594529

research-article2015
TAR0010.1177/1753465815594529Therapeutic Advances in Respiratory DiseaseKeeratichananont et al.

Therapeutic Advances in Respiratory Disease Original Research

Diagnostic yield and safety of sputum

Ther Adv Respir Dis

2015, Vol. 9(5) 217­–223

induction with nebulized racemic salbutamol DOI: 10.1177/

1753465815594529

versus hypertonic saline in smear-negative

© The Author(s), 2015.
Reprints and permissions:
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pulmonary tuberculosis
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Warangkana Keeratichananont, Thanapon Nilmoje, Suriya Keeratichananont

and Jedsada Rittatorn

Abstract
Objectives: The aim of the study was to compare the diagnostic yield and safety profile of
sputum induction (SI) with nebulized racemic salbutamol solution versus hypertonic saline in
smear-negative pulmonary tuberculosis (TB).
Methods: The prospective study was conducted at Songklanagarind Hospital, Thailand.
Suspected smear-negative pulmonary TB cases were recruited and randomized to receive
SI with either nebulized racemic salbutamol solution or 3% sodium chloride (NaCl) solution.
Induced sputum was examined with the acid-fast bacilli (AFB) smear test and cultured for
Mycobacterium tuberculosis. The efficacy and adverse events of SI were analyzed.
Results: A total of 59 patients received SI with nebulized racemic salbutamol solution and 53
received 3% NaCl solution. There was no significant difference between the two groups in the
average quantity of induced sputum (1.3 ± 0.1 versus 1.2 ± 0.2 ml, p = 0.5). The percentages
of positive AFB smear and TB cultures in the salbutamol group were 15% and 22%, and 13%
and 17% in the 3% NaCl group (p = 0.5), respectively. Racemic salbutamol solution could
increase the TB diagnostic yield similarly to 3% NaCl, but incurred less chest tightness (5%
versus 15%) and bronchospasm (0% versus 11.3%, p = 0.02) compared with 3% NaCl.
Conclusions: SI by nebulized racemic salbutamol solution offers equal benefits to 3%
NaCl solution in increasing both sputum quantity and diagnostic yield in smear-negative
patients suspected of having pulmonary TB. Nebulized racemic salbutamol does not produce
bronchospasm and chest tightness occurs less frequently than with 3% NaCl. Therefore,
SI with nebulized racemic salbutamol solution should be considered as a good alternative
Correspondence to:
noninvasive diagnostic tool for the diagnosis of pulmonary TB when hypertonic saline is Warangkana
Keeratichananont, MD
unavailable or contraindicated. Assistant Professor,
Division of Respiratory and
Respiratory Critical Care
Keywords:  hypertonic saline, salbutamol nebulizer, smear-negative pulmonary tuberculosis, Medicine, Department
of Medicine, Faculty
sputum induction of Medicine, Prince of
Songkla University, Hat
Yai, Songkhla 90110,
Thailand
tik25572014@hotmail.com
Introduction with bronchoalveolar lavage (BAL), and lung Thanapon Nilmoje, MD
Pulmonary tuberculosis (TB) continues to be a biopsy. These procedures have their own limitations Jedsada Rittatorn, MD
major worldwide health problem. Delayed diagno- such as invasiveness, need for patient cooperation, Department of Medicine,
Songklanagarind Hospital,
sis and untreated patients lead to further disease cost, and availability of the institutional and local Prince of Songkla
progression and transmission [WHO, 2012; Golub expertise needed. Therefore sputum collection for University, Songkhla,
Thailand
et al. 2006]. Intensified case finding is therefore a Ziehl–Neelsen staining for acid-fast bacilli (AFB) Suriya Keeratichananont,
key intervention and still presents a major chal- detection remains the mainstay of diagnosis MD
Department of Medicine,
lenge. There are many tools for making a TB diag- [American Thoracic Society and the Centers of Bangkok Hospital Hat Yai,
nosis, for example, gastric washing, bronchoscopy Disease Control and Prevention, 2000]. However, Songkhla,Thailand

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Therapeutic Advances in Respiratory Disease 9(5)

