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Background: To develop efficient selection strategies and improve the discovery of promising ligands, it is highly desirable to analyze the sequence composition of naïve phage display libraries and monitor the evolution of their peptide content during successive rounds of amplification. In the current study, we performed a comparative analysis of the compositional features in different lots of the same naïve phage display library and monitored alterations in their peptide compositions during three rounds of amplification. Methods: We conducted three rounds of duplicate serial amplification of two different lots of the Ph.D.™-12 phage display library. DNA from the samples was subjected to Next-Generation Sequencing (NGS) using an Illumina platform. The NGS datasets underwent a variety of bioinformatic analyses using Python and MATLAB scripts. Results: We observed substantial heterogeneity in the sequence composition of the two lots indicated by differences in the enhanced percentage of wildtype clones, reduced diversity (number of unique sequences), and increased enrichment factors (EFs) during amplification as well as by observing no common sequence between lots and decreased number of common sequences between the naïve library and the consecutive rounds of amplification for each lot. We also found potential propagation-related target-unrelated peptides (TUPs) with the highest EFs in the two lots, which were displayed by the fastest-propagating phage clones. Furthermore, motif analysis of the most enriched subpopulation of amplified libraries led to the identification of some motifs hypothesized to contribute to the increased amplification rates of the respective phage clones. Conclusion: Our results highlight tremendous heterogeneity in the peptide composition of different lots of the same type of naïve phage display library, and the divergent evolution of their compositional features during amplification rounds at the amino acid, peptide, and motif levels. Our findings can be instrumental for phage display researchers by bringing fundamental insights into the vast extent of non-uniformity between phage display libraries and by providing a clear picture of how these discrepancies can lead to different evolutionary fates for the peptide composition of phage pools, which can have profound impacts on the outcome of phage display selections through biopanning. [ABSTRACT FROM AUTHOR]

Introduction: Phage WO represents the sole bacteriophage identified to infect Wolbachia , exerting a range of impacts on the ecological dynamics and evolutionary trajectories of its host. Given the extensive prevalence of Wolbachia across various species, phage WO is likely among the most prolific phage lineages within arthropod populations. To examine the diversity and evolutionary dynamics of phage WO, we conducted a screening for the presence of phage WO in Wolbachia -infected cricket species from China. Methods: The presence of phage WO was detected using a PCR-based methodology. To elucidate the evolutionary forces driving phage WO diversity, analyses of intragenic recombination were conducted employing established recombination techniques, and horizontal transmission was investigated through comparative phylogenetic analysis of the phages and their hosts. Results and discussion: Out of 19 cricket species infected with Wolbachia , 18 species were found to harbor phage WO. Notably, 13 of these 18 cricket species hosted multiple phage types, with the number of types ranging from two to 10, while the remaining five species harbored a single phage type. Twelve horizontal transmission events of phage WO were identified, wherein common phage WO types were shared among different Wolbachia strains. Notably, each phage WO horizontal transfer event was associated with distinct Wolbachia supergroups, specifically supergroups A, B, and F. Previous studies have found that four Wolbachia strains infect two to five species of crickets. However, among these cricket species, in addition to the shared phage WO types, all harbored species-specific phage WO types. This suggests that Wolbachia in crickets may acquire phage WO types through horizontal viral transfer between eukaryotes, independent of Wolbachia involvement. Furthermore, nine putative recombination events were identified across seven cricket species harboring multiple phage types. These findings suggest that horizontal transmission and intragenic recombination have played a significant role in the evolution of the phage WO genome, effectively enhancing the diversity of phage WO associated with crickets. [ABSTRACT FROM AUTHOR]

Background: Bacteriophages are neutralized by the sera of patients undergoing phage therapy (PT), particularly during local or concomitant local and oral phage administration in bone infections, soft tissue infections, or upper respiratory tract infections. Methods: The antiphage activity of the sera (AAS) level of 27 patients with bacterial infections such as bone infections, soft tissue infections, or upper respiratory tract infections undergoing PT was performed using the plate phage neutralization test. Results: Our preliminary results suggest that high levels of antiphage antibodies appear late in the treatment period, at the earliest in the 3rd–8th week of PT. Patients with bone infections treated locally with the S. aureus phage Staph_1N and patients with upper respiratory tract infections administered locally and orally with the S. aureus phage Staph_A5L had elevated levels of antiphage antibodies in sera during PT. In parallel to these results, it has been shown that a strong antiphage humoral response does not prevent a positive outcome of phage treatment. Conclusions: The earliest time point at which high levels of antiphage antibodies in sera appear during local and oral PT is day 21 of therapy. We noticed that the high level of antiphage antibodies in sera occurring during local or both local and oral PT was correlated with the type of infection and phage type. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is a major global threat to human health, and phage therapy has emerged as a promising strategy for treating infections caused by multidrug-resistant pathogens. In this study, we isolated and characterized a Pseudomonas lytic phage, PaTJ, from wastewater. PaTJ belongs to the phage family Mesyanzhinovviridae, and is featured by short latency (30 min) and large burst size (103 PFU per infected cell). Our investigation revealed that PaTJ utilizes the type IV Pili (T4P) as a receptor. Transcriptome analysis of PaTJ infected host at latent stage showed distinct expression patterns of PaTJ encoding genes involved in replication and structure assembly, without expression of the majority of toxic accessory genes responsible for phage release. In addition, host bacteria exhibited specific induction of host metabolism-related genes in response to the PaTJ's infection. Furthermore, our findings demonstrated the PaTJ's potential in degrading biofilms. This work sheds light on the multifaceted impact of this lytic phage PaTJ on P. aeruginosa, presenting potential applications in both gene expression modulation and biofilm management. [ABSTRACT FROM AUTHOR]

We isolated and characterized the novel polyvalent T-even type bacteriophage vB_SenS_Jbel from wastewater using an enrichment of three different Salmonella strains. The vB_SenS_Jbel virions have prolate icosahedral capsids approximately 100 nm long and 80 nm wide. The genome consists of linear, double-stranded DNA that is 165,566 bp long. Analysis of the genome and structure of vB_SenS_Jbel indicates that it belongs to the genus Tequatrovirus of the family Straboviridae. This novel polyvalent phage can infect Escherichia coli and multiple Salmonella and Shigella species through its unique tail fiber structure. [ABSTRACT FROM AUTHOR]

Fast, accurate, and affordable bacterial identification methods are paramount for the timely treatment of infections, especially in resource‐limited settings (RLS). However, today, only 1.3% of the sub‐Saharan African diagnostic laboratories are performing clinical bacteriology. To improve this, diagnostic tools for RLS should prioritize simplicity, affordability, and ease of maintenance, as opposed to the costly equipment utilized for bacterial identification in high‐income countries, such as matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS). In this work, we present a new high‐throughput approach based on a simple wide‐field (864 mm2) lensless imaging system allowing for the acquisition of a large portion of a Petri dish coupled with a supervised deep learning algorithm for identification at the bacterial colony scale. This wide‐field imaging system is particularly well suited to RLS since it includes neither moving mechanical parts nor optics. We validated this approach through the acquisition and the subsequent analysis of a dataset comprising 252 clinical isolates from five species, encompassing some of the most prevalent pathogens. The resulting optical morphotypes exhibited intra‐ and interspecies variability, a scenario considerably more akin to real‐world clinical practice than the one achievable by solely concentrating on reference strains. Despite this variability, high identification performance was achieved with a correct species identification rate of 91.7%. These results open up some new prospects for identification in RLS. We released both the acquired dataset and the trained identification algorithm in publicly available repositories. [ABSTRACT FROM AUTHOR]

Background: Periprosthetic Joint Infection (PJI) following hip and knee arthroplasty is a catastrophic complication in orthopaedic surgery. It has long been a key focus for orthopaedic surgeons in terms of prevention and management. With the increasing incidence of antibiotic resistance in recent years, finding more targeted treatment methods has become an increasingly urgent issue. Bacteriophage Therapy (BT) has emerged as a promising adjunctive treatment for bone and joint infections in recent years. It not only effectively kills bacteria but also demonstrates significant anti-biofilm activity, garnering substantial clinical interest due to its demonstrated efficacy and relatively low incidence of adverse effects. Purpose: This review aims to systematically evaluate the efficacy and safety of bacteriophage therapy in treating PJI following hip and knee arthroplasty, providing additional reference for its future clinical application. Methods: Following predefined inclusion and exclusion criteria, our team conducted a systematic literature search across seven databases (PubMed, Embase, Web of Science, Cochrane Library, ClinicalTrials.gov, CNKI, and WanFang Database). The search was conducted up to May 2024 and included multiple clinical studies on the use of bacteriophage therapy for treating PJI after hip and knee arthroplasty to assess its efficacy and safety. Results: This systematic review included 16 clinical studies after screening, consisting of 15 case reports and one prospective controlled clinical trial, involving a total of 42 patients with PJI treated with bacteriophage therapy. The average patient age was 62.86 years, and 43 joints were treated, with patients undergoing an average of 5.25 surgeries. The most common pathogen in these infections was Staphylococcus aureus, accounting for 18 cases. 33 patients received cocktail therapy, while nine were treated with a single bacteriophage preparation. Additionally, all patients underwent suppressive antibiotic therapy (SAT) postoperatively. All patients were followed up for an average of 13.55 months. There were two cases of recurrence, one of which resulted in amputation one year postoperatively. The remaining patients showed good recovery outcomes. Overall, the results from the included studies indicate that bacteriophage therapy effectively eradicates infectious strains in various cases of PJI, with minimal side effects, demonstrating promising clinical efficacy. Conclusion: In the treatment of PJI following hip and knee arthroplasty, bacteriophages, whether used alone or in combination as cocktail therapy, have shown therapeutic potential. However, thorough preoperative evaluation is essential, and appropriate bacteriophage types and treatment regimens must be selected based on bacteriological evidence. Future large-scale, randomized controlled, and prospective trials are necessary to validate the efficacy and safety of this therapy. [ABSTRACT FROM AUTHOR]

