Tissue and Blood Residing Nematodes Filarial worm Trichina worm Lecture 6 Medical Parasitology Course (MLAB 362) Dr. Mohamed A. El-Sakhawy 1 Common Characteristics Biohelminth Need intermediate host Location (residing site) Tissue and blood Ovoviviparous (larviparous) adult female deposit larvae 2 1
Filariasis Filariasis is caused by nematodes (roundworms) that inhabit the lymphatics and subcutaneous tissues. 8 main species infect humans. 3 of these are responsible for most of the morbidity due to filariasis: Wuchereria bancrofti and Brugia malayi cause lymphatic filariasis, and Onchocerca volvulus causes onchocerciasis (river blindness). The other five species are Loa loa, Mansonella perstans, M. streptocerca, M. ozzardi, and Brugia timori. (The last species also causes lymphatic filariasis.) 3 Geographic Distribution of filaria Among the agents of lymphatic filariasis, Wuchereria bancrofti is encountered in tropical areas worldwide; Brugia malayi is limited to Asia; The agent of river blindness, Onchocerca volvulus, occurs mainly in Africa, with additional foci in Latin America and the Middle East. Loa loa and Mansonella streptocerca are found in Africa; Mansonella perstans occurs in both Africa and South America; and Mansonella ozzardi occurs only in the American 4 continent. 2
Filaria 2 types of filaria I- Lymphatic filaria Wuchereria bancrofti Brugia malayi II- Tissue filaria Subcutaneous tissue Onchocerca volvulus: river blindness Loa loa: subcutaneous swelling Peritoneal cavity (Mansonella perstans) All species are transmitted by insect vectors 8 species could infect human being 5 Life Cycles of Filaria Infective larvae are transmitted by infected biting arthropods during a blood meal. 1- The larvae migrate to the appropriate site of the host's body, where they develop into microfilariae-producing adults. 2- The adults dwell in various human tissues where they can live for several years. The agents of lymphatic filariasis reside in lymphatic vessels and lymph nodes; Onchocerca volvulus in nodules in subcutaneous tissues; Loa loa in subcutaneous tissues,; Brugia malayi and Wuchereria bancrofti in lymphatics; Mansonella streptocerca in the dermis and subcutaneous tissue; Mansonella ozzardi apparently in the subcutaneous tissues; and M. perstans in body 6 cavities and the surrounding tissues. 3
3- The female worms produce microfilariae which circulate in the blood, except for those of Onchocerca volvulus and Mansonella streptocerca, which are found in the skin, and O. volvulus which invade the eye. 4- The microfilariae infect biting arthropods: mosquitoes for the agents of lymphatic filariasis; blackflies [Simulium] for Onchocerca volvulus; midges for Mansonella perstans and M. streptocerca; and both midges and blackflies for Mansonella ozzardi; and deerflies [Chrysops] for Loa loa). 5- Inside the arthropod, the microfilariae develop in 1 to 2 weeks into infective filariform (third-stage) larvae. 7 Lymphatic filaria Wuchereria bancrofti Brugia malayi 8 4
Life Cycle of Brugia malayi 9 Life Cycle of Brugia malayi The typical vector for Brugia malayi filariasis are mosquito species. During a blood meal, an infected mosquito introduces third-stage filarial larvae onto the skin of the human host, where they penetrate into the bite wound. They develop into adults that commonly reside in the lymphatics. The adult worms resemble those of Wuchereria bancrofti but are smaller. Female worms measure 43 to 55 mm in length by 130 to 170 μm in width, and males measure 13 to 23 mm in length by 70 to 80 μm in width. Adults produce microfilariae, measuring 177 to 230 μm in length and 5 to 7 μm in width, which are sheathed and have nocturnal periodicity. 10 5
The microfilariae migrate into lymph and enter the blood stream reaching the peripheral blood. A mosquito ingests the microfilariae during a blood meal. After ingestion, the microfilariae lose their sheaths and work their way through the wall of the proventriculus and cardiac portion of the midgut to reach the thoracic muscles. There the microfilariae develop into first-stage larvae and subsequently into third-stage larvae. The third-stage larvae migrate through the hemocoel to the mosquito's proboscis and can infect another human when the mosquito takes a blood meal. 11 Wuchereria bancrofti and Brugia malayi 6
Morphology I Adult: White and thread-like. Two rings of small papillae on the head. Female:5~10cm in length Male: 2.5~4cm and a curved tail with two copulatory spicules. Morphology II Microfilaria: 177~296 µm in length, a sheath with free endings. Bluntly rounded anteriorly and tapers to a point posteriorly. A nerve ring with no nuclei at anterior 1/5 of the body. Wuchereria bancrofti Brugia malayi 7
