Dual-function antimicrobial-antibiofilm peptide hybrid to tackle biofilm-forming Staphylococcus epidermidis.

BACKGROUND: Due to their resistance and difficulty in treatment, biofilm-associated infections are problematic among hospitalized patients globally and account for 60% of all bacterial infections in humans. Antibiofilm peptides have recently emerged as an alternative treatment since they can be effectively designed and exert a different mode of biofilm inhibition and eradication. METHODS: A novel antibiofilm peptide, BiF, was designed from the conserved sequence of 18 α-helical antibiofilm peptides by template-assisted technique and its activity was improved by hybridization with a lipid binding motif (KILRR). Novel antibiofilm peptide derivatives were modified by substituting hydrophobic amino acids at positions 5 or 7, and both, with positively charged lysines (L5K, L7K). These peptide derivatives were tested for antibiofilm and antimicrobial activities against biofilm-forming Staphylococcus epidermidis and multiple other microbes using crystal violet and broth microdilution assays, respectively. To assess their impact on mammalian cells, the toxicity of peptides was determined through hemolysis and cytotoxicity assays. The stability of candidate peptide, BiF2_5K7K, was assessed in human serum and its secondary structure in bacterial membrane-like environments was analyzed using circular dichroism. The action of BiF2_5K7K on planktonic S. epidermidis and its effect on biofilm cell viability were assessed via viable counting assays. Its biofilm inhibition mechanism was investigated through confocal laser scanning microscopy and transcription analysis. Additionally, its ability to eradicate mature biofilms was examined using colony counting. Finally, a preliminary evaluation involved coating a catheter with BiF2_5K7K to assess its preventive efficacy against S. epidermidis biofilm formation on the catheter and its surrounding area. RESULTS: BiF2_5K7K, the modified antibiofilm peptide, exhibited dose-dependent antibiofilm activity against S. epidermidis. It inhibited biofilm formation at subinhibitory concentrations by altering S. epidermidis extracellular polysaccharide production and quorum-sensing gene expression. Additionally, it exhibited broad-spectrum antimicrobial activity and no significant hemolysis or toxicity against mammalian cell lines was observed. Its activity is retained when exposed to human serum. In bacterial membrane-like environments, this peptide formed an α-helix amphipathic structure. Within 4 h, a reduction in the number of S. epidermidis colonies was observed, demonstrating the fast action of this peptide. As a preliminary test, a BiF2_5K7K-coated catheter was able to prevent the development of S. epidermidis biofilm both on the catheter surface and in its surrounding area. CONCLUSIONS: Due to the safety and effectiveness of BiF2_5K7K, we suggest that this peptide be further developed to combat biofilm infections, particularly those of biofilm-forming S. epidermidis.

Cymbopogon citratus L. essential oil as a potential anti-biofilm agent active against antibiotic-resistant bacteria isolated from chronic rhinosinusitis patients.

Chronic rhinosinusitis (CRS) is long-term inflammation of the sinuses that can be caused by infection due to antibiotic-resistant bacteria. Biofilm developed by microbes is postulated to cause antibiotic treatment failure. Thus, the anti-biofilm activities of seven Thai herbal essential oils (EOs) against antibiotic-resistant bacteria isolated from CRS patients was investigated. Lemongrass (Cymbopogon citratus L.) EO showed the most effective antibiofilm activity against Klebsiella pneumoniae, Pseudomonas aeruginosa and Staphylococcus epidermidis grown as biofilm. GC-MS analysis found that myrcene was the major bioactive compound. Pretreatment with lemongrass EO significantly inhibited biofilm formation of all bacterial strains in more than 50% of cases. Furthermore, confocal microscopy analysis revealed the biofilm-disrupting activity of lemongrass EO against the biofilm matrix of all these bacterial species and also increased P. aeruginosa swarming motility with no toxicity to human cells. These results suggest that lemongrass EO has promising clinical applications as an anti-biofilm agent for CRS patients.

iAMY-SCM: Improved prediction and analysis of amyloid proteins using a scoring card method with propensity scores of dipeptides.

