bluuebunny/crossref_metadata_2025 huggingface dataset:
Is the taste acuity affected by oral dryness in primary Sjögren’s syndrome patients?
Authors: Al‐Ezzi Minan , Khan Khalid , Tappuni Anwar R.
Published: December 2019

Objectives

Taste disturbance is a symptom of primary Sjögren's syndrome (pSS) of unknown aetiology. This study's objectives were (a) to compare taste function in pSS vs. healthy subjects; (b) to establish whether there is an association between the taste acuity and oral dryness and/or the neurosensory threshold; and (c) to assess the impact of taste dysfunction on the quality of life (QoL).

Methodology

This study was conducted on 65 pSS females and 62 healthy volunteers. The gustatory function was tested with taste strips test. Visual analogue scale was used for self‐assessment of taste function. The electrogustometer was used to assess the neurosensory threshold. The oral dryness was assessed by the Clinical Oral Dryness Score, salivary flow rate and Xerostomia Inventory. The general and oral health‐related QoL and mental health well‐being were assessed using validated questionnaires.

Results

The pSS group had significantly impaired taste function, but this was not correlated with oral dryness. There was an association between taste acuity and the neurosensory threshold (β = −0.5, 95% CI = −0.2 to −0.1). The QoL was not impacted by taste dysfunction (p > .5).

Conclusion

The results suggest that taste impairment in pSS is associated with neurosensory dysfunction and is unlikely to be due to oral dryness.
Harnessing the Potential Synergistic Interplay Between Photosensitizer Dark Toxicity and Chemotherapy
Authors: Baglo Yan , Sorrin Aaron J. , Liang Barry J. , Huang Huang‐Chiao
Published: December 2019

The combination of photodynamic therapy and taxol‐ or platinum‐based chemotherapy (photochemotherapy) is an effective and promising cancer treatment. While the mechanisms of action of photochemotherapy are actively studied, relatively little is known about the cytotoxicity and molecular alterations induced by the combination of chemotherapy and photosensitizers without light activation in cancer cells. This study investigates the interplay between the photosensitizer benzoporphyrin derivative (BPD) without light activation and cisplatin or paclitaxel in two glioblastoma lines, U87 and U251. The combination effect of BPD and cisplatin in U87 cells is slightly synergistic (combination index, CI = 0.93), showing 1.8‐ to 2.6‐fold lower half‐maximal inhibitory concentrations (IC50) compared to those of individual drugs. In contrast, combining BPD and paclitaxel is slightly antagonistic (CI = 1.14) in U87 cells. In U251 cells, the combinations of BPD and cisplatin or paclitaxel are both antagonistic (CI = 1.24 and 1.34, respectively). Western blotting was performed to investigate changes in the expression levels of YAP, TAZ, Bcl‐2 and EGFR in U87 and U251 cells treated with BPD, cisplatin and paclitaxel, both as monotherapies and in combination. Our study provides insights into the molecular alterations in two glioma lines caused by each monotherapy and the combinations, in order to inform the design of effective treatments.
Re/Imagined Community: Neoliberalism, Human Rights, and Officials’ Accounts of the Salvadoran Transnation
Authors: Hallett Miranda Cady
Published: December 2019

Under neoliberal globalization, configurations of persons, legal regimes, and nation‐states transform. Some states explicitly employ a strategy of transnational nation‐building; for example, El Salvador is now an exemplary transnational state. With at least a quarter of the population living outside the national territory, the figure of the emigrant is significant in nationalist imaginaries. This article explores the process of imagining the transnation within one Salvadoran state institution—the Viceministry for Salvadorans Abroad—by examining upper‐and mid‐level state officials’ accounts of the relationship between emigrants and the nation‐state. State officials navigate and deploy global discourses around neoliberal development, on the one hand, and human rights for migrants, on the other hand, as they narrate their way to nationalist visions within the constraints of global political economy. The article looks comparatively at the ways officials frame the institution's central vision and role: under the right‐wing Alianza Republicana Nacionalista (ARENA) party administration in the late 2000s, and a few years later under the leadership of the leftist Frente Farabundo Martí para la Liberación Nacional (FMLN) party in the early 2010s. Under both administrations, viceministry officials navigated a fraught geopolitical field and worked diligently to engage the symbolic and material potentiality of cross‐border migrant citizens.
Hospitality's Prowess: Performing Sahrāwī Sovereignty in Refugee Camps
Authors: Solana Vivian
Published: December 2019

The Sahrāwī movement for national liberation—known as the Polisario Front—has been organizing into the Sahrāwī Arab Democratic Republic in refugee camps since 1976. This article describes rhetorical and administrative strategies that muster the labor of hospitality carried out in Sahrāwī households to advance the legitimacy of the Polisario Front. The khayma (tent, household, or family), a customary domain of female power and symbol of hospitality among the Sahrāwī (and beyond), bears a metonymic relationship with the project of a Sahrāwī revolutionary nationalism. Examining how hospitality serves as a nexus of articulation between different orders of the political, it argues that practices of hospitality among Sahrāwī refugees constitute the expression of a form of popular sovereignty and revolutionary practice.
Speak Out on Poverty: Hearing, Inaudibility, and Citizenship in Post‐Apartheid South Africa
Authors: Rousseau Nicky
Published: December 2019

