Change over Time in the Risk of Death among Japanese COVID-19 Cases Caused by the Omicron Variant Depending on Prevalence of Sublineages.

To assess temporal changes to the risk of death in COVID-19 cases caused by the Omicron variant, we calculated age-standardized case fatality rates (CFR) in patients aged ≥40 years over nine diagnostic periods (3 January to 28 August 2022) in ten Japanese prefectures (14.8 million residents). Among 552,581 study subjects, we found that there were 1836 fatalities during the isolation period (up to 28 days from date of onset). The highest age-standardized CFR (0.85%, 95% confidence interval (CI):0.78-0.92) was observed in cases diagnosed in the second 4-week period (January 31 to February 27), after which it declined significantly up to the 6th 4-week period (0.23%, 95% CI: 0.13-0.33, May 23 to June 19). The CFR then increased again but remained at 0.39% in the eighth period (July 18 to August 28). The CFR in cases with the BA.2 or BA.5 sublineages in the age range 60-80 years was significantly lower than that with BA.1 infections (60 years: 0.19%, 0.02%, 0.053%, respectively; 70 years: 0.91%, 0.33%, 0.39%; ≥80 years: 3.78%, 1.96%, 1.81%, respectively). We conclude that the risk of death in Japanese COVID-19 patients infected with Omicron variants declined through February to mid-June 2022.

Genomic characterization and the prevalence of a novel copiparvovirus in wild sika deer (Cervus nippon) in Japan.

A preliminary metagenomic analysis of the virome of wild sika deer (Cervus nippon) blood in Japan resulted in the identification of a novel parvovirus. The virus was closest, but only 44.7-60.7% identical to 17 reported strains belonging to the genus Copiparvovirus within the subfamily Parvovirinae, over the near-entire genomic sequence. The sika deer copiparvovirus DNA was detected in 15% (31/206) of sika deer captured in 7 prefectures of Japan, and a region-dependent prevalence of 0-66.7% was noted, with a biased distribution in the southern part of Japan. The observed biased distribution of sika deer copiparvovirus may be due to the habitat density of deer and the number of ticks, which might play a role in the transmission of the virus.

Identification of hepatitis E virus in wild sika deer in Japan.

Hepatitis E virus (HEV) is a zoonotic agent mainly transmitted through the consumption of uncooked or undercooked meat products derived from infected animals. In Japan, domestic pigs and wild boars are the major animal reservoirs, and whether or not deer are an HEV reservoir remains controversial. We analyzed 395 serum and 199 liver samples from 405 sika deer (Cervus nippon) caught in the wild between 1997 and 2020 in 11 prefectures of Japan for markers of HEV infection. Overall, 17 deer had anti-HEV IgG (4.3%), while 1 (0.2%) had HEV RNA (genotype 3b), indicating the occurrence of ongoing HEV infection in wild deer in Japan. An analysis of the complete HEV genome (deJOI_14) recovered from a viremic deer in Oita Prefecture revealed only 88.8% identity with the first HEV strain in sika deer (JDEER-Hyo03L) in Japan, being closest (96.3%) to the HEV obtained from a hepatitis patient living in the same prefecture. Of note, the deJOI_14 strain was 8.7-9.0% different from the wild boar HEV strains obtained in the same habitat and the same year, suggesting that difference in infected HEV strains between boar and deer may be explained by the limited possibility of close contact with each other, although boars are a known source of HEV infection. Increased numbers of hepatitis E cases after consumption of raw or undercooked meat products of wild deer have been reported in Japan. These results suggest a low but nonnegligible zoonotic risk of HEV infection in wild deer in this country.

Association of urinary vanin-1 with kidney function decline in hypertensive patients.

Previously, the authors reported the utility of urinary vanin-1 as an early biomarker of kidney injury in spontaneously hypertensive rats and in humans. However, little is known about whether urinary vanin-1 can be used to predict the clinical outcome. This study aimed to evaluate the predictive power of urinary vanin-1 based on kidney function decline in hypertensive patients. The authors measured urinary vanin-1 in 147 patients at the baseline and examined its association with the incidence of ≥20% decline in the estimated glomerular filtration rate (eGFR) using the Cox regression analysis. The mean age of the patients averaged 72.9 ± 8.2 years, and 39% were women. Median (interquartile range) urinary vanin-1 was 0.33 (0-2.6) ng/mg Cr During a median follow-up of 12 months, 14 patients showed kidney function decline. A higher urinary vanin-1 level was associated with an increased risk of kidney function decline (hazard ratio, 9.87; 95% CI, 1.11-87.5) (p = .04) in the fully adjusted model. In conclusion, urinary vanin-1 is an independent risk factor for kidney function decline in hypertensive patients and it could be useful in clinical settings. The underlying pathophysiologic mechanisms warrant additional investigation.

