Study on the zoonotic cycle of tegumentary leishmaniasis in an endemic area of a metropolitan region in the Northeastern region of Brazil

ABSTRACT

This study was conducted to characterize the transmission cycle of the tegumentary leishmaniasis (TL) in an old colonization area at Pernambuco State, Brazil. The aims were to identify autochthonous cases, sandflies fauna, domestic animals as possible reservoir hosts and the Leishmania species involved in this endemic area. A total of 168 suspected human cases of TL and 272 domestic animals (canine, feline, equine, goat, and sheep) were included. The sandflies were captured and identified by species. Patients were predominantly male and the average age was 37+18.1 years old. Of 85 patients who had skin lesions, 25.6% of them had direct positive smears for TL and 34 isolates were identified as Leishmania (Viannia) braziliensis. The confirmation for TL diagnosed by molecular detection (PCR) was almost three times more sensitive than the direct test [p < 0.001; PR = 2.72] associated with clinical examination. The Kappa test on PCR between two different specimens, biopsy, and skin lesion swab was 60.8% (p < 0.001). More than 200 specimens of sandflies (80 males and 159 females) were captured and identified as Lutzomyia whitmani (99.6%) and Lu. evandroi (0.4%). The detection of L. (V.) braziliensis by Real-Time PCR in the blood of a captured fed female was positive in 59.3% of Lu. whitmani. Of the 272 domestic animals included, 61.76% were male (n = 168). Thirty-six animals (13.2%) had lesions compatible with TL (34 dogs, 1 cat and 1 sheep) and 3 of them, all dogs, had lesions on the snout, showing destruction of cartilage and mucosa. The study suggests the participation of domestic animals as possible reservoirs. However, further studies are necessary to better understand the transmission cycle and take recommended measures in order to control the disease.

Transmission cycle; Eco-epidemiology; Tegumentary leishmaniasis; Leishmania (Viannia) braziliensis

INTRODUCTION

Tegumentary leishmaniasis (TL) is a zoonotic parasitic disease disseminated in the Old and New Worlds, endemic in 92 countries, with an estimate of one million new cases per year¹,². In Brazil, there were 15,484 new cases in 2019³. In the Americas, TL is transmitted through the bite of female phlebotomine sandflies of the Lutzomyia genus. The clinical forms of the disease are cutaneous (localized, disseminated, diffuse, recidivans cutis) and mucocutaneous, and in some cases it may evolve to spontaneous healing, except for the diffuse form¹,⁴.

Between 2007 and 2017, 235,301 cases were reported in all regions of Brazil and the North and Northeastern regions were the most affected with 101,332 and 72,395 cases, respectively⁵. In the Northeast, the predominance of cases occurs in locations with different agricultural plantations, some remnants of the Atlantic rainforest and secondary vegetation, whose aspects contribute to the spread of phlebotomines and the enzootic maintenance cycle with the involvement of wild and synanthropic reservoir hosts⁶. The modification of the natural environment caused by anthropic action leads to the adaptation of vectors and reservoirs with the zoonotic disease cycle in the peridomicile areas, affecting both humans and domestic animals⁷,⁸.

In Brazil, there are seven different predominant species of Leishmania, and Leishmania (Viannia) braziliensis is the predominant species. This species causes the most severe form of the disease, which can compromise the patient’s mucosa and cartilage. It is also the predominant etiologic agent of the disease in Pernambuco (PE)⁴,⁶,⁸,⁹.

The clinical diagnosis is based on anamnesis of the patient with clinical suspicion, epidemiological data (individual residing or transiting in an endemic area) and laboratory tests⁴,⁶. The gold standard method for the laboratory diagnosis of TL is the direct parasitological examination by microscopy on a slide with impression of the biopsy or scraping of the lesion⁶. Molecular tests, such as PCR and Real-Time PCR (qPCR), are very important in diagnosis because of their high sensitivity and specificity¹⁰.

The aim of this study was to characterize the zoonotic cycle through the description of several aspects of the eco-epidemiology of TL in an endemic area of the Northeast of Brazil. It also aimed to identify autochthonous cases, characterize phlebotomine fauna and domestic animals as possible involved reservoir hosts.

MATERIALS AND METHODS

A cross-sectional study was conducted in the Moreno municipality (Metropolitan Region of Recife, PE, Brazil), in the Atlantic Forest area, located at an altitude of 96 meters, with a rainy tropical climate and an average annual rainfall of 1,309.9 mm, and approximately 63,000 inhabitants, where 17% are in urban households¹¹. The majority of the studied region is covered by a monoculture of sugar cane or banana plantations.