15 -Phenomen which the number of microfilariae in peripherial blood is very low density during daytime, but increase from evening to midnight and reach the greatest density at 10p.m to 2 a.m. -May be related to cerebral activity and vasoactivity of pulmonary vessels. 8
Characteristic of life cycle Host: Mosqutoes (intermediate host) Human (final host) Location: Lymphatics and lymph nodes Infective stage: Infective larvae Transmission stage: Microfilariae Diagnostic stage: Microfilariae Lymphatic filariasis often consists of asymptomatic microfilaremia. Some patients develop lymphatic dysfunction causing lymphedema and elephantiasis(in limb or scrotum). Pulmonary tropical eosinophilia syndrome, with nocturnal cough and wheezing, fever, and eosinophilia. 9
Location (adult): lymphatic system W.b: superficial and deeper e.g. lower limbs, groin, scrotum, etc. B.m: superficial e.g. mainly in lower limbs Infective stage: filariform larva Infection route: mosquito inoculation Discharge stage: microfilaria 19 Intermediate host & vector: mosquito W.b: Culex (Anopheles) B.m: Anopheles (Aedes) Mf show nocturnal periodicity Nocturnal periodicity Mfs appear in the peripheral blood in high density during the night, but hide in the pulmonary capillaries during the daytime while the host is awaken. W.b: 10 Pm ~ 2 Am B.m: 8 Pm ~ 4 Am 20 10
Lymphatic filariasis: elephantiasis is the last consequence of the swelling of limbs and scrotum. 21 Early hydrocoel in a Tanzanian man with W. bancrofti infection 22 11
Lymphatic filariasis: elephantiasis of scrotum. Genital manifestations are frequent in W. bancrofti infections while they are rare during B.malayi infections. 23 elephantiasis Hydrocele testis 24 12
Laboratory Diagnosis of lymphatic filaria Etiological examination Stained thick blood smear: first choice of methods Blood drop microscopy: used in the field Lymph node biopsy 25 Loa loa (Loaiasis or Loiasis) (The African Eye Worm) Distribution: West and Central parts of tropical Africa. Adult Morphology : as described before in the introduction of filaria but with tuberculated cuticle at both ends. 13
Life cycle: Habitat: 1. Subcutaneous tissues of man, free, migrating in chest, back, scalp, axilla, groin and may be seen under the conjunctiva (hence the common name eye worm). Life cycle of Loa loa 14
2. Intermediate host (vector) : Female Chrysops fly (Deer fly), day biting insect W. Bancrofi ). (transmission as in 3. Reservoir host : Monkeys. 4. The infective stage is the filariform larvae in the mouth of chrysops. 5. The Microfilaria has the following characters : 250 x 5. The sheath is tight. body curves are kinky. The tail is S-shaped and full of nuclei. The periodicity is diurnal with maximum at noon. 15
Microfilaria of Loa loa Pathogenesis & Symptomatology : Disease : Loiasis, Loaiasis Wandering of adult worms may be observed in loose connective tissues such as eye lids, conjunctiva, breast & scrotum creeping sensation, irritation and itching, congestion and oedema of eye lids. It can be seen moving under the conjunctiva or crossing the bridge of the nose, but it doesn t cause blindness. 16
Loiasis of the eye 2. Calabar swellings : allergic reaction to worm metabolites. It is named after a town in Eastern Nigeria. It is a transient swelling appears suddenly and disappears gradually within 2-3 days without suppuration. The commonest sites are forearm, hands, wrist, elbows and ankle. It is due to hypersensitivity (allergic reaction) to the adult worms, microfilariae and their toxic metabolites. 17
3. Complications are due to invasion of the worms to ectopic foci causing encephalopathy due to involvement of C.N.S where microfilariae can be detected in cerebrospinal fluid (C.S.F.). 4. Arthritis: Acute arthritis with joint effusion may occur. 5. Also hydrocele, glomerulonephritis, endomyocardial fibrosis and retinopathy can occur. Diagnosis: 1. Clinical: By observing the adult worm crossing the bridge of the nose or under the conjunctiva, together with history of calabar swelling. 2. Laboratory: Blood examination, a drop of the peripheral blood is taken during the day time (10 am. to 2 pm.) and is examined fresh for motile microfilariae and stained thick smear for fixed microfilariae. 18
3. History of calabar swelling. 4. Serological and intradermal test 5. Eosinophilia& leucocytosis. Control : Treatment of infected persons. Protection by screening or by fly repellents when visiting forests in endemic area. Chrysops control is difficult because it breeds in swampy areas of forest. 19
