RESUMO
The severity of Entamoeba histolytica infection is determined by host immunology, pathogen virulence, and the intestinal environment. Conventional research for assessing pathogen virulence has been mainly performed using laboratory strains, such as a virulent HM-1: IMSS (HM-1) and an avirulent Rahman, under various artificial environmental conditions because of the difficulties of axenic isolation of the clinical strains. However, it is still unclear whether scientific knowledge based on laboratory strains are universally applicable to the true pathogenesis. Hereby, we performed transcriptomic analysis of clinical strains from patients with different degrees of disease severity, as well as HM-1 under different conditions. Even after several months of axenization, Clinical strains show the distinct profile in gene expression during in vitro passage, moreover, difference between any 2 of these strains was much greater than the changes on the liver challenge. Interestingly, 26 DEGs, which were closely related to the biological functions, were oppositely up- or down regulated between virulent Ax 19 (liver abscess) and avirulent Ax 11 (asymptomatic carrier). Additionally, RNAseq using laboratory strain (HM1) showed more than half of genes were differently expressed between continuously in vitro passaged HM1 (in vitro HM1) and periodically liver passaged HM1 (virulent HM1), which was much greater than the changes on the liver passage of virulent HM1. Also, transcriptomic analysis of a laboratory strain revealed that continuous environmental stress enhances its virulence via a shift in its gene expression profile. Changes in gene expression patterns on liver abscess formation were not consistent between clinical and laboratory strains.
Assuntos
Amebíase , Disenteria Amebiana , Entamoeba histolytica , Abscesso Hepático , Expressão Gênica , Humanos , Índice de Gravidade de DoençaRESUMO
Entamoeba moshkovskii, according to recent studies, appears to exert a more significant impact on diarrhoeal infections than previously believed. The efficient identification and genetic characterization of E. moshkovskii isolates from endemic areas worldwide are crucial for understanding the impact of parasite genomes on amoebic infections. In this study, we employed a multilocus sequence typing system to characterize E. moshkovskii isolates, with the aim of assessing the role of genetic variation in the pathogenic potential of E. moshkovskii. We incorporated 3 potential genetic markers: KERP1, a protein rich in lysine and glutamic acid; amoebapore C (apc) and chitinase. Sequencing was attempted for all target loci in 68 positive E. moshkovskii samples, and successfully sequenced a total of 33 samples for all 3 loci. The analysis revealed 17 distinct genotypes, labelled M1M17, across the tested samples when combining all loci. Notably, genotype M1 demonstrated a statistically significant association with diarrhoeal incidence within E. moshkovskii infection (P = 0.0394). This suggests that M1 may represent a pathogenic strain with the highest potential for causing diarrhoeal symptoms. Additionally, we have identified a few single-nucleotide polymorphisms in the studied loci that can be utilized as genetic markers for recognizing the most potentially pathogenic E. moshkovskii isolates. In our genetic diversity study, the apc locus demonstrated the highest Hd value and π value, indicating its pivotal role in reflecting the evolutionary history and adaptation of the E. moshkovskii population. Furthermore, analyses of linkage disequilibrium and recombination within the E. moshkovskii population suggested that the apc locus could play a crucial role in determining the virulence of E. moshkovskii.
Assuntos
Entamoeba , Tipagem de Sequências Multilocus , Marcadores Genéticos , Entamoeba/genética , Entamoeba/classificação , Entamoeba/isolamento & purificação , Humanos , Entamebíase/parasitologia , Entamebíase/epidemiologia , Genótipo , Polimorfismo de Nucleotídeo Único , Variação Genética , FilogeniaRESUMO
Epidemiological studies on amoebic infections are complicated by morphological overlap between the pathogenic E. histolytica, the commensal E. dispar and the amphizoic E. moshkovskii, necessitating molecular identification. The present study developed a simple and economical 18S PCR-RFLP method for the simultaneous detection and differentiation of the three species. PCR products were differentiated by Tat1 restriction digestion generating three different RFLP patterns. Validation was conducted by screening 382 faecal samples from human patients from Kolkata, India, hospitalized for diarrhoea. Analysis indicated that the PCR-RFLP could successfully differentiate between the three species and was confirmed by sequence analysis. This method could prove useful for clinical and epidemiological studies of amoebiasis.
