The influence of manual semen collection in male trained dogs (Canis familiaris), in the presence or absence of a female in estrus, on the concentrations of cortisol, oxytocin, prolactin and testosterone.

Sex pheromones are chemical substances secreted into the environment that affect the physiology and behavior of recipients. Females use these compounds during oestrus to attract males, which leads to attempts of mating. This study evaluates the influence of manual semen collection in male dogs, in the presence or absence of a female in estrus, on the blood concentrations of cortisol (CRT), oxytocin (OXT), prolactin (PRL) and testosterone (T), as hormones involved both in the physiology of reproduction and stress. Ten male dogs were used in Experiment 1 to measure the serum and plasma concentrations of the aforementioned hormones in the absence of semen collection. Subsequently in the same animals, the concentrations of these hormones were evaluated before and after semen collection in the presence (Exp. 2) or in absence of a female in estrus (Exp. 3). No significant changes in hormone concentration caused by the semen collection were found, either with, or without the presence of female in estrus. Obtained results suggest that the procedure of manual semen collection in dogs, probably due to its passive character, does not stimulate endocrine glands to secrete hormones, and the process of ejaculation is probably controlled by neural pathway. The lack of effect of semiochemical stimulation to the CRT, PRL, OXT and T level, could be caused by a short contact with female during semen collection. Further studies on involvement of the hormones during the process of natural mating, especially preceded by long courtships, similar to that observed under natural conditions, should shed a light on the physiology of mating and the connection between the endocrine system and semiochemical stimulation in dogs.

Overnight Corticosterone and Gene Expression in Mouse Hippocampus: Time Course during Resting Period.

The aim of the experiment was to test the effect of an elevated level of glucocorticoids on the mouse hippocampal transcriptome after 12 h of treatment with corticosterone that was administered during an active phase of the circadian cycle. Additionally, we also tested the circadian changes in gene expression and the decay time of transcriptomic response to corticosterone. Gene expression was analyzed using microarrays. Obtained results show that transcriptomic responses to glucocorticoids are heterogeneous in terms of the decay time with some genes displaying persistent changes in expression during 9 h of rest. We have also found a considerable overlap between genes regulated by corticosterone and genes implicated previously in stress response. The examples of such genes are Acer2, Agt, Apod, Aqp4, Etnppl, Fabp7, Fam107a, Fjx1, Fmo2, Galnt15, Gjc2, Heph, Hes5, Htra1, Jdp2, Kif5a, Lfng, Lrg1, Mgp, Mt1, Pglyrp1, Pla2g3, Plin4, Pllp, Ptgds, Ptn, Slc2a1, Slco1c1, Sult1a1, Thbd and Txnip. This indicates that the applied model is a useful tool for the investigation of mechanisms underlying the stress response.

The Case of Atypical Sexual Attractiveness in a Male Domestic Dog-A Case Study.

During the ovarian cycle in domestic dogs, females do not accept males during the first days of estrus but become attractive to males from the beginning of proestrus, with this attractiveness persisting until the end of the estrus phase. It is believed that increased estradiol is responsible for the female attractiveness to the males. In this paper we describe the case of strong, but atypical attractiveness of a castrated male to various, adult, intact males, influenced by the emitted semiochemical signals. Any significant changes in the level of hormones typically involved in the process connected with estrus and responsible for sexual arousal in the males were assessed. The case animal was a 4 year old castrated male Border Collie that was extremely attractive to various males, which presented high levels of sexual arousal, with intensive sniffing and licking of the preputial area, specific vocalization, increased salivation and, finally, mating attempts. Clinical examination of the castrated male revealed a lack of testes in the scrotum and abdominal cavity confirmed by USG. Laboratory tests indicated basal levels of estradiol, testosterone, and progesterone (15.23 pg/mL, <0.05 ng/mL, 0.25 ng/mL), and sex was confirmed via cytogenetic and molecular analysis. Chemical analysis (HS-SPME) of the urine indicated a huge similarity to the profile obtained from a bitch in estrus, with an elevated level of acetophenone, which has been previously postulated in the literature as being a characteristic of the estrus phase in female domestic dogs. This case presented very atypical sexual attractiveness, particularly when taking into account the basal levels of hormones which, according to current knowledge, are responsible for the creation of attractiveness. As a hypothesis requiring verification, we propose the idea of involvement of other hormones in the creation of incidental attractiveness or increased production of compounds responsible for attractiveness (sex pheromones) resulting from metabolic events unrelated to reproductive processes. To our knowledge it is the first described case presenting this phenomenon, which, with more detailed study, could shed new light on the process of creation of sexual attraction in the domestic dog.

