Theme: Laboratory diagnosis of leptospirosis and boerrelia infections

Taxonomia Order: Spirochaetales Family: Spirochaetaceae –Genera: Treponema, Borrelia Family: Leptospiraceae –Genus: Leptospira

Cell structure

Leptospira Silver staining Burry stain with Indian ink

Life cycle of Leptospira

Leptospirosis

Weil's disease

Laboratory Diagnosis of Leptospirosis Diagnostic testSpecimenMethod of Examination MicroscopyBlood, tissueDark field microscopy Direct immunofluorescence Silver stain CultureBlood, liquorPositive during first 7 days UrinePositive after first week SerologySerumMicroscopic agglutination test Hemagglutination test ELISA CFT Nucleic acid probes Blood, tissuePCR

Agglutination and lysis test: 1- control; 2-4 – positive test.

Borrelia Borrelia recurrentis –louse-borne or epidemic relapsing fever B.duttoni, B.persica, B. hispanica, B.caucasica, and B. turicatae –tick-borne or endemic relapsing fever. Borrelia burgdorferi, B.afzelii, B.garini, Borrelia burgdorferi. –Lyme disease

Morphology

Transmission of tick-borne or endemic relapsing fever Reservoir: rodents Vector: ticks

Diagnosis of relapsing fever DiseaseMethodSpecimenTime Epidemic relapsing fever Microscopic examination 1.Dark-field microscopy 2.Burry stain 3.Giemsa stain Bloodacute stage Serology Rickenberg-Brusine test indirect IF, PHAT, CFT Serum from the 2 nd – 3 rd weeks Endemic relapsing fever Microscopic examination 1. Dark-field microscopy 2. Burry stain 3. Giemsa stain Bloodduring illness Biological method Inoculation of guinea pig Bloodduring illness

Life cycle of Borrelia burgdorferi

Cycle of B.burgdorferi in nature

Erythema migrans rash of a patient with Lyme disease

DIAGNOSIS of Lyme disease Serological tests: - ELISA. - IF. PCR.

Proper Tick Removal 1.Use fine-point tweezers or tick removal tool 2.Grasp close to skin 3.Pull gently 4.Wash area with soap, water and antiseptic Do not use gasoline or match Do not use Vaseline Never use products such as soap or finger nail polish to try and remove a tick - this will trigger injection of the salivary glands and disease causing microbes into the feeding source 19 PPT

How should ticks be removed to reduce the chance of infection? Just brush off ticks that have not attached If ticks have attached to host: –Grasp tick at point nearest attachment site with flat tweezers or fingers, apply gentle, constant tugging –Mouthparts will release after about a minute –If residual mouthparts are left in skin, leave those alone –Mouthparts will extrude from skin naturally over time Nightly tick check may reduce transmission –Takes h for B. burgdorferi to move from tick to host

TESTS 1. A man was admitted to the hospital on the 5th day of disease that manifested itself by jaundice, muscle aching, chill, nose bleedings. In course of laboratory diagnostics a bacteriologist performed darkfield microscopy of the patients blood drop. Name a causative agent of this disease: A. Leptospira interrogans B. Borrelia dutlonii C. Calymmatobacterium granulomatis D. Bartonella bacilloformis E. Rickettsia mooseri

2. A 33 year old patient was delivered to the infectious diseases department on the 7-th day of disease. He complained about great weakness, high temperature, pain in the lumbar area and leg muscles, icteritiousness, dark color of urine, headache. The acute disease started with chill, body temperature rise up to 40 o C, headache, pain in the lumbar area and sural muscles. Icterus turned up on the 4th day, nasal and scleral haemorrhages came on the 5th day. Fever has lasted for 6 days. Diuresis ml. What is the most probable diagnosis? A. Leptospirosis B. Typhoid fever C. Virus A hepatitis D. Sepsis E. Yersiniosis

3. A 25-year-old patient was delivered to an infectious diseases unit on the 3rd day of illness with complaints of headache, pain in lumbar spine and gastrocnemius muscles, high fever, chill. Objectively: condition of moderate severity. Scleras are icteric. Pharynx is hyperemic. Tongue is dry with dry brown coating. Abdomen is distended. Liver is enlarged by 2 cm. Spleen is not enlarged. Palpation of muscles, especially gastrocnemius muscles, is painful. Urine is dark in color. Stool is normal in color. The most likely diagnosis is: A. Leptospirosis B. Infectious mononucleosis C. Malaria D. Viral hepatitis A E. Yersiniosis