Short communication: Antiviral activity of subcritical water extract of Brassica juncea against influenza virus A/H1N1 in nonfat milk

Subcritical water extract (SWE) of Brassica juncea was studied for antiviral effects against influenza virus A/H1N1 and for the possibility of application as a nonfat milk supplement for use as an “antiviral food.” At maximum nontoxic concentrations, SWE had higher antiviral activity against influenza virus A/H1N1 than n-hexane, ethanol, or hot water (80°C) extracts. Addition of 0.5 mg/mL of B. juncea SWE to culture medium led to 50.35% cell viability (% antiviral activity) for Madin-Darby canine kidney cells infected with influenza virus A/H1N1. Nonfat milk supplemented with 0.28 mg/mL of B. juncea SWE showed 39.62% antiviral activity against influenza virus A/H1N1. Thus, the use of B. juncea SWE as a food supplement might aid in protection from influenza viral infection.     

A(H1N1)流感病毒及抗病毒新药的筛选

A(H1N1)流感病毒是2009年3～4月在美国和墨西哥发现的一种流感病毒变异的新病毒株.这类流感疫情的蔓延引起了世界各国和世界卫生组织的严重关注.本文介绍了A(H1N1)流感新病毒株及感染这种病毒患者的症状,A(H1N1)流感病毒的致命力和传播,流感病毒变异对抗病毒药的抗药性,以及1918年流感大流行病毒聚合酶特性,人流感病毒和禽流感病毒聚合酶的结晶结构及其在感染中的作用.据此,提出了抗流感病毒药的筛选思路和研究方向,为抗流感病毒新药的设计和开发提供有益的参考.     

A(H1N1)流感病毒及抗病毒新药的筛选

A(H1N1)流感病毒是2009年3~4月在美国和墨西哥发现的一种流感病毒变异的新病毒株。这类流感疫情的蔓延引起了世界各国和世界卫生组织的严重关注。本文介绍了A(H1N1)流感新病毒株及感染这种病毒患者的症状,A(H1N1)流感病毒的致命力和传播,流感病毒变异对抗病毒药的抗药性,以及1918年流感大流行病毒聚合酶特性,人流感病毒和禽流感病毒聚合酶的结晶结构及其在感染中的作用。据此,提出了抗流感病毒药的筛选思路和研究方向,为抗流感病毒新药的设计和开发提供有益的参考。     

Anti‐influenza virus activity of extract of Japanese wasabi leaves discarded in summer

BACKGROUND: Japanese wasabi (Wasabia japonica) is now habitually used as a spice in some kinds of Japanese foods, and its pungent taste and flavor are preferred. Generally, rhizomes and winter leaves are used as a spice and for processed foods such as pickled wasabi. Since the leaf area of summer leaves is far greater than that of winter leaves, they are not used for food, and are discarded. Thus, we need to develop an effective use for summer leaves. We investigated anti‐influenza virus activity in these summer leaves as a new function. RESULTS: Seventy percent ethanol extracts of leaves harvested in July exhibited a high replication inhibition rate (98% or higher) in the type A strain (AH1N1, A/shimane/48/2002), its subtype (AH3N2, A/shimane/122/2002), and type B strain (B/shimane/2/2002). The extracts of summer leaves exhibited the same anti‐influenza virus activity as winter leaves, and showed a stronger activity than stems, roots, and rhizomes. CONCLUSION: A potent anti‐influenza virus activity was discovered in summer leaves of Japanese wasabi. The ethanol extracts inhibited influenza virus replication regardless of the hemagglutinin antigen type. Therefore, such extracts are expected to be a promising source of a novel anti‐influenza virus agent. Copyright © 2008 Society of Chemical Industry     

甲型H1N1流感病毒三维空间结构预测

采用从头计算思想,用改良的遗传算法对甲型H1N1流感病毒的蛋白质空间结构进行预测研究。基于蛋白质空间结构的HP模型,构建甲型H1N1流感病毒蛋白质空间结构的3DHP模型,并利用改良的遗传算法找到最小自由能结构,从而预测得到甲型H1N1流感病毒蛋白质三维空间结构。利用蛋白质空间结构数据建立距离矩阵,通过相关性分析和显著性检验,表明预测结构与已知结构存在高度的一致性。该模型提供了一种快速预测甲型H1N1流感病毒结构的方法。     

