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Calcification of cartilage by hydroxyapatite is a hallmark of osteoarthritis and its deposition strongly correlates with the severity of osteoarthritis. However, no effective strategies are available to date on the prevention of hydroxyapatite deposition within the osteoarthritic cartilage and its role in the pathogenesis of this degenerative condition is still controversial. Therefore, the present work aims at uncovering the pathogenic mechanism of intra-cartilaginous hydroxyapatite in osteoarthritis and developing feasible strategies to counter its detrimental effects. With the use of in vitro and in vivo models of osteoarthritis, hydroxyapatite crystallites deposited in the cartilage are found to be phagocytized by resident chondrocytes and processed by the lysosomes of those cells. This results in lysosomal membrane permeabilization (LMP) and release of cathepsin B (CTSB) into the cytosol. The cytosolic CTSB, in turn, activates NOD-like receptor protein-3 (NLRP3) inflammasomes and subsequently instigates chondrocyte pyroptosis. Inhibition of LMP and CTSB in vivo are effective in managing the progression of osteoarthritis. The present work provides a conceptual therapeutic solution for the prevention of osteoarthritis via alleviation of lysosomal destabilization.

Streptococcus suis (S. suis) is a Gram-positive bacterium and the main culprit behind zoonotic outbreaks, posing a serious threat to public health. The prevalent strains in China are mainly of sequence types (ST) 1 and 7, with few cases of human infections caused by other sequence type being reported. This study presents the first isolation of a ST25 strain from the blood of a septicemic patient. A 57-year-old febrile patient was admitted to a hospital in Hainan of China, diagnosed as septicemia and hepatic dysfunction. A strain of S. suis was isolated from blood culture and confirmed to be serotype 2 and ST25 through 16S rRNA sequencing and whole-genome sequencing, and its genome was further analyzed for gene functions and presence of drug resistance genes. The full-length genome of strain HN28 spans 2,280,124 bp and encodes a total of 2291 proteins. Genes annotated in COG, GO, KEGG, CAZy, and PHl databases accounted for 75.38 %, 69.14 %, 55.35 %, 4.58 %, and 11.87 % of the total predicted proteins, respectively. Virulence factor analysis revealed the presence of seven putative virulence genes in strain HN28. Analysis using the CARD database identified 51 resistance genes in HN28, alongside abundant exocytosis systems. These findings underscore the occurrence of S. suis infections in humans caused by less common ST, emphasizing the need for enhanced epidemiological investigations and monitoring of S. suis infections in the human population.

Abstract Brain-derived extracellular vesicles participate in interorgan communication after traumatic brain injury by transporting pathogens to initiate secondary injury. Inflammasome-related proteins encapsulated in brain-derived extracellular vesicles can cross the blood‒brain barrier to reach distal tissues. These proteins initiate inflammatory dysfunction, such as neurogenic heterotopic ossification. This recurrent condition is highly debilitating to patients because of its relatively unknown pathogenesis and the lack of effective prophylactic intervention strategies. Accordingly, a rat model of neurogenic heterotopic ossification induced by combined traumatic brain injury and achillotenotomy was developed to address these two issues. Histological examination of the injured tendon revealed the coexistence of ectopic calcification and fibroblast pyroptosis. The relationships among brain-derived extracellular vesicles, fibroblast pyroptosis and ectopic calcification were further investigated in vitro and in vivo. Intravenous injection of the pyroptosis inhibitor Ac-YVAD-cmk reversed the development of neurogenic heterotopic ossification in vivo. The present work highlights the role of brain-derived extracellular vesicles in the pathogenesis of neurogenic heterotopic ossification and offers a potential strategy for preventing neurogenic heterotopic ossification after traumatic brain injury. Brain-derived extracellular vesicles (BEVs) are released after traumatic brain injury. These BEVs contain pathogens and participate in interorgan communication to initiate secondary injury in distal tissues. After achillotenotomy, the phagocytosis of BEVs by fibroblasts induces pyroptosis, which is a highly inflammatory form of lytic programmed cell death, in the injured tendon. Fibroblast pyroptosis leads to an increase in calcium and phosphorus concentrations and creates a microenvironment that promotes osteogenesis. Intravenous injection of the pyroptosis inhibitor Ac-YVAD-cmk suppressed fibroblast pyroptosis and effectively prevented the onset of heterotopic ossification after neuronal injury. The use of a pyroptosis inhibitor represents a potential strategy for the treatment of neurogenic heterotopic ossification.

