Transcript Traditional Practices and Antibacterial Activity of Rhus vulgaris and

Chewing sticks: Socio-cultural considerations
and anti-streptococcal activity of some plants
Authors: Odongo C.O1, Musisi L.N2, Waako P1,
Obua C1
1
Department of Pharmacology & Therapeutics, School of Biomedical Sciences, College of Health
Sciences, Makerere University
2 Department of Microbiology, Faculty of Veterinary Medicine, Makerere University
Background
• Dental self-care is poor and plaque-associated dental
diseases are widespread in Uganda (Okullo, 2004; Wandera & Ntwatwa, 2003; Muwazi et al; 2002, Tirwomwe et al; 1987)
• Professional dental care services are scarce and
expensive, Needs of the majority are unmet – limited
professionals (National Oral Health Policy, 2007)
• Need to explore and develop affordable, yet effective
methods of disease prevention
• Good oral hygiene (with plaque removal) is essential in the
prevention of dental diseases
• Traditional toothbrushes (chewing sticks) have been in use for
ages and clinical studies have demonstrated their efficacy
(Almas & Al-Zeid, 2005; Al-Otaibi et al., 2003; Darout et al., 2000; Aderinokun, 1999)
• Some chewing sticks contain compounds with antibacterial or
anti-inflammatory activity (Prashant et al., 2007; Ndukwe et al., 2005; Ooshima et
al., 2000; Hu et al., 2000; Almas, 1999; Sote & Wilson, 1995)
• No studies of this kind have been done on local chewing sticks
despite reported use (Katende et al., 1998; Kokwaro, 1976, Eggeling, 1952)
Aims of the study
1. Identify and document plants from which chewing sticks
are prepared in a community in Buganda
2. Describe the preparation and use of chewing sticks
3. Evaluate antibacterial activity of aqueous extracts of
two of the most reported plants against wild cultures
of Streptococcus mutans.
Methods and Materials
Study design: Qualitative study of mixed design
• community survey using Key Informant Interviews,
• field walks and laboratory experimental assays
Study sites and selection method
• 1st: Sango & Buwanda villages (Mpigi district), conveniently selected
• 2nd: Makerere University,
Depts. of Pharmacology & Therapeutics (CHS) and Microbiology (FVM).
Selection of Key informants
• Local Council 1 Chairperson: permission and recommend “Village
Elders” as key informants.
Plant collection and identification
• Plants identified and collected with aid of Key
Informants
• Demonstrations of chewing stick preparation and use
• Vouchers prepared & stored in field, approx. 500g of
branches of Rhus vulgaris & Lantana trifolia taken for
analysis
• Taxonomic identification done by two independent
botanists, deposits at herbarium (Makerere University)
Processing & formulation of standard test solutions
• Method of Prashant et al, 2007, (& Almas, 1999).
• Briefly: 2 day sun-dried sticks crushed, weights of 5g, 10g, 50g,
• @ soaked in 100ml of de-ionised water at 4o C for 48 hrs.
• Filtrates of 5%, 10%, 50% obtained.
Modifications:
• Crushed fresh, Open air-dried for 2 weeks in cool shade,
• Only 50g used, Extraction (48 hrs at 2o C)
• Filtrates passed thru bacterial membrane filters (0.45 µm pores,
Ministart®, UK).
Assumption: extraction process progressed until there was saturation
of the aqueous phase
Isolation of Strep. mutans
• Dental plaque - transported to lab in Stuart’s medium (Sterilin®,
COPAN innovations, UK)
• Isolation on Mitis-Salivarious agar (Harold & Johannes, 1975).
• Pure plates prepared and confirmed with biochemical tests.
Antibacterial assays
• Agar-well diffusion used on Blood agar (Baker & Silverton, 1985).
• 24 hr old culture spread on three plates using sterile cotton
buds, 4 wells (4 mm deep) made onto @ plate using borers (5 mm
diameter).
• Wells were labeled and filled sequentially using separate sterile
o
pipette tips, incubated aerobically at 37 C for 48 hrs.
• Experiments set in triplicate , run concurrently.
• Extracts considered active if inhibition zone was 8mm or more
(Baker & Silverton, 1985)
Data Analysis:
• Qualitative Content Analysis (Abramson & Abramson, 1999),
• MS Excel program (bioassay results)
RESULTS
Table 1: Socio-demographic characteristics of key
informants from Sango and Buwanda villages, Mpigi district
Characteristic
Male
Female
Mean age (yrs)
55 (n=12)
59 (n=9)
Subsistence farmer
8
8
Commercial farmer
1
Office clerk b
3
Teacher b
1
Policeman b
1
Herbalist
1
3
Retail shop keeper
5
1
Consent to audio records
5
1
Occupation a
a: some key informants had multiple occupations
b: these were retired civil servants
12
Table 2a: Details of plants identified as sources of chewing
sticks in Sango and Buwanda villages, Mpigi district
Botanical name
Family
Parts used
Voucher /
accession no.
Rhus vulgaris Meikle
Anacardiaceae
Branch
37789
Mangifera indica L.
Anacardiaceae
Branch
37794
Draceana fragrans (L.)
Ker Gawl
Agavaceae
Stem
37793
Lantana trifolia L.
Verbenaceae
Branch
37792
Acacia hockii De Wild
Fabaceae
Branch
37791
Citrus sinensis (L.) Osb. Rutaceae
Branch
37790
Citrus limonia (L.) Osb.
