Contactless Video-Based Heart Rate Monitoring of a Resting and an Anesthetized Pig.

Heart rate (HR) is a vital bio-signal that is relatively easy to monitor with contact sensors and is related to a living organism's state of health, stress and well-being. The objective of this study was to develop an algorithm to extract HR (in beats per minute) of an anesthetized and a resting pig from raw video data as a first step towards continuous monitoring of health and welfare of pigs. Data were obtained from two experiments, wherein the pigs were video recorded whilst wearing an electrocardiography (ECG) monitoring system as gold standard (GS). In order to develop the algorithm, this study used a bandpass filter to remove noise. Then, a short-time Fourier transform (STFT) method was tested by evaluating different window sizes and window functions to accurately identify the HR. The resulting algorithm was first tested on videos of an anesthetized pig that maintained a relatively constant HR. The GS HR measurements for the anesthetized pig had a mean value of 71.76 bpm and standard deviation (SD) of 3.57 bpm. The developed algorithm had 2.33 bpm in mean absolute error (MAE), 3.09 bpm in root mean square error (RMSE) and 67% in HR estimation error below 3.5 bpm. The sensitivity of the algorithm was then tested on the video of a non-anaesthetized resting pig, as an animal in this state has more fluctuations in HR than an anaesthetized pig, while motion artefacts are still minimized due to resting. The GS HR measurements for the resting pig had a mean value of 161.43 bpm and SD of 10.11 bpm. The video-extracted HR showed a performance of 4.69 bpm in MAE, 6.43 bpm in RMSE and 57% in . The results showed that HR monitoring using only the green channel of the video signal was better than using three color channels, which reduces computing complexity. By comparing different regions of interest (ROI), the region around the abdomen was found physiologically better than the face and front leg parts. In summary, the developed algorithm based on video data has potential to be used for contactless HR measurement and may be applied on resting pigs for real-time monitoring of their health and welfare status, which is of significant interest for veterinarians and farmers.

An Approach towards Motion-Tolerant PPG-Based Algorithm for Real-Time Heart Rate Monitoring of Moving Pigs.

Animal welfare remains a very important issue in the livestock sector, but monitoring animal welfare in an objective and continuous way remains a serious challenge. Monitoring animal welfare, based upon physiological measurements instead of the audio-visual scoring of behaviour, would be a step forward. One of the obvious physiological signals related to welfare and stress is heart rate. The objective of this research was to measure heart rate (beat per minutes) in pigs with technology that soon will be affordable. Affordable heart rate monitoring is done today at large scale on humans using the Photo Plethysmography (PPG) technology. We used PPG sensors on a pig's body to test whether it allows the retrieval of a reliable heart rate signal. A continuous wavelet transform (CWT)-based algorithm is developed to decouple the cardiac pulse waves from the pig. Three different wavelets, namely second, fourth and sixth order Derivative of Gaussian (DOG), are tested. We show the results of the developed PPG-based algorithm, against electrocardiograms (ECG) as a reference measure for heart rate, and this for an anaesthetised versus a non-anaesthetised animal. We tested three different anatomical body positions (ear, leg and tail) and give results for each body position of the sensor. In summary, it can be concluded that the agreement between the PPG-based heart rate technique and the reference sensor is between 91% and 95%. In this paper, we showed the potential of using the PPG-based technology to assess the pig's heart rate.

Analysis of fluoroquinolones in dusts from intensive livestock farming and the co-occurrence of fluoroquinolone-resistant Escherichia coli.

Fluoroquinolones are important therapeutics in human and veterinary medicine. This study aimed to retrospectively analyse sedimentation dusts from intensive-livestock-farming barns for fluoroquinolones and investigate the association between resistant Escherichia coli and the detected drugs. Sedimentation-dust samples (n = 125) collected (1980-2009) at 14 barns of unknown-treatment status were analysed by HPLC and tandem-mass spectroscopy to detect enrofloxacin, ciprofloxacin, marbofloxacin, and difloxacin. Recent microbiological data were included to investigate the relationship between fluoroquinolone presence and fluoroquinolone-resistant E. coli. Fifty-nine dust samples (47%) from seven barns contained fluoroquinolone residues. Up to three different fluoroquinolones were detected in pig and broiler barns. Fluoroquinolone concentrations ranged from 10-pg/mg to 46-ng/mg dust. Fluoroquinolone-resistant E. coli were isolated from four barns. Of all the dust samples, 22% contained non-susceptible isolates. Non-susceptible isolate presence in the dust was significantly associated (p = 0.0283) with detecting the drugs, while drug detection increased the odds (4-fold) of finding non-susceptible E. coli (odds ratio = 3.9877, 95% CI: 1.2854-12.3712). This retrospective study shows that fluoroquinolone usage leads to dust contamination. We conclude that farmers and animals inhale/swallow fluoroquinolones and fluoroquinolone-resistant bacteria due to drug application. Furthermore, uncontrolled drug emissions via air exhausted from the barns can be assumed.

