Recombinase polymerase amplification combined with Pyrococcus furiosus Argonaute for fast Salmonella spp. testing in food safety.

Foodborne illness caused by Salmonella spp. is one of the most prevalent public health problems globally, which have brought immeasurable economic burden and social impact to countries around the world. Neither current nucleic acid amplification detection method nor standard culture method (2-3 days) are suitable for field detection in areas with a heavy burden of Salmonella spp. Here, we developed a highly sensitive and accurate assay for Salmonella spp. detection in less than 40 min. Specifically, the invA gene of Salmonella spp. was amplified by recombinase polymerase amplification (RPA), followed by Pyrococcus furiosus Argonaute (PfAgo)-based target sequence cleavage, which could be observed by a fluorescence reader or the naked eye. The assay offered the lowest detectable concentration of 1.05 × 101 colony forming units/mL (CFU/mL). This assay had strong specificity and high sensitivity for the detection of Salmonella spp. in field samples, which indicated the feasibility of this assay.

Correlating Young's Modulus with High Thermal Conductivity in Organic Conjugated Small Molecules.

Attaining elevated thermal conductivity in organic materials stands as a coveted objective, particularly within electronic packaging, thermal interface materials, and organic matrix heat exchangers. These applications have reignited interest in researching thermally conductive organic materials. The understanding of thermal transport mechanisms in these organic materials is currently constrained. This study concentrates on N, N'-dioctyl-3,4,9,10-perylenedicarboximide (PTCDI-C8 ), an organic conjugated crystal. A correlation between elevated thermal conductivity and augmented Young's modulus is substantiated through meticulous experimentation. Achievement via employing the physical vapor transport method, capitalizing on the robust C═C covalent linkages running through the organic matrix chain, bolstered by π-π stacking and noncovalent affiliations that intertwine the chains. The coexistence of these dynamic interactions, alongside the perpendicular alignment of PTCDI-C8 molecules, is confirmed through structural analysis. PTCDI-C8 thin film exhibits an out-of-plane thermal conductivity of 3.1 ± 0.1 W m-1  K-1 , as determined by time-domain thermoreflectance. This outpaces conventional organic materials by an order of magnitude. Nanoindentation tests and molecular dynamics simulations elucidate how molecular orientation and intermolecular forces within PTCDI-C8 molecules drive the film's high Young's modulus, contributing to its elevated thermal conductivity. This study's progress offers theoretical guidance for designing high thermal conductivity organic materials, expanding their applications and performance potential.

Comparison of Urine and Genital Samples for Detecting Human Papillomavirus (HPV) in Clinical Patients.

Background: Human papillomavirus (HPV) is the main cause of cervical cancer. The aim of the present study was to investigate HPV DNA detection and genotyping on paired genital and urine samples and to evaluate if urine samples could be used to monitor HPV infection. Methods: Study subjects were recruited from one local hospital in Guangdong of China from September 1, 2011, to June 30, 2012. They were invited to participate if they have taken an HPV genotyping assay for clinical diagnosis of the genital-urinary disease or for a health check-up 3-5 days ago. DNA was extracted from paired genital and urine samples; genotyping was performed with the GenoArray assay. Results: A total of 250 patients were recruited, which included 203 females and 47 males. Our results showed that the overall agreement on HPV status between the paired samples was 77.1% (155/201, 95% CI: 0.713-0.829) for females, with a kappa value of 0.523 (95% CI: 0.469-0.632), while the agreement was extremely low in the paired male samples. As to individual genotyping, the greatest agreement was found for HPV16 type-specific identification in females (96.02%, 0.933-0.987), followed by the other 12 high oncogenic risk (HR-HPV) types, while the agreement for low-risk HPV detection is poor (κ < 0.6). Agreement between paired samples showed that HPV detection had a significantly greater concordance in the samples obtained in females than males (p = 0.002). Moreover, the agreement for low-risk HPV detection was significantly lower as compared to HR-HPV detection (48.1% vs. 62.3%, p = 0.044). Conclusion: Despite reduced sensitivity, HPV detection in urine closely represents the same trend that is seen with genital sampling. Urine appears to be an appropriate surrogate sample for HPV DNA detection in women with very limited access to healthcare, while the utility of urine for HPV DNA detection in males is less certain.

