Viable State Research Articles

Quantitative Detection of VBNC State Pseudomonas aeruginosa Contributing to Accurate Assessment of Microbial Inactivation in Drinking Water Disinfection

In harsh environments, bacteria often enter a viable but nonculturable (VBNC) state, which cannot be detected using heterotrophic plate counting (HPC). Importantly, VBNC bacteria can potentially resuscitate under favorable conditions, posing a risk to drinking water safety. This study introduces an innovative approach, combining improved quantitative polymerase chain reaction (qPCR) with propidium monoazide (PMA) dye and HPC to accurately quantify VBNC Pseudomonas aeruginosa (P. aeruginosa). The method was applied to assess the ability of various disinfection techniques to induce P. aeruginosa into the VBNC state. Different disinfection methods, including ultraviolet radiation (UV), sodium hypochlorite (NaClO), and peracetic acid (PAA), significantly reduced bacterial culturability (>99.9%), with the majority entering the VBNC state. Notably, under favorable conditions, UV-induced VBNC cells were resuscitated faster than those induced by NaClO. VBNC P. aeruginosa exhibited relatively high intracellular adenosine triphosphate (ATP) levels, indicating ongoing metabolic activity. Scanning electron microscopy (SEM) reveals that some bacteria maintained cellular integrity for UV and PAA treatment, while evident membrane disruption was observed after NaClO disinfection. This study represents a significant advancement in quantitatively detecting VBNC state P. aeruginosa, contributing to an accurate assessment of microbial inactivation during drinking water disinfection.

Ultrasound-assisted activation of PAW residual radicals in the concurrent elimination of ARB and ARGs: Process efficiency, mechanism and implication

Methicillin-resistant Staphylococcus aureus (MRSA) and antibiotic resistance gene (mecA) are considered emerging contaminants that have been extensively identified in aqueous environments due to their high resistance to disinfectants. Herein, we report for the first time the inactivation of MRSA through a sequential application of plasma-activated water and ultrasound (PAW-SU). While the impact of PAW alone on bacterial culturability and mecA integrity was negligible, the combined PAW-SU treatment outperformed the inactivation method involving PAW with direct US (PAW-DU). It exhibited a shorter lag phase and a higher maximum inactivation rate constant. We numerically demonstrated that sonolysis processes induced by external ultrasound sources considerably improved the intracellular and extracellular reactive oxygen and nitrogen species (RONS) of MRSA in PAW system, thereby increasing the PAW reactivity and applicability. The highest inactivation efficiency of PAW-SU can be attributed to the synergistic effect of enhanced intracellular RONS (i.e., peroxynitrite and singlet oxygen) and membrane disruption induced by US, resulting in irreparable oxidative damage to the Save our Soul response and energy metabolic system. In particular, the elevated arginine metabolites in MRSA cells were primarily responsible for the formation of the viable but nonculturable state (VBNC) bacteria associated with high resistance to PAW system. Surprisingly, a decrease in arginine synthesis capacity was observed to expedite the flow of the tricarboxylic acid cycle, accelerating the detrimental effects of PAW-SU on intracellular components in VBNC bacteria. Finally, the limited regrowth potential of injured MRSA caused by cyclic inactivation of PAW-SU process was further confirmed in wastewater from meat and dairy models and therefore, advocates the possible real-world applications.

Intestinal Translocation of Live Porphyromonas gingivalis Drives Insulin Resistance.

