Lipidomics of homeoviscous adaptation to low temperatures in Staphylococcus aureus utilizing exogenous straight-chain unsaturated fatty acids.

It is well established that Staphylococcus aureus can incorporate exogenous straight-chain unsaturated fatty acids (SCUFAs) into membrane phospho- and glyco-lipids from various sources in supplemented culture media and when growing in vivo during infection. Given the enhancement of membrane fluidity when oleic acid (C18:1Δ9) is incorporated into lipids, we were prompted to examine the effect of medium supplementation with C18:1Δ9 on growth at low temperatures. C18:1Δ9 supported the growth of a cold-sensitive, branched-chain fatty acid (BCFA)-deficient mutant at 12°C. Interestingly, we found similar results in the BCFA-sufficient parental strain, supported by the fact that the incorporation of C18:1Δ9 into the membrane increased membrane fluidity in both strains. We show that the incorporation of C18:1Δ9 and its elongation product C20:1Δ11 into membrane lipids was required for growth stimulation and relied on a functional FakAB incorporation system. Lipidomics analysis of the phosphatidylglycerol and diglycosyldiacylglycerol lipid classes revealed major impacts of C18:1Δ9 and temperature on lipid species. Growth at 12°C in the presence of C18:1Δ9 also led to increased production of the carotenoid pigment staphyloxanthin. The enhancement of growth by C18:1Δ9 is an example of homeoviscous adaptation to low temperatures utilizing an exogenous fatty acid. This may be significant in the growth of S. aureus at low temperatures in foods that commonly contain C18:1Δ9 and other SCUFAs in various forms. IMPORTANCE: We show that Staphylococcus aureus can use its known ability to incorporate exogenous fatty acids to enhance its growth at low temperatures. Individual species of phosphatidylglycerols and diglycosyldiacylglycerols bearing one or two degrees of unsaturation derived from the incorporation of C18:1Δ9 at 12°C are described for the first time. In addition, enhanced production of the carotenoid staphyloxanthin occurs at low temperatures. The studies describe a biochemical reality underlying membrane biophysics. This is an example of homeoviscous adaptation to low temperatures utilizing exogenous fatty acids over the regulation of the biosynthesis of endogenous fatty acids. The studies have likely relevance to food safety in that unsaturated fatty acids may enhance the growth of S. aureus in the food environment.

Growth of Staphylococcus aureus in the presence of oleic acid shifts the glycolipid fatty acid profile and increases resistance to antimicrobial peptides.

Staphylococcus aureus readily adapts to various environments and quickly develops antibiotic resistance, which has led to an increase in multidrug-resistant infections. Hence, S. aureus presents a significant global health issue and its adaptations to the host environment are crucial for understanding pathogenesis and antibiotic susceptibility. When S. aureus is grown conventionally, its membrane lipids contain a mix of branched-chain and straight-chain saturated fatty acids. However, when unsaturated fatty acids are present in the growth medium, they become a major part of the total fatty acid composition. This study explores the biophysical effects of incorporating straight-chain unsaturated fatty acids into S. aureus membrane lipids. Membrane preparations from cultures supplemented with oleic acid showed more complex differential scanning calorimetry scans than those grown in tryptic soy broth alone. When grown in the presence of oleic acid, the cultures exhibited a transition significantly above the growth temperature, attributed to the presence of glycolipids with long-chain fatty acids causing acyl chain packing frustration within the bilayer. Functional aspects of the membrane were assessed by studying the kinetics of dye release from unilamellar vesicles induced by the antimicrobial peptide mastoparan X. Dye release was slower from liposomes prepared from cells grown in oleic acid-supplemented cultures, suggesting that changes in membrane lipid composition and biophysics protect the cell membrane against peptide-induced lysis. These findings underscore the intricate relationship between the growth environment, membrane lipid composition, and the physical properties of the bacterial membrane, which should be considered when developing new strategies against S. aureus infections.

Lipidomics of homeoviscous adaptation to low temperatures in Staphylococcus aureus utilizing exogenous straight-chain unsaturated fatty acids over biosynthesized endogenous branched-chain fatty acids.

