Real‐time mid‐infrared imaging of living microorganisms

The speed and efficiency of quantum cascade laser‐based mid‐infrared microspectroscopy are demonstrated using two different model organisms as examples. For the slowly moving Amoeba proteus, a quantum cascade laser is tuned over the wavelength range of 7.6 µm to 8.6 µm (wavenumbers 1320 cm^–1 and 1160 cm^–1, respectively). The recording of a hyperspectral image takes 11.3 s whereby an average signal‐to‐noise ratio of 29 is achieved. The limits of time resolution are tested by imaging the fast moving Caenorhabditis elegans at a discrete wavenumber of 1265 cm^–1. Mid‐infrared imaging is performed with the 640 × 480 pixel video graphics array (VGA) standard and at a full‐frame time resolution of 0.02 s (i.e. well above the most common frame rate standards). An average signal‐to‐noise ratio of 16 is obtained. To the best of our knowledge, these findings constitute the first mid‐infrared imaging of living organisms at VGA standard and video frame rate.

     

Impact of Early Initiation of Antiretroviral Therapy in Patients with Acute HIV Infection in Vienna,Austria

BackgroundIt is unclear whether antiretroviral therapy (ART) should be initiated during acute HIV infection. Most recent data provides evidence of benefits of early ART.MethodsWe retrospectively compared the clinical and immunological course of individuals with acute HIV infection, who received ART within 3 months (group A) or not (group B) after diagnosis.ResultsAmong the 84 individuals with acute HIV infection, 57 (68%) received ART within 3 months (A) whereas 27 (32%) did not receive ART within 3 months (B), respectively. Clinical progression to CDC stadium B or C within 5 years after the diagnosis of HIV was less common in (A) when compared to (B) (P = 0.002). After twelve months, both the mean increase in CD4+ T cell count and the mean decrease in viral load was more pronounced in (A), when compared to (B) (225 vs. 87 cells/μl; P = 0.002 and -4.19 vs. -1.14 log10 copies/mL; P<0.001). Twenty-four months after diagnosis the mean increase from baseline of CD4+ T cells was still higher in group A compared to group B (251 vs. 67 cells/μl, P = 0.004).ConclusionsInitiation of ART during acute HIV infection is associated with a lower probability of clinical progression to more advanced CDC stages and significant immunological benefits.     

Updating the proteome of the uncultivable hemotrophic Mycoplasma suis in experimentally infected pigs

Mycoplasma suis belongs to the hemotrophic mycoplasmas that are associated with acute and chronic anemia in a wide range of livestock and wild animals. The inability to culture M. suis in vitro has hindered its characterization at the molecular level. Since the publication of M. suis genome sequences in 2011 only one proteome study has been published. Aim of the presented study was to significantly extend the proteome coverage of M. suis strain KI_3806 during acute infection by applying three different protein extraction methods followed by 1D SDS‐PAGE and LC‐MS/MS. A total of 404 of 795 M. suis KI_3806 proteins (50.8%) were identified. Data analysis revealed the expression of 83.7% of the predicted ORFs with assigned functions but also highlights the expression of 179 of 523 (34.2%) hypothetical proteins with unknown functions. Computational analyses identified expressed membrane‐associated hypothetical proteins that might be involved in adhesion or host–pathogen interaction. Furthermore, analyses of the expressed proteins indicated the existence of a hexose‐6‐phosphate‐transporter and an ECF transporter. In conclusion, our proteome study provides a further step toward the elucidation of the unique life cycle of M. suis and the establishment of an in vitro culture. All MS data have been deposited in the ProteomeXchange with identifier PXD002294 ( http://proteomecentral.proteomexchange.org/dataset/PXD002294 ).     

The Effects of Lung Protective Ventilation or Hypercapnic Acidosis on Gas Exchange and Lung Injury in Surfactant Deficient Rabbits

Background

Permissive hypercapnia has been shown to reduce lung injury in subjects with surfactant deficiency. Experimental studies suggest that hypercapnic acidosis by itself rather than decreased tidal volume may be a key protective factor.

Objectives

To study the differential effects of a lung protective ventilatory strategy or hypercapnic acidosis on gas exchange, hemodynamics and lung injury in an animal model of surfactant deficiency.

Methods

30 anesthetized, surfactant-depleted rabbits were mechanically ventilated (FiO₂ = 0.8, PEEP = 7cmH₂O) and randomized into three groups: Normoventilation-Normocapnia (NN)-group: tidal volume (Vt) = 7.5 ml/kg, target PaCO₂ = 40 mmHg; Normoventilation-Hypercapnia (NH)-group: Vt = 7.5 ml/kg, target PaCO₂ = 80 mmHg by increasing FiCO₂; and a Hypoventilation-Hypercapnia (HH)-group: Vt = 4.5 ml/kg, target PaCO₂ = 80 mmHg. Plasma lactate and interleukin (IL)-8 were measured every 2 h. Animals were sacrificed after 6 h to perform bronchoalveolar lavage (BAL), to measure lung wet-to-dry weight, lung tissue IL-8, and to obtain lung histology.

Results

PaO₂ was significantly higher in the HH-group compared to the NN-group (p<0.05), with values of the NH-group between the HH- and NN-groups. Other markers of lung injury (wet-dry-weight, BAL-Protein, histology-score, plasma-IL-8 and lung tissue IL-8) resulted in significantly lower values for the HH-group compared to the NN-group and trends for the NH-group towards lower values compared to the NN-group. Lactate was significantly lower in both hypercapnia groups compared to the NN-group.

