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CLASSICAL AND NEW EMERGING ENTERIC
PATHOGENS ASSOCIATED WITH
CHILDHOOD DIARRHEA IN
TRIPOLI-LIBYA
Prof. Khalifa Sifaw Ghenghesh
Faculty of Medicine, Tripoli University
Libya
Introduction
• In developing countries, infectious diarrhea is associated
with high rates of morbidity and mortality, mainly in
childhood.
• Classical enteric pathogens
– Salmonella sp., Shigella sp., enteropathogenic Escherichia coli
(EPEC), enteroinvasive E. coli (EIEC),Vibrio cholerae and
rotavirus.

• New emerging enteric pathogens
– bacteria (e.g., Campylobacter spp., enterohemorrhagic
Escherichia coli [EHEC], and enteroaggregative E. coli [EAEC]),
– viruses (e.g., norovirus, adenovirus, and astrovirus), and
– parasites (e.g., Cryptosporidium sp.).

• In Libya, rotavirus and Salmonella have been documented
as major causative agents of childhood diarrhea.
Agents of Infective Acute Diarrhea in Libyan Children
1980-2007
Agent

Rate of Occurrence

Bacterial
1. Enteropathogenic E. coli
3. Salmonella sp.
4. Shigella sp.
5. Yersinia enterocolitica
6. Campylobacter sp.
7. Aeromonas sp.

4 - 11 %
6 -25 %
4 - 10 %
<1 %
2-6%
0 - 15 %

Parasitic
1. Entamoeba histolyitca/disapr
2. Giardia lamblia
3. Cryptosporidium

12 %
1 - 18 %
13 %

Rotavirus

24 - 31 %

Multiple agents

12 - 14.5 %
• Major drawbacks of previous data from Libya:
– Available data based on few studies and some of
them dealt with single or few pathogens.
– No available data for some of the classical and new
emerging enteric pathogens that include:
•
•
•
•
•
•
•

Enterotoxigenic E. coli (ETEC)
Enteroinvasive E. coli (EIEC)
Enterohemorrhagic E. coli (EHEC)
Entroaggregative E. coli (EAEC)
Norovirus
Adenovirus
Astrovirus

– Available data on enteric protozoa obtained using wet
mount techniques.
• Not sensitive
• Do not differentiate between E. histolytica and E. dispar.
Patients and Methods
• Stool samples:
– 239 children (102 females) with diarrhea,
– a few days to 5 years,
– Outpatient Clinics of Aljala Children’s Hospital and Alkhadra
Hospital
– February-October 2008.

• Clinical history by MDs after informed consent from a parent
or guardian:
– Clinical symptoms include fever, vomiting, and dehydration were
recorded.
– Also recorded type of feeding (breast, artificial, or both).
– Other.

• Microbiology:
– Stools examined for bacterial, viral and parasitic agents enteric
pathogens using a combination of culture, enzyme-immunoassay,
and polymerase chain reaction (PCR) methods.
Results
Enteric pathogens isolated from 239 diarrheic
children in Tripoli, Libya.
Enteric pathogen (s)

No. (%)
positive

Single

89 (37.2)

Multiple

33 (13.8)

Total

122 (51.0)

Diarrheagenic Escherichia coli

27 (11.2)

Salmonella spp.

19 (7.9)

Shigella spp.

1 (0.8)

Campylobacter spp.

7 (2.9)

Aeromonas spp.

10 (4.2)

Rotavirus

32 (13.4)

Norovirus

37 (15.5)

Adenovirus

17 (7.1)

Astrovirus

4 (1.7)

Cryptosporidium spp.

5 (2.1)

Entamoeba histolytica

2 (0.8)

Giardia lamblia

3 (1.3)

No pathogen detected

117 (49)
Enteric Bacterial Pathogens
Diarrheagenic Escherichia coli1

27 (11.2)

-

pCVD432 (EAEC)

10 (4.1)

-

eaeA (EPEC/EHEC)

9 (3.8)

-

ipaH (EIEC)

4 (1.6)

-

pCDV432 + ipaH

2 (0.8)

-

eaeA + ipaH

1 (0.4)

-

pCVD432 + eaeA

1 (0.4)

Salmonella spp.

19 (7.9)

-

Salmonella group B

3 (1.3)

-

Salmonella group C1

1 (0.4)

-

Salmonella group C2

13 (5.4)

-

Salmonella group D1

2 (0.8)

Shigella spp.