In 1998, Speak Out on Poverty held hearings across South Africa shortly after the Truth and Reconciliation Commission (TRC) completed eighteen months of highly publicized, nationwide hearings at which victims testified. Speak Out challenged the TRC's focus on overt political violations, seen to occlude forms of structural violence central to apartheid's policy and practice, as well as longer legacies of colonialism. Reading Speak Out alongside the TRC puts pressure on supposed differences between official truth commissions or tribunals and those run by civil society. Discussing Speak Out in relation to the TRC signaled more than a set of comparisons. In a time of transition, Speak Out spoke from within and against the noise of the TRC. It aimed to make poverty and inequality the nation's priority rather than reconciliation, or at least to challenge notions of reconciliation that did not have inequity and poverty at its center. This intention failed to come to fruition, and Speak Out seemed paradoxically inaudible in public debate. In exploring this conundrum, this article considers particular strategic and organizational choices by Speak Out. These shaped the discursive terrain of operations and subjectivities in ways that unintentionally enabled inaudibility and constrained the organizers’ transformative intentions.
Making welfare conditional: A street‐level perspective
Authors: van Berkel Rik
Published: December 2019

This article looks at how welfare conditionality is delivered at the street level. It argues that the street‐level delivery of welfare conditionality is structured by policies, the governance context in which workers deliver welfare conditionality, the organization in which they work, and the occupation they are part of. Characteristics of these contexts present street‐level workers with a variety of signals and incentives that direct their decision making. The article elaborates on this proposition on the basis of a review of academic studies analysing the street‐level delivery of various aspects of welfare conditionality: the use of sanctions, service personalization, and the treatment of vulnerable clients. The review shows that context characteristics together have a significant impact on the street‐level transformation of welfare conditionality policies into practices. Street‐level decision making concerning the use of sanctions is far more complex than can be captured by a perspective on street‐level workers as merely policy implementers. Sanctioning practices are sometimes harsher, sometimes more lenient than policies lead us to expect. The “soft” side of welfare conditionality—represented by service personalization—is often under pressure at the street level, potentially strengthening welfare conditionality's tough side. This affects vulnerable jobseekers most: Street‐level studies show that the balance between disciplining and enabling aspects of welfare‐to‐work is most at risk for more vulnerable groups. The article concludes that the contextual pressures street‐level workers have to deal with in their daily work hardly reflect the “delicate equilibrium” that they need to deliver welfare conditionality in a professional, responsive, and responsible way.
Coagulation abnormalities in dengue and dengue haemorrhagic fever patients
Authors: Hassan Jawad , Borhany Munira , Abid Madiha , Zaidi Uzma , Fatima Naveena , Shamsi Tahir
Published: December 2019

Objective

The aim of this study was to assess abnormality of coagulation and anticoagulation parameters in dengue fever patients and the impact of these tests on the management of patients.

Background

Dengue fever is endemic in Pakistan with seasonal rise in cases. Morbidities and mortalities are proportionately reported to be increasing and associated with disseminated intravascular coagulation resulting in haemorrhagic or thrombotic manifestations in patients having deranged coagulation profiles.

Methods/Materials

This observational and descriptive study was conducted on confirmed Dengue patients at the National Institute of Blood Diseases during the years 2013 to 2016. Patients of all age groups were included in this study. Results were analysed by SPSS version 23.

Results

A total of 200 patients were selected with the mean age being 28.68 years (±13.28) and male predominance (147/200). The mean platelet count, haemoglobin and haematocrit at base line for bleeders and non‐bleeders showed significant results, where platelet count at baseline for bleeders was 24 000, whereas for non‐bleeders it was 29 000 and it showed significant correlation with bleeding (P‐value .027). Platelets were transfused to 76 (38%) patients. However, none of the specialised haemostasis parameters beside the platelet count correlated with bleeding, requiring platelet transfusions.

Conclusion

Our study showed a significant association of platelet counts, haemoglobin and haematocrit with bleeding. It can be concluded that coagulation and anticoagulation profiles will not benefit the management of dengue patients and in countries like Pakistan, it will only add to the economic burden on the patients.
Cellular perspectives for improving mesophyll conductance
Authors: Lundgren Marjorie R. , Fleming Andrew J.
Published: December 2019

After entering the leaf, CO2 faces an intricate pathway to the site of photosynthetic fixation embedded within the chloroplasts. The efficiency of CO2 flux is hindered by a number of structural and biochemical barriers which, together, define the ease of flow of the gas within the leaf, termed mesophyll conductance. Previous authors have identified the key elements of this pathway, raising the prospect of engineering the system to improve CO2 flux and, thus, to increase leaf photosynthetic efficiency. In this review, we provide a perspective on the potential for improving the individual elements that contribute to this complex parameter. We lay particular emphasis on generation of the cellular architecture of the leaf which sets the initial boundaries of a number of mesophyll conductance parameters, incorporating an overview of the molecular transport processes which have been proposed as major facilitators of CO2 flux across structural boundaries along the pathway. The review highlights the research areas where future effort might be invested to increase our fundamental understanding of mesophyll conductance and leaf function and, consequently, to enable translation of these findings to improve the efficiency of crop photosynthesis.
Cervids as sentinel‐species for tick‐borne encephalitis virus in Norway ‐ A serological study
Authors: Paulsen Katrine M. , das Neves Carlos G. , Granquist Erik G. , Madslien Knut , Stuen Snorre , Pedersen Benedikte N. , Vikse Rose , Rocchi Mara , Laming Ellie , Stiasny Karin , Andreassen Åshild K.
Published: December 2019

