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1.
Medicine (Baltimore) ; 100(21): e26091, 2021 May 28.
Artículo en Inglés | MEDLINE | ID: mdl-34032746

RESUMEN

INTRODUCTION: This work reports a patient with recurrent renal calculi subjected to three surgeries in half a year to be in the same position, and the high-throughput sequencing data showed different species in the renal pus and urine samples, which suggested that partial renal infection or stone formation can be judged by the bacteria in urine. PATIENT CONCERNS: The female patient aged 43 years was referred to the authors' department on April 13, 2020, due to left waist pain and fever for 3 days. DIAGNOSIS: Kidney stones and hydronephrosis were determined by a urinary system computed tomography scan. INTERVENTIONS: On April 20, 2020 and June 15, 2020, the patient was successfully treated with left percutaneous nephrolithotomy twice under general anesthesia. An investigation on the health and eating habits of the patient within 6 months was completed at the last admission. The components of the second renal calculus sample were analyzed with an infrared spectrum analyzer. The third renal stone (renal pus, triplicates) was subjected to microbial metagenome sequencing, and urine samples before and after surgery were subjected to 16S RNA sequencing by SEQHEALTH (Wuhan, China). OUTCOMES: After percutaneous nephrolithotomy, the left kidney stones were basically cleared, stone analysis revealed that the main components were calcium oxalate monohydrate, silica, and a small amount of calcium oxalate dehydrate. Although the urine samples exhibited differences, the renal pus and urine sample shared a single species. CONCLUSION: It is not clear that the prospects of partial renal infection or stone formation can be judged by the bacteria in urine.


Asunto(s)
Infecciones por Helicobacter/diagnóstico , Hidronefrosis/diagnóstico , Cálculos Renales/diagnóstico , Nefrolitotomía Percutánea/efectos adversos , Infecciones Urinarias/diagnóstico , Adulto , ADN Bacteriano/aislamiento & purificación , Femenino , Infecciones por Helicobacter/complicaciones , Infecciones por Helicobacter/microbiología , Infecciones por Helicobacter/cirugía , Helicobacter pylori , Humanos , Hidronefrosis/microbiología , Hidronefrosis/cirugía , Cálculos Renales/microbiología , Cálculos Renales/cirugía , Metagenoma/genética , ARN Ribosómico 16S/genética , Recurrencia , Reoperación , Infecciones Urinarias/complicaciones , Infecciones Urinarias/microbiología , Infecciones Urinarias/cirugía
2.
Nat Commun ; 12(1): 3186, 2021 05 27.
Artículo en Inglés | MEDLINE | ID: mdl-34045458

RESUMEN

Long-term infection of the airways of cystic fibrosis patients with Pseudomonas aeruginosa is often accompanied by a reduction in bacterial growth rate. This reduction has been hypothesised to increase within-patient fitness and overall persistence of the pathogen. Here, we apply adaptive laboratory evolution to revert the slow growth phenotype of P. aeruginosa clinical strains back to a high growth rate. We identify several evolutionary trajectories and mechanisms leading to fast growth caused by transcriptional and mutational changes, which depend on the stage of adaptation of the strain. Return to high growth rate increases antibiotic susceptibility, which is only partially dependent on reversion of mutations or changes in the transcriptional profile of genes known to be linked to antibiotic resistance. We propose that similar mechanisms and evolutionary trajectories, in reverse direction, may be involved in pathogen adaptation and the establishment of chronic infections in the antibiotic-treated airways of cystic fibrosis patients.


Asunto(s)
Antibacterianos/farmacología , Fibrosis Quística/complicaciones , Farmacorresistencia Microbiana/genética , Evolución Molecular , Infecciones por Pseudomonas/tratamiento farmacológico , Pseudomonas aeruginosa/genética , Adaptación Fisiológica/efectos de los fármacos , Adaptación Fisiológica/genética , Antibacterianos/uso terapéutico , Proteínas Bacterianas/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Fibrosis Quística/tratamiento farmacológico , Fibrosis Quística/inmunología , Fibrosis Quística/microbiología , Análisis Mutacional de ADN , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Evolución Molecular Dirigida , Farmacorresistencia Microbiana/efectos de los fármacos , Regulación Bacteriana de la Expresión Génica , Aptitud Genética/efectos de los fármacos , Genoma Bacteriano , Humanos , Pulmón/inmunología , Pulmón/microbiología , Pruebas de Sensibilidad Microbiana , Mutación , Fenotipo , Infecciones por Pseudomonas/inmunología , Infecciones por Pseudomonas/microbiología , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/aislamiento & purificación , Esputo/microbiología
3.
Int J Mol Sci ; 22(9)2021 Apr 29.
Artículo en Inglés | MEDLINE | ID: mdl-33946648