Association between a low dose of proton pump inhibitors and kidney function decline in elderly hypertensive patients: a retrospective observational study.

OBJECTIVES: Proton pump inhibitors (PPIs) are widely used for acid suppression therapy. Recently, PPI use was reported to be associated with chronic kidney disease (CKD); however, whether a low dose of PPIs is associated with CKD remains unknown. METHODS: This retrospective observational study included hypertensive patients who visited Kenwakai Hospital between 2017 and 2019. Renal parameters, such as the estimated glomerular filtration rate (eGFR) and serum creatinine (Scr), were extracted from medical records and compared between three years before treatment and the baseline. PPI use was assessed as cumulative exposure for three years. RESULTS: The study population included 152 patients (57.9% men; mean age, 74.5 years). Of those, 35.5% were PPI users (low dose, 17.1%; high dose, 18.4%). A significant decrease in eGFR and an increase in Scr were observed between three years before treatment and the baseline in the high-dose PPI group but not the non-use or low-dose PPI groups. CONCLUSIONS: Our results suggest that a low dose of PPIs may be safe in clinical settings, but further prospective studies are needed to clarify our findings.

Urinary vanin-1 associated with chronic kidney disease in hypertensive patients: A pilot study.

Hypertension and chronic kidney disease (CKD) are serious interrelated public health problems. Despite the monitoring and control of high blood pressure, symptoms of CKD are not usually apparent in its early stages. Previously, we reported the utility of urinary vanin-1 as an early biomarker of kidney injury in spontaneously hypertensive rats, but it remains unknown whether urinary vanin-1 is associated with CKD in humans. In this study, we estimated associations between urinary vanin-1 and parameters of kidney function in a cross-sectional study of hypertensive patients. We measured concentrations of vanin-1 using spot urine from 147 adult hypertensive patients (mean age, 72.8 years; 39.5% women). Patients were divided into 2 groups based on the median of the estimated glomerular filtration rate (eGFR). The group with eGFR < 60 mL/min per 1.73 m2 showed significantly higher levels of urinary vanin-1 than those with eGFR ≥ 60 mL/min per 1.73 m2. On univariate analysis, urinary vanin-1 as well as neutrophil gelatinase-associated lipocalin (NGAL) showed significant negative correlations with eGFR; however, multivariate analysis revealed that urinary vanin-1, but not NGAL, significantly correlated with eGFR. In addition, urinary vanin-1 had a significant positive correlation with the urinary protein-to-creatinine ratio (UPCR) (r = 0.21; P = .021) and albumin-to-creatinine ratio (UACR) (r = 0.61; P < .01). In conclusion, urinary vanin-1 is associated with lower eGFR and higher UPCR and UACR, and might be a potential marker of decreased kidney function in hypertensive patients. Further studies are needed to confirm these findings.

Prevalence and genotype/subtype distribution of hepatitis E virus (HEV) among wild boars in Japan: Identification of a genotype 5 HEV strain.

To further investigate the prevalence of hepatitis E virus (HEV) infection and characterize HEV genomes among Japanese wild boars (Sus scrofa leucomystax), 1880 boars captured in 17 prefectures in Japan from 2013 to 2019 were studied. Overall, anti-HEV IgG was detected in 8.9 % and HEV RNA was detected in 3.9 % of boars, which was comparable with our previous studies during 2003-2013 (10.3 % and 3.5 %, respectively). Among 74 boar HEV strains obtained from infected boars in the present study, 50 (68 %) were classified into genotype 3 (3a and 3b), 23 (31 %) were classified into genotype 4 (4i), and the remaining strain (wbJGF_19-1) was classified into genotype 5. The wbGF_19-1 strain shared 92.7 % identity over the entire genome with the prototype genotype 5 strain (JBOAR135-Shiz09). The identification of the second genotype 5 HEV strain in a place that is located only 100 km from the site at which JBOAR135-Shiz09 was identified, suggests that genotype 5 HEV circulates within a relatively close range in Japan. Genetically similar HEV strains forming a clade were identified from wild boars living in each area during the observation period of 11-13 years, although the nucleotide sequence changed gradually, accounting for up to 3.4-3.6 % within the 412-nucleotide ORF2 sequence. Eight groups of boars with a cluster of HEV infections were observed, consisting of two, three or four infected offspring, presumably born to the same mother or offspring with their mother. These results suggest that wild boars continue to be important reservoirs for HEV infection in humans in Japan.

