Microsporidial keratoconjunctivitis - first outbreak in Japan.

BACKGROUND: Most cases of microsporidial keratoconjunctivitis are found in the Southern hemisphere. Our purpose was to investigate the first outbreak of microsporidial keratoconjunctivitis in Japan among healthy, immunocompetent soccer players from the same team during a 1-month period. CASE PRESENTATION: This study is an observational case series. The medical records were analyzed for five cases with microsporidial keratoconjunctivitis who presented within September 2022. All five cases were males between 28 and 36 years old. These previously healthy individuals belonged to the same football team. Their eyes were considered susceptible to contaminated water or dirt from the turf at game and practice sites. All cases involved unilateral conjunctivitis, with scattered round white lesions that showed positive fluorescein staining in the corneal epithelium. All cases experienced diminution of vision in the affected eye. In three cases, direct smears showed spores of approximately 2-3 µm in diameter. Polymerase chain reaction (PCR) analysis of corneal scrapes revealed partial amplification of microsporidial 18 S ribosomal RNA gene in four cases. Sequences of PCR products from all four cases showed 100% identity with strains of Vittaforma corneae previously reported from an outbreak in Singapore. All cases were treated with topical therapy, including voriconazole, fluorometholone, and levofloxacin. Four eyes underwent corneal scraping. After treatment, all eyes healed without residual opacities. CONCLUSIONS: Only a few sporadic case reports of this disease have previously been reported in Japan. We detected V. corneae in our case series, representing what appears to be the first outbreak of microsporidial keratoconjunctivitis in Japan. Exposure to contaminated water or soil, in addition to inadequate sanitary facilities, represents a potential source of infection. Further investigations to clarify the characteristics of microsporidia seem warranted.

Amebiasis as a sexually transmitted infection: A re-emerging health problem in developed countries.

Amebiasis, which is caused by Entamoeba histolytica (E. histolytica), is the second leading cause of parasite-related death worldwide. It manifests from asymptomatic carriers to severe clinical conditions, like colitis and liver abscesses. Amebiasis is commonly seen in developing countries, where water and food are easily contaminated by feces because of the poor sanitation. However, a recently challenge in many developed countries is the increase in domestic cases of invasive amebiasis as a sexually transmitted infection (STI amebiasis). In contrast to food-/ waterborne transmission of E. histolytica in developing countries, transmission of STI amebiasis occurs directly through human-to-human sexual contact (e.g., men who have sex with men and people who engage in oral-anal sex); in this setting, asymptomatic infected individuals are the main reservoir of E. histolytica. The Development of screening methods for the early diagnosis of asymptomatic E. histolytica infection is the key to epidemiologic control. Moreover, delay in diagnosis of severe cases (e.g., fulminant amebiasis) leads to death even in developed countries. It is also important to increase clinical awareness of domestically transmitted STI amebiasis in the clinical settings. This review considers the changing epidemiology and clinical manifestations of STI amebiasis, and finally discusses the future strategies for the better practice.

Identification of asymptomatic Entamoeba histolytica infection by a serological screening test: A cross-sectional study of an HIV-negative men who have sex with men cohort in Japan.

BACKGROUND: Amebiasis, caused by Entamoeba histolytica, is spreading in developing countries and in many developed countries as a sexually transmitted infection. Here, we evaluated the efficacy of serological screening to identify asymptomatic E. histolytica infection as a potential epidemiological control measure to limit its spread. METHODOLOGY/PRINCIPAL FINDINGS: This cross-sectional study was carried out between January and March 2021 in an HIV-negative men who have sex with men (MSM) cohort at the National Center for Global Health and Medicine. Serological screening was performed using a commercially available ELISA kit. For seropositive individuals, we performed stool polymerase chain reaction (PCR) to determine current E. histolytica infection. We performed E. histolytica serological screening of 312 participants. None had a history of E. histolytica infection prior to the study. The overall E. histolytica seropositivity was 6.7% (21/312), which was similar to that found by the rapid plasma reagin test (17/312). We identified current infection in 8 of 20 seropositive participants (40.0%) by stool PCR. CONCLUSIONS/SIGNIFICANCE: Our serological screening approach constitutes a potentially practical epidemiological strategy. Active epidemiological surveys, in combination with an effective screening strategy for asymptomatically infected individuals, should be applied to help reduce sexually transmitted E. histolytica infections.

The application of shotgun metagenomics to the diagnosis of granulomatous amoebic encephalitis due to Balamuthia mandrillaris: a case report.

