RESUMO
A semi-quantitative multiplex PCR assay for the diagnosis of bacterial vaginosis (BV) was evaluated in a prospective study in a population of Dutch women with complaints of abnormal vaginal discharge. The PCR targets Gardnerella vaginalis, Atopobium vaginae, Megasphaera phylotype 1, Lactobacillus crispatus and Lactobacillus iners. Together with a short questionnaire, a vaginal swab for PCR and vaginal smear for microscopy were taken by their general practitioner or gynaecologist. Data from 151 women (median age 32) were available. Nugent Score (NS) was used to classify the samples and 83 samples were classified as normal (NS 0-3), 13 as intermediate (NS 4-6), and 55 as bacterial vaginosis (NS 7-10). In women with a NS of 7-10, PCR detected Gardnerella vaginalis, Atopobium vaginae and Megasphaera phylotype 1 in respectively, 96 %, 87 % and 60 %, whereas in women with a NS of 1-3 these species were detected in 27 %, 6 % and 2 % (P <0.001). A ratio of Lactobacillus crispatus over Lactobacillus iners of <1 (as calculated from the quantification cycle value (Cq)) was present in women with a NS of 7-10 in 66 % versus 33 % in women with a NS of 1-3 (P <0.001). The BV-PCR displayed a sensitivity of 92 % and specificity of 96 % with a positive predictive value of 94 % and a negative predictive value of 95 %. The Lactobacillus-index improved the correct classification of samples where only one of the other bacterial species was detected. Compared to the Nugent Score this multiplex qPCR offers a convenient tool for performing observer independent diagnosis of BV.
Assuntos
Reação em Cadeia da Polimerase em Tempo Real/métodos , Vagina/microbiologia , Descarga Vaginal/diagnóstico , Vaginose Bacteriana/diagnóstico , Adolescente , Adulto , DNA Bacteriano/genética , DNA Fúngico/genética , Feminino , Humanos , Pessoa de Meia-Idade , Países Baixos , Estudos Prospectivos , Inquéritos e Questionários , Esfregaço Vaginal , Adulto JovemRESUMO
Starting August 2012, an increase in Cryptosporidium infections was reported in the Netherlands, the United Kingdom and Germany. It represented a 1.8 to 4.9-fold increase compared to previous years. Most samples were C. hominis IbA10G2. A casecontrol study was performed in the Netherlands but did not identify an endemic source. A casecase study in the north of England found travel abroad to be the most common risk factor.
Assuntos
Criptosporidiose/epidemiologia , Cryptosporidium/genética , Fezes/parasitologia , Adolescente , Adulto , Distribuição por Idade , Estudos de Casos e Controles , Criança , Criptosporidiose/parasitologia , Cryptosporidium/classificação , Cryptosporidium/isolamento & purificação , Feminino , Genótipo , Alemanha/epidemiologia , Humanos , Técnicas Imunoenzimáticas , Incidência , Masculino , Países Baixos/epidemiologia , Reação em Cadeia da Polimerase , Fatores de Risco , Estações do Ano , Distribuição por Sexo , Reino Unido/epidemiologia , Adulto JovemRESUMO
BACKGROUND: Dientamoeba fragilis in children has been associated with gastrointestinal symptoms, like abdominal pain and diarrhea. The mechanism underlying these symptoms in children with D. fragilis remains unclear. We hypothesized that concomitant microbial alterations, which have been described in other parasitic infections, may be associated with gastrointestinal symptoms in D. fragilis. METHODS: In this case-control study performed in 2 centers, 19 children referred to a pediatrician because of gastrointestinal symptoms and with a positive fecal PCR for D. fragilis were included as cases. We included 19 healthy children as controls and matched for age and gender, selected from an existing cohort of 63 children. A PCR for D. fragilis was performed on fecal samples of the 19 controls to assess D. fragilis carriership in this asymptomatic group. Microbiota was analyzed with the IS-pro technique, and the intestinal microbiota composition and diversity were compared between the 2 groups. RESULTS: Microbiota of children with D. fragilis and gastrointestinal symptoms did not significantly differ in terms of composition and diversity compared with controls, both on phylum and species level. In the asymptomatic controls, a positive fecal PCR for D. fragilis was found in 16 of 19 (84.2%). CONCLUSION: Intestinal microbiota does not seem to play a key role in the presence of clinical symptoms in children with D. fragilis. The pathogenicity of D. fragilis and pathophysiologic pathways underlying the development of gastrointestinal symptoms remains yet to be clarified.
