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Ultrasonographic appearance of clinically healthy testicles and epididymides of rams
Theriogenology 59 (2003) 1959±1972
Ultrasonographic appearance of clinically healthy testicles and epididymides of rams P.G. Gouletsoua,*, G.S. Amiridisa, P.J. Crippsb, T. Lainasc, K. Deligiannisd, P. Saratsise, G.C. Fthenakisa a
Veterinary Faculty, University of Thessaly, P.O. Box 199, GR-43100 Karditsa, Greece Faculty of Veterinary Medicine, University of Liverpool, Neston, South Wirral, CH64 7TE, UK c Veterinary Service of Karditsa, GR-43100 Karditsa, Greece d National Agricultural Research Foundation, GR-43100 Karditsa, Greece e Faculty of Veterinary Medicine, Aristotle University of Thessaloniki, GR-54006 Thessaloniki, Greece b
Received 3 June 2002; received in revised form 7 August 2002; accepted 6 September 2002
Abstract The clinically healthy testicles and epididymides of 31 rams were imaged inside and outside the breeding period, by using a real time ultrasound scanner. A scanning technique based on multiple imaging planes from the caudal and the lateral surface of the genitalia was employed. Optimum imaging was achieved by using a 6.0 MHz frequency sector transducer. The testicular parenchyma appeared homogeneous with a coarse medium echo-pattern. The mediastinum testis was present in 87% of rams and 77% of testicles; its median echogenicity score was 2 (range: 0±3) among rams aged 13 months or older and 1 among rams aged less than 13 months (P 0:001). The tail of the epididymis was always clearly visible; it appeared less echoic than the testicular parenchyma and with a heterogeneous structure. The epididymal body was not visible, whilst the epididymal head was consistently partially imaged. The pampiniform plexus was clearly imaged as a dome-shaped structure masking the upper part of the head of the epididymis. The scrotal septum was seen in lateral sonograms as a highly echogenic line between the testicles. The scrotal skin formed a thick hyper-echoic peripheral structure. # 2002 Elsevier Science Inc. All rights reserved. Keywords: Sheep reproduction; Ultrasound; Testicle; Epididymis; Normal; Sheep
1. Introduction Fertility evaluation of rams, which includes an examination of the genitalia and an assessment of libido, should be carried out in ¯ocks before the beginning of the breeding * Corresponding author. Tel.: 30-24410-66071; fax: 30-24410-66077. E-mail address: [email protected] (P.G. Gouletsou).
0093-691X/02/$ ± see front matter # 2002 Elsevier Science Inc. All rights reserved. PII: S 0 0 9 3 - 6 9 1 X ( 0 2 ) 0 1 2 5 9 - 1
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season. It may also take place during selection of replacement ram-lambs, before the private purchase of a ram, or when investigating a reproductive problem in a ¯ock [1±3]. Ultrasonography presents an image of the internal architecture of soft tissues, and could provide visualization of abnormal testicular and epididymal features, thus, complimenting the clinical examination. However, the ultrasonographic appearance of testicles and epididymides of rams, although it can be particularly useful, has not been extensively documented. We are aware of one report only, that of Ahmad et al. [4], who examined only ®ve rams younger than 1 year and described the ultrasonographic features of their genitalia. The work described in this paper is the ®rst part of a project on the ultrasonography of ovine male genitalia. The objectives of this work were (i) to establish scanning techniques for ultrasound of ram genitalia and (ii) to describe the normal ultrasonographic appearance of these organs. In this way, reference standards would be available, in order to image and interpret abnormal features subsequently. 2. Materials and methods 2.1. Study design Thirty-one Karagouniko-breed rams, maintained under general husbandry conditions, were included in the study and allocated into one of the following groups: Group A with rams (n 15) aged 13 months or older, used for mating, Group B with rams (n 11) aged less than 13 months, prepared for the forthcoming breeding season, or Group C with rams (n 5) aged 13 months or older, kept for semen collection. In the ®rst part of the study, the animals were examined clinically and ultrasonographically within the reproductive season; rams into Group C, were scanned twice, before and after two ejaculations. Subsequently, in 18 of these rams (six of Group A, seven of Group B and ®ve of Group C), a second clinical and ultrasonographic examination was carried out outside the reproductive season. 2.2. Clinical examination Initially, a general health examination was performed in the rams. Clinical examination of the genitalia followed and was carried out as previously described [9], with the ram restrained in the cast position. The scrotum was observed for the presence of lesions. The testicles were palpated, compressed and compared to each other; their size, temperature, shape and consistency were assessed and recorded. They were squeezed gently throughout to detect pain reaction. Free movement of the testicles into the scrotum and lack of other masses therein were con®rmed. The epididymides were palpated, starting from the head, continuing to the body and the tail, and compared to each other. Their size and resilience were checked. The spermatic cords were palpated as far as possible into the abdomen. 2.3. Ultrasonographic imaging Ultrasonographic imaging of the testicles (testis) and the epididymides (epididymis) was carried out with the ram in the standing position, by using an ultrasound scanner
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(AMI B7, Alliance Medical, Que., Canada) ®tted with a variable frequency (2.6, 3.2, 4.0, 6.0 MHz) sector transducer. Initially, various frequencies were tested, in order to select the one providing better images; ®nally the 6.0 MHz frequency was used for the whole work. The animal was restrained by two assistants, one of whom also lifted its tail; no sedatives were used. Following scrotal hair trimming, the testicles were pulled downwards within the scrotum and maintained in that position. The testicles were not grasped, but were left pending; however, the examiner's left hand was placed on the surface opposite to the one where the transducer was applied on, in order to stabilize the organs. Initially, the left testicle and epididymis were imaged. The probe was placed on the caudal surface of the testicle along its longitudinal axis (sagittal plane) and was moved from left to right to monitor the testicular parenchyma and the body of the epididymis (corpus epididymidis); 40, 80 and 120 mm scanning depths were used. Then, the probe was moved upwards, in order to image the head of the epididymis (caput epididymidis) and the pampiniform plexus (pampiniform plexus) and downwards to image the tail of the epididymis (cauda epididymidis). In each testicle, images in up to three ®elds/levels were obtained (Fig. 1). The probe was then placed in a position perpendicular to the long axis and dorsal sections of the testicle, the epididymis and the spermatic cord (funiculus) were taken, starting from the upper part downwards (Fig. 2 section T1). Again, 40, 80 and 120 mm scanning depths were used. The whole procedure was repeated after placing the probe on the left lateral surface of the testicle (Fig. 2 section T2, Fig. 3). Subsequently, the whole procedure was repeated for the right testicle.
