INTRODUCTION
Leishmaniasis, a sandfly-borne disease, is endemic in different parts of Iran. Around 20,000 cases of leish- maniases are estimated to occur annually in three differ- ent epidemiological forms, i.e. two cutaneous forms, viz.
anthroponotic cutaneous leishmaniasis (ACL) and zoonotic cutaneous leishmaniasis (ZCL), and one visceral form; viz. zoonotic visceral leishmaniasis (ZVL)1. Accord- ing to a report by the World Health Organization, Islamic Republic of Iran has been designated as one of the six countries in which more than two-thirds of new cases of cutaneous leishmaniasis (CL) occur2. ZCL is endemic in rural settings of 17 out of 31 provinces of the country1, and the endemicity within these areas extend from
hypoendemic to hyperendemic foci. ZCL is becoming alarming both in the context of geographical and statisti- cal characteristics. An officially published data has dis- closed that in a 12-yr period (1998–2009) the ZCL cases have increased from 18,560 to 24,5863. Also, the inci- dence of CL in different provinces of Iran during 2001–
2011 has been computed to be 0.002 to 1.337 per 1000 population4.
Golestan province, located in southeastern border of the Caspian Sea and bordered by Turkmenistan to the north, is recognized as a hypoendemic focus of ZCL5. Over the past decade, 500 to 1780 ZCL cases have been reported from the province every year (Golestan Health Center, unpublished data), and the incidence of the dis- ease during the year 2010–12 were calculated as 55.2,
Ecoepidemiological characteristics of a hypoendemic focus of zoonotic cutaneous leishmaniasis in north Iran (southeast of Caspian Sea)
M.D. Agh-Atabay
1, A. Sofizadeh
2, G.M. Ozbaki
3, N. Malaki-Ravasan
4, M.R. Ghanbari
5&
O. Mozafari
61Department of Environment, Golestan Head Office, Gorgan; 2Infectious Diseases Research Center, Golestan University of Medical Sciences, Gorgan; 3Gonbad-e kavoos Health Center, Golestan University of Medical Sciences, Gorgan; 4Malaria and Vector Research Group, Biotechnology Research Center, Pasteur Institute of Iran, Tehran; 5School of Public Health, Golestan University of Medical Sciences, Gorgan;
6Department of Traditional Persian Medicine, School of Medicine, Mazandaran University of Medical Sciences, Sari, Iran
ABSTRACT
Background & objectives: Leishmaniasis is an endemic parasitic disease reported sporadically throughout Iran.
Golestan province is one of the 17 zoonotic cutaneous leishmaniasis (ZCL) foci. In this study, ecoepidemiological characteristics of ZCL were investigated in Gonbad-e Kavus County, Golestan province of Iran.
Methods: This cross-sectional study was carried out in three endemic villages and one ZCL-free village by exploiting a stratified random sampling methodology in 2013. To understand particular time-allocation and activity patterns, sandflies were entrapped twice per month using sticky traps from May to October 2013. Records of daily maximum and minimum temperatures (°C) obtained from the meteorological station were used to calculate accumulated degree days (ADD) for Phlebotomus papatasi sandflies using an online software, viz. degree-day calculator, available from the website of Integrated Pest Management, University of California. The computational ADD and expected stage appearance for P. papatasi sandflies were evaluated along with the phenological events. Human infection indices were investigated in the study areas as well.
Results: In the entomological survey, a total of 13 different sandfly species were identified. Sergentomyia sintoni (66.8%) and P. papatasi (20%) were observed to be the most dominant species. Monthly activity of the different species of sandflies started from early May to mid-October, with two peaks observed in early July and mid- September, and the peak of hourly activity occurred at night (1900–2000 hrs). According to the phenological study, population dynamics of P. papatasi was completely in agreement with ADD required for the various stages of their growth. Epidemiologically, the highest incidence of ZCL was observed in the age group of 30–34 yr (29.4%).
