Molecular analysis of accessory gene regulator functionality and virulence genes in Staphylococcus aureus derived from pediatric wound infections

Staphylococcus aureus is a major human pathogen causing infections with high morbidity and mortality in both healthcare and community settings. The accessory gene regulator (Agr) is a key genetic element controlling the expression of numerous virulence factors in S. aureus. The significance of a functional Agr system in clinical S. aureus isolates derived from pediatric wound infections is still unclear. Therefore, the present study was conducted to identify virulence genes and determine Agr functionality from this cohort of patients. A total of 48 S. aureus wound isolates were collected from patients referred to Tehran Children's Medical Center Hospital from April 2017 to April 2018. In addition, in vitro antimicrobial susceptibility of the isolates was assessed using the disk diffusion and E-test methods. Conventional PCR was performed for the detection of toxins (tsst-1, hla, hlb, hld, eta, etb, etd, edin-A, edin-B, edin-C) and Agr typing (agrI, agrII, agrIII, agrIV). Agr functionality was assessed by quantitative reverse transcriptase real-time PCR (qRT-PCR). All S. aureus isolates were found to be susceptible to linezolid and vancomycin. The most frequently detected toxin gene was eta (100%), and the most prevalent Agr type was agrIII (56.3%). Importantly, qRT-PCR revealed that Agr was functional in 28 (58%) of wound isolates. Consequently, our data suggests that a functional Agr system may not be required for the development of S. aureus wound infections.</p

Unveiling promising immunogenic targets in Coxiella burnetii through in silico analysis: paving the way for novel vaccine strategies

Abstract Background Coxiella burnetii, an intracellular pathogen, serves as the causative agent of zoonotic Q fever. This pathogen presents a significant threat due to its potential for airborne transmission, environmental persistence, and pathogenicity. The current whole-cell vaccine (WCV) utilized in Australia to combat Q fever exhibits notable limitations, including severe adverse reactions and limited regulatory approval for human use. This research employed the reverse vaccinology (RV) approach to uncover antigenic proteins and epitopes of C. burnetii, facilitating the development of more potent vaccine candidates. Methods The potential immunogenic proteins derived from C. burnetii RSA493/Nine Mile phase I (NMI) were extracted through manual, automated RV, and virulence factor database (VFDB) methods. Web tools and bioinformatics were used to evaluate physiochemical attributes, subcellular localization, antigenicity, allergenicity, human homology, B-cell epitopes, MHC I and II binding ratios, functional class scores, adhesion probabilities, protein-protein interactions, and molecular docking. Results Out of the 1850 proteins encoded by RSA493/NMI, a subset of 178 demonstrated the potential for surface or membrane localization. Following a series of analytical iterations, 14 putative immunogenic proteins emerged. This collection included nine proteins (57.1%) intricately involved in cell wall/membrane/envelope biogenesis processes (CBU_0197 (Q83EW1), CBU_0311 (Q83EK8), CBU_0489 (Q83E43), CBU_0939 (Q83D08), CBU_1190 (P39917), CBU_1829 (Q83AQ2), CBU_1412 (Q83BU0), CBU_1414 (Q83BT8), and CBU_1600 (Q83BB2)). The CBU_1627 (Q83B86 ) (7.1%) implicated in intracellular trafficking, secretion, and vesicular transport, and CBU_0092 (Q83F57) (7.1%) contributing to cell division. Additionally, three proteins (21.4%) displayed uncharacterized functions (CBU_0736 (Q83DJ4), CBU_1095 (Q83CL9), and CBU_2079 (Q83A32)). The congruent results obtained from molecular docking and immune response stimulation lend support to the inclusion of all 14 putative proteins as potential vaccine candidates. Notably, seven proteins with well-defined functions stand out among these candidates. Conclusions The outcomes of this study introduce promising proteins and epitopes for the forthcoming formulation of subunit vaccines against Q fever, with a primary emphasis on cellular processes and the virulence factors of C. burnetii

Molecular analysis of accessory gene regulator functionality and virulence genes in Staphylococcus aureus derived from pediatric wound infections

Staphylococcus aureus is a major human pathogen causing infections with high morbidity and mortality in both healthcare and community settings. The accessory gene regulator (Agr)is a key genetic element controlling the expression of numerous virulence factors in S. aureus. The significance of a functional Agr system in clinical S. aureus isolates derived from pediatric wound infections is still unclear. Therefore, the present study was conducted to identify virulence genes and determine Agr functionality from this cohort of patients. A total of 48 S. aureus wound isolates were collected from patients referred to Tehran Children's Medical Center Hospital from April 2017 to April 2018. In addition, in vitro antimicrobial susceptibility of the isolates was assessed using the disk diffusion and E-test methods. Conventional PCR was performed for the detection of toxins (tsst-1, hla, hlb, hld, eta, etb, etd, edin-A, edin-B, edin-C)and Agr typing (agrI, agrII, agrIII, agrIV). Agr functionality was assessed by quantitative reverse transcriptase real-time PCR (qRT-PCR). All S. aureus isolates were found to be susceptible to linezolid and vancomycin. The most frequently detected toxin gene was eta (100%), and the most prevalent Agr type was agrIII (56.3%). Importantly, qRT-PCR revealed that Agr was functional in 28 (58%)of wound isolates. Consequently, our data suggests that a functional Agr system may not be required for the development of S. aureus wound infections.</p

Quality Control Strategy for Automated CBC: A Laboratory Point of View Deducted from an Internal Study Organised in an Emergency Laboratory

