Search Results (77)

Sternal bursitis is an underexplored lesion in poultry, often overlooked in microbiological diagnostics. In this study, we characterized 36 Escherichia coli isolates recovered from sternal bursitis in broiler chickens, combining phenotypic antimicrobial susceptibility testing, PCR-based screening, and whole genome sequencing (WGS). The genetic analysis revealed a diverse population spanning 15 sequence types, including ST155, ST201, and ST58. Resistance to tetracycline and ciprofloxacin was common, and several isolates carried genes encoding β-lactamases, including blaTEM-1B. Chromosomal mutations associated with quinolone and fosfomycin resistance (e.g., gyrA p.S83L, glpT_E448K) were also identified. WGS revealed a high number of virulence-associated genes per isolate (58–96), notably those linked to adhesion (fim, ecp clusters), secretion systems (T6SS), and iron acquisition (ent, fep, fes), suggesting strong pathogenic potential. Many isolates harbored virulence markers typical of ExPEC/APEC, such as iss, ompT, and traT, even in the absence of multidrug resistance. Our findings suggest that E. coli from sternal bursitis may act as reservoirs of resistance and virulence traits relevant to animal and public health. This highlights the need for including such lesions in genomic surveillance programs and reinforces the importance of integrated One Health approaches. Full article

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease affecting motor neurons with a phenotypic and genetic heterogeneity and elusive molecular mechanisms. With the present pilot study, we investigated different genetic mutations (C9orf72, TARDBP, and KIF5A) associated with ALS by generating induced pluripotent stem cells (iPSCs) from peripheral blood of ALS patients and healthy donors. iPSCs showed the typical morphology, expressed stem cell markers both at RNA (OCT4, SOX2, KLF4, and c-Myc) and protein (Oct4, Sox2, SSEA3, and Tra1-60) levels. Moreover, embryoid bodies expressing the three germ-layer markers and neurospheres expressing neural progenitor markers were generated. Importantly, the transcriptomic profiles of iPSCs and neurospheres were analyzed to highlight the differences between ALS patients and healthy controls. Interestingly, the differentially expressed genes (DEGs) shared across all ALS iPSCs are linked to extracellular matrix, highlighting its importance in ALS progression. In contrast, ALS neurospheres displayed widespread deficits in neuronal pathways, although these DEGs were varied among patients, reflecting the disease’s heterogeneity. Overall, we generated iPSC lines from ALS patients with diverse genetic backgrounds offering a tool for unravelling the intricate molecular landscape of ALS, paving the way for identifying key pathways implicated in pathogenesis and the disease’s phenotypic variability. Full article

Background/Objectives: Klebsiella pneumoniae ST307 is emerging as a significant global high-risk antimicrobial-resistant (AMR) clone with a notable capacity to acquire and disseminate resistance genes. However, there is limited research on the pathogenicity, virulence, and adaptation of ST307 strains and on the clinical characteristics of infected patients. Methods: In this study, a carbapenem-resistant K. pneumoniae (CRKP) ST307 strain named U989 was isolated from a urine culture of a hospitalized patient in Volos, Greece, in July 2024. Whole-genome sequencing was performed to identify resistance genes to β-lactams bla_KPC-2, bla_CTX-M-15, bla_TEM-1B, bla_OXA-1, bla_OXA-10, bla_SHV-106, and bla_VEB-1 and resistance genes to other antibiotics. Results: A genomic analysis also revealed the presence of virulence factors such as iutA, clpK1, fyuA, fimH, mrkA, Irp2, and TraT and an IncFiB(pQil)/IncFII(K) replicon, which harbors the bla_KPC-2 gene. Additionally, the transposable element Tn4401 was identified as a key vehicle for the mobilization of the bla_KPC-2 resistance gene. Finally, this is the report of a high-risk CRKP ST307 clone expressing KPC-2, SHV-106, CTX-M-15, and VEB-1 bla genes in Greece. Conclusions: The coexistence of these resistance genes in addition to aminoglycoside, quinolone, and other resistance genes results in difficult-to-treat infections caused by respective carrier strains, often requiring the use of last-resort antibiotics and contributing to the global challenge of antimicrobial resistance. Full article