Fast, accurate identification and characterization of amyloid proteins at a large-scale is essential for understating their role in therapeutic intervention strategies. As a matter of fact, there exist only one in silico model for amyloid protein identification using the random forest (RF) model in conjunction with various feature types namely the RFAmy. However, it suffers from low interpretability for biologists. Thus, it is highly desirable to develop a simple and easily interpretable prediction method with robust accuracy as compared to the existing complicated model. In this study, we propose iAMY-SCM, the first scoring card method-based predictor for predicting and analyzing amyloid proteins. Herein, the iAMY-SCM made use of a simple weighted-sum function in conjunction with the propensity scores of dipeptides for the amyloid protein identification. Cross-validation results indicated that iAMY-SCM provided an accuracy of 0.895 that corresponded to 10-22% higher performance than that of widely used machine learning models. Furthermore, iAMY-SCM achieving an accuracy of 0.827 as evaluated by an independent test, which was found to be comparable to that of RFAmy and was approximately 9-13% higher than widely used machine learning models. Furthermore, the analysis of estimated propensity scores of amino acids and dipeptides were performed to provide insights into the biophysical and biochemical properties of amyloid proteins. As such, this demonstrates that the proposed iAMY-SCM is efficient and reliable in terms of simplicity, interpretability and implementation. To facilitate ease of use of the proposed iAMY-SCM, a user-friendly and publicly accessible web server at http://camt.pythonanywhere.com/iAMY-SCM has been established. We anticipate that that iAMY-SCM will be an important tool for facilitating the large-scale prediction and characterization of amyloid protein.

Anserine/Carnosine-Rich Extract from Thai Native Chicken Suppresses Melanogenesis via Activation of ERK Signaling Pathway.

Skin hyperpigmentation is an aesthetic problem that leads to psychosocial issues. Thus, skin whitening agents from agro- and poultry-industrial co-products are considered high economic value ingredients of interest for sustainable application. Therefore, this study aimed to determine the cosmeceutical potential of anserine/carnosine-rich chicken extract (ACCE) from the Thai native chicken Pradu Hang Dam Mor Kor 55 (PD) meat. The chemical composition was identified and quantified using the HPLC-UV method. Then, the antioxidation potential of the extract was compared to that of L-anserine and L-carnosine, using 1,1-diphenyl-2-picrylhydrazyl assay and shikonin-induced production of reactive oxygen species in CCD-986Sk cell models, and the anti-melanogenesis effect in the MNT-1 melanoma cell line model was investigated. Furthermore, related mechanisms were identified using colorimetric tyrosinase assay and the Western blot technique. The ACCE was composed of L-anserine and L-carnosine as two major constituents. In a dose-dependent manner, ACCE, L-anserine, and L-carnosine manifested significant antioxidation potential and significant reduction of melanin production. Activation of the extracellular signal-regulated kinase (ERK) signaling pathway and inhibition of tyrosinase activity of ACCE were demonstrated as the mechanisms of the anti-melanogenesis effect. In conclusion, ACCE has been revealed as a potential cosmeceutical agent due to its antioxidation and anti-melanogenic activity in association with L-anserine and L-carnosine composition and biomolecular regulating ability. Therefore, further studies and development should be considered to support the utilization of anserine/carnosine-rich chicken extract in the cosmetic industry for economic value creation and sustainability.

Nationwide Surveillance and Molecular Characterization of Critically Drug-Resistant Gram-Negative Bacteria: Results of the Research University Network Thailand Study.

