Good-quality Sputum Samples Research Articles

Stool Xpert MTB/RIF as a possible diagnostic alternative to sputum in Africa: a systematic review and meta-analysis.

Worldwide, COVID-19 pandemic lead to a large fall in the number of newly reported TB cases. In sub-Saharan Africa, microbiological diagnosis of TB is generally based on smear microscopy and Xpert MTB/RIF on sputum samples, but good quality sputum samples are often difficult to obtain, leading clinicians to rely on more invasive procedures for diagnosis. Aim of this study was to investigate pooled sensitivity and specificity of Xpert MTB/RIF on stool samples compared to respiratory microbiological reference standards in African countries. Four investigators independently searched PubMed, SCOPUS, and Web of Science until 12th October 2022, then screened titles and abstracts of all potentially eligible articles. The authors applied the eligibility criteria, considered the full texts. All the studies reported the data regarding true positive (TP), true negative (TN), false positive (FP) and false negative (FN). Risk of bias and applicability concerns were assessed with the Quadas-2 tool. overall, among 130 papers initially screened, we evaluated 47 works, finally including 13 papers for a total of 2,352 participants, mainly children. The mean percentage of females was 49.6%, whilst the mean percentage of patients reporting HIV was 27.7%. Pooled sensitivity for Xpert MTB/RIF assay for detecting pulmonary tuberculosis was 68.2% (95%CI: 61.1-74.7%) even if characterized by a high heterogeneity (I2=53.7%). Specificity was almost 100% (99%, 95%CI: 97-100%; I2 = 45.7%). When divided for reference standard, in the six studies using sputum and nasogastric aspirate the accuracy was optimal (AUC = 0.99, SE = 0.02), whilst in the studies using only sputum for tuberculosis detection the AUC was 0.85 (with a SE = 0.16). The most common source of bias was exclusion of enrolled patients in the analysis. Our study confirms that, in Africa, stool Xpert MTB/RIF may be a useful rule-in test for children above and below 5 years of age under evaluation for pulmonary tuberculosis. Sensitivity increased substantially when using both sputum and nasogastric aspirate as reference samples.

332. Broad range PCR of pleural fluid improves diagnosis of bacterial pneumonia complicated by parapneumonic effusion

Abstract Background Bacterial pneumonia complicated by a para pneumonic effusion is frequently encountered in hospitalized patients. However, identifying the etiology of pneumonia remains elusive. Published data report rates of bacteremia ranging from 13-26% among patients with pneumonia complicated by parapneumonic effusion. Good quality sputum samples are difficult to obtain in children. Culture of pleural fluid is helpful, but most patients are already pretreated before a specimen is obtained limiting utility. Nucleic acid assays of pleural fluid have reported sensitivities from 40-80%. We wanted to review our experience with pleural fluid nucleic acid assay at Akron childrens hospital Methods Using help from our laboratory data support team, we retrospectively reviewed the records of all pleural fluid isolates that were sent out to Washington University Medical Center for 16 S rRNA sequencing (broad range polymerase chain reaction assay). The review covered a 8-year time period ranging from January 2014 to March 2022. Negative blood and pleural fluid cultures were a prerequisite before the send out for nucleic acid assay. Results A total of 29 pleural fluid samples were sent out for 16S rRNA sequencing. 22 of these had a detectable target. This translated into a yield of about 75%. The following table summarizes our findings. Conclusion In our experience, nucleic acid assay of the pleural fluid is valuable in making a bacteriological diagnosis of pneumonia with para pneumonic effusion. Its benefit remains in patients pretreated with antibiotics. For the individual patient it may help narrow antibiotic therapy. At an institutional level it would allow for developing guidelines for empiric antibiotic management. However, in most institutions’ nucleic acid assay of pleural fluid remains a send out test and optimizing workflows to reduce turnaround time remains a challenge. Disclosures All Authors: No reported disclosures.

Expiratory Technique versus Tracheal Suction to Obtain Good-Quality Sputum from Patients with Suspected Lower Respiratory Tract Infection: A Randomized Controlled Trial.

