Leishmania and helminths.

Two Brazilian cohort studies describe the frequency of helminth infections in patients with TL [5,12]. The first study recruited 120 patients with TL in a village health post in a rural area of Bahia state [5]. Only patients with cutaneous forms of leishmaniasis were included (maximum 4 lesions on maximum 2 body regions). The Leishmania species was not determined, but the predominant species in this region is known to be L. braziliensis. Study participants provided three stool samples for parasitological assays (sedimentation, Baermann, and Kato-Katz methods). Of the 120 patients with TL, 106 (88%) were diagnosed with a helminth infection. Seventy-three percent of the study participants were infected with more than one helminth species at the same time. The most common helminths in this study were Ancylostoma duodenale, Trichuris trichiura, Ascaris lumbricoides, Schistosoma mansoni, and Strongyloides stercoralis.

The second study was done in an urban area in the state of Rio de Janeiro [12]. This was a retrospective cohort study of 109 TL patients who received antimony therapy in a referral centre between 2004 and 2006: there were 99 cases of cutaneous and 10 of mucocutaneous leishmaniasis. All included patients had a parasitologically confirmed diagnosis of leishmaniasis. The species was typed in samples from 47 patients; they were all L. braziliensis. Parasitological examination of stool samples using sedimentation, Kato-Katz, and Baermann-Moraes methods was routinely performed during the study period. Sixteen (15%) out of 109 TL patients had helminth infections. The most frequent helminths were Ancylostomidae, A. lumbricoides, S. stercoralis, S. mansoni, and T. trichiura [12].

Leishmania and other Trypanosomatidae.

The existence of coinfection with T. cruzi was proven in Argentina in 1996 [33]. Seven (58%) out of 12 patients with TL were diagnosed with T. cruzi infection based on specific serological tests. In three of the seven coinfected patients, the presence of T. cruzi could be proven with a direct parasitological technique (i.e., xenodiagnosis using Triatoma infestans nymphs). Six additional studies confirmed, based on specific serological and molecular techniques, that T. cruzi coinfection is frequent in TL patients from Salta in northern Argentina [31, 34–37,43], where the seroprevalence of T. cruzi in rural populations is estimated to range between 4% and 30% [31,91]. In all these studies, the coinfected patients had clinical TL but no signs of cardiac abnormalities typical of Chagas disease at the time of recruitment. The largest study included 330 patients with TL caused by L. braziliensis or L. amazonensis and found coinfection with T. cruzi in 135 (41%) of them [36].

Coinfection with T. cruzi has also been found in other Latin American countries [30,32,39,40]. One study in a hospital in Los Yungas in Bolivia recruited 28 patients with TL caused by L. braziliensis complex, L. mexicana complex, or both and obtained positive PCR results for T. cruzi in 22 (79%) [32]. In Paraguay, 8 (8%) out of 101 patients with clinical TL coming from the Caazapá and Alto Paraná departments were suspected of carrying T. cruzi [39].

The largest prevalence study was done in Brazil and reported on the frequency of coinfection of L. braziliensis, L. infantum (syn. L. chagasi), and T. cruzi in a sample of 1,100 apparently healthy people living in fast-growing villages in the outskirts of São Luiz City, the capital of Maranhão State [30]. Diagnosis of Leishmania and Trypanosoma infections was based on serology and molecular testing of blood samples. Forty-one subjects (4%) were diagnosed with L. braziliensis infection only, 35 (3%) with T. cruzi only, 50 (5%) with L. chagasi only, 17 (2%) had L. braziliensis together with L. chagasi, 7 (1%) had L. chagasi together with T. cruzi, and 11 (1%) had L. braziliensis together with T. cruzi. None of the study participants had signs of past or present TL, visceral leishmaniasis, or Chagas disease.

Leishmania and human T-lymphotropic virus 1.