Abstract Heterotopic ossification (HO) comprises the abnormal formation of ectopic bone in extraskeletal soft tissue. The factors that initiate HO remain elusive. Herein, we found that calcified apoptotic vesicles (apoVs) led to increased calcification and stiffness of tendon extracellular matrix (ECM), which initiated M2 macrophage polarization and HO progression. Specifically, single‐cell transcriptome analyses of different stages of HO revealed that calcified apoVs were primarily secreted by a PROCR+ fibroblast population. In addition, calcified apoVs enriched calcium by annexin channels, absorbed to collagen I via electrostatic interaction, and aggregated to produce calcifying nodules in the ECM, leading to tendon calcification and stiffening. More importantly, apoV‐releasing inhibition or macrophage deletion both successfully reversed HO development. Thus, we are the first to identify calcified apoVs from PROCR+ fibroblasts as the initiating factor of HO, and might serve as the therapeutic target for inhibiting pathological calcification.

Abstract Some biomolecules involved in pathological calcification have been identified. However, the exact mechanism in which this intricate process occurs remains unknown. Extracellular DNA (exDNA) has recently been recognized as a partaker in this ectopic phenomenon. Extracellular DNA acts as an intercellular messenger that transmits information and orchestrates complex inflammatory responses. Changes in the morphology or function of exDNA may trigger calcification under pathological conditions. In the present review, recent advances on how DNA is released into the extracellular milieu to become exDNA will be highlighted in conjunction with how exDNA directly and indirectly contributes to the progression of pathological calcification. Emphasis is placed on the “gluing” effect of neutrophil extracellular traps that act as a bridge between inflammation and pathological calcification. Manipulation of exDNA may open new vistas for the development of enterprising strategies that prevent or treat pathological calcification.

Abstract On October 23, 2020, a 69‐year‐old Chinese female patient was admitted to Yuncheng Hospital due to a history of postmenopausal bleeding and lower abdominal pain for 5 months. The HPV test and pathology results indicated the presence of independent HPV in primary serous carcinoma of the uterine cervix. The genetic testing identified variants of uncertain significance (PAX8 p.Tyr 410 Ter and TP53 p.Asn 247 Ile), microsatellite instability stable (MSI‐S), tumor mutational burden (TMB) 7.33Muts/Mb, and an elevated tumor neoantigen burden. Before undergoing radical hysterectomy treatment, the patient exhibited a positive response to three cycles of intravenous docetaxel (100 mg/3 h) and carboplatin (450 mg/1 h). Following the surgery, she received an additional three cycles of docetaxel (100 mg/3 h) and carboplatin (500 mg/1 h), accompanied by 25 cycles of radiation therapy (DT 46Gy/2Gy/23f). Concurrently, cisplatin (450 mg/1 h) was administered. As of now, the patient has achieved 20 months of disease‐free survival.

