RESUMEN
Human granulocytic anaplasmosis (HGA) is a tick-borne infection caused by Anaplasma phagocytophilum. Only seven cases of HGA have been reported in Japan to date. We report the case of a 61-year-old female farmer who developed HGA with rash and rhabdomyolysis. The patient had fever and erythema covering the entire body, including the palms. An induration with an eschar was observed on the right leg, indicating that the patient had been bitten by a tick. Elevated serum creatinine and creatinine kinase levels and hematuria indicated rhabdomyolysis. We suspected Japanese spotted fever, a tick-borne illness caused by Rickettsia Japonica, and administered minocycline and ciprofloxacin for a week. Transient neutropenia and thrombocytopenia were observed, but the symptoms improved. Polymerase chain reaction (PCR) and antibody tests for R. japonica and Orientia tsutsugamushi, which causes scrub typhus, were both negative. The PCR test for severe fever with thrombocytopenia syndrome virus was also negative. Antibodies against A. phagocytophilum-related proteins were detected by western blotting, indicating seroconversion of IgG with paired serum samples, and the patient was diagnosed with HGA. HGA should be suspected in acute febrile patients with a history of outdoor activity and cytopenia, with or without a rash. A testing system and the accumulation of cases in Japan are necessary for the early diagnosis and appropriate treatment of HGA.
RESUMEN
In retrospective analyses, we report 3 febrile patients in Japan who had seroconversion to antibodies against Ehrlichia chaffeensis antigens detected by using an immunofluorescence and Western blot. Our results provide evidence of autochthonous human ehrlichiosis cases and indicate ehrlichiosis should be considered a potential cause of febrile illness in Japan.
Asunto(s)
Ehrlichia chaffeensis , Ehrlichiosis , Humanos , Ehrlichia , Estudios Retrospectivos , Japón/epidemiología , Ehrlichiosis/epidemiología , Antígenos Bacterianos , Anticuerpos AntibacterianosRESUMEN
Anaplasma phagocytophilum, an obligate intracellular bacterium that propagates within host granulocytes, is considered to modify the host intracellular environment for pathogenesis. However, the mechanism(s) underlying such host modifications remain unclear. Here, we aimed to investigate the relation between A. phagocytophilum and endoplasmic reticulum (ER) stress in THP-1 cells. A. phagocytophilum activated the three ER stress sensors: inositol-requiring enzyme-1 (IRE1), protein kinase RNA-like endoplasmic reticulum kinase (PERK), and activating transcription factor-6 (ATF6). IRE1 activation occurred immediately after host cell invasion by A. phagocytophilum; however, the activated IRE1-induced splicing of X-box-binding protein 1 was not promoted during A. phagocytophilum infection. This suppression was sustained even after the doxycycline-mediated elimination of intracellular A. phagocytophilum. IRE1 knockdown accelerated A. phagocytophilum-induced apoptosis and decreased intracellular A. phagocytophilum. These data suggest that A. phagocytophilum utilizes IRE1 activation to promote its own intracellular proliferation. Moreover, PERK and ATF6 partially mediated A. phagocytophilum-induced apoptosis by promoting the expression of CCAAT/enhancer-binding protein homologous protein, which induces the transcription of several proapoptotic genes. Thus, A. phagocytophilum possibly manipulates the host ER stress signals to facilitate intracellular proliferation and infection of surrounding cells before/after host cell apoptosis.
Asunto(s)
Anaplasma phagocytophilum/patogenicidad , Apoptosis/inmunología , Ehrlichiosis/inmunología , Estrés del Retículo Endoplásmico/inmunología , Interacciones Microbiota-Huesped/inmunología , Factor de Transcripción Activador 6/inmunología , Línea Celular , Ehrlichiosis/microbiología , Endorribonucleasas/inmunología , Humanos , Proteínas Serina-Treonina Quinasas/inmunología , Proteína 1 de Unión a la X-Box/inmunología , eIF-2 Quinasa/inmunologíaRESUMEN
Staphylococcal enterotoxin A (SEA) functions both as superantigens that stimulate non-specific T cell proliferation as well as potent gastrointestinal toxins. We previously reported that (-)-epigallocatechin gallate (EGCG) binds to SEA. Therefore, the ability of EGCG to inhibit SEA toxin activity was examined. As a result, EGCG significantly decreased SEA-induced expression and production of interferon gamma (IFN-γ). In addition, EGCG inhibited SEA-induced spleen cell proliferation. To investigate the role of the galloyl group in EGCG on SEA cytotoxicity in more detail, the effect of the binding of a hydroxyl group at position 3 of the galloyl group in EGCG to SEA on SEA cytotoxicity was examined using two methylated EGCG. SEA cytotoxicity was significantly controlled in both (-)-3''-Me-EGCG and (-)-4''-Me-EGCG. These results suggest that EGCG inhibits toxic activity via direct interaction with SEA or without any interaction with SEA. The binding affinity between SEA and EGCG under in vivo conditions was examined using a model solution. Although after treatment under acidic and alkaline conditions, the presence of protein and the digestive tract model solution, EGCG still interacted with SEA. Our studies are the first to demonstrate the effect of the binding of EGCG to SEA on toxin activity.