Conclusion

Whereas hypercapnic acidosis may have some beneficial effects, a significant effect on lung injury and systemic inflammatory response is dependent upon a lower tidal volume rather than resultant arterial CO₂ tensions and pH alone.     

Phthalates Are Metabolised by Primary Thyroid Cell Cultures but Have Limited Influence on Selected Thyroid Cell Functions In Vitro

Phthalates are plasticisers added to a wide variety of products, resulting in measurable exposure of humans. They are suspected to disrupt the thyroid axis as epidemiological studies suggest an influence on the peripheral thyroid hormone concentration. The mechanism is still unknown as only few in vitro studies within this area exist. The aim of the present study was to investigate the influence of three phthalate diesters (di-ethyl phthalate, di-n-butyl phthalate (DnBP), di-(2-ethylhexyl) phthalate (DEHP)) and two monoesters (mono-n-butyl phthalate and mono-(2-ethylhexyl) phthalate (MEHP)) on the differentiated function of primary human thyroid cell cultures. Also, the kinetics of phthalate metabolism were investigated. DEHP and its monoester, MEHP, both had an inhibitory influence on 3''-5''-cyclic adenosine monophosphate secretion from the cells, and MEHP also on thyroglobulin (Tg) secretion from the cells. Results of the lactate dehydrogenase-measurements indicated that the MEHP-mediated influence was caused by cell death. No influence on gene expression of thyroid specific genes (Tg, thyroid peroxidase, sodium iodine symporter and thyroid stimulating hormone receptor) by any of the investigated diesters could be demonstrated. All phthalate diesters were metabolised to the respective monoester, however with a fall in efficiency for high concentrations of the larger diesters DnBP and DEHP. In conclusion, human thyroid cells were able to metabolise phthalates but this phthalate-exposure did not appear to substantially influence selected functions of these cells.     

Archaeal and Bacterial Communities Associated with the Surface Mucus of Caribbean Corals Differ in Their Degree of Host Specificity and Community Turnover Over Reefs

Comparative studies on the distribution of archaeal versus bacterial communities associated with the surface mucus layer of corals have rarely taken place. It has therefore remained enigmatic whether mucus-associated archaeal and bacterial communities exhibit a similar specificity towards coral hosts and whether they vary in the same fashion over spatial gradients and between reef locations. We used microbial community profiling (terminal-restriction fragment length polymorphism, T-RFLP) and clone library sequencing of the 16S rRNA gene to compare the diversity and community structure of dominant archaeal and bacterial communities associating with the mucus of three common reef-building coral species (Porites astreoides, Siderastrea siderea and Orbicella annularis) over different spatial scales on a Caribbean fringing reef. Sampling locations included three reef sites, three reef patches within each site and two depths. Reference sediment samples and ambient water were also taken for each of the 18 sampling locations resulting in a total of 239 samples. While only 41% of the bacterial operational taxonomic units (OTUs) characterized by T-RFLP were shared between mucus and the ambient water or sediment, for archaeal OTUs this percentage was 2-fold higher (78%). About half of the mucus-associated OTUs (44% and 58% of bacterial and archaeal OTUs, respectively) were shared between the three coral species. Our multivariate statistical analysis (ANOSIM, PERMANOVA and CCA) showed that while the bacterial community composition was determined by habitat (mucus, sediment or seawater), host coral species, location and spatial distance, the archaeal community composition was solely determined by the habitat. This study highlights that mucus-associated archaeal and bacterial communities differ in their degree of community turnover over reefs and in their host-specificity.     

Analysis of Genes Involved in Body Weight Regulation by Targeted Re-Sequencing

Introduction

Genes involved in body weight regulation that were previously investigated in genome-wide association studies (GWAS) and in animal models were target-enriched followed by massive parallel next generation sequencing.

Methods

We enriched and re-sequenced continuous genomic regions comprising FTO, MC4R, TMEM18, SDCCAG8, TKNS, MSRA and TBC1D1 in a screening sample of 196 extremely obese children and adolescents with age and sex specific body mass index (BMI) ≥ 99^th percentile and 176 lean adults (BMI ≤ 15^th percentile). 22 variants were confirmed by Sanger sequencing. Genotyping was performed in up to 705 independent obesity trios (extremely obese child and both parents), 243 extremely obese cases and 261 lean adults.

Results and Conclusion

We detected 20 different non-synonymous variants, one frame shift and one nonsense mutation in the 7 continuous genomic regions in study groups of different weight extremes. For SNP Arg695Cys (rs58983546) in TBC1D1 we detected nominal association with obesity (p_TDT = 0.03 in 705 trios). Eleven of the variants were rare, thus were only detected heterozygously in up to ten individual(s) of the complete screening sample of 372 individuals. Two of them (in FTO and MSRA) were found in lean individuals, nine in extremely obese. In silico analyses of the 11 variants did not reveal functional implications for the mutations. Concordant with our hypothesis we detected a rare variant that potentially leads to loss of FTO function in a lean individual. For TBC1D1, in contrary to our hypothesis, the loss of function variant (Arg443Stop) was found in an obese individual. Functional in vitro studies are warranted.