Temporal evolution of bacterial species and their antimicrobial resistance characteristics in wound infections of war-related injuries in Ukraine from 2014 to 2023.

BACKGROUND: This study continues surveillance of antimicrobial resistance associated with combat injuries in Ukraine. AIM: To compare species composition, antibiotic resistance profiles, and emergence of new resistance genes between 2014-2020 and 2022-2023. METHODS: Retrospective multicentre microbiological survey in Ukrainian hospitals. Antibiotic susceptibility, whole genome sequencing, and MLST typing were conducted on 154 organisms obtained from 125 casualties from 2022-2023. FINDINGS: The data reveals a predominance of gram-negative bacteria, particularly Acinetobacter baumannii (35.7%), Pseudomonas aeruginosa (14.9%), and Klebsiella pneumoniae (20.7%). High levels of carbapenem resistance was observed among A. baumannii (meropenem 72,2% [(39/54) 95% CI 58.4-83.5], imipenem 66.7% [(36/54) 95% CI 52.5-78.9]), Klebsiella (meropenem 90.6% [(29/32) 95% CI 75.0-98.0], imipenem 81.2% [(26/32) 95% CI 63.6-92.8]), and P. aeruginosa (meropenem 47.8% [(11/23) 95% CI 26.8-69.4], imipenem 60.8% [(14/23) 95% CI 38.5-80.3]) strains. A. baumannii ST-78 and ST-400 were prevalent from 2014 to 2020, while 5 strains of ST-1077 were newly identified. Pseudomonas aeruginosa strains showed diversity across 16 sequence types (STs), with ST-773 increasing in frequency and new STs emerging, but lacking carbapenemase genes. K. pneumoniae exhibited increased genetic diversity over time, with three STs from 2014 to 2020 and six new STs, including blaNDM-1, blaOXA-48, and blaKPC2 carriers, in recent years. CONCLUSION: There is a growing prevalence of multidrug resistant isolates from globally distributed sequence types.

Single-cell guided prenatal derivation of primary fetal epithelial organoids from human amniotic and tracheal fluids.

Isolation of tissue-specific fetal stem cells and derivation of primary organoids is limited to samples obtained from termination of pregnancies, hampering prenatal investigation of fetal development and congenital diseases. Therefore, new patient-specific in vitro models are needed. To this aim, isolation and expansion of fetal stem cells during pregnancy, without the need for tissue samples or reprogramming, would be advantageous. Amniotic fluid (AF) is a source of cells from multiple developing organs. Using single-cell analysis, we characterized the cellular identities present in human AF. We identified and isolated viable epithelial stem/progenitor cells of fetal gastrointestinal, renal and pulmonary origin. Upon culture, these cells formed clonal epithelial organoids, manifesting small intestine, kidney tubule and lung identity. AF organoids exhibit transcriptomic, protein expression and functional features of their tissue of origin. With relevance for prenatal disease modeling, we derived lung organoids from AF and tracheal fluid cells of congenital diaphragmatic hernia fetuses, recapitulating some features of the disease. AF organoids are derived in a timeline compatible with prenatal intervention, potentially allowing investigation of therapeutic tools and regenerative medicine strategies personalized to the fetus at clinically relevant developmental stages.

Treatment of a contained rupture of a splenic artery pseudoaneurysm with direct thrombin injection.

Splenic artery pseudoaneurysm is a rare and potentially fatal condition. In the present report, we describe the case of a 50-year-old woman with chronic pancreatitis who presented with worsening abdominal pain. Computed tomography demonstrated a 3.5-cm splenic artery pseudoaneurysm of the mid-splenic artery. The patient underwent attempted endovascular repair of the pseudoaneurysm that was unsuccessful. Open conversion revealed an inaccessible splenic artery due to chronic pancreatitis that resulted in dense retroperitoneal fibrosis, and repair was achieved via direct thrombin injection under ultrasound guidance of the pseudoaneurysm and splenectomy. The patient recovered well, and computed tomography at 3 days postoperatively revealed complete thrombosis of the pseudoaneurysm.

A Method-Comparison Study Highlighting the Disparity between Osseous- and Skin-Based Measures of Foot Mobility.

