Immunomodulator-Based Enhancement of Anti Smallpox Immune Responses

Background

The current live vaccinia virus vaccine used in the prevention of smallpox is contraindicated for millions of immune-compromised individuals. Although vaccination with the current smallpox vaccine produces protective immunity, it might result in mild to serious health complications for some vaccinees. Thus, there is a critical need for the production of a safe virus-free vaccine against smallpox that is available to everyone. For that reason, we investigated the impact of imiquimod and resiquimod (Toll-like receptors agonists), and the codon-usage optimization of the vaccinia virus A27L gene in the enhancement of the immune response, with intent of producing a safe, virus-free DNA vaccine coding for the A27 vaccinia virus protein.

Methods

We analyzed the cellular-immune response by measuring the IFN-γ production of splenocytes by ELISPOT, the humoral-immune responses measuring total IgG and IgG2a/IgG1 ratios by ELISA, and the TH1 and TH2 cytokine profiles by ELISA, in mice immunized with our vaccine formulation.

Results

The proposed vaccine formulation enhanced the A27L vaccine-mediated production of IFN-γ on mouse spleens, and increased the humoral immunity with a TH1-biased response. Also, our vaccine induced a TH1 cytokine milieu, which is important against viral infections.

Conclusion

These results support the efforts to find a new mechanism to enhance an immune response against smallpox, through the implementation of a safe, virus-free DNA vaccination platform.     

Incidence and Risk Factors for Respiratory Syncytial Virus and Human Metapneumovirus Infections among Children in the Remote Highlands of Peru

Introduction

The disease burden and risk factors for respiratory syncytial virus (RSV) and human metapneumovirus (MPV) infections among children living in remote, rural areas remain unclear.

Materials and Methods

We conducted a prospective, household-based cohort study of children aged <3 years living in remote rural highland communities in San Marcos, Cajamarca, Peru. Acute respiratory illnesses (ARI), including lower respiratory tract infection (LRTI), were monitored through weekly household visits from March 2009 through September 2011. Nasal swabs collected during ARI/LRTI were tested for RSV, MPV, and other respiratory viruses using real-time RT-PCR. Incidence rates and rate ratios were calculated using mixed effects Poisson regression.

Results

Among 892 enrolled children, incidence rates of RSV and MPV ARI were 30 and 17 episodes per 100 child-years, respectively. The proportions of RSV and MPV ARI that presented as LRTI were 12.5% and 8.9%, respectively. Clinic visits for ARI and hospitalizations were significantly more frequent (all p values <0.05) among children with RSV (clinic 41% and hospital 5.3%) and MPV ARI (38% and 3.5%) when compared with other viral infections (23% and 0.7%) and infections without virus detected (24% and 0.6%). In multivariable analysis, risk factors for RSV detection included younger age (RR 1.02, 95% CI: 1.00-1.03), the presence of a smoker in the house (RR 1.63, 95% CI: 1.12-2.38), residing at higher altitudes (RR 1.93, 95% CI: 1.25-3.00 for 2^nd compared to 1^st quartile residents; RR 1.98, 95% CI: 1.26-3.13 for 3^rd compared to 1^st quartile residents). Having an unemployed household head was significantly associated with MPV risk (RR 2.11, 95% CI: 1.12-4.01).

Conclusion

In rural high altitude communities in Peru, childhood ARI due to RSV or MPV were common and associated with higher morbidity than ARI due to other viruses or with no viral detections. The risk factors identified in this study may be considered for interventional studies to control infections by these viruses among young children from developing countries.     

Molecular and Phenotypic Characterization of Staphylococcus epidermidis Isolates from Healthy Conjunctiva and a Comparative Analysis with Isolates from Ocular Infection