Periodontitis has been emphasized as a risk factor of insulin resistance-related systemic diseases. Accumulating evidence has suggested a possible "oral-gut axis" linking oral infection and extraoral diseases, but it remains unclear whether periodontal pathogens can survive the barriers of the digestive tract and how they play their pathogenic roles. The present study established a periodontitis mouse model through oral ligature plus Porphyromonas gingivalis inoculation and demonstrated that periodontitis aggravated diet-induced obesity and insulin resistance, while also causing P. gingivalis enrichment in the intestine. Metabolic labeling strategy validated that P. gingivalis could translocate to the gastrointestinal tract in a viable state. Oral administration of living P. gingivalis elicited insulin resistance, while administration of pasteurized P. gingivalis had no such effect. Combination analysis of metagenome sequencing and nontargeted metabolomics suggested that the tryptophan metabolism pathway, specifically indole and its derivatives, was involved in the pathogenesis of insulin resistance caused by oral administration of living P. gingivalis. Moreover, liquid chromatography-high-resolution mass spectrometry analysis confirmed that the aryl hydrocarbon receptor (AhR) ligands, mainly indole acetic acid, tryptamine, and indole-3-aldehyde, were reduced in diet-induced obese mice with periodontitis, leading to inactivation of AhR signaling. Supplementation with Ficz (6-formylindolo (3,2-b) carbazole), an AhR agonist, alleviated periodontitis-associated insulin resistance, in which the restoration of gut barrier function might play an important role. Collectively, these findings reveal that the oral-gut translocation of viable P. gingivalis works as a fuel linking periodontitis and insulin resistance, in which reduction of AhR ligands and inactivation of AhR signaling are involved. This study provides novel insight into the role of the oral-gut axis in the pathogenesis of periodontitis-associated comorbidities.

Unveiling the PCB biodegradation potential and stress survival strategies of resuscitated strain Pseudomonas sp. HR1

The exploration of resuscitated strains, facilitated by the resuscitation promoting factor (Rpf), has substantially expanded the pool of cultivated degraders, enhancing the screening of bio-inoculants for bioremediation applications. However, it remains unknown whether these resuscitated strains can re-enter the viable but non-culturable (VBNC) state and the specific stress conditions that trigger such a transition. In this work, the whole genome, and polychlorinated biphenyl (PCB)-degrading capabilities of a resuscitated strain HR1, were investigated. Notably, the focus of this exploration was on elucidating whether HR1 would undergo a transition into the VBNC state when exposed to low temperature and PCBs, with and without the presence of heavy metals (HMs). The results suggested that the resuscitated strain Pseudomonas sp. HR1 harbored various functional genes related to xenobiotic biodegradation, demonstrating remarkable efficiency in Aroclor 1242 degradation and strong resistance against stress induced by low temperature and PCBs. Nevertheless, when exposed to the combined stress of low temperature, PCBs, and HMs, HR1 underwent a transition into the VBNC state. This transition was characterized by significant decreases in enzyme activities and notable changes in both morphological and physiological traits when compared to normal cells. Gene expression analysis revealed molecular shifts underlying the VBNC state, with down-regulated genes showed differential expression across multiple pathways and functions, including oxidative phosphorylation, glycolysis, tricarboxylic acid cycle, amino acid metabolism, translation and cytoplasm, while up-regulated genes predominantly associated with transcription regulation, membrane function, quorum sensing, and transporter activity. These findings highlighted the great potential of resuscitated strains as bio-inoculants in bioaugmentation and shed light on the survival mechanisms of functional strains under stressful conditions, which should be carefully considered during bioremediation processes.

Stress response of Salmonella Newport with various sequence types toward plasma-activated water: Viable but nonculturable state formation and outer membrane vesicle production

This study aims to investigate the response of Salmonella Newport to plasma-activated water (PAW), a novel disinfectant that attracts attention due to its broad-spectrum antimicrobial efficacy and eco-friendliness. In this work, we demonstrated that S. Newport of different sequence types (STs) could be induced into the viable but nonculturable (VBNC) state by PAW treatment. Notably, a remarkable 99.96% of S. Newport ST45 strain entered the VBNC state after a 12-min PAW treatment, which was the fastest observed among the five S. Newport STs (ST31, ST45, ST46, ST166, ST2364). Secretion of outer membrane vesicles was observed in ST45, suggesting a potential strategy against PAW treatment. Genes related to oxidative stress (sodA, katE, trxA), outer membrane proteins (ompA, ompC, ompD, ompF) and virulence (pagC, sipC, sopE2) were upregulated in the PAW-treated S. Newport, especially in ST45. A reduction of 38–65% in intracellular ATP level after PAW treatment was observed, indicating a contributor to the formation of the VBNC state. In addition, a rapid method for detecting the proportion of VBNC cells in food products based on pagC was established. This study contributes to understanding the formation mechanism of the VBNC state in S. Newport under PAW stress and offers insights for controlling microbial risks in the food industry.