RESUMEN

Growing evidence highlights the crucial role of gut microbiota in affecting different aspects of obesity. Considering the ability of deep transcranial magnetic stimulation (dTMS) to modulate the cortical excitability, the reward system, and, indirectly, the autonomic nervous system (ANS), we hypothesized a potential role of dTMS in affecting the brain-gut communication pathways, and the gut microbiota composition in obesity. In a hospital setting, 22 subjects with obesity (5 M, 17 F; 44.9 ± 2.2 years; BMI 37.5 ± 1.0 kg/m2) were randomized into three groups receiving 15 sessions (3 per week for 5 weeks) of high frequency (HF), low frequency (LF) dTMS, or sham stimulation. Fecal samples were collected at baseline and after 5 weeks of treatment. Total bacterial DNA was extracted from fecal samples using the QIAamp DNA Stool Mini Kit (Qiagen, Italy) and analyzed by a metagenomics approach (Ion Torrent Personal Genome Machine). After 5 weeks, a significant weight loss was found in HF (HF: -4.1 ± 0.8%, LF: -1.9 ± 0.8%, sham: -1.3 ± 0.6%, p = 0.042) compared to LF and sham groups, associated with a decrease in norepinephrine compared to baseline (HF: -61.5 ± 15.2%, p < 0.01; LF: -31.8 ± 17.1%, p < 0.05; sham: -35.8 ± 21.0%, p > 0.05). Furthermore, an increase in Faecalibacterium (+154.3% vs. baseline, p < 0.05) and Alistipes (+153.4% vs. baseline, p < 0.05) genera, and a significant decrease in Lactobacillus (-77.1% vs. baseline, p < 0.05) were found in HF. Faecalibacterium variations were not significant compared to baseline in the other two groups (LF: +106.6%, sham: +27.6%; p > 0.05) as well as Alistipes (LF: -54.9%, sham: -15.1%; p > 0.05) and Lactobacillus (LF: -26.0%, sham: +228.3%; p > 0.05) variations. Norepinephrine change significantly correlated with Bacteroides (r2 = 0.734; p < 0.05), Eubacterium (r2 = 0.734; p < 0.05), and Parasutterella (r2 = 0.618; p < 0.05) abundance variations in HF. In conclusion, HF dTMS treatment revealed to be effective in modulating gut microbiota composition in subjects with obesity, reversing obesity-associated microbiota variations, and promoting bacterial species representative of healthy subjects with anti-inflammatory properties.


Asunto(s)
Microbioma Gastrointestinal , Obesidad/microbiología , Obesidad/terapia , Estimulación Magnética Transcraneal/métodos , Adulto , Anciano , Sistema Nervioso Autónomo/fisiopatología , Bacterias/clasificación , Bacterias/genética , Bacterias/aislamiento & purificación , Biodiversidad , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Método Doble Ciego , Heces/microbiología , Femenino , Microbioma Gastrointestinal/genética , Microbioma Gastrointestinal/fisiología , Humanos , Masculino , Persona de Mediana Edad , Modelos Biológicos , Norepinefrina/sangre , Obesidad/fisiopatología , ARN Ribosómico 16S/genética , Factores de Tiempo , Pérdida de Peso , Adulto Joven
4.
BMC Infect Dis ; 21(1): 463, 2021 May 21.
Artículo en Inglés | MEDLINE | ID: mdl-34020607

RESUMEN

BACKGROUND: Streptococcus pyogenes causes a profound global burden of morbidity and mortality across its diverse clinical spectrum. To support a new controlled human infection ('challenge') model seeking to accelerate S. pyogenes vaccine development, we aimed to develop an accurate and reliable molecular method for quantifying bacterial load from pharyngeal swabs collected during experimental human pharyngitis. METHODS: Combined sequential RNA + DNA extraction from throat swabs was compared to traditional separate RNA-only and DNA-only extractions. An emm-type specific qPCR was developed to detect the emm75 challenge strain. Results from the qPCR were compared to culture, using throat swab samples collected in a human challenge study. RESULTS: The qPCR was 100% specific for the emm75 challenge strain when tested against a panel of S. pyogenes emm-types and other respiratory pathogens. Combined RNA + DNA extraction had similar yield to traditional separate extractions. The combined extraction method and emm75 qPCR had 98.8% sensitivity compared to culture for throat swabs collected from challenge study participants. CONCLUSIONS: We have developed a reliable molecular method for measuring S. pyogenes bacterial load from throat swabs collected in a controlled human infection model of S. pyogenes pharyngitis. TRIAL REGISTRATION: NCT03361163 on 4th December 2017.


Asunto(s)
Antígenos Bacterianos/genética , Carga Bacteriana , Proteínas de la Membrana Bacteriana Externa/genética , Faringitis/microbiología , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Infecciones Estreptocócicas/microbiología , Streptococcus pyogenes/genética , Adulto , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Estudios de Seguimiento , Voluntarios Sanos , Humanos , ARN Bacteriano/genética , ARN Bacteriano/aislamiento & purificación , Sensibilidad y Especificidad , Streptococcus pyogenes/clasificación , Streptococcus pyogenes/aislamiento & purificación
5.
Nat Commun ; 12(1): 2267, 2021 04 15.
Artículo en Inglés | MEDLINE | ID: mdl-33859184

RESUMEN

Studies in humans and laboratory animals link stable gut microbiome "enterotypes" with long-term diet and host health. Understanding how this paradigm manifests in wild herbivores could provide a mechanistic explanation of the relationships between microbiome dynamics, changes in dietary resources, and outcomes for host health. We identify two putative enterotypes in the African buffalo gut microbiome. The enterotype prevalent under resource-abundant dietary regimes, regardless of environmental conditions, has high richness, low between- and within-host beta diversity, and enrichment of genus Ruminococcaceae-UCG-005. The second enterotype, prevalent under restricted dietary conditions, has reduced richness, elevated beta diversity, and enrichment of genus Solibacillus. Population-level gamma diversity is maintained during resource restriction by increased beta diversity between individuals, suggesting a mechanism for population-level microbiome resilience. We identify three pathogens associated with microbiome variation depending on host diet, indicating that nutritional background may impact microbiome-pathogen dynamics. Overall, this study reveals diet-driven enterotype plasticity, illustrates ecological processes that maintain microbiome diversity, and identifies potential associations between diet, enterotype, and disease.


