bluuebunny/crossref_metadata_2025 huggingface dataset:
Queen bee gut microbiota extends honeybee lifespan by inhibiting insulin signaling
Authors: Wang Hongfang , Chen Wenfeng , Lei Li , Zhang Wei , Liu Zhenguo , Wang Ying , Xu Baohua
Published: March 2024

Queen and worker bees are natural models for aging research, as their lifespans vary considerably independent of genetic variation. Investigating the reasons why queens live longer than workers is of great significance for research on the universal processes of aging in animals. The gut microbiome has received attention as a vital regulator of host health, while its precise role in honeybee aging needs further investigation. The effects and mechanisms behind the relationship between gut microbiota and worker lifespan were measured by transplanting queen bee gut bacteria (QG) and worker bee gut bacteria (WG) into microbiota-free (MF) workers. The transplantation of QG to MF bees significantly extended the workers’ lifespans compared with MF and WG bees. Untargeted metabolomics identified 49 lifespan-related differential metabolites, and Kyoto Encyclopedia of Genes and Genomes analysis of these revealed three lifespan-related metabolic pathways: insulin/insulin-like growth factor signaling, immune, and ketone body metabolism pathways. Further verification showed that QG inhibited the expression of insulin-like peptides (ILPs), and the expression of ILPs was lower in natural queens than in natural workers. QG transplantation also stimulated the expression of antioxidant genes and lowered oxidative damage products in natural queen bees. However, gut microbiota transplantation failed to mimic the immune properties and ketone body metabolism profiles of natural queens and workers. Concisely, QG could increase the antioxidant capacity to extend lifespan by inhibiting insulin signaling. These findings may help determine the mechanisms behind queen longevity and provide further insights into the role of gut symbionts.

IMPORTANCE

Queen and worker bees share the same genetic background but have vastly different lifespans. The gut microbiome regulates host health, suggesting that differences in lifespan between queen and worker bees could be related to gut bacteria. Herein, we used an innovative method to transplant gut microbiota from adult queen or worker bees to microbiota-free bees. The transplantation of queen gut microbiota to microbiota-free bees extended their lifespan. Insulin/insulin-like growth factor signaling, a highly conserved metabolic pathway related to lifespan, displayed identical expression profiles in natural queen bees and microbiota-free bees transplanted with queen microbiota. This finding significantly expands our understanding of the relationships between intestinal bacteria, host health, and the biology of aging.
The white rot basidiomycete
Gelatoporia subvermispora
produces fatty aldehydes that enable fungal manganese peroxidases to degrade recalcitrant lignin structures
Authors: Kapich Alexander N. , Suzuki Hideki , Hirth Kolby C. , Fernández-Fueyo Elena , Martínez Angel T. , Houtman Carl J. , Hammel Kenneth E.
Published: March 2024

The ability of some white rot basidiomycetes to remove lignin selectively from wood indicates that low molecular weight oxidants have a role in ligninolysis. These oxidants are likely free radicals generated by fungal peroxidases from compounds in the biodegrading wood. Past work supports a role for manganese peroxidases (MnPs) in the production of ligninolytic oxidants from fungal membrane lipids. However, the fatty acid alkylperoxyl radicals initially formed during this process are not reactive enough to attack the major structures in lignin. Here, we evaluate the hypothesis that the peroxidation of fatty aldehydes might provide a source of more reactive acylperoxyl radicals. We found that
Gelatoporia subvermispora
produced
trans-
2-nonenal,
trans-2-
octenal, and n-hexanal (a likely metabolite of
trans-
2,4-decadienal) during the incipient decay of aspen wood. Fungal fatty aldehydes supported the
in vitro
oxidation by MnPs of a nonphenolic lignin model dimer, and also of the monomeric model veratryl alcohol. Experiments with the latter compound showed that the reactions were partially inhibited by oxalate, the chelator that white rot fungi employ to detach Mn
3+
from the MnP active site, but nevertheless proceeded at its physiological concentration of 1 mM. The addition of catalase was inhibitory, which suggests that the standard MnP catalytic cycle is involved in the oxidation of aldehydes. MnP oxidized
trans-
2-nonenal quantitatively to
trans-
2-nonenoic acid with the consumption of one O
2
equivalent. The data suggest that when Mn
3+
remains associated with MnP, it can oxidize aldehydes to their acyl radicals, and the latter subsequently add O
2
to become ligninolytic acylperoxyl radicals.

IMPORTANCE

The biodegradation of lignin by white rot fungi is essential for the natural recycling of plant biomass and has useful applications in lignocellulose bioprocessing. Although fungal peroxidases have a key role in ligninolysis, past work indicates that biodegradation is initiated by smaller, as yet unidentified oxidants that can infiltrate the substrate. Here, we present evidence that the peroxidase-catalyzed oxidation of naturally occurring fungal aldehydes may provide a source of ligninolytic free radical oxidants.
Testing for SARS-CoV-2: lessons learned and current use cases
Authors: Theel Elitza S. , Kirby James E. , Pollock Nira R.
Published: March 2024

