RESUMO
To rapidly identify Mycobacterium and Nocardia spp. without costly probes, we had implemented capillary electrophoresis (CE) in polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis to analyze their 65-kDa heat shock protein (hsp65) gene. The PCR-RFLP analysis with CE (PRACE) involved only one restriction enzyme, HaeIII, and a single electrophoretic separation less than 10 min. Full-range (10-200 bp) RFLP patterns of 12 less common Mycobacterium and 7 Nocardia spp. were investigated. A good agreement was observed between the sizes of restriction fragments resolved by CE and the real sizes deduced from sequence analysis. Including hsp65 gene patterns of 12 Mycobacterium spp. published earlier, differentiation was distinct among 24 Mycobacterium and 7 Nocardia spp. Some closely related species exhibiting similar biochemical characteristics could be well discriminated by an extra HaeIII digestion site. Thus, PRACE offers a nonprobe alternative for rapid identification of various cultured Mycobacterium and Nocardia to the species level.
Assuntos
Proteínas de Bactérias/genética , Chaperoninas/genética , Eletroforese Capilar/métodos , Mycobacterium/classificação , Nocardia/classificação , Polimorfismo de Fragmento de Restrição , Chaperonina 60 , DNA Bacteriano/genética , Desoxirribonucleases de Sítio Específico do Tipo II , Mycobacterium/genética , Nocardia/genética , Reação em Cadeia da PolimeraseRESUMO
Sepsis is a life-threatening medical condition. Salidroside, a substance isolated from Rhodiola rosea, possesses antioxidant and anti-inflammatory properties. The effect and mechanism of salidroside on sepsis-induced acute lung injury still remains to be well clarified. Here, we investigated the effect and mechanism of salidroside on septic mouse models and explored the role of salidroside-upregulated SIRT1. Salidroside inhibited the inflammatory responses and HMGB1 productions in bacterial lipopolysaccharide (LPS)-treated macrophages and mice. Salidroside could also reverse the decreased SIRT1 protein expression in LPS-treated macrophages and mice. Salidroside also alleviated the sepsis-induced lung edema, lipid peroxidation, and histopathological changes and the mortality, and improved the lung PaO2/FiO2 ratio in cecal ligation and puncture (CLP)-induced septic mice. Salidroside significantly decreased the serum TNF-α, IL-6, NO, and HMGB1 productions, pulmonary inducible NO synthase (iNOS) and phosphorylated NF-κB-p65 protein expressions, and pulmonary HMGB1 nuclear translocation in CLP septic mice. Moreover, sepsis decreased the SIRT1 protein expression in the lungs of CLP septic mice. Salidroside significantly upregulated the SIRT1 expression and inhibited the inflammatory responses in CLP septic mouse lungs. These results suggest that salidroside protects against sepsis-induced acute lung injury and mortality, which might be through the SIRT1-mediated repression of NF-κB activation and HMGB1 nucleocytoplasmic translocation.
Assuntos
Lesão Pulmonar Aguda/prevenção & controle , Glucosídeos/farmacologia , Proteína HMGB1/metabolismo , NF-kappa B/metabolismo , Fenóis/farmacologia , Sepse/complicações , Sirtuína 1/metabolismo , Lesão Pulmonar Aguda/etiologia , Lesão Pulmonar Aguda/metabolismo , Animais , Regulação para Baixo/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Fitoterapia , Células RAW 264.7 , Rhodiola/química , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacosRESUMO
Intestinal parasitic infections (IPIs) among schoolchildren in Republic of Marshall Islands (RMI) largely remains unknown, thus investigation on IPIs status to establish the baseline data is urgently needed. This cross-sectional study intended to investigate the current IPIs status and associated risk factors among schoolchildren at capital of RMI. Single stool sample from 400 schoolchildren (207 boys and 193 girls) aged 9.73±2.50 yrs old was examined by employing merthiolate-iodine-formaldehyde concentration method. Demographic characteristics, uncomfortable symptoms and risk factors were obtained by questionnaires investigation. The overall prevalence of IPIs in schoolchildren was 22.8% (91/400), of them 24.2% harbored at least 2 different parasites. Notably, the majority was infected by waterborne protozoan parasites (82.4%, 75/91). Nine different intestinal parasites have been identified, of which six were pathogenic including Hook worm, Trichuris trichiura, Enterobius vermicularis, Entamoeba histolytica/dispar, Giardia intestinalis and Blastocystis hominis. Schoolchildren who ever complained dizziness or headache showed a significant higher prevalence of pathogenic IPIs than those who did not (p<0.05). Schoolchildren who lived in urban area than rural area had higher chance to acquire pathogenic IPIs (p=0.03). However, none of risk factors were identified to be associated with pathogenic IPIs.