Microbiological diagnostics of good-quality sputum samples are fundamental for infection control and targeted treatment of lower respiratory tract infections (LRTI). This study aims to compare the expiratory technique and tracheal suction on the quality of sputa from adults acutely hospitalized with suspected LRTI. We performed an open-label, randomized controlled trial. Patients were randomized to sputum sampling by tracheal suction (standard care) or the expiratory technique. The primary outcome was quality of sputum evaluated by microscopy and was analysed in the intention-to-treat population. The secondary outcomes were adverse events and patients experience. In total, 280 patients were assigned to tracheal suction (n = 141, 50.4%) or the expiratory technique (n = 139, 49.6%). Sputum samples were collected from 122 (86.5%) patients with tracheal suction and 67 (48.2%) patients with expiratory technique. Good-quality sputa were obtained more often with tracheal suction than with expiratory technique (odds ratio 1.83 [95% CI 1.05 to 3.19]; p = 0.035). There was no statistical difference in adverse events (IRR 1.21 [95% CI, 0.94 to 1.66]; p = 0.136), but patient experience was better in the expiratory technique group (p < 0.0001). In conclusion, tracheal suction should be considered a routine procedure in emergency departments for patients with suspected LRTI.

Microbiologic and virulence characteristics of Moraxella catarrhalis isolates from Zambian children presenting with acute pneumonia.

Moraxella catarrhalis is one of the bacterial pathogens associated with childhood pneumonia, but its clinical importance is not clearly defined. This study aimed to investigate the microbiologic and virulence characteristics of M. catarrhalis isolates obtained from children with pneumonia in Lusaka, Zambia. This retrospective, cross-sectional study analyzed 91M. catarrhalis isolates from induced sputum samples of children less than 5 years of age with pneumonia enrolled in the Pneumonia Etiology Research for Child Health study in Lusaka, Zambia between 2011 and 2014. Bacteria identification and virulence genes detection were performed by PCR and DNA sequencing, while antimicrobial susceptibility testing was determined by the Kirby-Bauer method. All the M. catarrhalis isolates were obtained from good-quality sputum samples and were the predominant bacteria. These isolates harbored virulence genes copB (100%), ompE (69.2%), ompCD (71.4%), uspA1 (92.3%), and uspA2 (69.2%) and were all β-lactamase producers. They showed resistance to ampicillin (100%), amoxicillin (100%), trimethoprim-sulfamethoxazole (92.3%), ciprofloxacin (46.2%), chloramphenicol (45.1%), erythromycin (36.3%), tetracycline (25.3%), cefuroxime (11.0%), and amoxicillin-clavulanate (2.2%), with 71.4% displaying multi-drug resistant phenotype but all susceptible to imipenem (100%). This study showed that M. catarrhalis isolates were the predominant or only bacterial isolates from the sputum samples analyzed. The findings provide supportive evidence for the pathogenic potential role of this bacterium in pediatric pneumonia. High multidrug resistance was also observed amongst the isolates, which can result in affected patients not responding to standard treatment, leading to prolonged illness, increased healthcare costs, and risk of death.

The need to prioritise childhood tuberculosis case detection

The need to prioritise childhood tuberculosis case detection

Can sputum gram stain be used to predict lower respiratory tract infection and guide empiric antimicrobial treatment: Experience from a tertiary care hospital

BackgroundThe mortality associated with lower respiratory tract infection is high. Indiscriminate use of antimicrobials leads to alteration of respiratory tract flora and development of multi-drug resistance. Rapid diagnostic tests to confirm infection can guide the clinicians about antimicrobial treatment. So the present study was planned to evaluate the role of direct gram stain examination as a rapid and simple test to help clinicians for appropriate patient management. MethodsThe present study was conducted on 1000 respiratory specimens which were processed using conventional microbiological techniques. Gram stain smear and culture results were compared statistically to assess the sensitivity, specificity, positive and negative predictive value. The agreement between gram stain smear examination and culture was calculated using kappa statistics. ResultsPotential pathogens were obtained from 28 of 209 deeply coughed out sputum samples (13.3%) and from 19 of 315 saliva mixed sputum samples (6%). Out of 473 tracheal aspirates, 115 (24.3%) had potential pathogens. The sensitivity for predicting infection was higher for good quality sputum samples (54%) as compared to poor quality sputum samples (37%). The gram stain and culture of tracheal samples had a good agreement for predicting infection whereas there was only moderate agreement for sputum sample. ConclusionGram stain smear examination from respiratory samples can be used to guide empiric antibiotic therapy pending final culture sensitivity results if the attending physicians ensure appropriate sample collection and transport. In absence of these supportive measures smear examination should not be relied upon for empiric treatment.

