FMT for Recurrent C. Difficile Infection
FMT for Recurrent C. Difficile Infection
Recurrent Clostridium difficile infections (rCDI) are a real challenge for the health-care system. Recently, faecal microbiota transplantation (FMT) has been recognised as an effective treatment for rCDI. In this study, we developed a simple and practical, one-step protocol for the preparation of frozen faecal inoculum for FMT and evaluated the efficacy of the frozen preparation in treating rCDI. The previously frozen inoculum was equally effective in treating rCDI as the fresh inoculum with the resolution of symptoms in 22 of 23 (96%) and 25 of 26 (96%) of patients, respectively, at 12 weeks after FMT. Our result is in agreement with the two previous studies showing that frozen faecal suspensions can be applied for FMT without the loss of efficacy. The preparation of multiple transplants from the donated faeces and freeze storage of the ready-to-use suspensions simplified the practical aspects of FMT without loss of efficacy or safety.
The limitations of our study are that it was a retrospective review of patients, who had undergone FMT and that there were small number of patients in the different treatment groups. The strengths are that we used the same clinical setting for FMT as we have used previously in our hospital i.e. using the same protocols for colonic lavage and the colonoscopic route for FMT. Also the patient group, that underwent FMT by using the previously frozen faeces, was comparable to the patient group that was treated by using fresh faeces. Therefore, we consider the comparison of outcomes in the two treatment groups highly reliable.
In this study, the success rate of FMT in treating rCDI was slightly higher than in earlier reports. Differences in results may arise from the route of administration of FMT, patient characteristics and selection, and the C. difficile strains causing the infections. In the patient selection, we avoided FMT with patients with continuous antibiotic treatment for other indication than CDI. Those patients we treated with 10 days fidaxomicin or with indefinite vancomycin instead of FMT. The higher success rate in this study may also have been affected by low prevalence of the hypervirulent C. difficile ribotype 027, which has a higher tendency for recurrence.
Faecal microbiota transplantation restores the diversity and composition of the disrupted intestinal microbiota and subsequently suppresses CDI. Gut functionality is normalised within days and generally, the treatment effect is long-term, but post-FMT antibiotic courses for other causes can lead to the recurrence of CDI. In this study cohort, all recurrences of CDI after FMT developed after antibiotic therapy, interestingly also after vancomycin. We are currently characterising the microbiota of both the universal donors and patients receiving either fresh or previously frozen faeces from the universal donors in an attempt to identify the crucial therapeutic bacteria.
Faecal microbiota transplantation is, albeit being an effective treatment, rather laborious and time consuming partly due to the lengthy process of donor screening and sample processing. In this study, two healthy volunteers donated faeces for the frozen faecal preparations, which were used to treat CDI of 23 patients i.e. we used split donation approach for FMT. The use of split donation and preparation of frozen transplants has obvious benefits, as noted previously. Firstly, the use of split donation and preparation of frozen transplants is efficient, because typically more than one transplant can be prepared from a single donation. The use of a pool of screened universal donors and the storing possibility also decreases the donation times per individual donor. Secondly, overall costs savings are obtained due to reduced costs associated with screening of potential donors. Further, the benefits include improved access to FMT for patients for whom no family members or friends are available as a potential donor. Therefore, the use of stored, pre-screened frozen donor faeces can simplify some practical aspects of this treatment and make FMT available to more patients.
We developed, for the preparation of frozen faecal material to be used for FMT, a one-step protocol, which is practical and does not necessitate the use of specific laboratory facilities. The sole requirement is a −80 °C freezer. The previously frozen faeces showed similar efficacy as fresh faeces in treating rCDI, which is indicative for the good survival of commensal bacteria in the frozen preparation, most notably the species that are essential for the treatment effect. The vast majority of intestinal bacteria are anaerobic and until today, a significant proportion of these bacteria are as yet uncultured. Anaerobic members of the intestinal microbiota are generally presumed to be very sensitive to oxygen and strictly anaerobic techniques have been instrumental in the cultivation of such bacteria. The simple protocol used in this study to prepare the frozen faecal transplants allows fast handling and freezing, and thereby avoids the excessive exposure of fastidious anaerobes to oxygen without the need of working under anoxic atmosphere. By the fast preparation, we succeeded to preserve sufficient numbers and diversity of bacteria for effective FMT. Recently, Youngster et al. presented a simple protocol for preparing frozen transplants under ambient air (not nitrogen) and achieved resolution of rCDI in 14 of 20 (70%) of patients after a single FMT with the frozen transplant. The success rate is lower than in this study (96%), but as the patient numbers are small in both studies, no conclusions can be drawn on the superiority of either preparation protocol. In this study, the frozen preparations were stored up to 16 weeks before used for FMT with successful clinical outcome. Thus, the frozen transplants seem to stand for a relatively long storage time at −80 °C. Later, we have used frozen preparations with up to 7 months storage time and thus far the clinical outcomes in treating rCDI have been similar (data not shown).