Сирийская экономика и санкции: ситуативная адаптация

War, loss of control over part of the territory, and economic sanctions have had a devastating effect on Syria’s economy. However, the country has managed to survive. This has been possible in part due to the measures of the Syrian government, private business, and the population to resist and adapt to economic sanctions. The adaptation strategy had not been planned, and the measures the state, private business, and the population have taken were not coordinated, unlike, for example, in Iran’s case, where strategies are developed and then announced based on future economic risks. Because of this, this strategy of counteraction and adaptation to economic sanctions may be termed as adaptive. The most serious economic sanctions on the Syrian economy have been imposed by the US, the EU, and the League of Arab States. They have restricted Syrian access to goods, technology, financial instruments and products, investments, and even in some cases humanitarian aid from the countries that initiated the sanctions. It is a very difficult task to identify the real Syrian losses caused by the economic sanctions, but it can be accurately asserted, based on the analysis in this article, that they do not allow the country to move forward to the recovery phase. State measures of adaptation to economic sanctions were supposed to become a buffer that would allow the national economy not only to survive, but also to move from stagnation to economic growth. However, it has to be stated that the situational strategy of adaptation to economic sanctions allowed the national economy to remain in a viable state, however, it does not allow it to move from recession to growth.

Preservación pulmonar: trasplantes sin límite de tiempo ni distancia

One of the great obstacles for organ transplantation is the time in which the ischemic organ can be preserved in a viable state before being implanted. With the use of the standard static cold preservation technique (4°C) preservation times are limited to 6-8hours, a time known as cold ischemia time. This begins from the moment the “cross-clamp” or complete cessation of circulation in the donor is performed until the organ is irrigated in the recipient.Preclinical and clinical studies have recently been published demonstrating benefits of graft conservation at 10°C compared to standard storage, in addition to allowing ischemia times to be prolonged with the same safety standards for the recipient. These new lines of development aim to facilitate the logistics of lung transplantation, transforming it from an emergency surgery to a semi-elective surgical procedure, a change that has demonstrated benefits in surgical safety and even in implant survival results.

A Nation’s Right to Exist

In justifying Russian aggression against Ukraine, President Vladimir Putin asserts that Ukraine is neither a distinct nation nor a viable state. In response, this essay will establish a Christian account of Ukraine’s right to self-defense not only via just war criteria but also in relation to its purpose theologically as a nation-state. This essay, after reviewing Christian ethical positions that either reject or embrace the nation-state, draws on the Niebuhr brothers and Karl Barth to develop key theological criteria for judging whether particular nation-states are true to their divine purpose. While affirming the role of the nation-state in protecting a people’s distinct identity, Christians will call on the nation-state to direct its people outward, toward their shared humanity with the world’s other nations. The essay concludes that the Ukrainian nation-state is fulfilling these criteria, whereas the Russian nation-state is not.

Umbilical Vascular Thromboembolism: High-Risk Factors, Diagnosis, Management, and Pregnancy Outcomes: A Scoping Review.

Umbilical vascular thromboembolism is a rare condition that can lead to serious consequences such as fetal hypoxia, fetal growth restriction, and even stillbirth. However, there is currently a lack of research on the pathology, pathogenesis, clinical management, and prognosis of this condition. Therefore, the purpose of this article is to analyze this condition's high-risk factors, clinical characteristics, pregnancy management, and discuss its corresponding pregnancy outcomes. Databases such as PubMed are searched using the relevant keywords of umbilical vascular thromboembolism in worldwide. And related information is analyzed such as maternal risk factors, fetal risk factors, umbilical cord and placental risk factors, and pregnancy outcomes. The literature search yields 113 articles, 64 of which meet the inclusion criteria for umbilical vascular thromboembolism. There are 4 retrospective cohort studies and 8 case series, the rest are all case reports. A total of 262 cases of umbilical vascular thromboembolism are found. The most common maternal complications and fetal related risk factors are diabetes (25 cases, 9.5%) and stillbirths (106 cases, 40.5%), respectively. Among these 262 cases, 98 (37.4%) cases are found by prenatal ultrasound to have umbilical vascular thromboembolism and the fetus is in a viable state with complete clinical information. In addition, considering the effectiveness and safety of low molecular weight heparin in thromboembolic conditions, twenty-four patients of umbilical artery thromboembolism attempted to use low molecular weight heparin during observation. Maternal diabetes was the highest risk factor for this condition. When umbilical artery thromboembolism occurs, the incidence of stillbirth increases. Premature patients with this condition can continue their pregnancy under close external monitoring. However, due to the small sample size, further research is needed.