Specific IgA against Pseudomonas aeruginosa in severe COPD.

BackgroundThe bronchial mucosa is protected by a specialized immune system focused on the prevention of colonization and infection by potentially pathogenic microorganisms (PPMs). Immunoglobulin A (IgA) is the principal antibody involved in this mechanism. A defective immune barrier may facilitate the recurrent presence of PPMs in COPD.PurposeThe aim of this study was to determine IgA-mediated bronchial specific immune responses against Pseudomonas aeruginosa in stable patients with severe disease.MethodsCOPD patients with good-quality sputum samples obtained during stability were included and classified according to the presence or absence of chronic bronchial colonization by P. aeruginosa. Levels of specific IgA for P. aeruginosa in sputum were determined by ELISA and expressed as ratios, using the pooled level of 10 healthy subjects as reference (optical density450 patient/control).ResultsThirty-six stable COPD patients were included, 15 of whom had chronic colonization by P. aeruginosa. Levels of specific IgA against P. aeruginosa in stable non-colonized patients were lower than those in healthy subjects (IgA ratio: median =0.15 [interquartile range {IQR} 0.05–0.36]). Colonized patients had higher levels, (1.56 [IQR 0.59–2.79]) (p<0.001, Mann–Whitney U test), with figures equivalent but not exceeding the reference value.ConclusionIgA-based immune response against P. aeruginosa was low in severe COPD patients. Levels of specific IgA against this microorganism were higher in colonized patients, but did not attain clear-cut levels above the reference. An impaired local response against P. aeruginosa may favor chronic colonization and recurrent infections in severe COPD.

Functional Metagenomics of the Bronchial Microbiome in COPD.

The course of chronic obstructive pulmonary disease (COPD) is frequently aggravated by exacerbations, and changes in the composition and activity of the microbiome may be implicated in their appearance. The aim of this study was to analyse the composition and the gene content of the microbial community in bronchial secretions of COPD patients in both stability and exacerbation. Taxonomic data were obtained by 16S rRNA gene amplification and pyrosequencing, and metabolic information through shotgun metagenomics, using the Metagenomics RAST server (MG-RAST), and the PICRUSt (Phylogenetic Investigation of Communities by Reconstruction of Unobserved States) programme, which predict metagenomes from 16S data. Eight severe COPD patients provided good quality sputum samples, and no significant differences in the relative abundance of any phyla and genera were found between stability and exacerbation. Bacterial biodiversity (Chao1 and Shannon indexes) did not show statistical differences and beta-diversity analysis (Bray-Curtis dissimilarity index) showed a similar microbial composition in the two clinical situations. Four functional categories showed statistically significant differences with MG-RAST at KEGG level 2: in exacerbation, Cell growth and Death and Transport and Catabolism decreased in abundance [1.6 (0.2–2.3) vs 3.6 (3.3–6.9), p = 0.012; and 1.8 (0–3.3) vs 3.6 (1.8–5.1), p = 0.025 respectively], while Cancer and Carbohydrate Metabolism increased [0.8 (0–1.5) vs 0 (0–0.5), p = 0.043; and 7 (6.4–9) vs 5.9 (6.3–6.1), p = 0.012 respectively]. In conclusion, the bronchial microbiome as a whole is not significantly modified when exacerbation symptoms appear in severe COPD patients, but its functional metabolic capabilities show significant changes in several pathways.

Evaluation of the implementation of the Xpert® MTB/RIF assay in Fiji.

