Discrepant Prevalence and Incidence of Leishmania Infection between Two Neighboring Villages in Central Mali Based on Leishmanin Skin Test Surveys

Apart from a single report, the last publication of cutaneous leishmaniasis (CL) in Mali dates back more than 20 years. The absence of information on the current status of CL in Mali led us to conduct a cohort study in Kemena and Sougoula, two villages in Central Mali from which cases of CL have been recently diagnosed by Mali''s reference dermatology center in Bamako. In May 2006, we determined the baseline prevalence of Leishmania infection in the two villages using the leishmanin skin test (LST). LST-negative individuals were then re-tested over two consecutive years to estimate the annual incidence of Leishmania infection. The prevalence of Leishmania infection was significantly higher in Kemena than in Sougoula (45.4% vs. 19.9%; OR: 3.36, CI: 2.66–4.18). The annual incidence of Leishmania infection was also significantly higher in Kemena (18.5% and 17% for 2007 and 2008, respectively) than in Sougoula (5.7% for both years). These data demonstrate that the risk of Leishmania infection was stable in both villages and confirm the initial observation of a significantly higher risk of infection in Kemena (OR: 3.78; CI: 2.45–6.18 in 2007; and OR: 3.36; CI: 1.95–5.8 in 2008; P<0.005). The absence of spatial clustering of LST-positive individuals in both villages indicated that transmission may be occurring anywhere within the villages. Although Kemena and Sougoula are only 5 km apart and share epidemiologic characteristics such as stable transmission and random distribution of LST-positive individuals, they differ markedly in the prevalence and annual incidence of Leishmania infection. Here we establish ongoing transmission of Leishmania in Kemena and Sougoula, Central Mali, and are currently investigating the underlying factors that may be responsible for the discrepant infection rates we observed between them.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00344084","term_id":"NCT00344084"}}NCT00344084     

Sandflies (Diptera: Psychodidae) in a focus of visceral leishmaniasis in White Nile, Sudan

Visceral leishmaniasis (VL) has been known to occur since the 1980s on the western bank of the White Nile River (Central Sudan), 150 km south of Khartoum, and has resulted in high mortality. The most recent outbreak of the disease in this area began in 2006. Entomological surveys were carried out during May 2008, June 2010 and May and July 2011 in the White Nile area. Sandflies were collected using Centers for Disease Control light traps and sticky oil traps in the village of Kadaba and the nearby woodland. Phlebotomus females were dissected for the presence of Leishmania promastigotes. A total of 17,387 sandflies, including six species of Phlebotomus and 10 species of Sergentomyia, were identified. The Phlebotomus species recorded were Phlebotomus orientalis, Phlebotomus papatasi, Phlebotomus bergeroti, Phlebotomus duboscqi, Phlebotomus rodhaini and Phlebotomus saevus. P. orientalis was collected in both habitats. The relative abundance of P. orientalis in the woodland habitat was higher than that recorded in the village habitat. In the woodland habitat, there was a notable increase in the relative abundance of P. orientalis during the surveys conducted in 2008 and 2010 compared to 2011. None of the 311 P. orientalis females dissected were infected with Leishmania promastigotes, although relatively high parous rates were recorded in both habitats. Based on the distribution of P. orientalis recorded in this study, this species is the most likely vector of VL in the endemic focus in the White Nile area. Further investigation is required to elucidate the seasonal abundance and distribution of the vector, as well as the transmission season of VL in both habitats so that appropriate control strategies for the vector can be designed.     

Incrimination of Phlebotomus kandelakii and Phlebotomus balcanicus as vectors of Leishmania infantum in Tbilisi, Georgia