Three small studies in Colombia, Peru, and Iran reported a low frequency of human T-lymphotropic virus 1 (HTLV-1) infection in patients with TL. The number of study participants with TL ranged from 4 to 92, and the frequency of HTLV-1 infection ranged from 0% to 4% in subgroups with different forms of TL (subclinical or clinical, acute or chronic) [87–89]. A fourth study, from Mashhad in Iran, also failed to confirm a clear link between these two infections. These authors reported that 8 out of 100 HTLV-1–infected candidate blood donors mentioned a history of cutaneous leishmaniasis, which was not significantly different from the frequency reported by 100 HTLV-1–negative candidate blood donors [90].

Leishmania and other pathogens.

One study from Salta in northern Argentina looked into several coinfections at the same time [37]. In a series of 93 patients with parasitologically confirmed cutaneous (n = 50) or mucocutaneous (n = 43) leishmaniasis, 37% had one or more coinfection, i.e., intestinal parasites (n = 2), T. cruzi (n = 25), Toxoplasma gondii (n = 2), Paracoccidioides brasiliensis (n = 2), Coccidioides posadasii (n = 1), M, tuberculosis (n = 3), and/or Treponema pallidum (n = 4). The authors described that the frequency of coinfections was higher in patients with mucosal forms of leishmaniasis than in those with cutaneous leishmaniasis [37].

Our search retrieved no studies on the frequency of other coinfecting pathogens in TL patients or the general population, although there were some case reports and series. Therefore, we can only report on the absolute number of human cases with coinfection mentioned in the literature. We found reports of 16 cases of concurrent coinfection of Leishmania with T. gondii, 4 with Sporothrix schenckii, 10 with P. brasiliensis, 1 with Cryptococcus laurentii, 9 with M. tuberculosis, 25 with M. leprae, 1 with M. ulcerans, and 1 with Burkholderia pseudomallei (Table 1).

Types of interaction.

Coinfections may influence the immune response during TL in several different ways: through actions on local phagocytes, innate immune mechanisms, the balance between effector and regulatory T-cell subsets, and the capacity of macrophages to kill Leishmania amastigotes (Fig 2).

Download:

• PPT
  PowerPoint slide
• PNG
  larger image
• TIFF
  original image

Fig 2. Immune responses during TL and the potential for interference through coinfection: A means to focus new research.

(A) Leishmania parasite transmission during sandfly bite initiates TL. Local phagocyte function (including neutrophils, macrophages, and dendritic cells) may be affected by coinfections affecting skin homeostasis. Furthermore, coinfection may affect the nature of preexisting immunity to sandfly saliva and/or the local response to sandfly/parasite proteins. (B) Innate immune mechanisms regulated by stromal cells, dendritic cells, and innate lymphoid cells may all be influenced by the microenvironment created by local or systemic coinfection. (C) Changes to innate immunity or immunological cross-reactivity may influence the balance between effector (Th1, Th2, and Th17) and regulatory (R) T-cell subsets, leading to altered control of parasite load and/or altered immunopathology. (D) Coinfections may directly or indirectly alter macrophage intracellular signalling, affecting the intracellular survival of Leishmania independently of any effects on the specific T-cell response. ILC, innate lymphoid cell; Th, T helper cell; TL, tegumentary leishmaniasis.

https://doi.org/10.1371/journal.pntd.0006125.g002

There is considerable evidence supporting the roles of various key phagocyte populations (dermal macrophages, monocyte-derived macrophages and dendritic cells, and neutrophils) in the establishment of infection and first-line defence against Leishmania [92]. There is also a growing body of literature indicating that the functional attributes of these phagocytes can be influenced by products introduced during transmission (e.g., sandfly salivary proteins or parasite-derived immunomodulators) [93–95] or by changes in skin homeostasis (e.g., driven by pathologic coinfection or changes to the commensal microbiota) [96,97]. One study in mice showed that resident skin commensals were critical to promoting protective effector T-cell responses to L. major [98] and thus act as potent immunomodulatory coinfections necessary for the control of TL. However, specific publications about how phagocytes engaged in TL control may be affected by other pathogens or skin microbiota are currently lacking. Likewise, coinfection-associated changes in the function of innate lymphoid cells or mesenchymal stromal cells, although readily predicted from the literature, have yet to be shown to be relevant in established models of TL.

