Collection of tissue and culture samples from the canine reproductive tract

Definitive diagnosis of reproductive tract infection or other disease often requires sampling of tissue, either for culture or histopathology. Indications, sample collection technique, possible side-effects and interpretation of results are reviewed. Pertinent facts include: (1) collection of uterine biopsy specimens via laparotomy was associated with higher yield of diagnostic samples and fewer side-effects than other less invasive techniques; (2) vaginal culture samples should be collected from the anterior vagina to minimize number of contaminants in the sample; (3) collection of culture samples from the anterior vagina during proestrus or estrus, in the presence of discharge originating in the uterus, was a non-invasive technique for assessment for uterine infection; (4) samples for bacterial culture from mucosal surfaces, including the vagina and penis, must be quantitated to allow interpretation, with moderate to heavy growth of any single aerobic bacterial organism relevant; (5) mycoplasma and ureaplasma are part of the normal flora of the genitourinary tract in dogs and bitches and, because most laboratories cannot give reliable quantitative results, interpretation of positive results often is difficult; (6) collection of prostatic tissue samples for cytology or culture was more likely to yield a correct diagnosis than submission of ejaculated prostatic fluid.     

Struvite calculus in the vagina of a bottlenose dolphin (Tursiops truncatus)

On 27 January 2000, a struvite calculus was observed in the vagina during necropsy of a 138-cm-long female bottlenose dolphin (Tursiops truncatus) collected from the Stono River, South Carolina (USA). Vaginal calculi have been reported in other species of cetaceans but not in bottlenose dolphins. Urinary tract infection might have been an underlying cause of the calculus. While urinary tract inflammation was not detected by light microscopic evaluation of sections of the urinary tract, it is conceivable that sufficient time had lapsed following voiding of the calculus through the urethra for urinary tract infection to have resolved. To further define the prevalence and significance of urolithiasis, prosectors of dead stranded marine mammals are encouraged to closely observe their urinary and genital tracts for calculi and to submit them for quantitative analysis.     

A multiplex biomarker approach for the diagnosis of transitional cell carcinoma from canine urine

Transitional cell carcinoma (TCC), the most common cancer of the urinary bladder in dogs, is usually diagnosed at an advanced disease stage with limited response to chemotherapy. Commercial screening tests lack specificity and current diagnostic procedures are invasive. A proof of concept pilot project for analyzing the canine urinary proteome as a noninvasive diagnostic tool for TCC identification was conducted. Urine was collected from 12 dogs in three cohorts (healthy, urinary tract infection, TCC) and analyzed using liquid chromatography tandem mass spectrometry. The presence of four proteins (macrophage capping protein, peroxiredoxin 5, heterogeneous nuclear ribonucleoproteins A2/B, and apolipoprotein A1) was confirmed via immunoblot. Of the total 379 proteins identified, 96 were unique to the TCC group. A statistical model, designed to evaluate the accuracy of this multiplex biomarker approach for diagnosis of TCC, predicted the presence of disease with 90% accuracy.     

Vestibular, vaginal and urethral relationships in spayed and intact normal dogs

The purpose of this study was to evaluate the urogenital anatomy in sound spayed and intact dogs. Fourteen intact and 12 spayed sound dogs had their lower urogenital tract evaluated with conventional vaginourethrography, computed tomography (CT) vaginourethrography and uroendoscopy under general anesthesia. Measurements for each of the three imaging modalities were made. Measurements included vaginal length, vaginal height, vaginal width, vestibule length, vestibule height, vestibule width, cingulum height, cingulum width, cingulum area, urethral length, urethral height, urethral width, angle of urethra to vestibule and angle of vagina to vestibule. Group comparisons were made between sound spayed and intact dogs. In general, most measurements were greater in intact dogs compared to spayed dogs regardless of imaging modality utilized. Group-weight interactions (P<0.01) were found on conventional vaginourethrograms (vaginal height and length and vestibule height, length and width) and CT vaginourethrograms (vaginal length, vestibule height, length and width, cingulum area, urethral width and angle of vagina to vestibule). A three-way interaction (P<0.01) for vaginal length, vaginal height and vestibule height was also identified. Vestibule length and height, cingulum height and urethral length were larger (P<0.01) with increasing dog weight and in intact dogs for both conventional and CT vaginourethrograms. No difference in vestibulovaginal ratio was observed between intact dogs and spayed dogs (P=0.0221). One of 14 intact dogs and 8 of 12 spayed dogs had vestibulovaginal ratios <0.33 on conventional vaginourethrograms. Anatomical differences in the lower urogenital tract between spayed and intact dogs emphasizes the need to establish normal reference ranges for spayed and intact dogs.     