The sample of autochthonous human cases included residents in endemic locations, who sought the primary health care provided by Programa Saude da Familia (PSF - Family Health Program) between 2015 and 2018. After the anamnesis, data were collected for clinical and epidemiological diagnosis, and clinical samples by biopsy or lesion scraping. The samples were analysed for diagnostic confirmation at the Laboratory of Immunoparasitology of Aggeu Magalhaes Institute/Fiocruz-PE.

Case suspect: individuals with compatible clinical signs, such as the presence of a cutaneous ulcer, with a granular background and infiltrated edges, from an endemic area or someone who had visited it.

Case definition: individuals with compatible clinical signs, epidemiological data and positive parasitological or molecular tests.

The parasitological diagnosis was performed by visualizing the parasite in the smear through an optical microscope⁶, and the lesion fragments or scrapings obtained were inoculated in culture medium and in hamsters (Mesocricetus auratus) for the in vivo isolation of Leishmania spp.⁸. The identification and characterization of the parasite was previously carried out through the analysis of the electrophoretic profile with isoenzymes (MLEE)⁸,¹², Leishmania Collection of the Oswaldo Cruz Institute/FIOCRUZ, Rio de Janeiro - CLIOC/RJ. The samples of domestic animals residing in Engenho Jardim and Engenho Cumaru (Moreno) included canines, felines, horses, goats, and sheep examined by a veterinarian from the Municipal Health Surveillance Center between 2017 and 2018. Blood and swab samples from ocular conjunctiva were collected and, for those who had active lesion, scarification, puncture, or swab of skin lesion was performed.

The sandflies were captured between January 2017 and December 2018 inside the houses where human or suspected cases of TL were notified. For these captures, the light traps model CDC (Centers for Disease Control) was installed in areas of banana plantations and in the remnants of the Atlantic Forest, from the twilight period onwards¹³. The specimens were separated by gender and morphological identification according to Young and Duncan¹⁴ and stored in ethanol 70% at room temperature until the DNA extraction.

Each entire female body specimen was submitted for molecular testing by qPCR to detect natural infection. For the molecular tests, DNA extraction and purification from human biological samples were performed using the commercial kit QIamp^® DNA mini kit (Qiagen, Valencia, USA) according to the protocol of the manufacturer. The extraction of DNA was performed individually according to Solano et al.¹⁵.

In order to confirm and verify the extraction efficiency, some random samples had DNA quantified on the NanoDrop spectrophotometer (Themo Scientific^®, model 2000/2000c, Waltham, Massachusetts, USA).

The DNA from human samples and from the reference strain CLIOC IOC/L0566 (MHOM/BR/75/M2903) of L. (V.) braziliensis was amplified by PCR, using oligonucleotides B1/B2 target of Leishmania kDNA of the Viannia subgenus, performed according to de Bruijn and Barker¹⁶. Visualization of the amplification products was done by electrophoresis in 1% agarose gel, stained with ethidium bromide¹⁷. Visualization of the DNA bands occurred in a photo documentation system for electrophoresis gel (L-PIX Touch–Loccus, Cotia, SP, Brazil).

For molecular diagnosis in domestic animals, DNA from blood samples, swab from lesion secretion and from ocular conjunctiva were amplified by qPCR, as well as for the phlebotomine. The kDNA fragments of the subgenus (Viannia) used as qPCR target had 138 base pairs and were amplified using a reference thermocycler (QuantiStudio 5 Applied Biosystems^®, Waltham, Massachusetts, USA), using Software 7500 (version 2.0.5, Applied Biosystems^®). The qPCR reactions followed the conditions of Cavalcanti et al.¹⁸.

Weather data on temperature, relative humidity and precipitation from the Pernambuco Water and Climate Agency were analyzed and compared with the monthly density of the collected sandflies.

Statistical analysis of frequencies, Kappa tests, Pearson’s and Yates’ chi-square tests and Fisher’s Exact tests were performed using the IBM SPSS Statistical Software (Version 20.0, IBM SPSS, Armonk, NY, USA). Comparison of proportions was calculated in MedCalc Statistical software (version 20.110 MedCalc Software, Ostend, Belgium), considering α = 5%. The risk of positivity (PR) was calculated using the prevalence ratio and respective 95% confidence intervals (CI). The calculation to analyze the leukocyte and plasma samples submitted to qPCR was performed according to Medronho et al.¹⁹.

The study in humans was approved by the Ethics Committee of University Fernando Pessoa (SSD-18072016), with written informed consent. The study on domestic animals was approved by the Ethics Committee on the Use of Animals of the Aggeu Magalhaes Institute/Oswaldo Cruz Foundation in Pernambuco (115/2017), with free and informed consent of the research by the tutors.

RESULTS

The aspect of the endemic region studied can be viewed in Figure 1, where a house can be seen among remains of the Atlantic Forest and sugarcane and banana plantations.

Figure 1
Panoramic view of Cumaru Mill in Moreno municipality.

