Effect of Nigella sativa Extract on Bacteriology Parameter of
Multibacillar leprosy Patients Administering MDT-WHO: Three
Months Observation
Dian Kusuma Dewi Ramadhani, Renni Yuniati, Subakir, Asih Budi Astuti, Falah Faniyah
and Prasetyowati Subchan
Faculty of Medicine, Diponegoro University, Semarang, Indonesia
Keywords: multibacillar leprosy, Nigella sativa, bacterial index, morphology index, MDT-WHO
Abstract: MDT-WHO implementation has decreased leprosy global prevalence. However, number of new cases
detected is still high. Therefore, new appropriate strategy, besides early detection and WHO-MDT treatment
which are the main strategies in leprosy eradication, is needed. The aim of this study was to investigate the
effects of Nigella sativa extract on bacteriology parameters of multibacillar leprosy patients administering
MDT-WHO. It was Double Blind - Randomized Controlled Trial study which was conducted in Jepara,
Central Java Province, Indonesia during February to June 2016. This study was performed on 60 eligible
multibacillar leprosy patients administering MDT-WHO. Bacterial index (BI) was measured using Ridley
logarithmic scale, whereas morphology index (MI) was measured as percentage of solid bacteria to the all
forms of bacteria. There was no subject who dropped out. The characteristics of subjects were similar between
the two groups, except for the MDT duration at baseline, where MDT duration in control group was longer.
There was no significant difference in BI proportion and MI proportion in the two groups after treatment.
Average BI in month 2 and 3 tend to be lower than control although not statistically significant. There were
significant differences of average BI decline between control and treatment groups in month 2 (p=0.000) and
month 3(p=0.005). There were statistically significant differences of average MI and average MI decline
between control and treatment groups in month 1 (p=0,000, p= 0.001, respectively). This study demonstrated
that Nigella sativa extract affected average MI and BI decline differences. Its efficacy as adjuvant therapy
might be confounded by duration of MDT-WHO.
1 INTRODUCTION
Leprosy is still world burden. Leprosy is infectious
disease that does not only affect medical condition,
but also social, economy, culture, and national
defense aspects. In most of the cases, leprosy does not
cause disability when it first manifests. Conditions
contributing to disability and deformity are
preventable when early detection can be performed
(Kemenkes RI, 2012).
Multi Drug Therapy recommended by World
Health Organization (MDT-WHO) has decreased
leprosy global prevalence from 5.2 million cases in
1980 to 200,000 cases in 2014. However, new cases
detected in 2014 were still high that is about 220,000-
250,000 new cases are diagnosed every year (Smith
& Aerts, 2014). Indonesia is a country with the third
highest leprosy prevalence in the world after India
and Brazil. Central Java is the province which has the
second highest new cases of leprosy in Indonesia
(Kemenkes RI, 2012). Therefore, new appropriate
strategy, besides early detection and WHO-MDT
treatment which are the main strategies in leprosy
eradication, is needed. To date, there are some studies
that have investigated micronutrients as adjuvant
therapy in leprosy treatments as well as to prevent
contact from getting infection (Smith & Aerts, 2014;
Vazquez et al., 2014; Rahfiludin, 2011).
Lepromatous or multibacillar (MB) leprosy has
lower cellular immunological reaction against M.
leprae. Multibacillar leprosy was characterized by
high bacillary load and high number of skin lesions.
Thus, untreated MB patients are the main source of
leprosy transmission (Walker & Lockwood, 2006).
The presence of acid-fast bacilli on skin tissue
scrapings is considered as one of the main signs of
Ramadhani, D., Yuniati, R., Subakir, ., Astuti, A., Faniyah, F. and Subchan, P.
Effect of Nigella Sativa Extract on Bacteriology Parameter of Multibacillar Leprosy Patients Administering MDT-WHO: Three Months Observation.