Assuntos
Amebíase , Entamoeba histolytica , Entamoeba , Entamebíase , Humanos , Entamoeba/genética , Polimorfismo de Fragmento de Restrição , Reação em Cadeia da Polimerase/métodos , Fezes/química , DNA de Protozoário/genética , DNA de Protozoário/análise , Entamoeba histolytica/genéticaRESUMO
Amoebiasis is an infection caused by enteric protozoa, most commonly Entamoeba histolytica, and is globally considered a potentially severe and life-threatening condition. To understand the impact of the parasite genome on disease outcomes, it is important to study the genomes of infecting strains in areas with high disease prevalence. These studies aim to establish correlations between parasite genotypes and the clinical presentation of amoebiasis. We employ a strain typing approach that utilizes multiple loci, including SREHP and three polymorphic non-coding loci (tRNA-linked array N-K2 and loci 1-2 and 5-6), for high-resolution analysis. Distinct clinical phenotype isolates underwent amplification and sequencing of studied loci. The nucleotide sequences were analysed using Tandem Repeats Finder to detect short tandem repeats (STRs). These patterns were combined to assign a genotype, and the correlation between clinical phenotypes and repetitive patterns was statistically evaluated. This study found significant polymorphism in the size and number of PCR fragments at SREHP and 5-6 locus, while the 1-2 locus and NK2 locus showed variations in PCR product sizes. Out of 41 genotypes, two (I6 and I41) were significantly associated with their respective disease outcomes and were found in multiple isolates. We observed that I6 was linked with a symptomatic outcome, with a statistically significant p-value of 0.0183. Additionally, we found that I41 was associated with ALA disease outcome, with a p-value of 0.0089. Our study revealed new repeat units not previously reported, unveiling the genetic composition of E. histolytica strains in India, associated with distinct disease manifestations.
Assuntos
Entamoeba histolytica , Entamebíase , Humanos , Entamebíase/parasitologia , Polimorfismo Genético , Entamoeba histolytica/genética , Fenótipo , Repetições de MicrossatélitesRESUMO
BACKGROUND: Amoebozoa is a eukaryotic supergroup composed of unicellular and multicellular amoebic protozoa (e.g. Acanthamoeba, Dictyostelium, and Entamoeba). They are model organisms for studies in cellular and evolutionary biology and are of medical and veterinary importance. Despite their importance, Amoebozoan genome organization and genetic diversity remain poorly studied due to a lack of high-quality reference genomes. The slime mold Dictyostelium discoideum is the only Amoebozoan species whose genome is available at the chromosome-level. RESULTS: Here, we provide a near-chromosome-level assembly of the Entamoeba histolytica genome, the second semi-completed Amoebozoan genome. The availability of this improved genome allowed us to discover inter-strain heterogeneity in ploidy at the near-chromosome or sub-chromosome level among 11 clinical isolates and the reference strain. Furthermore, we observed ploidy-independent regulation of gene expression, contrary to what is observed in other organisms, where RNA levels are affected by ploidy. CONCLUSIONS: Our findings offer new insights into Entamoeba chromosome organization, ploidy, transcriptional regulation, and inter-strain variation, which will help to further decipher observed spectrums of virulence, disease symptoms, and drug sensitivity of E. histolytica isolates.
Assuntos
Entamoeba histolytica , Genoma de Protozoário , Cromossomos/genética , Entamoeba histolytica/genética , Genes de Protozoários , Ploidias , Proteínas de Protozoários/genéticaRESUMO
Intestinal amebiasis is a major cause of diarrhea. However, research on host-amebae interactions has been hampered owing to a lack of appropriate animal models. Recently, a mouse model of intestinal amebiasis was established, and using it, we reported that Entamoeba moshkovskii colonized the intestine in a manner similar to that of the pathogenic Entamoeba histolytica In this study, we evaluated the protective mechanisms present against amebae using this model. CBA/J mice infected with E. histolytica had a persistent infection without apparent symptoms. In contrast, E. moshkovskii-infected mice rapidly expelled the ameba, which was associated with weight loss, diarrhea, and intestinal damage characterized by apoptosis of intestinal epithelial cells (IECs). Expression of NKG2D on intestinal intraepithelial lymphocytes (IELs) and IFN-γ-producing cells in Peyer's patches were significantly induced after infection with E. moshkovskii but not with E. histolytica IFN-γ-deficient mice infected with E. moshkovskii showed no obvious symptoms. Notably, none of these mice expelled E. moshkovskii, indicating that IFN-γ is responsible not only for intestinal symptoms but also for the expulsion of amebae. Furthermore, apoptosis of IECs and expression of NKG2D on IELs observed in E. moshkovskii-infected mice did not occur in the absence of IFN-γ. In vivo blocking of NKG2D in mice infected with E. moshkovskii enabled ameba to survive longer and remarkably reduced apoptotic IECs. Our results clearly demonstrate a novel protective mechanism exerted by IFN-γ against intestinal amebae, including induction of cytotoxicity of IELs toward IECs.
