Antimicrobial resistance-Do we share more than companionship with our dogs?

AIMS: To investigate and compare antimicrobial resistance genes (ARGs) in faeces from cohabiting dogs and owners. METHODS AND RESULTS: DNA from faecal samples from 35 dogs and 35 owners was screened for the presence of 34 clinically relevant ARGs using high throughput qPCR. In total, 24 and 25 different ARGs were present in the dog and owner groups, respectively. The households had a mean of 9.9 ARGs present, with dogs and owners sharing on average 3.3 ARGs. ARGs were shared significantly more in households with dogs over 6 years old (3.5, interquartile range 2.75-5.0) than in households with younger dogs (2.5, interquartile range 2.0-3.0) (p = 0.02). Dogs possessed significantly more mecA and aminoglycoside resistance genes than owners. CONCLUSIONS: Dogs and owners can act as reservoirs for a broad range of ARGs belonging to several antimicrobial resistance classes. A modest proportion of the same resistance genes were present in both dogs and owners simultaneously, indicating that ARG transmission between the dog and human gut is of minor concern in the absence of antimicrobial selection. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides insight into the common dog and human gut resistomes, contributing to an improved knowledge base in risk assessments regarding ARG transmission between dogs and humans.

Microtiter plate cultivation of oleaginous fungi and monitoring of lipogenesis by high-throughput FTIR spectroscopy.

BACKGROUND: Oleaginous fungi can accumulate lipids by utilizing a wide range of waste substrates. They are an important source for the industrial production of omega-6 polyunsaturated fatty acids (gamma-linolenic and arachidonic acid) and have been suggested as an alternative route for biodiesel production. Initial research steps for various applications include the screening of fungi in order to find efficient fungal producers with desired fatty acid composition. Traditional cultivation methods (shake flask) and lipid analysis (extraction-gas chromatography) are not applicable for large-scale screening due to their low throughput and time-consuming analysis. Here we present a microcultivation system combined with high-throughput Fourier transform infrared (FTIR) spectroscopy for efficient screening of oleaginous fungi. RESULTS: The microcultivation system enables highly reproducible fungal fermentations throughout 12 days of cultivation. Reproducibility was validated by FTIR and HPLC data. Analysis of FTIR spectral ester carbonyl peaks of fungal biomass offered a reliable high-throughput at-line method to monitor lipid accumulation. Partial least square regression between gas chromatography fatty acid data and corresponding FTIR spectral data was used to set up calibration models for the prediction of saturated fatty acids, monounsaturated fatty acids, polyunsaturated fatty acids, unsaturation index, total lipid content and main individual fatty acids. High coefficients of determination (R2 = 0.86-0.96) and satisfactory residual predictive deviation of cross-validation (RPDCV = 2.6-5.1) values demonstrated the goodness of these models. CONCLUSIONS: We have demonstrated in this study, that the presented microcultivation system combined with rapid, high-throughput FTIR spectroscopy is a suitable screening platform for oleaginous fungi. Sample preparation for FTIR measurements can be automated to further increase throughput of the system.

Temporal analysis shows relaxed genetic erosion following improved stocking practices in a subarctic transnational brown trout population.

Maintaining standing genetic variation is a challenge in human-dominated landscapes. We used genetic (i.e., 16 short tandem repeats) and morphological (i.e., length and weight) measurements of 593 contemporary and historical brown trout (Salmo trutta) samples to study fine-scale and short-term impacts of different management practices. These had changed from traditional breeding practices, using the same broodstock for several years, to modern breeding practices, including annual broodstock replacement, in the transnational subarctic Pasvik River. Using population genetic structure analyses (i.e., Bayesian assignment tests, DAPCs, and PCAs), four historical genetic clusters (E2001A-D), likely representing family lineages resulting from different crosses, were found in zone E. These groups were characterized by consistently lower genetic diversity, higher within-group relatedness, lower effective population size, and significantly smaller body size than contemporary stocked (E2001E) and wild fish (E2001F). However, even current breeding practices are insufficient to prevent genetic diversity loss and morphological changes as demonstrated by on average smaller body sizes and recent genetic bottleneck signatures in the modern breeding stock compared to wild fish. Conservation management must evaluate breeding protocols for stocking programs and assess if these can preserve remaining natural genetic diversity and morphology in brown trout for long-term preservation of freshwater fauna.

Genetic changes caused by restocking and hydroelectric dams in demographically bottlenecked brown trout in a transnational subarctic riverine system.

