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Sand fly fauna (Diptera: Psychodidae) from the Goytacazes National Forest and surrounding areas of southeastern Brazil Thieres Marassati das Virgens1, Helder Ricas Rezende2, Israel de Souza Pinto3*, and Aloísio Falqueto4 Instituto Federal de Educação, Ciência e Tecnologia do Espírito Santo, Campus Linhares, Av. Filogônio Peixoto S/N, Bairro Aviso, CEP 29901-291, Linhares, Espírito Santo, Brazil 2 Núcleo de Entomologia e Malacologia do Espírito Santo, Secretaria de Estado da Saúde, Rua Pedro Zangrande 381, Bairro Jardim Limoeiro, CEP 29164-020, Serra, Espírito Santo, Brazil 3 Laboratório de Biologia Molecular de Insetos, Instituto Oswaldo Cruz, FIOCRUZ, Av. Brasil 4365, Manguinhos, CEP 21040-360, Rio de Janeiro, Rio de Janeiro, Brazil, [email protected] 4 Unidade de Medicina Tropical, Departamento de Patologia, Universidade Federal do Espírito Santo, Av. Marechal Campos 1468, Maruípe, CEP 29043-900, Vitória, Espírito Santo, Brazil 1
Received 9 July 2014; Accepted 20 November 2014 ABSTRACT: Most studies of the sand fly fauna in southeastern Brazil are conducted in the peridomiciliary environment of leishmaniasis endemic regions. Therefore, to increase the knowledge about diversity and richness of sand fly conservation areas, we describe here the sand fly fauna from the National Forest of Goytacazes (NFG), state of Espírito Santo, Brazil, and its surroundings areas. We also used sand fly fauna records from eight conservations units within the state of Espírito Santo to understand the similarity and relationships among them. The sand flies were simultaneously collected from June, 2008 to May, 2009 in two different environments: a preserved environment represented by the NFG and a modified environment represented by a peridomicile. To establish the similarity among the conservation units, we used a method very similar to parsimony analysis of endemism. We collected 2,466 sand fly specimens belonging to 13 species. Pressatia choti and Nyssomyia intermedia were the most abundant sand fly species. Ny. intermedia is a known vector of Leishmania braziliensis and epidemiological surveillance must be conducted in the area. We discuss aspects regarding the diversity of sand flies as well as the risk of transmission of Leishmania parasites in the area. We also provide for the first time a hypothesis of similarity relationships among conservation units within the state of Espírito Santo. Journal of Vector Ecology 40 (1): 28-35. 2015. Keyword Index: Phlebotominae, biodiversity, Atlantic forest, leishmaniasis, parsimony, endemism.
INTRODUCTION Despite its degradation and deforestation, the Atlantic Forest still remains one of the hotspots of vertebrate biodiversity in the world (Myers et al. 2000, Orme et al. 2005). Since vertebrates are the food source for hematophagous insects, we can also predict that the Atlantic forest biome has high species richness of insects such as fleas, mosquitoes, and phlebotomine sand flies. The phlebotomine sand flies (Diptera: Psychodidae) are the main leishmaniasis vectors along the Brazilian states situated on the Atlantic coast, such as the state of Espírito Santo. Historically, almost all the studies of sand fly fauna in this state were conducted in disturbed areas of endemic leishmaniasis regions as a result of the medical and veterinary importance of these flies (Barros et al. 1985, Ferreira et al. 2001, Pinto et al. 2010a, Pinto et al. 2012a). Only recently, some studies were conducted in conservation units next to leishmaniasis endemic (Pinto et al. 2012b) or nonendemic (Pinto et al. 2012b) areas. These last studies highlighted the importance of sand fly surveys in preserved environments, which present higher species richness than in disturbed areas. In Espírito Santo, there have been only two surveys on sand fly fauna that were simultaneously conducted in a conservation unit and in a surrounding area to compare sand fly fauna between the two different environments, also recording higher species richness
in the preserved environment (Virgens et al. 2008, Ferreira et al. 2013). Nevertheless, some of the conservation units situated in Espírito Santo still lack an inventory of sand fly fauna. Furthermore, there is also no information about the similarity among conservation units within the state of Espírito Santo based on sand fly species records. Therefore, our aim was to describe, for the first time, the sand fly fauna from the National Forest of Goytacazes and surrounding areas of southeastern Brazil. In addition, we used sand fly fauna records from conservations units of the state of Espírito Santo to understand the similarity and the relationships among them. MATERIALS AND METHODS Study area The study was carried out simultaneously in two different environments: 1) a preserved environment represented by the National Forest of Goytacazes (NFG), and 2) a modified environment represented by a peridomicile with some domestic animals, such as dogs, chickens, and pigs. The NFG (19º 28’ 01” S; 40º 04’ 18” W; elevation of 10 m [a.s.l.]) is a federal conservation unit situated in the municipality of Linhares, state of Espírito Santo, southeastern Brazil (Figure
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Figure 1. Collection sites (square: peridomiciliary environment; circle: forest environment represented of the National Forest of Goytacazes) in the Atlantic Forest, state of Espírito Santo, southeastern Brazil. 1). Currently, the NFG covers an area of 1,424 hectares and is a remnant of the Atlantic Forest on the Doce River alluvial floodplain (Rolim et al. 2006). Almost all the NFG area is covered by a secondary forest, but the vegetation is dense with trees as tall as 30 m that present moderate overlapping canopies. The KöppenGeiger climate classification for the region is tropical monsoon (Peel et al. 2007). The average temperatures range from 11 to 18° C in the colder months (June to August) and from 30 to 34° C in the warmer months (December to March), with annual rainfall of approximately 1,200 mm (Hijmans et al. 2005). Sand fly captures The sand flies were collected simultaneously in the modified and preserved environments from June, 2008 to May, 2009. For each night and environment, we used one illuminated Shannon trap and two CDC (Center for Disease Control) automatic light traps. For each environment, one of us collected sand flies with an aspirator during the first three h of the night (18:00 to 21:00) in the illuminated Shannon trap, while one automatic light trap was installed up to one m above ground level and the other in the tree canopy (at ~10 m height). The automatic light traps worked during the night (18:00 to 21:00). We spent 36 collection hours for each trap and a total of 108 collection hours for each environment.
The sand flies collected were stored in 70% ethanol and mounted on glass slides using the technique of Barretto and Coutinho (1940). We used the classification of Galati (1995, 2003) and the abbreviation of generic names of Marcondes (2007). We identified all the species according to morphological characters with the taxonomic key of Galati (2003). The sample material was deposited in the entomology collection of the Department of Tropical Medicine at the Federal University in Espírito Santo. Climate data (rainfall, relative humidity, and mean temperature) were obtained from a station of the National Institute of Meteorology situated at the Espírito Santo state Institute for Research, Technical Assistance, and Rural Extension (INCAPER), municipality of Linhares, which borders the NFG. Statistical analysis We evaluated the specific richness (S), equitability (J), and Shannon’s diversity index for each environment, kind of trap, and vertical stratum. These analyses were performed in PAST (Hammer et al. 2001). We used the Mann-Whitney test to verify differences between the numbers of sand fly specimens collected by the CDC automatic trap on the ground level and on the tree canopy for the two environmental samplings. We used the Spearman non-
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parametric test to evaluate the correlation between climate data and the most abundant species for each environment and vertical stratum. We considered the differences to be significant at p < 0.05. These analyses were performed in Biostat 5.0.
