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Use of an oligonucleotide array for laboratory diagnosis of bacteria responsible for acute upper respiratory infections.

ABSTRACT: We developed a diagnostic array of oligonucleotide probes targeting species-specific variable regions of the genes encoding topoisomerases GyrB and ParE of respiratory bacterial pathogens. Suitable broad-range primer sequences were designed based on alignment of gyrB/parE sequences from nine different bacterial species. These species included Corynebacterium diphtheriae, Fusobacterium necrophorum, Haemophilus influenzae, Legionella pneumophila, Moraxella catarrhalis, Mycoplasma pneumoniae, Staphylococcus aureus, Streptococcus pneumoniae, and Streptococcus pyogenes. Specific probe sequences were selected by comparative analysis against the European Bioinformatics Database, as well as gyrB/parE sequences generated for this study. To verify specificity, at least six initial oligonucleotide probe sequences per bacterial species were tested by hybridization on a solid glass support using culture collection strains as templates. Finally, three oligonucleotide probes per bacterial species were utilized to examine 65 middle ear fluid and 29 throat swab samples. The sensitivities of the developed assay compared to classic culture from middle ear fluid samples for H. influenzae, M. catarrhalis, and S. pneumoniae were 96 (93 for culture), 73 (93 for culture), and 100% (78% for culture), respectively. No cross-reactivity with bacterial species belonging to the normal oral flora was observed when the 29 throat swab samples were studied. The sensitivity of the assay to detect S. pyogenes from these samples was 93% (80% for culture). These results provide a proof of concept for the diagnostic use of microarray technology based on broad-range topoisomerase gene amplification, followed by hybridization and specific detection of bacterial species.

SUBMITTER: Roth SB

PROVIDER: S-EPMC516371 | biostudies-literature | 2004 Sep

REPOSITORIES: biostudies-literature

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Use of an oligonucleotide array for laboratory diagnosis of bacteria responsible for acute upper respiratory infections.

Roth Stina B SB Jalava Jari J Ruuskanen Olli O Ruohola Aino A Nikkari Simo S

Journal of clinical microbiology 20040901 9

We developed a diagnostic array of oligonucleotide probes targeting species-specific variable regions of the genes encoding topoisomerases GyrB and ParE of respiratory bacterial pathogens. Suitable broad-range primer sequences were designed based on alignment of gyrB/parE sequences from nine different bacterial species. These species included Corynebacterium diphtheriae, Fusobacterium necrophorum, Haemophilus influenzae, Legionella pneumophila, Moraxella catarrhalis, Mycoplasma pneumoniae, Staph ...[more]

PMID: 15365022

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Project description:OBJECTIVES:To investigate if use of antibiotics was associated with bacterial complications following upper respiratory tract infections (URTIs). DESIGN:Ecological time-trend analysis and a prospective cohort study. SETTING:Primary, outpatient specialist and inpatient care in Stockholm County, Sweden. All analyses were based on administrative healthcare data on consultations, diagnoses and dispensed antibiotics from January 2006 to January 2016. MAIN OUTCOME MEASURES:Ecological time-trend analysis: 10-year trend analyses of the incidence of URTIs, bacterial infections/complications and respiratory antibiotic use. Prospective cohort study: Incidence of bacterial complications following URTIs in antibiotic-exposed and non-exposed patients. RESULTS:The utilisation of respiratory tract antibiotics decreased by 22% from 2006 to 2015, but no increased trend for mastoiditis (p=0.0933), peritonsillar abscess (p=0.0544), invasive group A streptococcal disease (p=0.3991), orbital abscess (p=0.9637), extradural and subdural abscesses (p=0.4790) and pansinusitis (p=0.3971) was observed. For meningitis and acute ethmoidal sinusitis, a decrease in the numbers of infections from 2006 to 2015 was observed (p=0.0038 and p=0.0003, respectively), and for retropharyngeal and parapharyngeal abscesses, an increase was observed (p=0.0214). Bacterial complications following URTIs were uncommon in both antibiotic-exposed (less than 1.5 per 10 000 episodes) and non-exposed patients (less than 1.3 per 10 000 episodes) with the exception of peritonsillar abscess after tonsillitis (risk per 10 000 tonsillitis episodes: 32.4 and 41.1 in patients with no antibiotic treatment and patients treated with antibiotics, respectively). CONCLUSIONS:Bacterial complications following URTIs are rare, and antibiotics may lack protective effect in preventing bacterial complications. Analyses of routinely collected administrative healthcare data can provide valuable information on the number of URTIs, antibiotic use and bacterial complications to patients, prescribers and policy-makers.