Tick‐borne encephalitis virus (TBEV) is the causative agent of tick‐borne encephalitis (TBE). TBEV is one of the most important neurological pathogens transmitted by tick bites in Europe. The objectives of this study were to investigate the seroprevalence of TBE antibodies in cervids in Norway and the possible emergence of new foci, and furthermore to evaluate if cervids can function as sentinel animals for the distribution of TBEV in the country. Serum samples from 286 moose, 148 roe deer, 140 red deer and 83 reindeer from all over Norway were collected and screened for TBE immunoglobulin G (IgG) antibodies with a modified commercial enzyme‐linked immunosorbent assay (ELISA) and confirmed by TBEV serum neutralisation test (SNT). The overall seroprevalence against the TBEV complex in the cervid specimens from Norway was 4.6%. The highest number of seropositive cervids was found in south‐eastern Norway, but seropositive cervids were also detected in southern‐ and central Norway. Antibodies against TBEV detected by SNT were present in 9.4% of the moose samples, 1.4% in red deer, 0.7% in roe deer, and nil in reindeer. The majority of the positive samples in our study originated from areas where human cases of TBE have been reported in Norway. The study is the first comprehensive screening of cervid species in Norway for antibodies to TBEV, and shows that cervids are useful sentinel animals to indicate TBEV occurrence, as supplement to studies in ticks. Furthermore, the results indicate that TBEV might be spreading northwards in Norway. This information may be of relevance for public health considerations and supports previous findings of TBEV in ticks in Norway.
Costos de calidad como estrategia de gestión en el central azucarero Trujillo, S.A., Venezuela
Authors: Montilla Morelia , Alizo Stephania , Salazar Diyerson , Rivas Nellitza
Published: December 2019

El propósito de esta investigación es construir una aproximación teórica que relacione los costos de calidad con las estrategias de gestión en el Central Azucarero Trujillo S.A., Venezuela. Para tal fin, se realizó un estudio descriptivo con diseño de campo. Para la recolección de la información se utilizó la entrevista estructurada, validada por tres expertos. Los resultados fueron analizados con matrices de triangulación. Se concluye que la entidad incurre en costos de calidad, destacando los de prevención y evaluación, tienen establecidas estrategias orientadas al mejoramiento continuo de la entidad; no obstante, se evidencia la falta de consideración de la información suministrada por los costos de calidad en el establecimiento de estrategias de gestión. Se propone una matriz que facilite comprender la participación que pueda generar cada una de las categorías de los costos de calidad con respecto a las estrategias de gestión llevadas a cabo en el Central Azucarero Trujillo S.A., con el propósito de fortalecer su productividad y operatividad.
Three new eriophyid mites of Phyllocoptini (Acari: Eriophyidae) from China

Authors: Zhang Qi , Yao Liang-fei , Xue Xiao-feng
Published: December 2019

Three new species of the tribe Phyllocoptini (Acari: Eriophyidae) from China, are described and illustrated. They are Phyllocoptes gaotainensus sp. n. on Populus euphratica (Salicaceae), Calepitrimerus anomalus sp. n. on Paeonia anomala (Paeoniaceae) and Paraepitrimerus stellarus sp. n. on Stellaria sp. (Caryophyllaceae). Based on the characteristics of dorsal and ventral annuli smooth, P. gaotainensus sp. n. may represent a deutogyne form. All three new species are vagrant on the lower leaf surface. No damage to the host was observed.
Multimodal method for the scattering by an array of plates connected to an elastic half-space
Authors: Maurel Agnès , Pham Kim
Published: December 2019

An accurate and numerically inexpensive method is presented to calculate the reflection of in-plane waves at the free surface of an elastic substrate supporting a periodic array of plates. The method is based on the expansions of the elastic fields on the pseudo-periodic modes in the substrate and on the Lamb modes in the plates. These expansions involve unknown amplitudes which are determined by simple matrix inversion when accounting for the boundary conditions at the surface of the substrate and at the free edges of the plates. Exemplifying results are reported in a wide range of frequency covering two resonances of the plates which are analyzed: a flexural resonance of slender bodies and the quasi-resonance of the so-called edge mode. The convergence of the method is inspected quantitatively; it is shown that it is good in view of the fact that the stress is singular at the corners of the plates in contact with the substrate. In particular, the scattering coefficients converge as 1/Ns3/2 with Ns the truncation of the expansions.
Stepping on the Mat: A Novice Practitioner’s Experiences Conducting a Mental-Training Workshop for Mixed Martial Arts Athletes
Authors: Taber Jamie , Longshore Kat
Published: December 2019

The current case reports a multimodal group intervention implemented at a mixed martial arts (MMA) school. Participants in the workshop included athletes of a range of belt ranks and experience levels, demonstrating an interest in mental-skills training from not only competitive but also recreational athletes. Goal setting, arousal regulation, self-talk, and imagery were covered, with a short informative lecture and an activity for each skill. Feedback was generally positive, with participants indicating that they had learned a lot and suggesting more workshops in the future, as well as more sport-specific examples. For the student-trainee, being new to leading workshops and being a member of the population were important; for the supervisor, taking on that role and working in an unfamiliar sport were novel experiences. Efficacy research should be conducted with this kind of intervention for recreational MMA athletes, but feedback demonstrated an interest in mental skills for this population.
“Clean Athlete” or “Drug Cheat and a Jerk”? A Comparative Analysis of the Framing of an Athlete Conflict in Australian and Chinese Print Media
Authors: Li Bo , Scott Olan K.M. , Sharpe Stirling , Xu Qingru , Naraine Michael
Published: December 2019