Improvement of Blood Pressure Control by Adherence Check in Patients With Apparent Treatment-Resistant Hypertension: A Case Series.

Adherence to medications is an important challenge while treating chronic disease such as resistant hypertension, which is defined as uncontrolled blood pressure (BP) despite treatment with more than 3 antihypertensive drugs to achieve targets. It is possible that poor adherence is the most significant contributor to rates of pseudo-resistance among treated hypertensive patients. In this report, we describe 4 patients with apparent treatment-resistant hypertension, who received intervention to promote adherence by pharmacists who set the prescribed medicines in a weekly medication calendar and conducted a weekly pill count. The results showed that the intervention of pharmacists to medication adherence improved systolic BP in patients with apparent treatment-resistant hypertension; however, further controlled trials are required to strengthen supporting evidence.

A synthetic male-specific sterilization system using the mammalian pro-apoptotic factor in a malaria vector mosquito.

Conditional cell death systems are useful for various aspects of basic science with a wide range of applications, including genetic pest control. We recently demonstrated that expression of the mammalian pro-apoptotic factor, B-cell leukaemia/lymphoma 2-associated X protein (Bax), can induce apoptosis in specific tissues by using tissue specific promoters in silkworm and mosquito. Here, we newly identified a functional promoter in the Asian malaria vector, Anopheles stephensi, which enables gene expression specifically in the testis. We produced a transgenic mosquito line that expresses mouse Bax under the control of this testis-specific promoter. Transgenic mosquito males exhibited aberrant testes without functional sperm and complete sterility, whereas transgenic females maintained normal fecundity. Despite their abnormal testes, the transgenic males maintained normal function of male accessory glands and typical mating behaviour. As a result of mating with these males, females showed refractoriness to further mating. These results suggest that transgenic males induce female sterility via mating. The mosquito is one of the most important disease vectors, and the control of their population benefits global public health. Thus, this Bax-mediated synthetic male-specific sterilization system could be applied to population control of mosquitoes.

Upper gastrointestinal pathophysiology due to mouse malaria Plasmodium berghei ANKA infection.

BACKGROUND: Epigastric pain, vomiting, and other gastrointestinal problems are among the most important symptoms of malaria infection as they suggest the possibility that the condition is serious. Pathophysiologies such as gastric mucosal changes and delayed gastric emptying have been reported in serious cases of malaria infection. However, it is unclear whether or not pathophysiological involvement of the upper gastrointestinal tract occurs in Plasmodium berghei ANKA (PbA)-infected mice. METHODS: PbA-infective Anopheles mosquitoes were used to infect mice via the natural route of infection. Fifteen PbA-C57BL/6 mice were used as a cerebral malaria model and the same numbers of PbA-BALB/c mice were used as a cerebral malaria-resistant model, and then we investigated the pathophysiological involvement of the stomach and small intestine. RESULTS: On day 8 post infection, six PbA-C57BL/6 mice showed cerebral malaria and nine others had uncomplicated infection. All the PbA-C57BL/6 mice on that same day showed severe weight loss with multiple, red gastric patches and changes to the course of the small intestine with villus goblet cell enlargement. In addition, cerebral malaria cases showed gastric gas retention with submucosal edema and small intestinal shortening. In PbA-BALB/c mice, overextension of the stomach and gas retention were evident from week 2 after PbA infection, as well as changes to the course of the small intestine and mesenteric thinning with fragility. CONCLUSIONS: We described the upper gastrointestinal pathophysiology representing new findings directly linked to malarial severity and subsequent death in PbA-infected mice as a mouse model of malaria infection.

Baculovirus-Induced Fast-Acting Innate Immunity Kills Liver-Stage Plasmodium.