ABSTRACT Rarely has the vast diversity of bacteria on Earth been profiled, particularly on inaccessible plateaus. These uncultured microbes, which are also known as “microbial dark matter,” may play crucial roles in maintaining the ecosystem and are linked to human health, regarding pathogenicity and prebioticity. The plateau pika (Ochotona curzoniae) is a small burrowing steppe lagomorph that is endemic to the Qinghai-Tibetan Plateau and is a keystone species in the maintenance of ecological balance. We used a combination of full-length 16S rRNA amplicon sequencing, shotgun metagenomics, and metabolomics to elucidate the species-level community structure and the metabolic potential of the gut microbiota of the plateau pika. Using a full-length 16S rRNA metataxonomic approach, we clustered 618 (166 ± 35 per sample) operational phylogenetic units (OPUs) from 105 plateau pika samples and assigned them to 215 known species, 226 potentially new species, and 177 higher hierarchical taxa. Notably, 39 abundant OPUs (over 60% total relative abundance) are found in over 90% of the samples, thereby representing a “core microbiota.” They are all classified as novel microbial lineages, from the class to the species level. Using metagenomic reads, we independently assembled and binned 109 high-quality, species-level genome bins (SGBs). Then, a precise taxonomic assignment was performed to clarify the phylogenetic consistency of the SGBs and the 16S rRNA amplicons. Thus, the majority of the core microbes possess their genomes. SGBs belonging to the genus Treponema, the families Muribaculaceae, Lachnospiraceae, and Oscillospiraceae, and the order Eubacteriales are abundant in the metagenomic samples. In addition, multiple CAZymes are detected in these SGBs, indicating their efficient utilization of plant biomass. As the most widely connected metabolite with the core microbiota, tryptophan may relate to host environmental adaptation. Our investigation allows for a greater comprehension of the composition and functional capacity of the gut microbiota of the plateau pika. IMPORTANCE The great majority of microbial species remain uncultured, severely limiting their taxonomic characterization and biological understanding. The plateau pika (Ochotona curzoniae) is a small burrowing steppe lagomorph that is endemic to the Qinghai-Tibetan Plateau and is considered to be the keystone species in the maintenance of ecological stability. We comprehensively investigated the gut microbiota of the plateau pika via a multiomics endeavor. Combining full-length 16S rRNA metataxonomics, shotgun metagenomics, and metabolomics, we elucidated the species-level taxonomic assignment of the core uncultured intestinal microbiota of the plateau pika and revealed their correlation to host nutritional metabolism and adaptation. Our findings provide insights into the microbial diversity and biological significance of alpine animals.

Mussels can form tough and long-lasting adhesions to organic and inorganic surfaces in saline and impactive severe aquatic environments. Similar to mussel adhesion, dentin bonding occurs in a wet environment. However, unlike mussels, it is difficult to achieve long-lasting bonds with dentin. Moreover, water is considered a major hindrance in dentin bonding. Inspired by the synergistic effect of cationic lysine (Lys) and catechol on the elimination of the hydration layer during mussel adhesion, a catechol- and Lys-functionalized polymerizable polymer (catechol–Lys–methacrylate [CLM]) was synthesized to replicate the complex synergy between amino acids and catechol. The bond-promoting potential of 5 ​mg/mL CLM primer was confirmed using an in vitro wet dentin-bonding model, which was characterized by an improvement in bond strength and durability. CLM can adhere to wet demineralized dentin, with Lys acting as a molecular vanguard to expel water. Subsequently, a myriad of interfacial interactions can be obtained by introducing the catechol group into the interface. Additionally, tough and long-lasting adhesion, similar to that formed by mussels, can be achieved by grafting CLM onto type I collagen via covalent bonds, hydrogen bonds, Van der Waals interactions, and cation–π interactions, which can enhance the mechanical and chemical stability of collagen, increase the enzymatic resistance of collagen, and provide additional physical/chemical adhesion to dentin bonds. Catechol- and cationic Lys-functionalized polymers can improve the stability of the resin–dentin interface under wet conditions.

ObjectiveThis study aimed to investigate the clinical value of ultrasonography combined with tumor markers in the diagnosis and prediction of recurrence of borderline ovarian tumors (BOTs) and analyze the value of the combination of two different auxiliary examinations in the diagnosis and prediction of recurrence of BOTs.MethodsHere, 221 patients with BOTs confirmed by postoperative pathology were enrolled. Their clinical data, including the ultrasonography features, tumor markers, and clinicopathological data, were retrospectively analyzed.ResultsThe statistical data of the 221 cases with BOTs were as follows: 94 (42.5%) with left-sided lesions, 102 (46.2%) with right-sided lesions, and 25 (11.3%) with bilateral lesions. Moreover, 93 cases (42.1%) had a borderline serous tumor, 110 (49.8%) had a borderline mucinous tumor, 12 (5.4%) had a borderline serous mucinous tumor, 2 (0.9%) had a borderline endometrioid tumor, 1 (0.5%) had a borderline Brenner tumor, and 2 (0.9%) had a clear cell BOT. There were 104 cases (47.1%) with a tumor diameter of ≤10 cm and 117 cases (52.9%) with a tumor diameter of >10 cm as suggested by ultrasonography. There were 89 cases (40.3%) with septation, 44 (19.9%) with papilla, and 97 (43.9%) with blood flow as demonstrated by ultrasonography. Carbohydrate antigen 125 (CA 125) was elevated in 132 cases (59.7%), and CA 19-9 was elevated in 52 cases (23.5%).ConclusionIn general, BOTs are difficult to diagnose preoperatively and have a certain recurrence rate. Ultrasonography combined with CA 125 and CA 19-9 is significant for the preoperative diagnosis and selection of surgical modality for BOTs and could be used as a guideline to achieve good preoperative preparation and avoid secondary surgery.

