Fine structure of the reproductive system of <Emphasis Type="Italic">Chrysomya megacephala</Emphasis> (Diptera: Calliphoridae): the external sexual organ

The fine structure of the reproductive system of Chrysomya megacephala (Fabricius), a blow fly species of medical importance, was studied using scanning electron microscopy (SEM) to contribute detailed morphological information on the external sexual organs or genitalia of this species. Analysis of the male genitalia or hypopygium revealed that the epandrium is a broad organ resembling a crescent shape, while the ejaculatory apodeme and aedeagal apodeme show similarity in their lengths. The cercus is significantly longer than the surstylus with its apical end being more or less rounded. Dense patches of long bristles, morphologically similar to the sensilla chaetica and sensilla trichodea, were found along the lower half of the cercus. The surstylus has a stout triangular shape, and the proximal half is greatly endowed with sensilla thought to be sensilla chaetica and sensilla trichodea. The aedeagus per se is prominent and has a clavate shape formed by the base theca and elongated phallus. The vesica of the phallus is a smooth bilobed structure, which curves inward. The juxta and juxta process are armed with many rows of strong spines resembling thorns in their appearance. The harpe is slender, recurved anteriorly, and distally pointed like a sickle. Regarding the female genitalia or ovipositor, SEM observation of the supraanal plate revealed it to be sclerotized and more or less triangular-shaped, lying between a pair of short cerci. Several types of sensilla were found on the supraanal plate, subanal plate and cercus, including the sensilla trichodea, sensilla basiconica, sensilla placodea, and probably, sensilla styloconica. The possible functions of sensilla distributed in particular regions of the fly genitalia are discussed.     

Ultrastructure of eggshell of<Emphasis Type="Italic"> Chrysomya nigripes</Emphasis> Aubertin (Diptera: Calliphoridae)

We examined the eggshell structure of the blowfly, Chrysomya nigripes Aubertin, a species of forensic importance, using scanning electron microscopy. A relatively wide plastron region was located dorsally. It extended almost the entire length of the egg and bifurcated slightly, close to the micropyle, which is a deep cavity surrounded by an elevated, fine, wavy rim. The chorionic sculpture had an hexagonal pattern, with a smooth and elevated boundary. The ruptured eggshell showed the outermost exochorion, membranous outer endochorion and pillar layer, which also had an hexagonal pattern on a perforated mesh pillar layer. This study provides a greater database on the egg of this forensically significant blowfly.     

Morphology and developmental rate of blowflies <Emphasis Type="Italic">Chrysomya megacephala</Emphasis> and <Emphasis Type="Italic">Chrysomya rufifacies</Emphasis> in Thailand: application in forensic entomology

The larval morphology and developmental rate of Chrysomya megacephala (F.) and Chrysomya rufifacies (Macquart), the two most forensically important blowfly species in Thailand, are presented. Morphological comparison of the third instar of both species revealed different characteristics (e.g., body appearance, cephalopharyngeal skeleton, dorsal cuticular spines between the prothorax and mesothorax, and feature of the posterior spiracle), thereby, allowing correct identification. A data analysis was conducted in Chiang Mai province, Northern Thailand during 2000–2001 on the developmental rate of both flies under natural ambient temperature and a natural light–dark photoperiod. The results indicated that larvae of C. megacephala developed more rapidly in April, pupariation initiated at 84 h at temperatures averaging 31.4°C, and the larvae grew slower in the rainy season and winter. Similarly, rapid development of C. rufifacies larvae appeared in the summer, with a pupariation period as short as 96 h in June (average temperature 27.4°C). Analysis of the median body length of C. megacephala and C. rufifacies larvae in different seasons of the years 2000–2001 in Thailand revealed that both species developed rapidly in the summer; pupariation of C. rufifacies initiated at 144 h, while C. megacephala initiated pupariation at 156 h. This information is potentially useful for estimating the postmortem interval of a corpse in forensic investigations, where the corpse becomes infesting with these fly species. This work has been presented at the 21st Pacific Science Congress held in Okinawa, Japan, in 12–18 June, 2007.     