Interpretation & conclusion: Higher incidence of the disease was seen in the northern and the northeastern parts of the province bordered by Turkmenistan. The most effective alternative for controlling the disease in this hypo- endemic focus is health education regarding personal protection from infected P. papatasi. The long-term studies are imperative for better understanding and estimation of the factors affecting ZCL transmission in the endemic areas.
Key words Golestan province; Iran; phenology; Phlebotomus papatasi; zoonotic cutaneous leishmaniasis
98.2 and 38.2 per 100,000 population respectively6. The northern and northeastern areas of the province, especially the Gonbad-e Kavus County, are important hotspots for the disease, and inhabitants of these areas are readily at risk of the bites of infected sandflies7. Within the same period (2010–12), the number of new cases in Gonbad-e Kavus County were reported to be 616, 1168 and 376 respectively, with ZCL incidence per 100,000 population as 199.9 (2010), 335 (2011) and 110.23 (2012)6.
Previous epidemiological studies conducted in this province have indicated that ZCL is caused by the kinetoplastid parasite Leishmania major, transmitted mainly by the bite of Phlebotomus papatasi Scopoli (Diptera: Psychodidae), whereas Rhombomys opimus (Great gerbil) and Meriones libycus (Libyan jird) serve as the main and secondary mammalian reservoir hosts respectively5, 8–9.
Gonbad-e Kavus County has six districts and 250 villages; out of which three districts and 53 villages are located in ZCL endemic areas. In each of the districts, there is one center for the treatment of CL, supervised by the physicians. Additionally, for effective preventive and control measures, rodenticides are distributed across ev- ery kilometer within each village.
Overall, 44 sandfly species, including 26 Phleboto- mus and 18 Sergentomyia have been reported in previous studies in Iran. Among them, natural leptomonad infec- tions have been reported in 13 species (10 Phlebotomus and three Sergentomyia)1.
Geographical distribution and evolution of the vec- tors depend mainly on climatic factors such as air tem- perature, relative humidity and rainfall. Relatively, air temperature plays a crucial role than the other factors, and this highly influences the metabolic rate, oviposition and survival (longevity) of immature and mature stages of the vectors10–12. The development of poikilothermic insects follows a predictable process based on heat13. The amount of heat needed for the growth and development of insects is predictable and quantifiable, and is measured in terms of Degree-day. Degree-day is a measure of the amount of heat that accumulates above a specified tem- perature during a 24 h period14.
Modeling and simulation of vector population dynam- ics and the environmental factors affecting them, enable scientists to predict and monitor the epidemiological pa- rameters of a putative vector-borne disease. Parameters such as development time, the number of generations per year, the potential ability of population increment, distribution along temperature gradients and the probable outbreak of a disease can be inferred from these models12, 15–16.
There is persistent risk of ZCL infection in the north- ern and northeastern parts of Golestan province. This is attributed to climatic factors and hypoendemic identity of the Golestan foci. Accordingly, periodic surveillance of ZCL in these foci is essential. In this study, ecoepidemiolo-gical characteristics of ZCL were investi- gated in Gonbad-e Kavus County, Golestan province in the year 2013.
MATERIAL & METHODS Study area
A cross-sectional study was conducted in the four ran- domly selected villages—Oochghoei (55°14'42.905"E 7°44'37.749"N); Fadavi (54°4958.501"E 37°34' 23.899"N); Gharemakher (54°58'6.003"E 37°31' 45.313"N); and Abozar (54°56'56.484"E 37°15' 38.647"N) of Gonbad-e Kavus County, Golestan province, Iran from May to October 2013. The Golestan province comprises a variety climatic zones spanning from sub-temperate to arid conditions, covering areas like deserts, forests and sea bor- der. The major landform characterized by the province are plains, and >2/3 of the arid and semiarid climates, and the remaining 1/3 have a mild climatic condition. It is bordered by the Caspian Sea and Mazandaran province to the west, Semnan province to the south, North Khorasan province to the east and Turkmenistan to the north (Fig. 1). The Gonbad-e Kavus County comprising an area of 5071.32 km2 and a population of 3,32,940 is located in the north- eastern part of the Golestan province. The average maxi- mum and minimum temperature reported are 31 and 8.3 °C respectively, and the mean annual relative
Fig. 1:Districts map of Golestan province, Iran showing the study villages in Gonbad-e Kavus County.
