Fish muscle mercury concentration and bioaccumulation fluctuate year-round - Insights from cyprinid and percid fishes in a humic boreal lake.

Boreal lakes demonstrate pronounced seasonality, where the warm open-water season and subsequent cold and ice-covered season dominate natural cycles. While fish muscle total mercury concentration (mg/kg) [THg] is well documented in open-water summer months, there is limited knowledge on the ice-covered winter and spring mercury dynamics in fish from various foraging and thermal guilds. This year-round study tested how seasonality influences [THg] and its bioaccumulation in three percids, perch (Perca fluviatilis), pikeperch (Sander lucioperca), ruffe (Gymnocephalus cernua), and three cyprinids, roach (Rutilus rutilus), bleak (Alburnus alburnus), and bream (Abramis brama) in deep boreal mesotrophic Lake Pääjärvi, southern Finland. Fish were sampled and [THg] was quantified in the dorsal muscle during four seasons in this humic lake. Bioaccumulation regression slopes (mean ± STD, 0.039 ± 0.030, range 0.013-0.114) between [THg] and fish length were steepest during and after spawning and shallowest during autumn and winter for all species. Fish [THg] was significantly higher in the winter-spring than summer-autumn in all percids, however, not in cyprinids. The lowest [THg] was observed in summer and autumn, likely due to recovery from spring spawning, somatic growth and lipid accumulation. Fish [THg] was best described by multiple regression models (R2adj: 52-76%) which included total length and varying combinations of seasonally changing environmental (water temperature, total carbon, total nitrogen, and oxygen saturation) and biotic factors (gonadosomatic index, and sex) in all species. The seasonal variation in [THg] and bioaccumulation slopes across multiple species suggests a need for standardized sampling seasons in long-term monitoring to avoid any seasonality bias. From the fisheries and fish consumption perspective in seasonally ice-covered lakes, monitoring of both winter-spring and summer-autumn would improve knowledge of [THg] variation in fish muscle.

Effects of a germ-free environment on gut immune regulation and diabetes progression in non-obese diabetic (NOD) mice.

AIMS/HYPOTHESIS: Microbial factors influence the development of diabetes in NOD mice. Studies in germ-free animals have revealed important roles of microbiota in the regulation of Th17 and forkhead box P3 (FOXP3)(+) T regulatory (Treg) activation in the intestine. However, the effects of intestinal microbiota in immune regulation and diabetes development in NOD mice are still poorly understood. METHODS: A colony of germ-free NOD mice was established to evaluate the effects of intestinal microbiota on regulatory immunity in the gut, and on the development of insulitis and diabetes in NOD mice. RESULTS: Diabetes developed in roughly equal numbers in germ-free and specific pathogen-free NOD mice. Insulitis was accentuated in germ-free NOD mice; yet insulin preservation was unaltered. Germ-free NOD mice showed increased levels of Il17 (also known as Il17a) mRNA in the colon, and of Th17 and Th1 cells in the mesenteric and pancreatic lymph nodes, while Foxp3 mRNA and FOXP3(+) Tregs were reduced. In the islet infiltrates, FOXP3(+)CD4(+) T cells were slightly increased in germ-free mice. B cells appeared less activated in the peritoneum and were less abundant in islet infiltrates. CONCLUSIONS/INTERPRETATION: These results indicate that lack of intestinal microbiota promotes an imbalance between Th1, Th17 and Treg differentiation in the intestine. This imbalance is associated with accelerated insulitis, but intact recruitment of FOXP3(+) Tregs into islets, suggesting: (1) a microbial dependence of local induction of Treg in the gut and draining lymph nodes; but (2) a potentially compensatory function of naturally occurring Tregs in the islets, which may help control diabetogenic T cells.

Patient-reported complications associated with Campylobacter jejuni infection.

This study aimed to investigate the occurrence of complications, especially musculoskeletal symptoms, after sporadic Campylobacter jejuni enteritis of domestic origin in Finland. This multi-centre cross-sectional study was conducted during a seasonal peak in 2002. Questionnaires were sent to Campylobacter-positive patients, representing different geographical areas, 2 months after collection of positive stool samples. Medical records were viewed in several cases. Besides antimicrobial susceptibility testing C. jejuni isolates were serotyped. A total of 235 patients (58%) returned the questionnaire and 201 C. jejuni-positive patients were finally included in the study. Musculoskeletal symptoms associated with C. jejuni enteritis were frequent (39%); joint pain was most commonly reported (81%). The incidence of reactive arthritis was 4% and that of Achilles enthesopathy and/or heel pain was 9%. Stomach ache during enteritis was associated with the later development of joint pain. Antimicrobial treatment was common but did not prevent complications.

