bluuebunny/crossref_metadata_2025_split huggingface dataset:
The
Caulobacter
NtrB-NtrC two-component system bridges nitrogen assimilation and cell development
Authors: North Hunter , McLaughlin Maeve , Fiebig Aretha , Crosson Sean 
Published: October 2023

ABSTRACT

A suite of molecular sensory systems enables
Caulobacter
to control growth, development, and reproduction in response to levels of essential elements. The bacterial enhancer-binding protein (bEBP) NtrC and its cognate sensor histidine kinase, NtrB, are key regulators of nitrogen assimilation in many bacteria, but their roles in
Caulobacter
metabolism and development are not well defined. Notably,
Caulobacter
NtrC is an unconventional bEBP that lacks the σ
54
-interacting loop commonly known as the GAFTGA motif. Here we show that deletion of
Caulobacter crescentus ntrC
slows cell growth in complex medium and that
ntrB
and
ntrC
are essential when ammonium is the sole nitrogen source due to their requirement for glutamine synthetase expression. Random transposition of a conserved IS3-family mobile genetic element frequently rescued the growth defect of
ntrC
mutant strains by restoring transcription of the
glnBA
operon, revealing a possible role for IS3 transposition in shaping the evolution of
Caulobacter
populations during nutrient limitation. We further identified dozens of direct NtrC-binding sites on the
C. crescentus
chromosome, with a large fraction located near genes involved in polysaccharide biosynthesis. The majority of binding sites align with those of the essential nucleoid-associated protein, GapR, or the cell cycle regulator, MucR1. NtrC is therefore predicted to directly impact the regulation of cell cycle and cell development. Indeed, loss of NtrC function led to elongated polar stalks and elevated synthesis of cell envelope polysaccharides. This study establishes regulatory connections between NtrC, nitrogen metabolism, polar morphogenesis, and envelope polysaccharide synthesis in
Caulobacter
.

IMPORTANCE

Bacteria balance cellular processes with the availability of nutrients in their environment. The NtrB-NtrC two-component signaling system is responsible for controlling nitrogen assimilation in many bacteria. We have characterized the effect of
ntrB
and
ntrC
deletion on
Caulobacter
growth and development and uncovered a role for spontaneous IS element transposition in the rescue of transcriptional and nutritional deficiencies caused by
ntrC
mutation. We further defined the regulon of
Caulobacter
NtrC, a bacterial enhancer-binding protein, and demonstrate that it shares specific binding sites with essential proteins involved in cell cycle regulation and chromosome organization. Our work provides a comprehensive view of transcriptional regulation mediated by a distinctive NtrC protein, establishing its connection to nitrogen assimilation and developmental processes in
Caulobacter
.
Bacteriophage steering of
Burkholderia cenocepacia
toward reduced virulence and increased antibiotic sensitivity
Authors: Ruest Marta K. , Supina Brittany S. I. , Dennis Jonathan J. 
Published: October 2023

ABSTRACT

Antibiotic resistance in bacteria is a growing global concern and has spurred increasing efforts to ﬁnd alternative therapeutics, such as the use of bacterial viruses, or bacteriophages. One promising approach is to use phages that not only kill pathogenic bacteria but also select phage-resistant survivors that are newly sensitized to traditional antibiotics, in a process called “phage steering.” Members of the bacterial genus
Burkholderia
, which includes various human pathogens, are highly resistant to most antimicrobial agents, including serum immune components, antimicrobial peptides, and polymixin-class antibiotics. However, the application of phages in combination with certain antibiotics can produce synergistic effects that more effectively kill pathogenic bacteria. Herein, we demonstrate that
Burkholderia cenocepacia
serum resistance is due to intact lipopolysaccharide (LPS) and membranes, and phage-induced resistance altering LPS structure can enhance bacterial sensitivity not only to immune components in serum but also to membrane-associated antibiotics such as colistin.