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Similarity analysis To establish the similarity among conservation units within the state of Espírito Santo, we used eight conservation areas as Operational Geographic Units (OGUs) (Crovello 1981) and sand fly species records were used as characters (Table 1). The sand fly distribution data for the conservation units were compiled from Virgens et al. (2008), Pinto et al. (2010 a,b), Pinto et al. (2012 a,b), Ferreira et al. (2013), and Núcleo de Entomologia da Secretaria do Estado da Saúde do Espírito Santo (unpublished data). The conservation units can be grouped into three kinds using the elevation as a classification criterion: 1) highland conservation units (830 m a.s.l.]) - Lombardia Biological Station and Caparaó National Park; 2) intermediate (500 m a.s.l. < elevation < 750 m a.s.l.) - Duas Bocas Biological Reserve and Santa Lúcia Biological Station; and 3) lowlands conservation units (< 400 m a.s.l.) - National Forest of Goytacazes, National Forest of Rio Preto, Natural Monument of Pontões Capixabas, and Sooretama Biological Reserve. We coded the absence of a species as a primitive character (0) and its presence as a derived character (1) of an OGU. For the tree rooting, we used a hypothetical locality having no taxa, a procedure known as Lundberg rooting following Rosen (1988) and Costa et al. (2000). Fifty-two species occurrences were used as characters. All characters were treated as equally weighted and unordered in the similarity analyses. The data matrix (Table 1) was built in Mesquite (Maddison and Maddison 2006). The data were analyzed using the maximum parsimony optimality criterion from PAUP* 4.0b10 (Sinauer, Sunderland, MA) and the interface PaupUp 1.0.3.1 for Microsoft Windows 95/NT. We conducted a heuristic search using tree-bisection-reconnection as a branch swapping algorithm with a starting tree obtained through stepwise addition held with one tree at each step and computed a strict consensus of the most parsimonious trees. All the trees were inspected using FigTree v1.3.1.
overall male/female sex ratio was 0.88 and, for captures using the Shannon modified trap and the CDC traps, the ratios were 0.78 and 1.33, respectively. The number and sex of the species collected with the Shannon trap and the CDC trap for both environments are presented in Tables 2 and 3, respectively. Pressatia choti was the most abundant sand fly species collected with 720 specimens (29.2%), followed by Ny. intermedia, 716 specimens (29.0%). Pr. choti was also the most abundant species, followed by Ny. intermedia for each stratum analyzed for the two environments sampled (Table 3). Therefore, only these two species were used in the statistical analyses. The number of Ny. intermedia collected in the peridomiciliary environment was significantly higher than in the forest environment. There is no significant difference between the number of collected specimens belonging to Pr. choti and Ny. intermedia between the ground level and the tree canopy level for each environment. In the forest environment, Pr. choti was negatively correlated with monthly rainfall (R = -0.76, p = 0.00) and Ny. intermedia was negatively correlated with monthly rainfall (R = -0.70, p = 0.01) and monthly average temperature (R = -0.76, p = 0.00). In the peridomicile, Pr. choti was negatively correlated with monthly rainfall (R = -0.73, p = 0.00) and Ny. intermedia was negatively correlated with monthly average temperature (R = -0.65, p = 0.02). Regarding the similarity analysis, we found two most parsimonious trees (Figure 2), 92 steps-long each. Thirty-six characters were parsimony-informative and the overall Consistency Index (excluding uninformative characters) was 0.4737, and the Retention Index was 0.4030. The two highland conservation units (Lombardia Biological Station and Caparaó National Park) composed a clade in a tree. In the other tree, the Lombardia Biological Station is a sister group of the Caparaó National Park. The two conservation units with intermediate elevation (Duas Bocas Biological Reserve and Santa Lúcia Biological Station) composed another clade. The lowlands conservation units were paraphyletic, since the Sooretama Biological Reserve was a sister group of the clade composed of the Duas Bocas Biological Reserve and the Santa Lúcia Biological Station.
RESULTS
DISCUSSION
We collected 2,466 sand fly specimens belonging to 13 species, nine genera, and four sub-tribes: Brumptomyiina - Brumptomyia cunhai (Mangabeira 1942); Lutzomyiina - Evandromyia callipyga (Martins and Silva 1965), Evandromyia costalimai (Mangabeira 1942), Evandromyia sallesi (Galvão and Coutinho 1939), Migonemyia migonei (França 1920), Pintomyia fischeri (Pinto 1926), Pressatia choti (Floch and Abonnenc 1941) and Pressatia equatorialis (Mangabeira 1942); Psychodopygiina - Nyssomyia intermedia (Lutz and Neiva 1912), Psathyromyia pascalei (Coutinho and Barreto 1941), Psathyromyia pelloni (Sherlock and Alencar 1959) and Psychodopygus hirsutus (Mangabeira 1942); and Sergentomyiina - Micropygomyia schreiberi (Martins, Falcão and Silva 1975). The number of sand flies collected was higher in the peridomiciliary environment (n = 2,001) than in the forest environment. However, the species diversity and richness were higher in the forest than in the peridomiciliary environment. The
The sand fly fauna recorded in the National Forest of Goytacazes was similar to that of forested regions of northern Espírito Santo, with an abundance of Pr. choti (Virgens et al. 2008, Pinto et al. 2012b). Considering only the species from the modified environment sampled near the NFG, Ny. intermedia and Mg. migonei were also abundant. This seems to be a pattern along the state of Espírito Santo, as pointed out by different works conducted in modified environments (Barros et al. 1985, Falqueto et al. 1991, Ferreira et al. 2001, Virgens et al. 2008, Pinto et al. 2010a,b). These last two species are incriminated in the transmission of Leishmania braziliensis Vianna, 1911 in endemic areas of cutaneous leishmaniasis along the Brazilian Atlantic coast (Gomes et al. 1983, Rangel et al. 1984, Teodoro et al. 1993, PitaPereira et al. 2005). Therefore, despite the absence of leishmaniasis records close to the NFG to date, it is important to maintain epidemiological surveillance in the area. The number of Mg. migonei specimens collected was lower in