Fig. 1. Sagittal ultrasonographic plane: (a) in situ and (b) schematic representation. (i) Pampiniform plexus; (ii) epididymal head; (iii) testicular parenchyma; (iv) mediastinum testis; (v) epididymal tail; (vi) scrotal septum; (vii) epididymal body.
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Fig. 2. Dorsal ultrasonographic plane: section T1, caudal to cranial; section T2, lateral to medial: (a) in situ and (b) schematic representation. (i) Pampiniform plexus; (ii) epididymal head; (iii) testicular parenchyma; (iv) mediastinum testis; (v) epididymal tail; (vi) scrotal septum; (vii) epididymal body.
Finally, paired transverse sections of the genitalia were taken by moving the probe from the upper part of the genitalia downwards, by using 40±120 mm scanning depth. The testicles, epididymides and spermatic cords were compared to each other (Fig. 2 section T2).
Fig. 3. Transverse ultrasonographic plane: (a) in situ and (b) schematic representation. (i) Pampiniform plexus; (ii) epididymal head; (iii) testicular parenchyma; (iv) mediastinum testis; (v) epididymal tail; (vi) scrotal septum; (vii) epididymal body.
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We classi®ed structures observed within the testicular parenchyma into one of the following three categories; moderately echogenic structures with de®ned edges (score 1), structures highly echogenic in relation to the surrounding tissues with sharp and clear edges (score 2) and diffuse echogenic structures (score 3). 2.4. Semen collection and evaluation From Group C rams, semen was collected into an arti®cial vagina. The volume of the ejaculate was measured and the semen was quickly transferred into a test tube held at 38 8C inside a water-bath. The colour of the semen was observed and recorded. Semen motility and wave motion were scored according to the vigor category, by using the standards of Mylne et al. [5]. Semen concentration was visually assessed and scored according to the standards of Mylne et al. [5]. 2.5. Post-mortem examination After the second clinical and ultrasonographic examination, carried out outside the breeding period, seven rams (four Group A and three Group B, selected at random) were euthanatised. Structures had been observed into the parenchyma of 10 testicles of these rams. The genitalia were removed and dissected. Initially, the scrotal skin was removed. Subsequently, the testicles and epididymides were dissected transversally and divided into a caudal and cranial half; then the halves were dissected dorsally and 5-mm thick slices were cut. The features of the dissected genitalia were recorded. 2.6. Data management and analysis All data were entered into an Excel spreadsheet (Microsoft Corporation, Redmond, WA, USA). Statistical analysis was performed using Minitab (Minitab Inc., State College, PA, USA) and Epi-Info (CDC, Atlanta, GA, USA). Relationships between categorical variables were examined using the Chi-squared and Fisher Exact tests as appropriate. The Wilcoxon Signed Rank test was used to compare ®ndings in the left and the right testicles of the same animal, as well as in the two occasions of ultrasonographic examination. For comparisons between median scores, the Kruskal±Wallis test was used. Statistical signi®cance was de®ned as P < 0:05 against a two-sided null hypothesis of no difference. 3. Results 3.1. Clinical examination No problems were recorded during the clinical examination of the rams into the study. Furthermore, no clinical abnormalities were detected in their genitalia. On palpation, the testicles were ®rm with a homogeneous consistency; neither deformities, hardness or
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inclusions were palpated, nor pain reaction was recorded. The testicles were moving freely into the scrotum. The epididymides and the spermatic cords were clearly palpated. 3.2. Ultrasonographic ®ndings Optimum ultrasonographic imaging was achieved by using the 6.0 MHz frequency. In order to scan each testicle, as well as the body of the epididymis, an 80 mm scanning depth was optimum. A 40 mm scanning depth was used in order to view more details inside the parenchyma. A 40 or 80 mm depth was used to scan the spermatic cords and the epididymal head and tail. Images of both testicles and epididymides were obtained by using an 80 or 120 mm scanning depth, depending on the size of the genitalia. When examined inside the reproductive season, the parenchyma appeared homogeneous with a coarse medium echo-pattern, independently of the axis or surface used, in all the testicles scanned. The only structure observed in most of these clinically healthy testicles, was the mediastinum testis (mediastinum testis) (MT). In sagittal and transverse planes, it was observed as a central linear hyper-echoic structure of varying echogenicity, extending up to three quarters of the testicular length. In caudal to cranial dorsal planes, it was observed as a central linear structure, extending up to one ®fth of the testicular width. In lateral to medial dorsal planes, it was observed as a central focal structure on the transverse plane. If observed, this structure was always seen in all three sections imaged in a particular testicle, although it was seen better in transverse planes (Figs. 4, 5 and 6b, d). The MT was observed in 27 (87%) rams: 14 Group A, 8 Group B and 5 Group C, and in 77% of testicles examined; in 21 (68%) rams it was observed in both testicles. MT was absent from one or both testicles in 7 of the 18 rams aged less than 25 months and 2 of the 13 rams aged 25 months and over; however, this difference was not statistically signi®cant (P 0:15). There was a strong relationship between a ram having a MT on one testicle and having it on the other as well (P < 0:01), whilst there was no strong evidence (P 0:38) to suggest that the structure was present more often on the left or the right testicle. The median combined echogenicity score was 2 among rams aged 13 months or older (range: 0±3) and 1 among rams aged less than 13 months (range: 0±2). This difference was statistically signi®cant (P 0:001). The tail of the epididymis was clearly visible independently of the axis or surface used; it appeared less echoic than the testicular parenchyma and with a heterogeneous structure. The epididymal body could not be imaged. The full length of the head of the epididymis could not be always imaged clearly, because the pampiniform plexus masked its upper part; the larger part of the head of the epididymis was always clearly imaged and of echogenicity similar to that of the tail (Figs. 6a, c and 7). The pampiniform plexus was clearly imaged as a dome-shaped structure masking the upper part, though not always to the same extent, of the head of the epididymis. It was easily distinguished from the testicular parenchyma, as it appeared less echoic than that; it also contained numerous unechoic rounded or loop-like areas, representing the small spermatic veins (Figs. 6a and 7a, c). The spermatic cords were imaged as far as the transducer could be advanced; their echotexture was similar to the pampiniform plexus. The scrotal septum (septum scroti) was seen in dorsal (section T2) sonograms as a highly echogenic line between the testicles (Fig. 6d). In dorsal (section T1) sonograms it could not
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Fig. 4. Sonograms of clinically healthy rams: testicular parenchyma without mediastinum testis (a) or with moderately echogenic (b), highly echogenic (c) or diffuse (d) mediastinum testis, sagittal plane (tp: testicular parenchyma, mt: mediastinum testis, s: scrotum).
