The objective of this study was to develop a rapid and precise detection technique for por-cine Senecavirus A(SVA)employing reverse transcription recombinase polymerase amplification(RT-RAA)in conjunction with CRISPR Cas13a technology.Additionally,the study aimed to opti-mize the assay's reaction conditions to enhance amplification efficiency.Eight RT-RAA primer sets were designed based on the conserved gene sequence of porcine SVA,and a series of reaction condi-tions were evaluated to refine the RT-RAA reaction system.Subsequently,CRISPR-derived RNA(crRNA)sequences were developed and selected to construct the RT-RAA-CRISPR reaction sys-tem.The method's specificity was determined by examining six prevalent porcine pathogenic nucleic acids,while its sensitivity was assessed using SVA cRNA standards quantified by digital PCR.The method's stability and the consistency of clinical sample analysis were also evaluated.The findings revealed that the optimized RT-RAA and CRISPR reaction systems exhibited the highest amplifi-cation efficiency at a reaction temperature of 37 ℃.Among the eight crRNAs,five were identified as exhibiting the strongest detection signals.The formulated RT-RAA-CRISPR Cas13a method demonstrated exceptional specificity,showing no cross-reactivity with other common porcine disea-ses,including ASFV,PRRSV,PEDV,PCV2,CSFV,and PRV.The method achieved high sensitivi-ty,detecting as low as 0.86 copies/μL of SVA,and exhibited stable fluorescence output,robust re-producibility,and the ability to complete clinical sample analysis within 50 minutes.Consistency e-valuation with six positive and 58 negative samples indicated 100％agreement in outcomes.These results substantiate that the study successfully established a rapid and specific RT-RAA-CRISPR Cas13a detection method for the on-site identification of porcine Senecavirus A,demonstrating high specificity and sensitivity,and holds promise for application in SVA monitoring and control initia-tives.

The objective of this study was to develop a rapid and precise detection technique for por-cine Senecavirus A(SVA)employing reverse transcription recombinase polymerase amplification(RT-RAA)in conjunction with CRISPR Cas13a technology.Additionally,the study aimed to opti-mize the assay's reaction conditions to enhance amplification efficiency.Eight RT-RAA primer sets were designed based on the conserved gene sequence of porcine SVA,and a series of reaction condi-tions were evaluated to refine the RT-RAA reaction system.Subsequently,CRISPR-derived RNA(crRNA)sequences were developed and selected to construct the RT-RAA-CRISPR reaction sys-tem.The method's specificity was determined by examining six prevalent porcine pathogenic nucleic acids,while its sensitivity was assessed using SVA cRNA standards quantified by digital PCR.The method's stability and the consistency of clinical sample analysis were also evaluated.The findings revealed that the optimized RT-RAA and CRISPR reaction systems exhibited the highest amplifi-cation efficiency at a reaction temperature of 37 ℃.Among the eight crRNAs,five were identified as exhibiting the strongest detection signals.The formulated RT-RAA-CRISPR Cas13a method demonstrated exceptional specificity,showing no cross-reactivity with other common porcine disea-ses,including ASFV,PRRSV,PEDV,PCV2,CSFV,and PRV.The method achieved high sensitivi-ty,detecting as low as 0.86 copies/μL of SVA,and exhibited stable fluorescence output,robust re-producibility,and the ability to complete clinical sample analysis within 50 minutes.Consistency e-valuation with six positive and 58 negative samples indicated 100％agreement in outcomes.These results substantiate that the study successfully established a rapid and specific RT-RAA-CRISPR Cas13a detection method for the on-site identification of porcine Senecavirus A,demonstrating high specificity and sensitivity,and holds promise for application in SVA monitoring and control initia-tives.