PURPOSE: This study examined the validity of standard clinical measures of arch height mobility, midfoot width mobility (MWM), and foot mobility magnitude (FMM) relative to skin-based and osseous measures derived from radiographs. METHODS: Skin-based clinical indices of foot mobility were calculated from standard, caliper-based measures of foot length, midfoot width, and dorsal arch height of the left limb of 20 healthy participants (8-71 yr) during non-weight-bearing and weight-bearing. Skin-based radiographic and osseous indices were derived from concurrent anteroposterior and lateral radiographs. Agreement between skin-based clinical and skin-based radiographic measures of foot mobility with those of osseous measures was investigated using the Bland and Altman approach. RESULTS: Foot mobility indices derived from clinical measures were significantly higher (20%-50%) than skin-based radiographic measures ( P < 0.01), which were, in turn, significantly higher (200%-250%) than osseous measures ( P < 0.01). Clinical measures demonstrated significant levels of proportional bias compared with radiographic measures of foot mobility ( P < 0.01). The contribution of osseous movement to skin-based clinical measures of mobility was highly variable between individuals, ranging between 19% and 81% for arch height mobility, between 4% and 87% for MWM, and between 14% and 75% for FMM. The limits of tolerance for clinical measures of foot mobility ranged from ±3.2 mm for MWM to ±6.6 mm for measures of FMM. The limits of tolerance for skin-based clinical and skin-based radiographic measures were generally larger than osseous movement with weight-bearing. CONCLUSIONS: Skin-based measures of foot mobility, whether clinical or radiographic methods, are not interchangeable and are poor indicators of osseous mobility. Although further research regarding the utility of osseous measures is warranted, these findings strongly caution against the use of skin-based clinical measures of foot mobility in clinical and research settings.

Generation of human gastric assembloids from primary fetal organoids.

PURPOSE: Understanding human gastric epithelium homeostasis remains partial, motivating the exploration of innovative in vitro models. Recent literature showcases the potential of fetal stem cell-derived organoids in developmental and disease modelling and translational therapies. To scale the complexity of the model, we propose to generate assembloids, aiming to increase gastric maturation to provide new structural and functional insights. METHODS: Human fetal gastric organoids (fGOs) were expanded in 3D Matrigel cultures. Confluent organoid cultures were released from the Matrigel dome and resuspended in a collagen I hydrogel. Subsequently, the organoid mixture was seeded in a ring shape within a 24-well plate and allowed to gelate. The structure was lifted in the medium and cultured in floating conditions, allowing for organoid self-assembling into a gastric assembloid. After 10 days of maturation, the assembloids were characterized by immunostaining and RT-PCR, comparing different fetal developmental stages. RESULTS: Successful generation of human fetal gastric assembloids (fGAs) was achieved using spontaneous self-aggregation within the collagen I hydrogel. Immunostaining analysis of early and late fGAs showed the establishment of apico-basal cell polarity, secretion of gastric mucins, and the presence of chromogranin A in both samples. Transcriptional markers analysis revealed distinct disparities in markers associated with mature cell types between late and early fetal stages. CONCLUSIONS: fGOs can reliably be generated from human fetal samples. This pioneering assembloid approach paves the way for advancing our comprehension of human gastric epithelium homeostasis and its perturbation, offering a better in vitro platform for the study of gastric epithelial development and therapeutic translation.

Hypoxia regulate developmental coronary angiogenesis potentially through VEGFR2- and SOX17-mediated signaling.

Background: Coronary vessels in embryonic mouse heart arises from multiple progenitor population including sinus venosus (SV), endocardium, and proepicardium. ELA/APJ signaling is shown to regulate coronary growth from SV pathway within the subepicardium, whereas VEGF-A/VEGF-R2 pathways is implicated to regulate coronary growth from endocardium pathway. Our previous study show hypoxia as a potential signaling cue to stimulate overall coronary growth and expansion within the myocardium. However, the role of hypoxia and its downstream signaling pathways in the regulation of coronary vessel development is not known. In this study, we investigated the role of hypoxia in coronary vessel development and have identified SOX17- and VEGF-R2-mediated signaling as a potential downstream pathway of hypoxia in the regulation of coronary vessel development. Results: We show that hypoxia gain-of-function in the myocardium through upregulation of HIF-1α disrupts the normal pattern of coronary angiogenesis in developing mouse hearts and displays phenotype that is reminiscent of accelerated coronary growth. We show that VEGF-R2 expression is increased in coronary endothelial cells under hypoxia gain-of-function in vivo and in vitro . Furthermore, we show that SOX17 expression is upregulated in developing mouse heart under hypoxia gain-of-function conditions, whereas SOX17 expression is repressed under hypoxia loss-of-function conditions. Furthermore, our results show that SOX17 loss-of-function disrupts normal pattern of coronary growth. Conclusion: Collectively, our data provide strong phenotypic evidence to show that hypoxia might regulate coronary growth in the developing mouse heart potentially through VEGF-R2- and SOX17-mediated downstream signaling pathways.