Induced sputum in the diagnosis of childhood community-acquired pneumonia.

BACKGROUND: The usefulness of induced sputum in searching for causative agents of pneumonia in children has not been studied. METHODS: The study involved 101 children, aged 6 months to 15 years, treated for community-acquired pneumonia at Turku University Hospital (Turku, Finland) from January 2006 to April 2007. Nasopharyngeal aspirate samples were first collected through both nostrils. Sputum production was then induced by inhalation of 5.0% hypertonic saline for 5-10 min and a sputum sample was either aspirated or expectorated. The presence and amount of bacteria and viruses in paired nasopharyngeal aspirate and sputum specimens was analysed and compared using semiquantitative bacterial culture and quantitative PCR techniques. RESULTS: A good quality sputum specimen was obtained from 76 children. The possible causative agent was found in 90% of cases. Streptococcus pneumoniae (46%) and rhinovirus (29%) were the most common microbes detected. Newly discovered viruses human bocavirus and human metapneumovirus were detected in 18% and 13% of the children, respectively. One-quarter of all bacterial findings were only detected in sputum, and the amount of bacteria in the remainder of the sputum specimens compared with nasopharyngeal aspirate was higher in 14% and equal in 70%. The amount of rhinovirus in sputum was higher than in nasopharyngeal aspirate in 82%. CONCLUSIONS: Sputum induction provides good quality sputum specimens with high microbiological yield in children with community-acquired pneumonia. Induced sputum analysis can be useful in the microbiological diagnosis of childhood community-acquired pneumonia.

Eradication of methicillin-resistant Staphylococcus aureus from a health center ward and associated nursing home.

BACKGROUND: Long-term health care facilities have been recognized as reservoirs of multiresistant bacterial strains, especially methicillin-resistant Staphylococcus aureus (MRSA). Efforts to control MRSA in this setting usually have been only partially effective. We describe herein the eradication of epidemic MRSA from a Finnish health care center ward and affiliated nursing home. METHODS: The methods to control MRSA included (1) contact isolation precautions, (2) screening for asymptomatic carriage, (3) eradication of carriage, and (4) education of staff on hygienic measures. The first 6 patients with MRSA-positive findings were referred without delay to the Infectious Diseases Unit of the adjacent university hospital for eradication treatment. Later, an isolation unit of 6 rooms was founded in the health care center, where the MRSA-colonized patients were nursed as a separate cohort until they, in succession, were referred to the Infectious Diseases Unit for decolonization. RESULTS: From May 20 through August 17, 1993, the epidemic MRSA strain was isolated from 8 long-term patients on the 40-bed ward of the health care center, 4 of the 59 residents of the nursing home, and 1 member of the staff. Eradication of carriage was successful in all except 1 patient with dementia, who was nursed in contact isolation in the health care center until his death 21 months later. CONCLUSIONS: It is possible to eradicate MRSA from a long-term health care facility even after 13 cases by applying strict control measures. Our experience may be valuable in the future decision-making process for control of new and more challenging multiresistant bacteria, eg, vancomycin-resistant strains of MRSA.

Spectrum of enteropathogens detected by the FilmArray GI Panel in a multicentre study of community-acquired gastroenteritis.

The European, multicentre, quarterly point-prevalence study of community-acquired diarrhoea (EUCODI) analysed stool samples received at ten participating clinical microbiology laboratories (Austria, Finland, France, Germany, Greece, Ireland, Italy, Portugal, Romania, and the UK) in 2014. On four specified days, each local laboratory submitted samples from ≤20 consecutive patients to the Austrian Study Centre for further testing with the FilmArray GI Panel (BioFire Diagnostics, Salt Lake City, UT, USA). Of the 709 samples from as many patients received, 325 (45.8%) tested negative, 268 (37.8%) yielded only one organism, and 116 (16.4%) yielded multiple organisms. Positivity rates ranged from 41% (30 of 73 samples) in France to 74% (59 of 80 samples) in Romania. With the exception of Entamoeba histolytica and Vibrio cholerae, all of the 22 targeted pathogens were detected at least once. Enteropathogenic Escherichia coli, Campylobacter species, toxigenic Clostridium difficile, enteroaggregative E. coli, norovirus and enterotoxigenic E. coli were the six most commonly detected pathogens. When tested according to local protocols, seven of 128 positive samples (5.5%) yielded multiple organisms. Overall, the FilmArray GI Panel detected at least one organism in 54.2% (384/709) of the samples, as compared with 18.1% (128/709) when testing was performed with conventional techniques locally. This underlines the considerable potential of multiplex PCR to improve routine stool diagnostics in community-acquired diarrhoea. Classic culture methods directed at the isolation of specific pathogens are increasingly becoming second-line tools, being deployed when rapid molecular tests give positive results. This optimizes the yield from stool examinations and dramatically improves the timeliness of diagnosis.

