Focus Of Zoonotic Cutaneous Leishmaniasis Research Articles

Novel identification of Leishmania major in Hemiechinus auritus and molecular detection of this parasite in Meriones libycus from an important foci of zoonotic cutaneous leishmaniasis in Iran

One of the well-known foci of zoonotic cutaneous leishmaniasis (ZCL) in Iran is Turkemen Sahara, which is located in north eastern Iran. ZCL is a disease of mammals, and humans can become infected as accidental hosts. Many researchers have argued that Rhombomys opimus is the only main reservoir host of ZCL in this region of the Golestan province. No other rodents or mammals are thought to host or have been reported to host Leishmania parasites in this region. This research was designed and developed to isolate, detect and firmly identify Leishmania parasites in mammals and rodents other than R. opimus. Wild mammals were caught from gerbil burrows. Leishmania parasites were detected to assess the infection of reservoir hosts in 2010. Each genomic DNA sample was screened for Leishmania infection via nested PCR and sequencing using the internal transcribed spacer ribosomal DNA (ITS-rDNA) identification protocol for parasites. The greatest number of infections (8/19) were found in Meriones libycus. One in three infections was found in Hemiechinus auritus, and this is the first report of infection in this species. Only Leishmania major was definitively identified and unambiguously typed in M. libycus and H. auritus. The infection rates in these two wild mammals were not significantly different, and no other gerbil parasites were detected in M. libycus or H. auritus at our study site. Recent findings of Leishmania turanica in R. opimus and failures to detect L. turanica in M. libycus may be attributable to unidentified Leishmania infections in two M. libycus due to unreadable sequences. These cases may represent mixed infections by L. major and L. turanica. The assumptions that gerbil parasites can be co-infectors provide a starting point for the identification of the causative and potential parasites responsible for the frequent infections that are mainly mediated via sandfly vectors.

High density of Leishmania major and rarity of other mammals' Leishmania in zoonotic cutaneous leishmaniasis foci, Iran

Only Leishmania major is well known as a causative agent of zoonotic cutaneous leishmaniasis (ZCL) in Iran. Our objective was to find Leishmania parasites circulating in reservoir hosts, sand flies and human simultaneously. Sand flies, rodents and prepared smears of humans were sampled. DNA of Leishmania parasites was extracted, and two fragments of ITS-rDNA gene amplified by PCR. RFLP and sequencing were employed to identify Leishmania parasites. Leishmania major and L. turanica were identified unequivocally by targeting and sequencing ITS-rDNA from humans, rodents and sand flies. The new Leishmania species close to gerbilli (GenBank Accession Nos. EF413076; EF413087) was discovered only in sand flies. Based on parasite detection of ITS-rDNA in main and potential reservoir hosts and vectors and humans, we conclude that at least two Leishmania species are common in the Turkmen Sahra ZCL focus. Phylogenetic analysis proved that the new Leishmania is closely related to Leishmania mammal parasites (Leishmania major, Leishmania turanica, Leishmania gerbilli). Its role as a principal agent of ZCL is unknown because it was found only in sand flies. Our findings shed new light on the transmission cycles of several Leishmania parasites in sand flies, reservoir hosts and humans.

Laboratory diagnosis, clinical manifestations, epidemiological situation and public health importance of cutaneous leishmaniasis in Shushtar County, Southwestern Iran