Oral probiotics can resolve urogenital infections

We report the first clinical evidence that probiotic lactobacilli can be delivered to the vagina following oral intake. In 10 women with a history of recurrent yeast vaginitis, bacterial vaginosis (BV) and urinary tract infections, strains Lactobacillus rhamnosus GR-1 and Lactobacillus fermentum RC-14 suspended in skim milk and given twice daily for 14 days, were recovered from the vagina and identified by morphology and molecular typing within 1 week of commencement of therapy. In six cases of asymptomatic BV or intermediate BV (based upon Nugent scoring) was resolved within 1 week of therapy.     

Use of Lactobacillus to prevent infection by pathogenic bacteria

This review focuses on the use and potential of Lactobacillus to prevent infections of the urogenital and intestinal tracts. The presence and dominance of Lactobacillus in the vagina is associated with a reduced risk of bacterial vaginosis and urinary tract infections. The mechanisms appear to involve anti-adhesion factors, by-products such as hydrogen peroxide and bacteriocins lethal to pathogens, and perhaps immune modulation or signaling effects. The instillation of Lactobacillus GR-1 and B-54 or RC-14 strains into the vagina has been shown to reduce the risk of urinary tract infections, and improve the maintenance of a normal flora. Ingestion of these strains into the gut has also been shown to modify the vaginal flora to a more healthy state. In addition, these strains inhibit the growth of intestinal, as well as urogenital pathogens, colonize the gut and protect against infections as shown in mice. Other probiotic strains, such as Lactobacillus GG, have been shown to prevent and treat gastroenteritis caused by rotavirus and bacteria. Given that lactobacilli are not the dominant commensals in a gut which comprises around 10(10) organisms, much work is still needed to define the mechanisms whereby GR-1, RC-14, GG and other strains contribute to health restoration and maintenance. Such critically important studies will require the medical science community to show a willingness to turn away from pharmaceutical remedies as the only solution to health and disease.     

Ultrasonographic evaluation of the pre-pubertal development of the reproductive tract in beef heifers

To study the development of the reproductive tract in heifers, the ovaries, uterus, cervix and vagina were examined by transrectal ultrasonography every 2 weeks, from 2 to 60 weeks after birth. First ovulation occurred at 63.7 +/- 1.1 weeks of age. Ovarian dimensions increased rapidly from 2 to 14 weeks of age, and increased again after 34 weeks of age (P<0.05). The size of the largest ovarian follicles increased from 8 to 14 weeks of age, from 38 to 42 weeks of age, and finally from 52 to 60 weeks of age (P<0.05). The number of follicles > or =3 mm in diameter tended to increase from 6 to 14 weeks of age (P<0.10) and increased significantly from 6 to 60 weeks of age (P<0.05). Mean numerical pixel values of the ovarian images decreased from 4 to 26 weeks of age, and then rose to 44 weeks of age (P<0.05). Diameter of the uterine body, cervix and vagina increased from 2 to 20-24 weeks of age, and again after 32 weeks of age (P<0.05). Mean numerical pixel values for the uterus and vagina decreased initially (uterus: 4-8 weeks and vagina: 6-22 weeks of age) and then increased (uterus: 14-42 weeks and vagina: 22-32 weeks of age; P<0.05). Pixel heterogeneity showed a consistent peak at 20-22 weeks of age for the uterus, cervix and vagina (P<0.05). In summary, in the heifer calf, the marked growth of the reproductive tract in the first few months of age, and prior to first ovulation, reflects phases of increased ovarian follicle (> or =3 mm in diameter) numbers and size. Ultrasonographic image analysis revealed patterns of numerical pixel values and heterogeneity that may be useful in determining important stages of growth and differentiation of the reproductive system.     