Cat owners' perception on having a pet cat during the COVID-19 pandemic.

Restrictions during the COVID-19 pandemic may affect the lives of pet animals. We aimed to assess the impact of the COVID-19 pandemic on the welfare of pet cats across countries, as well as the owners' experience in relation to their cat(s). An online survey containing 21 questions was distributed internationally. Questions were related to information about the cat and its behaviour and health, changes in the household due to the pandemic, and how the owner related to the cat. The survey was completed by 324 respondents from 25 different countries. The survey showed that keeping pet cats during the COVID-19 pandemic did not create problems for the owners, except some difficulties in acquiring veterinary care. The majority (67.3%) of respondents reported no changes in their cats' behaviour. When behavioural changes were reported, they were mostly of a positive nature. Owners who took more measures in relation to the cat to prevent the spread of the virus, e.g., reduced contact, were 1.8 times more likely to report changes in the behaviour of their cats but also 3.8 times more likely to report difficulties related to the care of the cat during the pandemic. Two-third of the respondents indicated a reduction of their own psychological tensions due to having a cat during the pandemic. We concluded that the welfare of pet cats, as reported by the owners, was not adversely affected by the pandemic and the human-cat interaction had positive effects on the owners during the lockdown.

Changes in the health and behaviour of pet dogs during the COVID-19 pandemic as reported by the owners.

The COVID-19 pandemic affects human health, movement and behaviour, and this may consequently influence the behaviour and health of their pets. The aim of this study was to assess the impact of the COVID-19 pandemic on pet dogs' behaviour, as reported by their owners, as well as the owners' experience in relation to their dogs. We hypothesized that dog owners who underwent lockdown or quarantine would indicate more behavioural changes in their dogs and experience support in having a dog during the pandemic. An international online survey asked dog owners questions regarding their household, the dog, and dog-related changes during the pandemic. A total of 688 surveys, collected May-June 2020, were analysed. Respondents were from across Europe (87 %), the Americas (9%), and Asia and Australia (together 4%). Data were analysed in GLMM models with a binary distribution and country included as random variable. The main predictor variable was whether the respondent experienced lockdown (300 respondents, 44 %), quarantine (76 respondents, 11 %) or no restrictions (312 respondents, 45 %). Respondents who underwent lockdown or quarantine were 1.8 times more likely to report behavioural changes in their dogs (p = 0.02), with more negative behavioural changes in the dogs reported by respondents in lockdown than expected by chance. However, overall behavioural changes were more often positive (30 %) than negative (24 %). Respondents in lockdown were 2.6 times more likely to report health changes in their dogs (p = 0.02). The dog was perceived as a source of support during the pandemic: 65 % of the respondents indicated reduced tension due to their dog and 47 % indicated that the ability to walk the dog outside was another benefit. Advantages were reported more by respondents in lockdown and quarantine as compared to respondents who did not face these restrictions (p < 0.001). Difficulties in dog care were increased for respondents who experienced lockdown or quarantine (p < 0.01) and those who had no garden as compared to those who did (p < 0.001). One-third of the respondents took dog-related measures to prevent the spread of the coronavirus, and this was associated with more behavioural changes and more difficulties. In conclusion, lockdown, and to a lesser extent quarantine, may influence the dogs' behaviour and health, or the owners' awareness of it, and can contribute to a perceived tension-reduction in the owners.

The Role of Urine in Semiochemical Communication between Females and Males of Domestic Dog (Canis familiaris) during Estrus.

This study aimed to assess the mechanisms of semiochemical signal detection in dogs. In the first experiment, five males were exposed to volatile semiochemicals emitted by a live female in estrus and the female's urine sample collected during estrus. The odor of canine food and clean air were used as controls. In the second experiment, 25 males could directly sniff and lick the urine samples from females in estrus, from females in anestrus, from males and from humans, placed in a lineup. Sniffing, licking and salivation, as well as keeping dogs at different distances from the source of odor, were recorded in both experiments. Experiment 1 showed that food odor was sniffed by males longer than estrous urine. Volatile semiochemicals from females in estrus evoked interest in males but without visual cues did not cause overt symptoms of sexual arousal. In Experiment 2, the estrous urine evoked interest in males and provoked significantly longer sniffing. Licking accompanied by salivation was observed in all instances only during direct contact with estrous urine. The results suggest a complex character of detection of female reproductive status, in which both volatile and nonvolatile compounds emitted by females and present in female urine are involved.