A large-scale surveillance is an important measure to monitor the regional spread of antimicrobial resistance. We prospectively studied the prevalence and molecular characteristics of clinically important Gram-negative bacilli, including Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii complex (ABC), and Pseudomonas aeruginosa, from blood, respiratory tract, urine, and sterile sites at 47 hospitals across Thailand. Among 187,619 isolates, 93,810 isolates (50.0%) were critically drug resistant, of which 12,915 isolates (13.8%) were randomly selected for molecular characterization. E. coli was most commonly isolated from all specimens, except the respiratory tract, in which ABC was predominant. Prevalence of extended-spectrum cephalosporin resistance (ESCR) was higher in E. coli (42.5%) than K. pneumoniae (32.0%), but carbapenem-resistant (CR)-K. pneumoniae (17.2%) was 4.5-fold higher than CR-E. coli (3.8%). The majority of ESCR/CR-E. coli and K. pneumoniae isolates carried blaCTX-M (64.6% to 82.1%). blaNDM and blaOXA-48-like were the most prevalent carbapenemase genes in CR-E. coli/CR-K. pneumoniae (74.9%/52.9% and 22.4%/54.1%, respectively). In addition, 12.9%/23.0% of CR-E. coli/CR-K. pneumoniae cocarried blaNDM and blaOXA-48-like. Among ABC isolates, 41.9% were extensively drug resistant (XDR) and 35.7% were multidrug resistant (MDR), while P. aeruginosa showed XDR/MDR at 6.3%/16.5%. A. baumannii was the most common species among ABC isolates. The major carbapenemase gene in MDR-A. baumannii/XDR-A. baumannii was blaOXA-23-like (85.8%/93.0%), which had much higher rates than other ABC species. blaIMP, blaVIM, blaOXA-40-like, and blaOXA-58-like were also detected in ABC at lower rates. The most common carbapenemase gene in MDR/XDR-P. aeruginosa was blaIMP (29.0%/30.6%), followed by blaVIM (9.5%/25.3%). The findings reiterate an alarming situation of drug resistance that requires serious control measures.

Helicobacter pylori GroEL Seropositivity Is Associated with an Increased Risk of Opisthorchis viverrini-Associated Hepatobiliary Abnormalities and Cholangiocarcinoma.

Despite the synergistic effect of Opisthorchis viverrini and Helicobacter pylori co-infection on pathogenesis of severe hepatobiliary abnormalities (HBA) including advanced periductal fibrosis and replace with cholangiocarcinoma (CCA) have been established, the immune response to H. pylori in O. viverrini infected population has never been explored. Hence, this study aimed to investigate the antibody responses to 2 immunogenic H. pylori proteins in O. viverrini-infected patients with HBA and CCA. The risk analysis by multinomial logistic regression revealed that GroEL seropositivity was associated with higher risks of hepatobiliary abnormalities and CCA with adjusted odds ratios (95% confidence intervals) of 2.11 (95% CI=1.20-3.71, P=0.008) and 2.13 (95% CI=1.21-3.75, P=0.009), respectively. These findings indicate that GroEL seropositivity might be a biomarker for early detection of O. viverrini associated HBA and CCA.

iDPPIV-SCM: A Sequence-Based Predictor for Identifying and Analyzing Dipeptidyl Peptidase IV (DPP-IV) Inhibitory Peptides Using a Scoring Card Method.

The inhibition of dipeptidyl peptidase IV (DPP-IV, E.C.3.4.14.5) is well recognized as a new avenue for the treatment of Type 2 diabetes (T2D). Until now, peptide-like DDP-IV inhibitors have been shown to normalize the blood glucose concentration in T2D subjects. To the best of our knowledge, there is yet no computational model for predicting and analyzing DPP-IV inhibitory peptides using sequence information. In this study, we present for the first time a simple and easily interpretable sequence-based predictor using the scoring card method (SCM) for modeling the bioactivity of DPP-IV inhibitory peptides (iDPPIV-SCM). Particularly, the iDPPIV-SCM was developed by employing the SCM method together with the propensity scores of amino acids. Rigorous independent test results demonstrated that the proposed iDPPIV-SCM was found to be superior to those of well-known machine learning (ML) classifiers (e.g., k-nearest neighbor, logistic regression, and decision tree) with demonstrated improvements of 2-11, 4-22, and 7-10% for accuracy, MCC, and AUC, respectively, while also achieving comparable results to that of the support vector machine. Furthermore, the analysis of estimated propensity scores of amino acids as derived from the iDPPIV-SCM was performed so as to provide a more in-depth understanding on the molecular basis for enhancing the DPP-IV inhibitory potency. Taken together, these results revealed that iDPPIV-SCM was superior to those of other well-known ML classifiers owing to its simplicity, interpretability, and validity. For the convenience of biologists, the predictive model is deployed as a publicly accessible web server at http://camt.pythonanywhere.com/iDPPIV-SCM. It is anticipated that iDPPIV-SCM can serve as an important tool for the rapid screening of promising DPP-IV inhibitory peptides prior to their synthesis.

