Distribution of virulence genes in clinical isolates of hospital-associated and community-associated methicillin-susceptible Staphylococcus aureus from Terengganu, Malaysia.

Staphylococcus aureus is a common bacterial pathogen known to cause various kinds of infections due to its repertoire of virulence factors. This study aimed to investigate the distribution of 19 types of virulence genes among clinical isolates of methicillin-susceptible S. aureus (MSSA) using the polymerase chain reaction. A total of 109 MSSA isolates, i.e., 63 hospital-associated (HA) and 46 community-associated (CA) were collected from Hospital Sultanah Nur Zahirah, the main tertiary hospital in Terengganu, Malaysia, from July 2016 to June 2017. The most frequent virulence genes detected were hla (78.9%, n=86) and hld (78.0%, n=85) encoding hemolysins, lukED (56.9%, n=62) encoding leukotoxin ED, followed by seb (26.6%, n=29) and sea (24.8%, n=27) encoding enterotoxins. Among 34 (31.2%) isolates carrying six or more virulence genes, only five were multidrug resistant (MDR) while the remaining isolates were susceptible. Significant associations were discovered between the hld gene with CA-MSSA (p=0.016) and the seo gene with HA-MSSA (p=0.023). However, there is no significant association between virulence genes among the different types of infection. The clinical MSSA isolates in Terengganu showed high prevalence and high diversity of virulence gene carriage.

A unique combination of natural fatty acids from Hermetia illucens fly larvae fat effectively combats virulence factors and biofilms of MDR hypervirulent mucoviscus Klebsiella pneumoniae strains by increasing Lewis acid-base/van der Waals interactions in bacterial wall membranes.

Introduction: Hypervirulent Klebsiella pneumoniae (hvKp) and carbapenem-resistant K. pneumoniae (CR-Kp) are rapidly emerging as opportunistic pathogens that have a global impact leading to a significant increase in mortality rates among clinical patients. Anti-virulence strategies that target bacterial behavior, such as adhesion and biofilm formation, have been proposed as alternatives to biocidal antibiotic treatments to reduce the rapid emergence of bacterial resistance. The main objective of this study was to examine the efficacy of fatty acid-enriched extract (AWME3) derived from the fat of Black Soldier Fly larvae (Hermetia illucens) in fighting against biofilms of multi-drug resistant (MDR) and highly virulent Klebsiella pneumoniae (hvKp) pathogens. Additionally, the study also aimed to investigate the potential mechanisms underlying this effect. Methods: Crystal violet (CV) and ethidium bromide (EtBr) assays show how AWME3 affects the formation of mixed and mature biofilms by the KP ATCC BAA-2473, KPi1627, and KPM9 strains. AWME3 has shown exceptional efficacy in combating the hypermucoviscosity (HMV) virulent factors of KPi1627 and KPM9 strains when tested using the string assay. The rudimentary motility of MDR KPM9 and KP ATCC BAA-2473 strains was detected through swimming, swarming, and twitching assays. The cell wall membrane disturbances induced by AWME3 were detected by light and scanning electron microscopy and further validated by an increase in the bacterial cell wall permeability and Lewis acid-base/van der Waals characteristics of K. pneumoniae strains tested by MATS (microbial adhesion to solvents) method. Results: After being exposed to 0.5 MIC (0.125 mg/ml) of AWME3, a significant reduction in the rudimentary motility of MDR KPM9 and KP ATCC BAA-2473 strains, whereas the treated bacterial strains exhibited motility between 4.23 ± 0.25 and 4.47 ± 0.25 mm, while the non-treated control groups showed significantly higher motility ranging from 8.5 ± 0.5 to 10.5 ± 0.5 mm. Conclusion: In conclusion, this study demonstrates the exceptional capability of the natural AWME3 extract enriched with a unique combination of fatty acids to effectively eliminate the biofilms formed by the highly drug-resistant and highly virulent K. pneumoniae (hvKp) pathogens. Our results highlight the opportunity to control and minimize the rapid emergence of bacterial resistance through the treatment using AWME3 of biofilm-associated infections caused by hvKp and CRKp pathogens.

Antibiofilm and Antivirulence Potentials of 3,2'-Dihydroxyflavone against Staphylococcus aureus.

