Plastic microbiome development in a freshwater ecosystem.

To understand biological interactions of plastic litter in freshwater ecosystems, as well the potential effects of plastics on ecosystem processes, studies of the activity and composition of plastic-associated microbial communities are needed. The physical properties and chemical composition of plastic polymers are key components of plastic product design, and may also select for distinct microbial biofilms colonizing plastic litter. We monitored growth and succession of biofilm communities on plastic substrates of common morphotypes (i.e., hard, soft, foam, and film) and a natural surface (i.e., an unglazed ceramic tile) incubated in an urban stream. We measured biofilm biomass, metabolism, extracellular enzyme activity, and bacterial, fungal and algal community composition over four weeks during primary succession. Results demonstrated a general increase in biofilm biomass and enzymatic activity corresponding to carbon, nitrogen and phosphorus metabolism during biofilm development for all substrate types. We observed higher respiration rates and negative net ecosystem productivity on foam and tile surfaces in comparison to hard, soft and film plastic surfaces. Biofilm bacterial, fungal and algal assemblages showed few significant differences in composition among substrates. However, all microbial communities changed significantly in composition over time. While substrate type was not the major factor driving biofilm composition and activity, these data show plastic litter in streams is well colonized by an active and dynamic biofilm community. As plastic litter is increasing across all types of aquatic ecosystems, it should be considered a medium for biologically active organisms that contribute to key ecosystem processes.

Pyrosequencing of IDH1 and IDH2 mutations in brain tumors and non-neoplastic conditions.

The molecular profiling of brain tumors, including testing for MGMT promoter methylation and chromosome 1p/19q deletion, can provide both diagnostic and prognostic information that may guide treatment. Isocitrate dehydrogenase (IDH) mutation testing is a recent addition to this armamentarium of molecular pathology tools that similarly provides both diagnostic (eg, glioma vs. gliosis) and prognostic information. Herein, we describe a pyrosequencing-based approach to IDH1 and IDH2 mutation testing and its application to 139 neoplastic and non-neoplastic central nervous system specimens. Several technical issues encountered in the development of the assay, particularly with regard to the optimization of the sequencing reaction, are described. Mutations in IDH1 codon 132 or IDH2 codon 172 were identified in 31.2% of all screened cases and 46.2% of screened World Health Organization grade I to IV gliomas (n=93), with mutations arising exclusively in grade II to IV oligodendroglial, astrocytic, or mixed oligoastrocytic neoplasms. Examination of the relationship between the mutation status and other pertinent variables demonstrated a significant male predominance among IDH1-mutated gliomas, most notably in grade III to IV astrocytic neoplasms. A significant association between IDH1/IDH2 mutation and 1p/19q deletion was also seen (Kendall τ coefficient=0.26, P=0.018), although several cases with 1p/19q deletion were IDH1/IDH2 wild type.

Comparison of 2 line blot assays for defining HPV genotypes in oral and oropharyngeal squamous cell carcinomas.

Patients with invasive oral and oropharyngeal squamous cell carcinomas infected with human papillomaviruses (HPV) demonstrate improved survival. HPV detection in tumors may assist in risk stratification of patients and in guiding optimum treatment. Two reverse line blot assays [Linear Array (LA) and INNO-LiPA (LiPA)] were evaluated for detection of HPV genotypes in paraffin-embedded biopsies. Overall, 82.4% of 131 biopsies were HPV+ by LiPA versus 61.1% by LA (κ = 0.32). Completely concordant results were observed in 52.7% of cases: 18 negative and 51 with exactly the same genotype(s). An additional 13 cases had partial agreement. These 82 completely or partially concordant cases revealed a high rate of HPV positivity (78.0%), primarily involving HPV16 (90.6%). HPV+ tumors occurred preferentially in the oropharynx, especially tonsils, with trends for male patients and poor differentiation. Significant differences in these associations were found when LA and LiPA results were analyzed independently. No relationships were found between tumor HPV status and tobacco or alcohol use.

Proteasome degradation of GRK2 during ischemia and ventricular tachyarrhythmias in a canine model of myocardial infarction.

Arrhythmia-prone subepicardial border zone (EBZ) tissue demonstrates decreased G protein receptor kinase 2 (GRK2) activity and increased sensitivity to isoproterenol 6-24 h after coronary artery ligation (CAL) in the dog. With the use of a semiquantitative immunofluorescence technique, the relative fluorescence intensity (RF) of GRK2 in EBZ decreased to 24% of that in a remote site (RS) (P < 0.01, n = 30 cells from 3 dogs), whereas GRK5 RF did not change. Confocal studies of cardiac tissue from transgenic mice overexpressing GRK2 validated the use of a semilogarithmic relationship between RF and GRK2 activity. As shown with the use of quantitative real-time RT-PCR, both GRK2 and GRK5 mRNA were not decreased at 24 h in EBZ (n = 6 dogs) relative to RS control, indicating that the decrease of GRK2 in the EBZ is likely due to posttranscriptional degradation following CAL. Pretreatment of six dogs with the selective proteasome inhibitor bortezomib provided 100% (EBZ) and 50% (infarct) protection against loss of GRK2 at 24 h. There was an absence of rapid (>300 beats/min) and very rapid (>360 beats/min) ventricular triplets that are highly predictive of sudden cardiac death during ECG monitoring in the bortezomib-pretreated animals in contrast to nonpretreated infarcted animals. We have demonstrated that the dramatic decrease in GRK2 in cardiac ischemic tissue can be largely blocked by prior proteasome blockade and that this is associated with significant cardioprotection against malignant ventricular tachyarrhythmias.