Nano-modified viruses prime the tumor microenvironment and promote the photodynamic virotherapy in liver cancer.

BACKGROUND: As of 2020, hepatocellular carcinoma (HCC), a form of liver cancer, stood as the third most prominent contributor to global cancer-related mortality. Combining immune checkpoint inhibitors (ICI) with other therapies has shown promising results for treating unresectable HCC, offering new opportunities. Recombinant adeno-associated viral type 2 (AAV2) virotherapy has been approved for clinical use but it efficacy is stifled through systemic administration. On the other hand, iron oxide nanoparticles (ION) can be cleared via the liver and enhance macrophage polarization, promoting infiltration of CD8+ T cells and creating a more favorable tumor microenvironment for immunotherapy. METHODS: To enhance the efficacy of virotherapy and promote macrophage polarization towards the M1-type in the liver, ION-AAV2 were prepared through the coupling of ION-carboxyl and AAV2-amine using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC)/N-hydroxysulfosuccinimide (Sulfo-NHS). Efficacy after systemic delivery of ION-AAV2 in an orthotopic HCC model was evaluated. RESULTS: After 28 days, the tumor weight in mice treated with ION-AAV2 was significantly reduced by 0.56-fold compared to the control group. The ION-AAV2 treatment led to an approximate 1.80-fold increase in the level of tumor associated M1-type macrophages, while the number of M2-type macrophages was reduced by 0.88-fold. Moreover, a proinflammatory response increased the population of tumor-infiltrating CD8+ T cells in the ION-AAV2 group. This transformation converted cold tumors into hot tumors. CONCLUSIONS: Our findings suggest that the conjugation of ION with AAV2 could be utilized in virotherapy while simultaneously exploiting macrophage-modulating cancer immunotherapies to effectively suppress HCC growth.

Repolarization of M2 to M1 Macrophages Triggered by Lactate Oxidase Released from Methylcellulose Hydrogel.

Deregulated proliferation of tumors is generally associated with altered energy metabolism. A high rate of anaerobic glycolysis in solid tumors contributes to an acidification of pH to ∼6.7-7.2 in the tumor microenvironment and lactate accumulation. Macrophages in the tumor microenvironment can be educated by tumor cells. Tumor-derived lactate induces the polarization of M2 macrophages and promotes tumor invasion and metastasis. However, a particular challenge is to sustain lactate depletion. We propose that the repolarization of the tumor-supportive M2 macrophage to the tumor-suppressive M1 macrophage after the depletion of lactate by lactate oxidase (LOX) released from the hydrogels in the tumor microenvironment may enhance the antitumor treatment efficacy.

Effect of fetal hemodynamics on growth in fetuses with single ventricle or transposition of the great arteries.

OBJECTIVES: As birth weight is a critical predictor of outcome in neonates with congenital heart defect (CHD), the common problem of poor fetal growth in this population is clinically important. However, it is not well understood and the impact of fetal hemodynamics on fetal growth and birth weight in those with CHD has not been assessed. In this study, we sought to evaluate the association between combined cardiac output (CCO) and fetal middle cerebral artery (MCA) and umbilical artery (UA) pulsatility indices (PIs) and fetal growth in different subgroups of CHD, and to study the effects of fetal hemodynamics on late gestational weight gain. We hypothesized that fetuses with CHD will have lower CCO and be smaller at birth. METHODS: This was a retrospective review of fetal echocardiograms from 67 fetuses diagnosed with hypoplastic left heart syndrome (HLHS, n = 30), non-HLHS single ventricle (SV) (n = 20) or dextrotransposition of the great arteries (d-TGA, n = 17), compared with normal controls (n = 42). CCO was calculated using valvar area, velocity-time integral and heart rate and indexed to estimated fetal weight. MCA- and UA-PI were calculated using systolic, diastolic and mean velocities. Fetal biometry was recorded. Regression models were used to study trends in CCO, MCA- and UA-PI and fetal biometry over gestational age. To evaluate fetal weight gain in late gestation, Z-scores of estimated fetal weight at 30 weeks and birth weight were compared. Regression analysis was used to determine the associations of CCO, indexed CCO and MCA- and UA-PI at 30 weeks with birth weight, length and head circumference Z-scores, in addition to weight gain late in gestation. The gestational age of 30 weeks was chosen based on previous studies that found evidence of poor weight gain in fetuses with CHD in late gestation, starting at around that time. RESULTS: CCO increased with gestation in all four groups but the rate was slower in fetuses with HLHS and in those with SV. MCA-PI was lower in fetuses with HLHS compared with in those with non-HLHS-SV throughout gestation, suggesting different cerebral blood distribution. At the end of gestation, rate of fetal weight gain slowed in those with HLHS and in those with SV (similar to CCO curves), and head circumference growth rate slowed in all groups but controls. CCO, indexed CCO and MCA- and UA-PI did not correlate with any of the birth measurements or with weight gain late in gestation in fetuses with CHD. CONCLUSIONS: We found no associations of CCO or MCA- and UA-PI with late gestational weight gain or biometry at birth in fetuses with CHD. This does not support fetal hemodynamics as the primary driver of suboptimal fetal growth in fetuses with SV. Future research could further explain genetic and placental abnormalities that may affect fetal growth in those with CHD. Copyright © 2017 ISUOG. Published by John Wiley & Sons Ltd.