Staphylococcus aureus, particularly drug-resistant strains, poses significant challenges in healthcare due to its ability to form biofilms, which confer increased resistance to antibiotics and immune responses. Building on previous knowledge that several flavonoids exhibit antibiofilm activity, this study sought to identify a novel flavonoid capable of effectively inhibiting biofilm formation and virulence factor production in S. aureus strains including MRSA. Among the 19 flavonoid-like compounds tested, 3,2'-dihydroxyflavone (3,2'-DHF) was identified for the first time as inhibiting biofilm formation and virulence factors in S. aureus with an MIC 75 µg/mL. The antibiofilm activity was further confirmed by microscopic methods. Notably, 3,2'-DHF at 5 µg/mL was effective in inhibiting both mono- and polymicrobial biofilms involving S. aureus and Candida albicans, a common co-pathogen. 3,2'-DHF reduces hemolytic activity, slime production, and the expression of key virulence factors such as hemolysin gene hla and nuclease gene nuc1 in S. aureus. These findings highlight the potential of 3,2'-DHF as a novel antibiofilm and antivirulence agent against both bacterial and fungal biofilms, offering a promising alternative to traditional antibiotics in the treatment of biofilm-associated infections.

Integrating multi-wet laboratory diagnostics to study staphylococci in animals in Uganda.

BACKGROUND: Several diagnostic environments in Uganda lack real-time, robust and high-throughput technologies for comprehensive typing of microbes, which is a setback to infectious disease surveillance. This study combined various wet laboratory diagnostics to understand the epidemiology of pathogenic staphylococci isolated from animals in Uganda and the implications for global health security priorities. METHODS: A retrospective study was conducted employing records and pathogenic staphylococci (from animals) archived at the Central Diagnostic Laboratory (CDL), Makerere University, Uganda, between January 2012 and December 2019. The bacteria were speciated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and tested for virulence factors [beta lactamases, lecithinase, deoxyribonuclease (DNase), haemolysins] and resistance to ten antimicrobials of clinical and veterinary relevance. Tetracycline and methicillin resistance genes were also tested. RESULTS: The prevalent diseases were mastitis in cattle and skin infections in dogs. Of the 111 staphylococci tested by MALDI-TOF MS, 79 (71.2%) were Staphylococcus aureus, 27 (24.3%) were Staphylococcus pseudintermedius and 5 (4.5%) were Staphylococcus schleiferi. All these strains expressed haemolysins. The prevalence of strains with lecithinase, penicillinase, cephalosporinase and DNase was 35.9% (14/39), 89.7% (35/39), 0.0% (0/39) and 87.2% (34/39), respectively. Staphylococci were primarily resistant to early penicillins (over 80%), tetracycline (57.7%), and chloramphenicol (46.2%). Minimal resistance was noted with cloxacillin (0.0%), ciprofloxacin (9.6%), and cefoxitin (3.8%). The prevalence of multidrug resistance (MDR) was 78.8% for general staphylococci, 82.2% for S. aureus, 73.1% for S. pseudintermedius, and 60.0% for S. schleiferi. Multidrug resistant staphylococci were significantly more prevalent in the cattle isolates than in the dog isolates (P < 0.05). The prevalence of methicillin-resistant staphylococci (MRS) tested by resistance to cefoxitin and mecA carriage was 3.8%. These four strains were all isolated from dog skin infections. The tetK gene was the most predominant (35.4%), followed by tetM (25.0%). CONCLUSION: In resource-constrained settings, the approach of integrated diagnostics promises sustainable disease surveillance and the addressing of current capacity gaps. The emergence of MRS (zoonotic bacteria) in companion animals creates a likelihood of reduced treatment options for related human infections, a threat to global health.

Whole-genome sequencing of multidrug-resistant Klebsiella pneumoniae with capsular serotype K2 isolates from mink in China.

BACKGROUND: Klebsiella pneumoniae is a zoonotic opportunistic pathogen, and also one of the common pathogenic bacteria causing mink pneumonia. The aim of this study was to get a better understanding of the whole-genome of multi-drug resistant Klebsiella pneumoniae with K2 serotype in China. This study for the first time to analyze Gene Ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, resistance and virulence genes of Klebsiella pneumoniae in mink. RESULTS: The isolate was Klebsiella pneumoniae with serotype K2 and ST6189 by PCR method. The string test was positive and showed high mucus phenotype. There was one plasmid with IncFIB replicons in the genome. The virulence factors including capsule, lipopolysaccharide, adhesin, iron uptake system, urease, secretory system, regulatory gene (rcsA, rcsB), determinants of pili adhesion, enolase and magnesium ion absorption related genes. The strain was multi-drug resistant. A total of 26  resistance genes, including beta-lactam, aminoglycosides, tetracycline, fluoroquinolones, sulfonamides, amide alcohols, macrolides, rifampicin, fosfomycin, vancomycin, diaminopyrimidines and polymyxin. Multidrug-resistant efflux protein AcrA, AcrB, TolC, were predicted in the strain. CONCLUSION: It was the first to identify that serotype K2 K. pneumonia with ST6189 isolated from mink in China. The finding indicated that hypervirulent and multi-drug resistant K. pneumoniae was exist in Chinese mink. The whole-genome of K. pneumoniae isolates have importance in mink farming practice.