Unveiling the mechanisms of black phosphorus nanosheets-induced viable but non-culturable state in Bacillus tropicus

The release of black phosphorus (BP) nanosheets has raised concerns regarding potential ecological risks. Previous studies have confirmed their toxicity to bacteria, but discrepancies were observed between results obtained from...

The Viable but Non-Culturable (VBNC) State, a Poorly Explored Aspect of Beneficial Bacteria.

Many bacteria have the ability to survive in challenging environments; however, they cannot all grow on standard culture media, a phenomenon known as the viable but non-culturable (VBNC) state. Bacteria commonly enter the VBNC state under nutrient-poor environments or under stressful conditions. This review explores the concept of the VBNC state, providing insights into the beneficial bacteria known to employ this strategy. The investigation covers different chemical and physical factors that can induce the latency state, cell features, and gene expression observed in cells in the VBNC state. The review also covers the significance and applications of beneficial bacteria, methods of evaluating bacterial viability, the ability of bacteria to persist in environments associated with higher organisms, and the factors that facilitate the return to the culturable state. Knowledge about beneficial bacteria capable of entering the VBNC state remains limited; however, beneficial bacteria in this state could face adverse environmental conditions and return to a culturable state when the conditions become suitable and continue to exert their beneficial effects. Likewise, this unique feature positions them as potential candidates for healthcare applications, such as the use of probiotic bacteria to enhance human health, applications in industrial microbiology for the production of prebiotics and functional foods, and in the beer and wine industry. Moreover, their use in formulations to increase crop yields and for bacterial bioremediation offers an alternative pathway to harness their beneficial attributes.

Transcriptomic analysis of Lacticaseibacillus paracasei Zhang in transition to the viable but non-culturable state by RNA sequencing.

Some bacteria enter the viable but non-culturable (VBNC) state to survive harsh environmental conditions and external stresses. This alters cell physiology and has implications for the food industry as some bacteria, such as lactobacilli, undergo similar changes during food processing. This study aimed to investigate the transcriptomic changes of a probiotic strain, Lacticaseibacillus paracasei Zhang (L. paracasei Zhang), upon transition to the VBNC state using high throughput RNA sequencing (RNA-seq). Bacteria were inoculated into the de Man, Rogosa, and Sharpe medium and maintained at low temperature and pH to induce cell transition to the VBNC state. Cells were harvested for analysis at five stages of VBNC induction: 0, 3, 30, and 180 days after induction and 210 days when the cells entered the VBNC state. Our results showed that the expression of 2,617, 2,642, 2,577, 2,829, and 2,840 genes was altered at these five different stages. The function of differentially expressed genes (DEGs, compared to healthy cells collected at day 0) and their encoded pathways were analyzed by the Gene Ontology Consortium and the Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses. A total of 10 DEGs were identified in cells that entered the VBNC state: five continuously upregulated (LCAZH_0621, LCAZH_1986, LCAZH_2038, LCAZH_2040, and LCAZH_2174) and five continuously downregulated (LCAZH_0024, LCAZH_0210, LCAZH_0339, LCAZH_0621, and LCAZH_0754). This study proposes a molecular model of the VBNC mechanism in L. paracasei Zhang, highlighting that changes in cell metabolism improve substrate utilization efficiency, thereby enhancing bacterial survival under adverse conditions. These data may be useful for improving the survival of probiotics in industrial food processing.