Assuntos
Dientamoeba/genética , Dientamebíase/parasitologia , Gastroenteropatias/parasitologia , Microbioma Gastrointestinal/genética , Dor Abdominal , Estudos de Casos e Controles , Criança , Pré-Escolar , Estudos de Coortes , Diarreia/parasitologia , Dientamoeba/patogenicidade , Fezes/parasitologia , Variação Genética , HumanosRESUMO
OBJECTIVE: Short-course aminoglycosides as adjunctive empirical therapy to ß-lactams in patients with a clinical suspicion of sepsis are used to broaden antibiotic susceptibility coverage and to enhance bacterial killing. We quantified the impact of this approach on 30-day mortality in a subset of sepsis patients with a Gram-negative bloodstream infection. METHODS: From a prospective cohort study conducted in seven hospitals in the Netherlands between June 2013 and November 2015, we selected all patients with Gram-negative bloodstream infection (GN-BSI). Short-course aminoglycoside therapy was defined as tobramycin, gentamicin or amikacin initiated within a 48-hour time window around blood-culture obtainment, and prescribed for a maximum of 2 days. The outcome of interest was 30-day all-cause mortality. Confounders were selected a priori for adjustment using a propensity score analysis with inverse probability weighting. RESULTS: A total of 626 individuals with GN-BSI who received ß-lactams were included; 156 (24.9%) also received aminoglycosides for a median of 1 day. Patients receiving aminoglycosides more often had septic shock (31/156, 19.9% versus 34/470, 7.2%) and had an eight-fold lower risk of inappropriate treatment (3/156, 1.9% versus 69/470, 14.7%). Thirty-day mortality was 17.3% (27/156) and 13.6% (64/470) for patients receiving and not receiving aminoglycosides, respectively; yielding crude and adjusted odds ratios for 30-day mortality for patients treated with aminoglycosides of 1.33 (95% CI 0.80-2.15) and 1.57 (0.84-2.93), respectively. CONCLUSIONS: Short-course adjunctive aminoglycoside treatment as part of empirical therapy with ß-lactam antibiotics in patients with GN-BSI did not result in improved outcomes, despite better antibiotic coverage of pathogens.
Assuntos
Aminoglicosídeos/administração & dosagem , Infecções por Bactérias Gram-Negativas/tratamento farmacológico , Sepse/microbiologia , beta-Lactamas/administração & dosagem , Idoso , Idoso de 80 Anos ou mais , Aminoglicosídeos/uso terapêutico , Terapia Combinada , Feminino , Infecções por Bactérias Gram-Negativas/mortalidade , Humanos , Masculino , Pessoa de Meia-Idade , Países Baixos , Estudos Prospectivos , Sepse/tratamento farmacológico , Sepse/mortalidade , Análise de Sobrevida , Resultado do Tratamento , beta-Lactamas/uso terapêuticoRESUMO
OBJECTIVES: Current guidelines for the empirical antibiotic treatment predict the presence of third-generation cephalosporin-resistant enterobacterial bacteraemia (3GCR-E-Bac) in case of infection only poorly, thereby increasing unnecessary carbapenem use. We aimed to develop diagnostic scoring systems which can better predict the presence of 3GCR-E-Bac. METHODS: A retrospective nested case-control study was performed that included patients ≥18 years of age from eight Dutch hospitals in whom blood cultures were obtained and intravenous antibiotics were initiated. Each patient with 3GCR-E-Bac was matched to four control infection episodes within the same hospital, based on blood-culture date and onset location (community or hospital). Starting from 32 commonly described clinical risk factors at infection onset, selection strategies were used to derive scoring systems for the probability of community- and hospital-onset 3GCR-E-Bac. RESULTS: 3GCR-E-Bac occurred in 90 of 22 506 (0.4%) community-onset infections and in 82 of 8110 (1.0%) hospital-onset infections, and these cases were matched to 360 community-onset and 328 hospital-onset control episodes. The derived community-onset and hospital-onset scoring systems consisted of six and nine predictors, respectively. With selected score cut-offs, the models identified 3GCR-E-Bac with sensitivity equal to existing guidelines (community-onset: 54.3%; hospital-onset: 81.5%). However, they reduced the proportion of patients classified as at risk for 3GCR-E-Bac (i.e. eligible for empirical carbapenem therapy) with 40% (95%CI 21-56%) and 49% (95%CI 39-58%) in, respectively, community-onset and hospital-onset infections. CONCLUSIONS: These prediction scores for 3GCR-E-Bac, specifically geared towards the initiation of empirical antibiotic treatment, may improve the balance between inappropriate antibiotics and carbapenem overuse.