Abstract The discovery of new viruses is important for predicting their potential threats to the health of humans and other animals. A novel picornavirus was identified from oral, throat, and anal swab samples collected from belugas (Delphinapterus leucas), from Dalian Sun Asia Tourism Holding Co., China, between January and December 2018, using a metagenomics approach. The genome of this novel PicoV-HMU-1 strain was 8197 nucleotides (nt) in length, with a open reading frame (from 1091 to 8074 nt) that encoded a polyprotein precursor of 2328 amino acids. Moreover, the genomic length and GC content of PicoV-HMU-1 were within the ranges found in other picornaviruses, and the genome organization was also similar. Nevertheless, PicoV-HMU-1 had a lower amino acid identity and distinct host species compared with other members of the Picornaviridae family. Phylogenetic trees were constructed based on the P1 and 3D amino acid sequences of PicoV-HMU-1 along with representative members of the Picornaviridae family, which showed that PicoV-HMU-1 was related to unclassified bat picornaviruses groups. These findings suggest that the PicoV-HMU-1 strain represents a potentially novel genus of picornavirus. These data can enhance our understanding of the picornavirus genetic diversity and evolution.

It is well known that the chitosan (CS) properties such as molecular weight (Mw) and deacetylation degree (DD) are crucial for siRNA delivery. In the present study, we have unintentionally found that the transfection medium has much more substantial effect, which has not been reported before. Specifically, the transfection efficiency was considerably increased in Hank’s Balanced Saline Solution (HBSS), with various Mw (30–150 kD) and DD (85–95%) combinations in different cell types. In addition, even though the CS/siRNA nanoparticles physical parameters such as diameter size and polydisperse index were disturbed by cell culture medium or serum, a relatively high knockdown efficiency could still be obtained in HBSS. Moreover, the highly efficient silencing in HBSS even did not require pre-mixing to formulate the CS/siRNA nanoparticles. In conclusion, our findings have confirmed a neglected but crucial important factor of transfection medium in the chitosan mediated siRNA delivery.

Abstract Background This study aimed to investigate the clinical and pathological characteristics, and the recurrence and prognostic factors of borderline ovarian tumors (BOTs). Methods The data of 286 patients admitted to hospital and followed up for more than ten months were analyzed retrospectively to study the clinicopathological characteristics and related factors of recurrence. Results The median age of the patients was 42.06 ± 14.97 years, and the duration of the follow-up ranged from 10–109 months. During the follow-up period, 40 patients had a recurrence. Of these patients, 36 were ≤ 40 years, and patients with premenopausal recurrence accounted for 20.5% (36/176). In patients undergoing conservative treatment or radical operations, the recurrence rates were 21.3% and 1.8%, respectively, and they were 13.4% (36/268) in patients at Federation International of Gynecology and Obstetrics (FIGO) stage I, and 22.2% (4/18) in patients at an advanced stage. Postoperative pathology revealed that 40 patients had micropapillary tumors, among whom ten patients (25%) had a recurrence, and 19 patients had complications with interstitial infiltration. Of these 19 patients, six had a recurrence (31.5%). Another 22 patients had complications with calcified sand bodies; among these, eight patients (36.4%) had a recurrence. All the differences were statistically significant (P