蛋类禽流感病毒检测报告

了解丽水市禽蛋标本禽流感病毒的感染状况。方法 对随机抽样的690只鸡、鸭、鸽蛋及126份农贸市场外环境对照标本采用荧光定量PCR方法检测禽流感病毒H7N9、H5和H9亚型核酸。结果 690只禽蛋标本H7N9、H5及H9亚型禽流感病毒核酸检测均为阴性,126份外环境对照标本H7N9亚型禽流感病毒核酸检测均为阴性,H5和H9亚型阳性标本各2份,阳性率为3.17%。结论 禽蛋中H7N9、H5及H9亚型禽流感病毒污染率低,但丽水市禽间存在H5和H9亚型禽流感的传播。各有关部门应加强合作,采取切实有效措施,防止人感染禽流感。     

Mumefural and related HMF derivatives from Japanese apricot fruit juice concentrate show multiple inhibitory effects on pandemic influenza A (H1N1) virus

Fruit-juice concentrate of Japanese apricot (Prunus mume Sieb. et Zucc.) has been shown to be effective against influenza A infection in MDCK cells. In this study, we isolated five components from the fruit-juice concentrate of Japanese apricot, 5-(hydroxymethyl)-2-formylfuran (HMF), 1-[5-(2-formylfuryl)methyl]dihydrogen 2-hydroxypropane-1,2,3-tricarboxylate (mumefural, MF), 2-[5-(2-formylfuryl)methyl]dihydrogen 2-hydroxypropane-1,2,3-tricarboxylate (MF‘), 1-[5-(2-formylfuryl)methyl]hydrogen 1-hydroxyethane-1,2-dicarboxylate (MA1) and 2-[5-(2-formylfuryl)methyl]hydrogen 1-hydroxyethane-1,2-dicarboxylate (MA2), and investigated their inhibitory activities against the novel influenza A/Narita/1/2009 (H1N1) pandemic virus hemagglutinin and neuraminidase functions, which are essential for viral attachment and budding, respectively. An hemagglutination inhibition assay indicated that MF and MF‘ were effective at minimum hemagglutination concentrations of 3.1 and 6.3 mM, respectively. An inhibition study for sialidase activity of the neuraminidase spike showed that MF was the most active anti-sialidase compound with an IC₅₀ value of 0.21 ± 0.01 mM, followed by MA2 (IC₅₀, 0.71 ± 0.09 mM), MA1 (IC₅₀, 1.64 ± 0.31 mM) and MF‘(IC₅₀, 1.62 ± 0.22 mM). Furthermore, MF was shown to inhibit the growth of the pandemic virus in a dose-dependent manner (62 ± 3% inhibition at 5 mM). The results suggest that MF, a citric acid ester linked to HMF at the 1-position of the propane backbone, might be a lead compound for the development of anti-influenza A inhibitors.     

新疆琐琐葡萄提取物抗流感病毒A(H1N1)亚型作用研究

目的对新疆琐琐葡萄提取物抗流感病毒A(H1N1)亚型作用进行研究。方法1.采用狗肾传代细胞培养法和MTT法研究新疆琐琐葡萄多糖、琐琐葡萄总黄酮、琐琐葡萄总三萜对抗流感病毒A(H1N1)型作用。2.采用Read-muench法计算流感病毒A(H1N1)型在狗肾传代细胞中的半数感染量。3.通过观察细胞病变(CPE)、计算细胞病变CPE抑制率、MTT法测定药物对细胞的保护率等来评价琐琐葡萄多糖、总黄酮、总三萜的抗流感病毒效果。结果琐琐葡萄总黄酮的最大无毒作用浓度(TC0)为20μg/ml,琐琐葡萄总三萜的TC0为1500μg/ml。体外细胞试验中,琐琐葡萄黄酮、多糖提取物表现出对病毒的直接抑制作用,但是琐琐葡萄三萜提取物和利巴韦林(阳性对照)此作用不明显。体外细胞试验中,琐琐葡萄黄酮、多糖、三萜提取物均表现出对感染细胞后病毒的抑制作用,三萜的抑制效果要强于黄酮和多糖,利巴韦林的抑制作用强于黄酮、多糖、三萜。结论新疆特色植物资源—琐琐葡萄提取物抗流感病毒A(H1N1)亚型的作用是多途径的,琐琐葡萄总黄酮、琐琐葡萄多酚、琐琐葡萄三萜分别表现出对流感病毒A(H1N1)亚型感染细胞前、感染细胞后的抑制作用,以及直接抑制作用...     

