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Understanding the functions of human transcriptional regulatory genes SP110 and SP140 during Mycobacterium tuberculosis infection is crucial; in a mouse model, homologous genes Sp110 and Sp140 have been shown to negatively regulate inflammatory response genes, including the type I interferon (IFN) response. The reduction of these genes in mice is associated with susceptibility to M. tuberculosis infection and the development of necrotizing granulomatous lesions. To investigate the involvement of SP110 and SP140 in human inflammatory response, we analyzed their regulatory manner in THP-1 macrophages infected with M. tuberculosis. Genome-wide transcriptional profiling revealed that the depletion of SP110 and/or SP140 impaired the induction of gene expression associated with inflammatory responses, including IFN response genes, although it had little effect on the intracellular proliferation of M. tuberculosis. By contrast, genes related to phosphorylation were upregulated in infected macrophages with SP110 and/or SP140 knockdown, but downregulated in infected control macrophages without their knockdown. Reverse transcription-quantitative PCR and ELISA further confirmed the impairment of the induction of IFN response genes by the depletion of SP110 and/or SP140 in M. tuberculosis-infected macrophages. These findings suggest that human SP110 and SP140 act as positive regulators for genes associated with inflammatory responses in M. tuberculosis-infected macrophages. IMPORTANCE: Tuberculosis (TB) is one of the most serious infectious diseases, with high morbidity and mortality worldwide. C3HeB/FeJ mice are widely utilized for evaluating anti-TB drugs because their drug sensitivity and pathology during M. tuberculosis infection resemble those of human TB, including the development of necrotizing granulomas. Downregulation of the transcriptional regulatory genes Sp110 and Sp140 in C3HeB/FeJ mice has been demonstrated to activate gene expression associated with inflammatory responses during M. tuberculosis infection, resulting in susceptibility to the infection. Here, we examined the regulatory manner of SP110 and SP140 using transcriptomic analysis in M. tuberculosis-infected human macrophages. Depletion of SP110 and/or SP140 in M. tuberculosis-infected THP-1 macrophages impaired the induction of gene expression associated with inflammatory responses, including interferon response genes, compared with that in control macrophages. These results suggest that human SP110 and SP140 act as positive regulators for genes associated with inflammatory responses upon M. tuberculosis infection.
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We report the case of a 42-year-old man with bronchiectasis who had a history of infertility treatment for obstructive azoospermia. Young's syndrome was suspected based on the triad of obstructive azoospermia, sinusitis, and bronchiectasis. He had normal electron microscopy findings, normal nasal nitric oxide levels (116 nL/min), and no situs inversus. However, we found compound heterozygous variants in CFAP221. This led to a diagnosis of primary ciliary dyskinesia (PCD). Distinguishing PCD from Young's syndrome in patients with the triad of obstructive azoospermia, sinusitis, and bronchiectasis is challenging. Young's syndrome may be a phenotype of PCD.
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OBJECTIVE: Primary ciliary dyskinesia (PCD) is a relatively rare genetic disorder that affects approximately 1 in 20,000 people. Approximately 50 genes are currently known to cause PCD. In light of differences in causative genes and the medical system in Japan compared with other countries, a practical guide was needed for the diagnosis and management of Japanese PCD patients. METHODS: An ad hoc academic committee was organized under the Japanese Rhinologic Society to produce a practical guide, with participation by committee members from several academic societies in Japan. The practical guide including diagnostic criteria for PCD was approved by the Japanese Rhinologic Society, Japanese Society of Otolaryngology-Head and Neck Surgery, Japanese Respiratory Society, and Japanese Society of Pediatric Pulmonology. RESULTS: The diagnostic criteria for PCD consist of six clinical features, six laboratory findings, differential diagnosis, and genetic testing. The diagnosis of PCD is categorized as definite, probable, or possible PCD based on a combination of the four items above. Diagnosis of definite PCD requires exclusion of cystic fibrosis and primary immunodeficiency, at least one of the six clinical features, and a positive result for at least one of the following: (1) Class 1 defect on electron microscopy of cilia, (2) pathogenic or likely pathogenic variants in a PCD-related gene, or (3) impairment of ciliary motility that can be repaired by correcting the causative gene variants in iPS cells established from the patient's peripheral blood cells. CONCLUSION: This practical guide provides clinicians with useful information for the diagnosis and management of PCD in Japan.