1 (0.8)

Campylobacter spp.
- C. jejuni
- C. coli

7 (2.9)
6 (2.5)
1 (0.4)

Aeromonas spp.

10 (4.2)

1est

and

encoding

eltB

genes

for
enterotoxigenic E. coli (ETEC)
were not detected.
Clinical symptoms associated with enteric pathogens isolated from
diarrheic children in Tripoli, Libya
Agents

Cases
detected

No. (%) positive
Fever
Dehydration1
62(69.9)
30(33.7)
24(72.7)
13(39.4)
86(70.4)
43(35.2)

Single
Multiple
Total

89
33
122

Vomiting
73(82.8)
30(90.9)
103(84.4)

Diarrheagenic Escherichia coli

27

25(92.6)

20(74.1)

10(37.0)

Salmonella spp.

19

15(78.9)

15(78.9)

4(21.1)

Shigella spp.
Campylobacter spp.

1
7

1(100)
3(42.9)

1(100)
4(57.1)

0(0.0)
0(0.0)

Aeromonas spp.

10

10(100)

6(60)

5(50)

Rotavirus2

32

32(100)

24(75)

17(53.1)

Norovirus

37

34(91.9)

23(62.2)

9(24.3)

Adenovirus

17

14(82.4)

10(58.8)

7(41.2)

Astrovirus

4

3(75)

4(100)

2(50)

Cryptosporidium spp.

5

2(40)

4(80)

2(40)

Entamoeba histolytica

2

2(100)

2(100)

2(100)

Giardia lamblia

3

2(66.7)

3(100)

1(33.3)

117

95(81.2)

87(74.4)

2(1.7)

No pathogen detected

1Dehydration

associated
with
diarrheic children
positive for single,
multiple and total
enteric
pathogens
compared
with
children with no
pathogen detected.
2Rotavirus

associated with dehydration
compared
with norovirus.
Distribution of enteric pathogens isolated from diarrheic children in Tripoli
according to seasons of the year covered in the study.
Spring
(n=47)

Summer
(n=112)

Autumn
(n=66)

Total
(n=239)

Single1

22(46.8)

36(32.1)

31(48.5)

89(37.2)

Multiple
Total2

7(14.9)
29(61.7)

16(14.3)
52(46.4)

10(15.2)
41(62.1)

33(13.8)
122(51)

Diarrheagenic E. coli
Salmonella spp.
Shigella spp.
Campylobacter spp.

7(14.9)
2(4.3)
0(0.0)
0(0.0)

9(8)
11(9.8)
1(0.9)
6(5.4)

11(16.7)
6(9.1)
0(0.0)
1(1.5)

27(11.3)
19(7.9)
1(0.4)
7(2.9)

Aeromonas spp.3
Rotavirus4

5(10.6)
16(34)

5(4.5)
3(2.7)

0(0.0)
13(19.9)

10(4.2)
32(13.4)

Norovirus5
Adenovirus

3(6.4)
3(6.4)

19(17)
9(8)

15(22.7)
5(7.6)

37(15.5)
17(7.1)

Astrovirus
Cryptosporidium spp.
Entamoeba histolytica
Giardia lamblia
No pathogen detected

0(0.0)
1(2.1)
1(2.1)
0(0.0)
16(34)

3(2.7)
4(3.6)
0(0.0)
2(1.9)
59(52.7)

1(1.5)
0(0.0)
0(0.0)
1(1.5)
38(54.5)

4(1.7)
5(2.1)
2(0.8)
3(1.3)
117(49)

Enteric pathogen (s)

1,2,4Single

and total
pathogens and
rotavirus detected
more in spring and
autumn compared with
summer. 3Aeromonas
detected more in spring
compared with
autumn. 5Norovirus
detected more in
autumn compared with
spring.
Resistance of Salmonella, Shigella and Aeromonas species isolated from Libyan
children with diarrhea to antimicrobial agents

Antimicrobial agent
Ampicillin (10µg)
Amoxicillin/clavulanic
acid (30µg)
Imipenem (10µg)
Chloramphenicol (30µg)
Cephalothin (30µg)
Ceftriaxone (30µg)
Ceftazidime (10ug)
Cefepime (30µg)
Aztreonam (30µg)
Ciprofloxacin (5µg)
Nalidixic acid (30µg)
Streptomycin (10µg)
Tetracycline (30µg)
Trimethoprim/
sulfamethoxazole (25µg)

