BACTERIAL ISOLATES FROM CEREBROSPINAL FLUID OF SUSPECTED ACUTE MENINGITIS IN NIGERIAN CHILDREN.    [Original Research]
Nwadioha SI, FMCPath1,  Onwuezube I, FMCPath2,  Egesie J O, FMCPath3,  Kashibu E, FMLS4 ,  Nwokedi EOP, FMCPath1
1. Department of Medical Microbiology, College of Health Sciences, Benue State University, Makurdi, Nigeria
2. Department of Medical Microbiology, University of Uyo Teaching Hospital, Uyo Akwa, Ibom state, Nigeria
3. Department of Haematology, Jos University Teaching Hospital, Jos, Nigeria
4. Department of Microbiology, Aminu Kano Teaching Hospital, Kano, Nigeria.

[emedpub – International Infectious Diseases :   Vol 1:8]                  [Date of Publication: 06.03.2011]
ISSN 2231-6019

June 3, 2011 at 11:15 AM

Abstract

Objectives: To determine etiology of acute bacterial meningitis in Nigerian children and their antibiotic susceptibility pattern.

Methods: A retrospective study was conducted with a review of cerebrospinal fluid culture reports of suspected acute meningitis in pediatric patients aged 0-15 years between October 2006 and October 2009 in Aminu Kano Teaching Hospital Kano, Nigeria.

Results: A positive culture bacterial isolation rate of 3.3% (n=50/1500) with prevalence of Streptococcus pneumoniae (24%), Neisseria meningitidis(22%), Escherichia coli(16%), Haemophilus  influenzae (14%), Group B streptococci(8%) and Enterococci(8%) which were susceptible to ceftriaxone(96%), cefotaxime(95%) and ciprofloxacin(93%) across  the bacterial isolates. Neonates were 55% (n=6.8/12.4) most at risk.

Conclusion: Neonates are most at risk of acute bacterial meningitis. In the absence of antibiotic susceptibility report, ceftriaxone should be considered as a first choice antibiotic for empirical treatment of acute meningitis among children in Nigeria.

Introduction.

Meningitis is a medical emergency that requires urgent rational antibiotic therapy, especially in neonates and young infants1. It accounted for 2.7% of infant deaths among hospitalized children in Ilorin and 3.4% of post-neonatal deaths in Sokoto in Nigeria 2. Bacterial meningitis has a global incidence of about 20 -100 cases per 100,000 live births during the newborn period3.   Signs and symptoms of meningitis in infants may include fever or hypothermia (temperature may even be normal), poor feeding, irritability or lethargy, seizure, rash, tachypnea or apnea, jaundice, bulging fontanelle (in late stage), vomiting or diarrhea, or altered sleep pattern 2. When a toxic newborn or young infant presents with fever and lethargy or irritability, it is important to consider the diagnosis of meningitis even if the classic localizing signs and symptoms are absent 1.

The gold standard for diagnosis of meningitis is the laboratory testing. However, medical emergency demands an urgent rational antibiotic therapy to be instituted until culture results are ready. The common bacterial etiology reported for acute bacterial meningitis in neonates includes Group B streptococcus, Escherichia  coli, Klebsiella spp., Enterococcus spp., Listeria monocytogenes, Streptococcus pneumoniae; in the age group of one to six months, S. pneumoniae, Haemophilus influenzae, Neisseria meningitidis, E. coli, L. monocytogenes; in age group above six months, the prevalent bacterial agents include Neisseria meningitidis and S. pneumoniae4. In determining likely etiology of meningitis in children, it is therefore important for clinicians to consider the age, the underlying host factors, and the season of the year.

In such a life threatening emergency as meningitis, knowledge of epidemiology and antimicrobial susceptibility pattern of common pathogens in a geographic area helps in the choice of antibiotics. A regular, periodic surveillance of bacterial profile and their antibiotic susceptibility pattern is imperative for appropriate case management, infection control, and rational use of antibiotics.

