Evidence that sustained growth suppression of intestinal anaerobic bacteria reduces the risk of acute graft-versus-host disease after sibling marrow transplantation.

The influence of intestinal bacterial decontamination on the occurrence of grades II to IV acute graft-versus-host disease (GVHD) was retrospectively analyzed in 194 predominantly adult patients treated by genotypically identical sibling marrow transplantation under conditions of strict protective isolation and intestinal antimicrobial decontamination. Forty-five patients (23%) developed acute GVHD and univariate analysis identified four features that significantly increased the risk for this reaction: chronic myeloid leukemia as the underlying disease, as compared with all other disease categories (P < .0001); female marrow donors for male recipients, as compared with other gender combinations (P < .005); ineffective, as compared with sustained growth suppression of intestinal anaerobic bacteria (P < .006); and methotrexate as the sole immunoprophylactic compound, as compared with cyclosporine containing regimens (P < .05). Using the duration of anaerobic growth suppression as a time-dependent explanatory variable, proportional hazards regression analysis confirmed these features as independent predictors for acute GVHD with relative risk estimates of 1.9 (95% confidence interval [CI], 1.3 to 2.7) for the immunoprophylactic regimen (P < .0004), of 1.8 (95% CI, 1.3 to 2.5) for the underlying disease (P < .0005), of 1.7 (95% CI, 1.2 to 2.5) for anaerobic decontamination (P < .002), and of 1.3 (95% CI, 1.1 to 1.6) for the donor/recipient gender combination (P < .008), respectively. Best subset selection modeling also identified the quality of anaerobic decontamination as the third most important predictor for acute GVHD, when all four significant features were included. Estimates of acute GVHD stratified by the quality of anaerobic bacterial growth suppression showed a strong influence of anaerobic decontamination in patients burdened by at least one of the other unfavorable factors (P < .009). In conclusion, this study provides strong evidence that sustained growth suppression of intestinal anaerobic bacteria after clinical sibling marrow transplantation can independently modulate the occurrence of grades II to IV acute GVHD, which is in concordance with previous results from animal transplantation models. Antimicrobial chemotherapy specifically targeted to the intestinal anaerobic bacterial microflora may be complementarily useful in preventing acute GVHD and should be investigated in a prospective trial.

Prevention of infection in acute leukemia.

In a randomized study comparing cotrimoxazole plus colistin with ciprofloxacin, each in combination with nonabsorbable antimycotics, the incidence of major infections in terms of septicemias and pneumonias as well as of minor infections and episodes of unexplained fever (FUO) was higher in patients treated with ciprofloxacin. In cases of microbiologically documented infections, gram-positive cocci dominated by far. In surveillance cultures of oral washings and of feces, gram-negative enterobacteria were only rarely detected; however, large numbers of cultures were positive for Acinetobacter species. There were four cases of documented Pneumocystis carinii pneumonia in patients not receiving cotrimoxazole. The incidence of documented mycotic infections as well as the detection of fungi in surveillance cultures was similar in both treatment groups. A decrease in the number of adverse events, especially of allergic reactions, could not be achieved by the administration of ciprofloxacin. In conclusion, cotrimoxazole plus colistin in combination with nonabsorbable antimycotics remains the standard regimen for prevention of infection in patients with acute leukemia undergoing aggressive remission induction therapy. A detailed analysis of study II will be prepared for publication.

Ciprofloxacin and norfloxacin for selective decontamination in patients with severe granulocytopenia.

In a randomized multicenter study, ciprofloxacin and norfloxacin, each in two different dose regimens and in combination with non-absorbable antimycotics, were administered to 51 patients with acute leukaemia undergoing aggressive remission induction chemotherapy for infection prevention. Both drugs showed an effective elimination of gram-negative potential pathogens and Staphylococcus aureus not affecting the anaerobic flora of the gastrointestinal tract. A low incidence of side effects and a satisfactory patient compliance could be observed. A daily dosage of 1,000 mg ciprofloxacin or 800 mg norfloxacin is recommended for infection prevention in severely granulocytopenic patients.

Infections after experimental cadaver bone marrow transplantation in beagle dogs. Transplantations with and without selective gastrointestinal decontamination.