Assuntos
Entamoeba histolytica/imunologia , Interferon gama/imunologia , Intestinos/imunologia , Intestinos/patologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Animais , Apoptose/imunologia , Modelos Animais de Doenças , Entamebíase/imunologia , Entamebíase/parasitologia , Células Epiteliais/imunologia , Interações Hospedeiro-Parasita/imunologia , Inflamação/imunologia , Interferon gama/genética , Intestinos/parasitologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Knockout , Nódulos Linfáticos Agregados/imunologia , Linfócitos T/imunologiaRESUMO
BACKGROUND: Although there have been several reports of treating large post-endoscopic submucosal dissection (ESD) ulcers by covering them with a polyglycolic acid sheet (PGAs), this approach presents problems regarding PGAs delivery. This study assessed the usefulness of a device delivery station system (DDSS) to evaluate the appropriate and rapid PGAs coating method with DDSS. METHODS: Thirty-nine of 41 patients who were diagnosed with early gastric cancer over 20 mm in diameter and pathologically diagnosed with well-differentiated adenocarcinoma were randomly allocated to the following two groups according to delivery method: the conventional PGAs delivery group (C group) (n = 19) and the new DDSS group (DDSS group) (n = 20). The primary outcome was the coating area per minute in the C group and DDSS group (cm2/min). RESULTS: There were significant differences in the coating time (min), with values of 34.1 (15.0-60.7) vs. 16.85 (11.5-27.2) min for the C group and DDSS group, respectively (p = 0.001). There was also a significant difference in coating area per minute, with values of 0.261 (0.02-1.00) and 0.96 (0.173-2.06) cm2/min for the C group and DDSS group, respectively (p = 0.001). There were four cases of post-ESD bleeding (1-7 days after ESD) in the C group compared with 0 in the DDSS group, which represented a significant difference (p = 0.030). CONCLUSIONS: The DDSS was very useful for rapidly delivering and tightly attaching a PGAs to control post-ESD bleeding. TRIAL REGISTRATION: University Hospital Medical Network (UMIN) 000026377.
Assuntos
Sistemas de Liberação de Medicamentos/instrumentação , Ressecção Endoscópica de Mucosa/efeitos adversos , Ácido Poliglicólico , Complicações Pós-Operatórias/terapia , Úlcera Cutânea/terapia , Cicatrização/efeitos dos fármacos , Adenocarcinoma/diagnóstico , Adenocarcinoma/cirurgia , Idoso , Idoso de 80 Anos ou mais , Ressecção Endoscópica de Mucosa/métodos , Desenho de Equipamento , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Complicações Pós-Operatórias/etiologia , Estudos Prospectivos , Úlcera Cutânea/etiologia , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/cirurgia , Fatores de Tempo , Resultado do TratamentoRESUMO
A congenital abnormality of the inferior vena cava is said to be an anatomical risk factor for venous thromboembolism. In this report, we present a case of a patient with a left duplicated common iliac vein who developed a venous thromboembolism following total abdominal hysterectomy and bilateral salpingo-oophorectomy. Only 2 items were risk factors for thromboembolism: age of ≥40 years and open surgery duration of ≥30 min; no congenital abnormalities of the inferior vena cava or thrombotic factors were observed. Thus, it was suspected that the duplicated common iliac vein could have caused the venous thromboembolism.