Kano state of Nigeria in sub- saharan Africa is located in the meningitis belt where  seasonal  meningococcal epidemics occur  during the dry season i.e. from December to June each year. Therefore, this study was conducted to review the etiology and the antimicrobial susceptibility of the bacterial pathogens in childhood acute meningitis in  Aminu Kano Teaching Hospital, a tertiary healthcare facility located in the meningitis belt.

Patients and Methods.

A retrospective study was conducted for pediatric patients in the age group of <15 years, who presented with suspected acute bacterial meningitis between 2006 and 2009 in Aminu Kano Teaching Hospital, Nigeria. Formal approval for the study was obtained from the hospital ethics committee. Cerebrospinal fluid (CSF) samples from Special Care Baby Unit (SCBU), Emergency Pediatric Unit (EPU) and Pediatric wards were collected by the attending physicians with meticulous aseptic technique. CSF samples of varying quantity of 3 to 5 ml (3ml for neonates) were dispatched to the laboratory as quickly as possible, since undue delay might result in poor recovery of some pathogens, such as meningococci, and also cause the disintegration of leukocytes. If delay of few hours was unavoidable, the specimens were best kept in an incubator at 37° C.  CSF samples were categorized into different age groups: A, B and C ( 0-28 days , >28 days to <6 years,  and 6 years to <15 years, respectively).

MACROSCOPY: The specimens were examined for presence of turbidity and any sign of contamination with blood. Normal CSF was clear and colorless like water. A yellow color CSF might result from a previous cerebral hemorrhage.

MICROSCOPY:

Cell Count. The leukocytes in the CSF were counted by microscopic observation of well mixed fluid in a slide counting chamber. Cell count of normal CSF was taken as 0-5 leukocytes /mm3, mainly lymphocytes, although in neonates leukocytes up to 30/mm3 mainly lymphocytes were taken as normal 4. With slight contamination with blood, excess of 1 leukocyte per 1000 erythrocytes was suggestive of meningitis 4.

Gram stain. After taking CSF for the cell count, the remainder was spun to sediment any cells and bacteria. The film of sediment was stained by Gram’s stain.

Culture. CSF was seeded heavily on to culture media (blood agar, chocolate agar and enrichment broth thioglycolate) for incubation for 48 hours in humid air plus 5-10% CO2. With suspicion of brain abscess, a further blood agar plate was seeded for incubation for 2-5 days in anaerobic environment according to standard procedures 4.  Organisms isolated were identified by conventional methods 4.  Antibiotic susceptibility tests were done against locally available antibiotics by using disk diffusion method in accordance with CLSI, 2006 criteria, and interpreted accordingly 5. Penicillin (10 IU), cloxacillin (5µg), ampicillin (10 µg), chloramphenicol (10 µg), gentamicin (10µg), ciprofloxacin (5µg), ceftriaxone (30µg), ceftazidime (30 µg), cefotaxime (30 µg), clavulinate+amoxicillin (20 µg) were used.   Controlled strains namely, Staphylococcus aureus ATTC 25923, Escherichia coli ATTC 25922 and Pseudomonas aeruginosa ATTC 27853 were used.

Blood samples from peripheral veins were collected for blood culture at the same time as CSF tap. In neonates, swabs of the throat, nose, ear and umbilicus were also sent for cultures.

CHEMISTRY: The supernatant from centrifuged CSF was tested for its content of glucose and protein. Normal CSF contains 2.2- 4 mmol glucose/ liter ( about 60% of the plasma glucose value) and 0.15- 0.4 g protein /liter (in neonates up to 1.5 g protein /liter). In purulent bacterial meningitis, the glucose concentration was reduced (0-2 mmol) and protein concentration increased (0.5-3g/liter). The results were analysed using statistical package for the social sciences(SPSS) 11.0.

Results.