168 human cases with suspected TL were included, aged between 5 and 74 years old (mean 37±18.1), with the majority (63.1%) being male. Of the suspected cases, 112 (66.7%) were confirmed to have TL, 48 (28.6%) did not have the disease and 8 (4.8%) had no laboratorial confirmation.

The clinical forms found were localized cutaneous (92.9%), recidivans cutis (3.6%), disseminated (2.7%), and one that evolved to scar (0.8%). Most patients had a single skin lesion (81.2%) and the remaining had more than ten (multiple) lesions, with diameters between 0.5×0.5 cm and 9.0×7.5 cm and evolution periods between 15 days and 10 months. As for the location of the lesion, the majority (75.9%) occurred in the lower and upper limbs, while the others were affected in different regions of the body. Lesions of the type ulcerated (75.6%) were presented as plaque, crusted, verrucous, erythematous, disseminated, recidivans cutis and mucocutaneous, while the others healed spontaneously. Two patients had coinfection (pyoderma gangrenosum and psoriasis). Some patients had other diseases, such as vascular disease, stasis ulcer, tuberculosis, impetigo, fungi, chronic eczema, and folliculitis. After the diagnosis of TL, all patients received specific treatment.

The results of the laboratorial tests are shown in Table 1. It was found that the molecular test by PCR identified 54.8% and the parasitological test by direct research was found in 25.6% of cases, respectively. The concordance of the PCR test between samples collected by tissue fragment (biopsy) and lesion swab was 60.8% (Kappa value; p < 0.001).

Table 1
Results of laboratory tests on human samples.

Laboratory tests (n = 168)	Positive result n (%)	Negative result n (%)	Not carried out or contaminated n (%)
Parasitological
- Direct search (smear slide)	43 (25.6)	42 (25.0)	83 (49.4)
- In vivo culture	20 (11.9)	23 (13.7)	125 (74.4)
- In vitro culture	14 (8.3)	5 (3.0)	149 (88.7)
Molecular
- Conventional PCR	92 (54.8)	55 (32.7)	21 (12.5)

The relation between gender, age, and skin color did not show any statistical differences in PR. Having complete or incomplete primary education is a protective factor for infection by L. (V.) braziliensis compared to illiterates (PR = 0.75; [CI 95%: 0.58-0.99]). The type of occupation seems to be associated with an increased risk of having the disease, but it did not reach statistical significance (Table 2).

Table 2
Characterization of the sample of cases with suspected TL according to the diagnosis and respective risk of positivity with a confidence interval of 95%.

Characteristics	TL diagnosis
	Positive n (%)	Negative n (%)	Value of p	PR** [CI: 95%]	Total (%)
Gender (n = 160)
Male	69 (61.6)	29 (60.4)	0.971	1.02 [0.82-1.25]	98 (61.2)
Female	43 (38.4)	19 (39.6)		1.0	62 (38.8)
Age group (n = 158)*
5 to 18	19 (17.0)	10 (21.8)		1.0	29 (18.4)
19 to 29	24 (21.4)	7 (15.2)	0.773	1.18 [0.85-1.64]	31 (19.6)
30 to 49	41 (36.6)	18 (39.1)		1.06 [0.78-1.45]	59 (37.3)
50 to 74	28 (25.0)	11 (23.9)		1.10 [0.79-1.52]	39 (24.7)
Skin color (n = 160)
White	16 (14.3)	8 (16.7)		1.0	24 (15.0)
Mixed ethnicity	84 (75)	33 (68.8)	0.694	1.27 [0.5-3.26]	117 (73.1)
Black	12 (10.7)	7 (14.6)		0.86 [0.24-3.02]	19 (11.9)
Level of education (n = 160)
Illiterate	27 (24.1)	8 (16.7)		1.0	35 (21.9)
Incomplete or complete elementary school	39 (34.8)	28 (58.3)	0.021	0.75 [0.58-0.99]	67 (41.9)
Completed high school or more	46 (41.1)	12 (25.0)		1.03 [0.82-1.29]	58 (36.2)
Occupation (n = 160)
Agricultural professions	34 (30.3)	18 (37.5)		1.08 [0.43-2.68]	52 (32.5)
Other professions (indoor or mixed environment)	57 (50.9)	18 (37.5)	0.293	1.81 [0.75-4.39]	75 (46.9)
No profession (children and students)	21 (18.8)	12 (25.0)		1.0	33 (20.6)

• *There was no information for 2 individuals; **PR – risk of positivity calculated as prevalence ratio.

Of all the 239 specimens (80 males and 159 females) of phlebotomine sandfly captured, almost all were identified as Lu. whitmani, and one female was Lu. evandroi.