Experimental transplantations of cadaver bone marrow (BMT) in beagle dogs were performed to evaluate the problems and potentials in a preclinical setting. The effectiveness of selective decontamination of the gut (SD) and gnotobiotic surveillance in preventing infections during longer aplastic periods was investigated. Three groups of dogs were compared. Group A: controls. Group B: dogs with BMT, without SD and irregular gnotobiotic surveillance. Group C: dogs with BMT, with SD and regular gnotobiotic surveillance. The intestinal colonization of normal healthy beagles shows similarities as well as dissimilarities to the human intestinal microflora. Aerobic potentially pathogenic organisms do not colonize the gut of healthy beagles under our keeping conditions. SD resulted in a significant decrease in infections with Escherichia coli and Plesiomonas. Infections with anaerobes, as well as bacterial infections of the respiratory tract were, however, not prevented. The intestinal colonization in dogs of group C with Clostridium difficile is another obvious effect of SD. The infections encountered during the study indicate the importance of the "take" for the clinical significance and outcome of intestinal colonization with potentially pathogenic organisms. In order to reduce the drug burden of BMT patients, we consider the elimination of routine SD after BMT not to be superior to gnotobiotic surveillance and germ-specific short term decontamination.

Allogeneic bone marrow transplantation for acute leukaemia or chronic myeloid leukaemia in the fifth decade of life.

To determine the influence of advanced age on long-term survival after allogeneic bone marrow transplantation (BMT), the probability of survival and the frequency of transplantation-associated complications were analysed retrospectively in 20 patients with acute leukaemia (AL) or chronic myeloid leukaemia (CML), who were 40-49 years of age (median 44.5 years) at the time of transplant. The results of this patient group were compared to those of 32 patients aged 30-39 years (median 33.5 years) with AL or CML, who also underwent BMT during the same period of time. The overall actuarial survival of the two age groups was comparable with 44% and 41% at 5.9 and 5.6 years, respectively. Patients with standard risk criteria (i.e. HLA-genotypically identical sibling donor, 1st chronic phase of CML or 1st remission of AL) showed a higher probability of survival in both groups (62% at 5.9 years in older patients and 59% at 5.5 years in younger patients, respectively). In contrast, actuarial survival in patients who underwent BMT at an advanced stage of their disease or with marrow from a partially HLA-compatible donor was significantly inferior (P = 0.04). The cumulative incidence of acute and chronic graft-versus-host disease was low in older patients (27%), who received marrow from an HLA-identical sibling donor. The most frequent cause of death was interstitial pneumonia, occurring in seven of the older patients (35%) and in seven of the younger patients (22%). This difference, however, was not statistically significant. Our results indicate that allogenic marrow transplantation in the fifth decade of life might be associated with a tolerable risk of transplantation-related complications. This treatment modality may therefore be regarded as first-line therapy for patients in 1st remission of AL or first chronic phase of CML, who show a normal performance status. The same applies to older patients in advanced stages of disease, since the results are comparable to those achieved in the younger patient group.

Bone marrow transplantation in acute leukemia.

In Essen 142 bone marrow transplantations were carried out between December 1975 and February 1985. In 74 cases the indication was acute leukemia in relapse (n = 23) or in first or consecutive remission (n = 51). The conditioning regimen consisted of cyclophosphamide or the combination of cyclophosphamide and total body irradiation. All patients were treated under strict gnotobiotic care. To mitigate the risk of CMV infections, intravenous CMV-hyperimmune globulin and CMV-negative blood products have been applied routinely for 2 years. MTX was used as prophylaxis against GvHD. In the prognostically unfavorable group of acute leukemia in relapse, only one patient showed long-term survival. In this patient, leukemic relapse occurred 6 years after transplantation. The survival rate of AML patients grafted during the first remission is 55% (16/29) with a median observation time of 41 months. For patients grafted in first or consecutive remission of acute lymphoblastic leukemia, the survival rate is 50% (7/14) with a maximal observation time of 34 months. The overall incidence of GvHD in patients at risk was 28% in aplastic anemia, 26% in AML, 9% in ALL, and 63% in CML. In aplastic anemia, no patient developed an interstitial pneumonia. In leukemia, the risk of fatal interstitial pneumonia was 34%.

Prevention of infection in acute leukemia: a prospective randomized study on the efficacy of two different drug regimens for antimicrobial prophylaxis.

In a prospective study patients with acute leukemia undergoing remission induction therapy were randomized to receive either a regimen of non-absorbable antimicrobial drugs (colistin and neomycin) or of absorbable and non-absorbable drugs (trimethoprim-sulfamethoxazole [TMP-SMZ] and colistin) for antibacterial prophylaxis. For antifungal prophylaxis patients in both groups were given oral amphotericin B. The proportion of patients without acquired infections and the median of study time to the first acquired infection did not differ significantly between the two treatment groups (p greater than 0.05). Septicemias occurred in nine out of 49 recipients of colistin and neomycin and in one out of 56 patients receiving TMP-SMZ and colistin (p = 0.03). Localized infections and fever episodes without proven infections were equally distributed between the two groups. The incidence of febrile days and of days on parenteral antibiotic therapy was significantly lower in the group given TMP-SMZ and colistin (p less than 0.05). The duration of severe granulocytopenia and thrombocytopenia did not differ significantly between the two groups (p greater than 0.05).