Six Extensively Drug-Resistant Bacteria in an Injured Soldier, Ukraine.

Blood and surveillance cultures from an injured service member from Ukraine grew Acinetobacter baumannii, Klebsiella pneumoniae, Enterococcus faecium, and 3 distinct Pseudomonas aeruginosa strains. Isolates were nonsusceptible to most antibiotics and carried an array of antibiotic resistant genes, including carbapenemases (blaIMP-1, blaNDM-1, blaOXA-23, blaOXA-48, blaOXA-72) and 16S methyltransferases (armA and rmtB4).

APJ+ cells in the SHF contribute to the cells of aorta and pulmonary trunk through APJ signaling.

Outflow tract (OFT) develops from cardiac progenitor cells in the second heart field (SHF) domain. APJ, a G-Protein Coupled Receptor, is expressed by cardiac progenitors in the SHF. By lineage tracing APJ+SHF cells, we show that these cardiac progenitors contribute to the cells of OFT, which eventually give rise to aorta and pulmonary trunk/artery upon its morphogenesis. Furthermore, we show that early APJ â€‹+ â€‹cells give rise to both aorta and pulmonary cells but late APJ â€‹+ â€‹cells predominantly give rise to pulmonary cells. APJ is expressed by the outflow tract progenitors in the SHF but its role is unclear. We performed knockout studies to determine the role of APJ in SHF cell proliferation and survival. Our data suggested that APJ knockout in the SHF reduced the proliferation of SHF progenitors, while there was no significant impact on survival. In addition, we show that ectopic overexpression of WNT in these cells disrupted aorta and pulmonary morphogenesis from OFT. Overall, our study has identified APJ â€‹+ â€‹progenitor population within the SHF that give rise to aorta and pulmonary trunk/artery cells. Furthermore, we show that APJ signaling stimulates proliferation of these cells in the SHF.

Building gut from scratch - progress and update of intestinal tissue engineering.

Short bowel syndrome (SBS), a condition defined by insufficient absorptive intestinal epithelium, is a rare disease, with an estimated prevalence up to 0.4 in 10,000 people. However, it has substantial morbidity and mortality for affected patients. The mainstay of treatment in SBS is supportive, in the form of intravenous parenteral nutrition, with the aim of achieving intestinal autonomy. The lack of a definitive curative therapy has led to attempts to harness innate developmental and regenerative mechanisms to engineer neo-intestine as an alternative approach to addressing this unmet clinical need. Exciting advances have been made in the field of intestinal tissue engineering (ITE) over the past decade, making a review in this field timely. In this Review, we discuss the latest advances in the components required to engineer intestinal grafts and summarize the progress of ITE. We also explore some key factors to consider and challenges to overcome when transitioning tissue-engineered intestine towards clinical translation, and provide the future outlook of ITE in therapeutic applications and beyond.

Regenerative medicine for childhood gastrointestinal diseases.

Several paediatric gastrointestinal diseases result in life-shortening organ failure. For many of these conditions, current therapeutic options are suboptimal and may not offer a cure. Regenerative medicine is an inter-disciplinary field involving biologists, engineers, and clinicians that aims to produce cell and tissue-based therapies to overcome organ failure. Exciting advances in stem cell biology, materials science, and bioengineering bring engineered gastrointestinal cell and tissue therapies to the verge of clinical trial. In this review, we summarise the requirements for bioengineered therapies, the possible sources of the various cellular and non-cellular components, and the progress towards clinical translation of oesophageal and intestinal tissue engineering to date.

Patient-level costing analysis of paediatric short bowel syndrome care in a specialist tertiary centre.

PURPOSE: To undertake a pilot study estimating patient-level costs of care for paediatric short bowel syndrome (SBS) from the healthcare provider perspective. METHODS: A pilot group of patients with anatomical SBS was selected at a single specialist tertiary centre in the United Kingdom. The Patient Level Information and Costing System (PLICS) was used to extract costing data for all hospital-based activities related to SBS, from the implementation of PLICS in 2016 to April 2021. Patient-specific and pooled data were reported descriptively in per patient-year terms. RESULTS: Five patients had full PLICS data available for the 5-year study period and 2 patients had 4 years of data. The median cost for hospital care of SBS was £52,834 per patient-year (range £1804-£331,489). The key cost drivers were inpatient beds, pharmacy, and staffing costs, which made up > 60% of annual costs. In the first 3 years following index admission (n = 2), there was a steady decline in the annual cost of care to a level comparable with patients with established SBS. CONCLUSION: Patient-level cost of care analysis for SBS is feasible using PLICS. Hospital-related costs vary widely between and within individual patients over time. Key drivers of cost are related to complications of SBS.