Skin collagen can be accurately quantified through noninvasive optical method: Validation on a swine study.

BACKGROUND/PURPOSE: Diffuse reflectance spectroscopy (DRS) is a noninvasive optical technology characterized by relatively low system cost and high efficiency. In our previous study, we quantified the relative concentration of collagen for the individual keloid patient. However, no actual value of collagen concentration can prove the reliability of collagen detection by our DRS system. METHODS: Skin-mimicking phantoms were prepared using different collagen and coffee concentrations, and their chromophore concentrations were quantified using the DRS system to analyze the influence of collagen and other chromophores. Moreover, we used the animal study to compare the DRS system with the collagen evaluation of biopsy section by second-harmonic generation (SHG) microscopy at four different skin parts. RESULTS: In the phantom study, the result showed that coffee chromophore did not severely interfere with collagen concentration recovery. In the animal study, a positive correlation (r=.902) between the DRS system and collagen evaluation with SHG microscopy was found. CONCLUSIONS: We have demonstrated that the DRS system can quantify the actual values of collagen concentration and excluded the interference of other chromophores in skin-mimicking phantoms. Furthermore, a high positive correlation was found in the animal study with SHG microscopy. We consider that the DRS is a potential technique and can evaluate skin condition objectively.

Theoretical modeling and experimental demonstration of Raman probe induced spectral dip for realizing a superluminal laser.

We have demonstrated experimentally a Diode-Pumped Alkali Laser (DPAL) with a Raman resonance induced dip in the center of the gain profile, in order to produce an anomalous dispersion, necessary for making the laser superluminal. Numerical calculations match closely with experimental results, and indicate that the laser is operating superluminally, with the group index far below unity (~0.00526) at the center of the dip. The estimated factor of enhancement in the sensitivity to cavity length perturbation is ~190, approximately equaling the inverse of the group index. This enhancement factor can be made much higher via optimal tuning of parameters. Such a laser has the potential to advance significantly the field of high-precision metrology, with applications such as vibrometry, accelerometry, and rotation sensing.

Cryopreserved human umbilical cord patch for in-utero spina bifida repair.

OBJECTIVE: To identify a patch system to repair surgically created spina bifida in a sheep model for its efficacy in healing the skin defect, protecting the underlying spinal cord and reducing the Chiari II malformation. METHODS: Spina bifida was created surgically in 16 fetuses from eight timed-pregnant sheep at gestational age of 75 days. Two fetuses did not survive the procedure. Repeat hysterotomy was performed at 95 days' gestation to cover the defect with either biocellulose film with underwater adhesive (BCF-adhesive) (n = 7) or human umbilical cord with suture (HUC-suture) (n = 7). Three fetuses without formation of the defect served as reference controls. The skin healing was examined by direct visualization after a planned Cesarean section at term, followed by histological analysis using hematoxylin and eosin and Masson's trichrome stains. Mid-sagittal sections of the fetal cranium and upper cervical spine were analyzed by a pediatric neuroradiologist who was blinded to the type of patch received. RESULTS: Three fetuses that received the BCF-adhesive and six fetuses that received the HUC-suture survived to term for final analysis. As a result of dislodgment of the BCF-adhesive, all spina bifida defects repaired using BCF-adhesive were not healed and showed exposed spinal cord with leakage of cerebrospinal fluid. In contrast, all spinal defects repaired by HUC-suture were healed with complete regrowth of epidermal, dermal and subdermal tissue components, with no exposed spinal cord. The maximal skin wound width was 21 ± 3.6 mm in the BCF-adhesive group but 3 ± 0.8 mm in the HUC-suture group (P < 0.001). The spinal cord area (P = 0.001) and the number of anterior horn cells (P = 0.03) was preserved to a greater degree in the HUC-suture group than in the BCF-adhesive group, whilst psammoma bodies, signifying neuronal degeneration, were only observed in the BCF-adhesive group. Anatomic changes, indicative of Chiari II malformation, were seen in all three fetuses of the BCF-adhesive group but in none of the HUC-suture group (P < 0.01). CONCLUSION: Cryopreserved umbilical cord graft is a promising regenerative patch for intrauterine repair of spina bifida.