The gene spectrum of thalassemia in Yangjiang of western Guangdong Province.

Background: Thalassemia presents a higher incidence in southern China. The objective of this study is to analyze the genotype distribution of thalassemia in Yangjiang, a western city of Guangdong Province in China. Methods: The genotypes of suspected cases with thalassemia were tested by PCR and reverse dot blot (RDB). Unidentified rare thalassemia genotypes of the samples were further ascertained by PCR and direct DNA sequencing. Results: Among 22467 suspected cases with thalassemia, 7658 cases were found with thalassemia genotypes using our PCR-RDB kit. Among these 7658 cases, 5313 cases were found with α-thalassemia (α-thal) alone, --SEA/αα was the most common genotype, accounting for 61.75% of α-thal genotypes, and the following mutations were found: α3.7/αα, -α4.2/αα, αCSα/αα, αWSα/αα, and αQSα/αα. A total of 2032 cases were found with ß-thalassemia (ß-thal) alone. ßCD41-42/ßN, ßIVS-II-654/ßN, and ß-28/ßN accounted for 80.9% of all ß-thal genotypes, and the following genotypes were found: ßCD17/ßN, ßCD71-72/ßN, and ßE/ßN. Compound heterozygotes of ß-thal and ß-thalassemia homozygotes were identified in 11 and five cases, respectively, in this study. α-thal combined with ß-thal was identified in 313 cases, showing 57 genotype combinations of the coincidence of both Hb disorders; one extreme patient had a genotype of --SEA/αWSα and ßCD41-42/ß-28. In addition, four rare α-mutations (--THAI, HKαα, Hb Q-Thailand, and CD31 AGG>AAG) and six rare ß-mutations (CD39 CAG>TAG, IVS-Ⅱ-2 (-T), -90(C>T), Chinese Gγ+(Aγδß)0, CD104 (-G), and CD19 A>G) were also found in this study population. Conclusion: This study provided detailed genotypes of thalassemia in Yangjiang of western Guangdong Province in China and reflected the complexity of genotypes in this high-prevalence region, and this would be valuable for diagnosis and counseling for thalassemia in this area.

Effect of Sintering Temperature and Solution Treatment on Phase Changes and Mechanical Properties of High-Nitrogen Stainless Steel Prepared by MIM.

High-nitrogen stainless steel (HNSS) has been widely concerned and studied owing to its excellent mechanical, corrosion resistance, and biocompatibility properties. A series of HNSS was prepared by metal injection molding (MIM) using gas atomized Fe-Cr-Mn-Mo-0.3 N duplex stainless steel powders. Both sintering and solution treatments were carried out in an N2 atmosphere. The effects of nitrogen distribution and phase transformation on the mechanical properties of MIM HNSS during sintering and solution were studied. The results show that as the sintering temperature increased, the sample density increased, but the total nitrogen content decreased. Nitrogen and Cr2N concentration gradients along the cross-section of as-sintered samples were formed after cooling. The high nitrogen content promotes the decomposition of γ: γsaturated translated to γ and Cr2N. Meanwhile, the low Mn content in austenite also decomposes γ: γ translated to α and Cr2N. After solution treatment, a single γ phase was obtained for samples sintered at 1200 to 1320 °C. For solution treatment samples sintered at 1320 and 1350 °C, their tensile strength was 988.76 and 1036.12 MPa; yield strength was 615.61 and 636.14 MPa, and elongation was 42.58 and 40.08%, respectively. These values vastly exceeded the published MIM HNSS values.

Development of an HPV Genotype Detection Platform Based on Aggregation-Induced Emission (AIE) and Flow-Through Hybridization Technologies.