ObjectiveTo investigate as a descriptive and cross sectional study on epidemiology, the laboratory diagnosis and clinical manifestation of Cutaneous Leishmaniasis (CL) in Shushtar County. MethodsThe investigation was carried out on referrals from different districts of Shushtar County and was diagnosed based on their ulcer clinical manifestations with CL using laboratory method. This study was actively developed over the period of 2008 to 2010. Surface of a clean slide was pressed on to the scraped part of the lesions and moved along to produce a smear on the slide. The slides were fixed with methanol, stained with standard Giemsa, and examined carefully under high magnification for an hour or more to look for amastigotes by a trained person. Two smears were prepared and examined from each patient and data were collected using different correlative patients characteristics to analyze information. The essential records including residential area, type and location of lesion, age, sex, the number of ulcers, season, and the history of traveling to endemic areas for this disease were carefully gathered. ResultsParasites presumed to be Leishmania major (L. major) based on the existence of a large vacuole in the cytoplasm. Totally 206 patients were referred to the central laboratory and their manifestation lesions were clinically examined. Of these, 64 cases (31.1%) were female and 142 cases (68.9%) were male. The findings documented that the rate of incidence of CL was in fall 61 cases (29/6%) and winter 80 cases (38/8%), which is one of the most important characteristics of zoonotic CL. Sixty-eight cases (33%) had ulcers on hands, 63 cases (30.6%) on feet and 30 cases (14.6%) on hands and feet. A total of 120 patients (58.3%) had only one ulcer. But 70 cases (34%) had 2 ulcers. The highest incidence was found among patients with two age groups of 20-29 years old (43.2%) and 10-19 years old (18%). The most cases resided in rural areas with 164 cases (79.6%). ConclusionsAnalyzing of the composed results revealed that this area could be considered as one of the endemic foci of zoonotic cutaneous leishmaniasis caused by L. major. In view of the high prevalence rate and distribution of leishmaniasis in Iran, it is in necessity of data gathering about vectors and reservoirs of each district for prevention. Designing control programs play prominent role in this case. With respect to the increased prevalence of CL in Shushtar districts, health care observers should pay further attention on preventing.

Molecular characterization of Leishmania spp. in reservoir hosts in endemic foci of zoonotic cutaneous leishmaniasis in Iran

Zoonotic cutaneous leishmaniasis (ZCL) is an expanding disease and a public health issue in Iran. In the present study, rate of natural infection of rodent populations with Leishmania was investigated in six endemic foci including 28 villages in Golestan, Esfahan, Yazd, Fars, Khuzestan and Ilam provinces. A total of 593 rodents were captured and identified as Rhombomys opimus (n = 325), Meriones libycus (n = 171), Meriones persicus (n = 27), Tatera indica (n = 37), Nesokia indica (n = 12), Rattus rattus (n = 13) and Mus musculus (n = 8). Microscopic examinations of Giemsa-stained smears showed that 108 out of 593 (18.2%) rodents were infected with Leishmania spp., whereas infection of 186 out of 593 (31.4%) rodents with Leishmania was then confirmed by ITS1-PCR. The highest rate of infection was found in R. opimus (prevalence of 35%) and M. libycus (31%). Based on Restriction Fragment Length Polymorphism (RFLP), 145 (78%) of 186 samples detected as Leishmania DNA were identified as L. major, 8 (4%) samples as L. turanica and 33 (18%) as mixed infection (L. major and L. turanica). Samples from infected rodents were inoculated subcutaneously at tail base of BALB/c mice. In 35 of them, nodules and ulcers containing amastigotes appeared at the inoculation site. The samples prepared from infected rodents were cultured in NNN medium and only two samples werepositive. Rhombomys opimus, M. libycus, M. persicus, T. indica and N. indica were confirmed as reservoir hosts of ZCL in the studied regions. Leishmania major infection was usually accompanied L. turanica in naturally infected gerbils (R. opimus and M. libycus) in Golestan, Esfahan and Fars provinces.

Molecular characterization of sandflies and Leishmania detection in main vector of zoonotic cutaneous leishmaniasis in Abarkouh district of Yazd province, Iran

ObjectiveTo assess molecular characterization, distribution, seasonal activities of sandfly species and Leishmania parasites infecting them for this zoonotic cutaneous leishmaniasis focus. MethodsThe collections were carried out in 2009–2011 using CDC traps, Sticky Papers and manual aspirator in and around the villages in Abarkouh district. Individual sandflies were characterized by PCR amplification and sequencing of fragments of their mitochondrial cytochrome b gene. Leishmania parasite infections within sandflies were performed by targeting Cyt b, ITS-rDNA, k-DNA and microsatellite genes. ResultsThe PCR assays detected only Leishmania major (L. major). All infections (30) were found in the abundant and widespread vector Phlebotomus papatasi (P. papatasi). Small numbers of other sandfly species were also screened for infections, but none was found. Sergentomyia sintoni and P. papatasi were the predominant members in all locations of this district and in all habitats throughout the trapping season. Only five other sandfly species were found, namely Phlebotomus ansari, Phlebotomus caucasicus, Phlebotomus sergenti, Sergentomyia dentata and Sergentomyia merviney. ConclusionsIn the current survey, the only infections detected are of L. major in females of P. papatasi (30 out of 190). The rates of infection of P. papatasi by L. major are not significantly different in compare with other locations in Iran with no diversity of parasite strains. Zoonotic cutaneous leishmaniasis may have emerged only recently in Abarkouh district, and the reason may well be the instability of the transmission cycles there.