BACKGROUND: Granulomatous amoebic encephalitis (GAE) is an infrequent and fatal infectious disease worldwide. Antemortem diagnosis in this condition is very difficult because clinical manifestations and neuroimaging are nonspecific. CASE PRESENTATION: A 60-year-old Japanese woman was admitted with a chief complaint of left homonymous hemianopsia. Brain-MRI showed extensive necrotizing lesions enhanced by gadolinium, in the right frontal lobe, right occipital lobe, and left parietal lobe. Epithelioid granulomas of unknown etiology were found in the biopsied brain specimens. Shotgun metagenomic sequencing using a next-generation sequencer detected DNA fragments of Balamuthia mandrillaris in the tissue specimens. The diagnosis of granulomatous amoebic encephalitis was confirmed using an amoeba-specific polymerase chain reaction and immunostaining on the biopsied tissues. CONCLUSIONS: Shotgun metagenomics is useful for the diagnosis of central nervous system infections such as GAE wherein the pathogens are difficult to identify.

Use of PCR in the diagnosis of pericardial amebiasis: a case report and systematic review of the literature.

BACKGROUND: Entamoeba histolytica (E. histolytica) is rarely identified as a cause of amebic pericarditis. We report a case of amebic pericarditis complicated by cardiac tamponade, in which the diagnosis was missed initially and was made retrospectively by polymerase chain reaction (PCR) testing of a stored sample of pericardial fluid. Furthermore, we performed a systematic review of the literature on amebic pericarditis. CASE PRESENTATION: A 71-year-old Japanese man who had a history of sexual intercourse with several commercial sex workers 4 months previously, presented to our hospital with left chest pain and cough. He was admitted on suspicion of pericarditis. On hospital day 7, he developed cardiac tamponade requiring urgent pericardiocentesis. The patient's symptoms temporarily improved, but 1 month later, he returned with fever and abdominal pain, and multiple liver lesions were found in the right lobe. Polymerase chain reaction of the aspiration fluid of the liver lesion and pericardial and pleural fluid stored from the previous hospitalization were all positive for E. histolytica. Together with the positive serum antibody for E. histolytica, a diagnosis of amebic pericarditis was made. Notably, the diagnosis was missed initially and was made retrospectively by performing PCR testing. The patient improved with metronidazole 750 mg thrice daily for 14 days, followed by paromomycin 500 mg thrice daily for 10 days. CONCLUSIONS: This case suggests that, although only 122 cases of amebic pericarditis have been reported, clinicians should be aware of E. histolytica as a potential causative pathogen. The polymerase chain reaction method was used to detect E. histolytica in the pericardial effusion and was found to be useful for the diagnosis of amebic pericarditis in addition to the positive results for the serum antibody testing for E. histolytica. Because of the high mortality associated with delayed treatment, prompt diagnosis should be made.

Clinical Features and Gut Microbiome of Asymptomatic Entamoeba histolytica Infection.

BACKGROUND: Entamoeba histolytica infection is a sexually transmitted disease in some developed countries. Asymptomatic infection often occurs and can be a source of transmission; however, limited data are available regarding the pathogenesis of E. histolytica. METHODS: This was a single-center, cross-sectional study. Specimens were prospectively collected from patients with clinically suspected cases. Entamoeba histolytica infection was defined as a case in which the identification of E. histolytica was confirmed by polymerase chain reaction (PCR) of a clinical specimen. Data from asymptomatic cases were compared with those from symptomatic invasive cases. RESULTS: Sixty-four E. histolytica-infected cases, including 13 asymptomatic cases, were identified during the study period. Microbiological diagnosis was made by endoscopic sampling in 26.6% of these cases (17/64). Endoscopy identified macroscopically visible lesions in all cases; however, the sensitivity of histopathology on biopsy samples was low (45.5%) compared with PCR (94.7%). In asymptomatic cases, infection sites were limited around the proximal colon; moreover, trophozoites were frequently identified at infection sites whereas cystic forms were commonly detected in stools. Gut microbiome analyses showed more uniform composition in asymptomatic cases than in symptomatic invasive cases, which were represented by a relatively high abundance of Ruminococcaceae, Coriobacteriaceae, and Clostridiaceae, and a low abundance of Streptococcaceae. CONCLUSIONS: These results indicate that the encystation and attenuation of E. histolytica are highly affected by the intestinal contents, including the gut microbiome.