Genetic mutations can cause life-threatening diseases such as cancers and sickle cell anemia. Gene detection is thus of importance for disease-risk prediction or early diagnosis and treatment. Apart from genetic defects, gene detection techniques can also be applied to gene-related diseases with high risk to human health such as human papillomavirus (HPV) infection. HPV infection has been strongly linked to cervical cancer. To achieve a high-throughput HPV gene detection platform, the flow-through hybridization system appears to be one of the commercialized diagnostic techniques for this purpose. The flow-through hybridization technique is based on a vacuum-guided flow of DNA fragments which is continuously directed toward the oligoprobes that are immobilized on the testing membrane. However, the conventional colorimetric method and signal read-out approach suffers a problem of low sensitivity. On the contrary, fluorescence approaches allow more sensitive detection and broad sensing ranges. In this work, a fluorescent dye HCAP, which possesses aggregation-induced emission (AIE) properties and is responsive to alkaline phosphatase, was developed and applied to the flow-through hybridization platform to achieve HPV genome diagnosis of clinical samples. Also, an automatic membrane reader was constructed based on the AIE-based diagnosis platform which can identify the diagnostic result of patient DNA with a total concordance rate of 100% in the clinical trial.

Influence of Hydrogen Reduction on the Properties of Porous High-Nitrogen Austenitic Stainless Steel.

This work explores the impact of hydrogen reduction on sintering and nitriding of porous high-nitrogen austenitic stainless steel (HNASS) processed via powder metallurgy. A temperature-resolved hydrogen reduction (temperature range of 700-1250 °C) was performed to evaluate the phase composition of porous HNASS. The systematic microstructure was characterized by a scanning electron microscope (SEM) with energy disperse spectroscopy (EDS), X-ray diffraction (XRD), and X-ray photoelectron spectroscopy (XPS). The compressive mechanical properties and electrochemical corrosion behavior of the unreduced and reduced samples were discussed. Samples reduced in hydrogen at 1100 °C and 1250 °C show better compressive properties while still retaining good corrosion resistance. Reduction of oxide facilitates sintering thus improves the compressive properties. Increasing the content of solute nitrogen and reducing the precipitation of nitride can effectively improve the corrosion resistance of porous HNASS.

Clinical validation of a single-tube PCR and reverse dot blot assay for detection of common α-thalassaemia and ß-thalassaemia in Chinese.

OBJECTIVE: To evaluate a novel reverse dot blot assay for the simultaneous detection six types of common α-thalassaemia alleles (three deletional and three common non-deletional mutations) and 19 types of common ß-thalassaemia alleles in a Chinese population. METHODS: Genomic DNA samples were collected from three hospitals in southern China. The novel thalassaemia gene assay involved one multiplex polymerase chain reaction amplification system and one round of hybridization. Each of the clinically validated DNA samples was re-tested using the new multiplex polymerase chain reaction/reverse dot blot assay II (M-PCR/RDB II) assay in a double-blind manner. RESULTS: A total of 1060 unrelated study participants, including 829 patients with thalassaemia and 231 healthy control subjects, were analysed. The whole PCR and RDB procedures were completed in 260 min. All the samples, including heterozygous thalassaemia, homozygous thalassaemia and compound heterozygous thalassaemia, were correctly genotyped, yielding 100% concordance with the reference assays. HKαα/--SEA and HKαα/-α4.2, which were not included in the detection panel, yielded a contradictory result with this new assay. CONCLUSION: The novel M-PCR/RDB II assay was simple, rapid and accurate, suggesting that it could be used for the genetic screening and clinical diagnosis of common α-thalassaemia and ß-thalassaemia variants in Chinese populations.

Reverse Transcription Recombinase-Aided Amplification Assay With Lateral Flow Dipstick Assay for Rapid Detection of 2019 Novel Coronavirus.