Assuntos
Antibacterianos/efeitos adversos , Bacteriemia/diagnóstico , Bacteriemia/etiologia , Cefalosporinas/efeitos adversos , Infecções por Enterobacteriaceae/diagnóstico , Enterobacteriaceae/efeitos dos fármacos , Idoso , Antibacterianos/uso terapêutico , Bacteriemia/epidemiologia , Bacteriemia/microbiologia , Estudos de Casos e Controles , Cefalosporinas/uso terapêutico , Infecção Hospitalar/sangue , Infecção Hospitalar/diagnóstico , Infecção Hospitalar/tratamento farmacológico , Infecção Hospitalar/microbiologia , Infecções por Enterobacteriaceae/sangue , Infecções por Enterobacteriaceae/etiologia , Infecções por Enterobacteriaceae/microbiologia , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Estudos Retrospectivos , Fatores de RiscoRESUMO
BACKGROUND: The effects of the implementation of a new Dutch hepatitis B virus (HBV) vaccination strategy (1991) for expatriates on HBV vaccination status and HBV infection prevalence were evaluated in a group of 864 expatriates returning from HBV-endemic areas. METHODS: During a routine medical examination at the participating medical centres Dutch expatriates were asked to complete a questionnaire and to donate a serum sample for HBV testing. Blood was tested for antibodies against the hepatitis B core (anti-HBc) and surface antigens (anti-HBs). The serological data were related to information gathered on aspects of residence, sexual risk behaviour and occupational risks. RESULTS: A significantly higher percentage of expatriates (37%) were vaccinated compared to a previous study in 1987-1989 (14%). However, the percentage of expatriates with HBV infection markers (5%) had not decreased significantly. Moreover, the risk for HBV infection, as determined with a questionnaire, was still affected by well-known risk factors such as homosexual contacts (odds ratio [OR] = 6.6, 95% CI: 1.7-26), more than five casual local partners (OR = 3.6, 95% CI: 1.2-11) and more than five occupational accidents in the last 3 years (OR = 20, 95% CI: 2-187). Detailed analysis of the vaccination status indicated that especially young female expatriates with low risk behaviour (65%) were protected, while older male expatriates with high risk behaviour were less protected (20%). CONCLUSION: We conclude that the new vaccination strategy has resulted in a higher percentage of expatriates protected. However, only a small proportion was reached of those at highest risk for HBV infection.
Assuntos
Vacinas contra Hepatite B/administração & dosagem , Hepatite B/prevenção & controle , Viagem , Vacinação/estatística & dados numéricos , Adulto , África , Doenças Endêmicas/prevenção & controle , Feminino , Pessoal de Saúde/estatística & dados numéricos , Hepatite B/diagnóstico , Hepatite B/epidemiologia , Humanos , América Latina , Masculino , Países Baixos/epidemiologia , Países Baixos/etnologia , Doenças Profissionais/epidemiologia , Doenças Profissionais/prevenção & controle , Vigilância da População , Prevalência , Medição de Risco , Assunção de Riscos , Testes Sorológicos , Parceiros Sexuais , Fatores de TempoRESUMO
From a prospective cohort study on tuberculosis/human immunodeficiency virus (TB/HIV) interaction in Addis Ababa, Ethiopia, drug susceptibility results were available for 94 TB patients (46% HIV-infected). Resistance to one or more drug(s) was detected in 21 (22.3%) and multidrug resistance in five (5.3%) patients. Occurrence of resistance was not related to HIV status or outcome after 24 months of follow-up. However, among HIV-infected TB patients who died during follow-up, survival time in those with a resistant Mycobacterium tuberculosis strain was significantly shorter compared to those with a sensitive strain (6 vs. 13 months). Early detection of drug resistance and timely treatment change can therefore have a positive impact on survival in HIV-infected TB patients.