During the study period, the presence of the sandfly species occurred predominantly in the months of August 2017 and July 2018. At first, the room temperature varied between 20 and 28 ºC, the relative humidity between 51 and 70.4% and the rainfall was 123.2 mm (with annual average of 1,619.8 mm). Temperature average in the capture of July 2018, varied between 22 ºC and 26 ºC, the relative humidity between 57 and 79% and the rainfall was 4.0 mm (with an annual average of 1,331.4 mm).

In the peridomicile, phlebotomine infestation was more common in chicken coops (92%) than in stables, but, in general, shelters for domestic animals were close to homes and with poor hygiene conditions.

The specimens collected were submitted to qPCR for confirmation of the infection by L. (V.) braziliensis. Among those classified as fed (with visible blood in the abdomen, n = 27), 59.3% (n = 16) were positive for the parasite. Among unfed females (no visible blood on the abdomen, n = 132), 33.3% were positive (n = 44) for L. (V.) braziliensis. The average concentration level of DNA of L. (V.) braziliensis among the fed female sandflies was 10.07+28.28 fg (ranging from 0 to 119.669 fg) and, in the non-fed, 2.79+11.63 fg (ranging from 0 to 119.669 fg). Among all captured females (visibly fed and not fed), the infection rate was 37.7%.

Of the 272 domestic animals included in the study, 212 were dogs (Canis familiaris), 21 were cats (Felis cattus), 33 were horses/donkeys (Equus caballus /Equus asinus), 5 were goats (Capra aegagrus hircus) and 1 was a sheep (Ovis aries). They had different ages, breeds, and sizes. More than 70% of all animals were medium or large and only 13.2% (34 dogs, 1 cat and 1 sheep) had skin lesions.

Of the 272 animals in the study, 255 blood samples and 192 ocular conjunctival swabs were collected and submitted to qPCR. The positivity of the skin lesion smear in qPCR from domestic animals is described in Table 3. It was found that 174 animals (64%) were positive for L. (V.) braziliensis, of which 52.6% were asymptomatic. Of the animals with a suspicious lesion, 5 dogs were positive for sporotrichosis and 31 were positive for TL. In the parallel analysis between the qPCR results and the presence or absence of a skin lesion, there was 87.3% concordance of the negative results and 12.6% of the positive results (Table 4).

Table 3
qPCR results for detection of Leishmania spp. in blood samples, conjunctival swab or lesion swab from domestic animals residing in the study regions.

Domestic animal	Biological sample	Positive Result n (%)	Not detected n (%)	Not collected n (%)	Total
Dog	Blood	141 (66.5)	63 (29.7)	8 (3.8)	212
	Swab	38 (17.9)	115 (54.3)	59 (27.8)
Cat	Blood	11 (52.4)	1 (4.8)	9 (42.8)	21
	Swab	5 (23.8)	8 (38.1)	8 (38.1)
Horse Donkey	Blood	19 (57.6)	14 (42.4)	0 (0.0)	33
	Swab	5 (15.1)	16 (48.5)	12 (36.4)
Goat	Blood	2 (40)	3 (60)	0 (0.0)	5
	Swab	1 (20)	3 (60)	1 (20)
Sheep	Blood	1 (100)	0 (0.0)	0 (0.0)	1
	Swab	0 (0.0)	1 (100)	0 (0.0)
Total	Blood	174 (64)	81 (29.8)	17 (6.2)	272
	Swab	49 (18)	143 (52.6)	80 (29.4)

Table 4
qPCR result as a function of the existence, or not, of a skin lesion compatible with TL.

qPCR (parallel analysis)*	Skin lesion
	Yes n (%)	No n (%)	Total n (%)
Positive	24 (12.6)	167 (87.4)	191 (100)
Negative	9 (12.7)	62 (87.3)	70 (100)
Total	33 (12.6)	229 (87.4)	262** (100)

• *Parallel analysis means that every type of biological sample was analyzed in parallel: when all qPCR results were negative, the overall “qPCR” result was negative; when at least one qPCR result in any biological sample was positive, the overall “qPCR” result was positive; **it was not possible to obtain qPCR results of 10 animals.

The comparison of positivity between the different biological samples collected from domestic animals showed greater positivity in the swab from the skin lesion (85.7%) than in whole blood (73.3%) (Table 5).

Table 5
Comparison of positivity between different biological samples of domestic animals.

qPCR - Type of biological sample	Value of p	Chi-squared	Difference	Confidence interval 95%	Positivity in qPCR
					Whole blood* (leukocytes + plasma) = 73.3%
Whole blood vs. conjunctival swab	<0.001	98.40	47.5%	38.71-55.12	Conjunctival Swab = 25.8%
Whole blood vs. skin lesion swab	0.464	0.54	12.4%	-24.97-25.46	skin lesion swab 85.7%

• *Parallel analysis means that every type of biological sample was analyzed in parallel: when all qPCR results were negative, the overall “qPCR” result was negative; when at least one qPCR result in any biological sample was positive, the overall “qPCR” result was positive.