Asunto(s)
Búfalos/microbiología , Enfermedades Transmisibles/veterinaria , Conducta Alimentaria/fisiología , Microbioma Gastrointestinal/inmunología , Animales , Búfalos/fisiología , Enfermedades Transmisibles/epidemiología , Enfermedades Transmisibles/microbiología , ADN Bacteriano/aislamiento & purificación , Heces/microbiología , Firmicutes/genética , Firmicutes/aislamiento & purificación , Incidencia , Metagenómica , Filogenia , Planococcaceae/genética , Planococcaceae/aislamiento & purificación , Prevalencia , ARN Ribosómico 16S/genética , Sudáfrica/epidemiología , Simbiosis/inmunología
6.
Nutrients ; 13(4)2021 Apr 13.
Artículo en Inglés | MEDLINE | ID: mdl-33924396

RESUMEN

BACKGROUND: Few preclinical studies have shown that Knee osteoarthritis (KOA) is linked to gut microbiome dysbiosis and chronic inflammation. This pilot study was designed to look at the gut microbiome composition in KOA patients and normal individuals with or without vitamin D deficiency (VDD, serum vitamin D <30 ng/mL). METHODS: This pilot study was conducted prospectively in 24 participants. The faecal samples of all the participants were taken for DNA extraction. The V3-V4 region of 16s rRNA was amplified, and the library was prepared and sequenced on the Illumina Miseq platform. RESULTS: The mean (±SD) age was 45.5 (±10.2) years with no defined comorbidities. Of 447 total Operational Taxonomic Units (OTUs), a differential abundance of 16 nominally significant OTUs between the groups was observed. Linear discriminate analysis (LEfSe) revealed a significant difference in bacteria among the study groups. Pseudobutyrivibrio and Odoribacter were specific for VDD, while Parabacteroides, Butyricimonas and Gordonibacter were abundant in the KOA_VDD group, and Peptococcus, Intestimonas, Delftia and Oribacterium were abundant in the KOA group. About 80% of bacterial species were common among different groups and hence labelled as core bacterial species. However, the core microbiome of KOA and VDD groups were not seen in the KOA_VDD group, suggesting that these bacterial groups were affected by the interaction of the KOA and VDD factors. CONCLUSION: Parabacteroides, Butyricimonas, Pseudobutyrivibrio, Odoribacter and Gordonibacter are the predominant bacteria in vitamin D deficient patients with or without KOA. Together these results indicate an association between the gut microbiome, vitamin D and knee osteoarthritis.


Asunto(s)
Disbiosis/complicaciones , Microbioma Gastrointestinal/inmunología , Osteoartritis de la Rodilla/inmunología , Deficiencia de Vitamina D/inmunología , Adulto , ADN Bacteriano/aislamiento & purificación , Disbiosis/diagnóstico , Disbiosis/inmunología , Disbiosis/microbiología , Heces/microbiología , Microbioma Gastrointestinal/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Persona de Mediana Edad , Osteoartritis de la Rodilla/microbiología , Filogenia , Proyectos Piloto , ARN Ribosómico 16S/genética , Vitamina D/sangre , Deficiencia de Vitamina D/sangre , Deficiencia de Vitamina D/diagnóstico , Deficiencia de Vitamina D/microbiología
7.
Nat Commun ; 12(1): 2126, 2021 04 09.
Artículo en Inglés | MEDLINE | ID: mdl-33837203

RESUMEN

There is accumulating evidence that the lower airway microbiota impacts lung health. However, the link between microbial community composition and lung homeostasis remains elusive. We combine amplicon sequencing and bacterial culturing to characterize the viable bacterial community in 234 longitudinal bronchoalveolar lavage samples from 64 lung transplant recipients and establish links to viral loads, host gene expression, lung function, and transplant health. We find that the lung microbiota post-transplant can be categorized into four distinct compositional states, 'pneumotypes'. The predominant 'balanced' pneumotype is characterized by a diverse bacterial community with moderate viral loads, and host gene expression profiles suggesting immune tolerance. The other three pneumotypes are characterized by being either microbiota-depleted, or dominated by potential pathogens, and are linked to increased immune activity, lower respiratory function, and increased risks of infection and rejection. Collectively, our findings establish a link between the lung microbial ecosystem, human lung function, and clinical stability post-transplant.


Asunto(s)
Rechazo de Injerto/microbiología , Trasplante de Pulmón/efectos adversos , Pulmón/microbiología , Microbiota/inmunología , Neumonía Bacteriana/microbiología , Adulto , Aloinjertos/inmunología , Aloinjertos/microbiología , Bacterias/genética , Bacterias/inmunología , Bacterias/aislamiento & purificación , Bacterias/patogenicidad , Carga Bacteriana/inmunología , Técnicas Bacteriológicas , Líquido del Lavado Bronquioalveolar/microbiología , Broncoscopía , ADN Bacteriano/aislamiento & purificación , Femenino , Rechazo de Injerto/diagnóstico , Rechazo de Injerto/inmunología , Humanos , Tolerancia Inmunológica , Estudios Longitudinales , Pulmón/inmunología , Masculino , Metagenómica , Microbiota/genética , Persona de Mediana Edad , Neumonía Bacteriana/diagnóstico , Neumonía Bacteriana/inmunología , Estudios Prospectivos , ARN Ribosómico 16S/genética
8.
Int J Mol Sci ; 22(8)2021 Apr 16.
Artículo en Inglés | MEDLINE | ID: mdl-33923762

