Botulinum toxin type A in treating early-stage patients with small-angle acute acquired comitant esotropia

AIM:To investigate botulinum toxin A(BTXA)efficacy on small-angle(≤25Δ)acute acquired concomitant esotropia(AACE)in early-stage patients.METHODS:The electronic medical record data of AACE patients during March 2019 and June 2023 were collected in this retrospective and hospital-based cohort study.A total of 72 small-angle AACE patients received BTXA extraocular muscle injection.Patients were grouped by onset-to-treatment time(Group A:≤6mo,Group B:>6mo).Deviation of esotropia,eye alignment and stereopsis were analyzed at the period of pre/post-injection(1wk,1,3,and 6mo).Orthophoria rate at 6mo(horizontal deviation<10Δand binocular single vision)were considered as outcome index.RESULTS:There were no significant baseline differences(P>0.05)between two groups except onset-to-treatment time(2mo vs 11mo,P<0.001).Higher orthophoria rates were in Group A at last follow-up(94.74%vs 73.53%,P=0.013).Post-BTXA deviations of two groups at 1mo showed no difference(P>0.05);while in 3 and 6mo Group A was significantly smaller than group B(all P<0.001).No statistically significant differences were observed among all post-BTXA deviations of near and distance in Group A.In Group B,deviation at 3mo(near:2Δvs 0,P<0.001;distance:4Δvs 0,P<0.001)and 6mo(near:6Δvs 0,P<0.001;distance:6Δvs 0,P<0.001)was significant increased compared to deviation at 1wk after treatment.Group A showed better stereopsis recovery in last follow-up compared to Group B(80″vs 200″,P=0.002).Both groups obtained improved stereopsis after treatment(Group A:80″vs 300″,P<0.001;Group B:200″vs 300″,P=0.037).CONCLUSION:BTXA is effective for AACE with small deviation(≤25Δ)in early stage.Delayed treatment(>6mo)may reduce BTXA efficacy.Early BTXA intervention benefits long-term eye alignment and stereopsis recovery.

Betulinic acid protects against ovarian impairment by decreasing F-2 toxin-induced oxidative stress and inflammation associated with the downregulation of p38 expression in mice

F-2 toxin is an estrogenic mycotoxin that causes reproductive disorders in animals.Betulinic acid(BA)is a natural pentacyclic lupane-structure triterpenoid that has diverse pharmacological activities.In this study,the antioxidative and anti-inflammatory effects of BA and its underlying mechanism are explored in F-2 toxin-triggered mouse ovarian damage.We found that BA alleviated the F-2 toxin-induced ovarian impairment by stimulating follicle growth,reducing inflammatory cell infiltration,repairing damaged mitochondria and endoplasmic reticulum.Simultaneously,BA not only reversed F-2 toxin-induced reduction of follicle stimulating hormone(FSH)and luteinizing hormone(LH)levels in the serum,but also restrained the protein expression of the estrogen receptors a(ERa)and ERβ.Moreover,BA restored the balance of F-2 toxin-induced ovarian redox system disorders.Subsequently,we found that 0.25 mg/kg BA played an anti-inflammatory role in the F-2 toxin-induced ovarian impairment by decreasing interleukin-1β(IL-1β).IL-6,and tumor necrosis factor-α(TNF-α)mRNA expression,as well as inhibiting p38 protein expression.These data demonstrated that BA exerts its protective effect on F-2 toxin-induced ovarian oxidative impairment and inflammation by inhibiting p38 expression,which implies a natural product-based medicine to ameliorate F-2 toxin-caused female reproductive toxicity and provides a detoxifying method for food contaminated by mycotoxin.