Asunto(s)
Catequina/análogos & derivados , Enterotoxinas/toxicidad , Animales , Catequina/química , Catequina/farmacología , Proliferación Celular/efectos de los fármacos , Citocinas/biosíntesis , Citocinas/genética , Interacciones Farmacológicas , Enterotoxinas/farmacología , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Concentración de Iones de Hidrógeno , Ratones , Estructura Molecular , Pancreatina , Pepsina A/farmacología , Unión ProteicaRESUMEN
We found Rickettsia raoultii infection in 6/261 brucellosis-negative patients with fever of unknown origin in brucellosis-endemic Inner Mongolia, China. We further identified Hyalomma asiaticum ticks associated with R. raoultii, H. marginatum ticks associated with R. aeschlimannii, and Dermacentor nuttalli ticks associated with both rickettsiae species in the autonomous region.
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Vectores Arácnidos/microbiología , Ixodidae/microbiología , Rickettsia/aislamiento & purificación , Rickettsiosis Exantemáticas/epidemiología , Animales , China/epidemiología , Humanos , Rickettsia/genética , Rickettsiosis Exantemáticas/microbiologíaRESUMEN
Staphylococcal enterotoxin A (SEA) is a toxin protein, and is the most common cause of staphylococcal food poisoning. Polyphenols, such as catechins, are known to interact with proteins. In this study, we investigated the binding of catechins to SEA using SPR (Biacore), Fourier transform infrared spectroscopy (FT-IR), isothermal titration calorimetry (ITC), and protein-ligand docking. We found that (−)-epigallocatechin gallate (EGCG) could strongly bind to SEA. According to thermodynamic parameters, a negative ΔG indicated that the interaction between EGCG and SEA was spontaneous, and the electrostatic force accompanied by hydrophobic binding forces may play a major role in the binding. Data from Western blot analysis and docking simulation suggest that the hydroxyl group at position 3 of the galloyl group in the catechin structure was responsible for binding affinity with the Y91 of the A-6 region of SEA active sites. Our results provide further understanding of the binding interactions between catechins and SEA, and the inhibition of toxin activities by catechins.
Asunto(s)
Catequina/metabolismo , Enterotoxinas/metabolismo , Calorimetría , Dominio Catalítico , Catequina/química , Enterotoxinas/química , Simulación del Acoplamiento Molecular , Unión Proteica , Espectroscopía Infrarroja por Transformada de Fourier , Resonancia por Plasmón de Superficie , TermodinámicaRESUMEN
In this study, we examined the inhibitory effects of 14 food additives derived from polyphenol samples on staphylococcal enterotoxin A (SEA) production and biofilm formation by Staphylococcus aureus. Tannic acid AL (TA), Purephenon 50 W (PP) and Polyphenon 70A (POP) at 0.25 mg/mL and Gravinol®-N (GN), Blackcurrant polyphenol AC10 (BP), and Resveratrol-P5 (RT) at 1.0 mg/mL significantly decreased SEA production by S. aureus C-29 (p < 0.05). TA, GN, BP, and RT significantly inhibited the expression of the sea gene in S. aureus C-29 (p < 0.05), while suppression attempts by PP and POP proved unsuccessful. After result analysis, it can be derived that TA, GN, BP, and RT inhibit the production of SEA. Of the six samples, each one significantly inhibited biofilm formation (p < 0.05). Food additives derived from polyphenols have viability to be used as a means to inhibit the enterotoxin production and control the biofilm formation of foodborne pathogens.
