Use of quantitative molecular diagnostic methods to identify causes of diarrhoea in children: a reanalysis of the GEMS case-control study.
Diarrhoea is the second leading cause of mortality in children worldwide, but establishing the cause can be complicated by diverse diagnostic approaches and varying test characteristics. We used quantitative molecular diagnostic methods to reassess causes of diarrhoea in the Global Enteric Multicenter Study (GEMS).



GEMS was a study of moderate to severe diarrhoea in children younger than 5 years in Africa and Asia. We used quantitative real-time PCR (qPCR) to test for 32 enteropathogens in stool samples from cases and matched asymptomatic controls from GEMS, and compared pathogen-specific attributable incidences with those found with the original GEMS microbiological methods, including culture, EIA, and reverse-transcriptase PCR. We calculated revised pathogen-specific burdens of disease and assessed causes in individual children.



We analysed 5304 sample pairs. For most pathogens, incidence was greater with qPCR than with the original methods, particularly for adenovirus 40/41 (around five times), Shigella spp or enteroinvasive Escherichia coli (EIEC) and Campylobactor jejuni o C coli (around two times), and heat-stable enterotoxin-producing E coli ([ST-ETEC] around 1·5 times). The six most attributable pathogens became, in descending order, Shigella spp, rotavirus, adenovirus 40/41, ST-ETEC, Cryptosporidium spp, and Campylobacter spp. Pathogen-attributable diarrhoeal burden was 89·3% (95% CI 83·2-96·0) at the population level, compared with 51·5% (48·0-55·0) in the original GEMS analysis. The top six pathogens accounted for 77·8% (74·6-80·9) of all attributable diarrhoea. With use of model-derived quantitative cutoffs to assess individual diarrhoeal cases, 2254 (42·5%) of 5304 cases had one diarrhoea-associated pathogen detected and 2063 (38·9%) had two or more, with Shigella spp and rotavirus being the pathogens most strongly associated with diarrhoea in children with mixed infections.



A quantitative molecular diagnostic approach improved population-level and case-level characterisation of the causes of diarrhoea and indicated a high burden of disease associated with six pathogens, for which targeted treatment should be prioritised.