Media coverage in China and Australia examined a conflict between 2 Olympic swimmers, Chinese Sun Yang and Australian Mack Horton, during the 2016 Rio Olympic Games. While both swimmers performed well, there were several conflicts between the 2 leading to both nations’ media coverage portraying the foreign athlete negatively. An analysis of 398 print-media articles revealed there were sharp differences between the 2 nations in both the amount of coverage and the valence of the information sources. From a theoretical perspective, the framing of this conflict showed an “us vs. them” dichotomy, suggesting that both countries’ coverage was strongly influenced to protect the reputation and honor of the home athlete. Coverage in both nations was markedly different, suggesting a home-nation favoritism. Implications for sport communicators are discussed.
Instagram as Exemplar: Examining the Presence and Likability of Subjects and Statistics in March Madness Posts
Authors: Hahn Dustin A.
Published: December 2019

While many sport fans gravitate to new media, questions remain regarding what they are consuming. Specifically, this study addresses a nascent gap in sport communication research by identifying the presentation form and subject portrayals of 443 Instagram posts during 2018 college basketball tournaments and measuring subsequent likability of such depictions. Results yield 4 important findings. Primarily, in contrast to early exemplification research, evidence suggests that some audiences “like” base-rate information. In addition, while Instagram is known as a photo platform, posts in this population were most often videos, and memes were liked more than any other presentation form. Next, while this content analysis identifies an old problem in a new domain—that female athletes are shunned in favor of male athletes—it demonstrates that likability in this medium emerges as equal for male and female subjects and sports. Finally, related to subject demographics, findings demonstrated racial disparities and concerning statistics for likability of minority subjects. Implications for exemplification theory and social media producers in sport are discussed along with limitations and directions for future research in this burgeoning arena.
Sporting Women and Social Media: Sexualization, Misogyny, and Gender-Based Violence in Online Spaces
Authors: Kavanagh Emma , Litchfield Chelsea , Osborne Jaquelyn
Published: December 2019

This study investigated gender-based violence targeting high-profile women in virtual environments through the case of women’s tennis. Using a netnographic approach and the lens of third-wave feminism, 2 popular social media platforms (Facebook and Twitter) were analyzed to examine social commentary and fan interaction surrounding the top-5-seeded female tennis players during the Wimbledon Tennis Championships. Athletes were exposed to violent interactions in a number of ways. Four themes were identified through data analysis: threats of physical violence, sexualization that focused on the female physical appearance, sexualization that expressed desire and/or proposed physical or sexual contact, and sexualization that was vile, explicit, and threateningly violent in a sexual or misogynistic manner. Findings demonstrate how social media provides a space for unregulated gender-based cyberhate targeting high-profile women in their workplace in a way that traditional sport media does not.
Reevaluating human colonization of the Caribbean using chronometric hygiene and Bayesian modeling
Authors: Napolitano Matthew F. , DiNapoli Robert J. , Stone Jessica H. , Levin Maureece J. , Jew Nicholas P. , Lane Brian G. , O’Connor John T. , Fitzpatrick Scott M.
Published: December 2019

Bayesian-modeled colonization estimates demonstrate episodic dispersals into the Caribbean that support the southward route hypothesis.
Genome-wide analyses reveal the role of noncoding variation in complex traits during rice domestication
Authors: Zheng X. M. , Chen J. , Pang H. B. , Liu S. , Gao Q. , Wang J. R. , Qiao W. H. , Wang H. , Liu J. , Olsen K. M. , Yang Q. W.
Published: December 2019

During rice domestication, selection for genome-wide change in the expression of long noncoding RNAs led to improved grain traits.
Targeting NASH with OxPL neutralization
Authors: Ozcan Lale
Published: December 2019

Accumulation of oxidized phospholipids (OxPLs) contribute to NASH development.
Heterogeneity in the Attachment and Uptake Mechanisms of the Legionnaires’ Disease Bacterium,
Legionella pneumophila
, by Protozoan Hosts
Authors: Harb Omar S. , Venkataraman Chandrasekar , Haack Bradley J. , Gao Lian-Yong , Kwaik Yousef Abu
Published: December 2019

Invasion and intracellular replication of
Legionella pneumophila
within protozoa in the environment plays a major role in the transmission of Legionnaires’ disease. Intracellular replication of
L. pneumophila
within protozoa occurs in a rough endoplasmic reticulum (RER)-surrounded phagosome (Y. Abu Kwaik, Appl. Environ. Microbiol. 62:2022–2028, 1996). Since the subsequent fate of many intracellular pathogens is determined by the route of entry, we compared the mechanisms of attachment and subsequent uptake of
L. pneumophila
by the two protozoa
Hartmannella vermiformis
and
Acanthamoeba polyphaga
. Our data provide biochemical and genetic evidence that the mechanisms of attachment and subsequent uptake of
L. pneumophila
by the two protozoan hosts are, in part, different. First, uptake of
L. pneumophila
by
H. vermiformis
is completely blocked by the monovalent sugars galactose and
N
-acetyl-
d
-galactosamine, but these sugars partially blocked
A. polyphaga
. Second, attachment of
L. pneumophila
to
H. vermiformis
is associated with a time-dependent and reversible tyrosine dephosphorylation of multiple host proteins. In contrast, only a slight dephosphorylation of a 170-kDa protein of
A. polyphaga
is detected upon infection. Third, synthesis of
H. vermiformis
proteins but not of
A. polyphaga
proteins is required for uptake of
L. pneumophila
. Fourth, we have identified
L. pneumophila
mutants that are severely defective in attachment to
A. polyphaga
but which exhibit minor reductions in attachment to
H. vermiformis
and, thus, provide a genetic basis for the difference in mechanisms of attachment to both protozoa. The data indicate a remarkable adaptation of
L. pneumophila
to attach and invade different protozoan hosts by different mechanisms, yet invasion is followed by a remarkably similar intracellular replication within a RER-surrounded phagosome and subsequent killing of the host cell.
Morphological and Molecular Identification of
Trichoderma
Isolates on North American Mushroom Farms
Authors: Castle Alan , Speranzini Donna , Rghei Nezar , Alm Glen , Rinker Dan , Bissett John
Published: December 2019

