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The Veterinary Journal The Veterinary Journal 176 (2008) 102–114 www.elsevier.com/locate/tvjl

Postpartum uterine disease and dairy herd reproductive performance: A review Stephen J. LeBlanc * Population Medicine, Ontario Veterinary College, University of Guelph, Guelph, ON, Canada N1G 2W1 Accepted 18 December 2007

Abstract This paper reviews the causes, impact, treatment, and prevention of retained placenta (RP), metritis, and endometritis in dairy cows. The occurrence of each of these diseases largely depends on immune function in the transition period. Retained placenta affects 5–10% of calvings and greatly increases the risk of metritis and endometritis. More field studies are needed to validate criteria for treatment of metritis, but cows with at least two of RP, fever, dullness, and fetid uterine discharge appear to merit treatment with systemic antibiotics. Clinical endometritis affects 15–20% of cows at 4–6 weeks postpartum; an additional 30–35% have subclinical endometritis between 4 and 9 weeks postpartum. Under specific conditions, treatment of cows with endometritis improved pregnancy rate. Systematic use of prostaglandin F2a at 5 and 7 weeks postpartum may improve pregnancy rate. The economic benefit of efforts to identify and treat endometritis is herd-specific. Ó 2008 Elsevier Ltd. All rights reserved. Keywords: Retained placenta; Metritis; Endometritis; Prostaglandin; Transition cows

Introduction Reproduction is one of the key pillars of dairy production. Many dairy herds do not achieve their targets for reproductive performance and incur substantial economic opportunity cost. Despite the fact that postpartum uterine disease is only one component of reproductive performance, and that it generally has secondary importance to insemination efficiency (LeBlanc, 2005), it has traditionally occupied a substantial amount of veterinarians’ attention. Additional data are required to inform evidence-based medicine and rational health management interventions. Reproductive performance is linked to health in the weeks immediately before and after calving, and timely achievement of subsequent pregnancy in turn has a substantial impact on profitability (De Vries, 2006). If reproductive or health management practices cannot be shown to result

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1090-0233/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.tvjl.2007.12.019

in lower death losses, fewer associated diseases, more milk sold, or a higher herd pregnancy rate, they should be changed or removed. This paper reviews current information and questions on the causes, impact, treatment, and prevention of retained placenta (RP), metritis, and endometritis in dairy cows.

Peripartum physiology and immune function Various aspects of immune function are suppressed in dairy cows from 1 to 2 weeks prepartum until 2–3 weeks postpartum (Kehrli et al., 1989a,b; Mallard et al., 1998). The precise causes of impaired immune function in transition cows are unclear, although the peripartum drop in energy, vitamin and mineral intake, negative energy balance and mobilization of body fat and protein, dramatic changes in progesterone and estrogen levels in late gestation, and the massive increase in cortisol level at calving appear to contribute (Goff and Horst, 1997; Kehrli et al., 1999; Ingvartsen, 2006).

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The hormonal and energy effects of lactation appear to exert an immunosuppressive effect beyond those associated with parturition itself (Kimura et al., 1999). Cows in greater negative energy balance have more pronounced impairment of at least some immune functions (Hammon et al., 2006). Cows with RP or metritis have earlier and more profound impairments of innate immunity, starting several weeks before disease occurs (Gunnink, 1984; Gilbert et al., 1993; Cai et al., 1994; Hammon et al., 2006). Retained placenta Pathogenesis and risk factors Retained placenta is failure to pass the placenta within 24 h postpartum (Kelton et al., 1998). Although various authors define RP at 12 or 24 h, among cows that passed the placenta within 24 h, 95% had passed it within 12 h, so the distinction is not important (Van Werven et al., 1992). If RP occurs, the membranes are retained for 7 days on average (Eiler, 1997). RP is objectionable to the people who work with cows, and may also enhance bacterial contamination of the uterus and impair involution (Laven and Peters, 1996). In a summary of 50 reports on the incidence of RP (Kelton et al., 1998), the median lactational incidence rate (LIR) was 8.6%. The risk factors associated with RP include twins, dystocia, stillborn calf, induced parturition, abortion, milk fever, and increasing age, as well as conflicting seasonal effects (Sandals et al., 1979; Correa et al., 1993; Grohn and Rajala-Schultz, 2000). The key event in the pathogenesis of RP is a failure of prompt breakdown of the cotyledon–caruncle attachment after delivery of the calf. Lack of uterine motility to expel the placenta plays little or no role in the occurrence of RP (Eiler, 1997); cows with RP have normal to increased uterine activity in the days after calving (Frazer, 2005). Failure of placental detachment appears to be largely mediated by failure of the immune system to successfully degrade the plancentomes at the end of pregnancy. Cows in a greater degree of negative energy balance prepartum, as evidenced by higher non-esterified fatty acid (NEFA) concentration were 80% more likely to have RP, and accounting for the effect of NEFA, those with lower circulating vitamin E were at greater risk of RP (LeBlanc et al., 2004). This supports the notion that premature, or severe negative energy balance impairs immune function, which in turn makes RP more likely (Goff and Horst, 1997), but it also underlines the fact that the development of RP is multifactorial. Inbreeding may also contribute to failure of a vigorous immune response (Joosten et al., 1991). There is persuasive evidence that cows with RP have substantially lower leukocyte (particularly neutrophil) chemotaxis and oxidative burst activity, and lower interleukin (IL)-8 concentrations than cows without RP, and that these differences are present for up to 2 weeks prepartum, as well as at parturition (Gunnink, 1984; Kimura et al., 2002). Recently, it has been

