Regorafenib is a tyrosine kinase inhibitor (TKI) approved in metastatic gastrointestinal stromal tumor (GIST), colorectal cancer, and hepatocarcinoma. Anyway, the toxicity profile of Regorafenib standard schedule is associated with poor compliance and a high rate of discontinuation. For this reason, there is a growing need for a Regorafenib personalized schedule emerging from the scientific community.The aim of this case series was to describe the experience of our sarcoma referral center with the continuous administration of Regorafenib as an alternative regimen to treat metastatic GIST patients.We retrospectively collected clinical, pathological, and radiological data of patients with metastatic GIST treated with daily personalized Regorafenib at a single tertiary referral center from May 2021 to December 2022.We identified three patients fulfilling the inclusion criteria. The average follow-up since the start of Regorafenib was 19.1 months (12–25 months). All three patients had started a standard third-line Regorafenib schedule according to guidelines. The reasons for switching to a continuous schedule were as follows: exacerbation of symptoms during week-off treatment in the first patient, a serious adverse event (AE) in the second patient, and a combination of both conditions in the third. After switching, none of the patients reported severe AEs, and they improved control of tumor-related symptoms. Two of the patients experienced disease progression after 16 months (9 months of which is continuous schedule) and 12 months (8.1 months of which is continuous schedule) of Regorafenib, respectively; the third patient is still receiving continuous Regorafenib at the time of writing, with a progression-free survival of 25 months (14 months after the modified schedule start).With a similar efficacy and lower toxicities, a daily, personalized Regorafenib schedule seems to be a promising alternative to the standard regimen for metastatic GIST patients, including the frail ones. Further prospective analyses are needed to confirm the safety and efficacy of such regimen. Despite overall rarity, gastrointestinal stromal tumors (GISTs) represent the most common subtype of mesenchymal tumors, with an incidence of 1.5/100,000 people/year worldwide. The median age at diagnosis is mid-60 years of age, with an equal distribution between men and women  is recommended in high-risk patients according to the risk assessment classifications and to the mutational pattern –6. In moHowever, approximately 20% of patients present with metastases at diagnosis , 8 and uFinally, Regorafenib represents the standard third-line treatment, based on the results of a phase 3 trial (GRID) , in whicThe efficacy of Regorafenib, an oral multi-TKI able to inhibit several kinases, including VEGFR1 to 3, TEK, KIT, RET, RAF1, BRAF, PDGFR and FGFR, is unfortunately afflicted by a high incidence of drug-related adverse events (AEs) and often requires personalized dose adaptations . MoreoveUnfortunately, resistance to Regorafenib eventually occurs. Resistance to anti-angiogenetic agents can be a consequence of genetic/epigenetic modifications in cancer cells and/or in tumor endothelial cells . In ordeBased on these biological and clinical data, we present our experience with a continuous schedule of Regorafenib.We retrospectively collected clinical, pathological, and radiological data of patients with metastatic GIST treated with continuous Regorafenib 120 mg/day after failure or intolerance to Imatinib and Sunitinib at Humanitas Research Hospital from May 2021 to December 2022. We anonymously collected data through the clinical records in an electronic database. All the patients signed an informed consent to the clinical research according to the institutional requirements.We included three patients who fulfilled the inclusion criteria. In the same period of time, no other GIST patient received Regorafenib standard schedule in our institute. Patients’ characteristics are summarized in In April 2018 a 50-year-old man without relevant comorbidities accessed the Emergency Room due to abdominal increased volume, pain, and worsening of performance status (PS) according to the Eastern Cooperative Oncology Group (ECOG). An abdominal ultrasound showed a large mass of 17 × 14 cm and a CT scan confirmed the lesion associated with peritoneal localizations. A fine needle biopsy allowed the diagnosis of GIST presumably from the small bowel, with a mitotic index of 10/35 HPF (high power field). The tumor was stained positive for CD34, DOG1, and Caldesmon. The molecular pattern showed a mutation in KIT exon 11 .He received first-line therapy with Imatinib since May 2018, with an almost immediate symptoms relief and improvement of PS. After a month, a CT scan showed a trend to a reduction of the known lesions and the best overall response (BOR) as partial response (PR) occurred after 14 months of treatment, with subsequent disease progression (PD) after 21 months. Therefore, second line-therapy with Sunitinib was started. The BOR occurred after 4 months as stable disease (SD) according to Response Evaluation Criteria in Solid Tumors (RECIST) criteria version 1.1  came to our institute in 2018 with a diagnosis of gastric GIST with liver metastases. According to his medical history, he had undergone a total gastrectomy for a gastric GIST in another hospital in 2010. The histopathologic report had confirmed the diagnosis with a positivity for CD117, CD34, and DOG1 and a mitotic index of 18/25 HPF. Molecular analysis had shown a mutation in KIT exon 11 [W557-v559>Y]. The first CT scan performed after surgery had revealed a single liver metastasis, so a systemic treatment with Imatinib had been started, achieving a complete response (CR). After a treatment discontinuation of a few months, multifocal liver progression had occurred, so the patient had restarted Imatinib, obtaining disease control.In April 2018, the patient, motivated by the long-lasting disease stability, accessed our institute to be evaluated for surgery and underwent multiple liver resections. The pathologic report detected four metastases of GIST, and the molecular pattern showed a mutation in KIT exon 11 [W557_V559>Y] and a new mutation in c-KIT exon 17 [D816G].A new liver relapse occurred 2 months after surgery and Imatinib was gradually restarted, obtaining a metabolic CR after 1 month of treatment and a morphological SD after 6 months. After a further liver oligoprogression, the patient underwent a new liver resection, with histological confirmation of two metastases with the same mutational pattern of the previously resected ones. Nearly immediately after surgery, a liver relapse was detected and a new line of treatment with Sunitinib was started, with a PFS of 9 months and SD as BOR obtained after 2 months. At the time of PD, the CT scan detected pulmonary, liver, and new peritoneal lesions. Third-line treatment with Regorafenib was started 160 mg/day with the classic schedule. The patient interrupted the therapy after only 1 month because of a symptomatic (G2) hypothyroidism due to an autoimmune thyroiditis that warranted steroids and hormone replacement. Over the Regorafenib interruption, a stereotactic body radiotherapy was performed on all the known metastases. After 2 months, Regorafenib was cautiously restarted at 80 mg/day and then gradually increased until reaching a 160 mg/day standard schedule. Due to gastrointestinal toxicities , the dose was reduced to 120 mg/day d1–21 q28, with a morphologic SD and a metabolic CR after 3 months. As a consequence of an intestinal bleeding and G3 anemia that required blood transfusions, Regorafenib was interrupted. At the complete recovery, considering the medical history of the patient, the previous toxicities, and the low BMI, we proposed the resumption of Regorafenib with a personalized continuous schedule of 80 mg/day. This schedule has been well tolerated, without requiring new interruptions and with no more AEs other than G1 (hand skin reaction), obtaining SD. The patient is still progression-free after 25 months of treatment  and in subjective and objective good clinical conditions.A 63-year-old man came to our institute in 2021 for a second opinion for an ileal GIST with metachronous metastases. As significant comorbidities, he had had a myocardial infarction in 1977 and a stroke without neurological sequelae in 2007. In 2019, an abdominal large mass of 13 × 4.4 × 15 cm was detected due to abdominal pain, and in October 2019, surgery of the lesion was performed in another center. The pathology report diagnosed a high-risk [Mettienen et al. , having achieved initial hematologic recovery and a clinical stabilization, in agreement with the patient in August 2021, standard schedule Regorafenib was started and progressively increased up to 120 mg/day d1–21 q28. The patient experienced a mild clinical improvement and radiologic SD, but reported a worsening of the symptoms  over the week-off treatment. Moreover, owing to G3 anemia that required multiple blood transfusions, the treatment had to be interrupted and restarted at the recovery with a personalized schedule. Thus, we proposed a Regorafenib continuous schedule (120 mg/day), obtaining a good tolerance and no more AEs ≥G2, with the only AEs being G1 anemia and G1 hand–foot syndrome. Global clinical conditions significantly improved with subjective wellbeing, radiologic SD as BOR after 1 month, and metabolic PR after 5 months. A radiologic and clinical PD occurred after 12 months of therapy with Regorafenib  Regorafenib, and the patient was hospitalized for best supportive care. Unfortunately, his conditions, compromised by abdominal pain and bowel obstruction, did not allow any further treatment and he passed away 2 months after the discharge.Regorafenib is approved as a third-line therapy in metastatic GIST, but it presents a challenging toxicity profile often requiring a personalization of therapeutic schedule. In the pivotal phase III GRID trial , 98% of Similar results were also reported in the phase III CORRECT trial  of