The effect of different antibody affinities on ELISA absorbance and titer.

Twenty-one rabbit serum samples were assayed for IgG antibodies against 3-iodo-5-nitrophenyl-epsilon-amino-n-caproic acid (NIP-cap) by equilibrium dialysis and enzyme-linked immunosorbent assay (ELISA). The results of ELISA, expressed either as absorbances or titers, did not correlate to antibody concentration or average affinity measured by the equilibrium dialysis. Affinity distributions of the antibodies within each sample were determined by equilibrium dialysis in 30 antigen concentrations. Thus, it was possible to study the correlation of ELISA results with separate ranges of antibody affinities. ELISA absorbances measured at a low sample dilution (1:80) correlated with high affinity antibodies (affinity greater than 8 X 10(-6) M-1). Use of high antigen density increased this correlation. End-point titers obtained at a low cut-off absorbance level tended to correlate to a larger range of affinities (affinity greater than 6 X 10(4) M-1). ELISA measures antibodies of different affinities depending on how the results are expressed. This can be utilized in qualitative characterization of the measured antibodies.

Lymphoid cell chimerism and transplantation tolerance induced by bursal and postbursal cells.

Transplantation of allogeneic bursal cells into cyclophosphamide-treated, immunodeficient chickens is a useful experimental model for analyzing the mechanisms of transplantation tolerance, especially because transplanted bursal cells do not produce graft-versus-host disease. In this study we have determined B-lymphoid chimerism in various lymphoid organs after transplantation of allogeneic bursal stem cells or postbursal cells, and used a variety of tests to determine presence of immunological tolerance. Transplanted bursal stem cells induced a state of stable chimerism that could easily be detected in peripheral blood and other lymphoid organs. Chimerism induced by postbursal cells was low in peripheral blood, but clearly observable in other lymphoid organs, especially in spleen and thymus. Both bursal and postbursal cells induced specific unresponsiveness to donor-line alloantigens. Bursal cell recipients accepted donor line skin grafts--and their graft-versus-host reactivity, as assayed by embryonal splenomegaly, and mixed lymphocyte reactivity against donor line alloantigens were significantly decreased. Despite differences in chimerism, a strong transplantation tolerance was readily induced with bursal stem cells and with postbursal cells.

T cell function in chickens bursectomized at 60 hours of incubation.

Chickens surgically bursectomized in ovo (Bx) at 60 hr of embryonic development offer a unique model to study selectively the influence of the bursa of Fabricius on thymus-dependent immune functions because the lymphoid cells of these animals develop in the total absence of the bursal microenvironment. The Bx chickens have been shown to be unable to respond to antigenic stimulation by specific antibody production. In the present study, we have characterized different aspects of T-cell-mediated immunity in Bx chickens. Our results indicate the existence of a normal, functional T cell system in these animals. Peripheral blood leukocytes from Bx chickens were able to induce normal graft-versus-host reaction and mixed lymphocyte reaction. The capacity of peripheral blood T cells from Bx animals to produce interleukin-2 was indistinguishable from that of their normal counterparts, suggesting normal function of T helper cells. We demonstrate that peripheral blood leukocytes from Bx birds have normal in vitro proliferative responses to nonspecific T cell mitogens, concanavalin A, and phytohemagglutinin, and to a specific antigen, keyhole limpet hemocyanin. The inability of the Bx chickens to respond to specific antigens is therefore restricted to B cells and to production of specific antibodies. Our findings indicate that the bursa of Fabricius is not necessary for the development of thymus-dependent immune functions and support the suggestion that the specific function of the bursa is the creation of antibody diversity.

