Surveillance of Getah virus in mosquitoes and racehorses from 2016 to 2019 at a training center in Ibaraki Prefecture, Japan, a site of several previous Getah virus outbreaks.

Mosquitoes and EDTA-treated blood samples from febrile racehorses were investigated for Getah virus infection from 2016 to 2019 at the Miho Training Center, where several outbreaks of Getah virus have occurred. We collected 5557 mosquitoes and 331 blood samples from febrile horses in this study. The most frequently captured mosquito species was Culex tritaeniorhynchus (51.

A method of sequential liquid dispensing for the multiplexed genetic diagnosis of viral infections in a microfluidic device.

In this study, we introduce polydimethylsiloxane (PDMS)-based microfluidic devices capable of sequential dispensing of samples into multiple reaction microchambers in a single operation to provide a fast and easy sample-to-answer platform for multiplexed genetic diagnosis of multiple viral infectious diseases. This approach utilizes the loop-mediated isothermal amplification (LAMP) method to amplify and detect specific nucleic acid (DNA/RNA) targets. We present a microfluidic flow control theory for sequential liquid dispensing phenomena, which provides design guidelines for device optimization.

Detection and discrimination of multiple strains of Zika virus by reverse transcription-loop-mediated isothermal amplification.

Background: Monitoring both invasion of Zika virus disease into free countries and circulation in endemic countries is essential to avoid a global pandemic. However, the difficulty lies in detecting Zika virus due to the large variety of mutations in its genomic sequence. To develop a rapid and simple method with high accuracy, reverse transcription-loop-mediated isothermal amplification (RT-LAMP) was adopted for the detection of Zika virus strains derived from several countries.

Intensive diagnostic management of coronavirus disease 2019 (COVID-19) in academic settings in Japan: challenge and future.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), first emerged in Wuhan, China, and has spread globally to most countries. In Japan, the first COVID-19 patient was identified on January 15, 2020. By June 30, the total number of patients diagnosed with COVID-19 reached 18,000.

Seroprevalence and associated factors of Toxoplasma gondii among HIV-infected patients in Tokyo: A cross sectional study.

HIV infection, in particular in patients with developing AIDS, carries a risk of causing toxoplasmosis with encephalitis, which is mostly caused by a form (bradyzoite) of the protozoan parasite Toxoplasma gondii. HIV/AIDS in Japan has been recognized as a serious health issue in recent years. In this study, to elucidate T.

Seroprevalence of Toxoplasma gondii in wild sika deer in Japan.

Toxoplasmosis is a food-borne infection that is widespread around the world, causing congenital disorders and opportunistic infections. Ingestion of undercooked meat is one of the risk factors for infection with the causative agent, Toxoplasma gondii. Japanese people occasionally eat rare meat as a traditional cuisine style called "Sashimi".

Distribution of tick-borne diseases in Japan: Past patterns and implications for the future.

The rapid geographical spread of tick-borne diseases (TBDs) worldwide has recently provoked significant concerns amongst public health authorities. Tick-borne pathogens are maintained in enzootic cycles involving ticks and wild animal hosts, with epizootic spread to other mammals, including livestock and humans. Despite the increasing public health concern, current TBD diagnostic tests and treatments are inadequate, and predictive models of future risks posed by TBDs are limited by the heterogeneity of environmental, vector, and host factors, even in neighboring regions.

Plant hormone cytokinins control cell cycle progression and plastid replication in apicomplexan parasites.

Cytokinins are plant hormones that are involved in regulation of cell proliferation, cell cycle progression, and cell and plastid development. Here, we show that the apicomplexan parasites Toxoplasma gondii and Plasmodium berghei, an opportunistic human pathogen and a rodent malaria agent, respectively, produce cytokinins via a biosynthetic pathway similar to that in plants. Cytokinins regulate the growth and cell cycle progression of T.

A host cell membrane microdomain is a critical factor for organelle discharge by Toxoplasma gondii.

Host cell microdomains are involved in the attachment, entry, and replication of intracellular microbial pathogens. Entry into the host cell of Toxoplasma gondii and the subsequent survival of this protozoan parasite are tightly coupled with the proteins secreted from organelle called rhoptry. The rhoptry proteins are rapidly discharged into clusters of vesicles, called evacuoles, which are then delivered to parasitophorous vacuoles (PVs) or nucleus.

