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2.  Rectal Douching and Implications for Rectal Microbicides among Populations Vulnerable to HIV in South America: A Qualitative Study 
While gel-formulated Rectal Microbicides (RM) are the first to enter clinical trials, rectal douching in preparation for anal intercourse is a common practise, thus RMs formulated as douches may be a convenient alternative to gels. Nonetheless, little is known about potential users’ thoughts regarding douche-formulated RMs or rectal douching practises, data needed to inform the advancement of douche-based RMs. This qualitative study examined thoughts regarding douches, their use as a RM and current douching practises among men who have sex with men and transgender women.
Ten focus groups and 36 in-depth interviews were conducted (N=140) to examine the overall acceptability of RM, of which one component focused on rectal douching. Focus groups and interviews were recorded, transcribed verbatim and coded; text relating to rectal douching was extracted and analysed. Sociodemographic information was collected using a self-administered questionnaire.
Support for a douche-formulated RM centred on the possibility of combined pre-coital hygiene and HIV protection, and it was believed that a deeply-penetrating liquid douche would confer greater HIV protection than a gel. Drawbacks included rectal dryness; impracticality and portability issues; and, potential side effects. Non-commercial douching apparatus use was common and liquids used included detergents, vinegar, bleach, lemon juice and alcohol.
A douche-formulated RM while desirable and perceived as more effective than a gel-formulated RM also generated questions regarding practicality and side-effects. Of immediate concern were the non-commercial liquids already being used which likely damage rectal epithelia, potentially increasing HIV infection risk. Pre-coital rectal douching is common and a RM formulated as such is desirable, but education on rectal douching practices is needed now.
PMCID: PMC4035233  PMID: 23966338
rectal douching; microbicides; HIV prevention; MSM; transgender women; South America
3.  A Review on Hemeoxygenase-2: Focus on Cellular Protection and Oxygen Response 
Hemeoxygenase (HO) system is responsible for cellular heme degradation to biliverdin, iron, and carbon monoxide. Two isoforms have been reported to date. Homologous HO-1 and HO-2 are microsomal proteins with more than 45% residue identity, share a similar fold and catalyze the same reaction. However, important differences between isoforms also exist. HO-1 isoform has been extensively studied mainly by its ability to respond to cellular stresses such as hemin, nitric oxide donors, oxidative damage, hypoxia, hyperthermia, and heavy metals, between others. On the contrary, due to its apparently constitutive nature, HO-2 has been less studied. Nevertheless, its abundance in tissues such as testis, endothelial cells, and particularly in brain, has pointed the relevance of HO-2 function. HO-2 presents particular characteristics that made it a unique protein in the HO system. Since attractive results on HO-2 have been arisen in later years, we focused this review in the second isoform. We summarize information on gene description, protein structure, and catalytic activity of HO-2 and particular facts such as its cellular impact and activity regulation. Finally, we call attention on the role of HO-2 in oxygen sensing, discussing proposed hypothesis on heme binding motifs and redox/thiol switches that participate in oxygen sensing as well as evidences of HO-2 response to hypoxia.
PMCID: PMC4127239  PMID: 25136403
4.  Efficient Production of Bacillus thuringiensis Cry1AMod Toxins under Regulation of cry3Aa Promoter and Single Cysteine Mutations in the Protoxin Region 
Applied and Environmental Microbiology  2013;79(22):6969-6973.
Bacillus thuringiensis Cry1AbMod toxins are engineered versions of Cry1Ab that lack the amino-terminal end, including domain I helix α-1 and part of helix α-2. This deletion improves oligomerization of these toxins in solution in the absence of cadherin receptor and counters resistance to Cry1A toxins in different lepidopteran insects, suggesting that oligomerization plays a major role in their toxicity. However, Cry1AbMod toxins are toxic to Escherichia coli cells, since the cry1A promoter that drives its expression in B. thuringiensis has readthrough expression activity in E. coli, making difficult the construction of these CryMod toxins. In this work, we show that Cry1AbMod and Cry1AcMod toxins can be cloned efficiently under regulation of the cry3A promoter region to drive its expression in B. thuringiensis without expression in E. coli cells. However, p3A-Cry1Ab(c)Mod construction promotes the formation of Cry1AMod crystals in B. thuringiensis cells that were not soluble at pH 10.5 and showed no toxicity to Plutella xylostella larvae. Cysteine residues in the protoxin carboxyl-terminal end of Cry1A toxins have been shown to be involved in disulfide bond formation, which is important for crystallization. Six individual cysteine substitutions for serine residues were constructed in the carboxyl-terminal protoxin end of the p3A-Cry1AbMod construct and one in the carboxyl-terminal protoxin end of p3A-Cry1AcMod. Interestingly, p3A-Cry1AbMod C654S and C729S and p3A-Cry1AcMod C730S recover crystal solubility at pH 10.5 and toxicity to P. xylostella. These results show that combining the cry3A promoter expression system with single cysteine mutations is a useful system for efficient expression of Cry1AMod toxins in B. thuringiensis.