Establishment of human cerebral organoid systems to model early neural development and assess the central neurotoxicity of environmental toxins

Human brain development is a complex process,and animal models often have significant limitations.To address this,researchers have developed pluripotent stem cell-derived three-dimensional structures,known as brain-like organoids,to more accurately model early human brain development and disease.To enable more consistent and intuitive reproduction of early brain development,in this study,we incorporated forebrain organoid culture technology into the traditional unguided method of brain organoid culture.This involved embedding organoids in matrigel for only 7 days during the rapid expansion phase of the neural epithelium and then removing them from the matrigel for further cultivation,resulting in a new type of human brain organoid system.This cerebral organoid system replicated the temporospatial characteristics of early human brain development,including neuroepithelium derivation,neural progenitor cell production and maintenance,neuron differentiation and migration,and cortical layer patterning and formation,providing more consistent and reproducible organoids for developmental modeling and toxicology testing.As a proof of concept,we applied the heavy metal cadmium to this newly improved organoid system to test whether it could be used to evaluate the neurotoxicity of environmental toxins.Brain organoids exposed to cadmium for 7 or 14 days manifested severe damage and abnormalities in their neurodevelopmental patterns,including bursts of cortical cell death and premature differentiation.Cadmium exposure caused progressive depletion of neural progenitor cells and loss of organoid integrity,accompanied by compensatory cell proliferation at ectopic locations.The convenience,flexibility,and controllability of this newly developed organoid platform make it a powerful and affordable alternative to animal models for use in neurodevelopmental,neurological,and neurotoxicological studies.

Severe Botulism after Intragastric Botulinum Toxin-A Injection: A Case Series

Intragastric botulismus toxin-A (BoNT-A) is one of the new approaches in the treatment of obesity. We aimed to contribute to the literature by presenting the clinical features, laboratory findings and treatment responses of iatrogenic botulism cases due to intragastric BoNT-A administered in our clinic. All detailed medical information was obtained by accessing the medical records of the patients who were hospitalized and followed up and treated between September 2022 and December 2022, and the diagnosis of A.05.1 Botulism was entered according to ICD-10, and whose clinical findings were compatible with botulism disease and who underwent intragastric BoNT-A application beforehand. These records were obtained by examining this information. 10 patients who developed botulism after intragastric BoNT-A application between 01/09/2022 and 28/02/2023 were followed up in our clinic. All of the patients were women. The mean age was 35. The mean hospital stay was 9 days. Only 1 of our cases required intensive care. Good response to treatment was accepted as a complete or near-complete improvement in the clinical findings of the patients and all of them had a good response to treatment. Intragastric BoNT-A administration is a procedure that requires careful indication with a profit/loss calculation considering the potential side effects. In addition, attention should be paid to dilution rates and dose amounts.

Effect of Purified Paper Wasp Ropalidia marginata Venom Toxins on Different Biomolecules in Mice Serum

This study evaluated the effects of purified paper wasp Ropalidia marginata venoms on various biomolecules in the blood serum of albino mice. Changes in the concentration of some important macromolecules, i.e., proteins, free amino acids, uric acid, cholesterol, pyruvic acid, total lipids and glucose were noted down. These alterations were measured after intraperitoneal injection of 40% and 80% 24-hour LD50 purified Ropalidia marginata venom toxin. Serum total protein levels were found to decrease to 78% after 6 hrs, while serum free amino acid levels were significantly increased to 117% 6 hrs after venom injection compared to control. It was also found that serum uric acid levels increased to 138% after 8 hrs of venom injection compared to control. The increase in serum cholesterol i.e. (101% and 106%) and pyruvic acid increased significantly to a maximum value of 106% after 6 hrs of treatment at 40% LD50. Glycogen levels in the gastrocnemius muscle were found to decrease significantly (p-0.05) to 43% and 92% at LD50 after injection of purified Ropalidia marginata venom after 8 h and 80% at LD50 compared to control. Moreover, up to 71% and 81% were obtained at 10 hrs of treatment with the same dose. In the present study, the purified toxins significantly changed the levels of biomolecules in blood serum, indicating their wider effects on cellular physiology due to toxic effects and stress on the animal. These toxins can be good antigens and stimulate immune responses in experimental mice.

