Seroepidemiology of Leptospira infection in slaughtered cattle in Gauteng province, South Africa.

Leptospirosis is an important economical disease of livestock globally, especially in Asia, the Caribbean, and the African continent. Its presence has been reported in a wide range of livestock. However, information on leptospirosis in South Africa is scanty. We conducted a cross-sectional study in 11 randomly selected abattoirs to determine the seroprevalence and risk factors for leptospirosis in slaughtered cattle in Gauteng province, South Africa. During abattoir visits to selected abattoirs, blood samples were collected from 199 cattle and demographic data obtained on the slaughtered animals. The microscopic agglutination test (MAT) was performed on all sera using a 26-serotype panel using cutoff titer ≥ 1:100. Animal- and abattoir-level risk factors were investigated for their association with seropositivity for leptospirosis. The seroprevalence of leptospirosis in the cattle sampled was 27.6% (55/199). The predominant serogroups detected in seropositive cattle were Sejroe (sv. Hardjo) (38.2%) and Mini sv. Szwajizak) (14.5%) but low to Canicola (sv. Canicola) (1.8%) and Pomona (sv. Pomona) (1.8%). The differences were statistically significant (P < 0.05). Of the five variables investigated, only one (abattoirs) had statistically significantly (P < 0.001) differences in the seroprevalence of leptospirosis among abattoirs. The study documented for the first time in South Africa, the occurrence of serogroups Sejroe (Hardjo bovis strain lely 607), Tarassovi, Hebdomadis, and Medanensis in slaughtered cattle. It was concluded that six of the nine serovars (representing seven serogroups) of Leptospira spp. circulating in cattle population in South Africa are not vaccine serogroups. The clinical, diagnostic, and public health importance of the findings cannot be ignored.

Leptospira in breast tissue and milk of urban Norway rats (Rattus norvegicus).

Leptospirosis is a zoonosis caused by bacteria of the genus Leptospira. The disease is globally distributed and a major public health concern. The Norway rat (Rattus norvegicus) is the main reservoir of the pathogen in urban slums of developing and developed countries. The potential routes of intra-specific leptospire transmission in rats are largely unknown. Herein, we identified pathogenic Leptospira spp. in breast tissue and milk of naturally infected rats. We examined kidney, breast tissue and milk from 24 lactating rats for the presence of leptospires using immunofluorescence, immunohistochemistry, polymerase chain reaction (PCR) and scanning electronic microscopy. All 24 rats had evidence for Leptospira in the kidneys, indicating chronic carriage. The majority of kidney-positive rats had detectable leptospires in milk (18, 75%) and breast tissue (16, 67%), as evidenced by immunofluorescence assay and immunohistochemistry. Four (17%) milk samples and two (8%) breast tissue samples were positive by quantitative real-time PCR. Scanning electron microscopy confirmed the presence of leptospires in breast tissue. No major pathological changes in breast tissue were found. This study, for the first time, identified leptospires in the milk and breast tissue of wild Norway rats, suggesting the possibility of milk-borne transmission of leptospirosis to neonates.

Comparison of hematopoietic and immune recovery after autologous bone marrow or blood stem cell transplants.

We studied hematopoietic and immune recovery in 40 subjects receiving autologous bone marrow (ABMT) or blood stem cell transplants (ABSCT). Supportive care, transplant-related morbidity, duration of hospitalization and cost were also considered. ABSCT was associated with more rapid recovery of all hematopoietic lineages than was ABMT. However, kinetics of immune recovery were similar between the groups. In the ABSCT group, there was a correlation between numbers of blood progenitor cells infused and the rate of hematopoietic recovery. The accelerated hematopoietic recovery following ABSCT correlated with less morbidity, fewer transfusions, briefer hospitalization and lower cost than ABMT.

Comparison of four serum-free, cytokine-free media for analysis of endogenous erythroid colony growth in polycythemia vera and essential thrombocythemia.

