Introduction: Giardia lamblia is a flagellated protozoan and the etiological agent of giardiasis, a leading cause of epidemic and sporadic diarrhoea globally. The clinical significance and public health impact of giardiasis underscores the need for robust methodologies to investigate and manage this pathogen. This study systematically reviews the main methodologies described in the literature for studying the life cycle of G. lamblia, focusing on isolation, purification, axenization, excystation, and encystation processes. Methods: A systematic literature review was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA 2020) statement. The search was performed in the MEDLINE, ScienceDirect, and Web of Science databases using the following search equation: “("axenization" OR "isolation" OR "excystation" OR "encystation" OR "purification") AND ("method") AND ("giardia")". Results: A total of 39 studies were included in the review, revealing 56 methods for isolation and purification, 7 methods for excystation, and 3 methods for axenization and encystation. Isolation and purification methods exhibited significant variability, often involving two phases: an initial separation using simple techniques such as filtration and centrifugation, followed by a purification phase using a density gradient for faecal samples and immunomagnetic separation for water samples or nucleic acid extraction. The effectiveness of these methods differed depending on the source and sample type, highlighting the need for standardized protocols to ensure consistent and reliable results. Conclusion: Methods for the isolation and purification of G. lamblia exhibit notable variability and lack uniformity compared with the more consistent methods that re used for other life cycle stages. These findings underscore the urgent need for the development of standardized methodologies to enhance the reproducibility and reliability of research outcomes in this field.

Alphaviruses cause mosquito-borne erythrogenic diseases. Sindvis virus (SINV) circulates in an enzootic cycle between its vectors and birds. In humans, SINV triggers musculoskeletal syndromes characterised by fever, rash, acute and chronic polyarthritis, etc. Few studies examining the virus's impact on the nervous/immune systems are available. SINV often causes cerebral infection, so understanding the brain's response is crucial. The SH-SY5Y cell line, from neuroblastoma, is used in many neurobiological and virological studies. This study aimed to understand viral infection in vitro and examine immune markers using SINV infection in SH-SY5Y cells.

Due to the lack of adequate knowledge of SINV, optimization was necessary for all experimental designs. After the virus stock preparation, the median tissue culture infectious dose was used to determine the multiplicity of infection (MOI). Taqman-probe-based RT-qPCR was designed for the SINV nucleic acid, and immunofluorescence-based ds-RNA antibody staining was used to reinforce the virus replication. Apoptotic markers were also investigated by means of immunofluorescence. To monitor specific immune gene expressions (pattern recognition receptors (PRRs), regulator gene, cytokines) in a time-dependent manner (3/6/12/16/24/30 hr), RT-qPCR was also performed.

SINV infected the cells, and after 24 hr, a cytopathogenic effect and virus replication could be observed. SINV induced the caspase-dependent apoptotic pathway. Most of the investigated genes, including PRRs, (TLR-3/7, RIG1/MDA5), a regulator (b-catenin), and inflammatory (IL-1b, IL-6, TNFa), and antiviral (IL-10, IFNβ) genes, exhibited a consistent induction up to 12/16 hr; by the end of the 30 hr, their expression mainly (e.g., RIG1/MDA5, IFNβ) decreased compared with those of our controls. We can conclude that in vitro, SINV can be transmitted even in small quantities, and an inflammatory environment can be developed, which later leads to cell death.

These preliminary results allow us to gain insights into SINV–SH-SY5Y interactions, which are better understood by examining them in much finer molecular detail.

Arnica montana L. is renowned for its biological properties, including its antimicrobial effects, which are attributed to the presence of sesquiterpene lactones or flavonoids. The aim of this study was to assess the antimicrobial activity of ethanolic (EtE) and aqueous (AqE) extracts of Arnica montana L. against a reference strains of microbes, employing a double broth dilution method (concentration of 7.81-30000 µg/mL); identifiyng MICs and MBC/MFCs. Amoxicillin, nystatin, and 70% ethanol were used as positive controls. The MBC(MFC)/MIC ratios were used to assess the activity of EtE/AqE (<4 bactericidal/fungicidal; ≥4 -static). As a result, a differential antimicrobial activity was demonstrated in MIC=1000-2000 µg/mL and MIC=8000-16000 µg/mL for Gram(+) bacteria for EtE and AqE, respectively. EtE was more effective against Salmonella Typhimurium, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Proteus mirabilis (MIC=1000-2000 µg/mL), compared to AqE (MIC>16000 µg/mL). EtE was bactericidal against staphylococci and Enterobacteriaceae rods, while AqE was bactericidal against Bacillus cereus. Unlike EtE, which was effective to most Candida spp. (MIC=500-800 µg/mL), AqE had no antifungal properties.