Identification and Expression of the Conotoxin Homologous Genes in the Giant Triton Snail (Charonia tritonis)

Conotoxin(CTX)is a small peptide toxin,and plays crucial role in anesthetizes prey,preys and defends competitors.Charonia tritonis is the natural predator of the crown of thorns starfish(CoTS)and the protector of coral reefs.It plays an important role in the coral reef ecosystem.However,the types of toxins produced by tritons and the molecular mechanisms of toxin secretion of C.tritonis are unknown.In the present study,the four conotoxin homologous genes(CTXs)from C.tritonis were identified.Species and conotoxin superfamily phylogenetic tree indicated that CTX-1(CL2216.Contig2)and CTX-4(Unigene 58438_All)belong to the C superfamily,CTX-2(Unigene 66414_All)belong to the V superfamily,and CTX-3(Unigene 21408_All)belong to the B1 superfamily.CTXs were highly expressed in salivary gland,liver and digestive gland.The predation control experiment revealed that the expressions of CTXs were significantly different in salivary gland,liver and digestive gland.The results indicated that CTXs may participate in the process of C.tritonis predating CoTs,and provided a scientific basis for further studying the toxins secretion mechanism of C.tritonis.

Identification of Monitoring Organ in Bivalves for Early Warning of Paralytic Shellfish Toxins Accumulation

Bivalve farming plays a dominant role in mariculture in China.Paralytic shellfish toxins(PSTs)can be accumulated in bivalves and cause poisoning the consumers.A sensitive detection of PSTs can provide early warning to decrease poisoning events in bivalve consuming.PSTs are traditionally examined using the whole soft-tissues.However,PSTs accumulation varies dramatically in different tissues of bivalves.Some tough tissues/organs(such as mantle),which account for a large proportion of the total soft body,exhibit a lower accumulation of PSTs and make the toxin extraction time-and reagent-consuming,potentially decreasing the accuracy and sensitivity of PSTs monitoring in bivalves.To develop a sensitive and cost-effective approach for PSTs examination in massively farmed bivalves,we fed three commercially important bivalves,Yesso scallop Patinopecten yessoensis,Pacific oyster Crassostrea gigas,and blue mussel Mytilus edulis with PSTs-producing dinoflagellate Alexandrium catenella,and detected PSTs concentration in different tissues.For all three bivalve species,the digestive gland accumulated much more PSTs than other tissues,and the digestive gland’s toxicity was significantly correlated with the PSTs toxicity of the whole soft-tissues,with r^(2)=0.94,0.92,and 0.94 for Yesso scallop,Pacific oyster,and blue mussel,respectively.When the toxicity of the whole soft-tissues reached 80μgSTXeq(100g)^(−1),the regulatory limit for commercial shellfish,the digestive gland’s toxicity reached 571.48,498.90,and 859.20μgSTXeq(100g)^(−1) in Yesso scallop,Pacific oyster,and blue mussel,respectively.Our results indicate that digestive gland can be used for the sensitive and cost-effective monitoring of PSTs in bivalves.

Prospective observational study to assess the validity of the functional disability score in patients with blepharospasm,hemifacial spasm and synkinesis treated with botulinum toxin injection

