Identification, antifungal susceptibility and scanning electron microscopy of a keratinolytic strain of Rhodotorula mucilaginosa: a primary causative agent of onychomycosis

Onychomycosis is a dermatological problem of high prevalence that mainly affects the hallux toenail. Onychomycosis caused by the yeast Rhodotorula mucilaginosa was identified using colony morphology, light microscopy, urease and carbohydrate metabolism in a 57-year-old immunocompetent patient from Rio de Janeiro, Brazil. High-resolution scanning electron microscopy of nail fragments, processed by a noncoating method, led to the observation with fine detail of the structures of both nail and fungus involved in the infection. Yeasts were mainly found inside grooves in the nail. Budding yeasts presented a spiral pattern of growth and blastoconidia were found in the nail groove region. Keratinase assays and keratin enzymography revealed that this isolate was highly capable of degrading keratin. Antifungal susceptibility tests showed that the fungus was susceptible to low concentrations of amphotericin B and 5-flucytosine and resistant to high concentrations of fluconazole, itraconazole, voriconazole and terbinafine. These findings showed data for the first time concerning the interaction of R. mucilaginosa in toenail infection and suggest that this emerging yeast should also be considered an opportunistic primary causative agent of onychomycosis.     

Microsurgical toenail transfer to the hand

Free nonvascularized toenail grafts have been used to reconstruct congenital or traumatic nail defects of the thumb or finger. Unfortunately, these transfers often result in deformity or atrophy. To avoid these undesirable results, microsurgical free vascularized toenail transfer was performed in 10 patients, 3 for congenital nail absence and 7 for traumatic nail defects. Patient age averaged 17 years (range 2 to 32 years). In contrast with previous reports, the whole big or second toenail complex without pulp was used in reconstruction. All 10 nails were successfully transferred with complete survival. No digits required reexploration. There were no donor- or recipient-site problems. Follow-up averaged 3 years, with a range of 14 months to 5 years and 4 months. Appropriate nail growth occurred in the congenital patients. No atrophy of the nail complex was found as long as sufficient bony support was present (9 of 10 cases). Whole free vascularized toenail transfers for reconstruction of congenital and traumatic nailbed defects achieve excellent aesthetic results while maintaining normal hand function.     

Ingrowing toenails: an evaluation of two treatments.

Most of the procedures used for treating ingrowing toenails cause considerable discomfort and have high failure rates. This study evaluated two methods of treatment: (a) a simple procedure, and (b) angular phenolisation. Patients were seen in a special toenail clinic and were assessed for severity and duration of symptoms. Those with no permanent deformity of the nail fold and with only minor infection were treated by the simple procedure. The nail was nicked and torn down to expose the infected nail fold. The patients were then taught to clean the area, spray it with povidone-iodine dry powder, and pack the nail fold with a twist of cotton-wool. Patients with recurrent or severe ingrowing toenails were treated by angular phenolisation. One hundred patients were treated by the simple procedure and 61 of these had had no recurrence after six months. A total of 280 phenolisations were carried out over 18 months and 272 were successful. The treatments described are simple, effective, and well tolerated and should be considered as alternatives to traditional treatment.     

Correction of pincer-nail deformity using dermal grafting

Pincer-nail syndrome has been described as distortion in the shape of the nails with excessive transverse curvature of the plate that increases from proximal to distal, leading to pinching and loss of soft tissue in the affected digit, resulting in severe pain. Many treatments have been recommended, but an effective long-term method that preserves the nail matrix has not been described. A method of dermal grafting under the nail matrix is described, and the results of treatment of six digits are reported. Five women and one man with an average age of 52 were treated. The affected digit was the thumb in four patients and the great toe in two patients. Follow-up averaged 25 1/2 months. The results were good in all cases with only one side of one nail remaining slightly curved. Pain was relieved in all cases, and complete adherence of the new nail plate occurred. Dermal grafting seems to provide excellent long-term treatment of the pincer-nail deformity with preservation of the nail matrix.     

Human nails as a biomarker of arsenic exposure from well water in Inner Mongolia: comparing atomic fluorescence spectrometry and neutron activation analysis.