This study suggests that the extracts and essences of A. montana have several beneficial properties such as antibacterial and antifungal effects. It highlights that essence extracts (EtEs) are more potent than aqueous extracts (AqEs) and can serve as natural antimicrobial agents. The raw material is mainly used in topical preparations due to its high toxicity, but can also be taken orally in low concentrations as a homeopathic remedy. The results of this study may pave the way for further research into activity against other microbial strains, and the development of new formulations for different applications. However, the challenge is to optimise solvent concentrations that are most suitable for biological experiments, particularly for alcohol extracts, which are good for solubilisation but can be cytotoxic if not carefully controlled.

Conventional pesticides commonly used in modern agriculture pose significant risks to plant health by disrupting soil microbiota, including beneficial plant growth-promoting bacteria. These non-target effects contribute to long-term soil degradation and ecological imbalances, ultimately undermining plant productivity and sustainability. Consequently, there is an urgent need to identify safer and more sustainable pest management alternatives that support plant health. Volatile phytochemicals, known for their bioactivity against plant pests, offer a promising solution due to their reduced environmental impact and lower toxicity.

This study evaluated the toxicity of dodecan-1-ol, a volatile compound with nematicidal properties, on crop-associated bacterial species of importance for plant growth and soil health: Bacillus megaterium, Pseudomonas azotoformans, Paenibacillus zeisoli, and Burkholderia phymatum. Direct-contact bioassays in 96-well microplates were used to monitor bacterial growth spectrophotometrically. For comparison, the conventional pesticides emamectin benzoate and oxamyl were tested under identical conditions.

Dodecan-1-ol inhibited bacterial growth by a maximum of 20%, significantly lower than the 93% and 11% inhibition observed with emamectin benzoate and oxamyl, respectively. These results were obtained at a concentration of 0.02 mg/mL for all compounds. Importantly, at this concentration, dodecan-1-ol achieves its lowest maximal effective concentration (EC₁₀₀) against plant-parasitic nematodes, whereas emamectin benzoate and oxamyl exhibit limited nematicidal efficacy and require substantially higher application rates in agricultural settings.

By combining effective nematicidal activity with minimal impact on plant-beneficial bacteria, this study highlights the potential of dodecan-1-ol to support plant health while addressing pest control needs. These findings emphasize its role as a safer and more sustainable alternative to conventional pesticides, with promising applications in integrated pest and soil management strategies to promote healthy crop growth and long-term agricultural sustainability.

Introduction: Our systematic review and meta-analysis address gaps in UV-C viral inactivation protocols in liquid solutions to understand both the influencing characteristics of viruses and related experimental setups. Key issues include inconsistent reporting of experimental setups, virus identification, terminology, and dose–effectiveness relationships, which have all bee examined in the literature (Chiappa et al., 2021; Freeman et al., 2022; Masjoudi et al., 2021; Raeiszadeh & Adeli, 2020). A clear distinction between "inactivation" and "disinfection" is crucial. We propose defining complete inactivation as an undetectable viral load while preserving viral and protein structures to improve research applications.

Methods: A PICO search (April 10, 2024) across five databases (N=2814) yielded 1,686 records after duplicates were removed adhering to the PRISMA, GRADE, and MECIR protocols. We reviewed 309 full texts, with 33 studies meeting our inclusion criteria for strain-specific viruses, defined UV-C setups, and quantifiable viral reductions. Random-effects meta-regression of 18 studies analyzed strain-specific inactivation, while Spearman correlation was used to examine terminology usage.

Results: UV-C effectively (254nm being the most effective) reduced viral loads but faced inconsistencies in experimental setups, viral conditions, and dosages. Virus-specific dose–response relationships varied, leading us to focus on only SARS-CoV-2 strains for further calculations. Our semantic analysis revealed the interchangeable use of "inactivation" and "disinfection". The issues identified in our meta-analysis—0-0 distortions, dose saturation, and PFU/TCID inconsistencies—necessitate non-linear models and data refinement. Despite the variability, the preliminary regression (R² = 0.49) suggested predictable outcomes.

Conclusions: We propose a reporting framework to improve reliability and cross-study comparability. Inactivation studies must start with higher viral loads to account for plaque assay and TCID₅₀ detection limits, and the “delivered dose” should only be reported when measured; otherwise, “calculated dose” is correct. Reports must include TCID/plaque assay details, log-transformed data (e.g., log₃ = base 3), and at least five data points for inactivation curves. "Inactivation" should be used for laboratory studies maintaining viral structures, while "disinfection" suits hospitals and water treatments.