Bone marrow transplantation in severe aplastic anemia and acute or chronic leukemia.

In Essen 121 bone marrow transplantations were carried out. The indications were severe aplastic anemia (n = 18), acute leukemia in relapse (n = 20), acute leukemia in remission (n = 46) or chronic myeloid leukemia (n = 37). The conditioning regimen consisted of cyclophosphamide or the combination of cyclophosphamide and total body irradiation. All patients were treated under strict gnotobiotic care. To mitigate the risk of CMV infections intravenous CMV-hyperimmunoglobulin and CMV-negative blood products have been applied routinely since two years. MTX was used as prophylaxis against GVH-disease. In case of severe aplastic anemia 13 patients (72%) are still alive with a median observation time of 24 months. In the prognostically unfavourable group of acute leukemia in relapse only one patient showed long term survival. In this patient leukemic relapse occurred six years after transplantation. The survival rate of AML patients grafted during the first remission is 55% (15/27) with a median observation time of 40 months. For patients grafted in first or consecutive remission of ALL the survival rate is 42% (5/12) with a maximal observation time of 29 months. Out of 37 patients grafted because of CML, eight were in an advanced stage of the disease. 13 patients are still alive, the maximal observation time is 37 months. The overall incidence of GVHD in patients at risk was 28% in aplastic anemia, 26% in AML, 9% in ALL and 63% in CML. In aplastic anemia no patient developed an interstitial pneumonia. In leukemia the risk of fatal interstitial pneumonia was 34%.

Bone marrow transplantation for chronic granulocytic leukaemia.

Twenty-one patients with chronic granulocytic leukaemia underwent marrow transplantation. The donors were human-lymphocyte antigen-identical siblings in 19 cases. In the remaining 2 cases the donor was a parent in one and an identical twin in the other. The preparatory regimen included cyclophosphamide and 8.6 Gy total body irradiation given at either a dose of 0.1 Gy/min or 0.04 Gy/min. Five patients were in the accelerated phase of the disease, one was in remission following blast crisis, and the rest were all in the chronic phase. After chemotherapy and irradiation, all patients received bone marrow transplants. To date, nine patients are still alive, with a median survival of 64 days (range 28-683 days). One patient continued to have leukaemic cells and in another, the leukaemia recurred 18 months following transplantation. Interstitial pneumonitis was the cause of death of eight patients (38%). Graft-versus-host disease occurred in ten patients (47%).

[Gnotobiotic studies on SPF mice in relation to a study of tumor development in the colon after cholecystectomy]. / Gnotobiotische Untersuchungen an SPF-Mäusen im Rahmen einer Studie zur Tumorentwicklung im Kolon nach Cholezystektomie.

The relation between cholecystectomy and colon carcinogenesis has not been fully elucidated. As bacteria may be involved in the carcinogenic process, we investigated the effect of cholecystectomy and dimethylhydrazine (DMH) administration to SPF NMRI mice with regard to tumour genesis and bacterial colonisation of the intestine. It results from this study that cholecystectomy does not influence tumour genesis and that 6-7 months post operationem and DMH administration tumours and bacteria originally not found in the animals develop: clostridia, eubacteria spec. which cannot be differentiated and E. lentum. Theses changes appear in group II of mice (laparotomy and DMH) and group III (cholecystectomy and DMH), but not in group I (controls). From the results of this study we cannot conclude whether the tumours or the new bacteria appeared first. Biochemical investigations of C. innocuum, C. paraputrificum and C. tertium indicated that these bacteria metabolised bile acids by a specific metabolic step only but not produced carcinogenic substances themselves. If bacteria are involved in tumorgenesis, different species may be involved producing a carcinogenic environment by metabolic chain reactions. We know of such a bacterial collaboration in anaerobic infections.

Laminar air flow versus barrier nursing in marrow transplant recipients.

Forty-eight patients with acute leukaemia in relapse (n = 14), acute leukaemia in complete remission (n = 19), chronic myeloid leukaemia (n = 8) or severe aplastic anaemia (n = 7) received a marrow transplant. The first 26 patients were nursed in laminar-air-flow plastic isolators while the next 22 patients were treated in barrier nursing rooms. Gnotobiotic parameters and morbidity in the 2 groups are compared. Good decontamination of the gastro-intestinal tract was obtained using either of the 2 isolation techniques. The incidence of bacterial and mycotic infections, as well as the supportive care required by the patients was almost equal in both groups. Our results also suggest that the incidence of graft versus host disease may decrease with efficient decontamination of the patients.