A well-known paradigm in immunity relates to the opposing effects of interferon-gamma (IFNγ) and interleukin-4 (IL-4) with regard to control of L. major lesion development in mice [99,100]. Whereas C57BL/6 mice self-heal under the control of IFNγ, BALB/c mice succumb to Leishmania infection in an IL-4–dependent manner. These counteracting cytokines were identified as the products of different subsets of CD4⁺ T helper (Th) cells (Th1 and Th2). The finding that these Th subsets/cytokines have different roles in the control of helminth versus Leishmania infection led to the notion that differing infections may skew T-cell immunity in polarised directions [100,101].

The included studies that contribute information about the interactions between Leishmania and specific other pathogens are summarised below per coinfecting agent. Most of these reports are based on research in animal models (n = 22), while only a few (n = 5) provide an extensive immunological characterisation of human coinfection. Most of the possible interaction mechanisms outlined in Fig 2 have not been covered yet by the specific literature about TL and coinfections included in this review.

Helminths.

The effect of helminth coinfection on the course of TL has been studied in mice models [7,23–29] and described in human patients [5,12,22], with mixed findings. Some of the studies in mice concluded that in the presence of helminth infection, the time between experimental infection with Leishmania and development of skin lesions increased [26,27], while others found that this prepatent period decreased [23] or remained unchanged [28]. The conclusions were also divided about the size of the TL lesions, finding larger [7], smaller [27], or similar lesions [25,28] in mice with helminth coinfection. One study with extended follow-up (16 weeks) showed that the impact of helminth coinfection on lesion growth was time dependent [26]. These divergent findings may be partly due to the parasites used in the experiments (S. mansoni or Litomosoides sigmodontis, with L. mexicana or L. major) and the time between the two experimental infections [23,26,27].

When it comes to explaining the effects of helminth coinfection on the course of TL, one experimental study suggested that the Th2 responses induced by helminth infection had systemic effects that down-regulated the initial, local Th1 response to Leishmania [26]. In contrast, several other studies found that helminth infection did not interfere with the generation of Leishmania-specific Th1-type responses [24,25,27–29]. Furthermore, two groups used in vitro models to show that macrophages from helminth-infected mice were impaired in their ability to kill Leishmania [7,26]. Three studies in mice also evaluated whether TL altered the course of helminth infections, but no measurable effect was reported [24,26,28].

Two cohort studies in Brazil compared the characteristics of TL in patients with and without helminthiasis [5,12]. The studies were conducted in Rio de Janeiro and Bahia, where L. braziliensis is predominant and pentavalent antimony is the recommended treatment. The study in Bahia enrolled 120 patients with cutaneous forms of TL (including 106 [88%] with helminthiasis), and the study in Rio de Janeiro enrolled 109 patients with cutaneous and mucocutaneous forms of TL (including 16 [15%] with helminthiasis). The helminths detected were A. duodenale, T. trichiura, A. lumbricoides, S. mansoni, and S. stercoralis. Both studies reported that the time to heal under pentavalent antimony treatment was longer for patients with TL and helminth infection than for patients with TL only [5,12]. The study in Rio de Janeiro also found significant associations of helminth coinfection with mucosal leishmaniasis and poor response to treatment [12].

Trypanosoma.