Distribution of the usp gene in uropathogenic Escherichia coli isolated from companion animals and correlation with serotypes and size-variations of the pathogenicity island

Uncomplicated urinary tract infection (UTI) caused by uropathogenic Escherichia coli(UPEC) is a serious problem not only among humans but also in companion animals such as dogs and cats. The uropathogenic specific protein gene (usp ) is preferentially distributed in UPEC isolates from dogs and cats compared with the distribution of usp in E. coli strains from feces of healthy dogs and cats and this pattern of distribution resembles that observed in human UPEC strains. The UPEC strains from companion animals share common O serotypes like O1, O2, O4, O6, O16, O18, O22, O25 and O75 as those reported for human UPEC. The size variation of the pathogenicity island that includes usp in UPEC from dogs and cats was almost similar to those seen in human UPEC. We propose that dogs and cats are the alternative reservoirs for UPEC strains that are associated with human UTI.     

Lactobacillus rhamnosus L60, a potential probiotic isolated from the human vagina

The vagina has been increasingly viewed as an "ecosystem" whose normal microflora help protect it from invading pathogens, including those that cause urinary tract infections and sexually transmitted diseases. We tested new strains of lactobacilli as potential probiotics for maintenance of urogenital tract health, as well as prevention and therapy of urogenital infections. A strain of lactobacilli isolated from the vagina of nonpregnant, healthy, premenopausal women was identified as Lactobacillus rhamnosus L60 by 16S rDNA sequence homology. L60 was evaluated for antimicrobial activity, in vitro antibiotic resistance, autoaggregation, surface hydrophobicity, co-aggregation with other bacterial species, hydrogen peroxide (H(2)O(2)) production, and bacterial adherence. It displayed a wide spectrum of antimicrobial activity against urogenital pathogens, and resistance to antibiotics commonly prescribed for infections caused by these pathogens. L60 produced H(2)O(2), adhered to vaginal epithelial cells, co-aggregated with Escherichia coli and Candida albicans, and displayed self-aggregation. In view of these characteristics, L60 is considered a potential probiotic, and will be further evaluated for preventive and therapeutic application locally in the vaginal tract.     

Luteinizing hormone and follicle-stimulating hormone receptors and their transcribed genes (mRNA) are present in the lower urinary tract of intact male and female dogs

In dogs, one of the side effects of neutering is the development of urinary incontinence. The relationship between neutering and urinary incontinence caused by acquired urethral sphincter mechanism incompetence (USMI) has been reported. Recently, GnRH analogue treatment that suppresses elevated plasma gonadotrophin concentrations post-spaying has been successfully used in incontinent bitches. These data and the fact that non-gonadal tissues may contain receptors for LH (LHR) and FSH (FSHR) suggest that there might be a functional relationship between gonadotrophins and the lower urinary tract in dogs. This study aimed to investigate the presence of LHR and FSHR in the lower urinary tract of intact male and female dogs. Four regions of the lower urinary tract, i.e. (i) body of the bladder, (ii) neck of the bladder, (iii) proximal urethra and (iv) distal urethra were collected from 10 healthy dogs (5 males and 5 anoestrous females). In situ hybridization and immunohistochemistry were performed to characterise the presence of receptor mRNA and receptor protein. Staining was rated semi-quantitatively, incorporating both the distribution and intensity of specific staining. The distribution of receptor expression in different tissue layers (epithelium, subepithelial stroma and muscle) in each region was statistically analyzed. Luteinizing hormone receptor and FSHR mRNA and protein were present in all four regions and in three tissue layers of males and females. Irrespective of region and layer, female dogs expressed significantly higher expression for LHR mRNA (P<0.001), LHR protein (P<0.05) and FSHR protein (P<0.001). The expression of LHR and FSHR mRNA and protein was not uniform and depended on region, tissue layer and gender. The expression of LHR mRNA was higher in the bladder, compared to the urethra (P<0.05). The FSHR mRNA significantly increased from the bladder to the urethra. Protein expression for LHR and FSHR was highest in the proximal urethra (P<0.05). The overall expression for LHR and FSHR at both mRNA and protein levels was highest in the epithelium, intermediate to low in the subepithelial stroma and muscle. A significant interaction between region and tissue layer showed that mRNA and protein expression for LHR and FSHR decreased from the bladder to the urethra in the epithelium and subepithelial stroma. In contrast, it gradually increased from the bladder to the urethra in the muscle. In conclusion, the present study showed that both mRNA and protein for LHR and FSHR were expressed in the canine lower urinary tract, and the expression levels varied between genders and among regions and tissue layers. The presence of these receptors suggests that gonadotrophins have a role in the physiology and/or pathology of the lower urinary tract function in the dog.     