Scent lineups compared across eleven countries: Looking for the future of a controversial forensic technique.

A scent lineup is generally a procedure whereby a dog's alerting behavior is used to establish that the dog detects two scents, one from a crime scene and one from a suspect, as deriving from the same person. The aim of this article is to compare methodologies of using dogs in scent lineups as a means of identifying perpetrators of crimes. It is hoped that this comparative approach, looking at countries where the method is currently or has in the past been used, will help determine what issues should be addressed in order to assure that the scent lineup will have a future as a forensic technique. Participants from eleven countries-Belgium, The Czech Republic, Finland, France, Germany, Hungary, Lithuania, The Netherlands, Poland, Russia, and the U.S.-completed a survey questionnaire regarding key aspects of the scent lineup procedures used by the police in their countries. Although there was broad overlap on certain matters, such as the use of control and zero trials, collection of decoy scents from individuals of similar gender and race as the suspect, materials for holding scent, frequency of cleaning and changing stations, and use and timing of rewards, there were significant differences in the degree of blindness required, who calls an alert (handler or experimenter), and whether handlers can work with more than one dog. The gap between recommendations and results available from the scientific literature and procedures used in police practice was greater for some countries than others, even taking into account that some scientific methodologies might be expensive or impractical given agency resources. The authors make recommendations about how to go forward if scent lineups are to remain a valid forensic technique.

Identification of putative volatile sex pheromones in female domestic dogs (Canis familiaris).

More than hundred urine samples, vaginal secretions, vulval and anal imprints as well as anal sac secretions, collected during various phases of the ovarian cycle, were evaluated with the HS-SPME/GC-MS technique. The results indicate there were differences among samples of urine collected during particular phases of the ovarian cycle. Unequivocally, the amount of carbonyl aromatic compounds, such as acetophenone (hypnone) and benzaldehyde, increased during the period of proestrus and estrus of the estrous cycle. This was accompanied by increases in methylketones, e.g., 2-octanone, 2-pentanone and 3-hexanone. Simultaneously, amounts of sulfide compounds (1-methylotiopropane; 1-methylotiobutane, 1-methylotipentane and dimethyl trisulfide) decreased during the period of estrus and abruptly increased in diestrus. These observations suggest a possible dual mechanism of interaction between males and females during and subsequent to the mating period, including the existence of both attracting and repelling signals. No significant changes were detected in samples other than urine. Further studies including a proteomic approach as well as behavioral assessments are suggested to identify the complete range of estrual semiochemical signals and to verify the semiochemical activities of identified substances.

Do olfactory behaviour and marking responses of Konik polski stallions to faeces from conspecifics of either sex differ?

The aim of the study was to assess whether adult stallions differentiate their olfactory and marking behaviour towards the excreta of their potential male rivals and mares. Four Konik polski stallions were individually exposed simultaneously to their own and each others' faeces as well as faeces of mares in oestrus vs. dioestrus. Five series of 30min observation sessions were conducted in a round pen each on two consecutive days from March to July, totaling in 5h observation per stallion. Stallions sniffed and displayed flehmen reaction towards mares' faeces significantly longer (P<0.01) than towards stallions' faeces. No significant differences were found in marking by defecations upon stallions' vs. mares' faeces. The stallions urinated exclusively on mares' faeces. Sniffing, flehmen, defecations and urinations performed towards faeces of mares in oestrus vs. dioestrus did not differ significantly. The results showed that stallions differentiate their olfactory behaviour towards excreta of conspecifics of different sex but perhaps not towards faeces of females in different reproductive status. This for the first time establishes that stallions exhibit different marking behaviour upon stallions' vs. mares' faeces. It is hypothesized that marking of other stallions' faeces by defecation may advertise presence of the stallion, whereas marking of mares' faeces by urination may serve to mask the presence of a mare to prevent potential rivals from locating the mare in an area where mares' faeces are found.