tet(X4)-positive IncHI1 plasmids are widely prevalent in various bacteria. To understand their transmission characteristics, we analyzed two extensively drug-resistant (XDR) Escherichia coli strains isolated from pet dog feces in Henan Province, China. Strain T28R harbored tet(X4)-positive IncHI1, IncF18:A-:B-, and mcr-1-positive IncI2 plasmids, while T16R carried tet(X4)-positive IncHI1, F16:A-:B-, and mcr-1-positive IncX4 plasmids. Four representative fusion plasmids, pT28R-F1, pT28R-F2, pT28R-F3, and pT16R-F1, in transconjugants were analyzed using WGS and PCR mapping. The results showed that IS26 from the IncF18:A-:B--plasmid attacked the conjugative transfer-associated genes trhc and rsp on the IncHI1 plasmid, generating pT28R-F1 and pT28R-F2. pT28R-F3 was generated through ISCro1- and ISCR2-mediated homologous recombination, deleting the Tra1 region of the IncHI1 plasmid. T16R-F1 emerged from ISCR2- and IS1B-mediated homologous recombination, losing transfer regions of parental plasmids. Notably, fusion plasmids lost the temperature sensitivity of the IncHI1 plasmid, with conjugation frequencies between 1.57 × 10⁻⁴ and 3.84 × 10⁻⁵ at 28 °C and 37 °C. The findings suggest that tet(X4)-positive IncHI1 plasmids could be mobilized with the assistance of conjugative helper plasmids and that fusion events enhance the adaptability of these plasmids, thus facilitating the spread of antibiotic resistance, posing a growing public health threat. Full article

The limited availability of primary normal human epidermal keratinocyte (NHEK) has hampered the large-scale implementation of skin models in biomedical, toxicological, and pharmaceutical research. Therefore, in this study, we aimed to establish an induced pluripotent stem cell (iPSC)-derived epidermal skin model that is not limited by donor type and cell lifespan, and evaluate whether it is equivalent to the primary NHEK-derived reconstructed epidermal skin model (RHE) for skin irritation testing. The results show that high expression of OCT4, SOX2, KLF4, c-MYC, and SSEA-4, TRA-1-60, TRA-1-81 indicated that iPSCs were successfully generated from human fibroblasts in vitro. The expression levels of ectoderm or KC marker genes CGB, IVL, KRT10, KRT14, TP63, and TBP were close to those of NHEKs. This result confirms that iPSCs were successfully differentiated into iPSC-KCs. The expression levels of iPSC-derived-RHE in FLG (60), AQP3 (151), CLDN1 (30.6), IVL (209), KRT5 (39.3), KRT10 (39.2), TSLP (99), IL-6 (53.1), IL-8 (79.4), and TNF-a (91.5) were significantly higher than those in RHE. These results indicate that iPSC-derived RHE has extremely strong vitality and renewal capacity. Meanwhile, there was no significant difference between iPSC-derived RHE and SkinEthic in predicting skin irritation, which means that our iPSC-derived RHE performed well in the test. iPSC-derived RHE can replace other skin models for skin irritation testing related to cosmetics. This technology has the potential to generate an unlimited number of genetically identical skin models and improve the reproducibility of experiments. Full article

The genes that encode the universal stress protein (USP) family domain (pfam00582) aid the survival of bacteria in specific host or habitat-induced stress conditions. Genome sequencing revealed that the genome of Helicobacter pylori, a gastric cancer pathogen, typically contains one USP gene, while related helicobacters have one or two distinct USP genes. However, insights into the functions of Helicobacteraceae (Helicobacter and Wolinella) USP genes are still limited to inferences from large-scale genome sequencing. Thus, we have combined bioinformatics and visual analytics approaches to conduct a more comprehensive data investigation of a set of 1045 universal stress protein sequences encoded in 1014 genomes including 785 Helicobacter pylori genomes. The study generated a representative set of 183 USP sequences consisting of 180 Helicobacter sequences, two Wolinella succinogenes sequences, and a sequence from a related campylobacteria. We used the amino acid residues and positions of the 12 possible functional sites in 1030 sequences to identify 25 functional sites patterns for guiding studies on functional interactions of Helicobacteraceae USPs with ATP and other molecules. Genomic context searches and analysis identified USP genes of gastric and enterohepatic helicobacters that are adjacent or in operons with genes for proteins responsive to DNA-damaging oxidative stress (ATP-dependent proteases: ClpS and ClpA); and DNA uptake proteins (natural competence for transformation proteins: ComB6, ComB7, ComB8, ComB9, ComB10, ComBE, and conjugative transfer signal peptidase TraF). Since transcriptomic evidence indicates that oxidative stress and the presence of virulence-associated genes regulate the transcription of H. pylori USP gene, we recommend further research on Helicobacter USP genes and their neighboring genes in oxidative stress response and virulence of helicobacters. To facilitate the reuse of data and research, we produced interactive analytics resources of a dataset composed of values for variables including phylogeography of H. pylori strains, protein sequence features, and gene neighborhood. Full article