Arsenic (As) is found naturally in the geological strata within the Ba Men Region of Inner Mongolia, China. A study was conducted to compare the total As measurements from two analytical techniques: instrumental neutron activation analysis (INAA) and atomic fluorescence spectrometry (AFS), and to verify nails as an exposure biomarker in this population. In 1999, nail and water samples were collected in a pilot study. Fingernails and toenails were pooled from 32 participants and analysed for total As by both INAA and AFS. Mean nail As values were 14.8+/-2.4 and 19.4+/-2.8 microg g-1 (+/-SEM) for INAA and AFS, respectively. Results from these two methods were significantly correlated (r=0.93, p<0.0001). In 2000, a second study was conducted and INAA was used to measure total As in toenails from 314 Ba Men residents. Well water samples were collected from 121 households and analysed by AFS. A significant correlation was observed between toenail and well water As (r=0.84, p<0.0001). Based on the results, INAA was significantly correlated with AFS and proved to be a reliable measure of nail As levels. In this population, toenail samples are a useful internal As exposure biomarker from drinking water sources.     

Invasive external otitis: review of 12 cases.

Invasive external otitis is an infection caused by Pseudomonas aeruginosa that often occurs in elderly people with diabetes. Twelve cases that illustrate the problems associated with the clinical recognition and successful outcome of the condition were reviewed. The patients'' average age was 62.5 years, and they had been ill for an average of 1.8 months before admission to hospital. Predisposing factors included diabetes, swimming in a warm climate and the use of a hearing aid. Radionuclide bone scanning and surgical exploration revealed pathognomonic findings. Initial therapy was often suboptimal: one or more relapses occurred in seven of the patients. All of the patients were cured without relapse after a minimum of 4 weeks of therapy with tobramycin plus an anti-Pseudomonas penicillin. The average duration of the illness was 3.9 months. The outcome in invasive external otitis should be excellent if the condition is diagnosed early and appropriate therapy is instituted.     

Fluconazole Non-susceptible <Emphasis Type="Italic">Cryptococcus neoformans</Emphasis>, Relapsing/Refractory Cryptococcosis and Long-term Use of Liposomal Amphotericin B in an AIDS Patient

The treatment of cryptococcosis is hampered by inefficacy or intolerance to the recommended antifungal agents. A patient diagnosed with AIDS had multiple relapses of cryptococcal infection, which became refractory to antifungal agents during the course of therapy. During the follow-up, the patient developed renal toxicity due to amphotericin B use and non-susceptibility of isolated Cryptococcus neoformans to fluconazole was detected. Thereafter, antifungal treatment was performed exclusively with liposomal amphotericin B, reaching a cumulative dose of 19,180 mg over 46 months. The final relapse of cryptococcosis occurred during the maintenance phase with liposomal formulation in a once-weekly dose. Measurement of the minimum serum concentrations of amphotericin B, determined sequentially before and after this relapse, suggested the importance of monitoring drug levels when the liposomal formulation is used for a long period.     

Human nails as a biomarker of arsenic exposure from well water in Inner Mongolia: comparing atomic fluorescence spectrometry and neutron activation analysis

Abstract

Arsenic (As) is found naturally in the geological strata within the Ba Men Region of Inner Mongolia, China. A study was conducted to compare the total As measurements from two analytical techniques: instrumental neutron activation analysis (INAA) and atomic fluorescence spectrometry (AFS), and to verify nails as an exposure biomarker in this population. In 1999, nail and water samples were collected in a pilot study. Fingernails and toenails were pooled from 32 participants and analysed for total As by both INAA and AFS. Mean nail As values were 14.8±2.4 and 19.4±2.8 µg?g^?1 (±SEM) for INAA and AFS, respectively. Results from these two methods were significantly correlated (r=0.93, p<0.0001). In 2000, a second study was conducted and INAA was used to measure total As in toenails from 314 Ba Men residents. Well water samples were collected from 121 households and analysed by AFS. A significant correlation was observed between toenail and well water As (r=0.84, p<0.0001). Based on the results, INAA was significantly correlated with AFS and proved to be a reliable measure of nail As levels. In this population, toenail samples are a useful internal As exposure biomarker from drinking water sources.     

Short term treatment of dermatophyte onychomycosis with terbinafine.