Background:Benign essential blepharospasm(BEB),aberrant facial nerve degeneration and hemifacial spasm(HFS)are all examples dystonia which,though not life-threatening,can have a significant impact on patient quality of life.The need for reliable self-rating surveys to monitor functional disability is fundamental.The Blepharospasm Disability Index(BSDI)is already a widely utilised and validated selfrating score for blepharospasm whilst the functional disability score(FDS)requires further validation.The principle aim of this study is to repeat validation of the FDS against the BSDI,which has been validated by several groups since its original description but only in patients with BEB.Methods:A randomised blinded prospective cohort study was conducted at a single unit on 38 patients with BEB,aberrant facial nerve degeneration and HFS.Patients were blinded to complete the FDS followed by the BSDI or the BSDI followed by the FDS with a 30-minute interval.Results:Both the FDS and BSDI were found to be reliable with high internal consistency and test-retest reliability.Both scales were also found to be moderately correlated with the Jankovic disease severity score.Conclusions:This study is the first to use the FDS as a rating scale in patients with HFS and aberrant facial nerve degeneration.It is also the first study to formally validate the FDS as an acceptable rating scale for patients with dystonia and in particular it provides validation for its use in patients with HFS and aberrant facial nerve degeneration.

Systematic evaluation of the efficacy and safety of botulinum toxin type A in the treatment of spastic cerebral palsy

Background:Botulinum toxin type A(BTX-A)is a neuromuscular blocking agent.BTX-A inhibits acetylcholine release,causes neuromuscular transmission impairment,and decreases muscle spasms.Objective:To explore the efficacy and safety of BTX-A injection in the treatment of spastic cerebral palsy through systematic evaluation and to provide a reference for the clinical use of BTX-A.Methods:We used“Cerebral palsy”and“BTX-A”as the subject terms and used a combination of subject terms and free words for the search.We searched 7 databases,including CNKI,Wanfang,VIP,Sinomed,PubMed,Embase,and Web of science.Based on the inclusion and exclusion criteria,we screened the articles by reading their titles,abstracts,and full texts and finally included relevant literature for systematic evaluation.Result:A total of 93 papers were systematically evaluated,revealing that BTX-A injection treatment can effectively reduce muscle tone,increase joint mobility,improve gait and motor posture,and enhance gross motor movements in patients with spastic cerebral palsy.The benefits of BTX-A treatment can be sustained for 3–6 months,with motor ability improvement lasting up to 1 year.Combining BTX-A treatment with rehabilitation or external fixation therapy can enhance its efficacy.However,the effectiveness of BTX-A treatment is influenced by several factors,such as the dosage,number of injections,and patient age.Adverse reactions to BTX-A treatment are typically mild and can be relieved within 1–2 weeks.Conclusion:BTX-A injection is relatively safe but reversible.

李佃贵教授防治胆结石经验

浊毒理论是中医学体系中的重要组成部分,李佃贵教授将浊毒论与临床紧密结合,提出浊毒致病观。以“浊毒论”为基础,探讨胆结石的病机及治则治法,认为脾失升清,胃失降浊,肝失疏泄,胆失清净是本病发生的基础;浊毒内蕴,胆腑炼汁为石为核心病机;病位在肝胆,与脾胃密切相关。在胆结石的发生发展过程中,五脏失调均会引起浊毒内蕴,故治疗当化浊解毒,以求胆清汁净;提出化浊解毒的治疗总则,与肝相关则养阴柔肝,与胆相关则清热化湿,与脾胃相关则健脾祛湿,与气相关则疏肝利胆,与血相关则活血化瘀,坚积难去则软坚散结。

康乃馨抗尖孢镰刀菌无性系诱变技术

枯萎病是康乃馨鲜切花种植过程中较为严重的真菌病害之一,该病的病原菌是尖孢镰刀菌康乃馨专化型(Fusarium oxysporum f.sp.dianthi),培育和合理种植抗病品种是防控康乃馨枯萎病最有效的方法之一。应用植物体细胞无性系变异及真菌毒素加压筛选技术,可以定向培育康乃馨抗病育种材料并加快抗病品种选育速度,为康乃馨抗病育种提供新的思路。为获得抗枯萎病的康乃馨育种中间材料,以感枯萎病的康乃馨多花品种紫蝴蝶组培苗为试验材料,诱导愈伤组织并进行悬浮培养建立悬浮培养系,再用甲基磺酸乙酯(EMS)诱变后添加尖孢镰刀菌毒素粗提液筛选抗病细胞系。结果表明,诱导愈伤组织最适宜的培养基是Murashig-Skoog培养基+麦草畏1.0 mg/L;筛选出EMS最佳处理组合为0.4%处理4 h;在80.0%的粗毒素培养基上培养10 d是康乃馨抗尖孢镰刀菌无性系筛选较适宜的选择压;诱导康乃馨再生植株较好的激素组合是苄氨基腺嘌呤(BA)0.5 mg/L+噻苯隆(TDZ) 0.1 mg/L+萘乙酸(NAA)0.1 mg/L;经人工接种尖孢镰刀菌进行抗病性鉴定后发现,紫蝴蝶抗病无性系的病情指数为45,为中抗水平。