RESUMEN

The efficiency of existing cell lysis methods to isolate nucleic acids from diverse bacteria varies depending on cell wall structures. This study tested a novel idea of using broad-spectrum antimicrobial peptides to improve the lytic efficiency of hard-to-lyse bacteria and characterized their differences. The lysis conditions of Staphylococcus aureus using recombinant porcine myeloid antimicrobial peptide 36 (PMAP-36), a broad-spectrum pig cathelicidin, was optimized, and RNA isolation was performed with cultured pellets of ten bacterial species using various membranolytic proteins. Additionally, three other antimicrobial peptides, protegrin-1 (PG-1), melittin, and nisin, were evaluated for their suitability as the membranolytic agents of bacteria. However, PMAP-36 use resulted in the most successful outcomes in RNA isolation from diverse bacterial species. The amount of total RNA obtained using PMAP-36 increased by ~2-fold compared to lysozyme in Salmonella typhimurium. Streptococci species were refractory to all lytic proteins tested, although the RNA yield from PMAP-36 treatment was slightly higher than that from other methods. PMAP-36 use produced high-quality RNA, and reverse transcription PCR showed the efficient amplification of the 16S rRNA gene from all tested strains. Additionally, the results of genomic DNA isolation were similar to those of RNA isolation. Thus, our findings present an additional option for high quality and unbiased nucleic acid isolation from microbiomes or challenging bacterial strains.


Asunto(s)
Péptidos Catiónicos Antimicrobianos/química , ARN Bacteriano/química , Staphylococcus aureus/química , Péptidos Catiónicos Antimicrobianos/farmacología , Fraccionamiento Celular/métodos , Fraccionamiento Celular/normas , ADN Bacteriano/química , ADN Bacteriano/aislamiento & purificación , ARN Bacteriano/aislamiento & purificación , Staphylococcus aureus/efectos de los fármacos
9.
Methods Mol Biol ; 2278: 21-29, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33649945

RESUMEN

Rapid and efficient protocols aimed at the isolation and purification of DNA for the purpose of downstream applications, such as cloning, PCR, Southern blotting, or sequencing, are essential for genetic, biochemical, and molecular biological analyses of a given bacterium. The protocols herein presented provide a robust and efficient method for the isolation of chromosomal and plasmid DNA from Bifidobacterium strains by organic extraction. The methods are simple, and the yield, purity, and quality of the DNA are adequate to perform various downstream applications including next-generation sequencing.


Asunto(s)
Bifidobacterium/genética , ADN Bacteriano/genética , Plásmidos/genética , Cromosomas Bacterianos/genética , ADN Bacteriano/aislamiento & purificación , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Plásmidos/aislamiento & purificación , Secuenciación Completa del Genoma/métodos
10.
Medicine (Baltimore) ; 100(10): e24915, 2021 Mar 12.
Artículo en Inglés | MEDLINE | ID: mdl-33725850

RESUMEN

RATIONALE: Antibiotic resistance poses a challenge for Helicobacter pylori eradication treatment. Current guidelines strongly recommend avoiding repeated treatments with the same antibiotic to prevent the emergence of drug resistance. However, for penicillin-allergic patients with recurrent H. pylori eradication failures, avoiding repeated treatments with the same antibiotic severely limits the choice of treatment. PATIENT CONCERNS: A 47-year-old woman with a penicillin allergy for whom 2 previous levofloxacin and bismuth-based therapies had failed. DIAGNOSIS: H. pylori infection. INTERVENTIONS: Agar dilution susceptibility testing and gene sequence analysis was performed to confirm levofloxacin susceptibility again. Therefore, we treated her with a 14-day regimen consisting of levofloxacin (500 mg once daily), furazolidone (100 mg twice daily), colloidal bismuth pectin (220 mg twice daily), and esomeprazole (20 mg twice daily). OUTCOMES: The patient was successfully treated with a third levofloxacin and bismuth-based regimen. LESSONS: Antibiotics included in previous failed therapies need not be eliminated if no antibiotic resistance is found on antimicrobial susceptibility testing.


Asunto(s)
Antibacterianos/farmacología , Gastritis/tratamiento farmacológico , Infecciones por Helicobacter/tratamiento farmacológico , Helicobacter pylori/efectos de los fármacos , Levofloxacino/farmacología , Antibacterianos/uso terapéutico , Bismuto/uso terapéutico , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Hipersensibilidad a las Drogas/diagnóstico , Hipersensibilidad a las Drogas/inmunología , Farmacorresistencia Bacteriana/genética , Quimioterapia Combinada/métodos , Esomeprazol/uso terapéutico , Femenino , Furazolidona/uso terapéutico , Mucosa Gástrica/patología , Gastritis/diagnóstico , Gastritis/microbiología , Gastritis/patología , Infecciones por Helicobacter/diagnóstico , Infecciones por Helicobacter/microbiología , Infecciones por Helicobacter/patología , Helicobacter pylori/genética , Helicobacter pylori/aislamiento & purificación , Humanos , Levofloxacino/uso terapéutico , Persona de Mediana Edad , Penicilinas/inmunología , Penicilinas/uso terapéutico , Recurrencia , Retratamiento/métodos , Resultado del Tratamiento
11.
Medicine (Baltimore) ; 100(10): e24970, 2021 Mar 12.
Artículo en Inglés | MEDLINE | ID: mdl-33725864