A survey of potential vector sand flies was conducted in the neighboring suburban communities of Vake and Mtatsminda districts in an active focus of visceral Leishmaniasis (VL) in Tbilisi, Georgia. Using light and sticky-paper traps, 1,266 male and 1,179 female sand flies were collected during 2006-2008. Five Phlebotomus species of three subgenera were collected: Phlebotomus balcanicus Theodor and Phlebotomus halepensis Theodor of the subgenus Adlerius; Phlebotomus kandelakii Shchurenkova and Phlebotomus wenyoni Adler and Theodor of the subgenus Larroussius; Phlebotomus sergenti Perfil'ev of the subgenus Paraphlebotomus. Phlebotomus sergenti (35.1%) predominated in Vake, followed by P. kandelakii (33.5%), P. balcanicus (18.9%), P. halepensis (12.2%), and P. wenyoni (0.3%). In Mtatsminda, P. kandelakii (76.8%) comprised over three fourths of collected sand flies, followed by P. sergenti (12.6%), P. balcanicus (5.8%), P. halepensis (3.7%), and P. wenyoni (1.1%). The sand fly season in Georgia is exceptionally short beginning in early June, peaking in July and August, then declining to zero in early September. Of 659 female sand flies examined for Leishmania, 12 (1.8%) specimens without traces of blood were infected including 10 of 535 P. kandelakii (1.9%) and two of 40 P. balcanicus (5.0%). Six isolates were successfully cultured and characterized as Leishmania by PCR. Three isolates from P. kandelakii (2) and P. balcanicus (1) were further identified as L. infantum using sequence alignment of the 70 kDa heat-shock protein gene. Importantly, the sand fly isolates showed a high percent identity (99.8%-99.9%) to human and dog isolates from the same focus, incriminating the two sand fly species as vectors. Blood meal analysis showed that P. kandelakii preferentially feeds on dogs (76%) but also feeds on humans. The abundance, infection rate and feeding behavior of P. kandelakii and the infection rate in P. balcanicus establish these species as vectors in the Tbilisi VL focus.     

Molecular detection of Leishmania tropica in field caught Phlebotomus sand flies (Diptera: Psychodidae) from cutaneous leishmaniasis endemic focus of Pakistan

Cutaneous Leishmaniasis is endemic in tribal district Khyber for last more than one decade. The causative agent Leishmania tropica is known but sand fly species responsible for the transmission of disease still needs to be investigated. A total of 2647 Phlebotomus females belonging to 11 species were divided into 435 batches and subjected to PCR for detection of Leishmania in sand flies. A total of 50 batches belonging to three species i.e. Phlebotomus sergenti, Phlebotomus papatasi and Phlebotomus alexandri were detected positive for Leishmania tropica. Overall minimum infection rate was 1.89% (50/2647). Highest minimum infection rate of 2.11% (39/1710) was observed for Phlebotomus sergenti followed by 1.21% (8/661) for Phlebotomus paptasi and 1.82% (3/165) for Phlebotomus alexandri. Both blood fed (38%) and unfed (62%) sand flies were detected positive for the parasite DNA. Positive specimens were collected throughout the active season, from all collection sites of the study area. Detection of Leishmania parasite in multiple species of Phlebotomus indicates the possible role of these species as vector of disease in the tribal district Khyber of Pakistan. It also indicates the probable complex transmission cycle of the disease involving multiple vector species in the study area. Devising a control strategy by focusing on these vector species, may reduce the disease burden in the cutaneous leishmaniasis endemic tribal district Khyber.     

Seasonal Variation in the Prevalence of Sand Flies Infected with Leishmania donovani

Background

Visceral Leishmaniasis (VL) is a life threatening neglected infectious disease in the Indian subcontinent, transmitted by the bite of female sand flies. Estimation of the infectivity in the vector population, collected in different seasons, may be useful to better understanding the transmission dynamics of VL as well as to plan vector control measures.

Methodology

We collected sand flies from highly endemic regions of Bihar state, India for one year over three seasons. The species of the sand flies were confirmed by species-specific PCR-RFLP. Leishmania donovani infection was investigated in 1397 female Phlebotomus argentipes using PCR, targeting the Leishmania specific minicircle of the kDNA region. Further, the parasitic load in the infected sand flies was measured using quantitative PCR.

Conclusion

Though sand flies were most abundant in the rainy season, the highest rate of infection was detected in the winter season with 2.84% sand flies infected followed by the summer and rainy seasons respectively. This study can help in vector elimination programmes and to reduce disease transmission.     

DNA hybridizations on squash-blotted sandflies to identify both Phlebotomus papatasi and infecting Leishmania major