40–60% of active pulmonary TB cases have nega- In 2002, 1 mg of oral salbutamol three times a day
tive AFB staining by self-expectoration (smear-neg- for 3 days was used to induce sputum in 289
ative pulmonary TB) due to a dry cough, scarce chest-symptomatic patients who had either a dry
sputum, or paucibacillary TB [Paggiaro et al. 2002; cough or scanty sputum [Yazdani et al. 2002]. It
Hong Kong Chest Survice/Tuberculosis Research showed that 88% of cases could produce ade-
Center Madras/British Medical Research Council, quate sputum after induction with oral salbuta-
1979; Narain et al. 1971; Hensler et al. 1961], thus mol and there was no reported adverse drug
noninvasive sputum induction (SI) with ultrasonic reaction (ADR). Recently, Ansari and colleagues
nebulized hypertonic saline (3% sodium chloride used nebulized levosalbutamol (R-isomer salbu-
[NaCl]) is a good alternative method for diagnosis tamol) at a dosage of 1.26 mg for 2 consecutive
in this patient group. days prior to collecting three sputum samples in
40 clinically suspected TB patients who had a dry
SI was first used by Hensler and colleagues cough or scant sputum [Ansari et  al. 2013].
[Hensler et  al. 1961], and hypertonic saline was Results showed sputum collection was successful
used to stimulate cough reflex, increase osmotic in 90% of patients and gave a positive AFB smear
pressure in the lower airways, draw more water in 77.5% of cases. Nebulized racemic salbutamol
into the lumen, dilute mucins in the airways, and (S-isomer salbutamol) was not only cheaper than
facilitate sputum expectoration. Previous studies levosalbutamol but was also a widely used beta2-
showed that SI with hypertonic saline could agonist in Thailand. Its efficacy and safety profile
improve both the quantity and quality of the col- was not considered to be inferior to levosalbuta-
lected sputum in 70–90% of cases [Paggiaro et al. mol for the treatment of acute asthma attack [Jat
2002]. Moreover, SI has been reported to be as and Khairwa, 2013]. Therefore, the present study
effective as gastric washing and bronchoscopy was conducted to evaluate the diagnostic yield
with BAL for TB diagnoses, and gives a diagnos- and adverse events of SI in nebulized racemic sal-
tic yield 2–41% higher than self-expectorated butamol versus hypertonic saline (3% NaCl) in
sputum collection [Gonzalez-Angulo et al. 2012; Thai patients who were suspected of smear-nega-
Hepple et al. 2012; Hatherill et al. 2009; Brown tive pulmonary TB.
et  al. 2007; Gupta and Seema, 2005; Bell et  al.
2003; Li et  al. 1999; Kawada et  al. 1996; Shata
et al. 1996; Anderson et al. 1995; Kim et al. 1984; Material and methods
Narain et al. 1971]. However, SI with hypertonic A prospective, randomized, patient-blinded, com-
saline itself can produce chest pain, tightness, parative study was conducted from August 2008
rapid breathing, and bronchospasm by activation to February 2012. The inclusion criteria were: (a)
of airway mast cells and sensory nerve endings patients at least 15 years old in Songklanagarind
with an incidence of 6–32% (especially in patients Hospital, Songkhla, Thailand, who presented
with asthma or chronic obstructive pulmonary with a nonproductive cough persisting for at least
disease [COPD]), despite pretreatment with 3 weeks or a dry cough or scanty sputum (< 2 ml
beta2-agonist inhalation [Geldenhuys et al. 2012, saliva) with prolonged fever; (b) radiologic find-
2014; Dunleavy et al. 2008; Makker and Holgate, ings compatible with active pulmonary TB; (c)
1993]. There is a report of death in one patient negative AFB smears on 3 consecutive days of
with asthma undergoing ultrasonic nebulized self-expectorated sputum after being well
hypertonic saline [Saetta et al. 1995], thus SI with instructed by a certified respiratory therapy tech-
hypertonic saline was contraindicated for patients nician. Patients diagnosed with asthma, COPD,
with a history of bronchospasm, and prophylactic pneumonia, lung cancer, bronchiectasis, uncon-
nebulized salbutamol is suggested in cases of trolled hypertension, cardiac arrhythmias, preg-
moderate to severe asthma where SI is required nancy, allergy to bronchodilators, undiagnosed
[Carlsten et  al. 2007; Delvaux et  al. 2004; cause of chest symptoms and abnormal imaging,
Pizzichini et al. 2002; Jones et al. 2001]. had received antituberculosis drugs within the
previous 2 months, or had a positive AFB smear
Beta2-agonists have been demonstrated to from self-expectoration were excluded from the
enhance mucociliary transport in healthy subjects study. All patients signed informed consent and
as well as patients with asthma and chronic bron- were randomized (using block randomization)
chitis. It decreases the tenacity of sputum and into two groups to receive SI via a face mask with
thereby facilitates easy expectoration [Bennett, either nebulized racemic salbutamol solution
2002; Yazdani et al. 2002; Mortensen et al. 1991]. (1 ml of 0.5% salbutamol solution plus 3 ml of

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 W Keeratichananont, T Nilmoje et al.

