RESUMO
OBJECTIVES: To review the utilization of prostate-specific antigen (PSA) testing in Winnipeg, a major Canadian city, and to compare PSA testing rates between Winnipeg and Calgary, another major Canadian city of comparable size. METHODS: PSA testing results were reviewed by year and age group. We focused our studies in years 2011 and 2016, for which census demographic data are available. RESULTS: In Winnipeg, the PSA testing rates (patients with one or two PSA tests divided by the male population) showed a declining trend over years from 2008 to 2017. For almost all age groups, PSA testing rates in 2016 decreased in comparison to those in 2011. For age older than 40 years, the relative percentage decreases were 14% to 20%.In 2011, Winnipeg PSA testing rates were consistently higher than those in Calgary for all age groups. For age older than 40 years, the relative percentage differences were 36% to 50%.In addition, 41% and 40% of patients in Winnipeg who underwent PSA testing were younger than 50 years or older than 69 years in 2011 and 2016, respectively. CONCLUSIONS: PSA testing utilization may be falling short of optimal rates. There is a need to reinforce the optimal use of clinical recommendations.
Assuntos
Detecção Precoce de Câncer , Antígeno Prostático Específico/sangue , Neoplasias da Próstata/diagnóstico , Adulto , Fatores Etários , Idoso , Canadá , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias da Próstata/sangueRESUMO
High-throughput screening can create the potential ability to screen large numbers of monoclonal antibodies (mAb) in a short time period. A major bottleneck in the hybridoma method for mAb development has historically been the inability to sift through large numbers of hybridoma culture supernatants to identify clones secreting mAbs of the desired specificity. Herein, we develop a homogeneous fluorometric microvolume assay technology (FMAT) and compare it to conventional ELISA screening techniques for monoclonal antibody against soluble protein toxin fragments of the Clostridium botulinum types A, B and E neurotoxin (BoNT) proteins. In total 8,744 hybridoma clones were screened to identify 29 stable hybridomas to neurotoxin binding domain; six of these would have been missed by ELISA alone. Screening of hybridoma supernatants on days 1 and 4 following cloning from semi-solid HAT agarose reveals that FMAT provides a reliable method for screening hybridoma clones to purified protein toxins. The homogeneous FMAT utilizes far less reagent (antigen and hybridoma supernatant) allowing for simultaneous screening against multiple serovariant antigens early in the hybridoma cloning cycle. This reduces costs for reagents and labour by lowering numbers of clones being maintained with undesired specificity. Furthermore, this assay easily accommodates replicate screening which facilitates identification of cross-reactivity to neurotoxin serotypes, thus readily identifying mAb to serovariant antigens. These findings have broad application in accelerating mAb development to serovariant cell-surface or bead bound targets without arraying devices. In summary, FMAT provides a reliable method for the screening of mAbs against C. botulinum neurotoxins.
Assuntos
Anticorpos Antibacterianos/análise , Anticorpos Monoclonais/análise , Toxinas Botulínicas/imunologia , Imunoensaio , Neurotoxinas/imunologia , Animais , Anticorpos Antibacterianos/imunologia , Anticorpos Monoclonais/imunologia , Especificidade de Anticorpos , Ensaio de Imunoadsorção Enzimática , Feminino , Hibridomas , Camundongos , Camundongos Endogâmicos BALB CRESUMO
In the spring of 2003, the Public Health Agency of Canada (then, Health Canada) partnered with several provincial/territorial and regional public health stakeholders to improve pan-Canadian public health surveillance, communications and response through the application of new technologies. This resulted in the creation of the Canadian Network for Public Health Intelligence (CNPHI), a comprehensive framework of applications and resources designed to fill critical gaps in Canada's national public health infostructure. Over the past four years, the CNPHI has evolved into Canada's only pan-Canadian public health information management system. With over 2000 registered users, the current CNPHI environment consists of more than 30 integrated applications and systems that can be loosely categorized into four functional groups: data exchange; data analysis and integration; communication, collaboration and coordination; and knowledge management. Despite poor data repositories, legacy information management systems, and the lack of standards and agreements, the CNPHI has demonstrated that much can be accomplished in these areas. Over the next decade, significant barriers impeding additional advances will be bridged through the implementation of the Electronic Health Record, and through ongoing efforts to address gaps in standards, and data- and information-sharing agreements. Together with new technologies coming on-line, opportunities to further enhance public health surveillance and response will be limited only by one's imagination.
