Biosorption is considered an effective technique for the treatment of heavy-metal-bearing wastewaters. In recent years, various biogenic products, including native and functionalized biopolymers, have been successfully employed in technologies aiming for the environmentally sustainable immobilization and removal of heavy metals at contaminated sites, including two commercially available heteropolysaccharides—xanthan and gellan. As biodegradable and non-toxic fermentation products, xanthan and gellan have been successfully tested in various remediation techniques. Here, to highlight their prospects as green adsorbents for water decontamination, we have reviewed their biosynthesis machinery and chemical properties that are linked to their sorptive interactions, as well as their actual performance in the remediation of heavy metal contaminated waters. Their sorptive performance in native and modified forms is promising; thus, both xanthan and gellan are emerging as new green-based materials for the cost-effective and efficient remediation of heavy metal-contaminated waters. Heavy metals are considered metallic elements  whose compounds can induce toxicity at a low level of exposure, thus posing a critical threat to human health ,2,3,4,5.Since heavy-metal-associated acute and chronic toxicity at low concentrations may pose a critical threat to human health , variousRegarding the issue of finding new green-based materials for the remediation of contaminated waters, biopolymers have emerged as a promising alternative to synthetic polymers, primarily due to their biodegradability, non-toxicity, and economic sustainability . This reXanthan and gellan biosynthesis comprises several steps , includiLike most bacterial surface heteropolysaccharides, xanthan is synthesized through a Wzy-dependent pathway, where the lipid-linked repeat units are built up on polyisoprenol phosphate at the cytoplasmic face of the inner membrane, then flipped to the periplasmic face of the inner membrane by a flippase and polymerized by a block-transfer mechanism involving the Wzy polymerase .Xanthomonas campestris pv. campestris B100 are the nucleotide sugars GDP-mannose, UDP-glucose, and UDP-glucuronic acid, which comprise the xanthan repeating units (in a molar ratio of 2:2:1).The direct precursors for xanthan production in gum cluster. The gum cluster comprises 12 genes that are involved in the assembly of the pentasaccharide repeating unit, its decoration with substituents, polymerization, translocation, and secretion [xanA and xanB, which are not included in the gum cluster. The xanA and xanB genes encode bifunctional enzymes phosphoglucomutase/phosphomannomutase, which convert glucose-6-phosphate and mannose-6-phosphate to their corresponding 1-phosphates, and phosphomannose isomerase/GDP-mannose pyrophosphorylase, respectively [xanA deletion restricted the capacity to synthesize the nucleotide, the monosaccharide composition of xanthan gum in Xanthomonas citri subsp. citri was affected. This resulted in the decrease of biofilm formation, sliding motility, and pathogenicity of this phytopathogenic Gram-negative proteobacterium [The building of pentasaccharide units is under the control of the 12 kb gene called the ecretion ,27. Howeectively . Since xacterium .The precursor biosynthesis is followed by the processes of building and decoration of pentasaccharide subunits at the cell envelope. Among others, the process is initiated by the GumD and GumM glycosyltransferases. While the former transfers glucose-phosphate residue from UDP-glucose to an undecaprenyl-phosphate C-55 lipid carrier molecule located at the cytoplasmic face of the membrane , the latgumB and gumC [Finished lipid carrier-linked repeating pentasaccharide units are then translocated by the putative flippase GumJ to the periplasmic face of the inner membrane, where it can be polymerized by GumE, a homolog to O-antigen polymerase Wzy , which tand gumC , whose oand gumC .Sphingomonas, include UDP-glucose, UDP-glucuronic acid, and deoxythymidine diphosphate (dTDP)-rhamnose [Sphingomonas spp., which are presumed to be involved in precursor synthesis, include phosphoglucomutase catalyzing the conversion of glucose-6-phosphate into a glucose-1-phosphate encoded by pgmG gene and glucose-1-phosphate uridylyltransferase (pyrophosphorylase) encoded by the ugpG gene [gel cluster (18-gene cluster required for gellan biosynthesis), also exhibited dTDP-glucose pyrophosphorylase activity in Sphingomonas elodea ATCC 31461 [The activated sugar precursors for biosynthesis of tetrasaccharide unite in gellan, a sphingan heteropolysaccharide produced by genera rhamnose . The enzgpG gene . The latCC 31461 . The bioCC 31461 .gelB, gelK, gelL, and gelQ encoding the glycosyl transferases [gelB, enables the transfer of UDP-glucose residue to an activated C55-isoprenylphosphate lipid carrier. The gene gelB is homologous to spsB from Sphingomonas sp. ATCC 31554 encoding glucosyl-isoprenylphosphate transferase [β-1,4-glucuronosyltransferase, structurally homologous to E. coli N-acetyl glucosaminyl transferase MurG, which catalyzes the transfer of UDP-glucuronate residue to activated lipid link glucose [gelL (homologous to spsL) and gelQ of the gellan cluster, which most likely enable the incorporation of the third (glucose) and fourth (rhamnose) sugars to the tetrasaccharide repeating unit of the gellan exopolysaccharide, respectively [The genes involved in the sequential assembly of repeating tetrasaccharide units of the gellan backbone include sferases . The prinsferase . The Gel glucose . The remectively ,43. The gelS and gelG (homologues to Wzx and Wzy) located in the gel cluster region IV are likely involved in the repeating unit translocation across the inner membrane and polymerization of gellan, respectively. Its chain-length-regulating protein, a polysaccharide co-polymerase, is encoded by the genes gelC and gelE, with the latter reportedly having an ATP-binding activity in vitro [gelD [The genes in vitro . Finallyro [gelD .There are various emerging molecular techniques which address heavy metal pollution and which, via genetic engineering, exploit some of the metal-related features of bacterial metabolism, e.g., synthesis of the specific metal-chelating polymers . EngineeIdentifying the components responsible for the pollutant binding as well as the understanding of the molecular machinery of their biosynthesis are both essential aspects for the successful implementation of the genetic engineering principles in enhancing the performance of xanthan and gellan in remediation of heavy metal-contaminated waters, as well as in reducing the costs for the polysaccharides’ production. Unfortunately, to the best of our knowledge, there is no molecularly oriented research on xanthan and gellan synthesis that directly relates to remediation of heavy metal contaminated sites. Still, the development of the low-cost, high-yield production of recombinant biopolysaccharides is highly beneficial for food, pharmaceutical, and cosmetic industries; thus, a lot of progress has been made regarding the genetic engineering in these industrial areas.gelQ, gelK, gelL, and gelB) and produced the recombinant polysaccharide that was similar in structural properties with the native gellan. Transformation of the recombinant pBBR122-gelD plasmid into Sphingomonas paucimobilis cells and overexpression of the gelD in recombinant strain yielded twice as much gellan in comparison to a wild-type strain [pgmG, ugpG or gelK gene copies in recombinant plasmids had no positive effect on gellan productivity, while the simultaneous overexpression of pgmG and ugpG genes that are involved in precursors synthesis increased the overall production of gellan by 20%. Another approach, which reduces the cost of xanthan and gellan production and is unrelated to genes directly involved in polysaccharide synthesis, includes constructing mutants that produce more polysaccharides and which are capable of utilizing a cheap industrial by-product, e.g., construction of transconjugants which utilize whey- or lactose-containing media for xanthan production [In the effort to enhance gellan production, Manjusha et al.  successfe strain . Via expe strain . Since pe strain , their aoduction .It is reasonable to expect that the ever-growing versatility and precision in genetic engineering techniques will, in time, enable the modification of the xanthan and gellan biosynthesis machinery to an extent that the capacity of microorganisms to tolerate and accumulate heavy metals will be enhanced significantly, and they can be employed for the development of a green clean-up strategy that utilizes the recombinant polysaccharides for the removal of heavy metals from waters.Xanthomonas campestris belonging to the family Pseudomonadaceae [X. campestris is most commonly employed for industrial production of xanthan gum, there are several other strains of genus Xanthomonas which can produce xanthan, e.g., X. arbicola pv. juglandis, X. axonopodis pv. begonia, X. axonopodis pv. vesicatoria [X. pelargonii [Xanthan gum is an extracellular water-soluble heteropolysaccharide that is commercially produced from various carbon sources by fermentation in controlled conditions using a Gram-negative bacteria nadaceae . Being dnadaceae . Althougicatoria , and X. largonii .Sphingomonas, which belongs to the α-4 subclass of the Proteobacteria [Sphingomonas paucimobilis ATCC 31461 strain , a highly productive industrial strain of gellan gum. There have been other strains of S. paucimobilis reported to be capable of producing gellan gum [S. paucimobilis ATCC 31461 [S. azotifigens GL-1 [S. paucimobilis ATCC 31461, strain NK2000. Raghunandan and Kumar [Sphingomonas bacteria isolated from biodiesel-derived waste glycerol for their potential usage in the commercial production of gellan. Two isolates, S. yabuuchiae, and S. pseudosanguinis, displayed high waste glycerol degradation as well as gellan conversion rates. They demonstrated great potential in the cost-effective commercial production of gellan, and great prospects in the remediation of biodiesel-derived crude glycerol.Gellan (earlier referred to as polysaccharide S-60), a sphingan group of heteropolysaccharides, is secreted by members of the bacterial genus bacteria . It is mllan gum , includiCC 31461  and S. aens GL-1 . Variousens GL-1  developeX. campestris is sensitive to cadmium’s toxic effects. However, pre-exposure to low concentrations of cadmium(II) induced cross-protection against the lethal concentrations of cadmium(II) and zinc(II). Similar effects have been reported by Hrimpeng et al. [X. campestris to sublethal concentrations of arsenic(III), which induced the cross-protective response to CuOOH and peroxide. Cadmium(II) has been also shown to be a potent inducer of peroxide-scavenging enzymes [Xanthomonas in populations of plant-pathogenic bacteria [Exposure to heavy metals can induce various physiological responses in bacteria, such as alteration in gene expression related to the transport of metal cations and oxidative stress, or it can enhance the production of extracellular polymers capable of sequestering the toxicants. Banjerdkij et al.  noted thg et al.  in the c enzymes . These sbacteria . Howeverbacteria .Sphingomonas sp. LK11 strain has shown promising prospects in the remediation of contaminated environments, since it maintained steady growth during cadmium-induced stress. However, Li et al. [Sphingomonas sp. was biosorption, and the involvement of carboxyl and carbonyl functional groups of saccharides in cadmium immobilization via physical entrapment, ion exchange, and complexation has been suggested. Hence, the metal binding most likely occurred on extracellular polysaccharides. Furthermore, Ashour [X. campestris This physiological response of bacterium to heavy metal exposure can be viewed as a protective mechanism against cobalt’s toxic effects, since the sequestration of the metallic cations inhibits their entry into the cells [Also, i et al.  reported, Ashour  noted thhe cells .Xanthomonas spp., is expected to play a substantial role in metal removal from the water produced by the oil industry via binding the metal cations by forming the Werner-type complexes, since it possesses a number of hydroxyl and carboxyl groups [Sphingomonas genera in soil microbial communities at sites highly contaminated with heavy metals due to long-term smelting and mining operations [Xanthomonas bacteria possess effective enzymatic and non-enzymatic strategies to survive and proliferate in the presence of reactive oxygen species and, thus, have shown great tolerance to heavy-metal-dependent induction of oxidative stress in cells [copAB operon identified in some of Xanthomonas species that regulate efflux of copper [The Involvement of xanthan and gellan in heavy metal cation sequestration by living bacteria is plausible, since both polymers exhibit anionicity, and they are packed with negatively charged functional groups . For exal groups . Howeverl groups  and thatl groups , the immerations  renders in cells , their pf copper .S. paucimobilis [Xanthomonas to reduce mercury(II), thus accelerating the mercury volatilization [Since bacterial sensitivity to heavy metals and their specific growing and nutritional requirements  provide drawbacks to their feasibility, and xanthan and gellan have performed well in heavy metal immobilization, the utilization of separated bacterial polysaccharides in decontamination of wastewaters can be advantageous over living cells. This is most likely the reason why there are virtually no comprehensive studies on bioaccumulation of heavy metals by xanthan- or gellan-producing living bacteria. That being the case, the more complex kinetic and thermodynamic studies on heavy metal removal are available only for the dead cells, e.g., the report on copper(II) biosorption by non-living biomass of imobilis . Still, lization .Both the xanthan and gellan chains are considered stable in aqueous solutions and capable of reversibly binding the metallic ions. Their ability to bind cations is due to the presence of various active functional groups. FTIR spectra of xanthan gum revealed the presence of a prominent broad band typical for hydroxyl groups, but also carbonyl esters of acetyl and carboxylate groups . SimilarXanthomonas strain, as well as the operational conditions used [The xanthan molecule is composed of pentasaccharide repeating units which consist of a cellulose-like backbone with a trisaccharide side-chain of two mannose and one glucuronate residue attached to a β-D-glucose unit, with pyruvic acid linked to O(4) and O(6) of the terminal D-mannose  and an aons used .The native biosynthesized polymer of gellan consists of two residues of β-D-glucose, one of β-D-glucuronate and one of α-L-rhamnose, which provides hydroxyl groups, as well as the L-glyceryl and acetyl substituents positioned at O(2) and O(6) of D-glucose residue of the tetrasaccharide sequence .Bergmann et al.  suggesteThe character of the interaction of the active functional groups of the polysaccharide with metallic ions also affects the properties of polymers, e.g., polymer morphology and solution rheology . Some poX. campestris CCTCC M2015714 in the detoxication of lead(II)-, cadmium(II)-, and copper(II)-contaminated waters at acidic pH simulating the stomach pH. The xanthan showed excellent adsorption capacity by reducing the concentration of contaminants in an aqueous solution by 50% after one hour. Gellan gum’s performance in nickel(II) removal from the aqueous solutions was equally excellent since its sorption capacity was found to be superior in comparison to other gel-forming polymers, including agar, Ca-alginate, silica gel, polyacrylamide, and agarose [Although both xanthan and gellan are usually used in the food industry, they also show great prospects in the remediation of contaminated waters, since they provide various active functional groups that are capable of binding the heavy metals from aqueous solutions and removing them efficiently . Wang et agarose .There are numerous approaches to synthesizing the gellan- and xanthan-based sorbents that are applicable for the cost-effective and environmentally friendly treatment of wastewater and which improve the thermal and mechanical performance and low surface area of the native biopolymers . The hea−1 of zinc(II) at pH 5.0, while the presence of polyethylacrylate grafts increased the adsorption capacity by six-fold. Later, Pandey and Mishra [Modification by grafting synthetic polymers onto a native xanthan polysaccharide allows for overcoming some drawbacks of the native xanthan; also, it yields new materials with different structural characteristics and desirable properties . Pandey d Mishra  modified−1 for lead(II) was calculated using the Langmuir isotherm model, highlighting the excellent adsorption efficiency of the synthesized nanocomposite for lead removal from the complex industrial wastewaters. Promising results in lead(II) removal have been achieved also by cross-linking the xanthan and guar gums, where the metal cation was predominantly attached to carboxylic groups [The FTIR spectra revealed that the partial hydrolyzation of the product of polyacrylamide-grafted xanthan gum increased the abundance of carboxylic groups, thus suggesting the increase of anionic moieties in the polymer. This should have had a beneficial effect on the adsorbent performance in the immobilization of cationic heavy metals; however, the flocculation performance was affected negatively . This rec groups . Their a−1. This is significantly higher than other reported copper(II) removal capacities of grafted xanthan gum, e.g., 39.1 mg·g−1 for the xanthan gum-graft-2-acrylamido-2-methyl-1-propane sulfonic acid [−1 for xanthan gum modified with thionyl chloride followed by ethylenediamine modification [−1 for cadmium and lead cations, respectively.Zheng et al.  synthesinic acid  or 46.9 fication . A hybrification . The sor−1, respectively. Mama et al. [−1), iron (45.2 mg·g−1), and zinc (44.6 mg·g−1) cations; its good performance in the ternary sorption system highlighted the applicability of this novel composite for actual wastewater treatment.Inorganic fillers in cross-linked polymers, such as clay material, can lend biopolymer-specific chemical and physical properties and minimize chain mobility. Esmaeildoost et al.  attemptea et al.  prepared3O4@silica–xanthan gum composite that was successfully tested for lead(II) removal and showed promising results for resolving the critical pollution of lead-bearing battery industry wastewater [Another novel approach is the modification of biopolymers with ferromagnetic (nano)materials that allows the magnetic separation of contaminants bound to the adsorbent. This includes a Festewater . Mirza astewater  also demMagnetic nanoparticles are also advantageous in the remediation of heavy metals, since they can be tailored via surface functionalization for specific contaminants with organic compounds . Various−1, while exhibiting reasonable regenerative capacity.The cross-linked gellan was also used for the encapsulation of other biopolymers, such as alginate. Nussinovitch and Dagan  prepared22+), and the maximum sorption capacity of the novel biosorbent was 202.8 mg·g−1. The same research team also synthesized a gellan-gum-based adsorbent with less, a 97.9 mg·g−1 removal capacity for uranyl cations, but with improved gel strength, by reinforcing gellan gum microspheres using the carboxymethyl konjac glucomannan [Liang et al.  preparedcomannan .The research outcomes indicate that although natural xanthan and gellan provide plenty of carboxyl and hydroxyl groups for heavy metal immobilization, their adsorption capacity, selectivity, and mechanical properties  can be profoundly improved by the grafting of other functional groups or molecules onto the biopolysaccharide structure. Therefore, even in cases when xanthan- and gellan-derived adsorbents possess lower sorption capacities in comparison to other conventional or non-conventional adsorbents , there iTo summarize the presented outcomes, the practical application of xanthan and gellan polymers in heavy metal removal from aqueous media may be successful, yet uncertain. This is due to high competitiveness among available non-conventional adsorbents regarding their sorptive performance and costs. This is clearly exemplified in Pseudomonas veronii and attached the biopolymer mix onto zeolite granules. Bioaugmentation of mercury-contaminated soil with immobilized bacteria having mercury(II) reduction and volatilization capacity resulted in a sharp increase of gaseous elemental mercury emissions, with maximum flux rates exceeding 10 μg Hg·m2·h−1.Besides immobilization via sorptive interactions, the application of xanthan and gellan gums in heavy metal remediation is limited, primarily due to the non-degradative properties of heavy metals. There is a much wider variety of strategies for the decontamination of degradable organic pollutants using polymeric bioinspired materials . Still, 2PO4) [Xanthan and gellan are both viscous biopolymers; hence, as an additive, they can enhance soil strength. Chang et al.  demonstr2PO4) .A unique approach combining plant growth and amendment of xanthan gum has been suggested for remediation purposes of heavy-metal-contaminated soils by Ni et al. , who notBiopolymer-based materials are rapidly emerging as an effective green alternative to traditional adsorbents employed for heavy metal removal. As our review shows, among the other commercially available biopolymers, the heteropolysaccharides xanthan and gellan show great prospects in this regard. Unfortunately, the actual research on their performance in heavy metal removal is still in the initial stages and comprises developing new, more effective xanthan- or gellan-based adsorbents and their testing under optimized laboratory conditions. Nonetheless, to evaluate the viability and cost-effectiveness of xanthan and gellan’s employment in remediation technologies, experiments should be performed at a larger scale and should involve more complex matrices . However, no such studies are available to this day. Still, we hope it will not be left unchallenged and in time to come, this issue will be overcome by reporting an in situ large-scale experiment that would highlight the economic feasibility and excellent performance of xanthan- and gellan-based adsorbents for the removal of heavy metals from contaminated waters.


