High-peak-power electro-optically Q-switched laser with a gradient-doped Nd:YAG crystal.

We report on a high-peak-power electro-optically Q-switched laser emitting a near-diffraction-limited beam profile at 1064 nm by using a gradient-doped Nd:YAG crystal. The gradient-doped crystal features a unique combination of a reduced thermal lens effect through effectively spreading the heat load distribution within its volume. Its performance is compared with those of Nd:YAG crystals with uniform volume doping distribution operating in the Q-switched regime with the same laser configuration, demonstrating the higher average and peak power achievable with the gradient-doped crystal. The maximum average output power amounts to 6.9 W at a pulse repetition rate of 2 kHz, which corresponds to a maximum peak power of ∼585kW. Compared to homogeneous dopant crystals, the slope efficiency and average output power increased by 30.8% and 21.1%, respectively.

A rapid and convenient derivatization method for quantitation of short-chain fatty acids in human feces by ultra-performance liquid chromatography/tandem mass spectrometry.

RATIONALE: Short-chain fatty acids (SCFAs) are associated with intestinal microbiota and diseases in humans. SCFAs have a low response in mass spectrometry, and in order to increase sensitivity, reduce sample consumption, shorten analysis time, and simplify sample preparation steps, a derivatization method was developed. METHODS: We converted seven SCFAs into amide derivatives with 4-aminomethylquinoline. The reaction occurred for 20 min at room temperature. The analytes were separated on a reversed-phase C18 column and quantitated in the positive ion electrospray ionization mode using multiple reaction monitoring. Acetic acid-d4 was used as the stable-isotope-labeled surrogate analyte for acetic acid in the working solutions, while the other stable-isotope-labeled standards were used as internal standards (ISs). RESULTS: Method validation showed that the intra-day and inter-day precision of quantitation for the seven SCFAs over the whole concentration range was ≤3.8% (n = 6). The quantitation accuracy ranged from 85.5% to 104.3% (n = 6). Most important, the collected feces were vortexed immediately with ethanol. CONCLUSIONS: This study provides a new derivatization method for a precise, accurate, and rapid quantitation of SCFAs in human feces using ultra-performance liquid chromatography/tandem mass spectrometry. This method successfully determined the concentration of SCFAs in human feces and could assist in the exploration of intestinal microbiota and diseases.

A TaqMan probe based real-time PCR for the detection of Decapod iridescent virus 1.

Decapod iridescent virus 1 (DIV1) was proven to be the aetiological agent of a disease causing mass die-offs of shrimp, prawn and crayfish. The specific purpose of this study was to develop a new sensitive real-time PCR method for the specific detection of DIV1. A pair of primers that amplify a 142 bp fragment and a TaqMan probe were selected for the major capsid protein gene of DIV1. They were shown to be specific for DIV1 and did not react with other common shrimp pathogens or healthy shrimp DNA. The method could detect as virus levels as low as 1.2 copies of DIV1 plasmid DNA.

Dietary protein intake and risk of type 2 diabetes: a dose-response meta-analysis of prospective studies.

PURPOSE: The association between dietary protein intake and type 2 diabetes risk has been inconsistent in the previous epidemiological studies. We aimed to quantitatively assess whether dietary total, animal, and plant protein would be associated with type 2 diabetes risk. METHODS: A comprehensive literature review was conducted to identify related articles by searching PubMed, Embase, Web of Science, and Wiley Online Library through 20th March 2018. Generalized least squares for trend estimation and restricted cubic spline regression model were applied in the dose-response analysis. RESULTS: Eight publications with ten prospective cohorts of 34,221 type 2 diabetes cases were included. After adjustment of potential confounders, a 5% of energy increment from dietary total and animal protein intake was related to a 9% (1.04, 1.13; I2 = 42.0%) and 12% (95% CI 1.08, 1.17; I2 = 14.0%) higher risk of type 2 diabetes respectively. However, for plant protein, a significant U-shaped curve was observed with the most risk reduction at intake of about 6% of energy intake from plant protein intake (Pnonlinearity = 0.001). The results were robust in sensitivity analysis and no publication bias was detected. CONCLUSIONS: These findings indicate that the consumption of protein particularly animal protein may be associated with an increased risk of type 2 diabetes.

Magnesium lithospermate B improves the gut microbiome and bile acid metabolic profiles in a mouse model of diabetic nephropathy.