0.9% NaCl), or 4 ml nebulized 3% NaCl (with- et  al. 2013] However, negative AFB smears on 3
out prophylactic bronchodilator) under 100% consecutive days of self-expectorated sputum had
oxygen compressor nebulizer at a flow rate of 15 not been excluded before SI in this study.Therefore,
L/min for up to 20 min in a standard isolation an expected smear-positive SI by nebulized race-
negative pressure room. Within 60 min of nebuli- mic salbutamol was 40% (at α = 0.05). The total
zation, all patients were instructed to expectorate calculated sample size was 49 patients per group.
and a spot sputum specimen was collected in a The mean + standard deviation was used to
sterile container and measured. It was then exam- describe continuous data. Proportion (%) was
ined with Ziehl–Neelsen staining and cultured for used to describe categorical data. Student’s t-test
Mycobacterium tuberculosis (using Lowenstein– and the chi-square test were used to analyze con-
Jensen medium) by the blinded experienced labo- tinuous and categorical data, respectively.
ratory technicians. Vital signs (by automated Outcomes of interest were analyzed with SPSS
device), electrocardiography, and SI-related version 11 software and the results were consid-
adverse events, for example, palpitation, cardiac ered as statistically significant if the p value was less
arrhythmia, chest tightness, bronchospasm, than 0.05.
tachypnea, and oxygen desaturation (by finger
pulse oximeter) were closely observed at baseline,
continuously during the nebulization phase, and Results
at 15-min intervals for up to 1 h postnebulization A total of 147 participants were initially recruited
by the primary physicians who were blinded to into the study and randomized into two groups:
the composition of the SI. If symptomatic bron- 74 patients received SI with nebulized racemic
chospasm (expiratory wheezing with tachypnea salbutamol solution and 73 received nebulized
or oxygen desaturation, SO2 < 95%) occurred, 3% NaCl solution, respectively. A total of 35 cases
the rescue bronchodilator (nebulized salbutamol were excluded due to negative TB results by bron-
solution) was given, and the SI procedure stopped choscopy or other conditions were proven to be
for safety reasons and interpreted as a negative the cause of pulmonary disease (i.e. 12 lung can-
AFB smear result. cers, eight pneumonias, two bronchiectasis, and
13 inconclusive), which meant that 112 patients
Patients with a negative AFB smear by SI under- with pulmonary TB were analyzed in the study.
went bronchoscopy with BAL with or without Of these 59 received SI with nebulized racemic
transbronchial lung biopsy (TBB) within 1 week salbutamol solution and 53 received nebulized
for a definite diagnosis. The definitive diagnosis of 3% NaCl solution. There was no difference in the
active pulmonary TB cases depends on either the baseline characteristics between the two groups in
detection of M. tuberculosis from culture speci- terms of age, sex, underlying diseases, and pattern
mens (by induced sputum or bronchoscopy with abnormalities on chest X-ray (Tables 1 and 2). A
BAL) or TBB showing granulomatous inflamma- total of 84 cases (75%) were diagnosed as pulmo-
tion with clinical response to a full treatment nary TB by positive culture of induced sputum or
course of antituberculosis drugs. The study proto- BAL, and 25% were diagnosed by lung biopsy
col was approved by the ethics committee of the specimens (15 cases in the salbutamol group and
institution (IRB#50/372-045), and was con- 13 cases in the hypertonic saline group, respec-
ducted in accordance with the World Medical tively). There was no significant difference
Association Declaration of Helsinki and 2013 between the two groups in the average quantity of
good clinical practical guideline. [WMA, 2013]. induced sputum (1.3 ± 0.1 versus 1.2 ± 0.2 ml/
case, p = 0.5). The percentages of positive AFB
smear and positive TB culture from spot SI with
Statistical analysis the nebulized racemic salbutamol solution were
The sample sizes were calculated using two inde- 15% (nine cases) and 22% (13 cases), and 13%
pendent proportions with 80% power of detection. (seven cases) and 17% (nine cases) with the 3%
Previous studies showed that diagnostic yield in NaCl solution, respectively (p = 0.5) (Table 3).
pulmonary TB using nebulized 3% NaCl solution There was no statistically significant change in
for SI ranged from 2% to 41%. Thus an expected body temperature, heart rate, blood pressure, or
smear-positive SI by nebulized 3% NaCl in this oxygenation between the two groups. Neither pal-
study was 15%, whereas the diagnostic yield of pul- pitations nor arrhythmia were documented dur-
monary TB using nebulized levosalbutamol was ing and after the period of nebulization. However,
61% in the study of Ansari and colleagues [Ansari nebulized 3% NaCl significantly produced more

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Therapeutic Advances in Respiratory Disease 9(5)

Table 1.  Baseline characteristics of patients in the nebulized 3% sodium chloride solution and racemic
salbutamol solution groups.

