To understand the molecular mechanisms of parasitism in vivo, it is essential to elucidate how the transcriptomes of the human hosts and the infecting parasites affect one another. Here we report the RNA-seq analysis of 116 Indonesian patients infected with the malaria parasite Plasmodium falciparum (Pf). We extracted RNAs from their peripheral blood as a mixture of host and parasite transcripts and mapped the RNA-seq tags to the human and Pf reference genomes to separate the respective tags. We were thus able to simultaneously analyze expression patterns in both humans and parasites. We identified human and parasite genes and pathways that correlated with various clinical data, which may serve as primary targets for drug developments. Of particular importance, we revealed characteristic expression changes in the human innate immune response pathway genes including TLR2 and TICAM2 that correlated with the severity of the malaria infection. We also found a group of transcription regulatory factors, JUND, for example, and signaling molecules, TNFAIP3, for example, that were strongly correlated in the expression patterns of humans and parasites. We also identified several genetic variations in important anti-malaria drug resistance-related genes. Furthermore, we identified the genetic variations which are potentially associated with severe malaria symptoms both in humans and parasites. The newly generated data should collectively lay a unique foundation for understanding variable behaviors of the field malaria parasites, which are far more complex than those observed under laboratory conditions.

It has been previously reported that the production of interleukin‐6 (IL‐6) is often enhanced in systemic lupus erythematosus (SLE). The authors examined the secretion of IL‐6, tumour necrosis factor‐α (TNF‐α), granulocyte–macrophage colony‐stimulating factor, IL‐1α and IL‐4 by B cells and monocytes from lupus patients and compared this to the production in normal controls and in rheumatoid arthritis patients. IL‐6 production was increased an average of 3.4‐fold compared to that in normal subjects and 8.4‐fold compared to rheumatoid arthritis patients. In SLE, a strongly positive correlation was found between the levels of IL‐6 and TNF‐α (R = 0.8987, P = 0.002). Since production of both IL‐6 and TNF‐α is regulated by IL‐10, the enhancement of the production of these cytokines could reflect a defect in either IL‐10 production or responsiveness. However, spontaneous production of IL‐10 was enhanced in cultures of B cells and monocytes from lupus patients, compared to normal controls, the levels being increased 3.1‐ to 6‐fold for monocytes and B cells, respectively. The finding of increased secretion of these cytokines implies an abnormality in IL‐10‐mediated suppression in SLE. To assess this possibility, the authors examined recombinant human IL‐10‐mediated suppression of IL‐6 production by monocytes and B cells from lupus patients, compared to normal controls, and found that whereas IL‐10 caused a concentration‐dependent suppression of IL‐6 production in normal B cells and monocytes, this suppression was deficient in B cells and monocytes from lupus patients. In SLE, it therefore appears that there may be an intrinsic defect in IL‐10‐induced suppression of cytokine synthesis. This could explain the increased levels of IL‐10 and IL‐6 found in this condition, and may also be responsible for the characteristic polyclonal B‐cell activation that is seen.

The recent development of a nanopore-type portable DNA sequencer has changed the way we think about DNA sequencing. We can perform sequencing directly in the field, where we collect the samples. Here, we report the development of a novel method to detect and genotype tropical disease pathogens, using dengue fever as a model. By combining the sequencer with isothermal amplification that only requires a water bath, we were able to amplify and sequence target viral genomes with ease. Starting from a serum sample, the entire procedure could be finished in a single day. The analysis of blood samples collected from 141 Indonesian patients demonstrated that this method enables the clinical identification and serotyping of the dengue virus with high sensitivity and specificity. The overall successful detection rate was 79%, and a total of 58 SNVs were detected. Similar analyses were conducted on 80 Vietnamese and 12 Thai samples with similar performance. Based on the obtained sequence information, we demonstrated that this approach is able to produce indispensable information for etiologically analyzing annual or regional diversifications of the pathogens.