humidity is 65%, with a minimum of 53% in June and maxi- mum of 76% in February. The average annual rainfall is 772 mm established between February (maximum) and August (minimum). The main agricultural products are wheat, barley, water melon, sunflower, rice and cotton.
Sandfly collection
This entomological study was performed in three endemic villages (Oochghoei, Fadavi, and Gharemakher) and one ZCL-free village (Abozar) by utilizing the strati- fied random sampling from May to October 2013.
Sandflies were collected using 60 sticky traps (castor oil- coated white papers 20 × 30 cm) from indoors and out- doors (60 traps/village) in three replicates and twice per month. The traps were placed set before sunset and gath- ered before sunrise (1700–0700 hrs). Trapped sandflies were removed from sticky papers with needles, immersed in acetone to remove castor oil, and stored in microtubes containing 70% ethanol.
In total, six villages (Dashlibooron, Korand, Gharemakher, Oochghoei, Egriboghaz and Haji Ghoshan) in different parts of the Gonbad-e Kavus were selected for the investigation of the sandflies’ hourly activities. In each village, both indoors and outdoors, 10 sticky traps were set at dusk, and were replaced by new ones every hour until the next morning. The number of trapped sandflies was totaled and the subsequent data was recorded. The samples were mounted with Pouri’s solution on glass slides for species identification using specific morphological keys17. Comparing population dynamics and accumulated degree days (ADD) of P. papatasi
The relationship between population dynamics and accumulated degree days (ADD) of P. papatasi was evalu- ated. In the study, ADD needed for each life stage of P.
papatasi sandflies were calculated under laboratory con- ditions (26 ± 1°C and 80% RH)12.
Corresponding, ADDs were introduced to each egg, larval and pupal stages as 86.4, 154.75 and 142.29 de- gree days (DDs) respectively. For the pre-oviposition period, the calculated ADD ranged from 28.75 to 109.25 DD, for which the upper bound (109.25 DD) was consid- ered in the study. Maximum developmental threshold used in this study was 35°C and the minimum developmental thresholds (DZ) for each four sequential developmental stages of P. papatasi (11.60, 19.81, 17.63 and 20.25°C respectively) was adopted from Kasap and Alten18. Up- per and lower (DZ) temperatures define the thresholds at which no growth or development occurs in the species of interest. In the Iranian condition, sandflies overwinter in the stage of the IV larval instar17, hence, the IV larval
instar was considered as the starting point of growth for the first generation in the region, and 39 ADD during early spring is necessary to complete their development to pupae12. Putting these data together, records of daily maximum and minimum temperatures (°C) collected from the Gonbad-e Kavus Meteorological Station were used in calculation of ADD for P. papatasi using an online software, viz. degree-day calculator, available from the website of Integrated Pest Management, University of California. This was preformed to acquire the ADD re- quirements of each developmental stage and to compare them with the population dynamics (such as monthly ac- tivity) of P. papatasi. In the online software, horizontal cut-off and single triangle models were selected for de- fining the upper temperature threshold.
Studies on human infection
The epidemiological study was performed in the same four study villages. For the analysis of human infection indices, 100 households from each village (total 400 households) were selected. With the knowledge and con- sent of households, in total 1799 people were visited and were questioned and examined for the presence of ulcer(s) or scar(s). For each case, a form was filled to record the necessary information such as name, address, age, sex, number of ulcers or scars, site of ulcers or scars, date and place of acquiring the disease, etc. All participants were visited in June, September, December and March months of 2013. Ethical approval for the study was obtained by the Ethics Committee of Gorgan University of Medical Sciences, Iran and signed consents were obtained from the participants prior to commencing the survey.
