Molecular and functional analysis of the TOL plasmid pWWO from Pseudomonas putida and cloning of genes for the entire regulated aromatic ring meta cleavage pathway.

The genetic organization of the Pseudomonas putida plasmid pWWO-161, which encodes enzymes for the degradation of toluene and related aromatic hydrocarbons, has been investigated by transposition mutagenesis and gene cloning. Catabolic genes were localized to two clusters, one for upper pathway (hydrocarbon leads to carboxylic acid) enzymes and the other for lower pathway (carboxylic acid leads to tricarboxylic acid cycle) enzymes, that are separated by a 14-kilobase DNA segment. The physical organization of the catabolic genes thus reflects their functional organization into two regulatory blocks. The pWWO-161 DNA fragments Sst I fragment C and fragment D were cloned in a broad host range vector to produce plasmid pKT530. This hybrid encodes toluate oxygenase and all meta cleavage pathway enzymes, and it enables P. putida mt-2 and Escherichia coli K-12 cells to grow on m-toluate as sole carbon source. The pKT530 plasmid also carries xylS (a gene whose product has been postulated to regulate expression of the lower pathway genes) and the control sequences of the pathway that interact with this product, because catechol 2,3-oxygenase synthesis is specifically induced by m-toluate in both P. putida and E. coli. Evidence is presented that suggests the promoter operator of the meta pathway gene functions less effectively with the RNA polymerase or xylS product of E. coli than with the enzyme or product of P. putida.     

Pediatric traumatic brain injury: not just little adults

PURPOSE OF REVIEW: This review will update the reader on the most significant recent findings with regards to both the clinical research and basic science of pediatric traumatic brain injury. RECENT FINDINGS: The developing brain is not simply a smaller version of the mature brain. Studies have uncovered important distinctions of the younger brain after traumatic brain injury, including an increased propensity for apoptosis, age-dependent parameters for cerebral blood flow and metabolism, development-specific biomarkers, increased likelihood of early posttraumatic seizures, differential sensitivity to commonly used neuroactive medications and altered neuroplasticity during recovery from injury. Specifically, there is strong preclinical evidence for increased neuronal apoptosis in the developing brain being triggered by anesthetics and anticonvulsants, making it paramount that future studies more clearly delineate preferred agents and specific indications for use, incorporating long-term functional outcomes as well as short-term benefits. In addition, the young brain may actually benefit from therapeutic interventions that have been less effective following adult traumatic brain injury, such as decompressive craniectomy and hypothermia. SUMMARY: An increasing body of evidence demonstrates the importance of establishing age-dependent guidelines for physiological monitoring, pharmacological intervention, management of intracranial pressure and facilitating recovery of function.     

Potentiation of the function of Hageman factor fragments by high molecular weight kininogen.

Patients lacking high molecular weight (HMW) kininogen have profound abnormalities of the Hageman factor-dependent pathways of coagulation, kinin formation, and fibrinolysis. The ability of HMW kininogen to potentiate the Hageman factor fragments (HFf) activation of prekallikrein and Factor XI in plasma was studied. HFf only partially converted Factor XI to XIa and prekallikrein to kallikrein in plasma deficient in HMW kininogen (Williams trait), while enhanced activation of Factor XI and prekallikrein by HFf resulted after reconstitution with HMW kininogen. In a system using highly purified components, HMW kininogen increased the initial rate of prekallikrein activation whether the kallikrein formed was assayed by arginine esterase activity or kininforming ability. The potentiation of prekallikrein activation occurred over a 12-fold range of enzyme (HFf) concentration and was nonhyperbolic with respect to substrate (prekallikrein). HMW kininogen exerted its effect even in the absence of prekallikrein since the hydrolysis of acetylglycyl-lysine methyl ester by HFf was increased by HMW kininogen. These results suggest that one of the functions of HMW kininogen is to augment the catalytic action of HFf.     