Astroviruses infect human and animals and cause diarrhea, fever, and vomiting. In severe cases, these infections may be fatal in infants and juvenile animals. Previous evidence showed that humans in contact with infected animals can develop serological responses to astroviruses. Mamastrovirus 11 is a species of Mamastrovirus and was first reported in 2018. It was detected in the fecal samples of a California sea lion. The genome sequence of its capsid protein (CP) was submitted to GenBank. However, the genome sequence of its non-structural protein region was not elucidated. In the present study, we characterized the genome sequences of the novel astroviruses AstroV-HMU-1 and AstroV-like-HMU-2. These were obtained from California sea lions (Zalophus californianus) and walruses (Odobenus rosmarus) presenting with loose stools. A phylogenetic analysis revealed that the CP of AstroV-HMU-1 closely clustered with Mamastrovirus 11 while its RNA-dependent RNA polymerase (RdRp) and serine protease (SP) were closely related to the mink astrovirus in the genus Mamastrovirus. The genome of AstroV-HMU-1 provided basic information regarding the NS protein regions of Mamastrovirus 11. Recombination analyses showed that the genomes of Z. californianus AstroV-HMU-1, VA2/human and the mink astrovirus may have recombined long ago. The NS of AstroV-like-HMU-2 segregated from the Astroviridae in the deep root of the phylogenetic tree and exhibited 36% amino acid identity with other mamastroviruses. Thus, AstroV-like-HMU-2 was proposed as a member of a new genus in the unclassified Astroviridae. The present study suggested that that the loose stools of pinnipeds may be the result of occasional infection by this novel astrovirus. This discovery provides a scientific basis for future investigations into other animal-borne infectious diseases.

Bilayer zirconia restoration is one of the most commonly used restorations in dental practice, but the high frequency of the cohesive/adhesive fracture of veneered porcelain is still a problem. This paper focuses on the development of nanostructured glass-zirconia to improve the interface stability of dental zirconia substrate and veneered porcelain. A novel SiO2-Li2O-Al2O3 (SLA) glass was prepared and infiltrated into the surface of fully sintered dental zirconia to obtain nanostructured glass-zirconia structure. The prepared glass-zirconia was analyzed with scanning electron microscopes (SEM), energy dispersive spectroscopy (EDS) and X-ray diffraction spectroscopy (XRD). The wettability, roughness and 3D morphology of zirconia were altered, and shear bonding strength (SBS) test demonstrated almost double increase in SBS values of the nanostructured glass-zirconia structure. The failure modes and microstructure characteristics also verified the improved interfacial stability. This investigation provides a promising method for enhancing the structural stability of bilayer zirconia restorations.

Abstract The present study evaluated the effect of different high-power-laser surface treatments on the bond strength between resin cement and disilicate ceramic. Lithium disilicate ceramic specimens with truncated cones shape were prepared and divided into 5 groups: HF (hydrofluoric acid-etching), Er:YAG laser + HF, Graphite + Er:YAG laser + HF, Nd:YAG laser + HF, and Graphite + Nd:YAG laser + HF. The treated ceramic surfaces were characterized with scanning electron microscopy and surface roughness measurement. Hourglasses-shaped ceramic- resin bond specimens were prepared, thermomechanically cycled and stressed to failure under tension. The results showed that for both the factors “laser” and “graphite”, statistically significant differences were observed (p Nd:YAG (p

The Old World vulture may carry and spread pathogens for emerging infections since they feed on the carcasses of dead animals and participate in the sky burials of humans, some of whom have died from communicable diseases. Therefore, we studied the precise fecal microbiome of the Old World vulture with metataxonomics, integrating the high-throughput sequencing of almost full-length small subunit ribosomal RNA (16S rRNA) gene amplicons in tandem with the operational phylogenetic unit (OPU) analysis strategy. Nine vultures of three species were sampled using rectal swabs on the Qinghai-Tibet Plateau, China. Using the Pacific Biosciences sequencing platform, we obtained 54 135 high-quality reads of 16S rRNA amplicons with an average of 1442±6.9 bp in length and 6015±1058 reads per vulture. Those sequences were classified into 314 OPUs, including 102 known species, 50 yet to be described species and 161 unknown new lineages of uncultured representatives. Forty-five species have been reported to be responsible for human outbreaks or infections, and 23 yet to be described species belong to genera that include pathogenic species. Only six species were common to all vultures. Clostridium perfringens was the most abundant and present in all vultures, accounting for 30.8% of the total reads. Therefore, using the new technology, we found that vultures are an important reservoir for C. perfringens as evidenced by the isolation of 107 strains encoding for virulence genes, representing 45 sequence types. Our study suggests that the soil-related C. perfringens and other pathogens could have a reservoir in vultures and other animals.Emerging Microbes & Infections (2017) 6, e9; doi:10.1038/emi.2016.137; published online 22 February 2017