PMCID: PMC3811527  PMID: 24014526
5.  Bacillus thuringiensis Cry1A toxins are versatile proteins with multiple modes of action: two distinct pre-pores are involved in toxicity 
Biochemical Journal  2014;459(Pt 2):383-396.
Cry proteins from Bacillus thuringiensis are insecticidal PFTs (pore-forming toxins). In the present study, we show that two distinct functional pre-pores of Cry1Ab are formed after binding of the protoxin or the protease-activated toxin to the cadherin receptor, but before membrane insertion. Both pre-pores actively induce pore formation, although with different characteristics, and contribute to the insecticidal activity. We also analysed the oligomerization of the mutant Cry1AbMod protein. This mutant kills different insect populations that are resistant to Cry toxins, but lost potency against susceptible insects. We found that the Cry1AbMod-protoxin efficiently induces oligomerization, but not the activated Cry1AbMod-toxin, explaining the loss of potency of Cry1AbMod against susceptible insects. These data are relevant for the future control of insects resistant to Cry proteins. Our data support the pore-formation model involving sequential interaction with different midgut proteins, leading to pore formation in the target membrane. We propose that not only different insect targets could have different receptors, but also different midgut proteases that would influence the rate of protoxin/toxin activation. It is possible that the two pre-pore structures could have been selected for in evolution, since they have differential roles in toxicity against selected targets, increasing their range of action. These data assign a functional role for the protoxin fragment of Cry PFTs that was not understood previously. Most PFTs produced by other bacteria are secreted as protoxins that require activation before oligomerization, to finally form a pore. Thus different pre-pores could be also part of the general mechanism of action of other PFTs.
Two distinct functional pre-pore oligomers of the Cry1Ab insecticidal toxin are formed before membrane insertion. These oligomers are formed after binding of either the protoxin or the protease-activated toxin to the cadherin receptor. Both pre-pores have different characteristics and contribute to insecticidal activity.
PMCID: PMC3969221  PMID: 24456341
Bacillus thuringiensis; Cry toxin; mechanism of action; oligomerization; pore-forming toxin; pre-pore; ABCC2, ATP-binding cassette C2; ALP, alkaline phosphatase; APN, aminopeptidase N; BBMV, brush border membrane vesicle; 3d-Cry, three-domain Cry; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PFT, pore-forming toxin; PLB, planar lipid bilayer; SUV, small unilamellar vesicle
6.  Direct Observation of Reversible Electronic Energy Transfer Involving an Iridium Center 
Inorganic Chemistry  2014;53(5):2677-2682.
A cyclometalated iridium complex is reported where the core complex comprises naphthylpyridine as the main ligand and the ancillary 2,2′-bipyridine ligand is attached to a pyrene unit by a short alkyl bridge. To obtain the complex with satisfactory purity, it was necessary to modify the standard synthesis (direct reaction of the ancillary ligand with the chloro-bridged iridium dimer) to a method harnessing an intermediate tetramethylheptanolate-based complex, which was subjected to acid-promoted removal of the ancillary ligand and subsequent complexation. The photophysical behavior of the bichromophoric complex and a model complex without the pendant pyrene were studied using steady-state and time-resolved spectroscopies. Reversible electronic energy transfer (REET) is demonstrated, uniquely with an emissive cyclometalated iridium center and an adjacent organic chromophore. After excited-state equilibration is established (5 ns) as a result of REET, extremely long luminescence lifetimes of up to 225 μs result, compared to 8.3 μs for the model complex, without diminishing the emission quantum yield. As a result, remarkably high oxygen sensitivity is observed in both solution and polymeric matrices.
A cyclometalated iridium complex with an appended pyrene unit is reported to have a remarkably long excited-state lifetime (225 μs) due to an efficient reversible electronic energy transfer between the two chromophores, as evidenced by time-resolved spectroscopies. As a result, the complex is among the best iridium complexes for oxygen sensing at low concentration.
PMCID: PMC3950932  PMID: 24555716
7.  Weed Mapping in Early-Season Maize Fields Using Object-Based Analysis of Unmanned Aerial Vehicle (UAV) Images 
PLoS ONE  2013;8(10):e77151.
