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@#Objective To prepare rabbit polyclonal antibodies against pertussis toxin(PT) and develop a double antibody sandwich ELISA for quantitative determination of PT antigen,identify and apply the method.Methods The rabbit polyclonal antibody against PT was prepared by immunizing Chinchilla rabbit with PT using traditional method.The reaction conditions of ELISA system were optimized,the double antibody sandwich ELISA method for quantitative determi-nation of PT was developed,and the specificity,linearity,accuracy,precision and sensitivity were verified.The developed method was used to detect PT antigen content in fimbriae proteins(FIM) stock solution of samples during detoxification and other purification process of pertussis antigen.Results The working condition of double antibody sandwich ELISA for detection of PT antigen content was the coating concentration of PT rabbit polyclonal antibody of 1 μg/mL,and the enzyme-labeled antibody dilution of 1:8 000.This detection system showed specific reaction with PT purified protein,but had no cross reaction with filamentous hemagglutinin,diphtheria toxoid and tetanus toxoid;the linear detection range of the developed double antibody sandwich ELISA was within 25—400 ng/mL;the recovery rates of PT at high,moderate and low concentrations were 103.27%,91.48% and 103.52%,respectively;both the intra-and inter-coefficients of variation(CVs)were less than 10%;the sensitivity of the method was 20.719 ng/mL,and the detection limit was 41.438 ng/mL.Thirty-five batches of samples were detected under five different detoxification process conditions and at different sampling time points,and the changes of antigen content were all consistent with the trend of detoxification reaction.Conclusion The PT rabbit polyclonal antibody was successfully prepared,and a double antibody sandwich ELISA with high precision and accuracy was developed for the quantitative determination of PT antigen content,which can be used for the antigen content detection of chemically detoxified samples in the production process of component DPT vaccines.
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@#Pertussis is an ancient disease,and early vaccination has greatly reduced the incidence and mortality of pertussis.However,the pertussis resurgence has been reported worldwide in recent years.Further understanding of the epidemiological characteristics of pertussis is essential for the development of more effective pertussis vaccines and for better prevention and control and early elimination of pertussis.This paper mainly reviews the basic epidemiological characteristics,seroepidemiological characteristics,molecular epidemiological characteristics and prevention and control progress of pertussis at home and abroad.
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@#Objective To prepare rabbit polyclonal antibodies against pertussis toxin(PT) and develop a double antibody sandwich ELISA for quantitative determination of PT antigen,identify and apply the method.Methods The rabbit polyclonal antibody against PT was prepared by immunizing Chinchilla rabbit with PT using traditional method.The reaction conditions of ELISA system were optimized,the double antibody sandwich ELISA method for quantitative determi-nation of PT was developed,and the specificity,linearity,accuracy,precision and sensitivity were verified.The developed method was used to detect PT antigen content in fimbriae proteins(FIM) stock solution of samples during detoxification and other purification process of pertussis antigen.Results The working condition of double antibody sandwich ELISA for detection of PT antigen content was the coating concentration of PT rabbit polyclonal antibody of 1 μg/mL,and the enzyme-labeled antibody dilution of 1:8 000.This detection system showed specific reaction with PT purified protein,but had no cross reaction with filamentous hemagglutinin,diphtheria toxoid and tetanus toxoid;the linear detection range of the developed double antibody sandwich ELISA was within 25—400 ng/mL;the recovery rates of PT at high,moderate and low concentrations were 103.27%,91.48% and 103.52%,respectively;both the intra-and inter-coefficients of variation(CVs)were less than 10%;the sensitivity of the method was 20.719 ng/mL,and the detection limit was 41.438 ng/mL.Thirty-five batches of samples were detected under five different detoxification process conditions and at different sampling time points,and the changes of antigen content were all consistent with the trend of detoxification reaction.Conclusion The PT rabbit polyclonal antibody was successfully prepared,and a double antibody sandwich ELISA with high precision and accuracy was developed for the quantitative determination of PT antigen content,which can be used for the antigen content detection of chemically detoxified samples in the production process of component DPT vaccines
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@#Pertussis is an ancient disease,and early vaccination has greatly reduced the incidence and mortality of pertussis.However,the pertussis resurgence has been reported worldwide in recent years.Further understanding of the epidemiological characteristics of pertussis is essential for the development of more effective pertussis vaccines and for better prevention and control and early elimination of pertussis.This paper mainly reviews the basic epidemiological characteristics,seroepidemiological characteristics,molecular epidemiological characteristics and prevention and control progress of pertussis at home and abroad.