Assuntos
Veia Ilíaca/anormalidades , Malformações Vasculares/complicações , Trombose Venosa/etiologia , Fatores Etários , Anticoagulantes/uso terapêutico , Angiografia por Tomografia Computadorizada , Feminino , Humanos , Histerectomia/efeitos adversos , Veia Ilíaca/diagnóstico por imagem , Pessoa de Meia-Idade , Duração da Cirurgia , Flebografia/métodos , Fatores de Risco , Salpingo-Ooforectomia/efeitos adversos , Terapia Trombolítica , Resultado do Tratamento , Malformações Vasculares/diagnóstico por imagem , Trombose Venosa/diagnóstico por imagem , Trombose Venosa/tratamento farmacológicoRESUMO
BACKGROUND/AIMS: Multiple colorectal polyps with a diameter in the range of 10-19 mm are unable to be retrieved through a 3-mm endoscopic channel by the aspiration method. This study aims to assess the usefulness of Catcher Tag retrieval, which not only allows the accurate identification of the resected location but also enables the easiest retrieval in a short time without any special device. METHODS: One hundred thirty five patients (483 polyps) were diagnosed with colorectal neoplasm, and 64 patients (225 polyps) were enrolled and randomly allocated into the Net forceps group (NET) and the Catcher Tagged group (TAG). In TAG, 3 types of colored ring-threads were used to retrieve resected polyps. After local injection of natural saline, ring-threads were placed close to polyps. The primary outcome was the number of one-to-one correspondence locations (UMIN000020826). RESULTS: There was significant difference in one-to-one correspondence (p = 0.004). The average retrieval procedure time was 13.56 ± 3.47 (min) in NET and 3.55 ± 1.68 in TAG (p = 0.006). In NET, 1 polyp in each of 4 cases was lost during endoscopic mucosal resection and 2 polyps were lost in 1 case. In TAG, no polyp was lost (p = 0.016). CONCLUSION: The Catcher Tagged method is very useful for accurate one-to-one correspondence locations and pathological evaluation, and easy-to-retrieve multiple resected specimens.
Assuntos
Pólipos do Colo/cirurgia , Colonoscopia/métodos , Neoplasias Colorretais/cirurgia , Ressecção Endoscópica de Mucosa/métodos , Adulto , Pólipos do Colo/patologia , Colonoscopia/instrumentação , Neoplasias Colorretais/patologia , Ressecção Endoscópica de Mucosa/instrumentação , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Estudos Prospectivos , Instrumentos CirúrgicosRESUMO
Unique species of macaques are distributed across Sulawesi Island, Indonesia, and the details of Entamoeba infections in these macaques are unknown. A total of 77 stool samples from Celebes crested macaques (Macaca nigra) and 14 stool samples from pigs were collected in Tangkoko Nature Reserve, North Sulawesi, and the prevalence of Entamoeba infection was examined by PCR. Entamoeba polecki was detected in 97% of the macaques and all of the pigs, but no other Entamoeba species were found. The nucleotide sequence of the 18S rRNA gene in E. polecki from M. nigra was unique and showed highest similarity with E. polecki subtype (ST) 4. This is the first case of identification of E. polecki ST4 from wild nonhuman primates. The sequence of the 18S rRNA gene in E. polecki from pigs was also unique and showed highest similarity with E. polecki ST1. These results suggest that the diversity of the 18S rRNA gene in E. polecki is associated with differences in host species and geographic localization, and that there has been no transmission of E. polecki between macaques and pigs in the study area.
Assuntos
Entamoeba/genética , Entamoeba/isolamento & purificação , Entamebíase/parasitologia , Macaca/parasitologia , RNA Ribossômico 18S/genética , Suínos/parasitologia , Animais , Sequência de Bases , Conservação dos Recursos Naturais , DNA de Protozoário , Entamoeba/classificação , Entamoeba/citologia , Entamebíase/epidemiologia , Entamebíase/transmissão , Entamebíase/veterinária , Genes de Protozoários , Genoma de Protozoário , Indonésia/epidemiologia , Filogenia , Reação em Cadeia da Polimerase , Prevalência , Alinhamento de Sequência , Análise de Sequência de DNA , Especificidade da Espécie , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/parasitologiaRESUMO
We have proposed the revival of the name Entamoeba nuttalli for a virulent ameba strain, P19-061405, from a rhesus macaque and located it phylogenetically between E. histolytica and E. dispar. As E. nuttalli was originally described for an ameba found in a toque macaque in Sri Lanka, the prevalence and characteristics of Entamoeba species in wild toque macaques were examined. PCR analysis of 227 stool samples from six locations showed positive rates for E. nuttalli, E. dispar, and E. histolytica of 18.5%, 0.4%, and 0%, respectively. Fifteen E. nuttalli strains were cultured successfully from five locations. The 18S ribosomal RNA gene showed only three nucleotide differences in comparison with P19-061405 strain. In isoenzyme analysis, the pattern of hexokinase in Sri Lankan strains was different from that of P19-061405 strains and the difference was confirmed by analysis of the genes. Hepatic inoculation of one of the Sri Lankan E. nuttalli strains in hamsters resulted in amebic abscess formation and body weight loss. These results demonstrate that E. nuttalli is prevalent in wild toque macaques and that several characteristics of the strains are unique. We conclude that use of the name E. nuttalli is appropriate for the new Entamoeba species found in nonhuman primates.
