Leishmania (Viannia) naiffi: rare enough to be neglected?

Abstract

In the Brazilian Amazon, American tegumentary leishmaniasis (ATL) is endemic and presents a wide spectrum of clinical manifestations due, in part, to the circulation of at least seven Leishmania species. Few reports of Leishmania (Viannia) naiffi infection suggest that its occurrence is uncommon and the reported cases present a benign clinical course and a good response to treatment. This study aimed to strengthen the clinical and epidemiological importance of L. (V.) naiffi in the Amazon Region (Manaus, state of Amazonas) and to report therapeutic failure in patients infected with this species. Thirty Leishmania spp samples isolated from cutaneous lesions were characterised by multilocus enzyme electrophoresis. As expected, the most common species was Leishmania (V.) guyanensis (20 cases). However, a relevant number ofL. (V.) naiffi patients (8 cases) was observed, thus demonstrating that this species is not uncommon in the region. No patient infected withL. (V.) naiffi evolved to spontaneous cure until the start of treatment, which indicated that this species may not have a self-limiting nature. In addition, two of the patients experienced a poor response to antimonial or pentamidine therapy. Thus, either ATL cases due to L. (V.) naiffi cannot be as uncommon as previously thought or this species is currently expanding in this region.

Leishmania (Viannia) naiffi; therapeutic failure; clinical outcome; multilocus enzyme electrophoresis; Amazon Region

American tegumentary leishmaniasis (ATL) is highly endemic in the state of Amazonas (AM), Brazil. According to the Information System on Notifiable Diseases database, 18,675 Brazilian cases were reported in 2013, 8% of which occurred in the city of Manaus, AM (SVS/MS 2015). In this region, control of the disease is considered difficult because of the epidemiological characteristics and socioeconomic conditions associated with the sylvatic transmission cycle. In addition, environmental changes may influence infection dynamics, which makes control even more difficult. The circulation of at least seven Leishmania species has been observed in ATL in the Amazon Region. Leishmania (Viannia) guyanensisis the most prevalent species and is highly endemic in north of the Amazon River (Naiff et al. 1988, Grimaldi Jr et al. 1991, Lainson et al. 1994, Romero et al. 2001a, 2002a,Guerra et al. 2003). In this sense, several studies have shown that the frequency of circulating species in this area varies (Table I) and that Leishmania (V.) naiffi seems to be more consistently isolated from cutaneous lesions in patients from this region.

TABLE I
Frequency of Leishmania species associated with American tegumentary leishmaniasis in Amazon Region

Leishmaniaspecies	Cases (n)	Relative proportion within the study (%)	Region of endemicity state (city)	Reference
L. (Viannia) guyanensis	61	64.9	Amapá, Amazonas, Pará and Rondônia	Grimaldi Jr et al. (1991)
L. (V.) braziliensis	15	16
L. (V.) naiffi	10	10.6
L. (Leishmania) amazonensis	6	6.4
Not identified	2	2.1
L. (V.) guyanensis	69	97.2	Amazonas (Manaus)	Romero et al. (2002b)
L. (V.) braziliensis	2	2.8
L. (V.) braziliensis	16	53.3	Acre (Rio Branco)	da Silva et al. (2006)
L. (V.) lainsoni	12	40
L. (V.) guyanensis	1	3.3
Hybrid: L. (V.) lainsoni/L. (V.) naiffi	1	3.4
L. (V.) guyanensis	10	76.9	Amazonas (Manaus)	Figueira et al. (2008)
L. (V.) naiffi	3	23.1
L. (V.) guyanensis	8	80	Amazonas (Rio Preto da Eva)	Figueira et al. (2008)
L. (V.) naiffi	2	20
L. (V.) guyanensis	80	89	Amazonas (Rio Preto da Eva)	Figueira et al. (2014)
L. (L.) amazonensis	7	7.8
L. (V.) naiffi	3	3.3
L. (V.) braziliensis	11	25.6	Pará (lower Amazon Region)	Jennings et al. (2014)
L. (V.) guyanensis	6	13.9
L. (V.) shawi	4	9.3
L. (V.) shawi	7	16.3
L. (V.) lainsoni	6	13.9
L. (L.) amazonensis	2	4.7
Hybrid: L. (V.) guyanensis/ L. (V.) shawi shawi	7	16.3