High-speed modulation in ladder transitions in Rb atoms using high-pressure buffer gas.

Modulators using atomic systems are often limited in speed by the rate of spontaneous emission. One approach for overcoming this limit is to make use of a buffer gas such as Ethane, which causes rapid fine structure mixing of the P(1/2) and P(3/2) states, and broadens the absorption spectra of the D1 and D2 lines in alkali atoms. Employing this effect, we show that one can achieve high speed modulation using ladder transitions in Rubidium. We demonstrate a 100-fold increase, due to the addition of the buffer gas, in the modulation bandwidth using the 5S-5P-5D cascade system. The observed bandwidth of ~200 MHz is within a factor of 2.5 of the upper bound of ~0.51 GHz for the system used, and is limited by various practical constraints in our experiment. We also present numerical simulations for the system and predict that a much higher modulation speed should be achievable under suitable conditions. In combination with a tapered nano fiber or a SiN waveguide, it has the potential to be used for high-speed, low-power all-optical modulation.

Optically controlled waveplate at a telecom wavelength using a ladder transition in Rb atoms for all-optical switching and high speed Stokesmetric imaging.

We demonstrate an optically controlled waveplate at ~1323 nm using the 5S(1/2)-5P(1/2)-6S(1/2) ladder transition in a Rb vapor cell. The lower leg of the transitions represents the control beam, while the upper leg represents the signal beam. We show that we can place the signal beam in any arbitrary polarization state with a suitable choice of polarization of the control beam. Specifically, we demonstrate a differential phase retardance of ~180 degrees between the two circularly polarized components of a linearly polarized signal beam. We also demonstrate that the system can act as a Quarter Wave plate. The optical activity responsible for the phase retardation process is explained in terms of selection rules involving the Zeeman sublevels. As such, the system can be used to realize a fast Stokesmetric imaging system with a speed of ~3 MHz. When implemented using a tapered nano fiber embedded in a vapor cell, this system can be used to realize an ultra-low power all-optical switch as well as a Quantum Zeno Effect based all-optical logic gate by combining it with an optically controlled polarizer, previously demonstrated by us. We present numerical simulations of the system using a comprehensive model which incorporates all the relevant Zeeman sub-levels in the system, using a novel algorithm recently developed by us for efficient computation of the evolution of an arbitrary large scale quantum system.

Factor IX oligomerization underlies reduced activity upon disruption of physiological conditions.

Coagulation factor IX (FIX) is a serine protease that plays a pivotal role in the blood coagulation cascade. FIX deficiency leads to a blood clotting disorder known as haemophilia B. FIX, synthesized as a prepro-peptide of 461 amino acids, is processed and secreted into plasma. The protein undergoes numerous modifications, including, but not limited to glycosylation, γ-carboxylation and disulphide bond formation. Upon processing and limited proteolysis, the protein is converted into an active protease. Under physiological conditions, the FIX zymogen is a monomer. The purpose of this work was to analyse the conditions that may affect FIX monomeric state and promote and/or reduce oligomerization. Using native gel electrophoresis and size exclusion chromatography, we found that under decreased pH and ionic strength conditions, the FIX zymogen can oligomerize, resulting in the formation of higher molecular weight species, with a concomitant reduction in specific activity. Similarly, FIX oligomers formed readily with low bovine serum albumin (BSA) concentrations; however, increased BSA concentrations impeded FIX oligomerization. We hypothesize that normal blood physiological conditions are critical for maintaining active FIX monomers. Under conditions of stress associated with acidosis, electrolyte imbalance and low albumin levels, FIX oligomerization is expected to take place thus leading to compromised activity. Furthermore, albumin, which is commonly used as a drug stabilizer, may enhance the efficacy of FIX biological drugs by reducing oligomerization.

Comparison of cryopreserved amniotic membrane and umbilical cord tissue with dehydrated amniotic membrane/chorion tissue.