Assuntos
Entamoeba/enzimologia , Entamoeba/isolamento & purificação , Entamebíase/veterinária , Hexoquinase/metabolismo , Macaca/parasitologia , Doenças dos Macacos/parasitologia , Animais , Cricetinae , DNA de Protozoário/genética , Entamoeba/genética , Entamebíase/parasitologia , Genótipo , Hexoquinase/genética , Hexoquinase/isolamento & purificação , Isoenzimas , Masculino , Mesocricetus , Filogenia , Reação em Cadeia da Polimerase/métodos , Polimorfismo Genético , Prevalência , Sri LankaRESUMO
A 66-year-old Japanese male with a history of a rectal ulcer and rectovesical fistula following brachytherapy and radiotherapy for prostate cancer, who had undergone colostomy and vesicotomy presented with a painful peristomal ulcer of approximately 5 x 2.5cm adjacent to the direction of 6 o'clock of the stoma in his left lower abdomen. Although he was admitted to be treated with intravenous antibiotics and topical debridement, the ulcer was rapidly increasing. In the laboratory findings, WBC was 12,400/µL, CRP was 16.9 mg/dL, ESR was 105mm in the first hour. Contrast enhanced CT images showed a wide high density area of skin and subcutaneous tissue around the stoma and dillitation of the transverse and descending colon. Colonoscopy showed furred profound ulcers in the rectum. A biopsy from the ulcer floor submitted to histopathology showed necrotic tissue with a mixed inflammatory infiltrates mainly composed of neutrophils and lymphocytes in the dermis. We suspected pyoderma gangrenosum with an inflammatory bowel disease in the beginning. Although he was started on oral prednisolone 60 mg daily, the ulcer did not respond to treatment. Additional methylprednisolone pulse therapy, intravenous cyclosporine and granulocytapheresis were also ineffective. A biopsy specimen from the skin ulcer margin showed erythrophagocytosis by trophozoites of amebae which were identified on PAS stained slides. The PCR method and stool examination showed positive for Entamoeba histolytica (E. histolytica), but serum antibodies were negative. Within two weeks of treatment with oral metronidazole 2,250 mg/day and topical metronidazole ointment, resolution of the ulcer was observed, then the prednisolone dosage was tapered. A split-thickness skin graft was used to cover the ulcer with a successful result. Even though we originally misdiagnosed this case, we finally reached a diagnosis of amebiasis. It is important to take account of amebiasis in the differential diagnosis of intractable ulcers which can be contaminated by feces.
Assuntos
Antibacterianos/uso terapêutico , Disenteria Amebiana/tratamento farmacológico , Entamoeba histolytica/isolamento & purificação , Úlcera/tratamento farmacológico , Idoso , Desbridamento/métodos , Combinação de Medicamentos , Disenteria Amebiana/complicações , Disenteria Amebiana/diagnóstico , Fezes/microbiologia , Humanos , Masculino , Úlcera/diagnóstico , Úlcera/etiologiaRESUMO
Polymerase chain reaction (PCR) technique is being increasingly used for the microbiological diagnosis of Pneumocystis pneumonia (PCP). As PCR is highly sensitive, it can be positive even in a patient with Pneumocystis colonization. In this study, we evaluated whether the ß-d-glucan assay could be used to differentiate between PCP and Pneumocystis jirovecii colonization in immunocompromised patients with pulmonary infiltrates. We retrospectively evaluated data from 166 consecutive patients who underwent bronchoalveolar lavage for the diagnosis of PCP. Serum levels of ß-d-glucan in the negative, colonization, probable PCP, and definite PCP groups were 20.2 ± 6.3, 48.8 ± 15.9, 89.9 ± 20.2, 224.9 ± 25.9 pg/mL, respectively. The ß-D-glucan levels in the definite PCP group were significantly higher than those in the other 3 groups (p < 0.001). Serum ß-d-glucan levels in patients with either definite or probable PCP (173.1 ± 18.8 pg/mL) were significantly greater than those in patients with colonization who had positive PCR results but improved without anti-PCP treatment (p < 0.002). The cut-off level for discrimination was estimated to be 33.5 pg/mL, with which the positive predictive value was 0.925. These results indicate that ß-D-glucan is a useful marker to differentiate between PCP and Pneumocystis colonization. A positive ß-D-glucan assay result might be a good indication to begin anti-PCP treatment.