In our patient group, mild transient fever appeared for two patients receiving frozen stool, but no other significant side effects were observed. Although both patients were from the frozen transplant group, we consider that FMT itself rather than glycerol used in the frozen preparations caused the side-effect. Previously, transient self-limiting fever has been reported as a side-effect of FMT in case series of ulcerative colitis patients. The authors considered fever as a result of temporary systemic immune response to the applied bacteria, which we find a likely explanation also for our patients. In general, our results are in agreement with the previous reports considering FMT as a safe treatment for patients with CDI. Thus far, the benefits of FMT clearly outweigh the observed harms, but the potential risks of FMT have to be acknowledged. The risks of FMT are the risks of colonoscopy including perforation and infectious risks due to the administration of faeces from another person. The latter risks have been minimised by the careful pre-screening of donors. However, the transmission of contagious agents, including possible yet-unknown-agents that haven't been noticed in the donor screening or, which the donor has acquired between the screening tests and faecal donation cannot be definitely excluded. Thereby, there is a possibility of transferring infective agents, some of which may cause a disease even after a considerable time period after FMT. The long-term effects of FMT possibly modifying the risks of autoimmune diseases, allergies and functional intestinal diseases should also be addressed. Long-term follow-ups of FMT patients, particularly children and young adults, who have a longer time to develop side effects than elderly patients, are needed to address these issues.
The frozen faecal preparations included 17 g of glycerol as a cryoprotectant to improve bacterial viability upon freezing. Glycerol is widely used in pharmaceutical formulations including use as a laxative in suppositories or small-volume enemas. Glycerol is of low toxicity and its adverse effects in humans have been studied. After oral ingestion, glycerol is rapidly absorbed and metabolised and the adverse effects are primarily due to its dehydrating action at oral doses that are higher than 0.75 grams per kg of body weight (g/kg bw). In FMT via colonoscopy, the faecal preparation is infused into caecum and some part of the glycerol may be absorbed. The total glycerol dose is 17 g i.e. 0.34 g/kg bw in a 50 kg subject, which is less than half of the oral exposure levels, where dehydrating effects are seen. Thus, the used amount of glycerol does not represent a noteworthy risk for otherwise healthy adults. However, in diabetic patients, oral administration of glycerol increases blood glucose level slightly and therefore, some caution should be taken if glycerol-containing transplant is given to a diabetic patient, especially if duodenal infusion is used.
FMT is being considered as a treatment also for other intestinal disorders and its use is likely to increase over the coming years. The described protocol for frozen faecal preparations can significantly simplify clinical work by facilitating the banking of faeces from carefully selected and screened donors as frozen preparations that are readily available to be used as transplants when needed.
Discussion
Recurrent Clostridium difficile infections (rCDI) are a real challenge for the health-care system. Recently, faecal microbiota transplantation (FMT) has been recognised as an effective treatment for rCDI. In this study, we developed a simple and practical, one-step protocol for the preparation of frozen faecal inoculum for FMT and evaluated the efficacy of the frozen preparation in treating rCDI. The previously frozen inoculum was equally effective in treating rCDI as the fresh inoculum with the resolution of symptoms in 22 of 23 (96%) and 25 of 26 (96%) of patients, respectively, at 12 weeks after FMT. Our result is in agreement with the two previous studies showing that frozen faecal suspensions can be applied for FMT without the loss of efficacy. The preparation of multiple transplants from the donated faeces and freeze storage of the ready-to-use suspensions simplified the practical aspects of FMT without loss of efficacy or safety.
The limitations of our study are that it was a retrospective review of patients, who had undergone FMT and that there were small number of patients in the different treatment groups. The strengths are that we used the same clinical setting for FMT as we have used previously in our hospital i.e. using the same protocols for colonic lavage and the colonoscopic route for FMT. Also the patient group, that underwent FMT by using the previously frozen faeces, was comparable to the patient group that was treated by using fresh faeces. Therefore, we consider the comparison of outcomes in the two treatment groups highly reliable.
In this study, the success rate of FMT in treating rCDI was slightly higher than in earlier reports. Differences in results may arise from the route of administration of FMT, patient characteristics and selection, and the C. difficile strains causing the infections. In the patient selection, we avoided FMT with patients with continuous antibiotic treatment for other indication than CDI. Those patients we treated with 10 days fidaxomicin or with indefinite vancomycin instead of FMT. The higher success rate in this study may also have been affected by low prevalence of the hypervirulent C. difficile ribotype 027, which has a higher tendency for recurrence.
Faecal microbiota transplantation restores the diversity and composition of the disrupted intestinal microbiota and subsequently suppresses CDI. Gut functionality is normalised within days and generally, the treatment effect is long-term, but post-FMT antibiotic courses for other causes can lead to the recurrence of CDI. In this study cohort, all recurrences of CDI after FMT developed after antibiotic therapy, interestingly also after vancomycin. We are currently characterising the microbiota of both the universal donors and patients receiving either fresh or previously frozen faeces from the universal donors in an attempt to identify the crucial therapeutic bacteria.