Background: The emerging Coronavirus Disease-2019 (COVID-19) has challenged the public health globally. With the increasing requirement of detection for SARS-CoV-2 outside of the laboratory setting, a rapid and precise Point of Care Test (POCT) is urgently needed. Methods: Targeting the nucleocapsid (N) gene of SARS-CoV-2, specific primers, and probes for reverse transcription recombinase-aided amplification coupled with lateral flow dipstick (RT-RAA/LFD) platform were designed. For specificity evaluation, it was tested with human coronaviruses, human influenza A virus, influenza B viruses, respiratory syncytial virus, and hepatitis B virus, respectively. For sensitivity assay, it was estimated by templates of recombinant plasmid and pseudovirus of SARS-CoV-2 RNA. For clinical assessment, 100 clinical samples (13 positive and 87 negatives for SARS-CoV-2) were tested via quantitative reverse transcription PCR (RT-qPCR) and RT-RAA/LFD, respectively. Results: The limit of detection was 1 copies/µl in RT-RAA/LFD assay, which could be conducted within 30 min at 39°C, without any cross-reaction with other human coronaviruses and clinical respiratory pathogens. Compared with RT-qPCR, the established POCT assay offered 100% specificity and 100% sensitivity in the detection of clinical samples. Conclusion: This work provides a convenient POCT tool for rapid screening, diagnosis, and monitoring of suspected patients in SARS-CoV-2 endemic areas.

Fabrication of Micro-Structured LED Diffusion Plate Using Efficient Micro Injection Molding and Micro-Ground Mold Core.

In this paper, a new style of micro-structured LED (light-emitting diode) diffusion plate was developed using a highly efficient and precise hybrid processing method combined with micro injection molding and micro-grinding technology to realize mass production and low-cost manufacturing of LED lamps with excellent lighting performance. Firstly, the micro-structured mold core with controllable shape accuracy and surface quality was machined by the precision trued V-tip grinding wheel. Then, the micro-structured LED diffusion plate was rapidly fabricated by the micro injection molding technology. Finally, the influences of micro injection molding process parameters on the illumination of the micro-structured diffusion plate were investigated. The simulated optical results show that the illumination of the micro-structured diffusion plate can achieve a maximum value when the V-groove depth and V-groove angle are designed to be 300 µm and 60°, respectively. The experimental results indicate that the developed micro-structured diffusion plate may improve the illumination by about 40.82% compared with the traditional diffusion plate. The prediction accuracy of the designed light efficiency simulation method was about 90.33%.

Efficient and Precise Grinding of Sapphire Glass Based on Dry Electrical Discharge Dressed Coarse Diamond Grinding Wheel.

In this paper, in view of low grinding efficiency and poor ground surface quality of sapphire glass, the coarse diamond grinding wheel dressed by dry impulse electrical discharge was proposed to perform efficient and precise grinding machining of sapphire glass. The dry electrical discharge dressing technology was employed to obtain high grain protrusion and sharp micro-grain cutting edges. The influences of grinding process parameters such as wheel speed, depth of cut and feed speed on the ground surface quality, grinding force and grinding force ratio on sapphire glass were investigated, and the relationship between grinding force and ground surface quality was also revealed. The experimental results show that the grain protrusion height on the surface of a coarse diamond grinding wheel dressed by dry electrical discharge can reach 168.5 µm. The minimum line roughness Ra and surface roughness Sa of ground sapphire glass surface were 0.194 µm and 0.736 µm, respectively. In order to achieve highly efficient ground quality of sapphire glass, the depth of cut was controlled within 7 µm, and the wheel speed and feed speed were 3000-5000 r/min and 10-20 mm/min, respectively. The influences of feed speed and wheel speed on grinding force ratio were more significant, but the influence of depth of cut was little.

Fabrication of Micro-Structured Polymer by Micro Injection Molding Based on Precise Micro-Ground Mold Core.

Precise micro-grinding machining was proposed to fabricate regular and controllable micro-grooved array structures on the surface of mold cores to realize the mass production and manufacturing of micro-structured polymer components by micro injection molding in this paper. First, the 3D topographies and section profiles of micro-ground mold cores and micro-formed polymers with different micro-structure parameters were presented. Then, the surface roughness of mold cores and polymers were compared. Next, the relationships between machining accuracy of mold core ground by micro-grinding and filling rates of micro-structured polymer formed by micro injection molding were investigated. Finally, the influences of micro injection molding parameters on the filling rate of micro-structures polymer were investigated. It is shown that the micro-structured polymer can be effectively and rapidly fabricated using the proposed method. The experimental results indicate the highest form accuracy of the micro-grooved mold core and the filling rate of micro-structured polymer can reach to 4.05 µm and 99.30%, respectively. It is found that the filling rate of the micro-structured polymer roughly increased with increasing machining accuracy of the mold core. The injection pressure had the greatest influence on the filling rate of the injection formed polymer, while the melt temperature had the least influence.