Four experimental studies (in mice or squirrel monkeys) and one observational study in humans addressed the effect of Trypanosoma coinfection (Trypanosoma brucei or T. cruzi) on TL [46–49]. Experimental Chagas disease did not protect against leishmaniasis and vice versa [46], although there were elements of immune cross-reactivity [47]. For the studies evaluating the impact of Trypanosoma on time until Leishmania lesion development [46–49], the main finding was a reduction in lesion growth rate in coinfected animals. In some cases, protection from ulceration was reported [46,48,49]. Normal lesion growth returned once the Trypanosoma infection was treated [48]. In one study in squirrel monkeys, L. braziliensis coinfection was shown to block the increase in QRS interval, i.e., the depolarisation time of the cardiac ventricles, which is normally associated with T. cruzi infection. This finding led the authors to suggest that prior infection with Leishmania parasites might provide some protection against Chagas-related cardiopathy [46]. One human immunological study focused on T-cell responses and showed that TL patients coinfected with T. cruzi had a higher T-cell differentiation profile than patients with TL only [44].

Toxoplasma.

Experimental studies in mice suggest that toxoplasmosis affects the course of leishmaniasis and vice versa [51,52]. Albino mice that were infected first with L. major and 30 to 70 days later with T. gondii developed more severe forms of leishmaniasis than mice infected with L. major alone [51]. By contrast, the course of toxoplasmosis was more benign in coinfected mice than in those infected with Toxoplasma alone [51]. Another study showed a different type of interaction. Here, BALB/c mice were experimentally infected first with T. gondii and 5 days later with L. major. The acute toxoplasmosis induced a strong Th1 response, and the BALB/c mice that are normally susceptible to leishmaniasis developed a level of resistance comparable to that of C57BL/6 mice [52]. In human patients, such positive or negative interactions between toxoplasmosis and TL have not been reported yet, although one in vitro study found that T. gondii–specific T cells are recruited into L. braziliensis lesions and could influence TL pathogenesis locally [50].

Plasmodium.

Seven experimental studies assessed Plasmodium coinfection and TL [53–59]. In coinfection models of Plasmodium yoelii or Plasmodium berghei together with Leishmania enrietti, L. mexicana, or L. amazonensis in hamsters, C57BL/6 mice, and BALB/c mice, the coinfected animals had larger lesions than the animals with Leishmania infection only. There was also an adverse effect of leishmaniasis on the course of malaria because coinfected animals had increased parasitaemia and mortality compared with animals with Plasmodium infection only [53–58]. These effects may vary according to the Leishmania species because one study of P. yoelii in BALB/c mice reported different findings for L. amazonensis and L. braziliensis [59].

Sporothrix.

Coinfection with Sporothrix may occur when fungal spores are inoculated in a TL lesion. In Colombia, it was suggested that such inoculations occur when people lance their TL lesions using Sporothrix-contaminated thorns [60]. There is also a case report linking coinfection with Sporothrix to traumatic injury and TL reactivation (Koebner phenomenon) [61].

M. tuberculosis.

We found 9 studies (8 case reports and 1 cross-sectional study) describing 12 human patients with concurrent tuberculosis and TL (Table 1). Five out of these 12 patients had mucosal forms of TL, and 4 had other, nonlocalised forms; the type of TL was not described in 3 patients. Results of leishmanin skin tests (arguably an in vivo correlate of Th1 responses) were available for 6 coinfected patients: 5 were positive or strongly positive. More detailed analyses of T-cell responses were not performed. Some authors hypothesised that an episode of tuberculosis can trigger reactivation of latent leishmaniasis [65,67–69]. Others suggested that an underlying immune defect could lead to the development of several infectious diseases at the same time [70]. This was based on the study of one patient who had lepromatous leprosy, several leishmaniasis lesions, and miliary tuberculosis and in whom a reduced responsiveness to IL-12 was found [70].

M. leprae.

The search retrieved 12 case reports/series of human patients with concurrent leprosy and TL, but none of them contained evidence of a significant interaction between the two infections. Leprosy and TL are both caused by obligate intracellular organisms and involve a broad spectrum of clinical, histopathological, and immunological manifestations [6,70,73–83]. The paucibacillary/pauciparasitic type of disease (tuberculoid leprosy and localised cutaneous leishmaniasis) is at one pole of the spectrum and reflects effective T-cell immunity. At the other pole of the spectrum is the multibacillary/multiparasitic type of disease (lepromatous leprosy and diffuse cutaneous leishmaniasis), which occurs when the antigen-specific T-cell response is depressed [70,82–83].