Gene expression changes throughout the life cycle allow a bacterial plant pathogen to persist in diverse environmental habitats.

Bacterial pathogens exhibit a remarkable ability to persist and thrive in diverse ecological niches. Understanding the mechanisms enabling their transition between habitats is crucial to control dissemination and potential disease outbreaks. Here, we use Ralstonia solanacearum, the causing agent of the bacterial wilt disease, as a model to investigate pathogen adaptation to water and soil, two environments that act as bacterial reservoirs, and compare this information with gene expression in planta. Gene expression in water resembled that observed during late xylem colonization, with an intriguing induction of the type 3 secretion system (T3SS). Alkaline pH and nutrient scarcity-conditions also encountered during late infection stages-were identified as the triggers for this T3SS induction. In the soil environment, R. solanacearum upregulated stress-responses and genes for the use of alternate carbon sources, such as phenylacetate catabolism and the glyoxylate cycle, and downregulated virulence-associated genes. We proved through gain- and loss-of-function experiments that genes associated with the oxidative stress response, such as the regulator OxyR and the catalase KatG, are key for bacterial survival in soil, as their deletion cause a decrease in culturability associated with a premature induction of the viable but non culturable state (VBNC). This work identifies essential factors necessary for R. solanacearum to complete its life cycle and is the first comprehensive gene expression analysis in all environments occupied by a bacterial plant pathogen, providing valuable insights into its biology and adaptation to unexplored habitats.

New insights into survival strategies and PCB bioremediation potential of resuscitated strain Achromobacter sp. HR2 under combined stress conditions

The resuscitated strains achieved through the addition of resuscitation promoting factor (Rpf) hold significant promise as bio-inoculants for enhancing the bioremediation of polychlorinated biphenyls (PCBs). Nevertheless, the potential of these resuscitated strains to transition into a viable but non-culturable (VBNC) state, along with the specific stressors that initiate this transformation, remains to be comprehensively elucidated. In this study, a resuscitated strain HR2, obtained through Rpf amendment, was employed to investigate its survival strategies under combined stress involving low temperature (LT), and PCBs, in the absence and presence of heavy metals (HMs). Whole-genome analysis demonstrated that HR2, affiliated with Achromobacter, possessed 107 genes associated with the degradation of polycyclic aromatic compounds. Remarkably, HR2 exhibited effective degradation of Aroclor 1242 and robust resistance to stress induced by LT and PCBs, while maintaining its culturability. However, when exposed to the combined stress of LT, PCBs, and HMs, HR2 entered the VBNC state. This state was characterized by significant decreases in enzyme activities and notable morphological, physiological, and molecular alterations compared to normal cells. These findings uncovered the survival status of resuscitated strains under stressful conditions, thereby offering valuable insights for the development of effective bioremediation strategies.

Effect of RpoS on the survival, induction, resuscitation, morphology, and gene expression of viable but non-culturable Salmonella Enteritidis in powdered infant formula

Involvement of the transcriptional regulator RpoS in the persistence of viable but non-culturable (VBNC) state has been demonstrated in several species of bacteria. This study investigated the role of the RpoS in the formation and resuscitation of VBNC state in Salmonella enterica serovar Enteritidis CICC 21482 by measuring bacterial survival, morphology, physiological characteristics, and gene expression in wild-type (WT) and rpoS-deletion (ΔrpoS) strains during long-term storage in powdered infant formula (PIF). The ΔrpoS strain was produced by allelic exchange using a suicide plasmid. Bacteria were inoculated into PIF for 635-day storage. Survival, morphology, intracellular reactive oxygen species (ROS) levels and intercellular quorum sensing autoinducer-2 (AI-2) contents were regularly measured. Resuscitation assays were conducted after obtaining VBNC cells. Gene expression was measured using real-time quantitative polymerase chain reaction (qPCR). The results showed that RpoS and low temperature conditions were associated with enhanced culturability and recoverability of Salmonella Enteritidis after desiccation storage in low water activity (aw) PIF. In addition, the synthesis of intracellular ROS and intercellular quorum sensing AI-2 was regulated by RpoS, inducing the formation and resuscitation of VBNC cells. Gene expression of soxS, katG and relA was found strongly associated with RpoS. Due to the lack of RpoS factor, the ΔrpoS strain could not normally synthesize SoxS, catalase and (p)ppGpp, resulting in its early shift to the VBNC state. This study elucidates the role of rpoS in desiccation stress and the formation and resuscitation mechanism of VBNC cells under desiccation stress. It serves as the basis for preventing and controlling the recovery of pathogenic bacteria in VBNC state in low aw foods.