RESUMO
Antibodies against the protective antigen (PA) of Bacillus anthracis play a key role in response to infection by this important pathogen. The aim of this study was to produce and characterize monoclonal antibodies (mAbs) specific for PA and to identify novel neutralizing epitopes. Three murine mAbs with high specificity and nanomolar affinity for B. anthracis recombinant protective antigen (rPA) were produced and characterized. Western immunoblot analysis, coupled with epitope mapping using overlapping synthetic peptides, revealed that these mAbs recognize a linear epitope within domain 2 of rPA. Neutralization assays demonstrate that these mAbs effectively neutralize lethal toxin in vitro.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Bactérias/imunologia , Bacillus anthracis/imunologia , Toxinas Bacterianas/imunologia , Animais , Anticorpos Monoclonais/biossíntese , Sítios de Ligação de Anticorpos , Mapeamento de Epitopos , Epitopos , Camundongos , Camundongos Endogâmicos BALB C , Testes de Neutralização , Estrutura Terciária de ProteínaRESUMO
BACKGROUND: In the field of clinical mycobacteriology, Mycobacterium tuberculosis (MTB) can be a difficult organism to manipulate due to the restrictive environment of a containment level 3 (CL3) laboratory. Tests for rapid diagnostic work involving smears and molecular methods do not require CL3 practices after the organism has been rendered non-viable. While it has been assumed that after organism deactivation these techniques can be performed outside of a CL3, no conclusive study has consistently confirmed that the organisms are noninfectious after the theoretical 'deactivation' steps. Previous studies have shown that initial steps (such as heating/chemical fixation) may not consistently kill MTB organisms. METHODS: An inclusive viability study (n = 226) was undertaken to determine at which point handling of culture extraction materials does not necessitate a CL3 environment. Four different laboratory protocols tested for viability included: standard DNA extractions for IS6110 fingerprinting, crude DNA preparations for PCR by boiling and mechanical lysis, protein extractions, and smear preparations. For each protocol, laboratory staff planted a proportion of the resulting material to Bactec 12B medium that was observed for growth for 8 weeks. RESULTS: Of the 208 isolates initially tested, 21 samples grew within the 8-week period. Sixteen (7.7%) of these yielded positive results for MTB that included samples of: deactivated culture resuspensions exposed to 80 degrees C for 20 minutes, smear preparations and protein extractions. Test procedures were consequently modified and tested again (n = 18), resulting in 0% viability. CONCLUSIONS: This study demonstrates that it cannot be assumed that conventional practices (i.e. smear preparation) or extraction techniques render the organism non-viable. All methodologies, new and existing, should be examined by individual laboratories to validate the safe removal of material derived from MTB to the outside of a CL3 laboratory. This process is vital to establish in house biosafety-validated practices with the aim of protecting laboratory workers conducting these procedures.
Assuntos
Técnicas de Laboratório Clínico/normas , Contenção de Riscos Biológicos/normas , Viabilidade Microbiana , Mycobacterium tuberculosis , Contenção de Riscos Biológicos/métodos , Impressões Digitais de DNA , DNA Bacteriano/isolamento & purificação , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Saúde Ocupacional , Polimorfismo de Fragmento de Restrição , Medição de RiscoRESUMO
Sequencing of the 16S ribosomal DNA (rDNA) for identification of nontuberculous mycobacteria (NTM) has contributed to the establishment of more than 35 new species during the last decade. Increasingly, NTM are accepted as potential or proven pathogens. We identified, by 16S rDNA sequence analysis, slowly growing NTM isolates negative by AccuProbe (GenProbe, San Diego, CA) that previously were identified by using conventional biochemical techniques, to determine the accuracy of reporting AccuProbe-negative NTM prior to sequence-based identification. Of 82 strains, 30 were deemed novel. An attempt was made to determine the clinical importance of previously misidentified novel species. Clinical cases are described for a number of strains previously identified as Mycobacterium terrae complex, Mycobacterium scrofulaceum, and Mycobacterium avium complex. As sequence-based identification methods become more commonplace in clinical microbiology laboratories, there is a need to understand the significance of previously undescribed species, which often mimic and subsequently are identified as well-established species.