Elderly long-term care residents (ELTCRs) face considerable burden of infection, especially evident during the COVID-19 pandemic. The nutritional status of the host can influence susceptibility to infection by altering immune system integrity, therefore, nutrition-based interventions may be a viable complement to existing infection prevention measures.This scoping review sought to identify nutritional interventions and factors that have the strongest evidence to benefit ELTCRs, and thus best poised for rigorous clinical trial evaluation and subsequent implementation.A database search of OVID-Medline, OVID-Embase, and Web of Science was performed from 2011 to 2021 to identify nutritional intervention studies which attribute to changes in infection in contemporary ELTCR settings. Articles were screened in duplicate and data extraction completed by a single reviewer, while a second reviewer verified the data which was fitted to identify evidence for nutritional interventions related to reducing rates of infection among ELTCRs.The search identified 1018 studies, of which 11  satisfied screening criteria. Interventions that significantly reduced risk of infection included whey protein (any infection), Black Chokeberry (urinary tract infection), and vitamin D . Both zinc and a dedicated meal-plan significantly improved lymphocyte parameters. Vitamin D deficiency was associated with the development of respiratory tract infections. Probiotic and soy-based protein interventions did not significantly affect risk of infection or lymphocyte parameters, respectively.The current scoping review was effective in identifying the use of nutrition-based interventions for infection prevention among ELTCRs. In this study, some nutrition-based interventions were observed to significantly influence the risk of infection among ELTCRs. Nutritional interventions such as vitamin D (preventing deficiency/insufficiency), Black Chokeberry juice, zinc gluconate, whey protein, and varied and nutrient dense meal plans may be suitable for future rigorous clinical trial evaluation. Elderly long-term care residents (ELTCRs) face a considerable burden of infection, including high rates of respiratory tract infections , urinaryAmong ELTCRs, age-associated waning of systemic immunity (known as immunosenescense) may contribute towards increased risk and severity to infectious diseases . An optiIn practice, malnutrition, as determined by validated nutritional assessment tools, has been observed to increase the risk of serious infections  –22, urinThese observations are relevant to Canadian ELTCR cohorts, as nutrient intake deficits and/or malnutrition is reported to be widespread and persistent in long-term care facilities across the nation. A 2018 Canadian cross-sectional study conducted in thirty-two long-term care homes (n = 632 randomly selected residents) found that >50% of residents consumed inadequate amounts of vitamin D, E, K, B6, folate, calcium, and magnesium . AnotherTo our knowledge, no scoping review encompassing evidence for nutritional interventions in the realm of reducing rates of infection among ELTCRs has been published. A scoping review would facilitate the delineation of data on the effects of nutrition-based interventions to reduce the risk of infection among ELTCRs and facilitate in identifying areas of research to inform future experimental studies and pragmatic approaches to reduce the burden of infection in long-term care facilities. This present scoping review, through a detailed and structured literature search of peer reviewed articles, will attempt to describe the current evidence for the use of nutrition-based interventions to address infections in ELTCRs. The present scoping review is used to identify nutritional interventions and factors that have the strongest evidence to benefit this cohort, and thus, are best suited for rigorous clinical trial evaluation and subsequent implementation.To methodically map the extent, range, and nature of published, peer-reviewed, research related to nutrition-based interventions to reduce the risk of infection among ELTCRs , 33.To identify nutrition-based interventions that have demonstrated the capacity to significantly reduce the risk of infection and/or improve clinically relevant immune system parameters  among ELTCRs, which may be suitable for future rigorous clinical trial evaluation.To identify nutrition related parameters, evaluated in prospective cohort studies, demonstrating an association between undernutrition and/or nutrient deficiencies, and the risk of infection among ELTCRs.https://www.crd.york.ac.uk/prospero/) as it is not a systematic review.The process of a scoping review facilitates identifying existing peer-reviewed literature that provides information that has not been comprehensively reviewed or is complex and diverse in nature . A scopiThe scoping review was conducted using a structured approach presented and informed using guidelines for a preferred reporting of items for systematic reviews and meta-analyses (PRISMA) extension for Scoping Reviews (PRSIMA-ScR) as shown in  32]. A . A 32]. Published between 2011–2021.Full-text accessible for review.Published in English .Peer-reviewed published literature.Either experimental/interventional , phase I-II clinical trials) or longitudinal epidemiologic study.Included a primary intervention that was nutrition based  or a mixed intervention that included nutritional reporting.At least one outcome or measure was infection related, including risk of infection (any type of infection), immune system parameters , the need for antibiotic use for present infection or other opportunistic infections, length of hospital stay due to infection, other complications due to infection, recovery, and response to vaccination.Participants were elderly (≥ 65 years of age). Studies with mixed aged groups were allowed, if the majority (>50%) were ≥ 65 years. The cut off for age (≥ 65 years) was based on the conventional definition of an ‘elderly’ individual to be defined as a chronological age of 65 years .Participants resided in any type of long-term care facility, such as nursing homes, throughout the intervention period.The absence of a peer-review process.Unpublished results or literature.in vitro studies, conference abstracts/presentations/posters, editorials, and opinions.The following study designs: non-longitudinal observational studies, case reports, review, animal/pre-clinical/Did not include nutritional related primary intervention/s.Included participants that were primarily of community dwelling, hospitalised, with an acute care setting, in transitional centres, and/or were not within the guidelines of living in long-term care facility.Did not include outcome(s) or measure(s) related to infection.st, 2021, and both Ovid-Medline and Ovid-Embase on June 7th, 2021. Grey literature was not included as the intention of this scoping review was to specifically identify possible nutrition-based intervention candidates that can be evaluated and tested in future rigorous clinical trials. Thus, a peer-reviewed process for publication was essential and hence required. The primary reason for choosing 10 years (2011 to 2021) for all three databases was to ensure the collection would most closely reflect contemporary ELTCRs in the population, and thus would be most applicable and relevant to modern and future real-world cohorts. Nevertheless, an initial search with a 20 to 25-year time span (2001–2021 & 1996–2021) with the above-mentioned inclusion and exclusion criteria was implemented. This increased time frame included a very large number of citations. Following removal of duplicates, the remaining studies were not appropriate for nutrition-based interventions. Therefore, the rationale for a 10-year time span was adopted to reflect contemporary nutrition-based interventions among this cohort. Further, there is evidence of updates and revised guidelines pertaining to infection control for long-term and health care institutions in Canada : year of publication; study design; data collection period; sample size and characteristics; type of long-term care institution; country of origin; nutrition components details; infection related outcomes; infection related results; study limitations and strengths; and keywords related to the publication.An annotated bibliography was developed  with theExtracted data was organised into tables to present general characteristics of each included study. Results were organised by using the type of nutritional intervention relative to infection related outcomes.Database searches of Ovid-Medline, Ovid-Embase, and Web of Science identified 1018 articles (after de-duplication). Following inter-database de-duplication (n = 36 articles removed), 982 records were screened by title and abstract for inclusion, resulting in 952 articles being excluded. The remaining 30 records were screened via full text, resulting in 19 exclusions, yielding 11 articles for data extraction. Chain searching of the 30 articles recognized for full text review resulted in no additional eligible studies being identified. Detailed depiction of study identification, screening, eligibility assessment, and inclusion is presented as a PRISMA flow chart .Aronia melanocarpa) juice (n = 1) [All 11 articles selected for data extraction were conducted in high income countries with ten having membership with the organisation for economic co-operation and development (OECD) and one a non-OECD member. Nine of the identified studies were experimental –57 and t (n = 1) , dedicat (n = 1) , and zin (n = 1) .As described in 10 colony forming unit, CFU) product, delivered two times a day, beginning at the onset of antibiotic therapy until one-week post completion, did not reduce the frequency of antibiotic usage/prescriptions throughout the follow-up period [None of the three probiotic-based experimental studies reported a significant benefit of supplementation for any infection related outcome –51. A fip period .Lactobacillus casei Shirota (6.5 x 109 CFU) did not significantly reduce the number of days with respiratory infection symptoms (p = 0. 342), nor the number of participants presenting with respiratory symptoms (p = 0.325), based on univariate analysis [Lactobacillus casei Shirota did not significantly alter the risk of developing a severe respiratory tract infection, compared to placebo  (p = 0.073) [Compared to placebo, two bottles a day, for 176 days, of a fermented milk product containing analysis . Multipl= 0.073) . Further= 0.073) .Lactobacillus rhamnosus GG (10 billion CFU/capsule) did not significantly reduce the risk of developing a viral respiratory infection (PCR-confirmed), compared to placebo  [Six months of supplementation with two capsules a day of 32–1.31) . There w32–1.31) .In another vein, while it did not significantly affect an infection related outcome, supplementation with a multi-strain probiotic significantly reduced the number of antibiotic-associated diarrhea episodes (p = 0.022), possibly signalling an option to improve treatment tolerance and compliance .Two intervention studies provided nursing home residents with supplemental protein, yielding mixed results , 56.Twenty grams of whey protein, dissolved in 4.5 deciliter (dL) of juice, consumed in three equal portions a day for 6 months, did not significantly reduce risk of urinary tract infections, compared to control  (p = 0.053) . Over a 2 & MNA score ≤ 24) consuming a warm soy drink product  once a day, over a 24-week period, did not significantly improve lymphocyte counts, compared to control (p>0.05) [At risk residents (BMI ≤ 24 kg/m(p>0.05) .One intervention trial, with crossover, providing residents with a Black Chokeberry product reported mixed results for different infection related outcomes . DependiOne single-arm, pre-post, clinical trial providing dysphagic  residents with a needs-specific meal plan, which was comprised of food options that included recommended amounts