It is well established that Staphylococcus aureus can incorporate exogenous straight-chain unsaturated fatty acids (SCUFAs) into membrane phospho- and glyco-lipids from various sources in supplemented culture media, and when growing in vivo in an infection. Given the enhancement of membrane fluidity when oleic acid (C18:1Δ9) is incorporated into lipids, we were prompted to examine the effect of medium supplementation with C18:1Δ9 on growth at low temperatures. C18:1Δ9 supported the growth of a cold-sensitive, branched-chain fatty acid (BCFA)-deficient mutant at 12°C. Interestingly, we found similar results in the BCFA-sufficient parental strain. We show that incorporation of C18:1Δ9 and its elongation product C20:1Δ9 into membrane lipids was required for growth stimulation and relied on a functional FakAB incorporation system. Lipidomics analysis of the phosphatidylglycerol (PG) and diglycosyldiacylglycerol (DGDG) lipid classes revealed major impacts of C18:1Δ9 and temperature on lipid species. Growth at 12°C in the presence of C18:1Δ9 also led to increased production of the carotenoid pigment staphyloxanthin; however, this was not an obligatory requirement for cold adaptation. Enhancement of growth by C18:1Δ9 is an example of homeoviscous adaptation to low temperatures utilizing an exogenous fatty acid. This may be significant in the growth of S. aureus at low temperatures in foods that commonly contain C18:1Δ9 and other SCUFAs in various forms.

The Human STAT2 Coiled-Coil Domain Contains a Degron for Zika Virus Interferon Evasion.

The ability of viruses to evade the host antiviral immune system determines their level of replication fitness, species specificity, and pathogenic potential. Flaviviruses rely on the subversion of innate immune barriers, including the type I and type III interferon (IFN) antiviral systems. Zika virus infection induces the degradation of STAT2, an essential component of the IFN-stimulated gene transcription factor ISGF3. The mechanisms that lead to STAT2 degradation by Zika virus are poorly understood, but it is known to be mediated by the viral NS5 protein that binds to STAT2 and targets it for proteasome-mediated destruction. To better understand how NS5 engages and degrades STAT2, functional analysis of the protein interactions that lead to Zika virus and NS5-dependent STAT2 proteolysis were investigated. Data implicate the STAT2 coiled-coil domain as necessary and sufficient for NS5 interaction and proteasome degradation after Zika virus infection. Molecular dissection reveals that the first two α-helices of the STAT2 coiled-coil domain contain a specific targeting region for IFN antagonism. These functional interactions provide a more complete understanding of the essential protein-protein interactions needed for Zika virus evasion of the host antiviral response and identify new targets for antiviral therapeutic approaches. IMPORTANCE Zika virus infection can cause mild fever, rash, and muscle pain and in rare cases can lead to brain or nervous system diseases, including Guillain-Barré syndrome. Infections in pregnant women can increase the risk of miscarriage or serious birth defects, including brain anomalies and microcephaly. There are no drugs or vaccines for Zika disease. Zika virus is known to break down the host antiviral immune response, and this research project reveals how the virus suppresses interferon signaling, and may reveal therapeutic vulnerabilities.

The influence of low-carbohydrate diets on the metabolic response to androgen-deprivation therapy in prostate cancer.

BACKGROUND: Prostate cancer (PC) is the second most lethal cancer for men. For metastatic PC, standard first-line treatment is androgen deprivation therapy (ADT). While effective, ADT has many metabolic side effects. Previously, we found in serum metabolome analysis that ADT reduced androsterone sulfate, 3-hydroxybutyric acid, acyl-carnitines but increased serum glucose. Since ADT reduced ketogenesis, we speculate that low-carbohydrate diets (LCD) may reverse many ADT-induced metabolic abnormalities in animals and humans. METHODS: In a multicenter trial of patients with PC initiating ADT randomized to no diet change (control) or LCD, we previously showed that LCD intervention led to significant weight loss, reduced fat mass, improved insulin resistance, and lipid profiles. To determine whether and how LCD affects ADT-induced metabolic changes, we analyzed serum metabolites after 3-, and 6-months of ADT on LCD versus control. RESULTS: We found androsterone sulfate was most consistently reduced by ADT and was slightly further reduced in the LCD arm. Contrastingly, LCD intervention increased 3-hydroxybutyric acid and various acyl-carnitines, counteracting their reduction during ADT. LCD also reversed the ADT-reduced lactic acid, alanine, and S-adenosyl methionine (SAM), elevating glycolysis metabolites and alanine. While the degree of androsterone reduction by ADT was strongly correlated with glucose and indole-3-carboxaldehyde, LCD disrupted such correlations. CONCLUSIONS: Together, LCD intervention significantly reversed many ADT-induced metabolic changes while slightly enhancing androgen reduction. Future research is needed to confirm these findings and determine whether LCD can mitigate ADT-linked comorbidities and possibly delaying disease progression by further lowering androgens.