Epidemiological field studies on leishmaniasis have been hampered by the laborious, and often inefficient, methods used to assess the rates of infection of sandfly vectors (Diptera; Phlebotominae) by species of the causative disease organisms, protozoal parasites of the genus Leishmania (Kinetoplastida; Trypanosomatidae). We report the rapid and accurate identification of both sandfly vector (Phlebotomus (Phlebotomus) papatasi (Scopoli] and infecting Leishmania major Yakimov & Schokov by DNA hybridizations to squash-blotted sandflies. Large numbers of whole (infected) sandflies can be quickly squashed on to nylon hybridization filters and (following standard procedures) the filter-bound DNA can be hybridized sequentially to cloned, multicopy genomic sequences that are specific for species of Leishmania (kinetoplast DNA) or for the sandfly (ribosomal (r) DNA). Our sandfly probe consists of a 3.2 kb fragment of the intergenic 'non-transcribed' spacer of rDNA of P. papatasi that we have detected only in this species: it is present in all six geographically isolated populations tested (from Tunisia through to India) but cannot be detected in the morphologically similar P. (Phlebotomus) duboscqi Neveu-Lemaire, the vector of Leishmania major south of the Sahara; it also cannot be detected in Phlebotomus species of the subgenera Larroussius and Paraphlebotomus that together with P. papatasi are the dominant man-biting sandflies in north African foci of zoonotic cutaneous leishmaniasis, where (as in many arid regions of western Asia) P. papatasi is believed to be the sole vector of L. major.     

Phlebotomus (Adlerius) halepensis vector competence for Leishmania major and Le. tropica

In Eurasia, phlebotomine sandflies of the subgenus Adlerius (Diptera: Psychodidae) comprise about 20 known species. Some are suspected vectors of visceral leishmaniasis (VL) and at least one species has been implicated as a vector of cutaneous leishmaniasis (CL). We tested Phlebotomus (Adlerius) halepensis Theodor (Jordan strain) for CL vector competence, compared with three standard vectors: Phlebotomus (Phlebotomus) duboscqi N-L. from Senegal, Phlebotomus (Paraphlebotomus) sergenti Parrot from Turkey and the Neotropical Lutzomyia longipalpis (L. & N) (Jacobina strain). Sandfly females were membrane-fed on amastigote suspensions of Leishmania major Y. & S. and Le. tropica (Wright) (Kinetoplastida: Trypanosomatidae) and examined for parasite development 3, 6 and 10 days post-infection. Phlebotomus halepensis showed high susceptibility to both leishmanias, supporting typical suprapylarian parasite development similar to the other vectors. Phlebotomus halepensis infection rates were approximately 90% for Le. major and approximately 80% for Le. tropica, with high parasite densities. Development of infections was relatively fast, colonizing the thoracic midgut by 6 days post-bloodmeal in every case and reaching the stomodeal valve in >80% of flies. In late-stage infections, 10 days post-bloodmeal, nearly all P. halepensis females had cardia and stomodeal valve filled with very high numbers of parasites and some Le. tropica-infected females had promastigotes in the pharynx and proboscis. Host choice experiments in the laboratory showed that P. halepensis females fed readily on rat or rabbit and preferred the human forearm. In view of its vector competence and partial anthropophily, we infer that P. halepensis is a potential vector of cutaneous as well as visceral leishmaniases.     

The roles of Phlebotomus martini and P. celiae (Diptera: Phlebotominae) as vectors of visceral leishmaniasis in the Aba Roba focus, southern Ethiopia

Abstract. During field studies (December 1988 to April 1990) to determine the vectors of visceral leishmaniasis (kala-azar) in the Aba Roba (Segen Valley) focus of southern Ethiopia, a total of 40,770 sandflies (Diptera: Psychodidae: Phlebotominae) were collected and identified as six species of Phlebotomus and seventeen Sergentomyia spp. Nine of these species were anthropophilic (four Phlebotomus and five Sergentomyia spp.), the dominant being Phlebotomus (Synphlebotomus) martini, P.(S.)celiae and Sergentomyia (Sergentomyia) schwetzi. P. celiae adult populations increased during the rainy season. Of 2326 P. martini and 1044 P. celiae dissected, the parous rate was 29.9% and 24.4%, respectively. Leishmania promastigotes were detected in sixteen P. martini (eleven identified as L. donovani by isoenzyme analysis and/or a DNA probe) and in three P. celiae (two identified by DNA probe), giving overall infection rates of 0.7% (1.9% parous infection rate) and 0.3% (1.2% parous infection rate), respectively. Four isolates from P. martini were typed by their isoenzyme profile as L. donovani zymodeme MON-37 = LON-44, identical to one of the zymodemes isolated from patients in this focus. This is the first evidence that P. celiae is a vector and the first time that P. martini has been implicated in Ethiopia. Based on observations of abundance, seasonality and prevalence of infection of the two Synphlebotomus spp., it is concluded that P. martini is the major vector and P. celiae a secondary vector in the Aba Roba focus. The risk of infection with L. donovani is greatest during the wet season.     