Apoptosis in response to heat stress is positively associated with heat-shock protein 90 expression in chicken myocardial cells in vitro.

To determine heat-shock protein (Hsp)90 expression is connected with cellular apoptotic response to heat stress and its mechanism, chicken (Gallus gallus) primary myocardial cells were treated with the Hsp90 promoter, aspirin, and its inhibitor, geldanamycin (GA), before heat stress. Cellular viability, heat-stressed apoptosis and reactive oxygen species level under different treatments were measured, and the expression of key proteins of the signaling pathway related to Hsp90 and their colocalization with Hsp90 were detected. The results showed that aspirin treatment increased the expression of protein kinase B (Akt), the signal transducer and activator of transcription (STAT)-3 and p-IKKα/ß and the colocalization of Akt and STAT-3 with Hsp90 during heat stress, which was accompanied by improved viability and low apoptosis. GA significantly inhibited Akt expression and p-IKKα/ß level, but not STAT-3 quantity, while the colocalization of Akt and STAT-3 with Hsp90 was weakened, followed by lower cell viability and higher apoptosis. Aspirin after GA treatment partially improved the stress response and apoptosis rate of tested cells caused by the recovery of Akt expression and colocalization, rather than the level of STAT-3 (including its co-localization with Hsp90) and p-IKKα/ß. Therefore, Hsp90 expression has a positive effect on cellular capacity to resist heat-stressed injury and apoptosis. Moreover, inhibition of Hsp90 before stress partially attenuated its positive effects.

Aspirin upregulates αB-Crystallin to protect the myocardium against heat stress in broiler chickens.

We established in vivo and in vitro models to investigate the role of αB-Crystallin (CryAB) and assess the ability of aspirin (ASA) to protect the myocardium during prolonged heat stress. Thirty-day-old chickens were divided into three groups (n = 90): heat stress (HS, 40±1 °C); ASA(-)HS(+), 1 mg/kg ASA orally 2 h before heat stress; and ASA(+)HS(-), pretreated with aspirin, no heat stress (25 °C). Hearts were excised after 0, 1, 2, 3, 5, 7, 10, 15 and 24 h. Heat stress increased body temperature, though the ASA(-)HS(+) group had significantly higher temperatures than the ASA(+)HS(+) group at all time points. Compared to ASA(+)HS(+), the ASA(-)HS(+) group displayed increased sensitivity to heat stress. Pathological analysis revealed the ASA (+)HS(+) myocardium showed less severe changes (narrowed, chaotic fibers; fewer necrotic cells) than the ASA(-)HS(+) group (bleeding and extensive cell death). In vitro, ASA-pretreatment significantly increased primary chicken myocardial cell survival during heat stress. ELISAs indicated ASA induced CryAB in vivo to protect against heat stress-induced myocardial damage, but ASA did not induce CryAB in primary chicken myocardial cells. The mechanisms by which ASA induces the expression of CryAB in vivo and protects the myocardium during heat stress merit further research.

Lenti-siRNA Hsp60 promote bax in mitochondria and induces apoptosis during heat stress.

Hsp60 is a typical mitochondrial protein in eukaryotes, and is involved in facilitating the correction of misfolded protein back into the correct conformation. Previous, we identified aspirin-induced HSPs in response to heat stress [1]. To investigate whether Hsp60 can protect against death under heat stress, we used lenti-siRNA to knock down the expression of Hsp60. When exposed to heat stress, more apoptosis was observed with increasing exposure to heat stress, while necrosis was not affected. Furthermore, heat stress induced the loss of mitochondrial membrane potential (ΔΨm) and a significant increase of reactive oxygen species (ROS) produced in mitochondria as measured by TMRE and MitoSOXTM red. The loss of ΔΨm indicated a change in inner mitochondrial function. Real-time Quantitative PCR was used to investigate the mechanism by detecting mRNA expression profile of the inner mitochondrial membrane, including CypD, ANT, and PIC. Results showed no differences between lenti-siRNA Hsp60 and control. However, bax in the cytoplasm translocated to mitochondrial during heat stress and regulated the permeability of outer mitochondrial membrane. We hypothesize that Hsp60 can protect cardiac myocytes against apoptosis involving in outer mitochondrial membrane not the inner mitochondrial membrane under heat stress.