Baculovirus (BV), an enveloped insect virus with a circular dsDNA genome, possesses unique characteristics that induce strong innate immune responses in mammalian cells. In this study, we show that BV administration in BALB/c mice not only provides complete protection against a subsequent Plasmodium berghei sporozoite infection for up to 7 d after the injection but also eliminates existing liver-stage parasites completely. The elimination of sporozoites by BV was superior to that by primaquine, and this effect occurred in a TLR9-independent manner. At 6 h after BV administration, IFN-α and IFN-γ were robustly produced in the serum, and RNA transcripts of IFN-stimulated genes were markedly upregulated in the liver compared with control mice. The in vivo passive transfer of serum after BV administration effectively eliminated liver-stage parasites, and IFN-α neutralization abolished this effect, indicating that the BV liver-stage parasite-killing mechanism is downstream of the type I IFN signaling pathway. These findings provide evidence that BV-induced, fast-acting innate immunity completely kills liver-stage parasites and, thus, may lead to new malaria drug and vaccine strategies.

Malaria infectivity of xanthurenic acid-deficient anopheline mosquitoes produced by TALEN-mediated targeted mutagenesis.

Anopheline mosquitoes are major vectors of malaria parasites. When the gametocytes of the malaria parasite are transferred from a vertebrate to mosquitoes, they differentiate into gametes, and are fertilized in the midguts of mosquitoes. Xanthurenic acid (XA), a waste product of the ommochrome synthesis pathway, has been shown to induce exflagellation during microgametogenesis in vitro; however, it currently remains unclear whether endogenous XA affects the infectivity of anopheline mosquitoes to malaria parasites in vivo due to the lack of appropriate experimental systems such as a XA-deficient line. In the present study, we produced a XA-deficient line in Anopheles stephensi using transcription activator-like effector nuclease (TALEN)-mediated gene targeting (knockout) of the kynurenine 3-monooxygenase (kmo) gene, which encodes an enzyme that participates in the ommochrome synthesis pathway. The knockout of kmo resulted in the absence of XA, and oocyst formation was inhibited in the midguts of these XA-deficient mosquitoes, which, in turn, reduced sporozoite numbers in their salivary glands. These results suggest that endogenous XA stimulates exflagellation, and enhances the infectivity of anopheline mosquitoes to malaria parasites in vivo. The XA-deficient line of the anopheline mosquito provides a useful system for analyzing and understanding the associated factors of malaria gametogenesis in the mosquito midgut.

DAF-shielded baculovirus-vectored vaccine enhances protection against malaria sporozoite challenge in mice.

BACKGROUND: Previous studies have shown that the baculovirus-vectored vaccine based on the "baculovirus dual expression system (BDES)" is an effective vaccine delivery platform for malaria. However, a point of weakness remaining for use of this vaccine platform in vivo concerns viral inactivation by serum complement. In an effort to achieve complement resistance, the gene encoding the human decay-accelerating factor (hDAF) was incorporated into the BDES malaria vaccine expressing the Plasmodium falciparum circumsporozoite protein (PfCSP). RESULTS: The newly-developed BDES vaccine, designated BDES-sPfCSP2-Spider, effectively displayed hDAF and PfCSP on the surface of the viral envelope, resulting in complement resistance both in vitro and in vivo. Importantly, upon intramuscular inoculation into mice, the BDES-sPfCSP2-Spider vaccine had a higher protective efficacy (60%) than that of the control vaccine BDES-sPfCSP2-Spier (30%) against challenge with transgenic Plasmodium berghei sporozoites expressing PfCSP. CONCLUSION: DAF-shielded BDES-vaccines offer great potential for development as a new malaria vaccine platform against the sporozoite challenge.

Further Molecular Analysis of G6PD Deficiency Variants in Southern Vietnam and a Novel Variant Designated as G6PD Ho Chi Minh (173 A>G; 58 Asp>Gly): Frequency Distributions of Variants Compared with Those in Other Southeast Asian Countries.