Salmonella Shigella Aeromonas
(n=19)
(n=1)
(n=10)

Total
(n=30)

9 (47.3 )

1 (100)

10 (100)

20 (66.7)

1 ( 5.2)
0 (0.0)
1 ( 5.2)
11 (57.8)
0 (0.0)
0 (0.0)
0 (0.0)
0 (0.0)
12 (63.1)
16 (84.2)
15(78.9)
12 (63.1)

0 (0.0)
0 (0.0)
1 (100)
1 (100)
0 (0.0)
0 (0.0)
0 (0.0)
0 (0.0)
0 (0.0)
0 (0.0)
1 (100)
0 (0.0)

5 (50)
1 (10)
0 (0.0)
7 (70)
0 (0.0)
0 (0.0)
0 (0.0)
0 (0.0)
0 (0.0)
2 (20)
3 (30)
0 (0.0)

6 (20)
1 (3.3)
2 (6.6)
19 (63.3)
0 (0.0)
0 (0.0)
0 (0.0)
0 (0.0)
12 (40)
18 (60)
19 (63.3)
12 (40)

4 (21)

0 (0.0)

1(10)

5 (16.6)
Conclusion
• To our knowledge this is the first study to identify
norovirus, adenovirus, and astrovirus as enteric
pathogens causing children diarrhea in Libya.
• The findings of the present investigation indicate,
in addition to Salmonella and rotavirus, the enteric
pathogens norovirus, adenovirus and EAEC are
important causative agents of children diarrhea in
Tripoli.

• On
the
other
hand,
Cryptosporidium,
Campylobacter and Aeromonas spp. appear to play
a minor role in pediatric diarrheal disease in Tripoli.
• The
emergence
of
ciprofloxacin-resistant
Salmonella is a very serious health problem that
should be addressed by public health authorities.
• More studies are necessary in other major cities of
Libya as well as a dedicated national pathogenspecific surveillance system to identify various
etiologies of pediatric diarrhea to determine the
exact role of these enteric pathogens.
• Furthermore, introduction of a rotavirus vaccine
into the vaccination program in Libya to protect
the pediatric population is urgently needed.
• Rahouma A., Klena JD., Krema Z., Abobker AA.,
Treesh K., Franka E., Abusnena O., Shaheen HI.,
El Mohammady H., Abudher A., and Ghenghesh
KS. 2011. Enteric Pathogens Associated with
Childhood Diarrhea in Tripoli-Libya. American
Journal of Tropical Medicine and Hygiene; 84:
886-891.
• ghenghesh_micro@yahoo.com
• ghenghesh@yahoo.com

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Enteric pathogens-Tripoli-Libya