Rat spermatogenesis in vitro traced by quantitative flow cytometry.

In vitro differentiation of germ cells in rat seminiferous tubule segments at stages II-III of the epithelial cycle was studied. DNA flow cytometry was used for quantitation of absolute cell numbers from the cultured tubule segments that were compared to freshly isolated stages of the cycle, as identified by transillumination stereomicroscopy of the seminiferous tubules and phase-contrast microscopy of live cell squashes. Spermatogonia and spermatocytes from stages II-III showed normal morphological differentiation during 7 days in vitro. Round spermatids differentiated to Step 7 of spermiogenesis but Step 16 spermatids failed to develop. Acid phosphatase activity in the spermatogenic cells changed normally during the culture. As compared with freshly isolated control tubule segments, 35% of round spermatids and 42% of pachytene spermatocytes were present in culture after 7 days. The cell numbers recovered from defined stages by DNA flow cytometry were close to those found in morphometric studies. Flow cytometry is an efficient quantitation method for cells liberated from seminiferous epithelium. Spermatogonia, spermatocytes, and early spermatids are able to differentiate in vitro, but spermatids approaching the elongation (acrosome) phase, and particularly the maturation phase, fail to differentiate under present culture conditions.

Regulation of stages VI and VIII of the rat seminiferous epithelial cycle in vitro.

Plasminogen activator (PA) is secreted cyclically (in stages VII and VIII) by rat seminiferous tubules. To investigate whether this can be maintained and influenced in vitro, tubule segments from stages VI and VIII of the epithelial cycle were cultured for 3 days in chemically defined medium supplemented with testosterone, FSH, or a combination of testosterone, FSH, insulin and retinoic acid (4F). Morphological and flow cytometric analyses of stage VI tubules suggested a roughly normal differentiation to stage VIII. They developed an increased PA secretion on day 3 of culture. Stage VIII tubules, however, did not develop all the characteristics of stage XII. Step 8 spermatids did not elongate and step 19 spermatids failed to develop into spermatozoa. Secretion of PA on day 3 was not significantly different to that on day 1. The 4F combination very significantly stimulated PA secretion in both stages, but FSH alone was effective only in stage VIII. Most of the secreted PA had a molecular weight of 43,000 in both stages, suggesting that it is of urokinase type. The results suggest that stage VI is more able to differentiate in vitro for 3 days than stage VIII; the cyclic secretion pattern of PA was partially maintained in tubule segments from stage VI. Follicle-stimulating hormone had an effect on PA secretion only in stage VIII, whereas the 4F combination was stimulatory in both stages. The retinoic acid in this combination may be of importance in the regulation of PA secretion by seminiferous tubules.

Uracil-DNA glycosylase activity in chronic lymphoproliferative disorders.

The activity of uracil-DNA glycosylase, a repair enzyme for the excision of uracil from DNA, was studied in patients with chronic lymphoproliferative disorders and with malignant plasma cell dyscrasias. The biochemical assay was performed on mononuclear cells, isolated by density gradient centrifugation from peripheral blood, from bone marrow or from both. The activity of the uracil-DNA glycosylase of peripheral blood cells in 8/8 cases of myeloma and in 3/3 cases of Waldenström's macroglobulinemia was in the same range as in 22 non-hematological control patients, i.e. 2.4-25.1 U/mg of protein. Higher activities were found in 9/12 cases of chronic lymphocytic leukemia (CLL), in 2/4 cases of hairy cell leukemia (HCL), in 2/2 cases of chronic T-cell lymphocytosis and in the only case of small cell lymphocytic lymphoma. Follow-up of some CLL and HCL patients revealed that uracil-DNA glycosylase activity was fairly stable during the course of the disease. We conclude that malignant cells in chronic lymphoproliferative disorders are characterized by a normal or even increased capability to repair DNA, as exemplified by uracil-DNA glycosylase in this study.

Uracil-DNA glycosylase activity in human acute leukemia.