Redescription of <Emphasis Type="Italic">Lemuricola</Emphasis> (<Emphasis Type="Italic">Madoxyuris</Emphasis>) <Emphasis Type="Italic">bauchoti</Emphasis> (Nematoda,Oxyuridae) from <Emphasis Type="Italic">Lemur catta</Emphasis> in Madagascar

Lemuricola (Madoxyuris) bauchoti Chabaud, Brygoo et Petter, 1965 is redescribed from material collected from the ring-tailed lemur, Lemur catta, from the Beza Mahafaly Special Reserve in Madagascar using the scanning electron microscope. This is a new host record and the first oxyurid reported from the ring-tailed lemur. Previously, records of each species of the subgenus Madoxyuris have been restricted to a single host species, but the close relationship between these nematodes and their Strepsirrhini hosts will only be proven when additional records fill in the gaps in their distribution.     

Cysteine proteinases from promastigotes of <Emphasis Type="Italic">Leishmania</Emphasis> (<Emphasis Type="Italic">Viannia</Emphasis>) <Emphasis Type="Italic">braziliensis</Emphasis>

Leishmania (Viannia) braziliensis is the major causative agent of American tegumentary leishmaniasis, a disease that has a wide geographical distribution and is a severe public health problem. The cysteine proteinase B (CPB) from Leishmania spp. represents an important virulence factor. In this study, we characterized and localized cysteine proteinases in L. (V.) braziliensis promastigotes. By a combination of triton X-114 extraction, concanavalin A-affinity, and ion exchange chromatographies, we obtained an enriched fraction of hydrophobic proteins rich in mannose residues. This fraction contained two proteinases of 63 and 43 kDa, which were recognized by a CPB antiserum, and were partially sensitive to E-64 in enzymatic assays with the peptide Glu-Phe-Leu. In confocal microscopy, the CPB homologues localized in the peripheral region of the parasite. This data together with direct agglutination and flow cytometry assays suggest a surface localization of the CPB homologues. The incubation of intact promastigotes with phospholipase C reduced the number of CPB-positive cells, while anti-cross-reacting determinant and anti-CPB antisera recognized two polypeptides (63 and 43 kDa) derived from phospholipase C treatment, suggesting that some CPB isoforms may be glycosylphosphatidylinositol-anchored. Collectively, our results suggest the presence of CPB homologues in L. braziliensis surface and highlight the need for further studies on L. braziliensis cysteine proteinases, which require enrichment methods for enzymatic detection.     

Insecticidal properties of essential oils from <Emphasis Type="Italic">Lippia turbinata</Emphasis> and <Emphasis Type="Italic">Lippia polystachya</Emphasis> (Verbenaceae) against <Emphasis Type="Italic">Culex quinquefasciatus</Emphasis> (Diptera: Culicidae)

Mosquitoes are important vectors of diseases to humans and domestic animals. Chemical control of vectors remains a main resource for the prevention and control of vector-borne diseases. Due to the development of insecticide resistance and risks to human health and the environment of synthetic compounds, the search for alternative pesticides is encouraged. This work assessed the insecticidal activity of essential oils (EOs) from Lippia turbinata and L. polystachya from Argentina on Culex quinquefasciatus mosquitoes. EOs were extracted by hydro-distillation and analyzed with gas–liquid chromatography and mass spectrometry. The insecticidal activity against mosquito larvae, pupae, and adults were evaluated according to World Health Organization protocols. Concentrations ranking from 10 to 160 ppm were assessed at 1, 2, 3, and 24 h posttreatment. The composition of the EO of L. polystachya and L. turbinata were qualitatively similar, with α-thujone and carvone as main constituent; differences were mostly due to the proportion of each component. β-caryophyllene was also an important constituent of the EO of L. turbinata. Both EO were larvicidal at concentrations of 80 ppm or higher, but only L. turbinata was adulticidal. No pupal mortality was detected. The potential of these EOs for vector control is discussed.     