OBJECTIVE: To evaluate how the different processing methods cryopreservation and dehydration affect the structural integrity and biological composition of key signalling molecules within amniotic membrane and umbilical cord tissues. METHOD: We directly compared cryopreserved amniotic membrane (AM) and umbilical cord (UC) tissues with dehydrated amniotic membrane/chorion (dHACM) tissue using biochemical and functional assays including histological and histochemical staining, BCA, agarose gel electrophoresis, western blot, ELISA, and proliferation and cell death assays. RESULTS: Cryopreservation retains the native architecture of the AM/UC extracellular matrix and maintains the quantity and activity of key biological signals present in fresh AM/UC, including high molecular weight hyaluronic acid, heavy chain-HA complex, and pentraxin 3. In contrast, dehydrated tissues were structurally compromised and almost completely lacked these crucial components. CONCLUSION: The results presented here indicate that cryopreservation better preserves the structural and biological signaling molecules of foetal tissues.

Optically controlled polarizer using a ladder transition for high speed Stokesmetric Imaging and Quantum Zeno Effect based optical logic.

We demonstrate an optically controlled polarizer at ~1323 nm using a ladder transition in a Rb vapor cell. The lower leg of the 5S(1/2),F = 1->5P(1/2),F = 1,2->6S(1/2),F = 1,2 transitions is excited by a Ti:Sapphire laser locked to a saturated absorption signal, representing the control beam. A tunable fiber laser at ~1323 nm is used to excite the upper leg of the transitions, representing the signal beam. When the control beam is linearly polarized, it produces an excitation of the intermediate level with a particular orientation of the angular momentum. Under ideal conditions, this orientation is transparent to the signal beam if it has the same polarization as the control beam and is absorbed when it is polarized orthogonally. We also present numerical simulations of the system using a comprehensive model which incorporates all the relevant Zeeman sub-levels in the system, and identify means to improve the performance of the polarizer. A novel algorithm to compute the evolution of large scale quantum system enabled us to perform this computation, which may have been considered too cumbersome to carry out previously. We describe how such a polarizer may serve as a key component for high-speed Stokesmetric imaging. We also show how such a polarizer, combined with an optically controlled waveplate, recently demonstrated by us, can be used to realize a high speed optical logic gate by making use of the Quantum Zeno Effect. Finally, we describe how such a logic gate can be realized at an ultra-low power level using a tapered nanofiber embedded in a vapor cell.

Strongyloides stercoralis infection in an intestinal transplant recipient.

Strongyloides stercoralis is a helminth in tropical and subtropical areas. It may cause latent infection and progress to Strongyloides hyperinfection syndrome, which is associated with a high mortality rate. Transplant recipients under the treatment of immunosuppressant agents are at risk of severe S. stercoralis infection. According to related literature, most cases of S. stercoralis infection after solid organ transplantation are caused by reactivation of latent infections in the recipients, whereas only a few are acquired from the donors. We report on an intestinal transplant recipient who had S. stercoralis infection diagnosed by a larva of this parasite found in the stool from the ileostomy stoma 1 month after transplantation. The donor was considered the source of the infection because the donor was from an endemic area and had marked eosinophilia, and the recipient had no contact history or clinical manifestations related to the S. stercoralis infection before transplantation. The patient was treated with ivermectin and exhibited no evidence of infection after 7 months.

Potential targeting of the tumor microenvironment to improve cancer virotherapy.

In 2015, oncolytic virotherapy was approved for clinical use, and in 2017, recombinant adeno-associated virus (AAV) delivery was also approved. However, systemic administration remains challenging due to the limited number of viruses that successfully reach the target site. Although the US Food and Drug Administration (FDA) permits the use of higher doses of AAV to achieve greater rates of transduction, most AAV still accumulates in the liver, potentially leading to toxicity there and elsewhere. Targeting the tumor microenvironment is a promising strategy for cancer treatment due to the critical role of the tumor microenvironment in controlling tumor progression and influencing the response to therapies. Newly discovered evidence indicates that administration routes focusing on the tumor microenvironment can promote delivery specificity and transduction efficacy within the tumor. Here, we review approaches that involve modifying viral surface features, modulating the immune system, and targeting the physicochemical characteristics in tumor microenvironment to regulate therapeutic delivery. Targeting tumor acidosis presents advantages that can be leveraged to enhance virotherapy outcomes and to develop new therapeutic approaches that can be integrated with standard treatments.

High efficiency optical modulation at a telecom wavelength using the quantum Zeno effect in a ladder transition in Rb atoms.

