not hair

3/11/2020 MLS 602 EDWINA RAZAK 34 Subcutaneous Mycoses .Mycetoma . chromomycosis- .sporotrichosis – .Rhinosporidiosis- Rhinosporidium seeberi.

3/11/2020 MLS 602 EDWINA RAZAK 35