Unravelling Streptococcus agalactiae: Serotype distribution, virulence factors, obstetrics history and clinical presentation correlations in tertiary care hospitals of the western province of Sri Lanka.

PURPOSE: This study investigated to detect serotypes and virulence genes of Group B Streptococcus (GBS) isolated from pregnant women. METHODS: Forty-five samples of GBS isolates from January to August 2019 at antenatal clinics of 4 teaching hospitals in Western Province, Sri Lanka were included. Isolated GBS were carried to identify 9 serotypes by multiplex PCR. Different virulence determinants, including bac, rib and scp(B) have been detected by PCR. RESULTS: Among GBS-positive culture isolates most abundant serotype detected was type III 12/45 (26.7%) while serotype VII, VIII and IX were not seen. Furthermore, serotype Ia (15.6%); II (20%); V (17.8%); VI (15.6%); Ib (2.2%) and IV (2.2%) were identified. Among 5 rectal isolates, 1 isolate was serotype Ia, 2 isolates were serotype II and 2 isolates were serotype III. Forty (40/45) isolates expressed scpB gene (88.8%). Presence of rib gene was confirmed in 17.8%, bac in 13.3% isolates. ScpB, rib and bac were identified in 4.4% isolates, 8.9% isolates were scpB, rib positive and bac negative, 8.9% isolates were scpB, bac positive and rib negative. These three-virulence genes did not express in 8.9% isolates. ScpB gene was found once in serotype Ib and IV and all serotype VI expressed scpB gene. Rib gene was more common among serotype II and it was not found in serotype Ib, IV and VI. Bac gene was more common in serotype V and it was not found in serotype Ia, Ib and IV. There was not significant association between serotypes and virulence gene (p > 0.05). CONCLUSION: Serotype III is the most abundant serotype. In formulation of vaccine against GBS for Sri Lanka, serotype III should be targeted. Prevalence of vaccine candidate virulence protein such as ß antigens of the C protein (bac) and surface protein Rib (rib) genes were low in this study.

Antibiotic Sensitivity of Group B Streptococcus from Pregnant Mothers and Its Association with Resistance Genes.

OBJECTIVE: This study assessed the antibiotic susceptibility and characterized antibiotic resistance genes of group B Streptococcus (GBS) isolates from selected tertiary care hospitals in Western Province, Sri Lanka. METHODS: A descriptive cross-sectional study was carried out to determine antibiotic sensitivity of GBS among 175 pregnant women of >35 weeks of gestation attending antenatal clinics in four teaching hospitals. Low vaginal and rectal swabs were collected separately, and GBS was identified by standard microbiological methods. Antibiotic sensitivity and minimum inhibitory concentration (MIC) were performed according to Clinical and Laboratory Standards Institute (CLSI) guidelines. DNA was extracted from culture isolates, and antibiotic-resistant genes were identified by polymerase chain reaction using ermB, ermTR, mefA, and linB genes. RESULTS: GBS colonization in the study sample was 25.7% (45/175) with detection rate of 22.9% (40/175) and 2.9% (5/175) in vaginal and rectal samples, respectively. All isolates were susceptible to penicillin with an MIC range of 0.03-0.12 µg/mL. Six isolates (13.3%) were intermediate, and 11 isolates (24.4%) were resistant to erythromycin. There were 5 intermediately resistant isolates (11.1%) and 10 resistant isolates (22.2%) for clindamycin. Of them, seven had inducible clindamycin resistance (iMLSB). MIC range of erythromycin was 0.03-0.32 µg/mL and that of clindamycin was 0.06-0.32 µg/mL. ermB gene was detected in 7 (15.5%). ermTR gene was found in 16 (35.6%) and was significantly associated with iMLSB phenotype (p = 0.005). mefA gene was detected in two (4.4%) isolates, while linB gene was not detected in tested isolates. CONCLUSION: All isolates were sensitive to penicillin, and the most prevalent resistance genotype was ermTR in the study population.

Biofilm formation and antibiotic resistance among Coagulase Negative Staphylococcus species isolated from central venous catheters of intensive care unit patients.

