Since dirofilariasis infection is often asymptomatic, diagnosis most frequently occurs serendipitously during routine chest x-ray or physical examination.  Upon x-ray, the nodule is often misdiagnosed as a carcinoma.  In such cases, only after surgical removal of the nodule, is the nematode pathogen identified.  Upon removal of the worm, microscopic evaluation of tissue cross-sections and study of macroscopic characteristics of the worm serve as the main methods of dirofilariasis diagnosis.  Another important diagnostic factor is the geographic area where the infection is believed to have been acquired.  Even though dirofilariasis infection is rare, much of the medical community is in agreement that extrapulmonary dirofilariasis should be considered in the differential diagnosis of nodular inflammatory infection, and pulmonary dirofilariasis should be considered in the differential diagnosis of solitary pulmonary nodules.
    Human dirofilariasis infection, especially human pulmonary dirofilariasis, is often a benign and self-limiting condition that does not necessitate immediate treatment.  However, it is often confused with life-threatening diseases, (such as tumors, tuberculosis, fungal infections, and hamartomas), and invasive surgical procedures are generally required in order to arrive at a definitive diagnosis (Muro, et al).  An additional problem is that misdiagnosis occasionally leads to inappropriate and bodily harmful therapeutic interventions (Vakalis, et al).  For example, this may occur if dirofilariasis infection in the breast were mistaken for breast cancer and chemtherapy were prescribed to treat the cancer.

    Another incentive for the development of a diagnostic technique that does not rely on analysis of the actual worm specimen is the fact that diagnosis of specific Dirofilaria species based on worm morphology can be complicated by by tissue degeneration and poor specimen preparation.  For example, the longitudinal ridges of the cuticles of some Dirofilaria can be useful for morphological diagnosis, but they can also be a source of misdiagnosis.  These ridges are well defined in D. repens, D. tenuis, D. ursi, and D. subdermata, but they are absent in D. immitis and poorly developed in D. striata and D. lutrae.  Due to the long incubation periods of many dirofilariasis infections, the dead worm has much time to undergo morphological degeneration within the nodule.  The lysis of the cuticle of the dead worm by eosinophils causes the worm to have a scalloped appearance that can be mistaken for ridges in cross sectioned specimens (Gutierrez).  Thus, if morphological degeneration has occurred, it is easy to confuse one Dirofilaria species for another.
    It would be beneficial to develop noninvasive, reliable diagnostic tests for dirofilariasis infection so that misdiagnosis could be prevented, infected individuals could be informed of the nature of the infection before having to go through the invasive process of having the worm surgically removed, and accurate diagnosis of the specific Dirofilaria species for the purpose of epidemiological studies could consistently and easily be achieved.

Figure 3: Midpower view of same bulbar conjunctival biopsy as in Figure 2, showing degenerate pieces
of an immature Dirofilaria immitis worm in cross- and longitudinal-sections.
From emedicine.com: http://author.emedicine.com/ped/topic599.htm.

Diagnostic Methods

• Serological detection:  Serology is an attractive alternative to invasive diagnosis because it avoids surgery.  Serological detection of human antibodies to dirofilariasis has been reported, but with poor sensitivity and specificity.  The efficacy in using ELISA to quantitatively detect dirofilarial antibodies has been debated due to the presence of cross-reacting antibodies.  Also, measurement of circulating parasite antigens has not been promising in past studies, given that human infection is caused by a single worm and the quantity of antigens released by the worm is thought to be too low to be accurately detected.  Early attempts to detect Dirofilaria antigens in human serum samples from Puerto Rico using micro-ELISA were unsuccessful.  However, when these same samples were analyzed using a combination of ELISA and Western Blot, the researchers were able to make quantitative measurements of specific antibodies and to identify specific parasitic antigens involved in the human immune response to dirofilariasis infection (Villanueva and Rodriguez-Perez).  Also, more recent immunological studies have shown that an antibody response to the Dirofilaria organism develops and is strong enough to be detected (Muro, et al).  Therefore, despite some difficulties, there is potential for the development of specific immunodiagnostic methods for human dirofilariasis infection.
• Detection by PCR:  Early protocols for PCR-based detection of D. repens DNA failed to amplify dirofilariae DNA from formalin preserved human biopsies.  One report published in 1999, however, presented an improved PCR protocol and discussed its potential application to routine clinical diagnosis in humas and large scale epidemiological studies in dogs (Vakalis, et al).
• Histologic Examination:  Routine histologic stains generally are not effective for Dirofilaria identification.  However, the cuticle of nematodes contains chitin, which can be stained with non-specific whiteners such as calcofluor white.  In general, Dirofilaria specimens stained with calcofluor can be easily recognized in tissue sections (Garcia).  Identification of Dirofilaria based on histological sections is optimal when the nematode is alive at the time of fixing so that tissue degradation and morphological degeneration has not occurred.  The characteristic morphologic features that are examined include: diameter of body, thickness and structure of cuticle, presence or absence of external cuticular ridges, hypodermis, lateral chords, number and disposition of myoid tissue fibers, digestive tract, male or female sexual tubules, male or female genital cells, and in the rare cases of pregnant females, the microfilariae inside the uterus (Pampiglione, et al).

Figure 4: Histopathologic D. immitisspecimen obtained by wedge resection.
Image from April, 2001 volume of Chest.
 

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