Hyperspectral Sensing of Disease Stress in the Caribbean Reef-Building Coral,Orbicella faveolata - Perspectives for the Field of Coral Disease Monitoring

The effectiveness of management plans developed for responding to coral disease outbreaks is limited due to the lack of rapid methods of disease diagnosis. In order to fulfill current management guidelines for responding to coral disease outbreaks, alternative methods that significantly reduce response time must be developed. Hyperspectral sensing has been used by various groups to characterize the spectral signatures unique to asymptomatic and bleached corals. The 2010 combined bleaching and Caribbean yellow band disease outbreak in Puerto Rico provided a unique opportunity to investigate the spectral signatures associated with bleached and Caribbean yellow band-diseased colonies of Orbicella faveolata for the first time. Using derivative and cluster analyses of hyperspectral reflectance data, the present study demonstrates the proof of concept that spectral signatures can be used to differentiate between coral disease states. This method enhanced predominant visual methods of diagnosis by distinguishing between different asymptomatic conditions that are identical in field observations and photographic records. The ability to identify disease-affected tissue before lesions become visible could greatly reduce response times to coral disease outbreaks in monitoring efforts. Finally, spectral signatures associated with the poorly understood Caribbean yellow band disease are presented to guide future research on the role of pigments in the etiology.     

Induction and Analysis of Epithelial to Mesenchymal Transition

Epithelial to mesenchymal transition (EMT) is essential for proper morphogenesis during development. Misregulation of this process has been implicated as a key event in fibrosis and the progression of carcinomas to a metastatic state. Understanding the processes that underlie EMT is imperative for the early diagnosis and clinical control of these disease states. Reliable induction of EMT in vitro is a useful tool for drug discovery as well as to identify common gene expression signatures for diagnostic purposes. Here we demonstrate a straightforward method for the induction of EMT in a variety of cell types. Methods for the analysis of cells pre- and post-EMT induction by immunocytochemistry are also included. Additionally, we demonstrate the effectiveness of this method through antibody-based array analysis and migration/invasion assays.     

Frataxin from Psychromonas ingrahamii as a model to study stability modulation within the CyaY protein family

Adaptation of life to low temperatures influences both protein stability and flexibility. Thus, proteins from psychrophilic organisms are excellent models to study relations between these properties. Here we focused on frataxin from Psychromonas ingrahamii (pFXN), an extreme psychrophilic sea ice bacterium that can grow at temperatures as low as − 12 °C. This α/β protein is highly conserved and plays a key role in iron homeostasis as an iron chaperone. In contrast to other frataxin homologs, chemical and temperature unfolding experiments showed that the thermodynamic stability of pFXN is strongly modulated by pHs: ranging from 5.5 ± 0.9 (pH 6.0) to 0.9 ± 0.3 kcal mol^− 1 (pH 8.0). This protein was crystallized and its X-ray structure solved at 1.45 Å. Comparison of B-factor profiles between Escherichia coli and P. ingrahamii frataxin variants (51% of identity) suggests that, although both proteins share the same structural features, their flexibility distribution is different. Molecular dynamics simulations showed that protonation of His44 or His67 in pFXN lowers the mobility of regions encompassing residues 20–30 and the C-terminal end, probably through favorable electrostatic interactions with residues Asp27, Glu42 and Glu99. Since the C-terminal end of the protein is critical for the stabilization of the frataxin fold, the predictions presented may be reporting on the microscopic origin of the decrease in global stability produced near neutral pH in the psychrophilic variant. We propose that suboptimal electrostatic interactions may have been an evolutionary strategy for the adaptation of frataxin flexibility and function to cold environments.     

The Role of Vesicle Trafficking and Release in Oligodendrocyte Biology

Oligodendrocytes are a subtype of glial cells found within the central nervous system (CNS), responsible for the formation and maintenance of specialized myelin membranes which wrap neuronal axons. The development of myelin requires tight coordination for the cell to deliver lipid and protein building blocks to specific myelin segments at the right time. Both internal and external cues control myelination, thus the reception of these signals also requires precise regulation. In late years, a growing body of evidence indicates that oligodendrocytes, like many other cell types, may use extracellular vesicles (EVs) as a medium for transferring information. The field of EV research has expanded rapidly over the past decade, with new contributions that suggest EVs might have direct involvement in communications with neurons and other glial cells to fine tune oligodendroglial function. This functional role of EVs might also be maladaptive, as it has likewise been implicated in the spreading of toxic molecules within the brain during disease. In this review we will discuss the field’s current understanding of extracellular vesicle biology within oligodendrocytes, and their contribution to physiologic and pathologic conditions.