Prevalence, genotyping and risk factors of thermophilic Campylobacter spreading in organic turkey farms in Germany.

BACKGROUND: The need for organic food of animal origin has increased rapidly in recent years. However, effects of organic animal husbandry on food safety have not been rigorously tested especially in meat turkey flocks. This study provides for the first time an overview on the prevalence and genetic diversity of Campylobacter species (spp.) in five organic meat turkey farms located in different regions in Germany, as well as on potential risk factors of bacterial spreading. Thirty cloacal swabs as well as water samples and darkling beetles were collected from each flock and examined for the presence of Campylobacter by conventional and molecular biological methods. The isolates were genotyped by flaA-RFLP. RESULTS: Campylobacter spp. were detected in cloacal swabs in all 5 turkey flocks with prevalence ranged from 90.0 to 100 %. 13 cloacal swabs collected from birds in farm III and IV were harboured mixed population of thermophilic campylobacters. In total, from 158 Campylobacter isolated from turkeys 89 (56.33 %) were identified as C. coli and 69 (43.76 %) as C. jejuni. Three Campylobacter (2 C. jejuni and 1 C. coli) were detected in drinkers of two farms and 3 C. coli were isolated from darkling beetles of one farm. No Campylobacter were isolated from main water tanks. flaA-RFLP assay showed that turkey farms can harbour more than one genotype. In a single turkey two different genotypes could be detected. The genotypes of campylobacters isolated from water samples or beetles were identical with those isolated from turkeys. No effect was found of some environmental parameters [ammonia concentration (NH3), carbon dioxide concentration (CO2), relative humidity (RH) and air temperature)] on Campylobacter prevalence in organic turkey farms. Additionally, drinking water and darkling beetles might be considered as risk factors for the spreading of Campylobacter in turkey flocks. CONCLUSIONS: This study highlights the high prevalence and genotypic diversity of Campylobacter spp. isolated from organic turkey flocks. Further research is needed to assess other potential risk factors responsible for bacteria spreading in order to mitigate the spread of Campylobacter in organic turkey flocks by improving biosecurity control measures.

The association of Hsp90 expression induced by aspirin with anti-stress damage in chicken myocardial cells.

The protective effect of aspirin during exposure to heat stress in broiler chickens was investigated. We assayed pathological damage, expression and distribution of Hsp90 protein and hsp90 mRNA expression in chicken heart tissues after oral administration of aspirin following exposure to high temperature for varying times. Heat stress induced increases in plasma aspartate aminotransferase, creatine kinase and lactate dehydrogenase activities while causing severe heart damage, which was characterized by granular and vacuolar degeneration, nuclear shrinkage and even myocardium fragmentation in cardiac muscle fibers. After aspirin administration, myocardial cells showed fewer pathological lesions than broilers treated with heat alone. A high positive Hsp90 signal was always detected in the nuclei of myocardial cells from broilers treated with aspirin, while in myocardial cells treated with heat alone, Hsp90 in the nuclei decreased, as did that in the cytoplasm. Aspirin induced rapid and significant synthesis of Hsp90 before and at the initial phase of heat stress, and significant expression of hsp90 mRNA was stimulated throughout the experiment when compared with cells exposed to heat stress alone. Thus, specific pre-induction of Hsp90 in cardiovascular tissue was useful for resisting heat stress damage because it produced stable damage-related enzymes and fewer pathologic changes.

Characterization of Methicillin-Resistant Staphylococcus aureus Isolated from Healthy Turkeys and Broilers Using DNA Microarrays.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major human health problem and recently, domestic animals are described as carriers and possible reservoirs. Twenty seven S. aureus isolates from five turkey farms (n = 18) and two broiler farms (n = 9) were obtained by culturing of choana and skin swabs from apparently healthy birds, identified by Taqman-based real-time duplex nuc-mecA-PCR and characterized by spa typing as well as by a DNA microarray based assay which covered, amongst others, a considerable number of antibiotic resistance genes, species controls, and virulence markers. The antimicrobial susceptibility profiles were tested by agar diffusion assays and genotypically confirmed by the microarray. Five different spa types (3 in turkeys and 2 in broilers) were detected. The majority of MRSA isolates (24/27) belonged to clonal complex 398-MRSA-V. The most frequently occurring spa types were accordingly t011, t034, and t899. A single CC5-MRSA-III isolated from turkey and CC398-MRSA with an unidentified/truncated SCCmec element in turkey and broiler were additionally detected. The phenotypic antimicrobial resistance profiles of S. aureus isolated from both turkeys and broilers against 14 different antimicrobials showed that all isolates were resistant to ampicillin, cefoxitin, oxacillin, doxycycline, and tetracycline. Moreover, all S. aureus isolated from broilers were resistant to erythromycin and azithromycin. All isolates were susceptible to gentamicin, chloramphenicol, sulphonamides, and fusidic acid. The resistance rate against ciprofloxacin was 55.6% in broiler isolates and 42.1% in turkey isolates. All tetracycline resistant isolates possessed genes tetK/M. All erythromycin-resistant broiler isolates carried ermA. Only one broiler isolate (11.1%) carried genes ermA, ermB, and ermC, while 55.6% of turkey isolates possessed ermA and ermB genes. Neither PVL genes (lukF/S-PV), animal-associated leukocidin (lukM and luk-P83) nor the gene encoding the toxic shock syndrome toxin (tst1) were found in turkey and broiler isolates. In conclusion, the detection of MRSA in healthy turkeys and broilers with even additional antibiotic resistance markers is of major public health concern. The difference in antibiotic resistance and virulence markers between MRSA isolates from turkeys and broilers was addressed.