1966 P.G. Gouletsou et al. / Theriogenology 59 (2003) 1959±1972 Fig. 5. Sonograms of clinically healthy rams: testicular parenchyma without mediastinum testis (a) or with moderately echogenic (b), highly echogenic (c) or diffuse (d) mediastinum testis, dorsal plane section T1 (tp: testicular parenchyma, mt: mediastinum testis, s: scrotum).
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Fig. 6. Sonograms of clinically healthy rams, left and right genitalia imaged together: epididymal head and pampiniform plexus (a), testicular parenchyma (b), epididymal tail (c), dorsal plane section T1; testicular parenchyma and scrotal septum (d), dorsal plane section T2 (tp: testicular parenchyma, s: scrotum, eh: epididymal head, et: epididymal tail, pp: pampiniform plexus, ss: scrotal septum).
1968 P.G. Gouletsou et al. / Theriogenology 59 (2003) 1959±1972 Fig. 7. Sonograms of clinically healthy rams: epididymal head and pampiniform plexus (a), epididymal tail (b), sagittal plane; epididymal head and pampiniform plexus (c), dorsal plane section T1 (tp: testicular parenchyma, s: scrotum, eh: epididymal head, pp: pampiniform plexus, et: epididymal tail).
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Fig. 8. Sonograms of clinically healthy rams: scrotal skin and tunics, sagittal plane (tp: testicular parenchyma, s: scrotum, st: scrotal tunics).
be detected, whilst in its anatomical position a black shadow was always seen (Fig. 6b). The scrotal skin (cutis scroti) formed a thick hyper-echoic peripheral structure, encircling the genitalia imaged (Figs. 4±7). The parietal tunic (lamina parietalis) and the visceral tunic (lamina visceralis) were imaged only if ¯uid was viewed between them; if no ¯uid was obvious, then they could not be distinguished from the scrotal skin (Fig. 8). Previous ejaculation did not affect the echogenic pattern observed. In Group C rams, no difference in the ultrasonographic appearance of the genitalia was evident before and after semen collection. With the exception of the presence and appearance of the mediastinum testis as detailed above, no other difference was observed between the genitalia of the same animal. When the genitalia were imaged outside the reproductive period, the ultrasonographic appearance of the testicular and epididymal parenchyma were similar to those described above; however, they appeared slightly more echogenic. The MT was observed in both testicles of 14 (78%) rams: three of Group A, six of Group B and ®ve of Group C. In 25 of the 36 testicles scanned twice, the MT was observed on both occasions; in seven it was observed only in the ®rst, whilst in four only in the second; however, these differences were not statistically signi®cant (P 0:45). 3.3. Semen evaluation All ®ve rams of Group C provided a milky off-white coloured ejaculate with median volume of 1.5 ml (range: 1±2 ml). The median motility score of the semen was 5 (range: 4±5) and median semen concentration was 5 (range: 4±5). 3.4. Post-mortem ®ndings No gross pathological ®ndings were recorded in the testicles dissected. In all testicles, independently of the ultrasonographic image, a MT was always found during the post-
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mortem examination. This appeared as a linear structure of pearl-white colour, which appeared to be more cohesive than the surrounding testicular parenchyma. A small amount of ¯uid was always found within the vaginal cavity (cavum vaginale). 4. Discussion The clinical examination of the genitalia of rams is an integral part of preventive veterinary schemes in sheep ¯ocks. Although Bruere and West [3] consider that most abnormalities can be detected by means of this method, some concerns have been voiced recently about its accuracy [6]. Actually, in some countries, e.g. France and Spain, semen evaluation is considered to be of greater importance than clinical examination of the genitalia. However, semen evaluation is not applicable under practical conditions. It is expensive and requires access to a laboratory, which is not always near sheep farms. Furthermore, the rate of sperm production is reduced and the quality of semen is inferior outside the breeding season of sheep [7], when the ``pre-tupping'' examination of rams would be carried out; hence, misleading results might be obtained. Ultrasound scanning may be used to compliment the clinical examination of the genitalia of rams, thus, increasing its accuracy. Many veterinarians active in sheep practice use portable scanners for pregnancy detection in ewes; these may also be used for the examination of rams' genitalia. No detailed studies of the ultrasonographic appearance of the genitalia of rams have been published; in the only relevant paper, Ahmad et al. [4] described the examination of only ®ve rams younger than 1 year. We found ultrasound scanning to be useful for the examination of testicles and epididymides of rams. A clean, dry and dark place was required in order to use the ultrasonographic equipment. The procedure was accomplished with the animals in the standing position; casting them was not necessary. Restraint of the animal was accomplished without dif®culty; however, care should be taken to avoid damage of the equipment. The pendulous anatomy of the ovine male genitalia makes them easily accessible for ultrasonography. Scanning and interpretation of the ultrasonographic ®ndings did not present special dif®culties, as testicles are easy to access, do not have any other organs near them, they are symmetrical and easy to compare to each other. The use of a sector transducer was preferred, because it afforded good contact with the surface of the scrotum, capability of multi-frequency operation and better lateral resolution. Optimum ultrasonographic imaging of the testicles, which are super®cial organs, could be best effected by using a frequency of 5±7.5 MHz [8]. Smaller frequencies did not provide high quality resolution. The 6.0 MHz frequency combined good imaging of the testicular and epididymal parenchyma, with simultaneous full depth tissue penetration. The sector transducer also offered the advantage of imaging both testicles at the same sonogram. This is an important point to take into account, as comparison of the left and right testicles and epididymides of a ram is always required during their examination [2,3,9]. The linear probe and the high frequencies used by Ahmad et al. [4] did not allow concurrent imaging of the two testicles and comparison between them. With respect to the ultrasonographic appearance of the ovine genitalia, some of our ®ndings corroborate those previously reported from the study of Ahmad et al. [4].