Magnesium lithospermate B (MLB) is a new drug marketed in China to treat angina, but its low oral bioavailability limits its clinical application to the intravenous route. Paradoxically, orally administered low-dose MLB was found to alleviate kidney injury in diabetic nephropathy (DN) rats, but its mechanism of action remains unknown. In recent years, the kidney-gut axis has been suspected to be involved in kidney damage pathogenesis, potentially representing a non-classical pathway for pharmacologic intervention. To ascertain whether MLB targets the kidney-gut axis, streptozotocin (STZ)-treated mice were prepared as a mouse model of DN. The STZ mice were treated with MLB (50 mg kg-1 d-1, p.o.) for 8 weeks. Twenty-four-hour urinary albumin was detected to mirror kidney function. At week 4, 6, 8, feces were collected; bile acids (BAs) were quantified to examine the alterations in the BA metabolic profiles, and bacterial 16S rRNA gene fragments were sequenced to identify alterations in gut microbial composition. In STZ mice, 24-h urinary albumin levels and total fecal BAs, especially cholic acids (CAs) and deoxycholic acids (DCAs) were greatly increased, and the gut microbiome was dramatically shifted compared with control mice. Oral administration of MLB significantly decreased 24-h urinary albumin levels and total BAs, CAs and DCAs, and reversed CA:TCA (taurocholic acid) and DCA:CA ratios. It also changed the microbiome composition in STZ mice based on operational units. Thus the therapeutic effect of MLB on kidney injury might be attributed (at least partially) to its ability to modulate the disordered gut microbiome and BA metabolism.

The joint effects of major lifestyle factors on colorectal cancer risk among Chinese men: A prospective cohort study.

Previous studies have suggested individual healthy lifestyle factors are related to lower risk of colorectal cancer. Their joint effects, however, have rarely been investigated. We aimed to assess the combined lifestyle impact on colorectal cancer risk and to estimate the population attributable risks of these lifestyle factors. Using data from the Shanghai Men's Health Study (2002-2013), we constructed healthy lifestyle index composing the following lifestyle factors: smoking, alcohol consumption, diet, waist-hip ratio and exercise participation. Cox proportional hazards models were used to calculate the hazard ratios (HRs) and 95% confidence intervals (CIs). Over a median of 9.28 years' follow-up, 671 colorectal cancer cases occurred (400 colon cancer and 274 rectal cancer) among 59,503 men. Each increment of healthy lifestyle index was associated with a 17% lower risk of colorectal cancer (HR = 0.83, 95% CI: 0.78, 0.89), 10% of colon cancer (HR = 0.90, 95% CI: 0.83, 0.99) and 27% of rectal cancer (HR = 0.73, 95% CI: 0.66, 0.82). If all men in the cohort followed a lifestyle as defined by these five factors, 21% colorectal cancer cases would have been prevented (PAR = 21%, 95% CI: 4%, 36%). In conclusion, combined lifestyle factors are significantly related to lower risk of colorectal cancer and the effects are more pronounced on rectal cancer than on colon cancer.

Complete genome sequence of shrimp hemocyte iridescent virus (SHIV) isolated from white leg shrimp, Litopenaeus vannamei.

Infection with shrimp hemocyte iridescent virus (SHIV), a new virus of the family Iridoviridae isolated in China, results in a high mortality rate in white leg shrimp (Litopenaeus vannamei). The complete genome sequence of SHIV was determined and analyzed in this study. The genomic DNA was 165,809 bp long with 34.6% G+C content and 170 open reading frames (ORFs). Dotplot analysis showed that the longest repetitive region was 320 bp in length, including 11 repetitions of an 18-bp sequence and 3.1 repetitions of a 39-bp sequence. Two phylogenetic trees were constructed based on 27 or 16 concatenated sequences of proteins encoded by genes that are conserved between SHIV homologous and other iridescent viruses. The results of this study, suggest that SHIV should be considered a member of the proposed new genus "Xiairidovirus".

Detection and quantification of shrimp hemocyte iridescent virus by TaqMan probe based real-time PCR.