The emergence and worldwide dissemination of SARS-CoV-2 required both urgent development of new diagnostic tests and expansion of diagnostic testing capacity on an unprecedented scale. The rapid evolution of technologies that allowed testing to move out of traditional laboratories and into point-of-care testing centers and the home transformed the diagnostic landscape. Four years later, with the end of the formal public health emergency but continued global circulation of the virus, it is important to take a fresh look at available SARS-CoV-2 testing technologies and consider how they should be used going forward. This review considers current use case scenarios for SARS-CoV-2 antigen, nucleic acid amplification, and immunologic tests, incorporating the latest evidence for analytical/clinical performance characteristics and advantages/limitations for each test type to inform current debates about how tests should or should not be used.
Corrected and republished from: “Extracellular Vesicle Formation in
Cryptococcus deuterogattii
Impacts Fungal Virulence”
Authors: Castelli Rafael F. , Pereira Alana , Honorato Leandro , Valdez Alessandro , de Oliveira Haroldo C. , Bazioli Jaqueline M. , Garcia Ane W. A. , Klimeck Tabata D'Maiella Freitas , Reis Flavia C. G. , Camillo-Andrade Amanda C. , Santos Marlon D. M. , Carvalho Paulo C. , Zaragoza Oscar , Staats Charley C. , Nimrichter Leonardo , Fill Taícia P. , Rodrigues Marcio L.
Published: March 2024

Small molecules are components of fungal extracellular vesicles (EVs), but their biological roles are only superficially known.
NOP16
is a eukaryotic gene that is required for the activity of benzimidazoles against
Cryptococcus deuterogattii
. In this study, during the phenotypic characterization of
C. deuterogattii
mutants expected to lack
NOP16
expression, we observed a reduced EV production. Whole-genome sequencing, RNA-Seq, and cellular proteomics revealed that, contrary to our initial findings, these mutants expressed Nop16 but exhibited altered expression of 14 genes potentially involved in sugar transport. Based on this observation, we designated these mutant strains as Past1 and Past2, representing
p
otentially
a
ltered
s
ugar
t
ransport. Analysis of the small molecule composition of EVs produced by wild-type cells and the Past1 and Past2 mutant strains revealed not only a reduced number of EVs but also an altered small molecule composition. In a
Galleria mellonella
model of infection, the Past1 and Past2 mutant strains were hypovirulent. The hypovirulent phenotype was reverted when EVs produced by wild-type cells, but not mutant EVs, were co-injected with the mutant cells in
G. mellonella
. These results connect EV biogenesis, cargo, and cryptococcal virulence.
Single-domain antibodies reveal unique borrelicidal epitopes on the Lyme disease vaccine antigen, outer surface protein A (OspA)
Authors: Vance David J. , Basir Saiful , Piazza Carol Lyn , Willsey Graham G. , Haque H. M. Emranul , Tremblay Jacque M. , Rudolph Michael J. , Muriuki Beatrice , Cavacini Lisa , Weis David D. , Shoemaker Charles B. , Mantis Nicholas J.
Published: March 2024

Camelid-derived, single-domain antibodies (V
H
Hs) have proven to be extremely powerful tools in defining the antigenic landscape of immunologically heterogeneous surface proteins. In this report, we generated a phage-displayed V
H
H library directed against the candidate Lyme disease vaccine antigen, outer surface protein A (OspA). Two alpacas were immunized with recombinant OspA serotype 1 from
Borrelia burgdorferi sensu stricto
strain B31, in combination with the canine vaccine RECOMBITEK Lyme containing lipidated OspA. The phage library was subjected to two rounds of affinity enrichment (“panning”) against recombinant OspA, yielding 21 unique V
H
Hs within two epitope bins, as determined through competition enzyme linked immunosorbent assays (ELISAs) with a panel of OspA-specific human monoclonal antibodies. Epitope refinement was conducted by hydrogen exchange-mass spectrometry. Six of the monovalent V
H
Hs were expressed as human IgG1-Fc fusion proteins and shown to have functional properties associated with protective human monoclonal antibodies, including
B. burgdorferi
agglutination, outer membrane damage, and complement-dependent borreliacidal activity. The V
H
Hs displayed unique reactivity profiles with the seven OspA serotypes associated with
B. burgdorferi
genospecies in the United States and Europe consistent with there being unique epitopes across OspA serotypes that should be considered when designing and evaluating multivalent Lyme disease vaccines.
Salmonella
Enteritidis antitoxin DinJ inhibits NLRP3-dependent canonical inflammasome activation in macrophages
Authors: Gu Dan , Li Ang , Zang Xirui , Huang Tingting , Guo Yaxin , Jiao Xinan , Pan Zhiming
Published: March 2024

The inflammasome is a pivotal component of the innate immune system, acting as a multiprotein complex that plays an essential role in detecting and responding to microbial infections.
Salmonella
Enteritidis have evolved multiple mechanisms to regulate inflammasome activation and evade host immune system clearance. Through screening
S
. Enteritidis C50336Δ
fliC
transposon mutant library, we found that the insertion mutant of
dinJ
increased inflammasome activation. In this study, we demonstrated the genetic connection between the antitoxin DinJ and the toxin YafQ in
S
. Enteritidis, confirming their co-transcription. The deletion mutant Δ
fliC
Δ
dinJ
increased cell death and IL-1β secretion in J774A.1 cells. Western blotting analysis further showed elevated cleaved Caspase-1 product (p10 subunits) and IL-1β secretion in cells infected with Δ
fliC
Δ
dinJ
compared to cells infected with Δ
fliC
. DinJ was found to inhibit canonical inflammasome activation using primary bone marrow-derived macrophages (BMDMs) from
Casp
-/-
C57BL/6 mice. Furthermore, DinJ specifically inhibited NLRP3 inflammasome activation, as demonstrated in BMDMs from
Nlrp3