Study of the art: canine olfaction used for cancer detection on the basis of breath odour. Perspectives and limitations.

Experimental studies using trained dogs to identify breath odour markers of human cancer, published in the recent decade, have been analyzed and compared with the authors' own results. Particular published studies differ as regards the experimental setup, kind of odour samples (breath, urine, tumor tissue, serum), sample collection methods, dogs' characteristics and dog training methods as well as in results presented in terms of detection sensitivity and specificity. Generally it can be stated that trained dogs are able to distinguish breath odour samples typical for patients with lung cancer and other cancers from samples typical for healthy humans at a 'better than by chance' rate. Dogs' indications were positively correlated with content of 2-pentanone and ethyl acetate (r = 0.97 and r = 0.85 respectively) and negatively correlated with 1-propanol and propanal in breath samples (r = -0.98 and -0.87 respectively). The canine method has some advantages as a potential cancer-screening method, due to its non-invasiveness, simplicity of odour sampling and storage, ease of testing and interpretation of results and relatively low costs. Disadvantages and limitations of this method are related to the fact that it is still not known exactly to which chemical compounds and/or their combinations the dogs react. So far it could not be confirmed that dogs are able to sniff out early preclinical cancer stages with approximately the same accuracy as already diagnosed cases. The detection accuracy may vary due to failure in conditioning of dogs, decreasing motivation or confounding factors. The dogs' performance should be systematically checked in rigorous double-blind procedures. Recommendations for methodological standardization have been proposed.

Analysis of exhaled breath for disease detection.

Breath analysis is a young field of research with great clinical potential. As a result of this interest, researchers have developed new analytical techniques that permit real-time analysis of exhaled breath with breath-to-breath resolution in addition to the conventional central laboratory methods using gas chromatography-mass spectrometry. Breath tests are based on endogenously produced volatiles, metabolites of ingested precursors, metabolites produced by bacteria in the gut or the airways, or volatiles appearing after environmental exposure. The composition of exhaled breath may contain valuable information for patients presenting with asthma, renal and liver diseases, lung cancer, chronic obstructive pulmonary disease, inflammatory lung disease, or metabolic disorders. In addition, oxidative stress status may be monitored via volatile products of lipid peroxidation. Measurement of enzyme activity provides phenotypic information important in personalized medicine, whereas breath measurements provide insight into perturbations of the human exposome and can be interpreted as preclinical signals of adverse outcome pathways.

Efficacy of drug detection by fully-trained police dogs varies by breed, training level, type of drug and search environment.

Some recent publications claim that the effectiveness of police canine drug detection is uncertain and likely minimal, and that the deterrent effect of dogs on drug users is low. It is also claimed that more scientific evidence is needed to demonstrate to what extent dogs actually detect drugs. The aim of this research was to assess experimentally, but in actual training and testing environments used by the Polish police, how effective dogs trained by the police were at illicit substance detection depending on factors such as type of drug, dog breed, dog experience with the searching site, and drug odor residuals. 68 Labrador retrievers, 61 German shepherds, 25 Terriers and 10 English Cocker Spaniels, of both sexes in each breed, were used. Altogether 1219 experimental searching tests were conducted. On average, hidden drug samples were indicated by dogs after 64s searching time, with 87.7% indications being correct and 5.3% being false. In 7.0% of trials dogs failed to find the drug sample within 10min. The ranking of drugs from the easiest to the most difficult to detect was: marijuana, hashish, amphetamine, cocaine, heroin. German shepherds were superior to other breeds in giving correct indications while Terriers showed relatively poor detection performance. Dogs were equally efficient at searching in well-known vs. unknown rooms with strange (i.e., non-target novelty) odors (83.2% correct indications), but they were less accurate when searching outside or inside cars (63.5% and 57.9% correct indications respectively). During police examination trials the dogs made more false alerts, fewer correct indications and searching time was longer compared to the final stage of the training. The drug odor may persist at a site for at least 48h. Our experiments do not confirm the recent reports, based on drug users' opinions, of low drug detection efficiency. Usefulness of drug detection dogs has been demonstrated here, even if their effectiveness may not be 100%, but different factors have to be taken into consideration to assure maximum effectiveness.

Identification of volatile lung cancer markers by gas chromatography-mass spectrometry: comparison with discrimination by canines.