A Japanese case of amoebic meningoencephalitis initially diagnosed by cerebrospinal fluid cytology.

Microscopy can detect the presence of amoebic trophozoites in cerebrospinal fluid and tissue. The infection was confirmed in the present case by polymerase chain reaction and immunohistochemistry, but we were unable to achieve a cure. Our case rapidly progressed without any skin lesions.

Utility of the Rapid Antigen Detection Test E. histolytica Quik Chek for the Diagnosis of Entamoeba histolytica Infection in Nonendemic Situations.

Entamoeba histolytica infection is an increasingly common sexually transmitted infection in Japan. Currently, stool ova and parasite examination (O&P) is the only approved diagnostic method. Here, we assessed the utility of the commercially available rapid antigen detection test (Quik Chek) for E. histolytica A multicenter cross-sectional study was conducted. Stool samples that had been submitted for O&P were included. The samples were subjected to both Quik Chek and PCR, and the Quik Chek results were assessed in comparison with PCR as the reference standard. E. histolytica infection was confirmed in 5.8% (38/657) of the samples and comprised 20 diarrheal and 18 nondiarrheal cases. The overall sensitivity and specificity of Quik Chek were 44.7% (95% confidence interval, 30.1 to 60.3) and 99.8% (99.1 to 100), respectively. The sensitivity of Quik Chek was higher for diarrheal cases (60.0%) than for nondiarrheal cases (27.8%). Furthermore, the combined use of Quik Chek with O&P increased the sensitivity (78.9%), especially for diarrheal cases (up to 90%). The E. histolytica burden assessed by quantitative PCR was similar between Quik Chek-positive and -negative samples. The Quik Chek assay sensitivity was lower for cyst-containing stools than for trophozoite-containing stools, although it was shown that cultured E. histolytica clinical strains from Quik Chek-negative cyst-containing stools exhibited antigenicity in vitro The present study confirmed the high specificity of Quik Chek for E. histolytica infection. Combined use with O&P increased the sensitivity of detection, facilitating the use of Quik Chek in point-of-care settings in nonendemic situations.

Microsporidial keratitis retrospectively diagnosed by ultrastructural study of formalin-fixed paraffin-embedded corneal tissue: a case report.

BACKGROUND: The utility of formalin-fixed paraffin-embedded (FFPE) corneal tissue specimens for retrospective diagnosis of microsporidial keratitis was evaluated by transmission electron microscopy (TEM) analysis and the possible second case of microsporidial keratitis after Descemet stripping automated endothelial keratoplasty (DSAEK) was described. CASE PRESENTATION: A 68-year-old man presented with multiple crystalline opacities in the corneal stroma that progressed extremely slowly after DSAEK. Fungiflora Y staining of corneal scrapings from the affected regions revealed an oval microorganism. Topical voriconazole administration was ineffective and penetrating keratoplasty was performed. Histological and molecular analyses were carried out on the excised cornea. Ziehl-Neelsen staining revealed an acid-fast, oval organism that was visible by ultraviolet illumination after Fungiflora Y and Uvitex 2B staining, whereas periodic acid-Schiff and Grocott's staining did not yield any significant findings. Microsporidium was detected by TEM of FFPE tissue. Nosema or Vittaforma sp. was suspected as the causative microorganism by PCR of FFPE tissue and by the fact that those species are known to cause eye infection. The corneal graft has maintained transparency at 1 year and half postoperatively. CONCLUSIONS: This is the first known case of microsporidial keratitis diagnosed retrospectively by molecular and ultrastructural study of FFPE tissue, and the possible second case of microsporidial keratitis after DSAEK. Microsporidial keratitis should be considered when corneal opacity refractory to conventionally known therapy would occur after DSAEK. Our findings suggest that more microsporidial keratitis cases than have been reported to date can be identified by TEM or PCR examination of FFPE corneal specimens.

Pathogenic free-living amoebic encephalitis in Japan.

Over 600 cases of amoebic encephalitis caused by pathogenic free-living amoebas (Balamuthia mandrillaris, Acanthamoeba spp., and Naegleria fowleri) have been reported worldwide, and in Japan, 24 cases have been reported from the first case in 1976 up to 2018. Among these cases, 18 were caused by B. mandrillaris, four by Acanthamoeba spp., one by N. fowleri, and one was of unknown etiology. Additionally, eight cases were diagnosed with encephalitis due to pathogenic free-living amoebas before death, but only three cases were successfully treated. Unfortunately, all other cases were diagnosed by autopsy. These facts indicate that an adequate diagnosis is difficult, because encephalitis due to pathogenic free-living amoebas does not show typical symptoms or laboratory findings. Moreover, because the number of cases is limited, other cases might have been missed outside of those diagnosed by autopsy. Cases of encephalitis caused by B. mandrillaris have been reported from all over Japan, with B. mandrillaris recently isolated from soil in Aomori prefecture. Therefore, encephalitis caused by pathogenic free-living amoebas should be added to the differential diagnosis of encephalitis patients.