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Pruebas Genéticas , Síndrome de Kartagener , Humanos , Síndrome de Kartagener/diagnóstico , Síndrome de Kartagener/terapia , Síndrome de Kartagener/genética , Diagnóstico Diferencial , Cilios/ultraestructura , Cilios/patología , Japón , Dineínas Axonemales/genética , ProteínasRESUMEN
We present the case of a 58-year-old female patient with primary ciliary dyskinesia (PCD). She was born to parents with a consanguineous marriage. Chest computed tomography conducted at age 41 years indicated no situs inversus, and findings of bronchiectasis were limited to the middle and lingular lobes. Despite long-term macrolide therapy, bronchiectasis exacerbations frequently occurred. PCD was suspected because of the low nasal nitric oxide level (20.7 nL/min). Electron microscopy revealed outer and inner dynein arm defects, and a genetic analysis identified a homozygous single-nucleotide deletion in the DNAAF1 gene. Based on these results, the patient was diagnosed with PCD due to a biallelic DNAAF1 mutation.
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Primary ciliary dyskinesia (PCD) is a rare genetic disorder characterized by impaired motile cilia function, particularly in the upper and lower airways. To date, more than 50 causative genes related to the movement, development, and maintenance of cilia have been identified. PCD mostly follows an autosomal recessive inheritance pattern, in which PCD symptoms manifest only in the presence of pathogenic variants in both alleles. Several genes causing PCD have been recently identified that neither lead to situs inversus nor cause definitive abnormalities in ciliary ultrastructure. Importantly, the distribution of disease-causing genes and pathogenic variants varies depending on ethnicity. In Japan, homozygosity for a â¼27.7-kb deletion of DRC1 is estimated to be the most common cause of PCD, presumably as a founder mutation. The clinical picture of PCD is similar to that of sinobronchial syndrome, thus making its differentiation from diffuse panbronchiolitis and other related disorders difficult. Given the diagnostic challenges, many cases remain undiagnosed or misdiagnosed, particularly in adults. While no fundamental cure is currently available, lifelong medical subsidies are provided in Japan, and proper respiratory management, along with continued prevention and treatment of infections, is believed to mitigate the decline in respiratory function. Timely action will be necessary when specific treatments for PCD become available in the future. This narrative review focuses on variations in the disease status of PCD in a non-Western country.
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Trastornos de la Motilidad Ciliar , Adulto , Humanos , Japón/epidemiología , Trastornos de la Motilidad Ciliar/genética , MutaciónRESUMEN
Primary ciliary dyskinesia (PCD) is a genetic disease characterized by motile cilia dysfunction, mostly inherited in an autosomal recessive or X-linked manner. We herein report a 29-year-old woman with PCD caused by a heterozygous frameshift mutation due to a single nucleotide deletion in exon 3 of FOXJ1. Heterozygous de novo mutations in FOXJ1 have been reported as an autosomal-dominant cause of PCD. The patient had situs inversus, congenital heart disease, infertility, and hydrocephalus. However, the nasal nitric oxide level was normal. Long-term macrolide therapy was remarkably effective. This is the first case report of PCD caused by a FOXJ1 variant in Japan.