We found descriptions of five patients with lepromatous leprosy and localised TL [74,75,77–79]. In one of these cases, a man with lepromatous leprosy and mucosal leishmaniasis, skin reaction and IFNγ production against Leishmania antigens were strong, whereas the responses against M. leprae antigens were almost absent [78,79]. Therefore, despite the similarities in the pathogenesis of TL and leprosy, patients can have a divergent T-cell response to each pathogen, indicating a degree of compartmentalisation of T-cell immunity. Nonetheless, follow-up of one patient suggested that IL-10–mediated regulatory responses induced during leprosy may help control the immunopathology of mucosal leishmaniasis [78,79]. Twenty other patients described in the literature had disease manifestations of leprosy and TL that were not that far apart on the disease spectrum [6,70,73,74,76,80–82].

In addition to these naturally occurring combinations of TL and leprosy, we found descriptions of artificially induced coinfection [83,84]. In the 1950s and 1960s, it was common practice in some Leishmania-endemic areas to immunise people against leishmaniasis by the inoculation of live Leishmania tropica parasites (‘leishmanisation’). Two papers report on the clinical and histopathological evolution of 24 Israeli patients with lepromatous leprosy who received a vaccination with living Leishmania parasites. Twenty-three patients showed the classical clinical progression of cutaneous leishmaniasis at the site of inoculation. The authors suggested that this clinical response to vaccination was similar to that of people without leprosy [83]. One additional patient with lepromatous leprosy, described in a separate report, developed diffuse leishmaniasis after vaccination, but (also in this person) the lesions healed spontaneously. These observations also suggest that leprosy does not alter the course of TL or vice versa [84].

Clinical similarities complicating diagnosis.

A first diagnostic challenge occurs when there are clinical similarities between the lesions caused by Leishmania and some other pathogens. When one aetiological diagnosis is well established, a clinician may be tempted to attribute all the patient’s lesions to this one infection and stop examining the patient for symptoms and signs of other diseases. This may happen, for example, in patients with concurrent leprosy and leishmaniasis, particularly when patients have many skin lesions [82]. Furthermore, 2 case reports describe a year-long delay in the diagnosis of mucosal leishmaniasis because nasal symptoms were first attributed to leprosy [77,78]. Mucosal leishmaniasis can also be confused with mucosal manifestations of tuberculosis. Several authors have emphasised the importance of examining multiple samples from different skin lesions when coinfection is suspected [73–75,82]. Diagnosis of coinfection can become particularly challenging when more than one pathogen is present within the same lesion. Leishmania parasites have been found in skin or mucosal lesions together with S. schenckii, C. laurentii, M. tuberculosis, M. leprae, and M. ulcerans [6,60,61,64,65,85].

Biological similarities complicating diagnosis.

A second diagnostic challenge stems from the biological similarities between Leishmania parasites and other pathogens. This problem is well documented for Leishmania and T. cruzi, which are both kinetoplastid protozoa with antigenic similarities. When conventional serological tests are used for the diagnosis of Chagas disease, there is a problem of cross-reactivity with Leishmania. There have been several attempts to develop serological tests that differentiate Leishmania from T.cruzi infections [38,39,41,42] and to evaluate their diagnostic performance in settings in which both pathogens are endemic [42,43]. Tests using purified or recombinant specific antigens of T. cruzi, such as Ag163B6, Ag162B6/cruzipain, or shed acute phase antigen (SAPA), proved to be useful to identify true coinfections [41,42].

Issues with the interpretation of diagnostic test results.