Shrimp hemocyte iridescent virus (SHIV) is a recently reported shrimp virus, which threats the cultured white-leg shrimp Litopenaeus vannamei and can cause huge economic loss in shrimp farming industry in China. A quantitative real time polymerase chain reactio (qPCR) assay was developed using a TaqMan probe to detect and quantify SHIV. A pair of qPCR primers, which amplify a 188 bp DNA fragment, and a TaqMan probe were selected from ATPase gene (ORF114R) of the SHIV genome. The primers and TaqMan probe used in this assay were shown to be specific for SHIV and did not react with White spot syndrome virus (WSSV), Infectious hypodermal and hematopoietic necrosis virus (IHHNV), Hepatopancreatic parvovirus (HPV), Vibrio parahaemolyticus causing acute hepatopancreatic necrosis disease (VPAHPND), and Enterocytozoon hepatopenaei (EHP), or healthy shrimp DNA. The detection limit of the qPCR method was as low as 4 copies per reaction. The diagnostic sensitivity and the diagnostic specificity of the qPCR compared with nested PCR were 95.3% and 99.2%, respectively. The resulting standard curves showed high correlation coefficient values (R2 = 0.998) in the range of 4 × 109 to 4 × 100 DNA copies/reaction. Test of farm samples showed that SHIV was detected in L. vannamei, Fenneropenaeus chinensis and Macrobrachium rosenbergii contained SHIV ranged from 1.21E+02 to 7.95E+07 copies (µg DNA)-1. Quantitative detection of different tissues in artificial infected shrimp showed that haemolymph contained the highest SHIV load and muscle contained lowest SHIV load.

[Analysis of adverse reactions' factors to Danhong injection--nested case control study by using hospital centralized monitoring data].

To study the adverse reactions' factors to Danhong injection in the real world. A multi-center, large sample and prospective hospital centralized monitoring method was adopted, and 30 888 cases of Danhong injection from 37 national 3A hospitals were collected to carry out a nested case control design study. These cases were divided into adverse reaction group and non-adverse group. Single factor logistic regression and multiple factor logistic regression were used to analyze data, and investigate the correlation between adverse reaction and gender, allergy history, methods of administration, and combined drug use. One hundred and eight cases of adverse reactions in 30 888 patients were determined, with an incidence of 0.35%. The results showed that Danhong injection combined with other medication(potassium mendoxine magnesium, thymic peptide, celecoxib, fumarate bisoprolol) with history of adverse reactions including scephalosporin allergy and proprietary Chinese medicine allergies had more adverse reactions than the control group(P<0.05, estimated coefficient>0), indicating that these six factors were the risk factors for the adverse reaction of Danhong injection. The adverse reaction of Danhong injection combined with the aspirin was less than that in the control group(P<0.05, estimated coefficient<0), indicating that the aspirin was a non-risk factor for the adverse reaction of Danhong injection. All the above results indicate that the adverse factors to Danhong injection include scephalosporin allergy, patent Chinese medicine allergy, Danhong injection combined with medication(potassium mendoxine magnesium, thymic peptide, celecoxib, fumarate bisoprolol), suggesting special attention shall be paid in clinical application.

[Efficacy and safety of ribavirin aerosol in children with hand-foot-mouth disease].

OBJECTIVE: To evaluate the efficacy and safety of ribavirin aerosol in children with hand-foot-mouth disease (HFMD). METHODS: A randomized, double-blind, placebo-controlled trial was performed. A total of 119 children with mild HFMD were randomly divided into an observed group (n=59) and a control group (n=60). In the observed group, ribavirin aerosol was given four times within the first hour, followed by once every other hour for the remaining time of the day and day 2; from days 3 to 7, it was given 4 times per day, with 2-3 sprays every time, for 7 days. In the control group, placebo was given in the same way as in the observed group. Additionally, both groups used oral antiviral liquid. The scores of clinical symptoms including oral ulcer, skin rash, nasal congestion, runny nose, sneezing, cough, and fever before and after treatment were recorded to evaluate treatment outcomes. Throat swabs were taken before treatment and 5-7 days after treatment to measure viral load by RT-PCR and to compare the negative conversion rate between the two groups. RESULTS: Fifty-seven patients in the observed group and 56 patients in the control group were tested according to the original research design. After 5-7 days of treatment, the observed group had a significantly higher overall negative conversion rate of enterovirus than the control group (P<0.01). The overall marked response rate and overall response rate of the observed group were 89% and 89%, respectively, significantly higher than those of the control group (29% and 43%). During treatment, there were no adverse reactions such as dizziness, vomiting, and notable decreases in hemoglobin, white blood cells, and platelets in the two groups. CONCLUSIONS: Ribavirin aerosol can be effectively and safely used for treating mild HFMD. With low dosage and few adverse reactions, it holds promise for clinical application.

[Expression of BRCA1 and WWOX and their clinicopathologic implication in breast carcinomas occurring in young women].