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Introduction: It is assumed that host defense systems eliminating the pathogen and regulating tissue damage make a strong impact on the outcome of tuberculosis (TB) disease and that these processes are affected by rifampicin (RIF) resistance-conferring mutations of Mycobacterium tuberculosis (Mtb). However, the host responses to the pathogen harboring different mutations have not been studied comprehensively in clinical settings. We analyzed clinico-epidemiological factors and blood transcriptomic signatures associated with major rpoB mutations conferring RIF resistance in a cohort study. Methods: Demographic data were collected from 295 active pulmonary TB patients with treatment history in Hanoi, Vietnam. When recruited, drug resistance-conferring mutations and lineage-specific variations were identified using whole-genome sequencing of clinical Mtb isolates. Before starting retreatment, total RNA was extracted from the whole blood of HIV-negative patients infected with Mtb that carried either the rpoB H445Y or rpoB S450L mutation, and the total RNA was subjected to RNA sequencing after age-gender matching. The individual RNA expression levels in the blood sample set were also measured using real-time RT-PCR. Logistic and linear regression models were used to assess possible associations. Results: In our cohort, rpoB S450L and rpoB H445Y were major RIF resistance-conferring mutations [32/87 (36.8%) and 15/87 (17.2%), respectively]. H445Y was enriched in the ancient Beijing genotype and was associated with nonsynonymous mutations of Rv1830 that has been reported to regulate antibiotic resilience. H445Y was also more frequently observed in genetically clustered strains and in samples from patients who had received more than one TB treatment episode. According to the RNA sequencing, gene sets involved in the interferon-γ and-α pathways were downregulated in H445Y compared with S450L. The qRT-PCR analysis also confirmed the low expression levels of interferon-inducible genes, including BATF2 and SERPING1, in the H445Y group, particularly in patients with extensive lesions on chest X-ray. Discussion: Our study results showed that rpoB mutations as well as Mtb sublineage with additional genetic variants may have significant effects on host response. These findings strengthen the rationale for investigation of host-pathogen interactions to develop countermeasures against epidemics of drug-resistant TB.
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Síndrome de Kartagener , Humanos , Síndrome de Kartagener/genética , Expresión Génica , CiliosRESUMEN
MAFB, v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog B, has been identified as a candidate gene for early tuberculosis (TB) onset in Thai and Japanese populations. Here, we investigated the genome-wide transcriptional profiles of MAFB-knockdown (KD) macrophages infected with Mycobacterium tuberculosis (Mtb) to highlight the potential role of MAFB in host immunity against TB. Gene expression analysis revealed impaired type I and type II interferon (IFN) responses and enhanced oxidative phosphorylation in MAFB-KD macrophages infected with Mtb. The expression of inflammatory chemokines, including IFN-γ-inducible genes, was confirmed to be significantly reduced by knockdown of MAFB during Mtb infection. A similar effect of MAFB knockdown on type I and type II IFN responses and oxidative phosphorylation was also observed when Mtb-infected macrophages were activated by IFN-γ. Taken together, our results demonstrate that MAFB is involved in the immune response and metabolism in Mtb-infected macrophages, providing new insight into MAFB as a candidate gene to guide further study to control TB.
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Infection with Mycobacterium tuberculosis leads to the development of tuberculosis (TB) with the formation of granulomatous lesions. Foamy macrophages (FM) are a hallmark of TB granulomas, because they provide the primary platform of M. tuberculosis proliferation and the main source of caseous necrosis. In this study, we applied spatial multiomic profiling to identify the signatures of FM within the necrotic granulomas developed in a mouse model resembling human TB histopathology. C3HeB/FeJ mice were infected with M. tuberculosis to induce the formation of necrotic granulomas in the lungs. Using laser microdissection, necrotic granulomas were fractionated into three distinct regions, including the central caseous necrosis, the rim containing FM, and the peripheral layer of macrophages and lymphocytes, and subjected to proteomic and transcriptomic analyses. Comparison of proteomic and transcriptomic analyses of three distinct granulomatous regions revealed that four proteins/genes are commonly enriched in the rim region. Immunohistochemistry confirmed the localization of identified signatures to the rim of necrotic granulomas. We also investigated the localization of the representative markers for M1 macrophages in granulomas because the signatures of the rim included M2 macrophage markers. The localization of both macrophage markers suggests that FM in necrotic granulomas possessed the features of M1 or M2 macrophages. Gene set enrichment analysis of transcriptomic profiling revealed the upregulation of genes related to M2 macrophage activation and mTORC1 signaling in the rim. These results will provide new insights into the process of FM biogenesis, leading to further understanding of the pathophysiology of TB granulomas.
