Phlebotomus from Madagascar (Diptera: Psychodidae). III--Description of Phlebotomus (Anaphlebotomus) fontenillei n. sp

The male of Phlebotomus (Anaphlebotomus) fontenillei n. sp. is described from Namoroka area (Madagascar). Its belongs to the subgenus Anaphlebotomus: style with four spines, coxite without basal process and paramere with two branches. It shares with P. berentiensis an original and exclusive antennal formula: 2/III-XII which distinguishes them from P. fertei. P. fontenillei n. sp. differs mainly from P. berentiensis by about 40 setae in tuft on the ventral face of the coxite, the length of the genital ducts and the position of the spines on the style. Sequence of the second internal transcribed spacer (ITS2) of the ribosomal DNA (rDNA) is very informative: the male of P. fontenillei n. sp. cannot be linked to the female of P. huberti (male unknown) regarding the size of amplified DNA fragment (459 bp versus 600 respectively) and the high degree of variability. There are few differences (10 mutations) between the sequences of P. fontenillei n. sp. and P. berentiensis which are closely related species.     

Phlebotomus (Legeromyia) multihamatus subg. nov., sp. nov. from Gabon (Diptera: Psychodidae)

During a research project aimed at the study of the Culicinae fauna of Gabon and carried out in the National Park of La Lopé, we captured an unknown sandfly male specimen (genus Phlebotomus) by CDC miniature light trap belonging to a new species for Science. Furthermore, the originality of his genitalia does not allow us to include this species in one of the existing subgenus, thus in this paper we propose the creation of a new subgenus, as Phlebotomus (Legeromyia) multihamatus sp. nov., subg. nov. described from the National Park of La Lopé, through one male captured with CDC miniature light trap. A new species and a new subgenus of sandfly is characterised by a short style with three spines, a paramere wearing a basal hook as well as a basal pouch and the absence of basal lobe on the coxite. The originality of the genitalia of the male gives way to discussion about potential primary homologies between P. multihamatus sp. nov. and Phlebotomus (Abonnencius) fortunatarum, Phlebotomus (Anaphlebotomus) stantoni and Phlebotomus (Euphlebotomus) argentipes, which should be verified for future studies. The discovery of this new species in Gabon must encourage the study of sandflies in this country.     

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.     

Phlebotomus from Madagascar (Diptera: Psychodidae). II--Description of the female Phlebotomus (Anaphlebotomus) fertei Depaquit, Léger & Robert, 2002; description of the male and new data for the female Phlebotomus (Anaphlebotomus) berentiensis (Léger & Rodhain, 1978) comb. nov

The female of Phlebotomus (Anaphlebotomus) fertei Depaquit, Léger & Robert, 2002 is described from Madagascar. The spermathecae have separated ducts, a body with an enlarged-sclerified dilatation and a deriviated large bladder with thin walls and long head. P. fertei female has the following characters: long antennal segments, pharyngeal armature with small teeth and cibarium with 15 to 30 denticles, and very long legs. Its remarkable spermatheca is closely related to that of P. berentiensis. The status of P. (Anaphlebotomus) berentiensis (Léger & Rodhain, 1978) is revised into this paper. The female is redescribed and the male is described for the first time. The latter has an unusual antennal formula (2/III-XII), a cibarium with four teeth and about 30 denticles, no tuft of setae on the coxite, four spines on the style, including a very long proximal one. Males and females were linked to each other based on morphological and molecular characteristics (second internal transcribed spacer of the ribosomal DNA and ND4 gene of the mitochondrial DNA). The inclusion of these taxa into the subgenus Anaphlebotomus Theodor, 1948 is exclusively based on male characters as listed by Theodor (style with four spines, no basal lobe on the coxite, paramere with two or three branches). The very heterogeneous characters of the females do not allow their use for the definition of this subgenus, which is probably not homogeneous.     

Genetic polymorphism in sympatric species of the genus Phlebotomus, with special reference to Phlebotomus perniciosus and Phlebotomus longicuspis (Diptera, Phlebotomidae)

The Random Amplified Polymorphic DNA assay was used to study genetic variation within and between five Phlebotomus species belonging to three subgenera: P. (Larroussius) ariasi, P. (L.) longicuspis, P. (L.) perniciosus, P.(Paraphlebotomus) sergenti and P. (Phlebotomus) papatasi sympatric in southern Spain and proven vector of leishmaniasis. Two cluster analysis were proposed: one according to sandfly species and populations, the second according individual specimens of Phlebotomus perniciosus, Phlebotomus longicuspis s.l. and intermediate morphological specimens between these species. The results obtained are closely correlated with the taxonomy classically accepted for the subgenera and with the automatic classifications made by other authors which use morphological and isoenzymatic data. The validity of the species Phlebotomus longicuspis is also discussed.     

