bluuebunny/crossref_metadata_2025_split huggingface dataset:
Anaerobic benzene oxidation in
Geotalea daltonii
involves activation by methylation and is regulated by the transition state regulator AbrB
Authors: Bullows James E. , Kanak Alison , Shedrick Lawrence , Kiessling Christina , Aklujkar Muktak , Kostka Joel , Chin Kuk-Jeong 
Published: September 2024

ABSTRACT

Benzene is a widespread groundwater contaminant that persists under anoxic conditions. The aim of this study was to more accurately investigate anaerobic microbial degradation pathways to predict benzene fate and transport. Preliminary genomic analysis of
Geotalea daltonii
strain FRC-32, isolated from contaminated groundwater, revealed the presence of putative aromatic-degrading genes.
G. daltonii
was subsequently shown to conserve energy for growth on benzene as the sole electron donor and fumarate or nitrate as the electron acceptor. The
hbs
gene, encoding for 3-hydroxybenzylsuccinate synthase (Hbs), a homolog of the radical-forming, toluene-activating benzylsuccinate synthase (Bss), was upregulated during benzene oxidation in
G. daltonii
, while the
bss
gene was upregulated during toluene oxidation. Addition of benzene to the
G. daltonii
whole-cell lysate resulted in toluene formation, indicating that methylation of benzene was occurring. Complementation of σ
54
-
(deficient)
E. coli
transformed with the
bss
operon restored its ability to grow in the presence of toluene, revealing
bss
to be regulated by σ
54
. Binding sites for σ
70
and the transition state regulator AbrB were identified in the promoter region of the σ
54
-encoding gene
rpoN,
and binding was confirmed. Induced expression of
abrB
during benzene and toluene degradation caused
G. daltonii
cultures to transition to the death phase. Our results suggested that
G. daltonii
can anaerobically oxidize benzene by methylation, which is regulated by σ
54
and AbrB. Our findings further indicated that the benzene, toluene, and benzoate degradation pathways converge into a single metabolic pathway, representing a uniquely efficient approach to anaerobic aromatic degradation in
G. daltonii
.

IMPORTANCE

The contamination of anaerobic subsurface environments including groundwater with toxic aromatic hydrocarbons, specifically benzene, toluene, ethylbenzene, and xylene, has become a global issue. Subsurface groundwater is largely anoxic, and further study is needed to understand the natural attenuation of these compounds. This study elucidated a metabolic pathway utilized by the bacterium
Geotalea daltonii
capable of anaerobically degrading the recalcitrant molecule benzene using a unique activation mechanism involving methylation. The identification of aromatic-degrading genes and AbrB as a regulator of the anaerobic benzene and toluene degradation pathways provides insights into the mechanisms employed by
G. daltonii
to modulate metabolic pathways as necessary to thrive in anoxic contaminated groundwater. Our findings contribute to the understanding of novel anaerobic benzene degradation pathways that could potentially be harnessed to develop improved strategies for bioremediation of groundwater contaminants.
Molecular insights into the hydrolysis and transglycosylation of a deep-sea
Planctomycetota
-derived GH16 family laminarinase
Authors: Li Wei , Lin Shanshan , Wang Xianjie , Chen Shiting , Long Lijuan , Yang Jian 
Published: September 2024

ABSTRACT

The biochemical and structural characteristics of
Pt
Lam, a laminarinase from deep-sea
Planctomycetota
, have been extensively elucidated, unveiling the fundamental molecular mechanisms governing substrate recognition and enzymatic catalysis.
Pt
Lam functions as an exo-laminarinase with the ability to sequentially hydrolyze laminarin, cleaving glucose units individually. Notably,
Pt
Lam exhibits proficient transglycosylation capabilities, utilizing various sugar alcohols as acceptors, with lyxose, in particular, yielding exclusively transglycosylated products. Structural analysis of both apo-
Pt
Lam and its laminarin oligosaccharide-bound complex revealed significant conformational alterations in active residues upon substrate binding. Moreover, pivotal residues involved in substrate recognition were identified, with subsequent mutation assays indicating the contribution of positive subsites in modulating exo-hydrolysis and transglycosidic activities. These results enhance our comprehension of laminarin cycling mechanisms by marine
Planctomycetota
, while also providing essential enzyme components for laminarin hetero-oligosaccharide synthesis.