A Chromatographic Strip for Rapid Semi-quantitative Detection of H5 Subtype Avian Influenza Viruses in Poultry

An immunochromatographic strip was developed for rapid semi-quantitative of the H5 subtype of avian influenza viruses (H5 AIVs) in poultry. A specific monoclonal antibody 1C4 for H5 AIV hemagglutinin (HA) was produced and conjugated with colloidal gold as the detecting probes for the immunochromatographic strips, and a polyclonal antibody against H5N1 was used to firm the test line. Different from the traditional strips, four test lines with a same capture complex of different concentrations were designed on the nitrocellulose membrane. By contrast with the HA tests and the simulated infection experiment, the immune strip was proved to be specific for the detection of H5 AIV within 10 min and had the character to reflect the viruses infection degree in different organs of the infected poultry. It should be useful for diagnosing and semi-quantitative analysis of the H5 subtype of AIV.     

Characterization of N-glycan structures and biofunction of anti-colorectal cancer monoclonal antibody CO17-1A produced in baculovirus-insect cell expression system

Advantages of the baculovirus insect cell expression system for production of recombinant proteins include high capacity, flexibility, and glycosylation capability. In this study, this expression system was exploited to produce anti-cancer monoclonal antibody (mAb) CO17-1A, which recognizes the antigen GA733. The heavy chain (HC) and light chain (LC) genes of mAb CO17-1A were cloned under the control of P₁₀ and Polyhedrin promoters in the pFastBac™ dual vector, respectively. Gene expression cassettes carrying the HC and LC genes were transposed into a bacmid in Escherichia coli (DH10Bac). The transposed bacmid was transfected to Sf9 insect cells to generate baculovirus expressing mAb CO17-1A. Confocal immunofluorescence and Western blot analyses confirmed expression of mAb CO17-1A in baculovirus-infected insect cells. The optimum conditions for mAb expression were evaluated at 24, 48, and 72 h after the virus infection at an optimum virus multiplicity of infection of 1. Expression of mAb CO17-1A in insect cells significantly increased at 72 h after infection. HPLC analysis of glycosylation status revealed that the insect-derived mAb (mAb^I) CO17-1A had insect specific glycan structures. ELISA showed that the purified mAb^I from cell culture supernatant specifically bound to SW948 human colorectal cancer cells. Fluorescence-activated cell sorting analysis showed that, although mAb^I had insect specific glycan structures that differed from their mammalian counterparts, mAb^I similarly interacted with CD64 (FcγRI) and Fc of IgG, compared to the interactions of mammalian-derived mAb. These results suggest that the baculovirus insect cell expression system is able to express, assemble, and secrete biofunctional full size mAb.     

Inactivation of avian influenza virus by heat and high hydrostatic pressure

Avian influenza viruses threaten the life of domestic terrestrial poultry and contaminate poultry meat and eggs. Recently, these viruses rarely infected humans but had a high mortality rate in Southeast Asia, the Middle East, and Egypt. Thereby, these viruses caused high economic costs for production of poultry and health protection. We inactivated a highly pathogenic avian influenza A virus of subtype H7N7 in cell culture medium and chicken meat by heat and high hydrostatic pressure. Because heat and pressure inactivation curves of the H7N7 virus showed deviations from first-order kinetics, a reaction order of 1.1 had to be selected. A mathematical inactivation model has been developed that is valid between 10 and 60 degrees C and up to 500 MPa, allowing the prediction of the reduction in virus titer in response to pressure, temperature, and treatment time. Incubation at 63 degrees C for 2 min and 500 MPa at 15 degrees C for 15 s inactivated more than 10(5) PFU/ml, respectively. Thus, we suggest high-pressure treatment of poultry and its products to avoid the possible health threat by highly pathogenic avian influenza viruses.     