The SPSS software version 16 was employed for the calculation of prevalence and incidence indices for the different ages and villages. Since, ZCL lead to the devel- opment of stable immunity, the incidence of the disease was calculated without the inclusion of the individuals with scars.
RESULTS Entomological findings
A total of 2114 specimens (13 sandfly species, in- cluding seven Phlebotomus and six Sergentomyia) were collected from four study villages. Frequency, sex ratio, and species composition of the sandflies are shown in Table 1. Sergentomyia sintoni was the most dominant species (66.8%) and P. sergenti, S. tiberiadis and S. hodgsoni were the least abundant species (each com- prising 0.05%). The compositions of other species were P. papatasi (20%), P. mongolensis (5.8%), P. caucasicus
(2.1%), P. caucasicus group (including females of P.
mongolensis, P. caucasicus and P. andrejivi) (3.5%), P.
kazeruni (0.3%), P. brevis (0.09%), S. clydei (0.7%) and S. grekovi (0.3%), and S. antenatal (0.2%).
Sex ratio (male : female) varied from 1.17 for P. papatasi to 0.26 for S. sintoni (Table 2). There was no significant difference between seasonal prevalence pat- terns of sandflies in the four studied areas. The popula- tion of sandflies started increasing from early May, and continued till mid-October, with two peaks observed dur- ing early July and mid-September 2013 (Fig. 2). The peak activity period of the sandflies occurred at night, from 1900–2000 hrs (Table 3).
Phenological computations in this study, revealed that the optimum temperature needed for the development of P. papatasi was 193 days in a year and during this time, closely eight DDs were received daily by the sandflies out of the total DD (1478.07/193 = 7.66 DD/day). Com- putation of ADD for each generation (492.69 DD) con- firmed that P. papatasi were multivoltine, i.e. they pro- duced at least two to three generations per year. Table 4 illustrates the first occurrence of all developmental stages and general activities of P. papatasi, and this was based on DDs that have accumulated from the initial capture of sandflies during spring. It also provides the average num- ber of developmental days required to complete the given stage. Variation between generations does not occur.
Epidemiological characteristics of the ZCL
Studies on human infection: The prevalence and in- cidence of ZCL was calculated among 1799 residents in different study villages of the Gonbad-e Kavus County.
Fig. 2: Number of collected sandflies (seasonal abundance) in different months of 2013 study villages of Gonbad-e Kavus County, Iran.
Table 1. Sandfly specimens caught from the study areas of Gonbad-e Kavus, Iran in 2013 VillagesP. papatasiP. mongo-P. cauca-P. cauca-P. kaz-P. ser-P. brevisS. sintoniS. clydeiS. grekoviS. ant-S. tiber-S. hodg-Total lensissicussicus grouperunigentienataiadissoni MFMFMFMFMFMFMFMFMFMFMFMFMFMF Fadavi124119201003000000215100500010000003421128 Oochghoei62109037006506102038652011100116888 Gharemakher391311070050000004326403121000010946 Abozar60610000000000003280000000000092141 Total22919512204500730610202931119105523210017111403 Grand total424 (20)122 (5.8)45 (2.1)73 (3.5)6 (0.3)1 (0.05)2 (0.09)1412 (66.8)15 (0.7)7 (0.3)5 (0.2)1 (0.05)1 (0.05)2114 (100) (M+F) M–Male; F–Female; Figures in parentheses indicate percentages.