Introduction: The aim of this study was to determine the performance of the total testing process of complete blood count (CBC) on two different instruments in an emergency setting of a county hospital, and to design an appropriate internal quality control plan

In silico analysis of the substitution mutations and evolutionary trends of the SARS-CoV-2 structural proteins in Asia

OBJECTIVES: To address a highly mutable pathogen, mutations must be evaluated. SARS-CoV-2 involves changing infectivity, mortality, and treatment and vaccination susceptibility resulting from mutations. MATERIALS AND METHODS: We investigated the Asian and worldwide samples of amino-acid sequences (AASs) for envelope (E), membrane (M), nucleocapsid (N), and spike (S) proteins from the announcement of the new coronavirus 2019 (COVID-19) up to January 2022. Sequence alignment to the Wuhan-2019 virus permits tracking mutations in Asian and global samples. Furthermore, we explored the evolutionary tendencies of structural protein mutations and compared the results between Asia and the globe. RESULTS: The mutation analyses indicated that 5.81%, 70.63%, 26.59%, and 3.36% of Asian S, E, M, and N samples did not display any mutation. Additionally, the most relative mutations among the S, E, M, and N AASs occurred in the regions of 508 to 635 AA, 7 to 14 AA, 66 to 88 AA, and 164 to 205 AA in both Asian and total samples. D614G, T9I, I82T, and R203M were inferred as the most frequent mutations in S, E, M, and N AASs. Timeline research showed that substitution mutation in the location of 614 among Asian and total S AASs was detected from January 2020. CONCLUSION: N protein was the most non-conserved protein, and the most prevalent mutations in S, E, M, and N AASs were D614G, T9I, I82T, and R203M. Screening structural protein mutations is a robust approach for developing drugs, vaccines, and more specific diagnostic tools

Supplementary Material for: Evidence of Interruption of the comM Gene in a Large Series of Clinical Isolates of Multidrug-Resistant Acinetobacter baumannii

<p>Recent studies have recognized the ATPase-encoding <i>comM</i> gene as a hot spot for the integration of <i>Acinetobacter baumannii</i> resistance islands (RIs). Despite the circulation of high numbers of multidrug-resistant <i>A. baumannii </i>(MDR-AB) isolates in Middle East countries, no information is available about the interruption of <i>comM</i> and subsequent transposition into <i>comM</i> in isolates belonging to the global clones (GC) GC1, GC2, or GC3. In this study 401 <i>A. baumannii</i> isolates from hospitals in Tehran, Iran, were included. The resistance profile was determined by disc diffusion against 22 antibiotics. PCR was used to assess the GC type, presence of the <i>comM</i> gene, and the boundary junctions (J1 and J2) of RIs. Most of the MDR-AB isolates (384 of 388; 98%) and more than half of the susceptible <i>A. baumannii </i>isolates (9 of 13; 69%) had interrupted <i>comM</i> gene-carrying integrative elements. Among the isolates tested, 57 belonged to GC1, 86 to GC2, and 8 to GC3. A set of 250 isolates showed distinct patterns of allele-specific PCR for <i>ompA</i>, <i>csuE</i>, and <i>bla</i>_OXA-51-like genes. All but 2 of the GC1 isolates and 2 of the GC2 isolates contained interrupted <i>comM</i> genes. Four <i>A. baumannii</i> isolates harbored intact <i>comM</i>, but were multiply resistant to antibiotics. This study demonstrated that the <i>comM</i> gene is targeted by transposons in Iranian MDR-AB isolates belonging to different GCs. The data also showed that the carriage of interrupted <i>comM</i> is not exclusive to MDR isolates of <i>A. baumannii</i>.</p

Antibacterial activity of gold nanoparticles conjugated by aminoglycosides against A. baumannii isolates from burn patients

BACKGROUND AND OBJECTIVE: Today, resistance to multiple classes of antibiotics, and notably to the beta-lactam and aminoglycosides in A. baumannii is becoming great problem and it's necessitate to make new approach to combat with multidrug-resistant (MDR), extensive drug-resistance (XDR) or Pandrug-resistant (PDR) isolates. In this case new strategy and ways should be designed and introduce against such infections. Therefore the aim of present study was evaluation of antibacterial activity of nanoconjugate gentamicin and amikacin with gold against clinical isolates of A. baumannii that collected from burn wound infection. METHODS: Eighteen A. baumannii collected from burn wound infections. For confirmation and detection of aminoglycoside resistant genes PCR carried out. Gold nanoparticles and nanoconjugates prepared according to protocol. For evaluation of the nanoconjugate, Dynamic light cattering, Transmission electron microscopy and FTIR Analysis were carried out. Then, the antibacterial activity of nanoconjugates were done by using micro broth dilution method, Result: prevalence of aminoglycoside resistant genes were aacC1, aphA6, aadA1, aadB genes 55.5, 22.2, 38.8 and 22.2 respectively. Synthesis of bare nanoconjugates resulted nanoparticle in a size 10 nm. Amikacin bound to Gnps showed excellent antibacterial activity (94.5) and just one isolate was intermediate resistance. Also gentamicin bound to GNPS have good antimicrobial effect (50) contrast to gentamicin alone. CONCLUSION: our study showed that a combination of amikacin and gentamicin with Gnps has a significant antibacterial efficiency against clinical isolates of A. baumannii. Gnps can be used as extraordinary molecular carriers for the targeting, and delivery the of antibiotic molecules to the specific infection