Asunto(s)
Biopelículas/efectos de los fármacos , Enterotoxinas/biosíntesis , Aditivos Alimentarios/química , Polifenoles/química , Polifenoles/farmacología , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/metabolismo , Biopelículas/crecimiento & desarrollo , Enterotoxinas/genética , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Staphylococcus aureus/genética , Staphylococcus aureus/fisiologíaRESUMEN
We retrospectively confirmed 2 cases of human Anaplasma phagocytophilum infection. Patient blood samples contained unique p44/msp2 for the pathogen, and antibodies bound to A. phagocytophilum antigens propagated in THP-1 rather than HL60 cells. Unless both cell lines are used for serodiagnosis of rickettsiosis-like infections, cases of human granulocytic anaplasmosis could go undetected.
Asunto(s)
Anaplasma phagocytophilum/genética , Anaplasmosis/diagnóstico , Ehrlichiosis/diagnóstico , Anciano , Anaplasmosis/sangre , Anaplasmosis/tratamiento farmacológico , Anaplasmosis/inmunología , Anticuerpos Antibacterianos/sangre , Proteínas de la Membrana Bacteriana Externa/inmunología , ADN Bacteriano/sangre , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Ehrlichiosis/sangre , Ehrlichiosis/tratamiento farmacológico , Ehrlichiosis/inmunología , Genes Bacterianos , Células HL-60 , Humanos , Japón , Masculino , Persona de Mediana Edad , Técnicas de Diagnóstico Molecular , Tipificación Molecular , Filogenia , ARN Ribosómico 16S/genética , Estudios Retrospectivos , Pruebas SerológicasRESUMEN
Tick-borne infectious diseases pose a serious health threat in certain regions of the world. Emerging infectious diseases caused by novel tick-borne pathogens have been reported that are causing particular concern. Several tick-borne diseases often coexist in the same foci, and a single vector tick can transmit two or more pathogens at the same time, which greatly increases the probability of co-infection in host animals and humans and can lead to an epidemic of tick-borne disease. The lack of epidemiological data and information on the specific clinical symptoms related to co-infection with tick-borne pathogens means that it is not currently possible to accurately and rapidly distinguish between a single pathogen infection and co-infection with multiple pathogens, which can have serious consequences. Inner Mongolia in the north of China is endemic for tick-borne infectious diseases, especially in the eastern forest region. Previous studies have found that more than 10% of co-infections were in host-seeking ticks. However, the lack of data on the specific types of co-infection with pathogens makes clinical treatment difficult. In our study, we present data on the co-infection types and the differences in co-infection among different ecological regions through genetic analysis of tick samples collected throughout Inner Mongolia. Our findings may aid clinicians in the diagnosis of concomitant tick-borne infectious diseases.
Asunto(s)
Coinfección , Enfermedades Transmisibles Emergentes , Enfermedades por Picaduras de Garrapatas , Garrapatas , Humanos , Animales , Garrapatas/microbiología , Coinfección/epidemiología , Enfermedades por Picaduras de Garrapatas/epidemiología , Enfermedades por Picaduras de Garrapatas/microbiología , Bacterias/genética , China/epidemiologíaRESUMEN
Ehrlichia species are obligatory intracellular bacteria transmitted by arthropods, and some of these species cause febrile diseases in humans and livestock. Genome sequencing has only been performed with cultured Ehrlichia species, and the taxonomic status of such ehrlichiae has been estimated by core genome-based phylogenetic analysis. However, many uncultured ehrlichiae exist in nature throughout the world, including Japan. This study aimed to conduct a molecular-based taxonomic and ecological characterization of uncultured Ehrlichia species or genotypes from ticks in Japan. We first surveyed 616 Haemaphysalis ticks by p28-PCR screening and analyzed five additional housekeeping genes (16S rRNA, groEL, gltA, ftsZ, and rpoB) from 11 p28-PCR-positive ticks. Phylogenetic analyses of the respective genes showed similar trees but with some differences. Furthermore, we found that V1 in the V1-V9 regions of Ehrlichia 16S rRNA exhibited the greatest variability. From an ecological viewpoint, the amounts of ehrlichiae in a single tick were found to equal approx. 6.3E+3 to 2.0E+6. Subsequently, core-partial-RGGFR-based phylogenetic analysis based on the concatenated sequences of the five housekeeping loci revealed six Ehrlichia genotypes, which included potentially new Ehrlichia species. Thus, our approach contributes to the taxonomic profiling and ecological quantitative analysis of uncultured or unidentified Ehrlichia species or genotypes worldwide.
