Alternative oxidase in the branched mitochondrial respiratory network: an overview on structure, function, regulation, and role

Plants and some other organisms including protists possess a complex branched respiratory network in their mitochondria. Some pathways of this network are not energy-conserving and allow sites of energy conservation to be bypassed, leading to a decrease of the energy yield in the cells. It is a challenge to understand the regulation of the partitioning of electrons between the various energy-dissipating and -conserving pathways. This review is focused on the oxidase side of the respiratory chain that presents a cyanide-resistant energy-dissipating alternative oxidase (AOX) besides the cytochrome pathway. The known structural properties of AOX are described including transmembrane topology, dimerization, and active sites. Regulation of the alternative oxidase activity is presented in detail because of its complexity. The alternative oxidase activity is dependent on substrate availability: total ubiquinone concentration and its redox state in the membrane and O2 concentration in the cell. The alternative oxidase activity can be long-term regulated (gene expression) or short-term (post-translational modification, allosteric activation) regulated. Electron distribution (partitioning) between the alternative and cytochrome pathways during steady-state respiration is a crucial measurement to quantitatively analyze the effects of the various levels of regulation of the alternative oxidase. Three approaches are described with their specific domain of application and limitations: kinetic approach, oxygen isotope differential discrimination, and ADP/O method (thermokinetic approach). Lastly, the role of the alternative oxidase in non-thermogenic tissues is discussed in relation to the energy metabolism balance of the cell (supply in reducing equivalents/demand in energy and carbon) and with harmful reactive oxygen species formation.

Microbial profile of a kefir sample preparations: grains in natura and lyophilized and fermented suspension

Probiotics are supplementary foods developed by microbial strains that improve animal health beyond basic nutrition. Probiotics are consumed orally, regardless of being considered as normal inhabitants of the intestines, able to survive in enzimatic and biliary secretions. Kefir is a probiotic originated from the old continent, fermented by several bacteria and yeasts, encapsulated in a polyssacharide matrix, and resembles jelly grains. Kefir is also presented as its sourish product both in sugary or milky suspensions containing vitamins, aminoacids, peptides, carbohydrates, ethanol, and volatile compounds. Kefir is known to have a diverse microbial content depending on the country and fermentative substrates, which cause distinct probiotic effects. In this sense, the purpose of this work was to isolate, identify, and quantify the microbial content of a native sugary kefir sample (fermented suspension and lyophilized natural grains). Serial dilutions were plated on Rogosa agar (AR) and De Man, Rogosa and Sharpe (MRS), for Lactobacillus; Brain Heart Infusion (BHI), for total bacteria; Sabouraud-Dextrose-Agar (SDA), for yeasts and filamentous fungi; Thioglycolate Agar (TA), for Streptococcus, Acetobacteria and Leuconostoc; and Coconut Water Agar (CWA), and CWA supplemented with yeast extract (CWAY), for various genera. Genera and species for all strains were identified through biochemical reactions and specific API systems. The microbial profile of kefir was different from other sources of grains despite the presence of similar microorganisms and others which have not been reported yet. The data obtained with the CWA and CWAE media suggest that both substrates are alternative and salutary media for culture of kefir strains.

Antimonial drugs entrapped into phosphatidylserine liposomes: physicochemical evaluation and antileishmanial activity

Abstract: INTRODUCTION: Leishmaniasis is a disease caused by the protozoan Leishmania that resides mainly in mononuclear phagocytic system tissues. Pentavalent antimonials are the main treatment option, although these drugs have toxic side effects and high resistance rates. A potentially alternative and more effective therapeutic strategy is to use liposomes as carriers of the antileishmanial agents. The aims of this study were to develop antimonial drugs entrapped into phosphatidylserine liposomes and to analyze their biological and physicochemical characteristics. METHODS: Liposomes containing meglumine antimoniate (MA) or pentavalent antimony salt (Sb) were obtained through filter extrusion (FEL) and characterized by transmission electron microscopy. Promastigotes of Leishmania infantum were incubated with the drugs and the viability was determined with a tetrazolium dye (MTT assay). The effects of these drugs against intracellular amastigotes were also evaluated by optical microscopy, and mammalian cytotoxicity was determined by an MTT assay. RESULTS: Liposomes had an average diameter of 162nm. MA-FEL showed inhibitory activity against intracellular L. infantum amastigotes, with a 50% inhibitory concentration (IC50) of 0.9μg/mL, whereas that of MA was 60μg/mL. Sb-FEL showed an IC50 value of 0.2μg/mL, whereas that of free Sb was 9μg/mL. MA-FEL and Sb-FEL had strong in vitro activity that was 63-fold and 39-fold more effective than their respective free drugs. MA-FEL tested at a ten-times higher concentration than Sb-FEL did not show cytotoxicity to mammalian cells, resulting in a higher selectivity index. CONCLUSIONS: Antimonial drug-containing liposomes are more effective against Leishmania-infected macrophages than the non-liposomal drugs.