Helicobacter pylori Outer Membrane Vesicles: Biogenesis, Composition, and Biological Functions.

Helicobacter pylori has been recognized not only as a causative agent of a spectrum of gastroduodenal diseases including chronic gastritis, peptic ulcer, mucosa-associated lymphoid tissue lymphoma, and gastric cancer, but also as the culprit in several extra-gastric diseases. However, the association of H. pylori infection with extra-gastric diseases remains elusive, prompting a reevaluation of the role of H. pylori-derived outer membrane vesicles (OMVs). Like other gram-negative bacteria, H. pylori constitutively sheds biologically active OMVs for long-distance delivery of bacterial virulence factors in a concentrated and protected form, averting the need of direct bacterial contact with distant host cells to induce extra-gastric diseases associated with this gastric pathogen. Additionally, H. pylori-derived OMVs contribute to bacterial survival and chronic gastric pathogenesis. Moreover, the immunogenic activity, non-replicable nature, and anti-bacterial adhesion effect of H. pylori OMVs make them a desirable vaccine candidate against infection. The immunogenic potency and safety concerns of the OMV contents are challenges in the development of H. pylori OMV-based vaccines. In this review, we discuss recent advances regarding H. pylori OMVs, focusing on new insights into their biogenesis mechanisms and biological functions.

Antimicrobial resistance profile of Staphylococcus aureus isolated from patients, healthcare workers, and the environment in a tertiary hospital in Addis Ababa, Ethiopia.

Staphylococcus aureus infection and colonization in patients may be transmitted to healthcare providers and the environment and subsequently cause healthcare-associated infections in other patients. Pathogenic S. aureus strains produce virulence factors, such as Panton-Valentine Leukocidin (PVL), that contribute to the severity of infections and aid in their spread. The emergence of antimicrobial resistance (AMR) is additional concern with respect to S. aureus infection. In this study, the virulence genes and antibiotic resistance profiles of S. aureus were characterized from patients' clinical isolates, healthcare workers' (HCWs') nasal colonization screenings, and the environment at a tertiary healthcare hospital in Addis Ababa, Ethiopia. A total of 365 samples were collected from September 2021 to September 2022: 73 patients' clinical specimens, 202 colonization screenings from HCWs, and 90 hospital environment's swabs. Fifty-one (25.2%) HCW and 10/90 (11.1%) environment S. aureus isolates were identified. Among the 134 isolates, 10 (7.5%) were methicillin-resistant S. aureus (MRSA). Three (4.1%), five (9.8%), and two (20.0%) of the MRSA isolates were identified from the patients, HCWs, and the environment, respectively. Overall, 118 (88.1%) were ampicillin and penicillin resistant; 70 (52.2%) were trimethoprim sulfamethoxazole resistant; and 28 (20.9%) were erythromycin resistant. S. aureus isolates from patients were more resistant to antibiotics than isolates from HCWs or the hospital environment (p<0.05). A total of 92/134 (68.6%) isolates possessed the lukfF-PV gene, which was identified in 62 (85.0%), 26 (51.0%), and 4 (40.0%) of the patient, HCWs, and the environment, respectively. The proportion of lukfF-PV gene containing S. aureus isolated from patient samples was statistically significant. Four (40.0%) of the MRSA isolates also had the lukfF-PV gene. The identification of highly AMR and virulence factors from patients, HCWs and the environment is concerning. Further studies are needed to identify potential transmission links and improve infection prevention and control.

Tuber pathogenicity of Macrophomina phaseolina strain 3 a isolated from rotten cassava tuber from farm lands in Osogbo, Osun State, Nigeria, its virulence genes and ADMET properties.