DOI: 10.5220/0008149600090013
In Proceedings of the 23rd Regional Conference of Dermatology (RCD 2018), pages 9-13
ISBN: 978-989-758-494-7
Copyright
c
2021 by SCITEPRESS Science and Technology Publications, Lda. All rights reserved
9
leprosy and sufficient to establish a diagnosis of
leprosy. In 1982, WHO recommended the use of
MDT as a standard treatment for leprosy and
introduced an operational classification of leprosy
which differentiated leprosy into PB and MB based
on Reitz serum smear. Bacterial Index (BI) and
Morphological Index (MI) are used for the serum
Reitz smear assessments. Bacterial Index is a semi-
quantitative measure of leprosy bacillus density in the
smear. BI helps in determining the type of leprosy and
assessing the outcomes of the treatment. Morphology
Index is a percentage of the solid form to all forms of
leprosy bacillus. The Morphology Index is useful for
assessing the transmission and the outcomes of the
treatment and determining drug resistance
(Kemenkes RI, 2012).
Black cumin/ black seed (Nigella sativa) belongs
to Ramunculaceae family and is popular as herbal
medicine.(6) The efficacies of Nigella sativa that
have been widely known by the community included
as anti-parasite, antimicrobial, anti-inflammatory,
improving liver and kidney function, treatment for
respiratory and digestive disorders, and immune-
modulator. Some studies showed that continue
Nigella sativa administration can improve immune
responses. Nigella sativa oil has potential
potentiation effect to cellular immune response
through Th1 modulation and Th2 suppression,
meanwhile the other contents can suppress cell-
mediated immune system (humoral) (Salem, 2005;
Boskabady et al., 2011).
To our knowledge, there has not been any
research article evaluating the effect of Nigella sativa
extract on bacterial parameter in MB leprosy patients.
Therefore, this study sought to study the effects of
Nigella sativa on bacteriological parameters of MB
leprosy patients.
2 METHODS
The double blind-randomized control trial was
conducted from February to June 2016 in Donorejo
Hospital, Pati Regency, Central Java Province,
Indonesia. Donorejo Hospital was referral hospital for
leprosy.
Eligible subjects for this study were those who
gave their consent after thorough explanation and met
inclusion and exclusion criteria. Subjects were men
and women aged 20-60 years who had not received
MDT-WHO or had been on MDT-WHO for at least 1
month, did not have leprosy reaction, did not pregnant
or breastfeed, and did not consume any drugs with
immune-suppressant or immune-modulator effects in
the past 1 month. All subjects who did not follow
research protocols or experienced drug eruption due
to MDT and Nigella sativa are excluded.
Multibacillary leprosy was diagnosed according
to WHO classification. Ritz serum from both ear
lobules and active lesion were stained with acid-fast
stain and observed under microscope to obtain MI
and BI indexes. BI was calibrated using Ridley
algorithm and scaled from 0 to +6 points. MI was
counted from the percentage of solid mycobacterium
to all mycobacterium observed in the smear. MI was
expressed as percentage.
Subjects who were enrolled to this study were
randomized into two groups, i.e. treatment group and
controlled group. Subjects in controlled group were
asked to take two capsules of 500 mg Nigella sativa
oil, three times daily for 3 months. Subjects in control
group were asked to take placebo capsules in the same
frequency and duration. Each placebo capsules
contain 1000 mg lactose and had the same appearance
as Nigella sativa oil capsules. All of subjects received
WHO-MDT. Evaluation of BI and MI through Reitz
serum smear was performed in month 1, 2, and 3.
This study was approved by Health Ethics
Committee of Medical Faculty, Diponegoro
University and Kariadi Central Hospital, Semarang,
Indonesia.
3 RESULTS
During study period there were 65 patients who
clinically diagnosed having MB leprosy and only 60
subjects had positive Reitz serum. Therefore, only 60
patients were enrolled to this study. Through
randomization, the subjects were divided equally into
treatment and control groups. There was no dropped
out subject. All of subjects’ characteristics, except
MDT duration, were similar in the two groups. The
MDT duration in the treatment group (Mean ± SD,
1.67 ± 1.3) was significantly shorter than in the
control group (Mean ± SD, 2.53 ± 1.1), p = 0.007.
RCD 2018 - The 23rd Regional Conference of Dermatology 2018
10
Table 1. Average BI and MI in month 0, 1, 2, and 3
Treatment Control P
N Mean ± SD
Median
(Min – Max.)
N Mean ± SD
Median
(Min – Max.)