One Brazilian study found that 52 out of 107 patients with a definite diagnosis of sporothrichosis also had one or more positive immunological test results for leishmaniasis (leishmanin skin test, ELISA, or indirect immunofluorescence test) [62]. The diagnosis of TL could not be confirmed in this study because parasitological confirmation tests were negative (n = 24) or not done (n = 28). It was therefore not possible to distinguish between true coinfections, serological cross-reactions, or false-positive results of the leishmanin skin test due to an allergy to the diluent [62]. The authors emphasise that in such a setting, incorrect diagnoses of TL are possible in patients with sporotrichosis and that, even in the presence of suggestive clinical and epidemiological arguments together with positive immunological test results for TL, parasitological confirmation is still needed before patients are exposed to a toxic and possibly unnecessary TL treatment [62].

Treatment sequence.

The first therapeutic challenge in patients with coinfection is to determine the best sequence of the different treatments. Because helminth coinfection appears to increase the time to healing in patients with cutaneous leishmaniasis [5,12], it seems logical to assume that prompt diagnosis and treatment of helminth infections may improve the outcome of TL treatment. One randomised, double-blind, placebo-controlled trial in Bahia, Brazil, examined early versus deferred treatment of helminth coinfection [22]. This trial enrolled 90 patients with cutaneous leishmaniasis (most probably caused by L. braziliensis) and helminth coinfection (mainly hookworms, T. trichiura, A. lumbricoides, S. mansoni, and S. stercoralis). All participants were treated with intravenous antimony at 20 mg/kg/day for 20 days. The treatment group also received triple antihelminthic therapy with albendazole, ivermectin, and praziquantel at days 0 and 30 and placebo at day 60. The control group received placebo at days 0 and 30 and specific antihelminthic therapy based on stool test results on day 60. There was no significant difference between the two groups in the time to healing of the skin lesions: the median time to cure was 98 days in the treatment group and 88 days in the control group [22].

Treatment side effects.

When two infections are treated at the same time, the drug combinations may lead to increased intolerance or adverse effects. The combination of antimony with antituberculous drugs is feared, and we found a description of death due to renal failure that was attributed to the combined treatment [67]. The combination treatment for TL (with pentavalent antimony) and leprosy (with diaminodiphenyl sulfone plus rifampicin plus clofazimine) may also produce considerable side effects [6]. Furthermore, several authors have raised concerns about the use of antimonial treatment for TL in patients with Chagas disease [40,45]. Pentavalent antimony drugs are known to prolong QT time and cause arrhythmia; they are therefore contraindicated in patients with known heart disease. On the one hand, cardiomyopathy is a well-known clinical manifestation of Chagas disease, and therefore prudence is called for in patients with Leishmania–Trypanosoma coinfection [40,45].

Unexpected responses to treatment.

Some case reports discussed unexpected benefits of 1 treatment on 2 infections. For example, there was a report about a patient with chagasic cardiomyopathy and TL [45]. Amiodarone was used to control the patient’s ventricular arrhythmia and seemed to promote the healing of TL. The authors considered that amiodarone could have had an antileishmanial effect although they could not rule out the possibility that the use of amiodarone coincided with the healing of TL by chance [45].

Another interesting case was reported in Colombia [69]. A patient diagnosed with mucocutaneous leishmaniasis and pulmonary tuberculosis first received treatment for tuberculosis with rifampin, isoniazid, streptomycin, and pyrazinamide over a period of 7 months. The antimonial treatment was deferred because of concerns about the adverse effects of the combination of antituberculous and antimonial drugs. Despite the lack of specific antileishmanial treatment, when assessed 3 months after the end of antituberculous therapy, the mucosal lesions were fibrosed, scar tissue was evident, and the patient was biopsy culture negative. A similar observation was reported in Brazil, where the lesions of a patient with diffuse cutaneous leishmaniasis temporarily improved while the patient was receiving antituberculous therapy [66]. Some studies have suggested that streptomycin, isoniazid, and rifampin may have direct antileishmanial activity [66]. Alternatively, this response might reflect an interaction between TL and tuberculosis. For example, reduction of mycobacterial burden may release regulatory pressure within the immune system that also favours resolution of mucosal lesions, or antituberculous treatment may (re)activate host protective mycobacteria-specific T cells that cross-react with Leishmania antigens.