Clinical characteristics of patients

A total of 158 patients undergoing HSCT, aged from 9 months to 16 years, were enrolled in the present study between November 2019 and February 2021. The median patient age was 6.00 years, with 37.97% of patients being female and 96.84% being allogeneic HSCT recipients. The most common underlying diseases were hematological malignancies (40.51%), aplastic anemia (23.42%), primary immunodeficiency (17.72%), and inherited metabolic disorders (12.66%).

Ninety-five of the 158 (60.13%) patients tested positive for at least one GI pathogen before transplantation and were therefore classified as colonized patients. There were no significant differences regarding age, stem cell source or outcomes in colonized or non-colonized patients (Table 1).

Table 1 Clinical characteristics of patients colonized or not colonized with a gastrointestinal pathogen before HSCTEtiologies of diarrhea after transplantation

Eighty-four of the 158 (53.16%) patients developed diarrhea after transplantation (Fig. 1). In half of these patients, diarrhea was considered to be caused by a single factor, including infection (21/84, 25.00%), medication-induced diarrhea (14/84, 16.67%), GVHD (6/84, 7.14%), and engraftment syndrome (1/84, 1.19%). In the other 37 patients, diarrhea was due to multiple factors, including infection plus GVHD (13/84, 15.48%), infection plus medication-induced (22/84, 26.19%), infection plus GVHD plus medication-induced (1/84, 1.19%), and GVHD plus medication-induced (1/84, 1.19%) (Fig. 2a). In summary, the primary cause of post-transplantation diarrhea was infection (57/84, 67.86%), followed by medication (38/84, 45.24%) and GVHD (21/84, 25.00%) (Fig. 2b).

Fig. 2

Etiologies of diarrhea after transplantation. a Etiologies of diarrhea in 84 patients who experienced diarrhea episodes after transplantation; b etiologies of diarrhea that combined all factors, including infection, GVHD and medication; c the incidence of diarrhea and infectious diarrhea in colonized and non-colonized patients. GVHD graft-versus-host disease

To determine whether pre-transplantation colonization with GI pathogens has an impact on post-transplantation diarrhea, we compared the proportion of diarrhea patients in the colonized and non-colonized groups. The incidence of diarrhea was higher in the colonized patients (56/95, 58.95%) than in the non-colonized patients (28/63, 44.44%), although the difference was not significant (P = 0.074). However, there was a significant increase in the incidence of infection-induced diarrhea in the colonized group (47/95, 49.47%) compared to the non-colonized group (10/63, 15.87%) (P < 0.001, Fig. 2c).

Pathogens detected by FilmArray GI panel

In total, 95 patients were colonized by at least one GI pathogen pre-transplantation, resulting in 124 pathogens being detected. The most commonly detected pathogen was C. difficile, present in 62 (65.26%) patients. Other pathogens detected included EPEC (21/95, 22.11%), norovirus (19/95, 20.00%), EAEC (7/95, 7.37%), etc. (Fig. 3, left column). Fifty-nine patients tested positive for at least one GI pathogen 28 ± 7 days after transplantation. The most commonly detected pathogen was C. difficile, present in 38 (64.41%) patients, followed by norovirus (21/59, 35.59%) (Fig. 3, right column). Interestingly, we found that EPEC was only present in 1 of the 59 patients. No other pathogenic E. coli, including EAEC and EPEC, were detected in the 59 patients. These results indicated that the conditioning therapy as well as other medications used during the transplantation period may eliminate most colonizing bacterial pathogens, with the exception of C. difficile. In another way, viral pathogens were almost unaffected.

Fig. 3

Pathogens detected by FilmArray gastrointestinal (GI) panel in pre- and post-transplantation stool samples. The number of patients who had a clinically relevant post-transplantation diarrheal infection due to pre-transplantation colonization is shown for each pathogen (red part in the bar plot). EPEC enteropathogenic Escherichia coli, EAEC enteroaggregative E. coli, ETEC enterotoxigenic E. coli

Risk of post-transplantation diarrhea due to colonizing pathogens

We next assessed the proportions of patients who developed a diarrheal infection due to their colonized pathogens pre-transplantation. Overall, 45 of the 124 (36.29%) colonized pathogens detected in 39 patients led to post-transplantation diarrhea. In detail, 14 of 19 (73.68%) patients who were initially colonized with norovirus pre-transplantation developed a post-transplantation norovirus infection. Twenty-four of 62 (38.71%) patients colonized with C. difficile developed a diarrheal infection. Only 2 of 21 (9.52%) patients colonized with EPEC and 1 of 7 (14.29%) patients colonized with EAEC developed post-transplantation EPEC and EAEC infections, respectively. In addition, four patients were colonized with ETEC, and three patients were colonized with Giardia lamblia, astrovirus and Vibrio cholerae, but none of them developed post-transplantation diarrhea due to their colonized pathogens (Fig. 3, left column).

We also assessed the proportions of patients with different underlying diseases who developed infectious diarrhea due to their colonized pathogens. Twenty-four of 28 (85.71%) patients with primary immunodeficiency were colonized by at least one GI pathogen pre-transplantation. In 14 patients who developed infectious diarrhea post-transplantation, 11 of them (11/14, 78.57%) had colonization by pathogens before transplantation. Similarly, 41 of 64 (64.06%) patients with hematological malignancies were colonized with GI pathogens, and 19 of 23 (82.60%) patients who developed a diarrheal infection were colonized by pathogens (Fig. 4). On the other hand, patients with aplastic anemia and inherited metabolic disorders were much less frequently colonized with GI pathogens (35.14% and 50.00%, respectively, P < 0.001), and the risk of developing infectious diarrhea was also comparatively lower (28.57% and 37.50%, respectively, P = 0.005) (Fig. 4).

Fig. 4

Impact of different underlying diseases on pathogen colonization and subsequent development of infectious diarrhea due to the colonized pathogens. a The number and proportions of patients with different underlying diseases who were colonized with gastrointestinal pathogens pre-transplantation; b the number and proportions of patients in the colonized group who developed infectious diarrhea due to their colonized pathogens. HM hematological malignancies, PI primary immunodeficiency, AA aplastic anemia, IMD inherited metabolic disorders

Comparison of FilmArray GI-detected pathogens with clinic-confirmed pathogens detected during diarrhea episodes

In 57 patients who developed infectious diarrhea posttransplantation, 92 pathogens were detected by various methods and confirmed by clinicians. Norovirus and C. difficile were the most frequently detected diarrheal pathogens, present in 26 and 25 patients, respectively. EBV, adenovirus, and Enterococcus faecium were present in 7, 5, and 4 patients, respectively. Other pathogenic microbes, such as Salmonella spp., CMV, and HHV6, were less frequently detected (Table 2, left part).

Table 2 Pathogens detected during diarrhea episodes