IMPORTANCE

The ubiquitous
Planctomycetota
, with distinctive physiological traits, exert a significant influence on global carbon and nitrogen fluxes. Their intimate association with algae suggests a propensity for efficient polysaccharide degradation; however, research on glycoside hydrolases derived from
Planctomycetota
remains scarce. Herein, we unveil the GH16 family laminarinase
Pt
Lam from deep-sea
Planctomycetota
, shedding light on its catalytic mechanisms underlying hydrolysis and transglycosylation. Our findings elucidate the enzymatic pathways governing the marine laminarin cycle orchestrated by
Planctomycetota,
thereby fostering the exploration of novel polysaccharide hydrolases with promising practical implications.
Application and antagonistic mechanisms of atoxigenic
Aspergillus
strains for the management of fungal plant diseases
Authors: Wang Suyan , Wang Yanxia , Shi Xinchi , Herrera-Balandrano Daniela D. , Chen Xin , Liu Fengquan , Laborda Pedro 
Published: September 2024

ABSTRACT

This review covers, for the first time, all methods based on the use of
Aspergillus
strains as biocontrol agents for the management of plant diseases caused by fungi and oomycetes. Atoxigenic
Aspergillus
strains have been screened in a variety of hosts, such as peanuts, maize kernels, and legumes, during the preharvest and postharvest stages. These strains have been screened against a wide range of pathogens, such as
Fusarium
,
Phytophthora
, and
Pythium
species, suggesting a broad applicability spectrum. The highest efficacies were generally observed when using non-toxigenic
Aspergillus
strains for the management of mycotoxin-producing
Aspergillus
strains. The modes of action included the synthesis of antifungal metabolites, such as kojic acid and volatile organic compounds (VOCs), secretion of hydrolytic enzymes, competition for space and nutrients, and induction of disease resistance.
Aspergillus
strains degraded
Sclerotinia sclerotiorum
sclerotia, showing high control efficacy against this pathogen. Collectively, although two
Aspergillus
strains have been commercialized for aflatoxin degradation, a new application of
Aspergillus
strains is emerging and needs to be optimized.
Infections caused by
Haemophilus ducreyi:
one organism, two stories
Authors: Al-Tawfiq Jaffar A. , Spinola Stanley M. 
Published: September 2024

SUMMARY

Chancroid, a sexually transmitted infection caused by
Haemophilus ducreyi
, is characterized by painful genital ulcers (GU) and inguinal lymphadenitis.
H. ducreyi
was recently described as a major cause of non-sexually transmitted cutaneous ulcers (CU) on the lower legs in children in yaws-endemic regions. This review explores the relationship between CU and GU strains of
H. ducreyi
; their clinical presentation, diagnosis, epidemiology, and treatment; and how findings from a human challenge model relate to GU and CU. We contrast the decline of GU with the persistence of CU caused by
H. ducreyi
. Factors such as transmission dynamics, control, and elimination efforts are discussed. Syndromic management and targeted treatment of sex workers can eradicate chancroid, while skin colonization by CU strains and environmental factors may necessitate topical treatments or vaccination for CU eradication. Efforts should focus on identifying additional reservoirs of CU strains, improving hygiene, and eliminating asymptomatic colonization to eradicate this painful infection in children.
Palmitoylation of SARS-CoV-2 Envelope protein is central to virus particle formation
Authors: Wang Zhaohuan , Qiu Manman , Ji Yue , Chai Keli , Liu Chenxi , Xu Fengwen , Guo Fei , Tan Juan , Liu Ruikang , Qiao Wentao 
Published: September 2024

ABSTRACT

The Envelope (E) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an integral structural protein in the virus particles. However, its role in the assembly of virions and the underlying molecular mechanisms are yet to be elucidated, including whether the function of E protein is regulated by post-translational modifications. In the present study, we report that SARS-CoV-2 E protein is palmitoylated at C40, C43, and C44 by palmitoyltransferases zDHHC3, 6, 12, 15, and 20. Mutating these three cysteines to serines (C40/43/44S) reduced the stability of E protein, decreased the interaction of E with structural proteins Spike, Membrane, and Nucleocapsid, and thereby inhibited the production of virus-like particles (VLPs) and VLP-mediated luciferase transcriptional delivery. Specifically, the C40/43/44S mutation of E protein reduced the density of VLPs. Collectively, these results demonstrate that palmitoylation of E protein is vital for its function in the assembly of SARS-CoV-2 particles.