Assuntos
Mycobacterium/isolamento & purificação , RNA Ribossômico 16S/análise , Ribotipagem/métodos , Adulto , Idoso , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Mycobacterium/classificação , Mycobacterium/genética , Infecções por Mycobacterium/diagnóstico , RNA Ribossômico 16S/genética , Reprodutibilidade dos TestesRESUMO
Continuous monitoring systems for the detection of Mycobacterium tuberculosis are reported to have higher contamination rates than traditional radiometric technologies. Multiple decontamination methods have recently been reported in an attempt to optimize contamination rates for these systems. In this study, several decontamination methods for sputum were evaluated using viable colony count and flow cytometry. The decontamination protocols evaluated include N-Acetyl-L-Cysteine-Sodium Hydroxide (NALC-NaOH), modified Petroffs's method, and the Yamane procedure. Several parameters of the NALC-NaOH method were analyzed including final NaOH concentrations of 0.5-3%, NaOH exposure times of 0-30 min, and variations in resuspension media for the resultant pellet. All decontamination methods were performed on pooled and sterilized sputum seeded separately with either a mixture of common contaminating bacteria or M. tuberculosis H37Ra. Viability of organisms following decontamination was assessed by both colony counts and flow cytometric analysis. Flow cytometry viability assays utilized a combination of viability dyes and reference beads to determine viable organism concentrations. The results indicated that no decontamination method was clearly superior, however a concentration of 1-2% NaOH and an increase in the time of NaOH exposure to 30 min will effectively kill contaminating bacteria without significantly affecting the viability of M. tuberculosis H37Ra. While flow cytometry viability analysis did not directly correspond to viable colony counts, it was a useful tool for rapid viability analysis M. tuberculosis.
Assuntos
Citometria de Fluxo/métodos , Mycobacterium tuberculosis/isolamento & purificação , Hidróxido de Sódio/farmacologia , Escarro/microbiologia , Análise de Variância , Técnicas Bacteriológicas , Contagem de Colônia Microbiana , Humanos , Probabilidade , Sensibilidade e Especificidade , Manejo de Espécimes , Tuberculose Pulmonar/diagnósticoRESUMO
There is a global need to elucidate protective antigens expressed by the SARS-coronavirus (SARS-CoV). Monoclonal antibody reagents that recognise specific antigens on SARS-CoV are needed urgently. In this report, the development and immunochemical characterisation of a panel of murine monoclonal antibodies (mAbs) against the SARS-CoV is presented, based upon their specificity, binding requirements, and biological activity. Initial screening by ELISA, using highly purified virus as the coating antigen, resulted in the selection of 103 mAbs to the SARS virus. Subsequent screening steps reduced this panel to seventeen IgG mAbs. A single mAb, F26G15, is specific for the nucleoprotein as seen in Western immunoblot while five other mAbs react with the Spike protein. Two of these Spike-specific mAbs demonstrate the ability to neutralise SARS-CoV in vitro while another four Western immunoblot-negative mAbs also neutralise the virus. The utility of these mAbs for diagnostic development is demonstrated. Antibody from convalescent SARS patients, but not normal human serum, is also shown to specifically compete off binding of mAbs to whole SARS-CoV. These studies highlight the importance of using standardised assays and reagents. These mAbs will be useful for the development of diagnostic tests, studies of SARS-CoV pathogenesis and vaccine development.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Testes de Neutralização , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/imunologia , Animais , Antígenos Virais/imunologia , Western Blotting , Chlorocebus aethiops , Ensaio de Imunoadsorção Enzimática , Epitopos , Imunoglobulina G/análise , Imunoglobulina G/imunologia , Glicoproteínas de Membrana/imunologia , Camundongos , Nucleoproteínas/imunologia , Conformação Proteica , Glicoproteína da Espícula de Coronavírus , Células Vero , Proteínas do Envelope Viral/imunologiaRESUMO