of protein, were textured, more nutrient dense, and varied compared to standard meals, reported positive results . At the One intervention trial supplemented residents with a monthly high dose of vitamin D3 for 12 consecutive months, reporting overall positive results for most infection related outcomes . ResidenOne small intervention trial, involving residents with low baseline serum zinc concentrations, reported mixed results of zinc gluconate supplementation effects on immunity . Three mTwo observational, prospective cohort, studies were identified; one investigating the association between vitamin D deficiency and the risk of respiratory tract infections, hospitalizations, and any type of infection . The othThe prevalence of vitamin D deficiency was observed to be significantly higher in residents experiencing respiratory tract infections than those with no deficiency . A KaplaNeither the mini-nutritional assessment (MNA) tool, Nutritional Risk Screening 2002 (NRS) tool, or the Malnutrition Universal Screening Tool (MUST) at either the 6-month or 12-month time point was predictive of infection .In this scoping review, the described literature landscape included interventions that significantly reduced the risk of infection, encompassing whey protein (any infection) , Black CVitamin D is a well-documented modulator of both the innate and adaptive immune system with notable influence on antimicrobial peptide production, inflammatory cascade regulation, gene expression, and T-cell activity . ClinicaDietary protein supplies necessary organic compounds, termed amino acids , which sAronia melanocarpa) fruit is a rich source of bioactive phytochemicals, including anthocyanins, proanthocyanidins, flavonols, flavanols, and phenolic acids [Black Chokeberry  killing of virus-infected cells, and regulating immune responses . LymphopIn relation to this current scoping review, age-related changes in the human body are associated with a decline in immune system functions, which affect both innate and adaptive immunity and the provision of nutritional interventions may help prevent infectious sequelae . Diet isAlthough effective in many ways it is noteworthy that the following limitations were encountered in this scoping review. The protocol used for this scoping review was not registered with PROSPERO and the study included only peer reviewed published articles, while grey literature was not included within the area of interest. This approach was taken to minimize articles which are not a product of peer-review processes. Additionally, the other challenge with grey literature is that many resources may not be retained on an online platform to facilitate searchable solutions. The exclusion of non-longitudinal studies such as cross-sectional studies were a result of the impact of time required for the effectiveness of the nutrition-based intervention or supplement. Our scoping research review describes an elderly cohort living in long-term care facilities that undergo surveillance over time to document the incidence of an outcome (a reduction in infection) among exposed (nutrition-based intervention) and non-exposed participants. The temporality of a prospective or longitudinal study affords the cohort exposure time  with each participant observed repeatedly. We believe the time factor in such longitudinal designs is appropriate for this research work as it affords an explanation of causality. Case reports were excluded as they did not fit the designation of a trial as well ranking of validity is lower compared to experimental or prospective cohort studies. This scoping review only included recent studies published within the past 10 years. While this increases the relevance and applicability to modern cohorts, it does not exhaustively describe the overarching research landscape for this topic beyond this time frame.Review of the current literature was effective in identifying the use of nutrition-based interventions for infection prevention among ELTCRs. In this scoping review, some nutrition-based interventions were observed and significantly influenced the risk of infection among ELTCRs. The nutritional interventions best poised for immediate rigorous clinical trial evaluation and subsequent implementation for ELTCRs are vitamin D (preventing deficiency/insufficiency), Black Chokeberry juice, zinc gluconate, whey protein, and varied and nutrient dense meal plans. Probiotics are unlikely to be of much value for infection prevention/control in this specific at-risk population. Overall, there is minimal research for nutrition-based interventions for infection prevention among ELTCRs, warranting more intervention studies, beginning with the ones identified in this review.S1 File(PDF)Click here for additional data file.S1 Table(PDF)Click here for additional data file.


Immunosurveillance of cancer is mediated by T cell-based recognition of tumor-associated antigens, i.e., short peptides that are presented on the surface of cells on human leukocyte antigen (HLA) molecules. This encourages the analysis of the entirety of HLA-presented peptides, the so-called immunopeptidome, of malignant and benign cells, in order to identify novel therapeutic targets presented exclusively on malignant cells. In the present study, we aim to investigate the role of previously described immunopeptidome-defined antigen presentation in chronic lymphocytic leukemia (CLL) patients for clinical characteristics and disease outcome. We observed that higher yields of presented total and CLL-exclusive peptides were associated with a more favorable disease course, suggesting efficient immunosurveillance in a subgroup of patients and the possibility of further investigating T cell-based therapeutic approaches for CLL.Chronic lymphocytic leukemia (CLL) is characterized by recurrent relapses and resistance to treatment, even with novel therapeutic approaches. Despite being considered as a disease with low mutational burden and thus poor immunogenic, CLL seems to retain the ability of eliciting specific T cell activation. Accordingly, we recently found non-mutated tumor-associated antigens to play a central role in CLL immunosurveillance. Here, we investigated the association of total and CLL-exclusive HLA class I and HLA class II peptide presentation in the mass spectrometry-defined immunopeptidome of leukemic cells with clinical features and disease outcome of 57 CLL patients. Patients whose CLL cells present a more diverse immunopeptidome experienced fewer relapses. During the follow-up phase of up to 10 years, patients with an HLA class I-restricted presentation of high numbers of total and CLL-exclusive peptides on their malignant cells showed a more favorable disease course with a prolonged progression-free survival (PFS). Overall, our results suggest the existence of an efficient T cell-based immunosurveillance mediated by CLL-associated tumor antigens, supporting ongoing efforts in developing T cell-based immunotherapeutic strategies for CLL. Chronic lymphocytic leukemia (CLL), the most common leukemia in adults in western countries, is characterized by the clonal expansion of a population of atypical B cells, which accumulates in the blood, bone marrow, lymph nodes and spleen ,2,3. CLLDespite the improved management of CLL achieved over the past years , the per+ and CD4+ T cells, respectively [Despite being recognized as a low-mutational burden tumor entity , CLL appectively . We prevectively . Accordiectively ,24,25,26ectively ,28. So fectively ,29, wherectively . In thisImmunopeptidome data were retrieved from a previous publication . For immClinical and survival data of patients affected with CLL, whose malignant cells had been previously analyzed by immunopeptidome analysis , were cop values ≤ 0.05 were considered statistically significant.Data are displayed as box plots including median, 25th and 75th quartiles and min/max whiskers. Continuous data were tested for distribution and individual groups were tested by use of Kruskal–Wallis or Mann–Whitney U-test. If applicable correction for multiple comparison was done. Functional classification was performed of the source proteins of HLA class I-presented peptides by PANTHER 17.0 ,33. ProgWe analyzed the clinical, genetic and survival data of 57 CLL patients  in relatThe association of clinical characteristics of CLL patients with peptide presentation was investigated using mass spectrometry-derived immunopeptidome data of total as well as CLL-exclusive HLA class I- and HLA class II-restricted peptides. The presentation of peptides did not differ among patients with different Binet stages assessed at the primary diagnosis A. FurtheThe clinical phenotype of the patients was characterized in more detail and the presence of autoimmune phenomena , secondary tumors  and hypogammaglobulinemia was analyzed in relation to the immunopeptidome data. No difference in terms of total and CLL-exclusive number of peptides presented on CLL cells was observed among patients with or without any of these clinical features . p = 0.015 and p = 0.002, respectively, p = 0.042) . Patients with CLL cells presenting a more diverse immunopeptidome in terms of higher numbers of total and CLL-exclusive HLA class I-restricted peptides, experienced no more than one disease progression/recurrence compared to patients with a less diverse CLL-derived immunopeptidome who experienced multiple relapses (= 0.042) . Functiop = 0.027 and p = 0.022, respectively, Subsequently, we investigated the impact of immunopeptidome diversity on PFS and OS. For total and CLL-exclusive HLA class I-restricted peptide presentation, we observed a significantly improved PFS in patients with CLL cells presenting higher numbers of these peptides  presented higher numbers of total and CLL-exclusive HLA class I- as well as CLL-exclusive HLA class II-restricted peptides on their malignant cells compared to patients who present an aggressive disease course. Despite the previously reported stability of the immunopeptidome in CLL patients undergoing therapy, it could be hypothesized that the diversity of the immunopeptidome in CLL decreases in the disease course, due to its clonal evolution  and the In line with the observed effect on relapse numbers, we identified a prolonged PFS in CLL patients with a high number of different HLA class I-presented peptides  in comparison to patients with a less diverse CLL immunopeptidome. Despite the observed longer PFS of patients with a high number of HLA class I peptides, the HLA class I immunopeptdome showed no impact on patients’ OS after a follow-up of 10 years. To this regard, it is noteworthy that CLL is a chronic disease with OS ranging from 2 to 20 years ,4. There+ T cell-based immunosurveillance of CLL cells. In contrast, presentation of HLA class II-restricted peptides and, thus, CD4+ T cell-mediated immunosurveillance, which we recently found to associate with improved outcome in ovarian cancer [+ T cell responses and warrants further analysis in a larger cohort of CLL patients. Notably, an impaired immunogenicity, driven, e.g., by either HLA downmodulation or homozygosity of HLA alleles, is a well-known mechanism of tumor escape that leads to a poorer response to check-point inhibitors and shorter survival in B-cell lymphoma and solid tumors [In summary, the observations of this study together with previous functional characterization of CLL-associated peptides ,21 seem n cancer , appearsd tumors ,48. To td tumors , which sThese results corroborate our previous findings in CLL patients ,21, quesIn conclusion, our study demonstrates the association between the diversity of total and CLL-exclusive HLA class I-restricted peptides on CLL course and suggests a role of naturally presented HLA class I peptides for the immunosurveillance of CLL. This supports the further investigation of T cell-based therapies as complementary approach to the standard of care treatments, in order to improve patients’ quality of life and survival.


