D-allose and D-psicose reinforce the action of metronidazole on trichomonad.

The effects of D-allose and D-psicose on Tritrichomonas foetus were examined. They were cultured in F-bouillon medium including glucose, but had never increased when glucose was substituted to those sugars. When cultured in a medium including a dose of ED(50) metronidazole and those sugars, trichomonad density was significantly less than that in a medium with metronidazole only. D-Allose remarkably reinforced the action of metronidazole. This means there are some interactions between metronidazole and those sugars. Although the mechanism is not clear, by using those sugars for treatment with metronidazole, the drug dosage could be lowered and the development of drug resistance of trichomonad parasites might be prevented.

Implementation of a novel PCR based method for detecting malaria parasites from naturally infected mosquitoes in Papua New Guinea.

BACKGROUND: Detection of Plasmodium species in mosquitoes is important for designing vector control studies. However, most of the PCR-based detection methods show some potential limitations. The objective of this study was to introduce an effective PCR-based method for detecting Plasmodium vivax and Plasmodium falciparum from the field-caught mosquitoes of Papua New Guinea. METHODS: A method has been developed to concurrently detect mitochondrial cytochrome b (Cyt b) of four human Plasmodium species using PCR (Cytb-PCR). To particularly discriminate P. falciparum from P. vivax, Plasmodium ovale and Plasmodium malariae, a polymerase chain reaction-repeated fragment length polymorphism (PCR-RFLP) has further been developed to use with this method. However, due to limited samples number of P. ovale and P. malariae; this study was mainly confined to P. vivax and P. falciparum. The efficiency of Cytb-PCR was evaluated by comparing it with two 'gold standards' enzyme linked immunosorbent assay specific for circumsporozoite protein (CS-ELISA) using artificially infected mosquitoes; and nested PCR specific for small subunit ribosomal RNA (SSUrRNA) using field caught mosquitoes collected from three areas (Kaboibus, Wingei, and Jawia) of the East Sepic Province of Papua New Guinea. RESULTS: A total of 90 mosquitoes were artificially infected with three strains of Plasmodium: P. vivax-210 (n = 30), P. vivax-247 (n = 30) and P. falciparum (n = 30). These infected mosquitoes along with another 32 unfed mosquitoes were first checked for the presence of Plasmodium infection by CS-ELISA, and later the same samples were compared with the Cytb-PCR. CS-ELISA for P. vivax-210, P. vivax-247 and P. falciparum detected positive infection in 30, 19 and 18 mosquitoes respectively; whereas Cytb-PCR detected 27, 16 and 16 infections, respectively. The comparison revealed a close agreement between the two assays (kappa = 0.862, 0.842 and 0.894, respectively for Pv-210, Pv-247 and P. falciparum groups). It was found that the eight CS-ELISA-positive mosquitoes detected negative by Cytb-PCR were false-positive results. The lowest detection limit of this Cytb-PCR was 10 sporozoites. A highly concordance result was also found between nested PCR and Cytb-PCR using 107 field caught mosquitoes, and both tests concordantly detected P. falciparum in an Anopheles punctulatus mosquito collected from Kaboibus. Both tests thus suggested an overall sporozoite rate of 0.9% (1/107) in the study areas. Subsequently, PCR-RFLP efficiently discriminated P. falciparum from P. vivax for all of the Cytb-PCR positive samples. CONCLUSION: A single step PCR based method has been introduced here that is highly sensitive, efficient and reliable for identifying P. vivax and P. falciparum from mosquitoes. The reliability of the technique was confirmed by its ability to detect Plasmodium as efficiently as those of CS-ELISA and nested PCR. Application of the assay offers the opportunity to detect vector species of Papua New Guinea and may contribute for designing further vector control programmes.

Molecular phylogeography of Culex quinquefasciatus mosquitoes in central Bangladesh.