IMPORTANCE
In this study, we systematically examined the biochemistry of palmitoylation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) E protein and demonstrated that palmitoylation of SARS-CoV-2 E protein is required for virus-like particle (VLP) production and maintaining normal particle density. These results suggest that palmitoylated E protein is central for proper morphogenesis of SARS-CoV-2 VLPs in densities required for viral infectivity. This study presents a significant advancement in the understanding of how palmitoylation of viral proteins is vital for assembling SARS-CoV-2 particles and supports that palmitoyl acyltransferases can be potential therapeutic targets for the development of SARS-CoV-2 inhibitors.
Elucidating the molecular properties and anti-mycobacterial activity of cysteine peptidase domain of D29 mycobacteriophage endolysin
Authors: Gangakhedkar Rutuja , Jain Vikas 
Published: September 2024

ABSTRACT

Emergence of antibiotic resistance in pathogenic
Mycobacterium tuberculosis
(
Mtb
) has elevated tuberculosis to a serious global threat, necessitating alternate solutions for its eradication. D29 mycobacteriophage can infect and kill several mycobacterial species including
Mtb
. It encodes an endolysin LysA to hydrolyze host bacteria peptidoglycan for progeny release. We previously showed that out of the two catalytically active domains of LysA [N-terminal domain (NTD) and lysozyme-like domain], NTD, when ectopically expressed in
Mycobacterium smegmatis
(
Msm
), is able to kill the bacterium nearly as efficiently as full-length LysA. Here, we dissected the functioning of NTD to develop it as a phage-derived small molecule anti-mycobacterial therapeutic. We performed a large-scale site-directed mutagenesis of the conserved residues in NTD and examined its structure, stability, and function using molecular dynamic simulations coupled with biophysical and biochemical experiments. Our data show that NTD functions as a putative cysteine peptidase with a catalytic triad composed of Cys41, His112, and Glu137, acting as nucleophile, base, and acid, respectively, and showing characteristics similar to the NlpC/P60 family of cysteine peptidases. Additionally, our peptidoglycan hydrolysis assays suggested that NTD hydrolyzes only mycobacterial peptidoglycan and does not act on Gram-positive and Gram-negative bacterial peptidoglycans. More importantly, the combined activity of exogenously added NTD and sub-lethal doses of anti-mycobacterial drugs kills
Msm in vitro
and exhibits disruption of pre-formed mycobacterial biofilm. We additionally show that NTD treatment increases the permeability of antibiotics in
Msm
, which reduces the minimum inhibitory concentration of the antibiotics. Collectively, we present NTD as a promising phage-derived therapeutic against mycobacteria.

IMPORTANCE

Mycobacteriophages are the viruses that use mycobacteria as host for their progeny production and, in the process, kill them. Mycobacteriophages are, therefore, considered as promising alternatives to antibiotics for killing pathogenic
Mycobacterium tuberculosis
. The endolysin LysA produced by mycobacteriophage D29 plays an important role in host cell lysis and virion release. Our work presented here highlights the functioning of LysA’s N-terminal catalytic domain (NTD) in order to develop it as phage-derived small molecule therapeutics. We show that combined treatment of exogenously added NTD and sub-lethal doses of anti-mycobacterial drugs kills
M. smegmatis
, shows synergism by reducing the minimum inhibitory concentration of these antibiotics, and exhibits disruption of pre-formed mature biofilm. These outcomes and our detailed biochemical and biophysical dissection of the protein further pave the way toward engineering and development of NTD as a promising therapeutic against mycobacterial infections such as tuberculosis.
Alterations in circulating immunoregulatory proteins discriminate poor CD4 T lymphocyte trajectories in people with HIV on suppressive antiretroviral therapy
Authors: Moar Preeti , Bowler Scott , Landay Alan L. , Gianella Sara , Ndhlovu Lishomwa C. , Premeaux Thomas A. 
Published: September 2024