The expression of the DNA excision repair enzyme uracil-DNA glycosylase was investigated in bone marrow and peripheral samples from seven patients with acute lymphoblastic leukemia (ALL), from 17 patients with acute non-lymphocytic leukemia (ANLL), and from one patient with chronic granulocytic leukemia (CGL) in blast crisis. In addition, uracil-DNA glycosylase activities were determined in nine human leukemia/lymphoma cell lines. There was a clear correlation between the percentage of blast cells and the enzyme activity when mononuclear cell fractions from patient samples were analysed. The following uracil-DNA glycosylase activities were recorded (mean +/- S.D., number of samples): ALL = 45.6 +/- 14.8 U/mg of protein, N = 10; ANLL = 41.1 +/- 13.8 U/mg of protein, N = 22; CGL (blast crisis) = 44.7 U/mg of protein. The uracil-DNA glycosylase activity in nine human leukemia/lymphoma cell lines ranged from 35.2 to 66.0 U/mg of protein, and no striking differences were observed between the T-ALL, B-ALL, null cell ALL or myeloid lines. Similarly, the various biological features, such as the common ALL surface antigen, the terminal deoxynucleotidyl transferase enzyme, the sub-type of leukemia, chromosomal aberrations, or previous chemotherapy, did not apparently affect the expression of uracil-DNA glycosylase. We propose that the integrity of the genetic information is well protected by uracil-DNA glycosylase in different forms of leukemia, including cases with a low proportion of S-phase blasts, as assessed by flow cytometry in the present work. When compared to the activities in benign hematopoietic progenitor cells, studied previously in this laboratory, no big differences between the benign and malignant hematopoiesis were demonstrated. Hence, it is unlikely that selectivity of chemotherapy towards malignant vs benign hematopoietic growth could be based on the enzyme uracil-DNA glycosylase.

DNA aneuploidy in anaplastic carcinoma of the thyroid gland.

Nuclear DNA content of 19 cases of large cell or spindle cell anaplastic carcinomas of the thyroid was studied by flow cytometry. In 17 cases archival paraffin-embedded tissue and in 2 cases fresh material obtained by fine-needle aspiration biopsy was used for DNA content determination. Unequivocal evidence of DNA aneuploidy was obtained in 13 cases (68%). Two tumors contained more than one aneuploid stemline of cells. Eighteen (95%) of the patients died within 20 months after diagnosis, but 1 patient has survived for 13 years. The observed survival rate of the patients with carcinoma with a DNA index (DI) less than 1.30 was superior as compared with tumors with a DI greater than 1.30 (P = 0.007). All 7 patients with tumors with DIs greater than 1.30 died within seven months, whereas 5 of the 11 patients with tumors with DIs less than 1.30 survived longer than one year after the diagnosis.

Pure and mixed mucinous breast carcinomas: DNA stemline and prognosis.

The DNA stemline of 45 mucinous breast carcinomas was determined by flow cytometry using paraffin embedded archival tissue sections. The material consisted of 26 pure mucinous and 19 mixed mucinous carcinomas. The patients were followed up for at least 15 years or until death. Nearly all pure mucinous carcinomas had a normal DNA stemline (25 of 26) with only one aneuploid tumour. Mixed mucinous carcinomas had a DNA content resembling that of common ductal carcinoma with 11 aneuploid tumours. Aneuploid tumours tended to be of higher grade and stage than diploid tumours. The survival of patients with pure mucinous carcinoma was better than that of patients with mixed mucinous carcinoma. Mucinous carcinoma should be classified as such only if it is a pure mucinous carcinoma.

Determination of callus quantity in 4-week-old fractures of the rat tibia.

In clinical practice the quantity of fracture callus is usually estimated on the basis of radiographs. In experimental studies on fracture healing, more exact chemical measurements (total protein, DNA, and mineral contents) have been used to quantitate the amount of callus. It is not known, however, how these two parameters correlate with each other. In this study we used different (radiographic, histomorphometric, and chemical) methods to evaluate the quantity of fracture callus in 4-week-old tibial fractures of rats stabilized by intramedullary nailing. The wet and dry weights and the nitrogen, collagen, calcium, phosphorus, and DNA contents of the calluses showed no correlation with the radiographic size of the calluses. There was a strong correlation (p less than 0.001), however, between radiographic and histomorphometric callus sizes. The mass of callus and the chemically defined parameters of callus production thus indicate different properties of the healing bone.

Electrostimulation of rat callus cells and human lymphocytes in vitro.