Antimicrobial susceptibilities of Campylobacter jejuni and Campylobacter coli recovered from organic turkey farms in Germany.

The popularity of food produced from animals kept under an organic regimen has increased in recent years. In Germany, turkey meat consumption has increased. Despite several studies assessing the susceptibility of campylobacters to various antibiotics in poultry, no sufficient data exists regarding the antimicrobial resistance of campylobacters in organic-reared turkeys. This study provides information about antibiotic resistance in Campylobacter isolated from turkeys reared on organic farms in Germany. Ninety-six Campylobacter strains (41 C. jejuni and 55 C. coli) were isolated from different free-range turkey flocks. In vitro antimicrobial sensitivity testing was done using a broth microdilution test, and the presence of resistance genes to antibiotics (ciprofloxacin, tetracycline) was investigated. All Campylobacter isolates from organic turkeys (n = 96) were phenotypically sensitive to gentamicin, erythromycin, streptomycin, and chloramphenicol. In this study, the antibiotic susceptibilities of C. jejuni to ciprofloxacin, tetracycline, and naladixic acid were 56.0%, 51.3%, and 56.0%, respectively. In contrast, 44.0%, 73.0%, and 74.6% of C. coli isolates were resistant to tetracycline, ciprofloxacin, and nalidixic acid, respectively. Replacement of the Thr-86→Ile in the gyrA gene, and the presence of the tet(O) gene were the mainly identified resistance mechanisms against fluoroquinolones and tetracycline, respectively.These results also reinforce the need to develop strategies and implement specific control procedures to reduce the development of antimicrobial resistance.

Air samplings in a Campylobacter jejuni positive laying hen flock.

The air in laying hen houses contains high concentrations of airborne bacteria. The numbers of these bacteria can be influenced by the efficiency of the chosen sampling method. In the presented study, AGI-30 Impingers and the Coriolis(®)µ air Sampler were compared in terms of their efficiency in sampling aerobic mesophilic bacteria in a laying hen house. Measurements were conducted in a laying hen flock with high prevalences of C. jejuni in order to investigate if culturable cells of this organism can also be detected by the applied methods. Airborne dust was also analyzed for the presence of C. jejuni specific DNA to assess the possible occurrence of non-culturable C. jejuni in the hen house air. The numbers of mesophilic airborne bacteria ranged from 8 × 10(4) - 2 × 10(6) CFU/m(-3) when sampled using AGI-30 Impingers, and from 2 × 10(5) - 4 × 10(6) CFU/m -3 when sampled using a Coriolis(®)µ air Sampler. The concentrations detected simultaneously by both devices correlated well (rPearson = 0.755), but the Coriolis(®)µ air Sampler showed a significantly higher sampling efficiency (p<0.001). Although, the within flock prevalence of C. jejuni was high during the experiments (between 70-93%), neither of the air sampling methods could detect culturable C. jejuni from the air. However, C. jejuni specific DNA was detected in 15 out of 18 airborne dust samples by mapA PCR. Based on the results, it can be concluded that airborne culturable C. jejuni were not detectable, even with an efficient air sampler, because of their low concentration. Therefore, the risk of airborne infection to poultry workers on inhaling airborne C. jejuni seems negligible. Also, the transmission of culturable C. jejuni to neighboring farms by the airborne route is unlikely. Otherwise, the detection of airborne C. jejuni specific DNA suggests that non-culturable cells could appear in the hen house air, and in future it should be verified whether sampling stress of the air sampling methods could induce the non-culturable state.