-/-

and
Nlrc4

-/-

mice. Fluorescence resonance energy transfer (FRET) experiments confirmed the translocation of DinJ into host cells during infection. Finally, we revealed that DinJ could inhibit the secretion of IL-1β and IL-18
in vivo
, contributing to
S
. Enteritidis evading host immune clearance. In summary, our findings provide insights into the role of DinJ in modulating the inflammasome response during
S
. Enteritidis infection, highlighting its impact on inhibiting inflammasome activation and immune evasion.
Distinct dynamics and proximity networks of hub proteins at the prey-invading cell pole in a predatory bacterium
Authors: Remy Ophélie , Santin Yoann G. , Jonckheere Veronique , Tesseur Coralie , Kaljević Jovana , Van Damme Petra , Laloux Géraldine
Published: March 2024

In bacteria, cell poles function as subcellular compartments where proteins localize during specific lifecycle stages, orchestrated by polar “hub” proteins. Whereas most described bacteria inherit an “old” pole from the mother cell and a “new” pole from cell division, generating cell asymmetry at birth, non-binary division poses challenges for establishing cell polarity, particularly for daughter cells inheriting only new poles. We investigated polarity dynamics in the obligate predatory bacterium
Bdellovibrio bacteriovorus
, proliferating through filamentous growth followed by non-binary division within prey bacteria. Monitoring the subcellular localization of two proteins known as polar hubs in other species, RomR and DivIVA, revealed RomR as an early polarity marker in
B. bacteriovorus
. RomR already marks the future anterior poles of the progeny during the predator’s growth phase, during a precise period closely following the onset of divisome assembly and the end of chromosome segregation. In contrast to RomR’s stable unipolar localization in the progeny, DivIVA exhibits a dynamic pole-to-pole localization. This behavior changes shortly before the division of the elongated predator cell, where DivIVA accumulates at all septa and both poles.
In vivo
protein interaction networks for DivIVA and RomR, mapped through endogenous miniTurbo-based proximity labeling, further underscore their distinct roles in cell polarization and reinforce the importance of the anterior “invasive” cell pole in prey-predator interactions. Our work also emphasizes the precise spatiotemporal order of cellular processes underlying
B. bacteriovorus
proliferation, offering insights into the subcellular organization of bacteria with filamentous growth and non-binary division.

IMPORTANCE

In bacteria, cell poles are crucial areas where “hub” proteins orchestrate lifecycle events through interactions with multiple partners at specific times. While most bacteria exhibit one “old” and one “new” pole, inherited from the previous division event, setting polar identity poses challenges in bacteria with non-binary division. This study explores polar proteins in the predatory bacterium
Bdellovibrio bacteriovorus
, which undergoes filamentous growth followed by non-binary division inside another bacterium. Our research reveals distinct localization dynamics of the polar proteins RomR and DivIVA, highlighting RomR as an early “hub” marking polar identity in the filamentous mother cell. Using miniTurbo-based proximity labeling, we uncovered their unique protein networks. Overall, our work provides new insights into the cell polarity in non-binary dividing bacteria.
Peeling the onion: additional layers of regulation in the acid stress response
Authors: Kenney Linda J.
Published: March 2024

Bacteria are capable of withstanding large changes in osmolality and cytoplasmic pH, unlike eukaryotes that tightly regulate their pH and cellular composition. Previous studies on the bacterial acid stress response described a rapid, brief acidification, followed by immediate recovery. More recent experiments with better pH probes have imaged single living cells, and we now appreciate that following acid stress, bacteria maintain an acidic cytoplasm for as long as the stress remains. This acidification enables pathogens to sense a host environment and turn on their virulence programs, for example, enabling survival and replication within acidic vacuoles. Single-cell analysis identified an intracellular pH threshold of ~6.5. Acid stress reduces the internal pH below this threshold, triggering the assembly of a type III secretion system in
Salmonella
and the secretion of virulence factors in the host. These pathways are significant because preventing intracellular acidification of
Salmonella
renders it avirulent, suggesting that acid stress pathways represent a potential therapeutic target. Although we refer to the acid stress response as singular, it is actually a complex response that involves numerous two-component signaling systems, several amino acid decarboxylation systems, as well as cellular buffering systems and electron transport chain components, among others. In a recent paper in the
Journal of Bacteriology
, M. G. Gorelik, H. Yakhnin, A. Pannuri, A. C. Walker, C. Pourciau, D. Czyz, T. Romeo, and P. Babitzke (J Bacteriol 206:e00354-23, 2024,
https://doi.org/10.1128/jb.00354-23
) describe a new connection linking the carbon storage regulator CsrA to the acid stress response, highlighting new additional layers of complexity.
The intrinsically disordered linker in the single-stranded DNA-binding protein influences DNA replication restart and recombination pathways in
Escherichia coli
K-12
Authors: Sandler Steven J. , Bonde Nina J. , Wood Elizabeth A. , Cox Michael M. , Keck James L.
Published: March 2024