Characteristic 3% Sodium chloride Salbutamol p value

(n = 53) n (%) (n = 59) n (%)
Age (years) (mean ± standard deviation) 49.7 ± 15 49.6 ± 14 0.732
Male sex 25 (47.2) 33 (55.9) 0.354
Underlying diseases 0.997
Diabetes mellitus 5 (9.4) 6 (10.2)  
HIV 4 (7.5) 6 (10.2)  
Steroid use 3 (5.7) 3 (5.1)  
Others 7 (13.2) 7 (11.9)  

Table 2.  Patient chest film: location and pattern abnormality.

Characteristic 3% Sodium chloride Salbutamol p value

(n = 53) n (%) (n = 59) n (%)
Abnormal chest X-ray 0.993
location
Upper lobe 31 (58.5) 34 (57.6)  
Middle lobe 8 (15.1) 10 (16.9)  
Lower lobe 5 (9.4) 5 (8.5)  
Multilobar 9 (17.0) 10 (16.9)  
Chest X-ray pattern 0.995
Cavity 9 (17) 10 (16.9)  
Patchy 8 (15.1) 8 (13.6)  
Reticulonodular 20 (37.7) 21 (35.6)  
Nodular 5 (9.4) 6 (10.2)  
Reticular 11 (20.8) 14 (23.7)  

adverse events compared with the salbutamol with self-sputum collection [Paggiaro et al. 2002;
solution, such as chest tightness (15% versus 5%) Hong Kong Chest Survice/Tuberculosis Research
and symptomatic bronchospasm requiring rescue Center Madras/British Medical Research Council,
bronchodilator (11.3% versus 0%, p = 0.02) 1979; Narain et  al. 1971; Hensler et  al. 1961].
(Table 4). Six cases with symptomatic bronchos- However, hypertonic saline itself can produce
pasm in the 3% NaCl group were treated with chest tightness and bronchospasm requiring res-
nebulized salbutamol solution (1–2 doses/case). cue bronchodilator, and even death [Saetta et al.
Symptoms were relieved and the patients were 1995]. A few studies showed that beta2-agonists
discharged within 2–4 h following treatment. could also facilitate sputum expectoration [Ansari
et  al. 2013; Bennett, 2002; Yazdani et  al. 2002;
Mortensen et al. 1991]. In the present study, race-
Discussion mic salbutamol (S-isomer salbutamol) was used
We found that SI with nebulized racemic salbuta- for SI and the results showed that the nebulizer
mol solution offered equal benefits to nebulized solution of racemic salbutamol was as effective as
3% NaCl solution in the diagnostic yield for hypertonic saline in increasing both sputum quan-
smear-negative pulmonary TB and it did not incur tity and diagnostic yield of TB without any adverse
any ADRs. events. The nebulized form of salbutamol was pre-
ferred to an oral form because of the more rapid
It was known that SI with nebulized hypertonic onset of action and fewer side effects [National
saline could facilitate sputum expectoration and Asthma Education and Prevention Program,
increase the diagnostic yield of pulmonary TB, 2007]. Our study showed that nebulized S-isomer
especially in the case of negative AFB staining salbutamol could increase the diagnostic yield of

220 http://tar.sagepub.com
 W Keeratichananont, T Nilmoje et al.

Table 3.  Diagnostic yield of sputum induction with nebulized 3% sodium chloride solution versus nebulized
racemic salbutamol solution in suspected smear-negative pulmonary tuberculosis cases.

Characteristic 3% Sodium chloride Salbutamol p value

(n = 53) n (%) (n = 59) n (%)
Acid-fact bacilli positive 7 (13.2)   9 (15.3) 0.757
Culture positive 9 (17.0) 13 (22.0) 0.502
Acid-fact bacilli or 9 (17.0) 13 (22.0) 0.502
culture positive

Table 4.  Sputum induction-related adverse events.

