Pooled Sputum for Xpert MTB/RIF Testing: A Cost-Saving Strategy in Mwanza, Tanzania

Background: Tuberculosis (TB) diagnostics, such as Xpert MTB/RIF, is still limited by cost. Testing of pooled samples from presumptive TB patients has been thought as a cost-saving strategy to diagnose TB. We assessed the utility and cost-saving of pooled Xpert MTB/RIF testing strategy for the diagnosis of TB in Mwanza, Tanzania. Methods: Sputum samples from Presumptive TB patients were submitted to TB laboratory for routine diagnosis of TB using Xpert MTB/RIF. The TB results from the individual sputum samples were used as the reference standard and were concealed to the investigating laboratory technicians. The remainder of samples were collected serially and were pooled (5 samples per pool) for testing. The agreement of the results between individual sample testing against pooled sample testing and cost-savings was assessed. Results: A total of 250 sputum samples from presumptive TB patients were analyzed and 50 pools were made with each pool containing 5 samples. Of the 50 sputum pools made, Mycobacterium tuberculosis (MTB) was detected in 17 (34.0%) pools. Results from the individual sputum samples MTB/RIF testing were retrieved for all 250 samples and there were 28 (11.2%) samples in which MTB was detected whereas 222 (88.8%) samples had no MTB detected. Following re-analysis of positive pools, all 28 (100%) individual positive MTB samples were detected within the 17 positive pools, with 1 to 3 individual MTB positive samples per pool. The individual sputum samples were correctly identified by pooled sputum on Xpert MTB/RIF testing, with the sensitivity of 100% and specificity of 100%. Using pooling of sputum samples strategy, we saved 46.0% (115/250) of the cartridges. Conclusion: The pooled sputum testing strategy reduced cartridge costs by 46.0% and has the potential to increase the affordability of Xpert MTB/RIF testing in countries with limited resources, such as Tanzania.

EXAMINATION OF SPUTUM SEDIMENT PARAFFIN SECTION OF DIAGNOSIS OF LUNG CANCER

To establish the rapid and accurate methods for detecting cancer cells from the sputum of patients with lung cancer. Methods: Sputum sediment section examination and sputum smear examination were performed. For 142 cases of lung cancer patients diagnosed by bronchia fibrous lens and confirmed by pathologic diagnosis. Results: (a) The positive diagnosis rate of lung cancer cell was 71.83% (102/142) when using sputum sediment examination while that was 32.39% (46/142) when using sputum smear examination, when using sputum sediment examination jointly with sputum smear examination, the positive diagnosis rate of lung cancer was 91.55% (130/142). According to the positive diagnosis rate of lung cancer, the sputum sediment examination was significantly higher than sputum smear examination (P<0.001), joint examination of both of them was significantly higher than the single sputum sediment examination (P<0.001); (b) As to the histopathological types of lung cancers, 66 cases could be identified among 102 cases whom were confirmed by sputum sediment examination, the identification rate was 64.71%; and 8 cases could be identified among 46 cases whom were confirmed by sputum smear examination, with an identification rate of 17.39%, it was significantly lower than that by sputum sediment examination (P<0.001); (c) Immunohistochemistry technique could be used in the sections of sputum sediment to identify some histopathological types which could not be identified by routine HE stain, with an identification rate of 94.44% (34/36). Conclusion: the sputum sediment examination has some advantages compared to the sputum smear examination such as the specimen may include more materials, the higher cancer cell identification positive rate and immunohistochemistry method can be used to identify the histopathological type.

The Use of Sputum Dashboard and Dispatch Riders in Mitigating Delay in Sputum Samples’ Processing—Evidence from an Intervention in Lagos State, Nigeria

