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Journal of Ethnobiology and Ethnomedicine
In vitro antimicrobial activity of methanolic extracts of different Senna didymobotrya(Fresen.) H. S. Irwin & Barneby plant parts
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Full Title: In vitro antimicrobial activity of methanolic extracts of different Senna didymobotrya(Fresen.) H. S. Irwin & Barneby plant parts
Article Type: Research
Funding Information: Deutscher Akademischer Austauschdienst(DAAD) through in-country scholarshipand the National Commission for Science,Technology and Innovation (NACOSTI)through the Women Scientist ProgramCall II
M/s Pascaline Jeruto
Abstract: AbstractBackground: Herbal medicines are used widely for primary health care in Kenyaamong rural populations where modern medicines are not affordable. The flowers,roots, stems and leaves of Senna didymobotrya have both antifungal and antibacterialactivity. Decoctions or infusion are used to treat skin diseases, diarrhoea, malaria,venereal diseases and stomach problems.
Methods: Plants were collected from farmers' fields in western Kenya. Stem bark, rootbark, leaves, flowers and immature pods were dried and milled. Methanol was used asextractant. The extracts were reconstituted and incorporated into growth media toobtain 2.5%, 5%, 7.5% and 10%. Bioassays were carried out on Escherichia coli,Staphylococcus aureus, Trichophyton tonsurans and Candida albicans. The growth ofcultures on the plates was measured over a period of eight days for bacteria andsixteen days for fungi. The area under disease progress stairs was determined andsubjected to statistical analysis.
Results: The growth of S. aureus was completely inhibited by root bark at 2.5% and thestem bark at 7.5%. Immature pods, flowers and leaves extracts did not have muchsignificant effect. Growth on E. coli was completely inhibited at 7.5% by stem barkextracts while the root bark inhibited growth at 10%. Bioassays on C. albicans showedthat all plant part extracts did not have any significant effect on its growth. Growth of T.tonsurans was completely inhibited by immature pods extract at 10%, the leaves andflowers extracts inhibited the growth at 7.5%. The stem and root bark extracts inhibitedgrowth at 5 %.
Conclusions: Stem and root bark extracts of S. didymobotrya showed effectiveantimicrobial activities against S. aureus, E. coli and T. tonsurans at low dosages.There is need to carry our research on these plant part extracts to identify the activephytochemicals that contribute to their high efficacies as compared to other plant parts.On the conservation front, harvesting of root and stem barks may lead to depletion ofthe plant. Research should focus on the concentration of the active ingredients in theother plant parts so as to increase their efficacy since they regenerate every season.
Keywords: Medicinal plants, bacteria, fungi, Senna didymobotrya, diarrhoea, ringworm
Corresponding Author: Pascaline Jeruto, PhdUniversity of EldoretEldoret, Kenya, KENYA
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Corresponding Author's Institution: University of Eldoret
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First Author: Pascaline Jeruto, Phd
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Peter Arama, Phd
Beatrice Anyango, Phd
Teresia Akenga, Phd
Regina Nyunja, Phd
Daniel Khasabuli, Bsc
John Kamundia, MSc
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1
In vitro antimicrobial activity of methanolic extracts of different Senna didymobotrya
(Fresen.) H. S. Irwin & Barneby plant parts
Pascaline Jeruto1*
Department of Biological Sciences, School of Science, Eldoret University, P.O. Box 1125 –
30100, Eldoret, Kenya.
Peter Arama2: [email protected]
Department of Agricultural Economics and Agribusiness, School of Agriculture, Natural
Resources and Environmental Studies, Rongo University College, P.O. Box 103 – 40404,
Rongo, Kenya.
Beatrice Anyango3: [email protected]
Department of Biological Sciences, School of Biological and Physical Sciences, Jaramogi
Oginga Odinga University of Science and Technology, P.O. Box 210 – 40601, Bondo, Kenya.