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Table 1. Data matrix of 52 sand fly species used as characters for eight Operational Geographical Units (OGUs) to perform the similarity analysis. The letters represent the conservation units used as OGUs as follows: A) Sooretama Biological Reserve; B) Duas Bocas Biological Reserve; C) National Forest of Goytacazes; D) National Forest of Rio Preto; E) Lombardia Biological Station; F) Santa Lúcia Biological Station; G) Caparaó National Park; H) Natural Monument of Pontões Capixabas. O = OUTGROUP. The numbers refer to the characters’ state: 0) absent; 1) present. Characters Bichromomyia flaviscutellata Brumptomyia avellari Brumptomyia cardosoi Brumptomyia cunhai Brumptomyia figuereidoi Brumptomyia nitzulescui Brumptomyia troglodytes Evandromyia callipyga Evandromyia costalimai Evandromyia edwardsi Evandromyia grimaldii Evandromyia lenti Evandromyia sallesi Evandromyia sericea Evandromyia termitophila Evandromyia tupynambai Expapillata firmatoi Lutzomyia alencari Lutzomyia amarali Lutzomyia longipalpis Martinsmyia gasparviannai Micropygomyia capixaba Micropygomyia ferreirana Micropygomyia quinquefer Micropygomyia schreiberi Migonemyia migonei Nyssomyia intermedia Nyssomyia whitmani Nyssomyia yuilli yuilli Pintomyia bianchigalatiae Pintomyia fischeri Pintomyia mamedei Pintomyia misionensis Pintomyia monticola Pintomyia pessoai Pintomyia serrana Pressatia choti Pressatia equatorialis Psathyromyia lanei Psathyromyia lutziana Psathyromyia pascalei Psathyromyia pelloni Psathyromyia pestanai Psathyromyia shannoni Psychodopygus ayrozai Psychodopygus davisi Psychodopygus geniculatus Psychodopygus hirsutus Psychodopygus matosi Sciopemyia aff. microps Sciopemyia microps Trichopygomyia longispina
O 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
A 0 0 0 0 1 0 0 1 0 0 0 1 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 0 1 0 1 0 0 0 0 0 1 0 0 1 1 0 1 1 1 1 0 0 1 0 0 0
B 1 0 1 0 0 1 1 0 1 1 1 0 0 1 0 1 0 0 0 0 1 0 0 0 1 1 1 0 1 1 1 0 0 1 0 0 0 1 1 0 1 1 1 1 1 0 1 1 1 0 1 1
C 0 0 0 1 0 0 0 1 1 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 1 1 1 0 0 0 1 0 0 0 0 0 1 1 0 0 1 1 0 0 0 0 0 1 0 0 0 0
OGUs D 0 0 0 1 1 1 0 1 0 0 0 0 0 1 0 0 1 0 1 0 0 1 0 0 1 0 0 0 0 0 1 0 1 0 0 1 1 1 1 1 1 0 1 1 0 1 0 0 0 0 1 1
E 1 0 0 0 0 0 0 1 0 0 0 0 0 0 0 1 0 0 0 0 0 0 1 0 0 0 1 0 0 0 1 0 1 1 1 0 0 0 0 0 1 0 0 1 0 0 0 1 1 0 0 0
F 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 1 1 1 0 1 0 1 0 0 1 0 0 0 0 0 0 1 1 0 1 1 0 0 1 1 0 0 0
G 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 1 1 1 0 0 1 0 1 1 0 0 0 0 0 0 1 0 1 1 0 0 0 1 1 0 1 0
H 0 1 0 0 1 1 0 1 0 1 0 1 1 1 1 1 1 1 0 1 1 1 1 1 1 1 1 0 0 0 1 1 0 1 0 0 1 1 0 1 1 0 0 0 0 0 0 1 0 1 1 1
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Table 2. Number and sex of sand fly species collected with a Shannon trap from forest ambient and peridomiciliary ambient from June, 2008 to May, 2009 at the Goytacazes National Forest, municipality of Linhares, State of Espírito Santo, southeastern Brazil. Species ♂ Brumptomyia sp.* Evandromyia callipyga Evandromyia sallesi Evandromyia spp.** Micropygomyia schreiberi Migonemyia migonei Nyssomyia intermedia Pintomyia fischeri Pressatia choti Pressatia equatorialis Pressatia spp.*** Psathyromyia pelloni Total Species richness (S) Equitability (J) Shannon’s diversity (H)
Forest ♀ Total
1
13 1 7 9 81 5 1 118
1 7 4 3 10 7
115 146
8 0.65 1.36
7 17 4 17 16 81 5 115 1 264
Environment Peridomiciliary ♂ ♀ Total 1 1 2 2 1 1 5 5 1 1 2 24 23 47 169 436 605 44 14 58 447 447 44 44 455 455 731
936
8 0.54 1.13
1,667
♂ 3
14 25 176 53 528 49 1 849
♀ 1
Total
1 12 5 26 446 21
570 1,082
9 0.55 1.21
Total 1 3 1 12 19 51 622 74 528 49 570 1 1,931
*Brumptomyia sp. probably refers to females of Brumptomyia cunhai (Mangabeira, 1942); **Evandromyia spp. refers to females of Ev. callipyga (Martins and Silva, 1965) or Ev. costalimai (Mangabeira, 1942); *** Pressatia spp. refers to females of Pr. choti or Pr. equatorialis.
Figure 2. Dendrogram of similarity among eight conservation units within the state of Espírito Santo, southeastern Brazil.
32
Total
79 0.80 1.84
0.52
1.23
36 10
133
1
8
54
4
16
3
3
68
16
16
12
10
1
1
112
1
3
2
73
13
4
2
3
3
8
♂
1.56
0.65
11
89
1
2
44
3
24
4
1
7
3
♀
Total
201
2
5
44
2
73
3
37
8
3
7
3
3
3
8
T
62
55
1
6
♂
0.55
0.50
3
46
39
7
♀
39
55
1
13
T
1
64
1
23
43
♂
0.84
0.52
5
94
41
1
21
1
30
♀
41
1
64
2
44
1
30
43
T
1
119
2
29
43
♂
80
1
28
1
30
♀
Total
1
17
4
1
1
1
♂
80
1
2
2
119 112
3
57
1
30
43
T
2
67
21
3
1
6
♀
1
38
7
2
6
1
1
T
4
67
2
112
Ground
1
1
1
80
1
25
3
2
50
♂
1
57
4
31
1
1
2
33
♀
Canopy
Total
2
1
57
1
80
5
56
1
1
2
3
2
33
50
T
1
3
3
192
2
42
4
2
3
3
51
♂
3
192
6
94
8
3
8
3
3
33
51
T
1
2
2
5
124 124
4
52
4
1
8
33
♀
Total
1.08
0.67
5
1.02
0.46
9
1.41
0.59
11
1.34
0.55
11
226 194 140 334 141 100 241 164 130 294 306 229 535
Canopy
108 132
Ground
Peridomiciliary
*Brumptomyia sp. probably refers to females of Brumptomyia cunhai (Mangabeira, 1942); **Evandromyia spp. refers to females of Ev. callipyga (Martins and Silva, 1965) or Ev. costalimai (Mangabeira, 1942); *** Pressatia spp. refers to females of Pr. choti or Pr. equatorialis. The abbreviation of generic names follows the model proposed by Marcondes (2007).
S J H
2
2
28
16
2
1
1
1
3
3 1
2
3
7
T
2
3
♀
1
2
28
2
25
2
14
11
7
57
3
4
2
57
1
1
1
1
6
1
7
♂
1
T
1
6
♀
Canopy
1
♂
Ground
Forest
Br. cunhai Br. sp.* Ev. callipyga Ev. costalimai Ev. spp.** Mi. schreiberi Mg. migonei Ny. intermedia Pi. fischeri Pr. choti Pr. equatorialis Pr. spp.*** Pa. pascalei Ps. hirsutus
Species
Environment
Level
Table 3. Number and sex of sand fly species collected with CDC traps in two different strata from forest ambient and peridomiciliary ambient from June, 2008 to May, 2009 at the Goytacazes National Forest, municipality of Linhares, state of Espírito Santo, southeastern Brazil. (T = Total; S = species richness; J = equitability; H = Shannon’s diversity).
