Reproductive isolation in the Aegean Ophrys omegaifera complex (Orchidaceae)

The orchid genus Ophrys operates a system of sexual deception by which high specificity of pollination is attained. Reproductive isolation in Ophrys mainly rests upon prezygotic isolation mechanisms. The level of genetic separateness of Ophrys taxa with different pollinators is therefore likely determined by the fidelity of pollinators. The present study employs genetic fingerprinting to investigate this in the east Aegean Ophrys omegaifera s.l. complex, also including O. dryis, a west Mediterranean species of this complex. Ophrys fleischmannii, O. basilissa, and the west Mediterranean O. dyris, are found to be well-separated genetic entities whereas O. omegaifera s.str. and the putative hybrid taxon, O. sitiaca, are found to be genetically inseparable across their entire range of co-occurrence. This suggests that specific pollinators have high enough fidelity to act as effective isolating factors in east Aegean O. omegaifera s.l. as a whole, but that the situation in the species pair of O. sitiaca and O. omegaifera is likely to be more comple

Evidence for progenitor-derivative speciation in sexually deceptive orchids

Background and Aims Sexually deceptive orchids of the genus Ophrys use mimicry of pollinator females to attract specific pollinators. Pollinator shifts may drive speciation in Ophrys, since novel pollinators may in principle act as isolating factors immediately. It is thus possible that evolution of novel species occurs rapidly and with a progenitor-derivative pattern. The aims of this study are to compare genetic structure and diversity among widespread and geographically restricted Ophrys taxa, to test whether genetic structure is associated with specific pollinators, and to investigate whether any widespread species may have acted as a progenitor for the evolution of more restricted taxa. Methods Genetic differentiation and diversity were investigated in O. leucadica and O. cinereophila, the two taxa of the Ophrys fusca sensu lato complex widespread in the Aegean, and three geographically restricted taxa from Rhodes, O. attaviria, O. parvula and O. persephonae, all differing in their specific pollinators. This was done using amplified fragment length polymorphism (AFLP) DNA fingerprinting, and sequencing of the low-copy nuclear gene LEAFY (LFY). Key Results All taxa were found to be separate genetic entities, with O. leucadica forming two geographic groups from the west and east of the Aegean. Genetic structure was significantly shaped by pollinators and geography, and comparison of sequence and AFLP data revealed ancestral polymorphisms shared among several taxa. Among the sampled taxa, O. leucadica harbours the greatest genetic differentiation and geographic structure, and the highest genetic diversity. Part of the genome of O. parvula, endemic to Rhodes, may be derived from O. leucadica. Conclusions Pollinators probably influence the genetic structure of the investigated Ophrys species. The genetic pattern identified is consistent with O. leucadica being the oldest of the sampled taxa, making O. leucadica a candidate progenitor species from which more restricted taxa such as O. parvula may have evolve

Genetic variability in Brazilian Capsicum baccatum germplasm collection assessed by morphological fruit traits and AFLP markers.

<div><p><i>Capsicum baccatum</i> is one of the main pepper species grown and consumed in South America. In Brazil, it is commonly cultivated by family farmers, using mostly the genotypes bishop's hat genotypes (locally <i>cambuci</i>) and red chili pepper (<i>dedo-de-moça</i>). This study had the objective of characterizing 116 <i>C</i>. <i>baccatum</i> accessions from different regions of Brazil, based on morphological fruit descriptors and AFLP (Amplified Fragment Length Polymorphisms) markers. Broad phenotypic variability among the <i>C</i>. <i>baccatum</i> accessions was detected when using morphological fruit descriptors. The Ward modified location model (Ward-MLM) discriminated five groups, based mainly on fruit shape. Six combinations of AFLP primers detected polymorphism in 97.93% of the 2466 identified bands, indicating the high genetic variability in the accessions. The UPGMA coincided with the Bayesian clustering analysis and three large groups were formed, separating the wild variety <i>C</i>. <i>baccatum</i> var. <i>praetermissum</i> from the other accessions. There was no relation between genetic distance and geographical origin of the accessions, probably due to the intense exchange of fruits and seeds between farmers. Morphological descriptors used together with AFLP markers proved efficient in detecting the levels of genetic variability among the accessions maintained in the germplasm collections. These results can be used as an additional source of helpful information to be exploited in <i>C</i>. <i>baccatum</i> breeding programs.</p></div

Sustainable treatment of different high-strength cheese whey wastewaters: an innovative approach for atmospheric CO2 mitigation and fertilizer production

