Unusual evolutionary conservation of 5S rRNA pseudogenes inAspergillus nidulans: Similarity of the DNA sequence associated with the pseudogenes with the mouse immunoglobulin switch region

Summary AllAspergillus nidulans 5S rRNA pseudogenes known so far are the result of integration of an approx. 0.2-kbp-long DNA sequence into the 5S rRNA genes. This sequence, called block C, is present in at least five copies in theA. nidulans genome and seems to be associated either with 5S rRNA genes or pseudogenes. In contrast to the 78% sequence conservation of the C-block in pseudogenes, the truncated 5 halves of the pseudogenes are very highly conserved (96.9–100%). We postulate that the 5S rRNA pseudogenes are still a subject of concerted evolution. The C-block sequence shows similarity to the switch region of the mouse heavy chain immunoglobulin gene. A characteristic motif GGGTGAG is repeated several times in both sequences; the sequence conservation is 63%.     

Behavior of sperm nuclei in intact and bisected metaphase II mouse oocytes fertilized in the presence of colcemid

Our objective was to examine the developmental fate of sperm nuclei in oocytes fertilized under conditions of meiotic arrest. Therefore zona-free metaphase II oocytes and oocyte fragments (nucleate and anucleate) were fertilized in the presence of colcemid. In anucleate oocyte fragments, normal male pronuclei develop. In contrast, in intact oocytes and nucleate fragments sperm nuclei after initial decondensation undergo secondary condensation. This state is maintained as long as the oocytes are treated with colcemid. When the drug is removed 3 h after insemination, the meiotic spindle(s) is reconstructed, the second polar body(ies) is extruded, and a female pronucleus (or micronuclei) forms. At the same time the sperm nucleus decondenses again and transforms into a male pronucleus. In addition oocytes fertilized in the presence of colcemid could not be refertilized. These observations suggest that oocytes and oocyte fragments fertilized in the presence of colcemid undergo activation despite the failure of pronucleus formation. The inhibitory effect of colcemid on the formation of pronuclei is expressed only in the presence of oocyte chromosomes. We suggest that colcemid stabilizes factors responsible for chromosome condensation that are associated with oocyte chromosomes but not factors (whether the same or different) present in the cytoplasm.     

Transformation of sperm nuclei into male pronuclei in nucleate and anucleate fragments of parthenogenetic mouse eggs

Our objective was to examine the ability of nucleate and anucleate fragments of artificially activated mouse eggs to transform sperm nucleus into male pronucleus. To this end, zona-free oocytes in metaphase II were activated by ethanol and bisected into halves (one with the spindle, the other anucleate) either within 10 to 20 min (series A) or 3 or 5 hr later (series B). In series A, the fragments were inseminated 3,5, and 8 h after activation, and in series B. 3 and 5 h after activation. Both nucleate and anucleate fragments lose the capability of transforming sperm nucleus into fully formed pronucleus sometime between 3 and 5 h after activation. In 8 h old parthenogenetic fragments, the majority of sperm nuclei remain unchanged or begin decondensation but never reach the stage of an early pronucleus. In over 1/3 of anucleate fragments of this age group, sperm nuclei develop defectively: chromatin decondenses inside the persisting nuclear envelope. In other experimental groups, the incidence of these abnormal sperm nuclei varies between 0 and 10%. In general, the anuclcate fragments retain the capability to transform sperm nuclei (fully or partially) longer than their nuclear counterparts. This difference may be accounted for by a different level of substances required for pronuclcar growth (extrachromosomal constituents of the germinal vesicle and nuclear lamins): high and constant in the cytoplasm of anucleate egg halves and low and progressively decreasing in the nucleate halves because of their putative uptake by the female pronucleus. However, the cytoplasmic factors responsible for the initial stages of transformation (nuclear envelope breakdown, chromatin decondensation) become eventually inactivated both in the presence and in the absence of a female pronucleus.     

Fish biodiversity of Saint Peter and Saint Paulʼs Archipelago,Mid-Atlantic Ridge,Brazil: new records and a species database

