Evaluating the Impact of Breastfeeding on Rotavirus Antigenemia and Disease Severity in Indian Children

Abstract

Objectives: To evaluate the contribution of breastfeeding to Rotavirus (RV)-induced antigenemia and/or RNAemia and disease severity in Indian children (<2 yrs age).

Methods: Paired stool and serum samples were collected from (a) hospitalized infants with diarrhea (n = 145) and (b) healthy control infants without diarrhea (n = 28). Stool RV-antigen was screened in both groups by commercial rapid-test and enzyme immunoassay. The disease severity was scored and real-time-PCR was used for viral-load estimation. Serum was evaluated for RV-antigenemia by EIA and RV-RNAemia by RT-PCR. Data was stratified by age-group and breastfeeding status and compared.

Results: Presence of RV-antigenemia and RV-RNAemia was positively related with presence of RV in stool. Disease severity and stool viral-load was significantly associated with RV-antigenemia [(r = 0.74; CI:0.66 to 0.84; P<0.0001,R2 = 0.59) and (r = -0.55; CI:-0.68 to -0.39; P<0.0001,R2 = 0.31) respectively], but not with RV-RNAemia. There was significant reduction in RV-antigenemiarate in the breast-fed group compared to non-breastfed infants, especially in 0-6 month age group (P<0.001). Non-breastfed infants were at risk for RV-antigenemia with severe disease manifestations in form of high Vesikari scores correlating with high fever, more vomiting episodes and dehydration.

Conclusion: RV-antigenemia was common in nonbreastfed children with severe RV-diarrhea and correlated with stool RV-load and disease severity.

Fig 1. Percentage distribution of Rotavirus-positivity among hospitalalized infants <2 yrs age suffering from severe diarrhea in Patna.

(A-B) Temporal distribution of Rotavirus positivity: data of 2 consecutive years (A) and age wise stratified data among all patients (B). (C) Breast-feeding wise distribution of Rotavirus positivity among all patients.

Fig 2. Relation of breast feeding status with Rotavirus infection and related outcome.

(A) Breast-feeding wise distribution of Rotavirus positivity among all patients. (B, D-E) Age stratified data among breastfed and non-breastfed infants for Rotavirus positivity (B), Rotavirus antigenemia (D), and Rotavirus RNAemia (E). (C) Percentage positivity for Rotavirus antigenemia and Rotavirus RNAemia among breastfed and non-breastfed infants of the study.

Fig 3. Rotavirus antigenemia levels among the study groups.

(A-C) Data demonstrates level of Rotavirus antigenemia in acute-phase (A) and convalescent phase (B) sera in infants suffering from severe diarrhea in comparison with the healthy control group. Paired acute-phase and convalescent phase sera were tested for Rotavirus antigenemia levels, that is denoted by levels of Rotavirus antigen (optical density/O.D.) measured by RV-EIA at 450 nm X 1000 units, as mentioned in the text. Broken lines denote the cut-off value.

Fig 4. Rotavirus RNAemia levels among the study groups.

(A-C) Data demonstrates level of Rotavirus RNAemia in acute-phase (A) and convalescent phase (B) sera in infants suffering from severe diarrhea in comparison with the healthy control group. Paired acute-phase and convalescent phase sera were tested for Rotavirus RNAemia levels, denoted by levels of Rotavirus RNA in form of VP7 gene in reverse-transcriptase PCR calculated by band scanning software, as mentioned in the text.

Fig 5. Correlation of Rotavirus antigenemia and RNAemia with stool viral load.

(A-D) Correlation of stool viral load with Rotavirus antigenemia (A,B) and Rotavirus RNAemia (C,D) in acute-phase (A,C) and convalescent phase (B,D) serum of infants suffering from acute gastroenteritis and diarrhea. Paired acute-phase and convalescent phase sera were tested for (i) Rotavirus antigenemia levels, that is denoted by levels of Rotavirus antigen (optical density/O.D.) measured by RV-EIA at 450 nm X 1000 units and (ii) Rotavirus RNAemia levels, denoted by levels of Rotavirus RNA in form of VP7 gene in reverse-transcriptase PCR calculated by band scanning software, as mentioned in the text. Stool viral load is represented by C(t) values in real-time RT-PCR.

Fig 6. Correlation of Rotavirus antigenemia and RNAemia with disease severity.

(A-B) Correlation of disease severity with Rotavirus antigenemia (A) and Rotavirus RNAemia (B) in infants suffering from acute gastroenteritis and diarrhea.(C) Breastfeeding wise stratified data of disease severity among the breastfed and non-breastfed group of infants. Disease severity is represented by Vesikari scores (0–20) according to the 20 point Vesikari scoring system, as mentioned in the text.