Raw cheese whey wastewater (CWW) has been treated by means of FeCl3 coagulation-flocculation, NaOH precipitation, and Ca(OH)2 precipitation. Three different types of CWW were considered: without cheese whey recovery (CWW0), 60 % cheese whey recovery (CWW60), and 80 % cheese whey recovery (CWW80). Cheese whey recovery significantly influenced the characteristics of the wastewater to be treated: organic matter, solids, turbidity, conductivity, sodium, chloride, calcium, nitrogen, potassium, and phosphorus. Initial organic load was reduced to values in the interval of 60–70 %. Application of FeCl3, NaOH, or Ca(OH)2 involved additional chemical oxygen demand (COD) depletions regardless of the CWW used. Under optimum conditions, the combination of 80 % cheese whey recovery and lime application led to 90 % reduction in COD. Turbidity (99.8%), total suspended solids (TSS) (98–99 %), oils and fats (82–96 %), phosphorus (98–99 %), potassium (96–97 %), and total coliforms (100 %) were also reduced. Sludge generated in the latter process showed excellent settling properties. This solid after filtration and natural evaporation can be used as fertilizer with limitations due to its saline nature. In an innovative, low-cost, and environmentally friendly technology, supernatant coming from the Ca(OH)2 addition was naturally neutralized in 4–6 days by atmospheric CO2 absorption without reagent addition. Consequently, a final aerobic biodegradation step can be applied for effluent polishing. This technology also allows for some atmospheric CO2 mitigation. Time requirement for the natural carbonation depends on the effluent characteristics. A precipitate rich in organic matter and nutrients and depletions of solids, sodium, phosphorus, magnesium, Kjeldahl, and ammoniacal nitrogen were also achieved during the natural carbonation

Underestimation Of The Maximal Capacity Of The Mitochondrial Electron Transport System In Oligomycin-treated Cells

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)The maximal capacity of the mitochondrial electron transport system (ETS) in intact cells is frequently estimated by promoting protonophore-induced maximal oxygen consumption preceded by inhibition of oxidative phosphorylation by oligomycin. In the present study, human glioma (T98G and U-87MG) and prostate cancer (PC-3) cells were titrated with different concentrations of the protonophore CCCP to induce maximal oxygen consumption rate (OCR) within respirometers in a conventional growth medium. The results demonstrate that the presence of oligomycin or its A-isomer leads to underestimation of maximal ETS capacity. In the presence of oligomycin, the spare respiratory capacity (SRC), i.e., the difference between the maximal and basal cellular OCR, was underestimated by 25 to 45%. The inhibitory effect of oligomycin on SRC was more pronounced in T98G cells and was observed in both suspended and attached cells. Underestimation of SRC also occurred when oxidative phosphorylation was fully inhibited by the ATP synthase inhibitor citreoviridin. Further experiments indicated that oligomycin cannot be replaced by the adenine nucleotide translocase inhibitors bongkrekic acid or carboxyatractyloside because, although these compounds have effects in permeabilized cells, they do not inhibit oxidative phosphorylation in intact cells. We replaced CCCP by FCCP, another potent protonophore and similar results were observed. Lower maximal OCR and SRC values were obtained with the weaker protonophore 2,4-dinitrophenol, and these parameters were not affected by the presence of oligomycin. In permeabilized cells or isolated brain mitochondria incubated with respiratory substrates, only a minor inhibitory effect of oligomycin on CCCP-induced maximal OCR was observed. We conclude that unless a previously validated protocol is employed, maximal ETS capacity in intact cells should be estimated without oligomycin. The inhibitory effect of an ATP synthase blocker on potent protonophore-induced maximal OCR may be associated with impaired metabolism of mitochondrial respiratory substrates.113Sao Paulo Research Foundation (FAPESP) [11/50400-0, 10/51906-1, 13/07937-8]Brazilian National Council for Scientific and Technological Development (CNPq)CAPESFAPESP [12/51288-1]Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Seroprevalence of antibodies against SARS-CoV-2 among health care workers in a Portuguese hospital

"Background: Health care workers (HCW) are presumably exposed to a higher risk of infection by SARS-CoV-2 and could possibly represent a source of transmission to susceptible patients. Thus, characterization of SARS-CoV-2 infection among HCW is necessary to better understand the determinants of viral transmission and properly implement strategies to prevent dissemination and protect HCW and vulnerable patients. The aim of this study was to estimate the seroprevalence of antibodies against SARS-CoV-2 in a Portuguese tertiary hospital, in the period of July 2020 to March 2021, before the generalized use of the SARS-CoV-2 vaccine, characterize its evolution over time, and identify risk factors associated with seroconversion. Methods: HCW were approached to collect serum samples for qualitative SARS-CoV-2 antibody testing and completion of an online survey capturing demographics, previous symptoms, and details of health care and community exposure. Odds ratio with bivariable and multivariable logistic regression was used to assess characteristics associated with seroprevalence. Results: One thousand HCW were included for analysis. Two hundred nineteen HCW (22%) were seropositive for immunoglobulin G against SARS-CoV-2, and 166 (17%) were seropositive for immunoglobulin M, most of whom reported a previous diagnosis of SARS-CoV-2 infection. The risk factors associated with seroconversion included a previous COVID-19 diagnosis, contact with patients, occupational contact with colleagues, and outside contact. However, in a multivariate logistic regression analysis, only a previous diagnosis and outside contact were associated with seroconversion. Seropositivity decreased over time, especially 28 weeks after infection. Conclusion: HCWs have a high seroprevalence for SARS-CoV-2 infection, probably due to a combination of health care and community exposure. Seropositivity decreases over time, but further studies are needed to better understand our adaptive immune response.

