A multiplexed, confinable CRISPR/Cas9 gene drive can propagate in caged Aedes aegypti populations.

Aedes aegypti is the main vector of several major pathogens including dengue, Zika and chikungunya viruses. Classical mosquito control strategies utilizing insecticides are threatened by rising resistance. This has stimulated interest in new genetic systems such as gene drivesHere, we test the regulatory sequences from the Ae. aegypti benign gonial cell neoplasm (bgcn) homolog to express Cas9 and a separate multiplexing sgRNA-expressing cassette inserted into the Ae. aegypti kynurenine 3-monooxygenase (kmo) gene. When combined, these two elements provide highly effective germline cutting at the kmo locus and act as a gene drive. Our target genetic element drives through a cage trial population such that carrier frequency of the element increases from 50% to up to 89% of the population despite significant fitness costs to kmo insertions. Deep sequencing suggests that the multiplexing design could mitigate resistance allele formation in our gene drive system.

Quantitative intravital imaging for real-time monitoring of pancreatic tumor cell hypoxia and stroma in an orthotopic mouse model.

Pancreatic cancer is a lethal disease with few successful treatment options. Recent evidence demonstrates that tumor hypoxia promotes pancreatic tumor invasion, metastasis, and therapy resistance. However, little is known about the complex relationship between hypoxia and the pancreatic tumor microenvironment (TME). In this study, we developed a novel intravital fluorescence microscopy platform with an orthotopic mouse model of pancreatic cancer to study tumor cell hypoxia within the TME in vivo, at cellular resolution, over time. Using a fluorescent BxPC3-DsRed tumor cell line with a hypoxia-response element (HRE)/green fluorescent protein (GFP) reporter, we showed that HRE/GFP is a reliable biomarker of pancreatic tumor hypoxia, responding dynamically and reversibly to changing oxygen concentrations within the TME. We also characterized the spatial relationships between tumor hypoxia, microvasculature, and tumor-associated collagen structures using in vivo second harmonic generation microscopy. This quantitative multimodal imaging platform enables the unprecedented study of hypoxia within the pancreatic TME in vivo.

Assessing the Accuracy of Bioluminescence Image-Guided Stereotactic Body Radiation Therapy of Orthotopic Pancreatic Tumors Using a Small Animal Irradiator.

Stereotactic body radiation therapy (SBRT) has shown promising results in the treatment of pancreatic cancer and other solid tumors. However, wide adoption of SBRT remains limited largely due to uncertainty about the treatment's optimal fractionation schedules to elicit maximal tumor response while limiting the dose to adjacent structures. A small animal irradiator in combination with a clinically relevant oncological animal model could address these questions. Accurate delivery of X rays to animal tumors may be hampered by suboptimal image-guided targeting of the X-ray beam in vivo. Integration of bioluminescence imaging (BLI) into small animal irradiators in addition to standard cone-beam computed tomography (CBCT) imaging improves target identification and high-precision therapy delivery to deep tumors with poor soft tissue contrast, such as pancreatic tumors. Using bioluminescent BxPC3 pancreatic adenocarcinoma human cells grown orthotopically in mice, we examined the performance of a small animal irradiator equipped with both CBCT and BLI in delivering targeted, hypo-fractionated, multi-beam SBRT. Its targeting accuracy was compared with magnetic resonance imaging (MRI)-guided targeting based on co-registration between CBCT and corresponding sequential magnetic resonance scans, which offer greater soft tissue contrast compared with CT alone. Evaluation of our platform's BLI-guided targeting accuracy was performed by quantifying in vivo changes in bioluminescence signal after treatment as well as staining of ex vivo tissues with γH2AX, Ki67, TUNEL, CD31 and CD11b to assess SBRT treatment effects. Using our platform, we found that BLI-guided SBRT enabled more accurate delivery of X rays to the tumor resulting in greater cancer cell DNA damage and proliferation inhibition compared with MRI-guided SBRT. Furthermore, BLI-guided SBRT allowed higher animal throughput and was more cost effective to use in the preclinical setting than MRI-guided SBRT. Taken together, our preclinical platform could be employed in translational research of SBRT of pancreatic cancer.

