Test for Leptospirosis laus.ppt

GROUP 6Alagdon, Edsel

Gomez, Paul ArvinLaus, Abigail

BSMT 3-A

Tightly coiled, thin, flexible spirochetes with both ends hooked

Motile L. interrogans – all pathogenic strains L. biflexa – saprophytic strains Obligate aerobe Culture media – 14% (vol/vol) rabbit

serum• Fletcher’s semisolid• Stuart liquid• Ellinghausen-McCullough-Johnson-Harris (EMJH)

semisolid media• Polysorbate 80 (Tween 80)-albumin

L. interrogans serovar ictrohaemorrhagiae • rats

L. interrogans serovar Pomona and Hardjo • cattles

Reduced Phagocytosis in the host Cell-mediated sensitivity to

leptospiral antigen by the host Small amounts of endotoxins

produced by some strains

Route – small breaks in the skin or intact mucosa

Incubation period – 10-12 days Severe systemic disease (Weil’s

disease) includes renal failure, hepatic failure, and intravascular disease, and may result in death.

Specimen – CSF or Blood/SDS-treated blood (Acute phase)• Urine can be culture after the first week of

illness• Isolation Direct inoculation of 1 or 2 drops of freshly drawn

blood or CSF into CM and incubate the media in the dark at room temperature for 5 – 6 weeks at 28 ° C to 30 °C

Microscopic examination – Dark-field microscopy (tightly-coiled, rapidly motile spirochetes with hooked ends)

Urine (Specimen)• It should be inoculated immediately

because the acidity of the urine might harm the Leptospires.

• 1 or 2 drops of undiluted urine and 1:10 dilution of urine are added to 5 ml medium

• 200 ug/mL of 5-flurorouracil is added to prevent contamination of other bacteria

Direct Observation Insufficiently sensitive and extremely time

consuming Presence of fibrin strands in blood• In situ staining by immunofluorescence,

immunohistochemistry and sliver deposition Used in detection of Leptospires in

veterinary histopathology Replaced by PCR-based technology

Antigen Detection• EIA, RIA and chemiluminescence• Suffered from sensitivity problems and has

not gained widespread acceptance.

It is a molecular diagnostic procedure that has made possible the rapid and sensitive detection of leptospiral DNA in a variety of clinical samples.

The most frequent target sequences are 16S and 23S rRNA genes.

IgM antibodies are detected within a week after onset of disease.

IgG antibodies are detected a month or more after onset of disease.

IgM dot-ELISA assay – semiquantitative and it has 98% sensitivity and 90.6% specificity.

MAT – Microsopic Agglutination test• Gold standard for serological diagnosis.• Positive result – agglution of live or killed

Leptospires assessed by dark field microscopy.• It is most sensitive and specific serological test

available Microplate ELISA – detect Leptospira

IgM Abs LEPTO dipstick and LEPTO Dri-Dot Test

Leptospires is susceptible in vitro to antimicrobials streptomycin, tetracycline, doxycycline, and the macrolide antibmicrobics in vitro

Penicillin is considered beneficial and alters the course of the disease if treatment is initiated before the fourth day of illness.

Doxycycline shorten the course of illness in adults and reduce the incidence of convalascent leptospiuria.

Measurement of hemoglobin, total and differential white cell count, platelet count

Renal Function tests Serum bilirubin and serum

aminotransferase