Urinary oxytocin as a noninvasive biomarker of positive emotion in dogs

A reliable assay based on physiological parameters that does not require subjective input from the owners is required to assess positive emotions in dogs. In addition, when viewed from an animal welfare perspective, physiological parameters should be collected in a noninvasive manner. Oxytocin (OT) is a biomarker that may be associated with a calm, relaxed state, and positive emotion. We measured the time-lapse in the concentration of plasma OT relative to urinary OT using a radioimmunoassay with sufficient sensitivity and low variability, and examined the relationship between OT and cortisol. Six dogs were injected with exogenous OT intravenously to increase the blood OT concentration. As a result, the highest concentration of urinary OT occurred 1 h after the injection, although there was little change in urinary cortisol. Moreover, to evaluate the influence of stimuli on urinary OT and cortisol, we provided three stimuli of eating food, exercising and stroking, all of which were assumed to inspire a positive emotion in dogs, and significantly increased urinary OT concentrations. Our findings indicate that urinary OT might be useful as a noninvasive and objective biomarker of positive emotion in dogs.     

Surface properties of Lactobacillus strains. II. Adherence to tissue culture surfaces]

In the second part of this study, we evaluated the adherence properties of specific strains of Lactobacillus isolated from both the human vagina and gastrointestinal tracts. Lactobacilli taken from the vagina and GI tract were tested for their adherence to A431 vaginal tissue, and to CaCo-2 cells taken from the gastrointestinal tract. The Lactobacillus strains with the most marked adherence to the respective cell lines were examined under the electron microscope. These images revealed the presence of a substance morphologically resembling slime, which probably possess unknown active properties.     

A complex intersex condition in a Holstein calf

A case of disrupted embryonic development of the genital tract in a newborn Holstein calf is described. The physical examination of the calf evidenced several abnormalities, like atresia ani, rudimentary external genitalia and caudal vertebral agenesis. On necropsy, the excised genitalia consisted of bilateral streak gonads, apparently normal uterine tubes, a fluid-filled uterus, a long vagina and a very narrow clitoris-like structure covered with a discrete skin-fold. The urinary tract seemed normal and the urethra's opening was at the vestibule-vaginal junction. A cytogenetic analysis was requested. Karyotype revealed the existence of Y chromosome material in the two X chromosomes. However, the search for the sex-determining region Y (SRY) showed that this was an apparently absent gene. The histological examination of the gonads revealed the existence of ovarian dysplasia. Uterine sections evidenced the absence of the uterine epithelium, with only sporadic caruncles. Under microscopic examination, the uterine tubes and vagina structure was normal. The external genitalia sections revealed the existence of a skin-fold covering an erectile structure surrounding the urethra, a structure more similar to a penis than to a clitoris. This is an unusual situation of gonadal dysplasia combined with genital tract anomalies in cattle, probably associated to a genetic defect.     

Influence of mucosal cell origin on the in vitro adherence of Candida albicans: Are mucosal cells from different sources equivalent?