Here, we report the application of a portable sequencer, MinION, for genotyping the malaria parasite Plasmodium falciparum. In the present study, an amplicon mixture of nine representative genes causing resistance to anti-malaria drugs is diagnosed. First, we developed the procedure for four laboratory strains (3D7, Dd2, 7G8, and K1), and then applied the developed procedure to ten clinical samples. We sequenced and re-sequenced the samples using the obsolete flow cell R7.3 and the most recent flow cell R9.4. Although the average base-call accuracy of the MinION sequencer was 74.3%, performing >50 reads at a given position improves the accuracy of the SNP call, yielding a precision and recall rate of 0.92 and 0.8, respectively, with flow cell R7.3. These numbers increased significantly with flow cell R9.4, in which the precision and recall are 1 and 0.97, respectively. Based on the SNP information, the drug resistance status in ten clinical samples was inferred. We also analyzed K13 gene mutations from 54 additional clinical samples as a proof of concept. We found that a novel amino-acid changing variation is dominant in this area. In addition, we performed a small population-based analysis using 3 and 5 cases (K13) and 10 and 5 cases (PfCRT) from Thailand and Vietnam, respectively. We identified distinct genotypes from the respective regions. This approach will change the standard methodology for the sequencing diagnosis of malaria parasites, especially in developing countries.

The authors would like to correct the omission of DNA Data Bank of Japan accession number DRA001875 from the initial publication of this article. The data under this accession number are associated with the sequencing of RNA isolated from control samples (i.e., peripheral blood samples of healthy people and people with other infectious diseases). Please note the corrected Data access section in the text is as follows: "The sequencing data from this study have been submitted to the DNA Data Bank of Japan (DDBJ; http://www. ddbj.nig.ac.jp/) under accession numbers DRA000949 and DRA001875." The authors apologize for any inconvenience.

Infectious disease is still a major threat in the world today. Five decades ago, it was considered soon to be eradicated, but the adaptation of pathogens to environmental pressure, such as antimicrobials, encouraged the emergence and reemergence of infectious disease. The fight with infectious disease starts with prevention, diagnosis, and treatment. Diagnosis can be upheld by observing the cause of disease under the microscope or detecting the presence of nucleic acid and proteins of the pathogens. The molecular techniques span from classical polymerase chain reaction (PCR) to sequencing the nucleic acid composition. Here, we are reviewing the works have been undertaken to utilize a portable sequencer, MinION, in various aspects of infectious disease management.

No abstract

Dengue hemorrhagic fever (DHF) is an acute fever disease with high mortality and morbidity in many regions of the world. Leucopenia and thrombocytopenia are two of several laboratory findings that could be found in the course of DHF. This was an analytical retrospective study with a cross sectional design. Samples were patients diagnosed with DHF in Prof. Dr. R.D Kandou General Hospital Manado during the period of 2012. The inclusion criteria were patients <15 years, were diagnosed as DHF according to WHO 1997 criteria, and were examined for platelet and white blood cell count. This study used the medical record data which were analyzed statistically by using the Pearson's correlation test. There were 137 children with DHF during the period of 2012. Samples were 56 children that fulfiled the inclusion criteria. The Pearson correlation test showed a P value 0.801 and correlation coefficient r = -0.034 that indicated that there was a negative correlation which was not significant. Conclusion: There was no significant correlation between the number of thrombocytes and leukocytes in children with dengue hemorrhagic fever.Keywords: dengue hemorrhagic fever, leukocyte, thrombocyteAbstrak: Demam berdarah dengue (DBD) merupakan penyakit demam akut dengan morbiditas dan mortalitas yang tinggi di banyak daerah di dunia. Leukopenia dan trombositopenia merupakan dua temuan laboratorik yang sering ditemukan pada DBD. Penelitian ini bersifat analitik retrospektif dengan pendekatan potong lintang. Sampel penelitian ialah pasien anak yang terdiagnosis DBD di RSUP Prof. Dr. R.D. Kandou Manado selama periode tahun 2012. Pasien yang masuk dalam kriteria inklusi ialah pasien < 15 tahun, telah terdiagnosis menurut kriteria WHO 1997, serta melakukan pemeriksaan laboratorium trombosit dan leukosit. Penelitian ini menggunakan catatan rekam medik, dan untuk analisis statistik digunakan Pearson’s correlation test. Terdapat 137 anak dengan demam berdarah dengue pada periode 2012 dan 56 anak menjadi sampel dalam penelitian ini. Hasil penelitian melalui uji korelasi Pearson mendapatkan nilai P = 0,801 dan koefisien korelasi r = -0,034 yang berarti bahwa korelasi tidak bermakna, dengan kekuatan korelasi lemah dan arah korelasi negatif. Simpulan: Tidak terdapat hubungan yang bermakna antara jumlah trombosit dan leukosit pada pasien anak demam berdarah dengue.Kata kunci: demam berdarah dengue, leukosit, trombosit