花宝金教授调气解毒理论防治放射性肺炎的临证思路

放射性肺炎是由于辐射之火热毒邪袭肺,导致热毒内盛,气阴两虚,瘀血阻络,气机失调。其中热毒内盛,气机失调是核心病机。花宝金教授基于调气解毒理论,灵活运用清热解毒,益气养阴,活血化瘀,调畅气机等治法,同时注重局部与整体相结合,辨证辨病相结合,在放射性肺炎的预防期未病先防,急性期既病防变,恢复期瘥后防复,为防治放射性肺炎提供了新的临证思路,具有较好的临床价值。

适配体生物传感器检测细菌毒素的研究进展

细菌毒素是细菌产生的次生代谢产物,是人类健康的主要威胁之一。食品中细菌毒素的污染已成为人们迫切关注的主要食品安全问题之一。因此,开发出简便、快速、灵敏检测食品中细菌毒素的方法具有重要意义。核酸适体是体外筛选的单链寡核苷酸序列,能特异性识别靶标。具有易合成、易标记、易修饰、高灵敏度、高特异性和低分子量等优点,被广泛用于生物传感器中。本文简要介绍了细菌毒素的分类及危害,重点综述了基于光学和电化学技术的适配体生物传感器在细菌毒素检测和定量方面的最新进展及应用,以期为细菌毒素的检测提供研究基础。

2019-2021年中国近海有毒有害微藻和藻毒素分布数据集

中国近海包括渤海、黄海、东海和南海,海洋资源丰富,对国家经济发展有着重要战略意义。近年来,藻类及藻类产生的藻毒素造成的水体安全问题日益严重,中国近海有害藻华的发生频率和规模不断增加。本数据集汇总整理了2019-2021年中国近海有毒有害微藻和藻毒素调查数据,包括3429条微藻数据、33240条微藻分子物种数据、489条浮游植物中藻毒素数据、76条海水中藻毒素数据和440条经济物种体内藻毒素数据。提供数据的各研究团队通过项目内部和国际互校确保检测数据的准确性。为展示中国近海有毒有害微藻和藻毒素的分布情况,提供了115幅微藻分布图和29幅藻毒素分布图。本数据集可以为中国近海有毒有害微藻和藻毒素的时空分布规律研究、有害藻华的预防预警及防治等提供数据支撑。