Phlebotomine sand flies from Ouagadougou, Burkina Faso: first record of Phlebotomus (Larroussius) longicuspis south of the Sahara

During a brief entomological survey carried out in Ouagadougou, Burkina Faso, in March 2002, 178 phlebotomine sand flies (Diptera, Psychodidae) were caught using CDC miniature light traps. They were identified as Phlebotomus duboscqi Neveu-Lemaire, P. longicuspis Nitzulescu, Sergentomyia adleri (Theodor), S. affinis vorax (Parrot), S. antennata (Newstead), S. bedfordi (Newstead), S. christophersi (Sinton), S. fallax (Parrot), S. magna (Sinton) and S. schwetzi (Adler, Theodor & Parrot). This is the first record south of the Sahara of P. longicuspis sensu lato. This proven vector of Leishmania infantum Nicolle is a species closely related to P. perniciosus. It was identified by morphology and by the sequencing of a fragment of mitochondrial cytochrome b. There was 100% sequence homology with typical Tunisian specimens, and all characters identified the specimen as P. longicuspis sensu stricto. Epidemiological and biogeographical consequences are discussed.     

Bionomics of Phlebotomus papatasi (Diptera: Psychodidae) in an endemic focus of zoonotic cutaneous leishmaniasis in central Iran.

Following an epidemiological survey of zoonotic cutaneous leishmaniasis (ZCL) in several villages of Badrood, a rural district north of the city of Natanz, central Iran, Phlebotomus (Phlebotomus) papatasi Scopoli were found to be naturally infected with Leishmania (Leishmania) major zymodeme MON-26. Sand flies were collected and dissected biweekly from rodent burrows from May to October 2001. Leptomonad infection rates varied between 6.7% and 22.0%, being greatest in September, coinciding with peak activity of P. papatasi, two-three months before the highest incidence of ZCL human cases in November-December. The leptomonad infection rate was 1.1% of the 94 P. papatasi captured indoors. In ELISA testing of 520 P. papatasi blood meals during Sept. 2001 and Aug. 2002, the proportion giving positive reactions for human, sheep, cow, goat, rodent, and bird were 31.2%, 69.6%, 63%, 38.8%, 24.7%, and 21.8%, respectively. This report thus incriminates P. papatasi as the vector of zoonotic cutaneous leishmaniasis in this part of Iran.     

Phlebotomus sergenti (Parrot, 1917) identified as Leishmania killicki host in Ghardaïa, south Algeria

Since 2005, an outbreak of human cutaneous leishmaniasis (CL) in Gharda?a, south Algeria, was studied and one output of these investigations was the identification of two Leishmania species, Leishmania major and Leishmania killicki, as the CL causative agents. In the present study, we were curious to focus on sand fly fauna present in this area and detection of Leishmania-positive sand fly females. Sand flies (3717) were collected during two seasons using sticky papers and CDC light traps in urban, rural and sylvatic sites. Twelve Phlebotomus species were identified. Phlebotomus papatasi was dominant in the urban site while Phlebotomus sergenti and Phlebotomus riouxi/chabaudi were dominant in the sylvatic site. Out of 74 P. sergenti females captured by CDC light traps in the sylvatic site populated by Gharda?as' Gundi (Massoutiera mzabi), three ones were hosting Leishmania promastigotes. PCR-RFLP and sequencing of seven single-copy coding DNA sequences identified the promastigotes as L. killicki. Furthermore, laboratory experiments revealed that L. killicki isolate sampled from a CL patient inhabiting the studied region develop well in P. sergenti females. Our findings strongly suggest that the human cutaneous leishmaniases caused by L. killicki is a zoonotic disease with P. sergenti sand flies acting as hosts and vectors and gundi rodents as reservoirs.     