Background: The challenges in culturing Treponema pallidum have hindered molecular-biological analysis. This study aims to establish a molecular epidemiological analysis of syphilis among Japanese men who have sex with men (MSM) and to investigate the relationship between bacteremia and associated pathophysiology. Methods: We used whole blood specimens from syphilis-diagnosed individuals in Tokyo, collected between February 2019 and June 2022. All individuals were MSM, and most were people with HIV (97.2%). We used a multi-locus sequence typing (MLST) scheme for epidemiological analysis. Sequences for MLST (TP0136, TP0548, and TP0705) were obtained. Results: Out of 71 whole blood samples, 26 samples (36.6%) were positive for TP0136, and we sequenced three loci for MLST in 22 samples (31.0%). The most frequently detected sequence type (ST) was ST3 (n = 9), followed by ST6 (n = 6). Phylogenetic analysis revealed that 12 samples belonged to the SS14-like group (60%), and 8 samples belonged to the Nichols-like group (40%). Treponema pallidum subsp. endemicum (TEN), the cause of bejel was detected in three samples (12%). There was a significant association between TP0136 detection rate and C- reactive protein (CRP) (77.0% at a cut-off:0.5 mg/dL). Conclusion: Both SS14-like and Nichols-like strains were circulating concurrently, and TEN could have been sexually transmitted among MSM with HIV. Elevated CRP may indicate the presence of the pathogen in the blood. [ABSTRACT FROM AUTHOR]

Phage therapy is gaining increasing interest in the fight against critically antibiotic-resistant nosocomial pathogens. However, the narrow host range of bacteriophages hampers the development of broadly effective phage therapeutics and demands precision approaches. Here, we combine large-scale phylogeographic analysis with high-throughput phage typing to guide the development of precision phage cocktails targeting carbapenem-resistant Acinetobacter baumannii , a top-priority pathogen. Our analysis reveals that a few strain types dominate infections in each world region, with their geographical distribution remaining stable within 6 years. As we demonstrate in Eastern Europe, this spatiotemporal distribution enables preemptive preparation of region-specific phage collections that target most local infections. Finally, we showcase the efficacy of phage cocktails against prevalent strain types using in vitro and animal infection models. Ultimately, genomic surveillance identifies patients benefiting from the same phages across geographical scales, thus providing a scalable framework for precision phage therapy. [Display omitted] • A few CRAB types dominate infections in individual world regions • Phylogeography reveals stable strain composition for countries over 6 years • Spatiotemporal distribution informs the development of region-specific phage collections • Phage cocktails are effective against the most prevalent strain types in Eastern Europe By mapping the genetic diversity and geographic distribution of carbapenem-resistant Acinetobacter baumannii , bacteriophage cocktails were developed and designed to treat the maximum number of patients within a specific geographic region. [ABSTRACT FROM AUTHOR]

The food industry has incurred substantial losses from contamination by Pseudomonas fluorescens, emphasizing the critical importance of implementing effective control strategies. Phages are potential sterilizers due to their specific killing abilities and the difficulty bacteria face in developing resistance. However, a significant barrier to their development is the lack of diversity among phage types. In this study, we characterized a novel lytic P. fluorescens phage, named vB_PF_Y1-MI. Phage vB_PF_Y1-MI displayed a latent period of nearly 10 min and a high burst size of 1493 PFU/cell. This phage showed good activity over a wide range of temperature (up to 70 °C) and pH (3–12). The genome of phage vB_PF_Y1-MI spans 93,233 bp with a GC content of 45%. It encompasses 174 open-reading frames and 19 tRNA genes, while no lysogeny or virulence-associated genes were detected. Phylogenetic analysis positions it as a novel unassigned evolutionary lineage within the Caudoviricetes class among related dsDNA phages. Our study provides foundational insights into vB_PF_Y1-MI and emphasizes its potential as an effective biological control agent against P. fluorescens. This research offers crucial theoretical groundwork and technical support for subsequent efforts in preventing and controlling P. fluorescens contamination. [ABSTRACT FROM AUTHOR]

Acinetobacter baumannii (A. baumannii) poses a serious public health challenge due to its notorious antimicrobial resistance, particularly carbapenem-resistant A. baumannii (CRAB). In this study, we isolated a virulent phage, named P1068, from medical wastewater capable of lysing CRAB, primarily targeting the K3 capsule type. Basic characterization showed that P1068 infected the A. baumannii ZWAb014 with an optimal MOI of 1, experienced a latent period of 10 min and maintained stability over a temperature range of 4-37 °C and pH range of 3-10. Phylogenetic and average nucleotide identity analyses indicate that P1068 can be classified as a novel species within the genus Obolenskvirus of the Caudoviricetes class as per the most recent virus classification released by the International Committee on Taxonomy of Viruses (ICTV). Additionally, according to classical morphological classification, P1068 is identified as a T4-like phage (Myoviridae). Interestingly, we found that the tail fiber protein (TFP) of P1068 shares 74% coverage and 88.99% identity with the TFP of a T7-like phage (Podoviridae), AbKT21phiIII (NC_048142.1). This finding suggests that the TFP gene of phages may undergo horizontal transfer across different genera and morphologies. In vitro antimicrobial assays showed that P1068 exhibited antimicrobial activity against A. baumannii in both biofilm and planktonic states. In mouse models of intraperitoneal infection, P1068 phage protected mice from A. baumannii infection and significantly reduced bacterial loads in various tissues such as the brain, blood, lung, spleen, and liver compared to controls. In conclusion, this study demonstrates that phage P1068 might be a potential candidate for the treatment of carbapenem-resistant and biofilmforming A. baumannii infections, and expands the understanding of horizontal transfer of phage TFP genes. [ABSTRACT FROM AUTHOR]

AbstractAntimicrobial resistance (AMR) has been recognized as an important health crisis in the twenty first century. Type IV secretion systems (T4SSs) play key roles in the dissemination of AMR plasmids. Novel strategies that combat AMR problem by targeting T4SS sprung up in recent years. Here, we focus on the strategy of male-specific phages that could target and kill bacteria carrying conjugative AMR plasmids encoding T4SSs. We reviewed the recent advances in male-specific phages, including anti-conjugation mechanisms, clinical isolation and identification methods, classification and characteristics, in vitro and in vivo anti-conjugation efficacy and improving strategies. Male-specific phages constitute exciting candidates for developing sustainable anti-resistance biocontrol applications. [ABSTRACT FROM AUTHOR]

Background: Streptococcus mutans is studied for its acidogenic and aciduric characteristics, notably its biofilm formation in the presence of sucrose, toward its role in the caries process. Variations in both genotype and phenotype have been reported among clinical isolates of S. mutans. This study aimed to examine genotypic and phenotypic characteristics of S. mutans obtained from Thai children with varying caries statuses. Methods: We determined the presence of S. mutans and caries status in 395 children aged 3–4 years. From 325 children carrying S. mutans, we selected 90 with different caries statuses—caries-free (CF; n = 30), low severity of caries (LC; n = 30), or high severity of caries (HC; n = 30). Three isolates of S. mutans were taken from each child, thus, a total of 270 isolates were obtained. Multilocus sequence typing (MLST) was used to genotype the isolates and assess their clonal relationships. The properties, including biofilm formation, collagen binding, and acid production and tolerance were also evaluated. Results: Children with carious lesions showed a higher detection rate and number of S. mutans in saliva than those without caries. S. mutans from individuals with HC status showed the lowest biofilm formation ability, while this group had the highest detection rate of collagen-binding isolates. There was no difference in acid production or tolerance by caries status. Genotyping by MLST did not reveal any clone of S. mutans specific to CF status. This result remained even when we included MLST data from the open-access PubMLST database. MLST did identify clones containing only strains from caries-affected hosts, but tests of their phenotypic properties did not reveal any differences between S. mutans from these clones and clones that were from both caries-free and caries-affected children. Conclusions: The clonal relationships of S. mutans indicated by MLST were not associated with the status of dental caries in the host. [ABSTRACT FROM AUTHOR]