Aging Response and Precipitation Behavior after 5% Pre-Deformation of an Al-Mg-Si-Cu Alloy.

In this study, Al-1.00 Mg-0.65 Si-0.24 Cu alloy was solution heat-treated, water-quenched, and then pre-deformed for 5% before aging. The peak hardness and yield strength of the pre-deformed sample with subsequent artificial aging were similar to that of a T6 condition sample. It was also found that the pre-deformation treatment could inhibit the negative influence of natural aging to some degree. After seven days of natural aging, the pre-deformed sample obtained better peak hardness and yield strength upon artificial aging than the sample without pre-deformation. In addition, the pre-deformation treatment could reduce 50% of the artificial aging time to reach the peak aging condition compared with T6 treatment. For the peak aged condition in the pre-deformed sample, transmission electron microscopy (TEM) observation found two types of precipitates exhibited along the dislocations besides the ß″ precipitates in the Al matrix. Both precipitates had disordered atomic arrangements on the ordered subcell (Si network). The disordered precipitates occupied a number of Mg and Si atoms, resulting in less ß″ precipitates formed during artificial aging at 180 °C.

Microstructure and Properties of Porous High-N Ni-Free Austenitic Stainless Steel Fabricated by Powder Metallurgical Route.

Porous high-N Ni-free austenitic stainless steel was fabricated by a powder metallurgical route. The microstructure and properties of the prepared porous austenitic stainless steel were studied. Results reveal that the duplex stainless steel transforms into austenitic stainless steel after nitridation sintering for 2 h. The prepared high-N stainless steel consists of γ-Fe matrix and FCC structured CrN. Worm-shaped and granular-shaped CrN precipitates were observed in the prepared materials. The orientation relationship between CrN and austenite matrix is [011]CrN//[011]γ and (-1-11)CrN//(1-11)γ. Results show that the as-fabricated porous high-nitrogen austenitic stainless steel features a higher mechanical property than common stainless steel foam. Both compressive strength and Young’s modulus decrease with an increase in porosity. The 3D morphology of the prepared porous materials presents good pore connectivity. The prepared porous high-N Ni-free austenitic stainless steel has superior pore connectivity, a good combination of compressive strength and ductility, and low elastic modulus, which makes this porous high-N Ni-free austenitic stainless steel very attractive for metal foam applications.

[Rare thalassemia mutations among southern Chinese population].

OBJECTIVE: To detect rare types of thalassemia mutations among southern Chinese population. METHODS: Peripheral blood samples from 327 patients from various regions of southern China were collected. The patients were suspected as rare-type thalassemia for their inconsistency between hematological phenotypes and results of routine mutation screening. The samples were further analyzed with GAP-PCR and DNA sequencing. RESULTS: One hundred and eight cases were diagnosed as rare types of thalassemia. Among whom 10 rare α-globin gene mutations including --THAI, HKα, αααanti3.7, αααanti4.2, -α2.8, -α27.6, CD74 GAC>CAC (Hb Q-Thailand), CD30 (-GAG), CD31 AGG>AAG and CD118 (+TCA), and 12 rare ß-globin gene mutations including CD37 TGG>TAG, CD39 CAG>TAG/CD39 CAG>TAG, ß II-2 (-T), -90(C>T), -31(A>C), -88(C>T), CD7(-A), CD138(+T), CD89-93 (--AGTGAGCTGCACTG), CD54-58 (-TATGGGCAACCCT), Chinese G γ +(A γδß)0 and Vietnamese HPFH (HPFH-6) were identified. -88(C>T) (HBB: c.-138C>T) and CD39 CAG>TAG (HBB: c.118C>T) were discovered for the first time in Chinese population. CD7(-A) (HBB: c.23delA) and CD138(+T) (HBB: c.416_417insT) were new types of ß-globin gene mutations. CONCLUSION: The present study have enriched the mutation spectrum of thalassemia in southern China, which has provided necessary information for its diagnosis.