<Emphasis Type="Italic">Trypanosoma</Emphasis> (<Emphasis Type="Italic">Herpetosoma</Emphasis>) <Emphasis Type="Italic">kuseli</Emphasis> sp. n. (Protozoa: Kinetoplastida) in Siberian flying squirrels (<Emphasis Type="Italic">Pteromys volans</Emphasis>)

All trypanosome species classified in the subgenus Herpetosoma in sciurid hosts have been recorded from ground and tree squirrels to date, but not from any flying squirrels. We describe in this paper a novel trypanosome species, Trypanosoma (Herpetosoma) kuseli sp. n., from Siberian flying squirrels (Pteromys volans) imported from China, and compare it with T. (H.) otospermophili in Richardson's ground squirrels (Spermophilus richardsonii) and Columbian ground squirrels (Spermophilus columbianus) from the USA. Due to a short free flagellum, the new species appeared stumpy compared with T. otospermophili (length of free flagellum 7.0 +/- 0.8 microm, total length 32.1 +/- 0.8 microm, n = 13 and length of free flagellum 15.5 +/- 1.6 microm, total length 35.9 +/- 1.0 microm, n = 13, respectively). Another conspicuous morphological feature of the new species was an anteriorly positioned kinetoplast, found approximately at the midpoint between the nucleus and the posterior end. These characters have not been recorded from any squirrel Herpetosoma trypanosome species. Comparison of the nucleotide sequences of the small and large subunit rRNA genes indicated that T. kuseli sp. n. was more homologous to T. otospermophili than murid Herpetosoma species, such as T. grosi, T. lewisi, T. musculi, T. microti and T. evotomys.     

Winter activity of <Emphasis Type="Italic">Phlebotomus</Emphasis> (<Emphasis Type="Italic">Transphlebotomus</Emphasis>) <Emphasis Type="Italic">mascittii</Emphasis>, Grassi 1908 (Diptera: Psychodidae) on the island of Corsica

In the first week of February 2008 on the island of Corsica, 55 Phlebotomus (Transphlebotomus) mascittii, 39 males and 16 females, were caught in a railway tunnel. This is the first proof of the winter activity of the adult P. mascittii in Europe.     

Crossing experiment of <Emphasis Type="Italic">Anopheles maculatus</Emphasis> form K and <Emphasis Type="Italic">Anopheles willmori</Emphasis> (James) (Diptera: Culicidae)

We recently reported crossing experiments between Anopheles maculatus form K and five members of the Maculatus group to support the specific status of form K. In the present study, we performed further crosses between form K and a sixth species of the Maculatus group, Anopheles willmori (James). Low viability was observed in hybrid males and females. All hybrid males were sterile with atrophied testes, or partially sterile with abnormal spermatozoa. The hybrid females showed normal ovaries. The ovarian nurse cell polytene chromosomes of the F₁ hybrid females displayed complete asynapsis. Backcrossing showed low viability. All males were sterile with atrophied testes or partially sterile with abnormal spermatozoa, and the females showed varying degrees of atrophied ovaries. The results provide clear evidence that form K is not conspecific to A. willmori supporting previous studies that form K represents another species of the Maculatus group.     

Cercarial shedding from <Emphasis Type="Italic">Galba</Emphasis><Emphasis Type="Italic">truncatula</Emphasis> infected with <Emphasis Type="Italic">Fasciola</Emphasis><Emphasis Type="Italic">gigantica</Emphasis> of distinct geographic origins

Galba truncatula snails were experimentally infected with either of two different isolates of Fasciola gigantica, originating from Egypt or China, to determine the influence of these isolates on the characteristics of snail infections. The survival rates of G. truncatula on day 30 post-exposure were 90.0% and 60.2% in the Egyptian and Chinese groups, respectively. The frequency of cercaria-shedding snails within the Egyptian group was 79.8%, whereas in the Chinese group it was 22.4%. The parasite origin had a significant effect on the durations of the prepatent and patent periods. The mean number of cercariae shed from the Egyptian group was significantly greater than that shed from the Chinese group (a mean of 275.5 per cercaria-shedding snail compared with 29.0). These results could be explained by the fact that G. truncatula might be a natural intermediate host for F. gigantica in Egypt, and the greater adaptability of the Egyptian miracidia of F. gigantica to unusual snail hosts. These results demonstrate the influence of the geographic origin of the parasite on the success of trematodes infecting snails.     

Antiviral biflavonoids from <Emphasis Type="Italic">Radix Wikstroemiae</Emphasis> (<Emphasis Type="Italic">Liaogewanggen</Emphasis>)

Background  

Radix Wikstroemiae is a common Chinese herbal medicine. The ethyl acetate fraction of the ethanolic extract of W. indica possesses potent in vitro antiviral activity against respiratory syncytial virus (RSV). This study aims to identify the antiviral components of the active fraction.     