Tetrameric single-stranded (ss) DNA-binding proteins (SSBs) stabilize ssDNA intermediates formed during genome maintenance reactions in
Bacteria
. SSBs also recruit proteins important for these processes through direct SSB-protein interactions, including proteins involved in DNA replication restart and recombination processes. SSBs are composed of an N-terminal oligomerization and ssDNA-binding domain, a C-terminal acidic tip that mediates SSB-protein interactions, and an internal intrinsically disordered linker (IDL). Deletions and insertions into the IDL are well tolerated with few phenotypes, although the largest deletions and insertions exhibit some sensitivity to DNA-damaging agents. To define specific DNA metabolism processes dependent on IDL length,
ssb
mutants that lack 16, 26, 37, or 47 residues of the 57-residue IDL were tested for synthetic phenotypes with mutations in DNA replication restart or recombination genes. We also tested the impact of integrating a fluorescent domain within the SSB IDL using an
ssb::mTur2
insertion mutation. Only the largest deletion tested or the insertion mutation causes sensitivity in any of the pathways. Mutations in two replication restart pathways (PriA-B
1
and PriA-C) showed synthetic lethalities or small colony phenotypes with the largest deletion or insertion mutations. Recombination gene mutations
del(recBCD
) and
del(ruvABC
) show synthetic phenotypes only when combined with the largest
ssb
deletion. These results suggest that a minimum IDL length is important in some genome maintenance reactions in
Escherichia coli
. These include pathways involving PriA-PriB
1
, PriA-PriC, RecFOR, and RecG. The mTur2 insertion in the IDL may also affect SSB interactions in some processes, particularly the PriA-PriB
1
and PriA-PriC replication restart pathways.

IMPORTANCE

ssb
is essential in
Escherichia coli
due to its roles in protecting ssDNA and coordinating genome maintenance events. While the DNA-binding core and acidic tip have well-characterized functions, the purpose of the intrinsically disordered linker (IDL) is poorly understood.
In vitro
studies have revealed that the IDL is important for cooperative ssDNA binding and phase separation. However, single-stranded (ss) DNA-binding protein (SSB) variants with large deletions and insertions in the IDL support normal cell growth. We find that the PriA-PriB
1
and PriA-C replication restart, as well as the RecFOR- and RecG-dependent recombination, pathways are sensitive to IDL length. This suggests that cooperativity, phase separation, or a longer spacer between the core and acidic tip of SSB may be important for specific cellular functions.
Development of a sensor for disulfide bond formation in diverse bacteria
Authors: Dyotima  , Abulaila Sally , Mendoza Jocelyne , Landeta Cristina
Published: March 2024

In bacteria, disulfide bonds contribute to the folding and stability of proteins important for processes in the cellular envelope. In
Escherichia coli
, disulfide bond formation is catalyzed by DsbA and DsbB enzymes. DsbA is a periplasmic protein that catalyzes disulfide bond formation in substrate proteins, while DsbB is an inner membrane protein that transfers electrons from DsbA to quinones, thereby regenerating the DsbA active state. Actinobacteria including mycobacteria use an alternative enzyme named VKOR, which performs the same function as DsbB. Disulfide bond formation enzymes, DsbA and DsbB/VKOR, represent novel drug targets because their inhibition could simultaneously affect the folding of several cell envelope proteins including virulence factors, proteins involved in outer membrane biogenesis, cell division, and antibiotic resistance. We have previously developed a cell-based and target-based assay to identify molecules that inhibit the DsbB and VKOR in pathogenic bacteria, using
E. coli
cells expressing a periplasmic β-Galactosidase sensor (β-Gal
dbs
), which is only active when disulfide bond formation is inhibited. Here, we report the construction of plasmids that allows fine-tuning of the expression of the β-Gal
dbs
sensor and can be mobilized into other gram-negative organisms. As an example, when expressed in
Pseudomonas aeruginosa
UCBPP-PA14, which harbors two DsbB homologs, β-Gal
dbs
behaves similarly as in
E. coli
, and the biosensor responds to the inhibition of the two DsbB proteins. Thus, these β-Gal
dbs
reporter plasmids provide a basis to identify novel inhibitors of DsbA and DsbB/VKOR in multidrug-resistant gram-negative pathogens and to further study oxidative protein folding in diverse gram-negative bacteria.

IMPORTANCE

Disulfide bonds contribute to the folding and stability of proteins in the bacterial cell envelope. Disulfide bond-forming enzymes represent new drug targets against multidrug-resistant bacteria because inactivation of this process would simultaneously affect several proteins in the cell envelope, including virulence factors, toxins, proteins involved in outer membrane biogenesis, cell division, and antibiotic resistance. Identifying the enzymes involved in disulfide bond formation in gram-negative pathogens as well as their inhibitors can contribute to the much-needed antibacterial innovation. In this work, we developed sensors of disulfide bond formation for gram-negative bacteria. These tools will enable the study of disulfide bond formation and the identification of inhibitors for this crucial process in diverse gram-negative pathogens.
Oligomeric assembly of the C-terminal and transmembrane region of SARS-CoV-2 nsp3
Authors: Babot Marion , Boulard Yves , Agouda Samira , Pieri Laura , Fieulaine Sonia , Bressanelli Stéphane , Gervais Virginie
Published: March 2024