All Xpert® MTB/RIF tests performed in the three TB (tuberculosis) treatment centres in Fiji from June 2012 to February 2013. To determine 1) the number of Xpert tests performed in each centre, 2) the association between sputum quality and Xpert results, 3) the agreement of Xpert with acid-fast bacilli (AFB) smear microscopy and TB culture and 4) error rates. Retrospective review of records. A total of 415 Xpert tests were performed in the study period. Mycobacterium tuberculosis was detected in 69 (16.6%) samples. No rifampicin resistance was detected. M. tuberculosis was detected from 60 (18.7%) good-quality sputum samples. A total of 43 (10.4%) errors occurred during this period. M. tuberculosis was detected in 10 (2.9%) smear-negative specimens. There was a substantial and an almost perfect agreement between Xpert and AFB microscopy (κ = 0.793) and culture results (κ = 0.818), respectively. Although a good correlation between Xpert and the two tests were shown in the study, Xpert should not replace the routine first-line TB diagnostic tests used in Fiji for reasons related to logistics and sustainability. A further evaluation of the assay's performance is required over a longer time period to gauge its diagnostic value in detecting smear-negative, Xpert-positive cases in Fiji.

Infectious etiology of acute exacerbations in severe COPD patients

Since the new GOLD guidelines were implemented no data have been published about the etiology of acute exacerbations (AECOPD) in severe COPD patients with a different frequency of annual episodes. One hundred and eleven COPD patients (FEV1<50%) were prospectively followed up for a year. Good-quality sputum samples recovered during AECOPD were processed, including quantitative culture and PCR detection of atypical bacteria. A total of 188 sputum samples were obtained from AECOPD episodes. Forty patients had a single episode, and 71 patients had ≥2. In 128 episodes a single pathogen was isolated, while 42 episodes were polymicrobial (≥2 pathogens). Overall, the most frequent pathogen isolated was Pseudomonas aeruginosa (n=54), followed by Haemophilus influenzae (n=37), Streptococcus pneumoniae (n=31), Moraxella catarrhalis (n=29) and Staphylococcus aureus (n=12). P. aeruginosa was the most frequent in both groups of patients (35% and 27% in those with 1 and ≥2 AECOPD, respectively). H. influenzae was associated with patients with a single annual AECOPD (33% vs. 16%; P=0.006), while Enterobacteriaceae were associated with frequent exacerbators (0% vs. 12%; P<0.044). Overall, P. aeruginosa was the most frequent pathogen isolated from exacerbations. However, different bacterial etiology was observed depending on the number of annual episodes.

Differential Expression of Host Biomarkers in Saliva and Serum Samples from Individuals with Suspected Pulmonary Tuberculosis

The diagnosis of tuberculosis remains challenging in individuals with difficulty in providing good quality sputum samples such as children. Host biosignatures of inflammatory markers may be valuable in such cases, especially if they are based on more easily obtainable samples such as saliva. To explore the potential of saliva as an alternative sample in tuberculosis diagnostic/biomarker investigations, we evaluated the levels of 33 host markers in saliva samples from individuals presenting with pulmonary tuberculosis symptoms and compared them to those obtained in serum. Of the 38 individuals included in the study, tuberculosis disease was confirmed in 11 (28.9%) by sputum culture. In both the tuberculosis cases and noncases, the levels of most markers were above the minimum detectable limit in both sample types, but there was no consistent pattern regarding the ratio of markers in serum/saliva. Fractalkine, IL-17, IL-6, IL-9, MIP-1β, CRP, VEGF, and IL-5 levels in saliva and IL-6, IL-2, SAP, and SAA levels in serum were significantly higher in tuberculosis patients (P < 0.05). These preliminary data indicate that there are significant differences in the levels of host markers expressed in saliva in comparison to those expressed in serum and that inflammatory markers in both sample types are potential diagnostic candidates for tuberculosis disease.