Saint Peter and Saint Paul's Archipelago (SPSPA), one of the smallest and most isolated island groups in the world, is situated on the Mid-Atlantic Ridge, between Brazil and the African continent. SPSPA has low species richness and high endemism; nonetheless, the diversity of fishes from deep habitats (>30 m depth) had not been previously studied in detail. Several expeditions conducted between 2009 and 2018 explored the shallow and deep reefs of SPSPA using scuba, closed-circuit rebreathers, manned submersibles, baited remote underwater stereo-videos (stereo-BRUV) and fishing between 0 and 1050 m depth. These expeditions yielded 41 new records of fishes for SPSPA: 9 in open waters, 9 in shallow waters (0–30 m), 8 in mesophotic ecosystems (30–150 m) and 15 in deeper reefs (>150 m). Combined with literature records of adult pelagic, shallow and deep-reef species, as well as larvae, the database of the fish biodiversity for SPSPA currently comprises 225 species (169 recorded as adult fishes and 79 as larvae, with 23 species found in both stages). Most of them (112) are pelagic, 86 are reef-associated species and 27 are deep-water specialists. Species accumulation curves show that the number of fish species has not yet reached an asymptote. Whereas the number of species recorded in SPSPA is similar to that in other oceanic islands in the Atlantic Ocean, the proportion of shorefishes is relatively lower, and the endemism level is the third highest in the Atlantic. Twenty-nine species are listed as threatened with extinction. Observations confirm the paucity of top predators on shallow rocky reefs of the island, despite the presence of several pelagic shark species around SPSPA. Because all of the endemic species are reef associated, it is argued that the new marine-protected areas created by the Brazilian government do not ensure the protection and recovery of SPSPA's biodiversity because they allow exploitation of the most vulnerable species around the archipelago itself. This study suggests a ban on reef fish exploitation inside an area delimited by the 1000 m isobath around the islands (where all known endemics are concentrated) as the main conservation strategy to be included in the SPSPA management plan being prepared by the Brazilian government.     

A panel of recombinant proteins for the serodiagnosis of caseous lymphadenitis in goats and sheep

Caseous lymphadenitis (CLA) is a small ruminant disease characterized by the development of granulomatous lesions in superficial and internal lymph nodes, as well as in some organs, and causes significant economic losses worldwide. The aetiological agent of CLA is the bacterium Corynebacterium pseudotuberculosis; however, the commercially available diagnostic tools present problems with regard to specificity, which can lead to false-negative results. This study aimed to develop an indirect enzyme-linked immunosorbent assay (ELISA) for the detection of specific immunoglobulins in goats and sheep using recombinant C. pseudotuberculosis PLD, CP40, PknG, DtxR and Grx proteins. For validation of the ELISAs, 130 goat serum samples and 160 sheep serum samples were used. The best ELISA for goats was developed using a combination of PLD and CP40 as antigens at a 1:1 ratio, which presented 96.9% sensitivity and 98.4% specificity. The most effective ELISA for sheep presented 91% sensitivity and 98.7% specificity when recombinant PLD alone was used as the antigen. These ELISAs can be used as highly accurate tools in epidemiological surveys and for the serodiagnosis of C. pseudotuberculosis infection in goats and sheep.     

Molecular characterization of Mycobacterium tuberculosis isolates in a region of Brazil with a high incidence of tuberculosis

One hundred and seventy Mycobacterium tuberculosis clinical isolates were characterized by spoligotyping to evaluate the biodiversity of tubercle bacilli in a region of Brazil with a high incidence of tuberculosis (Pelotas and Rio Grande cities - Rio Grande do Sul State). The spoligotyping results were compared to the World Spoligotyping Database (Institut Pasteur de Guadeloupe), which contains data from >14,000 worldwide isolates of M. tuberculosis. The isolates clustered by spoligotyping were further characterized by IS6110-RFLP to confirm the clonal relationship. Sixty-six different spoligotypes were identified, grouping 125 of the isolates (74%). Approximately half of the isolates belonged to seven of the most frequently occurring spoligotypes in the database. Three shared types (with two or more isolates) not previously identified were given the type numbers 826, 827 and 863. An additional 45 spoligotypes were identified that did not match any existing database pattern. RFLP characterization reduced the number of isolates in most of the clusters, thereby showing a higher differentiation capacity than spoligotyping. These results highlight the importance of molecular epidemiology studies of tuberculosis in insufficiently studied regions with a high TB burden, in order to uncover the true extent of genetic diversity of the pathogen.     

Microarray analysis of vegetative phase change in maize

Vegetative phase change is the developmental transition from the juvenile phase to the adult phase in which a plant becomes competent for sexual reproduction. The gain of ability to flower is often accompanied by changes in patterns of differentiation in newly forming vegetative organs. In maize, juvenile leaves differ from adult leaves in morphology, anatomy and cell wall composition. Whereas the normal sequence of juvenile followed by adult is repeated with every sexual generation, this sequence can be altered in maize by the isolation and culture of the shoot apex from an adult phase plant: an 'adult' meristem so treated reverts to forming juvenile vegetative organs. To begin to unravel the as-yet poorly understood molecular mechanisms underlying phase change in maize, we compared gene expression in two juvenile sample types, leaf 4 and culture-derived leaves 3 or 4, with an adult sample type (leaf 9) using cDNA microarrays. All samples were leaf primordia at plastochron 6. A gene was scored as 'phase induced' if it was up- or downregulated in both juvenile sample types, compared with the adult sample type, with at least a twofold change in gene expression at a P-value of < or =0.005. Some 221 expressed sequence tags (ESTs) were upregulated in juveniles, and 28 ESTs were upregulated in adults. The largest class of juvenile-induced genes was comprised of those involved in photosynthesis, suggesting that maize plants are primed for energy production early in vegetative growth by the developmental induction of photosynthetic genes.