The "clostridial effect" of selective decontamination of the human gut with trimethoprim/sulphamethoxazole in neutropenic patients.

During 59 periods of hospitalisation, 39 patients with either acute myeloid leukemia (22), acute lymphatic leukemia (9), acute undifferentiated leukemia (1), blastic crisis of chronic myeloid leukemia (6) or high-grade malignant non-Hodgkin lymphoma (1) were subjected to aggressive polychemotherapy after selective decontamination of the gut. The patients were given an amphotericin B suspension in a dosage of 1.2 g/day for two days, after which one tablet of trimethoprim/sulphamethoxazole (TMP/SMZ) (160 mg TMP and 800 mg SMZ) t.i.d. was added to prevent endogenous infections by gram-negative aerobic bacteria or moulds and to maintain the "colonisation resistance" endowed by the anaerobes. During 16 of the 59 periods of hospitalisation, no potentially pathogenic aerobic bacteria were isolated. TMP/SMZ-resistant Escherichia coli were the etiological agent of septicemia in two patients, and resistant Klebsiella pneumoniae and Pseudomonas aeruginosa in two other patients. These bacteria were cultured from the patients' fecal samples prior to the development of septicemia. We observed that long-term prophylaxis with TMP/SMZ modified the normal aspect of the fecal biotop culture, not only by suppressing the aerobic gram-negative bacteria, but also by allowing certain clostridia to appear. We differentiated 207 clostridia from the fecal samples of 29 patients and observed a predominance of TMP/SMZ-resistant Clostridium difficile, Clostridium innocuum and Clostridium clostridiiforme. C. difficile was also isolated from the blood culture of a neutropenic patient treated with TMP/SMZ and proved to be very toxic in the Verocell culture.

[Megasphaera elsdenii--occurrence in 2,255 fecal samples of humans, chimpanzees and mice]. / Megasphaera elsdenii--über ihr Vorkommen in 2255 Stuhlproben von Menschen, Schimpansen und Mävsen.

In human gnotobiotic studies the number of cfu/g feces expresses the interspecies-specific relations within this biotop. In our study we tried to culture the resident anaerobe microflora diluting 1 g feces to 10(-6) - 10(-9), and using the cystein-blood-agar (Beerens mod. Werner) together with the dextrose-water-blue agar. M. elsdenii appear as big, smooth and yellow colonies, which may be recognised at "first sight" diagnosis. In a retrospective study of 2255 fecal samples we compared the isolation-rate of M. elsdenii in the range of greater than or equal to 10(7)/g feces from healthy people to that of persons with gastrointestinal troubles. M. elsdenii has not been isolated from fecal samples of healthy people but in 12% from those of adults and 7.4% from children, both suffering from gastrointestinal disorders. So we consider that M. elsdenii in the range of greater than or equal to 10(7)/g feces does not belong to the resident anaerobe flora of the intestine. Further studies have to establish wether its normal range may be 10(3)-10(4)/g feces like that of C. perfringens. Comparing the isolation-rate from patients with colon-carcinoma to that of patients with dermatological affections, we found a relation of 8% to 20%. The biochemical activities of our isolates were according to the Anaerobe Laboratory Manual. Additional tests e.g. API-Zym and others make us assume an activity inner the metabolism of fatty-acids and amino acids e.g. inner the aspartat and pyruvat-family.

[Bone marrow transplantation for aplastic anaemia and acute leukaemia (author's transl)]. / Knochenmarktransplantation bei Panmyelopathie und akuter Leukämie. Resultate des Westdeutschen Tumorzentrums Essen.

Twenty-six bone marrow transplantations were performed at the West Germany Tumour Centre at Essen between December 1975 and December 1981. Three patients had aplastic anaemia, 23 acute leukaemia. Of the leukaemia patients 13 had a recurrence and ten were in full remission at the time of the transplantation. Two of the patients with a recurrence received deep-frozen autologous marrow from the remission phase. One patient with aplastic anaemia had an identical twin as donour, while all other patients received allogeneic marrow from histocompatible family members. Eleven patients are still alive, all three of those with aplastic anaemia, one of those with leukaemia in a recurrence, and seven of those with acute leukaemia in full remission. Two of the survivors have a chronic graft-host reaction, but the others are symptom-free, after an observation period of up to four years. Main causes of death were leukemic recurrence (6), infection (5), and graft-host reaction (2). It would seem that at present bone marrow transplantation offers the best chance of a cure for severe aplastic anaemic and acute leukaemia in adults, but only if transplantation is undertaken early and not in the end-stage of the disease.