Parvoviruses are a diverse group of viruses that are capable of infecting a wide range of animals. In this study, we report the discovery of a novel parvovirus, tilapia parvovirus HMU-HKU, in the fecal samples of crocodiles and intestines of tilapia in Hainan Province, China. The novel parvovirus was firstly identified from crocodiles fed with tilapia using next-generation sequencing (NGS). Screening studies revealed that the prevalence of the novel parvovirus in crocodile feces samples fed on tilapia (75–86%) was apparently higher than that in crocodiles fed with chicken (4%). Further studies revealed that the prevalence of the novel parvovirus in tilapia feces samples collected at four areas in Hainan Province was between 40 and 90%. Four stains of the novel parvovirus were identified in this study based on sequence analyses of NS1 and all the four strains were found in tilapia in contrast only two of them were detected in crocodile feces. The nearly full-length genome sequence of the tilapia parvovirus HMU-HKU-1 was determined and showed less than 45.50 and 40.38% amino acid identity with other members of Parvoviridae in NS1 and VP1 genes, respectively. Phylogenetic analysis based on the complete helicase domain amino acid sequences showed that the tilapia parvovirus HMU-HKU-1 formed a relatively independent branch in the newly proposed genus Chaphamaparvovirus in the subfamily Hamaparvovirinae according to the ICTV’s most recent taxonomic criteria for Parvoviridae classification. Tilapia parvovirus HMU-HKU-1 likely represented a new species within the new genus Chaphamaparvovirus. The identification of tilapia parvovirus HMU-HKU provides further insight into the viral and genetic diversity of parvoviruses and its infections in tilapia populations need to be evaluated in terms of pathogenicity and production losses in tilapia farming.

Candida albicans is an opportunistic pathogenic yeast that predominantly causes invasive candidiasis. The conventional diagnosis of C. albicans infection depends on time-consuming, culture-based gold-standard methods. Here, a multiple cross displacement amplification (MCDA) assay, combined with a gold nanoparticle-based lateral flow biosensor (LFB) visualization method, was developed for the rapid detection of C. albicans. The internal transcribed spacer II, a region between 5.8 and 28 S fungal ribosomal DNA, is a C. albicans species-specific sequence that was used as the MCDA assay target. As an isothermal amplification method, the MCDA reaction with optimized conditions could be completed within only 40 min at a constant temperature (64°C). Then, the amplification reaction products could be visibly detected by a LFB without special equipment. The developed MCDA-LFB assay for C. albicans detection was a specific and accurate method, and could distinguish C. albicans from other pathogens. Just 200 fg of genomic DNA template from pure cultures of C. albicans could be detected using the MCDA-LFB method. The limit of detection (LOD) of the new method was more sensitive than that of both qPCR and loop-mediated isothermal amplification (LAMP). Of 240 clinical sputum samples, all of the C. albicans-positive (87/240) samples identified by the gold-standard method were successfully detected by the MCDA-LFB assay. Moreover, the true positive rate of the newly developed assay was not only higher than that of qPCR (100 vs. 86.2%), but also higher than that of LAMP (100 vs. 94.3%). Thus, the MCDA-LFB assay might be a simple, specific, and sensitive method for the rapid diagnosis of C. albicans in clinical samples.

