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In this study we determined some properties of the cholesterol oxidase from a Brevibacterium strain isolated from buffalos milk and identified the cholesterol degradation products by the bacterial cell. A small fraction of the enzyme synthesized by cells cultured in liquid medium for 7days was released into the medium whereas a larger fraction remained bound to the cell membrane. The extraction of this fraction was efficiently accomplished in 1 mM phosphate buffer, pH 7.0, containing 0.7% Triton X-100. The enzyme stability under freezing and at 45 o C was improved by addition of 20% glycerol. The optimum temperature and pH for the enzyme activity were 53°C and 7.5, respectively. The only steroidal product from cholesterol oxidation by the microbial cell and by the crude extract of the membrane-bound enzyme was 4-colesten-3-one. Chromatographic analysis showed that minor no steroidal compounds as well as 4-colesten-3-one found in the reaction media arose during fermentation process and were extracted together with the enzyme in the buffer solution. Cholesterol oxidation by the membrane-bound enzyme was a first order reaction type.

K lebsiella pneumoniae is an opportunistic bacteria that is a normal part of the nasopharyngeal and gastrointestinal tract microbiome of humans and animals (1). The hypermucoviscous variant of K. pneumoniae (hvKp), initially described in Southeast Asia, has emerged as a pathogen affecting young and healthy persons worldwide (2). The development of prominent polysaccharide capsules associated with capsular serotypes K1 or K2 have been reported as the major virulence determinants for human hvKp in liver abscesses, perhaps because it seems to protect the bacteria from phagocytosis and prevents destruction by bactericidal serum factors (2). K. pneumoniae strains have also been associated with a variety of diseases in animals, especially in Old World (Africa, Asia, and Europe) and New World (Oceana, North America, and South America) nonhuman primates (3-5). Sudden death or various clinical signs, including anorexia, prostration, fever, cough, dyspnea, mucopurulent discharge, meningitis, pneumonia, peritonitis, and sepsis are strongly associated with sporadic infections of K. pneumoniae in common marmosets research colonies (5,6). Despite the well-recognized zoonotic importance of hvKp and the public health risk of emerging multidrug-resistant strains (7-9), information is incomplete about the genotypic and phenotypic characterization of the etiologic agent essential to adequately diagnose and treat this pathogen in captive and wild nonhuman primates. The aim of this study was to report an epizootic among common marmosets in a wildlife rehabilitation center in Brazil and to describe the serotype, sequence typing, virulence properties, and resistance profile of the K. pneumoniae strains involved. The Study On February 10-11, 2019, a total of 11 captive marmosets (8 Callithrix penicillata, 2 C. jacchus, and 1 hybrid) died suddenly without clinical signs of disease. All of the animals were maintained in Parque Ecológico do Tietê, located in São Paulo municipality, São Paulo, Brazil, which is a center for receiving, rehabilitating, and referring wildlife. All animals had been in captivity 123-399 days. No new animals had been introduced into the cages in the previous 25 days. Each necropsy was performed <24 h after death in accordance with the Brazil Ministry of Health's guide for surveillance of epizootics in nonhuman primates (10). Tissue samples were preserved in phosphatebuffered formalin 10% and processed for routine histopathology and for 12 hours in refrigeration for microbiologic and molecular analyses. This study was approved by the Ethics Committee for the Use of