Assuntos
Antituberculosos/uso terapêutico , Infecções por HIV/complicações , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose Pulmonar/tratamento farmacológico , Adulto , Estudos de Casos e Controles , Farmacorresistência Bacteriana , Etiópia , Feminino , Seguimentos , Infecções por HIV/mortalidade , Humanos , Masculino , Estudos Prospectivos , Fatores de Tempo , Resultado do Tratamento , Tuberculose Pulmonar/mortalidade , Tuberculose Pulmonar/virologia , Adulto JovemRESUMO
The Netherlands Institute of Primary Health Care (NIVEL) has coordinated the activities of a sentinel surveillance network of 43 general practices since 1970. These practices care for 1% of the Dutch population, a sample representative of the national pop
RESUMO
During a three month prospective study, 229 in-patients with fever, admitted to St. Mary's Hospital, Mumias, were examined for bacterial and malarial causes of fever. Blood cultures taken from patients appeared to contain true pathogens in 51 (22%) cases. Nine different bacterial species were identified from positive blood cultures of which four predominated: Salmonella typhi (46%), Streptococcus pneumoniae (19%), Salmonella enteritidis (12%), Salmonella typhimurium (8%). S. enteritidis and S. typhimurium isolates were mostly multi-antibiotic resistant, compared to S. typhi isolates which were relatively susceptible to the antibiotics used in the hospital. Only 70% of the S. pneumoniae isolates were susceptible to penicillin. Among 227 patients in whom a thick blood-film for malaria parasites and HIV serology were performed, only 25 (11%) revealed malaria parasites. HIV-1 antibodies were detected in 51 (22%) patients. Without appropriate laboratory examinations, the majority of the diagnoses would have been missed and no optimal treatment would have been administered. This may increase resistance to antimalarials and antibiotics.
PIP: In developing countries, the presence of infectious diseases is diagnosed mainly on the basis of clinical symptoms and basic laboratory investigations. Such diagnosis, however, can lead to incorrect or overtreatment, resulting in an increased resistance to antibiotics. This study was conducted to characterize bacterial causes of community-acquired sepsis, in relation to malaria and HIV prevalence, and to assess whether isolated bacteria were susceptible to commonly used antibiotics. During the 3-month prospective study, 229 inpatients with fever admitted to St. Mary's Hospital, Mumias, were examined for bacterial and malarial causes of fever. Blood cultures taken from patients seemed to contain true pathogens in 51 (22%) cases. 9 different bacterial species were identified from positive blood cultures, of which the following predominated: Salmonella typhi (46%), Streptococcus pneumoniae (19%), Salmonella enteritidis (12%), and Salmonella typhimurium (8%). The S. enteritidis and S. typhimurium isolates were mainly multi-antibiotic resistant, compared to the S. typhi isolates which were relatively susceptible to the antibiotics used in the hospital. 70% of the S. pneumoniae isolates were susceptible to penicillin. Among the 227 patients upon whom a thick blood-film for malaria parasites and HIV serology were performed, 25 (11%) had malaria parasites. HIV-1 antibodies were detected in 51 (22%) patients. These study findings show how the culture of 1 blood sample and the performance of simple microbiological studies can facilitate the initiation of appropriate treatment with antibiotics.