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@#Objective To develop a kinetic chromogenic quantitative method for the determination of endotoxin content in intermediate of pertussis antigen,and to verify the method so as to better control the quality of diphtheria,tetanus,and pertussis vaccine(DTP vaccine).Methods A kinetic chromogenic assay[Limulus Amebocyte Lysate(LAL)]was developed to detect the endotoxin content in the intermediate products of pertussis antigens after detoxification,and verified for the linearity,specificity,accuracy,reproducibility and intermediate precision. The quantitative detection results of kinetic chromogenic assay were compared with those of gel method.Results The absolute value of the linear correlation coefficient(|r|)of the kinetic chromogenic assay was more than 0. 99;under the maximum effective multiple dilution,the interference test recovery of the intermediate was within 50% — 200%,and pertussis toxin(PT)diluted to 10,100 and 1 000 times,filamentous hemagglutinin(FHA)diluted to 3 000,5 000 and 10 000 times,and pertussis adhesin(PRN)diluted to 50,75 and 100 times had no interference effect on the experiment after detoxification;the accuracy verification recovery rates of PT,FHA and PRN were 125%,110% and 99% respectively;and the CVs of reproducibility verification were 7. 21%,8. 31% and 5. 84%,and the CVs of intermediate precision verification were 6. 04%,16. 29% and 12. 23%,respectively.The bacterial endotoxin content of the three batches of pertussis antigen intermediates detected by kinetic chromogenic assay was consistent with that verified by gel method,both of which were less than the limit of bacterial endotoxin in the intermediates of pertussis antigen after detoxification.Conclusion The developed kinetic chromogenic assay has good linearity,specificity,accuracy and precision with accurate detection results,which can be used to detect the endotoxin content in intermediate products of component pertussis antigen after detoxification.
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Objective To investigate the clinical features of hospitalized children with pertussis.Methods The clinical data of 87 hospitalized children with pertussis admitted to Women and Children's Hospital Affiliated to Xiamen University from January 2019 to December 2022 were retrospectively analyzed.According to the age of the children,they were divided into group A(≤3 months of age,35 cases),group B(4-12 months of age,36 cases)and group C(>12 months of age,16 cases).The clinical features of children at different ages were compared.Results The majority of children with pertussis were<1 year old(81.61%),and August was the month of high incidence.Spastic cough(72.41%)was the main clinical manifestation.Other manifestations included redness or cyanosis,vomiting after coughing,crow echo,etc.There were statistically significant differences in the incidence of spastic cough,redness or cyanosis and the contact history of patients with chronic cough among three groups(P<0.05).There were significant differences in the incidence of pneumonia and the average length of hospital stay among three groups(P<0.05).There was significant difference in C-reactive protein level among three groups(P<0.05).Among 87 children with pertussis,37 cases(42.53%)were infected with other pathogens,among which virus infection was the most common.The treatment is effective with macrolides and compound sulfamethoxazole.Conclusion Pertussis is more common in infants under 1 year of age,and the incidence is high in August.Chronic cough patients with close contact may be the source of infection.Spastic cough is the main clinical symptom of children;the younger,the more common symptoms such as spastic cough,redness and cyanosis,the higher incidence of pneumonia and the longer hospital stay.
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@#Objective To evaluate the influence of three kinds of software on the statistical results of titer detection of diphtheria tetanus acellular pertussis(DTaP)vaccine,so as to provide a basis for the selection of analysis software in daily detection. Methods The titers of pertussis,tetanus and diphtheria of DTaP vaccine were detected by the method specified in Chinese Pharmacopoeia(volume Ⅲ,2020 edition). PLA 3.0 software,Stati software and Excel calculation table compiled according to the General Rules 1431 Biological Verification Statistical Method(General Rules 1431)of Chinese Pharmacopoeia(volume Ⅲ,2020 edition)were used for the data statistics of the titer detection of DTaP vaccine,and the relative titer value was calculated. The 95% confidence limit interval of the above data was determined by using the mass reaction parallel line method,and the relative deviation was calculated. Results The relative titers of pertussis of DTaP vaccine calculated by three kinds of software were 99. 16%-100. 46%,and the relative deviations of 95% confidence interval were all less than 3%. The relative titers of tetanus ranged from 99. 67% to 100. 46%,with all the relative deviations of 95% confidence interval of less than 1%. The relative titers of diphtheria ranged from 97. 33% to 100. 03%,the relative deviations of the upper bound of 95% confidence interval were all less than 0,but those of the lower bound were large. Conclusion The three kinds of software can replace each other,among which Stati software and General Rules 1431 software are more suitable for some work scenarios without audit trail requirements,while PLA 3.0 software is more suitable for the quality control system in biological products enterprises.