Molecular Analysis of SARS-CoV-2 Lineages in Armenia

The sequencing of SARS-CoV-2 provides essential information on viral evolution, transmission, and epidemiology. In this paper, we performed the whole-genome sequencing of SARS-CoV-2 using nanopore and Illumina sequencing to describe the circulation of the virus lineages in Armenia. The analysis of 145 full genomes identified six clades (19A, 20A, 20B, 20I, 21J, and 21K) and considerable intra-clade PANGO lineage diversity. Phylodynamic and transmission analysis allowed to attribute specific clades as well as infer their importation routes. Thus, the first two waves of positive case increase were caused by the 20B clade, the third peak caused by the 20I (Alpha), while the last two peaks were caused by the 21J (Delta) and 21K (Omicron) variants. The functional analyses of mutations in sequences largely affected epitopes associated with protective HLA loci and did not cause the loss of the signal in PCR tests targeting ORF1ab and N genes as confirmed by RT-PCR. We also compared the performance of nanopore and Illumina short-read sequencing and showed the utility of nanopore sequencing as an efficient and affordable alternative for large-scale molecular epidemiology research. Thus, our paper describes new data on the genomic diversity of SARS-CoV-2 variants in Armenia in the global context of the virus molecular genomic surveillance.     

Interaction between muscles and fascia in the mystacial pad of whisking rodents

In whisking rodents, the mystacial pad is supplied with vibrissae and contains a collagenous skeleton that is a part of the snout fascia. The collagenous skeleton is composed of three interconnected layers: superficial, deep spongy mesh and subcapsular fibrous mat. We found that the first two layers contain diverse fascial structures, such as sheets of subcutaneous connective tissue, tendons, ligaments and follicular capsules which transmit muscle efforts to vibrissae and are thus involved in whisking. Subcapsular fibrous mat is built of oriented rostro-caudal wavy fibrils. It maintains spatial arrangement of whisker follicles, provides a quick response to deformation and connects entire mystacial pad to the skull. To move vibrissae, the forces of intrinsic muscles are applied directly to the capsules of the vibrissa follicles, whereas the forces of extrinsic muscles are applied to other parts of the collagenous skeleton, which transmit the forces to the capsules. According to the spatial distribution and anchoring sites of the muscles and fascia, extrinsic muscles provide vibrissa protraction or retraction by pulling the superficial layer of the collagenous skeleton rostral or caudal, respectively. Vibrissae can be also retracted when the efforts of extrinsic muscles are applied to the subcapsular fibrous mat. When the muscles relax, fascial structures return the vibrissae to their resting position. The deep spongy layer encompasses vibrissal follicles providing a uniform distribution of stresses and strains during whisking. In the mystacial pad, fascia is a dominant type of tissue that maintains the integrity of the vibrissa motor plant, translates muscular momentum to the vibrissae, and plays a role in vibrissae movements.     

The Effect of Sevoflurane and Desflurane on Upper Airway Reactivity

Background: Although bronchial reactivity can be assessed by changes in airway resistance, there is no well-accepted measure of upper airway reactivity during anesthesia. The authors used the stimulus of endotracheal tube cuff inflation and deflation to assess changes in airway reactivity in patients anesthetized with sevoflurane and desflurane.

Methods: Sixty-four patients classified as American Society of Anesthesiologists physical status I or II participated in this randomized, double-blind study. Patients were anesthetized with either sevoflurane or desflurane at 1.0 and 1.8 minimum alveolar concentration (MAC). The trachea was stimulated by inflating the endotracheal tube cuff. A blinded observer assessed the severity of patient response to the stimulus and changes in hemodynamic variables. The process was repeated at the second MAC treatment condition.

Results: At 1.0 MAC, patients anesthetized with desflurane had a more intense response and a greater likelihood of significant coughing and associated hemodynamic changes (both at P < 0.05). At 1.8 MAC, sevoflurane and desflurane both suppressed clinically significant responses to tracheal stimulation. Interrater reliability was excellent for this measure of upper airway reactivity (P < 0.001).     

Infectious complications of dialysis access devices

Infectious complications remain a major source of morbidity and mortality for patients with end-stage renal disease on dialysis. The majority of these complications are related to dialysis access devices, and as such represent a potentially modifiable risk factor. This article reviews the important infectious complications associated with dialysis access, including both hemodialysis and peritoneal dialysis. The discussion highlights the epidemiology, management, and prevention of dialysis access infections.