Klebsiella pneumoniae (K. pneumoniae) is a frequent pathogen causing nosocomial infections and outbreaks. We developed a multiple cross displacement amplification (MCDA) assay for the detection of K. pneumoniae, which can get the positive results within 40 minutes' isothermal amplification. Gold-nanoparticle lateral flow biosensor (LFB) and colorimetric indicators were used for the rapid readouts of MCDA amplification. The detection limit of this assay was 100 fg per reaction at 65°C, which was confirmed to be the optimal amplification temperature according to the real time turbidimeters. For specificity, all of the 30 clinical-source K. pneumoniae strains were positive for the MCDA, and all of the non-K. pneumoniae strains belonging to 31 different species were negative for this MCDA assay. To evaluate the practical applicability of this method, we assessed its detection limit for K. pneumoniae strains in sputum samples (24 CFU per reaction), and DNA templates of 100 sputum samples further underwent the MCDA-LFB tests. All of the sputum samples being positive for K. pneumoniae (30/100) with the culture method were successfully identified with the MCDA assay, the detection power of which was higher than that of polymerase chain reaction (PCR) (25/100). Thus, the MCDA test for K. pneumoniae combined with the gold nanoparticle LFB as the results readout scheme, are simple, specific, and sensitive methods for the rapid diagnosis of K. pneumoniae in clinical samples.

[Objectives] This study was conducted to investigate the molecular characteristics, genetic structure and evolutionary mechanism of cucurbit aphid-borne yellows virus (CABYV), which is of great significance for clarifying the epidemic laws of the disease in the field and formulating long-term sustainable control strategies. [Methods] 120 melon leaves suspected to be infected with CABYV were randomly collected from Ak.su City, Xinjiang. RT-PCR was used to detect the virus. MEGA 7.0 was used to construct a phylogenetic tree. DnaSP v5.10 was used to analyze the genetic diversity among different sequences, and RDP v.4.31 software was used to analyze possible recombination events of CABYV sequence. [Results] lire detection rate of CABYV was 31.67%. A new CABYV-2 was isolated, sequenced and cloned from these positive samples. The length of the genome is 5 497 bp, encoding six open reading frames. Compared with 23 CABYV strains isolated from different countries in NCBI database, CABYV-2 shared the highest homology with KR231942.1 and the lowest homology with JF939812.1. The results of evolutionär}' tree analysis showed that the 24 isolates were divided into two branches due to different geographical factors, including CABYV-2 and KR231942 1, which were clustered together and belonged to branch 1. The results of genetic diversity and neutrality tests showed that CABYV was highly variable and the population was expanding. The results of recombination analysis showed that there were two recombinants, EU636992.1 and KR231949.1, which exacerbated the variation of CABYV. [Conclusions]The CABYV isolate from Aksu City, Xinjiang had the closest relationship with the Korean isolate KR231942.1, and the farthest relationship with the isolates from European countries. There were large genetic differences between the isolates of branch 1 and branch 2, and the recombination of CABYV in different hosts aggravated the variation of CABYV. Recombination and negative selection may be important reasons for the genetic variation of CABYV. [ABSTRACT FROM AUTHOR]

Trench fever, caused by Bartonella quintana, is recognized as a re-emerging and neglected disease. Rapid and sensitive detection approaches are urgently required to monitor and help control B. quintana infections. Here, loop-mediated isothermal amplification (LAMP), which amplifies target DNA at a fixed temperature with high sensitivity, specificity and rapidity, was employed to detect B. quintana. Thirty-six strains, including 10 B. quintana, 13 other Bartonella spp., and 13 other common pathogens, were applied to verify and evaluate the LAMP assay. The specificity of the LAMP assay was 100%, and the limit of detection was 125 fg/reaction. The LAMP assay was compared with qPCR in the examination of 100 rhesus and 20 rhesus-feeder blood samples; the diagnostic accuracy was found to be 100% when LAMP was compared to qPCR, but the LAMP assay was significantly more sensitive (p < 0.05). Thus, LAMP methodology is a useful for diagnosis of trench fever in humans and primates, especially in low-resource settings, because of its rapid, sensitive detection that does not require sophisticated equipment. [ABSTRACT FROM AUTHOR]