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shown that intracellular calcium plays an important role in immune (mononuclear cell) function in dairy cows, suggesting a possible link between hypocalcaemia and immune function (Kimura et al. 2006). This association, more than lack of uterine contractions due to hypocalcemia, may explain the association between milk fever and RP. Additionally, the causal pathways to milk fever and RP likely share sub-optimal dry matter intake (DMI) as important components. Serum prostaglandin (PGF2a) metabolite (PGFM) was also elevated 1 week prior to parturition in cows that subsequently had RP, which was significantly earlier than in cows that did not have RP (Peter and Bosu, 1987); in the same study, the magnitude of the peripartum rise in serum cortisol was greatly increased in cows with RP. Impact Retained placenta is a symptom of a suboptimal transition period, but is only important to reproductive performance to the extent that it is a substantial risk factor for metritis and endometritis. The impact of RP ranges from none, to impaired reproductive performance, to progression to severe metritis with loss of production. Just as the occurrence of RP depends on immune function, so does the course once the condition has occurred. Retained placenta is associated with increased risk of subsequent ketosis, abomasal displacement and mastitis (Grohn et al., 1990; Oltenacu et al., 1990). The best analyses estimate that pregnancy rate in affected cows is reduced by approximately 15% relative to unaffected cows (Fourichon et al., 2000). It is likely that impaired reproductive performance occurs only if RP leads to the development of metritis or endometritis. Loss of milk production appears to be confined to those individuals that progress to clinical metritis (Fourichon et al., 1999), and RP itself appears not to increase culling risk (Grohn et al., 1998). Treatment It is important to clearly establish the objective of RP treatment. Many antimicrobial and hormonal treatments have been applied to cows with RP, generally without any reduction in risk of subsequent disease (such as displaced abomasum) or improvement in reproductive performance (Gilbert and Schwark, 1992; Stevens et al., 1995; Peters and Laven, 1996; Eiler, 1997; Drillich et al., 2006a). Despite occasional reports of benefit (Gross et al., 1986; Mollo et al., 1997), immediate postpartum treatments with oxytocin, PGF2a or calcium have generally failed to prevent RP (Stevens and Dinsmore, 1997; Hernandez et al., 1999), or hasten the passage of retained fetal membranes (Stevens et al., 1995; Frazer, 2005). Historically, manual removal of RP by manipulation and traction was practiced. There is no evidence that this practice produces beneficial results (Drillich et al., 2006b), and some evidence suggests that it is harmful