Assuntos
Portador Sadio/diagnóstico , DNA Fúngico/análise , Pneumocystis carinii , Pneumonia por Pneumocystis/diagnóstico , beta-Glucanas/sangue , Idoso , Biomarcadores/sangue , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/microbiologia , Portador Sadio/sangue , Diagnóstico Diferencial , Feminino , Humanos , Hospedeiro Imunocomprometido , Masculino , Pessoa de Meia-Idade , Pneumocystis carinii/genética , Pneumonia por Pneumocystis/sangue , Valor Preditivo dos Testes , Proteoglicanas , Estudos RetrospectivosRESUMO
BACKGROUND: Rapid and accurate diagnosis of toxoplasmosis is critical, particularly for immunocompromised patients. Several molecular methods could have value for toxoplasmosis diagnosis, but often require sophisticated and expensive equipment, and as such are impractical for use in resource-limited countries. Our study aimed to develop a new rapid diagnostic test for toxoplasmosis that can be used in developed countries as well as low- or middle-income countries. METHODS: Common primers for conventional loop-mediated isothermal amplification (LAMP) and the new LAMP DNA chromatography method were designed based on a 529-bp repeat present in Toxoplasma gondii genomic DNA. A total of 91 clinical samples from 44 patients suspected of having toxoplasmosis who were treated at several hospitals across Japan were tested using the new LAMP DNA chromatography method, conventional LAMP, and nested PCR and the sensitivity and specificity of the methods was compared. RESULTS: The LAMP DNA chromatography method showed better sensitivity and specificity (68.2% and 100%, respectively) compared with the nested PCR (45.4% and 100%, respectively) and conventional LAMP (63.6% and 100%, respectively) methods for diagnosis of toxoplasmosis in immunocompromised patients. LAMP DNA chromatography also has better sensitivity and specificity (75% and 100%, respectively) than nested PCR (50.0% and 93.5%, respectively) and conventional LAMP (62.5% and 100%, respectively) to diagnose toxoplasma encephalitis using CSF samples. CONCLUSION: We developed a LAMP DNA chromatography method to detect T. gondii DNA in clinical samples. This method also successfully detected T. gondii DNA in CSF from patients with toxoplasma encephalitis. This newly developed method can be a valuable rapid diagnostic test for toxoplasmosis in a range of settings, including resource-limited areas like those in low- or middle-income countries.
RESUMO
Entamoeba species infect humans and non-human primates, raising concerns associated with potential zoonotic transmission. Therefore, the prevalence of human Entamoeba infections is crucial for its management in areas, where macaques exhibit high infection rates. Previously, we demonstrated prevalent E. nuttalli infections in rhesus macaques in Kathmandu, Nepal. In this study, we surveyed Entamoeba infection among 185 schoolchildren from two schools visited by wild rhesus macaques to assess the risk of transmission. PCR-based screening for Entamoeba species identified E. coli in 13 % and E. dispar in 0.5 % of the human stool samples. However, E. nuttalli and E. chattoni infections, prevalent in macaques, were not detected in human samples. This suggests that Entamoeba spp. are not transmitted through macaques in the school environment. We surveyed the rhesus macaques living in the temple near schools as well as the rhesus and Assam macaques inhabiting Shivapri Nagarjun National Park, Kathmandu. Among the 49 macaque stool samples, E. chattoni, E. coli, E. nuttalli, and E. dispar were detected in 92 %, 86 %, 41 %, and 18 % of the samples, respectively. Notably, E. dispar infections in macaques were mostly prevalent in the temple. A sample isolated from Nagarujun showed an identical genotype at two tRNA-linked short tandem repeat loci to that of E. dispar isolated from humans, suggesting potential transmission from humans to macaques. Genotypic analysis of cultured E. nuttalli strains obtained from the macaques colonizing three locations demonstrated that the geographical distance rather than differences in macaque species played a crucial role in the genetic diversity of the parasites. The phylogenetic tree of E. nuttalli strains, including the previously isolated strains, reflected the geographical distribution of the isolation sites. This study sheds light on the intricate dynamics of Entamoeba transmission and genetic diversity in macaques and humans.
