Outbreaks are a risk to public health particularly when pathogenic, hypervirulent, and/or multidrug-resistant organisms (MDROs) are involved. In a hospital setting, vulnerable populations such as the immunosuppressed, intensive care patients, and neonates are most at risk. Rapid and accurate outbreak detection is essential to implement effective interventions in clinical areas to control and stop further transmission. Advances in the field of whole genome sequencing (WGS) have resulted in lowered costs, increased capacity, and improved reproducibility of results. WGS now has the potential to revolutionize the investigation and management of outbreaks replacing conventional genotyping and other discrimination systems. Here, we outline specific procedures and protocols to implement WGS into investigation of outbreaks in healthcare settings.

The study focused on the hunting practices and potentially pathogenic bacterial species among European fallow deer (Dama dama). Within a five-year period, three hunting grounds from Western Romania were examined. During this period, a total of 1881 deer were hunted, and 240 samples were collected by rectal and nasal swabbing from 120 carcasses. Bacterial strains were identified utilizing bacteriological assays and the Vitek® 2 Compact system. Notably, the Socodor hunting ground exhibited a significant difference in harvesting quotas between the bucks (Group M) and does/yearlings (Group F), favoring the latter. In the Chișineu Criș-Sălișteanca hunting ground, a likely correlation in harvesting quotas between the two groups was observed. The identified potentially pathogenic bacteria were Escherichia coli, Salmonella spp., Staphylococcus aureus, Listeria monocytogenes and Enterococcus faecium. These results highlight the importance of effectively managing the deer population and recognize the potential for Dama dama to spread zoonotic pathogens, emphasizing the necessity of adopting a One Health approach and maintaining ongoing surveillance of this game species\' population dynamics.

Bacteria (including disinfection- and antibiotic-resistant bacteria) are abundant in the consumer water cycle, where they may cause disease, and lead to biofouling and infrastructure damage in distributions systems, subsequently resulting in significant economic losses. Bacteriophages and their associated enzymes may then offer a biological control solution for application within the water sector. Lytic bacteriophages are of particular interest as biocontrol agents as their narrow host range can be exploited for the targeted removal of specific bacteria in a designated environment. Bacteriophages can also be used to improve processes such as wastewater treatment, while bacteriophage-derived enzymes can be applied to combat biofouling based on their effectiveness against preformed biofilms. However, the host range, environmental stability, bacteriophage resistance and biosafety risks are some of the factors that need to be considered prior to the large-scale application of these bacterial viruses. Characteristics of bacteriophages that highlight their potential as biocontrol agents are thus outlined in this review, as well as the potential application of bacteriophage biocontrol throughout the consumer water cycle. Additionally, the limitations of bacteriophage biocontrol and corresponding mitigation strategies are outlined, including the use of engineered bacteriophages for improved host ranges, environmental stability and the antimicrobial re-sensitisation of bacteria. Finally, the potential public and environmental risks associated with large-scale bacteriophage biocontrol application are considered, and alternative applications of bacteriophages to enhance the functioning of the consumer water cycle, including their use as water quality or treatment indicators and microbial source tracking markers, are discussed.

\'Candidatus Liberibacter asiaticus\', the putative causal agent of citrus greening disease is transmitted by the Asian citrus psyllid, Diaphorina citri in a propagative, circulative, and persistent manner. Unfortunately, \'Ca. L. asiaticus\' is not yet available in pure culture to carry out Koch\'s postulates and to confirm its etiology. When a pure culture is available, an assay to test its infectivity in both the insect vector and the plant host will be crucial. Herein, we described a transmission assay based on the use of phloem sap extracted from infected citrus plants and topical feeding to D. citri nymphs. Phloem sap was collected by centrifugation, diluted with 0.1 M phosphate buffer pH 7.4 containing 20% (w/v) sucrose and 0.1% ascorbic acid (w/v) as an antioxidant, and delivered to third through fifth instar nymphs by placing droplets on the mouthparts. Nymphs unfolded the stylets and acquired the phloem sap containing the bacterial pathogen. Nymphs were then placed onto Citrus macrophylla seedlings (ten nymphs/seedling) for an inoculation period of two weeks. A transmission rate of up to 80% was recorded at six months post-inoculation. The method could be a powerful tool to test the transmissibility of the bacterial pathogen after various treatments to reduce the viability of the bacteria or to block its transmission. In addition, it might be a potent assay to achieve Koch\'s postulates if a pure culture of \'Ca. L. asiaticus\' becomes available.