Natural infection and phylogenetic classification of Leishmania spp. infecting Rhombomys opimus, a primary reservoir host of zoonotic cutaneous leishmaniasis in northeast Iran

In the northeast and central parts of Iran, Rhombomys opimus (great gerbil) is the primary reservoir host of zoonotic cutaneous leishmaniasis (ZCL). This study used both parasitological and molecular methods to identify Leishmania spp. and their different haplotypes that were circulating in the great gerbil populations in ZCL foci from northeastern Iran. A cross-sectional study using microscopy, culturing and molecular methods was conducted to detect Leishmania parasites in 194 live R. opimus in ZCL foci from northeastern Iran during 2010-2011. Leishmania spp. were found in 38.1% (74/194) of the samples by microscopy and in 41.2% (80/194) by culturing. Small papules and skin thickening on the upper edge of the ears were observed in 25 (12. 9%) of the R. opimus. PCR-RFLP and PCR direct sequencing of internal transcribed spacer 1 (ITS1) rRNA showed similar infection rates for L. major and L. turanica in 60 eligible R. opimus, only one mixed infection containing both L. major and L. turanica was found. Phylogenetic analysis was conducted using the ITS1 sequences of 32 isolates that were successfully aligned by comparison of their base sequences with the ITS1 DNA sequence database using ClustalW and MEGA5. The samples were classified into monophyletic clusters (>97% bootstrap). Six haplotypes were observed for L. major and seven for L. turanica. In northeast Iran, L. major, and L. turanica naturally circulate in R. opimus, and L. major/L. turanica co-infections also exist. Phylogenetic analysis suggested that Leishmania spp. isolated from R. opimus are not a monophyletic group.

Low diversity of Leishmania parasites in sandflies and the absence of the great gerbil in foci of zoonotic cutaneous leishmaniasis in Fars province, southern Iran

Fars province lies to the south of the main foci of zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania major in central and northern Iran, and its ZCL foci might have emerged more recently because of different transmission cycles. However, there have been limited studies of the parasites infecting the regional sandfly vectors. The diversities of sandflies and the Leishmania parasites infecting female sandflies were assessed for different habitats in Fars ZCL foci, 2008-2009, using standardized sandfly trapping and characterization of ribosomal DNA haplotypes of parasites. Vector diversity was similar to that in the main Iran ZCL foci, but parasite diversity was lower. Only Le. major was detected in Fars sandflies. Most infections were found in the abundant vector Phlebotomus papatasi collected in rodent burrows. Infections were detected for the first time in Iran in the related P. bergeroti. The failure to detect other gerbil parasites (Le. turanica, Le. gerbilli) is explained by the absence of one reservoir host, the great gerbil Rhombomys opimus. Other gerbils (Meriones species) may be the regional reservoir hosts, but transmission modelling is required for a better understanding of the emergence and stability of ZCL foci in Fars and other Iranian provinces.

A survey of reservoir hosts in two foci of cutaneous leishmaniasis in Kerman province, southeast of Iran.

In the Old World, cutaneous leishmaniasis (CL) is zoonoses and natural vertebrate hosts of CL parasites are mammals. This study was carried out on natural infection rates of Leishmania parasites in reservoir hosts in one new focus of zoonotic cutaneous leishmaniasis (ZCL) and in suspected reservoir in an old focus of ACL in Iran. The sampling of rodents using Sherman traps was carried out and PCR technique was used for detection and identification of Leishmania species in Bahreman district, Kerman province, southeast of Iran. In addition, the smears were taken from suspicious lesions in stray dogs in the city of Kerman, the center of Kerman province. Simultaneously, pieces of lesion (1×1×1cm) were taken for further histopathological examination. Overall, 25 rodents were collected and identified, including Meriones libycus and Rhombomys opimus. Amastigotes were observed in 33% of the R. opimus by microscopic examination and indentified as Leishmania major by PCR technique. Four suspicious dogs out of 391 stray dogs showed no Leishmania species. To the best of our knowledge, this is the first isolation and identification of L. major from R. opimus in Kerman province, where ZCL has been present in recent years. Therefore, R. opimus is considered as the main animal reservoir host in Bahreman ZCL focus. In ACL focus such as the city of Kerman, dogs had no role in CL infection as reservoir host.