Green mold disease (causal agent,
Trichoderma
) has resulted in severe crop losses on mushroom farms worldwide in recent years. We analyzed 160 isolates of
Trichoderma
from mushroom farms for morphological, cultural, and molecular characteristics and classified these isolates into phenotypic groups. The most common group comprised approximately 40% of the isolates and was identified as a strain of
Trichoderma harzianum
. This group was consistently recovered from farms with severe green mold disease but not from farms with little or no problem. In addition, the strain identified as the major cause of green mold disease in Ireland and the United Kingdom grouped with these North American isolates in having very similar randomly amplified polymorphic DNA patterns.
Influence of Culture Conditions on Expression of the 40-Kilodalton Porin Protein of
Vibrio anguillarum
Serotype O2
Authors: Davey Michelle L. , Hancock Robert E. W. , Mutharia Lucy M.
Published: December 2019

Vibrio anguillarum
serotype O2 strains express a 40-kDa outer membrane porin protein. Immunoblot analysis revealed that antigenic determinants of the
V. anguillarum
O2 40-kDa porin were conserved within bacterial species of the genus
Vibrio
. The relative amounts of the
V. anguillarum
O2 40-kDa porin were enhanced by growth of
V. anguillarum
O2 in CM9 medium containing 5 to 10% sucrose or 0.1 to 0.5 M NaCl. In contrast, the levels of the porin were significantly reduced when cells were grown at 37°C, and a novel 60-kDa protein was also observed. However, the osmolarity or ionic concentration of the growth medium did not influence expression of the 60-kDa protein. Growth in medium containing greater than 0.6 mM EDTA reduced production of the
V. anguillarum
O2 40-kDa porin and enhanced levels of a novel 19-kDa protein. Thus, expression of the
V. anguillarum
O2 40-kDa porin was osmoregulated and possibly coregulated by temperature. The N-terminal amino acid sequence of the
V. anguillarum
O2 40-kDa protein and the effect of environmental factors on the cellular levels of the porin suggested that the
V. anguillarum
O2 40-kDa porin was functionally similar to the OmpC porin of
Escherichia coli
. However, pore conductance assays revealed that the
V. anguillarum
O2 40-kDa porin was a general diffusion porin with a pore size in the range of that of the OmpF porin of
E. coli.
Use of Subtractive Hybridization To Design Habitat-Based Oligonucleotide Probes for Investigation of Natural Bacterial Communities
Authors: Mau Margit , Timmis Kenneth N.
Published: December 2019

We describe a rapid oligonucleotide probe design strategy based on subtractive hybridization which yields probes for 16S rRNA or rRNA genes of individual members of microbial communities that are specific within the context of those communities. This strategy circumvents the need to sequence many similar or identical clones of dominant members of a community. Radioactively labeled subfragments of a cloned 16S rRNA gene sequence for which a probe is required (target) were hybridized with biotinylated total 16S ribosomal DNA (rDNA) amplified from the microbial community, and the hybrids formed were subsequently discarded. The remaining enriched fragments were used to screen a library consisting of cloned subfragments of the target sequence by colony hybridization in order to identify the variable regions of the 16S rRNA gene with the required specificity. The sequencing of random clones in one 16S rDNA library demonstrated that only those clones with 100% sequence identity with the probe fragment were detected by it. Moreover, sequencing of other, randomly selected, probe-positive clones revealed 100% sequence identity with the probe. Probes developed in this way tended to correspond to more variable regions of the 16S rRNA if the target sequences were similar to the sequences of other clones in the library and to less variable regions if the target sequences were phylogenetically isolated within the clone library. Although the absolute specificity of the latter probes, as assessed by comparison with available database sequences, was lower than the absolute specificity of the probes from the more variable regions, they were specific within the context of the environmental samples from which they were derived.
Anaerobic Killing of Oral Streptococci by Reduced, Transition Metal Cations
Authors: Dunning J. C. , Ma Y. , Marquis R. E.
Published: December 2019

Reduced, transition metal cations commonly enhance oxidative damage to cells caused by hydroperoxides formed as a result of oxygen metabolism or added externally. As expected, the cations Fe
2+
and Cu
+
enhanced killing of
Streptococcus mutans
GS-5 by hydroperoxides. However, unexpectedly, they also induced lethal damage under fully anaerobic conditions in a glove box with no exposure to O
2
or hydroperoxides from initial treatment with the cations. Sensitivities to anaerobic killing by Fe
2+
varied among the organisms tested. The oral streptococci
Streptococcus gordonii
ATCC 10558,
Streptococcus rattus
FA-1, and
Streptococcus sanguis
NCTC 10904 were approximately as sensitive as
S. mutans
GS-5.
Enterococcus hirae
ATCC 9790,
Actinomyces viscosus
OMZ105E, and
Actinomyces naeslundii
WVU45 had intermediate sensitivity, while
Lactobacillus casei
ATCC 4646 and
Escherichia coli
B were insensitive. Killing of
S. mutans
GS-5 in response to millimolar levels of added Fe
2+
occurred over a wide range of temperatures and pH. The organism was able to take up ferrous iron, but ferric reductase activity could not be detected. Chelators, uric acid, and thiocyanate were not effective inhibitors of the lethal damage. Sulfhydryl compounds, ferricyanide, and ferrocyanide were protective if added prior to Fe
2+
exposure. Fe
2+
, but not Fe
3+
, acted to reduce the acid tolerance of glycolysis by intact cells of
S. mutans
. The reduction in acid tolerance appeared to be related directly to Fe
2+
inhibition of F-ATPase, which could be assayed with permeabilized cells, isolated membranes, or F
1
enzyme separated from membranes. Cu
+
and Cu
2+
also inhibited F-ATPase and sensitized glycolysis by intact cells to acid. All of these damaging actions occurred anaerobically and thus did not appear to involve reactive oxygen species.
Role of the Air-Water-Solid Interface in Bacteriophage Sorption Experiments
Authors: Thompson Shawn S. , Flury Markus , Yates Marylynn V. , Jury William A.
Published: December 2019