RegorThe severe toxicity profile of Regorafenib across different cancers has also been highlighted by a systematic meta-analysis  that incThe published data were also confirmed in clinical practice as recently collected by Nannini et al. in a retrospective study conducted across several Italian sarcoma centers , 21, evaA continuous schedule has been rarely adopted in Nannini et al. , 21, altImatinib and Sunitinib treatment schedule is based on this rationale. The discontinuation of Imatinib results in early disease progression , so thatRegarding Regorafenib, a continuous schedule was explored in a phase I study showing a favorable clinical activity and safety profile . In a suPerhaps, to identify the correct dose and schedule of oral TKIs for every patient, a monitoring of drug plasma concentrations should be determined, as investigated in GIST patients on Imatinib  and receResistance to anti-angiogenic therapies is a significant challenge in oncology, as it leads to a lack of response and disease progression. It can develop due to genetic/epigenetic changes in cancer cells or endothelial cells and it can imply different mechanisms of tumor angiogenesis, as intussusceptive microvascular growth, vasculogenic mimicry, and vascular co-option. To overcome resistance, alternative therapeutic strategies have been explored, such as combining multiple anti-angiogenic drugs or anti-angiogenic drugs with immune checkpoint inhibitors, as it has been successfully done in other angiogenesis-dependent tumors such as renal cancer . In GISTAs for personalization of the cure, our cases support the previously reported data on continuous schedule Regorafenib, showing a comparable efficacy through a steady suppression of the oncogenic pathways and guaranteeing a better tolerance even in frail patients who had experienced serious AEs with standard schedule Regorafenib.Certainly, our analysis presents several limitations. Being a retrospective analysis, data can be affected by bias or missing. Secondly, the study was conducted at a single tertiary referral center, which may limit the generalizability of the findings to other populations or healthcare settings. Thirdly, the sample size is small, preventing us from applying statistical analysis and drawing robust conclusions. Lastly, the short follow-up may limit the assessability of long-term safety and efficacy of continuous schedule Regorafenib in this setting.Also, a personalized approach itself presents some limitations, such as the physician team’s expertise in identifying patients that could benefit the most from the schedule adjustment and the difficult generalizability of personalized schedules.On the other hand, a personalized schedule allows to take into account the patient’s needs and perspective, leading to an increased awareness of his cure plan and a better compliance.Our study provides initial evidence for the potential benefits of a continuous, personalized Regorafenib schedule, and these findings suggest that such a regimen may be a promising alternative to the standard, with similar efficacy and lower toxicities. However, further prospective studies with larger sample sizes and longer follow-up periods are needed to confirm the safety and efficacy of this treatment approach.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Internal Humanitas Research Hospital IRCCS committee. The ethics committee waived the requirement of written informed consent for participation.Conceptualization and first draft: AFB, MSG, RF. Data collection: MSG, RF, AL. Review: AFB, MSG, NG, AL, RF, LS, LVS, FD, FS, SLR, AS. Supervision: AFB, AS, AL. All authors contributed to the article and approved the submitted version.


Bletilla striata has the highest sensory evaluation score, with the key flavor substances ethyl caproate and N-heptanal containing 1.14% and 6.28%, respectively, and their ROAV values reaching 54.46 and 100.00. Additionally, the freeze-drying method can well retain flavonoids, polysaccharides, and phenolic components, while providing better antioxidant and antibacterial properties. The stove-drying method would make Bletilla striata slightly burnt and less flavorful and efficacious than freeze-drying; the air-drying method is difficult to retain the special odor and fragrance of Bletilla striata flowers and has the lowest sensory evaluation score, with the presence of volatile components with irritating and unpleasant odors such as pyrazine and 2-pentylfuran, while not showing better efficacy. In addition, steam fixation would destroy the morphology and flavor of Bletilla striata, lose polysaccharide and phenolic components, and reduce the efficacy of Bletilla striata scented tea, but could retain the flavonoid components well. In summary, direct freeze-drying without steam fixation is the best process for drying Bletilla striata scented tea, stove-drying without steam fixation is more economical and convenient in actual production and application, steam fixation and air-drying are not suitable as drying processes for Bletilla striata scented tea. This study analyzed the quality of Bletilla striata scented tea under different drying methods, promoted the further processing of Bletilla striata scented tea, and provided a reference for the comprehensive utilization of Bletilla striata scented tea.Flower tea is widely loved as a drink, especially for the beautiful and rich flowers of the orchid family, and the drying method for different flowers is also unique. GC-MS is widely used to study volatile substances to determine the quality of flower teas. The findings show that the freeze-drying method can retain the original aroma and flavor of  Bletilla striata (Thunb.) Reichb. f. is a rare Chinese medicinal herb of the orchid family in southwestern China, with the unique fragrance and flowering period of April to June. The flowers are light purple, which not only have high ornamental value, but also have certain pharmacological activities [Bletilla striata in Guizhou Province is more than 78 km2, and the yield of Bletilla striata flowers can exceed 60,000 kg per square kilometer [Bletilla striata scented tea has antibacterial, antioxidant, and other characteristics, and has potential anti-aging and antihypertensive activities [tivities ,2. At prilometer . Bletilltivities .Bletilla striata scented tea is widely accepted by people. Studies have found that the Bletilla striata flowers are rich in phenols, flavonoids, polysaccharides, and volatile components [Bletilla striata flowers are usually subjected to drying before transportation and storage. However, the capacity and technology of drying Bletilla striata flowers are insufficiently developed, which depends on the experience of self-harvesting and processing by local farmers. Without scientific guidance, such a raw drying process usually causes a waste of flowers, low quality, bad taste, and unstable batch-to-batch consistencies of scented tea. Some other drying methods like freeze-drying and stove-drying were also used in processing scented tea that better retain the natural fragrance, good taste, and potential therapeutic efficacy [Bletilla striata scented tea.Due to the unique fragrance and flavor, mponents ,6, whichmponents ,8,9. Bleefficacy . HoweverBletilla striata scented tea from three aspects: sensory, component, and effect. Bletilla striata scented tea after different drying methods was analyzed through sensory evaluation and gas chromatography–mass spectrometry (GC-MS). A variety of chemical substances, such as total flavonoids, polysaccharides, phenols, antioxidants, and antibacterial agents, was determined to reveal the relationship between Bletilla striata scented tea quality and different drying methods. This study advanced in-depth research and comprehensive utilization of Bletilla striata scented tea, which provided important practical guidance for its industrial production.This study explored the effects of different drying methods on Bletilla striata scented tea obtained under different drying methods was brewed, and the results are shown in Bletilla striata flowers in varying degrees, but No. 2 had some discoloration. Although No. 3 retained the original form of scented tea to some extent, its color faded to a brownish red. The color of No. 4 was slightly faded to light reddish brown, its flower shape was not good, and its fragrance was slightly light. No. 5 and No. 6 were close to each other, showing a black tea block with no fixed shape, and the special aroma of Bletilla striata flowers was not enough, but No. 5 showed a green tea aroma slightly, while No. 6 had no evident tea aroma. From the perspective of color, aroma, clarity, and taste of the tea soup, the No. 1 tea soup had a stronger aroma than the No. 2 tea soup, which better retained the original special aroma of the Bletilla striata flowers, and the aroma did not dissipate after brewing for 30 min. The No. 3 tea soup was yellow–brown, relatively clear, but the taste was not good, and the leaf bottom was relatively stretched, showing light brown. The No. 4 tea soup was pink, slightly light in aroma, slightly light in taste, and the leaf base was not stretched enough, showing a light pink color. The No. 5 tea soup was brownish red with a slight caramel flavor. After brewing for 30 min, the aroma was not evident, and the taste of the tea soup was not good. No. 6 was brownish yellow, slightly red, slightly turbid, not clear enough, the aroma of tea soup was not evident, the tea soup was sour and astringent, and the degree of the leaf base stretch was low.According to the sensory evaluation method of tea, the Bletilla striata flowers, while stove-drying will make the sugar aroma in the Bletilla striata flowers more prominent and slightly burnt, while air-drying is difficult to retain the aroma and taste of the Bletilla striata flowers. In addition, steam fixation will make the sugar in the flower more soluble and make the tea soup sweeter, but it will destroy the fragrance and shape of the flower and affect the appearance [After sensory evaluation, the results are as shown in