First report on susceptibility of wild <Emphasis Type="Italic">Aedes aegypti</Emphasis> (Diptera: Culicidae) using <Emphasis Type="Italic">Carapa guianensis</Emphasis> (Meliaceae) and <Emphasis Type="Italic">Copaifera</Emphasis> sp. (Leguminosae)

Oils of Carapa guianensis and Copaifera spp. are well known in the Amazonian region as natural insect repellent, and studies have reported their efficiency as larvicide against some laboratory mosquito species. However, in wild populations of mosquitoes, these oils have not yet been evaluated. Thus, the objective of this study was to investigate their efficiency as larvicide in wild populations of Aedes aegypti with a history of exposure to organophosphate. The susceptibility of larvae was determined under three different temperatures, 15°C, 20°C, and 30°C. For each test, 1,000 larvae were used (late third instar and early fourth instar—four replicates of 25 larvae per concentration). Statistical tests were used to identify significant differences. The results demonstrated that as the laboratory A. aegypti, the wild populations of A. aegypti were also susceptible to C. guianensis and Copaifera sp. oils. The lethal concentrations for Copaifera sp. ranged from LC₅₀ 47 to LC₉₀ 91 (milligrams per liter), and for C. guianensis, they were LC₅₀ 136 to LC₉₀ 551 (milligrams per liter). In relation to different temperature, the effectiveness of the oils on larvae mortality was directly related to the increase of temperature, and better results were observed for temperature at 25°C. The results presented here indicate the potential larvicidal activity of C. guianensis and species of Copaifera, in populations of A. aegypti from the wild. Therefore, the results presented here are very important since such populations are primarily responsible for transmitting the dengue virus in the environment.     

Subcellular localization of an extracellular serine protease in<Emphasis Type="Italic"> Leishmania</Emphasis> (<Emphasis Type="Italic">Leishmania</Emphasis>)<Emphasis Type="Italic"> amazonensis</Emphasis>

Extracellular proteolytic activity was detected in a Leishmania (L.) amazonensis culture supernatant and a 56-kDa protein was purified using (NH₄)₂SO₄ precipitation followed by affinity chromatography on aprotinin–agarose. A rabbit serum obtained against the 56-kDa extracellular serine protease was used in order to analyze its location in L. (L.) amazonensis parasites. Immunocytochemistry studies revealed that the enzyme is mainly found in the flagellar pocket and cytoplasmic vesicles of promastigote forms, whereas in amastigotes, it is located in electron-dense structures resembling megasomes. These results indicate that the 56-kDa serine protease is released into the extracellular environment through the flagellar pocket; and its intracellular location suggests either a correlated enzymatic activity or intracellular trafficking.     

<Emphasis Type="Italic">Leishmania</Emphasis> (<Emphasis Type="Italic">Viannia</Emphasis>) <Emphasis Type="Italic">shawi</Emphasis> purified antigens confer protection against murine cutaneous leishmaniasis

Objective  

Leishmania (Viannia) shawi was characterized only recently, and few studies concerning the immunogenic and protective properties of its antigens have been performed. The present study aimed to evaluate the protective potential of the five antigenic fractions isolated from L. (V.) shawi promastigotes in experimental cutaneous leishmaniasis.     

Occurrence of <Emphasis Type="Italic">Cryptosporidium hominis</Emphasis> in pigeons (<Emphasis Type="Italic">Columba livia</Emphasis>)

This study demonstrated the presence of Cryptosporidium hominis in pigeons for the first time. Previously, C. hominis had been cited only in another bird species, Branta canadiensis. The present findings suggest that pigeons may act as mechanical vectors for this protozoan.     