Prokaryotic taxonomy and functional diversity assessment of different sequencing platform in a hyper-arid Gobi soil in Xinjiang Turpan Basin, China.

Turpan Basin located in the eastern Xinjiang is a typical arid inland basin with extremely scarce water resources and a fragile ecosystem. Prokaryotic communities with unique genetic and physiological modifications can survive and function in such harsh environments, offering diverse microbial resources. However, numerous microbes can enter the viable but non-culturable state because of drought stress in the desert soil. In this work, next generation sequencing (NGS) technology based on DNA nanoball sequencing platform (DNBSEQ-G400) and sequencing-by-synthesis platform (NovaSeq 6000) were applied to analyze the prokaryotic diversity in three hyper-arid Gobi soils from Flaming Mountain, Toksun, and Kumtag. The comparison between two platforms indicated that DNBSEQ-G400 had better repeatability and could better reflect the prokaryotic community of this hyper-arid region. The diversity analysis based on DNBSEQ-G400 identified a total of 36 bacterial phyla, including Pseudomonadota, Bacteroidota, Bacillota, Actinomycetota, Methanobacteriota, Acidobacteriota, Nitrososphaerota, and Planctomycetota. The environmental factors, including soluble salt, available potassium, total nitrogen, and organic matter, were positively correlated with the abundance of most prokaryote. In addition, the prokaryotic community assembly in hyper-arid soil was well described by neutral-based models, indicating that the community assembly was mainly controlled by stochastic processes. Finally, the phylogenetic analysis of Actinomycetota proved that such extremophiles played an important role in the ecosystems they colonize. Overall, our result provides a reference for choosing the appropriate sequencing platform and a perspective for the utilization of soil microbial resources from hyper-arid regions.

Collaborative cytometric inter-laboratory ring test for probiotics quantification.

Probiotics are live microorganisms that, when administered in adequate amounts, confer a health benefit on the host. From this definition, accurate enumeration of probiotic products is a necessity. Nonetheless, this definition does not specify the methods for assessing such viability. Colony forming units is the de facto gold standard for enumerating viable in probiotic products. The notion of microbial viability has been anchored in the concept of cultivability, which refers to a cell's capacity to replicate and form colonies on agar media. However, there is a growing consensus that the term "viability" should not be exclusively tied to the ability to cultivate cells. For example, bacterial cells can exist in a Viable But Non-Culturable (VBNC) state, characterized by the maintenance of characteristics such as membrane integrity, enzymatic activity, pH gradients, and elevated levels of rRNA, despite losing the ability to form colonies. Herein we present the results of a collaborative inter-laboratory ring test for cytometric bacterial quantification. Specifically, membrane integrity fluorescence flow cytometry (FFC) method and the newer impedance flow cytometry (IFC) method have been used. Both methods interrogate single cells in solution for the presence of intact membranes. FFC exploits fluorochromes that reflect the presence or absence of an intact membrane. IFC probes membrane integrity in a label-free approach by detecting membrane-induced hindrances to the propagation of electricity. A performance ring-test and comparison design on the FFC method showed that the method is robust against the exchange of equipment, procedures, materials, and operators. After initial method optimization with assessments of rehydration medium, wake-up duration, and phase shift gating on the individual strains, the IFC method showed good agreement with the FFC results. Specifically, we tested 6 distinct species of probiotic bacteria (3 Lactobacillus and 3 Bifidobacterium strains) finding good agreement between FFC and IFC results in terms of total and live cells. Together, these results demonstrate that flow cytometry is a reliable, precise, and user-friendly culture-independent method for bacterial enumeration.