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Objective To understand the epidemiological characteristics and current situation of pertussis in Hubei Province from 2016 to 2021. Methods Data on the incidence of pertussis and immunization history of cases in Hubei Province from 2016 to 2021 were collected and descriptive epidemiological analysis was conducted. Results From 2016 to 2021, a total of 1 236 pertussis cases were reported in Hubei Province, with an average annual reported incidence rate of 0.35/100 000. The average annual reported incidence of pertussis in the age group ≤ 5 years old was 6.22/100 000, and the age group <1 year old reported the highest annual incidence rate (21.51/100 000). The proportion of pertussis among preschool children and students had increased significantly since 2020. Among the 1 111 cases with a known immunization history, 17.55% were under the age of vaccination , 41.85% were not vaccinated, and 17.46% had completed the whole course of vaccination. Conclusion Since 2016, the incidence of pertussis in Hubei Province has been on the rise. The risk of pertussis is higher in infants and young children who have not reached the age of vaccination and who have not been vaccinated in time according to the immunization program after reaching the age of vaccination. The timeliness and vaccination rate of DTP vaccine should be improved to reduce the risk of pertussis in infants and young children. In addition, more attention should be paid to the prevalence of pertussis among preschool children and students.
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For a long time, macrolides have been the first choice for the antibacterial treatment for pertussis.However, in the past decade, resistance to macrolide antimicrobials has been common in clinically isolated Bordetella pertussis in China, which is in contradiction with the recommended macrolide treatment.Therefore, Trimethoprim-Sulfamethoxazole (TMP-SMZ) is suggested as the first choice for antibacterial treatment for pertussis in China, with a dosage determined according to age and body weight, lasting 14 days.If TMP-SMZ cannot be used, full-dose and full-course β-lactam antimicrobials may be used, of which the effects should be assessed carefully.The impact of other antibacterial drugs, such as quinolones and tetracyclines, on the elimination of Bordetella pertussis should also be evaluated as soon as possible to treat adult pertussis and potential cases caused by drug-resistant bacteria in future.
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Objective:To determine the erythromycin resistance of Bordetella pertussis isolates and their ptxP1 and ptxP3 phenotypic composition and compare clinical manifestations of children with pertussis caused by the two types of strains. Methods:This was a cross-sectional study, the pertussis cases diagnosed using bacterial culture from January 2019 to December 2022 in Beijing Children′s Hospital and the First People′s Hospital of Wuhu were collected.Any suspected Bordetella pertussis colonies were identified by the slide agglutination test.The susceptibility of isolates to erythromycin was detected by the E-test and K-B test.The ptxP gene was amplified by polymerase chain reaction and sequenced to determine its genotype. t-test, Mann-Whitney U-test, Chi-square test and Fisher′s exact test were use to statistical analysis. Results:A total of 192 strains of Bordetella pertussis were identified, including 188 (97.9%) erythromycin-resistant strains.Among the 188 strains, 30.3%(57/188) belonged to the ptxP1 genotype and 69.7%(131/188) belonged to the ptxP3 genotype.In children aged below 1 year old, the incidence of paroxysmal cough caused by infection with the ptxP3 strain was higher than that with the ptxP1 strain (57.1% vs.29.4%, P<0.05), and children infected with the ptxP3 strain were more likely to develop apnea or asphyxia (23.8% vs.17.6%), post-tussive vomiting (44.4% vs.32.4%), whooping cough (72.0% vs.50.0%) and pneumonia or bronchitis (85.7% vs.73.5%) compared to those infected with the ptxP1 strain, but the differences were not statistically significant(all P>0.05). In children aged 1 year old and above, the white blood cell count of children infected with the ptxP1 strain was higher than that of infections with the ptxP3 strain [13.5(9.9, 24.5)×10 9/L, 10.3 (7.0, 16.4)×10 9/L, P<0.05], and children infected with the ptxP1 strain were more likely to contract other pathogen infections than those infected with the ptxP3 strain (17.4% vs.4.4%, P>0.05). Conclusions:ptxP3 erythromycin-resistant Bordetella pertussis has become the main pathogen of pertussis.Infants with pertussis caused by the ptxP3 erythromycin-resistant strain show more significant manifestations and a higher possibility of severe symptoms than those infected with the ptxP1 erythromycin-resistant strain.