Table 2. Species of sandflies caught from indoor and outdoor locations in the Gonbad-e Kavus County, Iran in 2013
Species Indoor Outdoor Total Grand total
Male Female Male Female Male Female
P. papatasi 110 (52.4) 100 (47.6) 119 (55.6) 95 (44.4) 229 (54) 195 (46) 424 (20)
P. mongolensis 49 (100) 0 (0) 73 (100) 0 (0) 122 (100) 0 (0) 122 (5.8)
P. caucasicus 23 (100) 0 (0) 22 (100) 0 (0) 45 (100) 0 (0) 45 (2.1)
P. caucasicus group 0 (0) 28 (100) 0 (0) 45 (100) 0 (0) 73 (100) 73 (3.4)
P. sergenti 1 (100) 0 (0) 0 (0) 0 (0) 1 (100) 0 (0) 1 (0.05)
P. brevis 2 (100) 0 (0) 0 (0) 0 (0) 2 (100) 0 (0) 2 (0.09)
P. kazeruni 0 (0) 1 (100) 0 (0) 5 (100) 0 (0) 6 (100) 6 (0.3)
S. sintoni 155 (18.9) 666 (81.1) 138 (23.4) 453 (76.6) 293 (20.7) 1119 (79.3) 1412 (66.8)
S. clydei 2 (50) 2 (50) 8 (72.7) 3 (27.3) 10 (66.7) 5 (33.3) 15 (0.7)
S. hodgsoni 0 (0) 1 (100) 0 (0) 0 (0) 0 (0) 1 (100) 1 (0.05)
S. grekovi 3 (75) 1 (25) 2 (66.7) 1 (33.3) 5 (71.4) 2 (28.6) 7 (0.3)
S. antenata 1 (50) 1 (50) 2 (66.7) 1 (33.3) 3 (60) 2 (40) 5 (0.2)
S. tiberiadis 0 (0) 0 (0) 1 (100) 0 (0) 1 (100) 0 (0) 1 (0.05)
Total 346 (30.2) 800 (69.8) 365 (37.7) 603 (62.3) 711 (33.6) 1403 (66.4) 2114 (100)
Figures in parentheses indicate percentages.
Table 3. Different species and number of captured sandflies in different hours of the night in the Gonbad-e Kavus County of Iran
Species Time (hrs) Total
1800– 1900– 2000– 2100– 2200– 2300– 2400– 0100– 0200– 0300– 0400– 0500–
1900 2000 2100 2200 2300 2400 0100 0200 0300 0400 0500 0600
P. papatasi 2 12 8 2 17 4 3 11 1 1 6 8 75
P. mongolensis 0 0 3 4 4 1 0 0 0 0 1 3 16
P. caucasicus 0 0 3 0 2 1 0 0 0 0 0 0 6
P. caucasicus group 0 2 0 0 3 0 0 0 0 0 0 1 6
P. kazeruni 0 0 0 0 0 0 0 0 0 0 0 1 1
S. sintoni 1 48 11 5 13 2 0 8 0 0 2 5 95
S. clydei 0 0 1 0 0 0 0 0 0 0 1 0 2
Total 3 62 26 11 39 8 3 19 1 1 10 18 201
Mean temperature (°C) 34 30.7 29.3 28 26.3 25.3 24.7 24.2 23.7 23.8 23.3 22.5
Relative humidity (%) 36.2 42.7 47 50.3 54.3 61.8 63.7 63.8 64.3 63.2 64.3 66
Table 4. The computational ADD and general activity of P. papatasi sandflies
Stage Computational data ADD Expected stage appearance
Temperature Lower Upper Starting Completion Days needed to
range (°C) threshold (°C) threshold (°C) date date complete
the stage
Larva 2 – 36 19.81 35 39 Mar 22, 2013 Apr 14, 2013 24
Pupa 12.4 – 34.8 17.63 35 142.29 Apr 15, 2013 May 13, 2013 29
Pre-oviposition 16.6 – 39.4 20.25 35 109.25 May 14, 2013 May 30, 2013 12
Egg 18.8 – 40.6 11.6 35 86.40 May 31, 2013 Jun 5, 2013 6
1st generation 376.94 71
Larva 18.6 – 40.8 19.81 35 154.75 Jun 6, 2013 Jun 23, 2013 18
Pupa 21.6 – 38.4 17.63 35 142.29 Jun 24, 2013 Jul 5, 2013 13
Pre-oviposition 23.8 – 36.6 20.25 35 109.25 Jul 6, 2013 Jul 16, 2013 11
Egg 23 – 39.2 11.6 35 86.40 Jul 17, 2013 Jul 22, 2013 5
2nd generation 492.69 47
Larva 23 – 38 19.81 35 154.75 Jul 23, 2013 Aug 5, 2013 14
Pupa 25 – 41.6 17.63 35 142.29 Aug 6, 2013 Aug 26, 2013 21
Pre-oviposition 22.8 – 41.4 20.25 35 109.25 Aug 27, 2013 Sep 7, 2013 11
Egg 21 – 42 11.6 35 86.40 Sep 8, 2013 Sep 13, 2013 5
3rd generation 492.69 51
Larva 16.4–37 19.81 35 115.75 Sep 14, 2013 Oct 6, 2013 24
Total 2 – 42 17.51 35 1478.07 193
ADD—Accumulated degree days.