Batch sorption experiments were carried out with the bacteriophages MS2 and φX174. Two types of reactor vessels, polypropylene and glass, were used. Consistently lower concentrations of MS2 were found in the liquid phase in the absence of soil (control blanks) than in the presence of soil after mixing. High levels of MS2 inactivation (∼99.9%) were observed in control tubes made of polypropylene (PP), with comparatively little loss of virus seen in PP tubes when soil was present. Minimal inactivation of MS2 was observed when the air-water interface was completely eliminated from PP control blanks during mixing. All batch experiments performed with reactor tubes made of glass demonstrated no substantial inactivation of MS2. In similar experiments, bacteriophage φX174 did not undergo inactivation in either PP or glass control blanks, implying that this virus is not affected by the same factors which led to inactivation of MS2 in the PP control tubes. When possible, phage adsorption to soil was calculated by the Freundlich isotherm. Our data suggest that forces associated with the air-water-solid interface (where the solid is a hydrophobic surface) are responsible for inactivation of MS2 in the PP control tubes. The influence of air-water interfacial forces should be carefully considered when batch sorption experiments are conducted with certain viruses.
Sequence Variation of the tRNA
Leu
Intron as a Marker for Genetic Diversity and Specificity of Symbiotic Cyanobacteria in Some Lichens
Authors: Paulsrud Per , Lindblad Peter
Published: December 2019

We examined the genetic diversity of
Nostoc
symbionts in some lichens by using the tRNA
Leu
(UAA) intron as a genetic marker. The nucleotide sequence was analyzed in the context of the secondary structure of the transcribed intron. Cyanobacterial tRNA
Leu
(UAA) introns were specifically amplified from freshly collected lichen samples without previous DNA extraction. The lichen species used in the present study were
Nephroma arcticum
,
Peltigera aphthosa
,
P. membranacea
, and
P. canina
. Introns with different sizes around 300 bp were consistently obtained. Multiple clones from single PCRs were screened by using their single-stranded conformational polymorphism pattern, and the nucleotide sequence was determined. No evidence for sample heterogenity was found. This implies that the symbiont in situ is not a diverse community of cyanobionts but, rather, one
Nostoc
strain. Furthermore, each lichen thallus contained only one intron type, indicating that each thallus is colonized only once or that there is a high degree of specificity. The same cyanobacterial intron sequence was also found in samples of one lichen species from different localities. In a phylogenetic analysis, the cyanobacterial lichen sequences grouped together with the sequences from two free-living
Nostoc
strains. The size differences in the intron were due to insertions and deletions in highly variable regions. The sequence data were used in discussions concerning specificity and biology of the lichen symbiosis. It is concluded that the tRNA
Leu
(UAA) intron can be of great value when examining cyanobacterial diversity.
Impaired Secretion of a Hydrophobic Cutinase by
Saccharomyces cerevisiae
Correlates with an Increased Association with Immunoglobulin Heavy-Chain Binding Protein (BiP)
Authors: Sagt C. M. J. , Müller W. H. , Boonstra J. , Verkleij A. J. , Verrips C. T.
Published: December 2019

This study focuses on the different efficiencies of secretion of two fungal cutinases by
Saccharomyces cerevisiae
, a wild-type cutinase (CY000) and a hydrophobic mutant cutinase (CY028). Both cutinases are placed under control of the
GAL7
promoter, by which the expression levels can be regulated. Wild-type cutinase was secreted at up to 25 mg per g (dry weight), while CY028 was secreted at a level of 2 mg per g (dry weight); this difference is nearly independent of the expression level. Pulse-chase experiments revealed that whereas CY000 cutinase is secreted, CY028 is irreversibly retained in the cell. Immunogold labelling followed by electron microscopy revealed colocalization of CY028 with immunoglobulin heavy-chain binding protein (BiP) in the endoplasmic reticulum (ER). The increase of wild-type cutinase expression did not result in higher levels of the molecular chaperone BiP, but BiP levels are raised by increased induction of the hydrophobic mutant cutinase. Immunoprecipitation studies showed that in contrast to the wild-type cutinase, the hydrophobic mutant cutinase interacts with BiP. These results indicate that the introduction of two exposed hydrophobic patches in cutinase results in a higher affinity for BiP which might cause the retention of this mutant cutinase in the ER.
Electron and Fluorescence Microscopy of Extracellular Glucan and Aryl-Alcohol Oxidase during Wheat-Straw Degradation by
Pleurotus eryngii
Authors: Barrasa J. M. , Gutiérrez A. , Escaso V. , Guillén F. , Martínez M. J. , Martínez A. T.
Published: December 2019