The use of remote imagery captured by unmanned aerial vehicles (UAV) has tremendous potential for designing detailed site-specific weed control treatments in early post-emergence, which have not possible previously with conventional airborne or satellite images. A robust and entirely automatic object-based image analysis (OBIA) procedure was developed on a series of UAV images using a six-band multispectral camera (visible and near-infrared range) with the ultimate objective of generating a weed map in an experimental maize field in Spain. The OBIA procedure combines several contextual, hierarchical and object-based features and consists of three consecutive phases: 1) classification of crop rows by application of a dynamic and auto-adaptive classification approach, 2) discrimination of crops and weeds on the basis of their relative positions with reference to the crop rows, and 3) generation of a weed infestation map in a grid structure. The estimation of weed coverage from the image analysis yielded satisfactory results. The relationship of estimated versus observed weed densities had a coefficient of determination of r2=0.89 and a root mean square error of 0.02. A map of three categories of weed coverage was produced with 86% of overall accuracy. In the experimental field, the area free of weeds was 23%, and the area with low weed coverage (<5% weeds) was 47%, which indicated a high potential for reducing herbicide application or other weed operations. The OBIA procedure computes multiple data and statistics derived from the classification outputs, which permits calculation of herbicide requirements and estimation of the overall cost of weed management operations in advance.
PMCID: PMC3795646  PMID: 24146963
8.  Bacillus thuringiensis ssp. israelensis Cyt1Aa enhances activity of Cry11Aa toxin by facilitating the formation of a pre-pore oligomeric structure 
Cellular microbiology  2007;9(12):2931-2937.
Bacillus thuringiensis ssp. israelensis (Bti) has been used worldwide for the control of dipteran insect pests. This bacterium produces several Cry and Cyt toxins that individually show activity against mosquitoes but together show synergistic effect. Previous work demonstrated that Cyt1Aa synergizes the toxic activity of Cry11Aa by functioning as a membrane-bound receptor. In the case of Cry toxins active against lepidopteran insects, receptor interaction triggers the formation of a pre-pore oligomer that is responsible for pore formation and toxicity. In this work we report that binding of Cry11Aa to Cyt1Aa facilitates the formation of a Cry11Aa pre-pore oligomeric structure that is capable of forming pores in membrane vesicles. Cry11Aa and Cyt1A point mutants affected in binding and in synergism had a correlative effect on the formation of Cry11Aa pre-pore oligomer and on pore-formation activity of Cry11Aa. These data further support that Cyt1Aa interacts with Cry11Aa and demonstrate the molecular mechanism by which Cyt1Aa synergizes or suppresses resistance to Cry11Aa, by providing a binding site for Cry11Aa that will result in an efficient formation of Cry11Aa pre-pore that inserts into membranes and forms ionic pores.
PMCID: PMC3700374  PMID: 17672866
9.  Configuration and Specifications of an Unmanned Aerial Vehicle (UAV) for Early Site Specific Weed Management 
PLoS ONE  2013;8(3):e58210.
A new aerial platform has risen recently for image acquisition, the Unmanned Aerial Vehicle (UAV). This article describes the technical specifications and configuration of a UAV used to capture remote images for early season site- specific weed management (ESSWM). Image spatial and spectral properties required for weed seedling discrimination were also evaluated. Two different sensors, a still visible camera and a six-band multispectral camera, and three flight altitudes (30, 60 and 100 m) were tested over a naturally infested sunflower field. The main phases of the UAV workflow were the following: 1) mission planning, 2) UAV flight and image acquisition, and 3) image pre-processing. Three different aspects were needed to plan the route: flight area, camera specifications and UAV tasks. The pre-processing phase included the correct alignment of the six bands of the multispectral imagery and the orthorectification and mosaicking of the individual images captured in each flight. The image pixel size, area covered by each image and flight timing were very sensitive to flight altitude. At a lower altitude, the UAV captured images of finer spatial resolution, although the number of images needed to cover the whole field may be a limiting factor due to the energy required for a greater flight length and computational requirements for the further mosaicking process. Spectral differences between weeds, crop and bare soil were significant in the vegetation indices studied (Excess Green Index, Normalised Green-Red Difference Index and Normalised Difference Vegetation Index), mainly at a 30 m altitude. However, greater spectral separability was obtained between vegetation and bare soil with the index NDVI. These results suggest that an agreement among spectral and spatial resolutions is needed to optimise the flight mission according to every agronomical objective as affected by the size of the smaller object to be discriminated (weed plants or weed patches).
PMCID: PMC3590160  PMID: 23483997
10.  Defensin Production by Human Limbo-Corneal Fibroblasts Infected with Mycobacteria 
Pathogens  2013;2(1):13-32.