DISCUSSION

According to the Health Surveillance Secretary of the Ministry of Health²⁰, the Pernambuco State recorded an average of 38 cases of TL annually. In 2015, there was a doubling of the number of cases in the Moreno municipality, which on average had 50 cases per year (unpublished data). The municipality is considered a low-risk area for TL in terms of infection: from 2018 to 2020, 35 cases were confirmed in the region. It had an average incidence of 11.67/year (between 2018 and 2020), with a ranging from 8 (in 2020) cases to 16 (in 2018). Despite the low-risk area for TL, Moreno is among the seven municipalities with the highest incidence of annual cases in the Pernambuco State²⁰.

In the present study, the individuals were mostly male, of an economically active age, with a pattern of populations vulnerable to infection by Leishmania spp. in rural areas²¹. The disease affects different ages, including children⁹,²²,²³. A previous study in the same population found a positivity of 22.4% with a positive Montenegro skin test for previous infection²⁴. Twenty years later, this study found a 66.7% positivity for TL infection (unpublished data).

Regarding the characteristics of the disease, several clinical forms of TL were identified, with a predominance of the simple ulcerated form, which is corroborated by other studies⁴,²¹. In Brazil, 90% of reported cases present the cutaneous form, whereas the mucocutaneous form varies from 3 to 6% of the cases, depending on the region²³,²⁵. Clinical manifestations are variable and dependent on the associated Leishmania species and the host’s immune response¹,⁶. Patients from the locations of this study presented co-infection with some bacteria species, pyoderma gangrenosum, psoriasis and other diseases, such as vascular diseases, stasis ulcer, impetigo, chronic eczema, folliculitis, STDs, tuberculosis — some with dermatological manifestations like TL. This makes the diagnosis of TL difficult⁶, hence the need for specific and sensitive diagnostic tests for TL²⁵.

Direct examination results were higher than those from the Leishmania isolates obtained in culture²⁶. The isolates obtained from hamsters (Mesocricetus auratus) had a higher positivity than the isolates from culture medium, but without statistical significance. The Leishmania species circulating in the region was identified in previous studies as L. (V.) braziliensis⁴,⁸,²⁷ and confirmed by specific qPCR tests. L. (V.) braziliensis is the predominant species throughout the country and is present in several municipalities in Pernambuco State⁴,⁸,⁹,²⁸.

The molecular tests used have high sensitivity and specificity, in addition to being fast and effective¹⁶,¹⁷,²⁹. PCR has detection sensitivity up to 1 fentogram (10-15g) of Leishmania DNA, which corresponds to 1/10 of the parasite¹⁶. This method has been standardized in tissue samples (invasive methods) to identify Leishmania species in subclinical cases of the disease, or with low parasite load, following up on treatment and distinction between active and past infections²⁹.

It was demonstrated that PCR is better to define the patient diagnosis of TL in all types of the analyzed biological samples. Therefore, the combination of minimally invasive sample collection procedures (such as the swab of lesion, salivary fluid, and blood) with molecular PCR test for diagnostic confirmation of TL is very important²⁵,²⁹-³¹. Especially when compared with classic parasitological tests with lower positivity¹⁰,²⁶,³² and need for invasive collection (lesion puncture and biopsy).

The concordance in PCR between the tissue fragments and swab samples from the lesion had a very significant index according to Cohen’s Kappa³¹, which demonstrates the effectiveness of the swab collection technique in relation to biopsy³¹. The divergence of results can be attributed to the low number of parasites in the lesion or in the exudate, as the parasites are not evenly distributed in the lesion, which can lead to false negatives³². Amplification of L. (V.) braziliensis DNA by PCR in blood samples from patients was found to be low compared to the tissue biopsy samples, due to the scarcity of circulating parasites in this biological sample³³.

The sandflies species most involved in the transmission in Brazil are Lu. whitmani, Lu. intermedia, Lu. umbratilis, Lu. wellcomei, Lu. flaviscutellata and Lu. migonei³⁴. In the present results, the predominance of Lu. whitmani was observed as the main species and probable vector in this region. Captured species were done in chicken coops and stables located in the peridomicile. The positivity to L. (V.) braziliensis in these sandflies measured by qPCR in the studied endemic region was very high. These results strongly indicate the natural infection by L. (V.) braziliensis in Lu. migonei. Among fed phlebotomine, positivity was 59.3% and in those without visible blood (not fed), 33.3%.