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Objective:To test the hypothesis that Pertussis toxin (PTX) can promote the occurrence of interstitial lung disease (ILD) in experimental autoimmune myositis (EAM) model and clarify the potential pathogenic mechanism.Methods:EAM mice model were induced by Skeletal muscle thomogenate with or without PTX, and the relationship between ILD phenotypes and neutrophil extracellular traps (NETs) infiltration was analyzed by histopathological and serological studies in EAM with PTX group and EAM without PTX group. Healthy mice were given PTX alone intraperitoneally to clarify whether NETs formation could be induced in vivo, and neutrophils separated from healthy human blood were intervened with PTX to induce NETs formation in vitro. The data was tested for normality using Shapiro-Wilk. Statistical methods and were analyzed using t-test or ANOVA, and multiple comparisons between different groups were tested using Tukey test. Results:Compared with EAM without PTX group, lung tissues in EAM with PTX group had multiple pathological changes similar to polymyositis/dermatomyositis-related ILD. Nonspecific interstitial pneumonia and usual interstitial pneumonia were the main pathological types. The pulmonary interstitial lesions were accompanied by significant infiltration of NETs; and serum NETs markers levels were obviously elevated in EAM with PTX group, compared with the control group [ n=5, (87±10) ng/ml], cfDNA levels were statistically significantly elevated in both the EAM without PTX group [ n=4, (115±27) ng/ml] and the EAM with PTX group [ n=7, (150±50) ng/ml] ( F=4.24, P=0.038); Cit-H3-DNA levels were elevated in the EAM without PTX group ( n=4, 0.24±0.09), and in the EAM EAM with PTX group ( n=6, 0.33±0.11) compared with the control group ( n=4, 0.13±0.02) ( F=6.21, P=0.016). After PTX intervention, serum cfDNA levels were higher in the PTX group [ n=3, (100±40) ng/ml] than in the control group [ n=3, (45±12) ng/ml, t=2.27, P=0.086]; PTX also induced neutrophils to form NETs in vitro. Conclusion:PTX may promote the development of ILD in EAM mice model by inducing the formation of NETs, indicating that EAM mice can serve as a model for targeting NETs to study the pathogenesis ILD.
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Introducción: La tosferina causada por la bacteria Bordetella pertussis, es una de las principales enfermedades prevenibles por vacunas en el mundo. En Panamá es una causa importante de morbimortalidad en la población infantil. A nivel nacional se registraron 20 casos en el 2018 y 108 en el 2019. El objetivo es describir el comportamiento clínico y epidemiológico de la infección en la población pediátrica. Materiales y métodos: Es un estudio descriptivo, retrospectivo y observacional. Se incluyeron pacientes de 0 a 14 años con diagnóstico de infección por B. pertussis. Las variables: edad, sexo, procedencia, comorbilidades, signos, síntomas, vacuna contra B. pertussis, hemograma, complicaciones, defunción. Resultados: La tasa hospitalaria fue de 5 casos por cada 10 000 hospitalizados. El sexo presentó una razón de 2:1. El grupo etario más afectado fue el de 2 a 3 meses. La tos fue la manifestación más frecuente. La comorbilidad que se observó con mayor frecuencia fue la desnutrición. El 56% de los pacientes no contaban con vacuna contra B. pertussis. La leucocitosis se observó en 66% de los casos, la linfocitosis en 16% y la trombocitosis en 34%. Tasa de mortalidad hospitalaria, por enfermedad específica de 1,4 casos por 10 000 hospitalizados. Tasa de letalidad, de 25 por cada 100 casos. Conclusiones: Se encontró predisposición en el sexo femenino. Los lactantes menores son el grupo más afectado. La mayoría de los pacientes y sus madres, no habían recibido vacunación contra B. pertussis. La letalidad observada fue mayor a la descrita en la literatura. (provisto por Infomedic International)
Introduction: Whooping cough caused by the Bordetella pertussis bacterium is one of the main vaccine-preventable diseases in the world. In Panama it is an important cause of morbidity and mortality in the child population. At the national level, 20 cases were registered in 2018 and 108 in 2019. The objective is to describe the clinical and epidemiological behavior of the infection in the pediatric population. Materials and methods: It's a descriptive, retrospective and observational study. Patients from 0 to 14 years old with a diagnosis of B. pertussis infection were included. The variables: age, sex, origin, comorbidities, signs, symptoms, vaccine against B. pertussis, blood count, complications, death. Results: The hospital rate was 5 cases per 10,000 hospitalized. Sex presented a ratio of 2:1. The most affected age group was 2 to 3 months. Cough was the most frequent manifestation. The most frequently observed comorbidity was malnutrition. 56% of the patients didn´t have a vaccine against B. pertussis. Leukocytosis was observed in 66% of cases, lymphocytosis in 16%, and thrombocytosis in 34%. Hospital mortality rate, by specific disease of 1.4 cases per 10,000 hospitalized. Fatality rate, 25 per 100 cases. Conclusions: A predisposition was found in the female sex. Younger infants are the most affected group. Most of the patients and their mothers had not received vaccination against B. pertussis. The observed lethality was higher than that described in the literature. (provided by Infomedic International)