BACKGROUND: To describe the demographic and geographic distribution of tuberculosis (TB) in Manitoba, thus determining risk factors associated with clustering and higher incidence rates in distinct subpopulations. METHODS: Data from the Manitoba TB Registry was compiled to generate a database on 855 patients with tuberculosis and their contacts from 1992-1999. Recovered isolates of M. tuberculosis were typed by IS6110 restriction fragment length polymorphisms. Bivariate and multivariate logistic regression models were used to identify risk factors involved in clustering. RESULTS: A trend to clustering was observed among the Canadian-born treaty Aboriginal subgroup in contrast to the foreign-born. The dominant type, designated fingerprint type 1, accounts for 25.8% of total cases and 75.3% of treaty Aboriginal cases. Among type 1 patients residing in urban areas, 98.9% lived in Winnipeg. In rural areas, 92.8% lived on Aboriginal reserves. Statistical models revealed that significant risk factors for acquiring clustered tuberculosis are gender, age, ethnic origin and residence. Those at increased risk are: males (p < 0.05); those under age 65 (p < 0.01 for each age subgroup); treaty Aboriginals (p < 0.001), and those living on reserve land (p < 0.001). CONCLUSION: Molecular typing of isolates in conjunction with contact tracing data supports the notion of the largest ongoing transmission of a single strain of TB within the treaty-status population of Canada recorded to date. This data demonstrates the necessity of continued surveillance of countries with low prevalence of the disease in order to determine and target high-risk populations for concentrated prevention and control measures.
Assuntos
Mycobacterium tuberculosis/isolamento & purificação , Tuberculose/epidemiologia , Adolescente , Adulto , Idoso , Técnicas de Tipagem Bacteriana , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Manitoba/epidemiologia , Pessoa de Meia-Idade , Epidemiologia Molecular , Mycobacterium tuberculosis/genética , Fatores de Risco , Tuberculose/transmissãoRESUMO
STUDY OBJECTIVE: To determine the frequency of analgesic use in children (5 to 17 years inclusive) who present to a pediatric emergency department with acute abdominal pain. METHODS: A retrospective medical record review of patients presenting to a children's hospital over a 1-year period with a chief complaint of abdominal pain and subsequently referred to the pediatric surgical service. The records were reviewed to determine emergency department analgesic use, patient disposition, and laparotomy rate. RESULTS: Two hundred ninety patients met our inclusion criteria. Of the patients seen initially by emergency physicians, 14.3% received analgesics, while those seen directly by the surgical service received analgesia 15.4% of the time. The laparotomy rate for the 290 patients was 46.6%. CONCLUSIONS: Analgesic use in children who present to the emergency department with acute abdominal pain and require a surgical consultation was very low, although half required a laparotomy. Prospective studies are needed to determine the efficacy and safety of analgesic use in this setting.
Assuntos
Dor Abdominal/tratamento farmacológico , Analgésicos/uso terapêutico , Doença Aguda , Adolescente , Criança , Pré-Escolar , Uso de Medicamentos/estatística & dados numéricos , Tratamento de Emergência/estatística & dados numéricos , Feminino , Humanos , Masculino , Estudos RetrospectivosRESUMO
Bacteremia due to Mycobacterium neoaurum, a rapidly growing mycobacterium, is described in a diabetic woman on hemodialysis. This is the first reported case of M neoaurum bacteremia in Canada. The organism initially grew on standard BacT/Alert SA aerobic blood cultures, and was subsequently positively identified using 16S rRNA sequence analysis. The present case serves to reinforce the need for a high index of clinical suspicion of infections caused by unusual microorganisms in the context of an immunocompromised host.