The low number of captured phlebotomine may be due to the use of herbicides and fire, as mentioned by Nasser and Will³⁵. These factors cause changes in the feeding habits when adapting to the environment modified by anthropic action. The climate changes also interfere with the biological behaviour of the fauna and, consequently, with their feeding role, modifying the frequency of blood meals³⁶.

According to Brito et al.²⁴, the expansion of agriculture and construction of houses close to the forests favors the risk of infection due the presence of domestic and synanthropic animals naturally infected close to the houses. On the other hand, the adaptation of phlebotomine to the modified environment increases transmissibility, as well as the expansion of the zoonotic cycle of TL through the contact between these hosts and the human population more frequently.

Lu. whitmani is the most important species in the transmission of TL in Brazil. Our results showed positivity for L. (V.) braziliensis in specimens collected from Lu. whitmani. An entomological survey in 2001 collected 444 specimens of Lu. whitmani in this municipality, also showing the predominance of this species in eleven other locations in the municipality (unpublished data). Between 2011 and 2012, another study confirmed the presence of the species in the intradomicile and peridomicile, with 3,071 specimens collected, of which 2,919 Lu. whitmani, 122 Lu. evandroi and 30 Lu. choti. Of the 37 sandflies species identified in Pernambuco, Lu. whitmani was the most predominant³⁷.

The domestic animals examined in this study showed a high positivity rate for TL through qPCR, especially in blood samples, which confirms the high prevalence of infection in the region, even with some asymptomatic animals. Dogs were the species with highest percentage of positivity in the blood. As they transit through more regions and different areas of the peridomicile, they may have a greater chance of being bitten. Equidae are also parasitized by Leishmania spp. Several studies have already confirmed the presence of L. (V.) braziliensis in these domestic animals residing in endemic areas⁹,³⁸,³⁹.

A sheep with a lesion like TL (uncommon symptomatic condition) was also identified, confirmed by direct examination for TL. A similar case occurred in South Africa, where a sheep presented a lesion that evolved to spontaneous healing⁷. According to Dantas-Torres⁴⁰, the diagnosis of asymptomatic domestic animals is important, as they may be reservoirs of Leishmania spp.

The knowledge of eco-epidemiology of the studied region, transmission control through early diagnosis and treatment are measures which collaborate with prevalence and incidence’s decrease of leishmaniasis, according to goals established by PAHO/WHO for 2030³. Also, the main measures of control for this disease, according to the Brazilian Ministry of Health are: avoiding living close to remnants of forest, netting on doors and windows, and applying repellent on the body⁶.

CONCLUSION

In conclusion, our findings demonstrate that the epidemiology of TL associated with L. (V.) braziliensis in this municipality of the Zona da Mata region has different characteristics, when compared with previous studies in the same region over time⁴,²⁵, to those observed in other regions of Brazil. In this sense, more studies are necessary, especially on the role of domestic animals, to support their real contribution to the disease transmission cycle and maintenance, and also to contribute to the adoption of TL control recommendations in endemic areas. It is also recommended that, within the scope of the Primary Health Care program and epidemiological surveillance, community health agents promote campaigns aimed to improve the spread of information on the transmission cycle, signs, and symptoms of disease and its care to the human populations living in the endemic areas of the country.