Laboratory Diagnosis of filaria Identification of microfilariae by microscopic examination is the most practical diagnostic procedure. Examination of blood samples will allow identification of microfilariae of Wuchereria bancrofti, Brugia malayi, Loa loa, Mansonella perstans, and M. ozzardi. It is important to time the blood collection with the known periodicity of the microfilariae. The blood sample can be a thick smear, stained with Giemsa or hematoxylin and eosin. For increased sensitivity, concentration techniques can be used. These include centrifugation of the blood sample lyzed in 2% formalin (Knott's technique), or filtration through a Nucleopore membrane. 39 Examination of skin snips will identify microfilariae of Onchocerca volvulus and Mansonella streptocerca. Skin snips can be obtained using a corneal-scleral punch, or more simply a scalpel and needle. The sample must be allowed to incubate for 30 minutes to 2 hours in saline or culture medium. Antigen detection using an immunoassay for circulating filarial antigens constitutes a useful diagnostic approach, because microfilaremia can be low and variable. A rapid-format immunochromatographic test. 40 20
Molecular diagnosis using PCR is available for W. bancrofti and B. malayi. Identification of adult worms is possible from tissue samples collected during nodulectomies (onchocerciasis), or during subcutaneous biopsies or worm removal from the eye (loiasis). Antibody detection is of limited value. Substantial antigenic cross reactivity exists between filaria and other helminths, and a positive serologic test does not distinguish between past and current infection. 41 a Infection with Wuchereria bancrofti: elephantiasis; b infection with Loa loa: eyelid swelling; c onchocercosis: cutaneous nodules caused by Onchocerca volvulus; d blindness caused by O. volvulus; e Trichinella spiralis; larvae in rat musculature; f larva migrans externa. 42 21
Trichina worm Trichinella spiralis قال تعانى بسى هللا انرح انرحيى )إ ا ح ر و ع ه ي ك ى ان ي ت ة و انذ و و ن ح ى ان خ ز ير و ي ا أ ه م ن غ ي ر هللا ب ه ف اض ط ر غ ي ر ب اغ و ل ع اد ف إ هللا غ ف ور ر ح يى ( ان حم - اآلية 115 43 Biological characteristics Zoonosis Ovoviviparous Adult & larva live in the same host individual Adult: small intestine Larva: striated muscle 44 22
Larva of Trichinella spiralis Trichinella spiralis larvae in muscle section. A higher power magnification 45 46 23
47 48 Life Cycle of T. spiralis Trichinellosis is acquired by ingesting meat containing cysts (encysted larvae) of Trichinella. After exposure to gastric acid and pepsin, the larvae are released from the cysts and invade the small bowel mucosa where they develop into adult worms (female 2.2 mm in length, males 1.2 mm; life span in the small bowel: 4 weeks). After 1 week, the females release larvae that migrate to the striated muscles where they encyst. Encystment is completed in 4 to 5 weeks and the encysted larvae may remain viable for several years. 24
Ingestion of the encysted larvae perpetuates the cycle. Rats and rodents are primarily responsible for maintaining the endemicity of this infection. Carnivorous/omnivorous animals, such as pigs or bears, feed on infected rodents or meat from other animals. Humans are accidentally infected when eating improperly processed meat of these carnivorous animals (or eating food contaminated with such meat). 49 Main Points of Life Cycle Infective stage: capsulated larva Infection route: eating raw or improperly cooked pork or its products No extra-hostal developing, but must change host to finish the life cycle exists many reservoir host Human acts as I.H and D.H 50 25
Pathogenesis Main pathogenic factor: larva Invasion stage (intestinal phase): minor digestive disturbance Migration stage (muscular phase): diarrhea, myalgias, abdominal pain, vomiting; weakness, blood eosinophilia; circumorbital edema; myositis; Muscular tenderness Encystment stage: recovery 51 Epidemiologiy Cosmopolitan, esp. in Europe and North-America China: due to raw pork consumption Diagnosis Based on the basis of clinical symptoms and history and eosinophilia Etiological diagnosis Biopsy of skeletal muscle Immunological diagnosis (antibody detection) 52 26
Control methods Hygienic education Properly cooking pork Low temperature storage of pork (all larvae are killed at -15ºC for 24hs) Scientific raising pigs: heat treatment of garbage used as pig food 53 27