Lipidomic and Ultrastructural Characterization of the Cell Envelope of Staphylococcus aureus Grown in the Presence of Human Serum.

Staphylococcus aureus can incorporate exogenous straight-chain unsaturated and saturated fatty acids (SCUFAs and SCFAs, respectively) to replace some of the normally biosynthesized branched-chain fatty acids and SCFAs. In this study, the impact of human serum on the S. aureus lipidome and cell envelope structure was comprehensively characterized. When S. aureus was grown in the presence of 20% human serum, typical human serum lipids, such as cholesterol, sphingomyelin, phosphatidylethanolamines, and phosphatidylcholines, were present in the total lipid extracts. Mass spectrometry showed that SCUFAs were incorporated into all major S. aureus lipid classes, i.e., phosphatidylglycerols, lysyl-phosphatidylglycerols, cardiolipins, and diglucosyldiacylglycerols. Heat-killed S. aureus retained fewer serum lipids and failed to incorporate SCUFAs, suggesting that association and incorporation of serum lipids with S. aureus require a living or nondenatured cell. Cytoplasmic membranes isolated from lysostaphin-produced protoplasts of serum-grown cells retained serum lipids, but washing cells with Triton X-100 removed most of them. Furthermore, electron microscopy studies showed that serum-grown cells had thicker cell envelopes and associated material on the surface, which was partially removed by Triton X-100 washing. To investigate which serum lipids were preferentially hydrolyzed by S. aureus lipases for incorporation, we incubated individual serum lipid classes with S. aureus and found that cholesteryl esters (CEs) and triglycerides (TGs) are the major donors of the incorporated fatty acids. Further experiments using purified Geh lipase confirmed that CEs and TGs were the substrates of this enzyme. Thus, growth in the presence of serum altered the nature of the cell surface with implications for interactions with the host.IMPORTANCE Comprehensive lipidomics of S. aureus grown in the presence of human serum suggests that human serum lipids can associate with the cell envelope without being truly integrated into the lipid membrane. However, fatty acids derived from human serum lipids, including unsaturated fatty acids, can be incorporated into lipid classes that can be biosynthesized by S. aureus itself. Cholesteryl esters and triglycerides are found to be the major source of incorporated fatty acids upon hydrolysis by lipases. These findings have significant implications for the nature of the S. aureus cell surface when grown in vivo Changes in phospholipid and glycolipid abundances and fatty acid composition could affect membrane biophysics and function and the activity of membrane-targeting antimicrobials. Finally, the association of serum lipids with the cell envelope has implications for the physicochemical nature of the cell surface and its interaction with host defense systems.

Bioinformatic analysis suggests potential mechanisms underlying parasitoid venom evolution and function.

Hymenopteran parasitoid wasps are a diverse collection of species that infect arthropod hosts and use factors found in their venoms to manipulate host immune responses, physiology, and behaviour. Whole parasitoid venoms have been profiled using proteomic approaches, and here we present a bioinformatic characterization of the venom protein content from Ganaspis sp. 1, a parasitoid that infects flies of the genus Drosophila. We find evidence that diverse evolutionary processes including multifunctionalization, co-option, gene duplication, and horizontal gene transfer may be acting in concert to drive venom gene evolution in Ganaspis sp.1. One major role of parasitoid wasp venom is host immune evasion. We previously demonstrated that Ganaspis sp. 1 venom inhibits immune cell activation in infected Drosophila melanogaster hosts, and our current analysis has uncovered additional predicted virulence functions. Overall, this analysis represents an important step towards understanding the composition and activity of parasitoid wasp venoms.