INTRODUCTION: The assay of endogenous erythroid colony formation (EEC), a characteristic of polycythemia vera and essential thrombocythemia, is not standardized. In this multicentric study, we tested four semisolid, serum-free, cytokine-free media based on either methylcellulose (M1, M2) or collagen (C1, C2) commercialized for the EEC assay. MATERIALS AND METHODS: Bone marrow mononuclear cells (BMMC) from 73 individuals (62 patients with either polycythemia vera (26), essential thrombocythemia (19), secondary polyglobuly (17) or chronic myeloid leukemia (2) and 11 healthy donors) were grown in parallel in the four media without, or with 0.01 U/ml erythropoietin (EPo). RESULTS: In all four media EEC formation was specific, as it was not observed in cultures of patients with secondary polyglobuly or chronic myeloid leukemia, nor of healthy donors. Analysis of fresh or MGG-stained collagen gel cultures allowed detection of EEC formation significantly more frequently than methylcellulose-based media; addition of 0.01 U/ml of EPo had little or no effect on EEC formation. Collagen-based medium C1 gave better results than the other media tested: the 'C1' EEC assay was positive for 68.2% of polycythemia vera cultures with significantly higher median EEC numbers (6.5/10(5) BMMC for patients with one major criteria of polycythemia vera and 19 and 21/10(5) BMMC for patients with two or three major criteria, respectively). Medium C1 was also better for essential thrombocythemia cultures with 47.4% of positive results but with a low median EEC number (6.7/10(5) BMMC). When associated with the ELISA dosage of serum EPo, the 'C1' EEC assay allowed confirmation or elimination of the diagnosis of polycythemia vera for 91% (20/22) of polyglobulic patients. CONCLUSION: We propose that serum-free collagen-based culture systems be considered to standardize the EEC assay, now part of the new criteria of polycythemia vera.

Comparison of sister-chromatid exchange induced by photoactivated 3-carbethoxypsoralen and 8-methoxypsoralen in human blood lymphocytes.

The induction of sister-chromatid exchange (SCE) by a photoactivated monofunctional derivative of psoralen, 3-carbethoxypsoralen (3-CPs) was compared with that of the bifunctional compound, 8-methoxypsoralen (8-MOP). Lymphocytes were exposed in vitro to a series of equimolar concentrations of the drugs as well as to increasing doses of long-wave ultraviolet light (UVA) and second-division metaphases examined for SCE. The drugs or UVA per se did not influence the incidence of SCE. However, combination of the drug and UVA exposure resulted in a dose-dependent increase in SCE and such elevation was less pronounced with 3-CPs as compared to 8-MOP. This difference between 3-CPs and 8-MOP could be due to the difference in the types of lesions induced/repaired in DNA.

Cloning properties of T lymphocyte subpopulations after treatment with 8-methoxypsoralen and UVA irradiation.

The cloning rate of PHA-stimulated T lymphocytes after treatment with 8-methoxypsoralen plus UVA irradiation described by Wunder and Reischmann (1983) gives a linear dose-effect relationship at low dosages. However, with increasing doses a flattening of the negative gradient occurs. This relationship deviates from the classical exponential curve which can be observed when fibroblasts are treated with mutagens and which is explainable by a 'recovery plateau' at lower dosages. In this study we show that some subpopulations of T lymphocytes, in particular the T-helper and T-suppressor cells, influence the overall dose-effect relationship. These isolated subpopulations exhibit varying sensitivities in comparison with their depleted cell populations. It may be assumed that heterogeneous cell populations exist within each isolated subpopulation which may be separated into further subclasses according to their specific sensitivity.

Further studies on compartmentalisation of DNA-topoisomerase I in Fanconi anemia tissue.

Previously we found increased DNA-topoisomerase I activity in the cytoplasmic fraction of a mature placenta from a child homozygously affected with Fanconi anemia (FA). Now determination of this enzyme was extended to one heterozygous and four homozygous FA fibroblast cultures in confluent stage, and one mature and one immature placenta from homozygously affected children. In all these cases enzyme activity was found in the cytoplasmic fraction in the range between 0.2 and 1.3 U/micrograms protein. For comparison six normal fibroblast cultures, five normal mature and two immature placentae were studied. The cytoplasmic fractions from all but one fibroblast culture, which showed trace activity, showed no detectable activity. The normal mature placentae had no measurable cytoplasmic activity, while the immature trisomy 18 placentae contained 0.1 and 0.27 U/microgram protein. In a homozygous FA fibroblast culture the total cellular distribution was determined; the cytoplasmic fraction contained 6.5% of the whole cellular activity.