Project description:BACKGROUND:Antibiotics are prescribed frequently for upper respiratory tract infections (URTIs) even though most URTIs do not require antibiotics. This over-prescription contributes to antibiotic resistance which is a major health problem globally. As physicians' prescribing behaviour is influenced by patients' expectations, there may be some opportunities to reduce antibiotic prescribing using patient-oriented interventions. We aimed to identify these interventions and to understand which ones are more effective in reducing unnecessary use of antibiotics for URTIs. METHODS:We conducted a systematic review by searching the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE (OVID), EMBASE (OVID), CINAHL, and the Web of Science. We included English language randomized controlled trials (RCTs), quasi-RCTs, controlled before and after studies, and interrupted time series (ITS) studies. Two authors screened the abstract/titles and full texts, extracted data, and assessed study risk of bias. Where pooling was appropriate, a meta-analysis was performed by using a random-effects model. Where pooling of the data was not possible, a narrative synthesis of results was conducted. RESULTS:We included 13 studies (one ITS, one cluster RCTs, and eleven RCTs). All interventions could be classified into two major categories: delayed prescriptions (seven studies) and patient/public information and education interventions (six studies). Our meta-analysis of delayed prescription studies observed significant reductions in the use of antibiotics for URTIs (OR = 0.09, CI 0.03 to 0.23; six studies). A subgroup analysis showed that prescriptions that were given at a later time and prescriptions that were given at the index consultation had similar effects. The studies in the patient/public information and education group varied according to their methods of delivery. Since only one or two studies were included for each method, we could not make a definite conclusion on their effectiveness. In general, booklets or pamphlets demonstrated promising effects on antibiotic prescription, if discussed by a practitioner. CONCLUSIONS:Patient-oriented interventions (especially delayed prescriptions) may be effective in reducing antibiotic prescription for URTIs. Further research is needed to investigate the costs and feasibility of implementing these interventions as part of routine clinical practice. SYSTEMATIC REVIEW REGISTRATION:PROSPERO CRD42016048007.

Project description:Early and accurate diagnosis of respiratory pathogens and associated outbreaks can allow for the control of spread, epidemiological modeling, targeted treatment, and decision making–as is evident with the current COVID-19 pandemic. Many respiratory infections share common symptoms, making them difficult to diagnose using only syndromic presentation. Yet, with delays in getting reference laboratory tests and limited availability and poor sensitivity of point-of-care tests, syndromic diagnosis is the most-relied upon method in clinical practice today. Here, we examine the variability in diagnostic identification of respiratory infections during the annual infection cycle in northern New Mexico, by comparing syndromic diagnostics with polymerase chain reaction (PCR) and sequencing-based methods, with the goal of assessing gaps in our current ability to identify respiratory pathogens. Of 97 individuals that presented with symptoms of respiratory infection, only 23 were positive for at least one RNA virus, as confirmed by sequencing. Whereas influenza virus (n = 7) was expected during this infection cycle, we also observed coronavirus (n = 7), respiratory syncytial virus (n = 8), parainfluenza virus (n = 4), and human metapneumovirus (n = 1) in individuals with respiratory infection symptoms. Four patients were coinfected with two viruses. In 21 individuals that tested positive using PCR, RNA sequencing completely matched in only 12 (57%) of these individuals. Few individuals (37.1%) were diagnosed to have an upper respiratory tract infection or viral syndrome by syndromic diagnostics, and the type of virus could only be distinguished in one patient. Thus, current syndromic diagnostic approaches fail to accurately identify respiratory pathogens associated with infection and are not suited to capture emerging threats in an accurate fashion. We conclude there is a critical and urgent need for layered agnostic diagnostics to track known and unknown pathogens at the point of care to control future outbreaks.

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Use of an oligonucleotide array for laboratory diagnosis of bacteria responsible for acute upper respiratory infections.

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Use of an oligonucleotide array for laboratory diagnosis of bacteria responsible for acute upper respiratory infections.

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