The rise of carbapenem-resistant Klebsiella pneumoniae (CRKP) presents a significant global challenge in clinical and healthcare settings, severely limiting treatment options. This study aimed to utilize a bacteriophage as an alternative therapy against carbapenem-resistant K. pneumoniae. A novel lytic N4-like Klebsiella phage, vB_kpnP_KPYAP-1 (KPYAP-1), was isolated from sewage. It demonstrated efficacy against the K62 serotype polysaccharide capsule of blaOXA-48-producing K. pneumoniae. KPYAP-1 forms small, clear plaques, has a latent period of 20 min, and reaches a growth plateau at 35 min, with a burst size of 473 plaque-forming units (PFUs) per infected cell. Phylogenetic analysis places KPYAP-1 in the Schitoviridae family, Enquatrovirinae subfamily, and Kaypoctavirus genus. KPYAP-1 employs an N4-like direct terminal repeat mechanism for genome packaging and encodes a large virion-encapsulated RNA polymerase. It lacks integrase or repressor genes, antibiotic resistance genes, bacterial virulence factors, and toxins, ensuring its safety for therapeutic use. Comparative genome analysis revealed that the KPYAP-1 genome is most similar to the KP8 genome, yet differs in tail fiber protein, indicating variations in host recognition. In a zebrafish infection model, KPYAP-1 significantly improved the survival rate of infected fish by 92% at a multiplicity of infection (MOI) of 10, demonstrating its potential for in vivo treatment. These results highlight KPYAP-1 as a promising candidate for developing phage-based therapies targeting carbapenemase-producing K. pneumoniae. [ABSTRACT FROM AUTHOR]

Shiga toxin (Stx)-producing Escherichia coli (STEC) causes serious gastrointestinal illness, including hemorrhagic colitis and hemolytic uremic syndrome. Two types of Stxs (Stx1 and Stx2) are known and both are encoded by bacteriophages (Stx phages), but the production of Stx2 is known to be a major risk factor for severe STEC infections. The production of Stx2, but not Stx1, is tightly coupled with the induction of Stx phages, and Stx2 production levels vary between STEC strains even within the same serotype. Here, we analyzed the genomic diversity of all Stx phages in 71 strains representing the entire O145:H28 lineage, one of the often highly pathogenic STECs, and the relationship between the variations in Stx phage genomes and the levels of Stx2 production by host strains. Our analysis reveals highly dynamic natures of Stx phages in O145:H28, including the independent acquisition of similar Stx phages by different sublineages, the recent transfer of Stx phage between different sublineages, and the frequent gain and loss of Stx phages in some sublineages. We also show the association of the Stx2 phage types with the Stx2 production levels of host strains: strains carrying short-tailed Stx2 phages exhibited significantly higher Stx2 production levels than those carrying long-tailed Stx2 phages. Detailed analyses of the Stx2 phage genomes revealed that both of short- and long-tailed phages exhibited sequence diversification and they were divided into two groups, respectively, based on the sequence similarity of the phage early region encoding genes responsible for phage induction, short-tailed phages contained early regions clearly different in genetic organization from those in long-tailed phages. Therefore, the variations in the early regions between short-and long-tailed Stx2 phages appeared to be linked to a striking difference in Stx2 production levels in their host strains. These results broaden our understanding of the diversification and dynamism of Stx phages in O145:H28 and the association of Stx2 phage types with the Stx2 production level in this STEC lineage. [ABSTRACT FROM AUTHOR]

Aim: This study aimed to evaluate the expression of tcdA, tcdB, and binary toxin genes (cdtA and cdtB) by Real-Time PCR and molecular typing of Clostridioides difficile isolated from patient diarrhea samples from Hamadan Hospitals, west of Iran. Background: The concentration of C. difficile toxins (CDTs) is associated with the severity of the disease and the mortality rate. Measuring CDT levels could provide a reliable and objective means of determining the severity of C. difficile infection (CDI). Methods: From November 2018 to September 2019, 130 diarrhea samples were collected from hospitalized patients in three hospitals in Hamadan. C. difficle isolates were detected by culture and PCR. The presence of the genes encoding the toxin was identified by PCR, whereas the measurement of toxin expression was conducted using a relative Real-Time PCR technique. Genetic linkage of the isolates was also assessed by Ribotyping and Repetitive Extragenic Palindromic (rep-PCR) methods. Results: Among 130 diarrhea samples, 16 (12.3%) were positive for C. difficile. Genes encoding cdtA and tcdB were detected in all isolates, and 8 (50%) and 6 (37.5%) isolates were positive for the cdtA and cdtB genes. Real-time PCR results showed different expression levels of the toxin genes. A significant increase in the expression of the tcdA gene was observed compared with the control strain (P<0.05). Besides, more expression of cdtA gene was observed in the strains compared with cdtB gene. Ribotyping and rep-PCR results showed high genetic diversity of C. difficile among hospitals investigated. Conclusion: We encountered toxigenic C. difficile strains with various toxin expression levels, ribotypes, and rep types based on the findings of this study. This indicated that various clones from various sources circulate in the hospitals and among patients. [ABSTRACT FROM AUTHOR]

Bacteriophages (phages) represent a unique category of viruses with a remarkable ability to selectively infect host bacteria, characterized by their assembly from proteins and nucleic acids. Leveraging their exceptional biological properties and modifiable characteristics, phages emerge as innovative, safe, and efficient delivery vectors. The potential drawbacks associated with conventional nanocarriers in the realms of drug and gene delivery include a lack of cell-specific targeting, cytotoxicity, and diminished in vivo transfection efficiency. In contrast, engineered phages, when employed as cargo delivery vectors, hold the promise to surmount these limitations and attain enhanced delivery efficacy. This review comprehensively outlines current strategies for the engineering of phages, delineates the principal types of phages utilized as nanocarriers in drug and gene delivery, and explores the application of phage-based delivery systems in disease therapy. Additionally, an incisive analysis is provided, critically examining the challenges confronted by phage-based delivery systems within the domain of nanotechnology. The primary objective of this article is to furnish a theoretical reference that contributes to the reasoned design and development of potent phage-based delivery systems. [ABSTRACT FROM AUTHOR]

Background: Enteric fever is caused by Salmonella enterica serovars Typhi (S. Typhi) and Paratyphi A, B, and C. It continues to be a significant cause of morbidity and mortality worldwide. In highly endemic areas, children are disproportionately affected, and antimicrobial resistance reduces therapeutic options. It is estimated that 2–5% of enteric fever patients develop chronic asymptomatic infection. These carriers may act as reservoirs of infection; therefore, the prospective identification and treatment of carriers are critical for long-term disease control. We aimed to find the frequency of Salmonella Typhi carriers in patients undergoing cholecystectomy. We also compared the detection limit of culturing versus qPCR in detecting S. Typhi, performed a geospatial analysis of the carriers identified using this study, and evaluated the accuracy of anti-Vi and anti-YncE in identifying chronic typhoid carriage. Methods: We performed a cross-sectional study in two centers in Pakistan. Gallbladder specimens were subjected to quantitative PCR (qPCR) and serum samples were analyzed for IgG against YncE and Vi by ELISA. We also mapped the residential location of those with a positive qPCR result. Findings: Out of 988 participants, 3.4% had qPCR-positive gallbladder samples (23 S. Typhi and 11 S. Paratyphi). Gallstones were more likely to be qPCR positive than bile and gallbladder tissue. Anti-Vi and YncE were significantly correlated (r = 0.78 p<0.0001) and elevated among carriers as compared to qPCR negative controls, except for anti-Vi response in Paratyphi A. But the discriminatory values of these antigens in identifying carriers from qPCR negative controls were low. Conclusion: The high prevalence of typhoid carriers observed in this study suggests that further studies are required to gain information that will help in controlling future typhoid outbreaks in a superior manner than they are currently being managed. Author summary: Enteric fever, caused by bacteria Salmonella Typhi or Paratyphi, is a serious global illness particularly affecting children in areas with inadequate hygiene and sanitation facilities. It is transmitted through the oral-fecal route. In Pakistan, prevalence of extensively drug resistant S. Typhi resistant strain further complicates its treatment. Some infected individuals develop chronic infections and become carriers of these pathogens. These asymptomatic carriers may potentially spread the disease by shedding the bacteria in urine or stool. To assess the frequency of the carrier state, we examined gallbladder specimens (bile, stones, and tissue) from patients who underwent gallbladder removal surgeries at Aga Khan University and Jinnah Post-graduate Medical Centre. We demonstrated that PCR had a higher sensitivity in detecting Salmonella carriage with a detection limit of 102 CFU/ml and identified 3.4% of typhoid carriers. Additionally, we investigated whether specific antibodies could identify these carriers using various serological tests. The results underscore the need for further research to promptly detect and appropriately treat this disease, thereby enhancing control and preventing future outbreaks. [ABSTRACT FROM AUTHOR]

