2.2 BACTERIAL STRAINS
Collected samples were inoculated into blood agar plates supplemented
with colistin nalidixic acid and incubated for 18-24 h at 37°C in a
candle jar enriched with 5% Carbon-dioxide. All greyish, smooth, small
and non-pigmented colonies with a visible zone of beta hemolysis
appearing 24 h after incubation were isolated, further incubation and
their reactivity to catalase evaluated. Colonies with a negative
catalase reactivity after further incubation were then isolated and used
for the confirmatory diagnosis using the Pastorex strep kit (BIO-RAD).
Colonies which agglutinated with the GBS latex reagent were considered
positive.
The resulting isolates were then used for antibiotic susceptibility
testing by the Kirby Bauer disc difusion method. The antibiotics was
tested and their respective diameters of inhibitions were measure and
compare to those of the French microbiology society as of 2018 (SFM ;
EUCAST)
2.3 ANTIMICROBIAL SUSCEPTIBILITY TESTING
Antimicrobial susceptibility testing was performed according to Clinical
and Laboratory Standard Institute Guidelines (CLSI, 2014) by disk
diffusion 14. A suspension of the test organism was
prepared by removing 3-5 colonies from a pure culture plate by
emulsifying in 3 ml of sterile physiological saline and was diluted with
saline until the turbidity of the suspension become matched with
turbidity standard equivalent to 0.5 McFarland and inoculated on
Muller-Hinton agar (MHA, Oxoid, England) with 5% sheep’s blood. After
the excess suspension was removed by gentle rotation of the swab against
the surface of the tube, the swab was then used to distribute the
bacteria evenly over the entire surface of MHA supplemented with 5%
sheep blood. The inoculated plates were left at room temperature to dry
for 3-5 minutes and a set of 6 antibiotic discs in each plate were
placed with the concentration of penicillin G (10µg), gentamicin (CN)
(10 µg), erythromycin (E) (15µg), clindamycin (DA) (2µg), tetracyclin
(10 µg), norfloxacin (10 µg), Chloramphenicol (C) (30µg), pristicin (10
µg), steptomycin (10 µg), rifampicin (10 µg) and incubated at 35-37 °C
with 5% CO2 atmosphere by candle jar for 18-24 hours.
The zone of growth inhibition was measured using rulers. The sizes of
the inhibition zones were graded according to the CLSI 2014 and
interpreted as susceptible, intermediate or resistant14 (All of the antibiotics used in the investigation
are product of Oxoid, England and HIMEDIA).