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(Bolinder et al., 1988). Several studies indicate that approximately 50–80% of cows with untreated RP will have a temperature >39.5 °C on at least 1 day within 10 days postpartum (Callahan and Horstman, 1987; Stevens et al., 1995; Stevens and Dinsmore, 1997; Drillich et al., 2003, 2006b). It is not clear if all of these require systemic antibiotic treatment. Several of these studies indicate that daily intrauterine (IU) infusions of 5 g oxytetracycline for as long as the RP is in place reduced the incidence of fever from approximately 50% of cows with RP to approximately 30% of affected cows. In another study, ceftiofur (1.1 mg/kg IM q 24 h for 5 days) in cows with RP and fever was as effective (67% absence of fever by 10 days in milk (DIM)) as a combination of systemic and IU ampicillin and manual removal of the placenta; there was no difference in reproductive performance between the two treatments (Drillich et al., 2003). Typically, 25–50% of cows with RP develop metritis. Recent studies consistently indicate that there is no advantage in reproductive performance to automatically treating all cases of RP with systemic antibiotics. There are no trials published in which cows with RP and fever (with or without fetid discharge) were left untreated, even for 1 or 2 days. Two published field studies (Risco and Hernandez, 2003; Drillich et al., 2006a) compared treatment with 1– 2 mg/kg ceftiofur of all cows with RP to treatment of cows with RP only if they progressed to having a fever. There were no differences in pregnancy rates or culling between the two approaches. Unless most cows with RP will eventually be selected for systemic treatment with antibiotics because they develop metritis, or production is exceptionally high in early lactation, current data indicate that it is economically preferable to selectively treat metritis cases rather than automatically treat all cows with RP with antibiotics. However, this approach depends on a management system and labour that may be relied upon to identify individuals with metritis early in the progression of the disease. Criteria to select cows that will benefit from treatment require further research. Definition of terms in uterine disease Histologically, the depth of inflammation of the uterine wall distinguishes metritis and endometritis. However, more clinically relevant definitions are needed. Case definitions in the literature are highly confounded by diagnostic method (observation or odour of discharge, fever, palpation, vaginoscopy, bacteriologic culture, biopsy, or cytology), as well as the interval postpartum at which diagnosis is made. The term metritis has been used across a broad range of intervals postpartum to describe a variety of heterogeneous conditions including puerperal metritis, endometritis, pyometra, and various subjective clinical findings that were assumed to correlate with bacterial infection of the uterus or slow uterine involution. Recently, case definitions have been proposed (Sheldon et al., 2006), and will be used here.

Normal uterine involution Understanding normal uterine involution is critical to provide context for accurate classification of postpartum findings as physiologic or pathologic. Lochia is normally passed until 14–23 DIM (Gier and Marion, 1968; Lewis, 1997; Mortimer et al., 1997). Cervical involution is slower than that of the uterus, such that by 15 DIM, the diameter of the cervix normally exceeds that of the horns of the uterus (Mortimer et al., 1997). The reported times for gross involution of the uterus and cervix vary considerably, from 25 to 47 days (Morrow et al., 1966; Gier and Marion 1968; Marion et al., 1968; Harrison et al., 1986). Complete microscopic involution takes longer than palpable involution, up to 42–50 days (Marion and Gier, 1959). The physiological role of PGF2a in the first month postpartum is unclear, but it might help to promote uterine contractility and so contribute to uterine involution. Serum PGFM is normally substantially elevated for 10–14 days postpartum (Peter and Bosu, 1987). In normal cows, the longer levels of PGFM remained elevated postpartum, the shorter the time to gross uterine involution (Lindell et al., 1982; Madej et al., 1984) and resumption of the normal estrous cycle (Madej et al., 1984). Conversely, in animals with purulent uterine discharge, PGFM levels were higher and remained elevated longer than in normal cows, yet involution was delayed (Lindell et al., 1982; Del Vecchio et al., 1994). Inflammation of the uterus, as would be produced by infection with pathogenic bacteria, may provoke an increased magnitude and duration of PGF2a release from the uterus (Lindell et al., 1982). There is evidence that after repeated administration of PGF2a the uterus becomes refractory (Rodriquez-Martinez et al., 1987), which may account for the apparent dose or duration-dependent association of PGF2a with uterine involution. Uterine immunology The normal uterus is able to efficiently clear bacterial infection and it is difficult to experimentally produce chronic uterine infection, even by IU infusion of Arcanobacterium pyogenes (Gilbert and Schwark, 1992). Once bacteria have gained access to the uterus, the most important component of uterine defence is non-specific phagocytosis by neutrophils (Hussain, 1989; Bondurant, 1999). There are interactions of reproductive hormones with uterine immune function. It is generally agreed that a high progesterone environment suppresses cervical mucus production, myometrial contractility, uterine gland secretion and the phagocytic activity of uterine neutrophils (Frank et al., 1983; Hussain, 1989; Bondurant, 1999), and is therefore permissive of at least short-term uterine infection. PGF2a is not only luteolytic but also appears to have proinflammatory actions that may enhance neutrophil function (Lewis, 2004). However, interactions between progesterone and PGF2a in uterine immune function in dairy cows are