BACKGROUND: Macrophomina phaseolina is a pathogen that causes an opportunistic disease that spreads by soil and seeds and affects more than 500 different plant species, like fruits, trees, and row crops. Mycotoxins, such as phaseolinic acid, and phaseolinone, are produced by M. phaseolina isolates in previous investigations; however, the production of these mycotoxins seems to vary depending on the host and the region. METHODS AND RESULTS: In this study, Macrophomina phaseolina strain 3 A was isolated from rotten cassava tuber and identified using the analysis of the sequences of the internal transcribed spacer region. The isolate was inoculated on a fresh healthy cassava tuber at 25 °C and tuber-rotting potential was monitored for 4 weeks. Virulence genes MPH_06603, MPH_06955, and MPH_01521 were determined with designed primers, and secondary metabolites were characterized by FTIR and GCMS. The rotten tuber effect was observed from the 2nd week of the experiment with severe tuber rot and weight reduction. The PCR showed the presence of MPH_06603 virulence gene. The GCMS showed N-Methylpivalamide (115.0 m/z), Butane, 1,4-dimethoxy- (119.0 m/z), and 5-Hydroxymethylfurfural (126.0 m/z) were the predominant metabolites produced by the pathogen. The compounds in the metabolites inhibit CYP3A4 enzymes, cause eye irritation, and Human Ether-a-go-go-related gene inhibition. CONCLUSION: This study revealed that M. phaseolina was responsible for the cassava tuber rot which leads to a lower yield of farm produce. The metabolites produced are toxic and unsafe for human consumption. It is suggested that farmers should destroy any cassava affected by this pathogen to prevent its toxic effects on humans and animals.

Molecular characterization, antimicrobial resistance and invasion of epithelial cells by Streptococcus agalactiae strains isolated from colonized pregnant women and newborns in Rio de Janeiro, Brazil.

AIMS: To evaluate the prevalence, molecular characteristics, antimicrobial susceptibility, and epithelial invasion of Streptococcus agalactiae strains isolated from pregnant women and newborns in Rio de Janeiro, Brazil. METHODS AND RESULTS: A total of 67 S. agalactiae isolates, 48 isolates from pregnant women and 19 from neonates, were analyzed. Capsular type Ia and V were predominant (35.8%/each). The multilocus sequence typing analysis revealed the presence of 19 STs grouped into 6 clonal complexes with prevalence of CC17/40.3% and CC23/34.3%. The lmb and iag virulence genes were found in 100% of isolates. Four S. agalactiae strains, belonging to CC17/ST1249 and CC23/ST23, were able to adhere to A549 respiratory epithelial cells. Antimicrobial resistance was verified mainly to tetracycline (85%), erythromycin (70.8%), and clindamycin (58.3%). Four S. agalactiae isolates were multidrug resistant. The resistance genes tested were found in 92.5% of isolates for tetM, 58.2% for ermB, 28.4% for mefAE, and 10.4% for tetO. CONCLUSION: The study showed a high prevalence of virulence and antimicrobial genes in S. agalactiae strains isolated from pregnant women and newborns, supporting the idea that continued surveillance is necessary to identify risk factors and perform long-term follow-up in pregnant women and neonates in Rio de Janeiro.

Whole-genome sequencing and pathogenicity analysis of Rhodococcus equi isolated in horses.

BACKGROUND: Rhodococcus equi (R. equi) is a Gram-positive zoonotic pathogen that frequently leads to illness and death in young horses (foals). This study presents the complete genome sequence of R. equi strain BJ13, which was isolated from a thoroughbred racehorse breeding farm in Beijing, China. RESULTS: The BJ13 genome has a length of 5.30 Mb and consists of a complete chromosome and a plasmid measuring 5.22 Mb and 0.08 Mb, respectively. We predicted 4,929 coding gene open reading frames, along with 52 tRNAs and 12 rRNAs. Through analysis of mobile genetic elements, we identified 6 gene islands and 1 prophage gene. Pathogenic system analysis predicted the presence of 418 virulence factors and 225 drug resistance genes. Secretion system analysis revealed the prediction of 297 secreted proteins and 1,106 transmembrane proteins. BJ13 exhibits genomic features, virulence-associated genes, potential drug resistance, and a virulence plasmid structure that may contribute to the evolution of its pathogenicity. Lastly, the pathogenicity of the isolated strain was assessed through animal experiments, which resulted in inflammatory reactions or damage in the lungs, liver, and spleen of mice. Moreover, by the 7th day post-infection, the mortality rate of the mice reached 50.0%, indicating complex immune regulatory mechanisms, including overexpression of IL-10 and increased production of pro-inflammatory cytokines like TNF-α. These findings validate the strong pathogenicity of the isolated strain and provide insights for studying the pathogenic mechanisms of Rhodococcus equi infection. CONCLUSIONS: The complete genome sequence of R. equi strain BJ13 provides valuable insights into its genomic characteristics, virulence potential, drug resistance, and secretion systems. The strong pathogenicity observed in animal experiments underscores the need for further investigation into the pathogenic mechanisms of R. equi infection.