Epithelial cells of the cornea and the conjunctiva constitutively produce antimicrobial peptides; however, the production of defensins by other cell types located around the eye has not been investigated. We analyzed the production of beta-defensins (hBD) and cathelicidin LL-37 during the infection of primary limbo-corneal fibroblasts with M. tuberculosis (MTB), M. abscessus (MAB), and M. smegmatis (MSM). The intracellular survival of each mycobacterium, the production of cytokines and the changes on the distribution of the actin filaments during the infection were also analyzed. Fibroblasts produce basal levels of hBD1 and LL-37 and under PMA stimulation they produce hBD2, hBD3 and overexpress hBD1 and LL-37. MAB induced the highest levels of hBD1 and LL-37 and intermediate levels of IL-6; however, MAB was not eliminated. In addition, MAB induced the greatest change to the distribution of the actin filaments. MTB also produced changes in the structure of the cytoskeleton and induced low levels of hBD1 and IL-6, and intermediate levels of LL-37. The balance of these molecules induced by MTB appeared to contribute to the non-replicative state observed in the limbo-corneal cells. MSM induced the lowest levels of hBD1 and LL-37 but the highest levels of IL-6; MSM was eliminated. The results suggest that mycobacterial infections regulate the production of antimicrobial peptides and cytokines, which in conjunction can contribute to the control of the bacilli.
PMCID: PMC4235707  PMID: 25436879
human beta-defensin; cathelicidin LL-37; fibroblasts; limbo-corneal cells; Mycobacterium abscessus; Mycobacterium tuberculosis; Mycobacterium smegmatis; fibroblast cytoskeleton; IL-6
11.  Diversity of commercial sex among men and male-born trans people in three Peruvian cities 
Culture, health & sexuality  2011;13(10):1207-1221.
In Peru, commercial sex involving men and male-born travestis, transgenders and transsexuals (CSMT) is usually represented as a dangerous practice carried out on the streets by people experiencing economic hardship and social exclusion. However, in reality little is known about the complexities of this practice in Peru. This paper presents findings from an ethnographic study of the characteristics, patterns and socio-cultural aspects of CSMT in three Peruvian cities. The study included participant observation in sex work venues and interviews with 42 sex workers and 25 key informants. We found that CSMT in Peru takes many forms (some not previously described in the country) and is practised in different places by people from various socioeconomic levels. In many cases, the practice appears linked to ideals of social mobility, migratory experiences and other economic activities. In addition, the increasing use of the internet and mobile phones has changed patterns of sex work in Peru. We review the implications of these findings for future research and public health interventions.
PMCID: PMC3244347  PMID: 21936651
prostitution; men; transsexual people; ethnography; Peru
12.  Macropinocytosis is responsible for the uptake of pathogenic and non-pathogenic mycobacteria by B lymphocytes (Raji cells) 
BMC Microbiology  2012;12:246.
The classical roles of B cells include the production of antibodies and cytokines and the generation of immunological memory, these being key factors in the adaptive immune response. However, their role in innate immunity is currently being recognised. Traditionally, B cells have been considered non-phagocytic cells; therefore, the uptake of bacteria by B cells is not extensively documented. In this study, we analysed some of the features of non-specific bacterial uptake by B lymphocytes from the Raji cell line. In our model, B cells were infected with Mycobacterium tuberculosis (MTB), Mycobacterium smegmatis (MSM), and Salmonella typhimurium (ST).
Our observations revealed that the Raji B cells were readily infected by the three bacteria that were studied. All of the infections induced changes in the cellular membrane during bacterial internalisation. M. smegmatis and S. typhimurium were able to induce important membrane changes that were characterised by abundant filopodia and lamellipodia formation. These membrane changes were driven by actin cytoskeletal rearrangements. The intracellular growth of these bacteria was also controlled by B cells. M. tuberculosis infection also induced actin rearrangement-driven membrane changes; however, the B cells were not able to control this infection. The phorbol 12-myristate 13-acetate (PMA) treatment of B cells induced filopodia and lamellipodia formation, the production of spacious vacuoles (macropinosomes), and the fluid-phase uptake that is characteristic of macropinocytosis. S. typhimurium infection induced the highest fluid-phase uptake, although both mycobacteria also induced fluid uptake. A macropinocytosis inhibitor such as amiloride was used and abolished the bacterial uptake and the fluid-phase uptake that is triggered during the bacterial infection.
Raji B cells can internalise S. typhimurium and mycobacteria through an active process, such as macropinocytosis, although the resolution of the infection depends on factors that are inherent in the virulence of each pathogen.
PMCID: PMC3559283  PMID: 23113903
Macropinocytosis; B lymphocytes; Raji cells; Mycobacterium smegmatis; Mycobacterium tuberculosis; Salmonella typhimurium; Cytoskeleton
13.  New fluorescent pH sensors based on covalently linkable PET rhodamines 
Talanta  2012;99(2-3):194-201.