RESUMO
BACKGROUND: Entamoeba moshkovskii is prevalent in developing countries and morphologically indistinguishable from pathogenic Entamoeba histolytica and nonpathogenic Entamoeba dispar. It is not known if E. moshkovskii is pathogenic. METHODS: Mice were intracecally challenged with the trophozoites of each Entamoeba spp. to test the ability to cause diarrhea, and infants in Bangladesh were prospectively observed to see if newly acquired E. moshkovskii infection was associated with diarrhea. RESULTS: E. moshkovskii and E. histolytica caused diarrhea and weight loss in susceptible mice. E. dispar infected none of the mouse strains tested. In Mirpur, Dhaka, Bangladesh, E. moshkovskii, E. histolytica, and E. dispar were identified in 42 (2.95%), 66 (4.63%), and 5 (0.35%), respectively, of 1426 diarrheal episodes in 385 children followed prospectively from birth to one year of age. Diarrhea occurred temporally with acquisition of a new E. moshkovskii infection: in the 2 months preceding E. moshkvskii-associated diarrhea, 86% (36 of 42) of monthly surveillance stool samples were negative for E. moshkovskii. CONCLUSIONS: E. moshkovskii was found to be pathogenic in mice. In children, the acquisition of E. moshkovskii infection was associated with diarrhea. These data are consistent with E. moshkovskii causing disease, indicating that it is important to reexamine its pathogenicity.
Assuntos
Diarreia/parasitologia , Entamoeba/isolamento & purificação , Entamebíase/parasitologia , Animais , Bangladesh/epidemiologia , Distribuição de Qui-Quadrado , DNA de Protozoário/química , DNA de Protozoário/genética , Diarreia/epidemiologia , Entamoeba/genética , Entamebíase/epidemiologia , Fezes/microbiologia , Humanos , Lactente , Recém-Nascido , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos CBA , Reação em Cadeia da Polimerase , Estudos Prospectivos , Organismos Livres de Patógenos EspecíficosRESUMO
In vitro excystation of cysts of microscopically identified Chilomastix mesnili and Retortamonas sp. isolated from Japanese macaques and Retortamonas sp. isolated from small Indian mongooses could be induced using an established protocol for Giardia intestinalis and subsequently by culturing with H2S-rich Robinson's medium supplemented with Desulfovibrio desulfuricans. Excystation usually began 2 h after incubation in Robinson's medium. DNA was isolated from excysted flagellates after 4 h of incubation or from cultured excysted flagellates. Phylogenetic analysis based on their 18S rRNA genes revealed that two isolates of C. mesnili from Japanese macaques belonged to the same cluster as a C. mesnili isolate from humans, whereas a mammalian Retortamonas sp. isolate from a small Indian mongoose belonged to the same cluster as that of an amphibian Retortamonas spp. isolate from a 'poison arrow frog' [sequence identity to AF439347 (94.9%)]. These results suggest that the sequence homology of the 18S rRNA gene of the two C. mesnili isolates from Japanese macaques was similar to that of humans, in addition to the morphological similarity, and Retortamonas sp. infection of the amphibian type in the small Indian mongoose highlighted the possibility of the effect of host feeding habitats.