DNase-mediated eDNA removal enhances D-LL-31 activity against biofilms of bacteria isolated from chronic rhinosinusitis patients.

Chronic rhinosinusitis (CRS) is a chronic infection of the nasal cavity and paranasal sinuses associated with the presence of a microbial biofilm. Extracellular DNA (eDNA) is an important component of the biofilm matrix. Antimicrobial peptides (AMPs) are natural peptides with the ability to kill microorganisms. D-LL-31 is a synthetic variant of the AMP cathelicidin with increased resistance to proteolytic breakdown. In this study it is shown for 3 clinical CRS isolates that treatment of 24 h biofilms with DNase I enhanced the antimicrobial activity of D-LL-31. Conversely, co-incubation of D-LL-31 at the IC50 value with exogenous DNA resulted in reduced antimicrobial activity. DNase I alone did not show antimicrobial activity against the isolates tested but caused dispersal of an established biofilm. Hence, the presence of eDNA in the biofilm matrix reduced AMP-mediated killing. These results suggest that combination therapy with proteolysis resistant AMP D-LL-31 and DNase could be considered for effective treatment of CRS.

D-LL-31 in combination with ceftazidime synergistically enhances bactericidal activity and biofilm destruction in Burkholderia pseudomallei.

Melioidosis is a severe disease caused by Burkholderia pseudomallei. The biofilm of B. pseudomallei acquires resistance to several antibiotics and may be related to relapse in melioidosis patients. Here, the killing activity of antimicrobial peptides (LL-37, LL-31) and the D-enantiomers (D-LL-37, D-LL-31) in combination with ceftazidime (CAZ) against B. pseudomallei 1026b, H777 and a biofilm mutant M10, derived from H777 grown under biofilm-stimulating conditions was observed. Using static conditions, D-LL-31 exhibited the strongest killing activity against the three isolates in a dose-dependent manner. IC50 values for D-LL-31 ranged from 1 to 6 µM, for isolates M10, H777, and 1026b, respectively. Moreover, D-LL-31 combined with CAZ synergistically decreased the IC50 values of the peptide and antibiotic and caused also disruption of biofilms of B. pseudomallei 1026b under flow conditions. Thus a combination of D-LL-31 and CAZ may enhance the efficacy of the currently used antibiotic treatments against B. pseudomallei.

DETECTION OF HELICOBACTER PYLORI AND VIRULENCE-ASSOCIATED GENES IN SALIVA SAMPLES OF ASYMPTOMATIC PERSONS IN NORTHEAST THAILAND.

The aims of the study were to develop nested-PCR (targeting vacA and cagA), SYBR green quantitative PCR (targeting 16S rDNA) tests and compared them with indirect fluorescent-monoclonal antibody (IFA) method for determination of the prevalence of Helicobacter pylori in 118 saliva samples from asymptomatic individuals in Khon Kaen, Thailand. Detection limit of both PCR-based assays was one cell. Prevalence of H. pylori in saliva samples was 55% based on the criterion of positivity of IFA test and one of the PCR-based methods or positivity of both PCR assays. Forty-nine percent of H. pylori detected carried cagA, encoding a cytotoxin associated with severe clinical outcomes. These results imply that the mouth may be an important reservoir for H. pylori, with nearly 50% of the virulent type that could possibly lead to gastroduodenal disease.

Ultrastructural effects and antibiofilm activity of LFchimera against Burkholderia pseudomallei.