Objective The aim of this study was to compare the performance of direct amplification of viral nucleic acid from transport medium to extracted nucleic acid for polymerase chain reaction (PCR), sequencing, and genotyping applications. Methods XpressAmp lysate and extracted total nucleic acid from viral transport medium containing nasopharyngeal specimens were evaluated across different molecular applications to determine performance characteristics. Results SARS-CoV-2 quantitative PCR and angiotensin-converting enzyme (ACE) genotyping assays worked well with XpressAmp lysate, almost equal with or better than extracted nucleic acid in some specimens. However, XpressAmp completely failed to perform in next-generation sequencing for strain typing. Both protocols failed to detect ACE2 expression in viral transport medium. Conclusion Direct amplification of viral nucleic acid from viral transport medium containing nasopharyngeal specimen works well for molecular assays with low thresholds of quality; however, it does have limitations with assays that require high quality nucleic acid for input. Use of the XpressAmp protocol significantly improves turnaround time and allows for easy ramp-up of PCR and genotyping assays. [ABSTRACT FROM AUTHOR]

Introduction: Methicillin-resistant Staphylococcus aureus (MRSA) has been known as an infectious pathogen worldwide since 1960. The epidemiological distribution of MRSA may have shifted due to outbreaks reported from several nations, making it more challenging to differentiate among CA-MRSA and HA-MRSA. It is currently important to develop a strain-based explanation for HA and CA-MRSA due to its distinct epidemiology, genetic profile, antibiogram, and quantifiable features. The study aimed to distinguish CA and HA-MRSA by Staphylococcal Cassette Chromosome mec (SCCmec) typing. Materials and Method: The study involved a total of 381 S. aureus isolates, which were processed in the department of Microbiology, JSS Hospital, Mysore. All isolates were confirmed as MRSA, initially by disk diffusion method using cefoxitin 30µg and oxacillin 1μg disk and later by using PCR technique for the detection of mecA-gene. All mecA-gene positive samples were amplified for SCCmec typing by multiplex PCR for detection of SCCmec type I, II, III, IVa, IVb, IVc, IVd, V and XI respectively. Results: PCR confirmed a total of 66% isolates as mecA-positive MRSA. Multiplex PCR method revealed only 53% isolates were SCCmec-typeable. The mainstream of the isolates belonged to SCCmec type IV (53.48%) and type V (44.18%), followed by type III (9.30%), type II (3%) and type I (1.16%) respectively. The study also demonstrated the presence of multiple SCCmec types in 10.46% of MRSA isolates. Staphylococcal cassette chromosome recombinase (ccr) typing determined only 43% of isolates were typeable. Conclusion: The study found that hospital-associated SCCmec type IV and type V were the most circulating strains in our healthcare setting. The research identified a few MRSA isolates with diverse SCCmec types. The presence of CAMRSA infection in in-patients and HA-MRSA infection in out-patients were also recognised. The situation indicates the spreading of hospital-acquired strains into the community and vice versa, necessitating the molecular characterisation of MRSA isolates in order to administer the appropriate antibiotic medication. [ABSTRACT FROM AUTHOR]

Acinetobacters pose a significant threat to human health, especially those with weakened immune systems. Type IV pili of acinetobacters play crucial roles in virulence and antibiotic resistance. Single-stranded RNA bacteriophages target the bacterial retractile pili, including type IV. Our study delves into the interaction between Acinetobacter phage AP205 and type IV pili. Using cryo-electron microscopy, we solve structures of the AP205 virion with an asymmetric dimer of maturation proteins, the native Acinetobacter type IV pili bearing a distinct post-translational pilin cleavage, and the pili-bound AP205 showing its maturation proteins adapted to pilin modifications, allowing each phage to bind to one or two pili. Leveraging these results, we develop a 20-kilodalton AP205-derived protein scaffold targeting type IV pili in situ, with potential for research and diagnostics. Here, the authors structurally characterise the interaction between Acinetobacter phage AP205 and the type IV Acinetobacter pili using cryo-electron microscopy, uncovering the mechanistic determinants of this interaction. [ABSTRACT FROM AUTHOR]

Objective To evaluate the performance of nitrate reductase assay (NRA), a rapid, colorimetric method for the determination of methicillin resistance in Staphylococcus aureus isolates obtained from the culture collection of the Akdeniz University Hospital Central Laboratory, Antalya, Türkiye. Materials and Methods Identification for all 290 S aureus isolates at the species level was performed via matrix-assisted laser desorption/ionization-time of flight. Isolates were tested with NRA for methicillin resistance. The cefoxitin broth microdilution (BMD) method recommended by the Clinical and Laboratory Standards Institute was used as the reference method in the study. S aureus ATCC 29213 and S aureus ATCC 43300 strains were used for quality control. Results According to Food and Drug Administration criteria, the category agreement between NRA and BMD was found to be 100%. The essential agreement between both methods was determined to be 96.20%. There is no minor, major, or extremely major discrepancy between both methods. Conclusion The results show that NRA is a rapid, practical, and reliable colorimetric method for detecting MRSA. [ABSTRACT FROM AUTHOR]

Bacteriophage therapy is a promising approach to treating bacterial infections. Research and development of bacteriophage therapy is intensifying due to the increase in antibiotic resistance and the faltering development of new antibiotics. Bacteriophage therapy uses bacteriophages (phages), i.e., prokaryotic viruses, to specifically target and kill pathogenic bacteria. The legal handling of this type of therapy raises several questions. These include whether phage therapeutics belong to a specially regulated class of medicinal products, and which legal framework should be followed with regard to the various technical ways in which phage therapeutics can be manufactured and administered. The article shows to which class of medicinal products phage therapeutics from wild type phages and from genetically modified (designer) phages do or do not belong. Furthermore, the article explains which legal framework is relevant for the manufacture and administration of phage therapeutics, which are manufactured in advance in a uniform, patient-independent manner, and for tailor-made patient-specific phage therapeutics. For the systematically coherent, successful translation of phage therapy, the article considers pharmaceutical law and related legal areas, such as genetic engineering law. Finally, the article shows how the planned legislative revisions of Directive 2001/83/EC and Regulation (EC) No 726/2004 may affect the legal future of phage therapy. [ABSTRACT FROM AUTHOR]

Bacteriophage therapy presents a promising avenue for combating antibiotic-resistant bacterial infections. Yet, challenges exist, particularly, the lack of a straightforward purification pipeline suitable for widespread application to many phage types, as some phages are known to undergo significant titer loss when purified via current techniques. Electrokinetic methods offer a potential solution to this hurdle, with nonlinear electrophoresis emerging as a particularly appealing approach due to its ability to discern both the size and shape of the target phage particles. Presented herein is the electrokinetic characterization of the mobility of nonlinear electrophoresis for two phages (SPN3US and ϕKZ) and three types of polystyrene nanoparticles. The latter served as controls and were selected based on their sizes and surface charge magnitude. Particle tracking velocimetry experiments were conducted to characterize the mobility of all five particles included in this study. The results indicated that the selected nanoparticles effectively replicate the migration behavior of the two phages under electric fields. Further, it was found that there is a significant difference in the nonlinear electrophoretic response of phages and that of host cells, as first characterized in a previous report, illustrating that electrokinetic-based separations are feasible. The findings from this work are the first characterization of the behavior of phages under nonlinear electrophoresis effects and illustrate the potential for the development of electrokinetic-based phage purification techniques that could aid the advancement of bacteriophage therapy. [ABSTRACT FROM AUTHOR]

Flavobacterium plurextorum is a potential fish pathogen of interest, previously isolated from diseased rainbow trout (Oncorhynchus mykiss) and oomycete‐infected chum salmon (Oncorhynchus keta) eggs. We report here the first complete genome sequence of F. plurextorum RSG‐18 isolated from the gut of Schlegel's black rockfish (Sebastes schlegelii). The genome of RSG‐18 consists of a circular chromosome of 5,610,911 bp with a 33.57% GC content, containing 4858 protein‐coding genes, 18 rRNAs, 63 tRNAs and 1 tmRNA. A comparative analysis was conducted on 11 Flavobacterium species previously reported as pathogens or isolated from diseased fish to confirm the potential pathogenicity of RSG‐18. In the SEED classification, RSG‐18 was found to have 36 genes categorized in 'Virulence, Disease and Defense'. Across all Flavobacterium species, a total of 16 antibiotic resistance genes and 61 putative virulence factors were identified. All species had at least one phage region and type I, III and IX secretion systems. In pan‐genomic analysis, core genes consist of genes linked to phages, integrases and matrix‐tolerated elements associated with pathology. The complete genome sequence of F. plurextorum RSG‐18 will serve as a foundation for future research, enhancing our understanding of Flavobacterium pathogenicity in fish and contributing to the development of effective prevention strategies. [ABSTRACT FROM AUTHOR]