pearance .Bletilla striata scented tea with different drying methods, including 34 alkanes, 2 olefin, 23 aromatic compounds, 5 esters, 10 alcohols, 13 aldehydes, 8 organic acids, and 8 other types. A total of 48, 55, 55, 51, 57 and 54 volatile components was detected from No. 1 to No. 6. It can be seen that the volatile components of Bletilla striata scented tea treated by different drying methods are different to some extent, and Bletilla striata scented tea is different from other scented tea in that its hydrocarbon compounds account for a high proportion of the total volatile components, from 19.97 μg/g~39.95 μg/g, and it is found that the mass fraction of hydrocarbon components is generally high after steam fixation, which may be due to the high mass fraction of polysaccharides in Bletilla striata flowers, and the high temperature process of steam fixation promotes the decomposition of polysaccharides and aliphatic hydrocarbons to produce more small molecular hydrocarbons [Bletilla striata scented tea on the whole because the threshold of alkanes is very high.As shown in ocarbons ,12, but Bletilla striata scented tea [Bletilla striata flowers, and aromatic components are also the main source of the special flavor of Bletilla striata flowers. From the results of GC-MS, steam fixation is not conducive to the retention of aromatic compounds, and the stove-drying method has the greatest difference before and after blanching. Ester composition is a representative component in flowers and fruits, which endows flowers and fruits with sweet flavor. Among them, ethyl caproate, the key flavor substance in esters, has the highest mass fraction in freeze-dried Bletilla striata scented tea. No ethyl caproate was detected in the No. 3 (air-dried) scented tea, resulting in the loss of this flavor. Alcohols are components with fresh and fruity aromas, among which phenylethanol and N-heptanol can provide the special fragrance of Bletilla striata scented tea, and phenylethanol is only detected in Bletilla striata scented tea without steam fixation. Because phenylethanol is a water-soluble alcohol component, it is speculated that it will be dissolved by a large amount of water and then lost in the process of steam fixation. However, N-heptanol was only detected in freeze-dried Bletilla striata scented tea. Aldehydes have a high mass fraction in Bletilla striata scented tea, among which 2-en-hexaldehyde, nonaldehyde, cinnamaldehyde, N-heptaldehyde, etc., provide modifications to the sweet and floral aroma of Bletilla striata scented tea. Acids generally have a rancid taste, but their threshold value is generally high, and their impact on the aroma of Bletilla striata scented tea is weak. Among the other ingredients, pyrazine was only detected in No. 3 (air-dried), which had an unpleasant and irritating odor.Aromatic compounds are a special kind of nted tea . Among tBletilla striata flower tea. Among them, heptanal has a similar sweet apricot and nut aroma; cinnamaldehyde has a cinnamon aroma and burning aroma; nonaldehyde provides a citrus aroma, fat aroma, and flower aroma; and ethyl caproate provides a fruit aroma. In addition, 2-en-hexaldehyde was the key flavor substance, except in No. 1 and No. 4, which indicated that stove-drying and air-drying can provide 2-en-hexaldehyde with a grass, fruit, and fat fragrance. However, 1-octen-3-ol was only used as the key flavor substance in No. 5 and No. 6, which indicated that the steam fixation has helped the production of 1-octen-3-ol, which had a green flavor, a vegetable flavor, and a mushroom flavor. The main flavoring substances with modification effects were p-methylphenol, phenylethanol, 2-pentylfuran, and pyrazine. Among them, p-methylphenol had a high mass fraction in the Bletilla striata scented tea without steam fixation, providing a tobacco aroma. Phenylethanol was only detected in the Bletilla striata scented tea without steam fixation, with the aroma of hyacinth and gardenia, while 2-pentylfuran and pyrazine were mainly detected in samples No. 3 and No. 6, which may produce unpleasant pungent odor.The ratio of the mass fraction of volatile flavor substances to their threshold value can more accurately reflect the contribution of this substance to aroma ,15. The The greater the ROAV, the greater the contribution to the overall aroma of the sample. The substance with 1 ≤ ROAV is defined as the key aroma substance of the sample, and the substance with 0.1 ≤ ROAV < 1 has an important modifying effect on the overall aroma.Bletilla striata scented tea.Through the ROAV results, it can be found that the aroma of scented tea is a complex special aroma, in which there are many key flavor substances and flavor substances with modification effects, and different drying methods and steam fixations, or not, have great impacts on the flavor of Flavonoids, sugars, and phenols are generally present in the flower, among which flavonoids, polysaccharides, and total phenols are generally its active components ,17,18. ABletilla striata scented tea was the lowest, which may be because the cells and related enzymes of scented tea do not completely lose their activity during the long time of air-drying, thus they will continue to produce respiration and consume the polysaccharide components. Whether freeze-drying or stove-drying, it greatly reduces the enzyme activity and saves a large number of polysaccharide components [As seen in mponents .Bletilla striata scented tea decreased significantly after steam fixation, which may lead to the degradation of total phenols due to the high temperature, as well as the dissolution of some water-soluble phenols after steam fixation. In addition, freeze-drying is the best way to retain total phenols, followed by stove-drying.As an important effective component in scented tea, total phenols mainly play the role of anti-tumor, anti-oxidation, antibacterial and improving immunity. From the results in Bletilla striata scented tea as a tea drink. From the result of Bletilla striata flowers occurred under high temperature during steam fixation. From the perspective of antioxidant activity, direct freeze-drying without steam fixation is the best drying method for Bletilla striata scented tea.Antioxidant activity is the most common and very important activity of general scented tea, which can delay the aging of human cells and remove free radicals from human body ,22. It iEscherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus were used for the evaluation of the antibacterial properties of Bletilla striata flowers treated with different drying methods. As shown in Pseudomonas aeruginosa, No. 1, No. 2, No. 3, and No. 4 had better effects, which may be due to the retained antibacterial substances without steam fixation. However, there was no significant difference in the inhibition of Staphylococcus aureus among the six groups, showing that drying methods had little effect on the components of anti-Gram-positive bacteria. For the inhibition of Escherichia coli, No. 1 was the best, No. 2, No. 3, No. 5 and No. 6 were the second, and No. 4 was the worst, indicating that freeze-drying can better retain the anti-Escherichia coli activity without steam fixation.Scented tea often has the function of clearing heat and detoxifying and is considered to have antibacterial effect in modern medicine . As the Bletilla striata were harvested from the Traditional Chinese medicine Resource Garden of Crop Research Institute of Guizhou in the middle of May 2022. All flowers were divided into 6 groups (>500 g per group). All chemicals were purchased from Sigma-Aldrich Ltd. unless otherwise stated and used without further purification . A DPPH free radical scavenging capacity test kit and an ABTS total antioxidant capacity test kit were purchased from Solarbio Life Science .Fresh flowers of Volatile substances were determined using 7890A-5975C GC-MS . Total flavonoids, total polysaccharides, and total phenols were determined using a HP8453 UV spectrophotometer (Aoyi Instruments (Shanghai) Co., Ltd., Shanghai, China). Freeze drying was determined using a CTFD-18S Freeze dryer . Drying was determined using a DHG oven .Bletilla striata flowers were rinsed with running water slightly to remove dust and soil and then placed in a cool and ventilated place, and water was drained until there was no evident damp feeling in the hands, numbered 1~6, respectively.Fresh Bletilla striata scented tea obtained in 1.3.1 on the steamer that has been aerated, and take it out after 3 min of blanching, while No. 1, No. 2 and No. 3 are not subject to blanching treatment. Freeze No. 1 and No. 4 Bletilla striata flowers in a refrigerator at −80 °C overnight, and then freeze them in a freeze dryer for 5 h. Put No. 2 and No. 5 Bletilla striata flowers in an oven at 50 °C for 12 h. Lay the No. 3 and No. 6 Bletilla striata flowers on the tray, and place them in a ventilated place to dry naturally for 5 days to obtain 6 groups of Bletilla striata scented tea, and put them in a refrigerator at −4 °C for cold storage.Put No. 4, No. 5 and No. 6 of the After placing the refrigerated samples at room temperature for 2 h and taking photos, evaluate the taste of the tea soup according to the Methods for Sensory Evaluation of Tea (GB/T 23776-2018). Accurately weigh 3.0 g of flower tea in a 150 mL review cup, add 150 mL of boiling water, cover and brew for 3 min, each 3 times in parallel, after brewing, filter out the tea broth in order of brewing in the evaluation tea bowl. The full score is 100 points, which is divided into five parts: appearance, aroma, taste, soup and leaf bottom evaluation. 