As for all single-stranded, positive-sense RNA (+RNA) viruses, intracellular RNA synthesis relies on extensive remodeling of host cell membranes that leads to the formation of specialized structures. In the case of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) coronavirus causing COVID-19, endoplasmic reticulum membranes are modified, resulting in the formation of double-membrane vesicles (DMVs), which contain the viral dsRNA intermediate and constitute membrane-bound replication organelles. The non-structural and transmembrane protein nsp3 is a key player in the biogenesis of DMVs and, therefore, represents an interesting antiviral target. However, as an integral transmembrane protein, it is challenging to express for structural biology. The C-terminus of nsp3 encompasses all the membrane-spanning, -interacting, and -remodeling elements. By using a cell-free expression system, we successfully produced the C-terminal region of nsp3 (nsp3C) and reconstituted purified nsp3C into phospholipid nanodiscs, opening the way for structural studies. Negative-stain transmission electron microscopy revealed the presence of nsp3C oligomers very similar to the region abutting and spanning the membrane on the cytosolic side of DMVs in a recent subtomogram average of the SARS-CoV-2 nsp3-4 pore (1). AlphaFold-predicted structural models fit particularly well with our experimental data and support a pore-forming hexameric assembly. Altogether, our data give unprecedented clues to understand the structural organization of nsp3, the principal component that shapes the molecular pore that spans the DMVs and is required for the export of RNA
in vivo
.

IMPORTANCE

Membrane remodeling is at the heart of intracellular replication for single-stranded, positive-sense RNA viruses. In the case of coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), this leads to the formation of a network of double-membrane vesicles (DMVs). Targeting DMV biogenesis offers promising prospects for antiviral therapies. This requires a better understanding of the molecular mechanisms and proteins involved. Three non-structural proteins (nsp3, nsp4, and nsp6) direct the intracellular membrane rearrangements upon SARS-CoV-2 infection. All of them contain transmembrane helices. The nsp3 component, the largest and multi-functional protein of the virus, plays an essential role in this process. Aiming to understand its structural organization, we used a cell-free protein synthesis assay to produce and reconstitute the C-terminal part of nsp3 (nsp3C) including transmembrane domains into phospholipid nanodiscs. Our work reveals the oligomeric organization of one key player in the biogenesis of SARS-CoV-2 DMVs, providing basis for the design of future antiviral strategies.
Human norovirus cultivation systems and their use in antiviral research
Authors: Hayashi Tsuyoshi , Kobayashi Sakura , Hirano Junki , Murakami Kosuke
Published: March 2024

Human norovirus (HuNoV) is a major cause of acute gastroenteritis and foodborne diseases, affecting all age groups. Despite its clinical needs, no approved antiviral therapies are available. Since the discovery of HuNoV in 1972, studies on anti-norovirals, mechanism of HuNoV infection, viral inactivation, etc., have been hampered by the lack of a robust laboratory-based cultivation system for HuNoV. A recent breakthrough in the development of HuNoV cultivation systems has opened opportunities for researchers to investigate HuNoV biology in the context of
de novo
HuNoV infections. A tissue stem cell-derived human intestinal organoid/enteroid (HIO) culture system is one of those that supports HuNoV replication reproducibly and, to our knowledge, is most widely distributed to laboratories worldwide to study HuNoV and develop therapeutic strategies. This review summarizes recently developed HuNoV cultivation systems, including HIO, and their use in antiviral studies.
Functionality of IAV packaging signals depends on site-specific charges within the viral nucleoprotein
Authors: Ciminski Kevin , Flore Viktoria , Jakob Celia , Mues Helen , Smedegaard Frederiksen Anne , Schwemmle Martin , Bolte Hardin , Giese Sebastian
Published: March 2024

The coordinated packaging of the segmented genome of the influenza A virus (IAV) into virions is an essential step of the viral life cycle. This process is controlled by the interaction of packaging signals present in all eight viral RNA (vRNA) segments and the viral nucleoprotein (NP), which binds vRNA via a positively charged binding groove. However, mechanistic models of how the packaging signals and NP work together to coordinate genome packaging are missing. Here, we studied genome packaging in influenza A/SC35M virus mutants that carry mutated packaging signals as well as specific amino acid substitutions at the highly conserved lysine (K) residues 184 and 229 in the RNA-binding groove of NP. Because these lysines are acetylated and thus neutrally charged in infected host cells, we replaced them with glutamine to mimic the acetylated, neutrally charged state or arginine to mimic the non-acetylated, positively charged state. Our analysis shows that the coordinated packaging of eight vRNAs is influenced by (i) the charge state of the replacing amino acid and (ii) its location within the RNA-binding groove. Accordingly, we propose that lysine acetylation induces different charge states within the RNA-binding groove of NP, thereby supporting the activity of specific packaging signals during coordinated genome packaging.

IMPORTANCE

Influenza A viruses (IAVs) have a segmented viral RNA (vRNA) genome encapsidated by multiple copies of the viral nucleoprotein (NP) and organized into eight distinct viral ribonucleoprotein complexes. Although genome segmentation contributes significantly to viral evolution and adaptation, it requires a highly sophisticated genome-packaging mechanism. How eight distinct genome complexes are incorporated into the virion is poorly understood, but previous research suggests an essential role for both vRNA packaging signals and highly conserved NP amino acids. By demonstrating that the packaging process is controlled by charge-dependent interactions of highly conserved lysine residues in NP and vRNA packaging signals, our study provides new insights into the sophisticated packaging mechanism of IAVs.
SP-CHAP, an endolysin with enhanced activity against biofilm pneumococci and nasopharyngeal colonization
Authors: Alreja Adit B. , Appel Amanda E. , Zhu Jinyi C. , Riley Sean P. , Gonzalez-Juarbe Norberto , Nelson Daniel C.
Published: March 2024