L. (V.) naiffi was first described by Lainson and Shaw (1989) after being isolated from an armadillo (Dasypus novemcinctus) in the state of Pará, northern Brazil. The few cases described in the literature usually associate L. (V.) naiffiwith low rates of virulence in humans. Thus, it has been described that the disease evolves with a benign clinical course and a good response to treatment (Naiff et al. 1991, Pratlong et al. 2002). Furthermore, two cases of spontaneous healing of leishmaniasis caused by L. (V.) naiffi were described (van der Snoek et al. 2009). To date, however, no association between L. (V.) naiffi and mucosal leishmaniasis has been observed. L. (V.) naiffi cutaneous leishmaniasis lesions are usually ulcerated, unique, small and located on the hands, arms or legs (Naiff et al. 1991). In the same way, experimental studies have shown that L. (V.) naiffi frequently causes discrete or even nonapparent infections on hamsters’ skin (Lainson & Shaw 1989). These findings were supported by in vitro analysis, which demonstrated that L. (V.) naiffi showed the lowest infection index and the highest nitric oxide production compared with other species of the Viannia subgenus (Campos et al. 2008). Then, the clinical and experimental information previously reported supported the idea that infection by L. (V.) naiffi commonly results in benign manifestations. Therefore, the present study aimed to strengthen the awareness of the clinical and epidemiological importance of L. (V.) naiffi in the Amazon Region and to report therapeutic failure associated with this species.

Thirty Leishmania spp samples were isolated from cutaneous lesions of patients from different surrounding areas of Manaus. Samples were collected during 2011-2013 at the Heitor Vieira Dourado Tropical Medicine Foundation (FMT-HVD), a reference centre for tropical diseases in AM. Skin lesion fragments obtained by biopsy were cultured in Novy-Neal-Nicolle medium and Leishmania was isolated. Parasites were sent to the Leishmania Collection from the Oswaldo Cruz Institute for species identification. This study was approved by the Research Ethical Committee of FMT-HVD (protocol 21273572). Identification of Leishmaniaspp isolates was based on multilocus enzyme electrophoresis (MLEE) and performed on agarose gel and allelic variations were tested for the following enzymes: 6-phosphogluconate dehydrogenase (EC1.1.1.44), glucose-6-phosphate dehydrogenase (E.C.1.1.1.49) and isocitrate dehydrogenase (E.C.1.1.1.42). The method was performed in accordance with the conditions described by Cupolillo et al. (1994).

Four species were identified: L. (V.) guyanensis 66.7% (20/30),L. (V.) naiffi 26.7% (8/30), Leishmania (Leishmania) amazonensis 3.3% (1/30) and Leishmania (V.) shawi 3.3% (1/30). As expected, the most common species was L. (V.) guyanensis. Surprisingly, none of the isolates were characterised as L. (V.) brazi- liensis. According to Romero et al. (2002b), cases of ATL caused byL. (V.) braziliensis are uncommon in nearby Manaus. Interestingly, the frequency of L. (V.) naiffi was 26.7% (8/30), which indicates that this species could be an etiologic agent of CL more frequent than expected in this region. These data corroborate those obtained by Figueira et al. (2008), who observed a relevant number of CL associated withL. (V.) naiffi in the municipalities of Rio Preto da Eva and Manaus.

Several methods have been used to define Leishmania species (Degrave et al. 1994, Romero et al. 2001a, Coelho et al. 2011). Nevertheless, MLEE remains one of the main tools for characterisingLeishmania because it reveals polymorphisms that express phenotypes of population variations and taxonomically classify the different species of this parasite (Cupolillo et al. 1994). Precise identification of Leishmania spp is fundamental to understanding the disease’s epidemiology; improving the current knowledge concerning its pathology and the control measures (Coelho et al. 2011). Thus, it is likely that the transmission of L. (V.) naiffi in the Amazon Region is more frequent than has been reported. In this connection, the CL patients infected byL. (V.) naiffi (n = 8) are described below. All of them were men with a mean age of 37.4 ± 13.7 years (median = 37.5 years). The mean days of duration was 30 ± 24.8 days [median = 30 days (95% confidence interval: 22.5 - 52.5)]. The number of lesions ranged from one-four. Four subjects were initially treated with antimonial and four individuals were initially treated with pentamidine (antimonial: 10-20 mg Sb⁺⁵/kg/day for 20/30 consecutive days; pentamidine: 3 doses of 4 mg/kg with an interval of 72 h between doses) (Table II). Two of the patients experienced a poor response to antimonial or pentamidine therapy.