Anti-virulence potential of iclaprim, a novel folic acid synthesis inhibitor, against Staphylococcus aureus.

Infections caused by Staphylococcus aureus pose a significant global public problem. Therefore, new antibiotics and therapeutic strategies are needed to combat this pathogen. This investigation delves into the effects of iclaprim, a newly discovered inhibitor of folic acid synthesis, on S. aureus virulence. The phenotypic and genotypic effects of iclaprim were thoroughly examined in relation to virulence factors, biofilm formation, and dispersal, as well as partial virulence-encoding genes associated with exoproteins, adherence, and regulation in S. aureus MW2, N315, and ATCC 25923. Then, the in vivo effectiveness of iclaprim on S. aureus pathogenicity was explored by a Galleria mellonella larvae infection model. The use of iclaprim at sub-inhibitory concentrations (sub-MICs) resulted in a reduction of α-hemolysin (Hla) production and a differential effect on the activity of coagulase in S. aureus strains. The results of biofilm formation and eradication assay showed that iclaprim was highly effective in depolymerizing the mature biofilm of S. aureus strains at concentrations of 1 MIC or greater, however, inhibited the biofilm-forming ability of only strains N315 and ATCC 25923 at sub-MICs. Interestingly, treatment of strains with sub-MICs of iclaprim resulted in significant stimulation or suppression of most virulence-encoding genes expression. Iclaprim did not affect the production of δ-hemolysin or staphylococcal protein A (SpA), nor did it impact the total activity of proteases, nucleases, and lipases. In vivo testing showed that sub-MICs of iclaprim significantly improves infected larvae survival. The present study offered valuable insights towards a better understating of the influence of iclaprim on different strains of S. aureus. The findings suggest that iclaprim may have potential as an anti-virulence and antibiofilm agent, thus potentially mitigating the pathogenicity of S. aureus and improving clinical outcomes associated with infections caused by this pathogen. KEY POINTS: • Iclaprim effectively inhibits α-hemolysin production and biofilm formation in a strain-dependent manner and was an excellent depolymerizing agent of mature biofilm • Iclaprim affected the mRNA expression of virulence-encoding genes associated with exoproteins, adherence, and regulation • In vivo study in G. mellonella larvae challenged with S. aureus exhibited that iclaprim improves larvae survival.

Genomic diversity, antibiotic resistance, and virulence in South African Enterococcus faecalis and Enterococcus lactis isolates.

This study presents the empirical findings of an in-depth genomic analysis of Enterococcus faecalis and Enterococcus lactis isolates from South Africa. It offers valuable insights into their genetic characteristics and their significant implications for public health. The study uncovers nuanced variations in the gene content of these isolates, despite their similar GC contents, providing a comprehensive view of the evolutionary diversity within the species. Genomic islands are identified, particularly in E. faecalis, emphasizing its propensity for horizontal gene transfer and genetic diversity, especially in terms of antibiotic resistance genes. Pangenome analysis reveals the existence of a core genome, accounting for a modest proportion of the total genes, with 2157 core genes, 1164 shell genes, and 4638 cloud genes out of 7959 genes in 52 South African E. faecalis genomes (2 from this study, 49 south Africa genomes downloaded from NCBI, and E. faecalis reference genome). Detecting large-scale genomic rearrangements, including chromosomal inversions, underscores the dynamic nature of bacterial genomes and their role in generating genetic diversity. The study uncovers an array of antibiotic resistance genes, with trimethoprim, tetracycline, glycopeptide, and multidrug resistance genes prevalent, raising concerns about the effectiveness of antibiotic treatment. Virulence gene profiling unveils a diverse repertoire of factors contributing to pathogenicity, encompassing adhesion, biofilm formation, stress resistance, and tissue damage. These empirical findings provide indispensable insights into these bacteria's genomic dynamics, antibiotic resistance mechanisms, and virulence potential, underlining the pressing need to address antibiotic resistance and implement robust control measures.

Salmonella Typhimurium exploits host polyamines for assembly of the type 3 secretion machinery.