100–90 points are excellent, 89–80 points are good, 79–70 points are average, 69–60 points are poor, and less than 60 points are not up to the requirements of tea drinking. The sensory evaluation team is composed of five members who have been trained in tea sensory evaluation.Place the refrigerated sample at room temperature for 2 h, grind the powder and pass it through a 200-mesh sieve, take a proper amount of sample and transfer it into a 15 mL extraction bottle, and seal it quickly. Age the SPME extraction fiber head to no impurity peak at 240 °C at the GC-MS sample inlet. Place the sample bottle on the solid phase microextraction device, set the temperature at 60 °C, and stir at 500 rpm. Put the sample bottle in the extraction device for preheating for 15 min, insert the SPME extraction head into the headspace part of the sample through the bottle cap, push out the fiber head, the extraction head is about 1.0 cm higher than the upper surface of the sample, and the headspace extraction is 50 min. Pull back the fiber head, pull out the extraction head from the sample bottle, insert the extraction head into the GC-MS sample inlet, push out the fiber head, analyze at 240 °C for 3 min, and then inject the sample for analysis.Chromatographic conditions: the chromatographic column is DB-WAX . The initial temperature of the chromatographic column is 40 °C for 2 min, rising to 85 °C at the rate of 3 °C/min for 2 min, rising to 110 °C at the rate of 2 °C/min for 2 min, rising to 160 °C at the rate of 5 °C/min for 1 min, rising to 220 °C at the rate of 5 °C/min for 5 min, gasification chamber temperature 240 °C, transmission line temperature 230 °C, carrier gas He, carrier gas flow rate 1 mL/min, no split flow.Mass spectrum conditions: EI source, electron energy 70 eV, ion source temperature 230 °C, quadrupole 150 °C, scanning mode Scan, scanning mass range 35 u~500 u.Qualitative analysis: MS database NIST11 and retention time are used for qualitative analysis of the detected components. The column loss peak should be deducted from the database screening results.N-decanol with a concentration of 100 μg/mL was used as the internal standard. The mass concentration of each component can be obtained by calculating the ratio of the peak area of each component to the peak area of the internal standard, then multiplying it with the mass concentration of the internal standard. The specific formula is as follows:Quantitative analysis: The NIST database is used to search, detect impurity peaks such as polyoxysilane (column loss component), and use the internal standard semi-quantitative method to carry out the quantitative results.ROAV was used to analyze the contribution of each volatile flavor substance to the overall aroma, and the odor activity value (OAV) was used to determine the substance that contributed the most to the aroma ,25, whicAccording to the results of OAV, the component that contributes the most to the aroma is defined as ROAVmax = 100, and other flavor substances are calculated by Equation (3):Put the refrigerated sample at room temperature for 2 h, beat the powder and pass it through a 200-mesh sieve, weigh 1.0 g accurately, add 20 mL of 80% ethanol, extract for 30 min by ultrasound, centrifuge and filter the residue to obtain the test solution, and determine the mass fraction of total flavanone according to the method in the Determination of Total Flavonoids in Exported Foods (SN/T 4592-2016). The mass fraction of the total polysaccharides shall be determined according to the method of the Phenol-Sulfuric Acid Method for the Determination of Crude Polysaccharides in Plant Derived Foods for Export SN/T 4260-2015), and the mass fraction of total phenols shall be determined according to the method of Zhang S. and Han X. [0-2015, ag min, centrifuge, filter the residue to obtain 20 mg/mL of test solution, and then use the dilution method to obtain 10, 5, 2, 1, and 0.5 mg/mL of test solution, and determine according to the methods described in the instructions of the total antioxidant capacity kit (DPPH method) and the total antioxidant capacity kit (ABTS method).Leave the refrigerated sample at room temperature for 2 h, beat the powder and pass it through a 200-mesh sieve, weigh 0.2 g accurately, add 10 mL of deionized water, ultrasonic extraction for 30× Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa as the test bacteria, the antibacterial effects of six samples were determined by the filter paper agar diffusion method and the co-culture method. Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa grow on the nutrient agar plate and can be stored for a period of time at 4 °C. When necessary, take the agar plate to room temperature, and rewarm it in a sterile environment for 40 min, then use the inoculation ring beside the flame to move a single bacterial colony into the LB broth, and shake it for 12 h at 37 °C to obtain the bacterial suspension, and compare it with the 0.5 # Maxwell tube (1.5 × 108 cfu/mL) dilute the above LB broth with normal saline to a similar turbidity, and then obtain a suspension of 108 bacteria. Weigh 10 mg of each sample and immerse it in 1 ml of PBS for 24 h.Using 8 cfu/mL of three bacterial suspensions, each 200 μL. Apply evenly on the nutrient agar plate, place the soaked sterile filter paper evenly on the agar plate, take PBS as the negative control, then incubate the plate upside down at 37 °C for 24 h, and record the diameter of the bacteriostatic ring.Immerse the sterile filter paper (diameter 6 mm) in the extract of two samples for 30 min, and absorb the above concentration of about 1 × 10The experiments were repeated three times, and the data were analyzed with the SPSS V22.0 software and plotted using origin 2018.Bletilla striata scented tea using sensory evaluation under different drying methods. Different drying methods indeed affected the shape, color, aroma, composition and efficacy of Bletilla striata scented tea. It was found that Bletilla striata scented tea had the highest score under the freeze-drying method without steam fixation. In addition, it was found that freeze-dried flower tea could better retain the original aroma and flavor of Bletilla striata flowers by making flowers slightly scorched. Through the analysis of the components in the scented tea, it was found that the aroma of the scented tea was a complex special aroma, in which there were many key flavor substances. The steam fixation and air-drying were not conducive to the retention of aromatic compounds, and, particularly, air-drying would lead to the loss of key flavor substances such as ethyl caproate. By contrast, the freeze-drying method benefits the retention of flavonoids, polysaccharides, and total phenols. From the perspective of efficacy, the freeze-drying method without steam fixation showed good effects in both antioxidant and antibacterial properties. In general, freeze-dried tea can better retain the flavor and effective components of Bletilla striata scented tea, and can provide better antioxidant and antibacterial effects. Stove-drying slightly changes the aroma of Bletilla striata scented tea to produce other flavors, which is more economical and convenient in actual production and application. Air-drying will reduce the quality of Bletilla striata scented tea. Steam fixation destroys the form and flavor of Bletilla striata scented tea, changes the effective components of Bletilla striata scented tea, and reduces the efficacy of Bletilla striata scented tea. In summary, direct freeze-drying without steam fixation is the best process for drying Bletilla striata scented tea, stove drying without steam fixation is more economical and convenient in actual production and application, and steam fixation and air-drying are not suitable as drying processes for Bletilla striata scented tea.This study studied


Aronia melanocarpa (Aronia) is a shrub with small berries, chokeberries. Chokeberries are claimed to possess health benefits due to a high content of polyphenols. Aronia is known to be extremely antioxidant; however, evidence for its health benefits is not established. This review gives an overview of the impact of Aronia on cardiometabolic risk factors and diseases.Seventeen studies on cardiometabolic risk factors and diseases were identified through a systematic search on PubMed, Embase, and Cochrane. Inclusion criteria were studies with Aronia as intervention, performed in individuals with cardiometabolic disease or risk factors, e.g., type 2 diabetes (T2D), cardiovascular disease, hypertension, dyslipidaemia, impaired glucose tolerance, overweight, central obesity and smoking. Four of these studies were applicable for a quantitative analysis.Aronia did not influence body weight, circulating triglycerides, total cholesterol, high-density lipoprotein (HDL) cholesterol, or blood pressure. The quantitative analysis revealed a mean reduction in blood glucose of 0.44 mmol/l (P=0.0001) in the treatment group compared with the control group suggesting that Aronia treatment may have a beneficial impact on blood glucose. In addition, treatment durations of 6 weeks to 3 months tended to decrease low-density lipoprotein (LDL) cholesterol, while shorter treatment durations had no effect on LDL cholesterol. The quantitative analysis did not provide data on long-term effects of Aronia on lipids.More long-term high-quality randomized controlled studies are needed to clarify if dietary supplementation with Aronia has beneficial effects on cardiometabolic diseases. ModifAronia melanocarpa is rich in polyphenols and compared to berries of other plants, its berries have the highest concentrations [Aronia melanocarpa may be effective in treatment and prevention of cardiometabolic diseases and related risk factors. Interestingly, no studies have reported adverse effects of Aronia melanocarpa consumption.trations . Due to Aronia melanocarpa is also called black chokeberry, which refers to the berry’s color and astringent taste. Aronia melanocarpa can be confused with Aronia prunifolia, which is a hybrid between Aronia melanocarpa and Aronia arbutifolia, so-called red chokeberry. These species all belong to the Aronia genus and Rosaceae family [Aronia melanocarpa berries, which will be referred to as Aronia throughout this review.e family . In thisAronia melanocarpa is a 1-2-meter-high deciduous bush [ous bush . Beside ous bush .in vitro [in vivo [Anthocyanins are highly antioxidant in vitro , and Aroin vitro  and in Rin vitro . During in vitro . However[in vivo .Sidor et