The influence of collecting mucosal cells from various anatomical sites, and varying the date of collection and cell donor on adhesion of Candida albicans to human epithelial cells was examined by using an in vitro adherence assay. Examination of buccal mucosal cells from twenty-four donors showed statistically significant differences in the number of attached yeasts between individuals. Sex did not exert a significant influence on adhesion. Examination of buccal mucosal cells from ten donors collected on five different dates revealed that yeast attachment to mucosal epithelial cells varied significantly within subjects across time. Epithelial cells from some donors manifested greater date-to-date variations in yeast adhesion than others. Adherence of Candida to mucosal cells from three anatomical sites (mouth, vagina and urinary tract) collected from ten different donors was also tested. Yeast adherence to buccal cells was highest, lowest using urinary tract cells, while vaginal epithelium was intermediate. Adherence to mucosal cells from three sites was significantly different both within and between individuals although some subjects manifested larger variations than others. These data suggest that the in vitro adherence of Candida albicans is influenced by mucosal cell donor, date of collection and body site of origin. Mucosal cells from different sources do not appear to be equivalent in receptiveness to C. albicans and this might explain some of the discrepancies observed when adhesion studies performed by different investigators are compared. The existing need for a more uniform methodology with which to pursue studies on fungal attachment to mucosal surfaces is emphasized.     

P-fimbriae trigger mucosal responses to Escherichia coli in the human urinary tract

Uropathogenic Escherichia coli elicit a host response that determines the severity of urinary tract infection (UTI). Specific adherence mechanisms allow the bacteria to initiate this process by targeting epithelial cells in the urinary tract mucosa. Epidemiological studies show a strong association of P-fimbriae with disease severity, suggesting that adherence mediated by these organelles has a direct effect on mucosal inflammation in vivo . The present study examined the ability of P-fimbriae to induce inflammation in the human urinary tract. Patients were subjected to intravesical inoculation with a non-fimbriated E. coli strain or transformants of this strain expressing P-fimbriae. The inflammatory response was analysed as a function of P-fimbrial expression. The P-fimbriated transformants invariably caused higher interleukin (IL)-8, IL-6 and neutrophil responses in the urinary tract than the ABU strain. Furthermore, loss of P-fimbrial expression in vivo was accompanied by a return to background levels of neutrophils, IL-6 and IL-8 in individual patients. The results demonstrate that the pap sequences confer on a non-fimbriated, avirulent strain the ability to induce a host response in the human urinary tract. P-fimbriae thus fulfil the 'molecular Koch–Henle postulates' linking a single virulence factor to host response induction.     

Clinical and Demographic Characteristics of Patients with Urinary Tract Hydatid Disease

Background

Human cystic echinococcosis (CE) is caused by flatworm larvae of Echinococcus granulosus and is endemic in many parts of the world. In humans, CE cysts primarily affect the liver and pulmonary system, but can also affect the renal system. However, the clinical manifestations of renal CE can be subtle, so healthcare professionals often overlook renal CE in differential diagnosis. In this study, we examined the clinical and demographic characteristics of patients with urinary tract CE and analyzed the diagnosis and treatment procedures for this disease.

Methods

The records of 19 consecutive renal CE patients who were admitted to the First Affiliated Hospital of Xinjiang Medical University from January 1983 to April 2011 were retrospectively reviewed. In all cases, CE of the urinary tract was confirmed by pathological examination and visual inspection during surgery.

Results

Fifteen patients were males and 4 were females. The most common symptoms were non-specific lower back pain and percussion tenderness on the kidney region. All patients were followed up for 9–180 months after surgery. None of the patients experienced a recurrence of renal CE, but 4 patients experienced non-renal recurrence of hydatid disease.

Conclusions

Hydatid cysts from E. granulosus are structurally similar in the liver and urinary tract. Thus, the treatment regimen for liver CE developed by the World Health Organization/Informal Working Group on Echinococcosis (WHO/IWGE) could also be used for urinary tract CE. In our patients, the use of ultrasound, computed tomography, serology, and clinical characteristics provided a diagnostic accuracy of 66.7% to 92.3%.     