RESUMO
The use of telepathology for clinical applications in Canada has steadily become more attractive over the last 10 years, driven largely by its potential to provide rapid pathology consulting services throughout the country regardless of the location of a particular institution. Based on this trend, the president of the Canadian Association of Pathologists asked a working group consisting of pathologists, technologists, and healthcare administrators from across Canada to oversee the development of guidelines to provide Canadian pathologists with basic information on how to implement and use this technology. The guidelines were systematically developed, based on available medical literature and the clinical experience of early adopters of telepathology in Canada. While there are many different modalities and applications of telepathology, this document focuses specifically on whole-slide imaging as applied to intraoperative pathology consultation (frozen section), primary diagnosis, expert or second opinions and quality assurance activities. Applications such as hematopathology, microbiology, tumour boards, education, research and technical and/or standard-related issues are not covered.
Assuntos
Bacteriemia/microbiologia , Infecções por Mycobacterium não Tuberculosas/microbiologia , Micobactérias não Tuberculosas/classificação , DNA Espaçador Ribossômico/genética , Humanos , Malaui , Micobactérias não Tuberculosas/genética , Filogenia , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Especificidade da Espécie , Tanzânia , Tailândia , Estados UnidosRESUMO
Vaccination with anthrax vaccine adsorbed (AVA) results in the production of protective antigen (PA) specific antibodies, which play an important protective role against anthrax toxins. Analyzing the specificity of serum antibodies generated in response to AVA vaccination can provide insight into the mechanisms of protective immunity against this important pathogen. The goal of this study was to develop a competitive enzyme linked immunosorbent assay (cELISA) to test human immune serum for antibodies specific for a known lethal toxin neutralizing epitope in PA. PA-specific antibodies in sera from individuals who received the six-dose AVA vaccine series competed for binding to immobilized PA with monoclonal antibody F20G75, which binds to a linear epitope in domain 2 of PA and neutralizes lethal toxin activity in vitro. These results suggest that antibodies in human AVA vaccinee serum recognize the same epitope as F20G75, or one in close proximity to it, and may serve a protective role against anthrax lethal toxin. This assay may be used for serological confirmation of successful immunization against anthrax and for the identification of antibodies in human vaccinee serum that recognize protective epitopes on PA.
Assuntos
Vacinas contra Antraz/imunologia , Anticorpos Antibacterianos/sangue , Especificidade de Anticorpos/imunologia , Antígenos de Bactérias , Toxinas Bacterianas , Ensaio de Imunoadsorção Enzimática/métodos , Mapeamento de Epitopos/métodos , Sequência de Aminoácidos , Vacinas contra Antraz/administração & dosagem , Anticorpos Monoclonais/genética , Antígenos de Bactérias/imunologia , Toxinas Bacterianas/imunologia , Ligação Competitiva , Humanos , Dados de Sequência Molecular , Peptídeos/genética , Estados UnidosRESUMO
Having the capacity to detect and identify pathogens that can be employed in a bioterror attack is critical from both a public health and defence perspective. Immunodiagnostic assays are useful tools for enhancing such detection capabilities. In order to develop an immunodiagnostic assay for the detection of Francisella tularensis, a murine monoclonal antibody (MAb) was developed, using the live vaccine strain (LVS) of F. tularensis as the inoculating antigen. A single MAb, F94G2-1, which is specific for the lipopolysaccharide (LPS) of this bacterium was developed and characterized. An indirect ELISA using purified LPS was effective in determining reactivity of the MAb against its target. An immunodotblot and a manually printed antigen microarray were also tested as suitable detection methods. Both assays showed that MAb F94G2-1 has excellent specificity for F. tularensis LPS and demonstrate the utility of using the same MAb in a variety of immunodiagnostic applications.