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not completely understood (Lewis, 2004). Uterine immune function appears to improve under the influence of estrogen (Frank et al., 1983; Bondurant, 1999). It is unclear whether estrogen causes an absolute increase in phagocytic and bactericidal activity, or whether the observed enhancements are simply relative to the situation under progesterone dominance (Hussain, 1989). In contrast, other studies (Subandrio et al., 2000) failed to find any consistent relationship between stage of the estrous cycle and uterine neutrophil function. Bacteriology of the postpartum uterus and pathogenesis of endometritis The uterus is routinely contaminated with bacteria in the early postpartum period (Elliott et al., 1968; Miller et al., 1980). Sheldon et al. (2004a) summarized four studies to illustrate that approximately 80–100% of cows have bacterial contamination of the uterus in the first 2 weeks postpartum. Infection with one or more of A. pyogenes, Escherichia coli, Pseudomonas spp., Streptococcus spp., Staphylococcus spp., Pasteurella multocida, Clostridium spp., Fusobacterium spp. and Bacteroides spp. is common in the first 2 weeks postpartum (Bondurant, 1999). By 3– 4 weeks postpartum, the number of bacteria and the variety of species has diminished substantially in healthy cows. The significance of bacterial culture from the postpartum uterus depends on the species isolated and the interval since calving. The bacterium that is consistently associated with chronic uterine inflammation is A. pyogenes (Bondurant, 1999). This opportunistic, Gram-positive facultative anaerobe is commonly present in mixed culture with a wide variety of organisms, but most often with the anaerobes Fusobacterium necrophorum and Prevotella melaninogenicus, E. coli or Streptococcus spp. (Studer and Morrow, 1978; Miller et al., 1980; Bonnett et al., 1991; Dohmen et al., 1995; Williams et al., 2005) with which A. pyogenes acts synergistically. Recent research indicates that uterine infection predominated by E. coli in the first week postpartum and A. pyogenes in the second week is associated with subsequent endometritis (Gilbert et al., 2007; Williams et al., 2007). The presence of A. pyogenes beyond 3 weeks postpartum is associated with purulent vaginal discharge (Studer and Morrow, 1978; Dohmen et al., 1995; Williams et al., 2005), persistent infection (Bonnett et al., 1991), elevated inflammation score in endometrial biopsies (Miller et al., 1980), and impaired reproductive performance (Studer and Morrow, 1978; Bonnett et al., 1993). These infections cause uterine inflammation, which is believed to result in delayed uterine involution and damage to embryos, causing impaired or delayed fertility (Bretzlaff, 1987; Gilbert, 1997). Additionally, uterine infection and inflammation are associated with some alteration in the pattern of follicle growth on ovaries ipsilateral to the previous pregnancy, and disruption of subsequent luteal phase durations (Mateus et al., 2002a,b; Sheldon et al., 2002b). Recent work has shown that cows with greater growth of uterine bacterial pathogens postpartum developed smaller