Asunto(s)
Proteínas Bacterianas/genética , Biodiversidad , ADN Bacteriano/genética , Ehrlichia/fisiología , Ehrlichiosis/diagnóstico , Garrapatas/microbiología , Animales , ADN Bacteriano/análisis , Ehrlichiosis/genética , Ehrlichiosis/parasitología , Humanos , Japón , FilogeniaRESUMEN
Non-pathogenic Rickettsia species LON strains closely related to an agent of Japanese spotted fever (JSF), R. japonica, were isolated in Japan from Haemaphysalis longicornis ticks in 2001. However, the biological properties of LONs in mammalian host cells are poorly understood. In this study, microscopic analysis showed that LONs in a mouse-derived L929 host cell line were rod shaped with sizes of 0.3-0.5 × 0.5-2.0 µm. Molecular analysis revealed the existence of a LON-specific disrupted open reading frame in R. japonica-related group-specific DNA regions. Growth kinetics of LON-2 and LON-13 strains analyzed by a quantitative real-time PCR showed 100-fold or more increment of LONs cultured in L929 host cells at 30°C and slightly less increment at 33°C, and 25-fold increment in human-derived THP-1 host cells at 35°C on day 7 (168 h) post infection. The generation times of the two LON strains cultured in L929 and THP-1 were estimated to be 9.4-12.9 h and 9.6-10.9 h, respectively. To our knowledge, this is the first report on the biological characteristics of Rickettsia sp. LON strains in mammalian cells, which may provide significant information for the experimental approaches for other rickettsiae.
Asunto(s)
Rickettsia/genética , Rickettsiosis Exantemáticas/microbiología , Garrapatas/microbiología , Animales , Línea Celular , ADN Bacteriano/aislamiento & purificación , Humanos , Ixodidae/microbiología , Japón , Ratones , Reacción en Cadena en Tiempo Real de la Polimerasa , Rickettsia/aislamiento & purificación , Células THP-1RESUMEN
BACKGROUND: Borrelia miyamotoi is a newly described relapsing fever spirochete transmitted by ixodid tick species. Little is known about the prevalence of B. miyamotoi infections in humans and ticks in Inner Mongolia, China. Therefore, we investigated the prevalence of B. miyamotoi in Ixodes persulcatus ticks, and we aimed to isolateB. miyamotoi from I. persulcatus from four regions of Greater Khingan, Inner Mongolia, China. METHODS: From May to June each year during the period 2016-2019, host-seeking adult I. persulcatus ticks were collected from vegetation. Genomic DNA was prepared from half of each tick body for PCR template, and the remaining half was used to cultivate B. miyamotoi in BSK-M medium. We employed quantitative real-time PCR (qPCR) to detect Borrelia DNA in the ticks and to calculate the prevalence of B. miyamotoi and infections with other borreliae. For characterization of the isolated B. miyamotoi, we performed draft genome sequencing and multilocus sequencing analysis (MLSA). RESULTS: A total of 2656 adult I. persulcatus ticks were collected. The overall prevalence of relapsing fever (RF) borreliae in ticks was 5.0% (134/2656) and that of Lyme disease (LD) borreliae was 43.8% (1164/2656). Co-infection with RF and LD borreliae was observed in 63 ticks (2.4%). Ticks that were positive for RF borreliae by qPCR were subjected to glycerophosphodiester diester phosphodiesterase gene (glpQ) PCR amplification and sequencing, through which we identified the RF borrelia specimens as B. miyamotoi. Furthermore, the B. miyamotoi strain Hetao-1 was isolated from I. persulcatus, and a draft genome sequence was obtained from the isolate. Sequencing determined the strain Hetao-1 genome to be approximately 906.1 kbp in length (28.9% average GC content), and MLSA identified the strain as ST633, which has previously been reported in Japan and Mongolia. CONCLUSION: We detected B. miyamotoi from I. persulcatus ticks collected in Inner Mongolia, and successfully isolated a B. miyamotoi strain. To our knowledge, this is the first study to culture a B. miyamotoi isolate from China. The data on the prevalence of B. miyamotoi and other borreliae in I. persulcatus ticks will be fundamental for future epidemiological studies of B. miyamotoi disease in Inner Mongolia.