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Mycobacterium tuberculosis , Tuberculosis Ganglionar , Animales , Granuloma/microbiología , Humanos , Pulmón/microbiología , Macrófagos/microbiología , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Endogámicos , Mycobacterium tuberculosis/genética , Necrosis , ProteómicaRESUMEN
The establishment of animal models reflecting human Mycobacterium avium complex (MAC) lung disease (LD) pathology has the potential to expand our understanding of the disease pathophysiology. However, inducing sustained infection in immunocompetent mice is difficult since MAC generally shows less virulence and higher genetic variability than M. tuberculosis. To overcome this hurdle, we developed a screening system for identifying virulent MAC strains using whole-genome sequencing (WGS). We obtained nine clinical strains from Mycobacterium avium complex lung disease (MAC-LD) patients and divided them into two groups to make the mixed strain inocula for infection. Intranasal infection with the strain mixture of both groups in BALB/c mice resulted in progressive infection and extensive granuloma formation in the lungs, suggesting the existence of highly pathogenic strains in each group. We hypothesized that the change in the abundance of strain-specific single-nucleotide variants (SNVs) reflects the change in bacterial number of each strain in infected lungs. Based on this hypothesis, we quantified individual strain-specific SNVs in bacterial DNA from infected lungs. Specific SNVs for four strains were detected, suggesting the pathogenicity of these four strains. Consistent with these results, individual infection with these four strains induced a high lung bacterial burden, forming extensive peribronchial granuloma, while the other strains showed a decreased lung bacterial burden. The current method combining mixed infection and WGS accurately identified virulent strains that induced sustained infection in mice. This method will contribute to the establishment of mouse models that reflect human MAC-LD and lead to antimycobacterial drug testing. IMPORTANCE To promote research on Mycobacterium avium complex (MAC) pathogenicity, animal models reflecting human progressive MAC lung disease (MAC-LD) are needed. Because there is high genetic and virulence diversity among clinical MAC strains, choosing a suitable strain is an important process for developing a mouse model. In this study, we developed a screening system for virulent strains in mice by combining mixed infection and whole-genome sequencing analysis. This approach is designed on the hypothesis that in vivo virulence of MAC strains can be examined simultaneously by comparing changes in the abundance of strain-specific single-nucleotide variants in the mouse lungs after infection with mixed strains. The identified strains were shown to induce high bacterial burdens and cause extensive peribronchial granuloma resembling the pulmonary pathology of human MAC-LD. The current method will help researchers develop mouse models that reflect human MAC-LD and will lead to further investigation of MAC pathogenicity.
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Coinfección , Enfermedades Pulmonares , Infección por Mycobacterium avium-intracellulare , Mycobacterium tuberculosis , Animales , Enfermedades Pulmonares/microbiología , Ratones , Complejo Mycobacterium avium/genética , Infección por Mycobacterium avium-intracellulare/microbiología , Mycobacterium tuberculosis/genética , NucleótidosRESUMEN
In mid-September 2019, a teenage Chinese male student and part-time waiter in Tokyo was diagnosed with multidrug-resistant (MDR) sputum smear-positive pulmonary tuberculosis (TB). This study describes the outbreak investigation of his friends and colleagues at the restaurant. We investigated 6 friends and 15 colleagues; 5 friends and 13 colleagues underwent interferon-γ release assay (IGRA). Of these, 3 friends (60.0%) and 4 colleagues (30.8%) were IGRA-positive. Each of the friends and colleagues was found to have MDR-TB (20% and 7.7%, respectively). Challenges during the investigation were the unavailability of regimens for latent TB infection (LTBI) for contacts with MDR-TB, budgetary constraints concerning implementing computed tomography (CT) scans for the contacts, frequent address changes of foreign-born patients and contacts, investigation during the coronavirus disease pandemic, and variations of alphabetical expression of the names of the patients and contacts, particularly for those from China. It is recommended that the national government officially adopt prophylaxis regimens for LTBI with MDR-TB, address the budgetary constraints regarding CT scans, and deploy liaison officers for coordinating investigations involving many foreign-born patients and contacts scattered in multiple municipalities. The names of foreign-born persons could more accurately be identified using both the alphabet and Chinese characters.