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.     

A new species of Phlebotomus subgenus Euphlebotomus (Psychodidae) from Afghanistan

Abstract. A new species of sandfly Phlebotomus (Euphlebotomus) caudatus sp. n. is described from three males captured at two points in mountains near Farah-Rud, Western Afghanistan.     

Phylogenetic analysis of Phlebotomus species belonging to the subgenus Larroussius (Diptera, psychodidae) by ITS2 rDNA sequences

In the genealogy of Phlebotomus (Diptera: Psychodidae), morphological analyses have indicated that the subgenus Larroussius is a monophyletic group which is most closely related to the subgenera Transphlebotomus and Adlerius. We conducted a phylogenetic analysis of the relationships among six representative species of the subgenus Larroussius and one species representatitive of the Phlebotomus subgenus, assessing sequences of the Second Internal Transcribed Spacer (ITS2) of the ribosomal RNA (rRNA). Three of the species (P. perniciosus, P. ariasi and P. perfiliewi perfiliewi) were collected in different parts of the Mediterranean area. The trees estimated from parsimony and neighbour-joining analyses supported the monophyly of the Larroussius subgenus inferred from the morphological analysis. According to our data, P. ariasi may be a sister group to the rest of the Larroussius subgenus, although additional sequence data are needed to confirm this observation. Our results suggest that P. perniciosus and P. longicuspis are distinct species, in spite of the fact that there are only slight morphological differences. The strict congruence between the phylogeny of the Larroussius subgenus inferred from the ITS2 sequences and that based on morphological studies further confirmed the ability of the spacer sequence to identify recently-derived affiliations.     

A taxonomic review of Phlebotomus (Idiophlebotomus) (Psychodidae)

Five known species (asperulus Quate & Fairchild, erebicolus Quate, pholetor Quate & Fairchild, sejunctus Quate, and stellae Quate) of the caverni-colous subgenus Phlebotomus (Idiophlebotomus) are discussed in relation to newly recognized species from West Malaysia and India. P. frondifer n.sp. and P. tubifer n.sp. are described and a key to the adults of all seven species is given. General features of the subgenus are discussed, with particular reference to the functional relationships between the specialized morphology of the mouthparts and the probable bat hosts of these species.     

Morphometric and molecular differentiation of Phlebotomus (Phlebotomus) sandflies

The closely related sandfly species of the subgenus Phlebotomus namely, Phlebotomus papatasi (Scopoli, 1786), Phlebotomus duboscqi Neveu‐Lemair, 1906 and Phlebotomus bergeroti Parrot, 1934 (Diptera: Psychodidae), are major vectors of Leishmania major (Kinetoplastida: Trypanosomatidae), the causative agent of cutaneous leishmaniasis in the Old World. Although allopatric in most of their distribution, the three species exist sympatrically in many places in central and eastern Sudan. Males of the three species can be distinguished using morphological characters; however, females are much harder to identify, thus complicating epidemiological studies. We carried out a morphometric and a molecular study to determine reliable morphological features and develop a polymerase chain reaction (PCR) assay for distinguishing females of these species. Males and females from each species were collected from sites in Sudan, East Africa and from one site in Mali, West Africa. Males were analysed morphologically and 20 characters and 10 character ratios were used in a stepwise discriminant analysis. This led to the identification of four characters with high discriminant loading scores sufficient for accurate male species identification. Male DNA was then used for the development of a PCR‐based species diagnostic based on the second internal transcribed spacer (ITS2) of the ribosomal DNA. A set of four primers was developed to generate fragment sizes that are specific to each species and can reliably identify females as well as hybrid DNA. Both the morphometric and the molecular findings of this study have important applications for studies of the epidemiology of cutaneous leishmaniasis.     