5针法A型肉毒杆菌毒素注射在治疗膀胱过度活动症中的应用研究

目的对比分析膀胱内注射A型肉毒杆菌毒素治疗膀胱过度活动症过程中,采用5针注射法与标准的20针注射法的疗效及安全性。方法回顾性分析2015年1月至2022年12月在杭州市第三人民医院泌尿外科接受膀胱内注射A型肉毒杆菌毒素治疗的48例膀胱过度活动症患者,根据注射针数分为两组,各24例,观察组采用5针注射法,对照组采用标准的20针注射法。分别记录两组患者治疗前后平均每日排尿次数、膀胱过度活动症分问卷(international consultation on incontinence questionnaire-overactive bladder,ICIQ-OAB)、膀胱过度活动症调查问卷(overactive bladder,OAB-Q)评分、视觉模拟评分(visual analogue scale,VAS)评分,以及治疗后患者全身状况改善问卷(patient generated index,PGI-I)评分、并发症发生率和重复注射意愿情况。结果两组患者年龄、性别、病程以及治疗前平均每日排尿次数和各评分基线数据等一般资料差异无统计学意义,具有可比性。所有患者均完成治疗,与治疗前相比,治疗后平均每日排尿次数、ICIQ-OAB、OAB-Q均有所改善(P<0.05),两组比较差异无统计学意义(P>0.05)。两组治疗后各评分数据、并发症发生率,差异均无统计学意义(P>0.05)。但是观察组患者更愿意接受再次注射(P<0.05)。结论5针法膀胱内注射A型肉毒杆菌毒素治疗膀胱过度活动症的疗效及安全性与标准的20针法相似,更容易被患者所接受,是一种安全有效的替代标准技术的方法。

2940 nm点阵激光联合A型肉毒毒素改善眶周皮肤老化的临床疗效

目的观察2940 nm点阵激光联合A型肉毒毒素改善眶周皮肤老化的临床疗效。方法将105例求美者随机分为3组:联合治疗组(35例),注射对照组(35例)和激光对照组(35例)。联合治疗组:先采用2940 nm点阵激光治疗1次,两周后采用A型肉毒毒素进行1次注射,间隔2个月再次使用2940 nm点阵激光治疗1次;注射对照组:仅采用A型肉毒毒素进行1次注射;激光对照组仅使用2940nm点阵激光治疗2次,每次间隔2个月。三组均随访26周,由医师及求美者对其静态和动态的眶周皱纹分别进行客观评价及满意度打分。结果采用皱纹严重等级量表(WSRS)统计分析3组求美者眶周皮肤老化均得到改善。面部动态皱纹的治疗效果:注射组4周有效率达到了93.3%,26周降到6.7%;激光组4周有效率为13.3%,26周达到了60%;联合组4周的有效率达到了93.3%,26周达到66.7%。面部静态皱纹的治疗效果:注射组4周有效率达到了73.3%,26周降到3.3%;激光组4周有效率为36.7%,26周达到了73.3%;联合组4周的有效率达到了83.3%,26周达到80%。结论2940nm点阵激光联合A型肉毒素改善眶周皮肤老化,起效快,效果显著,持续时间长,安全可靠,值得推广。

基于Bt毒素的杀虫蛋白理性设计与创新应用策略

Bt毒素是源于苏云金芽孢杆菌(Bacillusthuringiensis)的具有特殊杀虫功能的大分子蛋白,其制剂和转基因作物已广泛用于害虫防治,产生了巨大的经济和社会生态效益。围绕Bt毒素挖掘和提升其应用价值是持续研究的热点,特别是随着Bt毒素结构功能和作用机制日趋明晰,为其功能修饰和创新应用创造了条件,相关研究蓬勃发展,成效显著。大量研究表明,采用定点突变、结构域替换或融合以及抗独特型抗体模拟等策略,是理性设计活性更高、稳定性更强、杀虫谱更广、非靶标生物安全性更高甚至是可用于害虫抗药性治理的有别于母体Bt毒素的突变体、结构杂合体乃至功能效应物抗体等新型杀虫蛋白的有效手段;此外,采用催化毒素活化、驱动毒素靶向受体结合、促进毒素表达以及同源或异源杀虫材料复配或共表达的协同促效等创新增效策略,也是助推Bt毒素应用价值的重要手段。本文总结了Bt毒素结构功能和作用机制,梳理了基于Bt毒素功能修饰的突变体、结构杂合体以及功能效应物抗体等新型杀虫蛋白理性设计和基于Bt毒素功能增效的创新应用策略等相关研究进展,并结合作者团队在模拟Bt毒素杀虫功能效应物抗体靶向设计研发方面的最新成果,探讨了基于Bt毒素的杀虫蛋白理性设计与创新应用策略未来发展动向及潜在可行捷径,为相关研究提供较为全面的最新有价值的文献资料和启发思路。