A case report of granulomatous amoebic encephalitis by Group 1 Acanthamoeba genotype T18 diagnosed by the combination of morphological examination and genetic analysis.

BACKGROUND: The diagnosis of granulomatous amoebic encephalitis is challenging for clinicians because it is a rare and lethal disease. Previous reports have indicated that Acanthamoeba with some specific genotypes tend to cause the majority of human infections. We report a case of granulomatous amoebic encephalitis caused by Acanthamoeba spp. with genotype T18 in an immunodeficient patient in Japan after allogenic bone marrow transplantation, along with the morphological characteristics and genetic analysis. CASE PRESENTATION: A 52-year old man, who had undergone allogenic bone marrow transplantation, suffered from rapid-growing brain masses in addition to pneumonia and died within 1 month from the onset of the symptoms including fever, headache and disorientation. Infection with Acanthamoeba in the brain and lung was confirmed by histological evaluation; immunohistochemical staining and polymerase chain reaction analysis using autopsy samples also indicated the growth of Acanthamoeba in the brain. Gene sequence analysis indicated that this is the second documented case of infection with Acanthamoeba spp. with genotype T18 in a human host. Postmortem retrospective evaluation of cerebrospinal fluid sample in our case, as well as literature review, indicated that some cases of granulomatous amoebic encephalitis caused by Acanthamoeba may be diagnosable by cerebrospinal fluid examination. CONCLUSION: This case indicates that Acanthamoeba spp. with genotype T18 can also be an important opportunistic pathogen. For pathologists as well as physicians, increased awareness of granulomatous amoebic encephalitis is important for improving the poor prognosis along with the attempt to early diagnosis with cerebrospinal fluid.

An Acute Case of Granulomatous Amoebic Encephalitis-Balamuthia mandrillaris Infection.

A 74-year-old woman who exhibited drowsiness was referred to our hospital. Enhanced head magnetic resonance imaging (MRI) revealed multiple ring-enhancing lesions and lesions showing partial mild hemorrhaging. The patient gradually progressed to a comatose condition with notable brain deterioration of unknown cause on follow-up MRI. On day nine, the patient inexplicably died, although brain herniation was suspected. Autopsy and histopathology revealed numerous amoebic trophozoites in the perivascular spaces and within the necrotic tissue. Brain immunostaining tested positive for Balamuthia mandrillaris. Infection due to free-living amoeba is rare in Japan; however, it may increase in the near future due to unknown reasons.

The Molecular Detection of Cryptosporidium and Giardia in Sika Deer (Cervus Nippon Centralis) in Japan.

Fecal specimens (271 samples) from wild deer, Cervus nippon centralis, were collected from nine different areas in Japan; these samples were subjected to a real-time reverse transcription PCR for Cryptosporidium-and Giardia-specific 18S ribosomal RNA to investigate the prevalence of Cryptosporidium and Giardia infection. The incidence of Cryptosporidium and Giardia in the nine areas ranged from 0% to 20.0% and 0% to 3.4%, respectively. The prevalence of Cryptosporidium among male and female deer was 8.1% and 3.9%, respectively, while that of Giardia was 0.7% and 0.8%. Sequence analysis identified the Cryptosporidium deer genotype, Cryptosporidium bovis, Cryptosporidium ryanae and Cryptosporidium meleagridis from the sequence of Cryptosporidium-specific partial 18S ribosomal RNA and Giardia intestinalis assemblage A from the partial sequence of Giardia-specific 18S rRNA. The variation in regional prevalence indicates that Cryptosporidium infection depends on environmental factors, and that bovine Cryptosporidium was detected more frequently than cervine Cryptosporidium. These data suggest that wild deer might be a healthy carrier of bovine Cryptosporidium.

Underestimated Amoebic Appendicitis among HIV-1-Infected Individuals in Japan.