The aim of this study was to determine the level of infection of Ixodes ricinus ticks with pathogens (Borrelia spp., Rickettsia spp., and Anaplasma spp.) collected from Lacerta agilis and Zootoca vivipara lizards in the urban areas of Wrocław (SW Poland). The study was carried out in July-August 2020. Lizards were caught by a noose attached to a pole or by bare hands, identified by species, and examined for the presence of ticks. Each lizard was then released at the site of capture. Ticks were removed with tweezers, identified by species using keys, and molecular tests were performed for the presence of pathogens. From 28 lizards (17 specimens of Z. vivipara and 11 specimens of L. agilis) a total of 445 ticks, including 321 larvae and 124 nymphs, identified as I. ricinus were collected. A larger number of ticks were obtained from L. agilis compared to Z. vivipara. Molecular tests for the presence of pathogens were performed on 445 specimens of I. ricinus. The nested PCR method for the fla gene allowed the detection of Borrelia spp. in 9.4% of ticks, and it was higher in ticks from L. agilis (12.0%) than from Z. vivipara (1.0%). The RFLP method showed the presence of three species, including two belonging to the B. burgdorferi s.l. complex (B. lusitaniae and B. afzelii), and B. miyamotoi. The overall level of infection of Rickettsia spp. was 19.3%, including 27.2% in ticks collected from Z. vivipara and 17.0% from L. agilis. Sequencing of randomly selected samples confirmed the presence of R. helvetica. DNA of Anaplasma spp. was detected only in one pool of larvae collected from L. agilis, and sample sequencing confirmed the presence of (A) phagocytophilum. The research results indicate the important role of lizards as hosts of ticks and their role in maintaining pathogens in the environment including urban agglomeration as evidenced by the first recorded presence of (B) miyamotoi and (A) phagocytophilum in I. ricinus ticks collected from L. agilis. However, confirmation of the role of sand lizards in maintaining (B) miyamotoi and A. phagocytophilum requires more studies and sampling of lizard tissue.

Persistent infection caused by biofilm is an urgent in medicine that should be tackled by new alternative strategies. Low efficiency of classical treatments and antibiotic resistance are the main concerns of the persistent infection due to biofilm formation which increases the risk of morbidity and mortality. The gene expression patterns in biofilm cells differed from those in planktonic cells. One of the promising approaches against biofilms is nanoparticle (NP)-based therapy in which NPs with multiple mechanisms hinder the resistance of bacterial cells in planktonic or biofilm forms. For instance, NPs such as silver (Ag), zinc oxide (ZnO), titanium dioxide (TiO2), copper oxide (Cu), and iron oxide (Fe3O4) through the different strategies interfere with gene expression of bacteria associated with biofilm. The NPs can penetrate into the biofilm structure and affect the expression of efflux pump, quorum-sensing, and adhesion-related genes, which lead to inhibit the biofilm formation or development. Therefore, understanding and targeting of the genes and molecular basis of bacterial biofilm by NPs point to therapeutic targets that make possible control of biofilm infections. In parallel, the possible impact of NPs on the environment and their cytotoxicity should be avoided through controlled exposure and safety assessments. This study focuses on the biofilm-related genes that are potential targets for the inhibition of bacterial biofilms with highly effective NPs, especially metal or metal oxide NPs.

It is essential to have a thorough knowledge of the genetic variation among different strains of Xanthomonas citri pv. citri (Xcc), which is responsible for causing citrus bacterial canker. This understanding is important for studying disease characteristics, population structure, and evolution and ultimately for developing sustainable methods of control. A total of 48 strains obtained from citrus production areas in Burkina Faso in 2012, 2020, and 2021 were subjected to polymerase chain reaction (PCR) tests using specific primers. The aim was to examine the distribution of type 3 effectors (T3E) and determine the geographical origins of the strains. The examination of the distribution of type 3 non-transcription-activator-like effectors (TALEs) revealed a broader range of strains obtained in 2020 and 2021 than in 2012. However, all the strains possessed a shared set of three genes, specifically, XopE2, XopN, and AvrBs2. Furthermore, all examined effectors were observed in the Bobo-Dioulasso region. Regarding the characterization of TALEs, two profiles containing two to three TALEs were discovered. Profile 1, consisting of two TALEs, was found in 37 Xcc strains, whereas Profile 2, comprising three TALEs, was detected in 11 strains. Among the three TALEs (A, B, and C) that were identified, TALEs B and C were present in all the strains. The correlation matrix indicated a positive association between the T3E content of strains and the duration of their isolation. Principal component analysis revealed a limited organization of the strains under investigation.