IMPORTANCE
Bacteria frequently encounter selection pressure from both antibiotics and lytic phages, but little is known about the interactions between antibiotics and phages. This study provides new insights into the evolutionary trade-offs between phage resistance and antibiotic sensitivity. The creation of phage resistance through changes in membrane structure or lipopolysaccharide composition can simultaneously be a major cause of antibiotic sensitivity. Our results provide evidence of synergistic therapeutic efficacy in phage-antibiotic interactions and have implications for the future clinical use of phage steering in phage therapy applications.
Extracellular DNA enhances biofilm integrity and mechanical properties of mucoid
Pseudomonas aeruginosa
Authors: Ferguson Danielle L. , Gloag Erin S. , Parsek Matthew R. , Wozniak Daniel J. 
Published: October 2023

ABSTRACT

Pseudomonas aeruginosa
is one of the most common biofilm-forming pathogens responsible for lung infections of individuals with cystic fibrosis (CF).
P. aeruginosa
becomes tolerant to antimicrobials in the biofilm state and is difficult to treat. Production of extracellular polymeric substances (EPS), such as alginate and extracellular DNA (eDNA), can allow adherence to abiotic and biotic surfaces, antimicrobial evasion, and resilience to environmental pressures. Alginate-producing mucoid variants of
P. aeruginosa
are frequently isolated from CF airway samples and are associated with worsening patient outcomes. While eDNA is a major structural component of nonmucoid
P. aeruginosa
biofilms, the potential role of eDNA in mucoid biofilms is unclear. Here, we investigate how eDNA contributes to clinical mucoid biofilm physiology and integrity. We predicted that eDNA plays a structural and mechanical role in mucoid biofilms. To test this, we quantified biofilm eDNA in mucoid biofilms and used microscopy and rheology to visualize eDNA and detect changes in biofilm structure and mechanics upon DNaseI treatment. We showed that biofilm eDNA abundance is diverse across clinical mucoid strains and observed a temporal increase in foci of eDNA within intact mucoid biofilms. Increased cell dispersal and reduced biomass were also observed following DNaseI treatment of mucoid biofilms. Degradation of eDNA also impacted the mechanical integrity of mucoid biofilms by increasing the stiffness and decreasing the cohesion of the biofilm. These findings advance our understanding of clinical mucoid
P. aeruginosa
biofilms and facilitate the development of new approaches to target biofilms by exploiting the functions of EPS components.

IMPORTANCE

Understanding the role of eDNA in mucoid
Pseudomonas aeruginosa
biofilms will lead to therapeutic strategies that combat the biophysical and structural function of EPS for the eradication of bacteria in mucoid biofilms during chronic infections. This knowledge can be used to further identify unknown matrix component interactions within pathogenic biofilm-forming clinical isolates.
Rapid Aztreonam/Avibactam NP test for detection of aztreonam/avibactam susceptibility/resistance in Enterobacterales
Authors: Viguier Clément , Bouvier Maxime , Sadek Mustafa , Kerbol Auriane , Poirel Laurent , Nordmann Patrice 
Published: October 2023

ABSTRACT

Aztreonam-avibactam (AZA), a newly developed β-lactam/β-lactamase inhibitor combination
,
is a treatment option for infections due to carbapenem-resistant Enterobacterales (CRE), including metallo-ß-lactamase producers, regardless of additional production of broad-spectrum serine-ß-lactamases. However, AZA-resistance has already been reported in Enterobacterales and its early detection could be a valuable tool for faster and more accurate clinical decision-making. We therefore developed a rapid culture-based test for the identification of AZA resistance among multidrug-resistant Enterobacterales. The Rapid Aztreonam/Avibactam NP test is based on resazurin reduction when bacterial growth occurs in the presence of AZA at 8/4 µg/mL (protocol 1) or 12/4 µg/mL (protocol 2). Given the absence of guidelines on AZA susceptibility testing, two tentative breakpoints were indeed used to categorize AZA-susceptible isolates: ≤4 µg/mL in protocol 1 and ≤ 8 µg/mL in protocol 2. Bacterial growth was visually detectable by a blue-to-purple or blue-to-pink color change of the medium. A total of 78 enterobacterial isolates (among which 34 AZA-resistant and 13 AZA-resistant according to protocols 1 and 2, respectively) were used to evaluate the test performance using protocol 1 or protocol 2. The sensitivity and specificity of the test were found to be 100% and 97.7%, respectively, following protocol 1 and 100% and 100%, respectively, following protocol 2, in comparison with broth microdilution. All results were obtained within 4.5 hours corresponding to a time saving of ca. 14 hours compared with currently available methods for AZA susceptibility testing. The Rapid Aztreonam/Avibactam NP test is rapid, highly sensitive, specific, easily interpretable, and easy to implement in routine.
Lockdown!
Learning public health of infectious diseases through gameplay
Authors: Goh Evelyn , Szücs Anna , Loh Victor Weng Keong , Boucher Yann Felix 
Published: October 2023