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However, we do not agree with the conclusion of these authors that the mediastinum testis was a consistent ultrasonographic feature of ovine testicles; in our study, mediastinum testis was not always clearly imaged. Furthermore, we also do not agree that the head of the epididymis could not always be imaged, because it was masked by the pampiniform plexus; in our work, we always observed at least a large part of that structure. Various reasons might have accounted for this discrepancy of the results. For example, if the prevalence of mediastinum testis in rams aged 1 year or less is approximately 73%, as we found in our study, then the probability of ®nding it in ®ve out of ®ve randomly selected rams is around 0.21, hence the probability of one or more rams not having it is 0.79, i.e. the statistical power to detect its absence was not very great in the study of Ahmad et al. [4]. Alternatively, breed differences might have accounted for this discrepancy. The mediastinum testis is a mass of ®brous tissue containing numerous ®ne tubules in the central part of the gland; it consists of small connective tissue septa of varying thickness, which emerge from the deep surface of the tunica albuginea and extend into the interior of the testicle [10]. The mediastinum testis is found and imaged in healthy human [11], caprine [4], bovine [12], porcine [13] and canine [8] testicles. It consists of connective tissue, hence its presence and amount, re¯ected by the echogenicity score, would be expected to increase with age, as found in our study. Our ®ndings suggest that the presence or absence of mediastinum testis should not be evaluated as a ®nding of clinical signi®cance. The tail was the most consistently visualized epididymal structure and was less echoic than the testicular parenchyma; imaging of the entire head was dif®cult, whilst the body was not detectable. Similar ®ndings have been recorded by Ahmad et al. in rams [4], as well as in bulls [14] and in male goats [4]. However, in men the most clearly visible epididymal structure was the head, while the tail was non-de®nable [15±17]. This may be due to the anatomical differences between the pendulous testicles of ruminants and the spherical testicles of men. The small amount of extratesticular ¯uid detected in sonograms and recorded during the post-mortem examinations also, does not appear to be of clinical signi®cance. Similar ®ndings have been observed in men [16]. In rams maintained for semen production, ejaculation did not appear to affect the ultrasonographic pattern. The results of the present study indicate that ultrasonography is a reliable tool for the examination of the ram testicles and epididymides. By using the standards established in this work, our current research is focusing on the ultrasonographic appearance of various common testicular and epididymal abnormalities; the present ®ndings provide good standards, to which abnormal ®ndings are compared. Furthermore, the accuracy of this test over that of the clinical examination, in the detection of the most common abnormalities of the ovine male genitalia, is also investigated. References [1] Edgar DG. The place of ram testing in the sheep industry. NZ Vet J 1963;11:113±5. [2] Boundy T. Routine ram examination. In-Practice 1992;14:219±28. [3] Bruere AN, West DW. The sheep: health, disease & production. Palmerston North: Foundation for Veterinary Continuing Education of the New Zealand Veterinary Association; 1993.
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[4] Ahmad N, Noakes DE, Subandrio AL. B-mode real time ultrasonographic imaging of the testis and epididymis of sheep and goats. Vet Rec 1991;128:491±6. [5] Mylne MJA, Hunton JR, Buckrell BC. Arti®cial insemination of sheep. In: Youngquist RS, editor. Current therapy in large animal theriogenology. Philadelphia: Saunders; 1997. p. 585±94. [6] Beltsinger HJ. Make sure the pro®t is in the bag! Breeding soundness evaluation of the ram. In: Proceedings of the 5th International Sheep Veterinary Congress, Stellenbosch, South Africa; 2001. p. 250±1. [7] Malpaux B, Flores JA, Rubio MP, Pelletier J, Delgadillo JA, Bodin L, et al. Current status of seasonal reproduction control methods. In: Proceedings of the International Conference on Animal ReproductionÐ Satellite Meeting in Reproduction in Small Ruminants, Sandnes, Norway; 2000. p. 12±7. [8] Pugh CR, Konde LJ, Rark RD. Testicular ultrasound in the normal dog. Vet Radiol 1990;31:195±9. [9] Fthenakis GC, Karagiannidis A, Alexopoulos C, Brozos C, Saratsis P, Kyriakis S. Clinical and epidemiological ®ndings during ram examination in 47 ¯ocks in southern Greece. Prev Vet Med 2001;52:43±52. [10] Nickel R, Schummer A, Seiferle E. The viscera of the domestic animals. 2nd ed. Berlin: Paul Parey; 1979. [11] Lavoipierre AM. Ultrasound of the prostate and testicles. World J Surg 2000;24:198±207. [12] Hees H, Wrobel KH, Kohler T, Elmagd AA, Hees I. The mediastinum of the bovine testis. Cell Tissue Res 1989;255:29±39. [13] Cartee RE, Powe TA, Gray BW, Hudson RS, Kuhlers DL. Ultrasonographic evaluation of normal boar testicles. Am J Vet Res 1986;47:2543±8. [14] Pechman RD, Eilts BE. B-mode ultrasonography of the bull testicle. Theriogenology 1987;27:431±41. [15] Sample WF, Gottesman JE, Skinner DG, Ehrlich RM. Gray-scale ultrasound of the scrotum. Radiology 1978;127:225±8. [16] Leopold GR, Woo VL, Scheible FW, Nachtsheim D, Gosink BB. High-resolution ultrasonography of scrotal pathology. Radiology 1979;131:719±22. [17] Willscher MH, Conway JF, Daly KJ, GiGiacinto TM, Patten D. Scrotal ultrasonography. J Urol 1983;130:931±2.