Teresia Akenga1: [email protected]
Department of Chemistry, School of Science, Eldoret University, P.O. Box 1125 – 30100,
Eldoret, Kenya.
Regina Nyunja3: [email protected]
Department of Biological Sciences, School of Biological and Physical Sciences, Jaramogi
Oginga Odinga University of Science and Technology, P.O. Box 210 – 40601, Bondo, Kenya.
Daniel Khasabuli.4: [email protected]
Department of Botany, Faculty of Science, Maseno University, P.O. Box 333 – 40105, Maseno,
Kenya.
John Kamundia5: [email protected]
National Plant Breeding Research Center, Kenya Agricultural and Livestock Research
Organization, Private Bag, Njoro, Kenya
*Correspondence: [email protected]
1Department of Biological Sciences, School of Science, Eldoret University, P.O. Box 1125 –
30100, Eldoret, Kenya.
In vitro antimicrobial activity of methanolic extractsClick here to download Manuscript: Manuscript 1.docx Click here to view linked References
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Abstract
Background: Herbal medicines are used widely for primary health care in Kenya among rural
populations where modern medicines are not affordable. The flowers, roots, stems and leaves of
Senna didymobotrya have both antifungal and antibacterial activity. Decoctions or infusion are
used to treat skin diseases, diarrhoea, malaria, venereal diseases and stomach problems.
Methods: Plants were collected from farmers’ fields in western Kenya. Stem bark, root bark,
leaves, flowers and immature pods were dried and milled. Methanol was used as extractant. The
extracts were reconstituted and incorporated into growth media to obtain 2.5%, 5%, 7.5% and
10%. Bioassays were carried out on Escherichia coli, Staphylococcus aureus, Trichophyton
tonsurans and Candida albicans. The growth of cultures on the plates was measured over a
period of eight days for bacteria and sixteen days for fungi. The area under disease progress
stairs was determined and subjected to statistical analysis.
Results: The growth of S. aureus was completely inhibited by root bark at 2.5% and the stem
bark at 7.5%. Immature pods, flowers and leaves extracts did not have much significant effect.
Growth on E. coli was completely inhibited at 7.5% by stem bark extracts while the root bark
inhibited growth at 10%. Bioassays on C. albicans showed that all plant part extracts did not
have any significant effect on its growth. Growth of T. tonsurans was completely inhibited by
immature pods extract at 10%, the leaves and flowers extracts inhibited the growth at 7.5%. The
stem and root bark extracts inhibited growth at 5 %.
Conclusions: Stem and root bark extracts of S. didymobotrya showed effective antimicrobial
activities against S. aureus, E. coli and T. tonsurans at low dosages. There is need to carry our
research on these plant part extracts to identify the active phytochemicals that contribute to their
high efficacies as compared to other plant parts. On the conservation front, harvesting of root and
stem barks may lead to depletion of the plant. Research should focus on the concentration of the
active ingredients in the other plant parts so as to increase their efficacy since they regenerate
every season.
Keywords: Medicinal plants, bacteria, fungi, Senna didymobotrya, diarrhoea, ringworm
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Background
Herbal medicines have been used for many years in many parts of the world especially
developing countries [1]. Despite enormous advances in conventional medicines, traditional
medicines have been encouraged by the World Health Organization [2] as a building block of
primary healthcare [3], partly because some conventional drugs have failed to prove effective,
have serious side effects [4,5], or cannot cure certain new illnesses such as HIV and AIDS.
Large body of evidence has accumulated to demonstrate the promising potential of medicinal
plants used in various traditional, complementary and alternative systems of treatment of human
diseases [6].
In tropical countries like Kenya, modern medicines are not affordable to most of the rural
populations and W.H.O. estimates that 80% of the world’s population use botanical medicines
for primary health care [7]. The World Bank report further pointed out that in 2002, Kenya’s
ministry of health budget for medicine provided for only 30% of the population [2]. This left
70% (21 million) of the population who could not access the conventional medicine. The latter
population group was therefore left to rely on traditional medicines for their healthcare needs
[8]. The use of traditional medicines remains widespread in developing countries while the use
of complementary alternative medicine (CAM) is increasing rapidly in developed countries [7].