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first dominant follicles, which if ovulated, made smaller corpora lutea with lower circulating concentrations of progesterone (Williams et al., 2007). The development of metritis and endometritis has recently been summarized (Foldi et al., 2006). Recent in vitro studies (Donofrio et al., 2007) have suggested that bovine herpesvirus-4 might play a role in endometritis; this merits future investigation. There is now compelling evidence that both metritis and endometritis are associated with reduced feed intake, a more negative energy balance, and reduced immune function, and these differences are measurable beginning 2 weeks before calving, i.e., 3–7 weeks before the conditions are diagnosed (Urton et al., 2005; Hammon et al., 2006; Huzzey et al., 2007). Metritis Metritis (puerperal metritis) is inflammation of the uterus resulting in systemic signs of sickness, including fever, red-brown watery foul-smelling uterine discharge, dullness, inappetance, elevated heart rate, and low production (Sheldon et al., 2006). It occurs primarily in the first 7 days after calving. There is strong association between feed intake and cows’ behaviour and the subsequent development of metritis (Huzzey et al., 2007). Cows with severe metritis ate 2–6 kg DM less than healthy cows in the 2–3 weeks preceding the clinical signs of metritis (Huzzey et al., 2007). Among cows with metritis, E. coli and a variety of anaerobic bacteria are common isolates (Drillich et al., 2003). The largest risk factor for metritis is retained placenta, but other conditions that may impair feed intake and immune function also increase the risk of metritis. Affected cattle have moderate to severe illness, so there is little dispute that cows with metritis require systemic antibiotic treatment. There are data to indicate that 1 mg/kg ceftiofur maintains therapeutic concentrations against E. coli (Sheldon et al., 2004b) in uterine tissues in most, though not all treated animals (Okker et al., 2001; Drillich et al., 2006c). More importantly, field trials indicate that the treatments of choice for metritis include ceftiofur (1–2 mg/kg IM q 24 h) or procaine penicillin (21,000 IU/kg IM q 12–24 h) for 3–5 days (Smith et al., 1998; Drillich et al., 2001, 2006a; Chenault et al., 2004). Despite pharmacological evidence against achieving adequate uterine concentrations at label doses (Bretzlaff et al., 1983) a field study reported clinical efficacy of tetracycline at 10 mg/kg (Schmitt et al., 2001). Available field trial data indicate that addition of one dose of flunixin does not improve outcomes over the use of systemic antibiotics alone (Drillich et al., 2007). Clinical trials indicate that there is no benefit to IU oxytetracycline or ampicillin over systemic ceftiofur or penicillin alone (Smith et al., 1998; Drillich et al., 2003, 2006b). Treatment of metritis appears to be justified on the grounds of cow welfare and reduction of the probability of death in severe cases. However, the criteria for success are inconsistent. Ideally, the objective is to quickly return

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cows to their expected level of production with no other complications. From recent studies, the expected outcome is removal of fever in 67–77% of treated animals by 5–10 days after treatment; resolution of fetid odour may be more difficult (Drillich et al., 2001, 2006b; Chenault et al., 2004). There is a lack of data on the efficacy of treatment of metritis for prevention of subsequent related diseases (e.g. displaced abomasum) or for improvement of eventual reproductive performance. Furthermore, it is not clear whether systematic programs aimed at early diagnosis and treatment of metritis are effective in preventing progression to severe disease with losses in performance, or whether they result in treatment that is not medically or economically beneficial. For example, ceftiofur HCl (2.2 mg/kg IM q 24 h for 5 days) administered starting at 1 DIM to cows with RP without regard to body temperature significantly reduced the incidence of fever with fetid discharge, with no effect on time to pregnancy (Risco and Hernandez, 2003). In a separate study, administration of ceftiofur to multiparous cows at high risk for metritis in the first 3 DIM when fever was detected reduced the incidence of subsequent fetid discharge (Overton et al., 2003). In attempting early diagnosis of metritis, attention must be paid to normal uterine involution. For example, abundant red-brown lochia is normal in the first 2 weeks postpartum. Additionally, fever of 1–2 days duration is common in the first week after calving and is not well correlated with uterine infection (Sheldon et al., 2004a). Observation of cows’ attitude, appetite (if possible), and daily milk production are likely useful screening tests to select cows for further examination. Cows with RP should be inspected daily until the placenta is passed to detect progression to metritis. Measurement of rectal temperature is indicated in cows that are dull, inappetant, or producing less milk than expected. More large-scale field studies are needed to validate criteria for early treatment of metritis. For current practice, a reasonable protocol based on available information may be to treat cows with at least two symptoms of metritis (RP, rectal temperature >39.5 °C, dullness or inappetance, and fetid uterine discharge) with 3–5 days of systemic ceftiofur or penicillin. Endometritis Endometritis is inflammation of the uterus without systemic illness. It is characterized by muco-purulent or purulent uterine discharge associated with chronic bacterial infection of the uterus, occurring later than 3 weeks postpartum. Histologically, endometritis is characterized by disruption of the endometrial epithelium, infiltration of inflammatory cells and accumulations of lymphocytes, vascular congestion, and stromal edema (Bonnett et al., 1991; Bondurant, 1999). Diagnosis of endometritis The traditional clinical approach to diagnosis of endometritis has been to examine cows between 2 and 8 weeks