Assuntos
Técnicas de Laboratório Clínico , Infecções Comunitárias Adquiridas/microbiologia , Febre/microbiologia , Soroprevalência de HIV , HIV-1 , Malária/parasitologia , Sepse/microbiologia , Criança , Pré-Escolar , Infecções Comunitárias Adquiridas/sangue , Infecções Comunitárias Adquiridas/complicações , Resistência Microbiana a Medicamentos , Humanos , Lactente , Quênia , Malária/sangue , Malária/complicações , Estudos Prospectivos , Sepse/sangue , Sepse/complicaçõesRESUMO
OBJECTIVE: To determine the spread of respiratory infection with Mycoplasma pneumoniae in an institute for mentally disabled persons. DESIGN: Descriptive. METHODS: In the period from mid-April to mid-September in a certain year the transmission of M. pneumoniae in the facility was evaluated using questionnaires and laboratory investigations. The laboratory investigations consisted of an M. pneumoniae specific polymerase chain reaction (PCR) on throat swab specimens and detection of antibodies in serum. RESULTS: 21 Residents and 26 staff members from 2/36 units were involved in the initial investigation. 17 Persons had complaints of a (recent) respiratory infection (cough, malaise and fever). In 9 cases an M. pneumoniae infection was confirmed, in 5 cases by PCR and in 4 cases by serology. Two PCR positive persons had only complaints of coughing. During the investigation period 2 more persons were diagnosed with a respiratory infection due to M. pneumoniae. No new cases were found by investigation of contacts outside the facility. CONCLUSION: M. pneumoniae can cause an outbreak of M. pneumoniae respiratory infection in an institute for mentally disabled persons. Rapid detection of this pathogen is possible by PCR and is important for proper antibiotic therapy and epidemic-control measures.
Assuntos
Infecção Hospitalar/epidemiologia , Surtos de Doenças/prevenção & controle , Infecções por Mycoplasma/diagnóstico , Infecções por Mycoplasma/epidemiologia , Pessoas com Deficiência Mental/estatística & dados numéricos , Faringe/microbiologia , Infecções Respiratórias/diagnóstico , Infecções Respiratórias/epidemiologia , Adolescente , Adulto , Busca de Comunicante/métodos , Infecção Hospitalar/diagnóstico , Infecção Hospitalar/transmissão , Transmissão de Doença Infecciosa/prevenção & controle , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Infecções por Mycoplasma/transmissão , Mycoplasma pneumoniae/isolamento & purificação , Países Baixos/epidemiologia , Reação em Cadeia da Polimerase/métodos , Instituições Residenciais/estatística & dados numéricos , Infecções Respiratórias/transmissãoRESUMO
The first signs of influenza activity in the Netherlands during the 1999/2000 influenza season were the isolation of an influenza A (H3N2) virus in week 40 and of two more in week 43 of 1999. From week 50 onwards, a strong increase of the clinical influenza activity was observed which reached its peak in weeks 1 and 2 of 2000 and then rapidly declined. The clinical influenza activity was associated with the isolation of predominantly influenza A (H3N2) viruses. Near the end of the epidemic, influenza A (H1N1) and influenza B viruses were isolated sporadically. The antigenic properties of the influenza A (H3N2) viruses resembled those of the epidemic strains isolated in the previous season and the vaccine strain A/Sydney/5/97. This influenza season, influenza B viruses did not play a significant role and they matched the vaccine strain B/Yamanashi/166/98. In addition, a small number of influenza A (H1N1) viruses were isolated. Some of these viruses resembled the old variant of influenza A (H1N1) viruses, A/Bayern/7/95, whilst others showed a close antigenic relationship with the vaccine strain recommended for the next influenza season, A/New Caledonia/20/99. For the influenza season 2000/'01, it is recommended by the World Health Organization that the vaccines contain the following (or similar) virus strains: A/Moscow/10/99 (H3N2), A/New Caledonia/20/99 (H1N1) and B/Beijing/184/93.