The act of referring TB presumptive individuals after verbal screening to another health facility/diagnostic centres for testing often results in large loss to follow-up, owing to the many barriers people face in accessing care. The transportation of sputum samples is an efficient method of increasing access to tuberculosis (TB) diagnostics in areas where testing is not currently available. One of the major reasons for the low yield of TB cases in the field was delay in sputum sample transportation to diagnostic laboratories among other factors. Damien Foundation Belgium, carried out a unique sputum riders initiatives in 20 LGAs of Lagos state between Q3-2019 and Q4-2020. A total of 25 sputum dispatch riders across the 20 LGAs in the State who are monitored routinely on an electronic dashboard were engaged for sputum logistics. Therefore, this report highlights how the initiatives have contributed to increased TB case notification in a densely populated metropolitan city in Nigeria. Overall, 48,265 units of sputum samples were transported to the laboratory from the facilities 99.9% of results which were received from the laboratory by the dispatch riders. The contribution of sputum riders to the overall TB case detection by the project progressively increased from 9% when the intervention began to 57% as at Q4 2020. The transportation of sputum samples by motorcycle dispatch riders is an efficient method of increasing access to TB diagnostic services in communities where testing is not currently available.

Application of Inflammatory Markers in Induced Sputum in Stable Chronic Obstructive Pulmonary Disease Patients with Positive Bronchodilation Tests

Positive bronchodilation (BD) tests can be noticed in some stable chronic obstructive pulmonary disease (COPD) patients.The characteristics of airway inflammation in this entity remain unclear.Our study aimed to identify the characteristics of airway inflammation in stable COPD patients with positive BD tests.The airway inflammation was assessed in 88 patients with stable COPD using the examination of induced sputum in the aftermath of lung function and BD tests.Cellular counts and the levels of molecular markers including eosinophil cationic protein (ECP),myeloperoxidase (MPO),interleukin-5 (IL-5),and IL-8 were assayed by Wright's stain,Immuno-CAP system,and ELISA,RT-PCR.Among the 88 patients with stable COPD,20 (22.7%) showed positive BD tests.The values of eosinophils (4.7%±3.4%) and ECP (90.1±41.6 ng/mL) in induced sputum in stable COPD patients with positive BD tests were markedly elevated as compared with those in stable COPD patients with negative BD tests or in healthy controls (all P<0.05),but significantly lower than those in asthmatic patients (all P<0.01).The IL-5 in sputum supernatant was significantly decreased in stable COPD patients with positive BD tests as compared with the patients with asthma (12.5±7.8 vs.48.2±26.0 ng/mL;P<0.01).However,healthy controls exhibited similar concentrations of IL-5 in induced sputum with patients with stable COPD,whether with positive or negative BD tests (all P>0.05).Moreover,the values of neutrophils (61.8%±15.1%),MPO (574.0±111.8 ng/mL),and IL-8 (32.6±13.4 ng/mL) in induced sputum in stable COPD patients with positive BD tests were significantly higher than those in asthmatics or normal controls (all P<0.01).However,the values of the above inflammatory markers in induced sputum were similar among stable COPD patients with positive or negative BD tests (all P>0.05).The stable COPD patients with positive BD tests may present not only eosinophilic airway inflammation but also neutrophilic airway inflammation.

Detection of DNA Aneuploidy in Exfoliated Airway Epithelia Cells of Sputum Specimens by the Automated Image Cytometry and Its Clinical Value in the Identification of Lung Cancer

To evaluate the value of detecton of DNA aneuploidy in exfoliated airway epithelia cells of sputum specimens by the automated image cytometry for the identification of lung cancer, 100 patients were divided into patient group (50 patients with lung cancer) and control group (30 patients with tuberculosis and 20 healthy people). Sputum was obtained for the quantitative analysis of DNA content of exfoliated airway epithelial cells with the automated image cytometry, together with the examinations of brush cytology and conventional sputum cytology. Our results showed that DNA aneuploidy (DI>2.5 or 5c) was found in 20 out of 50 sputum samples of lung cancer, 1 out of 30 sputum samples from tuberculosis patients, and none of 20 sputum samples from healthy people. The positive rates of conventional sputum cytology and brush cytology were 16 % and 32 %, which was lower than that of DNA aneuploidy detection by the automated image cytometry (P<0.01,P>05). Our study showed that automated image cytometry, which uses DNA aneuploidy as a marker for tumor, can detect the malignant cells in sputum samples of lung cancer and it is a sensitive and specific method serving as a complement for the diagnosis of lung cancer.