The identification and differentiation of the Candida parapsilosis complex species by polymerase chain reaction-restriction fragment length polymorphism of the internal transcribed spacer region of the rDNA

Currently, it is accepted that there are three species that were formerly grouped under Candida parapsilosis: C. para- psilosis sensu stricto, Candida orthopsilosis, andCandida metapsilosis. In fact, the antifungal susceptibility profiles and distinct virulence attributes demonstrate the differences in these nosocomial pathogens. An accurate, fast, and economical identification of fungal species has been the main goal in mycology. In the present study, we searched sequences that were available in the GenBank database in order to identify the complete sequence for the internal transcribed spacer (ITS)1-5.8S-ITS2 region, which is comprised of the forward and reverse primers ITS1 and ITS4. Subsequently, an in silico polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was performed to differentiate the C. parapsilosis complex species. Ninety-eight clinical isolates from patients with fungaemia were submitted for analysis, where 59 isolates were identified as C. parapsilosis sensu stricto, 37 were identified as C. orthopsilosis, and two were identified as C. metapsilosis. PCR-RFLP quickly and accurately identified C. parapsilosis complex species, making this method an alternative and routine identification system for use in clinical mycology laboratories.

Communication Strategies Of The Nursing Team In The Aphasia After Cerebrovascular Accident

This is an exploratory, cross-sectional study of quantitative design that aimed to identify the communication strategies used and reported by the nursing staff in the care of aphasic patients after a stroke. The techniques used were the participant observation and interviews with 27 subjects of the nursing staff of neurological units in a general hospital. The most frequently mentioned strategies were gestures (100%), verbal communication (33.3%), written communication (29.6%) and the touch (18.5 %). Among the observed strategies, the gestures reached 40.7% and the touch was present in all situations, given its instrumental character essential to care. The findings show lack of knowledge of nonverbal, proxemics , kinesics and tacesics communication. No significant differences were observed among the professional categories depending on the length of experience with respect to the strategies reported by members of the nursing staff in the care for aphasic patients.


Skin cancer in rural workers: nursing knowledge and intervention

OBJECTIVETo identify the exposure of rural workers to the sun's ultraviolet radiation and pesticides; to identify previous cases of skin cancer; and to implement clinical and communicative nursing actions among rural workers with a previous diagnosis of skin cancer.METHODObservational-exploratory study conducted with rural workers exposed to ultraviolet radiation and pesticides in a rural area in the extreme south of Brazil. A clinical judgment and risk communication model properly adapted was used to develop interventions among workers with a previous history of skin cancer.RESULTSA total of 123 (97.7%) workers were identified under conditions of exposure to the sun's ultraviolet radiation and pesticides; seven (5.4%) were identified with a previous diagnosis of skin cancer; four (57.1%) of these presented potential skin cancer lesions.CONCLUSIONThis study's results enabled clarifying the combination of clinical knowledge and risk communication regarding skin cancer to rural workers.

Visual communication stimulates reproduction in Nile tilapia, Oreochromis niloticus (L.)