Staphylococcus epidermidis is a common commensal of healthy conjunctiva and it can cause endophthalmitis, however its presence in conjunctivitis, keratitis and blepharitis is unknown. Molecular genotyping of S. epidermidis from healthy conjunctiva could provide information about the origin of the strains that infect the eye. In this paper two collections of S. epidermidis were used: one from ocular infection (n = 62), and another from healthy conjunctiva (n = 45). All isolates were genotyped by pulsed field gel electrophoresis (PFGE), multilocus sequence typing (MLST), staphylococcal cassette chromosome mec (SCCmec), detection of the genes icaA, icaD, IS256 and polymorphism type of agr locus. The phenotypic data included biofilm production and antibiotic resistance. The results displayed 61 PFGE types from 107 isolates and they were highly discriminatory. MLST analysis generated a total of 25 STs, of which 11 STs were distributed among the ocular infection isolates and lineage ST2 was the most frequent (48.4%), while 14 STs were present in the healthy conjunctiva isolates and lineage ST5 was the most abundant (24.4%). By means of a principal coordinates analysis (PCoA) and a discriminant analysis (DA) it was found that ocular infection isolates had as discriminant markers agr III or agr II, SCCmec V or SCCmec I, mecA gene, resistance to tobramycin, positive biofilm, and IS256⁺. In contrast to the healthy conjunctiva isolates, the discriminating markers were agr I, and resistance to chloramphenicol, ciprofloxacin, gatifloxacin and oxacillin. The discriminant biomarkers of ocular infection were examined in healthy conjunctiva isolates, and it was found that 3 healthy conjunctiva isolates [two with ST2 and another with ST9] (3/45, 6.66%) had similar genotypic and phenotypic characteristics to ocular infection isolates, therefore a small population from healthy conjunctiva could cause an ocular infection. These data suggest that the healthy conjunctiva isolates do not, in almost all cases, infect the eye due to their large genotypic and phenotypic difference with the ocular infection isolates.     

How Well Does the World Health Organization Definition of Domestic Violence Work for India?

Domestic violence (DV) is reported by 40% of married women in India and associated with substantial morbidity. An operational research definition is therefore needed to enhance understanding of DV epidemiology in India and inform DV interventions and measures. To arrive at a culturally-tailored definition, we aimed to better understand how definitions provided by the World Health Organization and the 2005 India Protection of Women from Domestic Violence Act match the perceptions of behaviors constituting DV among the Indian community. Between September 2012 and January 2013, 16 key informant interviews with experts in DV and family counseling and 2 gender-concordant focus groups of lay community members were conducted in Pune, India to understand community perceptions of the definition of DV, perpetrators of DV, and examples of DV encountered by married women in Pune, India. Several key themes emerged regarding behaviors and acts constituting DV including 1) the exertion of control over a woman’s reproductive decision-making, mobility, socializing with family and friends, finances, and access to food and nutrition, 2) the widespread acceptance of sexual abuse and the influences of affluence on sexual DV manifestations, 3) the shaping of physical abuse experiences by readily-available tools and the presence of witnesses, 4) psychological abuse for infertility, dowry, and girl-children, and 5) the perpetration of DV by the husband and other members of his family. Findings support the need for a culturally-tailored operational definition that expands on the WHO surveillance definition to inform the development of more effective DV intervention strategies and measures.     

Bioarchaeological investigation of ancient Maya violence and warfare in inland northwest Yucatan,Mexico

This study investigates evidence of changes and continuities in ancient Maya violence and warfare in inland northwest Yucatan, Mexico from the Middle Preclassic (600–300 BC) to the Postclassic (AD 1050–1542) through bioarchaeological analysis of cranial and projectile trauma. It is hypothesized that the frequency of violence increases before the Classic Maya collapse and remains high during the Postclassic period. It is also hypothesized that the flat, open terrain was conducive to warfare and resulted in higher trauma frequencies than in other parts of the Maya area. Results show that the frequency of cranial trauma decreases before the Classic collapse and increases in the Postclassic, partially matching the expected chronological trends. The frequency of cranial trauma does not differ significantly from other Maya regions but the pattern does: for all periods, males have more healed injuries than females and they are concentrated on the left side of the anterior of the skull. Some injuries appear to be from small points hafted in wooden clubs. In addition, projectile trauma is evident in a scapula with an embedded arrowhead tip, the first such case reported in a Maya skeleton. Overall, these results suggest greater reliance on open combat and less on raids in this region compared with other parts of the Maya area, possibly due to the flat, open terrain, though the identification of perimortem trauma in both women and men indicates surprise raids on settlements were also practiced. Am J Phys Anthropol 154:140–151, 2014. © 2014 Wiley Periodicals, Inc.     

