Detection of severe fever with thrombocytopenia syndrome (SFTS) virus (SFTSV) during the early phase of the disease is important for appropriate treatment, infection control, and prevention of further transmission. The reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a nucleic acid amplification method that amplifies the target sequence under isothermal conditions. Here, we developed an RT-LAMP with a novel primer/probe set targeting a conserved region of the SFTSV L segment after extraction of viral RNA (standard RT-LAMP). Both the Chinese and Japanese SFTSV strains, including various genotypes, were detected by the standard RT-LAMP. We also performed RT-LAMP using the same primer/probe set but without the viral RNA extraction step (called simplified RT-LAMP) and evaluated the diagnostic efficacy. The sensitivity and specificity of the simplified RT-LAMP were 84.9% (45/53) and 89.5% (2/19), respectively. The simplified RT-LAMP can detect SFTSV in human sera containing >10^3.5 copies/mL viral RNA. The two RT-LAMP positive but quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) negative samples were positive in the conventional RT-PCR, suggesting that there was no false positive reaction in the RT-LAMP. Both the standard and simplified RT-LAMP are useful for detecting the SFTSV genome in patients during the early phase of the disease.

Malaria continues to be imported into Japan. To better assess the risk of imported malaria, we describe malaria species, suspected country/area of infection (accounting for the number of travelers), demographic characteristics, clinical manifestation, and healthcare access, based on the national surveillance data from 2006 to 2014. Among 557 cases, the median age was 33 years (range: 1–83 years), and 76% were male; 306 (55%) were classified as Japanese based on the reported name. The majority were Plasmodium falciparum infections (58%), followed by Plasmodium vivax infections (30%). Most P. falciparum cases were acquired in Africa and P. vivax in Asia/Oceania. Notification rates per 10,000 Japanese travelers for P. falciparum were highest for Africa, P. vivax were highest for Asia/Oceania, and high for both species for Papua New Guinea. Ten percent of the cases were clinically severe at the time of notification. Nearly 80% of severe cases were P. falciparum infections, and among P. falciparum cases, Japanese ethnicity was associated with severe case status (P = 0.03). Plasmodium falciparum cases among Japanese cases showed that older age (≥ 50 years) was associated with severe case status (odds ratio = 5.4; 95% confidence interval = 1.9–15.2), adjusted for sex and healthcare access. More informative assessments are possible by accounting for the number of travelers. Older Japanese represent an important demographic to target prevention and early treatment efforts for malaria.

JC polyomavirus (JCV) is the causative agent of progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the central nervous system in immunosuppressed patients. PML usually has a poor prognosis. Detection and quantification of the JCV genome in cerebrospinal fluid (CSF) is an efficacious tool for the diagnosis and management of PML, for which proper therapeutic interventions are required. A loop-mediated isothermal amplification (LAMP) assay was applied for the quantitative detection of JCV. The LAMP assay was evaluated for the efficacy in diagnosis of PML in comparison with the TaqMan-based quantitative real-time PCR (qPCR) assay using 153 CSF specimens collected from patients with suspected PML. The LAMP assay showed no cross-reactivity against other polyomavirus plasmids, viral DNA, and viral RNA, which causes encephalitis, and detected 1 copy of the standard DNA per reaction. Among 50 qPCR-positives, 42 specimens (containing JCV genome ranged from 3.2 × 100 to 3.2 × 106 copies/reaction) showed positive reactions and 8 specimens (containing 0.9 to 19.9 copies/reaction) showed negative in the LAMP assay. Furthermore, 3 of 103 qPCR-negative specimens showed positive reactions in the LAMP assay. The sensitivity, specificity, positive predictive value, and negative predictive values of the LAMP assay were 84% (42/50), 97% (100/103), 93% (42/45), and 93% (100/108), respectively. The kappa statistic was 0.83. The JCV loads determined by the LAMP assay showed a strong positive correlation with those determined by the qPCR assay for 33 specimens with copy numbers of ≥1 copies/reaction (r = 0.89). Additionally, the LAMP assay could monitor the JCV genome copy number in CSF for sequential samples equivalently to qPCR assay. The newly developed LAMP assay is highly specific against JCV and detect the JCV genome in the sample DNA containing 20 or more copies of JCV genome per reaction with 100% sensitivity (n = 29), which corresponds to ≥3 × 103 copies/mL of CSF. The LAMP assay is useful for the diagnosis and offers valuable information for the evaluation and management of PML in the clinical setting.

Herpes simplex virus type 1 (HSV-1) acyclovir (ACV) resistance is acquired by mutations in the viral thymidine kinase (TK) or DNA polymerase (DNApol) genes. We previously obtained an ACV-resistant clone (HSV-1_VZV_TK_clone α) by sequential passages of HSV-1_VZV-TK, a recombinant virus which lacked its endogenous TK activity and instead expressed the varicella-zoster virus (VZV) TK ectopically. HSV-1_VZV_TK_clone α had been generated using an HSV-1_BAC in the presence of increasing concentrations of ACV. The ACV-resistant clone bore normal TK and DNApol genes. Here, we deployed next-generation full-genome sequencing of HSV-1_VZV_TK_clone α and identified a single nucleotide substitution, resulting in a P597L missense mutation in the UL29 gene product, the ICP8 protein. Recombinant HSV-1 encoding a P597L ICP8 protein was generated, and its properties and ability to confer drug resistance were analyzed. No difference in virus growth and UL29 expression was observed between the mutant recombinant, the wild type, and a revertant mutant viral strain, and susceptibility tests of these strains to ACV and other drugs using Vero, HEL, and ARPE19 cells identified that the recombinant UL29 mutant virus was resistant only to ACV. These results indicate that ICP8 may be involved in the anti-herpesvirus drugs’ mechanism of action on HSV-1.