Reproductive fish behavior is affected by male-female interactions that stimulate physiological responses such as hormonal release and gonad development. During male-female interactions, visual and chemical communication can modulate fish reproduction. The aim of the present study was to test the effect of visual and chemical male-female interaction on the gonad development and reproductive behavior of the cichlid fish Nile tilapia, Oreochromis niloticus (L.). Fifty-six pairs were studied after being maintained for 5 days under one of the four conditions (N = 14 for each condition): 1) visual contact (V); 2) chemical contact (Ch); 3) chemical and visual contact (Ch+V); 4) no sensory contact (Iso) - males and females isolated. We compared the reproductive behavior (nesting, courtship and spawning) and gonadosomatic index (GSI) of pairs of fish under all four conditions. Visual communication enhanced the frequency of courtship in males (mean ± SEM; V: 24.79 ± 3.30, Ch+V: 20.74 ± 3.09, Ch: 0.1 ± 0.07, Iso: 4.68 ± 1.26 events/30 min; P < 0.05, two-way ANOVA with LSD post hoc test), induced spawning in females (3 spawning in V and also 3 in Ch+V condition), and increased GSI in males (mean ± SEM; V: 1.39 ± 0.08, Ch+V: 1.21 ± 0.08, Ch: 1.04 ± 0.07, Iso: 0.82 ± 0.07%; P < 0.05, two-way ANOVA with LSD post hoc test). Chemical communication did not affect the reproductive behavior of pairs nor did it enhance the effects of visual contact. Therefore, male-female visual communication is an effective cue, which stimulates reproduction among pairs of Nile tilapia.

Expression and purification of the immunogenically active fragment B of the Park Williams 8 Corynebacterium diphtheriae strain toxin

The construction of a hexahistidine-tagged version of the B fragment of diphtheria toxin (DTB) represents an important step in the study of the biological properties of DTB because it will permit the production of pure recombinant DTB (rDTB) in less time and with higher yields than currently available. In the present study, the genomic DNA of the Corynebacterium diphtheriae Park Williams 8 (PW8) vaccine strain was used as a template for PCR amplification of the dtb gene. After amplification, the dtb gene was cloned and expressed in competent Escherichia coli M15™ cells using the expression vector pQE-30™. The lysate obtained from transformed E. coli cells containing the rDTB PW8 was clarified by centrifugation and purified by affinity chromatography. The homogeneity of the purified rDTB PW8 was confirmed by immunoblotting using mouse polyclonal anti-diphtheria toxoid antibodies and the immune response induced in animals with rDTB PW8 was evaluated by ELISA and dermonecrotic neutralization assays. The main result of the present study was an alternative and accessible method for the expression and purification of immunogenically reactive rDTB PW8 using commercially available systems. Data also provided preliminary evidence that rabbits immunized with rDTB PW8 are able to mount a neutralizing response against the challenge with toxigenic C. diphtheriae.

Expression and purification of the non-tagged LipL32 of pathogenic Leptospira

Leptospirosis is a reemerging infectious disease and the most disseminated zoonosis worldwide. A leptospiral surface protein, LipL32, only occurs in pathogenic Leptospira, and is the most abundant protein on the bacterial surface, being described as an important factor in host immunogenic response and also in bacterial infection. We describe here an alternative and simple purification protocol for non-tagged recombinant LipL32. The recombinant LipL32(21-272) was expressed in Escherichia coli without His-tag or any other tag used to facilitate recombinant protein purification. The recombinant protein was expressed in the soluble form, and the purification was based on ion exchange (anionic and cationic) and hydrophobic interactions. The final purification yielded 3 mg soluble LipL32(21-272) per liter of the induced culture. Antiserum produced against the recombinant protein was effective to detect native LipL32 from cell extracts of several Leptospira serovars. The purified recombinant LipL32(21-272) produced by this protocol can be used for structural, biochemical and functional studies and avoids the risk of possible interactions and interferences of the tags commonly used as well as the time consuming and almost always inefficient methods to cleave these tags when a tag-free LipL32 is needed. Non-tagged LipL32 may represent an alternative antigen for biochemical studies, for serodiagnosis and for the development of a vaccine against leptospirosis.

Acute neuromuscular manifestations in a patient associated with ingesting octopus (Octopus sp.)