RESUMEN

ABSTRACT: The aim of this study was to discuss the correlation between the sulfamethoxazole-trimethoprim resistance of Shigella flexneri (S. flexneri) and the antibiotic resistance genes sul1, sul2, and sul3 and SXT element.From May 2013 to October 2018, 102 isolates of S. flexneri were collected from the clinical samples in Jinan. The Kirby-Bauer (K-B) test was employed to determine the antibiotic susceptibility of the S. flexneri isolates. The antibiotic resistance rate was analyzed with the WHONET5.4 software. The isolates were subject to the PCR amplification of the sul genes (sul1, sul2, and sul3) and the SXT element. On the basis of the sequencing results, the correlation between the sulfamethoxazole-trimethoprim resistance of the S. flexneri isolates and the sul genes was analyzed.The antibiotic resistance rates of the 102 S. flexneri isolates to ampicillin, streptomycin, chloramphenicol, tetracycline, and sulfamethoxazole-trimethoprim were 90.2%, 90.2%, 88.2%, 88.2%, and 62.7%, respectively. The antibiotic resistance rates of these isolates to cefotaxime, ceftazidime, and ciprofloxacin varied between 20% and 35%. However, these isolates were 100% susceptible to cefoxitin. Positive fragments were amplified from 59.8% (61/102) of the 102 S. flexneri isolates, the sizes of the sul1 and sul2 genes being 338 bp and 286 bp, respectively. The sequence alignment revealed the presence of the sul1 and sul2 genes encoding for dihydrofolate synthase. The carrying rate of the sul1 gene was 13.7% (14/102), and that of the sul2 gene was 48.0% (49/102). No target gene fragments were amplified from the 3 isolates resistant to sulfamethoxazole-trimethoprim. The sul3 gene and SXT element were not amplified from any of the isolates. The testing and statistical analysis showed that the resistance of the S. flexneri isolates to sulfamethoxazole-trimethoprim correlated to the sul1 and sul2 genes.The acquired antibiotic resistance genes sul1 and sul2 were closely associated with the resistance of the 102 S. flexneri isolates to sulfamethoxazole-trimethoprim.


Asunto(s)
Proteínas Bacterianas/genética , Proteínas Portadoras/genética , Disentería Bacilar/tratamiento farmacológico , Shigella flexneri/genética , Resistencia al Trimetoprim/genética , Combinación Trimetoprim y Sulfametoxazol/farmacología , Elementos Transponibles de ADN/genética , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Disentería Bacilar/microbiología , Heces/microbiología , Humanos , Pruebas de Sensibilidad Microbiana , Reacción en Cadena de la Polimerasa , Shigella flexneri/efectos de los fármacos , Shigella flexneri/aislamiento & purificación , Combinación Trimetoprim y Sulfametoxazol/uso terapéutico
12.
Medicine (Baltimore) ; 100(13): e25362, 2021 Apr 02.
Artículo en Inglés | MEDLINE | ID: mdl-33787640

RESUMEN

ABSTRACT: We investigated the vaginal flora diversity of preschool-aged (ie, 4-6-year-old) girls in southwest China.Fourteen preschool-aged girls were enrolled in this study. The statuses and differences in their vaginal flora were evaluated by Gram staining, bacterial culturing, and sequencing analysis.Gram staining and microbial culturing showed that the main vaginal flora of the preschool-aged girls were Gram-negative bacilli, whereas the main vaginal flora of healthy adult controls were large Gram-positive bacilli such as Lactobacillus crispatus. Shannon and Simpson indexes indicated that the bacterial diversity tended to decrease with age. The species abundance heat map showed that the vaginal microecology of the girls differed slightly at different ages but mainly comprised Pseudomonas, Methylobacterium, Sphingomona,s and Escherichia. The functional abundance heat map indicated that the bacterial functions increased with age.The vaginal microecology of preschool-aged girls differs from that of adults. A comprehensive understanding of the vaginal flora diversity of preschool-aged girls will aid in clinically diagnosing vulvovaginitis in preschool-aged girls.


Asunto(s)
Bacterias/aislamiento & purificación , Microbiota/genética , Vagina/microbiología , Vulvovaginitis/diagnóstico , Adulto , Factores de Edad , Bacterias/genética , Estudios de Casos y Controles , Niño , Preescolar , China , ADN Bacteriano/aislamiento & purificación , Femenino , Voluntarios Sanos , Humanos , Tipificación Molecular/métodos , Análisis de Secuencia de ADN , Frotis Vaginal , Vulvovaginitis/microbiología
13.
PLoS One ; 16(1): e0241732, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33406075

RESUMEN

Characterization of poultry microbiota is becoming increasingly important due to the growing need for microbiome-based interventions to improve poultry health and production performance. However, the lack of standardized protocols for sampling, sample processing, DNA extraction, sequencing, and bioinformatic analysis can hinder data comparison between studies. Here, we investigated how the DNA extraction process affects microbial community compositions and diversity metrics in different chicken respiratory sample types including choanal and tracheal swabs, nasal cavity and tracheal washes, and lower respiratory lavage. We did a side-by-side comparison of the performances of Qiagen DNeasy blood and tissue (BT) and ZymoBIOMICS DNA Miniprep (ZB) kits. In general, samples extracted with the BT kit yielded higher concentrations of total DNA while those extracted with the ZB kit contained higher numbers of bacterial 16S rRNA gene copies per unit volume. Therefore, the samples were normalized to equal amounts of 16S rRNA gene copies prior to sequencing. For each sample type, all predominant bacterial taxa detected in samples extracted with one kit were present in replicate samples extracted with the other kit and did not show significant differences at the class level. However, a few differentially abundant shared taxa were observed at family and genus levels. Furthermore, between-kit differences in alpha and beta diversity metrics at the amplicon sequence variant level were statistically indistinguishable. Therefore, both kits perform similarly in terms of 16S rRNA gene-based poultry microbiome analysis for the sample types analyzed in this study.