X-adrenoleukodystrophy (X-ALD) is a peroxisomal metabolic disorder caused by mutations in the ABCD1 gene encoding the peroxisomal ABC transporter adrenoleukodystrophy protein (ALDP). Similar mutations in ABCD1 may result in a spectrum of phenotypes in males with slow progressing adrenomyeloneuropathy (AMN) and fatal cerebral adrenoleukodystrophy (cALD) dominating most cases. Mouse models of X-ALD do not capture the phenotype differences and an appropriate model to investigate the mechanism of disease onset and progress remains a critical need. Here, we generated induced pluripotent stem cell (iPSC) lines from skin fibroblasts of two each of apparently healthy control, AMN, and cALD patients with non-integrating mRNA-based reprogramming. iPSC lines expanded normally and expressed pluripotency markers Oct4, SOX2, NANOG, SSEA, and TRA-1–60. Expression of markers SOX17, Brachyury, Desmin, OXT2, and beta tubulin III demonstrated the ability of the iPSCs to differentiate into all three germ layers. iPSC-derived lines from CTL, AMN, and cALD male patients were differentiated into astrocytes. Differentiated AMN and cALD astrocytes lacked ABCD1 expression and accumulated saturated very long chain fatty acids (VLCFAs), a hallmark of X-ALD, and demonstrated differential mitochondrial bioenergetics, cytokine gene expression, and differences in STAT3 and AMPK signaling between AMN and cALD astrocytes. These patient astrocytes provide disease-relevant tools to investigate the mechanism of differential neuroinflammatory response in X-ALD and will be valuable cell models for testing new therapeutics. Full article

Escherichia coli (E. coli) has the ability to induce clinical and subclinical mastitis in dairy cows, causing a huge loss for the dairy industry. In this study, 51 subclinical mastitis isolates and 36 clinical mastitis isolates from eight provinces of China between 2019 and 2021 were used to investigate the differences in their biological characteristics. The results showed that B1 (52.9%) and A (39.1%) were the predominant phylogroups; R1 (50.6%) was the predominant lipopolysaccharide (LPS) core type; and 44 STs (ST10 and ST58 were the most sequence-prevalent STs) and 2 new STs (ST14828 and ST14829) were identified; however, no significant difference was observed between the clinical and subclinical group strains. To compare the virulence gene differences between the clinical and subclinical mastitis-related isolates, 18 common virulence genes (including afaE, eaeA, papC, saa, sfa, ompA, aer, irp2, iucD, escV, sepD, east1, estB, stx2e, CNF1, cba, hlyA and traT) were determined using the PCR method. The results showed that the detection rates of traT, irp2 and iucD in clinical mastitis isolates were significantly higher than those in subclinical mastitis isolates (p ˂ 0.05). Meanwhile, subclinical-group E. coli had stronger biofilm formation abilities than the clinical group (p < 0.05) in 78 (89.7%) mastitis-related E. coli that could form biofilms. Furthermore, 87 mastitis-related E. coli showed severe resistance against tetracycline (37.9%), ampicillin (36.8%), streptomycin (34.5%) and cotrimoxazole (28.7%); their most prevalent resistance genes were blaCTX-M (33.3%), tetA (27.6%), sul2 (18.4%) and strB (28.7%). It was noteworthy that the clinical-group strains had a higher resistance against ampicillin and possessed higher amounts of the resistance gene blaCTX-M (p < 0.05) compared to the subclinical group. This study aims to provide references for preventing the E. coli isolates from inducing different types of mastitis. Full article

Background: Therapeutic strategies for methicillin-resistant Staphylococcus aureus (MRSA) are increasingly limited due to the ability of the pathogen to evade conventional treatments such as vancomycin and daptomycin. This challenge has shifted the focus towards novel strategies, including the resensitization of β-lactams, which are still used as first-line treatments for methicillin-susceptible Staphylococcus aureus (MSSA). To achieve this, it is essential to identify the secondary resistome associated with the clinically relevant β-lactam antibiotics. Methods: Transposon-Directed Insertion Site Sequencing (TraDIS) was employed to assess conditional essentiality by analyzing the depletion of mutants from a highly saturated transposon library of MRSA USA300 JE2 exposed to ½ minimal inhibitory concentration (MIC) of oxacillin or cefazolin. Results: TraDIS analysis led to the identification of 52 shared fitness genes involved in β-lactam resistance that are primarily linked to cell wall metabolism and regulatory systems. Among these, both known resistance factors and novel conditionally essential genes were highlighted. As proof of concept, transposon mutants corresponding to nine genes (sagB, SAUSA300_0657, SAUSA300_0957, SAUSA300_1683, SAUSA300_1964, SAUSA300_1966, SAUSA300_1967, SAUSA300_1692, and mazF) were grown in the presence of β-lactam antibiotics and their MICs were determined. All mutants showed significantly reduced resistance to β-lactam antibiotics. Conclusions: This comprehensive genome-wide investigation provides novel insights into the resistance mechanisms of β-lactam antibiotics, and suggests potential therapeutic targets for combination therapies with helper drugs. Full article