Asunto(s)
Borrelia/genética , Ixodes/microbiología , Fiebre Recurrente/microbiología , Animales , Técnicas de Tipificación Bacteriana , Borrelia/aislamiento & purificación , China/epidemiología , Monitoreo Epidemiológico , Genómica , Humanos , Tipificación de Secuencias Multilocus , Reacción en Cadena en Tiempo Real de la Polimerasa , Fiebre Recurrente/epidemiologíaRESUMEN
Angiogenesis is crucial for tumor growth and hematogenous metastasis. Specifically expressed and functional protein molecules in angiogenic endothelial cells, especially on the plasma membrane, may be molecular targets for antiangiogenic drugs and drug delivery systems (DDS) in cancer therapy. To discover such target molecules, we performed subcellular proteome analysis of human umbilical vein endothelial cells (HUVECs) treated with or without vascular endothelial growth factor (VEGF) using 2-dimensional difference in-gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-TOF/TOF-MS). Among the identified proteins, BiP/GRP78, a molecular chaperone, was highly expressed in the membrane/organelle fraction of HUVECs after VEGF treatment. The involvement of BiP in VEGF-induced angiogenesis was examined by RNA interference. BiP knockdown significantly suppressed VEGF-induced endothelial cell proliferation and VEGF-induced phosphorylation of extracellular-regulated kinase 1/2, phospholipase C-γ, and VEGF receptor-2 in HUVECs. Cell surface biotinylation analysis revealed that the cell surface expression of BiP was elevated in VEGF-activated HUVECs. Aiming to apply BiP to a target molecule in liposomal DDS, we developed liposomes modified with the WIFPWIQL peptide, which has been shown to bind to BiP, and investigated its potential for cancer therapy. The WIFPWIQL-modified liposomes (WIFPWIQL liposomes) were significantly taken up by VEGF-activated HUVECs as compared to peptide-unmodified liposomes. WIFPWIQL liposomes appeared to accumulate in tumor endothelial cells in vivo. WIFPWIQL liposomes containing doxorubicin significantly suppressed tumor growth and prolonged the survival of colon26 NL-17 carcinoma cell-bearing mice. In summary, BiP may regulate VEGF-induced endothelial cell proliferation through VEGFR-2-mediated signaling and be an effective target molecule for cancer antineovascular therapy.
Asunto(s)
Inhibidores de la Angiogénesis/administración & dosificación , Neoplasias del Colon/irrigación sanguínea , Neoplasias del Colon/tratamiento farmacológico , Proteínas de Choque Térmico/metabolismo , Neoplasias de la Próstata/irrigación sanguínea , Neoplasias de la Próstata/tratamiento farmacológico , Animales , Neoplasias del Colon/metabolismo , Doxorrubicina/administración & dosificación , Sistemas de Liberación de Medicamentos/métodos , Electroforesis en Gel Bidimensional , Chaperón BiP del Retículo Endoplásmico , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Técnicas de Silenciamiento del Gen , Proteínas de Choque Térmico/deficiencia , Proteínas de Choque Térmico/genética , Humanos , Liposomas/administración & dosificación , Liposomas/farmacocinética , Sistema de Señalización de MAP Quinasas , Masculino , Ratones , Neovascularización Patológica/tratamiento farmacológico , Oligopéptidos/administración & dosificación , Neoplasias de la Próstata/metabolismo , Proteómica , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Factor A de Crecimiento Endotelial Vascular/farmacologíaRESUMEN
Several physical and psychological stresses frequently become triggers for gastrointestinal disorders such as ulcer. In this study, we tried to identify serum proteins as potential biomarkers for the evaluation of stress-induced gastric ulcer. By proteomic analysis using rats with gastric ulcer induced by water immersion and restraint (WIR) stress as an animal model, we found quantitative changes in several serum proteins, including creatine kinase muscle M chain (CK-M) and apolipoprotein A-IV (ApoA4) in the stressed rats. On western blotting and enzyme-linked immunosorbent assay (ELISA), we confirmed that serum CK-M was remarkably increased by WIR stress. However, ApoA4 appeared to be decreased by fasting, but not WIR stress, which is usually applied prior to WIR stress. The findings suggest that these two serum proteins might be useful as biomarkers, CK-M for stress-induced gastric ulcer and ApoA4 for starvation.
