Identification of SARS-CoV-2 variants using viral sequencing for the Centers for Disease Control and Prevention genomic surveillance program.

BACKGROUND: The Centers for Disease Control and Prevention contracted with laboratories to sequence the SARS-CoV-2 genome from positive samples across the United States to enable public health officials to investigate the impact of variants on disease severity as well as the effectiveness of vaccines and treatment. Herein we present the initial results correlating RT-PCR quality control metrics with sample collection and sequencing methods from full SARS-CoV-2 viral genomic sequencing of 24,441 positive patient samples between April and June 2021. METHODS: RT-PCR confirmed (N Gene Ct value < 30) positive patient samples, with nucleic acid extracted from saliva, nasopharyngeal and oropharyngeal swabs were selected for viral whole genome SARS-CoV-2 sequencing. Sequencing was performed using Illumina COVIDSeq™ protocol on either the NextSeq550 or NovaSeq6000 systems. Informatic variant calling, and lineage analysis were performed using DRAGEN COVID Lineage applications on Illumina's Basespace cloud analytical system. All sequence data and variant calls were uploaded to NCBI and GISAID. RESULTS: An association was observed between higher sequencing coverage, quality, and samples with a lower Ct value, with < 27 being optimal, across both sequencing platforms and sample collection methods. Both nasopharyngeal swabs and saliva samples were found to be optimal samples of choice for SARS-CoV-2 surveillance sequencing studies, both in terms of strain identification and sequencing depth of coverage, with NovaSeq 6000 providing higher coverage than the NextSeq 550. The most frequent variants identified were the B.1.617.2 Delta (India) and P.1 Gamma (Brazil) variants in the samples sequenced between April 2021 and June 2021. At the time of submission, the most common variant > 99% of positives sequenced was Omicron. CONCLUSION: These initial analyses highlight the importance of sequencing platform, sample collection methods, and RT-PCR Ct values in guiding surveillance efforts. These surveillance studies evaluating genetic changes of SARS-CoV-2 have been identified as critical by the CDC that can affect many aspects of public health including transmission, disease severity, diagnostics, therapeutics, and vaccines.

The outcome of decompression alone for lumbar spinal stenosis with degenerative spondylolisthesis.

PURPOSE: Lumbar spinal stenosis in the presence of degenerative spondylolisthesis is generally treated by means of surgery. The role of lumbar decompression without fusion is not clear. Therefore, the aim of this study was to assess whether patients who undergo decompression alone have a favourable outcome without the need for a subsequent fusion. METHODS: This is a prospective cohort study with single blinding of 83 consecutive patients with lumbar stenosis and degenerative spondylolisthesis treated by decompression, without fusion, using a spinous process osteotomy. Blinded observers collected pre- and post-operative Oswestry Disability Index (ODI), EuroQol Five Dimensions (EQ-5D), and visual analogue scale (VAS) for back and leg pain scores prospectively. Failures for this study were those patients who required a subsequent lumbar fusion procedure at the decompressed levels. Statistical analysis was performed using paired t test and Mann-Whitney test. RESULTS: There were 36 males and 47 females with a mean age of 66 years (range 35-82). The mean follow-up was 36 months (range 19-48 months). The mean pre-operative ODI, EQ-5D, and VAS scores were 52 [standard deviation (SD) 18], 0.25 (SD 0.30), and 61 (SD 22), respectively. All mean scores improved post-operatively to 38 (SD 23), 0.54 (SD 0.34) and 36 (SD 27), respectively. There was a statistically significant improvement in all scores (p ≤ 0.0001). Nine patients (11 %) required a subsequent fusion procedure and five patients (6 %) required revision decompression surgery alone. CONCLUSION: Our study's results show that a lumbar decompression procedure without arthrodesis in a consecutive cohort of patients with lumbar spinal stenosis with degenerative spondylolisthesis had a significant post-operative improvement in ODI, EQ-5D, and VAS. The rate of post-operative instability and subsequent fusion is not high. Only one in 10 patients in this group ended up needing a subsequent fusion at a mean follow-up of 36 months, indicating that fusion is not always necessary in these patients.