In this work, a chromatographic method for identification of volatile organic compounds was compared with canine recognition. Gas chromatography and mass spectrometry (GC-TOF MS) were used for determination of concentrations of trace gases present in human breath. The technique enables rapid determination of compounds in human breath, at the parts per billion level. Linear correlations were from 0.83-234.05 ppb, the limit of detection was the range 0.31-0.75 ppb, and precision, expressed as relative standard deviation (RSD), was less than 10.00 %. Moreover, trained dogs are able to discriminate breath samples of patients with diagnosed cancer. We found a positive correlation between dog indications and the ethyl acetate and 2-pentanone content of breath (r = 0.85 and r = 0.97, respectively). The methods presented for detection of lung cancer markers in exhaled air could be used as a potential non-invasive tool for screening. In addition, the canine method is relatively simple and inexpensive in comparison with chromatography.

Do trained dogs discriminate individual body odors of women better than those of men?

Scent identification lineups using dogs are a potentially valuable forensic tool, but have been dismissed by some critics because of cases where a false identification was shown to have occurred. It is not known, however, why dogs appear to make more false indications to the odors of some persons than of others. In this study, human genders were compared as to the degree their individual odors are distinguishable or "attractive" to dogs. Six dogs were trained to smell an individual's hand odor sample and then find the matching hand odor sample in a lineup of five odors. Using one-gender lineups and two-gender lineups with different gender ratios, it was found that dogs trained for the study identified individual women's hand odors more accurately than those of men. It is hypothesized that this is either because of differences in chemical compounds making discrimination of women's odors easier, or because of greater "odor attractiveness" of women's scents to dogs.

Canine olfactory receptor gene polymorphism and its relation to odor detection performance by sniffer dogs.

The outstanding sensitivity of the canine olfactory system has been acknowledged by using sniffer dogs in military and civilian service for detection of a variety of odors. It is hypothesized that the canine olfactory ability is determined by polymorphisms in olfactory receptor (OR) genes. We investigated 5 OR genes for polymorphic sites which might affect the olfactory ability of service dogs in different fields of specific substance detection. All investigated OR DNA sequences proved to have allelic variants, the majority of which lead to protein sequence alteration. Homozygous individuals at 2 gene loci significantly differed in their detection skills from other genotypes. This suggests a role of specific alleles in odor detection and a linkage between single-nucleotide polymorphism and odor recognition efficiency.

Diagnostic accuracy of canine scent detection in early- and late-stage lung and breast cancers.

BACKGROUND: Lung and breast cancers are leading causes of cancer death worldwide. Prior exploratory work has shown that patterns of biochemical markers have been found in the exhaled breath of patients with lung and breast cancers that are distinguishable from those of controls. However, chemical analysis of exhaled breath has not shown suitability for individual clinical diagnosis. METHODS: The authors used a food reward-based method of training 5 ordinary household dogs to distinguish, by scent alone, exhaled breath samples of 55 lung and 31 breast cancer patients from those of 83 healthy controls. A correct indication of cancer samples by the dogs was sitting/lying in front of the sample. A correct response to control samples was to ignore the sample. The authors first trained the dogs in a 3-phase sequential process with gradually increasing levels of challenge. Once trained, the dogs' ability to distinguish cancer patients from controls was then tested using breath samples from subjects not previously encountered by the dogs. The researchers blinded both dog handlers and experimental observers to the identity of breath samples. The diagnostic accuracy data reported were obtained solely from the dogs' sniffing, in double-blinded conditions, of these breath samples obtained from subjects not previously encountered by the dogs during the training period. RESULTS: Among lung cancer patients and controls, overall sensitivity of canine scent detection compared to biopsy-confirmed conventional diagnosis was 0.99 (95% confidence interval [CI], 0.99, 1.00) and overall specificity 0.99 (95% CI, 0.96, 1.00). Among breast cancer patients and controls, sensitivity was 0.88 (95% CI, 0.75, 1.00) and specificity 0.98 (95% CI, 0.90, 0.99). Sensitivity and specificity were remarkably similar across all 4 stages of both diseases. CONCLUSION: Training was efficient and cancer identification was accurate; in a matter of weeks, ordinary household dogs with only basic behavioral "puppy training" were trained to accurately distinguish breath samples of lung and breast cancer patients from those of controls. This pilot work using canine scent detection demonstrates the validity of using a biological system to examine exhaled breath in the diagnostic identification of lung and breast cancers. Future work should closely examine the chemistry of exhaled breath to identify which chemical compounds can most accurately identify the presence of cancer.