Asunto(s)
Úlcera Gástrica/metabolismo , Estrés Psicológico/complicaciones , Animales , Apolipoproteínas A , Proteínas Sanguíneas/metabolismo , Úlcera Duodenal/metabolismo , Ensayo de Inmunoadsorción Enzimática , Inmersión , Masculino , Proteínas/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
ABSTRACT: The influence of muscle fiber direction (parallel or perpendicular) in relation to the inoculation surface on migration of Salmonella Enteritidis, Staphylococcus aureus, and Escherichia coli into raw chicken breasts was examined. Chicken breast samples with two types of surface fibers (running parallel or perpendicular to the surface) were inoculated with cultures of each bacterium. Inoculated samples were stored for 5 min, 1 h, or 24 h at 4°C. After storage, the samples were divided into segments, and bacterial counts were determined in different regions (inoculation surface, inoculation surface to 1 cm, 1 to 2 cm, 2 to 4 cm, and 4 to 6 cm). The migration of bacteria did not change at 5 min or 1 h regardless of fiber direction. However, after 24 h each bacterium was detected at 4 to 6 cm in the pieces of sample with a perpendicular muscle fiber surface cut. Although these bacteria were detected at 4 to 6 cm in samples with muscle fibers perpendicular to the inoculated surface, these results do not clearly indicate that bacteria migrated into the chicken breast. To monitor actual migration of bacteria into the chicken breast, the tops of the perpendicular muscle fibers of the breast sample were inoculated with bioluminescent E. coli Xen-14. Various regions of the breast sample (inoculation surface and cut surfaces at 1, 2, 4, and 6 cm) were stamped directly on growth medium. Culture revealed that the bacteria migrated directly under the contaminated site and dispersed along the surface of the chicken breast segments. More bacteria distributed laterally than migrated directly below the contamination site. These results suggest that the direction of the muscle fibers is a major factor influencing migration of pathogenic bacteria into chicken breast.
Asunto(s)
Neoplasias de la Mama , Salmonella enteritidis , Animales , Pollos , Recuento de Colonia Microbiana , Escherichia coli , Contaminación de Alimentos/análisis , Microbiología de Alimentos , Humanos , Carne , Fibras Musculares Esqueléticas , Staphylococcus aureusRESUMEN
Anaplasma phagocytophilum, which belongs to the order Rickettsiales, is an obligate intracellular bacterium and causes an emerging, tickborne, and febrile infectious disease, anaplasmosis, in humans and other mammals. This bacterium expresses a variety of 44-kDa immunodominant proteins encoded by the p44/msp2 multigene family on the surface for the purpose of avoiding the host immune defense due to the antigenic variation. In Japan, little is known about the molecular and biological features of A. phagocytophilum. In this study, we tried to characterize in detail the p44/msp2 multigene family of A. phagocytophilum from two tick species, Ixodes persulcatus and I. ovatus in Japan. A total of 174 amino acid sequences from the recombinant p44/msp2 clones after TA cloning of the amplicons obtained from the ticks were phylogenetically analyzed. The results showed that most of the clone sequences from I. ovatus were very similar to each other, but the sequences from I. persulcatus were diverse, and the sequences from the ticks were distinct from those from a wild deer that was previously reported. These findings suggest that Ixodes ticks are probably responsible for the transmission of certain genetic variants of A. phagocytophilum and that additional organism selection might occur in I. ovatus.