Bionomics of phlebotomine sand flies (Diptera: Psychodidae) in the province of Al-Baha, Saudi Arabia

The bionomics of phlebotomine sand flies (Diptera: Psychodidae) were studied for two successive years (January 1996-December 1997) at 12 collecting stations representing six sectors of the province of Al-Baha, Saudi Arabia. The predominant species was Phlebotomus bergeroti (41.7%), followed by lesser numbers of Phlebotomus sergenti (11%), Phlebotomus arabicus (10.6%), Sergentomyia tiberiadis (10.5%), Phlebotomus papatasi (10.2%), Sergentomyia antennata (9.6%), Phlebotomus alexandri (3%), Phlebotomus orientalis (2.3%) and Sergentomyia clydei (1.1%). The distribution of the collected species including species that are elsewhere known to act as vectors of human cutaneous leishmaniasis were distributed across different altitudes in Al-Baha. P. bergeroti, P. papatasi and P. arabicus were more abundant indoors; however, P. sergenti was more abundant outdoors. Sand fly populations exhibited three patterns of seasonal abundance in terms of their monthly activity. P. bergeroti, P. sergenti and P. arabicus were found to be naturally infected with Leishmania-like flagellates at an infection rate of 0.2%.     

Leishmania tropica in the black rat (Rattus rattus): persistence and transmission from asymptomatic host to sand fly vector Phlebotomus sergenti

Black rats (Rattus rattus) receiving Leishmania tropica injected intradermally into the ear were studied for the persistence of parasites and infectivity to natural sand fly vector. The mammalian host, the parasite, and the vector all originated from the endemic focus of Urfa, Turkey. Rats did not develop lesions or any apparent signs of disease, although at the site of inoculation they harboured live parasites capable of infecting sand flies. The number of L. tropica amastigotes detected in the inoculated ear by quantitative real-time PCR ranged from 5 x 10(3) to 10(6). Parasite DNA was also present in the tail and contralateral ear, sites distant from inoculation. After feeding on the ears of asymptomatic rats, Phlebotomus sergenti became infected with L. tropica. The average infection rate was 2.9%, and rats were infective for sand flies even 24 months post infection. The infectivity of the vertebrate host for insect vector was therefore not linked to the symptomatic stage of the infection. Such lack of correlation between clinical symptoms and infectivity to sand flies was reported previously for Leishmania infantum, the agent of visceral leishmaniasis; for species causing cutaneous leishmaniasis, however, this is the first evidence of transmission from a host without any visible cutaneous changes. If confirmed in the field, transmission from the asymptomatic host would be of great epidemiological significance.     

Bloodmeal digestion and Leishmania major infections in Phlebotomus duboscqi: effect of carbohydrates inhibiting midgut lectin activity

The carbohydrates galactosamine and heparin, previously shown to inhibit phlebotomine lectin activity in vitro, were fed to the sandfly Phlebotomus duboscqi Neveu-Lemaire (Diptera: Psychodidae) with blood, and the effects on mortality, fecundity, protease activity and susceptibility to Leishmania major Yakimoff & Schokhor (Kinetoplastida: Trypanosomatidae) were studied. Previous study revealed that galactosamine considerably enhanced the establishment of L. major infection in P. duboscqi and significantly increased parasite loads in late infections. This work demonstrates a similar but less pronounced effect of heparin. Heparin increased infection rates and parasite loads 3 and 9 days post-feeding but did not affect the location of Leishmania promastigotes and their anterior migration. Galactosamine supplement caused pronounced changes in bloodmeal digestion. It abolished the activity of alkaline proteases and trypsin, caused premature defecation of bloodmeal, increased mortality of female sandflies in days 1-4 post-feeding and decreased their fecundity. Heparin had a less pronounced effect on sandfly physiology. It lowered trypsin activity 12 and 72 h post-bloodmeal but did not alter defecation, mortality and oviposition. The data suggest that the enhancing effect of these carbohydrates on Leishmania infections in sandfly midgut could be explained by their interference with midgut proteases. The study supports the hypothesis that proteolytic activities of midgut proteases strongly influence the vector competence of sandflies.     

Experimental infection of Phlebotomus perniciosus and Phlebotomus tobbi with different Leishmania tropica strains

Cutaneous leishmaniasis due to Leishmania tropica is increasingly documented in Europe and the Middle East. Besides its specific vector, Phlebotomus sergenti, permissive Phlebotomus sand flies are suspected as potential vectors of L. tropica. We investigated the susceptibility of two widely distributed species, Phlebotomus perniciosus and Phlebotomus tobbi. Laboratory-reared sand flies were infected experimentally with L. tropica strains differing in lipophosphoglycan epitopes, geographical distribution and epidemiology. High infection rates, heavy parasite loads and fully developed late-stage infections including colonization of the stomodeal valve were observed in all parasite-vector combinations. Our findings demonstrate that P. perniciosus and P. tobbi are susceptible to different L. tropica strains and may play a role in their circulation in endemic foci of Europe, the Middle East and North Africa.     

