Equine Reproductive Procedures. Группа авторов
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Название: Equine Reproductive Procedures
Автор: Группа авторов
Издательство: John Wiley & Sons Limited
Жанр: Биология
isbn: 9781119555933
isbn:
The inner swab/brush is then advanced forward through the cap/guard.
Allow the swab/brush to be in contact with the endometrium and uterine secretions for 10–15 seconds and then reverse the procedure to remove the device.
Gentle rotation of the swab/brush may be indicated to pick up cells, but care should be taken to limit “heavy‐handed” movements of the uterine culture swab as this may predispose to breaking of the end of the swab or may be associated with mild hemorrhage and contamination of the swab with blood.
Once the culture instrument has been withdrawn from the external cervical os, a cupped hand should be placed over the device to limit vaginal cellular contamination to the swab/brush.
If a single swab was used, the swab should be gently rolled over a sterile microscope slide (and can then still be used for uterine culture). If a sterile slide is not available, then a second swab may be used to obtain a uterine culture.
If a single brush is used, a swab of the brush may be made for culture and then the brush rolled gently over a microscope slide. If the material discharged is thick, a second slide is placed over the first and gently pulled apart to create a smear.
It is also common practice to collect two samples from a mare; first a swab for microbial culture and then a brush sample for endometrial cytology.
Two slides are prepared from every sample in the event that one is broken prior to evaluation or if more than one stain is to be used.
Microscope slides should be air‐dried and may be fixed to preserve cellular architecture. The slides may then be stained with a modified Wright’s stain such as Diff‐Quik®.
Additional stains that may be considered are a Gram stain or a fungus‐specific stain such as Grocott methenamine silver (GMS) stain.
Interpretation
A cytology brush may yield significantly more cells than a swab for interpretation.
Distortion of cells may occur when a swab is rolled across a slide or during the collection of cells. This is more common if the swab is smeared instead of rolled across the slide.
Note any discharge present on the sterile sleeve when the gloved arm is pulled from the vagina. There should not be any purulent or hemorrhagic discharge, only the lubricant and clear mucus should be present.
Ensure that the culture device is intact and that no part of the instrument was left within the mare’s reproductive tract. Breakage of the swab/brush is more likely when the device is placed into a dependent uterus (older broodmare) and rotated, causing undue stress on the device tip.
Note any discharge on the tip of the culture device. It is not uncommon to see very mild hemorrhage on the culture device as it rubbed against the endometrium.
Microscopic examination is performed at 400× (10× eye piece and 40× objective) magnification for uterine endometrial cells (individual cells and rafts of cells), white blood cells, debris, red blood cells, bacteria, yeast, fungal organisms, and spermatozoa. Evaluation at 1,000× (10× eye piece and 100× oil immersion objective) may be required to confirm the presence of bacterial and fungal pathogens initially detected at 400×.
Evaluate slide quality (i.e., the presence of normal uterine epithelial cells as well as the presence and relative number of inflammatory cells (neutrophils, macrophages, lymphocytes, etc.) and any bacteria/fungi). If a low number of cells are present on the slide, the cytology should be repeated to obtain a diagnostic quality slide.
The greater the ratio of neutrophils: uterine epithelial cells, the greater the degree of inflammation or endometritis.
Neutrophils are primarily associated with acute inflammation, whereas macrophages, lymphocytes, and plasma cells are more common with chronic inflammation.
The presence of inflammatory cells does not necessarily indicate infectious endometritis. Inflammatory cells may be present in cases of pneumo‐uterus, reflux of urine into the uterus, or after normal procedures such as breeding, uterine lavage, or infusion. The case history and results of other diagnostic procedures should be considered in mares with a positive uterine cytology to differentiate infectious from non‐infectious endometritis.
The absence of inflammatory cells in a sample with an adequate number of normal endometrial cells generally indicates the absence of active inflammation. However, some bacteria, such as Escherichia coli, do not stimulate a large inflammatory response, whereas other bacteria, such as Streptococcus equi subspecies zooepidemicus are associated with a significant inflammatory response.
Uterine Epithelial Cells
Uterine epithelial cells (UECs) can range from cuboidal in anestrus to tall columnar during the estrous cycle. A majority of UECs are not ciliated, but ciliated cells are often observed in cytology preparations (Figure 17.1). A range of UECs may be noted, including large rafts of cells, intact individual cells, or disrupted UECs (Figure 17.2). A large number of disrupted or degenerate UECs may indicate improper sample preparation, handling, or storage prior to staining or may be associated with a chronic uterine infection.
Figure 17.1 Ciliated tall columnar epithelial cells normally found in non‐inflammatory uterine cytology.
Debris
Debris may be classified as none/minimal (<25% of the slide), mild (25–50% of the slide), or moderate/severe (>75% of the slide). The presence of moderate to severe debris has been associated with bacterial endometritis.
Figure 17.2 A raft of normal endometrial epithelial cells in an equine uterine cytology sample.
White Blood Cells
Neutrophils are the predominant white blood cells (WBCs) identified on uterine cytology preparations (Figure 17.3). Neutrophils are 10–12 μm in diameter (about twice as large as a red blood cell (RBC)), with a single nucleus that may be indented or divided into 3–5 lobes or segments.
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