Response of Sulfolobus solfataricus Dpo4 polymerase in vitro to a DNA G-quadruplex.

Repetitive DNA sequences support the formation of structures that can interrupt replication and repair, leading to breaks and mutagenesis. One particularly stable structure is G-quadruplex (G4) DNA, which is four-stranded and formed from tandemly repetitive guanine bases. When folded within a template, G4 interferes with DNA synthesis. Similar to non-duplex structures, DNA base lesions can also halt an advancing replication fork, but the Y-family polymerases solve this problem by bypassing the damage. In order to better understand how guanine-rich DNA is replicated, we have investigated the activity of the model Y-family polymerase, Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4), on guanine-rich templates in vitro. We find that Dpo4 progression on templates containing either a single GC-rich hairpin or a G4 DNA structure is greatly reduced and synthesis stalls at the structure. Human polymerase eta (hPol eta) showed the same pattern of stalling at G4; however, and in contrast to Klenow, hPol eta and Dpo4 partially synthesise into the guanine repeat. Substitution of the nucleotide selectivity residue in Dpo4 with alanine permitted ribonucleotide incorporation on unstructured templates, but this further reduced the ability of Dpo4 to synthesise across from the guanine repeats. The advancement of Dpo4 on G4 templates was highest when the reaction was supplied with only deoxycytidine triphosphate, suggesting that high-fidelity synthesis is favoured over misincorporation. Our results are consistent with a model where the Y-family polymerases pause upon encountering G4 structures but have an ability to negotiate some synthesis through tetrad-associated guanines. This suggests that the Y-family polymerases reduce mutagenesis by catalysing the accurate replication of repetitive DNA sequences, but most likely in concert with additional replication and structure resolution activities.

Acceptability of emergency contraception in Brazil, Chile, and Mexico: 1 - Perceptions of emergency oral contraceptives.

This article presents the results of a study on the acceptability of emergency contraception (EC) in Brazil, Chile, and Mexico. Opinions of potential users and possible providers were obtained through discussion groups and those of authorities and policy-makers through semi-structured interviews. Most participants had a positive opinion of EC, based on the view that it can help reduce unplanned pregnancy, adolescent pregnancy, and unsafe abortion. Several interviewees felt that all women should be informed about EC, while others viewed it as a method for special situations such as rape and unprotected first sexual intercourse. Concern was expressed that its introduction might be associated with a decrease in condom use, increase in sexually transmitted diseases, and irresponsible or promiscuous sexual behavior among adolescents. The need for EC was clearly perceived by most participants, leading to the conclusion that health authorities have the responsibility of implementing programs for its introduction. Training of health care personnel should include the discussion of reproductive health problems that could be prevented by EC.

Acceptability of emergency contraception in Brazil, Chile, and Mexico. 2 - Facilitating factors versus obstacles

Foram identificados fatores que poderiam facilitar ou dificultar a introdução da anticoncepção de emergência (AE) no Brasil, Chile e México. Foram levantadas informações sobre o contexto sócio-cultural, político e legal bem como sobre as características dos serviços da saúde reprodutiva. As opiniões de potenciais usuários e possíveis provedores foram obtidas por meio de grupos de discussão, e as das autoridades e outras pessoas influentes por intermédio de entrevistas semi-estruturadas. Os obstáculos incluíram: percepção da AE como abortiva; oposição da Igreja Católica; pouco reconhecimento dos direitos sexuais e reprodutivos; pouca educação sexual; e falta de sensibilidade frente às questões de gênero. Os facilitadores foram: percepção da AE como um método que poderia prevenir o aborto e a gravidez entre adolescentes e vítimas de estupro; interesse no método mostrado por potenciais usuárias, bem como por alguns provedores e autoridades. Parece possível reduzir as barreiras identificadas com o apoio dos segmentos da sociedade comprometidos com a melhora da saúde sexual e reprodutiva, e com o treinamento adequado dos provedores de saúde.