Objective To observe the value of ultrasonic biomicroscopy(UBM)imaging for diagnosing chestnut thorn foreign bodies in anterior segment.Methods A total of 70 patients who underwent UBM examination due to chestnut thorn injury or residue of chestnut thorn after removal and then operation were retrospectively enrolled.The distance of chestnut thorn foreign bodies from corneal limbus,and the subconjunctival depth and length of them were measured based on UBM imaging.The efficacy of UBM for diagnosing chestnut thorn foreign bodies in anterior segment was evaluated according to surgical findings,and the correlation of quantitative parameters of foreign bodies were analyzed.Results Among 70 cases of chestnut thorn foreign bodies in anterior segment,single ocular tissue(5/70,7.14％)involvement was detected in 5 cases,while multiple ocular tissues involvement(65/70,92.86％)were noticed in 65 cases according to UBM.Totally 109 foreign bodies were detected by UBM,among which 98 were consistent with surgical findings,with the diagnostic accuracy of 89.91％(98/109),while 11 foreign bodies were not found during operation,with the false positive rate of 10.09％(11/109).Meanwhile,4 foreign bodies found and removed with operation were not displayed with UBM,and the false negative rate of UBM was 3.92％(4/102).Among 98 chestnut thorn foreign bodies correctly diagnosed with UBM,10 located in stromal layer of cornea(10/98,10.20％)and 88 located in ocular wall(88/98,89.80％),including 43 subconjunctival(43/88,48.86％)and 45 sclerotic(45/88,51.14％)foreign bodies,with the median length of 0.200 mm.The distance of 88 foreign bodies in ocular wall from corneal limbus was(2.963±1.504)mm,and the subconjunctival depth was(0.785±0.388)mm.The distance of foreign bodies from corneal limbus were positively correlated with their subconjunctival depth(r,=0.361,P＜0.010),while their lengths were not correlated with their distance from corneal limbus and subconjunctival depth(both P＞0.05).Conclusion UBM imaging was valuable for clinical diagnosis and localization of chestnut thorn foreign bodies in anterior segment.

Objective To observe the value of ultrasonic biomicroscopy(UBM)imaging for diagnosing chestnut thorn foreign bodies in anterior segment.Methods A total of 70 patients who underwent UBM examination due to chestnut thorn injury or residue of chestnut thorn after removal and then operation were retrospectively enrolled.The distance of chestnut thorn foreign bodies from corneal limbus,and the subconjunctival depth and length of them were measured based on UBM imaging.The efficacy of UBM for diagnosing chestnut thorn foreign bodies in anterior segment was evaluated according to surgical findings,and the correlation of quantitative parameters of foreign bodies were analyzed.Results Among 70 cases of chestnut thorn foreign bodies in anterior segment,single ocular tissue(5/70,7.14％)involvement was detected in 5 cases,while multiple ocular tissues involvement(65/70,92.86％)were noticed in 65 cases according to UBM.Totally 109 foreign bodies were detected by UBM,among which 98 were consistent with surgical findings,with the diagnostic accuracy of 89.91％(98/109),while 11 foreign bodies were not found during operation,with the false positive rate of 10.09％(11/109).Meanwhile,4 foreign bodies found and removed with operation were not displayed with UBM,and the false negative rate of UBM was 3.92％(4/102).Among 98 chestnut thorn foreign bodies correctly diagnosed with UBM,10 located in stromal layer of cornea(10/98,10.20％)and 88 located in ocular wall(88/98,89.80％),including 43 subconjunctival(43/88,48.86％)and 45 sclerotic(45/88,51.14％)foreign bodies,with the median length of 0.200 mm.The distance of 88 foreign bodies in ocular wall from corneal limbus was(2.963±1.504)mm,and the subconjunctival depth was(0.785±0.388)mm.The distance of foreign bodies from corneal limbus were positively correlated with their subconjunctival depth(r,=0.361,P＜0.010),while their lengths were not correlated with their distance from corneal limbus and subconjunctival depth(both P＞0.05).Conclusion UBM imaging was valuable for clinical diagnosis and localization of chestnut thorn foreign bodies in anterior segment.

Animals ; Antibodies, Monoclonal ; pharmacokinetics ; therapeutic use ; Antibodies, Viral ; therapeutic use ; Hepatitis B virus ; immunology ; Histocompatibility Antigens Class I ; metabolism ; Humans ; Macaca fascicularis ; Mice ; Protein Binding ; Protein Engineering ; Receptors, Fc ; metabolism