Assuntos
Herpestidae , Parasitos , Retortamonadídeos , Humanos , Animais , Filogenia , Retortamonadídeos/genética , Herpestidae/genética , Macaca fuscata/genética , RNA Ribossômico 18S/genéticaRESUMO
BACKGROUND: Importance of the amphizoic amoeba Entamoeba moshkovskii is increasing in the study of amoebiasis as a common human pathogen in some settings. Limited studies are found on the genetic and phylogenetic characterization of E. moshkovskii from India; hence remain largely unknown. In this study, we determined the prevalence and characterized the E. moshkovskii isolates in eastern India. METHODS: A three-year systemic surveillance study among a total of 6051 diarrhoeal patients from ID Hospital and BC Roy Hospital, Kolkata was conducted for E. moshkovskii detection via a nested PCR system targeting 18S rRNA locus. The outer primer set detected the genus Entamoeba and the inner primer pair identified the E. moshkovskii species. The 18S rRNA locus of the positive samples was sequenced. Genetic and phylogenetic structures were determined using DnaSP.v5 and MEGA-X. GraphPad Prism (v.8.4.2), CA, USA was used to analyze the statistical data. RESULT: 4.84% (95%CI = 0.0433-0.0541) samples were positive for Entamoeba spp and 3.12% (95%CI = 0.027-0.036) were infected with E. moshkovskii. E. moshkovskii infection was significantly associated with age groups (X2 = 26.01, P<0.0001) but not with gender (Fisher's exact test = 0.2548, P<0.05). A unique seasonal pattern was found for E. moshkovskii infection. Additionally, 46.56% (95%CI = 0.396-0.537) were sole E. moshkovskii infections and significantly associated with diarrheal incidence (X2 = 335.5,df = 9; P<0.0001). Sequencing revealed that the local E. moshkovskii strains were 99.59%-100% identical to the prototype (GenBank: KP722605.1). The study found certain SNPs that showed a correlation with clinical features, but it is not necessarily indicative of direct control over pathogenicity. However, SNPs in the 18S rRNA gene could impact the biology of the amoeba and serve as a useful phylogenetic marker for identifying pathogenic E. moshkovskii isolates. Neutrality tests of different coinfected subgroups indicated deviations from neutrality and implied population expansion after a bottleneck event or a selective sweep and/or purifying selection in co-infected subgroups. The majority of FST values of different coinfected subgroups were <0.25, indicating low to moderate genetic differentiation within the subgroups of this geographical area. CONCLUSION: The findings reveal the epidemiological significance of E. moshkovskii infection in Eastern India as the first report in this geographical area and expose this species as a possible emerging enteric pathogen in India. Our findings provide useful knowledge for further research and the development of future control strategies against E. moshkovskii.
Assuntos
Amoeba , Coinfecção , Entamoeba histolytica , Entamoeba , Entamebíase , Humanos , Entamebíase/epidemiologia , Entamebíase/diagnóstico , RNA Ribossômico 18S/genética , Prevalência , Filogenia , Fezes , Diarreia/epidemiologia , Índia/epidemiologiaRESUMO
Objectives: Trichomonas vaginalis is the most prevalent sexually transmitted parasite worldwide. However, no surveillance system exists to monitor T. vaginalis cases and drug resistance in Japan. Methods: Cervical cytology vaginal swabs were collected from women with and without suspected symptoms of T. vaginalis infection; these swabs were used for the detection of T. vaginalis, human papillomavirus (HPV), and Candida albicans using specific polymerase chain reaction. Clinical isolates of T. vaginalis were subjected to metronidazole susceptibility tests using the previously reported minimal lethal concentration (MLC) and newly established half-maximal inhibitory concentration (IC50) values. Results: The prevalence of T. vaginalis in the study population was 4.2% (5/119; 95% confidence interval [Cl], 1.5-9.7). Additionally, asymptomatic infection constituted 60% (3/5) of all cases of T. vaginalis infection. All T. vaginalis-positive patients were coinfected with HPV but not C. albicans. Five clinical T. vaginalis isolates showed metronidazole susceptibility, which was evaluated using MLC values. The quantitative IC50 values revealed that two of these clinical isolates exhibited a decreased metronidazole susceptibility. Conclusion: This is the first study to demonstrate the prevalence of T. vaginalis in Japanese women. The IC50 values of metronidazole against T. vaginalis enabled the precise and quantitative evaluation of metronidazole-susceptible T. vaginalis.
RESUMO
Derivatives of 7-benzylidenenaltrexone (BNTX), which was recently reported to be an effective chloroquine (CQ)-resistance reverser, were synthesized and evaluated for their CQ-resistance reversing activities. The synthesized derivatives showed CQ-resistance reversing effects. They also reacted with glutathione (GSH) both enzymatically and chemically, and inhibited glutathione reductase activity. 7-Benzyl derivative, which was obtained by reduction of the olefin group in α,ß-unsaturated ketone structure of BNTX, also exhibited CQ-resistance reversing effect, but its potency was significantly lower than that of BNTX. These outcomes suggested that the decrease in GSH level could be one of the mechanisms of CQ-resistance reversing effects induced by BNTX derivatives.