Introduction

Nitrogen fertilization boosts crop yields but is inefficient due to the rapid activity of nitrifying bacteria, which leads to the loss of useful nitrogen forms. This requires the use of large amounts of fertilizers, leading to environmental pollution from compounds like NO₃ and N₂O. Although chemical inhibitors can improve fertilization, their impact on the environment and human health remains uncertain. A new strategy involves the use of bacteriophages (phages) that specifically target nitrifying bacteria.

Methods

Bacteriophages were isolated from sewage samples. The inhibition of nitrifying activity was tested in AOB medium and soil to monitor nitrite production with a colorimetric assay. The increase in bacteriophage particles during infection was assessed with qPCR. Controls without the addition of bacteriophages or bacteria were included for in vitro and soil samples. The persistence of infectious bacteriophages was analyzed in water suspensions and soil under different pH levels (3, 5, 6, 7, 8) and temperatures (4, 15, 22, 37 ^◦C) over 6 months.

Results

We isolated a bacteriophage (FNF-1) capable of infecting four species of Nitrosomonas. FNF-1 inhibited bacterial growth, resulting in a reduction in nitrite formation in phage-treated samples, both in vitro and in the soil. Meanwhile, FNF-1 increased by 2-log₁₀ units within 48 hours. Infectious FNF-1 remained stable in water suspensions for 6 months at 4, 15, 22, and 37 ^◦C and at pHs 6, 7, and 8 but was inactivated when exposed to pHs 3 and 5. In soils, FNF-1 infectivity showed a 2-log₁₀ reduction over 6 months.

Conclusions

This study presents a novel approach using bacteriophages to eliminate nitrifying bacteria. Bacteriophages as biocontrol agents offer an effective and environmentally friendly alternative to inhibit nitrification while overcoming the limitations of chemical products. While their application shows potential for improving agricultural fertilization, further research is required to scale up and optimize a product.

Fermented foods have been rapidly gaining popularity recently due to their use in medicine and dietetics. Kombucha is a fermented beverage containing numerous probiotic bacteria and their metabolites, which have beneficial effects on the digestive system. The purpose of this study was to determine the antimicrobial activity of kombucha containing probiotic strains against bacteria that cause gastrointestinal diseases.

The activity of kombucha was tested before fermentation and during fermentation on days 1, 2, 3, 7, 8, and 14 against reference strains of Gram-positive bacteria from the American Type Culture Collection (ATCC) such as methicillin-sensitive Staphylococcus aureus (MSSA) ATCC 29213, two methicillin-resistant Staphylococcus aureus (MRSA) strains (ATCC 43300 and ATCC BAA-1707), Bacillus cereus ATCC 10876, and Enterococcus faecalis ATCC 29212. The well-diffusion method was used for this assessment, and the antimicrobial activity of this beverage was evaluated by the zone of growth inhibition (mm) around the kombucha well.

The highest activity was observed after 14 days of kombucha fermentation against almost all tested bacteria with a zone of growth inhibition of 18-21 mm. The highest activity was observed against S. aureus ATCC 29213 and S. aureus ATCC 43300.

These data show that kombucha is a beverage that can support the treatment of food poisoning caused by pathogenic microorganisms such as methicillin-sensitive S. aureus (MRSA), methicillin-resistant S. aureus (MRSA), B. cereus, and E. faecalis.

Animal husbandry is a key promoter of antibiotic resistance (AMR) and contributes to the spread of AMR genes throughout the environment and food supply. The pig industry is one of the leading sectors in animal-based food production worldwide, with Brazil ranking fourth in terms of pork production and export. The emergence of extended-spectrum β-lactamase (ESBL)-producing bacteria in this sector represents a significant public health concern. These enzymes confer resistance to a broad range of β-lactams, threatening the effectiveness of antibiotic therapy for infections in both humans and animals. A systematic review was conducted to address the current knowledge of AMR within the Brazilian pig production chain. The presence of ESBL-encoding genes in bacteria isolated from pig feces, intestine, carcass, and urine, as well as pork-based food, was investigated. TEM-type genes were detected in 11 of the 12 studies, including blaTEM-1A (n=33 strains) and blaTEM-1B (n=44 strains). Both genes were found in Salmonella isolated from all sample categories. Additionally, blaTEM-1 was detected in E. coli (n=4 strains from feces) and Salmonella spp. (n=2 from carcass; n=5 from feces). CTX-type genes were detected in Proteus mirabilis (blaCTX-M-65; n=6 from food) and Salmonella (blaCTX-M-8; n=1 from feces; n=1 from pig carcass). blaCMY-2 was detected in Salmonella from feces (n=1) and carcass (n=1), whereas blaCMY-M2 was detected in E. coli (n=5) from pig feces. blaPSE-1 and blaSHV-1 genes were investigated using PCR in E. coli strains isolated from pig feces; however, these were not detected. In conclusion, the pig industry may contribute to the spread of ESBL-encoding genes, posing a public health risk. Data reveal a significant presence of blaTEM genes across various samples, alongside other ESBL-encoding gene types in the Brazilian pig production chain. These findings highlight the need for integrated strategies to control AMR, particularly to reduce risks that may arise from animal -based food production systems.