Chronic wounds have been a major factor of serious harm to global public health. At present, it is known that almost all chronic wounds contain biofilms, which seriously hinder the healing process. Removal of biofilms can effectively promote the healing of chronic wounds. As the study of wound biofilms deepens, many new treatment methods have emerged, thus bringing revolutionary means for the treatment of chronic wound biofilm. This review summarizes various methods for the treatment of chronic wound biofilm worldwide to provide a theoretical summary and practical basis for the selection of suitable wound biofilm treatment methods in clinical practice. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus sequence type (ST) 398 is a lineage affecting both humans and livestock worldwide. However, the mechanisms underlying its clonal evolution are still not clearly elucidated. We applied whole-genome sequencing (WGS) typing to 45 S. aureus strains from China and Canada between 2005 and 2014, in order to gain insight into their evolutionary pathway. Based on WGS phylogenetic analysis, 42 isolates were assigned to the human-associated clade (I/II-GOI) and 3 isolates to livestock-associated clade (IIa). Phylogeny of ϕSa3 sequences revealed five phage groups (Groups 1–5), with Group 1 carrying ϕSa3-Group 1 (ϕSa3-G1), Group 2 carrying ϕSa3-G2, Group 3 carrying ϕSa3-G3, Group 4 carrying ϕSa3-G4 and Group 5 lacking ϕSa3. ϕSa3-G1 was only found in strains that accounted for the most ancestral human clade I, while ϕSa3-G2, ϕSa3-G3 and ϕSa3-G4 were found restricted to sublineages within clade II-GOI. Some isolates of clade II-GOI were also found to be ϕSa3-negative or resistant to methicillin which are unusual characteristics for human-adapted isolates. This study demonstrated a strong association between phylogenetic grouping and phage type, suggesting an important role of ϕSa3 prophage in the evolution of human-adapted ST398 subclones. In addition, our results suggest that this subclone slowly began to adapt to animal hosts by losing ϕSa3 and acquiring methicillin resistance, which was observed in some strains of human-associated clade II-GOI, an intermediate human to livestock transmission clade. [ABSTRACT FROM AUTHOR]

We conducted laboratory experiments under varied solar radiation and salinity levels to investigate their influences on the natural attenuation of multiple promising microbial indicators including fecal bacteria and two types of bacteriophages. Inactivation coefficients were estimated and compared following first-order kinetics. Somatic coliphage was found to be the most resistant, while fecal bacteria exhibited higher susceptibility to both factors. The estimated inactivation coefficients of E. coli were applied to a 3D water quality model and validated with a daily basis monitoring dataset. The validation revealed high consistency among modelled and monitored concentrations, with a less than 1-log concentration difference. Further, the effect of actual solar radiation and salinity on E. coli inactivation after a rainfall event was calculated and compared. The results exhibited that solar radiation is a stronger influential factor. Simulation illustrated that lower-strength radiation exposure can limit E. coli inactivation, enabling them to survive up to one week after combined sewer overflow (CSO) discharge. The model revealed a promising capacity as a tool for the timely prediction of the CSO-induced severity of microbial contamination and associated risk, as well as associated natural attenuation; thus, this model can enhance the competency of public water managers for decision making. [ABSTRACT FROM AUTHOR]

Simple Summary: Breast cancer, the second-leading cause of mortality among women worldwide, is often diagnosed through invasive tissue sampling. This method may not fully represent the tumor's overall physiology, potentially leading to suboptimal treatment strategies and tumor recurrence. A non-invasive approach using MRI to quantify the hypoxic environment within the tumor could significantly enhance the accuracy of breast-cancer-subtype prediction. Our study introduces non-invasive imaging markers for hypoxia and related angiogenesis, utilizing hyperoxic-blood-oxygen-level-dependent (BOLD)-MRI and intravoxel-incoherent-motion (IVIM)-MRI techniques. These markers correlate with microvessel density and maturity as assessed by histology, enabling the distinction between the less aggressive luminal A and the more aggressive triple-negative breast cancers. Tumor neoangiogenesis is an important hallmark of cancer progression, triggered by alternating selective pressures from the hypoxic tumor microenvironment. Non-invasive, non-contrast-enhanced multiparametric MRI combining blood-oxygen-level-dependent (BOLD) MRI, which depicts blood oxygen saturation, and intravoxel-incoherent-motion (IVIM) MRI, which captures intravascular and extravascular diffusion, can provide insights into tumor oxygenation and neovascularization simultaneously. Our objective was to identify imaging markers that can predict hypoxia-induced angiogenesis and to validate our findings using multiplexed immunohistochemical analyses. We present an in vivo study involving 36 female athymic nude mice inoculated with luminal A, Her2+, and triple-negative breast cancer cells. We used a high-field 9.4-tesla MRI system for imaging and subsequently analyzed the tumors using multiplex immunohistochemistry for CD-31, PDGFR-β, and Hif1-α. We found that the hyperoxic-BOLD-MRI-derived parameter ΔR2* discriminated luminal A from Her2+ and triple-negative breast cancers, while the IVIM-derived parameter fIVIM discriminated luminal A and Her2+ from triple-negative breast cancers. A comprehensive analysis using principal-component analysis of both multiparametric MRI- and mpIHC-derived data highlighted the differences between triple-negative and luminal A breast cancers. We conclude that multiparametric MRI combining hyperoxic BOLD MRI and IVIM MRI, without the need for contrast agents, offers promising non-invasive markers for evaluating hypoxia-induced angiogenesis. [ABSTRACT FROM AUTHOR]

Phage therapy has shown great promise for the treatment of multidrugresistant bacterial infections. However, the lack of a thorough and organized understanding of phage-body interactions has limited its clinical application. Here, we administered different purified phages (Salmonella phage SE_SZW1, Acinetobacter phage AB_SZ6, and Pseudomonas phage PA_LZ7) intravenously to healthy animals (rats and monkeys) to evaluate the phage-induced host responses and phage pharmacokinetics with different intravenous (IV) doses in healthy animals. The plasma and the organs were sampled after different IV doses to determine the phage biodistribution, phage-induced cytokines, and antibodies. The potential side effects of phages on animals were assessed. A non-compartment model revealed that the plasma phage titer gradually decreased over time following a single dose. Repeated doses resulted in a 2-3 Log10 decline of the plasma phage titer at 5 min compared to the first dose, regardless of the type of phage administered in rats. Host innate immune responses were activated including splenic enlargement following repeated doses. Phage-specific neutralization antibodies in animals receiving phages were detected. Similar results were obtained from monkeys. In conclusion, the mammalian bodies were well-tolerant to the administered phages. The animal responses to the phages and the phage biodistribution profiles could have a significant impact on the efficacy of phage therapy. IMPORTANCE Phage therapy has demonstrated potential in addressing multidrugresistant bacterial infections. However, an insufficient understanding of phage-host interactions has impeded its broader clinical application. In our study, specific phages were administered intravenously (IV) to both rats and monkeys to elucidate phage-host interactions and evaluate phage pharmacokinetics (PK). Results revealed that with successive IV administrations, there was a decrease in plasma phage concentrations. Concurrently, these administrations elicited both innate and adaptive immune responses in the subjects. Notably, the observed immune responses and PK profiles exhibited variation contingent upon the phage type and the mammalian host. Despite these variations, the tested mammals exhibited a favorable tolerance to the IV-administered phages. This underscores the significance of comprehending these interactions for the optimization of phage therapy outcomes. [ABSTRACT FROM AUTHOR]

Intestinal microbiota, which plays an important role in human health by interacting with each other or with its hosts, is affected by many endogenous and exogenous factors. Any change in the composition and functionality of the intestinal microbiota, both in number and diversity, causes disruption of intestinal functioning and paves the way for many diseases. In this regard, many antimicrobial peptides, especially bacteriocins, synthesized by lactic acid bacteria are thought to be natural resources with a high potential for the protection of the intestinal microbiota and the treatment of intestinal diseases. Both the intestinal microbiota itself and many foodborne bacteria produce bacteriocins that can inhibit pathogenic microorganisms that cause serious health problems and regulate the intestinal microbiota. This review aims to provide a comprehensive overview of the intestinal microbiota, the properties of lactic acid bacteria, their bacteriocins, and the effects of bacteriocins on the human health. [ABSTRACT FROM AUTHOR]