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the forest environment. This same result was observed by different authors in the Atlantic forest biome (Cortez et al. 2007, Pinto et al. 2010b) as well as in the Amazon rainforest (Dias-Lima et al. 2002). However, the number of Mg. migonei specimens collected within the forest environment in the NFG was lower in the canopy level, different from the results found by Cortez et al. (2007) in an Atlantic forest area of northeastern Brazil, by Marcondes et al. (2001) in an Atlantic forest area of southern Brazil, and by Dias-Lima et al. (2002) in an Amazon rainforest area of northern Brazil. Despite the low number of Mg. migonei collected in the NFG, the higher number of specimens collected on the ground level of the forest can be epidemiologically important. Since Mg. migonei has a role in the transmission of L. braziliensis parasites, the higher numbers of this insect at ground level can favor the contact between the parasites and human hosts via vectors. Ps. hirsutus and Ev. sallesi were already found naturally infected with Leishmania parasites (Rangel et al. 1985, Saraiva et al. 2009), however, they deserve no special attention in the NFG, since they were collected in very low numbers and the role of these species in the transmission cycle of Leishmania parasites still remains unclear. The species diversity and richness were higher in the forested environment than in the modified environment. This was also reported by Virgens et al. (2008) at the Sooretama Biological Reserve. These authors discussed that this pattern can be explained by the greater variety of resting sites provided by vegetation cover. Sampling sand flies from another Atlantic Forest reserve in Espírito Santo, Pinto et al. (2010b) pointed out that, in addition to the great variety of resting sites, the availability of food sources influenced by a high diversity of vertebrates could also help to explain the high diversity and richness of species in the forest environment. The results of the similarity analyses allow us to infer that the sand fly fauna can be more similar at the same elevation level with one clade for the highland conservation units and another for the intermediate elevation conservation units. Unfortunately, the Sooretama Biological Reserve made the lowlands conservation units a paraphyletic group. This may be because the sand fly collections from the Sooretama Biological Reserve present insufficient sampling of its sand fly fauna compared with the other lowland conservation units. All the sand fly collections in the Sooretama Biological Reserve were performed using only modified Shannon traps (Virgens et al. 2008), while in the other lowland conservation units, automatic CDC traps were used (Pinto et al. 2012a,b, Ferreira et al. 2013). This sampling bias can explain the position of the Sooretama Biological Reserve in the trees. This hypothesis is also reinforced by the higher species richness recorded for the National Forest of Rio Preto and for the Natural Monument of Pontões Capixabas compared with the Sooretama Biological Reserve. The conservation level and larger forest area range of the Sooretama Biological Reserve is more similar to the conservation units of the intermediate and high lands, and it could explain its position in the trees. However, this corroborates the sampling bias hypothesis since the number of sand fly species is correlated to the extension of the forest cover range. Therefore, a more comprehensive sand fly survey from the Sooretama Biological Reserve is necessary, and more accurate methods are necessary to analyze the similarity among the conservation units of the state of Espírito Santo. In this work, we presented aspects of the diversity of sand flies
June 2015
in an Atlantic Forest conservation unit in the state of Espírito Santo and pointed out preferences of some species for the peridomiciliary environment. We also discussed aspects concerning the risk of transmission of Leishmania parasites in the area and, finally, we raised the first hypothesis of similarity relationships among conservation units within the state of Espírito Santo. Acknowledgments We thank Claudenice Cumin, Fabrício Iglesias Valente, Fernanda Passos, Janine Santana Santos, Kênia Carolina, Márcia Baiôco, Morena Andrade Liberato, and Tainara Viguini for their help with the field work. We thank Gustavo Rocha Leite for the map, Leony Wand Del Rey, manager of the NFG, for the logistic support during the field work, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for providing a fellowship to Israel S. Pinto, and the Federal Institute of Education, Science, and Technology of Espírito Santo (IFES) for the financial support for the revision of this article. REFERENCES CITED Barretto, M.P. and J.O. Coutinho. 1940. Processos de captura, dissecação e montagem de flebótomos. Ann. Facul. Med. de São Paulo 16: 173–187. Barros, G.C., P.A. Sessa, E.A. Mattos, V.R.D. Carias, W. Mayrink, J.T.A. Alencar, A. Falqueto, and A.C. Jesus. 1985. Foco de leishmaniose tegumentar nos municípios de Viana e Cariacica, estado do Espírito Santo, Brasil. Rev. Saúde Públ. 19: 146–153. Cortez A.M., V.P.M. Silva, P.V.S. Queiroz, H.T.A. Andrade, M.I.B. Loiola, and M.F.F.M. Ximenes. 2007. Vertical stratification and development aspects of phlebotomine sand flies (Diptera, Psychodidae) in an area of Atlantic Forest tree species in a metropolitan region in northeastern Brazil. J. Vector Ecol. 32: 336-341. Costa, L.P., Y.L.R. Leite, G.A.B. Fonseca, and M.T. Fonseca. 2000. Biogeography of South American forest mammals: endemism and diversity in the Atlantic Forest. Biotropica 32: 872-881. Crovello, T.J. 1981. Quantitative biogeography: an overview. Taxon 30: 563–575. Dias-Lima A., E.C. Bermúdez, J.F. Medeiros, and I. Sherlock. 2002. Estratificação vertical da fauna de flebótomos (Diptera: Psychodidae) numa floresta primária de terra firme da Amazônia Central, Estado do Amazonas, Brasil. Cad. Saúde Públ. 18: 823-832. Falqueto, A., P.A. Sessa, J.B.M. Varejão, G.C. Barros, H. Momen, and G. Grimaldi Jr. 1991. Leishmaniasis due to Leishmania braziliensis in Espírito Santo state, Brazil: further evidence on the role of dogs as a reservoir of infection for humans. Mem. Inst. Oswaldo Cruz 86: 499-500. Ferreira, A.L., P.A. Sessa, J.B.M Varejão, and A. Falqueto. 2001. Distribution of sand flies (Diptera: Psychodidae) at different altitudes in an endemic region of american cutaneous leishmaniasis in the state of Espírito Santo, Brazil. Mem. Inst. Oswaldo Cruz 96: 1061–1067. Ferreira A.L., A. Falqueto, G. Grimaldi, A.A. Peixoto, and I.S. Pinto. 2013. Ecological and epidemiological aspects of
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the sand fly (Diptera, Psychodidae) fauna of the National Monument of Pontões Capixabas, State of Espírito Santo, Southeastern Brazil. J. Med. Entomol. 50: 1215-1223. Galati, E.A.B. 1995. Phylogenetic systematics of Phlebotominae (Diptera, Psychodidae) with emphasis on American groups. Bol. Dir. Malariol. Saneam. Amb. 35: 133-142. Galati, E.A.B. 2003. Morfologia e Taxonomia. In: E.F. Rangel and R. Lainson (eds.) Flebotomíneos do Brasil. pp. 23–206. Fiocruz, Rio de Janeiro, iii + 367. Gomes A.C., E.X. Rabello, J.L.F. Santos, and E.A.B. Galati. 1983. Aspectos ecológicos da Leishmaniose Tegumentar Americana. 3. Observações naturais sobre o ritmo diário da atividade de Psychodopygus intermedius em ambiente florestal e extraflorestal. Rev. Saúde Públ. 17: 23-30. Hammer, Ø., D.A.T. Harper, and P.D. Ryan. 2001. PAST: Paleontological statistics software package for education and data analysis. Palaentol. Electronica 4: 9 pp. Hijmans, R.J., S.E. Cameron, J.L. Parra, P.G. Jones, and A. Jarvis. 2005. Very high resolution interpolated climate surfaces for global land areas. Int. J. Climatol. 25: 1965–1978. Maddison, W. P. and D. R. Maddison. 2006. Mesquite: a modular system for evolutionary analysis, version 1.12. (http:// mesquiteproject.org/mesquite/mesquite.html). Marcondes, C.B., L.G. Santos-Neto, and A.L. Lozovei. 2001. Ecology of Phlebotomine sandflies (Diptera, Psychodidae) in Brazilian Atlantic Forest. Rev. Soc. Bras. Med. Trop. 34: 255–260. Marcondes, C.B. 2007. A proposal of generic and subgeneric abbreviations for Phlebotomine sandflies (Diptera: Psychodidae: Phlebotominae) of the world. Entomol. News 118: 351–356. Myers, N., R.A. Mittermeier, C.G. Mittermeier, G.A.B. Fonseca, and J. Kent. 2000. Biodiversity hotspots for conservation priorities. Nature 403: 853-858. Orme C. D. L., R.G. Davies, M. Burgess, F. Eigenbrod, N. Pickup, V.A. Olson, A.J. Webster, D. Tzung-Su, P.C. Rasmussen, R.S. Ridgely, A.J. Stattersfield, P.M. Bennett, T.M. Blackburn, K.J. Gaston, and I.P.F. Owens. 2005. Global hotspots of species richness are not congruent with endemism or threat. Nature 463: 1016-1019. Peel, M.C., B.L. Finlayson, and T.A. McMahon. 2007. Updated world map of the Köppen-Geiger climate classification. Hydrol. Earth Syst. Sci. 11: 1633–1644. Pinto, I.S., C.B. Santos, G. Grimaldi Jr., A.L. Ferreira, and A. Falqueto. 2010a. American visceral leishmaniasis dissociated from Lutzomyia longipalpis (Diptera, Psychodidae) in the State of Espírito Santo, Brazil. Cad. Saúde Públ. 26: 365-372. Pinto, I.S., C.B. Santos, A.L. Ferreira, and A. Falqueto. 2010b. Richness and diversity of sand flies (Diptera, Psychodidae) in an Atlantic rainforest reserve in southeastern Brazil. J. Vector Ecol. 35: 325-332.