Description of Madathamugadia hiepei n. sp. (Nematoda: Splendidofilariinae), a parasite of a South African gecko,and its development in laboratory bred Phlebotomus dubosqi (Diptera: Psychodidae)

Madathamugadia hiepei n. sp., Splendidofilariinae, a parasite of a South African gecko Pachydactylus turneri is described together with its development obtained experimentally in Phlebotomus duboscqi (Diptera: Psychodidae: Phlebotominae). This new species differs from the two small, more highly evolved groups with a short tail and atrophied postcloacal papillae, the first group consisting of two Madagascan species, M. zonosauri and M. hopluri, parasites of the Gerrhosauridae and Iguanidae, and the second containing three species from the Ethiopian Region, M. huambensis, M. versterae and M. bissani, parasites of the Scincidae. It also differs from M. ineichi, the most primitive species of the genus (cuticularised buccal capsule, no atrophy of head papillae and largest number of precloacal papillae), a parasite of the Cordylidae in South Africa. M. hiepei is close to the two species parasitic in the Gekkonidae of the Mediterranean subregion, M. ivaschkini and M. wanjii, all three of which have a post-oesophageal vulva. However, the new species can be distinguished from the Mediterranean parasites by (a) the shorter oesophagus, (b) the number and position of the cloacal papillae and (c) the microfilaria. The three filariae of this group and M. ineichi, the only ones of which aspects of the life-cycles are known, experimentally develop in phlebotomine sand flies.     

Pseudocapillaria (Discocapillaria) margolisi n. subg., n. sp. (Nematoda: Trichuroidea) from freshwater fishes of West Bengal,India

A new nematode,Pseudocapillaria (Discocapillaria) margolisi n. subg., n. sp., from the freshwater fishesPuntius conchonius, P. sophore andAmblypharyngodon mola of West Bengal, India is described. The presence of a ventral lobe-like elevation with two papillae and the absence of a dorsal cuticular membrane on the male tail are referred to as distinguishing features of the new subgenus. The parasite is also characterised by small body size, short and non-annulated stichocytes, a distinctly expanded and folded lobular rim of the proximal end of the spicule, and eggs with protruding polar plugs, all features in which it differs distinctly from other species of the genusPseudocapillaria.     

Descriptions of Cosmocampus retropinnis sp. n., Minyichthys sentus sp. n. and Amphelikturus sp. (Pisces, Syngnathidae) from the Eastern Atlantic Region

Pelagic juvenile pipefishes, representing three genera previously unrecorded from the eastern Atlantic region, are reported from collections of the 1928–30 DANA expedition. Cosmocampus retropinnis sp. n., with dorsal-fin origin on tail and 6.75–8.0 subdorsal rings (S Morocco to Gambia), Minyichthys sentus sp. n., with patches of spinules on snout and 42 tail rings (W Mediterranean and Atlantic Ocean off N Morocco and Canary Is.), and Amphelikturus sp. (off Sierra Leone) are described and figured. The subgenus Minyichthys Herald and Randall is rediagnosed as a genus characterized, in part, by an 8-rayed caudal fin. A key is provided to the nine genera of Syngnathidae known from the eastern Atlantic region.     

ITS 2 sequences heterogeneity in Phlebotomus sergenti and Phlebotomus similis (Diptera,Psychodidae): possible consequences in their ability to transmit Leishmania tropica

An intraspecific study on Phlebotomus sergenti, the main and only proven vector of Leishmania tropica among the members of the subgenus Paraphlebotomus was performed. The internal transcribed spacer 2 (ITS2) sequences of 12 populations from 10 countries (Cyprus, Egypt, Italy, Lebanon, Morocco, Pakistan, Portugal, Spain, Syria, and Turkey) were compared. Samples also included three species closely related to P. sergenti: Phlebotomus similis (three populations from Greece and Malta), Phlebotomus jacusieli and Phlebotomus kazeruni. Our results confirm the validity of the taxa morphologically characterised, and imply the revision of their distribution areas, which are explained through biogeographical events. At the Miocene time, a migration route, north of the Paratethys sea would have been followed by P. similis to colonise the north of the Caucasus, Crimea, Balkans including Greece and its islands, and western Turkey. Phlebotomus sergenti would have followed an Asiatic dispersion as well as a western migration route south of the Tethys sea to colonise North Africa and western Europe. This hypothesis seems to be well supported by high degree of variation observed in the present study, which is not related to colonisation or to intra-populational variation. Two groups can be individualised, one oriental and one western in connection with ecology, host preferences and distribution of L. tropica. We hypothesise that they could be correlated with differences in vectorial capacities.     

Description and DNA barcoding of Tipula ( Pterelachisus) recondita sp. n. from the Palaearctic region (Diptera, Tipulidae)

Tipula (Pterelachisus) recondita Pilipenko & Salmela, sp. n. is described. The new species is collected from two localities: Finland, Kittilä (North boreal ecoregion) and Russia, Primorski kray (Zone of temperate broadleaf and mixed forests). Although variation in the structure of male hypopygium between the Finnish and Russian populations is observed, DNA barcode sequences differ only by three nucleotides (0.2 % K2P distance), supporting presence of one widespread species. K2P minimum distances between the new species and 17 other species of the subgenus range from 5.3 to 15.8 % (mean 8.8 %). The new species is forest-dwelling, known from an old-growth herb-rich forest (Finland) and Quercus mongolica forest (Russia). The new species is perhaps closest to Tipula (Pterelachisus) imitator Alexander and in lesser extent to Tipula (Pterelachisus) pauli Mannheims; the inner gonostylus of both species are illustrated.     

