OBJECTIVE:To describe an ultrasound-guided technique for anaesthesia of the pudendal nerve with evaluation of the anatomical spread of a bupivacaine-methylene blue solution in sheep. STUDY DESIGN:Prospective, experimental study. ANIMALS:A total of seven adult experimental sheep and one sheep cadaver. METHODS:The study had two phases. First, the anatomy and ultrasound imaging of the gluteal region, caudal to the region of tuber coxae was reviewed in one sheep cadaver. Second, in seven healthy sedated sheep, a linear transducer (8-13 MHz) was positioned on the lateral area of the hip to identify pudendal vessels at the level of the ischial spine ventral to the sacrotuberous ligament. The right and left pudendal nerves area were injected with 0.1 mL kg-1 of a 1:1 solution of bupivacaine-methylene blue. The ultrasound image quality was considered as excellent, good, or poor. Following euthanasia, both pudendal areas were dissected and the staining of the nerves assessed, expressed as the percentage of success (%) and compared with the chi-square test. RESULTS:The relevant ultrasound landmarks were visualized in both areas of all sheep, with an excellent visualization of the landmarks before and during injection in most cases (13/14). On dissection, methylene blue dispersion was observed surrounding the pudendal nerve, with at least 1 cm length in 13/14 cases (93%; the remainder partially stained). No evidence of dye was found around the sciatic nerve in any case nor was there any evidence of haematoma or faecal contamination within the punctured area. CONCLUSIONS AND CLINICAL RELEVANCE:Ultrasound-guided injection with 0.1 mL kg-1 in close proximity to the pudendal nerve at the ischial spine level resulted in a high success rate of nerve staining. This procedure may be a suitable technique for pudendal nerve blockade providing analgesia for common reproductive or perineal surgical procedures in sheep.

Introduction:Bovine Respiratory Disease (BRD) is a multifactorial condition and a major health and economic concern in dairy production. Methods:This study aimed to determine the prevalence of five key bacterial pathogens-Mannheimia haemolytica, Pasteurella multocida, Trueperella pyogenes, Mycoplasma bovis, and Histophilus somni-in Spanish dairy heifers and to evaluate the influence of seasonality, geographical location, farm size, and antibiotic use. In 2017, samples (deep nasopharyngeal swabs, transtracheal aspirates, and blood) were collected from 855 heifers (<12 months old) in 50 farms across Spain. Bacterial isolation and serological testing (ELISA) were performed. Results:Mycoplasma spp. showed the highest overall prevalence (26.7% at the individual level by culture; 75.7% of cultured farms), with PCR confirming M. bovis in 89% of Mycoplasma-positive farms. Serology revealed 16.3% individual-level positivity for M. bovis and 63% farm-level positivity. T. pyogenes was also notable, detected in 6.0% of animals (45% of farms). Lower isolation rates were observed for M. haemolytica (2.5%) and P. multocida (3.5%), while H. somni was not detected. Larger farm size, winter season, and certain regions were significantly associated with higher prevalence of M. bovis and T. pyogenes. Discussion:These findings underscore the need for improved management practices-such as better ventilation, reduced stocking density, and targeted vaccination-to mitigate BRD risk in high-prevalence settings.

Bovine abortions, often caused by infectious agents like Neospora caninum, inflict substantial economic losses. Studying host-pathogen interactions in pregnant cows is challenging, and existing cell cultures lack the intricate complexity of real tissues. To bridge the gap between in vitro and in vivo models, we explored the use of cryopreserved bovine placental explants. Building upon our successful development of protocols for obtaining, culturing, and cryopreserving sheep placental explants, we applied these methods to bovine tissues. Here, we compared fresh and cryopreserved bovine explants, evaluating their integrity and functionality over culture time. Additionally, we investigated their susceptibility to N. caninum infection. Our findings revealed that bovine explants deteriorate faster in culture compared to sheep explants, exhibiting diminished viability and function. Cryopreservation further exacerbated this deterioration. While fresh explants were successfully infected with N. caninum, parasite replication was limited. Notably, cryopreservation reduced infection efficiency. This pioneering work paves the way for developing ex vivo models to study reproductive pathogens in cattle. However, further optimization of the model is essential. These improved models will have the potential to significantly reduce the reliance on animals in research.