Oil spill causes mass mortality of sea snakes in the Gulf of Oman.

Oil spills in the marine environment inflict significant impacts on a wide diversity of marine fauna. Despite the abundance of literature describing these impacts on numerous species, no studies describe the impacts on sea snakes. In this study we report, for the first time, details of an oil spill which caused mass mortality of sea snakes. In this study, 39 sea snake mortalities from the Gulf of Oman, in particular, the coast of Kalba, Sharjah, UAE, were examined. The investigated sea snakes belong to four different species (Hydrophis platurus, H. lapemoides, H. spiralis and H. ornatus). The majority (84.6%) of sea snakes were observed to have oil covering 75-100% of their bodies. The majority (91.4%) of sea snakes were also observed with oil covering their snouts and eyes. A large proportion (25.8, 41.4 and 34.5%) of sea snakes were observed with oil in their mouth, esophagus and stomach.

Local and systemic responses to SARS-CoV-2 infection in children and adults.

It is not fully understood why COVID-19 is typically milder in children1-3. Here, to examine the differences between children and adults in their response to SARS-CoV-2 infection, we analysed paediatric and adult patients with COVID-19 as well as healthy control individuals (total n = 93) using single-cell multi-omic profiling of matched nasal, tracheal, bronchial and blood samples. In the airways of healthy paediatric individuals, we observed cells that were already in an interferon-activated state, which after SARS-CoV-2 infection was further induced especially in airway immune cells. We postulate that higher paediatric innate interferon responses restrict viral replication and disease progression. The systemic response in children was characterized by increases in naive lymphocytes and a depletion of natural killer cells, whereas, in adults, cytotoxic T cells and interferon-stimulated subpopulations were significantly increased. We provide evidence that dendritic cells initiate interferon signalling in early infection, and identify epithelial cell states associated with COVID-19 and age. Our matching nasal and blood data show a strong interferon response in the airways with the induction of systemic interferon-stimulated populations, which were substantially reduced in paediatric patients. Together, we provide several mechanisms that explain the milder clinical syndrome observed in children.

SARS-CoV-2 infection and replication in human gastric organoids.

COVID-19 typically manifests as a respiratory illness, but several clinical reports have described gastrointestinal symptoms. This is particularly true in children in whom gastrointestinal symptoms are frequent and viral shedding outlasts viral clearance from the respiratory system. These observations raise the question of whether the virus can replicate within the stomach. Here we generate gastric organoids from fetal, pediatric, and adult biopsies as in vitro models of SARS-CoV-2 infection. To facilitate infection, we induce reverse polarity in the gastric organoids. We find that the pediatric and late fetal gastric organoids are susceptible to infection with SARS-CoV-2, while viral replication is significantly lower in undifferentiated organoids of early fetal and adult origin. We demonstrate that adult gastric organoids are more susceptible to infection following differentiation. We perform transcriptomic analysis to reveal a moderate innate antiviral response and a lack of differentially expressed genes belonging to the interferon family. Collectively, we show that the virus can efficiently infect the gastric epithelium, suggesting that the stomach might have an active role in fecal-oral SARS-CoV-2 transmission.

Anatomy of an extensively drug-resistant Klebsiella pneumoniae outbreak in Tuscany, Italy.

A protracted outbreak of New Delhi metallo-ß-lactamase (NDM)-producing carbapenem-resistant Klebsiella pneumoniae started in Tuscany, Italy, in November 2018 and continued in 2020 and through 2021. To understand the regional emergence and transmission dynamics over time, we collected and sequenced the genomes of 117 extensively drug-resistant, NDM-producing K. pneumoniae isolates cultured over a 20-mo period from 76 patients at several healthcare facilities in southeast Tuscany. All isolates belonged to high-risk clone ST-147 and were typically nonsusceptible to all first-line antibiotics. Albeit sporadic, resistances to colistin, tigecycline, and fosfomycin were also observed as a result of repeated, independent mutations. Genomic analysis revealed that ST-147 isolates circulating in Tuscany were monophyletic and highly genetically related (including a network of 42 patients from the same hospital and sharing nearly identical isolates), and shared a recent ancestor with clinical isolates from the Middle East. While the blaNDM-1 gene was carried by an IncFIB-type plasmid, our investigations revealed that the ST-147 lineage from Italy also acquired a hybrid IncFIB/IncHIB-type plasmid carrying the 16S methyltransferase armA gene as well as key virulence biomarkers often found in hypervirulent isolates. This plasmid shared extensive homologies with mosaic plasmids circulating globally including from ST-11 and ST-307 convergent lineages. Phenotypically, the carriage of this hybrid plasmid resulted in increased siderophore production but did not confer virulence to the level of an archetypical, hypervirulent K. pneumoniae in a subcutaneous model of infection with immunocompetent CD1 mice. Our findings highlight the importance of performing genomic surveillance to identify emerging threats.