ABSTRACT
The COVID-19 pandemic has demonstrated the detrimental effects of a lack of understanding of public health measures. During the pandemic, lockdowns, social distancing, and mask mandates introduced by governments were met with skepticism, doubt, and an unwillingness to comply, increasing the extent of negative outcomes as a result. Albeit devastating, the pandemic has offered an invaluable opportunity to observe the correlation between the prevalence of public health education and compliance with public health measures during critical times. In this article, we describe a card game that was developed during the COVID-19 pandemic to educate the public (including children) about how specific public health measures address the pandemic and how global cooperation is essential in addressing even one country’s problems. The game can be used in primary, secondary, or tertiary education classrooms, initiating conversations about the topic and providing a basic understanding before more in-depth learning.
Teaching transferable skills in teamwork, accountability, goal setting, writing, and problem-solving in a non-major microbiology lab: the unknown bacteria experiment redefined
Authors: McKee Felicia , Wolf Josef D. , Simon Samantha , Floyd Jeanetta H. 
Published: October 2023

ABSTRACT
College to Career is a phrase that we often use to describe the skills and abilities that students should achieve while preparing for college and/or careers. To help prepare our students for their future careers, we developed a microbiology laboratory curriculum based on factors identified to improve college-to-career readiness. These factors include content knowledge, analyzing and interpreting data, accountability, goal setting, and teamwork. At the core of the design are inquiry and problem-based learning. This approach allows students to actively engage in the scientific process while collaborating with classmates and learning technical and transferable career skills. The curriculum includes microbiology laboratory skills, including plating, serial dilutions, and biochemical tests, with integrated opportunities for students to engage in critical thinking, analysis and interpretation of data, teamwork, goal setting, decision-making, and scientific writing.
Chimeric virus-like particles of human norovirus constructed by structure-guided epitope grafting elicit cross-reactive immunity against both GI.1 and GII.4 genotypes
Authors: Hou Ya Nan , Jin Yu Qin , Zhang Xue Feng , Tang Fang , Hou Jun Wei , Liu Zhao Ming , Han Zi Bo , Zhang Hao , Du Li Fang , Shao Shuai , Su Ji Guo , Liang Yu , Zhang Jing , Li Qi Ming 
Published: October 2023

ABSTRACT

Human norovirus (HuNoV) is the main cause of acute non-bacterial gastroenteritis worldwide. There is no vaccine currently available to prevent HuNoV infection. HuNoV is a highly mutated virus, and its genetic diversity and the lack of cross-protection between different genotypes hinder the broadly protective vaccine development. Among various genotypes, GI.1 is the prototype strain, and GII.4 currently predominates the prevalence of HuNoV. In this work, guided by the structural alignment of GI.1 and GII.4 HuNoV capsid proteins, several chimeric virus-like particles (VLPs) were designed to achieve cross-immunity against these two HuNoV genotypes. The neutralizing epitopes of HuNoV have been identified to be mainly located at the loop regions of VP1 protein exposed on the HuNoV surface. In this study, the exposed loops of GII.4 VP1 protein were grafted into the scaffold of GI.1 genotype to produce the chimeric VLPs. The designed chimeric VLPs were recombinantly expressed by the
Hansenula polymorpha
expression system developed by our laboratory. Mice were immunized with the chimeric VLPs plus aluminum adjuvant, and then the antibody responses were detected by using the enzyme-linked immunosorbent assay and the histo-blood group antigen-VLP interaction blocking assay. The experimental results show that two of the designed chimeric VLPs induced cross-reactive IgG and cross-blocking antibodies against both the parental GI.1 and GII.4 genotypes of HuNoV. The results also imply that the transplant site design is important to maintain the immunogenicity of foreign epitopes on the scaffold carrier. Our studies may provide a valuable strategy for the development of cross-reactive HuNoV vaccines.