Lactoferrin chimera (LFchimera), a hybrid peptide containing the two antimicrobial stretches of the innate immunity factor bovine lactoferrin, viz. LFampin265-284 and LFcin17-30, has strikingly high antimicrobial activity against the category B pathogen Burkholderia pseudomallei. The action mechanisms of LFchimera against B. pseudomallei is not fully understood. The aim of this study was to further investigate the effect of treated B. pseudomallei with LFchimera using (immune) electron microscopy. The effects of LFchimera on biofilm formation and against preformed biofilm of B. pseudomallei were also determined. After exposure to LFchimera, transmission electron microscopy revealed swelling of the periplasmic space of B. pseudomallei and a highly inhomogeneous electron density in the intracellular DNA region. Localization of LFchimera in B. pseudomallei using immunoelectron microscopy showed gold particles in intracellular structures without accumulation on the membranes. LFchimera also possessed stronger bactericidal activity than ceftazidime against B. pseudomallei grown in biofilm. Moreover, limited exposure of B. pseudomallei to LFchimera at subcidal concentration could reduce biofilm formation. Altogether, the results indicate that LFchimera possesses antibacterial and antibiofilm activities and can modulate B. pseudomallei colonization. Therefore, the efficacy of LFchimera merits further development of this agent for the therapy of melioidosis.

Synergistic effects of isomorellin and forbesione with doxorubicin on apoptosis induction in human cholangiocarcinoma cell lines.

BACKGROUND: Chemotherapy for advanced cholangiocarcinoma (CCA) is largely ineffective, but innovative combinations of chemotherapeutic agents and natural compounds represent a promising strategy. In our previous studies, isomorellin and forbesione, caged xanthones isolated from Garcinia hanburyi, were found to induce cell cycle arrest and apoptosis in CCA cell lines. The subject of our inquiry is the synergistic effect(s) of these caged xanthones with doxorubicin on growth inhibition and apoptosis induction in human CCA cell lines. METHODS: KKU-100, KKU-M139 and KKU-M156 cell lines and Chang cells were treated with either isomorellin or forbesione alone or in combination with doxorubicin. Cell viability was determined using the sulforhodamine B assay. The combined effects of plant compounds with doxorubicin were analyzed using the isobologram and combination index method of Chou-Talalay. Apoptosis was determined by ethidium bromide/acridine orange staining. Protein expressions were determined by Western blot analysis. RESULTS: Isomorellin or forbesione alone inhibited the growth of these CCA cell lines in a dose-dependent manner and showed selective cytotoxicity against CCA cells but not against Chang cells. Isomorellin/doxorubicin combination showed a synergistic growth inhibitory effect on KKU-M139 and KKU-M156 cells, while the forbesione/doxorubicin combination showed a synergistic growth inhibitory effect on KKU-100 and KKU-M139 cells. The percentages of apoptotic cells were significantly higher in the combined treatments than in the respective single drug treatments. The combined treatments strongly enhanced the expression of Bax/Bcl-2, activated caspase-9 and caspase-3, while suppressing the expression of survivin, procaspase-9 and procaspase-3, compared with single drug treatments. The degree of suppression of NF-κB activation mediated by a decrease in the expression of NF-κB/p65, a reduction of the pIκB-α level and an increase in the IκB-α protein level, was significantly higher in the combined treatment groups than in the single drug treatment groups. The degree of suppression of MRP1 protein expression was also significantly higher in the combined treatment than in the single drug treatment groups. CONCLUSION: The combinations of isomorellin/doxorubicin and forbesione/doxorubicin showed significant synergistic effects on the growth inhibition and apoptosis induction in KKU-M156 and KKU-100 cells. Caged xanthones may be useful adjunct treatments with chemotherapy for Opisthorchis viverrini (OV)-associated CCA.

Membrane-active mechanism of LFchimera against Burkholderia pseudomallei and Burkholderia thailandensis.