Faecal occurrence and emissions of livestock-associated methicillin-resistant Staphylococcus aureus (laMRSA) and ESbl/AmpC-producing E. coli from animal farms in Germany.

The occurrence of laMRSA (livestock-associated methicillin-resistant Staphylococcus aureus) and extended-spectrum beta-lactamase (ESBL) and/or plasmid-mediated AmpC beta-lactamase-producing (AmpC) Enterobacteriaceae in healthy livestock herds is known for some time.The spread of these bacteria in the environment is discussed critically.The object of this study was to determine the presence of these microorganisms in faeces of livestock as well as the discussion about a potential faecal emission. Therefore, faeces samples from 37 different MRSA positive livestock holdings were tested for MRSA. Furthermore, faeces samples from 50 farms with an unknown status regarding ESBL/AmpC-producing E. coli were screened for those resistant bacteria. LaMRSA was detected in samples of turkey (2/5, 40%) and broiler fattening farms (1/4, 25%) as well as in pig farms with higher detection frequencies in fattening farms (11/15, 73.3%) than in breeding farms (4/12, 33.3%). ESBL/AmpC-producing E. coli was found in all investigated eight broiler farms (100%), in nine out of 16 (56.3%) breeding pig as well as in six out of 10 (60%) dairy cattle herds and in seven of 16 (43.8%) fattening pig holdings. This presents the first detection of ESBL/AmpC-producing E. coli originating from healthy pigs, turkeys and broilers in Germany. In addition, samples of fertilized field surfaces were studied exemplarily for the presence of MRSA (n = 4) as well as ESBL/AmpC-producing E. coli (n = 2). Furthermore, slurry samples from four broiler and five pig farms were analysed for the latter. Both MRSA and ESBL/ AmpC-producing E. coli could be detected on the field surfaces, the last also in slurry samples. Faecal emissions from animal husbandry seem to be one possible route for the spread of these resistant microorganisms in the environment.

Occurrence of MRSA in air and housing environment of pig barns.

A high prevalence of MRSA among farm animals, especially pigs, has been observed for some time. However, knowledge on transmission routes of MRSA in livestock production is still scarce. Therefore, the aim of this study was to determine the occurrence of MRSA in pig house air as well as in samples from pigs and their housing environment in 27 MRSA positive pig barns of different sizes and production types. In 85.2% of all barns MRSA was detected in the animal house air. Impingement turned out to be a more sensitive sampling technique than filtration. Other environmental samples such as boot swabs or faeces showed prevalences of MRSA from 55.6% to 85.2% at sample level. The level of MRSA was 88.3% for pooled and 82.1% for single nasal swabs, in skin swabs the one was 87.7%, the others was 78.7%. Spa typing of isolates from air and nasal swabs showed predominantly spa types t011 and t034. MRSA prevalences in pigs as well as in various environmental samples were significantly higher in fattening farms than in breeding farms. This study provides good reference that there could be an airborne transmission of MRSA within pig herds indicating a potential contamination of the environment of barns.

Validation of a method for the detection of virulent Yersinia enterocolitica and their distribution in slaughter pigs from conventional and alternative housing systems.

Various methods have been described in the literature for the detection of virulent Yersinia enterocolitica in pigs. The risk factors for pig herd contamination have yet to be determined. The objective of this study was to validate a sensitive method for the detection of Y. enterocolitica and to describe the distribution of the bacteria in pigs at slaughter from conventional and alternative ("organic") housing systems. First, samples were collected from tonsils, caecum with caecal contents, and the caecal lymph nodes of 60 slaughter pigs. These samples were used to compare the sensitivity of six different laboratory culture methods either in common use or described in the literature with that of a polymerase chain reaction with two primer pairs (multiplex PCR). Then, only PCR was used to examine tonsils, caecum and caecal lymph nodes from two groups of slaughter pigs: 210 from six conventional fattening farms and 200 from three with alternative housing. The results of the multiplex PCR were positive in 28 cases. All culture methods proved inferior to PCR in sensitivity. In the second part of the study, PCR detected 36 (18%) positive pigs from alternative housing and 60 (29%) from conventional housing (p<0.05). The highest rate of Y. enterocolitica contamination was found in tonsils (11% alternative, 22% conventional; p<0.05), followed by caecum (5%, 11%) and lymph nodes (2%, 7%). The housing system appears to be one important factor in the prevalence of this common pathogen in pig herds, as we found important differences between the two systems studied here. In the conventional system, the main risk factors appeared to be sourcing pigs from different pig suppliers, use of commercial feed and transportation to slaughter.