A new class of rhodamines for the application as indicator dyes in fluorescent pH sensors is presented. Their pH-sensitivity derives from photoinduced electron transfer between non-protonated amino groups and the excited chromophore which results in effective fluorescence quenching at increasing pH. The new indicator class carries a pentafluorophenyl group at the 9-position of the xanthene core where other rhodamines bear 2-carboxyphenyl substituents instead. The pentafluorophenyl group is used for covalent coupling to sensor matrices by “click” reaction with mercapto groups. Photophysical properties are similar to “classical” rhodamines carrying 2′-carboxy groups. pH sensors have been prepared with two different matrix materials, silica gel and poly(2-hydroxyethylmethacrylate). Both sensors show high luminescence brightness (absolute fluorescence quantum yield ΦF≈0.6) and high pH-sensitivity at pH 5–7 which makes them suitable for monitoring biotechnological samples. To underline practical applicability, a dually lifetime referenced sensor containing Cr(III)-doped Al2O3 as reference material is presented.
► New rhodamine pH-indicators employing photoinduced electron transfer are prepared. ► Indicators are covalently immobilised to the sensor support. ► Obtained pH optrodes feature high brightness, excellent sensitivity and good operation stability. ► Dynamic range (pH 5–7) is particularly useful for biotechnological applications.
PMCID: PMC3484401  PMID: 22967541
pH sensor; Fluorescence; Rhodamine; Photoinduced electron transfer; Covalent dye coupling; “Click” chemistry
14.  Genital Tract Infections, Bacterial Vaginosis, HIV, and Reproductive Health Issues among Lima-Based Clandestine Female Sex Workers 
Sociodemographic and behavioral characteristics of 212 Peruvian female sex workers (FSWs) were analyzed. The association between genital tract infections (GTIs) and risk factors by multivariate analysis was evaluated. Eighty-eight percent of FSWs were diagnosed with at least one GTI (HSV-2 80.1%, BV 44.8%, candidiasis 9.9%, syphilis seropositivity 9.4%, Trichomonas vaginalis 2.4%, HIV seropositivity 2.4%). Reported condom use with clients was nearly universal (98.3%), but infrequent with husband/regular partners (7.3%). In multivariate analysis BV was negatively associated with more consistent condom use (PRR = 0.63, 95% CI, 0.42–0.96). Many had not visited a Sexually Transmitted Infection (STI) clinic or been tested for HIV in the past year (40.6%, 47.1%, resp.). Nonclient contraceptive use was low (57%) and induced abortion was common (68%). High GTI burden and abortions suggest that a services-access gap persists among marginalized FSWs. Continued health outreach programs and integrating family planning and reproductive health services into existing STI clinic services are recommended.
PMCID: PMC3395213  PMID: 22811592
16.  Particular characteristics of allergic symptoms in tropical environments: follow up to 24 months in the FRAAT birth cohort study 
Early wheezing and asthma are relevant health problems in the tropics. Mite sensitization is an important risk factor, but the roles of others, inherent in poverty, are unknown. We designed a birth-cohort study in Cartagena (Colombia) to investigate genetic and environmental risk factors for asthma and atopy, considering as particular features perennial exposure to mites, parasite infections and poor living conditions.
Pregnant women representative of the low-income suburbs of the city were randomly screened for eligibility at delivery; 326 mother-infant pairs were included at baseline and biological samples were collected from birth to 24 months for immunological testing, molecular genetics and gene expression analysis. Pre and post-natal information was collected using questionnaires.
94% of families were from the poorest communes of the city, 40% lacked sewage and 11% tap-water. Intestinal parasites were found as early as 3 months; by the second year, 37.9% of children have had parasites and 5.22% detectable eggs of Ascaris lumbricoides in stools (Median 3458 epg, IQR 975-9256). The prevalence of "wheezing ever" was 17.5% at 6 months, 31.1% at 12 months and 38.3% at 24 months; and recurrent wheezing (3 or more episodes) 7.1% at 12 months and 14.2% at 24 months. Maternal rhinitis [aOR 3.03 (95%CI 1.60-5.74), p = 0.001] and male gender [aOR 2.09 (95%CI 1.09 - 4.01), p = 0.026], increased risk for wheezing at 6 months. At 24 months, maternal asthma was the main predisposing factor for wheezing [aOR 3.65 (95%CI 1.23-10.8), p = 0.01]. Clinical symptoms of milk/egg allergy or other food-induced allergies were scarce (1.8%) and no case of atopic eczema was observed.