REFERENCES

• ¹
  Burza S, Croft SL, Boelaert M. Leishmaniasis. Lancet. 2018;392:951-70.
• ²
  World Health Organization. Leishmaniasis: status of endemicity of cutaneous leishmaniasis: 2020. [cited 2022 Aug 16]. Available from: https://apps.who.int/neglected_diseases/ntddata/leishmaniasis/leishmaniasis.html
  » https://apps.who.int/neglected_diseases/ntddata/leishmaniasis/leishmaniasis.html
• ³
  Pan American Health Organization. Leishmaniases: epidemiological report in the Americas. [cited 2022 Aug 16]. Available from: https://iris.paho.org/handle/10665.2/51742
  » https://iris.paho.org/handle/10665.2/51742
• ⁴
  Brito ME, Andrade MS, Dantas-Torres F, Rodrigues EH, Cavalcanti MP, Almeida AM, et al. Cutaneous leishmaniasis in northeastern Brazil: a critical appraisal of studies conducted in State of Pernambuco. Rev Soc Bras Med Trop. 2012;45: 425-9.
• ⁵
  Melo MG, Morais RC, Goes TC, Silva RP, Morais RF, Guerra JA, et al. Clinical and epidemiological profiles of patients with American cutaneous leishmaniasis from the states of Pernambuco and Amazonas, Brazil. Rev Soc Bras Med Trop. 2020;53:e20200083.
• ⁶
  Brasil. Ministério da Saúde. Departamento de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar. Brasília: Ministério da Saúde; 2017. [cited 2022 Aug 16] Available from: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
  » http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
• ⁷
  van der Lugt JJ, Carlyon JF, de Waal DT. Cutaneous leishmaniasis in a sheep. J S Afr Vet Assoc. 1992;63:74-7.
• ⁸
  Brito ME, Andrade MS, Mendonça MG, Silva CJ, Almeida EL, Lima BS, et al. Species diversity of Leishmania (Viannia) parasites circulating in an endemic area for cutaneous leishmaniasis located in the Atlantic rainforest region of northeastern Brazil. Trop Med Int Heal. 2009;14:1278-86.
• ⁹
  Brandão-Filho SP, Brito ME, Carvalho FG, Ishikaw EA, Cupolillo E, Floeter-Winter L, et al. Wild and synanthropic hosts of Leishmania (Viannia) braziliensis in the endemic cutaneous leishmaniasis locality of Amaraji, Pernambuco State, Brazil. Trans R Soc Trop Med Hyg. 2003;97:291-6.
• ¹⁰
  Pena HP, Belo VS, Xavier-Junior JC, Teixeira-Neto RG, Melo SN, Pereira DA, et al. Accuracy of diagnostic tests for American tegumentary leishmaniasis: a systematic literature review with meta-analyses. Trop Med Int Health. 2020;25:1168-81.
• ¹¹
  Instituto Brasileiro de Geografia e Estatística. Brasil/Pernambuco/Moreno. [cited 2022 Aug 16]. Available from: https://cidades.ibge.gov.br/brasil/pe/moreno/panorama
  » https://cidades.ibge.gov.br/brasil/pe/moreno/panorama
• ¹²
  Cupolillo E, Grimaldi G Jr, Momen H. A general classification of New World leishmania using numerical zymotaxonomy. Am J Trop Med Hyg. 1994;50:296-311.
• ¹³
  McNelly J. The CDC trap as a special monitoring tool. In: Proceedings of the Seventy-Sixth Annual Meeting of the New Jersey Mosquito Control Association. 1989:26-33. [cited 2022 Aug 16]. Available from: https://vectorbio.rutgers.edu/outreach/cdctrap.htm
  » https://vectorbio.rutgers.edu/outreach/cdctrap.htm
• ¹⁴
  Young DG, Duncan MA. Guide to the identification and geographic distribution of Lutzomyia sand flies in Mexico, the West Indies, Central and South America (Diptera: Psychodidae). Gainesville: Associated Publishers; 1994.
• ¹⁵
  Solano P, Duvallet G, Dumas V, Cuisance D, Cuny G. Microsatellite markers for genetic population studies in Glossina palpalis (Diptera: Glossinidae). Acta Trop. 1997;65:175-80.
• ¹⁶
  de Bruijn MH, Barker DC. Diagnosis of New World leishmaniasis: specific detection of species of the Leishmania braziliensis complex by amplification of kinetoplast DNA. Acta Trop. 1992;52:45-58.
• ¹⁷
  Green MR, Sambrook J. Molecular cloning: a laboratory manual. 4^th ed. New York: Cold Spring Harbor; 2012.
• ¹⁸
  Cavalcanti M,P Dantas-Torres F, Albuquerque SC, Morais R, Brito MC, Otranto D, et al. Quantitative real time PCR assays for the detection of Leishmania (Viannia) braziliensis in animals and humans. Mol Cell Probes. 2013;27:122-8.
• ¹⁹
  Medronho RA, Carvalho DM, Bloch KV, Luiz RR, Werneck GL, editores. Epidemiologia. São Paulo: Atheneu; 2002.
• ²⁰
  Brasil. Ministério da Saúde. DATASUS. Leishmaniose tegumentar americana: casos confirmados notificados no Sistema de Informação de Agravos de Notificação - Pernambuco. [cited 2022 Aug 16] Available from: http://tabnet.datasus.gov.br/cgi/tabcgi.exe?sinannet/cnv/ltape.def