Ultrasonographic appearance of clinically healthy testicles and epididymides of rams P.G. Gouletsoua,*, G.S. Amiridisa, P.J. Crippsb, T. Lainasc, K. Deligiannisd, P. Saratsise, G.C. Fthenakisa a
Veterinary Faculty, University of Thessaly, P.O. Box 199, GR-43100 Karditsa, Greece Faculty of Veterinary Medicine, University of Liverpool, Neston, South Wirral, CH64 7TE, UK c Veterinary Service of Karditsa, GR-43100 Karditsa, Greece d National Agricultural Research Foundation, GR-43100 Karditsa, Greece e Faculty of Veterinary Medicine, Aristotle University of Thessaloniki, GR-54006 Thessaloniki, Greece b
Received 3 June 2002; received in revised form 7 August 2002; accepted 6 September 2002
Abstract The clinically healthy testicles and epididymides of 31 rams were imaged inside and outside the breeding period, by using a real time ultrasound scanner. A scanning technique based on multiple imaging planes from the caudal and the lateral surface of the genitalia was employed. Optimum imaging was achieved by using a 6.0 MHz frequency sector transducer. The testicular parenchyma appeared homogeneous with a coarse medium echo-pattern. The mediastinum testis was present in 87% of rams and 77% of testicles; its median echogenicity score was 2 (range: 0±3) among rams aged 13 months or older and 1 among rams aged less than 13 months (P 0:001). The tail of the epididymis was always clearly visible; it appeared less echoic than the testicular parenchyma and with a heterogeneous structure. The epididymal body was not visible, whilst the epididymal head was consistently partially imaged. The pampiniform plexus was clearly imaged as a dome-shaped structure masking the upper part of the head of the epididymis. The scrotal septum was seen in lateral sonograms as a highly echogenic line between the testicles. The scrotal skin formed a thick hyper-echoic peripheral structure. # 2002 Elsevier Science Inc. All rights reserved. Keywords: Sheep reproduction; Ultrasound; Testicle; Epididymis; Normal; Sheep
1. Introduction Fertility evaluation of rams, which includes an examination of the genitalia and an assessment of libido, should be carried out in ¯ocks before the beginning of the breeding * Corresponding author. Tel.: 30-24410-66071; fax: 30-24410-66077. E-mail address: [email protected] (P.G. Gouletsou).
0093-691X/02/$ ± see front matter # 2002 Elsevier Science Inc. All rights reserved. PII: S 0 0 9 3 - 6 9 1 X ( 0 2 ) 0 1 2 5 9 - 1
1960
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season. It may also take place during selection of replacement ram-lambs, before the private purchase of a ram, or when investigating a reproductive problem in a ¯ock [1±3]. Ultrasonography presents an image of the internal architecture of soft tissues, and could provide visualization of abnormal testicular and epididymal features, thus, complimenting the clinical examination. However, the ultrasonographic appearance of testicles and epididymides of rams, although it can be particularly useful, has not been extensively documented. We are aware of one report only, that of Ahmad et al. [4], who examined only ®ve rams younger than 1 year and described the ultrasonographic features of their genitalia. The work described in this paper is the ®rst part of a project on the ultrasonography of ovine male genitalia. The objectives of this work were (i) to establish scanning techniques for ultrasound of ram genitalia and (ii) to describe the normal ultrasonographic appearance of these organs. In this way, reference standards would be available, in order to image and interpret abnormal features subsequently. 2. Materials and methods 2.1. Study design Thirty-one Karagouniko-breed rams, maintained under general husbandry conditions, were included in the study and allocated into one of the following groups: Group A with rams (n 15) aged 13 months or older, used for mating, Group B with rams (n 11) aged less than 13 months, prepared for the forthcoming breeding season, or Group C with rams (n 5) aged 13 months or older, kept for semen collection. In the ®rst part of the study, the animals were examined clinically and ultrasonographically within the reproductive season; rams into Group C, were scanned twice, before and after two ejaculations. Subsequently, in 18 of these rams (six of Group A, seven of Group B and ®ve of Group C), a second clinical and ultrasonographic examination was carried out outside the reproductive season. 2.2. Clinical examination Initially, a general health examination was performed in the rams. Clinical examination of the genitalia followed and was carried out as previously described [9], with the ram restrained in the cast position. The scrotum was observed for the presence of lesions. The testicles were palpated, compressed and compared to each other; their size, temperature, shape and consistency were assessed and recorded. They were squeezed gently throughout to detect pain reaction. Free movement of the testicles into the scrotum and lack of other masses therein were con®rmed. The epididymides were palpated, starting from the head, continuing to the body and the tail, and compared to each other. Their size and resilience were checked. The spermatic cords were palpated as far as possible into the abdomen. 2.3. Ultrasonographic imaging Ultrasonographic imaging of the testicles (testis) and the epididymides (epididymis) was carried out with the ram in the standing position, by using an ultrasound scanner
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(AMI B7, Alliance Medical, Que., Canada) ®tted with a variable frequency (2.6, 3.2, 4.0, 6.0 MHz) sector transducer. Initially, various frequencies were tested, in order to select the one providing better images; ®nally the 6.0 MHz frequency was used for the whole work. The animal was restrained by two assistants, one of whom also lifted its tail; no sedatives were used. Following scrotal hair trimming, the testicles were pulled downwards within the scrotum and maintained in that position. The testicles were not grasped, but were left pending; however, the examiner's left hand was placed on the surface opposite to the one where the transducer was applied on, in order to stabilize the organs. Initially, the left testicle and epididymis were imaged. The probe was placed on the caudal surface of the testicle along its longitudinal axis (sagittal plane) and was moved from left to right to monitor the testicular parenchyma and the body of the epididymis (corpus epididymidis); 40, 80 and 120 mm scanning depths were used. Then, the probe was moved upwards, in order to image the head of the epididymis (caput epididymidis) and the pampiniform plexus (pampiniform plexus) and downwards to image the tail of the epididymis (cauda epididymidis). In each testicle, images in up to three ®elds/levels were obtained (Fig. 1). The probe was then placed in a position perpendicular to the long axis and dorsal sections of the testicle, the epididymis and the spermatic cord (funiculus) were taken, starting from the upper part downwards (Fig. 2 section T1). Again, 40, 80 and 120 mm scanning depths were used. The whole procedure was repeated after placing the probe on the left lateral surface of the testicle (Fig. 2 section T2, Fig. 3). Subsequently, the whole procedure was repeated for the right testicle.