A few African phytomedicines are available in the international market. African medicinal
plants play a key role in basic healthcare, particularly in rural areas due to their accessibility and
affordability [9]. Modern medicines contain at least 25% of active ingredients derived from
plants [10].
The plant: Senna didymobotrya (Fresen.) H.S. Irwin & Barneby
Senna didymobotrya has been widely used as a medicinal plant, especially in East Africa, where
a decoction or infusion from the leaves, stems and roots is drunk as a laxative and purgative for
the treatment of abdominal pains, while in large quantities it is taken as an emetic [11]. In
Uganda, Rwanda and Burundi it is also taken to expel intestinal worms and to treat ringworm.
Pounded leaves and young stems are used in Kenya to treat skin diseases. The pulp is applied to
the skin [12]. The leaf sap is diluted in water and orally taken to treat diarrhoea and dysentery. It
is also taken as a diuretic, laxative, and emetic. The ash of burnt twigs is used to coat the inside
of gourds that are to be used for storing milk, as it is said to improve digestibility and
palatability. The milk can be kept in them for over a year [13]. According to [14], the pastoralists
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of West Pokot peel the bark, dry the stem and burn it into charcoal for preserving and flavouring
milk. Infusion from the leaves and the roots have been used by the Nandi community for
treatment of skin diseases, malaria, gonorrhoea, ringworms, cancer, emetic and as purgative to
remove excess bile [15.16]. The objective of this study was to assess, in vitro, the efficacy of
different concentrations of S. didymobotrya plant extracts on the bacterial species
Staphylococcus aureus, Escherichia coli and fungal species Candida albicans and Trichophyton
tonsurans.
Methods
Consent
This study was part of a larger project involving collection of Senna species from the country for
conservation in situ at the Jaramogi Oginga Odinga University of Science and Technology
botanical garden. The objectives of the study and the methods were discussed with the
communities where the samples were collected. Individuals from whom the samples were
collected were asked to agree for the plant samples to be obtained from their farms and to sign a
Consent Form.
Plant sample collection
Plant samples were collected from the three geographic regions in Kenya. These were Uyawi-
Miandhe in Siaya county, Ndurio in Nandi county and Rongai in Nakuru county. The samples
were collected ethically following standard procedures [17]. At each location, 20 mature plants
from a cluster were selected. From these plants, the leaves, flowers, stem bark, immature pods
and root barks were obtained. Each of the samples from a location was packed in individual
gunny bags then transported to Maseno University Horticultural Laboratory for drying at 25 - 300
C for two weeks. Samples were shredded using an electric hammer mill before grinding using
the laboratory Willy mill at 800 rpm.
Extraction of crude extract on plant materials was carried out using methanol solvent. About one
thousand (1000) grams of powdered dried plant part was obtained. At each time of extraction,
42.25 g was placed in soxhlet apparatus wrapped in a filter paper and distilled with 300 ml
methanol. The extract was concentrated by recovering the solvent using soxhlet apparatus until
the extract became semi-solid. The mixture was then transferred to the rotary vacuum evaporator
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placed in a water bath at 40oC. Samples were transferred to a beaker and then left in the open for
24 h for complete evaporation of the solvent. The crude extracts were put in a tightly screw
capped glass containers and stored in the refrigerator at 4°C until the time of use.
Three types of media: nutrient agar (NA), potato dextrose agar (PDA) and sabouraud dextrose
agar (SDA) were prepared according to manufacturers’ instructions. After autoclaving at121oC
for 15 minutes each medium was allowed to cool to 50oC. Using a sterile measuring cylinder,
100 ml was measured and poured into 100 ml Erlenmeyer flasks placed in the laminar flow
hood.