IMPORTANCE
Human norovirus (HuNoV) is highly infectious and can result in severe illnesses in the elderly and children. So far, there is no effective antiviral drug to treat HuNoV infection, and thus, the development of HuNoV vaccines is urgent. However, NoV evolves rapidly, and currently, at least 10 genogroups with numerous genotypes have been found. The genetic diversity of NoV and the lack of cross-protection between different genotypes pose challenges to the development of broadly protective vaccines. In this study, guided by structural alignment between GI.1 and GII.4 HuNoV VP1 proteins, several chimeric-type virus-like particles (VLPs) were designed through surface-exposed loop grafting. Mouse immunization studies show that two of the designed chimeric VLPs induced cross-immunity against both GI.1 and GII.4 HuNoVs. To our knowledge, this is the first designed chimeric VLPs that can induce cross-immune activities across different genogroups of HuNoV, which provides valuable strategies for the development of cross-reactive HuNoV vaccines.
A viral protein targets mitochondria and chloroplasts by subverting general import pathways and specific receptors
Authors: Sáiz-Bonilla María , Martín-Merchán Andrea , Pallás Vicente , Navarro Jose Antonio 
Published: October 2023

ABSTRACT
Dual targeting is a relatively little explored phenomenon by which some proteins are transported to two different subcellular compartments, such as chloroplasts and mitochondria. We previously showed that the R domain and the arm region of the melon necrotic spot virus coat protein could act as a dual-targeting peptide, which shares many relevant features in structure, amino acid composition, and functionality with that of the Thr-tRNA synthetases, the best-studied dual-targeted proteins by far. Here, we show that a viral coat protein hijacks the general pathways for organellar protein import and identifies the coat protein-interacting organellar translocon receptors. Our findings demonstrate for the first time that an exogenous dual-targeted protein uses the general Toc and Tom import systems to reach mitochondria and chloroplasts because it depends on the availability of NbToc75 and NbTom40. Interaction studies, subcellular localization analysis, and viral infection assays in silenced plants revealed the significance of plant receptors, such as the mitochondrion NbOm64 and chloroplast NbToc159A, in this transport. We also show that chloroplast protein translocation impairment could be interlinked with retrograde communication triggering a JA-based response that incidentally impairs viral infection. Future research may profit from our results to better comprehend cellular communication, protein import mechanisms, how preproteins interact with their receptors, and how plant pathogens manipulate host machinery to their ends.

IMPORTANCE
Many plant proteins and some proteins from plant pathogens are dually targeted to chloroplasts and mitochondria, and are supposed to be transported along the general pathways for organellar protein import, but this issue has not been explored yet. Moreover, organellar translocon receptors exist as families of several members whose functional specialization in different cargos is supposed but not thoroughly studied. This article provides novel insights into such topics showing for the first time that an exogenous protein, the melon necrotic spot virus coat protein, exploits the common Toc/Tom import systems to enter both mitochondria and chloroplasts while identifying the involved specific receptors.
Genome sequence ofHemileia vastatrixBerk. and Br. (Race I), the causal agent of coffee leaf rust, isolate from Risaralda, Colombia
Authors: Ángel C. Carlos A. , Marín-Ramírez Gustavo A. , Maldonado Carlos E. 
Published: October 2023

ABSTRACTCoffee leaf rust, caused by the fungusHemileia vastatrix(Basidiomycota; Pucciniomycota),is a devastating disease spread worldwide. To improve the available genomes, we use PacBio HiFi sequencing enhanced by Dovetail Omni-C chromatin conformation capture to assemble a highly contiguous 747.98 Mb genome of an isolate collected fromCoffea arabica.
Volatile profiling distinguishes
Streptococcus pyogenes
from other respiratory streptococcal species
Authors: Berna Amalia Z. , Merriman Joseph A. , Mellett Leah , Parchment Danealle K. , Caparon Michael G. , Odom John Audrey R. 
Published: October 2023