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Emigrantes e Inmigrantes , Tuberculosis Latente , Tuberculosis Resistente a Múltiples Medicamentos , Adolescente , Brotes de Enfermedades , Humanos , Japón/epidemiología , Tuberculosis Latente/tratamiento farmacológico , Masculino , Tokio/epidemiología , Tuberculosis Resistente a Múltiples Medicamentos/tratamiento farmacológico , Tuberculosis Resistente a Múltiples Medicamentos/epidemiologíaRESUMEN
Mycobacterium tuberculosis (Mtb) has different features depending on different geographic areas. We collected Mtb strains from patients with smear-positive pulmonary tuberculosis in Da Nang, central Vietnam. Using a whole genome sequencing platform, including genome assembly complemented by long-read-sequencing data, genomic characteristics were studied. Of 181 Mtb isolates, predominant Vietnamese EAI4_VNM and EAI4-like spoligotypes (31.5%), ZERO strains (5.0%), and part of EAI5 (11.1%) were included in a lineage-1 (L1) sublineage, i.e., L1.1.1.1. These strains were found less often in younger people, and they genetically clustered less frequently than other modern strains. Patients infected with ZERO strains demonstrated less lung infiltration. A region in RD2bcg spanning six loci, i.e., PE_PGRS35, cfp21, Rv1985c, Rv1986, Rv1987, and erm(37), was deleted in EAI4_VNM, EAI4-like, and ZERO strains, whereas another 118 bp deletion in furA was specific only to ZERO strains. L1.1.1.1-sublineage-specific deletions in PE_PGRS4 and PE_PGRS22 were also identified. RD900, seen in ancestral lineages, was present in majority of the L1 members. All strains without IS6110 (5.0%) had the ZERO spoligo-pattern. Distinctive features of the ancestral L1 strains provide a basis for investigation of the modern versus ancestral Mtb lineages and allow consideration of countermeasures against this heterogeneous pathogen.
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ADN Bacteriano , Variación Genética , Mycobacterium tuberculosis/genética , Filogenia , Tuberculosis Pulmonar/genética , Adulto , Femenino , Humanos , Masculino , Mycobacterium tuberculosis/aislamiento & purificación , Tuberculosis Pulmonar/epidemiología , Vietnam/epidemiologíaRESUMEN
Granule-associated killing molecules released from cytotoxic T lymphocytes participate as a crucial step in immunity against tuberculosis (TB), but the role of coordinated production remains controversial. Coordinated release of effector molecules in vitro after stimulating peripheral blood mononuclear cells (PBMCs) of active TB or HIV/TB coinfection patients with PPD, purified protein derivative of tuberculin and avirulent Mtb, H37Ra, an attenuated strain were investigated in association with clinical outcomes. Perforin, granzyme-B, granulysin and IFN-γ were measured using ELISA. Before anti-TB treatment, PBMCs of TB stimulated with PPD or H37Ra released higher perforin, granzyme-B, and granulysin levels than in HIV/TB and released significantly higher IFN-γ (p = 0.045, p = 0.022). Granulysin positively correlated with perforin in TB (p = 0.042, r = 0.385), HIV/TB coinfection (p = 0.003, r = 0.941) after PPD stimulation, and after H37Ra stimulation in TB (p = 0.005, r = 0.549), but negatively correlated with granzyme B in TB (p = 0.042, r = -0.386), HIV/TB coinfection (p = 0.042, r = 0.754) were noted. After anti-TB treatment, increased levels of perforin, granulysin and IFN-γ in TB or HIV/TB upon PPD or H37Ra stimulation, and decreased granzyme-B levels after PPD (p = 0.003) or H37Ra (p = 0.028) stimulation in TB were observed. These results suggest that granulysin may act synergistic with perforin and IFN-γ in TB, indicating its crucial function in host immunity to tuberculosis. Future studies with larger numbers of patients ought to be conducted in the future.
