Noninvasive Assessment of Chicken Egg Fertility during Incubation Using HSSE-GC-MS VOC Profiling.

Volatile organic compounds (VOCs) carry crucial information about chicken egg fertility. Assessing the fertility before incubation holds immense potential for poultry industry efficiency. Our study used headspace sorptive extraction-gas chromatography-mass spectrometry to analyze egg VOCs before and during the initial 12 incubation days. A total of 162 VOCs were identified. Hexanal was significantly higher in unfertilized eggs, whereas compounds such as propan-2-ol, propan-2-one, and carboxylic acids were higher in fertilized eggs. Furthermore, the obtained multiple logistic regression model outperformed the partial least-squares-discriminant analysis (PLS-DA) model, demonstrating lower complexity and superior performance. Fertile eggs were accurately identified in the validation set in 68-75% of the cases during the initial 4 days, to 85 and 100% on days 6 and 8. Finally, hierarchical cluster analysis in fertilized eggs revealed the clustering of VOCs of the same chemical class, indicative of their shared biochemical origin. This suggests a promising direction for future research aimed at understanding the biological information embedded in VOCs and their relationship to biochemical processes during embryo development.

Allantoic Fluid-Based qPCR for Early Onset In Ovo Sexing.

The day-old male chick culling remains a welfare issue in the poultry industry. Several governments have prohibited this practice, pushing hatcheries to seek alternatives. Although different solutions exist for solving this problem, sex determination during the embryo's incubation (in ovo sexing) is considered the most suitable one among the consumers and industry. However, to be industrialized, in ovo sexing technologies must meet several requirements: compatibility with all egg colors and early developmental stages while maintaining a high hatchability rate and accuracy at low cost and high throughput. To meet these requirements, we studied the use of the sexual genes HINTW (female-specific) and DMRT-1 (both sexes) at incubation days 6-9. By utilizing the quantitative polymerase chain reaction in allantoic fluid (AF) samples, our study confirmed female-specific HINTW detection on all days without any significant detrimental effects on embryo development. We achieved 95% sexing accuracy using the HINTW cycle threshold (Ct) alone and 100% accuracy rate when using Δλ values (difference between the HINTW and DMRT-1 Ct). In conclusion, the developed assay can provide information about AF as a sample for in ovo sexing and open new industrial possibilities for faster and cheaper assays.

Non-destructive egg breed separation using advanced VOC analytical techniques HSSE-GC-MS, PTR-TOF-MS, and SIFT-MS: Assessment of performance and systems' complementarity.

Over the past decade, advanced analytical techniques have been utilized to examine volatile organic compounds (VOCs) in eggs. These VOCs offer valuable insights into factors such as freshness, fertility, the presence of cracks, embryo sex, and breed. In our study, we assessed three mass spectrometry-based systems (headspace sorptive extraction gas chromatography-mass spectrometry; HSSE-GC-MS, proton transfer reaction time-of-flight-mass spectrometry; PTR-TOF-MS; and selected ion flow tube mass spectrometry; SIFT-MS) to analyze and identify VOCs present in intact hatching eggs from three distinct breeds (Dekalb white layer, Shaver brown layer, and Ross 308 broiler). The eggs were sampled on incubation days 2 and 8, to identify VOCs that distinguish breeds irrespective of incubation day. VOC measurements were conducted on 15 eggs per breed by placing them together with PDMS-coated stir bars inside inert Teflon® air sampling bags. After an accumulation period of 2 h, the headspace was analyzed using PTR-TOF-MS and SIFT-MS, while the VOCs adsorbed onto the stir bars were analyzed using GC-MS for additional compound identification. Partial least squares discriminant analysis (PLS-DA) models were constructed for breed differentiation, and variable selection was performed. As a result, 111 VOCs were identified using HSSE-GC-MS, with alcohols and esters being the most abundant. The PLS-DA models demonstrated the efficacy of breed discrimination, with the HSSE-GC-MS and the PTR-TOF-MS exhibiting the highest balanced accuracy of 95.5 % using a reduced set of 11 VOCs and 5 product ions, respectively. The SIFT-MS model had a balanced accuracy of 92.8 % with a reduced set of 11 product ions. Furthermore, complementarity was observed between HSSE-GC-MS, which primarily selected higher molecular weight VOCs, and PTR-TOF-MS and SIFT-MS. A higher correlation was found for compound abundances between the HSSE-GC-MS and the PTR-TOF-MS relative to the SIFT-MS, indicating that the PTR-TOF-MS was better suited to quantify specific compounds identified by the HSSE-GC-MS. Finally, the findings support the presence of VOCs originating from both synthetic and natural sources, highlighting the ability of the VOC analysis systems to non-destructively perform quality control and reveal differences in management practices or biological information encoded in eggs.

Trends in in ovo sexing technologies: insights and interpretation from papers and patents.