Faecal microbiota transplantation is, albeit being an effective treatment, rather laborious and time consuming partly due to the lengthy process of donor screening and sample processing. In this study, two healthy volunteers donated faeces for the frozen faecal preparations, which were used to treat CDI of 23 patients i.e. we used split donation approach for FMT. The use of split donation and preparation of frozen transplants has obvious benefits, as noted previously. Firstly, the use of split donation and preparation of frozen transplants is efficient, because typically more than one transplant can be prepared from a single donation. The use of a pool of screened universal donors and the storing possibility also decreases the donation times per individual donor. Secondly, overall costs savings are obtained due to reduced costs associated with screening of potential donors. Further, the benefits include improved access to FMT for patients for whom no family members or friends are available as a potential donor. Therefore, the use of stored, pre-screened frozen donor faeces can simplify some practical aspects of this treatment and make FMT available to more patients.
We developed, for the preparation of frozen faecal material to be used for FMT, a one-step protocol, which is practical and does not necessitate the use of specific laboratory facilities. The sole requirement is a −80 °C freezer. The previously frozen faeces showed similar efficacy as fresh faeces in treating rCDI, which is indicative for the good survival of commensal bacteria in the frozen preparation, most notably the species that are essential for the treatment effect. The vast majority of intestinal bacteria are anaerobic and until today, a significant proportion of these bacteria are as yet uncultured. Anaerobic members of the intestinal microbiota are generally presumed to be very sensitive to oxygen and strictly anaerobic techniques have been instrumental in the cultivation of such bacteria. The simple protocol used in this study to prepare the frozen faecal transplants allows fast handling and freezing, and thereby avoids the excessive exposure of fastidious anaerobes to oxygen without the need of working under anoxic atmosphere. By the fast preparation, we succeeded to preserve sufficient numbers and diversity of bacteria for effective FMT. Recently, Youngster et al. presented a simple protocol for preparing frozen transplants under ambient air (not nitrogen) and achieved resolution of rCDI in 14 of 20 (70%) of patients after a single FMT with the frozen transplant. The success rate is lower than in this study (96%), but as the patient numbers are small in both studies, no conclusions can be drawn on the superiority of either preparation protocol. In this study, the frozen preparations were stored up to 16 weeks before used for FMT with successful clinical outcome. Thus, the frozen transplants seem to stand for a relatively long storage time at −80 °C. Later, we have used frozen preparations with up to 7 months storage time and thus far the clinical outcomes in treating rCDI have been similar (data not shown).
In our patient group, mild transient fever appeared for two patients receiving frozen stool, but no other significant side effects were observed. Although both patients were from the frozen transplant group, we consider that FMT itself rather than glycerol used in the frozen preparations caused the side-effect. Previously, transient self-limiting fever has been reported as a side-effect of FMT in case series of ulcerative colitis patients. The authors considered fever as a result of temporary systemic immune response to the applied bacteria, which we find a likely explanation also for our patients. In general, our results are in agreement with the previous reports considering FMT as a safe treatment for patients with CDI. Thus far, the benefits of FMT clearly outweigh the observed harms, but the potential risks of FMT have to be acknowledged. The risks of FMT are the risks of colonoscopy including perforation and infectious risks due to the administration of faeces from another person. The latter risks have been minimised by the careful pre-screening of donors. However, the transmission of contagious agents, including possible yet-unknown-agents that haven't been noticed in the donor screening or, which the donor has acquired between the screening tests and faecal donation cannot be definitely excluded. Thereby, there is a possibility of transferring infective agents, some of which may cause a disease even after a considerable time period after FMT. The long-term effects of FMT possibly modifying the risks of autoimmune diseases, allergies and functional intestinal diseases should also be addressed. Long-term follow-ups of FMT patients, particularly children and young adults, who have a longer time to develop side effects than elderly patients, are needed to address these issues.
The frozen faecal preparations included 17 g of glycerol as a cryoprotectant to improve bacterial viability upon freezing. Glycerol is widely used in pharmaceutical formulations including use as a laxative in suppositories or small-volume enemas. Glycerol is of low toxicity and its adverse effects in humans have been studied. After oral ingestion, glycerol is rapidly absorbed and metabolised and the adverse effects are primarily due to its dehydrating action at oral doses that are higher than 0.75 grams per kg of body weight (g/kg bw). In FMT via colonoscopy, the faecal preparation is infused into caecum and some part of the glycerol may be absorbed. The total glycerol dose is 17 g i.e. 0.34 g/kg bw in a 50 kg subject, which is less than half of the oral exposure levels, where dehydrating effects are seen. Thus, the used amount of glycerol does not represent a noteworthy risk for otherwise healthy adults. However, in diabetic patients, oral administration of glycerol increases blood glucose level slightly and therefore, some caution should be taken if glycerol-containing transplant is given to a diabetic patient, especially if duodenal infusion is used.
FMT is being considered as a treatment also for other intestinal disorders and its use is likely to increase over the coming years. The described protocol for frozen faecal preparations can significantly simplify clinical work by facilitating the banking of faeces from carefully selected and screened donors as frozen preparations that are readily available to be used as transplants when needed.
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