Congenital toxoplasmosis in humans and in other mammalian species, such as small ruminants, is a well-known cause of abortion and fetal malformations. The calcium-dependent protein kinase 1 (CDPK1) inhibitor BKI-1748 has shown a promising safety profile for its use in humans and a good efficacy against Toxoplasma gondii infection in vitro and in mouse models. Ten doses of BKI-1748 given every other day orally in sheep at 15 mg/kg did not show systemic or pregnancy-related toxicity. In sheep experimentally infected at 90 days of pregnancy with 1000 TgShSp1 oocysts, the BKI-1748 treatment administered from 48 hours after infection led to complete protection against abortion and congenital infection. In addition, compared to infected/untreated sheep, treated sheep showed a drastically lower rectal temperature increase and none showed IgG seroconversion throughout the study. In conclusion, BKI-1748 treatment in pregnant sheep starting at 48 hours after infection was fully effective against congenital toxoplasmosis. Toxoplasmosis is a zoonotic disease and the first-line treatments for congenital Toxoplasma gondii infections in humans lack sufficient safety and efficacy. BKI-1748 is a lead compound and here we demonstrate its favorable safety and efficacy in sheep.

The histological findings associated to Besnoitia besnoiti infection were exhaustively studied in target tissues from experimentally and chronically infected calves. Calves were inoculated with 10(6) bradyzoites via intravenous, subcutaneous and intradermal route. Visible pathognomonic sclera cysts were observed in all infected animals. Tissue cysts were more abundant and lesions were more frequent in calves inoculated via intradermal. The most parasitized tissues were skin, including scrotum (40.81% of positive samples), nostril and nasal turbinate. Tissue cysts were already fully developed as the average tissue cyst diameter was 181.20 Microscopic lesions were mainly detected in skin samples, followed by reproductive and upper respiratory tracts. Mild lesions compatible with both acute (thrombus, oedema and inflammation) and chronic besnoitiosis (skin lesions, hyperkeratosis and dilated sweat glands) coexisted. Vascular damage and inflammation were more frequently observed in skin (including scrotum) followed by testicular parenchyma, epididymis and pampiniform plexus. Histological findings evidenced a subclinical chronic besnoitiosis.

Early Neospora caninum infection dynamics were investigated in pregnant heifers intravenously inoculated with PBS (G-Control) or 107 tachyzoites of high (G-NcSpain7)- or low (G-NcSpain1H)-virulence isolates at 110 days of gestation. Serial culling at 10 and 20 days post-infection (dpi) was performed. Fever was detected at 1 dpi in both infected groups (P < 0.0001), and a second peak was detected at 3 dpi only in G-NcSpain7 (P < 0.0001). At 10 dpi, Nc-Spain7 was detected in placental samples from one animal related to focal necrosis, and Nc-Spain7 transmission was observed, although no foetal lesions were associated with this finding. The presence of Nc-Spain1H in the placenta or foetuses, as well as lesions, were not detected at 10 dpi. At 20 dpi, G-NcSpain7 animals showed almost 100% positive placental tissues and severe focal necrosis as well as 100% transmission. Remarkably, foetal mortality was detected in two G-NcSpain7 heifers. Only one animal from G-NcSpain1H presented positive placental samples. No foetal mortality was detected, and lesions and parasite transmission to the foetus were not observed in this group. Finally, 100% of G-NcSpain7 heifers at 20 dpi presented specific antibodies, while only 60% of G-NcSpain1H animals presented specific antibodies at 20 dpi. In addition, earlier seroconversion in G-Nc-Spain7 was observed. In conclusion, tachyzoites from Nc-Spain7 reached the placenta earlier and multiplied, leading to lesion development, transmission to the foetus and foetal mortality, whereas Nc-Spain1H showed delayed infection of the placenta and no lesional development or transmission during early infection.