Clinical Indicators of Postoperative Bleeding in Bariatric Surgery.

The purpose of this study was to evaluate the relationship between patient, clinical and laboratory characteristics, and bleeding after bariatric surgery. A retrospective chart review was performed for all patients, who underwent a laparoscopic Roux-en-Y gastric bypass and laparoscopic sleeve gastrectomy at our institution between March 2012 and May 2014. In total, 788 patients were included in this study. Seventeen patients had postoperative bleeding. The demographics, comorbidities, and preoperative medications were similar between the groups. Mean postoperative hemoglobin in the bleeding group was significantly lower (94 vs. 126; P<0.001) with a larger decrease from the baseline value (-43 vs. -12; P<0.001). The mean heart rate (91 vs. 81; P<0.001) and its increase from baseline (12 vs. -0.01; P<0.001) were significantly different in the bleeding patients. Postoperative hemoglobin and heart rate were associated with bleeding but not systolic blood pressure or patient characteristics. Further research is needed to develop a robust predictive model.

Neonatal septicaemia, bacterial isolates and antibiogram sensitivity in Maiduguri North-Eastern Nigeria.

AIMS AND OBJECTIVES: The aim and objective of the study was to determine the incidence, bacterial isolates and the antibiogram sensitivity of the isolates in neonates with septicaemia. PATIENTS AND METHODS: The neonates with clinical diagnosis of neonatal septicaemia (NNS) were consecutively enrolled into our special care baby unit. The patients were investigated including blood cultures, cerebrospinal fluid cultures and urine among others. Data were analysed with Statistical Package for Social Sciences software version 16.0 (SPSS Inc., Chicago, IL, USA). RESULTS: Forty-six neonates (42.0%) had a positive blood culture, while 64 (58.0%) were blood culture-negative. Eighteen (39.1%) of those septicaemic neonates with positive blood culture were inborn, while 28 (60.9%) were outborn. The incidence of NNS was 5.9/1000 live births. The male-to-female ratio among septicaemic neonates was 1.9:1. The common risk factors for NNS were prolonged rupture of membrane, prematurity and low socioeconomic status of parents among others. Common clinical features were fever, poor feeding, excessive crying, tachypnoea and hepatomegaly. Staphylococcus aureus 16 (69.6%) and Streptococcus pyogenes 5 (21.8%) were the predominant Gram-positive organisms isolated whereas Escherichia coli 9 (39.1%) and Klebsiella pneumoniae 7 (30.4%) were the predominant Gram-negative organisms isolated. S. aureus was sensitive to cephalosporins and quinolones, but resistant to penicillins. E. coli and K. pneumoniae showed a high resistance (16.7% and 25.6%, respectively) to commonly used aminoglycoside such as gentamycin. CONCLUSION: The burden of NNS was high with high mortality in the study centre. The sensitivity pattern had remarkably changed; however, a combination of cephalosporins such as cefuroxime and gentamycin is still a good option.

Engineered female-specific lethality for control of pest Lepidoptera.

The sterile insect technique (SIT) is a pest control strategy involving the mass release of radiation-sterilized insects, which reduce the target population through nonviable matings. In Lepidoptera, SIT could be more broadly applicable if the deleterious effects of sterilization by irradiation could be avoided. Moreover, male-only release can improve the efficacy of SIT. Adequate methods of male-only production in Lepidoptera are currently lacking, in contrast to some Diptera. We describe a synthetic genetic system that allows male-only moth production for SIT and also replaces radiation sterilization with inherited female-specific lethality. We sequenced and characterized the doublesex (dsx) gene from the pink bollworm (Pectinophora gossypiella). Sex-alternate splicing from dsx was used to develop a conditional lethal genetic sexing system in two pest moths: the diamondback moth (Plutella xylostella) and pink bollworm. This system shows promise for enhancing existing pink bollworm SIT, as well as broadening SIT-type control to diamondback moth and other Lepidoptera.