Direct microscopy versus sputum cytology analysis and bleach sedimentation for diagnosis of tuberculosis: a prospective diagnostic study

BackgroundDiagnostic options for pulmonary tuberculosis in resource-poor settings are commonly limited to smear microscopy. We investigated whether bleach concentration by sedimentation and sputum cytology analysis (SCA) increased the positivity rate of smear microscopy for smear-positive tuberculosis.MethodsWe did a prospective diagnostic study in a Médecins Sans Frontières-supported hospital in Mindouli, Republic of Congo. Three sputum samples were obtained from 280 consecutive pulmonary tuberculosis suspects, and were processed according to WHO guidelines for direct smear microscopy. The remainder of each sputum sample was homogenised with 2.6% bleach, sedimented overnight, smeared, and examined blinded to the direct smear result for acid-fast bacilli (AFB). All direct smears were assessed for quality by SCA. If a patient produced fewer than three good-quality sputum samples, further samples were requested. Sediment smear examination was performed independently of SCA result on the corresponding direct smear. Positivity rates were compared using McNemar's test.ResultsExcluding SCA, 43.2% of all patients were diagnosed as positive on direct microscopy of up to three samples. 47.9% were diagnosed on sediment microscopy, with 48.2% being diagnosed on direct microscopy, sediment microscopy, or both. The positivity rate increased from 43.2% to 47.9% with a case definition of one positive smear (≥1 AFB/100 high power fields) of three, and from 42.1% to 43.9% with two positive smears. SCA resulted in 87.9% of patients producing at least two good-quality sputum samples, with 75.7% producing three or more. Using a case definition of one positive smear, the incremental yield of bleach sedimentation was 14/121, or 11.6% (95% CI 6.5-18.6, p = 0.001) and in combination with SCA was 15/121, or 12.4% (95% CI 7.1-19.6, p = 0.002). Incremental yields with two positive smears were 5/118, or 4.2% (95% CI 1.4-9.6, p = 0.062) and 7/118, or 5.9% (95% CI 2.4-11.8, p = 0.016), respectively.ConclusionsThe combination of bleach sedimentation and SCA resulted in significantly increased microscopy positivity rates with a case definition of either one or two positive smears. Implementation of bleach sedimentation led to a significant increase in the diagnosis of smear-positive patients. Implementation of SCA did not result in significantly increased diagnosis of tuberculosis, but did result in improved sample quality. Requesting extra sputum samples based on SCA results, combined with bleach sedimentation, could significantly increase the detection of smear-positive patients if routinely implemented in resource-limited settings where gold standard techniques are not available. We recommend that a pilot phase is undertaken before routine implementation to determine the impact in a particular context.

Insufficient quality of sputum submitted for tuberculosis diagnosis and associated factors, in Klaten district, Indonesia

BackgroundSputum smear microscopy is the standard diagnostic method for detection of smear positive pulmonary tuberculosis (TB). Insufficient quality of sputum might result in missing cases. In this study we aimed at assessing the quality of sputum in a district in Central Java and determining patient and health worker factors associated with submission of three good quality sputum samples.MethodsIn 16 health centers information was collected on the quality of sputum submitted by TB suspects, i.e. volume, color, and viscosity. TB suspects were interviewed to assess their knowledge of TB, motivation to provide sputum and whether they were informed why and how to produce a sputum sample. Health workers were interviewed to assess what information they provided to TB suspects about the reason for sputum examination, methods to produce sputum and characteristics of a good quality sputum sample. All health worker and patient factors were evaluated for association with sputum quality.ResultsOf 387 TB suspects, 294 (76.0%) could be traced and interviewed, and of 272 (70.3%) information about sputum quality was available. Of those 203 (74.6%) submitted three samples, 90 (33.1%) provided at least one good sample, and 37 (13.6%) provided three good quality sputum samples. Of the 272 TB suspects, 168 (61.8%) mentioned that information on the reason for sputum examination was provided, 66 (24.3%) remembered that they were informed about how to produce sputum and 40 (14.7%) recalled being informed about the characteristics of good quality sputum. Paramedics reported to provide often/always information on the importance of sputum examination, and when to produce sputum. Information on how to produce sputum and characteristics of a good sputum sample was less often provided. None of the studied patient characteristics or health worker factors was associated with providing good quality sputum.ConclusionA considerable number of TB suspects did not provide three sputum samples and a large number of sputum samples were of insufficient quality. Training of health workers in providing health education to the TB suspect about the reason for sputum examination and how to produce a good quality sputum sample should be a priority of the TB program.