Acceptability of emergency contraception in Brazil, Chile, and Mexico: 1 - Perceptions of emergency oral contraceptives

This article presents the results of a study on the acceptability of emergency contraception (EC) in Brazil, Chile, and Mexico. Opinions of potential users and possible providers were obtained through discussion groups and those of authorities and policy-makers through semi-structured interviews. Most participants had a positive opinion of EC, based on the view that it can help reduce unplanned pregnancy, adolescent pregnancy, and unsafe abortion. Several interviewees felt that all women should be informed about EC, while others viewed it as a method for special situations such as rape and unprotected first sexual intercourse. Concern was expressed that its introduction might be associated with a decrease in condom use, increase in sexually transmitted diseases, and irresponsible or promiscuous sexual behavior among adolescents. The need for EC was clearly perceived by most participants, leading to the conclusion that health authorities have the responsibility of implementing programs for its introduction. Training of health care personnel should include the discussion of reproductive health problems that could be prevented by EC

A 13-month multicenter clinical experience of a low-dose monophasic oral contraceptive containing 20 microg ethinylestradiol and 75 microg gestodene in Latin American women.

This prospective, multicenter study was conducted to evaluate the contraceptive reliability, cycle control and tolerability of a 21-day oral contraceptive regimen containing 20 microg ethinylestradiol and 75 microg gestodene in four Latin American countries (Mexico, Argentina, Brazil and Colombia). Participants took trial medication daily for 21 days. Contraceptive efficacy, cycle control and tolerability were evaluated over a period of 13 cycles. Efficacy data gathered from 5,109 treatment cycles were obtained from 393 participants. The trial medication proved to be an effective contraceptive and provided good cycle control. One pregnancy because of poor compliance was recorded. This resulted in a study Pearl index of 0.25. Forty-six percent of Latin American women reported one intracyclic spotting bleeding episode and 37.6% reported one intracyclic breakthrough bleeding (medium/excessive bleeding) episode during cycles 2-4 (primary target). Overall, intracyclic bleeding was reported in 41%. Overall, there was a trend towards a lower incidence of spotting in all the countries and this difference had statistical significance between Argentina and the others three countries (p < 0.05) during cycles 2-4. This trend was also apparent with respect to breakthrough bleeding, but again the difference did not achieve statistical significance. The discontinuation rate because of adverse events was low (3%); no serious adverse events were reported. More than 78% of the women in the four countries maintained constant body weight or lost weight (2 kg) during the study. The treatment effect on blood pressure was negligible. There were no appreciable changes in mean laboratory values over the course of the study.

Acceptability of emergency contraception in Brazil, Chile, and Mexico. 2 - Facilitating factors versus obstacles.

A multi-center study was performed in Brazil, Chile, and Mexico to identify factors that may facilitate or hinder the introduction of emergency contraception (EC) as well as perceptions concerning emergency contraceptive pills. Background information on the socio-cultural, political, and legal context and the characteristics of reproductive health services was collected. The opinions of potential users and providers were obtained through discussion groups, and those of authorities and policymakers through semi-structured interviews. Barriers to introduction included: perception of EC as an abortifacient, opposition by the Catholic Church, limited recognition of sexual and reproductive rights, limited sex education, and insensitivity to gender issues. Facilitating factors were: perception of EC as a method that would prevent abortion and pregnancy among adolescents and rape victims; interest in the method shown by potential users as well as by some providers and authorities. It appears possible to reduce barriers through support from segments of society committed to improving sexual and reproductive health and adequate training of health care providers.

Características del patrón de sangrado menstrual en un grupo de mujeres normales de Durango / Characteristics of menstrual bleeding pattern normal woman from Durango

Se estudiaron 1.250 episodios de sangrado en un total de 90 mujeres sanas, quienes registraron en calendarios menstruales únicamente variables objetivas como sangrado y manchado. El procesamiento de los datos se hizo según el procedimiento llamado "análisis del patrón menstrual" desarrollado por Rodríguez y col., el cual toma como unidad de estudio a la mujer. Otros parámetros fueron analizados por medio del método tradicional que utiliza como unidad de estudio el ciclo menstrual. El 81.1% de las mujeres estudiadas tenían entre 20 y 24 años de edad. En todos los casos se presentó un episodio de sangrado cada 30 días en promedio. El 90% de los casos permanecieron sangrando entre 6 y 15% de su periodo de referencia. La duración del ciclo menstrual fue entre 21 y 35 días en 93.6% del grupo total, con una duración del sangrado de uno a seis días en 99.2% del grupo total