OBJECTIVE: To explore the clinicopathologic characteristics and biological markers of breast carcinomas in young women. METHODS: Immunohistochemical SP method was used to study breast cancer susceptibility gene (BRCA1) and WWOX in breast carcinomas of patient ≤ 35 years of age (107 cases) and ≥ 60 years of age (112 cases). The findings were correlated with clinicopathological features. In addition, PCR amplification and direct sequencing were performed to detect the BRCA1 gene mutation of exons 2 and 20 using fresh frozen tissue samples in other 10 patients who were ≤ 35 years of age. RESULTS: The positive rate of BRCA1 protein expression was higher in the young age group [65.4% (70/107)] than that of the old age group [35.7% (40/112)]. ER, PR, HER2, and WWOX protein expression and proliferation marker Ki-67 were no statistically different in the two groups (all P > 0.05). BRCA1 expression was significantly correlated with pTNM and axillary lymph node metastasis (both P < 0.05), but not with ER, PR, HER2 and WWOX protein expression (all P > 0.05). Ki-67 and histological grading showed no statistical correlation (P > 0.05). WWOX protein expression showed no correlation with clinicopathologic characteristics (all P > 0.05). Mutation of exons 2 and 20 of the BRCA1 gene was not detected in any of 10 cases studied. CONCLUSION: BRCA1 cytoplasmic expression statistically correlates with the development and prognosis of breast cancer of young patients.

Caregiver-child interaction as an effective tool for identifying autism spectrum disorder: evidence from EEG analysis.

BACKGROUND: Autism Spectrum Disorder (ASD) is a complex neurodevelopmental disorder that affects individuals across their lifespan. Early diagnosis and intervention are crucial for improving outcomes. However, current diagnostic methods are often time-consuming, and costly, making them inaccessible to many families. In the current study, we aim to test caregiver-child interaction as a potential tool for screening children with ASD in clinic. METHODS: We enrolled 85 preschool children (Mean age: 4.90 ± 0.65 years, 70.6% male), including ASD children with or without developmental delay (DD), and typical development (TD) children, along with their caregivers. ASD core symptoms were evaluated by Childhood Autism Rating Scale (CARS) and Autism Diagnostic Observation Schedule-Calibrated Severity Scores (ADOS-CSS). Behavioral indicators were derived from video encoding of caregiver-child interaction, including social involvement of children (SIC), interaction time (IT), response of children to social cues (RSC), time for caregiver initiated social interactions (GIS) and time for children initiated social interactions (CIS)). Power spectral density (PSD) values were calculated by EEG signals simultaneously recorded. Partial Pearson correlation analysis was used in both ASD groups to investigate the correlation among behavioral indicators scores and ASD symptom severity and PSD values. Receiver operating characteristic (ROC) analysis was used to describe the discrimination accuracy of behavioral indicators. RESULTS: Compared to TD group, both ASD groups demonstrated significant lower scores of SIC, IT, RSC, CIS (all p values < 0.05), and significant higher time for GIS (all p values < 0.01). SIC scores negatively correlated with CARS (p = 0.006) and ADOS-CSS (p = 0.023) in the ASD with DD group. Compared to TD group, PSD values elevated in ASD groups (all p values < 0.05), and was associated with SIC (theta band: p = 0.005; alpha band: p = 0.003) but not IQ levels. SIC was effective in identifying both ASD groups (sensitivity/specificity: ASD children with DD, 76.5%/66.7%; ASD children without DD, 82.6%/82.2%). CONCLUSION: Our results verified the behavioral paradigm of caregiver-child interaction as an efficient tool for early ASD screening.

Apolipoprotein A-I mimetic peptide inhibits atherosclerosis by altering plasma metabolites in hypercholesterolemia.