The constitutions of the extracts were carried out according to methods of [18] and [19] with
modifications. Using analytical balance, 2.5 g, 5 g, 7.5 g and 10 g of crude extract was weighed
and placed in sterile beakers. Five millilitres of absolute methanol was then added to the solid
extract to reconstitute it. Each of the reconstituted extract was added into 100 ml of respective
medium in a conical flask, stirred and let to stand at 40-50oC. The negative control consisted of
media alone without addition of S. didymobotrya extracts. The positive control for the fungi
consisted of ketoconazole (Keto) incorporated at 10µg/ml of SDA and PDA [20]. The positive
control for the bacteria consisted of streptomycin (Str) incorporated at 1 g/L of nutrient agar [21].
The mixture was stirred and let to cool for 30 min with the flask mouths open under the laminar
flow hood. Medium from each flask was poured in four petri dishes (replicates).
The test microorganisms were chosen based on the ethnobotanical exploitation of the plant. The
test microorganisms were obtained from Kenya Medical Research Institute (KEMRI), Kisumu,
Kenya and consisted of two strains of bacteria and two strains of fungi
a) Bacteria: Staphylococcus aureus (ATCC 28923): Gram positive cocci
Escherichia coli (ATCC 25922): Gram negative rod
b) Fungus: Candida albicans (ATCC 1405)
Trichophyton tonsurans (ATCC 28942)
The two bacteria strains were cultured in nutrient agar. C. albicans was cultured in potato
dextrose agar (PDA) while T. tonsurans was grown in sabouraud dextrose agar (SDA).
The bacterial isolates were obtained from the petri dishes using a sterilized loop and placed in 10
ml of distilled water in a sterilized tube. It was stirred and then serially diluted from 10-1
to 10-5
.
The original sample was stored in a refrigerator at 4oC. One millilitre each was obtained from the
last two dilutions and plated on nutrient agar. The Petri dishes were incubated at 37oC for five
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days. The number of colonies developing in each plate was counted. Using this, the colony
forming units (cfu) in the original sample was determined and adjusted so as to obtain 108
cfu/ml.
A micropipette was used to obtain 50 µl of the inoculum. This was inoculated in the middle of
the petri dish which was then incubated at 37oC in a cooled incubator (Gallenkamp- IR 701SG).
The experiment was set with four replicates in a completely randomized design (CRD). Colony
growth was assessed at an interval of 2 days for 8 days. At each observation date, the diameter of
the colony growth was measured.
Fungal cultures of C. albicans and T.tonsurans were obtained from the plates and a spore
suspension of the respective isolate was prepared. Inoculum was placed in 50ml sterile distilled
water in a test tube. From this, serial dilution was made to 10-3
. Using a haemocytometer slide
the spore concentration in the last dilution was determined. The concentration was adjusted to108
spores/ml. From each spore suspension of the two fungal species, 50 µl of the inoculum was
inoculated as a drop on the respective agar medium. The experiment had four replicates in a
completely randomized design. Observations were made at an interval of four days. Four
observations were made in each experiment.
Data analysis
The diameter (mm) of the colony size of both bacterial and fungal cultures were used to calculate
the actual surface area (mm2) covered by the pathogen. The disease areas for the different
observations were summarized using the area under disease progress stairs (AUDPS) [22].
[
]
Where AUDPC is area under disease progress curve given with formula;
AUDPC =[½ Obs1t1 + Obs2t2+Obs3t3+½Obs4t4]
Where: t1, t2, t3 and t4 are the time intervals of the first, second, third and the fourth observation.
Obs1, Obs2, Obs3 and Obs4 are first, second, third and fourth observation respectively.
y1 and yn are assessments at the first and last observations respectively; D = tn-t1.
n is the total number of observation
The AUDPS was subjected to Analysis of Variance (ANOVA) and comparison of means was
carried out on all data using Statistical Analysis System (SAS). Means were separated by the use
of the least significant difference and compared at P = 0.05 for significance.