A total of 1500 CSF samples were collected, and 50 (3.3%) were detected as culture positive. The common bacterial isolates were Streptococcus pneumoniae (24%), Neisseria meningitidis (22%), Escherichia coli (16%), Haemophilus influenzae (14%),  Streptococcus agalactiae (8%) and Enterococci species (8%) (Table 1).

Bacterial isolates according to age groups; A (neonates), B (>28days to <6 years) and C (6years to < 15 years) were 6.8%, 4.0% and 1.6%, respectively. In neonates, the predominant bacteria were; E. coli, Streptococcus agalactiae and Enterococci species at 41%, 24% and 18%, respectively. Haemophilus influenzae, Neisseria meningitidis and Streptococcus pneumoniae predominated in group B at 32%, 27% and 27%, respectively. In group C, there was preponderance of Neisseria meningitidis and S. pneumoniae at 54% and 46%, respectively(Table 1).

Table 1. Cerebrospinal fluid bacterial isolates in three age groups in Aminu Kano Teaching Hospital, Oct.2006 –Oct.2009.

Bacteria                                                                    Age group

GroupA  (neonates) (%) GroupB (>1mo<6mo)(%) GroupC

(6mo<15yrs)(%)

Total

(%)

Escherichia coli 7 1 - 8(16.0)
Staphylococcus epidermidis 1 - - 1(2.0)
Klebsiella spp 1 - - 1(2.0)
Enterococcus spp 3 1 - 4(8.0)
Salmonella spp 1 1 - 2(4.0)
Streptococcus agalactiae 4 - - 4(8.0)
Neisseria meningitidis - 6 5 11(22.0)
Streptococcus pneumoniae - 6 6 12(24.0)
Haemophilus influenzae - 7 - 7(14.0)
Total Isolates 17 22 11 50(3.3)
Sterile(negative) cultures 233 528 689 1450(96.7)
Total cultures 250(16.6) 550(36.7) 700(46.7) 1500(100)

H. influenzae was susceptible to ceftriaxone (91%), cefotaxime (91%), ciprofloxacin (86%) and chloramphenicol (71%). Streptococcus pneumoniae was susceptible to ciprofloxacin (87%), ceftriaxone (87%), clavulinate-amoxycillin (78%) and penicillin (70%). Neisseria meningitidis was susceptible to ciprofloxacin (100%), ceftriaxone (100%), clavulinate-amoxycillin (82%) and penicillin (72%). E. coli was susceptible to ceftazidime (94%), cefotaxime (94%), ciprofloxacin (88%), and clavulinate-amoxycillin (81 %). Enterococci species were susceptible to ciprofloxacin (92%), clavulinate-amoxycillin (85%), ampicillin (80%) and gentamicin (77%) (Table 2).

Table 2. Bacterial isolates and antibiotic sensitivity pattern among study

population in Aminu Kano Teaching Hospital 2006-2009

DRUGS BACTERIAL                              ISOLATES
E.coli

16

(%)

S.epidermidis

3

(%)

Klebs-

iella

spp.

7

(%)

Strep. agalactiae

8

(%)

N.meningitidis

11

(%)

Salm-

onella

spp.

4

(%)

S.

pneum-

oniae

23

(%)

Entero

coccus

spp.

13

(%)

H.influenzae

35

(%)

PEN - 0(0) - 6(75) 8((72) - 18(70) 5(38) 10(29)
AMP 7(43) - 4(57) - 5(45) 2(50) 9(39) 11(80) 15(43)
CHLO 4(25) - 2(29) 2(25) 6(55) 2(50) 8(35) 4(31) 25(71)
CLOX - 2(67) - - - - - - -
GENT 9(56) 1(33) 4(57) 5(63) 5(45) 2(50) 11(48) 10(77) 12(34)
CIPRO 14(88) 3(100) 6(86) 8((100) 11(100) 4(100) 20(87) 12(92) 30(86)
CEFTR - 2(67) - 8(100) 11(100) 4(100) 20(87 - 32(91)
CEFTA 15(94) - 6(86) - - 4(100)- - - -
CEFO 15(94) - 6(86) - - - - - 32(91)
CLAV+AMOX 13(81) 2(67) 5(71) 5(63) 9(82) 4(100) 18(78) 11(85) -

PEN – penicillin, AMP – ampicillin, CHLO – chloramphenicol, CLOX – cloxacillin, GENT – gentamicin, CIPRO – ciprofloxacin, CEFTR – ceftriaxone, CEFTA -  ceftazidime, CEFO – cefuroxime, CLAV +AMOX – clavunated amoxicillin

-  – NOT DONE

Discussion.