The Feasibility of Sputum Transportation System in China: Effect of Sputum Storage on the Mycobacterial Detection

Sputum transportation from county-level to prefecture-level is an ideal strategy to cover the shortage of the laboratory capability in the resource-poor setting. Here, we firstly evaluated the feasibility of sputum transportation system in China by analyzing the culture and molecular diagnosis results from 1982 smear-positive patients with different delay in processing for culture.

Follow-up Study of Retreatment TB Patients with Sputum Smear and/or Culture Positive Two Years after They were Declared Cured with First-line Anti-TB Drugs in Shandong Province

This study aimed to learn the recurrence rate in the retreatment TB patients with sputum smear and/or culture positive （ss＋ and/or c＋） two years after they were declared cured, and to explore causes of recurrence in order to improve long-time treatment outcome. 5 cities were selected as research locations. Recurrence of TB was judged by chest X-ray examination together with sputum smear and culture examination.

Trends for Tuberculosis Treatment Outcomes, New Sputum Smear Positive Patients in Kwekwe District, Zimbabwe, 2007-2011: A Cohort Analysis

Introduction: Tuberculosis remains a public health problem in Zimbabwe. Understanding the treatment outcomes is an important proxy indicator on the performance of the tuberculosis program. This study analyzed treatment outcomes of new sputum smear positive tuberculosis patients in Kwekwe district. Materials and Methods: A retrospective records review of new sputum smear positive tuberculosis patients registered in the district tuberculosis register in Kwekwe was conducted. Treatment outcomes were categorized according to the national tuberculosis control program and multivariate logistic regression model was used. P values less than 0.05 were considered statistically significant. Results: From a total of 1115 new sputum smear positive tuberculosis patients, cure rate ranged from 40.8% to 62.8% and death rate of patients decreased from 11 (8.0%) in 2007 to 17 (5.3%) in 2011 (p = 0.016). However, defaulter rate increased from 10 (7.3%) in 2007 to 30 (9.3%) in 2011. In multivariate logistic model, HIV positive tuberculosis patients were more likely to experience (adjusted RR = 1.84, 95% CI: 1.10 - 3.08) unfavorable treatment outcome when compared to negative counterparts. Urban residents were also at risk 1.91 (95% CI: 1.14 - 3.20) unfavorable outcome compared to rural residents. Conclusion: The cure rate was low (ranged from 40.8% to 62.8%) and the defaulter rate needed urgent attention. The district should conduct defaulter tracing and follow up.

Occurrence of <i>N</i>-Acyl Homoserine Lactones in Extracts of Bacterial Strain of <i>Pseudomonas aeruginosa</i>and in Sputum Sample Evaluated by Gas Chromatography–Mass Spectrometry

This study presents a fast, accurate and sensitive technique using gas chromatography-mass spectrometry (GC-MS) for the identification and quantification of N-acyl homoserine lactones (AHLs) in the extracts of bacterial strain of Pseudomonas aeruginosa and sputum sample of a cystic fibrosis patient. This method involves direct separation and determination of AHLs by using GC-MS as simultaneous separation and characterization of AHLs were possible without any prior derivatiza-tion. Electron ionization resulted in a common fragmentation pattern with the most common fragment ion at m/z 143 and other minor peaks at 73, 57 and 43. The limit of detection for N-butanoyl, N-hexanoyl, N-octanoyl, N-decanoyl, N-dodecanoyl and N-tetradecanoyl homoserine lactones was 2.14, 3.59, 2.71, 2.10, 2.45 and 2.34 μg/L, respectively. The presence of AHLs in the culture of P. aeruginosa strain and spu-tum of a cystic fibrosis patient was achieved in selected ion monitoring (SIM) mode by using the prominent fragment at m/z 143.

Comparison of Sputum Smear Microscopy and Rapid Tuberculosisantibody Detection Test Kits for Diagnosis of Pulmonary Tuberculosis in Abia State, Nigeria