腹膜恶性肿瘤从三焦癌毒辨治的理论依据与临床应用

腹膜恶性肿瘤包括原发性腹膜癌和继发性腹膜恶性肿瘤,既往对其认识较少,随着外科肿瘤减灭术和腹腔热灌注化疗的兴起,腹膜恶性肿瘤的理念逐渐被接受,相关研究日益增多。结合历代古籍和临床经验,针对腹膜恶性肿瘤的特点,本文提出三焦为其病位,正虚癌毒为其病因,癌毒内结、三焦运化不畅为其病机;从三焦癌毒立法进行辨证论治,其治则为以攻毒为第一要素,以三焦为本,调达整体,并列举临床验案以资参考。

基于毒邪与络病学说探讨免疫炎症在慢性心力衰竭中作用

慢性心力衰竭(以下简称心衰),是各种心脏疾病的终末期阶段,其发病机制与免疫炎症密切相关。免疫细胞激活后,产生大量炎症因子,损伤心肌细胞,导致心肌纤维化、心室重构等病理改变。“毒损心络”是心衰发生、发展的主要病机,贯穿心衰发展的始终,以不同的形式影响心衰的发展及预后。该文通过查阅相关文献,基于毒邪与络病学说,从免疫炎症与心衰的关系及中药防治进行系统梳理,提出心衰早期,以心气虚为主,兼心气阴虚,尚未化毒、入络;心衰中期,心气亏虚为本,痰浊、瘀血、水饮留滞络脉,有化毒、入络趋势;心衰晚期,痰、瘀、水蕴结成毒,损伤心络。中药以复方治疗为主,多具有益气温阳、化瘀利水、解毒通络之功效,通过恢复M1/M2、Th1/Th2及Th17/Treg细胞之间的动态平衡,调节促炎/抗炎因子,从根本上祛除或缓解因毒邪产生的病理状态,减轻炎症,调整心之气血阴阳,达到扶正祛邪、保护心功能的作用,为临床防治心衰提供指导。

基于多交叉置换扩增和纳米生物传感技术快速检测肺炎支原体方法的建立

目的建立一种简单、灵敏、快速的肺炎支原体(MP)检测方法,并对其应用性进行验证和评价。方法利用多交叉置换扩增(MCDA)技术对肺炎支原体特异基因CARDS毒素基因进行扩增,利用侧流免疫层析生物传感(LFB)技术读取扩增结果,命名该方法为MP-MCDA-LFB。分析扩增反应在60~67℃(间隔1℃)的扩增效率,筛选最适反应温度;分析分别扩增10、20、30、40 min时能够检测到的最低核酸浓度,筛选最佳反应时间。利用10倍系列稀释的肺炎支原体核酸分析MP-MCDA-LFB方法的灵敏度和检测限,利用35株非肺炎支原体菌株分析MP-MCDA-LFB方法的特异性。利用MP-MCDA-LFB方法检测80份疑似MP感染的临床样本,并与RT-PCR法检测结果进行比较,分析MP-MCDA-LFB方法的临床应用性。结果MP-MCDA-LFB能够实现对肺炎支原体CARDS毒素基因的快速检测。其最佳反应温度为63℃,最短反应时间为40 min,整个检测过程可在1 h内。MP-MCDA-LFB方法具有较高的灵敏度和特异性,其检测限低至45 ng/L,与其他临床表现相似的病原体无交叉反应,特异性为100%。MP-MCDA-LFB方法从80份临床样本中检出45份阳性样本(56.3%),检出率与RT-PCR方法一致。结论本研究建立的以CARDS毒素基因为靶标的MP-MCDA-LFB检测方法具有简单、快速、灵敏度高、特异性强的优点,在基层医疗机构和现场检测具有较好的应用潜力。