     

A retrospective study of DNA prevalence of porcine parvoviruses in Mexico and its relationship with porcine circovirus associated disease

Worldwide, many emerging porcine parvoviruses (PPVs) have been linked to porcine circovirus-2 (PCV2) associated disease (PCVAD), which includes post-weaning multi-systemic wasting syndrome (PMWS), PCV2-related reproductive failure (PCV2-RF), as well as other syndromes. To determine the DNA prevalence of PPVs and their relationship with PMWS and PCV2-RF in Mexico, 170 formalin-fixed paraffin-embedded tissues were selected from archival collections to detect PPVs using a nested polymerase chain reaction. The tissues were composed of 50 PMWS cases, 20 age-matched tissues from healthy pigs, 56 PCV2-related reproductive failure (PCV2⁺-RF) cases, and 44 PCV2^--RF cases. Overall, PPV2 and PPV6 were the most prevalent species (90.0% and 74.7%, respectively). In 8–11 week old pigs, the highest prevalence was for PPV6 and PPV3. Concerning reproductive failure, the PCV2-affected farms had a significantly higher prevalence for PPV6 (61.6%) and PPV5 (36.4%) than the PCV2-unaffected farms (35.0% and 5.0%, respectively). The concurrent infection rate was high, being significant for PPV2/PPV4 and PPV1/PPV5 within the PMWS cases and for PPV6/PPV5 among the PCV2⁺-RF tissues. PPV5 showed a significant relationship with PMWS, whereas PPV5 and PPV6 were significant for PCVAD. The prevalence and coinfection rate of PPVs in Mexico were markedly higher than that described in other countries, denoting that PPV5 and PPV6 might have a potential role in PCVAD in Mexico. It is concluded that it is likely that the density population of pigs in Mexico is contributing to high PPV inter-species and PCV2 coinfections which might lead to a different pathogenic outcome.     

Pseudomembranous Candidiasis in HIV/AIDS Patients in Cali, Colombia

Candida albicans is the most frequently isolated yeast from the oral cavity of HIV/AIDS individuals. The use of fluconazole has increased the number of resistant or less-sensitive Candida species different than C. albicans. The purpose of this study was to identify the Candida species producing pseudomembranous candidiasis in patients suffering from AIDS, their relationship with CD4⁺ counts and their sensitivity to fluconazole and itraconazole. We studied 71 patients at a hospital in the city of Cali. Samples of white plaque were seeded on CHROMagar Candida, yeast identification was done with API 20C Aux, and susceptibility testing was determined by E test. Ninety-three yeast isolates were obtained, 52 single and 41 mixed. C. albicans was the most isolated, followed by C. glabrata. An increased frequency of isolates and variety of Candida species occurred in patients with a CD4⁺ cell count ≤100 cells/mm³ without significant differences (p = 0.29). The susceptibility study showed that 8 (8.6 %) isolates were resistant to fluconazole and 11 (11.8 %) to itraconazole, while 6 (8.8 %) C. albicans were simultaneously resistant. No association was found between the isolates of C. albicans or Candida species different than C. albicans and the use of fluconazole (p = 0.21). The results of this study indicate that in the tested population, fluconazole continues to be the best treatment option for oropharyngeal candidiasis in patients suffering from AIDS (HIV/AIDS); however, susceptibility tests are necessary in patients who present therapeutic failure.     

DNA Topological Context Affects Access to Eukaryotic DNA Topoisomerase I

Abstract

We have analyzed the reactivity of a 217 base pair segment of the intrinsically curved Crithidia fasciculata kinetoplast DNA towards eukaryotic DNA topoisomerase I. The substrates were open [linear fragment and nicked circle] and closed minidomains [closed relaxed circle and circles with linking differences of ?1 and ?2], We interpreted the results with the aid of a model that was used to predict the structures of the topoisomers. The modelling shows that the ΔLk(?l) form is unusually compact because of the curvature in the DNA. To determine the role of sequence-directed curvature in both the experimental and modeling studies, controls were examined in which the curved Crithidia sequence was replaced by an uncurved sequence obtained from the plasmid pBR322.