Identification and frequency distribution of Leishmania (L.) major infections in sand flies from a new endemic ZCL focus in southeast Iran

Zoonotic cutaneous leishmaniasis (ZCL) is an important public health challenge in Iran. It is often caused by the protozoan parasite, Leishmania major. This pathogen is principally transmitted by the infectious bites of adult female phlebotomine sand flies which belong to the subgenus Phlebotomus (Phlebotomus) Rondani and Berte (Diptera: Psychodidae). A recent outbreak of clinical ZCL cases in the rural district of Jask, Hormozgan province, southern Iran prompted the identification of sand flies naturally infected with L. major using polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA). A total of 8,123 wild sand flies were caught using sticky paper traps indoors and outdoors in seven villages of Jask from March 2007 to February 2008. About 120 trap-nights per month were carried out during peak seasonal density. Eight sand fly species of two distinct genera were morphologically identified. The most abundant species, Phlebotomus papatasi (≈60%) and Phlebotomus salehi (17%), had simultaneous population peaks in May and October. The anthropophilic index of P. papatasi was approximately twice that of P. salehi. PCR reactivity of L. major infections in P. salehi was weaker than those in P. papatasi. This is discussed with regard to their role in the natural transmission cycle of ZCL. This study provided the first PCR and ELISA evidence on P. salehi as a vector of L. major parasites in a new endemic region of ZCL in southeast Iran.

Epidemiological status of cutaneous leishmaniasis in Behbahan County, Khuzestan Province, SW Iran

Background: Leishmaniasis is a zoonotic infection inwhichparasites of the genus Leishmania are transmitted from rodents and small mammals to Phlebotomus species sand fly vectors. In Iran at least 20000 leishmaniasis cases per year have been reported. Cutaneous leishmaniasis (CL) is a major health problem in some parts of Iran, which exists as zoonotic and anthroponotic forms in different foci of the country. In recent years new foci of zoonotic CL were found in Iran. This disease is endemic in many rural areas, including Khuzestan Province, southwestern Iran. The main vector for humans is Phlebotomus papatasi, Scopoli 1786 (Diptera: Psychodidae). P. Papatasi is the predominant sand fly. Leishmania major is a causative agent of ZCL. Rodents are the main reservoirs of this disease. This study was carried out to determine the epidemiology and the status of CL in Behbahan County. Methods: This descriptive cross section study was done in Behbahan County from 2004-2008. We confirmed suspected cases by using stained smears. Data for this study were collected by means of a questionnaire. The data was analyzed by SPSS software. Results: A total of 172 cases of CL were reported in Behbahan County in 2004 (n=24), 2005 (n=33), 2006 (n=43), 2007 (n=34) and2008 (n=38).Most of the cases (55.2%)were found in themales. In 55 cases (32%) the ulcer was located on hand and the remainder were on foot (31.4%), face (19.2%) and trunk (17.4%) respectively. Eighty four patients (48.8%) had only one ulcer. But 88 cases (51.2%) had more than one ulcer. The highest incidence was found among patients with two age group of 20-29 (27.3%) and 10-19 (26.2%). Most frequencywas found in the rural areas (52.9%). The study also showed the rate of CL incidence during these 5 years had not been fixed andfluctuated from time to time (with the general rate of 0.18 per thousand). It was also evident that the highest rate of incidence of CL was in winter, spring, fall and summer, respectively. Conclusion: With respect to the increased prevalence of CL in Behbahan, health care observers should pay further attention on preventing the disease spread.

Molecular Detection of Leishmania major kDNA from Wild Rodents in a New Focus of Zoonotic Cutaneous Leishmaniasis in an Oriental Region of Iran