Genome-wide association study (GWAS) is the most popular approach to dissecting complex traits in plants, humans, and animals. Numerous methods and tools have been proposed to discover the causal variants for GWAS data analysis. Among them, linear mixed models (LMMs) are widely used statistical methods for regulating confounding factors, including population structure, resulting in increased computational proficiency and statistical power in GWAS studies. Recently more attention has been paid to pleiotropy, multi-trait, gene–gene interaction, gene–environment interaction, and multi-locus methods with the growing availability of large-scale GWAS data and relevant phenotype samples. In this review, we have demonstrated all possible LMMs-based methods available in the literature for GWAS. We briefly discuss the different LMM methods, software packages, and available open-source applications in GWAS. Then, we include the advantages and weaknesses of the LMMs in GWAS. Finally, we discuss the future perspective and conclusion. The present review paper would be helpful to the researchers for selecting appropriate LMM models and methods quickly for GWAS data analysis and would benefit the scientific society. Genome-wide association study (GWAS) is the most popular strategy for dissecting complex traits of agronomical significance and human diseases with the rising of cutting-edge microarray and next-generation sequencing (NGS) tools and the development of linear mixed models ,2. GWAS The major problems in GWASs are confounding factors, including population stratification, familial correlation, and relatedness among individuals ,20,21,22Recently, several papers have been published to highlight the importance of LMMs in GWAS ,30,31. TLMMs can solve different problems, including population stratification, family structure, cryptic relatedness, estimating polygenic effect, and missing heritability . The LMMs were originated to account for multiple levels of relatedness by using a kinship matrix, which greatly enhanced the performance of GWAS by reducing both the false-positive and the false-negative rates . LMMs iny is an Many LMM methods have been proposed and applied in GWAS according to the recent advancement ,23,27,32 defined  as folloStudies confirmed that the methods of controlling the population structure and the confounding factors had better performance than those that did not consider confounding factors ,21. For p-value for significance tests [p-values obtained by linear regression, then constructs a genetic similarity matrix involving SNPs until it detects the first minimum in the GC factor (λGC). The GRAMMAR-Gamma method has been proposed as an analytical estimate within the basis of the score test technique [RMLM (random-SNP-effect MLM) considers the SNP-effect as random and permits using Bonferroni correction to estimate the ce tests . The idece tests . ECMLM  extracted a small subgroup of SNPs and applied them in FaST-LMM . SUPER fEMMAX > EMMA > CMLM/P3D > Fast-LMM > GRAMMAR > GMMAT > RMLM > GRAMMER-Gamma. These orders are based on the most cited to less cited models, and this style is followed for all other cases to suggest the models that may be chosen by the researchers in this manuscript. The single-locus model and their respective software and packages for GWAS using LMM are given in Most of the methods perform a single-dimensional genome scan by testing one single marker at a time, where multiple test adjustments are needed for the cut-off value of the significance test. Several single-locus methods such as EMMAX , P3D 2020, FaST-Multi-locus methods consider all loci info together and do not need multiple test corrections due to the nature of multi-locus . Some muA multi-locus model can be written as the extended version of Equation (1) followed by the previous study, defined ,43 as foing loci . Resultsing loci . LMM-Lasing loci . It perming loci . It is ting loci . It can ing loci .mrMLM (multi-locus RMLM) uses markers selected from the RMLM method with a flexible selection criterion, and is more reliable in QTN discovery and more precise in the QTN effect estimation than the RMLM and EMMA . RecentlStepLMM (stepwise LMM) is a consistent, versatile, and computationally proficient method that can be applied to both GS (genomic selection) and GWAS . It usedBOLT-LMM > MLMM > FarmCPU > mrMLM > FASTmrEMMA > LMM-Lasso. Researchers could use these methods based on their research interests or data types. The multi-locus models and their respective software and packages for GWAS using LMM are given in jth phenotype. According to the single-locus LMM model in Equation (1), each phenotype follows a multivariate normal distribution with mean +jth phenotype. The details about the multi-traits model and variance-covariance matrix calculation can be found elsewhere [Multivariate LMMs are commonly used to assess the association between SNPs and multiple correlated phenotypes in genetics due to their effectiveness in controlling relatedness amongst samples . Many mulsewhere .Korte, Vilhjalmsson  initiallHowever, while various multivariate methods are proposed to discover variants linked to more than one phenotype, these existing approaches do not investigate the population structure . GAMMA , LRT-based VC studies ,19,27 arGEMMA > WOMBAT > BOLT-REML > MTMM > LIMIX. The widely used method’s recommendation could help the users and readers make a quick decision and save time in analyzing their GWAS data using a multivariate/multi-traits LMM model. Multi-trait/multivariate model and their respective software and packages for GWAS using LMM are given in k-th subject and h-th environment can be written by the following LMM [μ is the population mean; h-th ethnic population; i-th locus effect with coefficient i-th locus and the h-th environment with coefficient h-th environment with coefficient k-th individual in the h-th environment. The details about parameter estimation and test statistic for GG and G E interaction models can be found elsewhere [Though many optimistic results have been produced using the different methods in GWAS data analysis, it has been recognized that additive effects can elucidate only a portion of genetic variations . Epistaswing LMM :(4)ykh= lsewhere .p-values for the best model can be estimated after randomly permuting the traits under the assumption of familial correlation-free traits in this step [FAM-MDR, a multifactor dimensionality reduction technique, detects epistasis in minor or extensive pedigrees . It aggrhis step . Simulathis step . Zhang, his step  developehis step . A study showed that the LMMs were unable to control the inflation of test statistics for G × E but were only capable of handling population structure when considering the genetic relatedness in the model . To overx traits .G × E interaction can detect the genetic effects, which are avoided in the linear models, enhance the GWAS power, and give the fractional answer to the missing heritability ,103. AnoVarious phenotypes and environmental variables such as nutrition, physical exertion, or lifestyle covariates can help the G × E interaction study . The stuGWAS has been effectively used for discovering various genetic variants linked with complex traits/diseases . HoweverSMART  is based on the extension of LMM . This mo2 methods proposed using summary statistics GWAS data rather than individual-level data [2 has some benefits over CoMM. For example, CoMM-S2 is computationally more proficient than CoMM when using larger sample sizes. The authors showed that CoMM-S2 performed better when the cellular heritability was small [2 cannot be applied in a cross-tissue study. Additionally, CoMM-S2 cannot differentiate whether the discovered genes are only correlated with the complex traits or if they are genuine causal effects [Real data analysis demonstrated that CoMM could identify more genetically governed genes associated with complex traits deprived of excessive type I errors. However, CoMM is an effective method, but it uses individual-level GWAS data and cannot entirely use extensively existing summary statistics data in GWAS . CoMM-S2vel data . This meas small . However effects . 2 > LSMM. TWAS and longitudinal-related LMM models, software, and packages for GWAS are given in Numerous GWASs have been implemented in population cohorts that have repeated measures at multiple time points for each individual ,118,119,Many LMM-based packages have been developed in GWAS . DMU is Linear and logistic regression models can be fit using this software, which permits accommodating millions of regression models on a desktop by employing an effective execution, concurrent, and out-of-core data processing for big datasets. ASReml, a statistical package, utilizes LMMs by REML for big datasets with complex variance frameworks . Many vaSingle-RunKing, an efficient R software, has been proposed to speed up the GWAS by LMMs . It usesMany software and server-based tools have been developed for multi-omics data analysis in GWAS. Qxpak is a software-based mixed-model, which allows a versatile tool for QTL mapping in various populations, including cross-between inbred lines and within-population analysis . AssociaQTLNetwork is a widely used software for linkage mapping and visualizing the genetic structure for complex traits, where analytical populations are derived from a crossing of different inbred lines . It can MASTOR  has been proposed for genetic association mapping a quantitative attribute . It can PEPIS (Pipeline for estimating EPIStatic) has been proposed to estimate polygenic effects based on the LMM . PEPIS uOSCA was proposed to manage omics data from high-throughput trials in big cohorts and help analyze complex traits utilizing omics data . OSCA usLMMs are attractive because they can control population structure and explain polygenic information for typical single-variant analysis in GWAS ,23,27,65Recently, multi-traits association mapping has received more attention as these methods provide more power and in-depth knowledge for dissecting the genetic architecture of complex traits . Many unThe statistical power of the multi-traits LMMs increases across traits by combining small genetic effects  and consThere are many benefits of using G × G and G × E interaction methods, including the detection of the genetic effects which are missed in the linear models, enhancing the GWAS power, and giving the fractional answer to the missing heritability. However, different G × G and G × E methods have limitations. For example, GxEMM has several limitations, such as it is very computationally intensive, considering Gaussian random effects, which reduce power; and it does not correct for G–E correlation, which is a familiar origin of bias in the fixed effect situation . AdditioGene expression data and GWAS are incorporated by the TWAS to discover gene-traits associations. TWAS methods are needed to overcome the limitations of the other methods. For example, most of the GWAS hits are in non-coding regions, and their biological explanation is unclear. Additionally, all information from GWAS proposes that complex traits are frequently controlled by many variants with minor or moderate effects. In contrast, a prominent part of risk variants with minor effects remain unidentified . A TWAS However, there are some major restrictions on the current LMMs approaches. Firstly, LMMs are computationally costly and require a long time to analyze big datasets compared with simple models. For example, the run time and memory needed by LMM models are the scale as the cube and square of the cohort size, respectively . SecondlThe LMMs have been applied in most aspects of GWAS, including population stratification and relatedness, resulting in computational proficiency and increased statistical power in GWAS studies. However, genomes sequencing has rapidly increased due to the development of NGS, and genomic datasets are growing progressively . This coMoreover, artificial intelligence (AI) is growing quickly due to its robust and stable application for resolving problems in conventional computing methods . FurtherThis review introduced the available LMMs methods on GWAS, including single locus, multi-locus, multi-traits, TWAS, longitudinal GWAS, packages, and software in omics data. It provides a practical explanation and guides the reader to fundamental references that allow for an advanced methodological feature and better comprehension of GWAS. It also assists in finding appropriate LMM methods for dissecting complex traits in GWAS and further help to investigate these methods using diverse NGS and omics datasets. This review could guide both the new scholars and those desiring to update their knowledge in the field of GWAS by applying LMMs using the omics data. However, there is no unique and sophisticated software that users would like, including flexible and easy to use, combining different types of omics data, and which can handle big GWAS data analysis much faster than the existing methods. Necessary software and packages should be developed for analyzing big GWAS data sets and marker derivative kinship matrices. Overall, there is much scope to utilize the LMMs in diverse fields, including biostatistics, bioinformatics, and statistical genetics, which could be helpful for medical scientists, agriculturists, technologists, and data scientists to solve real-world problems.