Entamoeba histolytica is not a common causative agent of acute appendicitis. However, amoebic appendicitis can sometimes be severe and life threatening, mainly due to a lack of awareness. Also, its frequency, clinical features, and pathogenesis remain unclear. The study subjects were HIV-1-infected individuals who presented with acute appendicitis and later underwent appendectomy at our hospital between 1996 and 2014. Formalin-fixed paraffin-embedded preserved appendix specimens were reexamined by periodic acid-Schiff (PAS) staining and PCR to identify undiagnosed amoebic appendicitis. Appendectomies were performed in 57 patients with acute appendicitis. The seroprevalence of E. histolytica was 33% (14/43) from the available stored sera. Based on the medical records, only 3 cases were clinically diagnosed as amoebic appendicitis, including 2 diagnosed at the time of appendectomy and 1 case diagnosed by rereview of the appendix after the development of postoperative complications. Retrospective analyses using PAS staining and PCR identified 3 and 3 more cases, respectively. Thus, E. histolytica infection was confirmed in 9 cases (15.8%) in the present study. Apart from a significantly higher leukocyte count in E. histolytica-positive patients than in negative patients (median, 13,760 versus 10,385 cells/µl, respectively, P = 0.02), there were no other differences in the clinical features of the PCR-positive and -negative groups. In conclusion, E. histolytica infection was confirmed in 9 (15.8%) of the appendicitis cases. However, only 3, including one diagnosed after intestinal perforation, were diagnosed before the present analyses. These results strongly suggest there is frequently a failure to detect trophozoites in routine examination, resulting in an underestimation of the incidence of amoebic appendicitis.

The Development of a Novel, Validated, Rapid and Simple Method for the Detection of Sarcocystis fayeri in Horse Meat in the Sanitary Control Setting.

Sarcocystis fayeri (S. fayeri) is a newly identified causative agent of foodborne disease that is associated with the consumption of raw horse meat. The testing methods prescribed by the Ministry of Health, Labour and Welfare of Japan are time consuming and require the use of expensive equipment and a high level of technical expertise. Accordingly, these methods are not suitable for use in the routine sanitary control setting to prevent outbreaks of foodborne disease. In order to solve these problems, we have developed a new, rapid and simple testing method using LAMP, which takes only 1 hour to perform and which does not involve the use of any expensive equipment or expert techniques. For the validation of this method, an inter-laboratory study was performed among 5 institutes using 10 samples infected with various concentrations of S. fayeri. The results of the inter-laboratory study demonstrated that our LAMP method could detect S. fayeri at concentrations greater than 10(4) copies/g. Thus, this new method could be useful in screening for S. fayeri as a routine sanitary control procedure.

Copper Homeostasis for the Developmental Progression of Intraerythrocytic Malarial Parasite.

Malaria is one of the world's most devastating diseases, particularly in the tropics. In humans, Plasmodium falciparum lives mainly within red blood cells, and malaria pathogenesis depends on the red blood cells being infected with the parasite. Nonesterified fatty acids (NEFAs), including cis-9-octadecenoic acid, and phospholipids have been critical for complete parasite growth in serum-free culture, although the efficacy of NEFAs in sustaining the growth of P. falciparum has varied markedly. Hexadecanoic acid and trans-9-octadecenoic acid have arrested development of the parasite, in association with down-regulation of genes encoding copper-binding proteins. Selective removal of Cu+ ions has blockaded completely the ring-trophozoite-schizont progression of the parasite. The importance of copper homeostasis for the developmental progression of P. falciparum has been confirmed by inhibition of copper-binding proteins that regulate copper physiology and function by associating with copper ions. These data have provided strong evidence for a link between healthy copper homeostasis and successive developmental progression of P. falciparum. Perturbation of copper homeostasis may be, thus, instrumental in drug and vaccine development for the malaria medication. We review the importance of copper homeostasis in the asexual growth of P. falciparum in relation to NEFAs, copperbinding proteins, apoptosis, mitochondria, and gene expression.

Draft Genome Sequence of High-Temperature-Adapted Protochlamydia sp. HS-T3, an Amoebal Endosymbiotic Bacterium Found in Acanthamoeba Isolated from a Hot Spring in Japan.

Here, we report the draft genome sequence of high-temperature-adapted Protochlamydia sp. strain HS-T3, an environmental chlamydia. This bacterium is an amoebal endosymbiont, found in Acanthamoeba isolated from a hot spring in Japan. Strain HS-T3 readily grew in mammalian cells at 37°C, a characteristic not previously reported for environmental chlamydiae.

