Improvement of Bacillus subtilis Spore Enumeration and Label Analysis in Flow Cytometry.

The spores of Bacillus subtilis have already been proposed for different biotechnological and immunological applications; however, there is an increasing need for the development of methodologies that improve the detection of antigens immobilized on the surface of spores together with their quantification. Flow cytometry-based analyses have been previously proposed as fast, reliable, and specific approaches for detecting labeled cells of B. subtilis. Herein, we propose the use of flow cytometry to evaluate the display efficiency of a fluorescent antibody (FA) on the surface of the spore and quantify the number of spores using counting beads. For this, we used ethidium bromide as a DNA marker and an allophycocyanin (APC)-labeled antibody, which was coupled to the spores, as a surface marker. The quantification of spores was performed using counting beads since this technique demonstrates high accuracy in the detection of cells. The labeled spores were analyzed using a flow cytometer, which confirmed the coupling. As a result, it was demonstrated that DNA labeling improved the accuracy of quantification by flow cytometry, for the detection of germinated spores. It was observed that ethidium bromide was not able to label dormant spores; however, this technique provides a more precise determination of the number of spores with fluorescent protein coupled to their surface, thus helping in the development of studies that focus on the use of spores as a biotechnological platform in different applications.

Fatores de risco para perda de sensibilidade plantar em diabéticos: estudo caso-controle em ambulatório de endocrinologia / Risk factors for loss of plantar sensitivity in diabetics: a case-control study in endocrinology outpatient clinic

Introdução: Este estudo enfoca o problema da perda dasensibilidade plantar e seus fatores de risco, avaliadosretrospectivamente, em pacientes atendidos ambulatorialmenteno hospital universitário de João Pessoa, Paraíba,Brasil, onde muitos pacientes diabéticos são atendidos, masnão há conhecimento sobre a prevalência do risco dedesenvolvimento desse quadro. Objetivo: Avaliar fatoresde risco para perda de sensibilidade protetora plantar empacientes diabéticos atendidos no Hospital UniversitárioLauro Wanderley (HULW), e correlacionar esta perda comsintomas neuropáticos. Material e Métodos: Estudo casocontroleenvolvendo pacientes com sinais de neuropatiadiabética (casos), diabéticos sem neuropatia e pacientesnão-diabéticos (controles) atendidos nos ambulatórios deEndocrinologia do HULW. Aplicaram-se o Escore de SintomasNeuropáticos (ESN), a Escala Visual Analógica (EVA) e oteste do monofilamento de Semmes-Weinstein (10g).Resultados e Conclusão: A amostra foi de 90 pacientes,divididos em três grupos: 16 diabéticos tipo II neuropatas(casos; GDN), e por dois grupos-controle: grupo-controle 1com 42 diabéticos tipo II sem ND (GDNN) e grupo-controle 2com 32 adultos não-diabéticos (GND). Idade, duração do DMe valores de glicemia foram estatisticamente superiores noGDN. A odds ratio de 5 entre GDN e GDNN apontou para aassociação entre presença de neuropatia e tempo dediagnóstico de diabetes. O diagnóstico de NDP pelo examedo monofilamento não se relacionou com sintomasneuropáticos.

Introduction: This study evaluates retrospectively the lossof plant sensitivity and its risk factors in outpatients assistedin a university hospital in João Pessoa, Paraíba, Brazil, wheremany diabetic patients have been treated, but there is noknowledge about the prevalence of risk of developing thiscondition. Objective: To evaluate the risk factors for loss ofplantar protective sensitivity in diabetic outpatients at theUniversity Hospital Lauro Wanderley (HULW), as well as tocorrelate this loss with neuropathic symptoms. Material andMethods: This was a case-control study involving patientswith signs of diabetic neuropathy (cases), diabetic patientswithout neuropathy and non-diabetic patients (controls)attending the outpatient clinics of Endocrinology, HULW. Itwas applied the Neuropathy Symptom Score (ESN), VisualAnalogue Scale (VAS) and the test of Semmes-Weinsteinmonofilament (10 g). Results and Conclusion: The sampleconsisted of 90 patients divided into three groups: 16 type IIdiabetics with neuropathy (cases; GDN), and two controlgroups: a control-group 1 consisting of 42 type II diabeticswithout ND (GDND) and control-group 2 with 32 non-diabeticadults (GND). Age, diabetes duration and blood glucose levelswere significantly higher in GDN. An odds ratio of 5 wasfound for GDN and GDND, pointing to an association betweenthe presence of neuropathy and time since diabetes wasdiagnosed. The diagnosis of PDN by monofilament test wasnot related to neuropathic symptoms.