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Objective@#To evaluate the immune persistence following intradermal(ID) vaccination with diphtheria-tetanusacellular component pertussis and Sabin-derived inactivated poliovirus vaccine(DTacP-sIPV).@*Methods@#40 wistar rats were randomly assigned into four groups.Two test groups were injected intradermally with fractional-doses(1/5 and 1/10dose) of DTacP-sIPV(1/5D ID and 1/10D ID group);The positive control group was intramuscularly injected with full dose of DTacP-sIPV(full-dose IM group);The negative control group was injected with PBS intradermally.Wistar rats were immunized 3 times at 0,1 and 2 months and the blood samples were collected via tail vein 12 months after the last immunization and the serum samples were isolated.The titer of neutralizing antibody against poliovirus was detected by micro-neutralization test,and the titers of IgG antibodies against diphtheria toxin(DT),tetanus toxin(TT),pertussis toxin(PT),filamentous hemagglutinin(FHA) and pertactin(PRN) in rat serum were detected by indirect ELISA.The geometric mean titer(GMT)and positive rate of antibody were calculated.The rats were challenged with aerosolized B.pertussis for 30 min 12 months after the last immunization and determined for the white blood cell(WBC) count and colony-forming unit(CFU) in lung,trachea and nose at day 2,5 and 14 after challenge.@*Results@#Compared with the full-dose IM group,there was no significant difference in the positive rates of poliovirus type Ⅰ,Ⅱ and Ⅲ neutralizing antibodies between 1/5D ID and 1/10D ID groups(each P> 0.05) and the positive rates of all types of antibodies in the control group were 0.The positive rates of IgG antibodies against DT,TT,PT,FHA and PRN in 1/5D ID,1/10D ID and full-dose IM groups were all 100%,and those in control group were all 0.Compared with 2 d after challenge,the WBC counts of rats in control group increased significantly 5 d after aerosol challenge with B.pertussis(F=3.48,P <0.05),and then began to decrease,while those in other groups remained stable with time(F=0.14~1.30,P> 0.05).After aerosol challenge,the CFU in lungs of rats in control group was significantly higher than that in the other three groups(F=19.00~206.00,P<0.05),and B.pertussis was still detected 14 d after challenge;Except for the control group,the bacterial load in lungs of rats in the other three groups reached the peak 5d after challenge,the B.pertussis was basically cleared on the 14d,and there was no significant difference among the groups at each time point(F=1.14~1.25,P> 0.05).The bacterial load of trachea and nose in the control group was slightly higher than that in other groups at each time point,but the difference was not significant(F=0.71~3.54,P> 0.05).Except for the control group,the bacterial load in the trachea and nose of the other three groups were similar,and no significant difference was observed(F=0.75~3.41,P>0.05).@*Conclusion@#ID immunization with1/5 dose of DTacP-sIPV induced persistent protective antibodies against various components of the vaccine in rats.This study provided an experimental basis for the formulation of immunization strategy of ID immunization with fractional dose of DTacP-sIPV.
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@#Combined vaccine is a single vaccine preparation made by mixing two or more different organism or purified antigens by physical methods.The use of combined vaccine can reduce the number of immunization shots and improve the compliance of children and acceptance of parents;reduce the costs of transportation,storage and management;avoid missing vaccination and improve vaccination rate.At present,a variety of combined vaccines have been licensed abroad,which have good safety and immunogenicity;some combined vaccines have been put on the market in China,and many combined vaccines are in clinical trials.In recent years,with the successful development of component pertussis vaccine and inactivated poliovirus vaccines of Sabin strain Ⅰ,Ⅱ and Ⅲ in China,the combined vaccines based on diphtheria-tetanus-acellular pertussis vaccine(DTaP) have been greatly developed.In this paper,the research progress on combined vaccines based on DTaP,which have been licensed and in clinical trials at home and abroad,was reviewed,in order to provide ideas for the development of related combined vaccines in China.
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ObjectiveTo characterize the incidence of laboratory confirmed pertussis cases by immunization status in Ninghai County, and to provide evidence for improving pertussis prevention and control strategy in Ninghai County. MethodsData of reported pertussis cases in Ninghai County from 2018 to 2019 were collected through the National Notifiable Infectious Disease Reporting System. The immunization history of vaccines related to pertussis was collected through the Ningbo Immunization Planning Information Management System and field investigation. Descriptive epidemiological methods were used to analyze the collected data. ResultsThe respiratory secretions collected in the 64 cases tested positive for nucleic acid of Bordetella pertussis bacilli, of which 56.25% aged 0‒1 year. They were mainly pre-school children. No adolescent or adult case was reported. All cases had whooping cough. Of the cases, 78.13% had an increase in white blood cell count, 62.5% had an increase in lymphocyte percentage, 59.38% had completed three or more doses of pertussis vaccination, 69.39% had a time interval of >1 year between the onset and last dose of pertussis vaccination, and 31.25% reported close contact with pertussis patients before the onset. The misdiagnosis rate of pertussis was as high as 92.19%. In contrast, the prevalence of anti-pertussis IgG antibody in health population was 1.05%. ConclusionFurther improvement in the laboratory testing capacity can help clarify the diagnosis of pertussis. Generally, children aged 0-1 year are mainly affected, of which vast majority have received at least one dose of pertussis vaccination. Clinical symptoms are characterized by whooping cough. Additionally, elevated white blood cells and lymphocytes should be on the alert to pertussis infection.