OBJECTIVE--To evaluate the effect of short term treatment with terbinafine on dermatophytosis. DESIGN--Multicentre, randomised, double blind placebo controlled trial of 250 mg/day terbinafine for 12 weeks in dermatophyte onychomycosis. SETTING--Eight dermatology centres in the United Kingdom. PATIENTS--112 patients (mean age 44, range 19-78), 99 with mycologically proved toenail infections and 13 with fingernail infections, of whom eight were subsequently excluded and 19 failed to complete the study. INTERVENTION--Terbinafine 250 mg daily or placebo for 12 weeks. Follow up for 36 weeks after stopping treatment. MAIN OUTCOME MEASURES--Mycological cure (negative results on microscopy and culture) and clinical cure at the end of follow up, adverse events, and biochemical and haematological variables at monthly intervals during treatment. RESULTS--After follow up 82% (37/45) (95% confidence interval 68% to 92%) mycological cure and 69% clinical cure were recorded for evaluable patients treated with terbinafine for toenail infection and 71% (5/7) (30% to 96%) mycological cure and clinical cure for those treated for fingernail infection. The corresponding values for those treated with placebo were 12% (3% to 31%) mycological cure and no clinical cure for toenail infections and 33% (1% to 91%) mycological cure and no clinical cure for fingernail infections. On an intention to treat basis for toenail infections the figures were 73% (38/52) (58% to 85%) mycological cure for terbinafine compared with 6% (0% to 30%) for placebo (p less than 0.007). Two withdrawals were related to adverse events with terbinafine, and there were no significant abnormal laboratory test results. CONCLUSION--12 weeks'' terbinafine is effective and safe treatment for nail dermatophytosis.     

近端膨胀髓内钉与抗旋髓内钉治疗老年股骨转子间骨折的临床疗效分析

目的:探究近端膨胀髓内钉与近端螺旋刀片抗旋髓内钉治疗老年股骨转子间骨折疗效及躁动发生率比较。方法:病例来源为2012年5月至2013年10月收入我院的老年股骨转子间骨折患者60例,采用随机数字表随机分为两组,膨胀钉组30例,采用近端膨胀髓内钉的手术方法,PFNA组30例,采用近端螺旋刀片抗旋髓内钉的手术方法。手术结束后,对两组患者进行X射线片、手术时间及出血量、髋关节功能的比较,同时对比躁动发生率。结果:X线检查结果显示两组患者经手术后骨折处均复位良好,无并发症及内固定物的脱落及松动,两组比较无显著差异(P0.05);与PFNA组相比,膨胀组患者的手术时间及出血量明显降低,差异具有统计学意义(P0.05);两组患者术后3个月、6个月的Harris评分显示,与PFNA组相比,膨胀钉组的优良率明显升高,髋关节优于PFNA组,差异具有统计学意义(P0.05);膨胀钉组患者躁动率为10.00%低于PFNA组患者躁动率为33.33%,具有统计学意义(P0.05)。结论:可膨胀髓内钉具有手术时间较短、出血量少,更利于骨折的愈合,且躁动发生率较低,对股骨转子间骨折的疗效优于较近端螺旋刀片抗旋髓内钉,对临床有指导意义,值得临床推广。     

Onychomycosis Due to Neoscytalidium Treated with Oral Terbinafine, Ciclopirox Nail Lacquer and Nail Abrasion: A Pilot Study of 25 Patients

Background

Onychomycosis by Neoscytalidium constitutes chronic infection of the nails, and its frequency has increased in recent decades. Currently, no effective standard treatment exists and literature data remain scarce. This work aimed to conduct a pilot project of combined treatment for this infection.

Methods

Thirty patients were divided into three treatment groups: oral terbinafine plus ciclopirox nail lacquer twice a week; ciclopirox nail lacquer twice a week; and ciclopirox nail lacquer 5 days a week, all associated with nail abrasion when required, for 12 months, with 6 months posttreatment follow-up. Clinical and mycological criteria were used for evaluation.

Results

Twenty-five patients completed the study. Significant clinical lesion reduction in disease occurred in all three treatment groups: 21 patients (84 %) entered the study with more than 50 % of diseased nail plate, at the end of treatment, and at 6-month follow-up, 84 and 96 %, respectively, presented less than 25 % nail lesion. Negative microscopy was observed in 36 % of the patients at the end of treatment and in 24 % of the patients at 6-month follow-up. At treatment completion (12 months), culture was negative in 21 patients (84 %) and in 18 (72 %) at follow-up. It was not possible to establish any clinical or mycological statistical differences between groups (p > 0.05). Global medical evaluation upon treatment completion revealed that one patient (4 %) presented complete cure, 8 (32 %) presented partial cure, 16 (64 %) presented therapeutic failure. At the end of follow-up period, 6 patients (24 %) were considered to have recurrence/reinfection.

