Use of Larval Morphological Deformities in Chironomus plumosus (Chironomidae: Diptera) as an Indicator of Freshwater Environmental Contamination (Lake Trasimeno, Italy)

The mentum deformity incidence in Chironomus plumosus larvae to assess the environmental contamination level in Lake Trasimeno, Central Italy, was investigated. The survey lasted from May 2018 to August 2019. Fifty-one samplings were carried out: 34 in the littoral zone and 17 in the central zone. The deformity assessment was based on 737 and 2767 larval specimens of C. plumosus collected from the littoral and central zones, respectively. Comparison of the larval morphometric variables between normal and deformed specimens highlighted that the deformities did not cause alterations of the larval growth. The deformity incidence amounted to 7.22% in the whole Trasimeno's ecosystem, reaching 8.28% in the littoral zone and 6.94% in the central zone. Among the different seasonal cohorts, the spring cohort had overall the highest deformity value (11.41%). The deformity type assessment protocol highlighted that the most common deformity type was "round/filed teeth" (64%). The results of this 2018–2019 survey revealed a low deformity incidence, within the background range of relatively low-impacted freshwaters. Comparison with previous investigations (2000–2010) of the same habitat showed a clear decrease of the deformity incidence. This study further contributes to the evaluation of the mentum deformity in chironomids that represent an indicator endpoint of the anthropogenic contamination level in freshwaters

Comparative effect of glucagon and isoproterenol on hepatic glycogenolysis and glycolysis in isolated perfused liver

The effect of glucagon and isoproterenol (beta-adrenergic agonist) on hepatic glycogenolysis and glycolysis in isolated perfused liver was compared. The levels of isoproterenol and glucagon which promoted the maximal activation of glycogenolysis were 20 muM and 1nM respectively. However, glucagon (1 nM) not only increased glycogenolysis but also inhibited glycolysis. Because adenosine-3'-5'-cyclic monophosphate (cAMP) is a common second messenger to glucagon and isoproterenol, the level of cAMP that simulates the effect of these substances were investigated. The concentration of cAMP that inhibited glycolysis was five times higher (15 muM) than that which stimulated glycogenolysis (3 muM). Similar inhibition of glycolysis was obtained with cAMP agonists resistant to phosphodiesterases, i.e., 8-Br-cAMP and N6-monobutyryl-cAMP (6-MB-cAMP) at the concentration of 3 muM. Thus, apparently glucagon could produce higher cellular levels of cAMP than that obtained with the activation of beta-adrenergic receptors. The higher amount of cAMP could be enough to overcome the action of phosphodiesterases and penetrate in the cytosol creating a favourable gradient to inhibit the enzymes of glycolysis

Use of Larval Morphological Deformities in Chironomus plumosus (Chironomidae: Diptera) as an Indicator of Freshwater Environmental Contamination (Lake Trasimeno, Italy)

The mentum deformity incidence in Chironomus plumosus larvae to assess the environmental contamination level in Lake Trasimeno, Central Italy, was investigated. The survey lasted from May 2018 to August 2019. Fifty-one samplings were carried out: 34 in the littoral zone and 17 in the central zone. The deformity assessment was based on 737 and 2767 larval specimens of C. plumosus collected from the littoral and central zones, respectively. Comparison of the larval morphometric variables between normal and deformed specimens highlighted that the deformities did not cause alterations of the larval growth. The deformity incidence amounted to 7.22% in the whole Trasimeno’s ecosystem, reaching 8.28% in the littoral zone and 6.94% in the central zone. Among the different seasonal cohorts, the spring cohort had overall the highest deformity value (11.41%). The deformity type assessment protocol highlighted that the most common deformity type was “round/filed teeth” (64%). The results of this 2018–2019 survey revealed a low deformity incidence, within the background range of relatively low-impacted freshwaters. Comparison with previous investigations (2000–2010) of the same habitat showed a clear decrease of the deformity incidence. This study further contributes to the evaluation of the mentum deformity in chironomids that represent an indicator endpoint of the anthropogenic contamination level in freshwaters

Population Dynamics and Seasonal Patterns of <i>Chironomus plumosus</i> (Diptera, Chironomidae) in the Shallow Lake Trasimeno, Central Italy