Clinical validation of a novel real-time human papillomavirus assay for simultaneous detection of 14 high-risk HPV type and genotyping HPV type 16 and 18 in China.

In the present study, we describe the laboratory workflow and the clinical validation of a novel multiplex real-time PCR-based HPV assay in China. The cross-sectional validation analysis showed that this assay worked well for detection of 14 HR-HPV types and identification of HPV 16 and 18 in a single sensitive assay that is suitable for both clinical usage and high-throughput cervical screening purposes. We predict that this accurate, high-throughput and low-cost HPV assay can greatly reduce the heavy economic burden of HPV detection in China.

Epidemiologic characterization of human papillomavirus infection in rural Chaozhou, eastern Guangdong Province of China.

BACKGROUND: Human papilloma virus (HPV) infection was the main cause of cervical cancer. There were only a few reports and detailed data about epidemiological research of HPV infection in rural population of China. MATERIALS AND METHODS: The cervical cells of rural Chaozhou women were collected, and multiplex real time PCR was firstly performed to detect high-risk HPV (HR-HPV) infection, which could detect 13 types of HR-HPV (types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, and 68). Then, HPV-positive samples were typed by HPV GenoArray test. RESULTS: HR-HPV DNA was detected by multiplex real time-PCR in 3830 of 48559 cases (7.89%). There was a peak incidence in age of 55-60 years group, and a lower incidence in who lived in plain group compared with suburban, mountain and seashore group. 3380 cases of HPV positive sample were genotyped, 11.01% (372/3380) cases could not be classified, among the typed 3008 cases, 101 cases were identified without HR-HPV type infection, 2907 cases were infected with one HR-HPV type at least, the 6 most common HR-HPV types in descending order of infection, were type 52 (33.4%, 16 (20.95%), 58 (15.93%), 33 (9.94%), 68 (9.22%) and 18 (8.36%). The combined prevalence of HPV types 16 and 18 accounted for 28.52% of total infection. However, type 52 plus 58 presented 48.23% of total infection. 2209/2907 cases were infected with a single HPV type and 698/2907 cases were infected with multiple types, and multiple infection constituent ratio increased with age, with a peak incidence in age 55-60 years group. CONCLUSIONS: Our findings showed low prevalence of HPV vaccine types (16 and 18) and relatively high prevalence of HPV-52 and -58, support the hypothesis that the second-generation HPV vaccines including HPV-52 and -58 may offer higher protection for women in rural Guangdong Province.

Genital human papillomavirus screening by gene chip in Chinese women of Guangdong province.

BACKGROUND: Human papillomavirus (HPV) infections are associated with cervical cancer. There were only a few reports and detailed data about epidemiological research of HPV infection in general population of China. AIMS: To determine the prevalence of genital HPV infection in Chinese women of Guangdong province. METHODS: A total of 1705 women were screened by gene chip. All HPV-positive women were further examined by ThinPrep liquid-based cytology test (TCT), and the cervical biopsies of those women with positive HPV-DNA and abnormal TCT were collected for pathological diagnosis. RESULTS: The overall HPV prevalence was 9.03% (154 of 1705), and 72.3% (126 of 154) of total positive samples were high-risk types, with higher prevalence of types 52, 58, 16, 18 and CP8304. For women aged 51 years or older, the overall high-risk HPV prevalence was 12.2% (24 of 179), which was obviously higher than those of other age groups (P < 0.05). CONCLUSIONS: Our results showed that the HPV prevalence in Guangdong is very similar to the world level. Unlike most previous studies, our findings suggest that HPV prevalence increased with age, and that the predominant genotypes in this area were HPV 52 and 58.