High-Resolution Vertical Observation of Intracellular Structure Using Magnetically Responsive Microplates.

A vertical confocal observation system capable of high-resolution observation of intracellular structure is demonstrated. The system consists of magnet-active microplates to rotate, incline, and translate single adherent cells in the applied magnetic field. Appended to conventional confocal microscopes, this system enables high-resolution cross-sectional imaging with single-molecule sensitivity in single scanning.

Plant Hormone Salicylic Acid Produced by a Malaria Parasite Controls Host Immunity and Cerebral Malaria Outcome.

The apicomplexan parasite Toxoplasma gondii produces the plant hormone abscisic acid, but it is unclear if phytohormones are produced by the malaria parasite Plasmodium spp., the most important parasite of this phylum. Here, we report detection of salicylic acid, an immune-related phytohormone of land plants, in P.

Magnetically responsive microflaps reveal cell membrane boundaries from multiple angles.

A microflap system to incline adherent cells in the desired orientation is described. Inclination angles of cell-laden microflaps are precisely controlled by the applied magnetic field, enabling us to observe cell-membrane boundaries from multiple angles. This system is equipped with conventional microscopes, allowing clear focused images of cell-membrane boundaries to be obtained with high magnification.

Parylene mobile microplates integrated with an enzymatic release for handling of single adherent cells.

An approach for manipulating single adherent cells is developed that is integrated with an enzymatic batch release. This strategy uses an array of releasable microfabricated mobile substrates, termed microplates, formed from a biocompatible polymer, parylene. A parylene microplate array of 10-70 μm in diameter can be formed on an alginate hydrogel sacrificial layer by using a standard photolithographic process.

Detection of mutation by allele-specific loop-mediated isothermal amplification (AS-LAMP).

For effective control of pathogen-transmitting mosquitoes, precise surveillance data of mosquito distribution are essential. Recently, an increase of insecticide resistance due to the kdr mutation in Anopheles gambiae, a mosquito that transmits the malaria parasite, has been reported. With the aim of developing a simple and effective method for surveying resistant mosquitoes, LAMP was applied to the allele-specific detection of the kdr gene in An.

Intra-specific diversity of Serratia marcescens in Anopheles mosquito midgut defines Plasmodium transmission capacity.

A critical stage in malaria transmission occurs in the Anopheles mosquito midgut, when the malaria parasite, Plasmodium, ingested with blood, first makes contact with the gut epithelial surface. To understand the response mechanisms within the midgut environment, including those influenced by resident microbiota against Plasmodium, we focus on a midgut bacteria species' intra-specific variation that confers diversity to the mosquito's competency for malaria transmission. Serratia marcescens isolated from either laboratory-reared mosquitoes or wild populations in Burkina Faso shows great phenotypic variation in its cellular and structural features.

Multivariable analysis of host amino acids in plasma and liver during infection of malaria parasite Plasmodium yoelii.

Background: Malaria is the most significant human parasitic disease, and yet understanding of the energy metabolism of the principle pathogen, Plasmodium falciparum, remains to be fully elucidated. Amino acids were shown to be essential nutritional requirements since early times and much of the current knowledge of Plasmodium energy metabolism is based on early biochemical work, performed using basic analytical techniques, carried out almost exclusively on human plasma with considerable inter-individual variability.

Methods: In order to further characterize the fate of amino acid metabolism in malaria parasite, multivariate analysis using statistical modelling of amino acid concentrations (aminogram) of plasma and liver were determined in host infected with rodent malaria parasite, Plasmodium yoelii.

Development of an allele-specific, loop-mediated, isothermal amplification method (AS-LAMP) to detect the L1014F kdr-w mutation in Anopheles gambiae s. l.

Background: Malaria control relies heavily on treated bed nets and indoor residual spraying with pyrethroid insecticides. Unfortunately, the resistance to pyrethroid insecticides, mainly due to the kdr mutation, is spreading in the main malaria vector Anopheles gambiae s.l.