Wheezing is the most frequent phenotype during the first 24 months of life and is strongly associated with maternal asthma. At 24 months, the natural history of allergic symptoms is different to the "atopic march" described in some industrialized countries. This cohort is representative of socially deprived urban areas of underdeveloped tropical countries. The collection of biological samples, data on exposure and defined phenotypes, will contribute to understand the gene/environment interactions leading to allergy inception and evolution.
PMCID: PMC3331807  PMID: 22439773
Birth cohort study; Wheezing; Allergy; Asthma; Rhinitis; Eczema; Atopic march; The tropics; Parasite; Poverty; Cartagena; Latin America
17.  Differences in the expression of five senescence markers in oral cancer, oral leukoplakia and control samples in humans 
Oncology Letters  2012;3(6):1319-1325.
Oncogene-induced senescence (OIS) may be a response to oncogenic activation, acting as a natural barrier against carcinogenesis at a premalignant stage. Thus, numerous cells in premalignant lesions enter senescence, but none or few in malignant tumours. This event could be due to the loss of senescence pathway effectors, including p16 (INK4a)-pRb or ARF-p53. The aim of this study was to characterize and compare the expression of certain senescent markers between oral precancer and cancer tissue samples. The expression of cyclin D1, Rb, maspin, p53 and mouse double minute 2 (MDM2) was analyzed in 20 paraffin-embedded tissue samples of normal oral mucosa (NOM), 14 samples of oral leukoplakia without dysplasia (OLD−), 11 samples of leukoplakia with dysplasia (OLD+) and 15 samples of oral squamous cell carcinoma (OSCC) by immunohistochemistry in tissue arrays. The expression of p16-pRb pathway markers, cyclin D1, maspin and Rb, was more frequent in OLD+ samples than in OSCC samples, although a statistical significance was only observed for maspin (P=0.036). Cyclin D1 expression was also significantly more frequent in OLD− samples vs. NOM samples. For the ARF-p53 pathway, the expression of p53 and MDM2 was significantly more frequent in the OLD− samples compared to in the NOM ones. These findings may indicate a role for cellular senescence in oral carcinogenesis, considering maspin as a reliable senescence marker and prognostic factor in oral premalignant lesions.
PMCID: PMC3392562  PMID: 22783442
senescence; oral cancer; oral leukoplakia; cyclin D1; maspin; Rb; p53; MDM2
18.  Low-Replicating Viruses and Strong Anti-Viral Immune Response Associated with Prolonged Disease Control in a Superinfected HIV-1 LTNP Elite Controller 
PLoS ONE  2012;7(2):e31928.
To study the causes for the lack of clinical progression in a superinfected HIV-1 LTNP elite controller patient.
Methodology and Principal Findings
We studied host genetic, virological and immunological factors associated with viral control in a SI long term non progressor elite controller (LTNP-EC). The individual contained both viruses and maintained undetectable viral loads for >20 years and he did not express any of the described host genetic polymorphisms associated with viral control. None of four full-length gp160 recombinants derived from the LTNP-EC replicated in heterologous peripheral blood mononuclear cells. CTL responses after SI were maintained in two samples separated by 9 years and they were higher in breadth and magnitude than responses seen in most of 250 treatment naïve patients and also 25 controller subjects. The LTNP-EC showed a neutralization response, against 4 of the 6 viruses analyzed, superior to other ECs.
The study demonstrated that a strong and sustained cellular and humoral immune response and low replicating viruses are associated with viral control in the superinfected LTNP-EC.
PMCID: PMC3286446  PMID: 22384103
19.  Male Circumcision and Risk of HIV Acquisition among Men who have Sex with Men from the United States and Peru 
AIDS (London, England)  2011;25(4):519-523.
To assess the association between male circumcision, insertive anal sex practices, and HIV acquisition in a cohort of men who have sex with men (MSM).
Data were from 1824 HSV-2 seropositive, HIV seronegative MSM, 1362 (75%) from Peru and 462 (25%) from the US, who participated in a randomized placebo controlled trial of HSV-2 suppression for HIV prevention (HPTN 039). Circumcision status was determined by examination at enrollment. HIV testing was done every three months for up to 18 months. Partner-specific sexual behavior for up to the last three partners during the previous three months was analyzed.
There was no significant association between male circumcision and HIV acquisition in univariate analysis (RR=0.84, 95% CI 0.50–1.42). In a pre-specified multivariate analysis that assumed a linear relationship between the proportion of insertive acts and effect of circumcision on HIV acquisition, the interaction between circumcision and proportion of insertive acts was not significant (p=0.11). In an exploratory analysis that categorized behavior with recent partners by proportion of insertive acts (<60% or ≥60% insertive acts), circumcision was associated with a non-statistically significant 69% reduction in the risk of HIV acquisition (RR=0.31, 95% CI 0.06–1.51) among men who reported ≥60% of insertive acts with recent male partners.
