Antibody repertoire development in fetal and neonatal piglets. XX. B cell lymphogenesis is absent in the ileal Peyer's patches, their repertoire development is antigen dependent, and they are not required for B cell maintenance

The continuous ileal Peyer's patches (IPP) of sheep are regarded as a type of mammalian bursal equivalent where B cells diversify their repertoire in an Ag-independent fashion. Anatomically and developmentally similar IPP occur in swine. Resection of ～90% of the IPP in piglets at birth did not alter Ig levels in serum and secretions or retard diversification of the Ab repertoire when animals were maintained in isolators and colonized with a defined gut flora. Resection or sham surgery elevated IgG and IgA in serum and in lavage fluid from the gut, lung, and in saliva. No changes in the frequency of IgG-, IgA-, and IgM-containing cells in the spleen and peripheral lymph node were observed. Using an index that quantifies diversification of the VDJ repertoire, no differences were seen in three secondary lymphoid tissues between piglets lacking IPP and colonized controls, whereas both groups displayed >10-fold greater diversification than did late-term fetal piglets or piglets maintained germ-free. Somatic hypermutation was very low in fetal IPP and the IPP of germ-free piglets but increased 3- to 5-fold after colonization. D-J signal joint circles were not recovered in IPP, and V-DJ signal joint circles were 5-fold lower than in bone marrow and similar to those in thymus and spleen. We conclude that the porcine IPP are not a site of B cell lymphogenesis, do not undergo Ag-independent repertoire diversification, and are not primary lymphoid tissue since they are not required for maintenance of Ig levels in serum and secretions.     

JENUFA GEN. NOV.: A NEW GENUS OF COCCOID GREEN ALGAE (CHLOROPHYCEAE,INCERTAE SEDIS) PREVIOUSLY RECORDED BY ENVIRONMENTAL SEQUENCING1

The diversity of eukaryotic microorganisms is far from fully described, as indicated by the vast number of unassigned genotypes retrieved by environmental sequencing or metagenomics. We isolated several strains of unicellular green algae from algal biofilms growing on tree bark in a Southeast Asian tropical rainforest and determined them to be relatives of an unidentified lineage of environmental 18S rDNA sequences, thus uncovering its cellular identity. Light, confocal, and electron microscope observations and sequencing the 18S rRNA gene revealed that the strains represent two different species within an apparently new genus, described here as Jenufa gen. nov. Both species formed minute coccoid cells with an irregular globular outline, a smooth cell wall, and a single parietal chloroplast without a pyrenoid. The two species, described herein as J. perforata and J. minuta, differed in chloroplast morphology and cell wall structure. Phylogenetic analyses of 18S rRNA gene sequences showed a firm relationship between the two species and placed the Jenufa lineage in an unresolved position within the CS clade (Chlamydomonadales + Sphaeropleales) of the class Chlorophyceae, although possible affinities to the genus Golenkinia were suggested both by maximum‐likelihood (ML) and Bayesian methods. Furthermore, two almost identical environmental 18S rDNA sequences from an endolithic microbial community occurring in dolomite rock in the central Alps turned out to be specifically related to, yet apparently distinct from, the sequence of J. minuta, indicating the existence of an undescribed Jenufa species occurring in the temperate zone.     

The use of MLVA for Corynebacterium diphtheriae genotyping--preliminary studies

The complete genome sequence of strain NCTC 13129 C. diphtheriae were investigated in order to identify tandem repeats (VNTR). From 75 VNTR loci identified in the genome 14 were selected. Primers were designed and PCR conditions were optimized for amplification of the selected VNTR markers. Preliminary studies of usefulness of selected VNTR markers were conducted using a group of 28 C. diphtheriae strains. From 14 markers 8 were regarded as potentially useful. The diversity of individual markers ranged from 1 to 6 alleles (Simpson index from 0 to 0,746). No diversity were observed for 3 VNTR markers but it could be a results of too small group of strains analyzed in the tests. Simpson diversity index calculated for all the markers tested on 28 strains was 0,87. Results of the preliminary studies showed usefulness of MLVA for C. diphtheriae genotyping. Nevertheless, confirmation of reliability of the method should be done using a large group of strains. Moreover, the method should be compared with other genotyping methods.     