The SSM （sputum smear microscopy） and five immunochromatographic tuberculosis antibody detection tests （DiaSpot TB, Spodex TB, SD Rapid TB, Clinotech TB Screen and Precious One-step TB） were compared for diagnosis of active TB at the Leprosy and Tuberculosis Referral Hospital, Uzuakoli, Abia State, Nigeria. Sputum specimens from 150 study participants （male/female ratio, 0.81） were cultured on Lowenstein-Jensen slopes and direct smears were stained by Ziehl-Neelsen technique and examined by light microscopy. Sera were tested for anti-TB antibodies using the rapid TB tests. A total of 91 participants were culture positive, 79 （86.8%） for M. tuberculosis and 12 （13.2%） for nontuberculous mycobacteria. The sensitivity of SSM was 50% （95% CI： 39.0-61.0） and specificity was 92.3% （95% CI： 86.4-98.2） in those culture positive for M. tuberculosis. The sensitivity and specificity of the Rapid TB tests ranged from 24.1-39.2% and 78.4-87.8%, respectively. None of the five rapid TB tests had acceptable level of accuracy for diagnosis of active TB. The sensitivity of SSM though moderate is inadequate for long term TB control in this setting.

An HIV 1/2 point of care test on sputum for screening TB/HIV coinfection in central India-Will it work?

Objective:To determine whether the OraQuick(?).HIV-1/2 Assay(OraSure Technologies, Inc.,Bethlehem,PA,USA) in sputum is a valid tool for HIV surveillance among TB patients. Methods:A cross sectional study was carried out on sputa of patients diagnosed with tuberculosis.Sputa were tested for antibodies to HIV using OraQuick. HIV-1/2 Assay(OraSure Technologies,Inc.,Bethlehem,PA,USA).The results were compared with results of serum ELISA.Results:Compared to serum ELISA,the OraQuick(?).HIV-1/2 Assay in sputum specimens reported 90%sensitivity(9/10)and 100%specificity(307/307),with a positive predictive value of 100%(95%CI:66.37%-100.00%)and a negative predictive value of 99.68%(95%CI:98.20%-99.99%). Conclusions:This testing method may provide a useful strategy for conducting HIV surveillance in possible co-infected TB patients at peripheral centres.Since there is no investment on infrastructure,it may be possible for paramedical health professionals to carry out the test. particularly in areas with low HIV endemicity.

Adding Sputum Collection and Transportation Services for Early Identification TB Cases in Hard-to-Reach Difficult Terrain—Will It Help?

A community engagement project was designed and implemented to reach 10.6 million population in 13 districts of Uttarakhand state—India, to identify presumptive TB patients (PTBPs) and link to free diagnostics services, either through referral or sputum collection and transportation (SCT). The objective of this study is to determine additional yield of TB patients achieved by providing SCT services in comparison to referrals of PTBPs identified in hard-to-reach areas. This was before and after comparative study conducted between April-June 2014 (2Q2014-before) to July-September 2014 (3Q2014-after). SCT was advised for all PTBPs identified from community level interventions between July-September 2014 (3Q2014-after). In 3Q2014, additional 279 (34%) PTBPs were examined in the microscopy centre with an additional 36 (51%) new smear positive (NSP) patient diagnosed. A total of 46 NSP (128%) were contributed by SCT alone. Higher proportion (94%) of TB patients was initiated on treatment in 3Q2014 in comparison to 2Q2014 (91%). SCT as a strategy for early case detection in hard-to-reach difficult terrain yields positive results in comparison to mere referral of PTBPs.

Could Sputum Eosinophilia Act as a Predictive Test for Diagnosis of High Risk Bakery Workers to Asthma?

Background: Occupational flour inhalation has been a culprit in the start of several pulmonary diseases such as asthma. We examined the relationship between occupational wheat flour inhalation in bakery and supermarket employees with pulmonary functions, respiratory symptoms, and sputum eosinophilia to determine hyper-responsiveness in a cross-sectional study in Iran. Methods: 122 subjects from traditional bakeries and 137 subjects from supermarket employees were enrolled in the study. Flour exposure concentrations, respiratory signs, sputum analysis, and respiratory volumes and capacities were measured based on the standard methods. Results: Respirable concentration of flour in the bakery workers was two- to four-fold of ACGIH’s threshold limit value in which bread-bakers with 2.2 mg/m3 experienced maximum exposures. The supermarket employees were not exposed to flour dust. The respiratory volumes in both bakery and supermarket employees were in the normal range. However, the median of voluminal percentage in bakery workers except Forced Vital Capacity (FVC) decreased (p p < 0.05). In addition, we observed increased respiratory symptoms in the bakery workers, again more prevalent in the bread-bakers. There was a significant correlation between flour exposure concentration and sputum eosinophilia in which the percentage of eosinophilia in the bread-bakers was more than other bakery and supermarket employees. Although there were reductions in the respiratory volumes, the results indicated no obstructive spirometric pattern. Conclusions: Sputum eosinophilia might be a suitable screening method to detect airway hyper-responsiveness in workers exposed to known asthmogens.