An autopsy case of Balamuthia mandrillaris amoebic encephalitis, a rare emerging infectious disease, with a brief review of the cases reported in Japan.

Balamuthia mandrillaris is an amoeba found in fresh water and soil that causes granulomatous amoebic encephalitis. We report herein an autopsy case of B. mandrillaris amoebic encephalitis, which was definitely diagnosed by PCR. An 81-year-old man, who had Sjögren's syndrome, manifested drowsiness 2 months before his death with progressive deterioration. Neuroimaging demonstrated foci of T2- and fluid-attenuated inversion recovery high and T1 low-intensity with irregular post-contrast ring enhancement in the cerebral hemisphere, thalamus and midbrain. Pathologically, multiple hemorrhagic and necrotic lesions were found in the cerebrum, thalamus, midbrain, pons, medulla and cerebellum, which were characterized by liquefactive necrosis, marked edema, hemorrhage and necrotizing vasculitis associated with the perivascular accumulation of amoebic trophozoites, a few cysts, and the infiltration of numerous neutrophils and microglia/macrophages. The trophozoites were ovoid or round, 10-60 µm in diameter, and they showed foamy cytoplasm and a round nucleus with small karyosome in the center. The PCR and immunohistochemistry from paraffin-embedded brain specimens revealed angioinvasive encephalitis due to B. mandrillaris. Human cases of B. mandrillaris brain infection are rare in Japan, with only a few brief reports in the literature.

High-temperature adapted primitive Protochlamydia found in Acanthamoeba isolated from a hot spring can grow in immortalized human epithelial HEp-2 cells.

To elucidate how ancient pathogenic chlamydiae could overcome temperature barriers to adapt to human cells, we characterized a primitive chlamydia found in HS-T3 amoebae (Acanthamoeba) isolated from a hot spring. Phylogenetic analysis revealed the primitive species to be Protochlamydia. In situ hybridization staining showed broad distribution into the amoebal cytoplasm, which was supported by transmission electron microscopic analysis showing typical chlamydial features, with inclusion bodies including both elementary and reticular bodies. Interestingly, although most amoebae isolated from natural environments show reduced growth at 37°C, the HS-T3 amoebae harbouring the Protochlamydia grew well at body temperature. Although infection with Protochlamydia did not confer temperature tolerance to the C3 amoebae, the number of infectious progenies rapidly increased at 37°C with amoebal lysis. In immortalized human epithelial HEp-2 cells, fluorescence microscopic study revealed atypical inclusion of the Protochlamydia, and quantitative real-time polymerase chain reaction analyses also showed an increase in 16S ribosomal RNA DNA amounts. Together, these results showed that the Protochlamydia found in HS-T3 amoebae isolated from a hot spring successfully adapted to immortalized human HEp-2 cells at 37°C, providing further information on the evolution of ancient Protochlamydia to the present pathogenic chlamydiae.

Identification of photoactivated adenylyl cyclases in Naegleria australiensis and BLUF-containing protein in Naegleria fowleri.

Complete genome sequencing of Naegleria gruberi has revealed that the organism encodes polypeptides similar to photoactivated adenylyl cyclases (PACs). Screening in the N. australiensis genome showed that the organism also encodes polypeptides similar to PACs. Each of the Naegleria proteins consists of a "sensors of blue-light using FAD" domain (BLUF domain) and an adenylyl cyclase domain (AC domain). PAC activity of the Naegleria proteins was assayed by comparing sensitivities of Escherichia coli cells heterologously expressing the proteins to antibiotics in a dark condition and a blue light-irradiated condition. Antibiotics used in the assays were fosfomycin and fosmidomycin. E. coli cells expressing the Naegleria proteins showed increased fosfomycin sensitivity and fosmidomycin sensitivity when incubated under blue light, indicating that the proteins functioned as PACs in the bacterial cells. Analysis of the N. fowleri genome revealed that the organism encodes a protein bearing an amino acid sequence similar to that of BLUF. A plasmid expressing a chimeric protein consisting of the BLUF-like sequence found in N. fowleri and the adenylyl cyclase domain of N. gruberi PAC was constructed to determine whether the BLUF-like sequence functioned as a sensor of blue light. E. coli cells expressing a chimeric protein showed increased fosfomycin sensitivity and fosmidomycin sensitivity when incubated under blue light. These experimental results indicated that the sequence similar to the BLUF domain found in N. fowleri functioned as a sensor of blue light.