We demonstrate a high-efficiency optical modulator at ~1323 nm using the quantum Zeno effect in a ladder transition in a Rb vapor cell. The lower leg of the transitions represents the control beam while the upper leg of the transitions represents the signal beam. The cross-modulation of the signal beam transmission is observed as the control beam is intensity modulated, and is explained in terms of the quantum Zeno effect. We observe a modulation depth of near 100% at frequencies up to 1 MHz and demonstrate modulation at speeds up to 75 MHz, with a 3 dB bandwidth of about 5 MHz, limited by the homogeneous linewidth of the intermediate state. We also describe how much higher modulation speeds could be realized by using a buffer gas to broaden the transitions. We identify and explain the special conditions needed for optimizing the modulation efficiency. Numerical simulations of modulation at ~1 GHz are presented. The maximum modulation speed is found to scale with the pressure-broadened linewidth of the intermediate state, so that much higher speeds should be attainable.

Analysis of F9 point mutations and their correlation to severity of haemophilia B disease.

Haemophilia B is an X-linked recessive disorder caused by deficiency of functional coagulation factor IX, which results almost exclusively from mutations in the F9 gene. We sought to determine features, which could distinguish between mutations that cause severe disease symptoms from those that cause non-severe disease symptoms. Towards this objective, we have performed a statistical analysis of reported point mutations in F9. These include: potential local changes in mRNA free energy, codon usage, charge and type of mutated amino acid, location of the mutation with regard to protein secondary structure and functional domain and amino acids' evolutionary conservation scores. Wilcoxon signed-rank tests showed highly significant differences between severe and non-severe disease causing mutations in their effect on free energy of small mRNA fragments and evolutionarily conserved amino acids. Our results suggest that information at the mRNA level as well as conservation of the amino acid correlate well with disease severity. This study demonstrates that computational tools may be used to characterize the severity of haemophilia B associated with point mutations and suggests their utility in predicting the outcome of sequence changes in recombinant proteins.

Aqueous CdPbS quantum dots for near-infrared imaging.

Quantum dots (QDs) are semiconducting nanocrystals that have photoluminescent (PL) properties brighter than fluorescent molecules and do not photo-bleach, ideal for in vivo imaging of diseased tissues or monitoring of biological processes. Near-infrared (NIR) fluorescent light within the window of 700-1000 nm, which is separated from the major absorption peaks of hemoglobin and water, has the potential to be detected several millimeters under the surface with minimal interference from tissue autofluorescence. Here we report the synthesis and bioimaging demonstration of a new NIR QDs system, namely, CdPbS, made by an aqueous approach with 3-mercaptopropionic acid (MPA) as the capping molecule. The aqueous-synthesized, MPA-capped CdPbS QDs exhibited an NIR emission in the range of 800-950 nm with x(i) ≥ 0.3, where x(i) denotes the initial Pb molar fraction during the synthesis. Optimal PL performance of the CdPbS QDs occurred at x(i) = 0.7, which was about 4 nm in size as determined by transmission electron microscopy, had a rock salt structure and a quantum yield of 12%. Imaging of CdPbS QDs was tested in membrane staining and transfection studies. Cells transfected with CdPbS QDs were shown to be visible underneath a slab of chicken muscle tissue of up to 0.7 mm in thickness without the use of multiple-photon microscopy.

Functionalized polymer spheres via one-step photoinduced synthesis for antimicrobial activity and gene delivery.

Despite the fact that polystyrene (PS) spheres have been developed as polymeric carriers or matrices for various biomedical applications, the synthesis of PS spheres is time-consuming. This work describes the fabrication of a uniform PS sphere, coated with silver nanoparticles (Ag-PS), by simultaneous photoinduced polymerization and reduction fabricated using x-rays in aqueous solution without any initiator. The solution contains only styrene, silver ions (Ag(+)), and poly(vinyl pyrrolidone) (PVP) as a stabilizer. The proposed mechanism of the formation of the Ag-PS nanocomposite spheres involves the generation of radicals in the aqueous solution to induce PS polymerization and the reduction of Ag. The distribution of the sizes of the core PS spheres in the Ag-PS nanocomposite spheres was systematically examined as a function of irradiation time, concentration of styrene, and amount of PVP. Ag-PS nanocomposite spheres exhibit antimicrobial activity against bacteria (Escherichia coli and Staphylococcus aureus). Additionally, the cationic (vinylbenzyl)trimethylammonium (TMA) monomer was photopolymerized to form positively charged TMA-PS spheres as gene carriers with uniquely low cytotoxicity. Given these design advantages, the method proposed herein is simpler than typical approaches for synthesizing PS spheres with functionalized groups and PS spheres coated with Ag nanoparticles.