Bacterial pathogens utilize the factors of their hosts to infect them, but which factors they exploit remain poorly defined. Here, we show that a pathogenic Salmonella enterica serovar Typhimurium (STm) exploits host polyamines for the functional expression of virulence factors. An STm mutant strain lacking principal genes required for polyamine synthesis and transport exhibited impaired infectivity in mice. A polyamine uptake-impaired strain of STm was unable to inject effectors of the type 3 secretion system into host cells due to a failure of needle assembly. STm infection stimulated host polyamine production by increasing arginase expression. The decline in polyamine levels caused by difluoromethylornithine, which inhibits host polyamine production, attenuated STm colonization, whereas polyamine supplementation augmented STm pathogenesis. Our work reveals that host polyamines are a key factor promoting STm infection, and therefore a promising therapeutic target for bacterial infection.

Potential antivirulence and antibiofilm activities of sub-MIC of oxacillin against MDR S. aureus isolates: an in-vitro and in-vivo study.

BACKGROUND: Multi-drug resistant Staphylococcus aureus is one of the most common causes of nosocomial and community-acquired infections, with high morbidity and mortality. Treatment of such infections is particularly problematic; hence, it is complicated by antibiotic resistance, and there is currently no reliable vaccine. Furthermore, it is well known that S. aureus produces an exceptionally large number of virulence factors that worsen infection. Consequently, the urgent need for anti-virulent agents that inhibit biofilm formation and virulence factors has gained momentum. Therefore, we focused our attention on an already-approved antibiotic and explored whether changing the dosage would still result in the intended anti-virulence effect. METHODS: In the present study, we determined the antibiotic resistance patterns and the MICs of oxacillin against 70 MDR S. aureus isolates. We also investigated the effect of sub-MICs of oxacillin (at 1/4 and 1/8 MICs) on biofilm formation using the crystal violet assay, the phenol-sulphuric acid method, and confocal laser scanning microscopy (CLSM). We examined the effect of sub-MICs on virulence factors and bacterial morphology using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and electron microscopy, respectively. Moreover, we studied the effect of sub-MICs of oxacillin (OX) in-vivo using a wound infection model. RESULTS: Oxacillin at 1/2 MIC showed a significant decrease in bacterial viability, while 1/4 and 1/8 MICs had negligible effects on treated bacterial isolates. Treatment of MDR isolates with 1/4 or 1/8 MICs of oxacillin significantly reduced biofilm formation (64% and 40%, respectively). The treated MDR S. aureus with sub-MICs of OX exhibited a dramatic reduction in several virulence factors, including protease, hemolysin, coagulase, and toxic shock syndrome toxin-1 (TSST-1) production. The sub-MICs of OX significantly decreased (P < 0.05) the gene expression of biofilm and virulence-associated genes such as agrA, icaA, coa, and tst. Furthermore, oxacillin at sub-MICs dramatically accelerated wound healing, according to the recorded scoring of histological parameters. CONCLUSION: The treatment of MDR S. aureus with sub-MICs of oxacillin can help in combating the bacterial resistance and may be considered a promising approach to attenuating the severity of S. aureus infections due to the unique anti-biofilm and anti-virulence activities.

The Secreted Aminopeptidase of Pseudomonas aeruginosa (PaAP).

Pseudomonas aeruginosa is an opportunistic pathogen that causes severe infections in compromised hosts. P. aeruginosa infections are difficult to treat because of the inherent ability of the bacteria to develop antibiotic resistance, secrete a variety of virulence factors, and form biofilms. The secreted aminopeptidase (PaAP) is an emerging virulence factor, key in providing essential low molecular weight nutrients and a cardinal modulator of biofilm development. PaAP is therefore a new potential target for therapy of P. aeruginosa infections. The present review summarizes the current knowledge of PaAP, with special emphasis on its biochemical and enzymatic properties, activation mechanism, biological roles, regulation, and structure. Recently developed specific inhibitors and their potential as adjuncts in the treatment of P. aeruginosa infections are also described.

Occurrence, serotypes and virulence characteristics of Shiga toxin-producing and Enteropathogenic Escherichia coli isolates from dairy cattle in South Africa.