Epidemiology of invasive neonatal <Emphasis Type="Italic">Cronobacter</Emphasis> (<Emphasis Type="Italic">Enterobacter sakazakii</Emphasis>) infections

About 120–150 neonatal Cronobacter spp. (Enterobacter sakazakii) infections have been described. An analysis of current case numbers, epidemiological measures and risk factors is warranted. Data of microbiologically confirmed cases, published between 2000 and 2008, have been analysed statistically. More than 100 neonatal Cronobacter infections have been reported in this period. The overall lethality of the 67 invasive infections was 26.9%. The lethality of Cronobacter meningitis, bacteraemia and necrotising enterocolitis (NEC) was calculated to be 41.9% (P < 0.0001), <10% and 19.0% (P < 0.05), respectively. Logistic regression models (P < 0.0001) revealed a higher gestational age at birth and parentage not from Europe as significant factors for a higher reporting probability of neonatal Cronobacter meningitis. Neonates with Cronobacter meningitis not originating from North America have a higher risk for lethal outcome than other neonatal Cronobacter infections (P < 0.0001). Continental differences of risk factors for Cronobacter meningitis and for the lethal outcome of neonatal meningitis should be elucidated. Neonatal Cronobacter infections are mainly associated with the contamination of infant formula and of the relevant cleaning and preparation equipment. Eleven neonatal Cronobacter infections, not caused by contaminated infant formula, have been retrieved. Other environmental sources of infection should be considered. Consistent and sufficiently informative data of invasive neonatal Cronobacter infections should be recorded in a centralized reporting system.     

Larvicidal efficacy of <Emphasis Type="Italic">Jatropha curcas</Emphasis> and bacterial insecticide, <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis>, against lymphatic filarial vector<Emphasis Type="Italic">, Culex quinquefasciatus</Emphasis> Say (Diptera: Culicidae)

The present study explored the effects of Jatropha curcas leaf extract and Bacillus thuringiensis israelensis larvicidal activity against the lymphatic filarial vector, Culex quinquefasciatus. Wights were selected for investigating the larvicidal potential against the first to fourth instar larvae of the laboratory-reared mosquito species, C. quinquefasciatus Say, in which the major lymphatic filariasis was used. The medicinal plants were collected from the area around Bharathiar University, Coimbatore. The dried plant materials were powdered by an electric blender. From the powder, 100 g of the plant materials was extracted with 300 ml of organic solvents of methanol for 8 h, using a Soxhlet apparatus, and filtered. The crude plant extracts were evaporated to dryness in a rotary vacuum evaporator. The plant extract showed larvicidal effects after 24 h of exposure; however, the highest larval mortality was found in the leaf extract of methanol J. curcas against the first to fourth instar larvae of values LC₅₀ = 1.200%, 1.290%, 1.358%, and 1.448% and LC₉₀ = 2.094%, 2.323%, 2.444%, and 2.544% and B. thuringiensis israelensis against the first to fourth instar larvae of values LC₅₀ = 9.332%, 9.832%, 10.212%, 10.622% and LC₉₀ = 15.225%, 15.508%, 15.887%, and 15.986% larvae of C. quinquefasciatus, respectively. No mortality was observed in the control. These results suggest methanol extracts of J. curcas and B. thuringiensis israelensis have potential to be used as an ideal eco-friendly approach for the control of the major lymphatic filarial vector, C. quinquefasciatus.     

Occurrence of cystacanths of <Emphasis Type="Italic">Plagiorhynchus cylindraceus</Emphasis> (Acanthocephala) in the terrestrial isopods <Emphasis Type="Italic">Trachelipus squamuliger</Emphasis> and <Emphasis Type="Italic">Armadillidium vulgare</Emphasis> (Oniscidea) in Bulgaria

In total, 2097 individuals of Trachelipus squamuliger and 20 individuals of Armadillidium vulgare from four habitats (three woodland sites and one pasture) in the region of Stara Zagora, Bulgaria, were examined for the presence of cystacanths of Plagiorhynchus cylindraceus, a common acanthocephalan parasite of passerine birds. In T. squamuliger from woodland habitats, cystacanths were found with prevalence 4.0–9.3%, intensity 1–5 (mean 1.22–1.57) and mean abundance 0.057–0.113. No significant differences were observed between infections in males and females of T. squamuliger. None of the T. squamuliger individuals from the pasture examined was infected. Out of 48 infected females of T. squamuliger, only one had developed eggs (in agreement with previous studies revealing the negative effect of the cystacanths on the development of female gonads of woodlice). One individual of A. vulgare was infected with a single cystacanth. The occurrence of P. cylindraceus in T. squamuliger is a new host record.