The positive culture rate of bacterial isolates at 3.3% among children with suspected meningitis in this study was comparable to 3.5% reported from Maiduguri, north eastern Nigeria 1. However, it was much higher than 1.6% reported in a five-year study for children aged between 2 months and 14 years in Ilesa , southwestern Nigeria 1 and   0.69 % prevalence rate of pyogenic meningitis  in a general population of pediatric patients recorded in Nnewi, southeastern Nigeria 6. In a rural district hospital in Kenya, bacterial meningitis was reported among 1.3% of children on admission; 88% of whom were under 5 years of age7.   A low detection rate might result from intake of antibiotics before presentation for treatment at the hospital 8. A low bacterial isolation rate might as well result from depletion of delicate organisms like N. meningitidis, in the course of delay in transportation and testing of CSF sample.

In the study, bacterial isolation rate of 6.8% in neonates was higher than in post neonates and young infants (4%) , and older children (1.6%). Neonates had a predominance of Escherichia coli (41%), Group B streptococci (24%) and Enterococci (18%). These results are in agreement with other studies in Nigeria and Kenya, wherein newborns had the highest prevalence of bacterial meningitis compared to the other pediatric age groups 9,7,1. The prevalence of these pathogens in neonates is probably due to the immature immune system and permeability of the blood brain barriers. The predominance of E. coli, Group B-streptococcus and Enterococci species in neonates in the study might suggest that these infections were acquired during passage through colonized mothers’ vaginal vault 9.

Four percent prevalence rate of bacterial isolates was recorded in post- neonates and young infants in the study and had the predominance of Haemophilus influenzae (32%), Neisseria meningitidis (27%) and Streptococcus pneumoniae (27%). Some other previous studies also recorded predominance of H. influenzae, N. meningitidis , S. pneumoniae in this age group 10,1,8. The lack of demonstrable humoral antibody against H. influenzae capsulated Pittman b type in children is associated with increased incidence of meningitis 11,9 .

Older children recorded a 1.6% prevalence of bacterial isolates in the study and had a high predominance of Neisseria meningitidis (54%) and S. pneumoniae (46%), respectively. The prevalence of N. meningitidis in this age group was in agreement with other studies in Ife, southwestern Nigeria, Ilorin, north central Nigeria and Gambia, west Africa 12,11,2. Again, the importance of antibody is also a factor in this age group, since overcrowding in institutions is more likely to lead to disease. N. meningitidis has been associated with epidemic meningitis among children and young adults in crowded condition. Isolation of some other pathogens may suggest the presence of certain underlying conditions, such as otitis media, pneumonia, diabetes mellitus, and immunedeficiency.

The high incidence of sterile CSF (96.7%) may be due to pre-presentation antibiotic use as access to drugs, prescribed or otherwise is unrestricted in Nigeria13. This ‘culture-negative CSF’ debacle can be readily overcome if newer methods of diagnoses  which do not require the growth of live pathogens such as latex agglutination and polymerase chain reaction are incorporated into medical laboratory practice in the region. This problem of antibiotic misuse may lend credence to the increasing resistance to the common anti-meningitic drugs i.e., ampicillin, penicillin and chloramphenicol, in Nigeria13,1.  Ceftriaxone, cefotaxime, ciprofloxacin had the most effective anti-bacterial activity across all the bacterial isolates tested in the study. Ciprofloxacin is not routinely recommended for pediatric use except in special cases where the benefits outweigh the short term risk of joint toxicity such as in a condition like cystic fibrosis 14.  Emergence of resistant bacterial strains to conventional antibiotics such as chloramphenicol and penicillin have also been reported in other studies in Nigeria, Mumbai, and Trinidad 15,16,17,18 and have raised concern for spread of resistance  with resultant consequences i.e. in developing countries where the first line ceftriaxone is not readily available or affordable.