We conducted a survey of glucose-6-phosphate dehydrogenase (G6PD) deficiency among newborn babies at Tu Du Hospital, Ho Chi Minh, southern Vietnam. A total of 90 deficient babies were detected, including 85 in the Kinh ethnic group, 4 Chinese, and 1 in the K'Ho minority group. In the Kinh ethnic group, G6PD variants such as G6PD Viangchan (n=32), Kaiping (n=11), Canton (n=8), Chinese-5 (n=7), Union (n=5) and Quing Yuan (n=4) were detected. A variant with silent mutations at 1311 C>T and IVS11 nt 93 T>C was also detected in 17 cases. A novel mutation (173 A>G) in exon 4 with a predicted amino acid change of 58 Asp>Gly was also found in a Kinh newborn girl and her father, and it was designated as G6PD Ho Chi Minh. These findings demonstrated that the Kinh ethnic group in southern Vietnam has 8 different G6PD variants, indicating that the members of this group have many ancestors in terms of G6PD variants from Southeast Asia, China, and Oceania. We compared the frequency distribution of G6PD variants in the Kinh population with those of other Southeast Asian populations, and the Kinh population's distribution was quite similar to that in the Thai population, but differed from it by the absence of G6PD Mahidol.

Petit-High Pressure Carbon Dioxide stress increases synthesis of S-Adenosylmethionine and phosphatidylcholine in yeast Saccharomyces cerevisiae.

Petit-High Pressure Carbon Dioxide (p-HPCD) is a promising nonthermal technology for foods pasteurization. Cluster analysis of gene expression profiles of Saccharomyces cerevisiae exposed to various stresses exhibited that gene expression profile for p-HPCD stress (0.5MPa, 25°C) was grouped into a cluster including profiles for Sodium Dodecyl Sulfate and Roundup herbicide. Both are detergents that can disorder membrane structurally and functionally, which suggests that cell membrane may be a target of p-HPCD stress to cause cell growth inhibition. Through metabolomic analysis, amount of S-Adenosylmethionine (AdoMet) that is used as methyl donor to participate in phosphatidylcholine synthesis via phosphatidylethanolamine (PE) methylation pathway, was increased after p-HPCD treatment for 2h. The key gene OPI3 encoding phospholipid methyltransferase that catalyzes the last two steps in PE methylation pathway was confirmed significantly induced by RT-PCR. Transcriptional expression of genes (MET13, MET16, MET10, MET17, MET6 and SAM2) related to AdoMet biosynthesis was also significantly induced. Choline as the PC precursor and ethanolamine as PE precursor in Kennedy pathway were also found increased under p-HPCD condition. We also found that amounts of most of amino acids involving protein synthesis were found decreased after p-HPCD treatment for 2h. Moreover, morphological changes on cell surface were observed by scanning electron microscope. In conclusion, the effects of p-HPCD stress on cell membrane appear to be a very likely cause of yeast growth inhibition and the enhancement of PC synthesis could contribute to maintain optimum structure and functions of cell membrane and improve cell resistance to inactivation.

Inhibition of Malaria Infection in Transgenic Anopheline Mosquitoes Lacking Salivary Gland Cells.

Malaria is an important global public health challenge, and is transmitted by anopheline mosquitoes during blood feeding. Mosquito vector control is one of the most effective methods to control malaria, and population replacement with genetically engineered mosquitoes to block its transmission is expected to become a new vector control strategy. The salivary glands are an effective target tissue for the expression of molecules that kill or inactivate malaria parasites. Moreover, salivary gland cells express a large number of molecules that facilitate blood feeding and parasite transmission to hosts. In the present study, we adapted a functional deficiency system in specific tissues by inducing cell death using the mouse Bcl-2-associated X protein (Bax) to the Asian malaria vector mosquito, Anopheles stephensi. We applied this technique to salivary gland cells, and produced a transgenic strain containing extremely low amounts of saliva. Although probing times for feeding on mice were longer in transgenic mosquitoes than in wild-type mosquitoes, transgenic mosquitoes still successfully ingested blood. Transgenic mosquitoes also exhibited a significant reduction in oocyst formation in the midgut in a rodent malaria model. These results indicate that mosquito saliva plays an important role in malaria infection in the midgut of anopheline mosquitoes. The dysfunction in the salivary glands enabled the inhibition of malaria transmission from hosts to mosquito midguts. Therefore, salivary components have potential in the development of new drugs or genetically engineered mosquitoes for malaria control.

Application of copper-based ovitraps in local houses in West Sumatra, Indonesia: a field test of a simple and affordable larvicide for mosquito control.