Habitat discontinuity, anthropogenic disturbance, and overharvesting have led to population fragmentation and decline worldwide. Preservation of remaining natural genetic diversity is crucial to avoid continued genetic erosion. Brown trout (Salmo trutta L.) is an ideal model species for studying anthropogenic influences on genetic integrity, as it has experienced significant genetic alterations throughout its natural distribution range due to habitat fragmentation, overexploitation, translocations, and stocking. The Pasvik River is a subarctic riverine system shared between Norway, Russia, and Finland, subdivided by seven hydroelectric power dams that destroyed about 70% of natural spawning and nursing areas. Stocking is applied in certain river parts to support the natural brown trout population. Adjacent river segments with different management strategies (stocked vs. not stocked) facilitated the simultaneous assessment of genetic impacts of dams and stocking based on analyses of 16 short tandem repeat loci. Dams were expected to increase genetic differentiation between and reduce genetic diversity within river sections. Contrastingly, stocking was predicted to promote genetic homogenization and diversity, but also potentially lead to loss of private alleles and to genetic erosion. Our results showed comparatively low heterozygosity and clear genetic differentiation between adjacent sections in nonstocked river parts, indicating that dams prevent migration and contribute to genetic isolation and loss of genetic diversity. Furthermore, genetic differentiation was low and heterozygosity relatively high across stocked sections. However, in stocked river sections, we found signatures of recent bottlenecks and reductions in private alleles, indicating that only a subset of individuals contributes to reproduction, potentially leading to divergence away from the natural genetic state. Taken together, these results indicate that stocking counteracts the negative fragmentation effects of dams, but also that stocking practices should be planned carefully in order to ensure long-term preservation of natural genetic diversity and integrity in brown trout and other species in regulated river systems.

FTIR Spectroscopy for Evaluation and Monitoring of Lipid Extraction Efficiency for Oleaginous Fungi.

To assess whether Fourier Transform Infrared (FTIR) spectroscopy could be used to evaluate and monitor lipid extraction processes, the extraction methods of Folch, Bligh and Lewis were used. Biomass of the oleaginous fungi Mucor circinelloides and Mortierella alpina were employed as lipid-rich material for the lipid extraction. The presence of lipids was determined by recording infrared spectra of all components in the lipid extraction procedure, such as the biomass before and after extraction, the water and extract phases. Infrared spectra revealed the incomplete extraction after all three extraction methods applied to M.circinelloides and it was shown that mechanical disruption using bead beating and HCl treatment were necessary to complete the extraction in this species. FTIR spectroscopy was used to identify components, such as polyphosphates, that may have negatively affected the extraction process and resulted in differences in extraction efficiency between M.circinelloides and M.alpina. Residual lipids could not be detected in the infrared spectra of M.alpina biomass after extraction using the Folch and Lewis methods, indicating their complete lipid extraction in this species. Bligh extraction underestimated the fatty acid content of both M.circinelloides and M.alpina biomass and an increase in the initial solvent-to-sample ratio (from 3:1 to 20:1) was needed to achieve complete extraction and a lipid-free IR spectrum. In accordance with previous studies, the gravimetric lipid yield was shown to overestimate the potential of the SCO producers and FAME quantification in GC-FID was found to be the best-suited method for lipid quantification. We conclude that FTIR spectroscopy can serve as a tool for evaluating the lipid extraction efficiency, in addition to identifying components that may affect lipid extraction processes.

A Practical Solution for 77 K Fluorescence Measurements Based on LED Excitation and CCD Array Detector.

The fluorescence emission spectrum of photosynthetic microorganisms at liquid nitrogen temperature (77 K) provides important insights into the organization of the photosynthetic machinery of bacteria and eukaryotes, which cannot be observed at room temperature. Conventionally, to obtain such spectra, a large and costly table-top fluorometer is required. Recently portable, reliable, and largely maintenance-free instruments have become available that can be utilized to accomplish a wide variety of spectroscopy-based measurements in photosynthesis research. In this report, we show how to build such an instrument in order to record 77K fluorescence spectra. This instrument consists of a low power monochromatic light-emitting diode (LED), and a portable CCD array based spectrometer. The optical components are coupled together using a fiber optic cable, and a custom made housing that also supports a dewar flask. We demonstrate that this instrument facilitates the reliable determination of chlorophyll fluorescence emission spectra for the cyanobacterium Synechocystis sp. PCC 6803, and the green alga Chlamydomonas reinhardtii.