Members of Bacillus species are able to enhance the level of available phosphorus (P) for plant absorption through mechanisms of P solubilization and mineralization. In our study, B. subtilis PE7 showed P-solubilizing activity in simple phosphate broth (SPB) medium, and acetic acid, iso-butyric acid, and iso-valeric acid were major organic acids responsible for the increase in soluble P and decrease in pH of SPB medium. In addition, strain PE7 released phytase on phytase-screening agar (PSA) medium, and analysis of semi-quantitative reverse transcription and polymerase chain reaction (sqRT-PCR) revealed that the phyC gene expression was the highest at 1 day after incubation. A low concentration of KH₂PO₄ in SPB medium induced more biofilm formation than a high concentration of KH₂PO₄. Strain PE7 showed swimming and swarming motilities in TY and TrA agar media. Under P starvation, inoculation with higher cell numbers of strain PE7 enhanced biomass and nutrient acquisition by melon plants, resulting in higher values of growth parameters and nutrient contents. Moreover, the persistence of bacterial cells on the root surface and in the rhizosphere of melon plants indicated colonization of the plants by strain PE7. Due to its capacity for P solubilization and mineralization, B. subtilis PE7 could be utilized as an alternative to synthetic fertilizer for P deficient-stress management in crop plantation. Full article

Background/Objectives: We developed a multistage Plasmodium falciparum vaccine using a heterologous prime-boost immunization strategy. This involved priming with a highly attenuated, replication-competent vaccinia virus strain LC16m8Δ (m8Δ) and boosting with adeno-associated virus type 1 (AAV1). This approach demonstrated 100% efficacy in both protection and transmission-blocking in a murine model. In this study, we compared our LC16m8∆/AAV1 vaccine, which harbors a gene encoding Pfs25-PfCSP fusion protein, to RTS,S/AS01 (RTS,S) in terms of immune responses, protective efficacy, and transmission-blocking activity (TBA) in murine models. Methods: Mice were immunized following prime-boost vaccine regimens m8∆/AAV1 or RTS,S and challenged with transgenic Plasmodium berghei parasites. Immune responses were assessed via ELISA, and TB efficacy was evaluated using direct feeding assays. Results: m8∆/AAV1 provided complete protection (100%) in BALB/c mice and moderate (40%) protection in C57BL/6 mice, similar to RTS,S. Unlike RTS,S’s narrow focus (repeat region), m8∆/AAV1 triggered antibodies for all PfCSP regions (N-terminus, repeat, and C-terminus) with balanced Th1/Th2 ratios. Regarding transmission blockade, serum from m8∆/AAV1-vaccinated BALB/c mice achieved substantial transmission-reducing activity (TRA = 83.02%) and TB activity (TBA = 38.98%)—attributes not observed with RTS,S. Furthermore, m8∆/AAV1 demonstrated durable TB efficacy (94.31% TRA and 63.79% TBA) 100 days post-immunization. Conclusions: These results highlight m8∆/AAV1′s dual action in preventing sporozoite invasion and onward transmission, a significant advantage over RTS,S. Consequently, m8∆/AAV1 represents an alternative and a promising vaccine candidate that can enhance malaria control and elimination strategies. Full article

Hereditary spastic paraplegias are rare genetic disorders characterized by corticospinal tract impairment. Spastic paraplegia 83 (SPG83) is associated with biallelic mutations in the HPDL gene, leading to varied severities from neonatal to juvenile onset. The function of HPDL is unclear, though it is speculated to play a role in alternative coenzyme Q10 biosynthesis. Here, we report the generation of hiPS lines from primary skin fibroblasts derived from three SPG83 patients with different HPDL mutations, using episomal reprogramming. The patients’ clinical characteristics are carefully listed. The hiPS lines were meticulously characterized, demonstrating typical pluripotent characteristics through immunofluorescence assays for stemness markers (OCT4, TRA1-60, NANOG, and SSEA4) and RT-PCR for endogenous gene expression. Genetic integrity and identity were confirmed via Sanger sequencing and short tandem repeat analysis. These hiPS cells displayed typical pluripotent characteristics and were able to differentiate into neocortical neurons via a dual SMAD inhibition protocol. In addition, HPDL mutant neurons assessed via long-term culturing were able to achieve effective maturation, similarly to their wild-type counterparts. The HPDL hiPS lines we generated will provide a valuable model for studying SPG83, offering insights into its molecular mechanisms and potential for developing targeted therapies. Full article

