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PROJECT TOPIC- ISOLATION AND CHARACTERIZATION OF GROUP B STREPTOCOCCI (Streptococcus agalactiae) FROM CLINICAL AND NON- CLINICAL SOURCES IN ENUGU STATE

PROJECT TOPIC- ISOLATION AND CHARACTERIZATION OF GROUP B STREPTOCOCCI (Streptococcus agalactiae) FROM CLINICAL AND NON- CLINICAL SOURCES IN ENUGU STATE

 

ABSTRACT

In a comprehensive study to assess the rate of vaginal carriage of Group B Streptococci (GBS) in pregnant and non pregnant women,the impact on neonates and couples that visited infertility clinics in parts of Enugu state, a prevalence study was carried out between June 2007 and September 2008 involving analysis of a total of 590 samples (275 clinicals and 315 non-clinicals). All samples were cultured on a modified Islam medium and identification carried out by standard bacteriological methods. The 275 clinical samples, were cultured from 119 pregnant and 79 non-pregnant women, 52 neonates and 25 infertility couples (7 primary and 18 secondary) respectively seen at the Institute of Child Health, of the University Teaching Hospital Enugu and from Amblim Reference Laboratory where they were referred to from some infertility clinics in the area. Generated data were subjected to statistical analysis. Positive results showed that a total of 14(11.8%) were from pregnant women, while 6 (7.6 %) were from non-pregnant women, of which when tested statistically was found to be significant (P < 0.05). Positivity amongst pregnant women when correlated with parity and gestational age was found to be significant (R2 = 0.000738). Five (9.6%) neonates gave positive growth and 18 (72 %) samples for secondary infertility gave positive results. No GBS was isolated from primary infertility cases. There was statistical significant difference between the couples with secondary infertility and those with primary infertility (P < 0.05). Of the total of 315 non-clinical samples cultured consisting of 275 cattle udder samples from Fulani settlements in parts of Enugu state, 10 meat slabs from abbatoirs, 30 milk products “Fura-da-nunu” from hawkers from parts of Enugu state, 11(4.0%) and 25 (83.3 %) from the cattle udders and milk products were positive respectively for GBS. However, no statistically significant difference was observed between clinical and non-clinical samples in terms of recovery of GBS (P>0.05). Characterizations of the GBS isolates were carried out using sodium hippurate reactions, aesculin reactions, Christie, Atkin Muchin – Patterson (CAMP) test and definite haemolytic hydrolysis on blood agar. Isolates showed typical reactions for GBS. Pathological studies were conducted using three representatives each, selected from pregnant, non-pregnant, neonatal, infertility and non-clinical sample isolates using adult mice. Out of the fifteen adult mice used, 5 were pregnant (14days). Histological evidence of degenerative changes, inflammation and infiltration were recorded in the liver and uterus. All the pregnant mice aborted between 2-5days after inoculation of the organisms. The aborted infants gave positive isolation of GBS from various sites swabbed. This probably suggests some toxic effects by GBS and its probable effect on the uterine muscles and endometrium that may have contributed to the abortion episodes. The exact involvement of GBS in these clinical entities and the extent of the damage need further studies.

 

CHAPTER ONE
1.1 INTRODUCTION

Many years ago most human infections caused by aerobic Streptococci were attributed to Lancefield groups A and D or to the so called non-groupable viridans Streptococci. The first human isolates of the Group B Streptococci (GBS) were isolated from the vagina of post partum women (Lancefield and Hare, 1935). In 1938, Fry found group B Streptococci present in the vaginal cultures of both symptomatic and asymptomatic women at the time of delivery. Some of the symptomatic patients have the organism both in the vaginal swabs and blood cultures.
Despite these early associations with human infections, the group B Streptococci continued for many years to be considered as primarily an agent responsible for mastitis in cattle, goats and sheep with little human relevance and was therefore named Streptococcus agalactiae (Lancefield and Hare, 1935). Clinically, microbiological characteristics of the group B Streptococci are definitively differentiated from other Streptococci by Lancefield typing or by other serologic method such as counter immunoelectrophoresis. These techniques however, are not available in all clinical Laboratories.

Hence, in this study, presumptive identification of group B Streptococci is made by a number of other properties of these organisms as indicated below.
Morphologically, these human Group B Streptococci (GBS) strains usually appear pigmented. They show a narrow but definite zone of weak beta haemolysis unlike Group A Streptococci (Facklam and Smith, 1976), though, about 5 % of the human strains are not/slightly beta haemolytic and these are normally missed (Lancefield, 1935). About one third of GBS strains are Bacitracin sensitive by disk testing and are thus early confused with Group A Streptococci. The fluorescent antibody method available in most Laboratories for identifying group A beta-haemolytic Streptococci will differentiate these strains, (Lancefield, 1933).
Further studies suggest that by the use of haemolytic reactions on blood agar, sensitivity by the same bacitracin disk, bile aesculin reactions and hydrolysis of sodium hippurate that one can reliably detect most group B Streptococci in the clinical Laboratory (Facklam and Smith, 1976). These organisms fail to produce a bile aesculin reaction but nearly always hydrolyze sodium hippurate. CAMP (Christie Atkins Munch- Peterson) test, which is an abbreviation named after the initials of the three original authors (Christie R, Artkins N.E., Munch – Peterson), now served as a confirmatory test of this organism (GBS) (Christie et al., 1944).