Heritable, facultative symbionts are common in arthropods, often functioning in host defence. Despite moderately reduced genomes, facultative symbionts retain evolutionary potential through mobile genetic elements (MGEs). MGEs form the primary basis of strain‐level variation in genome content and architecture, and often correlate with variability in symbiont‐mediated phenotypes. In pea aphids (Acyrthosiphon pisum), strain‐level variation in the type of toxin‐encoding bacteriophages (APSEs) carried by the bacterium Hamiltonella defensa correlates with strength of defence against parasitoids. However, co‐inheritance creates difficulties for partitioning their relative contributions to aphid defence. Here we identified isolates of H. defensa that were nearly identical except for APSE type. When holding H. defensa genotype constant, protection levels corresponded to APSE virulence module type. Results further indicated that APSEs move repeatedly within some H. defensa clades providing a mechanism for rapid evolution in anti‐parasitoid defences. Strain variation in H. defensa also correlates with the presence of a second symbiont Fukatsuia symbiotica. Predictions that nutritional interactions structured this coinfection were not supported by comparative genomics, but bacteriocin‐containing plasmids unique to co‐infecting strains may contribute to their common pairing. In conclusion, strain diversity, and joint capacities for horizontal transfer of MGEs and symbionts, are emergent players in the rapid evolution of arthropods. [ABSTRACT FROM AUTHOR]

The article focuses on the detection of Streptococcus pyogenes in a pleural effusion from a patient with HIV and Hodgkin lymphoma. Topics include the use of May-Grünwald-Giemsa staining in identifying bacterial morphology, the confirmation of S. pyogenes through blood culture and Gram staining, and the whole-genome sequencing that revealed the presence of a hyaluronic acid capsule.

Bacteriophage (phage) therapy is being explored as a possible response to the antimicrobial resistance public health emergency. Administering a mixture of different phage types as a cocktail is one proposed strategy for therapeutic applications, but the optimal method for formulating phage cocktails remains a major challenge. Each phage strain has complex pharmacokinetic/ pharmacodynamic (PK/PD) properties which depend on the nano-scale size, target-mediated, self-dosing nature of each phage strain, and rapid selection of resistant subpopulations. The objective of this study was to explore the pharmacodynamics (PD) of three unique and clinically relevant anti-Pseudomonas phages after simulation of dynamic dosing strategies. The Hollow Fiber Infection Model (HFIM) is an in vitro system that mimics in vivo pharmacokinetics (PK) with high fidelity, providing an opportunity to quantify phage and bacteria concentration profiles over clinical time scales with rich sampling. Exogenous monotherapybolus (producing max concentrations of Cmax  =  7 log10 PFU/mL) regimens of phages LUZ19, PYO2, and E215 produced Pseudomonas aeruginosa nadirs of 0, 2.14, or 2.99 log10 CFU/mL after 6  h of treatment, respectively. Exogenous combination therapy bolus regimens (LUZ19  +  PYO2 or LUZ19  +  E215) resulted in bacterial reduction to <2 log10 CFU/mL. In contrast, monotherapy as a continuous infusion (producing a steady-state concentration of Css,avg  =  2 log10PFU/mL) was less effective at reducing bacterial densities. Specifically, PYO2 failed to reduce bacterial density. Next, a mechanism-based mathematical model was developed to describe phage pharmacodynamics, phage–phage competition, and phagedependent adaptive phage resistance. Monte Carlo simulations supported bolus dose regimens, predicting lower bacterial counts with bolus dosing as compared to prolonged phage infusions. Together, in vitro and in silico evaluation of the time course of phage pharmacodynamics will better guide optimal patterns of administration of individual phages as a cocktail. [ABSTRACT FROM AUTHOR]

Background and Aim: Escherichia coli (E.coli) is an essential bacterial indicator in the control of pharmaceutical quality and other similar fields. There are some biosensors designed for its detection based on electrochemical transduction methods. A biosensor with reduced graphene oxide was modified. Traditional methods are time-consuming and high prices, so in this study a new biosensor with modification of reduced graphene oxide (rGO) as a kind of carbon composition on glassy carbon electrode (GCE) with Au Nanoparticles (Au NPs) decoration was used for E. coli detection. Materials and Methods: Reduced graphene oxide (rGO) modified on glassy carbon electrode (GCE) and chronoamperometric and reduction methods were used for Au NPs decoration and it was completed with polyclonal E. coli antibody and 0.5 W/V% Bovine Serum Albumin (BSA) solution. Morphology and structure of rGO and Au NPs in GCE/rGO/Au NPs/ E. coli polyclonal antibody/ BSA biosensor were verified by SEM (Scanning Electron Microscope) during modification steps. E. coli detection in dissimilar samples was performed by Square-Wave Voltammetry (SWV) and Cyclic Voltammetry (CV) techniques which were set in 0.1M phosphate buffer solution (PBS) (pH 7.4) mixed with 0.5mM acetaminophen. In comparison with the biosensor, the classical method of detection was performed with serial dilutions of E. coli ATCC 8739 (1×101-1×108 CFU/mL) which was cultured on media. Results: Despite the two methods used to stabilize Au NPs, scanning electron microscope (SEM) images showed that the current difference did not increase due to gold particles. Its modification had no significant change in current and it was not a successful experiment for E.coli detection. Conclusion: Compared with the plate method, biosensors couldn't substitute with conventional methods for E. coli detection. [ABSTRACT FROM AUTHOR]

Acute diarrhea and cholera (AWD/C) result in more than 21000 to 143000 global mortality annually and are associated with Vibrio cholerae. The pathogen has shown increasing evolutionary/emerging dynamics linked with mobilome or ubiquitous nature of mobile integrative genetic and conjugative elements (MIGCE), however, such dynamics are rarely reported amongst somatic-antigen nonagglutinating Type-1/-139 V. cholerae (SA-NAG-T-1/139Vc). The study reports the genetic detection of mobilome-associated indices in SA-NAG-T-1/139Vc as a potential strategy for differentiating/discriminating emerging environmental bacteria. Presumptive V. cholerae isolates were retrieved from five water sources, while strains were characterized/serogrouped and confirmed using simplex and comparative-genomic-multiplex Polymerase Chain Reaction (PCR). Genomic island (GI-12det, GI-14det, GI-15det); Phages (TLC-phagedet, Kappa-phagedet) and ICEs of the SXT/R391 family genes (SXT/R391-ICEs integrase, SXT-Hotspot-IV, ICEVchInd5Hotspot-IV, ICEVchMoz10Hotspot-IV) were detected. Other rare ICE members such as the ICEVcBan8att gene and Vibrio Seventh Pandemic island detection (VSP-II Integrase, Prototypical VSP-II) were also detected. Results revealed that the 8.22% (61/742) SA-NAG-T-1/139Vc serogroup observed harbors the Vibrio Seventh Pandemic island integrase (34/61; 55.7%) and other rare genetic traits including: attB/attP (29/61; 47.5%, 14/61; 23%), integrative genetic elements (4/61; 6.56%), phage types (TLC-phagedet: 2/61; 3.28% and Kappa-phagedet: 7/61; 11.48%) as well as the integrase genes (INT1, Sul1, Sul2) (29/61: 47.5%; 21/61: 34.4%; 25/61: 41%). Such genetic detection of mobilome determinants/MIGCE suggests potential discriminatory tendencies amongst SA-NAG-T-1/139Vc which may be applied in mobilome typing of evolving/emerging environmental bacteria. The need to encourage the application of such mobilome typing indices and continuous study of these strains is suggestive of interest in controlling future potential emerging environmental strains. [ABSTRACT FROM AUTHOR]

Phage therapy is a promising method for the treatment of multidrug-resistant bacterial infections. However, its long-term efficacy depends on understanding the evolutionary effects of the treatment. Current knowledge of such evolutionary effects is lacking, even in well-studied systems. We used the bacterium Escherichia coli C and its bacteriophage ΦX174, which infects cells using host lipopolysaccharide (LPS) molecules. We first generated 31 bacterial mutants resistant to ΦX174 infection. Based on the genes disrupted by these mutations, we predicted that these E. coli C mutants collectively produce eight unique LPS structures. We then developed a series of evolution experiments to select for ΦX174 mutants capable of infecting the resistant strains. During phage adaptation, we distinguished two types of phage resistance: one that was easily overcome by ΦX174 with few mutational steps ("easy" resistance) and one that was more difficult to overcome ("hard" resistance). We found that increasing the diversity of the host and phage populations could accelerate the adaptation of phage ΦX174 to overcome the hard resistance phenotype. From these experiments, we isolated 16 ΦX174 mutants that, together, can infect all 31 initially resistant E. coli C mutants. Upon determining the infectivity profiles of these 16 evolved phages, we uncovered 14 distinct profiles. Given that only eight profiles are anticipated if the LPS predictions are correct, our findings highlight that the current understanding of LPS biology is insufficient to accurately forecast the evolutionary outcomes of bacterial populations infected by phage. [ABSTRACT FROM AUTHOR]