Microinjection of normal cell extracts into Fanconi anemia fibroblasts corrects defective scheduled DNA synthesis recovery after 8-methoxypsoralen plus UVa treatment.

Fanconi anemia (FA) cells show an increased sensitivity to 8-methoxypsoralen (8-MOP) plus UVa treatment; after an initial reduction of their semiconservative DNA synthesis rate, they do not recover like normal cells. We microinjected extracts from normal cells into FA fibroblasts from complementation group A and determined semiconservative DNA synthesis rates by autoradiography; the hampered recovery phase was completely restored.

Spontaneous 6-thioguanine-resistant lymphocytes in Fanconi anemia patients and their heterozygous parents.

The incidence of spontaneous 6-thioguanine-resistant (TGr) lymphocytes was studied in the peripheral blood collected from seven Fanconi anemia (FA) patients and five of their heterozygous parents using an autoradiographic or a lymphocyte cloning method. Five of the seven patients showed a significantly elevated incidence of TGr lymphocytes as compared to age- and sex-matched healthy controls. There was, however, no difference between FA heterozygotes and controls. These results suggest some variability among the patients similar to those reported in clinical and cytogenetic investigations. The basis for the increase in TGr cells in the patients is not known, but the inherent genomic instability reflected as increased frequencies of chromosomal aberrations is one possible explanation.

Response of lymphocytes from Fanconi's anemia patients and their heterozygous relatives to 8-methoxy-psoralene in a cloning survival test system.

Five homozygous patients with Fanconi's anemia and nine heterozygous relatives were studied for mutagen sensitivity by a new method. Lymphocyte cloning survival after treatment with several doses of the cross linking substance 8-methoxy-psoralene and exhaustive activation with UVa-light was used for precise measurement of the dose-response. Sensitivity of the homozygous lymphocytes was greatly increased; the degree of the cellular defects corresponded to the clinical course. In the two cases with highest sensitivity, the heterozygotes could be differentiated from normal probands by increased sensitivity, whereas this was not the case in three other families.

Fanconi's anemia: anomaly of enzyme passage through the nuclear membrane? Anomalous intracellular distribution of topoisomerase activity in placental extracts in a case of Fanconi's anemia.

In cells of Fanconi's anemia (FA) spontaneous breakage of chromosomes was first recognized by Schroeder et al. (1964). Sensitivity to bivalent alkylants has been found to be a constant feature, whereas low levels of several repair-related enzymes have been described in different FA cell lines. In a family with known FA, during a further pregnancy the prenatal diagnosis of the disease was made by cytogenetic analysis of amniotic cells. After birth the fresh placenta was extracted for further enzymologic analysis. An unusual distribution of DNA topoisomerase activity was noted: high in the cytoplasm and only a little activity in the nuclear sap. This contrasts with findings in normal placentae. Since amniotic cells, lymphocytes, and fibroblasts of this child exhibited both high spontaneous breakage of chromosomes and sensitivity to the bivalent alkylant, diepoxybutane, a correlation between the findings on cytogenetic and enzymologic levels is assumed. Whereas in other published cases, a true reduction of activities of enzymes involved in DNA replication and repair has been found, the present results suggest the interpretation that in our patient the genetic anomaly does not affect the level of synthesis of the enzyme itself, but the passage of the enzyme from the place of synthesis (the cytoplasm) to the substrate (inside the nucleus). A genetic anomaly of the nuclear membrane might be a possible explanation, or alternatively, a structural mutation of the enzyme at a site not affecting the catalytic activity, but affecting the membrane passage or intranuclear accumulation. Meanwhile, placentae of two other cases gave similar results, thus supporting our findings.

Impeded normal hematopoiesis in bone marrow of patients with multiple myeloma.

Insufficient output of mature blood cells frequently accompanies the typical impairments of late B cell development in multiple myeloma (MM). In a large group of previously untreated patients, bone marrow samples were analyzed and showed a general decrease of mononuclear cell (MNC) content. Colony growth of granulo-monocytic progenitors in short-term assays is compromised in a substantial number of patients at partly severe degrees, who at the same time show higher plasma cell content and belong to clinically more severe groups; the other patients show normal in vitro growth, contain less plasmocytes in the marrow and belong to varying degrees of aggressiveness. Thus a heterogeneity of the disease is emerging on the level of bone marrow cells which matches with high aggressiveness of the disease in one type. It can be speculated that in this type there are different underlying mutational events compared to the others: besides the characteristic changes in B cell differentiation, here the cellular defects have an impact on normal granulo-monocytic (and other) progenitor recruitment, which is absent in the other cases.