Branch
37788
Rutaceae
Table 2b: Names of chewing stick plants in some
local dialects in Uganda
Scientific
name
Luganda
Lusoga
Luo
Runyankore
Lugbara
Ateso
Mangifera
indica
Muyembe
Muyembe
Muyeme
Aeme
Omweembe
Mombe
Emwembe
Draceana
fragrans
Oluwanyi
Oluwanyi
Lantana
trifolia
Kayukiyuki
Kasekeranyonyi
Kapanga
Belwinyo
Bel awele
Omushekyera
Omuhuuki
Obolokoanya
Amwee
Elantaana
Acacia
hockii
Kasaana
Kasone
Oryang
Rugando
Ali
Ekisim
Rhus
vulgaris
Akakansokanso
Kakwansokwanso
Busojole
Awaca
Atyendwinyo
Omukanza
Obukaanja
Abilika
Epwatet
Ewaya
Citrus
sinensis
Omuchungwa
Omuchungwa
Macungwa
Mucunguwa
Omuchungwa
Citrus
limonia
Ennimu
Ennimu
Lemun
Ndimu
Endimu
Mugorogoro
Omugorora
Source: Katende et al., 1998 & Katende et al., 1999
14
Table 3: Frequency table showing percentage
reports of use of various chewing stick plants
Plants reported as chewing Number of times reported
stick sources
(n=21)q
Percentage reports (%)
Rhus vulgaris
16
76.2
Lantana trifolia
12
57.1
Mangifera indica
9
42.9
Enzzo f
9
42.9
Cassia hockii
7
33.3
Draceana fragrans
6
28.6
Citrus limonia
5
23.8
Citrus sinensis
2
9.5
q: most plants were reported more than once by various key informants
f: name in local dialect as it was not possible to obtain sample for identification
15
Why these plants?
• These plants yield chewing sticks with:
1.
Sufficient brush-like bristles
2.
Resilient to mechanical stress of tooth brushing
3.
Palatable
• High fiber content confers the firm & fibrous character common to all
• Users learnt of these plants (preparation & use) at an early stage in life
• No medicinal value was claimed by users
Are these plants available?
• The right chewing-stick plants have drastically reduced in number within the
domestic environs
(esp. Rhus vulgaris, Mangifera indica, Cassia hockii and Enzzo)
“Kakwansokwanso ta kya labika wano kumpi, aba mwetaaga ba mujja mu tale”
Reasons:
1. Rhus vulgaris roots boiled and drunk to treat abdominal pain in pregnant
women (Mubiru et al., 1990; Katende et al., 1998) and hypertension and sugar
disease (Mwebaza et al., 2005)
2.
Source of wood fuel
3.
Clearance of wild vegetation for habitation or agricultural land
(population pressure)
Preparation of chewing sticks
• Common method of preparation
• Stems and branches (0.5-1.0 cm) cut off, bark removed,
• One end chewed into a soft brush-like fringe
Usage and storage of chewing sticks
• Pressed against teeth & moved thru fine vertical & horizontal motions,
a tooth at a time so as to avoid gum injury
• Good manual dexterity & plenty time required to use correctly and effectively.
Used once or twice daily
• Stored anywhere safe from human or animal contact (e.g. stuck between tree
branches, grass thatched roofs, above door panels, inside ventilator bricks,
top of cupboards or wardrobes etc)
Reasons for use of the chewing stick:
1. Availability, cost free,
2. Does not require toothpaste,
3. Better oral access,
4. Avoids gum injury,
5. Synthetic bristles fall into the throat & cause cough
Reasons for use of the synthetic toothbrush:
1. Scarcity of chewing-stick plants (availability of synthetic toothbrushes),
2. Durability & better performance of the synthetic toothbrush,
3. Lack of time (elaborate preparation of chewing sticks),
4. Gum injury (there was a contradiction),
5. The chewing stick is an outdated tool.
Table 4: Summary results of inhibition diameters
of Rhus vulgaris and Lantana trifolia against
Strep. mutans on 5% sheep blood agar at 48 hrs
incubation
Extract / Control
Mean diameter (mm) 95% confidence
limits of mean
(± SD) n=3
Rhus vulgaris
Lantana trifolia
24.33 (± 0.58)
22.89-25.76
14.17 (± 0.29)
13.45-14.88
Benzyl penicillin
De-ionized water
30.67 (± 0.29)
30.45-31.88
0
-
Plate 1: Blood agar plates showing inhibition zones on labeled wells of Rhus vulgaris,
Lantana trifolia, benzyl penicillin and deionized water
Conclusions:
• Rhus vulgaris and Lantana trifolia chewing sticks are most
commonly used in oral care along with synthetic toothbrushes
• Their extracts are active against Streptococcus mutans
• Preparation and use is uniform within this community, not
associated with rituals or cultural beliefs
Recommendations
• Further studies to establish active principles
• Public awareness to promote conservation
Lantana trifoila
Rhus vulgaris
A key informant demonstrates preparation of a chewing stick from the
stem of Draceana fragrans
Acknowledgements
• Gulu University
• Laboratory staff of Pharmacology, Pharmacy and Veterinary
microbiology, (Mak.)
• Dr. Namaganda Mary (Department of Botany, Makerere University)
• Dr. Katuura Esther (Natural Chemotherapeutics Research Laboratory)
• Mr. Aloysius Lubega (translator & village guide)