Conclusions

The results obtained at the 6-month period of follow-up showed marked improvement (96 % of clinical improvement and 72 % of negative culture) of the patients treated for onychomycosis caused by Neoscytalidium in the three tested groups with no statistical differences between them. Multicentric studies with greater number of patients enrolled are necessary to confirm these results.     

甲板甲床部分切除联合苯酚烧灼术治疗嵌甲症的临床研究

目的:探讨甲板甲床部分切除联合苯酚烧灼术治疗嵌甲症的临床疗效。方法:选择武汉市中心医院2004年10月至2013年5月收治的115例148趾拇趾嵌甲进行研究。以2004年10月至2007年12月的患者53例(67趾)作为对照组行单纯甲板甲床部分切除术治疗,2008年1月至2013年5月的患者62例(81趾)作为观察组行甲板甲床部分切除联合苯酚烧灼术治疗。对比两组患者的出血、疼痛缓解和康复时间,了解一年后患者复发情况及满意度。结果:148趾手术伤口均一期愈合,观察组病例术后出血时间、疼痛缓解时间(1.85±0.42天、13.25±2.17小时)较对照组(2.69±0.53天、21.54±2.56小时)低,但术后恢复时间(11.32±2.37天)较对照组(8.93±2.06天)长,差异均有统计学意义(P0.05),观察组和对照组的复发率及总体满意率分别为6.15%、12.5%及97.06%、91.07%,两组差异并无统计学意义(P0.05)。结论:甲板甲床部分切除联合苯酚烧灼术治疗嵌甲症与单纯甲板甲床部分切除术的疗效相当,但可显著减少患者痛苦。     

Relapse of imported Plasmodium vivax malaria is related to primaquine dose: a retrospective study

ABSTRACT: BACKGROUND: Relapsing Plasmodium vivax infection results in significant morbidity for the individual and is a key factor in transmission. Primaquine remains the only licensed drug for prevention of relapse. To minimize relapse rates, treatment guidelines have recently been revised to recommend an increased primaquine dose, aiming to achieve a cumulative dose of [greater than or equal to]6 mg/kg, i.e. [greater than or equal to]420 mg in a 70 kg patient. The aims of this study were to characterize the epidemiology of P. vivax infection imported into Queensland Australia, to determine the rates of relapse, to investigate the use of primaquine therapy, and its efficacy in the prevention of relapse. METHODS: A retrospective study was undertaken of laboratory confirmed P. vivax infection presenting to the two major tertiary hospitals in Queensland, Australia between January 1999 and January 2011. Primaquine dosing was classified as no dose, low dose (<420 mg), high dose ([greater than or equal to]420 mg), or unknown. The dose of primaquine prescribed to patients who subsequently relapsed was compared to patients who did not relapse. RESULTS: Twenty relapses occurred following 151 primary episodes of P. vivax infection (13.2%). Relapses were confirmed among 3/21 (14.2%), 9/50 (18.0%), 1/54 (1.9%)and 7/18 (38.9%) of patients administered no dose, low dose, high dose and unknown primaquine dose respectively. High dose primaquine therapy was associated with significantly lower rates of relapse compared to patients who were prescribed low dose therapy (OR 11.6, 95% CI 1.5- 519, p = 0.005). CONCLUSIONS: Relapse of P. vivax infection is more likely in patients who received low dose primaquine therapy. This study supports the recommendations that high dose primaquine therapy is necessary to minimize relapse of P. vivax.     

Differential DNA methylation in umbilical cord blood of infants exposed to mercury and arsenic in utero

Mercury and arsenic are known developmental toxicants. Prenatal exposures are associated with adverse childhood health outcomes that could be in part mediated by epigenetic alterations that may also contribute to altered immune profiles. In this study, we examined the association between prenatal mercury exposure on both DNA methylation and white blood cell composition of cord blood, and evaluated the interaction with prenatal arsenic exposure. A total of 138 mother-infant pairs with postpartum maternal toenail mercury, prenatal urinary arsenic concentrations, and newborn cord blood were assessed using the Illumina Infinium Methylation450 array. White blood cell composition was inferred from DNA methylation measurements. A doubling in toenail mercury concentration was associated with a 2.5% decrease (95% CI: 5.0%, 1.0%) in the estimated monocyte proportion. An increase of 3.5% (95% CI: 1.0, 7.0) in B-cell proportion was observed for females only. Among the top 100 CpGs associated with toenail mercury levels (ranked on P-value), there was a significant enrichment of loci located in North shore regions of CpG islands (P = 0.049), and the majority of these loci were hypermethylated (85%). Among the top 100 CpGs for the interaction between arsenic and mercury, there was a greater than expected proportion of loci located in CpG islands (P = 0.045) and in South shore regions (P = 0.009) and all of these loci were hypermethylated. This work supports the hypothesis that mercury may be contributing to epigenetic variability and immune cell proportion changes, and suggests that in utero exposure to mercury and arsenic, even at low levels, may interact to impact the epigenome.     