LFchimera, a construct combining two antimicrobial domains of bovine lactoferrin, lactoferrampin265-284 and lactoferricin17-30, possesses strong bactericidal activity. As yet, no experimental evidence was presented to evaluate the mechanisms of LFchimera against Burkholderia isolates. In this study we analyzed the killing activity of LFchimera on the category B pathogen Burkholderia pseudomallei in comparison to the lesser virulent Burkholderia thailandensis often used as a model for the highly virulent B. pseudomallei. Killing kinetics showed that B. thailandensis E264 was more susceptible for LFchimera than B. pseudomallei 1026b. Interestingly the bactericidal activity of LFchimera appeared highly pH dependent; B. thailandensis killing was completely abolished at and below pH 6.4. FITC-labeled LFchimera caused a rapid accumulation within 15 min in the cytoplasm of both bacterial species. Moreover, freeze-fracture electron microscopy demonstrated extreme effects on the membrane morphology of both bacterial species within 1 h of incubation, accompanied by altered membrane permeability monitored as leakage of nucleotides. These data indicate that the mechanism of action of LFchimera is similar for both species and encompasses disruption of the plasma membrane and subsequently leakage of intracellular nucleotides leading to cell dead.

A heterodimer comprised of two bovine lactoferrin antimicrobial peptides exhibits powerful bactericidal activity against Burkholderia pseudomallei.

Melioidosis is a severe infectious disease that is endemic in Southeast Asia and Northern Australia. Burkholderia pseudomallei, the causative agent of this disease, has developed resistance to an increasing list of antibiotics, demanding a search for novel agents. Lactoferricin and lactoferrampin are two antimicrobial domains of lactoferrin with a broad spectrum of antimicrobial activity. A hybrid peptide (LFchimera) containing lactoferrampin (LFampin265-284) and a part of lactoferricin (LFcin17-30) has strikingly higher antimicrobial activities compared to the individual peptides. In this study, the antimicrobial activities of this chimeric construct (LFchimera1), as well as of another one containing LFcin17-30 and LFampin268-284, a shorter fragment of LFampin265-284 (LFchimera2), and the constituent peptides were tested against 7 isolates of B. pseudomallei and compared to the preferential antibiotic ceftazidime (CAZ). All isolates including B. pseudomallei 979b shown to be resistant to CAZ, at a density of 10(5) CFU/ml, could be killed by 5-10 µM of LFchimera1 within 2 h, while the other peptides as well as the antibiotic CAZ only inhibited the B. pseudomallei strains resulting in an overgrowth in 24 h. These data indicate that LFchimera1 could be considered for development of therapeutic agents against B. pseudomallei.

Vitamin D (1α,25(OH)2D3) supplementation minimized multinucleated giant cells formation and inflammatory response during Burkholderia pseudomallei infection in human lung epithelial cells.

Melioidosis is an infectious disease with high mortality rates in human, caused by the bacterium Burkholderia pseudomallei. As an intracellular pathogen, B. pseudomallei can escape from the phagosome and induce multinucleated giant cells (MNGCs) formation resulting in antibiotic resistance and immune evasion. A novel strategy to modulate host response against B. pseudomallei pathogenesis is required. In this study, an active metabolite of vitamin D3 (1α,25-dihydroxyvitamin D3 or 1α,25(OH)2D3) was selected to interrupt pathogenesis of B. pseudomallei in a human lung epithelium cell line, A549. The results demonstrated that pretreatment with 10-6 M 1α,25(OH)2D3 could reduce B. pseudomallei internalization to A549 cells at 4 h post infection (P < 0.05). Interestingly, the presence of 1α,25(OH)2D3 gradually reduced MNGC formation at 8, 10 and 12 h compared to that of the untreated cells (P < 0.05). Furthermore, pretreatment with 10-6 M 1α,25(OH)2D3 considerably increased hCAP-18/LL-37 mRNA expression (P < 0.001). Additionally, pro-inflammatory cytokines, including MIF, PAI-1, IL-18, CXCL1, CXCL12 and IL-8, were statistically decreased (P < 0.05) in 10-6 M 1α,25(OH)2D3-pretreated A549 cells by 12 h post-infection. Taken together, this study indicates that pretreatment with 10-6 M 1α,25(OH)2D3 has the potential to reduce the internalization of B. pseudomallei into host cells, decrease MNGC formation and modulate host response during B. pseudomallei infection by minimizing the excessive inflammatory response. Therefore, 1α,25(OH)2D3 supplement may provide an effective supportive treatment for melioidosis patients to combat B. pseudomallei infection and reduce inflammation in these patients.