Spinal Motocross Injuries in the United Kingdom.

BACKGROUND: Motocross is a form of motorcycle racing held on established off-road circuits and has been a recreational and competitive sport across the world for >100 years. In the United Kingdom alone, motocross has grown into a phenomenally ambitious and popular franchise. There are >200 motocross clubs across the country, permitting >900 events annually. PURPOSE: To assess the current trend of spine-related motocross injuries over the past 5 years. STUDY DESIGN: Descriptive epidemiology study. METHODS: Data were prospectively collected over 5 years (August 2010-August 2015) at our regional trauma and spine unit, regardless of whether the rider was performing the sport competitively or recreationally. RESULTS: During the study period, spine-related injuries were identified for 174 patients (age range, 6-75 years) who were directly referred to our department following recreational or competitive motocross, with most injuries being sustained within the early spring and summer months, representing the start of the motocross season. A significant number of injuries were in males (n = 203, 94%), with the majority of injuries occurring within the 21- to 30-year-old age group. A total of 116 (54%) injuries required operative treatment. The most common spinal injury was thoracolumbar burst fracture (n = 95), followed by chance fractures (n = 26). CONCLUSION: This data series emphasizes the prevalence and devastation of motocross-related spinal injuries in the United Kingdom and may serve in administering sanctions and guidelines to governing bodies of motocross. The spinal injuries that occur during motocross have significant capital connotations for regional spinal centers. The recent surge in motocross popularity is correlated with the number of injuries, which have increased over the past 5 years by almost 500%.

In vitro and in vivo optimization of impaction allografting by demineralization and addition of rh-OP-1.

Impaction allografting is a bone tissue engineering technique currently used in lower limb reconstruction orthopedic surgery. Our hypothesis was that biological optimization can be achieved by demineralization and addition of osteogenic protein-1 (OP-1) to the allograft. The objective of our in vitro study was to evaluate human mesenchymal stem cell (MSC) proliferation (Alamar Blue assay, titrated thymidine assay, total DNA Hoechst 33258, and scanning electron microscopy) and osteogenic differentiation (alkaline phosphatase assay) in two types of impacted carrier, namely, demineralized bone matrix (DBM) and insoluble collagenous bone matrix (ICBM), with or without OP-1. The objective in vivo was to compare the osteogenic potential of impacted DBM with or without OP-1, with that of impacted fresh frozen allograft (FFA), again with or without OP-1. DBM + OP-1 optimized osteoinduction and significantly improved (p < 0.05) proliferation and differentiation in comparison to the majority of all other graft preparation in vitro. In addition, DBM + OP-1 was significantly superior, with regard to osteogenesis, compared to the impacted FFA alone (p < 0.001), FFA + OP-1 (p = 0.01) and DBM alone (p = 0.02) in vivo. We propose that partial demineralization and addition of OP-1 provides a good method for improving the osteoinductive properties of fresh allograft currently used in the impaction grafting technique.

The potential adverse effects of aromatase inhibitors on wound healing: in vitro and in vivo evidence.

BACKGROUND: Estrogens and several other endogenous substances are recognised as being important in the process of wound healing. However, the effect of aromatase and aromatase inhibition in the wound healing process has yet to be fully defined. OBJECTIVE: A review of the in vitro and in vivo evidence on the effect of aromatase inhibition on wound healing. METHODS: The primary medical search engines used for the study were Ovid MEDLINE (1950 - March 2009) and EMBASE (1980 - March 2009) databases. RESULTS/CONCLUSION: The delayed healing of cutaneous wounds in aged individuals may in part reflect the decline in circulating levels of dehydroepiandrosterone (DHEA) and estrogens. The beneficial response on wound healing that DHEA and estrogen exert may be blocked by aromatase inhibition. Based on animal models, aromatase inhibitors may adversely affect cutaneous wound healing in the acute setting. So far, there have been no clinical trials investigating the adverse affect of aromatase inhibitors on the process of cutaneous wound healing in humans. Postmenopausal patients who take aromatase inhibitors as an adjunct to breast cancer therapy may, therefore, be at increased risk of delayed wound healing. Further studies are necessary to assess the extent of the effects on the wound healing process.