Shiga toxin-producing and Enteropathogenic Escherichia coli are foodborne pathogens commonly associated with diarrheal disease in humans. This study investigated the presence of STEC and EPEC in 771 dairy cattle fecal samples which were collected from 5 abattoirs and 9 dairy farms in South Africa. STEC and EPEC were detected, isolated and identified using culture and PCR. Furthermore, 339 STEC and 136 EPEC isolates were characterized by serotype and major virulence genes including stx1, stx2, eaeA and hlyA and the presence of eaeA and bfpA in EPEC. PCR screening of bacterial sweeps which were grown from fecal samples revealed that 42.2% and 23.3% were STEC and EPEC positive, respectively. PCR serotyping of 339 STEC and 136 EPEC isolates revealed 53 different STEC and 19 EPEC serotypes, respectively. The three most frequent STEC serotypes were O82:H8, OgX18:H2, and O157:H7. Only 10% of the isolates were classified as "Top 7" STEC serotypes: O26:H2, 0.3%; O26:H11, 3.2%; O103:H8, 0.6%; and O157:H7, 5.9%. The three most frequent EPEC serotypes were O10:H2, OgN9:H28, and O26:H11. The distribution of major virulence genes among the 339 STEC isolates was as follows: stx1, 72.9%; stx2, 85.7%; eaeA, 13.6% and hlyA, 69.9%. All the 136 EPEC isolates were eaeA-positive but bfpA-negative, while 46.5% carried hlyA. This study revealed that dairy cattle are a major reservoir of STEC and EPEC in South Africa. Further comparative studies of cattle and human STEC and EPEC isolates will be needed to determine the role played by dairy cattle STEC and EPEC in the occurrence of foodborne disease in humans.Please kindly check and confirm the country and city name in affiliation [6].This affiliation is correct.Please kindly check and confirm the affiliationsConfirmed. All Affiliations are accurate.

Virulence gene distribution and molecular epidemiological characteristics of carbapenem-resistant Klebsiella pneumoniae in the ICU. / ICUç¢³é’éœ‰çƒ¯è€è¯è‚ºç‚Žå ‹é›·ä¼¯èŒçš„æ¯’åŠ›åŸºå› åˆ†å¸ƒåŠåˆ†å­æµè¡Œç— å­¦ç‰¹å¾.

OBJECTIVES: The drug-resistant genes carried by carbapenem-resistant Klebsiella pneumoniae (CRKP) limit clinical treatment options, and its virulence genes severely affect patient prognosis. This study aims to investigate the distribution of virulence genes, capsular serotypes, and molecular epidemiological characteristics of CRKP in ICU, to understand the characteristics of CRKP infections in ICU, and to provide a scientific basis for effective monitoring and control of CRKP infections in ICU. METHODS: A total of 40 non-duplicate strains of CRKP isolated from the ICU of Guangdong Provincial People's Hospital between January 2021 and December 2022 were collected and analyzed. Whole-genome sequencing was used to analyze the distribution of resistance genes, virulence genes, and capsular serotypes of the strains. The sequences of 7 housekeeping genes of CRKP genome were uploaded to the Klebsiella pneumoniae (KPN)multilocus sequence typing (MLST) database to determine the sequence types (STs) of the strains. RESULTS: The age of the 40 ICU CRKP-infected patients was (69.03±17.82) years old, with various underlying diseases, and there were 20 patients with improved clinical outcome and 20 patients with death. The isolated strains primarily originated from mid-stream urine and bronchoalveolar lavage fluid. Whole-genome sequencing results revealed that the strains predominantly carried blaKPC-1 (29 strains, 72.5%) and blaNDM-1 (6 strains, 15.0%), with 5 strains carrying both blaKPC-1 and blaNDM-1. Various virulence genes were detected, among which the carriage rates of genes such as entA, entB, entE, entS, fepA, fepC, fepG, yag/ecp, and ompA reached 100%, while the carriage rates of genes such as entD, fimB, iroB, iroD, fes,and pla were low. The CRKP strains isolated from ICU were predominantly ST11 (27 cases, 67.5%), with KL64 being the main capsular serotype (29 cases, 72.5%). A total of 23 ST11-KL64 CRKP strains were detected, accounting for 57.5%. CONCLUSIONS: The main type of ICU CRKP is ST11-KL64, carrying various virulence genes, primarily those related to iron absorption. Furthermore, blaKPC has shifted from blaKPC-2 to blaKPC-1. Therefore, close monitoring of the molecular epidemiological changes of CRKP is necessary, and strict control measures should be implemented to effectively curb the occurrence of CRKP infections.

Retrospective analysis of molecular characteristics, risk factors, and outcomes in carbapenem-resistant Klebsiella pneumoniae bloodstream infections.