Evaluation of usefulness of the enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies to lipopolysaccharides of VTEC strains in patients suspected for Escherichia coli O104:H4 infection in Poland

The aim of the study was to investigate the presence of antibodies to lipopolysaccharides obtained by modified Boivin's method from E. coli serotype O104:H4 and O26, O103, O111, O121, O145, O157 in sera of 7 patients with acute diarrhea, suspected in clinical investigation for infection caused by E. coli O104:H4. Additionally, to determine the cut-off levels, the 75 sera from blood donors were tested. The high level of antibodies to LPS E. coli O104 was diagnosed in three patients from family outbreak caused by E. coli serotype O104:H4. In one of those patients, 7-years boy with HUS, we observed also a significant decrease of level of IgA, IgG and IgM antibodies in serum sample obtained in chronic phase of the disease. Furthermore, we showed that two others patients with clinical evidence of VTEC infection, not connected with this family outbreak, had a high level of antibodies to E. coli of other serotypes: one to O157 and one to O103. We did not observe presence of antibodies to LPS from E. coli O26, O111, O121 i O145 in the sera of tested patients. In conclusion, we confirmed that ELISA based on lipopolysaccharides obtained from different serotypes of E. coli may be helpful in laboratory diagnosis of infection caused by VTEC in humans.     

N9-substituted derivatives of kinetin: effective anti-senescence agents

The first isolated cytokinin, 6-furfurylaminopurine (kinetin or Kin), was identified almost 55 years ago. Its biological effects on plant cells and tissues include influences on such processes as gene expression, cell cycle, chloroplast development, chlorophyll biosynthesis, stimulation of vascular development, delay of senescence, and mobilization of nutrients. In the present study we prepared a series of eight N9-substituted Kin derivatives, and characterized them with available physicochemical methods such as CI+ mass spectrometry and ¹H NMR spectroscopy. All compounds were tested in three classical cytokinin bioassays: a tobacco callus assay, an Amaranthus assay, and a senescence assay with excised wheat leaves. The ability of the compounds to interact with Arabidopsis cytokinin receptors CRE1/AHK4 and AHK3 was tested in a bacterial receptor assay. Prepared derivatives with certain substitutions of the N9-atom of the purine moiety enhanced the cytokinin activity of the parent compound in the bioassays to a remarkable degree but negatively affected its perception by CRE1/AHK4 and AHK3. The ability of compounds to delay the senescence of excised wheat leaves in both dark and light conditions, was highly correlated with their ability to influence membrane lipid peroxidation, which is a typical symptom of senescence. Our results were corroborated by gene expression profiling of those genes involved in cytokinin metabolism and perception, plant senescence, and the stress response, and suggest that prepared kinetin derivatives might be used as potent anti-senescence agents.     

Frequency of detection of Ureaplasma urealyticum and Mycoplasma hominis in cervical canal and the Douglas pouch of infertile and fertile women

The group of organisms commonly referred to as genital mycoplasmas comprise species most often found in genitourinary tract of sexually active adults as common commensal inhabitants, or pathogens which can possibly cause many different pathologies like: non-gonococcal urethritis, bacterial vaginosis, cervicitis, endometritis or pelvic inflammatory disease. The problem of their morbidity and the possible influence they have on human fertility is still not clear. The aim of this study was to find out whether two investigated species- Ureaplasma urealyticum and Mycoplasma hominis can be detect more often in a group of infertile women. 74 women participated in the study and were assigned to one of 2 groups of patients: infertile women and fertile women without any sign of genital tract infection. Swabs from the cervical canal of the uterus and the fluid from the Douglas pouch were taken during the gynecological examination and laparoscopic procedure. Two diagnostic methods were used: biochemical method- commercial diagnostic kit- Mycoplasma IST 2 and PCR method. The results showed that Ureaplasma urealyticum and Mycoplasma hominis were detected among both fertile and infertile women with nearly the same frequency, much more often in cervical canal than in the Douglas pouch. Ureaplasma urealyticum was more common pathogen than Mycoplasma hominis in both groups and locations. The achieved results point out that the role of genital mycoplasmas in human infertility is still unclear and require further investigations.