Increasing evidence has suggested that nanoplastic pollution has become a global concern. More importantly, transgenerational toxicity can be induced by nanoplastics at predicted environmentally relevant doses (ERDs). Considering that amino modification could increase nanoplastic toxicity, we compared transgenerational neurotoxicity between pristine polystyrene nanoparticle (PS-NP) and amino-modified PS-NP (NH₂-PS-NP) in Caenorhabditis elegans. At 0.1–10 μg/L, NH₂-PS-NP caused more severe transgenerational toxicity on locomotion and neuronal development. Accompanied with a difference in transgenerational neuronal damage, compared to PS-NP (10 μg/L), NH₂-PS-NP (10 μg/L) induced more severe transgenerational activation of mec-4, crt-1, itr-1, and tra-3, which are required for the induction of neurodegeneration. Moreover, NH₂-PS-NP (10 μg/L) caused more severe transgenerational inhibition in expressions of mpk-1, jnk-1, dbl-1, and daf-7 than PS-NP (10 μg/L), and RNA interference (RNAi) of these genes conferred susceptibility to the toxicity of PS-NP and NH₂-PS-NP on locomotion and neuronal development. NH₂-PS-NP (10 μg/L) further caused more severe transgenerational activation of germline ligand genes (ins-3, ins-39, daf-28, lin-44, egl-17, efn-3, and lag-2) than PS-NP (10 μg/L), and RNAi of these ligand genes caused resistance to the toxicity of PS-NP and NH₂-PS-NP on locomotion and neuronal development. Our results highlighted more severe exposure risk of amino-modified nanoplastics at ERDs in causing transgenerational neurotoxicity in organisms. Full article

Background: Several Acinetobacter species live in different ecosystems, such as soil, freshwater, wastewater, and solid wastes, which has attracted considerable research interests in public health and agriculture. Methods: We assessed the distribution of Acinetobacter baumannii and Acinetobacter nosocomialis in three freshwater resources (Great Fish, Keiskemma, and Tyhume rivers) in South Africa between April 2017–March 2018. Molecular identification of Acinetobacter species was performed using Acinetobacter-specific primers targeting the recA gene, whilst confirmed species were further delineated into A. baumannii and A. nosocomialis. Similarly, virulence genes; afa/draBC, epsA, fimH, OmpA, PAI, sfa/focDE, and traT in the two Acinetobacter species were assessed. Results: Our finding revealed that 410 (48.58%) and 23 (2.7%) of the isolates were confirmed as A. baumannii and A. nosocomalis, respectively. Additionally, three hundred and eight (75.12%) A. baumannii and three (13.04%) A. nosocomialis exhibited one or more of the virulence genes among the seven tested. OmpA was the most prevalent virulence gene in A. baumannii in freshwater sources. Conclusions: The distribution of clinically important Acinetobacter species in the freshwater sources studied suggests possible contamination such as the release of hospital wastewater and other clinical wastes into the environment thereby posing a risk to public health. Full article

The ubiquitous presence of antimicrobial-resistant organisms and antimicrobial resistance genes (ARGs) constitutes a major threat to global public safety. Tetracycline (TET) is a common antimicrobial agent that inhibits bacterial growth and is frequently detected in aquatic environments. Although TET may display coselection for resistance, limited knowledge is available on whether and how it might influence plasmid-mediated conjugation. Subinhibitory concentrations (3.9–250 ng/mL) of TET promoted horizontal gene transfer (HGT) via the mobilizable plasmid pVP52-1 from the donor Vibrio parahaemolyticus NJIFDCVp52 to the recipient Escherichia coli EC600 by 1.47- to 3.19-fold. The transcription levels of tetracycline resistance genes [tetA, tetR(A)], conjugation-related genes (traA, traD), outer membrane protein genes (ompA, ompK, ompV), reactive oxygen species (ROS)-related genes (oxyR, rpoS), autoinducer-2 (AI-2) synthesis gene (luxS), and SOS-related genes (lexA, recA) in the donor and recipient were significantly increased. Furthermore, the overproduced intracellular ROS generation and increased cell membrane permeability under TET exposure stimulated the conjugative transfer of ARGs. Overall, this study provides important insights into the contributions of TET to the spread of antimicrobial resistance. Full article