Streptococcus pneumoniae
(
Spn
), a Gram-positive bacterium, is responsible for causing a wide variety of invasive infections. The emergence of multi-drug antibiotic resistance has prompted the search for antimicrobial alternatives. Phage-derived peptidoglycan hydrolases, known as endolysins, are an attractive alternative. In this study, an endolysin active against
Spn
, designated SP-CHAP, was cloned, produced, purified, biochemically characterized, and evaluated for its antimicrobial properties. Cysteine, histidine-dependent amidohydrolase/peptidase (CHAP) domains are widely represented in bacteriophage endolysins but have never previously been reported for pneumococcal endolysins. Here, we characterize the first pneumococcal endolysin with a CHAP catalytic domain. SP-CHAP was antimicrobial against all
Spn
serovars tested, including capsular and capsule-free pneumococci, and it was found to be more active than the most widely studied pneumococcal endolysin, Cpl-1, while not affecting various oral or nasal commensal organisms tested. SP-CHAP was also effective in eradicating
Spn
biofilms at concentrations as low as 1.56 µg/mL. In addition, a
Spn
mouse nasopharyngeal colonization model was employed, which showed that SP-CHAP caused a significant reduction in
Spn
colony-forming units, even more than Cpl-1. These results indicate that SP-CHAP may represent a promising alternative to combating
Spn
infections.

IMPORTANCE

Considering the high rates of pneumococcal resistance reported for several antibiotics, alternatives are urgently needed. In the present study, we report a
Streptococcus pneumoniae
-targeting endolysin with even greater activity than Cpl-1, the most characterized pneumococcal endolysin to date. We have employed a combination of biochemical and microbiological assays to assess the stability and lytic potential of SP-CHAP and demonstrate its efficacy on pneumococcal biofilms
in vitro
and in an
in vivo
mouse model of colonization. Our findings highlight the therapeutic potential of SP-CHAP as an antibiotic alternative to treat
Streptococcus pneumoniae
infections.
Is
Candida auris
the first multidrug-resistant fungal zoonosis emerging from climate change?
Authors: Garcia-Bustos Victor
Published: March 2024

The emergence and evolutionary path of
Candida
auris poses an intriguing scientific enigma. Its isolation from a pet dog’s oral cavity in Kansas, reported by White et al. (T. C. White, B. D. Esquivel, E. M. Rouse Salcido, A. M. Schweiker, et al., mBio 15:e03080-23, 2024,
https://doi.org/10.1128/mbio.03080-23
), carries significant implications. This discovery intensifies concerns about its hypothetical capacity for zoonotic transmission, particularly considering the dog’s extensive human contact and the absence of secondary animal/human cases in both animals and humans. The findings challenge established notions of
C. auris
transmissibility and underscore the need for further investigation into the transmission dynamics, especially zooanthroponotic pathways. It raises concerns about its adaptability in different hosts and environments, highlighting potential role of environmental and animal reservoirs in its dissemination. Critical points include the evolving thermal tolerance and the genetic divergence in the isolate. This case exemplifies the necessity for an integrated One Health approach, combining human, animal, and environmental health perspectives, to unravel the complexities of
C. auris
’s emergence and behavior.
The impact of orphan histidine kinases and phosphotransfer proteins on the regulation of clostridial sporulation initiation
Authors: Mehdizadeh Gohari Iman , Edwards Adrianne N. , McBride Shonna M. , McClane Bruce A.
Published: March 2024

Sporulation is an important feature of the clostridial life cycle, facilitating survival of these bacteria in harsh environments, contributing to disease transmission for pathogenic species, and sharing common early steps that are also involved in regulating industrially important solvent production by some non-pathogenic species. Initial genomics studies suggested that Clostridia lack the classical phosphorelay that phosphorylates Spo0A and initiates sporulation in
Bacillus
, leading to the hypothesis that sporulation in Clostridia universally begins when Spo0A is phosphorylated by orphan histidine kinases (OHKs). However, components of the classical
Bacillus
phosphorelay were recently identified in some Clostridia. Similar
Bacillus
phosphorelay components have not yet been found in the pathogenic Clostridia or the solventogenic Clostridia of industrial importance. For some of those Clostridia lacking a classical phosphorelay, the involvement of OHKs in sporulation initiation has received support from genetic studies demonstrating the involvement of several apparent OHKs in their sporulation. In addition, several clostridial OHKs directly phosphorylate Spo0A
in vitro
. Interestingly, there is considerable protein domain diversity among the sporulation-associated OHKs in Clostridia. Further adding to the emergent complexity of sporulation initiation in Clostridia, several candidate OHK phosphotransfer proteins that were OHK candidates were shown to function as phosphatases that reduce sporulation in some Clostridia. The mounting evidence indicates that no single pathway explains sporulation initiation in all Clostridia and supports the need for further study to fully understand the unexpected and biologically fascinating mechanistic diversity of this important process among these medically and industrially important bacteria.
Redefining development in
Streptomyces venezuelae
: integrating exploration into the classical sporulating life cycle
Authors: Shepherdson Evan M. F. , Elliot Marie A.
Published: March 2024

Two growth modes have been described for the filamentous
Streptomyces
bacteria. Their classic developmental life cycle culminates in the formation of dormant spores, where movement to new environments is mediated through spore dispersal. In contrast, exploratory growth proceeds as a rapidly expanding vegetative mycelium that leads to extensive surface colonization and is associated with the release of volatile compounds that promote alkalinization (and reduced iron bioavailability) of its surrounding environment. Here, we report that exploratory growth in
Streptomyces venezuelae
can proceed in tandem with classic sporulating development in response to specific nutritional cues. Sporulating exploration is not accompanied by a rise in environmental pH but has the same iron acquisition requirements as conventional exploration. We found that mutants that were defective in their ability to sporulate were unaffected in exploration, but mutants undergoing precocious sporulation were compromised in their exploratory growth and this appeared to be mediated through premature activation of the developmental regulator WhiI. Cell envelope integrity was also found to be critical for exploration, as mutations in the cell envelope stress-responsive extracytoplasmic function sigma factor SigE led to a failure to explore robustly under all exploration-promoting conditions. Finally, in expanding the known exploration-promoting conditions, we discovered that the model species
Streptomyces lividans
exhibited exploration capabilities, supporting the proposal that exploration is conserved across diverse streptomycetes.