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Results
S. aureus
Table 1: The area under disease progress stairs (AUDPS) of growth of S. aureus on culture
media impregnated with different concentrations of S. didymobotrya obtained from different
plant parts
Plant part Concentration (%)
0 2.5 5 7.5 10 Str
1 g/L
Immature pods 296.9 b 224.7 b 193.8 a 180.2 a 172.0 a 0
Leaves 307.6 b 195.6 c 180.2 a 188.9 a 155.8 a 0
Flowers 344.9 a 243.1 a 182.9 a 135.8 b 113.6 b 0
Stem bark 334.4 a 97.8 d 43.6 b 0 c 0 c 0
Root bark 304.2 b 0 e 0 c 0 c 0 c 0
LSD=16.8
Means followed by the same letter in a column are not significantly different (P>0.05) using
LSD
Results in table 1 indicated that the control (0%) had a mean range colony area colony growth of
296.9 – 344.9 with a mean colony area of 317.6. There were no colony growth of S. aureus
(Table 1) and E. coli (Table 2) observed on plates inoculated with streptomycin sulphate. When
media were impregnated with 2.5% concentration of the extract, there was no culture growth
observed on the medium containing root bark extract. This was also observed on medium
impregnated with 5%, 7.5% and 10% of the root bark extracts. Media impregnated with 2.5%
stem bark extract had mean colony area of 97.8. Media treated with immature pods, leaves and
flowers had lower colony growths as compared to the control. The same trend was observed at
5% where root extract had no culture growth on medium whilst stem bark had mean culture
growth area of 43.6. When media was impregnated with 7.5% and 10% of stem bark and root
bark extracts there was no growth of bacteria observed (Table 1).
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8
E. coli
Table 2. The area under disease progress stairs (AUDPS) of growth of E. coli on culture media
impregnated with different concentrations of S. didymobotrya obtained from different plant parts.
Plant part
Concentrations (%)
0 2.5 5 7.5 10 Str 1 g/L
Flowers 197.3 b 251.6 a 300.4 a 246.7 a 164.9 b 0
Immature pods 168.2 c 217.3 b 249.1 b 240.4 a 214.9 a 0
Leaves 229.1 a 201.8 b 198.7 c 189.1 b 182.2 b 0
Root bark 206.7 b 78.4 c 7.1 d 7.8 c 2.2 c 0
Stem bark 162.4 c 98.2 c 28.7 d 0 c 0 c 0
LSD = 23.61
Means followed by the same letter in a column are not significantly different (P>0.05) using
LSD
Table 2 shows the negative control treatments had AUDPS range of 162.4 – 229.1 with a mean
of 192.7. When medium was impregnated with 2.5% extract, root bark and stem bark had
AUDPS of 78.4 and 98.2 respectively. These were not significantly different (p>0.05). Flowers,
immature pods and leaf extracts had AUDPS range of 201.8 – 251.6. At concentration of 5% the
AUDPS for root bark was 7.1 whilst that of stem bark was 28.7. They were not significantly
different (P>0.05). The same trend was observed at concentrations of 7.5% and 10% whereby at
7.5% and 10%, there was no culture growth on the medium treated with stem bark. Root bark
extracts had AUDPS of 7.8 and 2.2 at 7.5% and 10% concentration respectively.
C. albicans
Table 3. The area under disease progress stairs (AUDPS) of growth of C. albicans on culture
media impregnated with different concentrations of S. didymobotrya obtained from different
plant parts.