The Anderson phage typing scheme has been successfully used worldwide for epidemiological surveillance of Salmonella enterica serovar Typhimurium. Although the scheme is being replaced by whole genome sequence subtyping methods, it can provide a valuable model system for study of phage-host interaction. The phage typing scheme distinguishes more than 300 definitive types of Salmonella Typhimurium based on their patterns of lysis to a unique collection of 30 specific Salmonella phages. In this study, we sequenced the genomes of 28 Anderson typing phages of Salmonella Typhimurium to begin to characterize the genetic determinants that are responsible for the differences in these phage type profiles. Genomic analysis of typing phages reveals that Anderson phages can be classified into three different groups, the P22-like, ES18-like and SETP3-like clusters. Most Anderson phages are short tailed P22-like viruses (genus Lederbergvirus); but phages STMP8 and STMP18 are very closely related to the lambdoid long tailed phage ES18, and phages STMP12 and STMP13 are related to the long noncontractile tailed, virulent phage SETP3. Most of these typing phages have complex genome relationships, but interestingly, two phage pairs STMP5 and STMP16 as well as STMP12 and STMP13 differ by a single nucleotide. The former affects a P22-like protein involved in DNA passage through the periplasm during its injection, and the latter affects a gene whose function is unknown. Using the Anderson phage typing scheme would provide insights into phage biology and the development of phage therapy for the treatment of antibiotic resistant bacterial infections. [ABSTRACT FROM AUTHOR]

Objective The aim of this study was to evaluate the effects of Pan Genera Detection (PGD) testing on reducing platelet product wastage and transfusion service costs. Methods We conducted a retrospective cross-sectional study comparing the number of platelet apheresis units wasted before (March 2017 to February 2019) and after (March 2019 to February 2021) PGD implementation. The PGD testing was performed before transfusion on days 6 and 7. Cost analysis considered the costs of platelet units wasted ($500.00/unit) and PGD test supplies and performance (estimated $26.50 per test). Paired samples t -test was used to compare platelet wastage pre- and post-PGD implementation. Results The number of wasted platelet units decreased from pre-PGD (419) to post-PGD (195), representing a significant decrease in platelet wastage from 17.5% to 9.2% (P  < .0001). During the post-PGD period, 366 and 133 units were tested on days 6 and 7, with 28 and 36 units discarded each day, allowing transfusion of an additional 302 platelet units. Costs from platelet wastage decreased from $209,500.00 pre-PGD to $97,500.00 post-PGD. Conclusion Our results showed that PGD testing effectively reduced platelet wastage, extended platelet availability, and reduced transfusion service costs. [ABSTRACT FROM AUTHOR]

On-pump cardiac surgery triggers sterile inflammation and postoperative complications such as postoperative atrial fibrillation (POAF). Hematopoietic somatic mosaicism (HSM) is a recently identified risk factor for cardiovascular diseases and results in a shift toward a chronic proinflammatory monocyte transcriptome and phenotype. The aim of this study was to assess the prevalence, characteristics, and impact of HSM on preoperative blood and myocardial myeloid cells as well as on outcomes after cardiac surgery. Blood DNA from 104 patients referred for surgical aortic valve replacement (AVR) was genotyped using the HemePACT panel (576 genes). Four screening methods were applied to assess HSM, and postoperative outcomes were explored. In-depth blood and myocardial leukocyte phenotyping was performed in selected patients using mass cytometry and preoperative and postoperative RNA sequencing analysis of classical monocytes. The prevalence of HSM in the patient cohort ranged from 29%, when considering the conventional HSM panel (97 genes) with variant allelic frequencies ≥2%, to 60% when considering the full HemePACT panel and variant allelic frequencies ≥1%. Three of 4 explored HSM definitions were significantly associated with higher risk for POAF. On the basis of the most inclusive definition, HSM carriers exhibited a 3.5-fold higher risk for POAF (age-adjusted OR: 3.5; 95% CI: 1.52-8.03; P = 0.003) and an exaggerated inflammatory response following AVR. HSM carriers presented higher levels of activated CD64+CD14+CD16− circulating monocytes and inflammatory monocyte-derived macrophages in presurgery myocardium. HSM is frequent in candidates for AVR, is associated with an enrichment of proinflammatory cardiac monocyte–derived macrophages, and predisposes to a higher incidence of POAF. HSM assessment may be useful in the personalized management of patients in the perioperative period. (Post-Operative Myocardial Incident & Atrial Fibrillation [POMI-AF]; NCT03376165) [Display omitted] [ABSTRACT FROM AUTHOR]

The marine bacterial family Oceanospirillaceae, is well-known for its ability to degrade hydrocarbons and for its close association with algal blooms. However, only a few Oceanospirillaceae-infecting phages have been reported thus far. Here, we report on a novel Oceanospirillum phage, namely, vB_OsaM_PD0307, which has a 44,421 bp linear dsDNA genome and is the first myovirus infecting Oceanospirillaceae. A genomic analysis demonstrated that vB_OsaM_PD0307 is a variant of current phage isolates from the NCBI data set but that it has similar genomic features to two highquality, uncultured viral genomes identified from marine metagenomes. Hence, we propose that vB_OsaM_PD0307 can be classified as the type phage of a new genus, designated Oceanospimyovirus. Additionally, metagenomic read mapping results have further shown that Oceanospimyovirus species are widespread in the global ocean, display distinct biogeographic distributions, and are abundant in polar regions. In summary, our findings expand the current understanding of the genomic characteristics, phylogenetic diversity, and distribution of Oceanospimyovirus phages. IMPORTANCE: Oceanospirillum phage vB_OsaM_PD0307 is the first myovirus found to infect Oceanospirillaceae, and it represents a novel abundant viral genus in polar regions. This study provides insights into the genomic, phylogenetic, and ecological characteristics of the new viral genus, namely Oceanospimyovirus. [ABSTRACT FROM AUTHOR]

Introduction: Diabetes is a non-contagious disease, but it can cause various complications. One of the most common complications of diabetes is diabetic ulcers. Diabetic ulcers are infections that occur in the legs of diabetics due to the destruction of the deepest skin tissue. Recent studies have reported the presence of Alcaligenes faecalis with extensive drug resistance (XDR) properties as a cause of diabetic ulcers. Bacteriophages are known to have the ability to infect bacteria specifically so that they can be used as an alternative solution for treating diabetic ulcers. The purpose of this study was to determine the characteristics of bacteriophages capable of infecting Alcaligenes faecalis bacteria. Material and methods: The method used is the spot test method, host range, and identification of nucleic acid types. Results: The results showed that the 6 bacteriophages isolated, namely AFaV1, AFaV2, AFaV3, AFaV4, AFaV5, and AFaV6, had cloudy plaques with a diameter of ±3 mm. AFaV1, AFaV2, and AFaV4 isolates could infect all bacteria used; they were Klebsiella pneumoniae, Escherichia coli, and Staphylococcus aureus. Meanwhile, bacteriophage isolates AFaV3, AFaV5, and AFaV6 could infect Klebsiella pneumoniae and Staphylococcus aureus bacteria only. The nucleic acid types of the 6 bacteriophage samples were dsDNA with band length > 1 Kb. Conclusions: The 6 isolates that were isolated had the ability to infect by forming a prophage that could inhibit the growth of Alcaligenes faecalis and other pathogenic bacteria in diabetic ulcers. [ABSTRACT FROM AUTHOR]

Phage therapy is now seen as a promising way to overcome the current global crisis in the spread of multidrug-resistant bacteria. However, phages are highly strain-specific, and in most cases one will have to isolate a new phage or search for a phage suitable for a therapeutic application in existing libraries. At an early stage of the isolation process, rapid screening techniques are needed to identify and type potential virulent phages. Here, we propose a simple PCR approach to differentiate between two families of virulent Staphylococcus phages (Herelleviridae and Rountreeviridae) and eleven genera of virulent Klebsiella phages (Przondovirus, Taipeivirus, Drulisvirus, Webervirus, Jiaodavirus, Sugarlandvirus, Slopekvirus, Jedunavirus, Marfavirus, Mydovirus and Yonseivirus). This assay includes a thorough search of a dataset comprising S. aureus (n = 269) and K. pneumoniae (n = 480) phage genomes available in the NCBI RefSeq/GenBank database for specific genes that are highly conserved at the taxonomic group level. The selected primers showed high sensitivity and specificity for both isolated DNA and crude phage lysates, which permits circumventing DNA purification protocols. Our approach can be extended and applied to any group of phages, given the large number of available genomes in the databases. [ABSTRACT FROM AUTHOR]