Circumcision does not have a significant protective effect against HIV acquisition among MSM from Peru and US, although there may be reduced risk for men who are primarily insertive with their male partners. This association needs to be investigated across diverse cohorts of MSM.
PMCID: PMC3120051  PMID: 21099672
HIV acquisition; male circumcision; men who have sex with men
20.  Density Functional Theory (DFT) Study of Triphenylamine-Based Dyes for Their Use as Sensitizers in Molecular Photovoltaics 
In this work we studied three dyes which are proposed for potential photovoltaic applications and named Dye7, Dye7-2t and Dye7-3t. The Density Functional Theory (DFT) was utilized, using the M05-2X hybrid meta-GGA functional and the 6–31+G(d,p) basis set. This level of calculation was used to find the optimized molecular structure and to predict the main molecular vibrations, the absorption and emission spectra, the molecular orbitals energies, dipole moment, isotropic polarizability and the chemical reactivity parameters that arise from Conceptual DFT. Also, the pKa values were calculated with the semi-empirical PM6 method.
PMCID: PMC3344223  PMID: 22605987
molecular structure; absorption spectrum; polarizability; chemical reactivity; dipole moment; triphenylamine; dye sensitizers
21.  Virtual Sensors for Designing Irrigation Controllers in Greenhouses 
Sensors (Basel, Switzerland)  2012;12(11):15244-15266.
Monitoring the greenhouse transpiration for control purposes is currently a difficult task. The absence of affordable sensors that provide continuous transpiration measurements motivates the use of estimators. In the case of tomato crops, the availability of estimators allows the design of automatic fertirrigation (irrigation + fertilization) schemes in greenhouses, minimizing the dispensed water while fulfilling crop needs. This paper shows how system identification techniques can be applied to obtain nonlinear virtual sensors for estimating transpiration. The greenhouse used for this study is equipped with a microlysimeter, which allows one to continuously sample the transpiration values. While the microlysimeter is an advantageous piece of equipment for research, it is also expensive and requires maintenance. This paper presents the design and development of a virtual sensor to model the crop transpiration, hence avoiding the use of this kind of expensive sensor. The resulting virtual sensor is obtained by dynamical system identification techniques based on regressors taken from variables typically found in a greenhouse, such as global radiation and vapor pressure deficit. The virtual sensor is thus based on empirical data. In this paper, some effort has been made to eliminate some problems associated with grey-box models: advance phenomenon and overestimation. The results are tested with real data and compared with other approaches. Better results are obtained with the use of nonlinear Black-box virtual sensors. This sensor is based on global radiation and vapor pressure deficit (VPD) measurements. Predictive results for the three models are developed for comparative purposes.
PMCID: PMC3522961  PMID: 23202208
virtual sensor; transpiration; nonlinear model; micro-lysimeter
22.  Computational Molecular Nanoscience Study of the Properties of Copper Complexes for Dye-Sensitized Solar Cells 
In this work, we studied a copper complex-based dye, which is proposed for potential photovoltaic applications and is named Cu (I) biquinoline dye. Results of electron affinities and ionization potentials have been used for the correlation between different levels of calculation used in this study, which are based on The Density Functional Theory (DFT) and time-dependent (TD) DFT. Further, the maximum absorption wavelengths of our theoretical calculations were compared with the experimental data. It was found that the M06/LANL2DZ + DZVP level of calculation provides the best approximation. This level of calculation was used to find the optimized molecular structure and to predict the main molecular vibrations, the molecular orbitals energies, dipole moment, isotropic polarizability and the chemical reactivity parameters that arise from Conceptual DFT.
PMCID: PMC3546675  PMID: 23443107
molecular structure; absorption spectra; polarizability; chemical reactivity; dipole moment; copper complex; dye-sensitized
Vaginosis bacteriana (VB) es una infección caracterizada por el cambio en la microflora de la vagina, asociándose a resultados adversos del embarazo y a la adquisición de infecciones de transmisión sexual (ITS), incluyendo el VIH. En este estudio se buscó la asociación entre el uso de duchas vaginales y otros factores de riesgos con VB.
Se usó un diseño observacional descriptivo transversal prospectivo, en 1,252 mujeres que asistieron al servicio de planificación familiar de tres hospitales nacionales (Dos de Mayo, Arzobispo Loayza, San Bartolomé) y el Instituto Materno Perinatal, durante el año 1997. Se utilizó un cuestionario estructurado donde se registraron variables socio demográficas y características del estilo de vida de las participantes. VB fue diagnosticada mediante el puntaje de Nugent. Se empleó análisis de regresión logística para calcular odds ratio (OR) e intervalos de confianza al 95%.
