**4. Discussion**

The sporotrichoid form (SF) of cutaneous leishmaniasis is defined by the development of palpable (and often visible) painless subcutaneous nodules distributed in a linear fashion along the lymphatics and extending proximally from a cutaneous lesion. In the Old World, sporo‐ trichoid leishmaniasis has mainly been associated with *L. major* ZCL [8, 9, 10, 11]. Its incidence ranges from 6% in Iran [12], to 10% in Saudi Arabia [10] and 11 to 22 % in Sudan [8, 9], whereas it seems to be less frequent in other *L. major* endemic areas like Turkey and Pakistan [13, 14].

In Tunisia, where ZCL is highly endemic, a previous study conducted by Masmoudi et al. [15] showed the SF to be common among patients originating from ZCL endemic areas in Sfax, Sidi Bouzid and Kasserine Governorates, with an incidence of 18%. In our study, which covered a much longer period and a higher number of patients, the incidence was much lower (1.7%). These contrasting results are difficult to explain. One of the factors could be a difference in the pathogenicity of infecting *Leishmania* strains according to the transmission areas. In a study conducted by Gaafar et al. [9] in Sudan and Saudi Arabia, it was shown that the incidence of the SF could be related to the causative *L. major* zymodeme. Yet, all characterized Tunisian *L. major* strains whatever obtained from patients, animal reservoirs or sand flies were found to belong to the single MON 25 zymodeme [2, 16, 17]. Nevertheless, we may consider that this zymodeme includes strains with a variable virulence, unevenly distributed over the trans‐ mission foci. An additional explanation may be the degree of immunization of infected populations, those living in emergent foci (like Sfax and Kasserine) being more susceptible to dissemination than those living in more ancient foci. In our series, nearly the 2/3 of patients with the SF came from Mahdia and Sousse Governorates where the ZCL epidemics occurred much later as compared to Kairouan Governorate (where the outbreak first arose) and Sidi Bouzid Governorate, now considered to include the more active foci in the region [1, 2].

The typing of strains obtained from 10 out of our 34 patients showed that most of them (9 strains) were, as expected, *L. major*. Interestingly the last one was *L. killicki*. This finding is not too surprising as the patient came from Ain Jloula, a recent emerging focus of *L. killicki* [2, 16]; and means that *L. killicki* may be a causal agent of SF.

Histopathologic studies show the subcutaneous nodules to consist of a granulomatous reaction with lymphocytes, plasma cells, macrophages and giant cells [9, 15, 18, 19]. They are supposed to represent an immunological reaction against the lymphatic spread of the parasite or its antigens [10, 11]. The factors that trigger lymphatic dissemination are unclear but some data suggest that the host immune status could influence this dissemination [10, 11]. Indeed, Sadeghian et al. [20] showed that the leishmanin skin test (LST) was negative in 72% of SF cases and that none was strongly positive in LST positive patients. They concluded that the SF of CL is at least partially due to a decrease in cellular immunity. However, the role of immunity should be considered with caution because if the immune response is actually involved, it cannot be the sole or the determinant factor as in this situation the age of patients with SF is expected to be lower than that of those with the common ZCL form, the children being much more affected. Yet, this is not the case in our study neither in that of Masmoudi et al. [15]. where the median age of patients with SF was higher as compared to patients with the common form (47.8 years and 26.8 years respectively).

**4. Discussion**

220 Leishmaniasis - Trends in Epidemiology, Diagnosis and Treatment

The sporotrichoid form (SF) of cutaneous leishmaniasis is defined by the development of palpable (and often visible) painless subcutaneous nodules distributed in a linear fashion along the lymphatics and extending proximally from a cutaneous lesion. In the Old World, sporo‐ trichoid leishmaniasis has mainly been associated with *L. major* ZCL [8, 9, 10, 11]. Its incidence ranges from 6% in Iran [12], to 10% in Saudi Arabia [10] and 11 to 22 % in Sudan [8, 9], whereas it seems to be less frequent in other *L. major* endemic areas like Turkey and Pakistan [13, 14].

In Tunisia, where ZCL is highly endemic, a previous study conducted by Masmoudi et al. [15] showed the SF to be common among patients originating from ZCL endemic areas in Sfax, Sidi Bouzid and Kasserine Governorates, with an incidence of 18%. In our study, which covered a much longer period and a higher number of patients, the incidence was much lower (1.7%). These contrasting results are difficult to explain. One of the factors could be a difference in the pathogenicity of infecting *Leishmania* strains according to the transmission areas. In a study conducted by Gaafar et al. [9] in Sudan and Saudi Arabia, it was shown that the incidence of the SF could be related to the causative *L. major* zymodeme. Yet, all characterized Tunisian *L. major* strains whatever obtained from patients, animal reservoirs or sand flies were found to belong to the single MON 25 zymodeme [2, 16, 17]. Nevertheless, we may consider that this zymodeme includes strains with a variable virulence, unevenly distributed over the trans‐ mission foci. An additional explanation may be the degree of immunization of infected populations, those living in emergent foci (like Sfax and Kasserine) being more susceptible to dissemination than those living in more ancient foci. In our series, nearly the 2/3 of patients with the SF came from Mahdia and Sousse Governorates where the ZCL epidemics occurred much later as compared to Kairouan Governorate (where the outbreak first arose) and Sidi Bouzid Governorate, now considered to include the more active foci in the region [1, 2].