Fig. 1. Sagittal ultrasonographic plane: (a) in situ and (b) schematic representation. (i) Pampiniform plexus; (ii) epididymal head; (iii) testicular parenchyma; (iv) mediastinum testis; (v) epididymal tail; (vi) scrotal septum; (vii) epididymal body.
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Fig. 2. Dorsal ultrasonographic plane: section T1, caudal to cranial; section T2, lateral to medial: (a) in situ and (b) schematic representation. (i) Pampiniform plexus; (ii) epididymal head; (iii) testicular parenchyma; (iv) mediastinum testis; (v) epididymal tail; (vi) scrotal septum; (vii) epididymal body.
Finally, paired transverse sections of the genitalia were taken by moving the probe from the upper part of the genitalia downwards, by using 40±120 mm scanning depth. The testicles, epididymides and spermatic cords were compared to each other (Fig. 2 section T2).
Fig. 3. Transverse ultrasonographic plane: (a) in situ and (b) schematic representation. (i) Pampiniform plexus; (ii) epididymal head; (iii) testicular parenchyma; (iv) mediastinum testis; (v) epididymal tail; (vi) scrotal septum; (vii) epididymal body.
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We classi®ed structures observed within the testicular parenchyma into one of the following three categories; moderately echogenic structures with de®ned edges (score 1), structures highly echogenic in relation to the surrounding tissues with sharp and clear edges (score 2) and diffuse echogenic structures (score 3). 2.4. Semen collection and evaluation From Group C rams, semen was collected into an arti®cial vagina. The volume of the ejaculate was measured and the semen was quickly transferred into a test tube held at 38 8C inside a water-bath. The colour of the semen was observed and recorded. Semen motility and wave motion were scored according to the vigor category, by using the standards of Mylne et al. [5]. Semen concentration was visually assessed and scored according to the standards of Mylne et al. [5]. 2.5. Post-mortem examination After the second clinical and ultrasonographic examination, carried out outside the breeding period, seven rams (four Group A and three Group B, selected at random) were euthanatised. Structures had been observed into the parenchyma of 10 testicles of these rams. The genitalia were removed and dissected. Initially, the scrotal skin was removed. Subsequently, the testicles and epididymides were dissected transversally and divided into a caudal and cranial half; then the halves were dissected dorsally and 5-mm thick slices were cut. The features of the dissected genitalia were recorded. 2.6. Data management and analysis All data were entered into an Excel spreadsheet (Microsoft Corporation, Redmond, WA, USA). Statistical analysis was performed using Minitab (Minitab Inc., State College, PA, USA) and Epi-Info (CDC, Atlanta, GA, USA). Relationships between categorical variables were examined using the Chi-squared and Fisher Exact tests as appropriate. The Wilcoxon Signed Rank test was used to compare ®ndings in the left and the right testicles of the same animal, as well as in the two occasions of ultrasonographic examination. For comparisons between median scores, the Kruskal±Wallis test was used. Statistical signi®cance was de®ned as P < 0:05 against a two-sided null hypothesis of no difference. 3. Results 3.1. Clinical examination No problems were recorded during the clinical examination of the rams into the study. Furthermore, no clinical abnormalities were detected in their genitalia. On palpation, the testicles were ®rm with a homogeneous consistency; neither deformities, hardness or
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inclusions were palpated, nor pain reaction was recorded. The testicles were moving freely into the scrotum. The epididymides and the spermatic cords were clearly palpated. 3.2. Ultrasonographic ®ndings Optimum ultrasonographic imaging was achieved by using the 6.0 MHz frequency. In order to scan each testicle, as well as the body of the epididymis, an 80 mm scanning depth was optimum. A 40 mm scanning depth was used in order to view more details inside the parenchyma. A 40 or 80 mm depth was used to scan the spermatic cords and the epididymal head and tail. Images of both testicles and epididymides were obtained by using an 80 or 120 mm scanning depth, depending on the size of the genitalia. When examined inside the reproductive season, the parenchyma appeared homogeneous with a coarse medium echo-pattern, independently of the axis or surface used, in all the testicles scanned. The only structure observed in most of these clinically healthy testicles, was the mediastinum testis (mediastinum testis) (MT). In sagittal and transverse planes, it was observed as a central linear hyper-echoic structure of varying echogenicity, extending up to three quarters of the testicular length. In caudal to cranial dorsal planes, it was observed as a central linear structure, extending up to one ®fth of the testicular width. In lateral to medial dorsal planes, it was observed as a central focal structure on the transverse plane. If observed, this structure was always seen in all three sections imaged in a particular testicle, although it was seen better in transverse planes (Figs. 4, 5 and 6b, d). The MT was observed in 27 (87%) rams: 14 Group A, 8 Group B and 5 Group C, and in 77% of testicles examined; in 21 (68%) rams it was observed in both testicles. MT was absent from one or both testicles in 7 of the 18 rams aged less than 25 months and 2 of the 13 rams aged 25 months and over; however, this difference was not statistically signi®cant (P 0:15). There was a strong relationship between a ram having a MT on one testicle and having it on the other as well (P < 0:01), whilst there was no strong evidence (P 0:38) to suggest that the structure was present more often on the left or the right testicle. The median combined echogenicity score was 2 among rams aged 13 months or older (range: 0±3) and 1 among rams aged less than 13 months (range: 0±2). This difference was statistically signi®cant (P 0:001). The tail of the epididymis was clearly visible independently of the axis or surface used; it appeared less echoic than the testicular parenchyma and with a heterogeneous structure. The epididymal body could not be imaged. The full length of the head of the epididymis could not be always imaged clearly, because the pampiniform plexus masked its upper part; the larger part of the head of the epididymis was always clearly imaged and of echogenicity similar to that of the tail (Figs. 6a, c and 7). The pampiniform plexus was clearly imaged as a dome-shaped structure masking the upper part, though not always to the same extent, of the head of the epididymis. It was easily distinguished from the testicular parenchyma, as it appeared less echoic than that; it also contained numerous unechoic rounded or loop-like areas, representing the small spermatic veins (Figs. 6a and 7a, c). The spermatic cords were imaged as far as the transducer could be advanced; their echotexture was similar to the pampiniform plexus. The scrotal septum (septum scroti) was seen in dorsal (section T2) sonograms as a highly echogenic line between the testicles (Fig. 6d). In dorsal (section T1) sonograms it could not
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Fig. 4. Sonograms of clinically healthy rams: testicular parenchyma without mediastinum testis (a) or with moderately echogenic (b), highly echogenic (c) or diffuse (d) mediastinum testis, sagittal plane (tp: testicular parenchyma, mt: mediastinum testis, s: scrotum).