Characteristic 3% Sodium chloride Salbutamol p value

(n = 53) n (%) (n = 59) n (%)
Overall adverse events 18 (34.0) 6 (10.2) 0.02
Bronchospasm 6 (11.3)  0  
Cough 4 (7.5) 3 (5.1)  
Chest tightness 8 (15.1) 3 (5.1)  
No event 35 (66.0) 53 (89.8)  

TB similarly to nebulized R-isomer salbutamol as two groups. Fourth, ultrasonic nebulization,

reported in the study by Ansari and colleagues which could increase the amount of sputum, was
[Ansari et  al. 2013]. However, the percentage of not used because it was costly and not available in
positive AFB smear within the first spot-induced Thailand. However, the study showed that the TB
sputum by nebulized S-isomer salbutamol was diagnostic yield could also be increased by the use
lower than the R-isomer salbutamol (15% versus of a compressor-type nebulizer. Fifth, this was a
60%, respectively), because all patients who had tertiary hospital-based study that was confined to
positive AFB smears on 3 consecutive days of self- the more complicated participants and might
expectorated sputum were initially excluded from explain why it had a relatively high number of
our study. Therefore, these results could not be adverse events with hypertonic saline. Therefore,
directly compared. SI by salbutamol was potentially vulnerable in the
tertiary hospital compared with the primary
Our study has several strengths. It was a prospec- healthcare setting. Lastly, there were no data on
tive, randomized comparative trial, and moreover, cumulative diagnostic yield of repeated SI and
the sample size was calculated; the diagnostic also patients with asthma or COPD were not
gold standard for active TB was applied; SI was recruited to this study due to ethical issues. As
used only after the failure of diagnosis by self- patients with asthma or COPD were at risk of
expectorated sputum as suggested by Geldenhuys developing symptomatic bronchospasm, they
and colleagues (real-life practice) [Geldenhuys could not be randomized to receive SI with hyper-
et al. 2012, 2014]; SI adverse events were also tonic saline. Therefore, further studies are needed
reported. However, there were limitations to our to confirm the efficacy of repeated SI with nebu-
study. First, an observational bias could occur in lized racemic salbutamol solution, and also to
our single-blind experimental design; however, all validate the safety profile of nebulized racemic
laboratory technicians were blinded to the SI salbutamol for SI in patients with asthma and
arm. Second, a crossover design was not applied COPD.
in the study because of concern about the impact
of carryover effects and washout periods between
each SI. Third, sample size calculation was based Conclusion
on a high estimated diagnostic yield of salbuta- SI by nebulized racemic salbutamol solution
mol; this might have meant that the calculated offers equal benefits to 3% NaCl solution in
sample size was inadequate to demonstrate a increasing both sputum quantity and diagnostic
small difference in diagnostic yield between the yield in smear-negative patients suspected

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Therapeutic Advances in Respiratory Disease 9(5)

of having pulmonary TB. SI using nebulized solution in exercise-induced bronchoconstriction.

racemic salbutamol does not produce bronchos- Chest 131: 1339–1344.
pasm and chest tightness occurs less frequently, Delvaux, M., Henket, M., Lau, L., Kange, P.,
therefore, it should be considered as a good alter- Bartsch, P., Djukanovic, R. et al. (2004) Nebulised
native noninvasive method for the diagnosis of salbutamol administered during sputum induction
pulmonary TB when hypertonic saline is unavail- improves bronchoprotection in patients with asthma.
able or contraindicated. Thorax 59: 111–115.

Dunleavy, A., Breen, R., Perrin, F. and Lipman, M.

Acknowledgment (2008) Is bronchodilation required routinely before
The authors would like to acknowledge The Royal diagnostic sputum induction? Evidence from studies
College of Physicians of Thailand for giving us with tuberculosis. Thorax 63: 473–474.
the Clinical Research Award in 2013.
Geldenhuys, H., Kleynhans, W., Buckerfield, N.,
Tameris, M., Gonzalez, Y., Mahomed, H. et al.
Funding (2012) Safety and tolerability of sputum induction
The author(s) received no financial support for in adolescents and adults with suspected pulmonary
the research, authorship, and/or publication of tuberculosis. Eur J Clin Microbiol Infect Dis 31: 529–537.
this article.
Geldenhuys, H., Whitelaw, A., Tameris, M., Van
As, D., Luabeya, K., Mahomed, H. et al. (2014)
Declaration of Conflicting Interests
A controlled trial of sputum induction and routine
The author(s) declared no potential conflicts of collection methods for TB diagnosis in a South
interest with respect to the research, authorship, African community. Eur J Clin Microbiol Infect Dis 33:
and/or publication of this article. 2259–2266.
Golub, J., Bur, S., Cronin, W., Gange, S., Baruch,
N., Comstock, G. et al. (2006) Delayed tuberculosis
diagnosis and tuberculosis transmission. Int J Tuberc
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