Relationships between sexual behaviour, dominant follicle area, uterus ultrasonic image and pregnancy rate in mares of two breeds differing in reproductive efficiency.

Weak or equivocal expression of oestrous behaviour, related to different level of mares' reactivity, may cause problems in oestrus detection and thus influence the reproductive efficiency. The aim of the study was to test whether a breed characterised by low pregnancy rate and high emotional reactivity (Thoroughbreds) differs in oestrous behaviour from a primitive breed with higher reproductive efficiency (Koniks). Additionally, the follicle size was examined to determine how it influences the intensity of oestrus and uterine ultrasonic image and whether the pregnancy rate differs in relation to the intensity of oestrous behaviour and the size of the dominant follicle(s) area on day preceding ovulation in both breeds. During four reproductive seasons the behaviour of 20 Konik polski (K) and 37 Thoroughbred (T) was observed during daily teasing. Simultaneously, the ultrasonic examinations of the reproductive system were carried out. The behaviour of mares was quantified by scoring on an 8-point scale (behavioural score, BS), according to increasing sexual receptivity. Cross-sectional follicular area (FA) was taken as a product of the two largest perpendicular follicular diameters and mean values for each breed were estimated on 1693 and 1982 mm2 for K and T mares, respectively (P < 0.05). Mares were classified according to the pooled area of dominant follicle (FA) during the preovulatory period: group A (FA < or = breed mean) and group B (FA > breed mean). Uterus image (UI) was scored (1-5) according to the increasing uterine echogenicity. The BS was higher (P < 0.01) in K mares (BS = 5.19) than in T mares (BS = 4.04). The BS was significantly related to increasing follicular area (FA). There was no breed difference in uterus image (UI) score. However, significant regression of UI on FA was found in K mares. The intensity of oestrus was positively related with UI (r = 0.29; P < 0.01) only in K mares, no such relationship was found in T mares. The pregnancy rate was significantly higher for K mares (88.5%) than for T mares (46.0%) and lower for T mares with less intense oestrous behaviour (29.4%) as compared to T mares with more intense oestrus (60.0%). No differences in pregnancy rate was found in mares belonging to A or B group of follicular area. The ascertained weaker oestrous behaviour in Thoroughbred was related to lower pregnancy rate. It is hypothesised that oestrus intensity may be the result of breed differences in the response of the neural system to follicular secretions, or may be an effect of higher incidence of multiple non-synchronic ovulation and/or higher sensitivity to stress in Thoroughbred mares.

Effectiveness of divergent selection for open-field activity in rabbits and correlated response for body weight and fertility.

Two lines deriving from the same rabbit stock were selected for 8 generations for high (H) or low (L) locomotor activity score in the open field (OFS). The divergent selection was most effective up to the 3rd generation in the H line and up to the 4th generation in the L line. In further generations a decrease of OFS in the H line and a "floor effect" (OFS = 0) in the L line were observed. The mean OFS increased significantly in consecutive trials in the H line, whereas this increase was non-significant in the L line. There was a negative and very high correlation between the latency to enter the open field and the OFS (-0.95 and -0.98 for the H and L line, respectively). The realized heritability of the OFS was 0.46 and 0.23 in generations 0-3 within the L and H line, respectively, and 0.44 and -0.06 in generations 0-8. As calculated on the basis of divergent selection, the heritability was 0.31 and 0.15 for generations 0-3 and 0-8, respectively. The L rabbits were heavier shortly before (4th wk, P < 0.001) and after (8th wk, P < 0.01) weaning, than those of the H line, whereas the H rabbits grew faster (P < 0.05) between the 4th and 20th wk of age. There was a tendency for decreasing weight gains in consecutive generations. Generally, a lower percentage of H females delivered litters than those of the L line, but this was due to a very low percentage of such females in the 3rd and 6th generations. It can be assumed that H and L lines represent different, i.e., active and passive, coping strategies. These lines of rabbits offer increased possibilities for physiologically and ethologically oriented studies, e.g., on the welfare of caged animals.