Morpology
Diagnostic Tests
Management and Therapy

MORPHOLOGY:

Babesia microti infection, Giemsa stained thin smear.
Intraerythrocytic protozoa measure 1-3 microns. The organisms resemble Plasmodium falciparum; however, Babesia parasites present two distinguishing features. They vary more in shape and in size and they do not produce pigment. The picture at left is a 67-year-old woman, status post-splenectomy, infection probably acquired in Long island (New York).

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DIAGNOSTIC TESTS:
Microscopy
Infection with Babesia. Giemsa stained thin smears. Note the tetrad on the left side of image A, a dividing form pathognomonic for Babesia. Note also the variation in size and shape of the ring stage parasites (compare A and B), and the absence of pigment. A 6-year-old girl, status post splenectomy for hereditary spherocytosis, infection acquired in the U.S.
A
B

The disease is diagnosed by the presence of the intracellular parasites in erythrocytes. Although these organisms are often mistaken for those of malaria, they can be distinguished on Giemsa stain of peripheral blood by the lack of gametocyte forms and the absence of intraerythrocytic pigment in Babesia-parasitized cells. Since the organism divides by budding, as opposed to schizogeny, characteristic tetrads may be seen identifying the organism as Babesia. Isolation of the organisms by inoculation of patient blood into hamsters or gerbils, also known as a xenodiagnosis, may also assist in diagnosis.

Molecular Methods
Agarose gel (2%) analysis of a PCR diagnostic test for detection of Babesia microti DNA. PCR was performed using a nested protocol with primers BAB1 and BAB4 (first round) and BAB2 and BAB3 (second round).

* Lane S: Molecular base pair standard (50-bp ladder). Black arrows show the size of standard bands.
* Lane 1: First step amplification with primers BAB1 and BAB4 of the nested PCR protocol for detection of B. microti in DNA extracted from whole blood. The specimen, serologically positive for B. microti, was submitted to the CDC by the American Red Cross. The red arrow shows the single-step PCR diagnostic band for B. microti (size: 238 bp).
* Lane2: Nested PCR with primers BAB2 and BAB3 using as template the product of the first step amplification. The blue arrow shows the nested PCR diagnostic band for B. microti (size: 154 bp). Please note the enhanced sensitivity of B. microti DNA detection with the nested reaction.

In some infections with intra-erythrocytic parasites, the morphologic characteristics observed on microscopic examination of blood smears do not allow an unambiguous differentiation between Babesia and Plasmodium. Moreover, potential blood donors may have subclinical symptoms and very low parasitemia, undetectable in blood smears. In such cases, the diagnosis can be derived from molecular techniques, such as PCR using the appropriate primers and single-step, or the more sensitive nested PCR technique. In addition, molecular approaches are very valuable in investigations of new Babesia variants (or species) observed in recent human infections in the US and in Europe.

Antibody Detection
Diagnosis of Babesia infection should be made by detection of parasites in patients' blood smears. However, antibody detection tests are useful for:
*detecting infected individuals with very low levels of parasitemia (such as asymptomatic blood donors in transfusion-associated cases),
*diagnosis after infection is cleared by therapy,
*and discrimination between Plasmodium falciparum and Babesia infection in patients whose blood smear examinations are inconclusive and whose travel histories cannot exclude either parasite.

The indirect fluorescent antibody test (IFA) using B. microti parasites as antigen detects antibodies in 88-96% of patients with B. microti infection. IFA antigen slides are prepared using washed, parasitized erythrocytes produced in hamsters. Patients' titers, or parasite concentrations, generally rise to >1:1024 during the first weeks of illness and decline gradually over 6 months to titers of 1:16 to1:256, but may remain detectable at low levels for a year or more. Specificity is 100% in patients with other tick-borne diseases or persons not exposed to the parasite. Cross-reactions may occur in serum specimens from patients with malaria infections, but generally titers are highest with the homologous antigen.

The extent of cross-reactivity between Babesia species is variable. A negative result with B. microti antigen for a patient exposed on the West Coast may be a false-negative reaction for Babesia infection. Individuals whose exposure could have occurred on the West Coast should be tested also for antibodies to the Babesia WA1 species, because of the lack of cross-reactivity with B. microti.
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MANAGEMENT AND THERAPY:
B. microti infections in patients with intact spleens are often self-limiting without treatment, although symptoms may persist for months with or without treatment. Because silent parasitemia may have prolonged symptoms and signs, treatment is advised for all patients infected with Babesia. Treatment with the combination of quinine sulfate (650 mg of salt orally tid) plus clindamycin* (600 mg orally tid or 1.2 g parenterally bid) for 7 to 10 days is usually effective but may not always eliminate parasites. The pediatric dose is 20 to 40 mg/kg per day for quinine sulfate and 25 mg/kg per day for clindamycin, both given in 3 divided doses over 7 to 10 days. Atovaquone* suspension (750 mg bid) plus azithromycin* (500 to 1000 mg/d) may be effective when quinine and clindamycin fail. Expecially severe infections with high-level B. microti parasitemia in asplenic patients have been successfully treated with exchange transfusions in addition to quinine and clindamycin*.

*These drugs are approved by the FDA but considered investigational for this purpose.
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All Giemsa stains are from http://www.dpd.cdc.gov/DPDx/HTML/Babesiosis.htm
The PCR test is also from http://www.dpd.cdc.gov/DPDx/HTML/Babesiosis.htm

Questions? Please e-mail me at juliet99@stanford.edu