Assuntos
Anticorpos Monoclonais/biossíntese , Francisella tularensis/imunologia , Lipopolissacarídeos/imunologia , Animais , Anticorpos Monoclonais/isolamento & purificação , Especificidade de Anticorpos , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/metabolismo , Lipopolissacarídeos/metabolismo , Camundongos , Camundongos Endogâmicos BALB CRESUMO
OBJECTIVES: The objectives of this study were to determine whether the administration of morphine to children with acute abdominal pain would impede the diagnosis of appendicitis and to determine the efficacy of morphine in relieving the pain. METHODS: This was a double-blind, randomized, placebo-controlled trial involving 5- to 16-year-old children who presented to the emergency department of a children's hospital with a chief complaint of acute abdominal pain that was thought by the pediatric emergency attending physician to require a surgical consultation. Subjects were randomized to receive intravenously administered morphine or normal saline solution. Clinical data and the emergency physician's confidence in his or her clinical diagnosis (0-100%) were recorded systematically with a standardized form. This was repeated 15 minutes after administration of the study medication. The surgeon assessed the child within 1 hour and completed a similar data collection sheet. Pain was assessed, with a color analog scale, before and after study medication administration. Each subject was monitored for 2 weeks after enrollment. RESULTS: One hundred eight children were enrolled; 52 received morphine and 56 received a placebo saline solution. There were no differences between groups in demographic variables or the degree of pain. There were no differences between groups in the diagnoses of appendicitis or perforated appendicitis or the number of children who were observed and then underwent laparotomy. The reduction in the mean pain score was significantly greater in the morphine group (2.2 vs 1.2 cm). The emergency physicians' and surgeons' confidence in their diagnoses was not affected by the administration of morphine. CONCLUSIONS: Our data show that morphine effectively reduces the intensity of pain among children with acute abdominal pain and morphine does not seem to impede the diagnosis of appendicitis.
Assuntos
Abdome Agudo/terapia , Analgésicos Opioides/administração & dosagem , Apendicite/diagnóstico , Morfina/administração & dosagem , Abdome Agudo/etiologia , Adolescente , Apendicite/complicações , Apendicite/cirurgia , Criança , Pré-Escolar , Erros de Diagnóstico , Método Duplo-Cego , Serviço Hospitalar de Emergência , Feminino , Humanos , Masculino , Medição da DorRESUMO
STUDY OBJECTIVE: We compare oral dexamethasone with placebo for the relief of pain in children with acute pharyngitis. METHODS: We performed a prospective, randomized, double-blind, placebo-controlled trial of children aged 5 to 16 years who presented to the emergency department with acute pharyngitis. Children rated their pain on a standardized color analog scale and had a rapid streptococcal antigen detection test performed to determine group assignment. Children were randomized to dexamethasone (0.6 mg/kg, maximum dose 10 mg) or placebo. Blinded research assistants called all families daily to determine pain scores until the point of complete pain relief. The primary outcome measures were the time to clinically significant pain relief and the time to complete pain relief. RESULTS: A total of 184 children were enrolled in the study. There were 85 children in the antigen-positive group, of whom 45 were randomized to dexamethasone and 40 to placebo. In children with group A beta-hemolytic streptococcal pharyngitis, the median time to clinically significant pain relief was 6 hours in the dexamethasone group versus 11.5 hours in the placebo group (P =.02; effect size of 5.5 hours with 95% confidence interval [CI] of 1.0 and 10.0 hours), and the time to complete pain relief was similar (36 hours for placebo versus 40 hours for dexamethasone, P =.86; effect size of 4.0 hours with 95% CI of -9.3 and 17.3 hours) in the placebo group. There were 99 children enrolled in the antigen-negative group, of whom 47 received dexamethasone and 52 received placebo. In this group, the median time to clinically significant pain relief was 13 hours in the dexamethasone group versus 9 hours in the placebo group (P =.32; effect size of 4 hours with 95% CI of -2 and 10 hours), and the time to complete pain relief was similar (48 hours for placebo versus 50 hours for dexamethasone, P =.61; effect size of 2 hours with 95% CI of -11.8 and 15.8 hours). CONCLUSION: For all children with acute pharyngitis, oral dexamethasone does not decrease the time to onset of clinically significant pain relief or time to complete pain relief. However, in the subset of children with positive antigen detection test results, there is a statistically significant improvement in time to onset of pain relief, but it is of marginal clinical importance.