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Pinto, I.S., A.L. Ferreira, V. Valim, F.S. Carvalho, G.M. Silva, A.L. Falcão, R. Dietze, and A. Falqueto. 2012a. Sand fly vectors (Diptera, Psychodidae) of American visceral leishmaniasis areas in the Atlantic Forest, State of Espírito Santo, southeastern Brazil. J. Vector Ecol. 37: 90-96. Pinto, I.S., J.F.R. Tonini, A.L. Ferreira, and A. Falqueto. 2012b. A brief inventory of sand flies (Diptera, Psychodidae) from the National Forest of the Rio Preto, state of the Espírito Santo, southeastern Brazil. Biota Neotrop. 12: 323-326. Pita-Pereira D, C.R. Alves, M.B. Souza, R.P. Brazil, A.L. Bertho, A.F. Barbosa, and C.C. Brito. 2005. Identification of naturally infected Lutzomyia intermedia and Lutzomyia migonei with Leishmania (Viannia) braziliensis in Rio de Janeiro (Brazil) revealed by a PCR multiplex non-isotopic hybridisation assay. Trans. R. Soc. Trop. Med. Hyg. 99: 905-913. Rangel E.F., N.A. Souza, E.D. Wermeliton, and A.F. Barbosa. 1984. Infecção natural de Lutzomyia intermedia Lutz & Neiva, 1912 em área endêmica de leishmaniose tegumentar americana no estado do Rio de Janeiro. Mem. Inst. Oswaldo Cruz 79: 395396. Rangel E.F., L. Ryan, R. Lainson, and J.J. Shaw. 1985. Observations on the sandfly (Diptera: Psychodidae) fauna of Além Paraíba, state of Minas Gerais, Brazil, and the isolation of a parasite of the Leishmania braziliensis complex from Psychodopygus hirsuta hirsuta. Mem. Inst. Oswaldo Cruz 80: 373-374. Rolim S.G., N.M. Ivanauskas, R.R. Rodrigues, M.T. Nascimento, J.M.L. Gomes, D.A. Folli, and H.T.Z. Couto. 2006. Composição Florística do estrato arbóreo da floresta estacional semidecidual na planície aluvial do rio Doce, Linhares, ES, Brasil. Acta Bot. Bras. 20: 549-561. Rosen, B.R. 1988. From fossils to Earth history: Applied historical biogeography. In: A.A. Myers and P.S. Giller (eds.). Analytical Biogeography: An Integrated Approach to the Study of Animal and Plant Distributions. pp. 437–481. Chapman and Hall, London. Saraiva L., G.M.L. Carvalho, C.M.F. Gontijo, P.F. Quaresma, A.C.V.M.R. Lima, A.L. Falcão, and J.D. Andrade Filho. 2009. Natural infection of Lutzomyia neivai and Lutzomyia sallesi (Diptera: Psychodidae) by Leishmania infantum chagasi in Brazil. J. Med. Entomol. 46: 1159-1163. Teodoro, U., V.L. Salvia-Filho, E.D. Lima, R.P. Spinosa, O.C. Barbosa, M.E.M.C. Ferreira, and M.V.C. Lonardoni. 1993. Observações sobre o comportamento de flebotomíneos em ecótopos florestais e extraflorestais, em área endêmica de leishmaniose tegumentar americana, no norte do Estado do Paraná, sul do Brasil. Rev. Saúde Públ. 27: 242–249. Virgens T.M., C.B. Santos, I.S. Pinto, K.S. Silva, F.C. Leal, and A. Falqueto. 2008. Phlebotomine sand flies (Diptera, Psychodidae) in an American tegumentary leishmaniasis transmission area in northern Espírito Santo state, Brazil. Cad. Saúde Públ. 24: 2969–2978.
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June 2015
Sand fly fauna (Diptera: Psychodidae) from the Goytacazes National Forest and surrounding areas of southeastern Brazil Thieres Marassati das Virgens1, Helder Ricas Rezende2, Israel de Souza Pinto3*, and Aloísio Falqueto4 Instituto Federal de Educação, Ciência e Tecnologia do Espírito Santo, Campus Linhares, Av. Filogônio Peixoto S/N, Bairro Aviso, CEP 29901-291, Linhares, Espírito Santo, Brazil 2 Núcleo de Entomologia e Malacologia do Espírito Santo, Secretaria de Estado da Saúde, Rua Pedro Zangrande 381, Bairro Jardim Limoeiro, CEP 29164-020, Serra, Espírito Santo, Brazil 3 Laboratório de Biologia Molecular de Insetos, Instituto Oswaldo Cruz, FIOCRUZ, Av. Brasil 4365, Manguinhos, CEP 21040-360, Rio de Janeiro, Rio de Janeiro, Brazil, [email protected] 4 Unidade de Medicina Tropical, Departamento de Patologia, Universidade Federal do Espírito Santo, Av. Marechal Campos 1468, Maruípe, CEP 29043-900, Vitória, Espírito Santo, Brazil 1
Received 9 July 2014; Accepted 20 November 2014 ABSTRACT: Most studies of the sand fly fauna in southeastern Brazil are conducted in the peridomiciliary environment of leishmaniasis endemic regions. Therefore, to increase the knowledge about diversity and richness of sand fly conservation areas, we describe here the sand fly fauna from the National Forest of Goytacazes (NFG), state of Espírito Santo, Brazil, and its surroundings areas. We also used sand fly fauna records from eight conservations units within the state of Espírito Santo to understand the similarity and relationships among them. The sand flies were simultaneously collected from June, 2008 to May, 2009 in two different environments: a preserved environment represented by the NFG and a modified environment represented by a peridomicile. To establish the similarity among the conservation units, we used a method very similar to parsimony analysis of endemism. We collected 2,466 sand fly specimens belonging to 13 species. Pressatia choti and Nyssomyia intermedia were the most abundant sand fly species. Ny. intermedia is a known vector of Leishmania braziliensis and epidemiological surveillance must be conducted in the area. We discuss aspects regarding the diversity of sand flies as well as the risk of transmission of Leishmania parasites in the area. We also provide for the first time a hypothesis of similarity relationships among conservation units within the state of Espírito Santo. Journal of Vector Ecology 40 (1): 28-35. 2015. Keyword Index: Phlebotominae, biodiversity, Atlantic forest, leishmaniasis, parsimony, endemism.