A two-arm analysis of the immune response to heterologous boosting of inactivated SARS-CoV-2 vaccines.

Several vaccine programs were introduced during the COVID-19 pandemic, which included inactivated virus, DNA viral vectors and mRNA vaccines. Booster programs are recommended, especially for those in high-risk groups. However, many of these booster programs involve heterologous vaccines. This study enrolled volunteers who first received two full-dose CoronaVac vaccinations before receiving heterologous boosters with DNA- and/or mRNA-vaccines for an additional 2 doses (n = 40) or an additional 3 doses (n = 16). Our results showed no difference in side effects, neutralizing antibodies, or T-cell responses for any of the heterologous vaccination programs. However, the neutralizing capacity and IFN-γ responses against the Omicron variant in volunteers who received 4 or 5 doses were improved. Polarization of peripheral memory T cells after stimulation in all booster groups with Omicron peptide showed an increased trend of naïve and central memory phenotypes of both CD4+ and CD8+ T cells, suggesting that exposure to Omicron antigens will drive T cells into a lymphoid resident T cell phenotype. Our data support a continuous vaccination program to maximize the effectiveness of immunity, especially in people at high risk. Furthermore, the number of boosting doses is important for maintaining immunity.

Large-scale comparative review and assessment of computational methods for phage virion proteins identification.

Phage virion proteins (PVPs) are effective at recognizing and binding to host cell receptors while having no deleterious effects on human or animal cells. Understanding their functional mechanisms is regarded as a critical goal that will aid in rational antibacterial drug discovery and development. Although high-throughput experimental methods for identifying PVPs are considered the gold standard for exploring crucial PVP features, these procedures are frequently time-consuming and labor-intensive. Thusfar, more than ten sequence-based predictors have been established for the in silico identification of PVPs in conjunction with traditional experimental approaches. As a result, a revised and more thorough assessment is extremely desirable. With this purpose in mind, we first conduct a thorough survey and evaluation of a vast array of 13 state-of-the-art PVP predictors. Among these PVP predictors, they can be classified into three groups according to the types of machine learning (ML) algorithms employed (i.e. traditional ML-based methods, ensemble-based methods and deep learning-based methods). Subsequently, we explored which factors are important for building more accurate and stable predictors and this included training/independent datasets, feature encoding algorithms, feature selection methods, core algorithms, performance evaluation metrics/strategies and web servers. Finally, we provide insights and future perspectives for the design and development of new and more effective computational approaches for the detection and characterization of PVPs.

SCMRSA: a New Approach for Identifying and Analyzing Anti-MRSA Peptides Using Estimated Propensity Scores of Dipeptides.

Staphylococcus aureus is deemed to be one of the major causes of hospital and community-acquired infections, especially in methicillin-resistant S. aureus (MRSA) strains. Because antimicrobial peptides have captured attention as novel drug candidates due to their rapid and broad-spectrum antimicrobial activity, anti-MRSA peptides have emerged as potential therapeutics for the treatment of bacterial infections. Although experimental approaches can precisely identify anti-MRSA peptides, they are usually cost-ineffective and labor-intensive. Therefore, computational approaches that are able to identify and characterize anti-MRSA peptides by using sequence information are highly desirable. In this study, we present the first computational approach (termed SCMRSA) for identifying and characterizing anti-MRSA peptides by using sequence information without the use of 3D structural information. In SCMRSA, we employed an interpretable scoring card method (SCM) coupled with the estimated propensity scores of 400 dipeptides. Comparative experiments indicated that SCMRSA was more effective and could outperform several machine learning-based classifiers with an accuracy of 0.960 and Matthews correlation coefficient of 0.848 on the independent test data set. In addition, we employed the SCMRSA-derived propensity scores to provide a more in-depth explanation regarding the functional mechanisms of anti-MRSA peptides. Finally, in order to serve community-wide use of the proposed SCMRSA, we established a user-friendly webserver which can be accessed online at http://pmlabstack.pythonanywhere.com/SCMRSA. SCMRSA is anticipated to be an open-source and useful tool for screening and identifying novel anti-MRSA peptides for follow-up experimental studies.