BI Index
Month 0 30 4.53 ± 0.97 4 (3 – 6) 30 3.87± 0.70 4 (3 – 5) 0.007
a
Month 1 30 2.43 ± 0.94 3 (1 – 4) 30 2.13± 0.86 2 (1 – 3) 0.188
a
Month 2 30 0.23± 0.63 0 (0 – 2) 30 0.47± 0.73 0 (0 – 2) 0.097
a
Month 3 30 0 0 (0 – 0) 30 0.03 ± 0.18 0 (0 – 1) 0.317
a
MI Index
Month 0 30 32.00± 27.72 15 (10 – 80) 30 12.00 ± 9.61 10 (0 – 60) 0,000
a
Month 1 30 6.33 ± 5.56 10 (0 – 20) 30 4.67± 5.07 0 (0 – 10) 0,000
a
Month 2 30 0 0 (0 – 0) 30 0 0 (0 – 0)
Month 3 30 0 0 (0 – 0) 30 0 0 (0 – 0)
Table 2. Average of BI and MI decline difference
Treatment Control P
Mean ± SD
Median
(Min – Max.)
Mean ± SD
Median
(Min – Max.)
IB month 1 2.10± 0.88 2 (0 – 3) 1.73± 0.69 2 (1 – 3) 0.064
a
IB month 2 4.30± 0.95 4 (2 – 6) 3.40± 0.86 3 (2 – 5) 0.000
a
IB month 3 4.53 ± 0.97 4 (3 – 6) 3.83 ± 0.70 4 (3 – 5) 0.005
a
IM month 1 25.67± 25.28 10 (0 – 70) 7.33± 9.80 10 (0 – 50) 0.001
a
IM month 2 0 0 (0 – 0) 0 0 (0 – 0)
IM month 3 0 0 (0 – 0) 0 0 (0 – 0)
There was no significant difference of bacterial
index proportion at baseline (month 0) between
treatment group and control group. There were BI
proportion changes in both groups in month 1, 2, and
3. However, the changes were not statistically
significant (data was not shown). The average BI of
treatment group in month 0 is significantly higher
from the control group (p= 0.007). However, in the
following months the difference became
insignificant, and even average BIs in treatment
group tended to be lower in month 2 and 3 as shown
in Table 1.
There was no significant difference of average BI
decline ( BI) in month 1 between two groups.
However, the comparison was statistically significant
in month 2 (p= 0,000) and month 3 (p= 0,005) as
shown in the Table 2.
At baseline there was statistically significant
difference of MI proportion between the two groups
(p= 0.037). After 1 month observation, significant
difference of MI proportion was not observed (p=
0.394). Statistics analysis could not be obtained in
month 2 and 3 due to all subjects in group treatment
were in MI 0% (data was not shown).
Average MI at baseline and month 1 were
significantly different between the two groups as
shown in the Table 1.
Average of MI decline
( IM) of treatment group in month 1 was
significantly higher than control group (p= 0.001) as
shown in the Table 2. Bivariate analysis could not be
obtained in month 2 and 3 due to average MI in both
groups were 0.
4 DISCUSSION
MDT duration between the two groups was
significantly different at baseline suggested that MDT
duration could confound interpretation of Nigella
sativa administration effect in this study. One or two
doses of rifampisin killed 10
5
organisms, decreased
organism numbers into 10
4
in MB leprosy, and
eliminated most of them. Rifampisin was the most
effective anti-leprosy drug and could decrease MI of
lepromatous leprosy into 0% in 5 weeks (Bryceson et
al., 1990). Therefore, significant MI and BI decline in
this study might be due to MDT or Nigella sativa
administration.
Administration of Nigella sativa capsules 1000
mg three times daily could significantly decrease the
average of BI decline difference on month 1, 2, and 3
compared to baseline. The average BI decline
difference in treatment group (4.53 ± 0.97) was
higher than the control group (3.83 ± 0.69). However,
Effect of Nigella Sativa Extract on Bacteriology Parameter of Multibacillar Leprosy Patients Administering MDT-WHO: Three Months
Observation
11
BI proportion difference between the two groups in
month 1, 2, and 3 were not statistically significant.
Those suggested that Nigella sativa did not result in
different BI outcomes.