Two-dimensional gel electrophoresis image registration using block-matching techniques and deformation models

Block-matching techniques have been widely used in the task of estimating displacement in medical images, and they represent the best approach in scenes with deformable structures such as tissues, fluids, and gels. In this article, a new iterative block-matching technique—based on successive deformation, search, fitting, filtering, and interpolation stages—is proposed to measure elastic displacements in two-dimensional polyacrylamide gel electrophoresis (2D–PAGE) images. The proposed technique uses different deformation models in the task of correlating proteins in real 2D electrophoresis gel images, obtaining an accuracy of 96.6% and improving the results obtained with other techniques. This technique represents a general solution, being easy to adapt to different 2D deformable cases and providing an experimental reference for block-matching algorithms.     

Characterization of genomic single nucleotide polymorphism via colorimetric detection using a single gold nanoprobe

Identification of specific nucleic acid sequences mediated by gold nanoparticles derivatized thiol-modified oligonucleotides (Au–nanoprobes) has been proven to be a useful tool in molecular diagnostics. Here, we demonstrate that, on optimization, detection may be simplified via the use of a single Au–nanoprobe to detect a single nucleotide polymorphism (SNP) in homo- or heterozygote condition. We validated this non-cross-linking approach through the analysis of 20 clinical samples using a single specific Au–nanoprobe for an SNP in the FTO (fat mass and obesity-associated) gene against direct DNA sequencing. Sensitivity, specificity, and limit of detection (LOD) were determined, and statistical differences were calculated by one-way analysis of variance (ANOVA) and a post hoc Tukey’s test to ascertain whether there were any differences between Au–nanoprobe genotyped groups. For the first time, we show that the use of a single Au–nanoprobe can detect SNP for each genetic status (wild type, heterozygous, or mutant) with high degrees of sensitivity (87.50%) and specificity (91.67%).     

Homo- and heterofermentative lactobacilli differently affect sugarcane-based fuel ethanol fermentation

Bacterial contamination during industrial yeast fermentation has serious economic consequences for fuel ethanol producers. In addition to deviating carbon away from ethanol formation, bacterial cells and their metabolites often have a detrimental effect on yeast fermentative performance. The bacterial contaminants are commonly lactic acid bacteria (LAB), comprising both homo- and heterofermentative strains. We have studied the effects of these two different types of bacteria upon yeast fermentative performance, particularly in connection with sugarcane-based fuel ethanol fermentation process. Homofermentative Lactobacillus plantarum was found to be more detrimental to an industrial yeast strain (Saccharomyces cerevisiae CAT-1), when compared with heterofermentative Lactobacillus fermentum, in terms of reduced yeast viability and ethanol formation, presumably due to the higher titres of lactic acid in the growth medium. These effects were only noticed when bacteria and yeast were inoculated in equal cell numbers. However, when simulating industrial fuel ethanol conditions, as conducted in Brazil where high yeast cell densities and short fermentation time prevail, the heterofermentative strain was more deleterious than the homofermentative type, causing lower ethanol yield and out competing yeast cells during cell recycle. Yeast overproduction of glycerol was noticed only in the presence of the heterofermentative bacterium. Since the heterofermentative bacterium was shown to be more deleterious to yeast cells than the homofermentative strain, we believe our findings could stimulate the search for more strain-specific antimicrobial agents to treat bacterial contaminations during industrial ethanol fermentation.     

Gluconacetobacter diazotrophicus PAL5 possesses an active quorum sensing regulatory system

The endophytic bacterium Gluconacetobacter diazotrophicus colonizes a broad range of host plants. Its plant growth-promoting capability is related to the capacity to perform biological nitrogen fixation, the biosynthesis of siderophores, antimicrobial substances and the solubilization of mineral nutrients. Colonization of and survival in these endophytic niche requires a complex regulatory network. Among these, quorum sensing systems (QS) are signaling mechanisms involved in the control of several genes related to microbial interactions, host colonization and stress survival. G. diazotrophicus PAL5 possesses a QS composed of a luxR and a luxI homolog, and produces eight molecules from the AHL family as QS signals. In this report data are provided showing that glucose concentration modifies the relative levels of these signal molecules. The activity of G. diazotrophicus PAL5 QS is also altered in presence of other carbon sources and under saline stress conditions. Inactivation of the QS system of G. diazotrophicus PAL5 by means of a quorum quenching strategy allowed the identification of extracellular and intracellular proteins under the control of this regulatory mechanism.