ABSTRACT

Despite the success in the management of HIV with antiretroviral therapy (ART), people with HIV (PWH) have a heightened state of immune activation and inflammation, and an estimated 10%–40% demonstrate poor CD4 T-cell reconstitution, thereby increasing their mortality and morbidity risk burden. Soluble immunoregulatory proteins that function in lymphocyte activation or inhibition are elevated in PWH and associate with T-cell dysfunction, HIV persistence, and are predictive of comorbid outcomes. Here, we measured a panel of 35 circulating immunoregulatory proteins in 116 PWH with variations in CD4 T-cell counts (poor CD4 trajectory: <200 cells/μl,
n
= 34 or immune competent: CD4 >500 cells/μl,
n
= 82) by Luminex. Participants were enrolled in the AIDS Clinical Trials Group Longitudinal Linked Randomized Trials cohort, had initiated ART on enrollment, and had been on suppressive ART for 1 year. Using non-parametric analysis, we found that the levels of CD276, ICOSL, BAFF, OX40, galectin-1, and galectin-9 were significantly higher in PWH with poor CD4 trajectories compared to individuals with immune-competent CD4 T-cell count. Notably, in logistic models, ICOSL and OX40 remained significant after adjusting for age and baseline plasma HIV RNA. Furthermore, Extreme Gradient Boosting machine learning models comprising co-stimulatory and inhibitory checkpoint proteins yielded high accuracy in classification of individuals with poor CD4 trajectories. In summary, we identified a novel signature of circulating immunoregulatory proteins indicative of poor CD4 trajectories that may serve as potential targets to monitor and manage immune perturbations more accurately in PWH during suppressive ART.

IMPORTANCE
It is essential to track immune perturbations related to insufficient CD4 T-cell recovery in PWH on suppressive ART as those with incomplete reconstitution are at a greater risk of non-AIDS-related morbidity and mortality. Several inflammatory soluble mediators have associated with poor immune reconstitution and adverse morbid outcomes in PWH, yet their implementation into routine clinical care to guide management remains inconsistent. Circulating immune checkpoint proteins have been linked to dysregulated immune pathways during suppressive ART and may serve as improved surrogate markers of clinical relevance. Here we investigate soluble lymphocyte-associated immunoregulatory proteins in virally suppressed PWH with no reported co-morbid outcomes and varying CD4 T-cell counts, to reveal underlying pathways that remain perturbed despite ART. This novel signature of immunoregulatory markers pertaining to poor CD4 T-cell trajectories uncover previously overlooked immune checkpoints as important targets for clinical monitoring of PWH in the setting of durable viral suppression by ART.
Genome-wide fitness analysis of
Salmonella enterica
reveals
aroA
mutants are attenuated due to iron restriction
in vitro
Authors: Rooke Jessica L. , Goodall Emily C. A. , Pullela Karthik , Da Costa Rochelle , Martinelli Nicole , Smith Chelsie , Mora Maria , Cunningham Adam F. , Henderson Ian R. 
Published: September 2024

ABSTRACT

Salmonella enterica
is a globally disseminated pathogen that is the cause of over 100 million infections per year. The resulting diseases are dependent upon host susceptibility and the infecting serovar. As
S. enterica
serovar Typhimurium induces a typhoid-like disease in mice, this model has been used extensively to illuminate various aspects of
Salmonella
infection and host responses. Due to the severity of infection in this model, researchers often use strains of mice resistant to infection or attenuated
Salmonella
. Despite decades of research, many aspects of
Salmonella
infection and fundamental biology remain poorly understood. Here, we use a transposon insertion sequencing technique to interrogate the essential genomes of widely used isogenic wild-type and attenuated
S
. Typhimurium strains. We reveal differential essential pathways between strains
in vitro
and provide a direct link between iron starvation, DNA synthesis, and bacterial membrane integrity.

IMPORTANCE

Salmonella enterica
is an important clinical pathogen that causes a high number of deaths and is increasingly resistant to antibiotics. Importantly,
S. enterica
is used widely as a model to understand host responses to infection. Understanding how Salmonella survives
in vivo
is important for the design of new vaccines to combat this pathogen. Live attenuated vaccines have been used clinically for decades. A widely used mutation,
aroA
, is thought to attenuate
Salmonella
by restricting the ability of the bacterium to access particular amino acids. Here we show that this mutation limits the ability of
Salmonella
to acquire iron. These observations have implications for the interpretation of many previous studies and for the use of
aroA
in vaccine development.
Genetic requirements for uropathogenic
E. coli
proliferation in the bladder cell infection cycle
Authors: Mediati Daniel G. , Blair Tamika A. , Costas Ariana , Monahan Leigh G. , Söderström Bill , Charles Ian G. , Duggin Iain G. 
Published: September 2024