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Objective@#To analyze the epidemiological characteristics of pertussis in Huzhou City, Zhejiang Province from 2012 to 2022, so as to provide insights into formulation of pertussis control measures.@*Methods@#The data of reported pertussis cases in Huzhou City from 2012 to 2022 were collected through the Infectious Disease Report Information System of Chinese Disease Prevention and Control Information System. The temporal, spatial and population distributions of pertussis cases in Huzhou City from 2012 to 2022 were analyzed using a descriptive epidemiological method.@*Results@#A total of 499 pertussis patients were reported in Huzhou City from 2012 to 2022, with mean annual reported incidence of 1.508/105, and no death was reported. The reported incidence of pertussis remained at a low level in Huzhou City from 2012 to 2021, all below 1/105, and increased to 12.625/105 in 2022. The reported incidence of pertussis appeared an overall tendency towards a rise in Huzhou City from 2012 to 2021 (Z=-29.261, P<0.001). The incidence of pertussis peaked from June to July, and a relatively higher incidence rate of pertussis was reported in Deqing (6.359/105) and Anji counties (1.725/105), while higher incidence was found among children at ages of <1 year (30.566/105), 4 years (31.896/105) and 5 years (29.485/105). @*Conclusion@#The reported incidence of pertussis was at a low level in Huzhou City from 2012 to 2021, and increased sharply in 2022. The incidence of pertussis peaked from June to July, was concentrated in Deqing and Anji counties and higher among infants under one year of age and preschool children at ages of 4 to 5 years.
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@#Objective To verify the applicability of Lowry2 method[the second method of Folin-phenol method of Chinese Pharmacopoeia(VolumeⅢ,2020 edition)]for the determination of protein content of intermediate products of component pertussis vaccine.Methods The standard and sample were precipitated and pretreated with deoxycholate and trichloroacetic acid to remove impurities,and then reacted with Folin-phenol,and the absorbance was measured at 750 nm wavelength.The standard curve was drawn with the concentration and absorbance of the standard,and the sample concentration was calculated. The method was verified for the specificity,linearity,accuracy and precision,and used to detect the protein content of intermediate products of three batches of component pertussis vaccine.Results There was no significant difference between the protein content in background buffer of intermediate product detected by this method and that in ultrapure water(t = 0. 277~1. 178,P = 0. 304~0. 795);The linear relationship of standard curve was good at the protein concentration of 0~50 μg/mL,each R~2> 0. 98;The spike recovery of accuracy verification was in the range of 90%~110%;The RSDs of repeatability and intermediate precision verification were both less than 3%. The detection results of protein content in the intermediate products of three batches of component pertussis vaccine all met the requirements of internal manufacturing and verification regulations.Conclusion Lowry2 method has good specificity,accuracy and precision,and can be used to determine the protein content of component pertussis intermediate products,which provides a basis for the applicability of Lowry2 method.
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ObjectiveTo investigate the expression of lysophosphatidic acid (LPA) in patients with liver cancer, as well as its influence on malignant biological behavior of liver cancer and related regulatory mechanism. MethodsFrom January 2016 to December 2022, 26 patients with liver cancer, 28 patients with liver cirrhosis, and 28 individuals undergoing physical examination were enrolled. ELISIA was used to measure the content of LPA in plasma and peritoneal effusion of the patients with liver cancer or liver cirrhosis accompanied by peritoneal effusion, and the content of LPA was measured in plasma of the normal population at the same time, so as to clarify the difference in the expression of LPA in different populations, such as the patients with liver cancer and those with liver cirrhosis. MTT cell proliferation assay and cell migration assay were used to observe the influence of LPA and its inhibitor pertussis toxin (PTX) on the proliferation, migration, and invasion of SMMC7721 cells. In order to investigate the effect of LPA on the expression of RhoA and its upstream and downstream molecules FAK and P53 after binding to its receptor, qPCR and Western blot were used to observe the effect of LPA on the mRNA and protein expression levels of P53, FAK, and RhoA in SMMC7721 cells. A one-way analysis of variance was used for comparison of the means of continuous data between multiple groups, and the SNK-q test was used for comparison between two groups. ResultsCompared with the patients with liver cirrhosis, the patients with liver cancer had a significantly higher concentration of LPA in plasma (4.99±0.55 μmol/L vs 2.63±0.43 μmol/L, P<0.05) and peritoneal effusion (5.19±0.63 μmol/L vs 2.91±0.46 μmol/L, P<0.05), and the patients with liver cancer also had