Histomorphometric study of the lower urogenital tract in pre- and post-menopausal women

In this study 22 histomorphometric parameters were used to quantify the histologic changes in the vagina, trigone of the bladder, and the proximal and distal urethra of 37 women aged between 18 and 82 years. The results of this quantitative study revealed a hormonally induced, age-independent atrophy of smooth muscle in the lamina propria of the mucosa and the longitudinal part of the muscular layer in the aforementioned anatomical sites in post-menopausal women. In the lamina propria of the mucosa of the lower urogenital tract in post-menopausal women hormonally induced quantitative changes of blood vessels were not observed. Senile atrophy of smooth muscles was only observed in the deeper part of the muscular layer of the vagina and the trigone of the urinary bladder and was less prominent than the hormonally induced smooth muscle atrophy.     

Canine cryptorchism and subsequent testicular neoplasia: case-control study with epidemiologic update

A retrospective study of 2,912 cryptorchid dogs identified 14 breeds with significantly high risk. Among six distinct closely interrelated breed groups (e.g., toy, miniature, and standard poodles), the risk in the smaller breed was always greater than that in the larger relative, suggesting that genetically influenced maldescent could be, in part, related to physical size or the rate of growth of the involved structures. Testicular tumors were diagnosed in 5.7% of the cryptorchid dogs; half had only Sertoli cell tumors, one-third had only seminomas. The relative risk for Sertoli cell tumor or seminoma was not directly related to a familial risk for cryptorchism. Using the health experience of a control population composed of male dogs with anal sac disease (N = 4,184), there is an estimated relative risk of 9.2 in cryptorchid dogs to develop a testis tumor (95% confidence interval, 5.9-14.3) and 4.2 in dogs with inguinal hernia (95% confidence interval, 1.8-9.5). Considering that the anatomical development of the genital tract, testis descent, and tunic relationships in dog are very similar to that in man, and that the associations of cryptorchism and inguinal hernia with testis neoplasms are also similar, the dog should be an excellent model system to further investigate the causes of human cryptorchism.     

The antimicrobial peptide cathelicidin protects the urinary tract against invasive bacterial infection

The urinary tract functions in close proximity to the outside environment, yet must remain free of microbial colonization to avoid disease. The mechanisms for establishing an antimicrobial barrier in this area are not completely understood. Here, we describe the production and function of the cathelicidin antimicrobial peptides LL-37, its precursor hCAP-18 and its ortholog CRAMP in epithelial cells of human and mouse urinary tract, respectively. Bacterial contact with epithelial cells resulted in rapid production and secretion of the respective peptides, and in humans LL-37/hCAP-18 was released into urine. Epithelium-derived cathelicidin substantially contributed to the protection of the urinary tract against infection, as shown using CRAMP-deficient and neutrophil-depleted mice. In addition, clinical E. coli strains that were more resistant to LL-37 caused more severe urinary tract infections than did susceptible strains. Thus, cathelicidin seems to be a key factor in mucosal immunity of the urinary tract.     

Do type 1 fimbriae promote inflammation in the human urinary tract?

Type 1 fimbriae have been implicated as virulence factors in animal models of urinary tract infection (UTI), but the function in human disease remains unclear. This study used a human challenge model to examine if type 1 fimbriae trigger inflammation in the urinary tract. The asymptomatic bacteriuria strain Escherichia coli 83972, which fails to express type 1 fimbriae, due to a 4.25 kb fimB-fimD deletion, was reconstituted with a functional fim gene cluster and fimbrial expression was monitored through a gfp reporter. Each patient was inoculated with the fim+ or fim- variants on separate occasions, and the host response to type 1 fimbriae was quantified by intraindividual comparisons of the responses to the fim+ or fim- isogens, using cytokines and neutrophils as end-points. Type 1 fimbriae did not promote inflammation and adherence was poor, as examined on exfoliated cells in urine. This was unexpected, as type 1 fimbriae enhanced the inflammatory response to the same strain in the murine urinary tract and as P fimbrial expression by E. coli 83972 enhances adherence and inflammation in challenged patients. We conclude that type 1 fimbriae do not contribute to the mucosal inflammatory response in the human urinary tract.