Asunto(s)
Anaplasma phagocytophilum/aislamiento & purificación , Proteínas de la Membrana Bacteriana Externa/genética , Ixodes/microbiología , Animales , Análisis por Conglomerados , ADN Bacteriano/química , ADN Bacteriano/genética , Japón , Datos de Secuencia Molecular , Familia de Multigenes , Filogenia , Análisis de Secuencia de ADN , Homología de SecuenciaRESUMEN
Anaplasma (formerly Ehrlichia) phagocytophilum, Ehrlichia chaffeensis, and Neorickettsia (formerly Ehrlichia) sennetsu are intracellular vector-borne pathogens that cause human ehrlichiosis, an emerging infectious disease. We present the complete genome sequences of these organisms along with comparisons to other organisms in the Rickettsiales order. Ehrlichia spp. and Anaplasma spp. display a unique large expansion of immunodominant outer membrane proteins facilitating antigenic variation. All Rickettsiales have a diminished ability to synthesize amino acids compared to their closest free-living relatives. Unlike members of the Rickettsiaceae family, these pathogenic Anaplasmataceae are capable of making all major vitamins, cofactors, and nucleotides, which could confer a beneficial role in the invertebrate vector or the vertebrate host. Further analysis identified proteins potentially involved in vacuole confinement of the Anaplasmataceae, a life cycle involving a hematophagous vector, vertebrate pathogenesis, human pathogenesis, and lack of transovarial transmission. These discoveries provide significant insights into the biology of these obligate intracellular pathogens.
Asunto(s)
Ehrlichia/genética , Ehrlichiosis/genética , Genómica/métodos , Animales , Biotina/metabolismo , Reparación del ADN , Ehrlichiosis/microbiología , Genoma , Humanos , Modelos Biológicos , Filogenia , Rickettsia/genética , GarrapatasRESUMEN
Anaplasma phagocytophilum, an agent of human granulocytic anaplasmosis, is an obligatory intracellular bacterium that dominantly produces P44 outer membrane proteins encoded by the p44/msp2 multigene family, which are major antigens for serodiagnosis. However, A. phagocytophilum antigens from cultures with different cell lines seem to have varying reactivities with sera. In this study, we performed RNA-seq to investigate the P44 expression of A. phagocytophilum propagated in 4 cell lines. In infected HL-60 cells, the P44-2b transcript was predominant in the first RNA-seq analysis (HL-60.1). However, the P44-23 transcript was predominant in the second RNA-seq analysis at 1 month after additional passages (HL-60.2). We further analyzed the P44 expression of A. phagocytophilum cultured in THP-1, NB4, and RF/6A cells through consecutive passages in the same cell lines for 1 year after transferring A. phagocytophilum from infected HL-60 cells to the respective cell lines. In the long-term cultures, P44-18, P44-78, and P44-51 were predominantly transcribed in infected THP-1, NB4, and RF/6A cells, respectively. Therefore, the predominant shifts of different P44-expressing transcripts of A. phagocytophilum might occur during cell culture even in the same cell line at different time points of sample harvest (HL-60.1 and HL-60.2), which may be attributed to host cell adaptation/selection/interaction.
Asunto(s)
Anaplasma phagocytophilum/crecimiento & desarrollo , Proteínas de la Membrana Bacteriana Externa/biosíntesis , Perfilación de la Expresión Génica , Animales , Proteínas de la Membrana Bacteriana Externa/genética , Línea Celular , Humanos , Macaca mulatta , Análisis de Secuencia de ARN , Pase SeriadoRESUMEN
Ehrlichiosis is a tick-borne bacterial disease caused by pathogens of the Ehrlichia genus. Although human ehrlichiosis has not been reported in Japan, Ehrlichia spp., which are closely related to Ehrlichia chaffeensis, were detected in several species of ixodid ticks. In this study, the presence of Ehrlichia spp. in ticks in Japan was studied by using isolation and molecular detection methods. In total, 1237 ticks were collected from vegetation in western, central, and eastern parts of Japan. The ticks were tested for detection of ehrlichial DNA with a nested polymerase chain reaction and/or isolation by inoculation of mice with the homogenate. Ehrlichial DNA was detected in 29 of these ticks. The ehrlichial DNAs, groEL and 16S rRNA genes, detected in Ixodes turdus showed a high similarity to those of E. chaffeensis with 94.7% and 99.2% identity, respectively. Ehrlichia sp. HF and Candidatus Neoehrlichia mikurensis were also detected in I. ovatus. Furthermore, Ehrlichia sp. HF was isolated from laboratory mice that were intraperitoneal inoculated with I. ovatus tick homogenate. Some ehrlichial agents detected in Ixodes ticks might be a previously unknown Ehrlichia species. In this study, Candidatus N. mikurensis was detected in I. ovatus ticks. Because I. ovatus is distributed widely and cases of its tick bite in humans are ubiquitously reported in Japan, there is a potential for ehrlichiosis to be endemic to Japan, necessitating intensive surveillance of this infectious disease.