INTRODUCTION Despite its degradation and deforestation, the Atlantic Forest still remains one of the hotspots of vertebrate biodiversity in the world (Myers et al. 2000, Orme et al. 2005). Since vertebrates are the food source for hematophagous insects, we can also predict that the Atlantic forest biome has high species richness of insects such as fleas, mosquitoes, and phlebotomine sand flies. The phlebotomine sand flies (Diptera: Psychodidae) are the main leishmaniasis vectors along the Brazilian states situated on the Atlantic coast, such as the state of Espírito Santo. Historically, almost all the studies of sand fly fauna in this state were conducted in disturbed areas of endemic leishmaniasis regions as a result of the medical and veterinary importance of these flies (Barros et al. 1985, Ferreira et al. 2001, Pinto et al. 2010a, Pinto et al. 2012a). Only recently, some studies were conducted in conservation units next to leishmaniasis endemic (Pinto et al. 2012b) or nonendemic (Pinto et al. 2012b) areas. These last studies highlighted the importance of sand fly surveys in preserved environments, which present higher species richness than in disturbed areas. In Espírito Santo, there have been only two surveys on sand fly fauna that were simultaneously conducted in a conservation unit and in a surrounding area to compare sand fly fauna between the two different environments, also recording higher species richness
in the preserved environment (Virgens et al. 2008, Ferreira et al. 2013). Nevertheless, some of the conservation units situated in Espírito Santo still lack an inventory of sand fly fauna. Furthermore, there is also no information about the similarity among conservation units within the state of Espírito Santo based on sand fly species records. Therefore, our aim was to describe, for the first time, the sand fly fauna from the National Forest of Goytacazes and surrounding areas of southeastern Brazil. In addition, we used sand fly fauna records from conservations units of the state of Espírito Santo to understand the similarity and the relationships among them. MATERIALS AND METHODS Study area The study was carried out simultaneously in two different environments: 1) a preserved environment represented by the National Forest of Goytacazes (NFG), and 2) a modified environment represented by a peridomicile with some domestic animals, such as dogs, chickens, and pigs. The NFG (19º 28’ 01” S; 40º 04’ 18” W; elevation of 10 m [a.s.l.]) is a federal conservation unit situated in the municipality of Linhares, state of Espírito Santo, southeastern Brazil (Figure
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Figure 1. Collection sites (square: peridomiciliary environment; circle: forest environment represented of the National Forest of Goytacazes) in the Atlantic Forest, state of Espírito Santo, southeastern Brazil. 1). Currently, the NFG covers an area of 1,424 hectares and is a remnant of the Atlantic Forest on the Doce River alluvial floodplain (Rolim et al. 2006). Almost all the NFG area is covered by a secondary forest, but the vegetation is dense with trees as tall as 30 m that present moderate overlapping canopies. The KöppenGeiger climate classification for the region is tropical monsoon (Peel et al. 2007). The average temperatures range from 11 to 18° C in the colder months (June to August) and from 30 to 34° C in the warmer months (December to March), with annual rainfall of approximately 1,200 mm (Hijmans et al. 2005). Sand fly captures The sand flies were collected simultaneously in the modified and preserved environments from June, 2008 to May, 2009. For each night and environment, we used one illuminated Shannon trap and two CDC (Center for Disease Control) automatic light traps. For each environment, one of us collected sand flies with an aspirator during the first three h of the night (18:00 to 21:00) in the illuminated Shannon trap, while one automatic light trap was installed up to one m above ground level and the other in the tree canopy (at ~10 m height). The automatic light traps worked during the night (18:00 to 21:00). We spent 36 collection hours for each trap and a total of 108 collection hours for each environment.
The sand flies collected were stored in 70% ethanol and mounted on glass slides using the technique of Barretto and Coutinho (1940). We used the classification of Galati (1995, 2003) and the abbreviation of generic names of Marcondes (2007). We identified all the species according to morphological characters with the taxonomic key of Galati (2003). The sample material was deposited in the entomology collection of the Department of Tropical Medicine at the Federal University in Espírito Santo. Climate data (rainfall, relative humidity, and mean temperature) were obtained from a station of the National Institute of Meteorology situated at the Espírito Santo state Institute for Research, Technical Assistance, and Rural Extension (INCAPER), municipality of Linhares, which borders the NFG. Statistical analysis We evaluated the specific richness (S), equitability (J), and Shannon’s diversity index for each environment, kind of trap, and vertical stratum. These analyses were performed in PAST (Hammer et al. 2001). We used the Mann-Whitney test to verify differences between the numbers of sand fly specimens collected by the CDC automatic trap on the ground level and on the tree canopy for the two environmental samplings. We used the Spearman non-
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parametric test to evaluate the correlation between climate data and the most abundant species for each environment and vertical stratum. We considered the differences to be significant at p < 0.05. These analyses were performed in Biostat 5.0.
June 2015
Similarity analysis To establish the similarity among conservation units within the state of Espírito Santo, we used eight conservation areas as Operational Geographic Units (OGUs) (Crovello 1981) and sand fly species records were used as characters (Table 1). The sand fly distribution data for the conservation units were compiled from Virgens et al. (2008), Pinto et al. (2010 a,b), Pinto et al. (2012 a,b), Ferreira et al. (2013), and Núcleo de Entomologia da Secretaria do Estado da Saúde do Espírito Santo (unpublished data). The conservation units can be grouped into three kinds using the elevation as a classification criterion: 1) highland conservation units (830 m a.s.l.]) - Lombardia Biological Station and Caparaó National Park; 2) intermediate (500 m a.s.l. < elevation < 750 m a.s.l.) - Duas Bocas Biological Reserve and Santa Lúcia Biological Station; and 3) lowlands conservation units (< 400 m a.s.l.) - National Forest of Goytacazes, National Forest of Rio Preto, Natural Monument of Pontões Capixabas, and Sooretama Biological Reserve. We coded the absence of a species as a primitive character (0) and its presence as a derived character (1) of an OGU. For the tree rooting, we used a hypothetical locality having no taxa, a procedure known as Lundberg rooting following Rosen (1988) and Costa et al. (2000). Fifty-two species occurrences were used as characters. All characters were treated as equally weighted and unordered in the similarity analyses. The data matrix (Table 1) was built in Mesquite (Maddison and Maddison 2006). The data were analyzed using the maximum parsimony optimality criterion from PAUP* 4.0b10 (Sinauer, Sunderland, MA) and the interface PaupUp 1.0.3.1 for Microsoft Windows 95/NT. We conducted a heuristic search using tree-bisection-reconnection as a branch swapping algorithm with a starting tree obtained through stepwise addition held with one tree at each step and computed a strict consensus of the most parsimonious trees. All the trees were inspected using FigTree v1.3.1.
overall male/female sex ratio was 0.88 and, for captures using the Shannon modified trap and the CDC traps, the ratios were 0.78 and 1.33, respectively. The number and sex of the species collected with the Shannon trap and the CDC trap for both environments are presented in Tables 2 and 3, respectively. Pressatia choti was the most abundant sand fly species collected with 720 specimens (29.2%), followed by Ny. intermedia, 716 specimens (29.0%). Pr. choti was also the most abundant species, followed by Ny. intermedia for each stratum analyzed for the two environments sampled (Table 3). Therefore, only these two species were used in the statistical analyses. The number of Ny. intermedia collected in the peridomiciliary environment was significantly higher than in the forest environment. There is no significant difference between the number of collected specimens belonging to Pr. choti and Ny. intermedia between the ground level and the tree canopy level for each environment. In the forest environment, Pr. choti was negatively correlated with monthly rainfall (R = -0.76, p = 0.00) and Ny. intermedia was negatively correlated with monthly rainfall (R = -0.70, p = 0.01) and monthly average temperature (R = -0.76, p = 0.00). In the peridomicile, Pr. choti was negatively correlated with monthly rainfall (R = -0.73, p = 0.00) and Ny. intermedia was negatively correlated with monthly average temperature (R = -0.65, p = 0.02). Regarding the similarity analysis, we found two most parsimonious trees (Figure 2), 92 steps-long each. Thirty-six characters were parsimony-informative and the overall Consistency Index (excluding uninformative characters) was 0.4737, and the Retention Index was 0.4030. The two highland conservation units (Lombardia Biological Station and Caparaó National Park) composed a clade in a tree. In the other tree, the Lombardia Biological Station is a sister group of the Caparaó National Park. The two conservation units with intermediate elevation (Duas Bocas Biological Reserve and Santa Lúcia Biological Station) composed another clade. The lowlands conservation units were paraphyletic, since the Sooretama Biological Reserve was a sister group of the clade composed of the Duas Bocas Biological Reserve and the Santa Lúcia Biological Station.