The most recent international guidelines regarding recurrent pregnancy loss (RPL) exclude most of the immunological tests recommended for RPL since they do not reach an evidence-based level. Comparisons for metanalysis and systematic reviews are limited by the ambiguity in terms of RPL definition, etiological and risk factors, diagnostic work-up, and treatments applied. Therefore, cohort heterogeneity, the inadequacy of numerosity, and the quality of data confirm a not standardized research quality in the RPL field, especially for immunological background, for which potential research application remains confined in a separate single biological layer. Innovative sequencing technologies and databases have proved to play a significant role in the exploration and validation of cancer research in the context of dataset quality and bioinformatics tools. In this article, we will investigate how bioinformatics tools born for large-scale cancer immunological research could revolutionize RPL immunological research but are limited by the nature of current RPL datasets. RPL is defined as two or more pregnancy losses before the 24th week of gestation, according to the most recent international guidelines of the ESHRE (European Society of Human Reproduction and Embryology) in 2017 , 4, and in vitro in women affected by RPL , anti-nuclear antibodies (ANA), anti-ds DNA and anti-thyroid-peroxidase (anti-TPO) antibody activation. Dysregulation of the maternal immune response to fetal/trophoblast antigens has been demonstrated, including through killer immunoglobulin-like receptor (KIR), human leukocyte antigen (HLA), mannose‐binding lectin (MBL) and H-Y antigen regulation , associated with M1, Th1 and Th17 dominance, along with a diminished Th2 responses inhibiting IL-10 and G-CSF production, could confer a lack of immunosuppression due to misregulation of Treg amounts . In addigulation , 15.ESHRE guidelines exclude most of the immunological tests recommended for RPL, likely because they do not reach an evidence-based level. This is due to the lack of a broader consensus in the RPL scientific research setting, for which most of the studies cannot be compared for meta-analysis and systematic review since their data are not homogeneous in terms of definition, etiological and risk factors, diagnostic work-up, and treatments. Yet, not all the members of the consensus beyond ESHRE guidelines 2017 perfectly agreed with this document, as claimed in the document itself ., testifImmune tolerance at fetal-maternal interface shares common pathways with immune escape in endometrial cancer (EC), which leads to cancer progression. Nevertheless, what is finely regulated and limited in time and space at the maternal-fetal interface in terms of immune tolerance, is conversely aberrant and irreversible in EC immune escape . To inveFrom a biotechnological context, the vast majority of the available RPL experimental data is based on either signal-based-omics  or targeted phenotypic profiles . These approaches, as shown by many meta-analyses, are suffering from a lack of reproducibility or are limited by a mere phenotypic description (such as the cytokinome) without the mechanistic and complete reconstruction of the tumor-host microenvironment , 20  or on a custom signature matrix built on single-cell data. For each sample, immune cell sub-population proportions have been estimated with the default setup .via t-test. Immune cell signatures have been computed through a weighted geometric mean.Microarray data was already quantile normalized. Single-cell data normalization is carried out by CIBERSORTx along with the generation of the signature matrix. CIBERSORTx and Seurat packages were used respectively for deconvolution and single-cell data preprocessing. Comparisons between RPL and controls were computed We implemented an immune-cell activity analysis based on the expression level of genes secreted by immunological cellular subtypes according to the most recent data available from Vallvé-Juanico et. Al . We genevs. 10.1% p-value: 0.002), and significantly lower proportion of M2 macrophages  were observed. These results are in line with previous reports on aberrant immune responses in RPL . Then, we structured the resulting data frame according to CIBERSORTx input constraints and completed the procedure as indicated in the CIBERSORTx manual. The new RPL signature matrix was used to perform the deconvolution (median RMSE: 0.91). Once again, no clustering and/or statistically significant differences in immunological populations proportions were found (Since the LM22 signature matrix has been validated only on TCGA solid tumor datasets we also attempted to design an RPL-specific signature matrix based on 3 decidual single-cell profiles. For this purpose, we extracted decidual cell expression levels and calculated the mean for each immune cellular subtype for each patient. Out of the 8 original populations, only 5 were sufficiently profiled (NK cells, Tregs and other CD4re found Table 2.Traditional experimental assays are time-consuming, expensive, and laborious in the heterogeneity of cancer characterization. Thus, more efficient approaches have been introduced in cancer research to improve the traditional research settings to reach a precision/personalized medicine level. The availability of high-dimensional data has significantly improved cancer research and led to the employment of advanced modeling techniques, such as machine learning and deep learning. Such methodological improvements driven by cancer research have strong potential for discovery of new and complex unknown features , but wilPublicly available data of RPL cohorts showed quite a few limitations for data accessibility and reanalysis, for either format incompatibility with standard bioinformatics tools or quantitative/qualitative limitations of the cohorts. While the Lucas et al. NGS  cohort pAlong with data availability limitations, one major limitation in RPL research is that separate cohorts cannot be compared nor aggregated due to the prominent level of heterogeneity in RPL definition criteria, diagnostic work-up to perform, etiological factors to investigate and their own definition, as described in Li J . et al. single gene approach and the scarcity in terms of numerosity and quality of RPL studies. Research, development, and the definition of standardized procedures should be considered to produce adequate RPL databases, so to enable the application of new methodological approaches. For instance, extensive transcriptomics studies on the decidua will enable to deconvolute and pinpoint the specific cell populations that are dysregulated in RPL and compare their behavior to the best-known mechanisms of the EC immunological microenvironment.Our attempt reinforces the limitations already expressed by ESHRE 2017 guidelines on the impossibility to reach an immunological evidenced-based level, which is imputable both to the inadequacy of the https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE165004 SRA SRP052612 https://www.ncbi.nlm.nih.gov/sra?term=SRP052612https://ngdc.cncb.ac.cn/gsa-human/browse/HRA000237.Publicly available datasets were analyzed in this study. This data can be found here: GEO Series GSE165004 VB and MB contributed to conception and design of the study. VB, MB, and MP explored the available datasets. EP, AP, and EV contributed to the clinical interpretation of the results. MB and MP performed the bioinformatics and statistical analysis. VB wrote the first draft of the manuscript. MB, and MP wrote sections of the manuscript. VB, MP, and MB reviewed the manuscript. All authors contributed to the article and approved the submitted version.


Subsequent data stratification according to newborn birth-weight categories, i.e., small (n = 39), appropriate (n = 50) or large (n = 49) for gestational age , showed an interaction between GDM and birth-weight categories for intermediate-density lipoproteins (IDL)-cholesterol content and IDL- and low-density lipoproteins (LDL)-triglyceride content, and the number of medium very low-density lipoproteins (VLDL) and LDL particles specifically in AGA neonates. Moreover, in a 2-year follow-up study, we observed that small LDL particles were independently associated with offspring obesity at 2 years (n = 103). Collectively, our data demonstrate that GDM disturbs triglyceride and cholesterol lipoprotein content across birth-weight categories, with AGA neonates born to GDM mothers displaying a profile more similar to that of adults with dyslipidemia. Furthermore, an altered fetal lipoprotein pattern was associated with the development of obesity at 2 years.Abnormal lipid metabolism is associated with gestational diabetes mellitus (GDM) and is observed in neonates with abnormal fetal growth. However, the underlying specific changes in the lipoprotein profile remain poorly understood. Thus, in the present study we used a novel nuclear magnetic resonance (NMR)-based approach to profile the umbilical cord serum lipoproteins. Two-dimensional diffusion-ordered 1H-NMR spectroscopy showed that size, lipid content, number and concentration of particles within their subclasses were similar between offspring born to control ( Fetal growth and development constitute a particularly vulnerable period in life that is greatly affected by the maternal environment. Prenatal exposure to nutritional stressors has been associated with fetal programming, which can impact both metabolism and physiology and, consequently, predispose to later development of cardiovascular and metabolic diseases, including obesity . In thisBirth-weight is strongly determined by neonatal fat mass and gestational age, and fetal growth disorders can result from impaired maternal and fetal lipid metabolism. In fact, the levels and composition of cord blood lipids, apolipoproteins and lipoproteins are affected by both maternal and fetal factors ,5,6. DisDiabetic pregnancies are associated with a higher incidence of fetal growth disorders, and there is evidence that disturbances in maternal metabolism strongly contribute to these situations . ChangesInterestingly, 1H-nuclear magnetic resonance (1H-NMR)-based analysis of lipoproteins has established that the number of LDL and HDL particles is a more powerful index of cardiovascular risk than classical cholesterol determinations, given the large variability in the amount of cholesterol per particle and in particle size . 