Mosquitoes in the Culex pipiens complex are a major vector of numerous parasitic and arboviral diseases. Here we report the phylogeography of a prevalent Culex mosquito, Cx. quinquefasciatus, from three locations in Bangladesh: Dhaka, Savar and Mymensingh. Sequence analysis of the genes encoding mitochondrial cytochrome oxidase subunit II, nuclear elongation factor-1 alpha, and acetylcholinesterase-2 revealed the lack of a population genetic structure among the three locations. Moreover, the highly divergent ribosomal internal transcribed spacer 2 suggests that this locus has not evolved in concert. The results further show evidence of historical introgression of internal transcribed spacer 2 from Cx. pipiens to Cx. quinquefasciatus of Bangladesh, and that the introgression occurred before Cx. quinquefasciatus had dispersed within this region. The study also reveals historical population expansion in this region, followed by a post-expansion Wolbachia sweep.

Genetic diversity in two sibling species of the Anopheles punctulatus group of mosquitoes on Guadalcanal in the Solomon Islands.

BACKGROUND: The mosquito Anopheles irenicus, a member of the Anopheles punctulatus group, is geographically restricted to Guadalcanal in the Solomon Islands. It shows remarkable morphological similarities to one of its sibling species, An. farauti sensu stricto (An. farauti s.s.), but is dissimilar in host and habitat preferences. To infer the genetic variations between these two species, we have analyzed mitochondrial cytochrome oxidase subunit II (COII) and nuclear ribosomal internal transcribed spacer 2 (ITS2) sequences from Guadalcanal and from one of its nearest neighbours, Malaita, in the Solomon Islands. RESULTS: An. farauti s.s. was collected mostly from brackish water and by the human bait method on both islands, whereas An. irenicus was only collected from fresh water bodies on Guadalcanal Island. An. irenicus is distributed evenly with An. farauti s.s. (Phi SC = 0.033, 0.38%) and its range overlaps in three of the seven sampling sites. However, there is a significant population genetic structure between the species (Phi CT = 0.863, P < 0.01; Phi ST = 0.865, P < 0.01 and FST = 0.878, P < 0.01). Phylogenetic analyses suggest that An. irenicus is a monophyletic species, not a hybrid, and is closely related to the An. farauti s.s. on Guadalcanal. The time estimator suggests that An. irenicus diverged from the ancestral An. farauti s.s. on Guadalcanal within 29,000 years before present (BP). An. farauti s.s. expanded much earlier on Malaita (texp = 24,600 BP) than the populations on Guadalcanal (texp = 16,800 BP for An. farauti s.s. and 14,000 BP for An. irenicus). CONCLUSION: These findings suggest that An. irenicus and An. farauti s.s. are monophyletic sister species living in sympatry, and their populations on Guadalcanal have recently expanded. Consequently, the findings further suggest that An. irenicus diverged from the ancestral An. farauti s.s. on Guadalcanal.

Identification of an eosinophil chemotactic factor from anopheline mosquitoes as a chitinase family protein.

Anopheline mosquitoes play an essential role in malaria transmission. The mosquito salivates copiously when probing for the location of a blood vessel. We found that the saliva of anopheline mosquitoes has chemotactic activity for naive eosinophils or neutrophils. The major eosinophil chemotactic component in saliva was shown to be one of the chitinase family proteins. A similar chitinase family protein was found also in the midgut of the anopheline mosquito. Production of antibodies to the chitinase family protein was generally observed in the sera of residents of a malaria endemic area. Both Plasmodium falciparum-infected and uninfected individuals had antibodies to chitinases. These results suggest that the chitinase family protein in mosquito saliva contributes to eliciting an inflammatory response of eosinophils in the host skin followed by antibody production in the host.

Intertidal snail-trematode communities on the southern Thailand before and after the South Asia tsunami.

Intertidal snail-trematode communities in southern Thailand were examined before and after the South Asia tsunami. Infection rates and species diversity of cercaria in the host snail Cerithidea in tidal zones did not change significantly from one year before to one month after the tsunami. However, the host snails C. quadrata, C. alata and C. obtusa disappeared from greatly damaged sites. It is important to follow up on the intertidal snail-trematode community recovery process after destruction of the intertidal ecosystem.

Mosquito salivary gland extracts induce EBV-infected NK cell oncogenesis via CD4 T cells in patients with hypersensitivity to mosquito bites.