TABLE II
Demographic and clinical data of patients infected withLeishmania (Viannia) naiffi

Patient ID	Gender	Age	Lesions (n)	Location of lesion	Disease durations (days)	Presence of amastigotes	Treatment	Outcome
1	M	36	1	Upper limb	30	+	Antimonial (20 days) + antimonial (30 days)	Failure
2	M	39	1	Lower limb	90	+	Pentamidine (3 doses) + pentamidine (3 doses)	Failure
3	M	59	1	Upper limb	60	+	Antimonial (20 doses)	Cure
4	M	30	1	Upper limb	15	+	Antimonial (20 doses)	Cure
5	M	29	4	Upper limb	20	+	Pentamidine (3 doses)	Cure
6	M	39	2	Lower limb	30	+	Pentamidine (3 doses)	Cure
7	M	52	2	Face	30	+	Antimonial (20 doses)	Cure
8	M	15	1	Lower limb	30	+	Pentamidine (3 doses)	Cure

• M: male; +: positive smears of scarifications.

Previous studies have suggested that the Leishmania species and the endemic area examined can be influenced by the cure rate (Romero et al. 2001b, Arevalo et al. 2007). In a recent study conducted in AM, the cure rates in L. (V.) guyanensis infection after treatment with antimonial or pentamidine were 55.5% and 58.1%, respectively (Neves et al. 2011). To date, all of the studies described in the literature have associated infection by L. (V.) naiffi with spontaneous healing or a good therapeutic response. However, the data obtained by our group showed therapeutic failure in a patient who was infected by L. (V.) naiffi and treated with pentamidine (R Vieira-Gonçalves, unpublished observations). Corroborating this finding, we herein illustrate two new CL cases that are associated with L. (V.) naiffi infection and presented pentamidine or antimonial-resistant lesions (Table II), which indicates that the clinical evolution of infection from this species may not be as favourable as described. No cases of spontaneously healing was seem probably because the short period of disease duration until the diagnosis establishment. The cases of L. (V.) naiffireported herein underline that this species may not have a self-limiting nature, as previously described (Naiff et al. 1991, van der Snoek et al. 2009), and could not be as uncommon as referenced (Grimaldi Jr et al. 1991,Figueira et al. 2014). The present results highlight the importance of characterising Leishmania spp in areas that exhibit a sympatric circulation of Leishmania spp parasites to define the epidemiological importance of each of these species to human disease. This knowledge can improve the surveillance and therapeutic approaches used to achieve a clinical cure.

ACKNOWLEDGEMENTS

To the technicians of the Leishmaniasis Management of FMT-HVD, specially to Yolanda F Noguth and Maria Rita Teixeira, and to Sabrina S Guimarães, for clinical assistance.

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• The funders had no role in study design, data collection and analysis, decision to publish or preparation of the paper.
• Financial support: IOC/FIOCRUZ, PAPESIV/VPPDT/FIOCRUZ, FAPERJ-APQ1 (E-26/110.497/2011), CNPq (458858/2014-5) AMD-C and EC are CNPq and FAPERJ (CNE) research fellow.

• Erratum

  Vol. 110 (6): 797-800, 2015.
  p. 797
  Financial support: IOC/FIOCRUZ, PAPESIV/VPPDT/FIOCRUZ, FAPERJ-APQ1 (E-26/110.497/2011), CNPq (458858/2014-5)
  should read:
  Financial support: IOC/FIOCRUZ, PAPESIV/VPPDT/FIOCRUZ, FAPERJ-APQ1 (E-26/110.497/2011), CNPq (458858/2014-5), FAPEAM/CNPq/PPP-FAPEAM (010/2011), MCT/CNPq (014/2011)

Publication Dates

• Publication in this collection
  Sept 2015

History

• Received
  27 Mar 2015
• Accepted
  14 July 2015