Asunto(s)
Pollos/microbiología , ADN Bacteriano , ADN Ribosómico , Microbiota , ARN Ribosómico 16S , Juego de Reactivos para Diagnóstico , Sistema Respiratorio/microbiología , Animales , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , ADN Ribosómico/genética , ADN Ribosómico/aislamiento & purificación , ARN Ribosómico 16S/genética , ARN Ribosómico 16S/aislamiento & purificación
14.
Food Microbiol ; 95: 103705, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33397623

RESUMEN

Amplicon sequencing approaches have been widely used in food bacterial ecology. However, choices regarding the methodology can bias results. In this study, bacterial communities associated with cold-smoked salmon products and their processing plant surfaces were monitored via sequencing of the V3-V4 region of the 16S rRNA gene. The impact of DNA extraction protocols, sampling methods (swabbing or sponging) and surface materials on bacterial communities were investigated. α and ß diversity analyses revealed that DNA extraction methods mainly influence the observed cold-smoked salmon microbiota composition. Moreover, different DNA extraction methods revealed significant differences in observed community richness and evenness. ß-Proteobacteria: Photobacterium, Serratia and Firmicutes: Brochothrix, Carnobacterium and Staphylococcus were identified as the dominant genera. Surface microbiota richness, diversity and composition were mainly affected by cleaning and disinfection procedures but not by DNA extraction methods. Surface community richness and evenness appeared higher when sampled by sponging compared to swabbing. ß-diversity analyses highlighted that surface topology, cleaning and disinfection and sampling devices seemed to affect the bacterial community composition. The dominant surface bacteria identified were mainly Flavobacteriaceae, ß-Proteobacteria and γ-Proteobacteria described as fish spoilers such as Acinetobacter, Pseudomonas and Shewanella. DNA extraction and sampling methods can have an impact on sequencing results and the ecological analysis of bacterial community structures. This study confirmed the importance of methodology standardization and the need for analytical validation before 16S rDNA metabarcoding surveys.


Asunto(s)
Bacterias/aislamiento & purificación , ADN Bacteriano/aislamiento & purificación , Productos Pesqueros/microbiología , Técnicas Genéticas , Microbiota , ARN Ribosómico 16S/aislamiento & purificación , Salmón/microbiología , Animales , Bacterias/clasificación , Bacterias/genética , Bacterias/metabolismo , ADN Bacteriano/genética , ADN Ribosómico/genética , ADN Ribosómico/aislamiento & purificación , Manipulación de Alimentos/instrumentación , ARN Ribosómico 16S/genética
15.
J Gastrointest Cancer ; 52(1): 365-368, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33492618

RESUMEN

PURPOSE: There is limited data regarding the fecal microbiome findings in patients with Lynch syndrome. We aimed to study the fecal micobiome of patients with Lynch syndrome with and without cancer. METHODS: We performed an observational study comparing the fecal microbiome of patients with Lynch syndrome (LS) with cancer with those without cancer. We included subjects older than 18 years with LS and excluded those with a history of colectomy or inflammatory bowel disease. We analyzed their fecal microbiome by 16S ribosomal subunit PCR amplification and performed comparative analyses. RESULTS: Eight patients were included: 3 of these with LS and cancer (LS-C) and 5 patients with LS and no cancer (LS-NC). We found non-significant differences at the phyla and genera level between the LS-C and LS-NC groups. At the phyla level, LS-C patients had a higher percentage of Bacteroidetes (42.2% vs. 28.5%; P = 0.068) and Verrucomicrobia (0.644% vs 0.0007%; P = 0.10), and a lower percentage of Firmicutes (48.3% vs. 65.4%; P = 0.078). At the genus level, LS-C patients had a higher rate of Akkermania (0.766% vs. 0.001%; P = 0.11). LS-C patients with endometrial cancer had a higher rate of Bacteroides (37.4% vs 17.3%; P = 0.10). LS-C patients had a lower rate of Pseudobutyrvibrio (0.74% vs. 2.71%; P = 0.10). CONCLUSIONS: The fecal microbiome of LS patients with extraintestinal cancer differs that of LS patients without cancer. Further studies are needed to explore microbiome changes in these high risk patients.


Asunto(s)
Neoplasias Colorrectales Hereditarias sin Poliposis/microbiología , Neoplasias Endometriales/microbiología , Microbioma Gastrointestinal/genética , Neoplasias Ováricas/microbiología , Adulto , Estudios de Casos y Controles , Neoplasias Colorrectales Hereditarias sin Poliposis/complicaciones , Neoplasias Colorrectales Hereditarias sin Poliposis/genética , ADN Bacteriano/aislamiento & purificación , Neoplasias Endometriales/genética , Heces/microbiología , Femenino , Humanos , Masculino , Mutación , Neoplasias Ováricas/genética , ARN Ribosómico 16S/genética
16.
Biosens Bioelectron ; 178: 113001, 2021 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-33493900