Portable X-ray fluorescence of zinc applied to human toenail clippings

Zinc is an essential trace element in humans. Zinc deficiency can result in a range of serious medical conditions which include effects on growth and development, the immune system, the central nervous system, and the gastrointestinal system. Diagnosis of zinc deficiency is often precluded by the lack of a noninvasive and reliable biomarker. Zinc concentration in nail is considered an emerging biomarker of zinc status in humans. Whether zinc in nail accurately reflects zinc status is beyond the scope of the current study, but is an important research question. The development of a portable method to quickly assess zinc concentration from a single nail clipping could be a useful advance. In this study, single toenail clippings from 60 individuals living in Atlantic Canada were measured for zinc using a portable X-ray fluorescence (XRF) technique. These samples were obtained from the Atlantic PATH cohort, part of the largest chronic disease study ever performed in Canada. Each toenail clipping was measured using three 300 s trials with a mono-energetic portable XRF system. Results were then assessed using two different approaches to the XRF analysis: (1) factory-calibrated zinc concentrations were output from each trial, and (2) energy spectra were analyzed for the characteristic X-rays resulting from zinc. Following the measurement of zinc using the non-destructive portable XRF method, the same clippings were measured for zinc concentration using the “gold standard” technique of inductively coupled plasma-mass spectrometry (ICP-MS). A linear equation of best fit was determined for the relationship between average XRF output zinc concentration and ICP-MS zinc concentration, with a correlation coefficient r = 0.60. Similarly, a linear equation of best fit was found for the relationship between a normalized XRF energy spectrum zinc signal and ICP-MS zinc concentration, with a correlation coefficient r = 0.68. Individual ICP-MS zinc concentrations ranged from 32 μg/g to 140 μg/g, with a population average of 85 μg/g. The results of this study indicate that portable XRF is a sensitive method for the measurement of zinc in a single nail clipping, and provides a reasonable estimation of zinc concentration. Further method development is required before portable XRF be considered a routine alternative to ICP-MS for the assessment of zinc in nail clippings.     

Onychomycosis in Tehran,Iran: Prevailing fungi and treatment with itraconazole

A total of 187 Patients with suspected onychomycosis were examined for causative fungal agents between 1996 and 1997. Laboratory examination confirmed onychomycosis in 115 patients, of which 97 cases were presented with positive microscopic and cultural examinations, and they were selected for itraconazole pulse therapy. From an etiological point of view, 48.4% of the nail infections, mainly toenail infections, were caused by dermatophytes, 43.3% were infected with Candida spp, specially infected fingernails, and 8.2% by non-dermatophytic molds. Trichophyton mentagrophytes var. interdigital and T. violaceum were the most prevalent species. Candida albicans and C. parapsilosis were the predominant species of the Genus Candida. Scopolariopsis brevicaulis was the most common non-dermatophyte molds observed. Female affected more frequently than male and in both sexes, those who were 30–49 years old, more infected. Toenails were affected more frequently than fingernails. In this study, itraconazole pulse therapy (400 mg daily) gave during the first week of per month for 3 months. The study included 51 patients with toenail onychomychosis (group 1) and 46 patients with fingernail infections (group 2). Patients were followed up for 9 months after the last treatment. Clinical response rates were 83% in the group 1, 95% in the group 2 at month 12; the corresponding mycological cure rates were 71 and 87%, respectively. This revised version was published online in June 2006 with corrections to the Cover Date.     

Quantification of manganese and mercury in toenail in vivo using portable X-ray fluorescence (XRF)

Background and objective: Toenail is an advantageous biomarker to assess exposure to metals such as manganese and mercury. Toenail Mn and Hg are in general analyzed by chemical methods such as inductively coupled plasma mass spectrometry and atomic absorption spectrophotometry. In this project, a practical and convenient technology—portable X-ray florescence (XRF)—is studied for the noninvasive in vivo quantification of manganese and mercury in toenail.