al. provide an overview of the composition of compounds in Aronia berries . The berPolyphenols occur naturally in plants as part of the plants’ defense system . In the Anthocyanins in Aronia are primarily cyanidin derivatives with O-linked sugar moieties in a 3’-position. These sugars include the monosaccharides xylose, arabinose, glucose, and galactose. Cyanidin-3-O-galactoside is the far most abundant anthocyanin ,25,28-31Polyphenols are volatile after harvest, and a large part is lost during processing. Among polyphenols, anthocyanins are most prone to degradation . AnthocyThe highest concentration of polyphenols is found in the pomace ,48,66,67Aronia berries contain various minerals , cardiovascular disease, hypertension, dyslipidemia, impaired glucose tolerance, overweight, central obesity, and smoking, were included independently of study duration, study design, and Aronia dosing. Outcomes were biomarkers of cardiometabolic diseases including but not limited to blood pressure, blood lipids, and inflammatory markers. However, studies with Aronia administered in a combination treatment with other substances and only constituted a minor amount were not included. Only studies with a control group were included in the quantitative analysis. Studies not performed in humans and studies reported in other languages than English as well as studies performed in humans without cardiometabolic risk or risk factors were excluded.Two persons (CBC and FBM) reviewed all studies for eligibility – first, based on title and abstract, and second, on the full article. Mean ± SD values for post-treatment were extracted for both intervention and control groups, along with the number of participants in each group. Cholesterol, triglyceride, and glucose concentrations were given in mmol/l and HbA1c in percentages.Due to a limited number of studies in this area, results from studies that could not be included in the quantitative analysis have been reported applying a narrative approach. These studies comprise quasi-designed studies and cross-over studies.The quality of the studies included in the quantitative analysis was assessed by 2 persons (CBC and FBM) using the Cochrane Collaboration’s tool for assessing risk of bias . The ris2 statistics were used to evaluate heterogeneity between studies. I2 values of 25%, 50%, and 75% are considered low, moderate, and high heterogeneity, respectively.We used RevMan 5.4 for statistical analyses of differences in the post-treatment mean ± SD values between the intervention and the control groups. The random effects model was used to determine the overall effect, which was given as mean difference ± 95 % CI for the studies and illustrated in a forest plot. I3The systematic search identified 1059 records after removal of duplicates, of which 26 were potentially relevant, as 2) individuals with metabolic syndrome (MS) and one cross-over study with 37 hypertensive participants did not observe weight loss after treatment with 300 mg to 2000 mg extract/ day for 2 months [2 upon treatment with Aronia juice corresponding to around 600 mg polyphenols per day for 3 months [2 at baseline lost an average of 1.1 kg/m2 and had a reduction of 4.2 cm waist circumference after treatment with 100 ml Aronia juice per day, corresponding to around 600 mg polyphenols, for 4 weeks [Three quasi-designed studies with a total of 88 obese  concentration after administration of Aronia extract or juice for 6 weeks to 2 months ,95,96. OSeven quasi-designed studies performed in individuals with hypercholesterolaemia, MS, hypertension or T2D found that Aronia extract administered in daily doses of 300 mg to 600 mg polyphenols for 4 weeks to 3 months lowered total cholesterol ,92,95,96Overall reduction in LDL cholesterol after longer-term treatment with Aronia has been reported ,92,95,96Evidence suggests that Aronia extract can increase plasma polyunsaturated fatty acid (PUFA) concentrations. In individuals with cardiovascular risk, 4 weeks Aronia supplementation resulted in an overall increase in plasma PUFA in a RCT . In contAronia has shown positive effects on blood pressure in quasi-experimental designed studies. Out of 7 studies, all but one observed positive effect on diastolic blood pressure (DBP), and in all but 2, in systolic blood pressure (SBP) after supplementation with Aronia corresponding to around 300 mg to 800 mg polyphenols per day administered for 4 weeks to 2 months ,88,92,95ex vivo study using platelets from persons with increased cardiovascular risk, Aronia was able to reduce coagulation [Evidence suggests that Aronia can inhibit coagulation by platelet-mediated actions. Its anticoagulant activity has not been examined directly in humans, but in an gulation .In an RCT, 6 weeks treatment with Aronia significantly reduced C-reactive protein (CRP) from 4.5 to 3.5 mg/l in patients with coronary artery disease . In 2 otRCTs also have investigated Aronia’s effect on other inflammatory markers with discordant results. Loo et al.  detectedIn individuals with cardiovascular risk, an Aronia supplementation of 300 mg polyphenols per day for 2 months resulted in decreased lipid peroxidation of 40% . In a RCIncreased circulating fibrinogen levels are believed to be correlated to enhanced risk of cardiovascular diseases . In a qu2) being at cardiovascular risk, e.g., former smoking, previous myocardial infarction, hypertension, or hypercholesterolaemia [The impact of Aronia on overweight has been studied in 3 RCTs including 190 overweight persons . Due to the high risk of bias in one of the studies [With respect to blood glucose, a controlled study with 21 individuals with T2D, observed that treatment with 200 ml Aronia juice for 3 months decreased fasting blood glucose from 13.3 mmol/l to 9.1 mmol/l and HbA1c from 9.4% to 7.5% . Importa studies , a new aThe quantitative analysis detected no significant decreases in triglycerides, total cholesterol, LDL cholesterol, or HDL cholesterol after consumption of Aronia compared with placebo as The quantitative analysis did not detect any overall effect on blood pressure after supplementation with Aronia compared to placebo as 4Only 3 studies have examined the effect of Aronia on cardiovascular diseases directly, e.g., T2D and coronary artery disease ,99,104, Regarding the applied doses, Aronia extracts provide polyphenols in concentrated form, which means that participants only had to consume 3 capsules of Aronia extract each day, while Aronia juice was administered in larger amounts of 150 m to 250 ml a day. However, both amounts seem applicable in daily life. It also should be underscored that there is no information about the storage of Aronia juice or extract including recommendations to trial subjects on how to store it. As previously mentioned, polyphenols in Aronia, especially anthocyanins, are known to degrade even at room temperature. Light exposure also increases the degradation rate . If the Regarding weight loss, overweight individuals do not seem to benefit from supplementation with Aronia for shorter periods than 2 months. The long-term efficacy of the supplementation is unknown, but obese persons may benefit from long-term supplementation with Aronia extract in doses of ≥ 500 mg polyphenols/day . It is aWhereas only one study has investigated the impact of Aronia in persons with established cardiovascular disease , severalDyslipidaemia is often present in individuals with cardiovascular diseases, and T2D and constitutes a significant risk for cardiac events . It is qUnhealthy lifestyle, e.g., obesity, smoking, and alcohol consumption increase concentrations of inflammatory markers, of which especially CRP, TNF-α and IL-6 may contribute to the development of insulin resistance and cardiometabolic diseases . Low-graOxidation of LDL has a detrimental impact on human health, but whether its formation is simply a pathological process or triggered by oxidative stress remain uncertain . The forHowever, statin treatment may result in adverse effects, which makes it desirable to discover alternative, effective treatments that could lower oxLDL without inducing side effects . oxLDL dKnowledge regarding the uptake and metabolism of the Aronia polyphenols is limited. Some research studies indicate a poor intestinal absorption, which could account for the limited and divergent health benefits found in the literature. Denev et al. reviewed the studies on the absorption of the anthocyanins in Aronia, and even after consumption of a rather large amount of Aronia, only a small fraction of the anthocyanins can be found in the blood, typically in metabolized forms of which the health benefits are unknown . ConversThe risk of bias assessment clearly indicates that the overall quality of the studies included in the quantitative analysis is low. Of the studies included in this analysis, only one  reports Unfortunately, Narauszewicz et al. did not use a power calculation, which raises the possibility that the studies are underpowered and unable to detect an effect . SimilarOne also should note that age is an important moderator of risk of cardiac events, and the risk is substantially increased after the age of 60 . In relaIn conclusion, only few studies have examined Aronia’s impact on cardiometabolic risk factors and diseases. Studies focusing on participants with cardiovascular diseases rather than with increased cardiovascular risk are almost lacking, and the same goes for trials with persons with diabetes where no data are available on the effects of Aronia on insulin secretion and sensitivity. In persons at even higher CVD risk, more pronounced effects may be present. Even though most results are inconsistent, evidence points towards beneficial effects of Aronia on blood glucose levels, but further high-quality RCT studies are needed to allow a clear conclusion. The specific mechanisms of action of polyphenols on cardiometabolic risk factors are not clear. Future studies should be carried out as high quality RCTs performed as long-term studies in patients with cardiometabolic diseases rather than risk factors.