Effect of Nebulised Acetylcysteine on Sputum Quality and Diagnostic Yield

ABSTRACTBackgroundAcetylcysteine acts as a mucolytic by cleaving disulfide bonds in mucoprotein and decreasing sputum viscosity. Mucolysis should lead to easier expectoration and improved sputum sample quality for microbiological analysis.AimTo compare an effect size for the efficacy of nebulised acetylcysteine and sodium chloride 0.9% in the procurement of good quality sputum samples.MethodA double‐blind, randomised, placebo‐controlled pilot study in the emergency department. Adults with probable lower respiratory tract infection requiring sputum sampling were enrolled. Patients were randomised to receive acetylcysteine 1 g or sodium chloride 0.9% (placebo) via nebuliser. Patients, emergency department and microbiology staff were blinded to the treatment allocation. The study endpoints were good quality sputum samples (&lt; 10 squamous cells per high‐powered field and &lt; 10 squamous cells plus &gt; 25 polymorphs per high‐powered field), sputum samples where a predominant pathogen was isolated and adverse effects of acetylcysteine and sodium chloride 0.9%.Results28 patients were randomised to the acetylcysteine (n = 14) and sodium chloride 0.9% (n = 14) groups. Four (29%) patients in each group produced good quality sputum samples based on squamous cells alone. 2 (14%) and 3 (21%) patients in the acetylcysteine and sodium chloride 0.9% groups, respectively, produced good quality samples based on squamous cells and polymorph counts. Predominant pathogens were isolated in 2 (14%) patients in each group. 2 (14%) and 5 (36%) patients in the acetylcysteine and sodium chloride 0.9% groups, respectively, reported bronchospasm and/or coughing.ConclusionThere was no detectable trend towards acetylcysteine improving sputum sample quality. A large scale trial is not warranted.

Bronchoscopy for Diagnosing TB in Children

TB is difficult to diagnose in children because of problems in obtaining good-quality sputum samples. Endobronchial TB (ETB) — tuberculous infection of

Quantitative PCR Assay Using Sputum Samples for Rapid Diagnosis of Pneumococcal Pneumonia in Adult Emergency Department Patients

Accurate diagnosis of pneumococcal pneumonia in the acute-care setting remains a challenge due to the inadequate sensitivity of conventional diagnostic tests. Sputum cultures, which are likely to have the highest diagnostic yields of all specimen types, have been considered unreliable, due to their inability to differentiate colonization from infection. Our objective was to evaluate the potential clinical utility of a rapid quantitative real-time PCR assay using sputum samples for Streptococcus pneumoniae in adult patients with community-acquired pneumonia (CAP). A prospective clinical observational study of consecutively enrolled emergency department patients with CAP was performed; only those patients with excess good-quality sputum samples were included for evaluation. Sputum samples were tested for the presence of S. pneumoniae by using a quantitative PCR that targets the pneumolysin gene. PCR findings were compared with those of a composite reference standard comprising Gram staining of sputum samples and sputum/blood cultures. The area under the curve (AUC) and a log-transformed threshold, which provides the maximal sensitivity and specificity, were calculated. Of 487 subjects enrolled, 129 were evaluable. Receiver operating characteristic curve analysis demonstrated an AUC of 0.87. Sensitivity and specificity were 90.0 percent and 80.0 percent, respectively; positive and negative predictive values were 58.7 percent and 96.2 percent, respectively. We have demonstrated that a quantitative rapid pneumolysin PCR assay has favorable accuracy for diagnosis of pneumococcal pneumonia in adult patients with CAP; this assay may be a useful diagnostic adjunct for clinicians, particularly those practicing in the acute-care setting, where rapid pathogen identification may assist in selection of the most appropriate antibiotics.