PROJECT TOPIC- ISOLATION AND CHARACTERIZATION OF GROUP B STREPTOCOCCI (Streptococcus agalactiae) FROM CLINICAL AND NON- CLINICAL SOURCES IN ENUGU STATE

The CAMP reaction was based on the observation by Christie et al, (1944) that a lytic phenomenom occurs when group B, but not other Streptococci, are grown in a zone of Staphylococcus β toxin activity on a sheep’s blood agar plate.
From the previous studies done in some parts of the world, it had been observed that numerous account in the past decade have established the association of group B Streptococci (GBS) with man. Here the organism is carried asymptomatically in various sites of the body, such as the vagina of pregnant women, urethra of males and the gastrointestinal tract (GIT) (Anthony and Okada, 1977).
Patients in all age groups may be infected with GBS, but the vast majority of infections caused by this pathogen occur among the very young, especially in infants, with a greater incidence among the neonates ≤ 5 days (Lippincott et al., 2001).There had been a microbiologic scientific adage that says ‘when thinking of group B Streptococci, think of group B for baby’, (Lippincott et al., 2001). Two distinct syndromes have been described that are associated with this GBS in infants namely: (1) Early-onset or acute onset; these are normally observed in neonates of ≤ 5 days old. This is often seen in premature neonates/or neonates born in the setting of serious maternal complications like in prolong rupture of membranes (Kosheleva and Zatsiorskaia, 1994).
(2) Late-onset; this shows non-specific findings of neonatal sepsis, bacteraemia and meningitis. These are very common while pulmonary symptoms are infrequent (Hoshina et al., 1994).
In addition to symptomatic bacteraemia, pnenmonia and meningitis, other clinical infections in infants have also been noted. These include otitis media, ethmoiditis, conjunctivitis, omphalitis, cellulitis, impetigo, emphysema, pericarditis, peritonitis, oesteomyelitis, arthritis and endocarditis (Hoshina et al., 1994).
Some studies have shown that about 25 % of women carry these bugs vaginally, and a baby can acquire these bacteria during delivery (Lise and Carl, 2004). In the same study done by Lise and Carl (2004), about 1 % of children born to mothers infected with group B Streptococci (GBS) develop bacteraemia and pneumonia within the first five days of life in the United States. In spite of intensive antibiotic therapy, such infections carry a mortality rate of 50–70 %. The carrier rates of this organism especially during pregnancy vary with localities and socio-economic backgrounds (Hickman et al., 2001).
The carrier rates documented from various parts of the world in birth canals range between 1.8 and 30 % (Nitzan. et al., 1980). Thus Olanishebe and Adetosoye (1986) in Ibadan showed a 1.6 % rate in asymptomatic pregnant women vaginal tract. The carrier rate documented from various parts of the world range between 1.8 % to about 30 % (Noah and Odugbemi, 1978). It is generally believed that more than 10 % of pregnant women are carriers of the organisms and are believed to transmit it to newborns and that up to 45,000 cases per year of post partum endometriosis in women following childbirth occur in different parts of the world (Schrag ,2002).
It is generally believed that more than 10 % of pregnant women are carriers of the organisms and are believed to transmit it to their newborns and that up to 45,000 cases per year of post partum endometriosis in women following childbirth occur in different parts of the world (Schrag et al., 2002). Hence, neonates from these women develop bacteremia in less than five days after birth which may result to neonatal meningitis or pneumonia. Whether this organism can indeed play a role in abortions of pregnancies and infertility has been a subject of intense research. Some studies have shown its significance especially in secondary infertility.
There is however, scarcity and uncoordinated information in this part of the country on group B Streptococci (GBS); hence this work.

1.2 AIMS AND OBJECTIVES

(1) To isolate GBS from gynaecological conditions by high vaginal swabs investigations.
(2) To isolate GBS from neonatal navel notch in infants with signs and symptoms suggestive of bacteremia.
(3) To assess the vaginal carrier status of GBS in pregnancy and women of child bearing age.
(4) To identify the presence of the organism from non-clinical sources like cattle and their milk products, and meat slabs in selected abattoirs in the area.
(5) To carry out antibiogram on the isolates
(6) To characterize the isolates by standard biochemical/ physiological investigations.
(7) To assess the virulence factors of GBS using animal models.

PROJECT TOPIC- ISOLATION AND CHARACTERIZATION OF GROUP B STREPTOCOCCI (Streptococcus agalactiae) FROM CLINICAL AND NON- CLINICAL SOURCES IN ENUGU STATE

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