BACKGROUND: The application of oviposition traps (ovitraps) is one of the currently available rational methods used in mosquito control campaigns because it eliminates the larval stage. However, the use of current larvicides is hampered by their cost and applicability. Therefore, a more economical and practical alternative is urgently needed. We previously reported that copper in liquid form is a promising candidate due to its potent larvicide properties in a laboratory setting, affordability, and availability. METHODS: In the present study, a field test was performed by randomly placing copper-filled plastic pots with a concentration of 10 ppm in 21 local houses in Painan City, West Sumatra, Indonesia. Three of these pots including a control were placed inside, while another two were placed outside each of the houses. RESULTS: After 14 days, a large number of dead first and second instar larvae of Aedes spp. were observed in the copper-filled pots. Larvae in the control pots were all viable and thriving. Unhatched eggs and pupae were detected in several pots in the copper-treated group but were excluded from the analysis. CONCLUSIONS: Our field data confirmed that copper is a potential larvicide for ovitraps, particularly in under-resourced areas.

Clinical categories of exaggerated skin reactions to mosquito bites and their pathophysiology.

Mosquito bites are skin irritating reactions, which usually resolve spontaneously without intensive medical care. However, in certain situations, mosquito bites may form a more vicious reaction, sometimes accompanying fever and systemic symptoms. In such cases, the presence of rare hematological disorders, abnormalities in eosinophils and/or association with Epstein-Barr virus (EBV) may underlie. Importantly, hypersensitivity to mosquito bites (HMB), which is characterized by necrotic skin reactions to mosquito bites with various systemic symptoms, is often observed in association with EBV infection and natural killer (NK) cell lymphoproliferative disorder. Exaggerated skin reaction to mosquito bites is also seen in Wells' syndrome. While strong Th2-skewing immune dysregulation is apparent in the patients, they also show robust CD4(+) T cell proliferation in response to mosquito salivary gland extracts, indicating close association between Wells' syndrome and mosquito bites. Similar skin reaction to mosquito bites is also noticed in certain types of B cell neoplasm, although the role of B cells in this peculiar reaction to mosquito bites is yet to be elucidated. In this review, we will discuss the current knowledge of exaggerated reaction toward mosquito bites seen in conjunction with these unique hematological disorders, and examine the scientific studies and observations reported in previous literatures to organize our current understanding of the pathogenesis of this distinct disorder.

Parasites resistant to the antimalarial atovaquone fail to transmit by mosquitoes.

Drug resistance compromises control of malaria. Here, we show that resistance to a commonly used antimalarial medication, atovaquone, is apparently unable to spread. Atovaquone pressure selects parasites with mutations in cytochrome b, a respiratory protein with low but essential activity in the mammalian blood phase of the parasite life cycle. Resistance mutations rescue parasites from the drug but later prove lethal in the mosquito phase, where parasites require full respiration. Unable to respire efficiently, resistant parasites fail to complete mosquito development, arresting their life cycle. Because cytochrome b is encoded by the maternally inherited parasite mitochondrion, even outcrossing with wild-type strains cannot facilitate spread of resistance. Lack of transmission suggests that resistance will be unable to spread in the field, greatly enhancing the utility of atovaquone in malaria control.

Detailed methodology for high resolution scanning electron microscopy (SEM) of murine malaria parasitized-erythrocytes.

Scanning electron microscopy (SEM) is a powerful tool used to investigate object surfaces and has been widely applied in both material science and biology. With respect to the study of malaria, SEM revealed that erythrocytes infected with Plasmodium falciparum, a human parasite, display 'knob-like' structures on their surface comprising parasitized proteins. However, detailed methodology for SEM studies of malaria parasites is lacking in the literature making such studies challenging. Here, we provide a step-by-step guide to preparing Plasmodium-infected erythrocytes from two mouse strains for SEM analysis with minimal structural deterioration. We tested three species of murine malaria parasites, P. berghei, P. yoelii, and P. chabaudi, as well as non-parasitized human erythrocytes and P. falciparum-infected erythrocytes for comparisons. Our data demonstrated that the surface structures of parasitized erythrocytes between the three species of murine parasites in the two different strains of mice were indistinguishable and no surface alterations were observed in P. falciparum-erythrocytes. Our SEM observations contribute towards an understanding of the molecular mechanisms of parasite maturation in the erythrocyte cytoplasm and, along with future studies using our detailed methodology, may help to gain insight into the clinical phenomena of human malaria.