Lancet Planetary Health Diet (PHD) or the Finnish Nutrition recommendations (FNR) and anthropometric changes is scarce. Especially, the role of the overall diet quality, distinct from energy intake, on weight changes needs further examination.Knowledge on the association between the EAT-To examine the association between diet quality and weight change indicators and to develop a dietary index based on the PHD adapted for the Finnish food culture.n = 4,371, 56% of women, aged 30−74 years at baseline). Dietary habits at the baseline were assessed with a validated food frequency questionnaire including 128−130 food items. We developed a Planetary Health Diet Score (PHDS) (including 13 components) and updated the pre-existing Recommended Finnish Diet Score (uRFDS) (including nine components) with energy density values to measure overall diet quality. Weight, height, and waist circumference (WC), and the body mass index (BMI) were measured at the baseline and follow-up, and their percentual changes during a 7-year follow-up were calculated. Two-staged random effects linear regression was used to evaluate β-estimates with 95% confidence intervals.The study population consisted of participants of two Finnish population-based studies (Adherence to both indices was relatively low (PHDS: mean 3.6 points (standard deviation [SD] 1.2) in the range of 0−13; uRFDS: mean 12.7 points (SD 3.9) in the range of 0−27). We did not find statistically significant associations between either of the dietary indices and anthropometric changes during the follow-up , BMI: β 0.05 , WC: β −0.08 ; uRFDS, weight: β 0.01 , BMI: β 0.01 , WC: β −0.02 ).No associations between overall diet quality and anthropometric changes were found, which may be at least partly explained by low adherence to the PHD and the FNR in the Finnish adult population. Lancet Planetary Health Diet in the context of different food cultures and health outcomes is required. We developed a dietary index to measure adherence to the planetary health diet in the Finnish adult population independently of energy intake.Further research of the recently published EAT-In this study, diet quality was not associated with anthropometric changes during a 7-year follow-up.Low adherence to diets, however, may attenuate the associations between diet quality and health outcomes.2) and 13% with obesity (BMI 30 kg/m2 and over)  were living with overweight (body mass index [BMI]: 25−29 kg/mnd over) . ObesityThe fundamental cause of weight gain is a long-term imbalance between energy intake and expenditure. However, specific foods and food groups such as refined grains, red and processed meat, and sugar-sweetened beverages are possible mediators suggested in the recent systemic review and meta-analysis of 43 reports based on 25 prospective studies . This con = 34) conducted from 1990 to 2016 in adults worldwide, and in a systematic review of observational and experimental studies (n = 38) conducted from 2015 to 2020 in adults worldwide, higher adherence to dietary guidelines has been associated with a lower risk for obesity  was inversely associated with waist circumference (WC) and body fat percentage , to enable global scientific targets for a healthy diet obtained from sustainable food systems , 84%  participated in a health examination and interviews in 2000, and 63% of the invited  took part in the health examination of the follow-up study in 2011  . DILGOM 2007 and 2014, a sub-study of the National FINRISK 2007 Health Study  were invited to DILGOM 2007, of which 80%  participated in a health examination. In 2014, 82% of the invited  participated in the follow-up   . The stud n = 6,3 took par Fig. 1) .n = 57). Furthermore, participants whose daily energy intake corresponded to 0.5% at either end of the energy intake distribution were excluded from the DILGOM 2007 study (n = 48) (n = 18). The analytical study sample for this study comprised 4,624 participants ; DILGOM 2007/2014, n = 1,192 ).All studies included health examinations with anthropometric measurements and self-administered questionnaires . For the present study, all participants with a baseline food frequency questionnaire (FFQ) and baseline and follow-up information on weight, height, and WC measured by trained research nurses were included. Furthermore, we harmonized both cohort studies based on age so that participants who were ≥30 years old at the baseline and ≤81 years old at the follow-up were included (excluding 237 participants). Pregnant women at the baseline or follow-up were also excluded ((n = 48) , and parAll studies were conducted according to the guidelines of the Declaration of Helsinki, and the research protocols were approved by the Ethics Committee of the Hospital District of Helsinki and Uusimaa. A written informed consent was obtained from all participants.The FFQ inquired about the participants’ habitual diet over the last 12 months. The FFQ was originally designed in 1996 and has been updated and repeatedly validated against food records in the Finnish adult populations –27. The ®) (The frequency of consuming each food item (128–130 depending on the study) in the FFQ was reported in nine categories . The portion size for each FFQ-item was fixed and appeared on the FFQ as natural units (e.g. slice or glass). The average daily consumption of foods (ingredient level) and intakes of nutrients and energy were calculated using the FINESSI in-house software, which utilizes the Finnish National Food Composition Database   . TherefoLancet Commission (≤100 g/day) was chosen for cut-off value in the index, as especially freshwater fish with relatively low environmental impact is emphasized in the Finnish food culture.The cut-off values were based either on the lower, upper, or mean daily consumption target levels of the PHD . When chWe updated the original Recommended Finnish Diet Score , which wTo analyze the diet quality distinctly from the energy intake more precisely, we standardized the food consumption for the energy intake by dividing the participant’s mean daily intakes of each index components in grams by the total mean daily energy intake of the participant (except the fat ratio). Then, points were given based on the quartiles of consumption as grams per energy unit of each component . For whole grains, vegetables, fruits and berries, fish, and fat ratio, the lowest quartile of intake was coded as 0 points, the second as 1 point, the third as 2 points, and the highest quartile of intake as 3 points. For red and processed meat, salt, sucrose, and alcohol, the scoring was opposite. Thus, the total score could range from 0 to 27 points, where a higher total score indicated better adherence to the FNR 2014.2). The WC was measured with a flexible, non-elastic measuring tape from the mid-point of the lowest rib bones and iliac crest to the nearest 0.1 or 0.5 cm.At both the baseline and follow-up, the weight, height, and WC were measured by trained research nurses at the study sites according to standardized international protocols with the participants wearing light clothing and no shoes . The heiThe absolute change in the weight, BMI, and WC during the follow-up was calculated as follows: follow-up measurement – baseline measurement, where positive values indicated weight gain and negative values indicated weight loss. The percentual change was calculated as the absolute weight, BMI, or WC change during the follow-up divided by baseline measure of weight, BMI, or WC. Percentual weight, BMI, and WC changes were further categorized into four groups: ≤−5.0% (weight loss), −4.9% to +4.9% (weight stable), +5.0% to +9.9% (moderate weight gain), and ≥+10.0% .Sociodemographic and lifestyle factors, for example, education, leisure-time physical activity, and smoking status, were assessed using self-administered questionnaires. Education was harmonized to three categories: low , medium (graduated from upper secondary school or was in the middle tertile), and high (graduated from university or university of applied sciences or was in the highest tertile). Leisure-time physical activity comprising the activity outside work was harmonized into three groups: inactive , moderately active , and active or very active . Smoking habits were classified into three categories: never smokers, former smokers, and current smokers.Men and women were combined in the analyses, since no significant interaction was found between the sexes when the association was analyzed between diet quality and anthropometric measures. Follow-up years were standardized to 7 years between the studies because of the different follow-up periods of the included datasets (7 and 11 years). Anthropometric changes during the 11 follow-up years of the Health 2000 studies were first divided by 11 and then multiplied by 7 to harmonize it with the follow-up period of the DILGOM study.The baseline characteristics of the participants for the studies are presented as means with standard deviations for continuous variables or as proportions for categorical variables. Pooled descriptive statistics are presented by anthropometric change categories separately for weight, BMI, and WC. Furthermore, pooled intakes of carbohydrates, protein, and fat as percentual intake of total energy intake, energy-standardized fiber intake, and each PHDS component by the PHDS scoring groups are presented as means with standard deviations. Mean dietary index scores with standard deviations are also presented by sex (men or women), education level (high or low), and smoking status (current smoker and non-smoker). Variables that did not satisfy the normality assumption were log transformed with the natural logarithm.P-values were pooled with a two-staged random effects linear regression  for each dietary index (PHDS and uRFDS) at the baseline as a continuous exposure variable, and baseline anthropometric measures  and percentual anthropometric changes  as continuous outcome variables. Then, study-specific β-estimates and 95% CI with gression . The hetgression .n = 724) as these conditions are commonly known medical conditions linked to weight changes . The analysis of energy under-reporting was based on the calculation of ratio of reported energy intake and predicted basal metabolic rate, where the ratio ≤1.14 was considered to indicate energy under-reporting  was adjusted for age, sex, and the baseline anthropometric measure  depending on which of these was examined. In addition, the weight change was adjusted with the participant’s baseline height information. In the second model (model 2), model 1 was further adjusted for the baseline information on education, leisure-time physical activity, and smoking status. Additionally, model 2 was further adjusted for