  • 1. CLASSICAL AND NEW EMERGING ENTERIC PATHOGENS ASSOCIATED WITH CHILDHOOD DIARRHEA IN TRIPOLI-LIBYA Prof. Khalifa Sifaw Ghenghesh Faculty of Medicine, Tripoli University Libya
  • 2. Introduction • In developing countries, infectious diarrhea is associated with high rates of morbidity and mortality, mainly in childhood. • Classical enteric pathogens – Salmonella sp., Shigella sp., enteropathogenic Escherichia coli (EPEC), enteroinvasive E. coli (EIEC),Vibrio cholerae and rotavirus. • New emerging enteric pathogens – bacteria (e.g., Campylobacter spp., enterohemorrhagic Escherichia coli [EHEC], and enteroaggregative E. coli [EAEC]), – viruses (e.g., norovirus, adenovirus, and astrovirus), and – parasites (e.g., Cryptosporidium sp.). • In Libya, rotavirus and Salmonella have been documented as major causative agents of childhood diarrhea.
  • 3. Agents of Infective Acute Diarrhea in Libyan Children 1980-2007 Agent Rate of Occurrence Bacterial 1. Enteropathogenic E. coli 3. Salmonella sp. 4. Shigella sp. 5. Yersinia enterocolitica 6. Campylobacter sp. 7. Aeromonas sp. 4 - 11 % 6 -25 % 4 - 10 % <1 % 2-6% 0 - 15 % Parasitic 1. Entamoeba histolyitca/disapr 2. Giardia lamblia 3. Cryptosporidium 12 % 1 - 18 % 13 % Rotavirus 24 - 31 % Multiple agents 12 - 14.5 %
  • 4. • Major drawbacks of previous data from Libya: – Available data based on few studies and some of them dealt with single or few pathogens. – No available data for some of the classical and new emerging enteric pathogens that include: • • • • • • • Enterotoxigenic E. coli (ETEC) Enteroinvasive E. coli (EIEC) Enterohemorrhagic E. coli (EHEC) Entroaggregative E. coli (EAEC) Norovirus Adenovirus Astrovirus – Available data on enteric protozoa obtained using wet mount techniques. • Not sensitive • Do not differentiate between E. histolytica and E. dispar.
  • 5. Patients and Methods • Stool samples: – 239 children (102 females) with diarrhea, – a few days to 5 years, – Outpatient Clinics of Aljala Children’s Hospital and Alkhadra Hospital – February-October 2008. • Clinical history by MDs after informed consent from a parent or guardian: – Clinical symptoms include fever, vomiting, and dehydration were recorded. – Also recorded type of feeding (breast, artificial, or both). – Other. • Microbiology: – Stools examined for bacterial, viral and parasitic agents enteric pathogens using a combination of culture, enzyme-immunoassay, and polymerase chain reaction (PCR) methods.
  • 7. Enteric pathogens isolated from 239 diarrheic children in Tripoli, Libya. Enteric pathogen (s) No. (%) positive Single 89 (37.2) Multiple 33 (13.8) Total 122 (51.0) Diarrheagenic Escherichia coli 27 (11.2) Salmonella spp. 19 (7.9) Shigella spp. 1 (0.8) Campylobacter spp. 7 (2.9) Aeromonas spp. 10 (4.2) Rotavirus 32 (13.4) Norovirus 37 (15.5) Adenovirus 17 (7.1) Astrovirus 4 (1.7) Cryptosporidium spp. 5 (2.1) Entamoeba histolytica 2 (0.8) Giardia lamblia 3 (1.3) No pathogen detected 117 (49)
  • 8. Enteric Bacterial Pathogens Diarrheagenic Escherichia coli1 27 (11.2) - pCVD432 (EAEC) 10 (4.1) - eaeA (EPEC/EHEC) 9 (3.8) - ipaH (EIEC) 4 (1.6) - pCDV432 + ipaH 2 (0.8) - eaeA + ipaH 1 (0.4) - pCVD432 + eaeA 1 (0.4) Salmonella spp. 19 (7.9) - Salmonella group B 3 (1.3) - Salmonella group C1 1 (0.4) - Salmonella group C2 13 (5.4) - Salmonella group D1 2 (0.8) Shigella spp. 1 (0.8) Campylobacter spp. - C. jejuni - C. coli 7 (2.9) 6 (2.5) 1 (0.4) Aeromonas spp. 10 (4.2) 1est and encoding eltB genes for enterotoxigenic E. coli (ETEC) were not detected.
  • 9. Clinical symptoms associated with enteric pathogens isolated from diarrheic children in Tripoli, Libya Agents Cases detected No. (%) positive Fever Dehydration1 62(69.9) 30(33.7) 24(72.7) 13(39.4) 86(70.4) 43(35.2) Single Multiple Total 89 33 122 Vomiting 73(82.8) 30(90.9) 103(84.4) Diarrheagenic Escherichia coli 27 25(92.6) 20(74.1) 10(37.0) Salmonella spp. 19 15(78.9) 15(78.9) 4(21.1) Shigella spp. Campylobacter spp. 1 7 1(100) 3(42.9) 1(100) 4(57.1) 0(0.0) 0(0.0) Aeromonas spp. 10 10(100) 6(60) 5(50) Rotavirus2 32 32(100) 24(75) 17(53.1) Norovirus 37 34(91.9) 23(62.2) 9(24.3) Adenovirus 17 14(82.4) 10(58.8) 7(41.2) Astrovirus 4 3(75) 4(100) 2(50) Cryptosporidium spp. 5 2(40) 4(80) 2(40) Entamoeba histolytica 2 2(100) 2(100) 2(100) Giardia lamblia 3 2(66.7) 3(100) 1(33.3) 117 95(81.2) 87(74.4) 2(1.7) No pathogen detected 1Dehydration associated with diarrheic children positive for single, multiple and total enteric pathogens compared with children with no pathogen detected. 2Rotavirus associated with dehydration compared with norovirus.