postpartum to identify animals that are not involuting normally. Many studies have focussed on attempts to identify cows that are ‘abnormal’ relative to an ill-defined timetable for normal involution, or to measure correlations among clinical findings in the absence of a gold standard such as pregnancy rate. The timing of examination should allow for the normal process of involution, yet also provide time for treatment and response prior to the start of the breeding period. Clinically, the challenge is to identify those cows that are truly at risk of impaired fertility, and to administer treatment that mitigates the problem. Data were collected from 1865 cows in 27 herds, including history of dystocia, twins, RP, or metritis (LeBlanc et al., 2002b). All cows were examined once between 20 and 33 DIM including external inspection, vaginoscopy, and palpation of the cervix, uterus and ovaries. Survival analysis was used to derive a case definition of endometritis based on factors associated with increased time to pregnancy. Clinical endometritis was diagnosed by the presence of purulent uterine discharge or cervical diameter >7.5 cm after 20 DIM, or mucopurulent discharge after 26 DIM. Given vaginoscopy, no diagnostic criteria based on palpation of the uterus had predictive value for time to pregnancy. The prevalence of clinical endometritis was 17%, with a range between herds of 5–26%. Vaginoscopy was required to identify 44% of these cases, but other methods of examining discharge from the cervix are likely at least as effective (e.g. manual examination of the vagina (Sheldon et al., 2002a) or use of the Metricheck device (McDougall et al., 2006). Although a single vaginoscopic examination may miss up to 9% of cows with uterine discharge (Kasimanickam et al., 2004), one of the above means of detection and classification of uterine discharge in the vagina is necessary to achieve acceptable diagnostic sensitivity for clinical endometritis. Cows that had twins, RP, or metritis, but not veterinary-assisted calving, were substantially more likely to subsequently have clinical endometritis (LeBlanc et al., 2002b). Using a similar case definition but with cows examined between 15 and 45 DIM, Whiteford and Sheldon (2005) reported that cows with milk fever resulting in recumbency at calving were more likely to have clinical endometritis. In contrast to LeBlanc et al. (2002b), other authors (Williams et al., 2005; McDougall et al., 2006) have reported that the uterine discharge category of ‘mucus with flecks of pus’ may be associated with reduced reproductive performance, at least by some measures of performance. A wider distribution of DIM at examination in the latter studies may explain the differences in results, because the significance of different categories of uterine discharge depends on the interval after calving at which they are identified (LeBlanc et al., 2002b). Among cows examined with the Metricheck device from 1 to 8 weeks postpartum, including cows with discharge classified as mucus with flecks of pus, the prevalence of endometritis was 21%, varying among herds from 10–31% (McDougall et al., 2006).

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Further large-scale investigations are warranted to refine the optimum time after calving and interpretive criteria for examination of cows for endometritis. Palpation of the uterus per rectum lacks accuracy as a tool to identify cows with reduced fertility (Miller et al., 1980; Kristula and Bartholomew, 1998). Given data on uterine discharge and cervical diameter, palpation of the uterus had no diagnostic value for pregnancy rate (LeBlanc et al., 2002b). Palpation correctly identified only 22% of cows predicted to have uterine infection, compared to uterine culture (Miller et al., 1980). Despite this, palpation is the most common means of diagnosis of endometritis. Vaginoscopy is more sensitive than simple external inspection for detection of purulent discharge (Dohmen et al., 1995; LeBlanc et al., 2002b). Vaginoscopy correctly predicted uterine infection in 59–82% of cases, with a stronger correlation with the presence of A. pyogenes (as well as F. necrophorum and Proteus spp.) as the character of the discharge became more purulent (Miller et al., 1980; Dohmen et al., 1995; Williams et al., 2005). Endometrial biopsy has been investigated as a diagnostic test for endometritis. Studer and Morrow (1978) found that histological evidence of inflammation was associated with increased days open, but this explained only 3% of the variation in this outcome. Bonnett et al. (1993) reported that biopsy correctly classified 84% of cows with respect to pregnancy by 120 DIM. However, biopsy has been associated with a detrimental effect on subsequent fertility (Miller et al., 1980; Bonnett et al., 1993).

rate, resulting in a 32 day increase in median time to pregnancy, and were 1.7 times more likely to be culled for reproductive failure than cows without endometritis (LeBlanc et al., 2002b). Cows with muco-purulent or purulent uterine discharge approximately 5 weeks before the start of breeding in a seasonal system were 10–19% less likely to be pregnant by 56 days into the breeding season, and the median time to pregnancy was 8–18 days longer than in cows with no or clear mucus in the vagina (McDougall et al., 2006).