Formaldehyde in hair straightening products: Rapid 1H NMR determination and risk assessment

Despite official regulations, the illegal use of formaldehyde‐containing or releasing hair straightening products has become a popular practice in Europe and high contents of formaldehyde in such products have been reported. In this study, a methodology utilizing ¹H NMR spectroscopy has been developed to measure the concentration of formaldehyde in hair straightening products. For sample preparation, a dilution and alkaline hydrolysis is required. The total formaldehyde content can then be quantified by a distinct peak of the CH₂ group of the methanediol molecule in the δ4.84–4.82 ppm range. The developed methodology was applied for the analysis of 10 hair straightening products. Seven of these products contained detectable amounts of formaldehyde that were higher than the maximum allowed concentration of 0.2%. The formaldehyde content of these products was found to be in the range 0.42–5.83% with an average concentration of 1.46%. The accuracy and reliability of the NMR results were confirmed by the EU reference photometric method. The air formaldehyde concentrations after application of hair straightening products were estimated in ranges 20–423 ppm and 1–18 ppm (for 1 and 24 m³ salon volume). A probabilistic exposure estimation using Monte Carlo simulation found the average formaldehyde concentration to be 6 ppm (standard deviation 15 ppm). All exposure scenarios considerably exceeded the safe level of 0.1 ppm. Our findings confirmed that the risk of cosmetic formulations with formaldehyde above 0.2% is not negligible, as these products may facilitate considerable exposure of formaldehyde for consumers especially for salon workers.     

Integrating silicon nanowire field effect transistor, microfluidics and air sampling techniques for real-time monitoring biological aerosols

Numerous threats from biological aerosol exposures, such as those from H1N1 influenza, SARS, bird flu, and bioterrorism activities necessitate the development of a real-time bioaerosol sensing system, which however is a long-standing challenge in the field. Here, we developed a real-time monitoring system for airborne influenza H3N2 viruses by integrating electronically addressable silicon nanowire (SiNW) sensor devices, microfluidics and bioaerosol-to-hydrosol air sampling techniques. When airborne influenza H3N2 virus samples were collected and delivered to antibody-modified SiNW devices, discrete nanowire conductance changes were observed within seconds. In contrast, the conductance levels remained relatively unchanged when indoor air or clean air samples were delivered. A 10-fold increase in virus concentration was found to give rise to about 20-30% increase in the sensor response. The selectivity of the sensing device was successfully demonstrated using H1N1 viruses and house dust allergens. From the simulated aerosol release to the detection, we observed a time scale of 1-2 min. Quantitative polymerase chain reaction (qPCR) tests revealed that higher virus concentrations in the air samples generally corresponded to higher conductance levels in the SiNW devices. In addition, the display of detection data on remote platforms such as cell phone and computer was also successfully demonstrated with a wireless module. The work here is expected to lead to innovative methods for biological aerosol monitoring, and further improvements in each of the integrated elements could extend the system to real world applications.     

Monitoring of Escherichia coli O157:H7 in food samples using lectin based surface plasmon resonance biosensor

A novel surface plasmon resonance (SPR) biosensor using lectin as bioreceptor was developed for the rapid detection of Escherichia coli (E. coli) O157:H7. The selective interaction of lectins with carbohydrate components from bacterial cells surface was used as the recognition principle for the detection. Five types of lectins from Triticum vulgaris, Canavailia ensiformis, Ulex europaeus, Arachis hypogaea, and Maackia amurensis, were employed to evaluate the selectivity of the approach for binding E. coli O157:H7 effectively. A detection limit of 3 × 10³ cfu mL^?1 was obtained for determination of E. coli O157:H7 when used the lectin from T. vulgaris as the binding molecule. Furthermore, the proposed biosensor was used to detect E. coli O157:H7 in real food samples. Results showed that the lectin based SPR biosensor was sensitive, reliable and effective for detection of E. coli O157:H7, which hold a great promise in food safety analysis.     