Determinants of Persistent Sputum Smear Positivity after Intensive Phase Chemotherapy among Patients with Tuberculosis at Rhodes Chest Clinic, Nairobi, Kenya

The prevalence of TB in sub-Sahara Africa has been reported as 511 per 100,000 populations and a mortality of 74 per 100,000 in year 2009. In the same period, incidence was estimated at 350 cases per 100,000. In this regard, the health system requires strengthening to respond to the rising cases of infection, drug resistance and quality of life lost while continuing to seek interventions that improve adherence to medication and case detection among those infected. Methods: This study sought to determine factors that are associated with sputum positivity after intensive phase of chemotherapy in people with tuberculosis. It was a retrospective case-control study conducted in Rhodes chest clinic, a City Council health unit in Nairobi that specializes in treatment of chest infections. The participants were sampled from clinic attendants who had completed two months of intensive phase TB chemotherapy and met inclusion criteria. Results: Seventy participants of whom 25 (36%) were sputum positive at the end of two (2) months intensive phase were included in the study. Skipping medication doses was significantly associated with sputum positivity (p = 0.01). Patients who were sputum positive at the end of the two-month period were more likely to have taken longer time before seeking treatment compared to those who were sputum negative by median (IQR) 8 (3 - 12) and 4 (3 - 8) weeks respectively although this difference was not significant (p = 0.09). Patients who had not disclosed their infection status had a two-fold possibility of remaining sputum positive at the end of intensive phase. Conclusion: Early diagnosis and treatment of TB, and adherence to medication were important factors that affect sputum conversion during intensive phase of TB treatment. Therefore, public health practitioners should advise patients to seek prompt diagnosis and treatment of signs and symptoms of tuberculosis.

Quality of Sputum Specimen Samples Submitted for Culture and Drug Susceptibility Testing at the National Tuberculosis Reference Laboratory-Uganda, July-October 2013

Setting: The Uganda National Tuberculosis Reference Laboratory (NTRL) in Kampala. Objective: The proportion of poor quality specimens received for drug susceptibility testing (DST) at the NTRL and factors contributing to poor specimen quality were assessed. Design: A cross-sectional study was conducted of sputum samples received at the NTRL from patients at high risk for multi-drug-resistant tuberculosis (MDR TB) during July-October 2013. Demographic, clinical, and bacte-riological data were abstracted from laboratory records. A poor quality sample failed to meet any one of four criteria: ≥3 milliliter (ml) volume, delivered within 72 hours, triple packaged, and non-salivary appearance. Results: Overall, 365 (64%) of 556 samples were of poor quality;89 (16%) were not triple packaged, 44 (8%) were <3 mls, 164 (30%) were not delivered on time, and 215 (39%) were salivary in appearance. Poor quality specimens were more likely to be collected during the eighth month of TB treatment (OR = 2.5, CI = 1.2 - 5.1), from the East or Northeast zones (OR = 2.2, CI = 1.1 - 4.8), and from patients who previously defaulted from treatment (OR = 1.9, CI = 1.1 - 3.2). Conclusion: The majority of sputum samples had poor quality. Additional efforts are needed to improve quality of samples collected at the end of treatment, from East and Northeast zones, and from patients who had previously defaulted.

DETECTION OF GENE MUTATION IN SPUTUM OF LUNG CANCER PATIENT

Lungcancerisacommonmalignanttumor,whichhasahighincidenceandmortalityrate.Therefore,itisnecessarytoseekanewmethodforthediagnosis,especiallytheearlydiagnosisoflungcancer.Thedevelopmentofmolecularbiologymakesthegenediagnosisoflungcancerpossible.PCR-SSCP...