Leishmania-sandfly interactions: an empirical field study

Phlebotomus papatasi is the sandfly vector of Leishmania major in the Jordan Valley. The objective of this study was to characterize vector-parasite relations in an active zoonotic focus. Seasonality and intensity of promastigote infection rates in female sandflies and the developmental stage of these hosts were established. On 153 trap-nights, 641 female P. papatasi were caught and examined. Of these, 48 (7.4%, range 12.9-4.8%) were infected with L. major promastigotes. Correlating the number of parasites with the gonotrophic age of the vector revealed that most infections initially are light ones, and those that survive in the vector generally prosper and proliferate. Comparing infection rates in parous and gravid flies revealed that similar proportions of gravid and parous flies are found infected. Thus, Leishmania infections do not appear to affect sandfly survival.     

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 Findings

An 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.

Conclusion

Our data suggest the possible involvement of P. duboscqi and potentially S. darlingi in the transmission of ZCL in Mali.     

A morphologically distinct Phlebotomus argentipes population from active cutaneous leishmaniasis foci in central Sri Lanka

Although the reported aetiological agent of cutaneous leishmaniasis (CL) in Sri Lanka is Leishmania donovani, the sandfly vector remains unknown. Ninety-five sandflies, 60 females and 35 males, collected in six localities in the district of Matale, central Sri Lanka, close to current active transmission foci of CL were examined for taxonomically relevant characteristics. Eleven diagnostic morphological characters for female sandflies were compared with measurements described for Indian and Sri Lankan sandflies, including the now recognised Phlebotomus argentipes sensu lato species complex. The mean morphometric measurements of collected female sandflies differed significantly from published values for P. argentipes morphospecies B, now re-identified as Phlebotomus annandalei from Delft Island and northern Sri Lanka, from recently re-identified P. argentipes s.s. sibling species and from Phlebotomus glaucus. Furthermore, analysis of underlying variation in the morphometric data through principal component analysis also illustrated differences between the population described herein and previously recognised members of the P. argentipes species complex. Collectively, these results suggest that a morphologically distinct population, perhaps most closely related to P. glaucus of the P. argentipess. I. species complex, exists in areas of active CL transmission. Thus, research is required to determine the ability of this population of flies to transmit cutaneous leishmaniasis.     

First report on natural infection of the Phlebotomus tobbi by Leishmania infantum in northwestern Iran

Visceral leishmaniasis (VL) is an important health problem in Ardebil, where it borders Azerbaijan in the northwestern Iran. In spite of the presence of both cutaneous and visceral leishmaniasis (CL and VL) in northwestern Iran, previous researches have consistently revealed the etiologic agent of VL in the region to be Leishmania infantum. This is the first report of natural infection of Phlebotomus tobbi with L. infantum in Bilesavar district in the northern part of Ardebil province bordering Azerbaijan. Polymerase chain reaction (PCR) of kDNA, ITS1-rDNA, and CPB genes of the parasite followed by restriction fragment length polymorphism (RFLP) and gene sequencing analyses revealed presence of L. infantum in six out of 433 tested female sand fly specimens. Although sand flies of P. tobbi were infrequent, two out of 32 (6.25%) females captured in the area were found infected with the parasite. Phlebotomus perfiliewi transcaucasicus, the known vector of VL in the area, were the most dominant species but only four out of 273 (1.47%) tested were infected with L. infantum. This study showed that P. tobbi similar to P. perfiliewi transcaucasicus could play a significant role in the transmission of the L. infantum. However more investigations are needed to demonstrate that L. infantum is the only species circulating in the focus.     

Post-engorgement dynamics of haemagglutination activity in the midgut of phlebotomine sandflies

Abstract. Haemagglutination activity (HA) was studied in gut extracts of both sexes of adults of six sandfly species. HA was sex dependent, with the activity in males more than 50 times lower man that of unfed females. In females, high HA was demonstrable in both the thoracic and abdominal midgut but not in the hindgut. In unfed flies the activity was similar in the midgut wall and the gut contents whereas, in fed females, a high increase was seen in the midgut contents. After blood-feeding, HA was elevated, reaching peak titres 2 days later and then falling to the base level or less immediately after defaecation. The magnitude of the HA response differed according to species, ranging from 2-fold in Lutzomyia carmelinoi up to 16-fold in Phlebotomus duboscqi. Quantitative differences between sandflies in their HA response may influence their ability to support the development of Leishmania spp.