Role of lipocalin 2 and its complex with matrix metalloproteinase-9 in oral cancer.

OBJECTIVES: Recent evidence demonstrated that lipocalin (LCN)2 is induced in many types of human cancer, while the detection of its complex with matrix metalloproteinase (MMP)-9 is correlated with the cancer disease status. We attempted to evaluate plasma expressions of LCN2, MMP-9, and their complex (LCN2/MMP-9) during the diagnostic work-up of patients with oral squamous cell carcinoma (OSCC) and investigated their correlations with disease progression. METHODS: In total, 195 patients with OSCC and 81 healthy controls were recruited. Expression levels of LCN2, MMP-9, and LCN2/MMP-9 were determined with immunoenzymatic assays. RESULTS: Patients with OSCC exhibited significantly higher levels of LCN2, MMP-9, and LCN2/MMP-9 compared with healthy controls (LCN2: P < 0.001; MMP-9: P < 0.001; LCN2/MMP-9: P < 0.01). Plasma levels of LCN2, MMP-9, and LCN2/MMP-9 in patients with OSCC were significantly correlated with each other and were associated with more-advanced clinical stages (P < 0.05) and/or a larger tumor size (P < 0.05), but were not associated with positive lymph-node metastasis or distal metastasis. CONCLUSION: Our results suggest that plasma levels of LCN2 and the LCN2/MMP-9 complex may be useful in non-invasively monitoring OSCC progression, while supporting their potential role as biomarkers of oral cancer disease status.

Anaerobic ammonium oxidation improvement via a novel capsule bioreactor.

This study developed a capsule bioreactor made from polyvinyl alcohol (PVA) to entrap anaerobic ammonium oxidation (ANAMMOX) bacteria for the removal of nitrogen from synthetic wastewater. Experimental results demonstrate that the ANAMMOX bacteria were entrapped inside the capsule and not washed out from the bioreactor. This method also avoids damaging the ANAMMOX bacteria during immobilization. The proposed bioreactor also has a shorter start-up period. The diffusion rate of ammonium and nitrite from the bulk solution through the capsule determined the rate of the ANAMMOX reaction. The optimal ratio of NO2(-)-N/NH4(+)-N was approximately 1 for the bioreactor. In addition, the capsule protected the ANAMMOX bacteria from nitrite inhibition. The bioreactor achieved a high stable nitrogen removal rate (5.8 Kg-N m(-3) day(-1)) at high nitrite levels (up to 600mg-N L(-1) in the influent). Therefore, the proposed capsule bioreactor has considerable potential for actual nitrogen removal applications.

Healthcare-associated COVID-19 outbreaks: a nationwide population-based cohort study.

BACKGROUND: Healthcare-associated coronavirus disease 2019 (COVID-19) has significant implications for patients, their companions and healthcare workers (HCWs). Controlling transmission in healthcare settings is critical to reduce deaths due to COVID-19. AIM: To describe the epidemiology and characteristics of healthcare-associated COVID-19 outbreaks and outbreak-related cases. METHODS: The investigation data for each healthcare-associated outbreak that occurred between 15th January 2020 and 31st July 2021 in Taiwan were analysed retrospectively. Confirmed outbreak-associated cases were categorized as HCW cases, patient companion cases or patient cases, and the characteristics of the confirmed cases were compared between these categories. FINDINGS: In total, 54 healthcare-associated COVID-19 outbreaks including 512 confirmed cases were reported. The median number of affected cases per outbreak was six [interquartile range (IQR) 2-12], and the median outbreak duration was 12 days (IQR 4.3-17.0). Only 5.7% and 0.2% of all confirmed cases were partially and fully vaccinated, respectively. Most outbreaks (90%, 48/54) occurred in May and June 2021. HCW cases, companion cases and patient cases accounted for 19.5%, 41.2% and 39.3% of the total cases. Patient cases were significantly older (median age 72 years, IQR 61-83) and had higher 30-day all-cause mortality (37.4%) than HCW cases (median age 41 years, IQR 28-58, 0%) and companion cases (median age 52 years; IQR 42-62, 1%). CONCLUSION: Healthcare-associated COVID-19 outbreaks have a critical impact on patients. Nevertheless, two-thirds of cases in the healthcare-associated outbreaks in this study comprised HCWs and companions. In order to effectively mitigate COVID-19 transmission in healthcare settings, multi-pronged infection prevention and control measures should be implemented and tailored for these three groups.