Assuntos
Surtos de Doenças/prevenção & controle , Vírus da Influenza A/isolamento & purificação , Vírus da Influenza B/isolamento & purificação , Vacinas contra Influenza , Influenza Humana/prevenção & controle , Influenza Humana/virologia , Previsões , Humanos , Influenza Humana/epidemiologia , Países Baixos/epidemiologia , Vigilância da PopulaçãoRESUMO
The first indication of influenza activity in the Netherlands in the 1998/'99 season was the isolation of an influenza B virus in week 47 of 1998. In subsequent weeks influenza activity slowly increased, reaching a peak in week 6 of 1999. After a gradual decline for three weeks a second peak was reached in week 8 of 1999. The first wave of influenza activity was primarily caused by influenza B viruses, whereas during the second wave predominantly influenza A viruses of the A/H3N2 subtype were isolated. The antigenic properties of the influenza A viruses resembled those of the viruses isolated in the previous season and the vaccine strain A/Sydney/5/97. The influenza B viruses did not completely match with B/Harbin/7/94 which is most commonly used for vaccine production. The vaccine, however, did provide good protection against the epidemic strains of influenza. This season influenza A/H1N1 viruses did not play a significant part and only a small number of viruses of this subtype were isolated at the end of the season. For the influenza season 1999/2000 it is recommended by the World Health Organization that the vaccines contain the following (or similar) virus strains: A/Sydney/5/97 (H3N2), A/Beijing/262/95 (H1N1) and B/Beijing/184/93.
Assuntos
Vacinas contra Influenza/provisão & distribuição , Influenza Humana/prevenção & controle , Influenza Humana/virologia , Orthomyxoviridae/patogenicidade , Guias como Assunto , Humanos , Influenza Humana/epidemiologia , Países Baixos/epidemiologia , Vigilância da População , Vacinas Combinadas/provisão & distribuição , Organização Mundial da SaúdeRESUMO
The 1997/'98 influenza season in the Netherlands was marked by influenza A/H3N2 activity which never reached a true epidemic level. There was no real peak activity either but a prolonged period of increased activity of approximately eight weeks with a maximum in week 13, when sentinel physicians reported 16.6 cases of influenza-like illness per 10,000 inhabitants. It was not until week 18 of 1998 that the influenza activity declined to baseline levels. During the season, almost exclusively influenza A/H3N2 viruses were isolated, of which the majority resembled the new strain influenza A/Sydney/5/97 (H3N2). Further analysis of these variant viruses revealed that, although there was some cross-reactivity with the vaccine strain (A/Nanchang/933/95), no optimal protection could be expected to be induced by the vaccine. Antigenic characterisation of the sporadic influenza A/H1N1 and influenza B viruses showed that these were related to the vaccine strains. As a result of these findings, the World Health Organization (WHO) recommended to change the H3N2 strain in the influenza vaccine for the season 1998/'99 to an influenza A/Sydney/5/97(H3N2)-like strain. Based on epidemiological data from other countries, it was also decided to change the influenza A/H1N1 component to an influenza A/Beijing/262/95 (H1N1)-like strain.
Assuntos
Surtos de Doenças/prevenção & controle , Vacinas contra Influenza/normas , Influenza Humana/epidemiologia , Vigilância da População/métodos , Feminino , Previsões , Humanos , Masculino , Países Baixos/epidemiologia , Orthomyxoviridae/patogenicidade , Vacinas Combinadas , Organização Mundial da SaúdeRESUMO
The performance of a multiplex real-time PCR for the detection of Blastocystis, Dientamoeba fragilis, Giardia lamblia, Cryptosporidium species and Entamoeba species in faecal samples was evaluated in an observational prospective study. Paediatric patients (0-18 years) presenting with gastrointestinal symptoms and suspected of having enteroparasitic disease were included. A questionnaire on gastrointestinal symptoms and the chosen treatment was completed at the start of the study and after 6 weeks. Of 163 paediatric patients (mean age, 7.8 years), 114 (70%) had a PCR-positive faecal sample. D. fragilis was detected most frequently, in 101 patients, followed by Blastocystis in 49. In faecal samples of 47 patients, more than one protozoan was detected, mainly the combination of D. fragilis and Blastocystis. Reported gastrointestinal symptoms were abdominal pain (78%), nausea (30%), and altered bowel habits (28%). Eighty-nine of the PCR-positive patients were treated with antibiotics. A significant reduction in abdominal pain was observed both in treated and in untreated patients. This study demonstrated that multiplex real-time PCR detects a high percentage of intestinal protozoa in paediatric patients with gastrointestinal symptoms. However, interpretation and determination of the clinical relevance of a positive PCR result in this population are still difficult.