Human cutaneous leishmaniasis is one of the most challenging public health issues in many tropical and subtropical countries of the world, including Iran. More than half (54%) of the new zoonotic cutaneous leishmaniasis (ZCL) cases among the Eastern Mediterranean countries were reported from Iran in 2008. The detection of Leishmania parasites in rodents is essential to incriminate them as probable reservoir hosts of ZCL infection. As a result of the annual detection of about 200-250 clinical ZCL cases in the Jask district of southern Iran, feral rodents were trapped, identified to species level, and examined for Leishmania presence by preparing routine blood smears on microscopic slides from 2007 to 2008. Overall, 27 Tatera indica, 17 Gerbillus nanus, 29 Meriones persicus, 26 M. hurrianae, and 7 M. libycus were identified. Females of T. indica, M. hurrianae, and G. nanus appeared to be naturally infected with the protozoan parasite, L. major. This is the first report of microscopic and molecular detection of this trypanosomatid parasite infecting these three rodents reported from Hormozgan province in southeast Iran. More than three-quarters (82%) of the parasite-infected rodents came from the eastern plain of this province, but none of the other rodents were found to be smear-positive or kinetoplast DNA-positive by PCR. M. hurrianae, G. nanus, and T. indica are therefore incriminated as three potential reservoir hosts of L. major in Oriental parts of Iran.

Baseline susceptibility of a wild strain of Phlebotomus papatasi (Diptera: Psychodidae) to DDT and pyrethroids in an endemic focus of zoonotic cutaneous leishmaniasis in Iran

Phelebotumus papatsi is considered to be the main vector of zoonotic cutaneous leishmaniasis as well as sand fly fever in Iran. There are several measures for vector control, with emphasis on insecticides. The objective of this study was to determine the baseline susceptibility of this vector to the commonly used insecticides in an endemic focus of the disease in central Iran. P. papatasi collected from the field were used for susceptibility status. Its baseline susceptibility to DDT and pyrethroids was assessed on about 6866 specimens collected from Badrood rural district, Esfahan Province, Iran, during the summer of 2010. The LT(50) and LT(90) values were measured according to the World Health Organisation (WHO) test using probit analysis and regression lines. Results of tests against female P. papatasi revealed LT(50) values to DDT 4%, permethrin 0.75%, deltamethrin 0.1%, cyfluthrin 0.15% and lambdacyhalothrin 0.05% of 1104.97, 182.35, 26.79, 15.42 and 1.48 s respectively. The figures for male P. papatasi were 973.51, 59.5, 4.4, 2.65 and 1.5. The results of this study provide a guideline for implementation of different vector control measures. Furthermore, guidelines are needed for monitoring and evaluation of insecticide susceptibility tests against sand flies.

First Detection of Leishmania major DNA in Sergentomyia (Spelaeomyia) darlingi from Cutaneous Leishmaniasis Foci in Mali

Background Leishmania major complex is the main causative agent of zoonotic cutaneous leishmaniasis (ZCL) in the Old World. Phlebotomus papatasi and Phlebotomus duboscqi are recognized vectors of L. major complex in Northern and Southern Sahara, respectively. In Mali, ZCL due to L. major is an emerging public health problem, with several cases reported from different parts of the country. The main objective of the present study was to identify the vectors of Leishmania major in the Bandiagara area, in Mali.Methodology/Principal FindingsAn entomological survey was carried out in the ZCL foci of Bandiagara area. Sandflies were collected using CDC miniature light traps and sticky papers. In the field, live female Phlebotomine sandflies were identified and examined for the presence of promastigotes. The remaining sandflies were identified morphologically and tested for Leishmania by PCR in the ITS2 gene. The source of blood meal of the engorged females was determined using the cyt-b sequence. Out of the 3,259 collected sandflies, 1,324 were identified morphologically, and consisted of 20 species, of which four belonged to the genus Phlebotomus and 16 to the genus Sergentomyia. Leishmania major DNA was detected by PCR in 7 of the 446 females (1.6%), specifically 2 out of 115 Phlebotomus duboscqi specimens, and 5 from 198 Sergentomyia darlingi specimens. Human DNA was detected in one blood-fed female S. darlingi positive for L. major DNA.ConclusionOur data suggest the possible involvement of P. duboscqi and potentially S. darlingi in the transmission of ZCL in Mali.