An outbreak of food poisoning in New South Wales (NSW) Australia in 2018, caused by Salmonella enterica serovar Enteritidis phage type 12 (PT12), was traced to eggs consumed from a NSW layer flock. This was the first report of Salmonella Enteritidis infection in NSW layer flocks, despite ongoing environmental monitoring. Clinical signs and mortalities were minimal in most flocks, although seroconversion and infection were demonstrated in some flocks. An oral dose–response challenge study with Salmonella Enteritidis PT12 was undertaken in commercial point-of-lay hens. Cloacal swabs collected at 3, 7, 10, and 14 days postinoculation and caeca, liver, spleen, ovary, magnum, and isthmus tissues collected at necropsy at either 7 or 14 days were processed for Salmonella isolation (AS 5013.10-2009 from ISO6579:2002). Histopathology was performed on the above tissues, as well as lung, pancreas, kidney, heart, and additional intestinal and reproductive tract tissues. Salmonella Enteritidis was consistently detected in cloacal swabs between 7 and 14 days postchallenge. The Salmonella Enteritidis PT12 isolate successfully colonized the gastrointestinal tract, liver, and spleen of all hens orally challenged with 107, 108, and 109Salmonella Enteritidis, and less consistently colonized their reproductive tracts. On histopathology, mild lymphoid hyperplasia in the liver and spleen, along with hepatitis, typhlitis, serositis, and salpingitis, was observed at 7 and 14 days postchallenge, with a greater proportion of affected birds in the two higher dose groups. Diarrhea and culture of Salmonella Enteritidis from heart blood were not detected in challenged layers. The NSW isolate of Salmonella Enteritidis PT12 was able to invade and colonize the birds' reproductive tracts as well as a wide range of other tissues, indicating the potential for these naive commercial hens to contaminate their eggs. La inoculación oral de gallinas ponedoras en el pico de postura con la cepa de Salmonella Enteritidis PT12 del brote en Nueva Gales del Sur causa infección, pero una histopatología mínima. Un brote de intoxicación alimentaria en Nueva Gales del Sur (NSW), Australia en 2018, causado por Salmonella enterica serovar Enteritidis fagotipo 12, se rastreó hasta los huevos consumidos de una parvada de ponedoras de NSW. Este fue el primer informe de infección por Salmonella Enteritidis en parvadas de ponedoras de NSW, a pesar del monitoreo ambiental continuo. Los signos clínicos y la mortalidad fueron mínimos en la mayoría de las parvadas, aunque se demostró seroconversión e infección en algunas parvadas. Se llevó a cabo un estudio de desafío oral para evaluar la dosis y su respuesta para Salmonella Enteritidis PT12 en gallinas ponedoras comerciales. Los hisopos cloacales recolectados a los tres, siete, diez y 14 días posteriores a la inoculación y los tejidos de ciego, hígado, bazo, ovario, magnum e istmos recolectados en la necropsia a los siete o 14 días se procesaron para el aislamiento de Salmonella (AS 5013.10-2009 del estándar ISO6579: 2002). Se realizó histopatología en los tejidos anteriormente mencionados, así como de pulmón, páncreas, riñón, corazón y tejidos intestinales y del tracto reproductivo adicionales. Salmonella Enteritidis se detectó consistentemente en hisopos cloacales entre los siete y 14 días después del desafío. El aislado de Salmonella Enteritidis PT12 colonizó con éxito el tracto gastrointestinal, el hígado y el bazo de todas las gallinas desafiadas por vía oral con dosis de 107, 108 y 109 de Salmonella Enteritidis, pero colonizó de manera menos consistente sus tractos reproductivos. En la histopatología, se observó hiperplasia linfoide leve en el hígado y el bazo, junto con hepatitis, tiflitis, serositis y salpingitis, a los siete y 14 días posteriores a la exposición, con una mayor proporción de aves afectadas en los dos grupos de dosis más altas. En las ponedoras desafiadas no se detectaron diarrea ni cultivo de Salmonella Enteritidis de sangre colectada del corazón. El aislamiento de Salmonella Enteritidis PT 12 de Nueva Gales del Sur pudo invadir y colonizar los tractos reproductivos de las aves, así como una amplia gama de otros tejidos, lo que indica el potencial de estas gallinas comerciales sin inmunidad para contaminar sus huevos. [ABSTRACT FROM AUTHOR]

• Phage cocktails are essential for phage biocontrol, but remain difficult to design. • More research is needed to develop predictive models of phage cocktail success. • Phage cocktails must be designed to limit off-target ecosystem effects. Current practices in water and wastewater treatment to control unwanted microbes have led to new problems, including health effects from disinfection byproducts, growth of opportunistic pathogens resistant to residual disinfectants (e.g., chlorine), and antibiotic resistance. These challenges are spurring interest in rethinking our practices of microbial control. Simultaneously, advances in molecular biology and computation power are driving renewed interest in using phages (viruses that infect bacteria) to precisely control microbial growth (aka, phage biocontrol). In this Making Waves article, I begin by reviewing the current state of research into phage cocktail design, emphasizing our limited understanding of the features of successful phage cocktails (combinations of multiple types of phages). I describe the state of modeling phage-bacteria interactions and underscore the need for increasing research efforts to predict phage cocktail success, a key gap slowing the application of phage biocontrol. I also detail how research must also focus on techniques for engineering more effective phages to offer a more rapid alternative to phage discovery from natural environments. In this way, phage cocktails comprised of phages with complementary infection strategies may be designed. The final area for increased research effort that I highlight is the need for phage cocktail design to account for possible unintended environmental effects, a risk that is increasingly acknowledged in phage ecology studies but mostly ignored by those developing phage biocontrol technologies. By focusing more research effort towards the areas necessary for intelligent phage cocktail design, we can accelerate the development of phage-based biocontrol in water systems and improve public health. [Display omitted] [ABSTRACT FROM AUTHOR]

Bacteriophages are ubiquitous organisms that can be specific to one or multiple strains of hosts, in addition to being the most abundant entities on the planet. It is estimated that they exceed ten times the total number of bacteria. They are classified as temperate, which means that phages can integrate their genome into the host genome, originating a prophage that replicates with the host cell and may confer immunity against infection by the same type of phage; and lytics, those with greater biotechnological interest and are viruses that lyse the host cell at the end of its reproductive cycle. When lysogenic, they are capable of disseminating bacterial antibiotic resistance genes through horizontal gene transfer. When professionally lytic—that is, obligately lytic and not recently descended from a temperate ancestor—they become allies in bacterial control in ecological imbalance scenarios; these viruses have a biofilm-reducing capacity. Phage therapy has also been advocated by the scientific community, given the uniqueness of issues related to the control of microorganisms and biofilm production when compared to other commonly used techniques. The advantages of using bacteriophages appear as a viable and promising alternative. This review will provide updates on the landscape of phage applications for the biocontrol of pathogens in industrial settings and healthcare. [ABSTRACT FROM AUTHOR]

The ability of bacteriophage (phage), abundant within the gastrointestinal microbiome, to regulate bacterial populations within the same micro-environment offers prophylactic and therapeutic opportunities. Bacteria and phage have both been shown to interact intimately with mucin, and these interactions invariably effect the outcomes of phage predation within the intestine. To better understand the influence of the gastrointestinal micro-environment on phage predation, we employed enclosed, in vitro systems to investigate the roles of mucin concentration and agitation as a function of phage type and number on bacterial killing. Using two lytic coliphage, T4 and PhiX174, bacterial viability was quantified following exposure to phages at different multiplicities of infection (MOI) within increasing, physiological levels of mucin (0–4%) with and without agitation. Comparison of bacterial viability outcomes demonstrated that at low MOI, agitation in combination with higher mucin concentration (>2%) inhibited phage predation by both phages. However, when MOI was increased, PhiX predation was recovered regardless of mucin concentration or agitation. In contrast, only constant agitation of samples containing a high MOI of T4 demonstrated phage predation; briefly agitated samples remained hindered. Our results demonstrate that each phage–bacteria pairing is uniquely influenced by environmental factors, and these should be considered when determining the potential efficacy of phage predation under homeostatic or therapeutic circumstances. [ABSTRACT FROM AUTHOR]

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Background: Infection with viruses, bacteria, or other pathogens can lead to inflammation of the meninges. Finding the pathogen and identifying the most common type is necessary for each country. Using multi-locus sequence typing (MLST), the aim of this study was to determine the genetic relationship among S. pneumoniae isolated from CSF in children with bacterial meningitis. Materials and methods: Fourteen isolates of S. pneumoniae from CSF in children with bacterial meningitis were included in this study. The seven housekeeping genes, primer, and analysis of the sequencing used in MLST were extracted from PubMLST. Results: The sequencing analysis showed four MLST types in the studied strains. The most frequent type is ST13649 and the least frequent are ST708 and ST285. Conclusion: Finding the bacterial sequence types (ST) enables comparing the ST in different, especially neighbouring, countries. [ABSTRACT FROM AUTHOR]