Rapid detection of plant diseases before they escalate can improve disease control. Our team has developed rapid nucleic acid extraction methods with microneedles (MN) and combined these with LAMP assays for pathogen detection in the field. In this work, we developed LAMP assays for early blight (Alternaria linariae, A. alternata, and A. solani) and bacterial spot of tomato (Xanthomonas perforans) and validated these LAMP assays and two previously developed LAMP assays for tomato spotted wilt virus and late blight. Tomato plants were inoculated and disease severity was measured. Extractions were performed using MN and LAMP assays were run in tubes (with hydroxynaphthol blue) on a heat block or on a newly designed microfluidic slide chip on a heat block or a slide heater. Fluorescence on the microfluidic chip slides was visualized using EvaGreen and photographed on a smartphone. Plants inoculated with X. perforans or tomato spotted wilt virus tested positive prior to visible disease symptoms, while P. infestans and A. linariae were detected at the time of visual disease symptoms. LAMP assays were more sensitive than PCR and the limit of detection was 1 pg of DNA for both A. linariae and X. perforans. The LAMP assay designed for early blight detected all three species of Alternaria that infect tomato and is thus an Alternaria spp. assay. This study demonstrates the utility of rapid MN extraction followed by LAMP on a microfluidic chip for rapid diagnosis of four important tomato pathogens.

Bacterial spot caused by Xanthomonas spp. is an economically important disease of pepper causing significant yield losses in Taiwan. Monitoring the pathogen population on a continuous basis is necessary for developing disease management strategies. We analyzed a collection of xanthomonad strains isolated from pepper in Taiwan between 1989 and 2019. Among the sequenced genomes, sixty-five were identified as Xanthomonas euvesicatoria and ten were X. perforans. Thirty-five X. euvesicatoria and ten X. perforans strains were copper tolerant, whereas only five X. euvesicatoria and none of the X. perforans strains were tolerant to streptomycin. Nine X. euvesicatoria strains were amylolytic, which is considered an unusual characteristic for X. euvesicatoria. Bayesian analysis of the population structure based on core gene SNPs clustered the strains into five clusters for X. euvesicatoria and three clusters for X. perforans. One X. perforans cluster, designated as TP-2019, appears to be a novel genetic cluster based on core genes, accessory gene content, and effector profile. This knowledge of pathogen diversity with whole genomic information will be useful in future comparative studies and in improving breeding programs to develop disease-resistant cultivars and other disease management options.

Antimicrobial resistance (AMR), caused by microbial infections, has become a major contributor to morbid rates of mortality worldwide and a serious threat to public health. The exponential increase in resistant pathogen strains including Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) poses significant hurdles in the health sector due to their greater resistance to traditional treatments and medicines. Efforts to tackle infectious diseases caused by resistant microbes have prompted the development of novel antibacterial agents. Herein, we present selenium and copper oxide monometallic nanoparticles (Se-MMNPs and CuO-MMNPs), characterized using various techniques and evaluated for their antibacterial potential via disc diffusion, determination of minimum inhibitory concentration (MIC), antibiofilm, and killing kinetic action. Dynamic light scattering (DLS), scanning electron microscopy (SEM/EDX), and X-ray diffraction (XRD) techniques confirmed the size-distribution, spherical-shape, stability, elemental composition, and structural aspects of the synthesized nanoparticles. The MIC values of Se-MMNPs and CuO-MMNPs against S. aureus and E. coli were determined to be 125 μg/mL and 100 μg/mL, respectively. Time-kill kinetics studies revealed that CuO-MMNPs efficiently mitigate the growth of S. aureus and E. coli within 3 and 3.5 h while Se-MMNPs took 4 and 5 h, respectively. Moreover, CuO-MMNPs demonstrated better inhibition compared to Se-MMNPs. Overall, the proposed materials exhibited promising antibacterial activity against S. aureus and E. coli pathogens.