Assuntos
Fezes/parasitologia , Enteropatias Parasitárias/diagnóstico , Técnicas de Diagnóstico Molecular/métodos , Reação em Cadeia da Polimerase Multiplex/métodos , Parasitos/classificação , Parasitos/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/métodos , Adolescente , Animais , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Enteropatias Parasitárias/parasitologia , Masculino , Parasitos/genética , Parasitologia/métodosRESUMO
Detection of intestinal protozoa by PCR methods has been described as being sensitive and specific, and as improving the diagnostic yield. Here we present the outcome of the transition from microscopy to molecular screening for detection of a select group of intestinal protozoa in faeces in our laboratory. Introduction of molecular screening for intestinal protozoa resulted in higher sensitivity, reduced hands-on-time, reduced time-to-results, leading to improved diagnostic efficiency.
Assuntos
Enteropatias Parasitárias/diagnóstico , Microscopia/métodos , Técnicas de Diagnóstico Molecular/métodos , Fezes/parasitologia , Humanos , Programas de Rastreamento/métodos , Países Baixos , Sensibilidade e Especificidade , Fatores de TempoRESUMO
Three methods for genotyping of Mycoplasma pneumoniae clinical isolates were applied to 2 reference strains and 21 clinical isolates. By a modified restriction fragment length polymorphism (RFLP) analysis of PCR products of the M. pneumoniae cytadhesin P1 gene, 5 subtypes were discriminated among 13 P1 type 1 strains and 3 subtypes were discriminated among 8 P1 type 2 strains. Sequence analysis of the 16S-23S rRNA gene spacer region and part of the 23S rRNA gene revealed one nucleotide difference in the intergenic spacer region in 3 of the 21 isolates. In the 23S rRNA gene sequence of the 8 P1 type 2 strains an extra adenosine was present, but it was absent from the 13 P1 type 1 strains. On the basis of M. pneumoniae genome sequence data, primers were designed to amplify large interrepeat fragments by long PCR, and these fragments were subsequently analyzed by RFLP analysis. Only two types, long PCR types 1 and 2, could be discriminated among the M. pneumoniae isolates. All P1 type 1 strains were assigned to long PCR type 1, and all P1 type 2 strains were assigned to long PCR type 2. These data obtained by three independent typing methods thus confirm the existence of two distinct M. pneumoniae genomic groups but expand the possibility of strain typing on the basis of variations within their P1 genes.
Assuntos
Mycoplasma pneumoniae/genética , Pneumonia por Mycoplasma/diagnóstico , Polimorfismo de Fragmento de Restrição , RNA Ribossômico 16S/genética , RNA Ribossômico 23S/genética , Adesinas Bacterianas/genética , Sequência de Bases , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , DNA Ribossômico/genética , Genótipo , Humanos , Mycoplasma pneumoniae/classificação , Mycoplasma pneumoniae/isolamento & purificação , Pneumonia por Mycoplasma/microbiologia , Reação em Cadeia da Polimerase , Sequências Repetitivas de Ácido NucleicoRESUMO
Mycoplasma pneumoniae strains traditionally are divided into two types, based on sequence variation in the P1 gene. Recently, however, we have identified 8 P1 subtypes by restriction fragment length polymorphism analysis. In the present study the P1 gene sequences of three P1 type 1 and two P1 type 2 M. pneumoniae strains were analyzed. A new P1 gene sequence in a type 1 strain with partial similarity to a recently reported variable region in the P1 gene of an M. pneumoniae type 2 strain (T. Kenri, R. Taniguchi, Y. Sasaki, N. Okazaki, M. Narita, K. Izumikawa, M. Umetsu, and T.Sasaki, Infect. Immun. 67:4557-4562, 1999) was identified. In addition, the P1 gene of the type 1 strain contained another region with nucleotide polymorphisms identical to a stretch in the P1 gene of one of our type 2 strains. These findings indicate that recombination between sequences specific for P1 type 1 and type 2 had occurred and that P1 type 1 and type 2 hybrid sequences can be present within the P1 gene of an individual strain. Identical or nearly identical variable P1 gene sequences were present in several repetitive regions outside the P1 gene locus in the genome of M. pneumoniae strain M129, implying recombination as a mechanism for generation of the P1 gene variation. Additionally, in the P1 gene sequences of four of the five strains studied, single-nucleotide polymorphisms different from the previously reported P1 type 1 and 2 characteristic sequences were identified. The polymorphic sites are candidate targets for genotyping of M. pneumoniae by direct sequencing of amplicons from clinical specimens.