Numerous researchers and institutions have been developing in ovo sexing technologies to improve animal welfare by identifying male embryos in an early embryonic stage and disposing of them before pain perception. This review gives a complete overview of the technological approaches reported in papers and patents by performing a thorough search using Web of Science and Patstat/Espacenet databases for papers and patents, respectively. Based on a total of 49 papers and 115 patent families reported until May 2023 worldwide, 11 technology categories were defined: 6 non-optical and 5 optical techniques. Every category was described for its characteristics while assessing its potential for application. Next, the dynamics of the publications of in ovo sexing techniques in both paper and patent fields were described through growth curves, and the interest or actual status was visualized using the number of paper citations and the actual legal status of the patents. When comparing the reported technologies in papers to those in patents, scientific gaps were observed, as some of the patented technologies were not reported in the scientific literature, e.g., ion mobility and mass spectrometry approaches. Generally, more diverse approaches in all categories were found in patents, although they do require more scientific evidence through papers or industrial adoption to prove their robustness. Moreover, although there is a recent trend for non-invasive techniques, invasive methods like analyzing DNA through PCR or hormones through immunosensing are still being reported (and might continue to be) in papers and patents. It was also observed that none of the technologies complies with all the industry requirements, although 5 companies already entered the market. On the one hand, more research and harmony between consumers, industry, and governments is necessary. On the other hand, close monitoring of the market performance of the currently available techniques will offer valuable insights into the potential and expectations of in ovo sexing techniques in the poultry industry.

Sputum and salivary protein biomarkers and point-of-care biosensors for the management of COPD.

Chronic obstructive pulmonary disease (COPD) has become one of the most fatal diseases of the century considering mortality and morbidity levels worldwide. This disease is an inflammatory response to environmental stress and tobacco smoking. Although spirometry is the gold-standard diagnostic test administrated in primary and secondary care, it often exhibits low accuracy in cases of predicting disease worsening and possible bias due to the operator, patient, and conditions. Recent developments in proteomics research suggest that the presence of protein biomarkers can aid in the accurate diagnosis and prediction of disease outcomes. This review presents the cutting-edge research progress in the area of protein biomarkers towards the management of COPD. The literature review was confined to protein biomarkers in saliva and sputum because testing these bodily fluids shows great promise for point-of-care (POC) testing due to its practicality, non-invasiveness and inexpensive handling and sampling. Although it is conclusive that more studies on sputum and saliva are needed, this review studies the promising clinical value of interleukin (IL)-6 and IL-8, matrix metalloproteinase (MMP)-8 and MMP-9, C-reactive protein (CRP), tumor necrosis factor-alpha (TNF-α), and neutrophil elastase (NE). Following the critical analysis of salivary and sputum biomarkers, the recent development of POC biosensors for the multiplexed detection of biomarkers is also reported. Overall, the review aims to explore the possibility for the future development of POC sensors for chronic lung disease management utilizing clinically relevant biomarkers in saliva and sputum.

A Fluorescence Sensing Method with Reduced DNA Typing and Low-Cost Instrumentation for Detection of Sample Tampering Cases in Urinalysis.

This work presents a method to unequivocally detect urine sample tampering in cases where integrity of the sample needs to be verified prior to urinalysis. The technique involves the detection of distinct patterns of a triplex short tandem repeats system in DNA extracted from human urine. The analysis is realized with single-dye fluorescence detection and using a regular smartphone camera. The experimental results had demonstrated the efficacy of the analytical approach to obtaining distinct profiles of amplicons in urine from different sample providers. Reproducibility tests with fresh and stored urine have revealed a maximum variation in the profiles within an interval of 5 to 9%. Cases of urine sample tampering via mixture were simulated in the study, and the experiments have identified patterns of mixed genotypes from dual mixtures of urine samples. Moreover, sample adulteration by mixing a non-human fluid with urine in a volume ratio over 25% can be detected. The low cost of the approach is accompanied by the compatibility of the technique to use with different DNA sample preparation protocols and PCR instrumentation. Furthermore, the possibility of realizing the method in an integrated microchip system open great perspectives to conducting sample integrity tests at the site of urine sample reception and/or at resource-limited settings.

Design and characterisation of a graphene-based FET for ultrasensitive detection of clinical biomarkers.

The high demand for point-of-care devices for the convenient detection and follow-up of chronic diseases is posing demands to the development of novel low-cost sensors. The chronic obstructive pulmonary disease (COPD) is one of the most worldwide spread diseases, due to cigarette smoking and air pollution. Owing to the unstable and spontaneous characteristics of this disease is essential to have a sensitive, rapid, and easy-to-use device for the detection of diseases biomarkers. The research of emerging materials such as graphene monolayer and perovskite may revolutionise the field of point-of-care devices. These materials can boost the sensitivity and specificity of the detection, and therefore the detection can be performed in samples taken non-invasively, such saliva, and with less sample quantity. A graphene field effect transistor (GFET) coated with PEDOT:PSS and perovskite, bring advantages to the photodetection field, due to the unique proprieties of 2D materials and the structure of perovskite. This work presents a study of material characteristics comprising a GFET, with perspective to detect biomarkers of COPD.