Assuntos
Enteropatias Parasitárias/epidemiologia , Animais , Blastocystis hominis/isolamento & purificação , Criança , Estudos Transversais , Entamoeba histolytica/isolamento & purificação , Enterobius/isolamento & purificação , Fezes/parasitologia , Feminino , Giardia lamblia/isolamento & purificação , Humanos , Enteropatias Parasitárias/parasitologia , Masculino , Micronésia/epidemiologia , Prevalência , Características de Residência , Fatores de Risco , Trichuris/isolamento & purificaçãoRESUMO
BACKGROUND: Breast milk is the main source of postnatal human cytomegalovirus (HCMV) infection. The aim of this study was to assess the relationship between HCMV load in breast milk and viral transmission to very low birth weight (VLBW) infants. METHODS: Breast-fed VLBW infants who were born to HCMV-seropositive mothers and who were managed in a neonatal intensive care unit were enrolled in the study. Blood from mothers and infants was tested for HCMV antibodies after birth. Breast milk was collected for viral culture and HCMV load measurement. Urine from the babies was obtained for HCMV-DNA detection. Symptoms of HCMV infection were recorded and evaluated by neonatologists. RESULTS: Of the 23 evaluated mothers during a 1-year period, 19 were HCMV seropositive; 17 of the women had detectable HCMV-DNA in their breast milk whey. Of the 23 infants born to the 19 seropositive mothers, 8 infants of 8 mothers had HCMV-DNA detected in the urine, indicating that they were infected, even though the breast milk was always frozen prior to feeding. Three infected infants had symptoms. At 4 weeks after delivery, the median viral load in breast milk from mothers of the 8 infected infants was significantly higher than that from mothers of the 15 noninfected infants (P = 0.04). HCMV was detectable in breast milk for a significantly longer period in mothers of infected infants (7.5 vs. 2.6 weeks P = 0.03). CONCLUSIONS: High HCMV load and prolonged virus excretion in breast milk are maternal risk factors for viral transmission to VLBW infants.
Assuntos
Infecções por Citomegalovirus/transmissão , Citomegalovirus/isolamento & purificação , Recém-Nascido de muito Baixo Peso , Transmissão Vertical de Doenças Infecciosas , Leite Humano/virologia , Complicações Infecciosas na Gravidez/virologia , Carga Viral , DNA Viral/genética , DNA Viral/isolamento & purificação , Feminino , Humanos , Recém-Nascido , Masculino , Reação em Cadeia da Polimerase/métodos , Gravidez , Fatores de Tempo , Urina/virologiaRESUMO
The improvement of sensitivity and differentiation in rapidly identifying a small amount of mycobacteria in sputum has significant implications for reducing tuberculosis transmission. We previously applied the conventional PCR and capillary electrophoresis (CE) to establish the restriction fragment length polymorphism (RFLP) pattern of mycobacterial 65-kDa heat shock protein (hsp65) gene from colony specimens. However, the previous analysis did not provide enough sensitivity for sputum specimens in which the limitation of analysis might be hindered by PCR inhibitors and primer-dimers formation during amplification. In the current study, nested PCR (nPCR) had been redesigned for PCR-RFLP analysis (PRA) of mycobacterial hsp65 gene using CE. The results show both Mycobacterium tuberculosis complex and mycobacteria other than tuberculosis could be identified in the presence of PCR inhibitors. The interference due to primer-dimers was also minimized. Based on the Poisson distribution, the repeatability of single DNA molecule detection was greatly affected by sampling probability and might be improved significantly by increasing the sample loading. The PRA using nPCR and CE is not only able to detect the individual mycobacterial DNA molecule but also potentially differentiate the species.