The incidence and prevalence of ZCL varied from 9.3 to 33.6 and from 509.3 to 718.7 per 1000 populations in endemic areas of the county respectively. The number of cases with scar and sores were 845 and 11 respectively, and these were evident on the trunk, foot, face and hand.
Scar wound were found in two peoples in the Abozar ZCL-free village. Detailed information on human infec- tions is presented in Table 5. Demographic information according to the various classified age groups has been given in Table 6. Reported cases to Gonbad-e Kavus Health Center showed that the incidence of the disease in Korand district was higher than in other districts in 2013 (Fig. 3).
DISCUSSION
In this study, we have described the entomological, phenological and epidemiological characteristics of the hypoendemic focus of ZCL in Gonbad-e Kavoos County, Golestan province of Iran during 2013.
In total, 13 species of sandflies were captured from indoors (54%) and outdoors (46%). The species compo- sition is similar to that reported in previous studies19–22, except for S. antennata which was observed for first time
in the county. Both P. papatasi and S. sintoni were found Fig. 3:The incidence of cutaneous leishmaniasis in districts of the Gonbad-e Kavus County, Iran in 2013.
Table 5. Epidemiological characteristics of cutaneous leishmaniasis in four study villages of the Gonbad-e Kavus County, Iran Screened People Location of scars People Location of sores Incidence Prevalence Village people scarswith > One Trunk Foot Hand Face with Trunk Foot Hand Face (1/1000) (1/1000)
organ sores
Oochghoei 432 218 38 60 39 38 43 2 2 0 0 0 9.3 509.3
Fadavi 487 346 42 17 103 79 105 4 0 1 2 1 28.4 718.7
Korand 428 279 78 125 24 15 37 5 2 1 1 1 33.6 705.6
Total of endemic area 1347 843 158 202 166 132 185 11 4 2 3 2 21.8 634
Abozar 452 2 0 0 0 2 0 0 0 0 0 0 0 4.4
Total 1799 845 158 202 166 134 185 11 4 2 3 2 11.5 475.8
Table 6. Incidence and prevalence of ZCL in different age groups in the Gonbad-e Kavus County, Iran
Age Screened People with Age-adjusted People with Age-adjusted Prevalence Incidence per
(yr) people sores incidence scars prevalence per 1000 1000 people
people
0–4 190 1 9.1 36 4.3 194.7 6.5
5–9 182 3 27.3 61 7.2 351.6 24.8
10–14 172 3 27.3 62 7.3 377.9 27.3
15–19 201 0 0 91 10.7 452.7 0
20–24 228 0 0 124 14.7 543.8 0
25–29 178 1 9.1 111 13.1 629.2 14.9
30–34 156 2 18.2 88 10.4 576.9 29.4
35–39 142 0 0 86 10.2 605.6 0
40 350 1 9.1 186 22 534.3 3.05
Total 1799 11 100 845 100 475.8 11.53
to be the dominant species in the present study which is in agreement with several previous studies completed in Iran20–21, 23–24, as well as in other countries25. In some studies conducted in the Kalaleh21; and Aliabad Katool22 counties, no difference in the density of these two spe- cies was reported, however, in the present study, the den- sity of S. sintoni was more than that of P. papatasi. Like- wise, S. sintoni was commonly occurring in the indoor and outdoor places (Table 2). Phlebotomus papatasi, how- ever, has been reported as a proven vector of ZCL in the Golestan province5. Sergentomyia sintoni, which has very wide distribution across Central Asia, Iraq, Afghanistan and Iran, has been described recently as the vector of liz- ard leishmaniasis in Iran and Afghanistan17, 19, 21.