IMPORTANCE

Streptomyces
bacteria have evolved diverse developmental and metabolic strategies to thrive in dynamic environmental niches. Here, we report the amalgamation of previously disparate developmental pathways, showing that colony expansion via exploration can proceed in tandem with colony sporulation. This developmental integration extends beyond phenotype to include shared genetic elements, with sporulation-specific repressors being required for successful exploration. Comparing this new exploration mode with previously identified strategies has revealed key differences (e.g., no need for environmental alkalinization), and simultaneously allowed us to define unifying requirements for
Streptomyces
exploration. The “reproductive exploration” phenomenon reported here represents a unique bet-hedging strategy, with the
Streptomyces
colony engaging in an aggressive colonization strategy while transporting a protected genetic repository.
Comparison of metagenomic and traditional methods for diagnosis of
E. coli
enteric infections
Authors: Royer C. , Patin N. V. , Jesser K. J. , Peña-Gonzalez A. , Hatt J. K. , Trueba G. , Levy K. , Konstantinidis K. T.
Published: March 2024

Diarrheagenic
Escherichia coli
, collectively known as DEC, is a leading cause of diarrhea, particularly in children in low- and middle-income countries. Diagnosing infections caused by different DEC pathotypes traditionally relies on the cultivation and identification of virulence genes, a resource-intensive and error-prone process. Here, we compared culture-based DEC identification with shotgun metagenomic sequencing of whole stool using 35 randomly drawn samples from a cohort of diarrhea-afflicted patients. Metagenomic sequencing detected the cultured isolates in 97% of samples, revealing, overall, reliable detection by this approach. Genome binning yielded high-quality
E. coli
metagenome-assembled genomes (MAGs) for 13 samples, and we observed that the MAG did not carry the diagnostic DEC virulence genes of the corresponding isolate in 60% of these samples. Specifically, two distinct scenarios were observed: diffusely adherent
E. coli
(DAEC) isolates without corresponding DAEC MAGs appeared to be relatively rare members of the microbiome, which was further corroborated by quantitative PCR (qPCR), and thus unlikely to represent the etiological agent in 3 of the 13 samples (~23%). In contrast, ETEC virulence genes were located on plasmids and largely escaped binning in associated MAGs despite being prevalent in the sample (5/13 samples or ~38%), revealing limitations of the metagenomic approach. These results provide important insights for diagnosing DEC infections and demonstrate how metagenomic methods can complement isolation efforts and PCR for pathogen identification and population abundance.

IMPORTANCE

Diagnosing enteric infections based on traditional methods involving isolation and PCR can be erroneous due to isolation and other biases, e.g., the most abundant pathogen may not be recovered on isolation media. By employing shotgun metagenomics together with traditional methods on the same stool samples, we show that mixed infections caused by multiple pathogens are much more frequent than traditional methods indicate in the case of acute diarrhea. Further, in at least 8.5% of the total samples examined, the metagenomic approach reliably identified a different pathogen than the traditional approach. Therefore, our results provide a methodology to complement existing methods for enteric infection diagnostics with cutting-edge, culture-independent metagenomic techniques, and highlight the strengths and limitations of each approach.
Coding-complete genome sequences of rabbit hemorrhagic disease virus type 2 detected in 2023 in Washington State indicate a divergent incursion into the United States
Authors: Pauszek Steven J. , Waters Kaitlyn , Mohamed Fawzi , Lakin Steven M. , Moran Karen , Minor Montariel , Creekmore Lynn , Makhdoomi Muzafar , Faburay Bonto
Published: March 2024

Three rabbit hemorrhagic disease virus type 2 (RHDV2) coding-complete genome sequences were obtained from domestic and wild rabbits in Washington State in June and July 2023. These three RHDV2 sequences are <82% identical to previous RHDV2 sequences in North America and likely indicate a discrete incursion.
Complete genome sequence of plasmid-bearing aerotolerant
Campylobacter jejuni
strain S2-20 isolated from retail chicken meat
Authors: Karki Anand B. , Delaborte Elise , Fakhr Mohamed K.
Published: March 2024

Complete genome sequencing of aerotolerant
Campylobacter jejuni
strain S2-20 revealed the presence of a chromosome of 1,695,449 bp and a plasmid of 49,741 bp that contains predicted antimicrobial resistance and type IV secretion system genes. The chromosome harbored several putative oxidative stress genes with potential roles in aerotolerance.
Whole-genome sequence of
Kosakonia cowanii
strain W006, isolated from seeds of
Triticum aestivum
L.
Authors: Chebotar Vladimir K. , Gancheva Maria S. , Chizhevskaya Elena P. , Keleinikova Oksana V. , Baganova Maria E. , Zaplatkin Alexander N. , Husainov Kharon A.
Published: March 2024