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The number of studies which evaluated interferon-gamma release assays (IGRAs) results after anti-tuberculosis (TB) treatment has been rapidly increasing. The aim of this study was to investigate the potential use of IGRAs (QFT-GIT, T-SPOT.TB, QFT-Plus) in assessing the response to anti-TB treatment. We searched all studies in English language published from 1 October 2011 to 18 November 2018 in PubMed, Web of Science, and Scopus. Our search included the term "tuberculosis treatment AND interferon-γ release assay". We included studies evaluating the performance of commercial IGRAs (including QFT-GIT, T-SPOT.TB and QFT-Plus) before and after the anti-TB treatment. We performed subgroup analysis based on the age (children, adults), type of TB (active, latent, active and latent, and contacts exposed to MDR defined as MDR LTBI), type of IGRAs (QFT-GIT and T-SPOT.TB), and follow-up interval (2, 3, 4, 6, 9 months). Of the 18 included studies, 12 used QFT-GIT for assessment of IGRA performance after therapy, 1 used T-SPOT.TB, and 3 used both QFT-GIT and T-SPOT.TB. Since then, only two studies have assessed the QFT-Plus performance during therapy. According to the results of the meta-analysis, the pooled rate of positive IGRAs (QFT-GIT and T-SPOT.TB) following anti-TB therapy was estimated at 76% [95% CI 70-81%] and no difference was found compared to the pooled positive rate of IGRAs before initiation of therapy which was 76% [95% CI 60-89%]. The subgroup analysis showed that the pooled rate of positive IGRAs (QFT-GIT and T-SPOT.TB) after anti-TB therapy was significantly higher in monitoring active TB subjects [80% (95% CI 74-88%)] than LTBI [71% (95% CI 70-81%)]. Available data are now sufficient to suggest that monitoring changes in the IGRAs (QFT-GIT and T-SPOT.TB) response during anti-TB treatment may have limited use in evaluating the effectiveness of treatment, while the monitoring changes in QFT-Plus during anti-tubercular treatment are recommended to determine treatment efficacy or for treatment monitoring. Further research is needed to establish the efficacy of this new assay as marker on a larger scale for treatment monitoring.
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Antituberculosos/uso terapéutico , Ensayos de Liberación de Interferón gamma , Tuberculosis/tratamiento farmacológico , Biomarcadores/metabolismo , Humanos , Interferón gamma/metabolismo , Ensayos de Liberación de Interferón gamma/métodos , Mycobacterium tuberculosis/efectos de los fármacos , Mycobacterium tuberculosis/inmunología , Juego de Reactivos para Diagnóstico , Resultado del Tratamiento , Tuberculosis/inmunologíaRESUMEN
BACKGROUND: Diffuse panbronchiolitis (DPB) is a sinopulmonary disease mainly affecting Asian populations. Primary ciliary dyskinesia (PCD) is a genetically heterogeneous disorder impairing ciliary structure and function. These two disorders are not easily distinguished by clinical signs and symptoms. METHODS: In 105 Japanese patients with recurrent sinopulmonary infection, initially diagnosed with DPB, and 37 patients with recurrent airway infection diagnosed in adulthood, the deletion allele of DRC1 or CCDC164, recently recognized as a pathogenic PCD gene variant, was searched using a multiplexed PCR-based method, and the deletion breakpoints and other variants around the gene were determined by Sanger sequencing and targeted resequencing. RESULTS: A large homozygous deletion in DRC1 was identified in three of the 142 patients. Furthermore, heterozygous carriers of the deletion with the same breakpoint were found with the allele frequency of 0.002 in the healthy Japanese population, indicating that this loss-of-function variant may be acting as a common mutation causing PCD in Japanese. CONCLUSION: PCD caused by the DRC1 defect is not readily identified by either high-speed video-microscopy or ciliary ultrastructure analysis, posing significant difficulties in reaching a correct diagnosis without the aid of genetic tests. Careful investigation of the causes of sinopulmonary diseases is warranted in Asian populations.
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Bronquiolitis/diagnóstico , Trastornos de la Motilidad Ciliar/diagnóstico , Eliminación de Gen , Pruebas Genéticas/métodos , Infecciones por Haemophilus/diagnóstico , Proteínas Asociadas a Microtúbulos/genética , Adulto , Cilios/ultraestructura , Trastornos de la Motilidad Ciliar/genética , Trastornos de la Motilidad Ciliar/microbiología , Diagnóstico Diferencial , Exones , Femenino , Homocigoto , Humanos , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa Multiplex/métodosRESUMEN
The mucosal glycocalyx of the ocular surface constitutes the point of interaction between the tear film and the apical epithelial cells. Membrane-associated mucins (MAMs) are the defining molecules of the glycocalyx in all mucosal epithelia. Long recognized for their biophysical properties of hydration, lubrication, anti-adhesion and repulsion, MAMs maintain the wet ocular surface, lubricate the blink, stabilize the tear film and create a physical barrier to the outside world. However, it is increasingly appreciated that MAMs also function as cell surface receptors that transduce information from the outside to the inside of the cell. A number of excellent review articles have provided perspective on the field as it has progressed since 1987, when molecular cloning of the first MAM was reported. The current article provides an update for the ocular surface, placing it into the broad context of findings made in other organ systems, and including new genes, new protein functions and new biological roles. We discuss the epithelial tissue-equivalent with mucosal differentiation, the key model system making these advances possible. In addition, we make the first systematic comparison of MAMs in human and mouse, establishing the basis for using knockout mice for investigations with the complexity of an in vivo system. Lastly, we discuss findings from human genetics/genomics, which are providing clues to new MAM roles previously unimagined. Taken together, this information allows us to generate hypotheses for the next stage of investigation to expand our knowledge of MAM function in intracellular signaling and roles unique to the ocular surface.