1H-NMR-Given the heterogeneity of growth patterns and the inconclusive findings in the cord blood lipoprotein profile of infants from GDM mothers, a comprehensive characterization of the main lipoproteins, including the assessment of the size and number of particles, is necessary to identify possible alterations in fetal lipoprotein metabolism and their potential consequences for fetal health. Thus, in the present study, we used the Liposcale test, a novel advanced lipoprotein assessment method based on 2D diffusion-ordered 1H-NMR , to examAll the mother–offspring pairs included in this study belonged to a carefully selected pre-birth cohort, and the mothers gave birth at the Department of Obstetrics of the Hospital Universitari de Tarragona Joan XXIII between June 2010 and May 2017. GDM and control women were recruited at their first prenatal visit to the Obstetrics Department and were followed until delivery. In this study, the participants were stratified according to the birth-weight category, and we included a similar proportion of infants born to mothers with GDM and control women who were AGA, SGA or LGA. The inclusion criteria at the end of pregnancy were: (1) singleton pregnancy, (2) accurate gestational age confirmed by an ultrasound examination before 20 weeks of gestation, (3) absence of fetal anomalies, (4) cord blood serum availability and (5) maternal lipid information from the third trimester exam. Neonates with major congenital anomalies, intrauterine infections or born from women with chronic and inflammatory diseases were excluded. One hundred and thirty-six neonate-and-mother pairs fulfilled these criteria (74 control pairs and 62 pairs with GDM) and were included in this study, which was performed in accordance with the tenets of the Declaration of Helsinki and whose protocol was reviewed and approved by the Hospital Universitari de Tarragona Research Ethics Board (ref: 243/2016). All participants provided informed consent before inclusion.All mothers were screened for GDM between 24–28 weeks of pregnancy following the Spanish Diabetes and Pregnancy Group recommendations . SubjectCare for GDM was managed according to the Spanish guidelines for diagnosis and therapy of GDM . Women wMaternal and umbilical cord blood samples were stored in a biobank collection along with the associated clinical data.2 and final weight/height (m)2, respectively. Similarly, gestational weight gain (GWG) was calculated as final weight—pre-pregnancy weight.Demographic and obstetric information on participants was collected via an interviewer-administered questionnaire, which paid particular attention to GDM risk factors. Maternal anthropometry included height, pre-pregnancy weight and weight at the end of pregnancy. Pre-pregnancy weight was self-reported and compared with the weight recorded in the first prenatal visit (before the 10th week of pregnancy) to ensure concordance. Pre-pregnancy body mass index (BMI) and final BMI were calculated as pre-pregnancy weight (kg)/height (m)3. Suprailiac skinfold thickness was measured within the first 48 h of life and was used to calculate the fat mass percentage. [Infant data included sex, gestational age, method of delivery and anthropometry. Neonatal length and weight were measured after delivery using a measuring board to the nearest 0.1 cm and a calibrated scale to the nearest 10 g. Ponderal index (PI) was calculated as birth weight (g)/length (cm)centage.  Neonatescentage. . InfantsHeight and weight information from birth up to 2 years of age was collected for 103 children. We defined obesity as a BMI ≥ 85th percentile according to age- and sex-specific BMI tables of the WHO growth standards .Umbilical cord blood was obtained immediately after delivery. Serum was immediately separated by centrifugation, divided into aliquots and stored at −80 °C until further analysis.Maternal fasting serum samples were obtained between gestational week 33 and 36 to determine glucose, triglycerides, total and HDL-cholesterol in an ADVIA 2400  autoanalyzer by standard enzymatic methods . LDL choCord blood serum samples were analyzed using the 2D diffusion-ordered 1H-NMR-based Liposcale test  . This tet-test or Mann–Whitney U test. One-way analysis of variance (ANOVA) was used to test differences among three or more groups. Potential interactions between GDM and birth-weight categories were assessed by two-way ANOVA followed by Bonferroni post hoc test to adjust for multiple comparisons. Spearman’s rank correlation coefficients were used for the analysis of the relationships between 1H-NMR-assessed lipoprotein profile and maternal and offspring metabolic and clinical variables. To control the false discovery rate (FDR), the Benjamini–Hochberg (B–H) procedure was used, and only those values significant with the B–H correction were considered [p < 0.05 was considered statistically significant.SPSS software v20.0  was used for statistical analysis. Data were presented as percentages for categorical variables, mean (±SD) for normally distributed continuous variables, and median (interquartile range) for non-normally distributed variables. Normal distribution of the data was tested with the Kolmogorov–Smirnov test. Non-normally distributed quantitative variables were used after log10 transformation, when required. For comparisons of proportions, differences between groups were analyzed using the chi-square test, while comparisons between normally and non-normally distributed quantitative variables were performed using unpaired nsidered . Logistip = 0.023) and fasting glucose  were lower in the GDM group of women treated only with diet compared to those who needed insulin therapy (data not shown). In the 1HNMR-assessed lipoprotein profile, no difference was observed between the two groups, except for a lower concentration of medium HDL-P in GDM treated with insulin compared with those treated with diet  (data not shown).Clinical and metabolic characteristics of the two groups, i.e., control and GDM, are shown in n = 62) and control (n = 74) mothers were categorized into three groups based on birth-weight categories according to age- and sex-weight specific charts (p = 0.321) (data not shown).One hundred and thirty-six neonates born to GDM  was highest in the SGA group and lowest in the LGA group, which was consistent with the differences observed among particle sizes . The number of LDL-P was lower in the SGA group than in the AGA and LGA groups, and the same distribution was observed for large and small LDL-P. The number of HDL-P was highest in the AGA group and showed an inverse U distribution when compared with the LGA and SGA groups. This phenomenon was observed specifically for small particles .While the cord blood lipoprotein profile was similar in offspring born to GDM and control mothers, we found some interactions when we assessed the effect of both birth-weight categories and GDM. AGA neonates born to GDM mothers had higher IDL-cholesterol and -triglyceride content, and higher LDL-triglyceride content than the SGA and LGA groups, in contrast to those born to control mothers who had lower concentrations. The same pattern was also observed with medium VLDL-P and LDL-P, which followed an inversed U distribution .Subsequently, we examined the relationship between the 1H-NMR-based lipoprotein profile and the maternal clinical and laboratory parameters as well as the neonatal outcomes. This analysis was conducted separately in both GDM and control groups, and results are shown in LDL-cholesterol concentrations were negatively associated with birth weight in the GDM group, while no association was observed in the control group. Cord blood insulin was strongly and positively associated with small and large LDL-P and LDL-cholesterol content in the control group, while a negative relationship was observed with VLDL-P and VLDL-triglyceride content in the GDM offspring.In both groups, neonatal adiposity was negatively correlated with cord blood VLDL-P, VLDL- and IDL-triglyceride content, and positively associated with cord blood LDL-cholesterol content and LDL-P, and these associations were stronger in the control group.2; p = 0.036), had higher birth weight  and were more exposed to GDM during the intrauterine life compared with normal-weight children . Additionally, they showed a higher number of cord blood small  and large LDL-P . No other differences were observed between the two groups. To further assess the independence of these associations, we performed logistic regression analysis. We found that small LDL-P were associated with infant obesity at 2 years after adjusting for potential confounders (p = 0.023), whereas large LDL-P showed a trend (p = 0.058)  . No diffBoth GDM and abnormal growth patterns have been associated with long-term adverse outcomes in offspring. Similarly, changes in the lipoprotein composition have been proposed as potential markers of cardiovascular diseases later in life. Taking advantage of a thorough lipoprotein profiling based on 1H-NMR, we showed for the first time that GDM modifies the umbilical cord blood lipoprotein profile in AGA neonates. In particular, GDM alters IDL lipoproteins, triglyceride content in LDL, and medium-size VLDL-P and LDL-P in those children. By contrast, GDM offspring belonging to the LGA and SGA groups have lipoprotein profiles more similar to those of controls. Additionally, we found that cord blood small LDL-P, known to be associated with atherosclerosis development, had predictive value for later obesity in the offspring.Both under and overnutrition in utero affects the lipoprotein profile of neonates ,5,6,7. SSince treatment with insulin at the end of pregnancy may activate placental nutrient transport to the fetus and promote placental fatty acid transfer , we asseDespite the differences observed in some lipoproteins and lipid particles in AGA infants of GDM women and controls, these differences were not seen in the rest of the lipoprotein particles or in the groups characterized by more severe alterations in growth, SGA and LGA, suggesting that in these cases, the pattern of growth and fat accumulation could dilute the effect produced by GDM. However, this has to be interpreted cautiously since evidence of cord blood lipoproteins as biomarkers for cardiovascular disease later in life is still scarce. In this regard, results in adult life are often extrapolated to fetal life, despite the fact that they might have a different interpretation. In fact, differences in lipoprotein composition between adults and fetuses have been described, including excess apoE on fetal HDL particles, which are large in size, lack paraoxonase I, and might have lower anti-oxidant capacity ,36, as wPrevious studies exploring the potential relationship between prenatal lipid metabolism and adverse metabolic outcomes in offspring have generated inconsistent results ,41,42,43There is evidence that an altered fetal lipoprotein profile is associated with aorta intima thickness in SGA and LGA neonates ,27, indiOne of the main limitations in observational studies is the inability to attribute causation between the observed associations. However, we considered several critical confounding variables to mitigate bias in the analysis. Thus, the main prenatal factors were addressed, and the groups were comparable for maternal BMI and birth-weight categories. Of note, to reach a sufficient sample size in the three birth-weight categories, the SGA and LGA groups were overrepresented, and further population-based studies are needed to determine the role of lipoprotein composition and subfractions in the pathogenesis of metabolic diseases in offspring. Furthermore, given the relevance of the placenta in the passage of nutrients and in the regulation of fetal metabolism and growth, it would be important to establish whether there is a parallelism between the lipoprotein pattern and placental metabolic pathways.The strengths of this study include a longitudinal birth cohort with almost complete maternal data that establish a temporal relationship between the outcome and the exposure to GDM. The novelty of the lipoprotein assessment, which allowed us to identify different fetal metabolic behaviors, is also a big asset in the experimental methods.GDM disturbs triglyceride and cholesterol lipoprotein concentrations across birth categories, with GDM-AGA neonates showing a profile more similar to that of adults with dyslipidemia and atherosclerosis than those born to normal glucose-tolerant mothers. Moreover, an altered fetal lipoprotein pattern is associated with obesity development at 2 years. Overall, these findings suggest that the fetal lipoprotein profile might be an early biomarker for the development of later diseases.