ABSTRACT

Uropathogenic
Escherichia coli
(UPEC) requires an adaptable physiology to survive the wide range of environments experienced in the host, including gut and urinary tract surfaces. To identify UPEC genes required during intracellular infection, we developed a transposon-directed insertion-site sequencing approach for cellular infection models and searched for genes in a library of ~20,000 UTI89 transposon-insertion mutants that are specifically required at the distinct stages of infection of cultured bladder epithelial cells. Some of the bacterial functional requirements apparent in host bladder cell growth overlapped with those for M9-glycerol, notably nutrient utilization, polysaccharide and macromolecule precursor biosynthesis, and cell envelope stress tolerance. Two genes implicated in the intracellular bladder cell infection stage were confirmed through independent gene deletion studies:
neuC
(sialic acid capsule biosynthesis) and
hisF
(histidine biosynthesis). Distinct sets of UPEC genes were also implicated in bacterial dispersal, where UPEC erupts from bladder cells in highly filamentous or motile forms upon exposure to human urine, and during recovery from infection in a rich medium. We confirm that the
dedD
gene linked to septal peptidoglycan remodeling is required during UPEC dispersal from human bladder cells and may help stabilize cell division or the cell wall during envelope stress created by host cells. Our findings support a view that the host intracellular environment and infection cycle are multi-nutrient limited and create stress that demands an array of biosynthetic, cell envelope integrity, and biofilm-related functions of UPEC.

IMPORTANCE

Urinary tract infections (UTIs) are one of the most frequent infections worldwide. Uropathogenic
Escherichia coli
(UPEC), which accounts for ~80% of UTIs, must rapidly adapt to highly variable host environments, such as the gut, bladder sub-surface, and urine. In this study, we searched for UPEC genes required for bacterial growth and survival throughout the cellular infection cycle. Genes required for
de novo
synthesis of biomolecules and cell envelope integrity appeared to be important, and other genes were also implicated in bacterial dispersal and recovery from infection of cultured bladder cells. With further studies of individual gene function, their potential as therapeutic targets may be realized. This study expands knowledge of the UTI cycle and establishes an approach to genome-wide functional analyses of stage-resolved microbial infections.
Diversity and impact of single-stranded RNA viruses in Czech
Heterobasidion
populations
Authors: Dálya László Benedek , Černý Martin , de la Peña Marcos , Poimala Anna , Vainio Eeva J. , Hantula Jarkko , Botella Leticia 
Published: September 2024

ABSTRACT

Heterobasidion annosum sensu lato
comprises some of the most devastating pathogens of conifers. Exploring virocontrol as a potential strategy to mitigate economic losses caused by these fungi holds promise for the future. In this study, we conducted a comprehensive screening for viruses in 98
H
.
annosum
s.l. specimens from different regions of Czechia aiming to identify viruses inducing hypovirulence. Initial examination for dsRNA presence was followed by RNA-seq analyses using pooled RNA libraries constructed from
H. annosum
and
Heterobasidion parviporum
, with diverse bioinformatic pipelines employed for virus discovery. Our study uncovered 25 distinct ssRNA viruses, including two ourmia-like viruses, one mitovirus, one fusarivirus, one tobamo-like virus, one cogu-like virus, one bisegmented narna-like virus and one segment of another narna-like virus, and 17 ambi-like viruses, for which hairpin and hammerhead ribozymes were detected. Coinfections of up to 10 viruses were observed in six
Heterobasidion
isolates, whereas another six harbored a single virus. Seventy-three percent of the isolates analyzed by RNA-seq were virus-free. These findings show that the virome of
Heterobasidion
populations in Czechia is highly diverse and differs from that in the boreal region. We further investigated the host effects of certain identified viruses through comparisons of the mycelial growth rate and proteomic analyses and found that certain tested viruses caused growth reductions of up to 22% and significant alterations in the host proteome profile. Their intraspecific transmission rates ranged from 0% to 33%. Further studies are needed to fully understand the biocontrol potential of these viruses
in planta
.

IMPORTANCE

Heterobasidion annosum sensu lato
is a major pathogen causing significant damage to conifer forests, resulting in substantial economic losses. This study is significant as it explores the potential of using viruses (virocontrol) to combat these fungal pathogens. By identifying and characterizing a diverse array of viruses in
H. annosum
populations from Czechia, the research opens new avenues for biocontrol strategies. The discovery of 25 distinct ssRNA viruses, some of which reduce fungal growth and alter proteome profiles, suggests that these viruses could be harnessed to mitigate the impact of
Heterobasidion
. Understanding the interactions between these viruses and their fungal hosts is crucial for developing effective, environmentally friendly methods to protect conifer forests and maintain ecosystem health. This study lays the groundwork for future research on the application of mycoviruses in forest disease management.