At baseline, there was significant difference of MI
proportion in both groups. However, after 1 month
therapy, the proportion comparison became not
statistically significant. This suggested that Nigella
sativa did not significantly decreased MI decline in
treatment group compared to MI decline in control
group.
Immunity mechanisms relied on Accessory
Immune System (AIS) reactivation which was
responsible for leprosy and stimulation of T or B
lymphocytes. T cells regulated T cytotoxic cells
induction, B lymphocyte function and macrophage
function by releasing cytokines. Th1 cells release
IFN- which would activate macrophages to kill
bacteria or to inhibit microbe growth and would
trigger T cytotoxic cell response resulting in self-
healing disease (Williams & Kupper, 2012). In
contrast, Th2 cells facilitated humoral immune
response and inhibited cellular immune response
which caused progressive infection. There were cross
regulatory cytokines. Th1 released IFN- which down
regulated Th2. Th2 released IL-4, IL-5, and IL-10
which inhibited and down regulated Th1 and
macrophages. (Williams et al., 2012; Abulafia &
Vignale, 2001).
Studies had demonstrated that Nigella sativa
extract had immune-modulator effect. Nigella sativa
had strong potentiation effect on cellular immunity
(mediated by T cell), as well as suppression effect on
humoral immunity which was mediated by B
lymphocytes. A study which was conducted more
than a decade ago, showed that Nigella sativa could
increase human immunity when was used regularly.
Most of subjects who administered Nigella sativa for
4 weeks showed increasing T CD4 to CD8 ratio by
55% and increasing natural killer cell function by
30% (Salem, 2005).
Boskabady et al. studied about Nigella sativa
immune-modulator effects in albumin desensitized
guinea pigs and observed the increase of IFN- and
the decrease of IL-10, suggested that Nigella sativa
had inhibition effect on Th2 cell and its cytokines and
stimulation effect on Th1 and its cytokines
(Boskabady, 2011). Study on cytomegalo-virus
infected murine showed that serum IFN- level was
associated with undetected virus on the 10
th
day
(Salem, 2000). Study on allergic rhinitis patients
showed that supplementation with Nigella sativa 2
gram daily for 30 days would increase PMN
fagocytosis activity and intracellular killing (Isik et
al., 2010).
In lepromatous leprosy, immune response against
M. leprae was dominated by T-suppressor cells
(CD8
+
, CD28
-
), with only small amount of CD4
+
.
Naive T cells commonly could not be activated
immediately. In this type of leprosy, high T2 activity
also observed, which should increase IL-4, IL-5, and
IL-10 productions. These cytokines would stimulate
B lymphocytes transformation into antibody-
secreting cells which produced antibody.(Bryceson et
al., 1990; Williams & Kupper, 2012; Abulafia &
Vignale, 2001) Therefore, Nigella sativa
supplementation to MB leprosy patients
administering MDT-WHO would improve patients’
prognosis.
It was possible to increase Nigella sativa clinical
efficacy and effect on lipid metabolism by means of
increasing the dose. However, it would increase the
gastrointestinal side effect. Daily 40 mg/kg Nigella
sativa administration was well tolerated by adults and
children. There was no side effect observed, except
for children who received 80 mg/kg dose (Kalus et
al., 2003) Other toxicity study demonstrated that 1
g/kg Nigella sativa supplementation for 28 days did
not elevate liver enzyme and did not cause toxicity
effect on the liver function (Dollah et al., 2013).
There were some limitations in this study. There
was inhomogeneity of MDT duration at baseline due
to the difficulty of finding new untreated cases as the
study subjects. In addition, in Donorojo Hospital,
Reitz serum examination were only performed on
serum from right and left ear lobules, whereas this
study only included patients from Donorejo Hospital.
The other limitation of this study was the study only
gave one supplementation dose to subjects.
5 CONCLUSIONS
This study had not proved Nigella sativa effect on
decreasing BI and MI of MB leprosy patients.
However, there was significant difference of average
MI and BI decline between treatment and control
group. Further studies should consider the subjects
homogeneity especially for MDT duration, enroll
new untreated cases only, use various Nigella sativa
doses, and be performed in larger field.
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Effect of Nigella Sativa Extract on Bacteriology Parameter of Multibacillar Leprosy Patients Administering MDT-WHO: Three Months
Observation
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