Severe hypersensitivity to mosquito bites (HMB) is characterized by intense local skin reactions and systemic symptoms such as high fever, lymphadenopathy, and hepatosplenomegaly. Patients with HMB often have natural killer (NK) cell lymphocytosis associated with Epstein-Barr virus (EBV) infection. Here we investigated whether mosquito bites have any influence on the oncogenesis of EBV-infected NK cells. We examined six HMB patients with EBV-infected NK cell lymphocytosis. We first demonstrated that CD4+ T cells, but not NK cells, proliferated well in response to mosquito salivary gland extracts (SGE), especially to SGE of Aedes albopictus. When NK cells were cocultured with autologous CD4+ T cells stimulated by mosquito SGE, the expression of viral oncogene latent membrane protein 1 (LMP1) was remarkably enhanced. Next, we stimulated mononuclear cells of the patients with mosquito SGE, and NK cell counts were monitored for 28 d. The counts changed little from initial levels in the culture with mosquito SGE, whereas they decreased steadily in the culture without the extracts. Furthermore, we detected LMP1 mRNA in the skin lesion induced by mosquito SGE. These results suggest that mosquito bites can induce expression of the viral oncogene LMP1 in NK cells via mosquito antigen-specific CD4+ T cells, which is involved in the oncogenesis of NK cells in vivo.

Three-dimensional reconstruction of the distribution of flame cells in a trematode cercaria.

In order to reveal the vertical distribution of flame cells in a trematode cercaria, horizontal serial semi-thin sections were reconstructed by an image analyzer and a manual procedure. The ascending main excretory tubes extended forward, meandering up and down at the digestive system level from the excretory bladder. At the posterior end of the ventral sucker, they curved dorsally, passed through the dorsal side, returning to the mid level at the anterior end of the ventral sucker. They then ran forward to the posterior part of the oral sucker, and turned back. The flame cells were more abundant in the ventral region than in the dorsal region. The majority of pairs of right and left flame cells were present at the same distance from the digestive system or ventral (or dorsal) surface. This method, using conventional techniques, has some advantages for determining the horizontal and vertical distributions of flame cells in cercaria and may, therefore, promote research on cercaria.

CD4+ T-lymphocyte-induced Epstein-Barr virus reactivation in a patient with severe hypersensitivity to mosquito bites and Epstein-Barr virus-infected NK cell lymphocytosis.

BACKGROUND: Natural killer (NK) cell lymphocytosis associated with Epstein-Barr virus (EBV) infection often shows severe hypersensitivity to mosquito bites (HMB) characterized by intense local skin reactions and systemic symptoms such as high fever, lymphadenopathy, and hepatosplenomegaly. However, the induction mechanism of HMB is still unclear. OBSERVATIONS: We investigated a typical case of HMB with EBV-positive NK cell lymphocytosis. CD4+ T cells dominantly infiltrated the site of the mosquito bite, while EBV-positive cells were few in comparison. CD4+ T cells, but not CD8+ T cells or NK cells, responded to the mosquito salivary gland extracts. Interestingly, coculturing of the NK cells and CD4+ T cells activated by mosquito extracts induced expression of EBV lytic-cycle proteins in the NK cells. Furthermore, the expression of BZLF1, a viral lytic-cycle transactivator, was detectable at the skin lesion induced by scratch patch testing with mosquito extract. The EBV DNA copy number levels in the plasma were elevated in systemic HMB symptoms compared with the normal condition. CONCLUSIONS: CD4+ T cells are important for the primary skin reaction to mosquito bites and might play a key role in reactivation of latent EBV infection in NK cells. This viral reactivation contributed to the pathogenesis of the infectious mononucleosis-like systemic symptoms of HMB in our present case.

A mathematical model for the transmission of Plasmodium vivax malaria.