The life cycle and host specificity of Psychodiella sergenti n. sp. and Ps. tobbi n. sp. (Protozoa: Apicomplexa) in sand flies Phlebotomus sergenti and Ph. tobbi (Diptera: Psychodidae)

Two new gregarines in the recently erected genus Psychodiella (formerly Ascogregarina), Psychodiella sergenti n. sp. and Psychodiella tobbi n. sp., are described based on morphology and life cycle observations conducted on larvae and adults of their natural hosts, the sand flies Phlebotomus sergenti and Phlebotomus tobbi, respectively. The phylogenetic analyses inferred from small subunit ribosomal DNA (SSU rDNA) sequences indicate the monophyly of newly described species with Psychodiella chagasi. Ps. sergenti n. sp. and Ps. tobbi n. sp. significantly differ from each other in the life cycle and in the size of life stages. The sexual development of Ps. sergenti n. sp. (syzygy, formation of gametocysts and oocysts) takes place exclusively in blood-fed Ph. sergenti females, while the sexual development of Ps. tobbi n. sp. takes place also in males and unfed females of Ph. tobbi. The susceptibility of Phlebotomus perniciosus, Phlebotomus papatasi, Ph. sergenti, Ph. tobbi, and Phlebotomus arabicus to both gregarines was examined by exposing 1st instar larvae to parasite oocysts. High host specificity was observed, as both gregarines were able to fully develop and complete regularly the life cycle only in their natural hosts. Both gregarines are considered as serious pathogens in laboratory-reared colonies of Old World sand flies.     

Parvicapsula bicornis n. sp. and P. limandae n. sp. (Myxozoa, Parvicapsulidae) in Pleuronectidae (Teleostei, Heterosomata) from Denmark

Two species of Parvicapsula were found in the kidney tubules and the urinary bladder of 2 pleuronectid fish from the northern Oresund, Denmark. The coelozoic, spherical, disporic trophozoites of both species are 10 to 12 pm in diameter. The myxospores of both species are elongate, asymmetrical and slightly curved, and have spherical polar capsules. Parvicapsula bicornis n. sp. (6-8 x 5-6 microm, polar capsule 2.5 microm in diameter) occurs in Pleuronectes platessa. The polar capsules of P. bicornis are arranged symmetrically on either side of the longitudinal axis and its spores differ from other species of Parvicapsula in having two 2-3 microm long posterior processes of different length. Parvicapsula limandae n. sp. (8-11 x 4-5 pm, polar capsule 1.6 microm in diameter) is found in Limanda limanda. The polar capsules are arranged along the longitudinal axis. It differs from Parvicapsula unicornis Kabata, 1962, recorded from L. limanda, in the arrangement of the polar capsules and in the absence of a posterior horn-like projection. The phylogenetic relationship between P. bicornis n. sp., P. limandae n. sp. and other Parvicapsula spp. was examined with their partial small subunit rDNA (SSU rDNA) sequences. P. limandae n. sp. and P. asymmetrica appear to be closely related, while P. bicornis n. sp. and P. minibicornis are the most divergent members of the genus.     

First record of Phlebotomine sandflies (Diptera: Psychodidae) in the Comoros Archipelago with description of Sergentomyia (Vattieromyia) pessoni n. sp. and S. (Rondanomyia) goodmani comorensis n. ssp

No Phlebotomine sandflies had ever been reported in the Comoros Archipelago, including the three islands of the Republic of the Union of Comoros (Grande Comore, Mohéli and Anjouan) and the French oversea department of Mayotte. During three field surveys carried out in 2003, 2007 and 2011, we provided the first record of Phlebotomine sandflies in this area. A total of 85 specimens belonging to three species were caught: a new species S. (Vattieromyia) pessoni n. sp. (two females from Grande Comore), a new subspecies of Sergentomyia (Rondanomyia) goodmani (80 specimens from Grande Comore and one from Anjouan) and Grassomyia sp. (two females from Mohéli). The individualisation of these taxa was inferred both from morphological criteria and sequencing of a part of the cytochrome b of the mitochondrial DNA. These taxa are closely related to Malagasy sandflies.