Late onset congenital adrenal hyperplasia due to 21-hydroxylase deficiency: revision of literature and preconception genetic study of five couples

The deficiency of the enzyme 21-hydroxylase (21-HO) is responsible for about 90% to 95% of all cases of congenital adrenal hyperplasia (HCSR). This disorder is one of the most frequent hereditary illnesses of autosomal recessive trait. The illness can be presented in two clinical forms: 1 - classic, subdivided in saltwasting form and simple virilizing form; 2 - nonclassic or late-onset. The severity of the illness is correlated with the degree of enzymatic activity of 21-HO, which depends on the type of mutation that occurs in gene CYP21A2. The late onset congenital adrenal hyperplasia is that one where the enzymatic blockade is less intense. The clinical is variable: precocious pubarche, polycystic ovaries, hirsutism, oligoamenorrhea, acne and infertility. Gold standard for the diagnosis of late onset congenital adrenal hyperplasia consists on the test of the tetracosactide, considering itself diagnostic positive when 17-hidroxiprogesterona (17-OHP) is higher of 10-15 ng per mL. Many patients don't need treatment; however, if necessary, such a treatment essentially consists on the Administration of glucocorticoids and antiandrogens. Considering that affected individuals and the carriers may have a severe mutation in CYP21A2, they can have descendents with the classic form (if their partner is also a carrier of a severe mutation), it is mandatory the genetic study of the couple. The authors illustrate the importance of this genetic study through five clinical cases, whose common link is the desire to have children, as well as the presence on the feminine element of late onset congenital adrenal hyperplasia. The genetic study of the couple allows adequate pre-conception counseling and also prevents the use of corticoids throughout the pregnancy (if there's no risk of descendents being affected with the classic form). This aspect must be had in account in programming the pregnancy, in order to prevent therapeutics and unnecessary distrusts

Genetic diversity among forty coffee varieties assessed by RAPD markers associated with restriction digestion

The genetic variability of 40 accessions of_C. arabica was evaluated using a combination of the random amplified polymorphic DNA (RAPD) technique and restriction digestion of genomic DNA. The genetic variability and the relatedness among all accessions were initially evaluated using 195 RAPD primers which revealed a very low level of genetic variation. To improve the efficiency in the detection of polymorphism, the genomic DNA of all accessions were submitted to digestion with restriction endonucleases prior to PCR amplification. A total of 24 primers combined with restriction digestion of DNA rendered 318 bands, of which 266 (83.65%) were polymorphic. The associations among genotypes were estimated using UPGMA-clustering analysis. The accessions were properly clustered according to pedigree and agronomic features. The ability to distinguish among coffee accessions was greater for RAPD plus restriction digestion than for RAPD alone, providing evidences that the combination of the techniques was very efficient for the estimation of genetic relationship among_C. arabica genotypes.Leandro Eugênio Cardamoni Diniz, Claudete de Fátima Ruas, Valdemar de Paula Carvalho, Fabrício Medeiros Torres, Eduardo Augusto Ruas, Melissa de Oliveira Santos, Tumoru Sera and Paulo Maurício Rua

Vaccination of Mice with Salmonella Expressing VapA: Mucosal and Systemic Th1 Responses Provide Protection against Rhodococcus equi Infection

Conventional vaccines to prevent the pneumonia caused by Rhodococcus equi have not been successful. We have recently demonstrated that immunization with Salmonella enterica Typhimurium expressing the VapA antigen protects mice against R. equi infection. We now report that oral vaccination of mice with this recombinant strain results in high and persistent fecal levels of antigen-specific IgA, and specific proliferation of the spleen cells of immunized mice in response to the in vitro stimulation with R. equi antigen. After in vitro stimulation, spleen cells of immunized mice produce high levels of Th1 cytokines and show a prominent mRNA expression of the Th1 transcription factor T-bet, in detriment of the Th2 transcription factor GATA-3. Following R. equi challenge, a high H2O2, NO, IL-12, and IFN-γ content is detected in the organs of immunized mice. On the other hand, TNF-α and IL-4 levels are markedly lower in the organs of vaccinated mice, compared with the non-vaccinated ones. The IL-10 content and the mRNA transcription level of TGF-β are also higher in the organs of immunized mice. A greater incidence of CD4+ and CD8+ T cells and B lymphocytes is verified in vaccinated mice. However, there is no difference between vaccinated and non-vaccinated mice in terms of the frequency of CD4+CD25+Foxp3+ T cells. Finally, we show that the vaccination confers a long-term protection against R. equi infection. Altogether, these data indicate that the oral vaccination of mice with S. enterica Typhimurium expressing VapA induces specific and long-lasting humoral and cellular responses against the pathogen, which are appropriately regulated and allow tissue integrity after challenge