In vitro proliferation and differentiation of human mesenchymal stem cells on hydroxyapatite versus human demineralised bone matrix with and without osteogenic protein-1.

BACKGROUND: Allografting is currently used in lower limb reconstruction surgery. Demineralised bone matrix (DBM) is more osteoinductive compared with allografts but lacks mechanical strength. Osteogenic protein-1 (OP-1) can improve the osteoinductivity of the allograft, however recent reports indicate significant allograft resorption when it is combined with OP-1. OBJECTIVES: Our hypothesis was that hydroxyapatite (HA) with human-mesenchymal stem cells (h-MSCs) and OP-1 (HA+h-MSCs+OP-1) has similar osteoinductive properties to human-DBM+h-MSCs. The objective was to evaluate h-MSC proliferation (by tritiated thymidine incorporation, total DNA Hoechst 33258 and scanning electron microscopy) and osteogenic differentiation (from alkaline phosphatase activity) in human demineralised bone matrix (h-DBM) and HA, with or without OP-1. RESULTS: H-MSC proliferation on HA+OP-1 was significantly higher compared with that on HA at all time points (p < 0.05) and compared with DBM alone [day 1, (198.4 vs 95.4) p = 0.042; day 14 (286.1 vs 119.9), p < 0.001]. H-MSC proliferation was higher in DBM+OP-1, at all time points compared with HA+OP-1 but the difference was not statistically significant (p > 0.05). H-MSC differentiation was significantly higher in HA+OP-1 compared with HA (p < 0.05) but not significantly different from diffferentiation on DBM alone (p > 0.05). Differentiation was significantly higher on DBM+OP-1 at all time points compared with HA (p < 0.05) and with HA+OP-1 [day 1, (21.1 vs 10.1) (p = 0.03); day 7 (39.4 vs 7.1) (p < 0.01); day 14 (40.2 vs 14.4) (p < 0.001)]. CONCLUSIONS: When HA+h-MSCs is combined with OP-1 in vitro its osteogenic potential is similar to that of DBM+h-MSCs alone which may be adequate for non-weight-bearing applications. Mechanical testing however is of great importance for weight-bearing applications and the in vivo testing of the composite graft HA+h-MSCs+OP-1 vs DBM+h-MSCs is recommended.

Enhancing the osteoinductive properties of hydroxyapatite by the addition of human mesenchymal stem cells, and recombinant human osteogenic protein-1 (BMP-7) in vitro.

Hydroxyapatite (HA) has been widely used as a bone graft substitute. In this study, we investigated whether the addition of osteogenic protein-1 (OP-1) further enhanced the weak osteoinductive properties of hydroxyapatite when loaded with human mesenchymal stem cells (h-MSCs). Over a 14 day period, cell proliferation in both groups was assessed qualitatively using SEM and quantitatively using alamar blue assay. Cell differentiation was also evaluated by measurement of ALP activity, which was expressed against total DNA. HA/MSC loaded with OP-1 demonstrated a statistically significant increase (p<0.001) in cell proliferation at all time points in comparison to unloaded samples. ALP activity per DNA was also significantly enhanced (p<0.001) in loaded samples when compared to unloaded controls. SEM demonstrated increased cellular attachment and proliferation into HA pores at all time points in the loaded samples. Our study suggests that the osteoinductive potential of HA can be improved in vitro by the combined incorporation of MSCs and OP-1.