The ligninolytic fungus
Pleurotus eryngii
grown in liquid medium secreted extracellular polysaccharide (87% glucose) and the H
2
O
2
-producing enzyme aryl-alcohol oxidase (AAO). The production of both was stimulated by wheat-straw. Polyclonal antibodies against purified AAO were obtained, and a complex of glucanase and colloidal gold was prepared. With these tools, the localization of AAO and extracellular glucan in mycelium from liquid medium and straw degraded under solid-state fermentation conditions was investigated by transmission electron microscopy (TEM) and fluorescence microscopy. These studies revealed that
P. eryngii
produces a hyphal sheath consisting of a thin glucan layer. This sheath appeared to be involved in both mycelial adhesion to the straw cell wall during degradation and AAO immobilization on hyphal surfaces, with the latter evidenced by double labeling. AAO distribution during differential degradation of straw tissues was observed by immunofluorescence microscopy. Finally, TEM immunogold studies confirmed that AAO penetrates the plant cell wall during
P. eryngii
degradation of wheat straw.
PCR Amplification and Species Determination of Microsporidia in Formalin-Fixed Feces after Immunomagnetic Separation
Authors: Dowd Scot E. , Gerba Charles P. , Enriquez F. Javier , Pepper Ian L.
Published: December 2019

The term microsporidia is used to describe several species of opportunistic protozoan parasites.
Encephalitozoon intestinalis
and
Enterocytozoon bieneusi
have been found in stools of more than 40% of AIDS patients with diarrhea. Diagnosis of infection with these small protozoans has been difficult, and until recently their occurrence has not been well documented. Formalin is widely used to preserve clinical specimens, but due to the nature of the fixation process, subsequent analysis, especially analysis by the PCR, is difficult. This study evaluated methods used to prepare formalin-fixed fecal specimens for PCR amplification of microsporidial DNA. Two methods were devised to allow PCR detection and subsequent identification of microsporidia in formalin-fixed fecal specimens to the species level. One method involved immunomagnetic separation to concentrate microsporidial spores from fecal specimens. In the second method Chelex resin (Bio-Rad, Hercules, Calif.) was used to remove inhibitory substances, followed by a DNA concentration step. Both methods resulted in reproducible, confirmed detection of microsporidia in formalinized fecal specimens and subsequent species determination by PCR sequencing. The detection sensitivity was two in vitro culture-derived spores (
Encephalitozoon intestinalis
) for the direct PCR. The reproducible detection sensitivity for DNA amplification from formalin-fixed fecal samples was 200 spores for either the Chelex method or the immunomagnetic bead separation method. Thus, we developed two methods for rapid, inexpensive detection of microsporidial spores in formalin-fixed fecal specimens.
Occurrence of
Vibrio vulnificus
Biotypes in Danish Marine Environments
Authors: Høi L. , Larsen J. L. , Dalsgaard I. , Dalsgaard A.
Published: December 2019

During the unusually warm summer in Denmark in 1994, 11 clinical cases of
Vibrio vulnificus
infection were reported. These reports initiated an investigation of the occurrence of
V. vulnificus
biotypes in Danish marine environments. Samples of coastal water, sediment, shellfish, and wild fish were analyzed by preenrichment in alkaline peptone water amended with polymyxin B (2.0 × 10
4
U/liter) followed by streaking onto modified cellobiose-polymyxin B-colistin agar.
V. vulnificus
-like colonies were tested with a
V. vulnificus
-specific DNA probe. Low densities of
V. vulnificus
were detected in water (0.8 to 19 CFU/liter) from June until mid-September and in sediment (0.04 to >11 CFU/g) from July until mid-November. The presence of
V. vulnificus
was strongly correlated with water temperature. However, we isolated
V. vulnificus
from water from a mussel farm at a lower temperature than previously reported (7°C). In 1 of the 13 locations studied,
V. vulnificus
was found in mussels in 7 of 17 samples analyzed; this is the first report of
V. vulnificus
in European shellfish.
V. vulnificus
was also isolated from gills, intestinal contents, and mucus from wild fish. Although biotyping of 706
V. vulnificus
strains isolated during our investigations revealed that the majority of the strains (99.6%) belonged to biotype 1, biotype 2 was detected in seawater at a low frequency (0.4%). Our findings provide further evidence that seawater can serve as a reservoir and might facilitate spread of
V. vulnificus
biotype 2 to eels, with subsequent spread to persons handling eels. In conclusion, our data demonstrate that
V. vulnificus
is ubiquitous in a temperate marine environment and that
V. vulnificus
biotype 2 is not strictly confined to eels.
Molecular Analysis of Hypervirulent Somatic Hybrids of the Entomopathogenic Fungi
Beauveria bassiana
and
Beauveria sulfurescens
Authors: Viaud Muriel , Couteaudier Yvonne , Riba Guy
Published: December 2019

Protoplast fusion of diauxotrophic mutants of a
Beauveria bassiana
entomopathogenic strain (Bb28) and a
Beauveria sulfurescens
toxinogenic strain (Bs2) produced hybrids which were significantly different from the parents in pathogenicity. Some of the hybrids were hypervirulent and killed insects more quickly than the Bb28 strain, probably because these hybrids had acquired the toxic activity of the Bs2 strain. By using six nuclear genes and a telomeric fingerprint probe, the molecular structures of the hybrids were studied. The results demonstrated the occurrence of parasexual events. Hybrids appeared to be diploid or aneuploid, with portions of the genome being heterozygous. A mitochondrial molecular marker indicated homoplasmy of the hybrids and inheritance of mitochondria from strain Bs2 or Bb28. The pathogenicities and the ploidies of the hybrids remained stable after passage through the host insect, showing that somatic hybridization provides an attractive method for the genetic improvement of biocontrol efficiency in the genus
Beauveria.
Leaching of Zinc Sulfide by
Thiobacillus ferrooxidans
: Experiments with a Controlled Redox Potential Indicate No Direct Bacterial Mechanism
Authors: Fowler T. A. , Crundwell F. K.
Published: December 2019