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Conjuntiva/metabolismo , Proteínas de la Membrana/genética , Mucinas/genética , Lágrimas/metabolismo , Animales , Células Epiteliales/metabolismo , Humanos , Proteínas de la Membrana/metabolismo , Ratones , Mucinas/metabolismoRESUMEN
Hanoi is the capital of Vietnam, one of the 30 countries with a high tuberculosis (TB) burden. Fundamental data on the molecular epidemiology of the disease is required for future TB management. To identify lineages and genotypes of Mycobacterium tuberculosis (Mtb), conventional genotyping data from clinical isolates of the Hanoi area was compared with whole genome sequencing (WGS) analysis from 332 of 470 samples. It was obtained from lineage-specific single nucleotide variants (SNVs), large sequence polymorphisms, spoligotyping, and variable number of tandem repeats (VNTR) analysis using mycobacterial interspersed repetitive unit (MIRU) and Japan anti-tuberculosis association (JATA) locus sets. This information was directly compared with results obtained from WGS. Mini-satellite repeat unit variants were identified using BLAST search against concatenated short read sequences, the RepUnitTyping tool. WGS analysis revealed that the Mtb strains tested are diverse and classified into lineage (L) 1, 2 and 4 (24.7, 57.2 and 18.1% respectively). The majority of the L2 strains were further divided into ancient and modern Beijing genotypes, and most of the L1 group were EAI4_VNM strains. Although conventional PCR-based genotyping results were mostly consistent with information obtained through WGS analysis, in-depth analysis identified aberrant deletions and spacers that may cause discordance. JATA-VNTR sets, including hypervariable loci, separated large Beijing genotypic clusters generated by MIRU15 into smaller groups. The distribution of repeat unit variants observed within 33 VNTR loci showed clear variation depending on the three lineages. WGS-based pairwise-SNV differences within VNTR-defined genotypic clusters were greater in L1 than in L2 and L4 (Pâ¯=â¯.001). Direct comparisons between results of PCR-based genotyping and in silico analysis of WGS data would bridge a gap between classical and modern technologies during this transition period, and provide further information on Mtb genotypes in specific geographical areas.
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Mycobacterium tuberculosis/genética , Tuberculosis/microbiología , Genotipo , Humanos , Repeticiones de Minisatélite , Filogenia , Reacción en Cadena de la Polimerasa , Vietnam , Secuenciación Completa del GenomaRESUMEN
Drug-resistant tuberculosis (TB) is a serious global problem, and pathogen factors involved in the transmission of isoniazid (INH)-resistant TB have not been fully investigated. We performed whole genome sequencing of 332 clinical Mycobacterium tuberculosis (Mtb) isolates collected from patients newly diagnosed with smear-positive pulmonary TB in Hanoi, Vietnam. Using a bacterial genome-wide approach based on linear mixed models, we investigated the associations between 31-bp k-mers and clustered strains harboring katG-S315T, a major INH-resistance mutation in the present cohort and in the second panel previously published in South Africa. Five statistically significant genes, namely, PPE18/19, gid, emrB, Rv1588c, and pncA, were shared by the two panels. We further identified variants of the genes responsible for these k-mers, which are relevant to the spread of INH-resistant strains. Phylogenetic convergence test showed that variants relevant to PPE46/47-like chimeric genes were significantly associated with the same phenotype in Hanoi. The associations were further confirmed after adjustment for the confounders. These findings suggest that genomic variations of the pathogen facilitate the expansion of INH-resistance TB, at least in part, and our study provides a new insight into the mechanisms by which drug-resistant Mtb maintains fitness and spreads in Asia and Africa.