RESUMEN

Amplification-based nucleic acid detection is widely employed in food safety, medical diagnosis and environment monitoring. However, conventional nucleic acid analysis has to be carried out in laboratories because of requiring expensive instruments and trained personnel. If people could do nucleic acid detection at home by themselves, the application of nucleic acid detection would be greatly accelerated. We herein reported a polypropylene (PP) bag-based method for convenient detection of nucleic acids in the oil-sealed space. The PP bag has three chambers which are responsible for lysis, washing and amplification/detection, respectively. After adding sample, nucleic acids are adsorbed on magnetic particles (MPs) and moved into these three chambers successively through immiscible oil channel by an external magnet. Combined with isothermal amplification, the PP bag can be incubated in a water bath or milk warmer and acted as a reaction tube. With highly specific CRISPR technology, Salmonella typhimurium (St) and SARS-CoV-2 can be visually detected in these PP bags within 1 h, indicating its potential household application. To further improve the reliability of nucleic acid testing at home, a logic decision method is introduced by detecting both target and endogenous reference gene. Positive/negative/invalid detection result can be obtained by chronologically adding the CRISPR reagents of target and endogenous reference gene. We anticipate that this PP bag can provide a novel toolkit for nucleic acid detection in people's daily life.


Asunto(s)
/métodos , /virología , Sistemas CRISPR-Cas , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificación de Ácido Nucleico/métodos , /aislamiento & purificación , Técnicas Biosensibles/instrumentación , Técnicas Biosensibles/métodos , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Microbiología de Alimentos , Humanos , Magnetismo , Técnicas de Diagnóstico Molecular/instrumentación , Técnicas de Amplificación de Ácido Nucleico/instrumentación , Polipropilenos , ARN Viral/genética , ARN Viral/aislamiento & purificación , Salmonella typhimurium/genética , Salmonella typhimurium/aislamiento & purificación
17.
Zoonoses Public Health ; 68(2): 121-130, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33428331

RESUMEN

The aim of this work was the establishment of a national laboratory sentinel surveillance service for human clinical Campylobacter in Ireland. This included detailed genomic molecular epidemiology of Campylobacter for 2019. For February-December 2019, 24 clinical microbiology laboratories in Ireland submitted all PCR/culture-positive clinical Campylobacter spp. specimens to Public Health Laboratory (PHL) Dublin one week out of every four. Antimicrobial susceptibility testing (AST) according to European Committee on Antimicrobial Susceptibility Testing (EUCAST) criteria was carried out for Campylobacter spp. isolates for ciprofloxacin, tetracycline and erythromycin. Batch whole genome sequencing (WGS) was carried out on cultures and analysis was performed to determine species, genotype, identify antimicrobial resistance (AMR) and virulence determinants and identify clusters. A total of 75 isolates and 366 PCR-positive stools were received, and 277 isolates recovered (55.7% recovery from stools). Of 257 isolates characterized by WGS, 86.4% (n = 222) were Campylobacter jejuni, 11.7% (n = 30) Campylobacter coli and 1.9% (n = 5) Campylobacter lari. There were 20 clonal complexes with ST-21 clonal complex most prevalent at 26.8% (n = 69). 50.5% (n = 140) of isolates were susceptible to all three antimicrobials tested. 39.3% (n = 109) isolates were ciprofloxacin resistant, 26.3% (n = 73) tetracycline resistant and two isolates erythromycin resistant. Congruence between phenotypic and genotypic AST was observed. There was 95.9% and 95.6% sensitivity and specificity for WGS to predict ciprofloxacin sensitivity and 98.6% and 99.5% sensitivity and specificity for WGS to predict tetracycline sensitivity. Virulence factors flaA, racR, ciaB and cdtB were detected in all isolates. WGS identified 31 potential clusters for public health alert. This sentinel surveillance of human campylobacteriosis in Ireland establishes the basis for a national reference service. Linking with other partners in a 'One Health' framework will help us better understand sources of infection to reduce disease burden and the threat of AMR.


Asunto(s)
Infecciones por Campylobacter/epidemiología , Infecciones por Campylobacter/microbiología , Campylobacter/aislamiento & purificación , Vigilancia de Guardia , Antibacterianos/farmacología , Campylobacter/efectos de los fármacos , Campylobacter/genética , Análisis por Conglomerados , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Farmacorresistencia Bacteriana , Regulación Bacteriana de la Expresión Génica , Genoma Bacteriano , Genotipo , Humanos , Irlanda/epidemiología , Factores de Virulencia/genética , Factores de Virulencia/metabolismo , Secuenciación Completa del Genoma
18.
Nat Commun ; 12(1): 161, 2021 01 08.
Artículo en Inglés | MEDLINE | ID: mdl-33420064

RESUMEN

Calf diarrhea is associated with enteric infections, and also provokes the overuse of antibiotics. Therefore, proper treatment of diarrhea represents a therapeutic challenge in livestock production and public health concerns. Here, we describe the ability of a fecal microbiota transplantation (FMT), to ameliorate diarrhea and restore gut microbial composition in 57 growing calves. We conduct multi-omics analysis of 450 longitudinally collected fecal samples and find that FMT-induced alterations in the gut microbiota (an increase in the family Porphyromonadaceae) and metabolomic profile (a reduction in fecal amino acid concentration) strongly correlate with the remission of diarrhea. During the continuous follow-up study over 24 months, we find that FMT improves the growth performance of the cattle. This first FMT trial in ruminants suggest that FMT is capable of ameliorating diarrhea in pre-weaning calves with alterations in their gut microbiota, and that FMT may have a potential role in the improvement of growth performance.