Brainstem auditory evoked potentials (BAEP) are useful to diagnose neurologic and genetic disorders in cattle but normal parameters and its application to detect changes in hearing are not demonstrated. The aims of this study were to develop a protocol for obtaining BAEP in Holstein calves and to establish the normal parameters of peak latencies, interpeak latencies (IPL) and waveforms of BAEP in this species. Ten male Holstein calves, aged 2.5 months and weighting 90.7 +/- 4.0 kg were used. All animals were previously sedated with 0.08 mg/kg xylazine hydrochloride administered intramuscularly. The BAEP analysis was recorded using the IHS SmartEP-ASSR program connected with electrode needles placed on the standardized locations of the animal's head. Monoaurally presented broadband click stimuli (0.1 ms) were used for BAEP stimulation, with contralateral masking of 30 dB SPL. Stimuli were presented at a rate of 35.5 pulses per second (PPS) and evoked responses were collected and averaged 450-1500 times for each sound intensity. Earphones were inserted into the ear canal to present the acoustic stimulus and to attenuate environmental noise. Peak latencies, IPL, waveforms and auditory thresholds were recorded and analyzed in the right ear and were like those described in previous papers in adult male bulls. Auditory thresholds obtained in all animals were 33 +/- 1.5 dB SPL. Peak latencies were 1.4 +/- 0.07 ms (Peak I), 2.4 +/- 0.10 ms (Peak II), 3.3 +/- 0.10 ms (Peak III), 4.4 +/- 0.19 ms (Peak IV) and 5.6 +/- 0.28 ms (Peak V). IPL were 1.9 +/- 0.12 ms (IPLI-III) 4.1 +/- 0.29 ms (IPL I-V), 2.2 +/- 0.19 ms (IPL II-V) and 2.2 +/- 0.19 ms (IPL III-V). Brainstem auditory evoked potentials are a minimally-invasive method to evaluate level of audition in Holstein calves. In addition, BAEP is a method that can be applied in young calves and it would work on-farm to select the animals with better conditions.

In a previous attempt, an experimental model of bovine besnoitiosis was established in calves that were intravenously inoculated with different doses of Besnoitia besnoiti tachyzoites. Despite the fact that all infected calves developed the acute stage of disease, only microscopic findings characteristic of chronic besnoitiosis were reported. In the present study, calves were inoculated by subcutaneous and intradermal routes with B. besnoiti tachyzoites with the aim of developing clinical signs and macroscopic lesions characteristic of chronic besnoitiosis. Nine 3-month-old male calves were randomly distributed into three groups of three animals each. Next, 106 tachyzoites were inoculated by either the subcutaneous (G1) or intradermal route (G2). The negative control group (G3) was inoculated with PBS. Daily clinical monitoring and regular blood collection were performed. At 70 days post-infection (pi), animals were euthanized, and tissues were collected to investigate lesions and parasites. Infected animals developed mild-moderate acute besnoitiosis characterized by lymphadenopathy from four days to 47 days pi, and sporadic fever peaks were only observed in one calf from G2. However, other clinical signs and macroscopic lesions characteristic of chronic besnoitiosis were not detected. Only nine tissue samples were B. besnoiti-DNA-positive, eight of which belonged to reproductive and respiratory tracts tissues from G1. Finally, the kinetics of the immune responses were similar in both infected groups. However, delayed and lower cellular and humoral immune responses were observed in G1 followed by G2 and were compared with intravenously inoculated calves. The differences observed among the three inoculation routes could be due to different effector mechanisms of the host early innate immune response against B. besnoiti. Accordingly, the inoculation route of B. besnoiti tachyzoites does not significantly influence the clinical outcome of the infection in calves. Thus, a further refinement of this experimental model of bovine besnoitiosis is needed to reproduce macroscopic lesions characteristic of chronic stage disease.

Early in vivo diagnosis of bovine besnoitiosis is crucial for the success of control programmes. However, diagnosis in acutely infected animals is hindered by the low sensitivity of the available serological tools. In this study, a novel ELISA to detect specific anti-Besnoitia besnoiti IgM antibodies was developed. The usefulness of this tool together with an avidity ELISA were studied with a well-coded sera panel from experimentally and naturally infected cattle. First, the kinetics of specific IgM levels were determined in experimentally infected calves during the acute and chronic infection. Next, IgM levels were determined in naturally infected cattle with either acute or chronic infection. Finally, the IgG avidity index was monitored in both experimentally and naturally infected cattle. Specific IgM antibodies were detected prior to specific IgG antibodies (7-19 days vs. 17-26 days post-infection). A prompt IgM response was associated with the end of the febrile stage in experimentally infected calves. Naturally and experimentally infected animals with acute clinical signs tested IgM-positive but IgG-negative, followed by IgG seroconversion 2-3 weeks later. Chronically infected cattle developed both IgM and IgG specific antibodies. Moreover, a progressive increase in the avidity index (AI) was observed in all experimentally infected calves during the course of the experimental trials. However, a low AI coincided with visible tissue cysts. Low avidity values were also detected when naturally infected cattle with acute clinical signs seroconverted, in contrast to a high AI detected in chronically infected cattle. In summary, IgM and avidity ELISAs improved the early in vivo diagnosis of bovine besnoitiosis. IgM-positive but IgG-negative results were indicative of an acute infection, whereas IgG positive results accompanied by low avidity values confirmed a recent infection.