Application of 16s rDNA Sequencing in the Analysis of Pathogenic Bacteria in Sputum of Severe Bacterial Pneumonia

Objective: 120 patients with severe pneumonia who were kept in the comprehensive ICU of our hospital in 2018 were selected, and 16s rDNA sequencing was performed to analyze the composition of pathogenic bacteria in the sputum of severe pneumonia. Methods: The sputum samples of patients with severe bacterial pneumonia were collected, and the diversity of pathogens in the samples was analyzed by polymerase chain reaction (PCR) amplification and high-throughput sequencing (16s rDNA PCR-DGGE). Results: Sequence showed that sputum samples contained a relatively large number of species, and there were many species that were not detected by sequencing. The dominant bacteria were Streptococcus, Sphingomonas, Corynebacterium, Denatobacteria, Aquobacteria, Acinetobacteria, Prevotella, Klebsiella, Pseudomonas, etc. Conclusion: Bacteria caused by sputum of severe bacterial pneumonia are complex and diverse, which provides new methods and ideas for individualized treatment of patients with severe pneumonia.

Application of 16S rDNA Sequencing Technology in the Analysis of Pathogenic Bacteria in Sputum of Severe Pneumonia

The diagnosis of pathogenic bacteria in severe pneumonia is difficult and the prognosis is poor. Its outcome is closely related to bacterial pathogenicity and the timeliness and pertinence of antibiotic treatment. Therefore, early diagnosis is of great significance to the prognosis of patients. Sputum examination and culture is the gold standard for the diagnosis of pathogens of severe pneumonia. However, due to the long time of bacterial culture, the early use of antibiotics, the change of bacteria species, mixed infection and other problems, the results of bacterial culture in sputum are often false negative. With the continuous application of new molecular biology techniques in clinical detection, the classification of bacteria and microorganisms has deepened from the identification of phenotypic characteristics to the classification of gene characteristics. Sequencing analysis with 16S rDNA sequencing technology has the characteristics of high sequencing flux, large amount of data obtained, short cycle, and can more comprehensively reflect the species composition of microbial community, real species distribution and abundance information. In this paper, 16S rDNA sequencing technology was used to analyze the bacterial population composition in the sputum of severe pneumonia, and to explore a new method of etiological diagnosis.

Correlation of Inflammatory Cells in Induced Sputum and Peripheral Blood of Patients with Acute Exacerbation of Chronic Obstructive Pulmonary Disease

Chronic obstructive pulmonary disease (COPD) is a major chronic respiratory disease worldwide. Inflammatory cells reflect the inflammatory situation both in peripheral blood and induced sputum. Their correlation has not been reported. The correlation between neutrophils (Neu), eosinophils (Eos), and lymphocyte (Lym) in induced sputum and that in peripheral blood of COPD patients was evaluated to explore the consistency of inflammatory cells in peripheral blood and induced sputum. Peripheral blood and induced sputum were collected from 437 patients with acute exacerbation of COPD (AECOPD) who were hospitalized in the Department of respiratory and critical care medicine, Inner Mongolia People’s Hospital. The correlation analysis was performed by Spearman correlation analysis. The ratios of Neu, Eos, and Lym in induced sputum were (79.15 ± 22.60)%, (5.23 ± 12.74)%, and (1.69 ± 2.66)%. The ratios of Neu, Eos, and Lym in peripheral blood were (63.29 ± 11.44)%, (2.99 ± 3.60)%, and (25.16 ± 10.19)%. The results showed that the ratios of Neu and Eos in induced sputum were significantly correlated with the proportion of corresponding cells in peripheral blood (P < 0.05). There was no correlation between the ratio of Lym and Leu in induced sputum and corresponding cells in peripheral blood (P > 0.05). In patients with AECOPD, the tendency of Neu and Eos in induced sputum was consistent with the corresponding cells in peripheral blood. Neu and Eos in induced sputum and peripheral blood reflected the degree of inflammation to guide the individualized medication of patients.

Bacteria spectrum of sputum patients with artificial airway and nursing countermeasures

Objective To investigate bacteria spectrum of the sputum collected from patients with artificial airway and explore the nursing strategy of preventing and reducing pulmonary infection. Methods 136 sputum samples are obtained by endotracheal tube suction hrough disposable closed suction catheter or brochoscopy sterile brush。