Prevalence and impact of endometritis

Intrauterine infusion of antimicrobials

The prevalence of endometritis in 43 studies ranged from 2.2% to 37.3%, with a median of 10.1% (Kelton et al., 1998). Unfortunately, in many studies the case definition was not explicit or did not account for normal lochia and involution, and it most cases it was not validated as being associated with economically important performance outcomes. Given the case definition of a localized uterine condition, there is no mortality attributable to endometritis, and there is no direct loss of milk production (Fourichon et al., 1999). A meta-analysis of 23 studies found that endometritis increased mean days open by 15, decreased the relative risk of pregnancy by 150 DIM by 31%, and reduced the rate at which cows became pregnant by 16% (Fourichon et al., 2000). The costs of endometritis include reproductive inefficiency, culling, treatment cost, milk discard, labour, and increased risk of residues in food products. It is very difficult to interpret and compare much of the data about the impact of endometritis because of the wide range of imprecise case definitions used, and failure to account for important covariates. This difficulty is compounded by the use of biased outcome measures and invalid statistical analyses. Using a validated case definition and accounting for parity, herd, and ovarian status, cows with clinical endometritis had a 27% relative reduction in pregnancy

Infusion of antimicrobials into the uterus is aimed at achieving high concentrations at the site of infection (Gustafsson, 1984; Gilbert and Schwark, 1992). In contrast to systemic administration, intrauterine administration achieves higher drug concentration in the endometrium, but little penetration to deeper layers of the uterus or other genital tissues (Masera et al., 1980; Bretzlaff et al., 1983). Substances reported to be used for IU infusion include tetracycline (Thurmond et al., 1993; Sheldon and Noakes 1998), penicillin (Thurmond et al., 1993), cephapirin (Dohmen et al., 1995; McDougall, 2001; LeBlanc et al., 2002c), chloramphenicol (Steffan et al., 1984), diluted Lugol’s iodine (Callahan and Horstman, 1987), gentamycin, spectinomycin, sulfonamides, nitrofurasone, iodine, and chlorhexidine (Gustafsson, 1984; Gilbert and Schwark, 1992). Most are not approved for IU use, and in many cases have no published information on withdrawal times. However, there is evidence to suggest that IU infusion of several antibiotics result in drug residues in milk (Kaneene et al., 1986; Dinsmore et al., 1996). Lugol’s iodine and oxytetracyline are irritating and are reported to cause coagulation necrosis of the endometrium (Gilbert and Schwark, 1992). In field trials, IU infusion of antibiotics in a variety of protocols to treat endometritis has generally failed to show any benefit in reproductive performance over PGF2a

Treatment of endometritis A wide variety of therapies for endometritis have been reported, including systemically or locally administered antibiotics, or systemically injected PGF2a. The general principle of therapy of endometritis is to reduce the load of pathogenic bacteria and enhance uterine defence and repair mechanisms, and thereby halt and reverse inflammatory changes that impair fertility. Many therapeutic trials suffered from a lack of negative controls, small numbers of animals leading to little statistical power, or both. Many investigations have used diagnostic criteria for endometritis that were not validated as having an impact on reproductive performance, making it impossible to discern the effects of treatment. Some studies have used clinical or bacteriological cure as the endpoint, rather than economically relevant measures of the probability and timing of pregnancy.