Sergentomyia antennata, which had been previously reported from western and southwestern localities of the country and from the Sistan and Baluchestan and Yazd provinces17,has been observed for the first time from Golestan province in this study.
Sergentomyia hodgsoni, which had formerly been reported from Sistan and Baluchestan, West Azerbaijan17 and Aliabad county in Golestan province22, was found to be less abundant in this study.
Species composition of the captured sandflies indi- cated that their diversity varied across the different land- forms, e.g. Fadavi village which is located in the plain region, showed less species diversity despite of more specimens collected, while in the mountainous villages such as Oochghoei and Gharemakher, the species diver- sity was comparatively higher even the numbers of speci- mens collected were less17. In the Abozar village (a re- portedly ZCL-free region) of Gonbad-e Kavus, both the number of sandflies and species diversity were low. Phle- botomus sergenti, S. tiberiadis, S. hodgsoni, P. kazeruni, P. brevis, S. clydei and S. antenata were the species col- lected only in the mountainous villages of Oochghoei and Gharemakher.
Comparison of species composition (especially P.
papatasi) with the ADD required for sandflies develop- ment showed that the sandflies overwintered as IV instar larvae. The ADD needed for the larvae to evade from diapause and pupate was 39 DD (during mid-April)12. These pupae would receive 142.29 ADD to emerge as adults. The first adult sandflies can be seen in early May within the region. They then received an amount of 109.25 ADD needed for pre-oviposition during late May.
Eggs required 86.4 ADD to hatch in early June. Lar- vae received the amount of 154.75 ADD by the end of June to pupate. Adults emerged in early July, as indi- cated in the chart showing the monthly activity of sandflies (Fig. 2/Table 4). The first peak of activity of sandflies
matched the calculated ADD in early July. Adult sandflies laid eggs by the end of July and larval-pupal duration passed through mid-August. The second peak of activity occurred at the end of August or early September in the region. The second peak of activity of sandflies in, Oochghoei and Gharemakher occurred in early Septem- ber, while in Fadavi, and Abozar, it was delayed by a few days in mid-September. This delay was negligible due to the conditions of sampling and the distance between the villages and the meteorological station. After receiving adequate ADD, the sandflies laid eggs which were trans- formed into larvae in late September. The larvae hurtled the cool weather and hibernated at this stage. Live adults of second generation disappeared from the region in early October.
Several earlier studies have indicated similar phenom- enon across the Iran. In a study by Bagheri et al22 in Aliabad Katool County in Golestan province, sandfly ac- tivity began in early June and continued until mid-Octo- ber with peak activity in August. Similarly, in a study by Oshaghi et al12, the monthly activity of sandflies began in mid-June and continued till late September with peak activity in mid-August.
Although, population diversity26 and environmental dissimilarities made the extended degree-day accumula- tions less accurate in forecasting the developmental stages of P. papatasi, its population dynamics remained compatible with amounts of ADD received by different generations12.
In a study conducted in the Jask county, sandflies activity started in late March which continued up to late December with two activity peaks, the first peak during May to June and the second peak from October to No- vember27. In another study conducted in the Bushehr County, activity of sandflies began in the first half of April and continued till the first half of December, with one activity peak during the first half of July28.