  • 10. Distribution of enteric pathogens isolated from diarrheic children in Tripoli according to seasons of the year covered in the study. Spring (n=47) Summer (n=112) Autumn (n=66) Total (n=239) Single1 22(46.8) 36(32.1) 31(48.5) 89(37.2) Multiple Total2 7(14.9) 29(61.7) 16(14.3) 52(46.4) 10(15.2) 41(62.1) 33(13.8) 122(51) Diarrheagenic E. coli Salmonella spp. Shigella spp. Campylobacter spp. 7(14.9) 2(4.3) 0(0.0) 0(0.0) 9(8) 11(9.8) 1(0.9) 6(5.4) 11(16.7) 6(9.1) 0(0.0) 1(1.5) 27(11.3) 19(7.9) 1(0.4) 7(2.9) Aeromonas spp.3 Rotavirus4 5(10.6) 16(34) 5(4.5) 3(2.7) 0(0.0) 13(19.9) 10(4.2) 32(13.4) Norovirus5 Adenovirus 3(6.4) 3(6.4) 19(17) 9(8) 15(22.7) 5(7.6) 37(15.5) 17(7.1) Astrovirus Cryptosporidium spp. Entamoeba histolytica Giardia lamblia No pathogen detected 0(0.0) 1(2.1) 1(2.1) 0(0.0) 16(34) 3(2.7) 4(3.6) 0(0.0) 2(1.9) 59(52.7) 1(1.5) 0(0.0) 0(0.0) 1(1.5) 38(54.5) 4(1.7) 5(2.1) 2(0.8) 3(1.3) 117(49) Enteric pathogen (s) 1,2,4Single and total pathogens and rotavirus detected more in spring and autumn compared with summer. 3Aeromonas detected more in spring compared with autumn. 5Norovirus detected more in autumn compared with spring.
  • 11. Resistance of Salmonella, Shigella and Aeromonas species isolated from Libyan children with diarrhea to antimicrobial agents Antimicrobial agent Ampicillin (10µg) Amoxicillin/clavulanic acid (30µg) Imipenem (10µg) Chloramphenicol (30µg) Cephalothin (30µg) Ceftriaxone (30µg) Ceftazidime (10ug) Cefepime (30µg) Aztreonam (30µg) Ciprofloxacin (5µg) Nalidixic acid (30µg) Streptomycin (10µg) Tetracycline (30µg) Trimethoprim/ sulfamethoxazole (25µg) Salmonella Shigella Aeromonas (n=19) (n=1) (n=10) Total (n=30) 9 (47.3 ) 1 (100) 10 (100) 20 (66.7) 1 ( 5.2) 0 (0.0) 1 ( 5.2) 11 (57.8) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 12 (63.1) 16 (84.2) 15(78.9) 12 (63.1) 0 (0.0) 0 (0.0) 1 (100) 1 (100) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 1 (100) 0 (0.0) 5 (50) 1 (10) 0 (0.0) 7 (70) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 2 (20) 3 (30) 0 (0.0) 6 (20) 1 (3.3) 2 (6.6) 19 (63.3) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 12 (40) 18 (60) 19 (63.3) 12 (40) 4 (21) 0 (0.0) 1(10) 5 (16.6)
  • 12. Conclusion • To our knowledge this is the first study to identify norovirus, adenovirus, and astrovirus as enteric pathogens causing children diarrhea in Libya. • The findings of the present investigation indicate, in addition to Salmonella and rotavirus, the enteric pathogens norovirus, adenovirus and EAEC are important causative agents of children diarrhea in Tripoli. • On the other hand, Cryptosporidium, Campylobacter and Aeromonas spp. appear to play a minor role in pediatric diarrheal disease in Tripoli.
  • 13. • The emergence of ciprofloxacin-resistant Salmonella is a very serious health problem that should be addressed by public health authorities. • More studies are necessary in other major cities of Libya as well as a dedicated national pathogenspecific surveillance system to identify various etiologies of pediatric diarrhea to determine the exact role of these enteric pathogens. • Furthermore, introduction of a rotavirus vaccine into the vaccination program in Libya to protect the pediatric population is urgently needed.
  • 14. • Rahouma A., Klena JD., Krema Z., Abobker AA., Treesh K., Franka E., Abusnena O., Shaheen HI., El Mohammady H., Abudher A., and Ghenghesh KS. 2011. Enteric Pathogens Associated with Childhood Diarrhea in Tripoli-Libya. American Journal of Tropical Medicine and Hygiene; 84: 886-891. • ghenghesh_micro@yahoo.com • ghenghesh@yahoo.com