[Residents of homes for the aged in Germany: characteristics of social structure and choice of nursing home]. / Altenheimbewohner in Deutschland: Sozialstrukturelle Charakteristika und die Wahl des Heims.

In this paper some important aspects of institutionalization in old age are studied. Compared with old people living in private households, people living in residential care facilities show some sociostructural particularities which are hints at possible factors related to institutionalization. The following analyses of the factors related to institutionalization in old age result in a conformity of objective and subjective reasons: poor health, lack of social network and bad housing conditions in combination with needs for care are essential factors related to institutionalization. In addition, the question of denominational choice of the residential care facility and the question about migration in old age is examined more closely. With regard to the religious community it is amazing that, in spite of secularization, there are major interdenominational differences. A further analysis of regional mobility suggests some sociostructural differences: women and widowed persons for example do not move over far distances (from the private household to the residential care facility), compared to men and married persons.

Anomalous plasma concentrations and impaired secretion of growth factors in Fanconi's anemia.

In Fanconi's anemia, which is known to be an autosomal recessive Mendelian trait with four complementary groups. In addition to stunning phenotypic variation at clinical and cellular levels, aplastic pancytopenia is a common feature. Since either an early block of differentiation in stem cells or their insufficient support by stromal functions could be an underlying factor, levels of stem cell factor (SCF) and cytokines have been measured in blood and in supernatants of monocytes after stimulation with granulocyte-macrophage colony stimulating factor (GM-CSF). In two of three FA patients, no GM-CSF was detectable, and simultaneously SCF was decreased to 8% and 15% of normal values. The combination of low SCF and GM-CSF may be implied in the pathogenesis of marrow aplasia, since comparison with W/Sl mice shows that impairment of the SCF/c-kit function alone has different effects. Also, this explains that treatment with GM-CSF in a recent study enhanced only leukogenesis and not all three lineages. In the third patient, both factors were normal, and here a different mechanism may act. In all three FA patients, interleukin 6 (IL-6) production in stimulated monocytes was decreased, which may hamper immune defense of infections in a nonspecific way.

Positive selection of CD34+ cells: a short review of the immunoadsorption methods currently available for experimental and clinical use. Report on the "2nd European Workshop on stem Cell Methodology," Mulhouse, France, May 3-7, 1993.

At the "2nd European Workshop on Stem Cell Methodology," held in Mulhouse, France, on May 3-7, 1993, part of the meeting was dedicated to the positive selection of CD34+ cells. All devices that are currently in use, or will be available in the near future, were explained and practically demonstrated using human cell populations by scientists involved in their development. In this paper, a review of these methods is given in the form of a short description, together with the data presented in Mulhouse and available from the literature.

Mature accessory cells influence long-term growth of human hematopoietic progenitors on a murine stromal cell feeder layer.

A recently described long-term culture system for early human progenitor cells was established with the murine preadipocyte stromal line FBMD-1 grown in 96-well plates; cobblestone areas formed by inoculated hematopoietic cells are determined in a limiting dilution setting after five weeks' culture. To compare the capacity of cobblestone-area-forming cell (CAFC) formation by bone marrow and leukapheresis products in this system, mononuclear cells (MNC) of both origins were cultured. As related to CD34+ cell content, CAFC yields after five weeks' culture were in the same range in bone marrow and leukapheresis stemming from patients with efficient mobilization of hematopoietic cells. In purified CD34+ cell fractions, the CAFC yield per inoculated cell number was considerably higher than in MNC; however, if the CAFC number was related to the inoculated CD34+ cell number in MNC and after purification, the yield was four to eight times decreased in purified fractions. Addition of the mature cells brought the CAFC yield back up to the numbers obtained in the unseparated MNC fraction. By contrast, slightly more advanced progenitors per CAFC were found in cultures of purified hematopoietic cells from both origins than in whole MNC. The results suggest that mature human accessory cells give noticeable support to recruitment of early progenitors on this feeder but lead to lower yield of GM progenitors.