The role of
Thiobacillus ferrooxidans
in bacterial leaching of mineral sulfides is controversial. Much of the controversy is due to the fact that the solution conditions, especially the concentrations of ferric and ferrous ions, change during experiments. The role of the bacteria would be more easily discernible if the concentrations of ferric and ferrous ions were maintained at set values throughout the experimental period. In this paper we report results obtained by using the constant redox potential apparatus described previously (P. I. Harvey and F. K. Crundwell, Appl. Environ. Microbiol. 63:2586–2592, 1997). This apparatus is designed to control the redox potential in the leaching compartment of an electrolytic cell by reduction or oxidation of dissolved iron. By controlling the redox potential the apparatus maintains the concentrations of ferrous and ferric ions at their initial values. Experiments were conducted in the presence of
T. ferrooxidans
and under sterile conditions. Analysis of the conversion of zinc sulfide in the absence of the bacteria and analysis of the conversion of zinc sulfate in the presence of the bacteria produced the same results. This indicates that the only role of the bacteria under the conditions used is regeneration of ferric ions in solution. In this work we found no evidence that there is a direct mechanism for bacterial leaching.
Purification, Characterization, and Substrate Specificity of a Novel Highly Glucose-Tolerant β-Glucosidase fromAspergillus oryzae
Authors: Riou Christine , Salmon Jean-Michel , Vallier Marie-Jose , Günata Ziya , Barre Pierre
Published: December 2019

Aspergillus oryzaewas found to secrete two distinct β-glucosidases when it was grown in liquid culture on various substrates. The major form had a molecular mass of 130 kDa and was highly inhibited by glucose. The minor form, which was induced most effectively on quercetin (3,3′,4′,5,7-pentahydroxyflavone)-rich medium, represented no more than 18% of total β-glucosidase activity but exhibited a high tolerance to glucose inhibition. This highly glucose-tolerant β-glucosidase (designated HGT-BG) was purified to homogeneity by ammonium sulfate precipitation, gel filtration, and anion-exchange chromatography. HGT-BG is a monomeric protein with an apparent molecular mass of 43 kDa and a pI of 4.2 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and isoelectric focusing polyacrylamide gel electrophoresis, respectively. Usingp-nitrophenyl-β-d-glucoside as the substrate, we found that the enzyme was optimally active at 50°C and pH 5.0 and had a specific activity of 1,066 μmol min−1mg of protein−1and aKmof 0.55 mM under these conditions. The enzyme is particularly resistant to inhibition by glucose (Ki, 1.36 M) or glucono-δ-lactone (Ki, 12.5 mM), another powerful β-glucosidase inhibitor present in wine. A comparison of the enzyme activities on various glycosidic substrates indicated that HGT-BG is a broad-specificity type of fungal β-glucosidase. It exhibits exoglucanase activity and hydrolyzes (1→3)- and (1→6)-β-glucosidic linkages most effectively. This enzyme was able to release flavor compounds, such as geraniol, nerol, and linalol, from the corresponding monoterpenyl-β-d-glucosides in a grape must (pH 2.9, 90 g of glucose liter−1). Other flavor precursors (benzyl- and 2-phenylethyl-β-d-glucosides) and prunin (4′,5,7-trihydroxyflavanone-7-glucoside), which contribute to the bitterness of citrus juices, are also substrates of the enzyme. Thus, this novel β-glucosidase is of great potential interest in wine and fruit juice processing because it releases aromatic compounds from flavorless glucosidic precursors.
Characterization of an Extremely Thermostable Restriction Enzyme,
Psp
GI, from a
Pyrococcus
Strain and Cloning of the
Psp
GI Restriction-Modification System in
Escherichia coli
Authors: Morgan Richard , Xiao Jian-ping , Xu Shuang-yong
Published: December 2019

An extremely thermostable restriction endonuclease,
Psp
GI, was purified from
Pyrococcus
sp. strain GI-H.
Psp
GI is an isoschizomer of
Eco
RII and cleaves DNA before the first C in the sequence 5′ ^CCWGG 3′ (W is A or T).
Psp
GI digestion can be carried out at 65 to 85°C. To express
Psp
GI at high levels, the
Psp
GI restriction-modification genes (
pspGIR
and
pspGIM
) were cloned in
Escherichia coli
. M.
Psp
GI contains the conserved sequence motifs of α-aminomethyltransferases; therefore, it must be an N4-cytosine methylase. M.
Psp
GI shows 53% similarity to (44% identity with) its isoschizomer, M.
Mva
I from
Micrococcus variabilis
. In a segment of 87 amino acid residues,
Psp
GI shows significant sequence similarity to
Eco
RII and to regions of
Sso
II and
Sty
D4I which have a closely related recognition sequence (5′ ^CCNGG 3′).
Psp
GI was expressed in
E. coli
via a T7 expression system. Recombinant
Psp
GI was purified to near homogeneity and had a half-life of 2 h at 95°C.
Psp
GI remained active following 30 cycles of thermocycling; thus, it can be used in DNA-based diagnostic applications.