A rational use of antibiotics especially in this pediatric group can not be over emphasized. The government programmes should comprehensively address the issue of proper distribution, dissemination and administration of conjugate vaccines and incorporation of the same in routine immunization schedule of children in our communities as practiced elsewhere in the world. Preventive antimicrobial treatment of pregnant women and a regular high vaginal swab for microbiologic study is advocated.

A limitation in the study includes loss of some fastidious bacteria leading to high culture negative CSF because of frequent inappropriate use of antibiotics by patients before presentation, delay in processing of CSF specimens and paucity of newer method of diagnoses which do not require the growth of live pathogens such as latex agglutination and polymerase chain reaction in the medical laboratory practice in the region.

Finally, in the absence of antibiotic susceptibility report, ceftriaxone should be considered as a first choice of reliable antibiotics for empirical treatment of meningitis in Kano.

References:

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2.      Sanya EO, Taiwo SS, Azeez O, Oluyaombo R. Bacterial Meningitis: Problems of empirical treatment in a Teaching Hospital in the tropics. Internet J  infect Dis 2007; 6(1): 1-4.

3.      World Health Organization. Detecting meningococcal meningitis epidemics in highly endemic African countries. Wkly Epidemiol Recd 2003; 78(33):294-296.

4. Forbes BA, Sahm DF, Weisfeld AS. Streptococcus Meningitis and other infections of the central nervous system. In: laboratory manual of Bailey & Scotts diagnostic microbiology. 12th Ed. Mosby Elsevier publication 2007, pp 907-916.

5. Clinical Laboratory Standard Institute. Performance standard for antimicrobial disk susceptibility tests; Approved standard -9th Ed. Supplement M2 – A9 2006, 26(1).

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11. Weber MW, Herman J, Jaffer S, Usen S, Ooaraugo A, Omsosighoc et al. Clinical predictors of bacterial meningitis in infants and young children in Gambia. Trop Med Int Hlth 2002; 7:722-731.

12. Onipede AO, Onayade AA, Elusanya JBE, Obiajunwa PO, Ogundare EOO, Olaniran OO et al. Invasive bacterial isolates from children with severe infections in a Nigerian hospital J Infect Dev Ctries 2009; 3(6):429-436.

13. Akuhwa RT, Alhaji MA, Bello MA, Okon KO. Susceptibility pattern of meningococcal meningitis outbreak in Nguru , Yobe state Nigeria. Internet j Trop med 2010 ; 7(1):1-4.

14. Nwadioha SI, Nwokedi EOP, Kashibu  E, Odimayo MS, Okwori EE. Bacterial isolates in blood cultures of children with suspected septicaemia in a Nigerian tertiary hospital. Internet  J Infect Dis 2010; 8(1) : 1-6. At http://www.ispub.com/journal/ the internet journal of infectious diseases.

15. WHO Standardized treatment of bacterial meningitis in Africa in epidemic and non epidemic situations. WHO/CDS/EPR/2007.3

16. Sonavane AE, Baradkar VP, Mathur M. Pattern and antibiotic susceptibility of bacteria isolated in clinically suspected cases of meningitis in children. J Paed Neurosci 2008;3:131- 133.

17. Orrett FA. Colonization with Group B streptococcus in pregnancy and outcome of infected neonates in Trinidad. Paed Int’l 2003; 45:319-323.

18. Matthew S, Guy ET. The diagnosis and management of acute bacterial meningitis in resource-poor settings. Lancet Neurol 2008;7(7):637-648.

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