Bovine besnoitiosis is continuing to spread in Europe. Therefore, the development of ruminant animal models of infection is urgently needed to evaluate therapeutic and prophylactic tools. Herein, we studied the effect of parasite dose and host age on the infection dynamics with Besnoitia besnoiti tachyzoites in cattle in two independent experimental infections. In experiment A, twelve 3-month-old male calves were inoculated intravenously with either three different doses of tachyzoites (G1: 10(8); G2: 10(7); G3: 10(6)) or with PBS (G4). In experiment B, six 14-month-old bulls were inoculated with 10(6) tachyzoites based on results obtained in experiment A. In both trials, clinical signs compatible with acute and chronic besnoitiosis were monitored daily; blood and skin samples were collected regularly for 70-115 days post-infection (pi). Finally, animals were killed, and tissues were collected for lesion and parasite detections. Infected animals developed mild-moderate signs compatible with acute besnoitiosis. Lymphadenopathy and fever were observed in both calves (from 12 hr until 7 days pi) and bulls (from 6 days until 9 days pi). Seroconversion was detected at 16-19 days pi, and antibody levels remained high. Infected animals did not developed characteristic clinical signs and macroscopic lesions of chronic besnoitiosis. However, successfully, parasite-DNA was detected in a reduced number of target tissues: conjunctiva, ocular sclera, epididymis, skin of the scrotum and carpus in calves (n = 10, 6 of which belonged to G3), and pampiniform plexus and testicular parenchyma in bulls. Remarkably, one tissue cyst and mild microscopic lesions were also detected. In summary, inoculated animals developed the acute besnoitiosis and chronic infection was evidenced by microscopic findings. However, our results suggest that tachyzoite dose and host age are not key variables for inducing clinical signs and macroscopic lesions characteristic of chronic besnoitiosis. Thus, a further refinement of this model should evaluate other parasite- and host-dependent variables.

Neospora caninum is one of the main causes of abortion in cattle, and recent studies have highlighted its relevance as an abortifacient in small ruminants. Vaccines or drugs for the control of neosporosis are lacking. Bumped kinase inhibitors (BKIs), which are ATP-competitive inhibitors of calcium dependent protein kinase 1 (CDPK1), were shown to be highly efficacious against several apicomplexan parasites in vitro and in laboratory animal models. We here present the pharmacokinetics, safety and efficacy of BKI-1553 in pregnant ewes and foetuses using a pregnant sheep model of N. caninum infection. BKI-1553 showed exposure in pregnant ewes with trough concentrations of approximately 4 mu M, and of 1 mu M in foetuses. Subcutaneous BKI-1553 administration increased rectal temperatures shortly after treatment, and resulted in dermal nodules triggering a slight monocytosis after repeated doses at short intervals. BKI-1553 treatment decreased fever in infected pregnant ewes already after two applications, resulted in a 37-50% reduction in foetal mortality, and modulated immune responses; IFN. levels were increased early after infection and IgG levels were reduced subsequently. N. caninum was abundantly found in placental tissues; however, parasite detection in foetal brain tissue decreased from 94% in the infected/untreated group to 69-71% in the treated groups. In summary, BKI-1553 confers partial protection against abortion in a ruminant experimental model of N. caninum infection during pregnancy. In addition, reduced parasite detection, parasite load and lesions in foetal brains were observed.

Experimental infections in pregnant sheep have been focused on studying the effect of the time of challenge on the outcome of N. caninum infection, whereas the impact of the dose and route of challenge has not been studied in depth. Therefore, clinical outcome, immune responses, parasite detection and burden, and lesion severity in placental tissues and foetal brains were investigated in 90-day-pregnant sheep inoculated intravenously with 105 (G1), 104 (G2), 103 (G3), or 102 (G4) tachyzoites or subcutaneously with 104 (G5) tachyzoites of the virulent Nc-Spain7 isolate and an uninfected group (G6). Comparing challenge doses, G1 was the only group that had 100% abortion. Likewise, IFNγ levels in G1 increased earlier than those in other intravenously infected groups, and IgG levels on day 21 post-infection (pi) were higher in G1 than those in other intravenously infected groups. Concerning vertical transmission, G1 shows a higher parasite burden in the foetal brain than did G2 and G3. Comparing routes of administration, no differences in foetal survival rate or parasite load in the foetal brain were found. Although G2 had higher IFNγ levels than G5 on day 10 pi, no differences were found in humoral immune responses. Because the outcome after intravenous infection with 105 tachyzoites was similar to that observed after intravenous infection with 106 tachyzoites used in a previous work (100% abortion and vertical transmission), we conclude that it may be reasonable to use 105 tachyzoites administered by the intravenous route in further experiments when assessing drugs or vaccine candidates.