BACKGROUND: Klebsiella pneumoniae (KP) is the second most prevalent Gram-negative bacterium causing bloodstream infections (BSIs). In recent years, the management of BSIs caused by KP has become increasingly complex due to the emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP). Although numerous studies have explored the risk factors for the development of CRKP-BSIs, the mortality of patients with KP-BSIs, and the molecular epidemiological characteristics of CRKP, the variability in data across different populations, countries, and hospitals has led to inconsistent conclusions. In this single-center retrospective observational study, we utilized logistic regression analyses to identify independent risk factors for CRKP-BSIs and factors associated with mortality in KP-BSI patients. Furthermore, a risk factor-based prediction model was developed. CRKP isolates underwent whole-genome sequencing (WGS), followed by an evaluation of microbiological characteristics, including antimicrobial resistance and virulence genes, as well as epidemiological characteristics and phylogenetic analysis. RESULTS: Our study included a total of 134 patients with KP-BSIs, comprising 50 individuals infected with CRKP and 84 with carbapenem-susceptible Klebsiella pneumoniae (CSKP). The independent risk factors for CRKP-BSIs were identified as gastric catheterization (OR = 9.143; CI = 1.357-61.618; P = 0.023), prior ICU hospitalization (OR = 4.642; CI = 1.312-16.422; P = 0.017), and detection of CRKP in non-blood sites (OR = 8.112; CI = 2.130-30.894; P = 0.002). Multivariate analysis revealed that microbiologic eradication after 6 days (OR = 3.569; CI = 1.119-11.387; P = 0.032), high Pitt bacteremia score (OR = 1.609; CI = 1.226-2.111; P = 0.001), and inappropriate empirical treatment after BSIs (OR = 6.756; CI = 1.922-23.753; P = 0.003) were independent risk factors for the 28-day mortality in KP-BSIs. The prediction model confirmed that microbiologic eradication after 6.5 days and a Pitt bacteremia score of 4.5 or higher were significant predictors of the 28-day mortality. Bioinformatics analysis identified ST11 as the predominant CRKP sequence type, with blaKPC-2 as the most prevalent gene variant. CRKP stains carried multiple plasmid-mediated resistance genes along with some virulence genes. Phylogenetic analysis indicated the presence of nosocomial transmission of ST11 CRKP within the ICU. CONCLUSIONS: The analysis of risk factors for developing CRKP-BSIs and the association between KP-BSIs and 28-day mortality, along with the development of a risk factor-based prediction model and the characterization of CRKP strains, enhances clinicians' understanding of the pathogens responsible for BSIs. This understanding may help in the timely administration of antibiotic therapy for patients with suspected KP-BSIs, potentially improving outcomes.

Molecular epidemiology of string test-positive Klebsiella pneumoniae isolates in Huzhou, China, 2020-2023.

Objective: This study used whole-genome sequencing (WGS) to explore the genetic diversity, virulence factors, and antimicrobial resistance determinants of string test-positive Klebsiella pneumoniae (KP) over a 4-year surveillance period in Huzhou, China. Methods: In total, 632 clinical isolates were collected via hospital surveillance from 2020 to 2023; 100 were positive in the string test and these 100 strains were subjected to antimicrobial susceptibility testing using an agar dilution method followed by WGS. Results: The resistance rates to cefotaxime (77.0%), trimethoprim-sulfamethoxazole (67.0%), and nalidixic acid (64.0%) were high. Multilocus sequence typing revealed high genetic diversity; there were 33 sequence types (STs) and 15 capsular serotypes. The most common ST was ST23 (16.0%) and the most common capsular serotype was K1 (22.5%). Virulome analysis revealed among-strain differences in virulence factors that affected bacterial adherence, efflux pump action, iron uptake, nutritional factors, metabolic regulation, the secretion system, and toxin production. The Kleborate strain-specific virulence scores of all 100 string test-positive KPs were derived: 28 strains scored 5, 28 scored 4, 21 scored 3, 12 scored 1, and 11 scored 0. All 77 strains with scores of 3 to 5 contained the iucA gene. The phylogeny based on whole-genome single nucleotide polymorphisms (wgSNPs) indicated high clonality; the string test-positive KP strains were grouped into six clades. Closely related isolates in each genetic cluster usually shared STs. Conclusion: The present study highlights the significance of the KP iucA gene in terms of hypervirulence and the diverse genotypes of string test-positive KP strains isolated in Huzhou hospitals.