The role of proboscis of the malaria vector mosquito Anopheles stephensi in host-seeking behavior.

Background: The proboscis is an essential head appendage in insects that processes gustatory code during food intake, particularly useful considering that blood-sucking arthropods routinely reach vessels under the host skin using this proboscis as a probe.

Results: Here, using an automated device able to quantify CO(2)-activated thermo (35°C)-sensing behavior of the malaria vector Anopheles stephensi, we uncovered that the protruding proboscis of mosquitoes contributes unexpectedly to host identification from a distance. Ablation experiments indicated that not only antennae and maxillary palps, but also proboscis were required for the identification of pseudo-thermo targets.

A single fluorescence-based LAMP reaction for identifying multiple parasites in mosquitoes.

Vector-borne diseases, such as malaria and lymphatic filariasis, are co-endemic in large parts of the world. To develop a multiplex amplification method for the simultaneous detection of multiple insect-borne infectious diseases, we used LAMP with fluorescently labeled primers to identify the SPECT2 gene of Plasmodium berghei and the cytochrome oxidase subunit I gene of Dirofilaria immitis in mosquitoes. This technique could detect as few as 100 P.

p38 MAPK-dependent phagocytic encapsulation confers infection tolerance in Drosophila.

Hosts employ a combination of two distinct yet compatible strategies to defend themselves against parasites: resistance, the ability to limit parasite burden, and tolerance, the ability to limit damage caused by a given parasite burden. Animals typically exhibit considerable genetic variation in resistance to a variety of pathogens; however, little is known about whether animals can evolve tolerance. Using a bacterial infection model in Drosophila, we uncovered a p38 MAP kinase-mediated mechanism of tolerance to intracellular bacterial infection as measured by the extent to which the host's survival rate increased or was maintained despite increasing bacterial burden.

Intrinsic tumor suppression and epithelial maintenance by endocytic activation of Eiger/TNF signaling in Drosophila.

Oncogenic alterations in epithelial tissues often trigger apoptosis, suggesting an evolutionary mechanism by which organisms eliminate aberrant cells from epithelia. In Drosophila imaginal epithelia, clones of cells mutant for tumor suppressors, such as scrib or dlg, lose their polarity and are eliminated by cell death. Here, we show that Eiger, the Drosophila tumor necrosis factor (TNF), behaves like a tumor suppressor that eliminates oncogenic cells from epithelia through a local endocytic JNK-activation mechanism.

Loop-mediated isothermal amplification applied to filarial parasites detection in the mosquito vectors: Dirofilaria immitis as a study model.

Background: Despite recent advances in our understanding of the basic biology behind transmission of zoonotic infectious diseases harbored by arthropod vectors these diseases remain threatening public health concerns. For effective control of vector and treatment, precise sampling indicating the prevalence of such diseases is essential. With an aim to develop a quick and simple method to survey zoonotic pathogen-transmitting vectors, LAMP (loop-mediated isothermal amplification) was applied to the detection of filarial parasites using a filarial parasite-transmitting experimental model that included one of the mosquito vectors, Aedes aegypti, and the canine heartworm, Dirofilaria immitis.

Rapid recruitment of innate immunity regulates variation of intracellular pathogen resistance in Drosophila.

Genetic variation in susceptibility to pathogens is a central concern both to medicine and agriculture and to the evolution of animals. Here, we have investigated the link between such natural genetic variation and the immune response in wild-type Drosophila melanogaster, a major model organism for immunological research. We found that within nine wild-type strains, different Drosophila genotypes show wide-ranging variation in their ability to survive infection from the pathogenic bacteria Listeria monocytogenes.

Rapid identification of virus-carrying mosquitoes using reverse transcription-loop-mediated isothermal amplification.

Mosquitoes are critical vectors in many arboviral transmission cycles. Considering the increasing incidence of arboviral infections throughout the world, monitoring of vector populations for the presence of an arbovirus could be considered an important initial step of risk assessment to humans and animals. In response to this need, increased efforts to develop rapid and reliable diagnostic techniques have been undertaken; a single-step reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay was developed to detect virus in vector mosquitoes (Aedes aegypti) using the Flock House Virus (FHV) as a model.