Junk food: Interspecific and intraspecific distinctions in marine debris ingestion by marine turtles.

The pervasiveness of marine debris is now considered one of the most persistent changes in marine environments. This study reports marine debris ingested by green sea turtles Chelonia mydas and loggerhead sea turtles Caretta caretta that stranded along the eastern coast of the Sharjah Emirate in the United Arab Emirates. We observed that both green and loggerhead sea turtles frequently ingest (Frequency of Occurrence: 75.0% and 57.1% respectively) high quantities of marine debris, particularly plastics. The results suggest that green sea turtles are more likely to ingest soft items such as threads and sheets while loggerheads are more likely to ingest hard items. When considering the quantity, frequency and nature of ingested marine debris as well as the physiology of specific species and age classes, green sea turtles, particularly younger specimens, ingest the greatest amount of marine debris.

Paediatric gastric organoids as a tool for disease modelling and clinical translation.

PURPOSE: Knowledge of gastric epithelial homeostasis remains incomplete, lacking human-specific models for study. This study establishes a protocol for deriving gastric epithelial organoids from paediatric gastric biopsies, providing a platform for modelling disease and developing translational therapies. METHODS: Full-thickness surgical samples and endoscopic mucosal biopsies were obtained from six patients. Gastric glands were isolated by a chemical chelation protocol and then plated in 3D culture in Matrigel® droplets in chemically defined medium. After formation, organoids were passaged by single cell dissociation or manual disaggregation. Cell composition and epithelial polarity of organoids were assessed by bright field microscopy and immunofluorescence analysis, comparing them to native paediatric gastric tissue. RESULTS: Gastric glands were successfully isolated from all six patients who were aged 4 months to 16 years. Gastric glands from all patients sealed to form spherical gastric organoids. These organoids could be passaged by manual disaggregation or single cell dissociation, remaining proliferative up to 1 year in culture. Organoids retained normal epithelial cell polarity, with the apical surface orientated towards the central lumen. Organoids expressed markers of mature gastric epithelial cell types, except for parietal cells. CONCLUSION: Gastric organoids can be reliably generated from paediatric biopsies and are a representative in vitro model for studying gastric epithelium.

An In Vitro Whole-Organ Liver Engineering for Testing of Genetic Therapies.

Explosion of gene therapy approaches for treating rare monogenic and common liver disorders created an urgent need for disease models able to replicate human liver cellular environment. Available models lack 3D liver structure or are unable to survive in long-term culture. We aimed to generate and test a 3D culture system that allows long-term maintenance of human liver cell characteristics. The in vitro whole-organ "Bioreactor grown Artificial Liver Model" (BALM) employs a custom-designed bioreactor for long-term 3D culture of human induced pluripotent stem cells-derived hepatocyte-like cells (hiHEPs) in a mouse decellularized liver scaffold. Adeno-associated viral (AAV) and lentiviral (LV) vectors were introduced by intravascular injection. Substantial AAV and LV transgene expression in the BALM-grown hiHEPs was detected. Measurement of secreted proteins in the media allowed non-invasive monitoring of the system. We demonstrated that humanized whole-organ BALM is a valuable tool to generate pre-clinical data for investigational medicinal products.

Technical Approach to Laparoscopic Examination of the Small Bowel in Gallstone Ileus.

BACKGROUND: Gallstone ileus is an infrequent cause of small bowel obstructions (SBO), accounting for only 0.1-5% of SBOs and 25% of nonstrangulating causes of SBO in the elderly population. There is scant literature available regarding the use of laparoscopy to treat gallstone ileus. Currently, much of the literature available is limited to case reports only. METHODS: A complete laparoscopic approach was utilized to manage a 65-year-old woman with morbid obesity who presented with gallstone ileus. With regard to our technical approach, we describe the technical approach that facilitates safe laparoscopic examination of the entire small bowel and can be applied to other acute care surgery cases involving small bowel pathology. RESULTS: The patient's postoperative course was complicated by new-onset atrial fibrillation which was treated medically with good response. She was safely discharged on postoperative day 2. CONCLUSION: Laparoscopy is a feasible option for the management of gallstone ileus and can lead to decreased morbidity compared to laparotomy. The technique described allows for laparoscopic examination of the entire small bowel.