a significantly higher level of plasma LPA than the normal population (4.99±0.55 μmol/L vs 1.61±0.39 μmol/L, P<0.05). The cell proliferation assay showed that LPA significantly promoted the proliferation of SMMC7721 cells, and cell proliferation rate increased with the increase in dose and time; in particular, the middle-and high-dose groups had a significantly higher proliferation rate than the control group (P<0.05). PTX inhibited the proliferative capacity of SMMC7721 cells in a time-dependent manner, and there was a significant difference between the groups (P<0.05). The proliferation rate of the 72-hour high-dose LPA group was 3.6 times that of the control group, while the proliferation rate of the PTX group was 0.6 times that of the control group; the proliferation rate of the 72-hour high-dose LPA+PTX group was 1.2 times that of the control group. In addition, LPA increased the migration ability of hepatoma cells, while PTX inhibited their migration, in a time-dependent manner, and there was a significant difference between the groups (P<0.05). The migration rate of the 72-hour high-dose LPA group was 3.09 times that of the control group, while the migration rate of the PTX group was 0.4 times that of the control group; the migration rate of the 72-hour high-dose LPA+PTX group was 0.99 times that of the control group. qPCR and Western blot showed that there were significant reductions in the mRNA and protein expression levels of P53 in SMMC7721 cells after LPA treatment, while there were significant increases in the mRNA and protein expression levels of FAK and RhoA; there was a significant difference between the LPA group and the control group (P<0.05). ConclusionThere is an abnormal increase in the expression of LPA in patients with liver cancer, and LPA can promote the proliferation of liver cancer cells and increase their migration ability. At the same time, LPA changes the expression levels of P53, FAK, and RhoA, which may be associated with the promotion of tumor development and progression by LPA.
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Objective:To analyse the clinical features and prognosis of pertussis in neonates and infants.Methods:The clinical data of neonates and infants with pertussis hospitalized in Children′s Hospital of Soochow University from September 2021 to September 2022 were retrospectively analyzed and grouped in terms of age, the severity of the disease, and whether a mixed infection, respectively.Results:A total of 40 infants with pertussis were analyzed.All cases showed improvement and were discharged after receiving active anti-infective treatment.In the neonatal group, higher rates of apnea and hyponatremia were observed compared to the non-neonatal group(all P<0.05).Additionally, peripheral blood leukocyte counts[20.9(15.0, 28.7)×10 9/L vs.16.6(11.3, 21.2)×10 9/L], neutrophil counts[4.6(3.7, 7.9)×10 9/L vs.3.2(2.1, 5.3)×10 9/L], γ-glutamyltransferase levels[78.0( 50.2, 109.4)U/L vs.22.5(15.1, 38.9)U/L], duration of hospitalization[21.5(16.8, 25.0)d vs.11.5(9.0, 19.8)d], and duration of oxygen use[7.0(0, 21.0)d vs.0(0, 2.3)d]were higher in the neonatal group than in the non-neonatal group(all P<0.05).However, the IgA level[0.02(0.02, 0.04)g/L vs.0.05(0.03, 0.09)g/L]was significantly lower in the neonatal group than in the non-neonatal group( P<0.05).In the severe group, the proportion of onset age of less than 3 months, fever, wheezing, shortness of breath, cyanosis after rough cough, apnea, decreased heart rate, wet rales on lung auscultation, respiratory failure, cardiac insufficiency, hyponatremia, CRP>8 mg/L, spotty/patchy shadows in the lungs, as well as the use of gammaglobulin, cardioactive drug and invasive ventilation, were higher than those in the non-severe group(all P<0.05).Furthermore, peripheral blood leukocyte counts[21.0(15.4, 37.4)×10 9/L vs.17.5(11.8, 21.2)×10 9/L], neutrophil counts[5.6(4.0, 10.7)×10 9/L vs.3.2(2.3, 4.6)×10 9/L], neutrophil to lymphocyte ratio[(0.6±0.4) vs.( 0.3±0.2)], systemic immune-inflammation index[237.5(109.5, 424.9) vs.135.9(75.4, 190.5)], γ-glutamyltransferase level[53.2(31.6, 87.4)U/L vs.29.5(15.2, 65.0)U/L], duration of oxygen use[18.0(12.8, 22.5)d vs.0(0, 0)d], and duration of hospitalization[24.5(21.8, 31.2)d vs.12.0(9.0, 16.8)d]were higher in the severe group than those in the non-severe group(all P<0.05).However, the IgA level[0.03(0.02, 0.04)g/L vs.0.05(0.03, 0.09)g/L]was significantly lower in the severe group than in the non-severe group( P<0.05).The mixed infection group had a longer duration of hospitalization and a higher proportion of fever than the single infection group(all P<0.05). Conclusion:Early detection of infantile pertussis can be challenging.Neonates with pertussis tend to experience severe symptoms, such as apnea, hyponatremia, elevated white blood cell count, and longer duration of oxygen use.Symptoms such as fever, wheezing, shortness of breath, decreased heart rate, wet lung rales, and spotty/patchy shadows in the lungs, as well as early elevated CRP, neutrophil to lymphocyte ratio, systemic immune-inflammation index, and decreased IgA levels are indicators of disease exacerbation.In mixed infections group, there is a higher proportion of fever.