RESULTS
DISCUSSION
We collected 2,466 sand fly specimens belonging to 13 species, nine genera, and four sub-tribes: Brumptomyiina - Brumptomyia cunhai (Mangabeira 1942); Lutzomyiina - Evandromyia callipyga (Martins and Silva 1965), Evandromyia costalimai (Mangabeira 1942), Evandromyia sallesi (Galvão and Coutinho 1939), Migonemyia migonei (França 1920), Pintomyia fischeri (Pinto 1926), Pressatia choti (Floch and Abonnenc 1941) and Pressatia equatorialis (Mangabeira 1942); Psychodopygiina - Nyssomyia intermedia (Lutz and Neiva 1912), Psathyromyia pascalei (Coutinho and Barreto 1941), Psathyromyia pelloni (Sherlock and Alencar 1959) and Psychodopygus hirsutus (Mangabeira 1942); and Sergentomyiina - Micropygomyia schreiberi (Martins, Falcão and Silva 1975). The number of sand flies collected was higher in the peridomiciliary environment (n = 2,001) than in the forest environment. However, the species diversity and richness were higher in the forest than in the peridomiciliary environment. The
The sand fly fauna recorded in the National Forest of Goytacazes was similar to that of forested regions of northern Espírito Santo, with an abundance of Pr. choti (Virgens et al. 2008, Pinto et al. 2012b). Considering only the species from the modified environment sampled near the NFG, Ny. intermedia and Mg. migonei were also abundant. This seems to be a pattern along the state of Espírito Santo, as pointed out by different works conducted in modified environments (Barros et al. 1985, Falqueto et al. 1991, Ferreira et al. 2001, Virgens et al. 2008, Pinto et al. 2010a,b). These last two species are incriminated in the transmission of Leishmania braziliensis Vianna, 1911 in endemic areas of cutaneous leishmaniasis along the Brazilian Atlantic coast (Gomes et al. 1983, Rangel et al. 1984, Teodoro et al. 1993, PitaPereira et al. 2005). Therefore, despite the absence of leishmaniasis records close to the NFG to date, it is important to maintain epidemiological surveillance in the area. The number of Mg. migonei specimens collected was lower in
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Table 1. Data matrix of 52 sand fly species used as characters for eight Operational Geographical Units (OGUs) to perform the similarity analysis. The letters represent the conservation units used as OGUs as follows: A) Sooretama Biological Reserve; B) Duas Bocas Biological Reserve; C) National Forest of Goytacazes; D) National Forest of Rio Preto; E) Lombardia Biological Station; F) Santa Lúcia Biological Station; G) Caparaó National Park; H) Natural Monument of Pontões Capixabas. O = OUTGROUP. The numbers refer to the characters’ state: 0) absent; 1) present. Characters Bichromomyia flaviscutellata Brumptomyia avellari Brumptomyia cardosoi Brumptomyia cunhai Brumptomyia figuereidoi Brumptomyia nitzulescui Brumptomyia troglodytes Evandromyia callipyga Evandromyia costalimai Evandromyia edwardsi Evandromyia grimaldii Evandromyia lenti Evandromyia sallesi Evandromyia sericea Evandromyia termitophila Evandromyia tupynambai Expapillata firmatoi Lutzomyia alencari Lutzomyia amarali Lutzomyia longipalpis Martinsmyia gasparviannai Micropygomyia capixaba Micropygomyia ferreirana Micropygomyia quinquefer Micropygomyia schreiberi Migonemyia migonei Nyssomyia intermedia Nyssomyia whitmani Nyssomyia yuilli yuilli Pintomyia bianchigalatiae Pintomyia fischeri Pintomyia mamedei Pintomyia misionensis Pintomyia monticola Pintomyia pessoai Pintomyia serrana Pressatia choti Pressatia equatorialis Psathyromyia lanei Psathyromyia lutziana Psathyromyia pascalei Psathyromyia pelloni Psathyromyia pestanai Psathyromyia shannoni Psychodopygus ayrozai Psychodopygus davisi Psychodopygus geniculatus Psychodopygus hirsutus Psychodopygus matosi Sciopemyia aff. microps Sciopemyia microps Trichopygomyia longispina
O 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
A 0 0 0 0 1 0 0 1 0 0 0 1 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 0 1 0 1 0 0 0 0 0 1 0 0 1 1 0 1 1 1 1 0 0 1 0 0 0
B 1 0 1 0 0 1 1 0 1 1 1 0 0 1 0 1 0 0 0 0 1 0 0 0 1 1 1 0 1 1 1 0 0 1 0 0 0 1 1 0 1 1 1 1 1 0 1 1 1 0 1 1
C 0 0 0 1 0 0 0 1 1 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 1 1 1 0 0 0 1 0 0 0 0 0 1 1 0 0 1 1 0 0 0 0 0 1 0 0 0 0
OGUs D 0 0 0 1 1 1 0 1 0 0 0 0 0 1 0 0 1 0 1 0 0 1 0 0 1 0 0 0 0 0 1 0 1 0 0 1 1 1 1 1 1 0 1 1 0 1 0 0 0 0 1 1
E 1 0 0 0 0 0 0 1 0 0 0 0 0 0 0 1 0 0 0 0 0 0 1 0 0 0 1 0 0 0 1 0 1 1 1 0 0 0 0 0 1 0 0 1 0 0 0 1 1 0 0 0
F 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 1 1 1 0 1 0 1 0 0 1 0 0 0 0 0 0 1 1 0 1 1 0 0 1 1 0 0 0
G 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 1 1 1 0 0 1 0 1 1 0 0 0 0 0 0 1 0 1 1 0 0 0 1 1 0 1 0
H 0 1 0 0 1 1 0 1 0 1 0 1 1 1 1 1 1 1 0 1 1 1 1 1 1 1 1 0 0 0 1 1 0 1 0 0 1 1 0 1 1 0 0 0 0 0 0 1 0 1 1 1
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June 2015
Table 2. Number and sex of sand fly species collected with a Shannon trap from forest ambient and peridomiciliary ambient from June, 2008 to May, 2009 at the Goytacazes National Forest, municipality of Linhares, State of Espírito Santo, southeastern Brazil. Species ♂ Brumptomyia sp.* Evandromyia callipyga Evandromyia sallesi Evandromyia spp.** Micropygomyia schreiberi Migonemyia migonei Nyssomyia intermedia Pintomyia fischeri Pressatia choti Pressatia equatorialis Pressatia spp.*** Psathyromyia pelloni Total Species richness (S) Equitability (J) Shannon’s diversity (H)
Forest ♀ Total
1
13 1 7 9 81 5 1 118
1 7 4 3 10 7
115 146
8 0.65 1.36
7 17 4 17 16 81 5 115 1 264
Environment Peridomiciliary ♂ ♀ Total 1 1 2 2 1 1 5 5 1 1 2 24 23 47 169 436 605 44 14 58 447 447 44 44 455 455 731
936
8 0.54 1.13
1,667
♂ 3
14 25 176 53 528 49 1 849
♀ 1
Total
1 12 5 26 446 21
570 1,082
9 0.55 1.21
Total 1 3 1 12 19 51 622 74 528 49 570 1 1,931
*Brumptomyia sp. probably refers to females of Brumptomyia cunhai (Mangabeira, 1942); **Evandromyia spp. refers to females of Ev. callipyga (Martins and Silva, 1965) or Ev. costalimai (Mangabeira, 1942); *** Pressatia spp. refers to females of Pr. choti or Pr. equatorialis.
Figure 2. Dendrogram of similarity among eight conservation units within the state of Espírito Santo, southeastern Brazil.
32
Total
79 0.80 1.84
0.52
1.23
36 10
133
1
8
54
4
16
3
3
68
16
16
12
10
1
1
112
1
3
2
73
13
4
2
3
3
8
♂
1.56
0.65
11
89
1
2
44
3
24
4
1
7
3
♀
Total
201
2
5
44
2
73
3
37
8
3
7
3
3
3
8
T
62
55
1
6
♂
0.55
0.50
3
46
39
7
♀
39
55
1
13
T
1
64
1
23
43
♂
0.84
0.52
5
94
41
1
21
1
30
♀
41
1
64
2
44
1
30
43
T
1
119
2
29
43
♂
80
1
28
1
30
♀
Total
1
17
4
1
1
1
♂
80
1
2
2
119 112
3
57
1
30
43
T
2
67
21
3
1
6
♀
1
38
7
2
6
1
1
T
4
67
2
112
Ground
1
1
1
80
1
25
3
2
50
♂
1
57
4
31
1
1
2
33
♀
Canopy
Total
2
1
57
1
80
5
56
1
1
2
3
2
33
50
T
1
3
3
192
2
42
4
2
3
3
51
♂
3
192
6
94
8
3
8
3
3
33
51
T
1
2
2
5
124 124
4
52
4
1
8
33
♀
Total
1.08
0.67
5
1.02
0.46
9
1.41
0.59
11
1.34
0.55
11
226 194 140 334 141 100 241 164 130 294 306 229 535
Canopy
108 132
Ground
Peridomiciliary
*Brumptomyia sp. probably refers to females of Brumptomyia cunhai (Mangabeira, 1942); **Evandromyia spp. refers to females of Ev. callipyga (Martins and Silva, 1965) or Ev. costalimai (Mangabeira, 1942); *** Pressatia spp. refers to females of Pr. choti or Pr. equatorialis. The abbreviation of generic names follows the model proposed by Marcondes (2007).