Assuntos
Anti-Inflamatórios/uso terapêutico , Dexametasona/uso terapêutico , Dor/tratamento farmacológico , Faringite/tratamento farmacológico , Doença Aguda , Administração Oral , Adolescente , Anti-Inflamatórios/administração & dosagem , Distribuição de Qui-Quadrado , Criança , Pré-Escolar , Dexametasona/administração & dosagem , Método Duplo-Cego , Serviço Hospitalar de Emergência , Feminino , Humanos , Masculino , Dor/etiologia , Faringite/complicações , Faringite/microbiologia , Placebos , Estudos Prospectivos , Estatísticas não Paramétricas , Resultado do TratamentoRESUMO
Eleven strains of a rapidly growing mycobacterium were isolated from patient specimens originating from various regions of the province of Ontario, Canada, over a 2-year period. Unique high-performance liquid chromatography (HPLC) and PCR-restriction enzyme pattern analysis (PRA) profiles initially suggested a new Mycobacterium species, while sequencing of the 16S rRNA gene revealed a sequence match with Mycobacterium sp. strain MCRO 17 (GenBank accession no. X93028), an isolate determined to be unique which is to date uncharacterized, and also a close similarity to M. elephantis (GenBank accession no. AJ010747), with six base pair variations. A complete biochemical profile of these isolates revealed a species of mycobacteria with phenotypic characteristics similar to those of M. flavescens. HPLC, PRA, and 16S rRNA sequencing of strain M. elephantis DSM 44368(T) and result comparisons with the clinical isolates revealed that these strains were in fact M. elephantis, a newly described species isolated from an elephant. All strains were isolated from human samples, 10 from sputum and 1 from an axillary lymph node.
Assuntos
Técnicas de Tipagem Bacteriana , Infecções por Mycobacterium/microbiologia , Mycobacterium/classificação , Mycobacterium/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Bases , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mycobacterium/isolamento & purificação , Ácidos Micólicos/análise , Fenótipo , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética , Mapeamento por Restrição/métodos , Análise de Sequência de DNARESUMO
A strain of a novel non-chromogenic mycobacterium was isolated from synovial tissue from a 68-year-old female with bursitis of her right elbow. The slowly growing strain had a unique PCR-restriction enzyme analysis (PRA) profile of the hsp65 gene and 16S rRNA gene sequence in comparison with other mycobacterium species. The most closely related species, as determined by 16S rRNA gene sequence analysis, are Mycobacterium malmoense, Mycobacterium marinum, Mycobacterium ulcerans and members of the Mycobacterium tuberculosis complex. The HPLC and biochemical profiles resembled those of Mycobacterium gastri, although differences were noted in the peak-height ratio of the HPLC pattern and the nitrate and pyrazinamidase tests. On the basis of PRA, HPLC, biochemical and 16S rRNA gene sequence analyses, the name Mycobacterium lacus sp. nov. is proposed for this potential pathogen. The type strain is strain NRCM 00-255(T) (= ATCC BAA-323(T) = DSM 44577(T)).
Assuntos
Proteínas de Bactérias , Mycobacterium/classificação , Idoso , Sequência de Bases , Bursite/microbiologia , Chaperonina 60 , Chaperoninas/genética , DNA Bacteriano/genética , DNA Ribossômico/genética , Farmacorresistência Bacteriana , Feminino , Genes Bacterianos , Humanos , Dados de Sequência Molecular , Mycobacterium/genética , Mycobacterium/isolamento & purificação , Mycobacterium/metabolismo , Infecções por Mycobacterium/microbiologia , Ácidos Micólicos/análise , Fenótipo , Filogenia , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Homologia de Sequência do Ácido Nucleico , Terminologia como AssuntoRESUMO
Molecular techniques are playing an important role in the diagnosis of nontuberculous mycobacterial infections. This case report describes a chronic soft tissue infection in an immunocompetent patient caused by a previously undescribed pigmented, rapidly growing Mycobacterium species, emphasizing the importance of clinical suspicion and effective laboratory techniques in the diagnosis and treatment of infection.