Bovine besnoitiosis, caused by the apicomplexan Besnoitia besnoiti, is a chronic and debilitating disease characterized by cutaneous and systemic manifestations that primarily affects adult beef cattle. Previous studies have reported that clinical besnoitiosisis is rare in calves. However, we isolated B. besnoiti from a chronically infected calf for the first time. The identity of the Besnoitia species was determined after parasite isolation and molecular genotyping. According to the results obtained in vitro the new isolate, named as Bb-Spain3, was characterized in a reproducible in vitro model and was categorized as a low invader and low prolific isolate with a slower lytic cycle compared to Bb-Spain 1 isolate. Specific traits that differentiate isolates obtained from adult animals from those infecting calves were not found. Next, we described the first case report of chronic besnoitiosis in a female calf less than 6 months-old with a low body condition. The disease was confirmed by the presence of specific anti-B. besnoiti antibodies and parasite detection in the skin. At post-mortem examination, tissue samples were collected for histological, immunohistochemical and molecular analyses. DNA-parasite was detected in 31 different calf's tissues, being the most highly parasitized tissues the skin and the respiratory and reproductive tracts. In addition, the parasite was also present in heart, eyes, lymph nodes and brain. The high parasite load, a wide intra-organic parasite distribution and the presence of both viable and degenerated cysts, were indicative of a rapid progression of the disease. This case report underlines the need to include the inspection of young animals in besnoitiosis control.

OBJECTIVE To compare the effectiveness of an ultrasound-guided paravertebral nerve blockade technique (UGPNB) with distal and proximal paravertebral nerve blockade techniques without ultrasound guidance (DPNB and PPNB, respectively) in calves. ANIMALS 4 calf cadavers and 7 healthy calves. PROCEDURES A suitable acoustic window was identified to facilitate access to the T13, L1, and L2 spinal nerves in cadavers and live calves. In cadavers, nerves were injected with dye under ultrasound guidance. In calves, the UGPNB, DPNB, and PPNB were performed in random order at 10-day intervals by injection of an anesthetic solution containing 2% lidocaine hydrochloride. Nociceptive withdrawal responses were assessed to determine the effects of the blockades. RESULTS In cadavers, nerve staining success rates (ie, ≥ 2-cm-long dye path) achieved with ultrasound guidance were 88% (T13 [ventral branch]), 75% (T13 and L1 [dorsal branches] and L1 and L2 [ventral branches]), and 38% (L2 [dorsal branch]). The nerves were each identified as a hyperechoic band in a longitudinal plane. In calves, the UGPNB, DPNB, and PPNB reduced the withdrawal response to the noxious stimulus, mainly in the dorsal-cranial, dorsal-caudal, and ventral-cranial areas of the flank. Overall, the UGPNB resulted in a better nociceptive cumulative score, administering only one half of the local anaesthetic dose, compared with findings for the DPNB and PPNB. However, time to perform the UGPNB was longer. CONCLUSIONS AND CLINICAL RELEVANCE The UGPNB evaluated may be an improved alternative to the DPNB and PPNB for provision of anesthesia for flank surgery in calves. However, effectiveness of the UGPNB should be evaluated in a clinical setting and in adult cattle.

Superficial nerves can be visualized through ultrasonography in the cattle and facilitate local anesthetic disposition around nerve structures. Expected advantages include a higher successful rate of nerve block improving the degree and duration of the block. Among others, conduction nerves of clinical interest in cattle include the paravertebral nerves, nerves of the epidural space , the brachial plexus , and the sciatic and femoral nerves , and nerves of the head.