1966 P.G. Gouletsou et al. / Theriogenology 59 (2003) 1959±1972 Fig. 5. Sonograms of clinically healthy rams: testicular parenchyma without mediastinum testis (a) or with moderately echogenic (b), highly echogenic (c) or diffuse (d) mediastinum testis, dorsal plane section T1 (tp: testicular parenchyma, mt: mediastinum testis, s: scrotum).
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Fig. 6. Sonograms of clinically healthy rams, left and right genitalia imaged together: epididymal head and pampiniform plexus (a), testicular parenchyma (b), epididymal tail (c), dorsal plane section T1; testicular parenchyma and scrotal septum (d), dorsal plane section T2 (tp: testicular parenchyma, s: scrotum, eh: epididymal head, et: epididymal tail, pp: pampiniform plexus, ss: scrotal septum).
1968 P.G. Gouletsou et al. / Theriogenology 59 (2003) 1959±1972 Fig. 7. Sonograms of clinically healthy rams: epididymal head and pampiniform plexus (a), epididymal tail (b), sagittal plane; epididymal head and pampiniform plexus (c), dorsal plane section T1 (tp: testicular parenchyma, s: scrotum, eh: epididymal head, pp: pampiniform plexus, et: epididymal tail).
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Fig. 8. Sonograms of clinically healthy rams: scrotal skin and tunics, sagittal plane (tp: testicular parenchyma, s: scrotum, st: scrotal tunics).
be detected, whilst in its anatomical position a black shadow was always seen (Fig. 6b). The scrotal skin (cutis scroti) formed a thick hyper-echoic peripheral structure, encircling the genitalia imaged (Figs. 4±7). The parietal tunic (lamina parietalis) and the visceral tunic (lamina visceralis) were imaged only if ¯uid was viewed between them; if no ¯uid was obvious, then they could not be distinguished from the scrotal skin (Fig. 8). Previous ejaculation did not affect the echogenic pattern observed. In Group C rams, no difference in the ultrasonographic appearance of the genitalia was evident before and after semen collection. With the exception of the presence and appearance of the mediastinum testis as detailed above, no other difference was observed between the genitalia of the same animal. When the genitalia were imaged outside the reproductive period, the ultrasonographic appearance of the testicular and epididymal parenchyma were similar to those described above; however, they appeared slightly more echogenic. The MT was observed in both testicles of 14 (78%) rams: three of Group A, six of Group B and ®ve of Group C. In 25 of the 36 testicles scanned twice, the MT was observed on both occasions; in seven it was observed only in the ®rst, whilst in four only in the second; however, these differences were not statistically signi®cant (P 0:45). 3.3. Semen evaluation All ®ve rams of Group C provided a milky off-white coloured ejaculate with median volume of 1.5 ml (range: 1±2 ml). The median motility score of the semen was 5 (range: 4±5) and median semen concentration was 5 (range: 4±5). 3.4. Post-mortem ®ndings No gross pathological ®ndings were recorded in the testicles dissected. In all testicles, independently of the ultrasonographic image, a MT was always found during the post-
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mortem examination. This appeared as a linear structure of pearl-white colour, which appeared to be more cohesive than the surrounding testicular parenchyma. A small amount of ¯uid was always found within the vaginal cavity (cavum vaginale). 4. Discussion The clinical examination of the genitalia of rams is an integral part of preventive veterinary schemes in sheep ¯ocks. Although Bruere and West [3] consider that most abnormalities can be detected by means of this method, some concerns have been voiced recently about its accuracy [6]. Actually, in some countries, e.g. France and Spain, semen evaluation is considered to be of greater importance than clinical examination of the genitalia. However, semen evaluation is not applicable under practical conditions. It is expensive and requires access to a laboratory, which is not always near sheep farms. Furthermore, the rate of sperm production is reduced and the quality of semen is inferior outside the breeding season of sheep [7], when the ``pre-tupping'' examination of rams would be carried out; hence, misleading results might be obtained. Ultrasound scanning may be used to compliment the clinical examination of the genitalia of rams, thus, increasing its accuracy. Many veterinarians active in sheep practice use portable scanners for pregnancy detection in ewes; these may also be used for the examination of rams' genitalia. No detailed studies of the ultrasonographic appearance of the genitalia of rams have been published; in the only relevant paper, Ahmad et al. [4] described the examination of only ®ve rams younger than 1 year. We found ultrasound scanning to be useful for the examination of testicles and epididymides of rams. A clean, dry and dark place was required in order to use the ultrasonographic equipment. The procedure was accomplished with the animals in the standing position; casting them was not necessary. Restraint of the animal was accomplished without dif®culty; however, care should be taken to avoid damage of the equipment. The pendulous anatomy of the ovine male genitalia makes them easily accessible for ultrasonography. Scanning and interpretation of the ultrasonographic ®ndings did not present special dif®culties, as testicles are easy to access, do not have any other organs near them, they are symmetrical and easy to compare to each other. The use of a sector transducer was preferred, because it afforded good contact with the surface of the scrotum, capability of multi-frequency operation and better lateral resolution. Optimum ultrasonographic imaging of the testicles, which are super®cial organs, could be best effected by using a frequency of 5±7.5 MHz [8]. Smaller frequencies did not provide high quality resolution. The 6.0 MHz frequency combined good imaging of the testicular and epididymal parenchyma, with simultaneous full depth tissue penetration. The sector transducer also offered the advantage of imaging both testicles at the same sonogram. This is an important point to take into account, as comparison of the left and right testicles and epididymides of a ram is always required during their examination [2,3,9]. The linear probe and the high frequencies used by Ahmad et al. [4] did not allow concurrent imaging of the two testicles and comparison between them. With respect to the ultrasonographic appearance of the ovine genitalia, some of our ®ndings corroborate those previously reported from the study of Ahmad et al. [4].