Assuntos
Adesinas Bacterianas/genética , Variação Genética , Mycoplasma pneumoniae/genética , Sequência de Aminoácidos , Sequência de Bases , Humanos , Dados de Sequência Molecular , Mycoplasma pneumoniae/classificação , Mycoplasma pneumoniae/metabolismo , Pneumonia por Mycoplasma/microbiologia , Reação em Cadeia da Polimerase , Padrões de Referência , Análise de Sequência de DNARESUMO
OBJECTIVES: To assess the performance of routine syphilis screening during 5 year follow up of Ethiopian factory workers, participating in a cohort study on HIV/AIDS. METHODS: Syphilis serology test results of factory workers, who each donated at least six blood samples were evaluated. Screening in 1997-8 had been performed by the Treponema pallidum particle agglutination (TPPA) assay and in 1999-2001 by the rapid plasma reagin (RPR) test. TPPA had been followed by RPR or RPR by TPPA, in case of a positive screening result. Samples of study subjects showing inconsistent sequential TPPA and/or RPR results were retested independently by three laboratory technicians. RESULTS: A total of 540 cohort participants (8.3% HIV positive at enrollment) donated 4,376 blood samples (mean 8.3 per subject). From 93 of the 176 participants with at least one positive TPPA result during follow up, 152 samples were retested by RPR and/or TPPA. Based on the revised syphilis test results, the 540 cohort participants were classified as having no (70.5%), past (20.6%), prevalent (6.9%), or incident (2.0%) syphilis. The RPR screening test was difficult to interpret and yielded 8.2% biological false positive (BFP) RPR results, or 3.2% if weak positive results were excluded. There was no correlation between HIV infection and BFP RPR reactions. Sample mix-ups were detected in 1.2%. CONCLUSION: Evaluation of routine syphilis screening as performed in a long term cohort study on HIV/AIDS in Ethiopia showed difficulties encountered in syphilis screening programmes such as a high percentage of BFP RPR, inconsistencies in interpretation of the RPR test, and sample mix ups. The findings stress the need to develop a syphilis screening assay that is easy to perform and interpret and to implement quality assurance programmes.
Assuntos
Infecções Oportunistas Relacionadas com a AIDS/diagnóstico , Sorodiagnóstico da Sífilis/normas , Sífilis/diagnóstico , Infecções Oportunistas Relacionadas com a AIDS/complicações , Adulto , Estudos de Coortes , Etiópia , Feminino , Humanos , Masculino , Programas de Rastreamento , Sensibilidade e Especificidade , Sífilis/complicaçõesRESUMO
During a 30-month prospective study in The Netherlands, the distribution of Mycoplasma pneumoniae and respiratory viruses among 1172 patients with acute respiratory infection (ARI) who were treated in the outpatient general practitioner setting was studied. M. pneumoniae, as detected by polymerase chain reaction analysis, was present in 39 (3.3%) patients. The infection rate was similar in all age groups. Nose and throat samples collected from 79 household contacts of M. pneumoniae-positive index patients revealed M. pneumoniae in 12 (15%) cases. The frequency of M. pneumoniae among household contacts of index patients treated with appropriate antibiotics and untreated index patients was similar. Nine of the 12 M. pneumoniae-positive household contacts were <16 years old (P=.02), and 4 (44%) of them did not develop ARI. Apparently, children are a relevant reservoir for M. pneumoniae.