Detection of 5 CFU/g of Escherichia coli O157:H7 on lettuce using activated charcoal and real-time PCR without enrichment

A sample treatment method which separates Escherichia coli O157:H7 from lettuce and removes PCR inhibitors allowing 5 CFU/g of target cells to be detected using real-time PCR is described. Lettuce leaves inoculated with E. coli O157:H7 were rinsed with 0.025% sodium dodecyl sulfate (SDS). In this study, there were two major factors that strongly affected the recovery of E. coli O157:H7 during sample preparation, the amount of bentonite coated activated charcoal used to remove PCR inhibitors and the agitated contact time of the samples with the coated charcoal. When 3.0 g of activated carbon coated with bentonite were mixed with target cell suspensions (30 ml) derived from 50 g of lettuce, a high recovery of E. coli O157:H7 (93%) was obtained. Sample agitation with bentonite coated activated charcoal for 15 min resulted in 95% recovery of E. coli O157:H7. When a commercial DNA purification resin was used for detection of E. coli O157:H7 without the use of the bentonite treated charcoal, the real-time PCR (Rti-PCR) failed to detect 1 × 10² CFU/g. In contrast, with the use of use of bentonite coated activated charcoal and a commercial DNA purifying resin together, Rti-PCR was able to detect 5 CFU of E. coli O157:H7/g of lettuce which was equivalent to 2.8 CFU/Rti-PCR. Such a successful detection level was the result of the bentonite coated activated charcoal’s ability to absorb the PCR inhibitors released from seeded lettuce during detachment. A standard curve was generated by plotting the Ct values against the log of CFU of target bacterial cells. A linear range of DNA amplification was exhibited from 5.0 × 10⁰ to 1.0 × 10⁴ CFU/g by using Rti-PCR.     

Quercetin,Hyperin, and Chlorogenic Acid Improve Endothelial Function by Antioxidant,Antiinflammatory, and ACE Inhibitory Effects

In recent years, the blueberry cultivation and processing industry developed quickly because blueberries are super‐fruit with healthy function. Blueberry leaves are byproducts of the blueberry industry, which are rich in bioactive phenolics, such as quercetin (Q), hyperin (H), and chlorogenic acid (C). This study investigated protective effects of 3 phenolics (Q, H, and C) from leaves of rabbiteye blueberry Vaccinium ashei on human umbilical vein endothelial cells. The results showed that all these 3 phenolics could improve endothelial function by inhibiting oxidative damage and proinflammatory cytokines caused by tumor necrosis factor‐α (TNF‐α). The cell vitalities of endothelial cells pretreated with Q, H, and C were higher than those stimulated with TNF‐α only. These phenolics could decrease reactive oxygen species and xanthine oxidase‐1 levels and increase superoxide dismutase and heme oxygenase‐1 levels in endothelial cells. They also could decrease the protein expressions of intercellular adhesion molecule‐1, vascular cell adhesion molecule‐1, and monocyte chemotactic protein‐1 induced by TNF‐α. In addition, Q, H, and C also exhibited vasodilatory effect by reducing the angiotensin I–converting enzyme (ACE) protein levels in endothelial cells. Mostly 3 phenolics exhibited bioactivities as a function of concentration, but the effects not always depended on the concentration. The antioxidant and antiinflammatory effects of Q seemed to be more pronounced than H; however, H exhibited higher cell vitalities. The results indicated that phenolics from rabbiteye blueberry leaves could be potential antioxidants, inflammation and ACE inhibitors, and rabbiteye blueberry leaves provide a new resources of phytochemicals beneficial for cardiovascular health.     

Effects of yogurt containing probiotics on respiratory virus infections: Influenza H1N1 and SARS-CoV-2

Respiratory virus infections are an escalating issue and have become common worldwide. Influenza and COVID-19 are typical infectious respiratory diseases, and they sometimes lead to various complications. In a situation in which no established drug or treatment exists, consumption of proper food might be beneficial in maintaining health against external infections. We studied the potential effects of mixtures of probiotic strains on various viral infections. The purpose of this study was to assess the ability of yogurt containing probiotics to reduce the risk of respiratory viruses such as influenza H1N1 and SARS-CoV-2 infection. First, we performed in vitro tests using infected Madin-Darby canine kidney (MDCK) and Vero E6 cells, to evaluate the potential effects of yogurt containing high-dose probiotics against influenza H1N1 and SARS-CoV-2 infection. The yogurt significantly reduced plaque formation in the virus-infected cells. We also performed in vivo tests using influenza H1N1-infected C57BL/6 mice and SARS-CoV-2-infected Syrian golden hamsters, to evaluate the potential effects of yogurt. Yogurt was administered orally once daily during the experimental period. Yogurt was also administered orally as pretreatment once daily for 3 wk before viral infection. Regarding influenza H1N1, it was found that yogurt caused an increase in the survival rate, body weight, and IFN-γ, IgG1, and IL-10 levels against viral infection and a decrease in the inflammatory cytokines TNF-α and IL-6. Although the SARS-CoV-2 copy number was not significantly reduced in the lungs of yogurt-treated SARS-CoV-2-infected hamsters, the body weights and histopathological findings of the lungs were improved in the yogurt-treated group. In conclusion, we suggest that consumption of yogurt containing probiotics can lead to beneficial effects to prevent respiratory viral infections.     