La edad promedio de las participantes fue 25.1 ± 4,7 años, el 23.4% tenían más de 11 años de educación. La prevalencía de VB fue 20,1%. Las mujeres que practicaban duchas vaginales tuvieron 2.28 veces (OR = 2.28, IC 95% [1.0–5.0]) mayor probabilidad de tener VB comparado con aquellas que no lo practicaban. Tener dos o más parejas sexuales estuvo asociado con 2.0 veces (OR =2.0, IC 95% [1.2–3.5]) mayor probabilidad de adquirir VB comparado con aquellas que habían tenido solo una pareja sexual. Las participantes que iniciaron una relación sexual a una edad temprana tuvieron 1.4 veces (OR=1.4, IC 95% [1.0 –1.9]) mayor probabilidad de adquirir VB.
El uso de duchas vaginales es un factor de riesgo de VB. Los programas destinados a la salud de la mujer deben abordar las repercusiones perjudiciales para la salud asociados con las duchas vaginales.
PMCID: PMC2995200  PMID: 21132048
Vaginosis bacteriana; duchas vaginales; parejas sexuales; edad a la primera relación sexual; factores de riesgo
24.  Use of the Yeast Pichia pastoris as an Expression Host for Secretion of Enterocin L50, a Leaderless Two-Peptide (L50A and L50B) Bacteriocin from Enterococcus faecium L50▿  
Applied and Environmental Microbiology  2010;76(10):3314-3324.
In this work, we report the expression and secretion of the leaderless two-peptide (EntL50A and EntL50B) bacteriocin enterocin L50 from Enterococcus faecium L50 by the methylotrophic yeast Pichia pastoris X-33. The bacteriocin structural genes entL50A and entL50B were fused to the Saccharomyces cerevisiae gene region encoding the mating pheromone α-factor 1 secretion signal (MFα1s) and cloned, separately and together (entL50AB), into the P. pastoris expression and secretion vector pPICZαA, which contains the methanol-inducible alcohol oxidase promoter (PAOX1) to express the fusion genes. After transfer into the yeast, the recombinant plasmids were integrated into the genome, resulting in three bacteriocinogenic yeast strains able to produce and secrete the individual bacteriocin peptides EntL50A and EntL50B separately and together. The secretion was efficiently directed by MFα1s through the Sec system, and the precursor peptides were found to be correctly processed to form mature and active bacteriocin peptides. The present work describes for the first time the heterologous expression and secretion of a two-peptide non-pediocin-like bacteriocin by a yeast.
PMCID: PMC2869124  PMID: 20348300
25.  HSV suppression reduces seminal HIV-1 levels in HIV-1/HSV-2 coinfected men who have sex with men (MSM) 
AIDS (London, England)  2009;23(4):479-483.
Suppressive HSV therapy can decrease plasma, cervical, and rectal HIV-1 levels in HSV-2/HIV-1 co-infected persons. We evaluated the effect of HSV-2 suppression on seminal HIV-1 levels.
Twenty antiretroviral therapy (ART)–naive HIV-1/HSV-2–MSM in Lima, Peru, with CD4 cell counts >200 cells/µL randomly received valacyclovir 500 mg twice daily or placebo for 8 weeks, then the alternative regimen for 8 weeks after a 2 week washout. Peripheral blood and semen specimens were collected weekly. Anogenital swab specimens for HSV DNA were self-collected daily and during clinic visits.
HIV-1 RNA was quantified in seminal and blood plasma by TaqMan real-time polymerase chain reaction (rt-PCR) or Roche Amplicor Monitor assays. HSV and seminal cytomegalovirus (CMV) were quantified by rt-PCR. Linear mixed models examined differences within subjects by treatment arm.
Median CD4 count of participants was 424 cells/µL. HIV-1 was detected in 71% of 231 semen specimens. HSV was detected from 29% and 4.4% of swabs on placebo and valacyclovir, respectively (P<0.001). Valacyclovir significantly reduced the proportion of days with detectable seminal HIV-1 (63% during valacyclovir vs. 78% during placebo, p=0.04). The quantity of HIV-1 in semen was 0.25 log10 copies/mL lower (95%CI −0.40 to −0.10, p=0.001) during the valacyclovir arm compared with placebo, a 44% reduction. CD4 count (p=0.32) and seminal cellular CMV quantity (p=0.68) did not predict seminal plasma HIV-1 level.
Suppressive valacyclovir reduced seminal HIV-1 levels in HIV-1/HSV-2 co-infected MSM not receiving ART. The significance of this finding will be evaluated in a trial with HIV-1 transmission as the outcome.
PMCID: PMC2707902  PMID: 19169140
seminal plasma; HIV-1; viral load; HSV-2; valacyclovir; herpes; blood plasma

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