Phenotypic and functional changes of T cell subsets after CoronaVac vaccination.

BACKGROUND: The pandemic coronavirus disease 2019 (COVID-19) is a major global public health concern and several protective vaccines, or preventive/therapeutic approaches have been developed. Sinovac-CoronaVac, an inactivated whole virus vaccine, can protect against severe COVID-19 disease and hospitalization, but less is known whether it elicits long-term T cell responses and provides prolonged protection. METHODS: This is a longitudinal surveillance study of SARS-CoV-2 receptor binding domain (RBD)-specific IgG levels, neutralizing antibody levels (NAb), T cell subsets and activation, and memory B cells of 335 participants who received two doses of CoronaVac. SARS-CoV-2 RBD-specific IgG levels were measured by enzyme-linked immunosorbent assay (ELISA), while NAb were measured against two strains of SARS-CoV-2, the Wuhan and Delta variants. Activated T cells and subsets were identified by flow cytometry. Memory B and T cells were evaluated by enzyme-linked immune absorbent spot (ELISpot). FINDINGS: Two doses of CoronaVac elicited serum anti-RBD antibody response, elevated B cells with NAb capacity and CD4+ T cell-, but not CD8+ T cell-responses. Among the CD4+ T cells, CoronaVac activated mainly Th2 (CD4+ T) cells. Serum antibody levels significantly declined three months after the second dose. INTERPRETATION: CoronaVac mainly activated B cells but T cells, especially Th1 cells, were poorly activated. Activated T cells were mainly Th2 biased, demonstrating development of effector B cells but not long-lasting memory plasma cells. Taken together, these results suggest that protection with CoronaVac is short-lived and that a third booster dose of vaccine may improve protection.

Assessment of in vitro activities of novel modified antimicrobial peptides against clarithromycin resistant Mycobacterium abscessus.

Mycobacterium abscessus (Mab) is one of the most drug resistant bacteria with a high treatment failure rate. Antimicrobial peptides (AMPs) are alternative therapeutic agents against this infection. This study was aimed to assess the in vitro activities of thirteen AMPs (S5, S52, S6, S61, S62, S63, KLK, KLK1, KLK2, Pug-1, Pug-2, Pug-3 and Pug-4) that have never been investigated against drug resistant Mab isolates. Only four novel modified AMPs (S61, S62, S63 and KLK1) provided the lowest minimum inhibitory concentration (MIC) values ranging from 200-400 µg/ml against the Mab ATCC19977 strain. These four potential AMPs were further tested with 16 clinical isolates of clarithromycin resistant Mab. The majority of the tested strains (10/16 isolates, 62.5%) showed ~99% kill by all four AMPs within 24 hours with an MIC <50 µg/ml. Only two isolates (12.5%) with acquired clarithromycin resistance, however, exhibited values <50 µg/ml of four potential AMPs, S61, S62, S63 and KLK1 after 3-days-incubation. At the MICs level, S63 showed the lowest toxicity with 1.50% hemolysis and 100% PBMC viability whereas KLK1 showed the highest hemolysis (10.21%) and lowest PBMC viability (93.52%). S61, S62 and S63 were further tested with clarithromycin-AMP interaction assays and found that 5/10 (50%) of selected isolates exhibited a synergistic interaction with 0.02-0.41 FICI values. This present study demonstrated the potential application of novel AMPs as an adjunctive treatment with clarithromycin against drug resistant Mab infection.