Plant part
Concentrations (%)
0 2.5 5 7.5 10
Keto
(0.10µg/ml)
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Immature pods 331.1 a 449.8 a 395.1 a 388.0 a 335.1 a 0
Leaves 339.6 a 370.2 b 367.1 a 373.3 ab 352.4 a 0
Flowers 334.2 a 377.3 b 389.3 a 367.6 ab 305.3 ab 0
Stem bark 371.6 a 260.0 c 280.9 b 302.7 c 286.7 b 0
Root bark 354. 7 a 267.6 c 264.9 b 328.9 bc 269.3 b 0
LSD= 47.9
Means followed by the same letter in a column are not significantly different (P>0.05) using
LSD
The negative control treatments had a mean range colony area colony growth range of 331.1-
371.6 with mean colony area of 346.2. There were no colony growth of C. albicans (Table 3)
and T. tonsorans (Table 4) observed on plates inoculated with Ketoconazole at 0.10µg/ml. The
media treated with stem bark and root bark had lower colony growth as compared to the control
(Table 3). When medium was impregnated with 2.5% extract, the stem bark and root bark had
AUDPS of 260.0 and 267.6 respectively. These were not significantly different (P≥0.05).
Leaves, flowers and immature pods had AUDPS range of 370.2- 449.8. At concentration 5%, the
AUDPS for stem bark was 280.9 whilst that of the root bark was 264.9. They were not
significantly different (P≥0.05). The same trend was observed at concentration of 10%.
Immature pods, leaves and flowers were not effective as from 2.5%, 5%, 7%, and 10% as
compared to the control (Table 3) above.
T. tonsurans
Table 4. The area under disease progress stairs (AUDPS) of growth of T. tonsurans on culture
media impregnated with different concentrations of S. didymobotrya obtained from different
plant parts.
Plant part
Concentrations (%)
0 2.5 5 7.5 10
Keto
(0.10µg/ml)
Immature pods 659.1 ab 508.4 a 430.2 a 91.6 a 0.0 a 0
Leaves 649.8 b 508.4 a 254.2 c 0.0 b 0.0 a 0
Flowers 626.2 c 468.4 b 298.2 b 0.0 b 0.0 a 0
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10
Stem bark 675.6 a 280.0 c 0.0 e 0.0 b 0.0 a 0
Root bark 646.2 bc 201.8 d 78.2 d 0.0 b 0.0 a 0
LSD= 22.5
Means followed by the same letter in a column are not significantly different (P>0.05) using
LSD
The results in Table 4 above indicated that the control (0%) had a mean range colony area of
colony growth of 675.6 - 626.2 with mean colony area of 652.5. The media treated with
concentrations of 2.5% of root bark extract showed the lowest growth of 201.8 while the stem
bark had the second lowest growth of 280.0. Immature pods, leaves and flower extracts had
lower colony growths as compared to the control. The same trend was observed at 5%. Media
impregnated with 5% of stem bark extracts showed no growth of T. tonsurans. This was also
observed on the media impregnated with 7.5% and 10% of the stem bark extracts. Root bark
extracts showed AUDPS value of 78.2 at 5%. At 7.5%, the immature pods had the least AUDPS
of 91.6 and the other parts had no growth. At concentration of 10%, there was no growth
observed in all media treated with the various extracts from the different parts of S.
didymobotrya.
Discussions
Bacterial bioassays
S. aureus
Attempts have been carried out by several authors in the world in managing S. aureus using
alternative and herbal medicine. The antibacterial activity of leaf juice and extracts of Moringa
oleifera determined in vitro using disc diffusion and minimum inhibitory concentrations
displayed a potential antibacterial activity against S. aureus [23]. The leaf aqueous extracts of
Euclayptus camaldulensis (Dehnh) using agar well diffusion technique was active against S.
aureus even from a low concentration of12 mg/ml. The zones of inhibition increased in a dose
dependent manner and it was bactericidal at 50mg/ml [24]. Results presented in Table 1 and
Table 5 showed that the 95% reduction of colony size was not achieved by the extracts from
immature pods, leaves, flowers at the highest concentration of 10% tested. Stem bark and root
bark extracts achieved 100% colony size reduction at the concentrations of 7.5% and 2.5%
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11
respectively. [25] reported that the antibacterial activity of S. didymobotrya leaves studied was
associated with the presence of phytochemicals which included tannins and alkaloids. It is
probable that the plant extracts from root and stem barks had higher concentrations of the
phytochemicals than the other plant parts tested.