In this study, we sequence, assemble, and annotate
Kosakonia cowanii
strain W006, isolated from seeds of
Triticum aestivum
L. W006 has a single circular chromosome of 4,788,099 bp and 4,466 genes, with a mean G +C content of 56.1%.
Complete genome sequence of
Escherichia coli
MP1
Authors: Pick Kat , Stothard Paul , Raivio Tracy L.
Published: March 2024

Here, we report the complete genome sequence of
Escherichia coli
strain MP1, consisting of one circular chromosome and one circular plasmid. Long-read assembly was performed using a consensus approach, followed by long- and short-read polishing, and gene annotation.
Complete genome sequence of
Paraburkholderia
sp. strain 22B1P capable of utilizing 3-chlorobenzoate as a carbon source
Authors: Moriuchi Ryota , Sano Rion , Fujii Shuma , Suzuki Yuito , Makita Miyune , Kawashima Yo , Shirakawa Takumi , Shindo Renki , Shinkai Tatsumi , Miura Kaede , Hirose Moka , Nakajima Momiji , Kurokawa Asahi , Chetia Rituparna , Hirokawa Chiharu , Suzuki Tomoko , Ito Yukiko , Murano Hiroki , Dohra Hideo , Ogawa Naoto , Kanesaki Yu
Published: March 2024

Paraburkholderia
sp. strain 22B1P utilizes 3-chlorobenzoate as a carbon source. Complete genome sequencing of strain 22B1P revealed two chromosomes and two plasmids. The genes involved in the conversion of 3-chlorobenzoate to 3-chlorocatechol and those involved in the conversion of 3-chlorocatechol to 3-oxoadipate were located on chromosomes 2 and 1, respectively.
Complete genome sequence of
Acidovorax temperans
strain LMJ isolated from a contaminated
Chlamydomonas reinhardtii
Tris-Acetate-Phosphate medium culture plate
Authors: Mitra Mautusi , Stanescu Ana
Published: March 2024

A heavy metal and xenobiotic-tolerant strain of
Acidovorax temperans
strain LMJ was isolated from a contaminated Tris-Acetate-Phosphate medium plate of a green micro-alga,
Chlamydomonas reinhardtii
. Here, we report the complete genome sequence of this strain to provide insights into its survival strategies and nearest taxonomic neighbor.
Diaphorobacter nitroreducens
synergize with oxaliplatin to reduce tumor burden in mice with lung adenocarcinoma
Authors: Ni Yalan , Li Rui , Shen Xiaoyu , Yi Deli , Ren Yilin , Wang Fudong , Geng Yan , You Qingjun
Published: March 2024

Lung adenocarcinoma (LADC) is the most common lung cancer and the leading cause of cancer-related deaths globally. Accumulating evidence suggests that the gut microbiota regulates the host response to chemotherapeutic drugs and can be targeted to reduce the toxicity of current chemotherapeutic agents. However, the effect of
Diaphorobacter nitroreducens
synergized with oxaliplatin on the gut microbiota and their impact on LADC have never been explored. This study aimed to evaluate the anti-cancer effects of
D. nitroreducens
, oxaliplatin, and their combined treatment on tumor growth in tumor-bearing mice. The composition of gut microbiota and the immune infiltration of tumors were evaluated by using 16S rRNA gene high-throughput sequencing and immunofluorescence, respectively. The inhibitory effect of the combination treatment with
D. nitroreducens
and oxaliplatin was significantly stronger than that of oxaliplatin alone in tumor-bearing mice. Furthermore, we observed that the combination treatment significantly increased the relative abundance of
Lactobacillus
and
Akkermansia
in the gut microbiota. Meanwhile, the combination treatment significantly increased the proportions of macrophage but decreased the proportion of regulatory T cells in the LADC tumor tissues of mice. These findings underscored the relationship between
D. nitroreducens
and the gut microbiota-immune cell-LADC axis, highlighting potential therapeutic avenues for LADC treatment.

IMPORTANCE

Oxaliplatin is widely used as an effective chemotherapeutic agent in cancer treatment, but its side effects and response rate still need to be improved. Conventional probiotics potentially benefit cancer chemotherapy by regulating gut microbiota and tumor immune infiltration. This study was novel in reporting a more significant inhibitory effect of
Diaphorobacter nitroreducens
on lung adenocarcinoma (LADC) cells compared with common traditional probiotics and validating its potential as an adjuvant therapy for LADC chemotherapy in mice. This study investigated the impact of
D. nitroreducens
combined with oxaliplatin on the gut microbiota and immune infiltration of tumors as a potential mechanism to improve anticancer effects.
Revisiting the dynamics of interacting vector-like dark energy
Authors: Rodriguez-Benites Carlos , Gonzalez-Espinoza Manuel , Otalora Giovanni , Alva-Morales Manuel
Published: March 2024

We revise the dynamics of interacting vector-like dark energy, a theoretical framework proposed to explain the accelerated expansion of the universe. By investigating the interaction between vector-like dark energy and dark matter, we analyze its effects on the cosmic expansion history and the thermodynamics of the accelerating universe. Our results demonstrate that the presence of interaction significantly influences the evolution of vector-like dark energy, leading to distinct features in its equation of state and energy density. We compare our findings with observational data and highlight the importance of considering interactions in future cosmological studies.