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However, we do not agree with the conclusion of these authors that the mediastinum testis was a consistent ultrasonographic feature of ovine testicles; in our study, mediastinum testis was not always clearly imaged. Furthermore, we also do not agree that the head of the epididymis could not always be imaged, because it was masked by the pampiniform plexus; in our work, we always observed at least a large part of that structure. Various reasons might have accounted for this discrepancy of the results. For example, if the prevalence of mediastinum testis in rams aged 1 year or less is approximately 73%, as we found in our study, then the probability of ®nding it in ®ve out of ®ve randomly selected rams is around 0.21, hence the probability of one or more rams not having it is 0.79, i.e. the statistical power to detect its absence was not very great in the study of Ahmad et al. [4]. Alternatively, breed differences might have accounted for this discrepancy. The mediastinum testis is a mass of ®brous tissue containing numerous ®ne tubules in the central part of the gland; it consists of small connective tissue septa of varying thickness, which emerge from the deep surface of the tunica albuginea and extend into the interior of the testicle [10]. The mediastinum testis is found and imaged in healthy human [11], caprine [4], bovine [12], porcine [13] and canine [8] testicles. It consists of connective tissue, hence its presence and amount, re¯ected by the echogenicity score, would be expected to increase with age, as found in our study. Our ®ndings suggest that the presence or absence of mediastinum testis should not be evaluated as a ®nding of clinical signi®cance. The tail was the most consistently visualized epididymal structure and was less echoic than the testicular parenchyma; imaging of the entire head was dif®cult, whilst the body was not detectable. Similar ®ndings have been recorded by Ahmad et al. in rams [4], as well as in bulls [14] and in male goats [4]. However, in men the most clearly visible epididymal structure was the head, while the tail was non-de®nable [15±17]. This may be due to the anatomical differences between the pendulous testicles of ruminants and the spherical testicles of men. The small amount of extratesticular ¯uid detected in sonograms and recorded during the post-mortem examinations also, does not appear to be of clinical signi®cance. Similar ®ndings have been observed in men [16]. In rams maintained for semen production, ejaculation did not appear to affect the ultrasonographic pattern. The results of the present study indicate that ultrasonography is a reliable tool for the examination of the ram testicles and epididymides. By using the standards established in this work, our current research is focusing on the ultrasonographic appearance of various common testicular and epididymal abnormalities; the present ®ndings provide good standards, to which abnormal ®ndings are compared. Furthermore, the accuracy of this test over that of the clinical examination, in the detection of the most common abnormalities of the ovine male genitalia, is also investigated. References [1] Edgar DG. The place of ram testing in the sheep industry. NZ Vet J 1963;11:113±5. [2] Boundy T. Routine ram examination. In-Practice 1992;14:219±28. [3] Bruere AN, West DW. The sheep: health, disease & production. Palmerston North: Foundation for Veterinary Continuing Education of the New Zealand Veterinary Association; 1993.
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[4] Ahmad N, Noakes DE, Subandrio AL. B-mode real time ultrasonographic imaging of the testis and epididymis of sheep and goats. Vet Rec 1991;128:491±6. [5] Mylne MJA, Hunton JR, Buckrell BC. Arti®cial insemination of sheep. In: Youngquist RS, editor. Current therapy in large animal theriogenology. Philadelphia: Saunders; 1997. p. 585±94. [6] Beltsinger HJ. Make sure the pro®t is in the bag! Breeding soundness evaluation of the ram. In: Proceedings of the 5th International Sheep Veterinary Congress, Stellenbosch, South Africa; 2001. p. 250±1. [7] Malpaux B, Flores JA, Rubio MP, Pelletier J, Delgadillo JA, Bodin L, et al. Current status of seasonal reproduction control methods. In: Proceedings of the International Conference on Animal ReproductionÐ Satellite Meeting in Reproduction in Small Ruminants, Sandnes, Norway; 2000. p. 12±7. [8] Pugh CR, Konde LJ, Rark RD. Testicular ultrasound in the normal dog. Vet Radiol 1990;31:195±9. [9] Fthenakis GC, Karagiannidis A, Alexopoulos C, Brozos C, Saratsis P, Kyriakis S. Clinical and epidemiological ®ndings during ram examination in 47 ¯ocks in southern Greece. Prev Vet Med 2001;52:43±52. [10] Nickel R, Schummer A, Seiferle E. The viscera of the domestic animals. 2nd ed. Berlin: Paul Parey; 1979. [11] Lavoipierre AM. Ultrasound of the prostate and testicles. World J Surg 2000;24:198±207. [12] Hees H, Wrobel KH, Kohler T, Elmagd AA, Hees I. The mediastinum of the bovine testis. Cell Tissue Res 1989;255:29±39. [13] Cartee RE, Powe TA, Gray BW, Hudson RS, Kuhlers DL. Ultrasonographic evaluation of normal boar testicles. Am J Vet Res 1986;47:2543±8. [14] Pechman RD, Eilts BE. B-mode ultrasonography of the bull testicle. Theriogenology 1987;27:431±41. [15] Sample WF, Gottesman JE, Skinner DG, Ehrlich RM. Gray-scale ultrasound of the scrotum. Radiology 1978;127:225±8. [16] Leopold GR, Woo VL, Scheible FW, Nachtsheim D, Gosink BB. High-resolution ultrasonography of scrotal pathology. Radiology 1979;131:719±22. [17] Willscher MH, Conway JF, Daly KJ, GiGiacinto TM, Patten D. Scrotal ultrasonography. J Urol 1983;130:931±2.