We have proposed a mathematical model for the transmission of Plasmodium vivax malaria quantitatively, which is adjusted to the infected region, Guadalcanal, in the Solomon Islands. The simulation of a transmission model will be instrumental in planning the malaria control strategy. A characteristic of the life cycle of P. vivax is that a sporozoite injected into the blood stream by a mosquito bite may sometimes stay in a hepatocyte as a hypnozoite. Therefore, we have incorporated a phenomenon of renewed infections caused by a relapse into the transmission model. Also through the simulations we have attempted to evaluate the decline in prevalence caused by the programs of selective mass drug administration (MDA) and vector control such as the distribution of permethrin-treated bednets. The simulations have indicated that the concentrated repetition of MDA at 1-week intervals would reduce the prevalence of vivax malaria swiftly in the beginning and would keep the parasite rate below 1% for a few years but the prevalence would increase thereafter. In contrast, the parasite rate would remain below 1% for a long time if a trial of 1 or 2 times MDA is accompanied with some reduction of the vectorial capacity by the enforcement of vector control. In any case, it is important to beware of relapse cases because even after the execution of MDA it takes a long time to decrease the proportion of hypnozoite carriers.

Variaciones de la secuencia de nucleótidos en el gen I de subunidad de cytocromo C oxidasa, en especies de Leishmania

Se realizó la determinación de las secuencias parciales de nucleótidos de genes COI de 8 especies de Leishmania como: L (Leishmania) amazonensis, L.(Viannia) equatorensis. L.(V) major-like, L. (L) mexicana, L.(V) major, L.(V) guayanensis, L.(V) brazilensis y L.(V) panamensis por el método de PCR, utilizando primers diseñados para genes COI de paramecium. El presente estudio demostró que los primers son útiles para la amplificación de los genes COI de 4 especies: L.(V) major, L(V) guayanensis, L(V) braziliensis y L.(V) panamensis. Se determinó una secuencia parcial de genes COI de las 4 especies.

Estudios preliminares sobre el diagnóstico de leishmaniasis cutánea utilizando muestras de biopsias de piel por reacción en cadena de polimerasa

Se estudiaron métodos de diagnóstico de leishmaniasis cutánea por reacción en cadena de la polimerasa (PCR), utilizando parásitos vultivados y muestras de biopsias de piel del Ecuador. Se prepararon template ADNs por ebullición por 10 minutos en soluciones Chelex al 5 por ciento. Los parásitos Leishmania fueron detectados por PCR, utilizando primers designados desdel el minicírculo (13A y 13B)y gen mini-exon (S-1629 y S-1630. Los primers primero mencionados amplificaron productos no específicos en ADN humano, y la sensibilidad de la reacción fue baja. Los últimos nunca amplificaron productos específicos aún en templete humano y posibilitaron la identificación a nivel de subgénero. Cuando se aumentó la sensibilidad...

Nuevos estudios sobre el tratamiento oral de la leishmaniasis cutánea, con la droga anti-malárica mefloquina (Mephaquin) en el Ecuador

Desde 1994, debido a la falta de antimoniales en ese año, probamos por primera vez las drogas antimaláricas mefloquina y artesunato, para tratar la leishmaniasis cutánea, y los resultados fueron muy sorprendentes, por su gran eficacia. Este artículo sigue al anterior, con el firme propósito de estudiar un número mayor de pacientes con una nueva dosis de mefloquina, primero, y luego hacer lo mismo con el artesunato, para obtener mayor evidencia de la mencionada capacidad curativa. Un grupo de 72 pacientes seleccionados (Grupos A,B y C), y un grupo al azar de 16 (D), fueron tratados con mefloquina, 100 mg después del desayuno y merienda por días días para adultos o niños con más de 45 kg de peso, y una dosis total...

Estudio sobre el tratamiento oral de la leishmaniasis cutánea con un derivado de la artemisina, al artesunato (Plasmotrim) en el Ecuador

Nuestro trabajo anterior reveló que las drogas antimaláricas, mefloquina (mephaquin) y artesunato (plasmotrim), son eficaces para la terapia oral de pacientes con leishmaniasis cutánea. A fin de determinar si estas drogas tienen efectos similares o diferentes es necesario desarrollar investigaciones más detalladas, basadas en casos adicionales de tratamiento, especialmente con el artesunato. Con este propósito, tratamos a 15 pacientes con leishmaniasis cutánea con artesunato. En el tratamiento se utilizó la siguiente dosificación: 1) para adultos con 45-59 kg de peso, y niños con más de 45 kg y/o mayores de 12 años, 100 mg de Plasmotrim (media tableta) después del desayuno y merienda por 10 días; y 2) para adultos...