PURPOSE: This study was conducted to determine the biofilm formation of coagulase negative Staphylococcus species (CoNS) isolated from patients with catheter related blood stream infection (CRBSI) and colonized central venous catheters (CVC) and their antibiotic susceptibility patterns and in situ biofilm formation of CVC tips. METHODS: Eighty-two CoNS isolated from intensive care unit (ICU) patients with CRBSI (n â€‹= â€‹8) or colonized CVC (n â€‹= â€‹74) were included. Species identification and antibiotic susceptibility test were done. All isolates were screened for biofilm formation using crystal violet and 3-(4,5-dimethylthiazole-2-yl)-2-5-diphenyl-2H-tetrazolium bromide (MTT) assays and categorized as strong or moderate biofilm formers. CVC tips were subjected to crystal violet stain and scanning electron microscopy (SEM) to detect in-situ biofilm formation. RESULTS: Staphylococcus haemolyticus (n â€‹= â€‹34; 41%) was the commonest to cause both CRBSI and CVC colonization. All 82 CoNS produced biofilms. Among them 77 (93.90%) were strong biofilm formers including all from CRBSI patients and 05 (6.10%) were moderate biofilm formers as detected by both methods. SEM showed bacteria adhered to surfaces of CVC tips with microbial-aggregates embedded in extracellular matrix. Mean crystal violet absorbance of CVC from CRBSI patients (0.6628) was significantly higher than colonized CVC (mean value 0.5592) (p â€‹= â€‹0.030). S. haemolyticus showed higher resistance to cloxacillin compared to other CoNS (p â€‹= â€‹0.039). CONCLUSION: Majority of CoNS isolated were strong biofilm formers. In-situ biofilm formation on CVC tips were significantly evident in CRBSI patients compared to CVC colonized patients. S. haemolyticus is the commonest to cause both CRBSI and CVC colonization and shows significantly higher cloxacillin resistance rate.

Distribution of mecA and qacA/B genes among coagulase negative staphylococci isolated from central venous catheters of intensive care unit patients.

PURPOSE: This study was designed to detect the prevalence of antibiotic and antiseptic resistance genes, mecA and qacA/B in coagulase negative Staphylococcus (CoNS) species isolated from intensive care unit patients with catheter related blood stream infections (CRBSI) or colonized central venous catheters (CVC). METHODS: Consecutive CoNS isolates from ICU patients with CRBSI or colonized central venous catheters were speciated and antibiotic susceptibilities were determined. The mecA and qacA/B genes were detected by polymerase chain reaction. RESULTS: Eighty-two CoNS isolates from ICU patients with CRBSI (n â€‹= â€‹8) or colonized CVC (n â€‹= â€‹74) were included. The mecA gene was detected in 62 CoNS isolates (76%). The commonest species isolated was S. haemolyticus (n â€‹= â€‹34; 41%) and 30 of these possessed mecA which was significantly higher compared to other CoNS species (p â€‹= â€‹0.036). The qacA/B gene was detected in 13 (16%) isolates. Eleven (13%) CoNS had both genes. A significant association was seen with the presence of mecA and resistance to cloxacillin (p â€‹< â€‹0.001) and erythromycin (p â€‹= â€‹0.046). Presence of qacA/B (p â€‹= â€‹0.007) or both mecA and qacA/B (p â€‹= â€‹0.014) was associated with a higher resistance to clindamycin. CONCLUSION: A considerably high prevalence of mecA and qacA/B genes as well as co-existence of both genes is noted among the CoNS isolated from ICU patients. This indicates the need of taking prompt actions in hospital acquired infection prevention including continuous surveillance.

In -vitro Biofilm Formation of Vaginal Isolates of Streptococcus agalactiae; Effect of pH and Culture Media

Abstract Background: Streptococcus agalactiae (GBS), a major cause of neonatal morbidity and mortality, is transmitted from mother to neonate via placenta or during birth. Biofilm formation is an important factor in GBS pathogenesis. This study aimed to determine effects of pH, different culture media and nutritional composition on in vitro biofilm forming ability of GBS isolated from pregnant women. Methods: A total of 30 confirmed isolates of GBS from pregnant women were tested for biofilm formation in Todd Hewitt Broth (THB) at pH 4.5,6 and 7. Ten of these isolates were tested for biofilm formation in growth media THB, brain heart infusion broth, tryptic soy broth, Mueller Hinton broth and nutrient broth. Further they were tested for influence of glucose on biofilm formation using crystal violet and MTT assay. Results: Of 30 GBS isolates strong biofilm formation (SBF) was observed at pH 7 in 56.6 %(n=17) while 36.6%(n=11) isolates showed weak biofilm formation (WBF). At pH 4.5, 43.3% (n=13) were non biofilm formers. In THB without glucose, all 10 isolates were SBF while THB with 1% glucose, 3(30%) isolates were SBF, 5(50%) isolates were moderate biofilm producers and 2(20%) isolates were WBF. Ten isolates tested in 5 types of growth media did not show statistically significant difference in biofilm forming ability. Conclusion: All tested vaginal GBS isolates were able to produce biofilms, maximum biofilm formation of GBS was at pH 7.0. and pH 4.5 is not favorable, thus in normal vaginal pH (3.5 - 4.5), GBS finds it difficult to grow biofilms.