Residual antibiotics in swine wastewater (SW) promote antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs), threatening the environment and human health. Anaerobic digestion (AD) is a promising green technology for treating SW, but the impact of antibiotics on the microbial community and resistome during AD is unclear.

To address this gap, anaerobic batch experiments treating SW containing different sulfonamides (SMs) were conducted, with samples collected for DNA extraction and analysis of the microbial community and antibiotic resistome. ARG-carrying mobile genetic elements (MGEs) were detected, and ARG hosts were identified using metagenomic binning.

The results showed a significant negative correlation between the Abundance-based Coverage Estimator (ACE) index and the batches, with notable variability in the Shannon index. Enterococcus emerged as the dominant genus under SMs stress. A total of 24 types and 440 subtypes of ARGs were identified, with SMs stress significantly increasing the abundance of sul1, sul2, and sul3. The highest ARG abundance was observed in the sulfamonomethoxine (SMM) group during the initial stage and in the sulfamerazine (SMR) group after long-term anaerobic culture. Plasmid identification revealed that ARGs on chromosomes are much higher than those on plasmids. However, ARGs are more likely to co-occur on plasmids. Hybrid assembly identified two complete integrons and two composite transposons. Binning reconstructed 34 and 33 medium- to high-quality ARGs host genomes from all samples, determining their taxonomic classification and potential pathogenicity.

In conclusion, SMs stress intermittently disrupts the microbial community structure during AD. ARG composition and distribution are influenced by antibiotic pressure and initial ARGs in seed sludge. Prolonged antibiotic exposure increases the risk of ARG mobilization via MGEs. Specific ARB without ARGs but with inherent resistance also require attention. This study offers new insights into the occurrence and impact of antibiotics during the AD treatment of SW.

INTRODUCTION

Natural products are an important source of therapeutic agents for their antimicrobial and prebiotic properties in an era of increasing antibiotic resistance. The antimicrobial and prebiotic properties of honeys from hawthorn, cherries, raspberries, almonds and apples were investigated.

METHODS

The antimicrobial activity of the honey, diluted 1:1 w: vol with deionized water and filtered, was assessed (20 mg/ml) by evaluating, through the crystal violet test, its ability to inhibit the biofilm of the high-risk pathogen Acinetobacter baumannii, and, through the MTT test, its capacity to affect the metabolism of the A. baumannii sessile cells. The prebiotic potentiality of the honey was assessed on Lactiplantibacillus plantarum and Lacticaseibacillus rhamnosus, to evaluate the capacity to affect their growth and to inhibit the biofilm of that pathogen.

RESULTS

The honeys inhibited the biofilm of A. baumannii with percentages ranging from 43.49% (cherry) to 59.43% (apple). The honeys did not affect the metabolism of the sessile cells of the pathogen (except cherry honey, 27.16%), so other inhibition mechanisms should be investigated. Rosaceae honey positively affected the growth of the two lactobacilli, with an increase ranging from 8-fold to even 13-fold (when L. rhamnosus grew in the presence of cherry honey), compared to the control, grown in MRS medium. The biofilm inhibitory activity of the supernatants of lactobacilli showed that all types of honey were more effective than the control, with inhibition percentages of up to 34.24% (supernatant of L. plantarum grown with apple honey), and was especially evident on the metabolism of sessile cells, as demonstrated by the high inhibition percentages observed (up to 61.31%), higher than the supernatant of the two strains conventionally grown.

CONCLUSIONS

Future work will focus on the effect of Rosaceae honey on other pathogenic bacteria and probiotics, with the aim of expanding knowledge of its biological properties and assessing the consequent possibilities of its use.