S J H
2
2
28
16
2
1
1
1
3
3 1
2
3
7
T
2
3
♀
1
2
28
2
25
2
14
11
7
57
3
4
2
57
1
1
1
1
6
1
7
♂
1
T
1
6
♀
Canopy
1
♂
Ground
Forest
Br. cunhai Br. sp.* Ev. callipyga Ev. costalimai Ev. spp.** Mi. schreiberi Mg. migonei Ny. intermedia Pi. fischeri Pr. choti Pr. equatorialis Pr. spp.*** Pa. pascalei Ps. hirsutus
Species
Environment
Level
Table 3. Number and sex of sand fly species collected with CDC traps in two different strata from forest ambient and peridomiciliary ambient from June, 2008 to May, 2009 at the Goytacazes National Forest, municipality of Linhares, state of Espírito Santo, southeastern Brazil. (T = Total; S = species richness; J = equitability; H = Shannon’s diversity).
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the forest environment. This same result was observed by different authors in the Atlantic forest biome (Cortez et al. 2007, Pinto et al. 2010b) as well as in the Amazon rainforest (Dias-Lima et al. 2002). However, the number of Mg. migonei specimens collected within the forest environment in the NFG was lower in the canopy level, different from the results found by Cortez et al. (2007) in an Atlantic forest area of northeastern Brazil, by Marcondes et al. (2001) in an Atlantic forest area of southern Brazil, and by Dias-Lima et al. (2002) in an Amazon rainforest area of northern Brazil. Despite the low number of Mg. migonei collected in the NFG, the higher number of specimens collected on the ground level of the forest can be epidemiologically important. Since Mg. migonei has a role in the transmission of L. braziliensis parasites, the higher numbers of this insect at ground level can favor the contact between the parasites and human hosts via vectors. Ps. hirsutus and Ev. sallesi were already found naturally infected with Leishmania parasites (Rangel et al. 1985, Saraiva et al. 2009), however, they deserve no special attention in the NFG, since they were collected in very low numbers and the role of these species in the transmission cycle of Leishmania parasites still remains unclear. The species diversity and richness were higher in the forested environment than in the modified environment. This was also reported by Virgens et al. (2008) at the Sooretama Biological Reserve. These authors discussed that this pattern can be explained by the greater variety of resting sites provided by vegetation cover. Sampling sand flies from another Atlantic Forest reserve in Espírito Santo, Pinto et al. (2010b) pointed out that, in addition to the great variety of resting sites, the availability of food sources influenced by a high diversity of vertebrates could also help to explain the high diversity and richness of species in the forest environment. The results of the similarity analyses allow us to infer that the sand fly fauna can be more similar at the same elevation level with one clade for the highland conservation units and another for the intermediate elevation conservation units. Unfortunately, the Sooretama Biological Reserve made the lowlands conservation units a paraphyletic group. This may be because the sand fly collections from the Sooretama Biological Reserve present insufficient sampling of its sand fly fauna compared with the other lowland conservation units. All the sand fly collections in the Sooretama Biological Reserve were performed using only modified Shannon traps (Virgens et al. 2008), while in the other lowland conservation units, automatic CDC traps were used (Pinto et al. 2012a,b, Ferreira et al. 2013). This sampling bias can explain the position of the Sooretama Biological Reserve in the trees. This hypothesis is also reinforced by the higher species richness recorded for the National Forest of Rio Preto and for the Natural Monument of Pontões Capixabas compared with the Sooretama Biological Reserve. The conservation level and larger forest area range of the Sooretama Biological Reserve is more similar to the conservation units of the intermediate and high lands, and it could explain its position in the trees. However, this corroborates the sampling bias hypothesis since the number of sand fly species is correlated to the extension of the forest cover range. Therefore, a more comprehensive sand fly survey from the Sooretama Biological Reserve is necessary, and more accurate methods are necessary to analyze the similarity among the conservation units of the state of Espírito Santo. In this work, we presented aspects of the diversity of sand flies
June 2015
in an Atlantic Forest conservation unit in the state of Espírito Santo and pointed out preferences of some species for the peridomiciliary environment. We also discussed aspects concerning the risk of transmission of Leishmania parasites in the area and, finally, we raised the first hypothesis of similarity relationships among conservation units within the state of Espírito Santo. Acknowledgments We thank Claudenice Cumin, Fabrício Iglesias Valente, Fernanda Passos, Janine Santana Santos, Kênia Carolina, Márcia Baiôco, Morena Andrade Liberato, and Tainara Viguini for their help with the field work. We thank Gustavo Rocha Leite for the map, Leony Wand Del Rey, manager of the NFG, for the logistic support during the field work, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for providing a fellowship to Israel S. Pinto, and the Federal Institute of Education, Science, and Technology of Espírito Santo (IFES) for the financial support for the revision of this article. REFERENCES CITED Barretto, M.P. and J.O. Coutinho. 1940. Processos de captura, dissecação e montagem de flebótomos. Ann. Facul. Med. de São Paulo 16: 173–187. Barros, G.C., P.A. Sessa, E.A. Mattos, V.R.D. Carias, W. Mayrink, J.T.A. Alencar, A. Falqueto, and A.C. Jesus. 1985. Foco de leishmaniose tegumentar nos municípios de Viana e Cariacica, estado do Espírito Santo, Brasil. Rev. Saúde Públ. 19: 146–153. Cortez A.M., V.P.M. Silva, P.V.S. Queiroz, H.T.A. Andrade, M.I.B. Loiola, and M.F.F.M. Ximenes. 2007. Vertical stratification and development aspects of phlebotomine sand flies (Diptera, Psychodidae) in an area of Atlantic Forest tree species in a metropolitan region in northeastern Brazil. J. Vector Ecol. 32: 336-341. Costa, L.P., Y.L.R. Leite, G.A.B. Fonseca, and M.T. Fonseca. 2000. Biogeography of South American forest mammals: endemism and diversity in the Atlantic Forest. Biotropica 32: 872-881. Crovello, T.J. 1981. Quantitative biogeography: an overview. Taxon 30: 563–575. Dias-Lima A., E.C. Bermúdez, J.F. Medeiros, and I. Sherlock. 2002. Estratificação vertical da fauna de flebótomos (Diptera: Psychodidae) numa floresta primária de terra firme da Amazônia Central, Estado do Amazonas, Brasil. Cad. Saúde Públ. 18: 823-832. Falqueto, A., P.A. Sessa, J.B.M. Varejão, G.C. Barros, H. Momen, and G. Grimaldi Jr. 1991. Leishmaniasis due to Leishmania braziliensis in Espírito Santo state, Brazil: further evidence on the role of dogs as a reservoir of infection for humans. Mem. Inst. Oswaldo Cruz 86: 499-500. Ferreira, A.L., P.A. Sessa, J.B.M Varejão, and A. Falqueto. 2001. Distribution of sand flies (Diptera: Psychodidae) at different altitudes in an endemic region of american cutaneous leishmaniasis in the state of Espírito Santo, Brazil. Mem. Inst. Oswaldo Cruz 96: 1061–1067. Ferreira A.L., A. Falqueto, G. Grimaldi, A.A. Peixoto, and I.S. Pinto. 2013. Ecological and epidemiological aspects of
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