Bill & Melinda Gates Foundation.
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA.
Jie
J
Liu
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA.
James A
JA
Platts-Mills
Center for Global Health Research, Kenya Medical Research Institute, Nairobi, Kenya.
Jane
J
Juma
Department of Paediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
Furqan
F
Kabir
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA.
Joseph
J
Nkeze
Medical Research Council Unit, Banjul, The Gambia.
Catherine
C
Okoi
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA.
Darwin J
DJ
Operario
International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR, B), Dhaka, Bangladesh.
Jashim
J
Uddin
International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR, B), Dhaka, Bangladesh.
Shahnawaz
S
Ahmed
Centro de Investigação em Saúde da Manhiça, Maputo, Mozambique; Barcelona Centre for International Health Research (CRESIB, Hospital Clinic-Universitat de Barcelona), Barcelona, Spain.
Pedro L
PL
Alonso
Medical Research Council Unit, Banjul, The Gambia.
Martin
M
Antonio
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA.
Stephen M
SM
Becker
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA.
William C
WC
Blackwelder
Global Disease Detection Division, Kenya Office of the US Centers for Disease Control and Prevention, Nairobi, Kenya.
Robert F
RF
Breiman
International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR, B), Dhaka, Bangladesh.
Abu S G
AS
Faruque
Global Disease Detection Division, Kenya Office of the US Centers for Disease Control and Prevention, Nairobi, Kenya.
Barry
B
Fields
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA.
Jean
J
Gratz
International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR, B), Dhaka, Bangladesh.
Rashidul
R
Haque
International Centre for Diarrhoeal Disease Research, Bangladesh (ICDDR, B), Dhaka, Bangladesh.
Anowar
A
Hossain
Medical Research Council Unit, Banjul, The Gambia.
M Jahangir
MJ
Hossain
Medical Research Council Unit, Banjul, The Gambia.
Sheikh
S
Jarju
Department of Paediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
Farah
F
Qamar
Department of Paediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
Najeeha Talat
NT
Iqbal
Medical Research Council Unit, Banjul, The Gambia.
Brenda
B
Kwambana
Centro de Investigação em Saúde da Manhiça, Maputo, Mozambique.
Inacio
I
Mandomando
Public Health Sciences, University of Virginia, Charlottesville, VA, USA.
Timothy L
TL
McMurry
Center for Global Health Research, Kenya Medical Research Institute, Nairobi, Kenya.
Caroline
C
Ochieng
Center for Global Health Research, Kenya Medical Research Institute, Nairobi, Kenya.
John B
JB
Ochieng
Center for Global Health Research, Kenya Medical Research Institute, Nairobi, Kenya.
Melvin
M
Ochieng
Global Disease Detection Division, Kenya Office of the US Centers for Disease Control and Prevention, Nairobi, Kenya.
Clayton
C
Onyango
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA.
Sandra
S
Panchalingam
Department of Paediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
Adil
A
Kalam
Department of Paediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
Fatima
F
Aziz
Department of Paediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
Shahida
S
Qureshi
National Institute of Cholera and Enteric Diseases, Kolkata, India.
Thandavarayan
T
Ramamurthy
Public Health Sciences, University of Virginia, Charlottesville, VA, USA.
James H
JH
Roberts
Medical Research Council Unit, Banjul, The Gambia.
Debasish
D
Saha
Centre pour le Développement des Vaccins, Bamako, Mali.
Samba O
SO
Sow
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA.
Suzanne E
SE
Stroup
National Institute of Cholera and Enteric Diseases, Kolkata, India.
Dipika
D
Sur
Centre pour le Développement des Vaccins, Bamako, Mali.
Boubou
B
Tamboura
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA.
Mami
M
Taniuchi
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA.
Sharon M
SM
Tennant
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA.
Deanna
D
Toema
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA.
Yukun
Y
Wu
Department of Paediatrics and Child Health, Aga Khan University, Karachi, Pakistan.
Anita
A
Zaidi
Department of Pediatrics, University of Virginia, Charlottesville, VA, USA.
James P
JP
Nataro
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Pediatrics, University of Maryland School of Medicine, Baltimore, MD, USA.
Karen L
KL
Kotloff
Center for Vaccine Development and Institute of Global Health, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Pediatrics, University of Maryland School of Medicine, Baltimore, MD, USA; Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA.
Myron M
MM
Levine
Division of Infectious Diseases and International Health, University of Virginia, Charlottesville, VA, USA. Electronic address: erh6k@virginia.edu.
Eric R
ER
Houpt
D000256:Adenoviridae / Q000302:isolation & purification / Q000472:pathogenicity; D000349:Africa / Q000453:epidemiology; D001208:Asia / Q000453:epidemiology; D001419:Bacteria / Q000302:isolation & purification / Q000472:pathogenicity; D001424:Bacterial Infections / Q000175:diagnosis; D002167:Campylobacter / Q000302:isolation & purification / Q000472:pathogenicity; D016022:Case-Control Studies; D002675:Child, Preschool; D060085:Coinfection; D017281:Cost of Illness; D003458:Cryptosporidium / Q000302:isolation & purification / Q000472:pathogenicity; D003967:Diarrhea / Q000453:epidemiology / Q000382:microbiology* / Q000821:virology*; D004926:Escherichia coli / Q000302:isolation & purification / Q000472:pathogenicity; D005260:Female; D006801:Humans; D015994:Incidence; D007223:Infant; D008297:Male; D012401:Rotavirus / Q000302:isolation & purification / Q000472:pathogenicity; D012760:Shigella / Q000302:isolation & purification / Q000472:pathogenicity; D014777:Virus Diseases / Q000175:diagnosis; D014780:Viruses / Q000302:isolation & purification / Q000472:pathogenicity
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