The typing of strains obtained from 10 out of our 34 patients showed that most of them (9 strains) were, as expected, *L. major*. Interestingly the last one was *L. killicki*. This finding is not too surprising as the patient came from Ain Jloula, a recent emerging focus of *L. killicki* [2, 16];

Histopathologic studies show the subcutaneous nodules to consist of a granulomatous reaction with lymphocytes, plasma cells, macrophages and giant cells [9, 15, 18, 19]. They are supposed to represent an immunological reaction against the lymphatic spread of the parasite or its antigens [10, 11]. The factors that trigger lymphatic dissemination are unclear but some data suggest that the host immune status could influence this dissemination [10, 11]. Indeed, Sadeghian et al. [20] showed that the leishmanin skin test (LST) was negative in 72% of SF cases and that none was strongly positive in LST positive patients. They concluded that the SF of CL is at least partially due to a decrease in cellular immunity. However, the role of immunity should be considered with caution because if the immune response is actually involved, it cannot be the sole or the determinant factor as in this situation the age of patients with SF is expected to be lower than that of those with the common ZCL form, the children being much more affected. Yet, this is not the case in our study neither in that of Masmoudi et al. [15]. where

and means that *L. killicki* may be a causal agent of SF.

In our study, females were more represented then males (F/M sex ratio = 2.1). A similar finding (F/M sex ratio = 1.7) was reported by Masmoudi et al. [15]; whereas in other previous studies, no significant association of subcutaneous nodules with sex was reported [9].

The time to appearance of nodules is variable. It ranges from a few days to a few weeks or months; nodules may even appear after complete healing of the primary cutaneous lesions [8], with a delay reaching 1 or 2 years in some reported cases [15, 21]. In most of our patients, the nodules appeared in less than one month after the onset of the primary lesions.

Among the factors that trigger the lymphatic dissemination of parasites away from the primary lesions, many reports suggest the intralesional antimonial treatment to play a role [9, 10, 11, 22]. The reason for that is unclear. Nevertheless, this factor appears not to be determinant as only 6 out of our 34 patients (17.6%) had received intralesional Glucantime®; in the remaining 28 patients the nodules appeared spontaneously. The same finding was reported in Masmoudi et al. study [15] where 74% of patients did not receive any Glucantime® infiltrations before the development of nodules.

The number of sporotrichroïd nodules is reported to be variable, but they often are multiple [10, 11, 19, 21]. In our patients the number ranged from 2 to > 10; most of patients had 2 to 4 nodules as shown in figure 1. In the series of Masmoudi et al. [15], this number reached 20 nodules in one patient, with a mean of 7 nodules.

When mentioned, the size of nodules is usually 1 to 2 cm [11, 15, 19]. In our patients, most nodules sized between 0.5 to 1 cm, the largest being 3 cm.

In our 34 patients, nodules mainly developed on forearms and arms; the lower limbs and other sites of the body were much less represented. This finding was expected as the distribution of nodules overall reflects that of primary lesions which mainly involved the upper limbs. This result is very similar to that of Masmoudi et al. [15] where nodules were located on upper limbs in 80% of cases, together with the primary ulcers. Nevertheless, other sites can be affected; indeed, in some of our patients, SN developed on the legs, the cheeks, the eyelids, the nose and the back.

The diagnosis of CL in all of our 34 patients was confirmed by the demonstration of amastigotes in ulcerated primary lesions, while SF was diagnosed on the basis of clinical criteria. In the 3 patients where the biopsy of nodules was performed, no parasites could be shown. According to previous reports, amastigotes may or may not be demonstrable on smears or impressed biopsies nodules [8, 10, 15, 19, 23]. The diagnosis of the SF is usually suspected when nodules develop in the vicinity of a previously confirmed primary CL lesion, and resolve under antimonial treatment. As stated above, the amastigotes may or may not be demonstrated in SN. Similar conditions mainly include sporotrichosis that, to our knowledge, has never been reported in Tunisia. Cutaneous tuberculosis and atypical mycobacteriosis can also be consid‐ ered but both conditions are unresponsive to antimonials.

**Figure 1.** Sporotrichoid nodules (arrows) on the forearm of a patient.

While in localized CL local treatment such as intralesional infiltrations of pentavalent anti‐ monials and /or cryotherapy may be helpful and sufficient, such treatment is of no value in disseminated disease where systemic antimonials must be started [23, 24]. In most reported patients with SF, systemic treatment was successful and outcome favorable [5, 14, 15], even though some patients were reported to be unresponsive to antimonials [11, 19, 22]. Thirty three (97%) of our patients responded well to systemic Glucantime® and SN together with primary lesions scared in 2 to 8 weeks. In the last patient, both primary ulcers and SN persisted up to 13 months. It is worth mentioning that this patient was receiving a long term corticotherapy and developed symptoms of antimonials toxicity which led to the interruption of the Glucan‐ time® cure.

### **5. Conclusion**

The SF of CL appears to be a rather rare event in Central Tunisian SCL foci. In patients affected, outcome is similar to uncomplicated cases.