the participant’s baseline total energy intake to observe the effect of energy residues on the results, even though the energy intake was already considered when both dietary indices were composed. In the first sensitivity analysis, we excluded from the model 2 all participants reporting back pain, arthritis, cancer, diabetes, and depression or any other mental illness , and the pooling of the cohorts was carried out with the R statistical computing program, version 4.1.1, package meta . A P-val2 and 97 cm, respectively. For women, the mean BMI was 26 kg/m2, and the mean WC was 86 cm. Furthermore, participants in the highest weight gain (incl. BMI and WC) categories (≥+10%) tended to be more often women, younger, highly educated, current smokers, and physically inactive in their leisure-time compared to participants in the weight loss or weight stable categories .The pooled data included 4,371 participants (56% women). The mean age across the studies at the baseline was 49 years . The parP < 0.05) except the protein intake (P = 0.11) and the sucrose component (P = 0.946) . For example, the mean daily intake of carbohydrates and fiber and the consumption of vegetables, fruits and berries, and legumes were positively associated with the PHDS, whereas the mean daily intake of fat and the mean daily consumption of red and processed meat, poultry, and dairy foods were inversely associated. Furthermore, the mean daily consumption of fish and fish products reached the cut-off value in almost every total score group (1−9 points); however, the mean daily consumption decreased by a higher total score (P < 0.05).In the pooled dataset, the mean score for PHDS was 3.6 (SD 1.2) ranging from 0 to 11 points (the highest 10 and 11 points were received by only one participant each) when theoretical maximum could have been 13 points . FurtherP < 0.05) (data not shown). For example, the mean daily consumption of whole grains, vegetables, fruits and berries, and fish and fish products was positively associated with the uRFDS, whereas the mean daily consumption of red and processed meat, alcohol, and sucrose was inversely associated.Furthermore, the uRFDS ranged from 3 to 25 points (maximum of 27 points) with a mean score of 13.5 points (SD 3.8) . For theP = 0.03; uRFDS: β −0.002 , P = 0.03) but not with the baseline weight  or the BMI . However, there were no statistically significant associations between either of the two dietary indices and changes in weight, BMI, or WC during the 7-year follow-up (P > 0.05) except for a negative significant association between percentual change in WC and uRFDS , P = 0.02, model 1) (P > 0.05).The diet quality evaluated with both dietary indices (PHDS and uRFDS) was negatively associated with the baseline WC measures in the pooled datasets in model 2 , model 1) . HoweverMoreover, the results remained similar after further adjustment of model 2 for the total energy intake or exclusion of energy under-reporters or participants reporting any of the possible confounding health problems from the data (data not shown).This is the first longitudinal study in adults to examine the association between overall diet quality, independently of energy intake, evaluated with two different dietary indices based on PHD or FNR 2014 and anthropometric changes. In general, the adherence to a healthy diet assessed with both dietary indices was relatively low. No association was found between either of the dietary indices and the anthropometric measures at the baseline or relative anthropometric changes in the standardized 7 years of follow-up in the Finnish adult population.n = 14,151 men and women, aged 35−74 years)   found an association between better adherence to the healthy Nordic diet at the baseline and decreased weight and BMI during a 7-year follow-up  . It was = 4,720) . Further . It wasLancet Commission expressed their recommendations as energy density values. In contrast to the PHD, the FNR 2014 emphasizes the importance of adequate energy intake based on a basal metabolic rate and moderate physical activity without the specific common energy intake recommendations included in FBDG  to target overconsumption considering both human health and environmental aspects . In othe in FBDG . Furthern = 22,421) observed low adherence to the index with a narrow score range, which complicated distributing the participants to groups based on the index scores  aspects of sustainability , it is the first global attempt to define clear targets to ensure the same possibilities for survival for all now and in the future , 44, 45.In this population-based longitudinal study, no associations between the overall diet quality and anthropometric changes were found, independent of energy intake. As energy intake is one of the key factors in weight management, excluding it may have attenuated our findings. Our findings could also be explained by the low adherence to dietary indices and narrow score ranges, for which detecting differences between participants was infeasible. As knowledge on the environmental aspects of nutrition has been rapidly rising in recent years, further research is needed with more recent dataset.Click here for additional data file.


Recently, human bocavirus (HBoV) has appeared as an emerging pathogen, with an increasing number of cases reported worldwide. HBoV is mainly associated with upper and lower respiratory tract infections in adults and children. However, its role as a respiratory pathogen is still not fully understood. It has been reported both as a co-infectious agent , and as an isolated viral pathogen during respiratory tract infections. It has also been found in asymptomatic subjects. The authors review the available literature on the epidemiology of HBoV, the underlying risk factors associated with infection, the virus’s transmission, and its pathogenicity as a single pathogen and in co-infections, as well as the current hypothesis about the host’s immune response. An update on different HBoV detection methods is provided, including the use of quantitative single or multiplex molecular methods (screening panels) on nasopharyngeal swabs or respiratory secretions, tissue biopsies, serum tests, and metagenomic next-generations sequencing in serum and respiratory secretions. The clinical features of infection, mainly regarding the respiratory tract but also, though rarely, the gastrointestinal one, are extensively described. Furthermore, a specific focus is dedicated to severe HBoV infections requiring hospitalization, oxygen therapy, and/or intensive care in the pediatric age; rare fatal cases have also been reported. Data on tissue viral persistence, reactivation, and reinfection are evaluated. A comparison of the clinical characteristics of single infection and viral or bacterial co-infections with high or low HBoV rates is carried out to establish the real burden of HBoV disease in the pediatric population. Human bocavirus  is a parvovirus that was isolated ten years ago, mainly affecting the lower respiratory and gastrointestinal tracts in childhood all over the world. It is a small, icosahedral, linear, non-enveloped, single-stranded DNA virus measuring between 18 and 26 nm ,4,5.HBoV1 has been found primarily in samples from the respiratory tract, responsible for mild to severe upper and lower RTIs; in contrast, the variants (HBoV 2–4) have been identified in gastrointestinal tract samples and seem to be involved in the pathogenesis of gastroenteritis . Their tBocavirus derives from a combination of the words bovine parvovirus and canine minute virus, which have similar genetic and amino-acid structures [The name ructures ,19.Bocavirus usually occurs in infants and children aged between 6 and 24 months, but sometimes it has been found in children older than 5 years and adults .HBoV can be detected alone or, more commonly, with other viruses causing respiratory or gastrointestinal infections, such as rhinoviruses, adenoviruses, noroviruses, and rotaviruses ,4; viralThe pathogenicity of HBoV remains questionable because often this agent seemed to be a harmless passenger rather than a true pathogen ,29. HoweGiven the potential for serious and even fatal consequences from such an infection, promptly diagnosing an acute HBoV1 infection is essential for pediatricians.Several clinical studies have identified HBoV1 as one of the most detected respiratory viruses in young children with RTIs and, as with other respiratory viruses, it is widespread in both low- and high-income countries ,3. Most Most literature reported HBoV infection as prevalent in children between 6 months and 2 years with the highest detection rate during the second year ,12,13,14Infection can occur at any time of the year, with the highest incidence rate in winter and spring ,18,19,20The prevalence of HBoV depends on climatic and geographical factors, the type of sample, and the detection method used.The current worldwide prevalence of HBoV has been estimated to be 6.3% for respiratory infections and 5.9% for gastrointestinal ones . HoweverA recent systematic review on the epidemiology of HBoV in the Middle East and North Africa showed a mean value of prevalence in children of 3.94%, with the lowest in Iran (0%) and the highest in Egypt (56.8%) . In othePCR screening on an English cohort of children with RTIs identified a 9% prevalence of HBoV1 [In Europe, a recent meta-analysis by Polo et al., including 35 studies on 32,656 subjects from 16 countries, showed that HBoV prevalence varied from 2.0% to 45.69% with a pooled estimate rate of 9.57% . More reof HBoV1 ; in Croaof HBoV1 , and, inof HBoV1 . The lowof HBoV1 , and KanOnly limited data are available on the prevalence of HBoV in asymptomatic individuals since most studies have focused on children with different clinical symptoms of infectious diseases. However, recent publications described the detection of viral genomes in about 5% of respiratory samples obtained from asymptomatic children ,34. NevePCR and serological results. Maternal antibodies have been detected in 90% of infants younger than 3 months old, after which seropositivity decreases and reaches a low level by the age of 6–12 months [The exact prevalence of acute HBoV infections can only be determined using serodiagnosis and combining at least the 2 months . Then, H2 months .Bocaviruses are small non-enveloped viruses, with linear ssDNA genome. Because the genome has a bilateral hairpin structure at both ends, the full length of its genome cannot be accurately determined, although it has been estimated to be around 4.7–5.7 kb (5543 nucleotides) .There are three open-reading frames : ORF1  encodes non-structural proteins NS1–4; ORF2 is a smaller middle reading frame which encodes a unique