The authors report neuromuscular manifestations in a 45-year-old woman after consuming octopus meat (Octopus sp.). The patient presented malaise, paresthesias in perioral and extremity areas, intense muscular weakness and arterial hypotension, followed by severe itch and disseminated cutaneous rash. Gastrointestinal manifestations and fever were not observed, reducing the probability of alimentary poisoning. The presence of muscular and neurological symptoms suggests neurotoxin action, which could have been ingested by the victim from the octopus salivary glands or from an accumulation of toxins in the meat, or by an unknown mechanism. There is little known about toxins of the Octopus genus and this communication is important alert to the possibility of poisoning in humans that eat octopus and its differentiation from alimentary poisonings arising from incorrect conservation of seafood.

Anatomic study of the dorsal arterial system of the hand

Historically, the dorsal arterial system of the hand received less attention than the palmar system. The studies concerning dorsal arterial anatomy present some controversies regarding the origin and presence of the dorsal metacarpal artery branches. Knowledge of the anatomy of dorsal metacarpal arteries is especially applied in the surgical planning for flaps taken from the dorsum of the hand. The purpose of this study is to analyze the arterial anatomy of the dorsum of the hand, compare our observations with those of previous studies from the literature, and therefore to define parameters for surgical planning for flaps supplied by the dorsal metacarpal arteries. METHOD: Twenty-six dissections were performed at the dorsum of the right hand of 26 cadavers by making a distal-based U-shaped incision. After catheterization of the radial artery at the wrist level, a plastic dye solution with low viscosity and quick solidification was injected to allow adequate exposure of even small vessels. The radial artery and its branches, the dorsal arterial arch, the dorsal metacarpal arteries, the distal and proximal communicating branches of the palmar system, and the distal cutaneous branches were carefully dissected and identified. RESULTS: The distal cutaneous branches originating from the dorsal metacarpal arteries were observed in all cases; these were located an average of 1.2 cm proximal from the metacarpophalangeal joint. The first dorsal metacarpal artery presented in 3 different patterns regarding its course: fascial, subfascial, and mixed. The branching pattern of the radial artery at the first intermetacarpal space was its division into 3 branches. We observed the presence of the dorsal arterial arch arising from the radial artery in 100% of the cases. The distance between the dorsal arterial arch and the branching point of the radial artery was an average of 2 cm. The first and second dorsal metacarpal arteries were visualized in all cases. The third and fourth dorsal metacarpal arteries were visualized in 96.2% and 92.3% of cases, respectively. There was proximal and distal communication between the dorsal arterial arch and the palmar system through the communicating branches contributing to the dorsal metacarpal artery formation. CONCLUSION: At the dorsum of the hand there is a rich arterial net that anastomoses with the palmar arterial system. This anatomical characteristic allows the utilization of the dorsal aspect of the hand as potential donor site for cutaneous flaps.

Daughter of time: the postmodern midwife (Part 1)

This article presents the notion of the postmodern midwife, defining her as one who takes a relativistic stance toward bio-medicine and other knowledge systems, alternative and indigenous, moving fluidly between them to serve the women she attends. She is locally and globally aware, culturally competent, and politically engaged, working with the resources at hand to preserve midwifery in the interests of women. Her informed relativism is most accessible to professional midwives but is also beginning to characterize some savvy traditional midwives in various countries. Thus the concept of the postmodern midwife can serve as a bridge across the ethnic, racial, and status gaps that divide the professional from the traditional midwife, and as an analytical focal point for understanding how the members of each group negotiate their identities and their roles in a changing world.

Comfort of the patient's family in an Intensive Care Unit related to welcoming

This study aimed to identify the level of comfort of families of patients in a critical health condition related to the welcoming practices performed by the hospital staff. Interviews were conducted with 250 relatives in hospitals of the state Bahia, using a Likert scale. Data were analyzed as percentages and quartiles. For nine of the 12 statements of the scale, most relatives scored their comfort level between very and totally comfortable, median of 4,revealing kindness, tranquility and friendly communication with family members. More than half of the sample scored its level as not at all to more or less comfortable, median of 3, for statements about demonstration of interest towards the relative by the staff and flexible visiting of the patient. The necessity of greater interest of the team in the condition and needs of the family was observed. Promoting comfort from the dimension of welcoming demands interdisciplinary actions grounded in humanistic philosophy, in which the nurse has an important role to play.