ABSTRACT

Sore throat is one of the most common complaints encountered in the ambulatory clinical setting. Rapid, culture-independent diagnostic techniques that do not rely on pharyngeal swabs would be highly valuable as a point-of-care strategy to guide outpatient antibiotic treatment. Despite the promise of this approach, efforts to detect volatiles during oropharyngeal infection have yet been limited. In our research study, we sought to evaluate for specific bacterial volatile organic compounds (VOC) biomarkers in isolated cultures
in vitro
, in order to establish proof-of-concept prior to initial clinical studies of breath biomarkers. A particular challenge for the diagnosis of pharyngitis due to
Streptococcus pyogenes
is the likelihood that many metabolites may be shared by
S. pyogenes
and other related oropharyngeal colonizing bacterial species. Therefore, we evaluated whether sufficient metabolic differences are present, which distinguish the volatile metabolome of Group A streptococci from other streptococcal species that also colonize the respiratory mucosa, such as
Streptococcus pneumoniae
and
Streptococcus intermedius
. In this work, we identified 27 discriminatory VOCs (
q
-values < 0.05), composed of aldehydes, alcohols, nitrogen-containing compounds, hydrocarbons, ketones, aromatic compounds, esters, ethers, and carboxylic acid. From this group of volatiles, we identify candidate biomarkers that distinguish
S. pyogenes
from other species and establish highly produced VOCs that indicate the presence of
S. pyogenes in vitro
, supporting future breath-based diagnostic testing for streptococcal pharyngitis.

IMPORTANCE

Acute pharyngitis accounts for approximately 15 million ambulatory care visits in the United States. The most common and important bacterial cause of pharyngitis is
Streptococcus pyogenesis
, accounting for 15%–30% of pediatric pharyngitis. Distinguishing between bacterial and viral pharyngitis is key to management in US practice. The culture of a specimen obtained by a throat swab is the standard laboratory procedure for the microbiologic confirmation of pharyngitis; however, this method is time-consuming, which delays appropriate treatment. If left untreated,
S. pyogenes
pharyngitis may lead to local and distant complications. In this study, we characterized the volatile metabolomes of
S. pyogenes
and other related oropharyngeal colonizing bacterial species. We identify candidate biomarkers that distinguish
S. pyogenes
from other species and provide evidence to support future breath-based diagnostic testing for streptococcal pharyngitis.
Investigating the determinants of iridolenticular contact area: a novel parameter for angle closure
Authors: Tun Tin A , Nongpiur Monisha Esther , Xu Benjamin Y , Wang Xiaofei , Tan Marcus , Quah Joanne Hui Min , Lim Hou-Boon , Cheng Ching Yu , Aung Tin 
Published: October 2023

Background/aims
To identify ocular determinants of iridolenticular contact area (ILCA), a recently introduced swept-source optical coherence tomography (SSOCT) derived parameter, and assess the association between ILCA and angle closure.

Methods
In this population-based cross-sectional study, right eyes of 464 subjects underwent SSOCT (SS-1000, CASIA, Tomey Corporation, Nagoya, Japan) imaging in the dark. Eight out of 128 cross-sectional images (evenly spaced 22.5° apart) were selected for analysis. Matlab (Matworks, Massachusetts, USA) was used to measure ILCA, defined as the circumferential extent of contact area between the pigmented iris epithelium and anterior lens surface. Gonioscopic angle closure (GAC) was defined as non-visibility of the posterior trabecular meshwork in two or more angle quadrants.

Results

The mean age of subjects was 62±6.6 years, with the majority being female (65.5%). 143/464 subjects (28.6%) had GAC. In multivariable linear regression analysis, ILCA was significantly associated with anterior chamber width (β=1.03, p=0.003), pupillary diameter (β=−1.9, p<0.001) and iris curvature (β=−17.35, p<0.001). ILCA was smaller in eyes with GAC compared with those with open angles (4.28±1.6 mm
2
vs 6.02±2.71 mm
2
, p<0.001). ILCA was independently associated with GAC (β=−0.03, p<0.001), iridotrabecular contact index (β=−6.82, p<0.001) or angle opening distance (β=0.02, p<0.001) after adjusting for covariates. The diagnostic performance of ILCA for detecting GAC was acceptable (AUC=0.69).

Conclusions
ILCA is a significant predictor of angle closure independent of other biometric factors and may reflect unique anatomical information associated with pupillary block. ILCA represents a novel biometric risk factor in eyes with angle closure.