This clinical report describes treatment of a patient with gingival pigmentation after the restoration of the right maxillary canine and first premolar with Ni-Crebased metal ceramic crowns and investigation of mechanisms of gingival discoloration. Histopathological observation and energy dispersive x-ray spectroscopy analysis of the retrieved pigmented gingiva revealed brown deposits both in the epithelial cells and along the basement membranes but no trace of Ni or Cr elements in the pigmented particles, indicating that the gingival pigmentation adjacent to the Ni-Crebased metal ceramic prosthesis may not be caused by direct deposition of the released ions. [ABSTRACT FROM AUTHOR]

Shigella and Salmonella are frequently isolated from various food samples and can cause human gastroenteritis. Here, a novel multiple endonuclease restriction real-time loop-mediated isothermal amplification technology (MERT-LAMP) were successfully established and validated for simultaneous detection of Shigella strains and Salmonella strains in only a single reaction. Two sets of MERT-LAMP primers for 2 kinds of pathogens were designed from ipaH gene of Shigella spp. and invA gene of Salmonella spp., respectively. Under the constant condition at 63°C, the positive results were yielded in as short as 12 min with the genomic DNA extracted from the 19 Shigella strains and 14 Salmonella strains, and the target pathogens present in a sample could be simultaneously identified based on distinct fluorescence curves in real-time format. Accordingly, the multiplex detection assay significantly reduced effort, materials and reagents used, and amplification and differentiation were conducted at the same time, obviating the use of postdetection procedures. The analytical sensitivity of MERT-LAMP was found to be 62.5 and 125 fg DNA/reaction with genomic templates of Shigella strains and Salmonella strains, which was consist with normal LAMP assay, and at least 10- and 100-fold more sensitive than that of qPCR and conventional PCR approaches. The limit of detection of MERT-LAMP for Shigella strains and Salmonella strains detection in artificially contaminated milk samples was 5.8 and 6.4 CFU per vessel. In conclusion, the MERT-LAMP methodology described here demonstrated a potential and valuable means for simultaneous screening of Shigella and Salmonella in a wide variety of samples. [ABSTRACT FROM AUTHOR]

Objectives: To evaluate the effectiveness of TRUShape13D Conforming Files, compared with Twisted Files, in reducing bacteria load from root canal walls, in the presence or absence of irrigant agitation. Methods: Extracted human premolars with single oval-shaped canals were infected with Enterococcus faecalis. Teeth in Group I (N = 10; NaOCl and QMix1 2in1 as respective initial and final irrigants) were subdivided into 4 subgroups: (A) TRUShape1 instrumentation without irrigant activation; (B) TRUShape1 instrumentation with sonic irrigant agitation; (C) Twisted Files without irrigant agitation; (D) Twisted Files with sonic irrigant agitation. To remove confounding factor (antimicrobial irrigants), teeth in Group II (N = 10) were irrigated with sterile saline, using the same subgroup designations. Specimens before and after chemomechanical debridement were cultured for quantification of colony-forming units (CFUs). Data from each group were analyzed separately using two-factor ANOVA and Holm-Sidak multiple comparison (α = 0.05). Canal wall bacteria were qualitatively examined using scanning electron microscopy (SEM) and light microscopy of Taylor-modified Brown and Brenn-stained demineralised sections. Results: CFUs from subgroups in Group I were not significantly different (P = 0.935). For Group II, both file type (P < 0.001) and irrigant agitation (P < 0.001) significantly affected log-reduction in CFU concentrations. The interaction of these two factors was not significant (P = 0.601). Although SEM showed reduced canal wall bacteria, bacteria were present within dentinal tubules after rotary instrumentation, as revealed by light microscopy of longitudinal root sections. Conclusions: TRUShape1 files removed significantly more canal wall bacteria than Twisted Files when used without an antibacterial irrigant; the latter is required to decontaminate dentinal tubules. Clinical significance: Root canal disinfection should not be focused only on a mechanistic approach. Rather, the rational choice of a rotary instrumentation system should be combined with the use of well-tested antimicrobial irrigants and delivery/agitation techniques to establish a clinically realistic chemomechanical debridement protocol. [ABSTRACT FROM AUTHOR]