Epidemiological aspects of cutaneous leishmaniasis in Ilam province, west of Iran (2000–2007)

ObjectiveTo investigate the epidemiology of cutaneous leishmaniasis in Ilam province between 2000 and 2007. MethodsThe data of this descriptive cross-sectional study included the existing data regarding the prevalence of cutaneous leishmaniasis in different seasons, years and districts which were reported to Ilam Provincial Health center between 2000 and 2007. ResultsThe analysis of the collected data revealed that Ilam province was one of the endemic foci of zoonotic cutaneous leishmaniasis caused by Leishmania major. The study also showed the rate of cutaneous leishmaniasis incidence with the general rate of 1.2 per one thousand populations. It was also evident that the highest rate of incidence of cutaneous leishmaniasis was in fall and winter, which is one of the most important characteristics of zoonotic cutaneous leishmaniasis. Ulcers were present in 5 161 Patients. Of these, 28.9% were female and 71.1% were male. 3 447 cases (66.8%) had ulcers on hand and foot, 1 213 cases (23.5%) on face and 501 cases (9.7%) on trunk. A total of 1 907 patients (36.9%) had only one ulcer. But 3 254 cases (63.1%) had more than one ulcer. The highest incidence was found among patients with two age groups of 20–29 years old (46.8%) and 10–19 years old (17.3%). ConclusionsWith respect to the increased prevalence of cutaneous leishmaniasis in Ilam province, health care observers should pay further attention on preventing the disease spread.

Isolation and detection of Leishmania species among naturally infected Rhombomis opimus, a reservoir host of zoonotic cutaneous leishmaniasis in Turkemen Sahara, North East of Iran

In Iran, three species of Leishmania have been incriminated as the causative agents of human leishmaniasis, Leishmania (L.) major, Leishmania tropica, and Leishmania infantum.Rhombomis opimus have been incriminated as a principal reservoirs of the parasitic protozoan Leishmania major, the causative agent of rural zoonotic cutaneous leishmaniasis (ZCL) in Iran. Rodents captured and examined to find Leishmania species using conventional methods including direct impression smear and microscopic observation inoculation samples to Balb/c and culture in NNN medium. Also molecular method was employed to detect Leishmania in rodents by amplifying a region of the ribosomal RNA amplicon of Leishmania (ITS1-5.8S rRNA–ITS2) using Nested PCR. Leshmania species were specified by DNA sequences. 36 (38.3%) of R. opimus were Leishmania positive using at least one conventional methods. Many more ITS-rDNA fragments were amplified from R. opimus but only 65 out of 74 PCR products contained enough DNA for direct sequencing or readable sequences. The PCR assays detected in Iranian R. opimus not only Leishmania major in 59 (79.7%) rodents but also Leishmania turanica in 6 (8.1%) rodents, another parasite of the great gerbil. These parasites were found in Turkemen Sahara, North East of Iran, in a focus of rural (ZCL). L. major and L. turanica in R. opimus firmly identified from Turkemen Sahara. Nine rodents with Leishmania infections unidentified which some were unreadable sequences, these could be mixed infections of L. major, L. turanica, Leishmania gerbillisensu lato and Leishmania close to L. gerbilli or a related species reported in sandflies previously from this location. The haplotypes of L. major and L. turanica were found to be identical to that of isolates of L. major and L. turanica from Iran and in GenBank elsewhere. R. opimus is probably the key reservoir in this ZCL focus because of its abundance and its infection rates with both L. major and L. turanica.

Nested PCR, RFLP and sequencing of nuclear ITS-rDNA genes detect Leishmania species in patients from an Iranian focus of zoonotic cutaneous leishmaniasis (ZCL)

Nested PCR, RFLP and sequencing of nuclear ITS-rDNA genes detect Leishmania species in patients from an Iranian focus of zoonotic cutaneous leishmaniasis (ZCL)

Geographical distribution and ecological features of the great gerbil subspecies in the main zoonotic cutaneous leishmaniasis foci in Iran

Objective To reveal subspecies composition of Rhombomy opimus ( R. opimus) in Iran. Methods In this study, field specimens of the gerbil were collected from all its geographical rang in northeast and central parts of Iran and identified on basis of morphological characteristics. Results Results revealed presence of two subspecies of R. opimus sodalis and R. opimus sargadensis in the country. The first subspecies with brown to hazel-nut color and a size bigger than a typical R. opimus was found only in Golestan province, an isolated colony which is located in plains of north slopes of Alborz Chain Mountains of the country, connected with the Turkmenistan. The subspecies of R. opimus sargadensis with a yellowish color and similar size as a typical R. opimus was found in all other areas of the great gerbil distribution in Iran. There was a distinct topographic difference but similar ectoparasites between colonies of the two subspecies in the study area. Conclusions Further ecological and genetic investigations are required for more detailed description of the R. opimus subspecies range and structure. The R. opimus – Phlebotomus papatasi-Leishmania major association and the ZCL severity and outcome in hosts are discussed.