Asunto(s)
Enfermedades de los Bovinos/terapia , Bovinos/crecimiento & desarrollo , Diarrea/terapia , Trasplante de Microbiota Fecal/veterinaria , Microbioma Gastrointestinal/genética , Animales , Bacteroidaceae/genética , Bacteroidaceae/aislamiento & purificación , Bovinos/microbiología , Enfermedades de los Bovinos/sangre , Enfermedades de los Bovinos/metabolismo , Enfermedades de los Bovinos/microbiología , ADN Bacteriano/aislamiento & purificación , Diarrea/sangre , Diarrea/metabolismo , Diarrea/microbiología , Heces/microbiología , Femenino , Estudios de Seguimiento , Genómica , Masculino , Metabolómica , ARN Ribosómico 16S/genética , Resultado del Tratamiento
19.
Gut ; 70(4): 698-706, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33431578

RESUMEN

OBJECTIVE: Although COVID-19 is primarily a respiratory illness, there is mounting evidence suggesting that the GI tract is involved in this disease. We investigated whether the gut microbiome is linked to disease severity in patients with COVID-19, and whether perturbations in microbiome composition, if any, resolve with clearance of the SARS-CoV-2 virus. METHODS: In this two-hospital cohort study, we obtained blood, stool and patient records from 100 patients with laboratory-confirmed SARS-CoV-2 infection. Serial stool samples were collected from 27 of the 100 patients up to 30 days after clearance of SARS-CoV-2. Gut microbiome compositions were characterised by shotgun sequencing total DNA extracted from stools. Concentrations of inflammatory cytokines and blood markers were measured from plasma. RESULTS: Gut microbiome composition was significantly altered in patients with COVID-19 compared with non-COVID-19 individuals irrespective of whether patients had received medication (p<0.01). Several gut commensals with known immunomodulatory potential such as Faecalibacterium prausnitzii, Eubacterium rectale and bifidobacteria were underrepresented in patients and remained low in samples collected up to 30 days after disease resolution. Moreover, this perturbed composition exhibited stratification with disease severity concordant with elevated concentrations of inflammatory cytokines and blood markers such as C reactive protein, lactate dehydrogenase, aspartate aminotransferase and gamma-glutamyl transferase. CONCLUSION: Associations between gut microbiota composition, levels of cytokines and inflammatory markers in patients with COVID-19 suggest that the gut microbiome is involved in the magnitude of COVID-19 severity possibly via modulating host immune responses. Furthermore, the gut microbiota dysbiosis after disease resolution could contribute to persistent symptoms, highlighting a need to understand how gut microorganisms are involved in inflammation and COVID-19.


Asunto(s)
Bacterias , Disbiosis , Microbioma Gastrointestinal/inmunología , Tracto Gastrointestinal , Inmunidad , Adulto , Bacterias/genética , Bacterias/inmunología , Bacterias/aislamiento & purificación , Proteína C-Reactiva/análisis , /diagnóstico , /inmunología , Citocinas/análisis , ADN Bacteriano/aislamiento & purificación , Disbiosis/epidemiología , Disbiosis/etiología , Disbiosis/inmunología , Disbiosis/virología , Femenino , Tracto Gastrointestinal/inmunología , Tracto Gastrointestinal/microbiología , Tracto Gastrointestinal/virología , Hong Kong , Humanos , Masculino , /aislamiento & purificación , Índice de Severidad de la Enfermedad , Transferasas/análisis
20.
Int J Cancer ; 148(1): 178-192, 2021 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-32803883

RESUMEN

Helicobacter pylori (H. pylori) are a primary factor in the pathogenesis of gastric cancer (GC); GC ranks third among cancer-related mortality. A clear understanding of the H. pylori genome factors underlying GC is necessary to develop more effective methods to prevent GC. A single-molecule real-time DNA sequencing-based H. pylori genome-wide association study analysis was performed using the H. pylori genome present in five early-stage GC (EGC) and five non-GC clinical DNA samples recovered from gastric washes. A total of 275 genes with 702 nucleotide variants (NVs) were found to be common to three or more patients with EGC but no non-GC patients (single-NV: 654/702, 93.2%; multi-NV: 40/702, 5.7%; deletion: 3/702, 0.4%; insertion: 3/702, 0.7%). Gene ontology analysis of H. pylori revealed that genes involved in the mitochondrial electron transport system, glycolytic processes and the TCA cycle were highly enriched. Cancer-related NVs were most frequently found in a member of the Helicobacter outer membrane protein family, hopL. In particular, one of the NVs in hopL was a novel six-nucleotide insertion (1159095̂1159096, TACTTC); this mutant was detected more frequently in a validation set of 50 additional EGC samples (22/50, 44.0%) than in 18 non-GC samples (3/18, 16.7%, P = .04). These results suggest that the hopL variant is associated with the development of GC and may serve as a genetic biomarker of H. pylori virulence and GC risk. Our assay can serve as a potent tool to expand our understanding of bacteria-associated tumorigenesis.


Asunto(s)
Mucosa Gástrica/patología , Infecciones por Helicobacter/microbiología , Helicobacter pylori/genética , Neoplasias Gástricas/microbiología , Anciano , Anciano de 80 o más Años , Proteínas Bacterianas/genética , Biomarcadores , Transformación Celular Neoplásica , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Femenino , Mucosa Gástrica/microbiología , Genoma Bacteriano , Infecciones por Helicobacter/patología , Helicobacter pylori/aislamiento & purificación , Helicobacter pylori/patogenicidad , Humanos , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Polimorfismo de Nucleótido Simple , Neoplasias Gástricas/diagnóstico , Neoplasias Gástricas/patología , Factores de Virulencia/genética , Secuenciación Completa del Genoma
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