Table 5. The mean Area Under Disease Progress Stairs (AUDPS), the 95% and 99% reductions
of the mean colony sizes of the four test organisms
Test organism Mean colony
size
95% reduction in
mean colony size
99% reduction in
mean colony size
S. aureus 317.6 15.9 3.2
E. coli 192.7 9.6 1.9
C. albicans 346.2 32.6 6.5
T. tonsurans 652.5 17.3 3.5
E. coli
There are numerous reports on the use of traditional plants for the treatment of diarrheal diseases.
For instance, through an ethnobotanical survey in Samburu community (Kenya), Ocimum suave,
Ormocarpum trachycarpum, Lannea triphylla, Salvodora persica, Solanum incanum, Teclea
simplicifolia, Thylachium africanum and Solanum aculeatissium were found to be used to treat
diarrhea [26]. According to [15], five plants; Justicia betonica, Cyathula schimperiana, Lannea
schimperi, Rhusna talensis and Periploca linearifolia were used in Nandi county, Kenya as anti-
diarrhea. In this study it was observed that just like in S. aureus, the extracts from flowers,
immature pods and leaves did not achieve 95% reduction in colony sizes at the highest 10%
extract concentration tested (Tables 2 and 5). Root bark extract achieved 95% colony size
reduction at concentration of 5% but did not achieve 99% reduction at 10% concentration. Stem
bark extracts achieved 100% colony size reduction at 7.5% concentration.The results obtained in
this research are in conformity with those obtained by [27], in which the plant roots were found
to inhibit the growth of E. coli with zone of inhibition of 13.33± 0.667 and this was associated
with the presence of phytochemicals like saponins, tannins, terpenoids, flavonoids, phenols,
alkaloids and steroidal rings. The crude extracts of S. didymobotrya showed activity that
surpassed those of the standard antibiotics suggesting that they could be used in treating diseases
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12
caused by E. coli with bactericidal value of 100mg/ml [24]. These authors [24] linked the results
to the presence of saponins and tannins that have been found useful in human diet for controlling
cholesterols and tannins have shown potential antiviral, antibacterial and antiparasitic effects.
This is an indication that the stem bark is more effective to E. coli. In other studies, [14] found
that methanolic extract of S. didymobotrya stem charcoal without the bark inhibited E. coli.
Findings in our study concurs with those of [28] who found that increased concentration of up to
15% methanol and aqueous extracts of Terminalia chebula inhibited growth of both E. coli and
S. aureus. Immature pods extracts showed higher colony counts compared to the control hence
an indication that the immature pods are not effective in management of E. coli and its related
illnesses. Cassia auriculata flower extracts exhibited significant activity against E. coli and S.
aureus [29]. This research found positive antibacterial activity of the leaves which concurs with
that of [25] where the leaf extract showed the highest average zone of inhibition against
Escherichia coli (12.17 mm) and Streptococcus pyogenes (11.67 mm). The leaf and root bark
extracts of Tabernaemontana stapfiana were the most active extracts that produced significant
activity against most of the test microorganisms [30]. The ability of these S. didymobotrya
extracts to be sensitive to both gram positive and gram negative bacteria is a clear indication of
their broad spectrum antimicrobial activity.
Fungal bioassays
C. albicans
Results presented in Tables 3 and 5 indicated that at 10% extract concentration none of the plant
parts extracts reduced the pathogen colony size by 95%. This means that S. didymobotrya plant
extracts were not effective against C. albicans. Other medicinal plants have been found to have
effective activity against C. albicans. The dichloromethane (DCM): methanol (1: 1) extracts of
Phyllanthus amarus and Phyllanthus odontadenius have been reported with strong antimicrobial
activity against C. albicans [31]. Decoctions of the leaves of Dodonaea viscosa var. angustifolia
(hopbush) have been used for the treatment of oral infections [32,33]. Glycyrrhiza glabra
(liquorice) and Polygala senega (Seneca snakeroot) are used extensively in Europe as treatment
for oral candidiasis as both plants as been reported to contain saponins compounds known to
possess antifungal activity [34]. Flavonoids isolated from Eysenhardtia texana and Termanalia
bellerica have been reported to possess antifungal activity against C. albicans [35]. Possibly S.