Leishmaniasis in the World Health Organization Eastern Mediterranean Region

Leishmaniasis represents a major public health problem in the Eastern Mediterranean Region (EMR) of the World Health Organization (WHO). Cutaneous and visceral leishmaniasis are mainly seen in 14 of the 22 countries of the region. In several of these countries outbreaks have an apparent tendency to occur at around 10-year intervals. In 2008, some 100 000 new cases of cutaneous leishmaniasis were reported. Foci of zoonotic cutaneous leishmaniasis, caused by Leishmania major, occur in Afghanistan, Egypt, Iran, Iraq, Jordan, Libya, Morocco, Palestine, Pakistan, Saudi Arabia, Sudan, Syria, Tunisia and Yemen. Anthroponotic cutaneous leishmaniasis, caused by L. tropica, occurs in Afghanistan, Iran, Iraq, Morocco, Pakistan, Saudi Arabia, Syria and Yemen. Anthroponotic visceral leishmaniasis, caused by L. donovani, occurs mainly in Sudan and Somalia. Zoonotic visceral leishmaniasis, caused by L. infantum, occurs in most countries of the region. In order to address the problem of leishmaniasis in the EMR, WHO is supporting ministries of health through a strategic plan focusing on (a) training programme managers and health workers on diagnosis and case management; (b) establishing a harmonized regional surveillance system; (c) creating a regional network of experts; (d) promoting political commitment of national governments.

Dynamics of Leishmania infection rates in Rhombomys opimus (Rodentia: Gerbillinae) population of an endemic focus of zoonotic cutaneous leishmaniasis in Iran

Zoonotic cutaneous leishmaniasis (ZCL) due to Leishmania major is a great public health problem in the Old World. Leishmania major is widely distributed in populations of rodents in arid and savannah regions. In this study, seasonal variation of natural infection with Leishmania parasites in Rhombomys opimus (Rodentia: Gerbillinae) population of an endemic focus of ZCL in Iran was monitored. The study was conducted from October 2007 to October 2008 in the central part of the country. Nested polymerase chain reaction (PCR) assay was used for the detection and identification of Leishmania parasites, and the results were confirmed by PCR-restriction fragment length polymorphism (RFLP). The results showed that Leishmania infection rate was 55.8% (29 out of 52 gerbils) using nested PCR. The highest and lowest Leishmania infection rates were observed in fall and summer, respectively. Gerbils that were found to be infected only with L. major were 5.8%, and that with Leishmania turanica were 23.1%. A mixed natural infection was seen in the rodents with L. major and L. turanica (21.2%), with L. major and L. gerbilli (1.9%), and with all the three species (3.9%). Leishmania major infection alone was seen in fall and winter whereas mixed infection of L. major and L. turanica was observed in all seasons except in summer. Leishmania turanica infection was observed throughout the year. It is concluded that L. major, L. gerbilli, and L. turanica circulate in the population of R. opimus in central part of Iran. Leishmania major infection is usually accompanied by L. turanica in naturally infected gerbils with the highest rate in fall. It is recommended that the role of L. turanica in the epidemiology and transmission of ZCL should be reconsidered.

Phlebotomus caucasicus and Phlebotomus mongolensis (Diptera: Psychodidae): indistinguishable by the mitochondrial cytochrome b gene in Iran

Diagnostic molecular markers for the females of Phlebotomus (Paraphlebotomus) caucasicus and P. mongolensis were sought by characterizing from individual Iranian specimens a gene fragment, namely mitochondrial cytochrome b, that had previously proven useful for the taxonomy of phlebotomine sandflies. Males of both species were used as reference material because their external genitalia provide the only diagnostic morphological characters. A phylogenetic analysis of the new sequences, and those previously reported for P. grimmi, found no support for recognizing more than one species (P. caucasicus s.l.) in Iran. Most of the genetic variation was geographical. An absence of lineage sorting was demonstrated, and it is proposed that any search for species-specific molecular markers for these three taxonomic species should be continued by applied biologists only if there is better evidence for associating any one of them with phenotypes important for understanding the transmission of Leishmania species in foci of zoonotic cutaneous leishmaniasis.