Transcranial direct current stimulation (tDCS)  also had a significant impact on post-stroke rehabilitation, with anodal stimulation  being more effective in improving spasticity in patients. This meta-analysis revealed moderate evidence that NIBS reduces spasticity after stroke and may promote recovery in stroke survivors. Future studies investigating the mechanisms of NIBS in addressing spasticity are warranted to further support the clinical application of NIBS in post-stroke spasticity.In recent years, the potential of non-invasive brain stimulation (NIBS) for the therapeutic effect of post-stroke spasticity has been explored. There are various NIBS methods depending on the stimulation modality, site and parameters. The purpose of this study is to evaluate the efficacy of NIBS on spasticity in patients after stroke. This systematic review and meta-analysis was conducted according to Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) guidelines. PUBMED (MEDLINE), Web of Science, Cochrane Library and Excerpta Medica Database (EMBASE) were searched for all randomized controlled trials (RCTs) published before December 2021. Two independent researchers screened relevant articles and extracted data. This meta-analysis included 14 articles, and all included articles included 18 RCT datasets. The results showed that repetitive transcranial magnetic stimulation (rTMS) (MD = −0.40, [95% CI]: −0.56 to −0.25,  Post-stroke spasticity, as a neurological manifestation with a typical syndrome of increased muscle tone, was reported to have a prevalence rate of up to 25% in stroke survivors . SpasticIn recent years, non-invasive brain stimulation (NIBS) has been actively explored in various diseases of the nervous system. Among various NIBS techniques, transcranial magnetic stimulation (TMS) and transcranial direct current stimulation (tDCS) are most often used to treat patients with post-stroke spasticity ,6. SpastCurrently, the effects of NIBS on post-stroke spasticity are contradictory. Although some studies have reported a beneficial effect of NIBS in the treatment of post-stroke spasticity ,12,13, oThis meta-analysis was performed in accordance with the PRISMA guidelines for systematic reviews and meta-analysis . The PICThe article search strategy is shown in All included RCTs were independently evaluated by two authors using the Cochrane risk of bias assessment tool . It inclFor each study that met the inclusion criteria, relevant information about experimental design and result analysis was extracted. All extracted information included research characteristics , treatment parameters  and main measurement results (MAS).A meta-analysis of the extracted studies was performed. Meta-analyses are useful for assessing the strength of evidence for treatment from multiple studies. The aim is to determine whether there is an effect, either positive or negative, and to obtain a single pooled estimate of effect rather than a single estimate of individual studies. In this meta-analysis, for each outcome related to continuous data, we calculated a pooled estimate and 95% confidence interval (CI) of the mean difference (MD) between the experimental and control groups after the intervention.2 index were used to assess the heterogeneity of all studies included in the meta-analysis. Statistical heterogeneity between these studies was calculated using Cochran’s Q test and the I2 index. An I2 index > 50% and p < 0.10 of the Cochran’s Q test indicated high heterogeneity, and the random-effects model was used; otherwise, the fixed-effects model was used. The results of all data analyses in this meta-analysis were shown by forest plots.This meta-analysis used RevMan 5.4  for statistical analysis. This was performed by entering the mean and standard deviation of all continuous data in each study into the software and calculating the mean difference (MD) of the 95% confidence interval (CI) to analyze the results. Cochran’s Q test and the IFunnel plots and Egger’s test to assess potential publication bias were applied. Still, because the number of studies included in each meta-analysis was less than 10, the funnel plot and Egger’s test could produce misleading results in this case . TherefoA total of 2482 publications were retrieved from two authors independently by searching the database. The search results are shown in Details of each study are provided in p < 0.01). The meta-analysis showed that there was no significant heterogeneity between the various studies   A.p < 0.01). However, although other studies had shown certain benefits, they did not reach statistical differences , and high-frequency rTMS (HF-rTMS), LF-rTMS combined with HF-rTMS and continuous theta-burst rTMS (cTBS) each had one study. The meta-analysis showed that compared with the control group, LF-rTMS had significant benefits for post-stroke spasticity, and the MAS was significantly reduced . However, stimulation of the affected hemispheres also had certain benefits but did not reach statistical differences . This meta-analysis showed that there was heterogeneity between different studies   A.p < 0.05); however, although cathode stimulation also had certain benefits, it did not reach a statistical difference   B.p < 0.01) and 1.2 mA  had significant effect on patients with post-stroke spasticity. However, the measurement results of other studies had changed but did not reach statistical differences  on post-stroke spasticity was also conducted.In terms of rTMS, the results of different subgroup analyses showed that LF-rTMS had a significant benefit in the unaffected hemispheres of patients with post-stroke spasticity B,C. ThisFor the stimulation types of tDCS, anodal stimulation has significant benefits for spasticity treatment in post-stroke patients B. In terIn the studies included in this meta-analysis, most of the brain regions stimulated by NIBS were the primary motor cortex ,27,28,30There are several different scales for assessing spasticity in post-stroke patients in rehabilitation studies. Currently, the MAS is used in most studies, and its main purpose is to evaluate abnormal muscle tone, while a small number of studies use the Modified Tardieu Scale (MTS) as a spasticity assessment tool . As the In patients after stroke, the balance between the two hemispheres of the brain is disrupted, resulting in hyperexcitability of the unaffected hemisphere and increased inhibition of the affected hemisphere . Most ofBased on this meta-analysis, the results of non-randomized controlled trials of NIBS for post-stroke spasticity were also discussed. At this stage, no other NIBS have been found in RCTs of patients with post-stroke spasticity, and new techniques still need to be explored in future studies.The results of the current meta-analysis are encouraging as they suggest that NIBS can promote rehabilitation in patients with post-stroke spasticity. At present, the NIBS applied to the field of post-stroke spasticity rehabilitation are mainly rTMS and tDCS. Other techniques, including transcranial alternating current stimulation (tACS) and transcranial ultrasound stimulation (TUS), still have limited evidence of significant variability in stimulation targets and stimulation parameters. Therefore, further in-depth study on the mechanism of action in the rehabilitation of post-stroke spastic patients is required. We hope that in the future, NIBS can be optimized and applied safely and efficiently to the rehabilitation of post-stroke spasticity.


Caenorhabditis elegans has helped revolutionize modern life sciences, leading to the pioneering work of programmed cell death, RNA interference (RNAi) and green fluorescent protein (GFP) and the wide-range applications of such technologies in biomedical areas. In the past decade, C. elegans has been increasingly used as a favorable and affordable in vivo animal model in pharmacological research and drug discovery, including target as well as lead identification. Indeed, C. elegans has already shown great power in the evaluation of potential therapeutics at both molecular and organismal levels owing to its short lifespan, relative simplicity and high degree of experimental tractability, which is further strengthened by significant conservation of disease genes and signaling pathways with humans. For example, transgenic C. elegans strains expressing human disease proteins have been generated to model neurodegenerative disorders such as Alzheimer’s, Parkinson’s and Huntington’s diseases, and have been used to screen for pharmacologically active compounds and to study their mechanisms of action . These studies not only exemplify the advantage of C. elegans in animal-based high-throughput screening but also provide an insight into the mechanisms of candidate compounds, demonstrating its transformative power in translational research for drug discovery and development. Due to its rapid life cycle and short lifespan, C. elegans can also be used to monitor long-term and even cross-generational outcomes of pharmacological interventions. Furthermore, C. elegans can also be conveniently used to study the overall efficacy, multitargeting capacity and potential interactions of component herbs of traditional complex formulae as recently demonstrated by formula dissection approaches  and α-synuclein and delay the onset of associated neurodegeneration in C. elegans models . The authors then showed that the lifespan-extending effect of complanatoside A was correlated with DAF-16/FOXO, SKN-1/Nrf2 and HSF-1. Interestingly, earlier studies have reported that astragalan, a polysaccharide isolated from the roots of Astragalus membranaceus, is able to alleviate neurotoxicity mediated by polyglutamine (polyQ) and extend lifespan of C. elegans in a DAF-16/FOXO-dependent fashion  imidazolium was identified to be effective against iron-induced oxidative stress in C. elegans, and the protective effect was shown to be associated with the heat shock transcription factor HSF-1 . The compound was further found to reduce the proteotoxicity in C. elegans models expressing the pathogenic proteins Aβ, α-synuclein or polyQ, including amelioration of Alzheimer’s disease-like paralysis and delay of age-related locomotion decline . These broad neuroprotective effects provide an important insight into the anti-neurodegenerative potential of the compound per se and a proof of concept for promising development of imidazole derivatives as novel therapies of age-related proteotoxic diseases. On the other hand, Mohankumar et al. found that α- and β-santalol, the natural sesquiterpenes from sandalwood, were able to stabilize the native state and potently inhibit the aggregation of transthyretin (TTR), another type of pathological proteins implicated in inherited amyloidosis. Specifically, the santalol isomers were shown to reduce wild-type and mutant TTR aggregates in C. elegans models expressing TTR fragments in body wall muscle cells and also extend both lifespan and healthspan of the nematodes by activating SKN-1/Nrf2, autophagy and proteasome pathways. Collectively these findings demonstrates their potential as therapeutic intervention against TTR amyloidogenesis and associated diseases .Wang and Zheng highlighted the advantages of C. elegans models in unravelling the microbe-host interactions that may regulate neurodegeneration, and identified pro- as well as anti-neurodegenerative microbial factors, including bacterial amyloid proteins. Therefore, targeting microbe-host interactions, e.g. targeting production of amyloid-like proteins in gut bacteria, is a potentially novel strategy in the discovery of neurodegenerative interventions as demonstrated in their screening work. In this regard, C. elegans can be conveniently used as animal models in high-throughput screening platforms for potential neurodegenerative therapeutics that target the microbe-host interactions (Wang and Zheng).Over recent years, it has emerged that the interactions between gut microbiome and the brain play an important role in neurodegeneration but when and how it happens remain largely unknown. C. elegans as an Emerging Model of Pharmacological Innovation” provided timely updates and state-of-the-art strategies through the expertise of contributing authors to explore the advantages of C. elegans as an alternative animal model in pharmacology. It is also our hope that this dedicated topic provides useful information and demonstrates important aspects justifying a broader use of this model animal in pharmacological research, including aging pharmacology, neuropharmacology, behavioral pharmacology, metabolic pharmacology, gut-brain interactions, pharmacogenomics, epigenetics and epigenome, target discovery, drug discovery, and ethnopharmacology.In summary, the article collection in the research topic “