Atresia coli is a congenital defect with disruption of the intestinal flow. Its etiopathogenesis is still unknown and the prognosis is reserved. This paper describes the case of a one week old Holstein Friesian calf that was referred to the HCVC, and was diagnosed with atresia coli . This confirmation was made by contrast radiography and the lesion was detected in dorsal portion of the spiral colon. A cecostomy was performed followed by a marsupialization of the intestinal cecum to the abdominal wall. After 36 hours of surgery, calf’s state worsens dramatically and for reasons of welfare, it was proceeded to sacrifice. The post-operative recovery is possible, even though the mortality rate is high. As in this case, this mortality rate increases when early diagnosis of atresia coli does not take place and there is peritonitis.

This study aimed to describe a suitable acoustic window to facilitate access to the sciatic and femoral nerves in calves and to study the effects of their blockade with local anaesthetics. The neuroanatomical and ultrasound (US) study was performed on the cadavers of 10 calves, and the effects of 2% lidocaine with epinephrine (0.2 mL/kg) were determined in five healthy calves. The sciatic nerve in the cadavers was easily visualised as a hyperechoic band distal to the femoral greater trochanter and caudal to the femoral shaft. The femoral nerve in the cadavers was not easily identified, and was visualised as a hyperechoic oval structure situated immediately medial to the psoas major muscle and lateral to the femoral artery. The sciatic nerve was stained by methylene blue, injected under US guidance, in 9/10 cases, and the femoral nerve was stained in 6/10 cases. Sciatic nerve blockade under US guidance produced adduction of the limb with metatarsophalangeal joint flexion, while the femoral nerve blockade produced reduced weight bearing. The sciatic nerve blockade produced a reduced response to the noxious stimulus, mainly in the phalanges, proximal and distal metatarsus, tarsus and tibia and, following the femoral nerve blockade, in the medial subarea of the femur. However, femoral nerve blockade produced a more variable degree of blockade. In conclusion, US -guided anaesthetic blockade of the sciatic nerve in calves may be considered for surgery in the distal pelvic limb, although further studies are necessary to determine its clinical application.

Live vaccines have emerged as one of the most potentially cost-effective measures for the control of bovine neosporosis. Previous studies have shown that Nc-Spain 1H is a naturally attenuated isolate of Neospora caninum and that immunisation with live Nc-Spain 1H tachyzoites generated a protective immune response in mice. The aim of this study was to evaluate the safety and efficacy of immunisation in cattle. N. caninum-seronegative heifers were immunised subcutaneously twice with 107 live Nc-Spain 1H tachyzoites prior to artificial insemination. No adverse reactions or negative effects on reproductive parameters were recorded following immunisation. In immunised and non-challenged heifers, no foetal deaths were observed, and none of the calves was congenitally infected. The efficacy against N. caninum-associated foetal death and vertical transmission was determined after challenge with high doses of the Nc-1 isolate at 70 and 135 days of gestation, respectively. After the challenge in early gestation, the immunisation induced a protection of 50% against foetal death. In addition, the microsatellite analysis performed in PCR-positive tissue samples from foetuses that died after challenge infection showed that the profiles corresponded to the challenge isolate Nc-1. A degree of protection against vertical transmission was observed after challenge at mid-gestation; calves from immunised heifers showed significantly lower pre-colostral Neospora-specific antibody titres than calves from the non-immunised/challenge group (P < 0.05). Strong antibody and interferon gamma responses were induced in the immunised heifers. This study indicates that the immunisation before pregnancy with the Nc-Spain 1H vaccine isolate appeared to be safe and reduced the occurrence of N. caninum-associated abortion and vertical transmission in experimentally infected cattle. In light of these encouraging results, the next step for testing this live attenuated candidate should be the assessment of its efficacy and safety in naturally infected cattle.

The aim of this study was to determine the efficacy of a concentrated combination of tiletamine-zolazepam [TZ, 0.53 mg/kg body weight (BW)], ketamine (Ket, 0.53 mg/kg BW), and detomidine (Det, 0.04 mg/kg BW) in the immobilization of free-range cattle for clinical procedures. The combination was administered intramuscularly to 53 animals. Anesthesia was reversed with the α2-adrenoceptor antagonist atipamezole. Locoregional anesthesia was provided with lidocaine when required. The TZKD combination induced suitable immobilization for minor surgical procedures or medical treatments. Anesthetic onset was rapid, taking a mean of 6.1 min [standard deviation (SD) 2.8 min]. The duration of anesthesia depended on the time of administration of the antagonist; the animals recovered in the standing position in 12.9 ± 8.9 min after the administration of atipamezole. The quality of anesthesia and analgesia were satisfactory. In conclusion, this TZKD combination can be used for both immobilization and minor surgical procedures in free-range cattle.