RESUMO
Objective:To investigate the clinical features of pertussis in children and analyze the risk factors of severe pertussis.Methods:The clinical data of 248 children with pertussis hospitalized in Hunan Children′s Hospital from March 2018 to March 2022 were analyzed retrospectively.According to the age at admission, the patients were divided into two groups: ≤3 months and > 3 months.According to the patient′s condition, they were classified into ordinary group and severe group.According to the pathogens detected, the children were divided into single infection group and mixed infection group.The independent sample t-test, chi- square test were used to analyze the clinical indexes of the infants in above groups. Results:(1)Of 248 hospitalized children with pertussis, 204 cases (82.2%) were less than 1 year old, 92 cases (37.0%) had contact with a coughing family member before, and 169 cases (68.1%) were unvaccinated.Among 248 children, 193 cases (77.8%) had an elevated white blood cell count, and 145 cases (58.4%) had mixed infections.The most common pathogen was respiratory syncytial virus [29/248(11.6%)]. About 173 cases (69.7%) had concurrent pneumonia, and 35 cases (14.1%) had pulmonary consolidation.(2)Compared with the group > 3 months of age, more patients in the group ≤3 months of age had contact with a coughing family member before, and suffered from cyanosis, dyspnea, respiratory failure, heart failure and pertussis encephalopathy ( χ2=4.612, 20.810, 7.882, 16.617, 13.740, 7.846, all P<0.05). The proportions of patients in the group ≤3 months of age required intensive care unit(ICU) hospitalization and mechanical ventilation were higher than those in the group > 3 months of age ( χ2=14.810, 21.436, all P<0.05). The mortality of the group ≤3 months of age was higher than that of the group >3 months of age ( χ2=12.016, P<0.05). Children ≤3 months of age had a higher WBC level [(27.83±27.70)×10 9/L vs.(23.34±15.28)×10 9/L, t=22.244, P<0.001], longer duration of spasmodic cough [(16.56±9.33) d vs.(15.06±6.16) d, t=10.145, P=0.002] and longer hospitalization time [(11.47±10.48) d vs.(9.48±4.80) d, t=20.050, P<0.001] than those >3 months of age.(3)Compared with the ordinary group, a higher proportion of children in the severe pertussis group were under 3 months old, and had not been vaccinated against pertussis vaccine ( χ2=14.803, 4.475, all P<0.05). The ratio of patients with dyspnea, an lymphocyte count/neutral cell(LC/NC) ratio <1, mixed infections, lung consolidation and pleural effusion in the severe pertussis group was higher than that in the ordinary group ( χ2=116.940, 43.625, 13.253, 106.370, 11.874, all P<0.05). The patients in the severe pertussis group had a higher WBC [(61.66±29.63)×10 9/L vs.(18.83±10.00)×10 9/L, t=112.580, P<0.001] and a lower LC (0.494±0.186 vs.0.676±0.132, t=13.752, P<0.001) than those in the ordinary group.(4)Compared with the single infection group, the proportions of children with fever, dyspnea, fine moist lung rales, an LC/NC ratio <1, and lung consolidation were higher in the mixed infection group ( χ2=8.909, 6.804, 7.563, 8.420, 12.458, all P<0.05). More children in the mixed infection group required ICU hospitalization and mechanical ventilation than those in the single infection group ( χ2=11.677, 7.397, all P<0.05). The mixed infection group had higher respiratory failure and death rates than the single infection group ( χ2=7.980, 4.267, all P<0.05). Compared with the single infection group, the mixed infection group had a higher WBC level [(27.73±24.13)×10 9/L vs.(21.25±14.65)×10 9/L, t=13.318, P<0.001], longer hospitalization time [(11.593±9.010) d vs.(8.339±4.047) d, t=17.283, P<0.001], and a smaller LC ratio (0.626±0.165 vs.0.684±0.132, t=7.997, P=0.005). (5) Logistic regression analysis showed that age ≤3 months, peak WBC and dyspnea were risk factors of severe pertussis. Conclusions:Hospitalized pertussis children are prone to pneumonia and pulmonary consolidation.Patients aged ≤3 months with a large WBC and dyspnea easily develop into severe pertussis.Monitoring blood routine is helpful for judging the severity of the disease.Mixed infections increase the incidence of complications and can impair the treatment effect.