  » http://tabnet.datasus.gov.br/cgi/tabcgi.exe?sinannet/cnv/ltape.def
• ²¹
  Pezente L, Benedetti M. Perfil epidemiológico da leishmaniose Tegumentar Americana no Estado de Roraima, Amazônia, Brasil, entre 2007 e 2016. Braz J Health Rev. 2019;2:1734-42.
• ²²
  Vasconcelos JM, Gomes CG, Sousa A, Teixeira AB, Lima JM. Leishmaniose tegumentar americana: perfil epidemiológico, diagnóstico e tratamento. Rev Bras Anal Clin. 2018;50:221-7.
• ²³
  World Health Organization. Working to overcome the global impact of neglected tropical diseases: first WHO report on neglected tropical diseases. [cited 2022 Aug 16] Available from: https://apps.who.int/iris/handle/10665/44440
  » https://apps.who.int/iris/handle/10665/44440
• ²⁴
  Brito ME, Silva CJ, Silva CM, Salazar PR, Coutinho JS, Reis LC, et al. Clinical epidemiological profile of American tegumentary leishmaniasis at the Pinto Sugar Mill in Moreno Municipality, Greater Metropolitan Recife, Pernambuco State, Brazil. Cad Saude Publica. 2008;24:2445-8.
• ²⁵
  Thakur S, Joshi J, Kaur S. Leishmaniasis diagnosis: an update on the use of parasitological, immunological and molecular methods. J Parasit Dis. 2020;44:253-72.
• ²⁶
  Corvalan FH, Sampaio RN, Brustoloni YM, Andreotti R, Lima-Junior MS. DNA identification of Leishmania (Viannia) braziliensis in human saliva from a patient with American cutaneous leishmaniasis. J Venom Anim Toxins Incl Trop Dis. 2011;17:98-102.
• ²⁷
  Moreira MA, Luvizotto MC, Garcia JF, Corbett CE, Laurenti MD. Comparison of parasitological, immunological and molecular methods for the diagnosis of leishmaniasis in dogs with different clinical signs. Vet Parasitol. 2007;145:245-52.
• ²⁸
  Rodrigues EH, Brito ME, Mendonça MG, Werkhäuser RP, Coutinho EM, Souza WV, et al. Evaluation of PCR for diagnosis of American cutaneous leishmaniasis in an area of endemicity in northeastern Brazil. Brazil J Clin Microbiol. 2002;40:3572-6.
• ²⁹
  Martins L, Alexandrino A, Guimarães G. Detection of Leishmania braziliensis DNA in American tegumentary leishmaniasis patients. Rev Saude Publica. 2010;44:571-4.
• ³⁰
  Brito ME, Almeida EL, Medeiros AC, Werkhäuser RP, Alexandre JL, Sá BS, et al. Leishmania (Viannia) braziliensis isolated from the saliva of patients in a cutaneous leishmaniasis-endemic area of northeastern Brazil. Mem Inst Oswaldo Cruz. 2018;113:e170250.
• ³¹
  Adams ER, Gomez MA, Scheske L, Rios R, Marquez R, Cossio A, et al. Sensitive diagnosis of cutaneous leishmaniasis by lesion swab sampling coupled to qPCR. Parasitology. 2014;141:1891-7.
• ³²
  Landis JR, Koch GG. The measurement of observer agreement for categorical data. Biometrics. 1977;33:159-74.
• ³³
  Shahbazi F, Shahabi S, Kazemi B, Mohebali M, Abadi AR, Zare Z. Evaluation of PCR assay in diagnosis and identification of cutaneous leishmaniasis: a comparison with the parasitological methods. Parasitol Res. 2008;103:1159-62.
• ³⁴
  Abbate JM, Maia C, Pereira A, Arfuso F, Gaglio G, Rizzo M, et al. Identification of trypanosomatids and blood feeding preferences of phlebotomine sand fly species common in Sicily, Southern Italy. PLoS One 2020;15:e0229536.
• ³⁵
  Nasser N, Will E. Perfil epidemiológico da Leishmaniose Tegumentar Americana no município de Blumenau-SC. Arq Catarin Med. 2017;46:28-38.
• ³⁶
  Silva CJ, Pereira SV, Apolinário EJ, Santos GL, Melo-Santos MA, Silva AF, et al. Culicidae fauna (Diptera: Culicidae) survey in urban, ecotonal and forested areas, from the Moreno municipality - Pernambuco State, Brazil. Rev Soc Bras Med Trop. 2018;51:523-7.
• ³⁷
  Sales KG, Costa PL, Morais RC, Otranto D, Brandão-Filho SP, Cavalcanti MP, et al. Identification of phlebotomine sand fly blood meals by real-time PCR. Parasit Vectors. 2015;8:230.
• ³⁸
  Guimarães VC, Costa PL, Silva FJ, Silva KT, Silva KG, Araújo AI, et al. Phebotomine sandflies (Diptera: Psychodidae) in São Vicente Férrer, a sympatric area curtaneous and visceral leishmaniasis in the state of Pernambuco, Brazil. Rev Soc Bras Med Trop. 2012;45:66-70.
• ³⁹
  Vieira AF, Alvarenga TF, Oliveira SF, Coelho EC, Teixeira MC, Moscon LA, et al. Leishmaniose em equídeos: revisão. Pubvet. 2020;14:1-5.
• ⁴⁰
  Dantas-Torres F, Simões-Mattos L, Brito FL, Figueredo LA, Faustino MA. Leishmaniose felina: revisão de literatura. Clin Vet. 2006;11:32-40.

Publication Dates

• Publication in this collection
  30 Sept 2022
• Date of issue
  2022

History

• Received
  11 Feb 2022
• Accepted
  26 July 2022