Reactivity of the Crithidia DNA [as analyzed both by the cleavage and the topoisomerization reactions] markedly varied among the DNA forms: (i) the hierarchy of overall reactivity observed is: linear fragment > nicked circular, closed circular [ΔLk(O)], interwound [ΔLk(?2)] > bent interwound [ΔLk(?l)]; (ii) the intensity of several cleavage positions differs among DNA forms.

The results show that eukaryotic DNA topoisomerase I is very sensitive to the conformation of the substrates and that its reactivity is modulated by the variation of the compactness of the DNA molecule. The C. fasciculata sequence contains a highly curved segment that determines the conformation of the closed circle in a complex way.     

Attenuated and Replication-Competent Vaccinia Virus Strains M65 and M101 with Distinct Biology and Immunogenicity as Potential Vaccine Candidates against Pathogens

Replication-competent poxvirus vectors with an attenuation phenotype and with a high immunogenic capacity of the foreign expressed antigen are being pursued as novel vaccine vectors against different pathogens. In this investigation, we have examined the replication and immunogenic characteristics of two vaccinia virus (VACV) mutants, M65 and M101. These mutants were generated after 65 and 101 serial passages of persistently infected Friend erythroleukemia (FEL) cells. In cultured cells of different origins, the mutants are replication competent and have growth kinetics similar to or slightly reduced in comparison with those of the parental Western Reserve (WR) virus strain. In normal and immune-suppressed infected mice, the mutants showed different levels of attenuation and pathogenicity in comparison with WR and modified vaccinia Ankara (MVA) strains. Wide genome analysis after deep sequencing revealed selected genomic deletions and mutations in a number of viral open reading frames (ORFs). Mice immunized in a DNA prime/mutant boost regimen with viral vectors expressing the LACK (Leishmania homologue for receptors of activated C kinase) antigen of Leishmania infantum showed protection or a delay in the onset of cutaneous leishmaniasis. Protection was similar to that triggered by MVA-LACK. In immunized mice, both polyfunctional CD4⁺ and CD8⁺ T cells with an effector memory phenotype were activated by the two mutants, but the DNA-LACK/M65-LACK protocol preferentially induced CD4⁺ whereas DNA-LACK/M101-LACK preferentially induced CD8⁺ T cell responses. Altogether, our findings showed the adaptive changes of the WR genome during long-term virus-host cell interaction and how the replication competency of M65 and M101 mutants confers distinct biological properties and immunogenicity in mice compared to those of the MVA strain. These mutants could have applicability for understanding VACV biology and as potential vaccine vectors against pathogens and tumors.     

Morphometric and molecular variation in concert: taxonomy and genetics of the reticulate Pyrenean and Iberian alpine spiny fescues (Festuca eskia complex,Poaceae)

The Iberian mountain spiny fescues are a reticulate group of five diploid grass taxa consisting of three parental species and two putative hybrids: F. × souliei (F. eskia × F. quadriflora) and F. × picoeuropeana (F. eskia × F. gautieri). Phenotypic and molecular studies were conducted with the aim of determining the taxonomic boundaries and genetic relationships of the five taxa and disentangling the origins of the two hybrids. Statistical analyses of 31 selected phenotypic traits were conducted on individuals from 159 populations and on nine type specimens. Molecular analyses of random amplified polymorphic DNA (RAPD) markers were performed on 29 populations. The phenotypic analyses detected significant differences between the five taxa and demonstrated the overall intermediacy of the F. × picoeuropeana and F. × souliei between their respective parents. The RAPD analysis corroborated the genetic differentiation of F. eskia, F. gautieri and F. quadriflora and the intermediate nature of the two hybrids; however, they also detected genetic variation within F. × picoeuropeana. These results suggest distinct origins for F. × picoeuropeana in the Cantabrian and Pyrenean mountains, with the sporadic Pyrenean populations having potentially resulted from recent hybridizations and the stabilized Cantabrian ones from older events followed by potential displacements of the parents. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 173 , 676–706.