An apolipoprotein A-I mimetic peptide, D-4F, has been shown to improve vasodilation and inhibit atherosclerosis in hypercholesterolemic low-density lipoprotein receptor-null (LDLr(-/-)) mice. To study the metabolic variations of D-4F ininhibiting atherosclerosis, metabonomics, a novel system biological strategy to investigate the pathogenesis, was developed. Female LDLr(-/-) mice were fed a Western diet and injected with or without D-4F intraperitoneally. Atherosclerotic lesion formation was measured, whereas plasma metabolic profiling was obtained on the basis of ultra-high-performance liquid chromatography in tandem with time-of-flight mass spectrometry operating in both positive and negative ion modes. Data were processed by multivariate statistical analysis to graphically demonstrate metabolic changes. The partial least-squares discriminate analysis model was validated with cross-validation and permutation tests to ensure the model's reliability. D-4F significantly inhibited the formation of atherosclerosis in a time-dependent manner. The metabolic profiling was altered dramatically in hypercholesterolemic LDLr(-/-) mice, and a significant metabolic profiling change in response to D-4F treatment was observed in both positive and negative ion modes. Thirty-six significantly changed metabolites were identified as potential biomarkers. A series of phospholipid metabolites, including lysophosphatidylcholine (LysoPC), lysophosphatidylethanolamine (LysoPE), phosphatidylcholine (PC), phatidylethanolamine (PE), sphingomyelin (SM), and diacylglycerol (DG), particularly the long-chain LysoPC, was elevated dramatically in hypercholesterolemic LDLr(-/-) mice but reduced by D-4F in a time-dependent manner. Quantitative analysis of LysoPC, LysoPE, PC, and DG using HPLC was chosen to validate the variation of these potential biomarkers, and the results were consistent with the metabonomics findings. Our findings demonstrated that D-4F may inhibit atherosclerosis by regulating phospholipid metabolites specifically by decreasing plasma long-chain LysoPC.

Effective components screening and anti-myocardial infarction mechanism study of the Chinese medicine NSLF6 based on "system to system" mode.

BACKGROUND: Shuanglong formula (SLF), a Chinese medicine composed of panax ginseng and salvia miltiorrhiza exhibited significant effect in the treatment of myocardial infarction (MI) in clinical. Because of the complex nature and lack of stringent quality control, it's difficult to explain the action mechanism of SLF. METHOD: In this study, we present a "system to system" (S2S) mode. Based on this mode, SLF was simplified successively through bioactivity-guided screening to achieve an optimized minimal phytochemical composition (new formula NSLF6) while maintaining its curative effect for MI. RESULTS: Pharmacological test combining with the study of systems biology show that NSLF6 has activity for treatment MI through synergistic therapeutic efficacies between total ginsenosides and total salvianolic acids via promoting cardiac cell regeneration and myocardial angiogenesis, antagonistic myocardial cell oxidative damage. CONCLUSIONS: The present S2S mode may be an effective way for the discovery of new composite drugs from traditional medicines.

[Tissue distribution of arsenic of liushen pills and realgar].

This study is to report the tissue distribution of arsenic after giving different doses of realgar and Liushen pills to Beagle dogs, in order to provide basis for the safety evaluation of Liushen pills. ICP-MS was used to measure arsenic concentration, and HPLC-ICP-MS was used to analyze arsenic speciation. The concentration of total arsenic and As(III) + DMA (arsenite + dimethylarsenic acid) increased with dosing of realgar. Total arsenic concentration in most tissues and As(III) + DMA concentration in all tissues of Liushen pills group are lower than that of realgar group, but AsB concentration in liver, spleen and kidney of Liushen pills group increased. The concentration of total arsenic showed a dose-dependent manner with dosage administered. It was indicated that components in Liushen pills can reduce solubility of arsenic in realgar, which may decrease toxicity of realgar.

Vaccine preventable infections and multiple organ dysfunction syndrome in critically ill children in China.

BACKGROUND: Multiple organ dysfunction syndrome (MODS) is a major cause of mortality in PICUs in industrialized countries. As developing countries become more advanced, PICU care comes into reach. The aim of this study was to examine MODS in a PICU in China in comparison with other countries. METHODS: A retrospective 5-year audit (2001-2006) of MODS cases in a 6-bed PICU in the Second Hospital of Lanzhou University, China. RESULTS: Of the 332 PICU patients, 176 (53%) had MODS; the mean age was 35 months, and 60% were <1 year of age. Infectious diseases were the predominant factor underlying MODS (128/176; 73%) with 24% attributable to potentially vaccine preventable infections (14 to Haemophilus influenzae b, 13 to Streptococcus pneumoniae, and 4 to Neisseria meningitidis). The case fatality rate for MODS was 49% with no difference between infectious or noninfectious causes (48% vs. 52%; P > 0.05). The frequency of organ dysfunction in MODS was in decreasing order: respiratory (95%), cardiovascular (89%), gastrointestinal (39%), neurologic (35%), hematologic (16%), renal (14%), and hepatic (4%) systems. Mortality rose with increasing number of organs involved. Gastrointestinal system failure was more common in MODS caused by infection than noninfection (45% vs. 23%; P <0.01) and had a higher mortality (72% vs. 34%; P < 0.001). The case fatality rate for MODS in this study was comparable to rates in other developing countries but higher than in industrialized countries. CONCLUSIONS: MODS is a major cause of mortality in PICU patients in China. In China, infection-related MODS PICU cases could potentially be reduced to rates closer to those in industrialized countries, if conjugated H. influenzae b and conjugated pneumococcal vaccines were introduced into routine practice.