Field sampling of littoral macrobenthos of the shallow Lake Trasimeno was conducted along 17 years (2005–2021) on 129 different occasions. This long-term field study deepens the knowledge concerning the life cycle of Chironomus plumosus (Diptera), the main responsible for summer chironomid swarms that adversely affect human littoral activities, providing useful information for its management. About 108,000 macrobenthic specimens were collected, belonging to Oligochaeta (Naididae) (62%), Diptera (Chironomidae) (37%), and only 1.5% to other invertebrate taxa. Eighteen chironomid taxa were found. The trend of chironomid density was not affected by C. plumosus, which showed a maximum increase in September. This peak is justified by the presence of large swarms of C. plumosus in late August in which the populations of the central area of Lake Trasimeno consistently participate. The larval density of this species did not increase over the 17 years. A detailed analysis of the sampled larvae and adult biomass catches from 2017 to 2020 reveals that four annual swarmings occurred: in April, July, August, and September–October. The water temperature remains higher than 20 °C during the night hours from the end of May to mid-September, strengthening the hypothesis of the three midge swarming cycles in the summer period until early autumn

Evaluation of the association between the JAK2 46/1 haplotype and chronic myeloproliferative neoplasms in a Brazilian population

OBJECTIVE: The JAK2 46/1 haplotype has recently been described as a major contributing factor to the development of myeloproliferative neoplasm, whether positive or negative forthe JAK2 V617F mutation. The G allele, identified by a single-nucleotide polymorphism known as JAK2 rs10974944, is part of the JAK2 46/1 haplotype. The aim of this study was to verify the association between the presence of the G allele and the development of BCR-ABL-negative chronic myeloproliferative neoplasms in our population. METHODS: Blood and oral mucosa swab samples were obtained from 56 patients of two local Brazilian hospitals who had previously been diagnosed with BCR-ABL-negative chronic myeloproliferative neoplasms. Blood samples from 90 local blood donors were used as controls. The presence of the G allele was assessed using a PCR-RFLP assay after extracting DNA from the samples. RESULTS: The presence of the G allele was strongly associated with the presence of BCR-ABL-negative chronic myeloproliferative neoplasms (p = 0.0001; OR = 2.674; 95% CI = 1.630-4.385) in the studied population. CONCLUSION: In agreement with previous reports, the JAK2 46/1 haplotype, represented in this study by the presence of the G allele, is an important predisposing factor in the oncogenetic development of these neoplasms in our population

Mutation of C20orf7 Disrupts Complex I Assembly and Causes Lethal Neonatal Mitochondrial Disease

Complex I (NADH:ubiquinone oxidoreductase) is the first and largest multimeric complex of the mitochondrial respiratory chain. Human complex I comprises seven subunits encoded by mitochondrial DNA and 38 nuclear-encoded subunits that are assembled together in a process that is only partially understood. To date, mutations causing complex I deficiency have been described in all 14 core subunits, five supernumerary subunits, and four assembly factors. We describe complex I deficiency caused by mutation of the putative complex I assembly factor C20orf7. A candidate region for a lethal neonatal form of complex I deficiency was identified by homozygosity mapping of an Egyptian family with one affected child and two affected pregnancies predicted by enzyme-based prenatal diagnosis. The region was confirmed by microcell-mediated chromosome transfer, and 11 candidate genes encoding potential mitochondrial proteins were sequenced. A homozygous missense mutation in C20orf7 segregated with disease in the family. We show that C20orf7 is peripherally associated with the matrix face of the mitochondrial inner membrane and that silencing its expression with RNAi decreases complex I activity. C20orf7 patient fibroblasts showed an almost complete absence of complex I holoenzyme and were defective at an early stage of complex I assembly, but in a manner distinct from the assembly defects caused by mutations in the assembly factor NDUFAF1. Our results indicate that C20orf7 is crucial in the assembly of complex I and that mutations in C20orf7 cause mitochondrial disease

Targeted proteomics to study mitochondrial biology

Targeted mass spectrometry in the selected or parallel reaction monitoring (SRM or PRM) mode is a widely used methodology to quantify proteins based on so-called signature or proteotypic peptides. SRM has the advantage of being able to quantify a range of proteins in a single analysis, for example, to measure the level of enzymes comprising a biochemical pathway. In this chapter, we will detail how to set up an SRM assay on the example of the mitochondrial protein succinate dehydrogenase [ubiquinone] flavoprotein subunit (mouse UniProt-code Q8K2B3). First, we will outline the in silico assay design including the choice of peptides based on a range of properties. We will further delineate different quantification strategies and introduce the reader to LC-MS assay development including the selection of the optimal peptide charge state and fragment ions as well as a discussion of the dynamic range of detection. The chapter will close with an application from the area of mitochondrial biology related to the quantification of a set of proteins isolated from mouse liver mitochondria in a study on mitochondrial respiratory flux decline in aging mouse muscle