didymobotrya extracts did not have sufficient amount of these biochemical agents to be effective
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against C. albicans. More studies need to be carried out to determine the quantities of the
biochemical in the plant parts.
T. tonsurans
Several pharmaceutical drugs have been used in tinea treatment which is subjected to a
confirmatory test by microscopy/culture. Drugs like terbinafine 250mg, nystatin, and use of
selenium or ketoconazole shampoo. Oral antifungal agents like terbinafine, fluconazole and
itraconazole are used in treatment of tinea capitis. Griseofulvin is known as the gold standard
therapy for tinea capitis and its efficacy is decreasing with time [36]. Results presented in this
study indicated that immature pods extracts reduced the colony size by 100% at the
concentration of 10%. Leaves and flower extracts reduced the colony size by 100% at the
concentration of 7.5% (Tables 4 and 5). Root bark and stem bark extracts reduced the colony size
by 100% at the concentrations of 7.5% and 5% respectively indicating that these plant parts had
higher concentrations of the biochemical agents that were effective against T. tonsurans. The
root extract was the best in suppression of colony growth and the immature pods extract was the
least effective. These results show that S. didymobotrya can be used in management of diseases
cause by T. tonsurans and related species.Pharmacological studies by various groups of
investigators have shown that S. didymobotrya possesses significant biological activity, such as
antibacterial, antibiofilm, antifungal and antioxidant properties. The antimicrobial activity of
plant extracts can be attributed to not only a single bioactive principle but also due to the
combined action of other compounds [37].Other authors have shown that some plant extracts
used in management of skin diseases in traditional communities’ possess antioxidant,
haemostatic, analgesic properties as well as immune stimulating activities [38,39].
Conclusion
S. didimobotrya stem bark and root barks have shown effective antimicrobial activities against S.
aureus, E. coli and T. tonsurans at low dosages. There is need to carry our research on these
plant part extracts to identify the active phytochemicals that contribute to their high efficacies as
compared to leaf, flower and immature pods extracts. On the conservation front, harvesting of
these plant parts may lead to depletion of S. didymobotrya because they are potentially
destructive unlike harvesting of the leaves, flowers and immature pods that regenerate every
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season. Research should focus on the concentration of the active ingredients in the leaves,
flowers and immature pods so as to increase their efficacy. It was observed that S. didymobotrya
plant extracts were not effective against C. albicans.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
PJ, BA and PA jointly initiated the project. PJ and DK carried out the laboratory
research/bioassays. PJ, RN and TA collected the plant samples from the field and prepared them
for the bioassays. JK analyzed and interpreted the data. BA, PA and PJ led the writing and final
approval of the paper. All authors read and approved the final manuscript.
Acknowledgements
We acknowledge financial support by Deutscher Akademischer Austauschdienst (DAAD)
through in-country scholarship and the National Commission for Science, Technology and
Innovation (NACOSTI) through the Women Scientist Program Call II. The Kenya Medical
Research Institute - Kisumu enabled us carry out this research by making available the test
microorganism. We highly appreciate the assistance of Prof. Netondo and Prof. Mang’uro of
Maseno University for allowing us to use the laboratory facilities at the Department of Botany
and Department of Chemistry respectively. We wish to thank the technical staff of both
laboratories for their technical support and also our colleagues at Jaramogi Oginga Odinga
University of Science and Technology collaborators who accorded us moral support and
assistance during the study. We thank all local communities in Nandi, Nakuru and Siaya counties
who contributed Senna plant specimens used in this study
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