[Raman spectra analysis of Nd : YAG single crystal and its nano-powder].

In the present paper, the authors measured the Raman spectra of YAG/Nd : YAG single crystal, Nd : YAG precursor and the powder sintered at different temperatures. The bands of these Raman spectra were assigned and analyzed. The results show that there is a structure transformation process in the course of sintering Nd : YAG precursor. The powder sintered at 700 degrees C was amorphous and it is of AlO4 tetrahedron structure. With the increase in sintering temperatures, the Raman spectra varied mainly in two respects. One is the decrease in FWHM with the increase in the bands intensity; the other is the bands shift. These should be due to the increase in the order degree of the interface component. Additionally, the difference in the lattice vibration modes between the powders sintered at 800 degrees C and the Nd : YAG single crystal powder was caused by the contribution of the interface component.

Description of a Natural Infection with Decapod Iridescent Virus 1 in Farmed Giant Freshwater Prawn, Macrobrachium rosenbergii.

Macrobrachium rosenbergii is a valuable freshwater prawn in Asian aquaculture. In recent years, a new symptom that was generally called "white head" has caused high mortality in M. rosenbergii farms in China. Samples of M. rosenbergii, M. nipponense, Procambarus clarkii, M. superbum, Penaeus vannamei, and Cladocera from a farm suffering from white head in Jiangsu Province were collected and analyzed in this study. Pathogen detection showed that all samples were positive for Decapod iridescent virus 1 (DIV1). Histopathological examination revealed dark eosinophilic inclusions and pyknosis in hematopoietic tissue, hepatopancreas, and gills of M. rosenbergii and M. nipponense. Blue signals of in situ digoxigenin-labeled loop-mediated isothermal amplification appeared in hematopoietic tissue, hemocytes, hepatopancreatic sinus, and antennal gland. Transmission electron microscopy of ultrathin sections showed a large number of DIV1 particles with a mean diameter about 157.9 nm. The virogenic stromata and budding virions were observed in hematopoietic cells. Quantitative detection with TaqMan probe based real-time PCR of different tissues in naturally infected M. rosenbergii showed that hematopoietic tissue contained the highest DIV1 load with a relative abundance of 25.4 ± 16.9%. Hepatopancreas and muscle contained the lowest DIV1 loads with relative abundances of 2.44 ± 1.24% and 2.44 ± 2.16%, respectively. The above results verified that DIV1 is the pathogen causing white head in M. rosenbergii. M. nipponense and Pr. clarkii are also species susceptible to DIV1.

Development and validation of a TaqMan RT-qPCR for the detection of convert mortality nodavirus (CMNV).

Covert mortality nodavirus (CMNV), an emerging RNA virus, is the pathogen of viral covert mortality disease (VCMD), which has emerged as a cause of serious losses in shrimp aquaculture in China. To improve VCMD diagnosis, a one-step, real-time TaqMan probe-based reverse transcription quantitative PCR (RT-qPCR) was developed in this study. The TaqMan RT-qPCR was optimized firstly, whereby the best results were obtained with 0.2 µM of each primer, 0.2 µM probe, and 0.5 µL Enzyme Mix II. The optimal reaction program was determined as 15 min at 51ºC for reverse transcription and 5 min at 95 ºC, followed by 40 cycles of denaturation at 94 ºC for 10 s, and annealing and extension at 52.7 ºC for 30 s. The optimized assay detected as little as 9.6 pg total RNA from CMNV-infected shrimp and 5.7 copies of the target plasmid. The RT-qPCR assay for CMNV with a high correlation coefficient (r2 = 0.996) was developed basing on the standard curve generated by plotting the threshold cycle values (y) against the common logarithmic copies (log10nc as x; nc is copy number) of pMD20-CMNV. The diagnostic sensitivity and specificity of this assay versus the previously reported RT-qPCR was 96.2% and 98.0%, respectively. This method is highly specific to CMNV, as it showed no cross-reactivity with other common shrimp viruses. It is anticipated that the newly developed and optimized RT-qPCR assay will be instrumental for the rapid diagnosis and quantitation of CMNV.