Asymmetrical pulsing low voltage current was supplied via electrodes to cultured rat fracture callus cells and human peripheral blood lymphocytes. The [3H]thymidine incorporation of the callus cells and 5-[125I]iodo-2'-deoxyuridine incorporation of the lymphocytes were determined. The growth pattern of callus cells (estimated by cellular density) did not respond to electrical stimulation. However, the uptake of [3H]thymidine was increased at the early phase of cell proliferation and inhibited at later phases of proliferation. The [3H]thymidine uptake of confluent callus cell cultures did not respond to electrical stimulation. Lymphocytes reacted in a similar way; stimulated cells took up more DNA precursor than control cells at the early phase of stimulation. During cell division, induced by the mitogens phytohemagglutinin and Concanavalin-A, the uptake of DNA precursor by stimulated cells was constantly inhibited. The results suggest that electrical stimuli affect the uptake mechanisms of cell membranes. The duality of the effect seems to be dependent on the cell cycle.

Eradication of the long-term carriage of methicillin-resistant Staphylococcus aureus in patients wearing dentures: a follow-up of 10 patients.

To cure the long-term carriage of methicillin-resistant Staphylococcus aureus (MRSA), eradication treatment was given to 10 patients wearing complete dentures. In addition to multiple body sites, MRSA was cultured from the dentures of six patients. The contaminated dentures were rebased and sterilized with heat in order to prevent recolonization. The patients did not use their dentures during the decolonization therapy. Subsequently, MRSA was eradicated from three of these patients but three others remained MRSA-positive despite at least two courses of combined systemic and topical eradication treatment. These particular patients had persistent stomatitis and their dentures were a poor fit, in poor condition and repeatedly grew MRSA. Eradication treatment was successful in the remaining four patients whose dentures were MRSA-negative. These results confirm that dentures may function as foreign bodies and sustain persistent nasopharyngeal. MRSA colonization. Therefore, we suggest that whenever eradication of MRSA is deemed necessary in cases of nasal, oral or pharyngeal carriage, heat treatment of the dentures should be included. Further comparative studies with larger patient populations are needed to evaluate the contribution of dentures to the long-term carriage of MRSA, as well as to assess the value of denture sterilization during the eradication course.

Flow cytometric DNA analysis of defined stages of rat seminiferous epithelial cycle during in vitro differentiation.

The stages of the rat seminiferous epithelial cycle have been isolated for flow cytometric analysis of DNA and for culture, using transillumination-assisted microdissection. Precise stages have been identified by phase contrast microscopy of live cell squashes from adjacent segments. Each stage of the cycle showed a characteristic flow cytometric pattern with haploid (1C), diploid (2C) and tetraploid (4C) peaks. Stages I to VIII of the cycle showed an additional hypofluorescent (0.25-0.70C) peak due to a reduced dye-binding capacity of maturation phase-spermatids at steps 15 through 19. The appearance of the hypofluorescent haploid peak coincided with the second nucleoprotein transition at step 15 of spermiogenesis and the homogeneous condensation of the chromatin seen in electron microscopy. As a concomitant of the formation of disulphide bonds during epididymal maturation, the fluorescence intensity decreased further to reach a relative value of 0.07C in the cauda epididymidis. The constant 1C peak was raised by round and elongating spermatids (steps 1-14), 2C by spermatogonia, secondary spermatocytes and Sertoli cells, and the 4C peak by primary spermatocytes and spermatogonia at G2 or M phase of the mitotic cycle. The proportion of each peak accurately reflected the relative proportion of cells in most stages of the cycle when compared with morphometric measurements of histologic preparations. DNA flow cytometry is a suitable method for quantitative evaluation of cultured seminiferous tubule segment DNA. Although the relative yield of the meiotic reductive divisions in vitro is comparable with that observed in vivo, steps 9 and 15 of spermiogenesis involving nucleoprotein transitions and spermiation itself did not occur under the present culture conditions.

Mutagen effects on rat seminiferous tubules in vitro: induction of meiotic micronuclei by adriamycin.

Mutagen effect on male germ cells can be analyzed by micronucleus induction during meiotic divisions. These can be followed in vitro by culturing seminiferous tubular segments from stages of the epithelial cycle that contain late pachytene and diakinetic primary spermatocytes. We studied the formation of micronuclei in this test system using adriamycin as a model mutagen. Micronuclei were induced in a dose-dependent manner at concentrations of 1-10 ng/ml that were far below the dose that caused morphologically or biochemically detectable cytotoxic effects. The meiotic micronucleus induction in vitro is a potentially sensitive test system of male germ cell mutagenesis.