Material and methods: The portable XRF method has advantages in that it does not require toenail clipping and it can be done in 3?min, which will greatly benefit human studies involving the assessment of manganese and mercury exposures. This study mainly focused on the methodology development and validation which includes spectral analysis, system calibration, the effect of toenail thickness, and the detection limit of the system. Manganese- and mercury-doped toenail phantoms were made. Calibration lines were established for these measurements.

Results: The results show that the detection limit for manganese is 3.65?μg/g (ppm) and for mercury is 0.55?μg/g (ppm) using 1?mm thick nail phantoms with 10?mm soft tissue underneath.

Discussion and conclusion: We conclude that portable XRF is a valuable and sensitive technology to quantify toenail manganese and mercury in vivo.     

Reconstruction of fingertip amputations with full-thickness perionychial grafts from the retained part and local flaps.

The treatment of fingertip amputations distal to the distal interphalangeal joint when the amputated part is saved is difficult and controversial. Both reattachment of the amputated portion as a composite graft and microvascular anastomosis are prone to failure in this distal location. The authors have evolved a reconstructive plan that uses the nail matrix, perionychium, and hyponychium of the amputated fingertip as a full-thickness graft when the amputation is between the midportion of the nail bed andjust proximal to the eponychial fold. Various flaps are used to lengthen and augment the finger pulp, and skeletal pinning is carried out as necessary. The charts of 15 patients who underwent this procedure over a 38 month period were evaluated retrospectively. Seven returned to the office for examination at least 1 year after the fingertip reconstruction described above; four others were interviewed by telephone. Nail deformity, fingertip sensation, and joint range of motion were evaluated, and the reconstructed fingertips were photographed in standardized views. In six of the seven patients seen in the office, aesthetic and functional results were judged as good by both patient and physician; one of the six had minimal nail curvature. The seventh patient had no nail growth, although finger length was retained and there was no functional disability. The four patients interviewed by phone reported normal fingertip use with no dysesthesias or cold intolerance; all had nail growth, although three patients described slight nail curvature that required care in trimming. The authors favor salvage of all perionychial parts when a distal fingertip amputation occurs. Reconstruction of the fingertip with grafting of the hyponychium, perionychium, and nail matrix from the amputated part combined with local flaps can provide a very satisfactory functional and aesthetic result.     

Therapeutic experience with netilmicin.

Netilmicin, a new aminoglycoside antibiotic, was used to treat 19 patients with urinary tract infection and 5 with systemic infection. The causal organisms were Escherichia coli (in 2), Klebsiella pneumoniae (in 4), Serratia marcescens (in 12) and Pseudomonas aeruginosa (in 7); 1 patient was infected with two of these organisms. All the isolates of causal organisms except one of Serratia were initially sensitive to netilmicin but many were resistant to other aminoglycosides. Sixteen of the urinary tract infections responded to netilmicin therapy, although relapse occurred in three patients. Two of the three patients with musculoskeletal infection responded to combined therapy with surgery and netilmicin; the other patient responded to the same regimen but with carbenicillin added. Netilmicin cured pneumonia in one patient but failed in the other patient with pneumonia, who had leukemia. Superinfection occurred in five patients with urinary tract infection. Adverse reactions to netilmicin were minor. Netilmicin may prove to be a useful agent, particularly for infections due to multiresistant Klebsiella or Serratia, or when prolonged aminoglycoside therapy is required.     

Report of two cases of acute myelogenous leukemia immunized with autologous leukemia-derived hybrid cells

Reports that immunizations with leukemia-derived hybrid cells prolonged the survival of leukemic mice led us to attempt an analogous approach in two adult patients with acute myeloid leukemia (AML). Hybrid cells were prepared from the pretreatment marrows of the newly-diagnosed patients with D98OR cells, in the first case, and with KR12 cells, in the second case. (D98OR and KR12 cells are human cell-lines.) Hybrids formed with KR12 cells expressed HLA antigens of both parental sources and some of the clonal isolates expressed myeloid-associated determinants. The immunizations were performed during the first complete clinical remission; the patients were demonstrably immunocompetent. Positive delayed type hypersensitivity responses to both (X-irradiated) hybrid cells and to (X-irradiated) autologous pretreatment marrow were observed following the immunizations. Mixed lymphocyte reactions toward autologous marrow were positive in one of the patients. In both, relapse occurred approximately two months after the first immunization and eight months after first diagnosis. The first patient remained in complete remission for two and one-half years following reinduction chemotherapy; reinduction chemotherapy was unsuccessful in the second patient.