A horizontal view of primary metabolomes in vegetative desiccation tolerance.

Vegetative desiccation tolerance (VDT), the ability of such tissues to survive the near complete loss of cellular water, is a rare but polyphyletic phenotype. It is a complex multifactorial trait, typified by universal (core) factors but with many and varied adaptations due to plant architecture, biochemistry and biotic/abiotic dynamics of particular ecological niches. The ability to enter into a quiescent biophysically stable state is what ultimately determines desiccation tolerance. Thus, understanding the metabolomic complement of plants with VDT gives insight into the nature of survival as well as evolutionary aspects of VDT. In this study, we measured the soluble carbohydrate profiles and the polar, TMS-derivatisable metabolomes of 7 phylogenetically diverse species with VDT, in contrast with two desiccation sensitive (DS) species, under conditions of full hydration, severe water deficit stress, and desiccation. Our study confirmed the existence of core mechanisms of VDT systems associated with either constitutively abundant trehalose or the accumulation of raffinose family oligosaccharides and sucrose, with threshold ratios conditioned by other features of the metabolome. DS systems did not meet these ratios. Considerable chemical variations among VDT species suggest that co-occurring but distinct stresses (e.g., photooxidative stress) are dealt with using different chemical regimes. Furthermore, differences in the timing of metabolic shifts suggest there is not a single "desiccation programme" but that subprocesses are coordinated differently at different drying phases. There are likely to be constraints on the composition of a viable dry state and how different adaptive strategies interact with the biophysical constraints of VDT.

The Problem of Recognition of Palestine in the context of the Palestinian People’s Realization of the Right to Self-Determination

The paper examines the problem of recognition of Palestine in modern international law. International documents and United Nations resolutions adopted on the issue of the conflict in Palestine are analyzed. The author, based on an analysis of international documents, concludes that Palestine meets all the criteria for statehood, but the Israeli occupation prevents the creation of a viable Palestinian state. Despite various efforts being made to reach a solution to the Palestinian issue, the peace process has reached a deadlock. This is facilitated by the policy and practice of the occupying power to consolidate the de facto position of the occupied territories in violation of its legal obligations. To date, Palestine is recognized by more than 138 states and the UN General Assembly has repeatedly confirmed the right to self-determination through resolutions. The efforts of the international community must be intensified to put an end to this irreconcilable conflict, which poses a threat to international peace and security.

Detection of Viable but Non-Culturable (VBNC)-Campylobacter in the Environment of Broiler Farms: Innovative Insights Delivered by Propidium Monoazide (PMA)-v-qPCR Analysis.

Campylobacteriosis cases in humans are of global concern, with high prevalence rates in the poultry reservoir considered the most important source of infection. Research findings show Campylobacters' ability to enter a viable but non-culturable (VBNC) state, remaining "viable" but unable to grow on culture media. We explored the persistence of VBNC states in specific environments, particularly at broiler farms, as this state may lead to an underestimation of the present Campylobacter prevalence. For VBNC detection, a propidium monoazide PMA-dye viability qPCR (v-qPCR) was used in combination with cultivation methods. We examined samples collected from broiler farm barns and their surroundings, as well as chicken manure from experimental pens. In addition, the tenacity of culturable and VBNC-Campylobacter was studied in vitro in soil and water. In a total of three visits, Campylobacter was not detected either culturally or by v-qPCR (no Campylobacter DNA) in the environment of the broiler farms. In four visits, however, VBNC-Campylobacter were detected both inside and outside the barns. The overall prevalence in environmental samples was 15.9% for VBNC-Campylobacter, 62.2% for Campylobacter DNA, and 1.2% for culturable C. jejuni. In the experimental pens, no cultivable C. jejuni was detected in chicken manure after 24 h. Strikingly, "VBNC-Campylobacter" persisted even after 72 h. "VBNC-Campylobacter" were confirmed in barn surroundings and naturally contaminated chicken manure. Laboratory studies revealed that VBNC-Campylobacter can remain intact in soil for up to 28 days and in water for at least 63 days, depending on environmental conditions.