These disagreements may be due to topology and weather conditions in the study area. In the southern ar- eas of the country (warmer climates), the length of sea- son and number of peak activity are longer and more than in the northern parts like Golestan province.
Overnight sticky trap collections of sandflies revealed that the first and second peak of activities occurred between 1900–2000 and 2200–2300 hrs respectively. Mean tem- perature of 31.7 and 26.3°C and relative humidity of 32.6 and 50.3% were recorded during this period respectively.
Sandfly activities reached zero at 0200–0400 hrs, which was later observed at a low level from 0400–0700 hrs.
Zoonotic cutaneous leishmaniasis is an endemic dis- ease in the Gonbad-e Kavus County and many people are
infected in the region annually29. During the period from 2010–12, the incidence rates of ZCL in the Gonbad-e Kavus County were 199.9, 335 and 110.23 per 100,000 populations respectively. The incidence of the disease in the study area increased from 55.2 in 2010 to 98.2 in 2011, and in Ramian County, it increased from 43.2 in 2009 to 92.2 in 2011 per 100,000 population6.
Proportion of acute ulcer was observed to be 9.1% in children <1 yr, 27.3% in children <7 yr and 36.4% in children >15 yr of age. The highest incidence of ZCL was observed in the age group of 30–34 yr (29.4%), indicating the hypoendemic characteristic of the Gonbad-e Kavus focus, which was in agreement with another study conducted in the Maraveh Tappeh of Golestan province30.
In the present study, highest scar rate was observed in the age group of 20–24 yr (14.7%) which differs from that reported in Haji Abad, Hormozgan province24 among the children of 10–14 yr (15%), and in Kashan County29 among 39–30 yr (21.5%) age group. Similar to our find- ings, the results of earlier studies carried out in Damghan, Kermanshah, Kashan and Gorgan counties reported the age group of 20–29 yr with maximum sores31–34.
Nosogeography of ZCL in Gonbad-e Kavus County indicated that the incidence is higher in Northern and Northeastern regions of the county (Korand and Atrak districts), which are bordered by Turkmenistan, rather than districts which are located in the west and south of the county.
According to a study conducted in Golestan prov- ince, ZCL incidences tend to be more prevalent in areas with high temperature, low relative humidity, low rain- fall, high evaporation rate and lower number of rainy days35. The reason could be further attributed to the Ac- cess to high quality of living conditions leading to the increase of rodent (Rhombomys opimus and Meriones libycus) serving as reservoirs of the disease in these areas.
ZCL wounds are generally seen on the hands and feet34. The results of this study showed that 43.7% of patients had wounds on their hands and feet (Totally, 70%
of patients have wounds in bare parts of body of skin).
Due to the shortened mouthpart appendages, sandflies are not capable of sucking blood through clothes, hence, ex- posed areas of the body are more vulnerable for attack, which was in agreement with other studies17, 32, 34, 36–37.
CONCLUSION
The epidemiological data generated through the study emphasize on the hypoendemic identity of the Gonbad-e
Kavus foci. However, the long-term studies are impera- tive for further evolutions of the factors affecting ZCL transmission in the studied areas. The best way to control the disease in areas with higher incidence, especially sur- rounding the borders with Turkmenistan, is health edu- cation for personal protection from infected P. papatasi.
Conflict of interest
The authors declare no conflict of interest with re- gards to this communication.
ACKNOWLEDGEMENTS
This study was the result of research projects approved by the Gonbad-e Kavus Research Center and Infectious Diseases Research Center of the Golestan University of Medical Sciences, Gorgan, Iran. We appreciate and are grateful to all officials and colleagues of the two Centers for financial support, guidance, and their valuable coop- eration.
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E-mail: a_sofizadeh@yahoo.com
Received: 28 January 2016 Accepted in revised form: 28 June 2016
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