Fate of Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella on fresh-cut celery

Illnesses from Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella have been associated with the consumption of numerous produce items. Little is known about the effect of consumer handling practices on the fate of these pathogens on celery. The objective of this study was to determine pathogen behavior at different temperatures under different storage conditions. Commercial fresh-cut celery was inoculated at ca. 3 log CFU/g onto either freshly cut or outer uncut surfaces and stored in either sealed polyethylene bags or closed containers. Samples were enumerated following storage for 0, 1, 3, 5, and 7 days when held at 4 °C or 12 °C, and after 0, 8, and 17 h, and 1, and 2 days when held at 22 °C. At 4 °C, all populations declined by 0.5–1.0 log CFU/g over 7 days. At 12 °C, E. coli O157:H7 and Salmonella populations did not change, while L. monocytogenes populations increased by ca. 0.5 log CFU/g over 7 days. At 22 °C, E. coli O157:H7, Salmonella, and L. monocytogenes populations increased by ca. 1, 2, or 0.3 log CFU/g, respectively, with the majority of growth occurring during the first 17 h. On occasion, populations on cut surfaces were significantly higher than those on uncut surfaces. Results indicate that populations are reduced under refrigeration, but survive and may grow at elevated temperatures.     

Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 feeding enhances humoral immune responses,which are suppressed by the antiviral neuraminidase inhibitor oseltamivir in influenza A virus–infected mice

Antiviral neuraminidase inhibitors, such as oseltamivir, zanamivir, and peramivir, are widely used for treatment of influenza virus infection. We reported previously that oseltamivir inhibits the viral growth cycle, ameliorates symptoms, and reduces viral antigen quantities. Suppressed viral antigen production, however, induces a reduction of acquired antiviral humoral immunity, and increases the incidence of re-infection rate in the following year. To achieve effective treatment of influenza virus infection, it is necessary to overcome these adverse effects of antiviral neuraminidase inhibitors. Feeding of yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) OLL1073R-1 is reported to have immune-stimulatory effects on influenza virus infection in mice and humans. In the present study, we assessed the effect of feeding L. bulgaricus OLL1073R-1 yogurt cultures (YC) on local and systemic humoral immune responses, which were suppressed by oseltamivir treatment, in mice infected with influenza A virus. Yogurt culture (1.14 × 10⁸ cfu/0.4 mL per mouse per day) or sterile water (vehicle) was administered by intragastric gavage for 35 d. At d 22, influenza A virus/Puerto Rico/8/34 (H1N1) (PR8; 0.5 pfu/15 μL per mouse) was instilled intranasally, followed immediately by oral administration of oseltamivir (50 μg/100 μL per mouse, twice daily) or 5% methylcellulose (100 μL/mouse) as a vehicle for 13 d. Titers of anti-PR8-specific IgG and IgA in serum and mucosal secretory IgA (S-IgA) and IgG in bronchoalveolar lavage fluid (BALF) were analyzed by ELISA at 14 d after infection. Oseltamivir significantly suppressed the induction of anti-PR8-specific IgG and IgA in serum and S-IgA and IgG in BALF after infection. Feeding YC mildly but significantly stimulated production of PR8-specific IgA in serum, S-IgA in BALF, and IgG in serum without changing the IgG_2a:IgG₁ ratio. We analyzed the neutralizing activities against PR8 in serum and BALF and found that oseltamivir also reduced protective immunity, and YC feeding abrogated this effect. The immune-stimulatory tendency of YC on anti-PR8-specific IgA and IgG titers in serum and BALF was also detected in mice re-infected with PR8, but the effect was insignificant, unlike the effect of YC in the initial infection.