nuclear protein, the nuclear phosphoprotein NP1, essential in viral DNA replication and mRNA processing ; and ORFThe virus can enter the host via the respiratory tract, through the bloodstream or via the fecal–oral route by direct ingestion . VerticaThe pathogenesis of human bocaviruses has been poorly studied . The HBoBesides the acute infection, HBoV is known to persist for a long time in lymphatic tissue and tissues afflicted with chronic sinusitis ,48.Current knowledge of the adaptive immune response, including both T- and B-cell-based cellular immunity evoked in response to HBoV infection, is inconclusive. HBoV viral capsid proteins (VP1 and VP2) have been described as immunodominant targets of the cellular and humoral immune response ,33. HighDuring the acute phase of the HBoV infection, the T-cell response is mediated mostly by CD4+ T cells rather than CD8+ T cells. The NK ratio has been found to be low, whereas the CD19+ CD23+ ratio, which is a marker of B-cell activation, is high, as expected for viral pathogenesis  and in vAfter the acute phase, HBoV may escape the host’s immune surveillance and establish a silent persistence. The immune-evasion strategies of HBoV involve type I IFN production or a TNF-a response. It has been demonstrated that HBoV nuclear phosphoprotein (NP1) suppressed IFN-β production by targeting the IRF-3 signalling pathway , while nRecent data published by Ivaska et al. analyzing tonsil tissue samples showed distinctively decreased T-helper17- and T-regulatory-type immune responses in local lymphoid tissue in HBoV-positive patients who underwent tonsillectomy. The authors demonstrated an inverse correlation between intra-tonsillar HboV1-DNA loads and cytokine expressions  . TogethePre-existing comorbidities which may represent a risk factor for a severe course of the disease , such asFurthermore, prematurity was reported in 11.6% of cases by Campelo et al., who, additionally, found that passive smoking was the principal risk factor in his cohort . HBoV1 wHBoV infection is described as affecting the lower respiratory tract predominantly ,57,58,59Escherichia coli or Salmonella sp.) have been frequently reported, with up to 77.6% of HBoV-positive children. However, the nature of the relationship between HBoV and gastroenteritis remains unclear [The variants (HBoV 2–4) are currently associated with gastrointestinal symptoms, the most frequent being the loss of appetite and vomiting, followed by diarrhea and nausea . Co-infe unclear .Fever has been frequently reported, varying from 68% by Tang et al. , 65% by The most reported complications related to bocavirus infection were increasing respiratory distress leading to acute respiratory failure, followed by dehydration . In 878 Liao et al. reviewed 12 children with HBoV1-related severe respiratory infection admitted to the ICU, including a case described by himself ,74,75,76Unfrequently, the central nervous system is affected in children with HBoV infections with symptoms including seizures, encephalitis, and encephalopathy ,79,80,81The severity of the illness could be remarkable. In the study by Akturk et al., 33.3% required PICU admission, and 20% were mechanically ventilated . SeveralIn several studies, the median hospitalization length (LOS) was seven days ,4. AlkhaCo-infection of HBoV with other respiratory viruses or bacteria has been reported at different rates. A high rate of co-detection is a well-recognized characteristic of bocavirus infection, and it is probably the result of prolonged shedding of HBoV1 in the nasopharynx, for weeks or even months ,85.The mixed viral infection rate is relatively high in most studies on NP swab samples, with rates of 38.3% in Australia , 47% in S. pneumoniae. In 69.6% of the Chinese HBoV-positive cases, a dual or triple bacterial co-infection has been detected, with S. pneumoniae and/or M. catarrhalis being the most commonly found , followed by H. influenzae for single-infection, and S. pneumoniae and S. maltophilia for mixed-infections (both with a detection rate of 5.4%) [S. pneumoniae was 29.9% in the Chinese ARIs Child Surveillance Study for the period 2009–2019 [S. pneumoniae (10.22%), H. influenzae (5.9%), and S. aureus in 10 children (1.8%) [Escherichia coli being the most frequent (29% of co-infection), followed by Klebsiella pneumoniae (20%); the authors also noted that the bacterial co-infections were significantly more frequent in infants less than 1-year-old [Mycoplasma Pneumoniae infections have been frequently found, for example, at a rate of 14.5% by Tang et al. [In several studies, a bacterial co-infection was found most frequently for 009–2019 , and 13.009–2019 . In the g et al. .Some authors note that clinical symptoms are more severe in cases with multiple infections than those with mono-infections ,91. EskiThere have, however, been cases of life-threatening diseases caused by a mono bocavirus infection ,72,93 anThe diagnosis of HBoV infection has so far mainly been based on the detection of viral genomes present in human respiratory samples, although serum, stool, and urine samples are also used. Techniques include different molecular assays  employing numerous sets of primers specific for the viral genes NP1, NS1, and VP1/2 . The mosStool samples may be the specimen of choice for patients with gastrointestinal symptoms. It is well-known that viral shedding can be prolonged, owing to the persistence of HBoV DNA in the nasopharynx for months and even up to a year, which can hamper initial diagnosis ,85,97,984 to 108) of HBoV1 DNA copies/mL of NP secretions, expressed as cycle threshold (Ct) values < 25, are suggestive of acute HBoV1 infection [p < 0.001) and single detection of HBoV1 [The presence of a high HBoV1 DNA load or mRNA in respiratory secretion is often considered a marker of viral activity, and can be used to distinguish acute infection from DNA persistence : high confection . Oldhoffof HBoV1 ,31,99.6 copies/mL) on NP secretions by quantitative PCR or HBoV1 mRNA, serum positive IgM, low IgG avidity, and/or >4-fold IgG titre [Serology has a higher specificity but lower sensitivity . It has gG titre . To overIn conclusion, no test alone is sufficient in all instances; for accurate HBoV1 diagnosis in severe infections, molecular assays and serology of paired serum samples should be combined and considered together with other laboratory data, clinical features, and the time of symptom onset ,100,102.C-reactive protein (CRP) and white blood cells are generally normal or just slightly elevated during acute HBoV infection. However, in the study by Tang et al., 36.7% of children with an HBoV single-infection showed elevated CRP levels . The plaChest radiography frequently shows peribronchial or interstitial infiltrates, hyperinflation, or atelectasis . In one Yet, there is no clinically approved specific treatment for bocavirus infection, and no comparative studies on antiviral drugs have been carried out. To date, there has been a single case report of treatment in an immunodeficient child who was treated with cidofovir, specific for herpesviruses; in this case, HHV-6 viremia decreased, and the HBoV infection was successfully eliminated .As for many viruses, supportive therapy remains the mainstay of treatment for HBoV. This includes providing oxygen for hypoxia and bronchodilators for patients with wheezing , and antAlkhaff et al., in a study on HBoV-infected Saudi Arabian children, noted that those with co-infections seem less likely to require an oxygen supply, and postulated that co-existence with other viruses could result in a better immune response, a less severe disease course, and a less likely need for oxygen . In contSteroids, specifically prednisolone, proved ineffective in treating children with bocavirus . HoweverIn most cases, even without bacterial co-infection, empirical intravenous antibiotics were administered at high rates ranging from 93% ,10,21,55Despite the high prevalence of pediatric HBoV infections worldwide, the virus is still not well-recognized by many clinicians. Failure to detect HBoV as a viral pathogen responsible for RTIs can lead to an inconclusive diagnosis, prolonged hospital stays, and unnecessary antibiotic use, ultimately contributing to a burden on local and global health and the economy. Expanding routine testing through viral respiratory panels which include HBoV could reduce this health burden.An early and accurate diagnosis of the etiological agent would be helpful for the effective treatment and adequate management of children with dyspnea and/or gastrointestinal diseases. Numerous studies confirm that the detection of HBoV DNA can persist for months; therefore, this does not necessarily indicate a recent infection. To certainly diagnose an acute infection of HBoV, therefore, required a combination of HBoV high DNA load or mRNA on NP secretions and one of the following three conditions: serum positive IgM, low specific IgG avidity, or >4-fold IgG titre .It has long been debated whether bocavirus was an autonomous pathogen or acted as a “passenger” since, in a significant percentage of cases, it has been found associated with other viruses and/or bacteria.However, although most HBoV infections mostly have a benign course, the infected children  could have a severe disease course at a not-marginal rate, requiring PICU admission and invasive ventilation ,21,81. OFuture lines of research should deepen our knowledge of the pathophysiology, replication mechanism, and persistence of HBoV in tissues such as the tonsils and adenoids.Another area of future investigation will be the development of more rigorous diagnostic methods, including sequence-independent amplification techniques combined with next-generation sequencing platforms that will enable the rapid and simultaneous detection of new viral agents, including HBoV.Further studies are also needed to clarify the influence of multiple viral infections on the severity of bronchiolitis and the clinical impact of HBoV in bronchiolitis cases. The high prevalence of HBoV in double and potentially life-threatening infections requires a new in-depth investigation to better understand the prognosis. Overall, a better understanding of the natural course of HBoV infection, the search for an experimental model, and the development of a specific treatment regimen to prevent HBoV infections including vaccines are needed to optimize its management.Finally, an additional analysis should be performed on the changing epidemiology of the common respiratory pathogens, including HBoV, during and after the COVID-19 pandemic when several non-pharmaceutical interventions such as wearing masks, hand hygiene, and social distancing have been taken for infection control.