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(Steffan et al., 1984; Olsen, 1996; Sheldon and Noakes, 1998) or no treatment (Steffan et al., 1984; Thurmond et al., 1993). Interestingly, studies questioning the utility of IU antibiotics are almost as old as the drugs themselves (Ulberg et al., 1952; Roberts, 1956). However, recently two large field studies have reported significantly improved reproductive performance in cows with validated cases of clinical endometritis (LeBlanc et al., 2002b,c) or with specific selection criteria (McDougall, 2001) that were treated with one dose of a commercially approved product containing cephapirin benzathine IU. In the former study, 316 cows (in 20 herds) with clinical endometritis were randomly assigned to receive cephapirin, PGF2a or no treatment. There was no benefit of treatment between 20 and 26 DIM. Cows that received cephapirin between 27 and 33 DIM became pregnant 60% faster than untreated controls. Pregnancy rate among cows that received PGF2a was not significantly different from either of the other groups. In the study in New Zealand, 690 cows (in 22 herds) with risk factors were randomly assigned at 41 ± 14 DIM to receive cephapirin IU or remain untreated. Among cows that had delivered a dead calf, had RP, or had purulent discharge from the vulva observed after 13 DIM, cows treated with cephapirin were approximately 2–3 times more likely to be pregnant by 56 days into the breeding season (McDougall, 2001). Prostaglandin F2a In cyclic cows, PGF2a causes luteolysis of a responsive corpus luteum (CL) resulting in decreased progesterone level and subsequent estrus, with increased estrogen level and myometrial contractions. These events are all plausibly favourable for clearance of uterine infection. The precise mechanism by which PGF2a resolves uterine infection is not known (Lewis, 2004). There is controversy about the possible effect of PGF2a other than to cause luteolysis and its consequential actions (Gilbert and Schwark, 1992), although PGF2a receptors are apparently present in the myometrium. There is some evidence that PGF2a may exert a direct short-term contractile effect on the uterus (Rodriquez-Martinez et al., 1987; Hirsbrunner et al., 1998). The hypothesis of an effect of PGF2a other than luteolysis is supported by several studies that have reported beneficial effects of PGF2a in the first month postpartum on reproductive performance parameters, in both normal and abnormal cows with low circulating progesterone levels (Steffan et al., 1984; Etherington et al., 1984; Young et al., 1984; Young and Anderson, 1986; McClary et al., 1989). However, other reports indicate that PGF2a is more effective when progesterone levels are high or a CL is palpable (Sheldon and Noakes, 1998; LeBlanc et al., 2002c). Among cows with dystocia, retained placenta or twins, there was no benefit in reproductive performance to three injections of PGF2a 1 week apart, starting at 3–10 DIM, relative to a single injection between 17 and 24 DIM

(Kristula and Bartholomew, 1998). This provides additional indirect evidence against a benefit of PGF2a in the absence of a CL. The optimum timing of administration of PGF2a for treatment of endometritis is unclear. The interval after calving until individual cows acquire a responsive CL is highly variable (Morrow et al., 1966; Benmrad and Stevenson, 1986) with approximately 20% of cows anovular at approximately 60 DIM (Opsomer et al., 2000; Walsh et al., 2006). Bonnett et al. (1990) found that a single administration of PGF2a at 26 ± 3 DIM resulted in decreased prevalence of vaginal discharge, decreased uterine size and decreased isolation of A. pyogenes at 40 DIM, independent of progesterone concentration at day 26. There is little evidence to support the use of PGF2a before 3 weeks postpartum (Kristula and Bartholomew, 1998; Hendricks et al., 2006). Gay and Upham (1994) reported that administration of PGF2a at approximately 25 DIM to clinically normal cows with a palpable CL significantly reduced first service pregnancy risk, although a mechanism to explain this observation is lacking. Tenhagen and Heuwieser (1999) found that biweekly administration of PGF2a from 22 to 28 DIM until first breeding at 50– 60 DIM reduced first service conception risk from 50% to 35%, in cows both with and without vaginoscopically diagnosed endometritis. Practically, there does not appear to be support for treating cows with or without endometritis with PGF2a until at least 4 weeks postpartum. Numerous studies have assessed the putative therapeutic effect of PGF2a in ‘abnormal’ cows in the first 5 weeks postpartum, all of which have failed to find statistically significant benefits in reproductive performance compared to untreated cows (Benmrad and Stevenson, 1986; Stevenson and Call, 1988; McClary et al., 1989; Archbald et al., 1990; Glanvill and Dobson, 1991; Risco et al., 1994). However, several showed a tendency for fewer days open in PGF2atreated cows (Benmrad and Stevenson, 1986; McClary et al., 1989; Risco et al., 1994), and all of these studies lacked statistical power. Additionally, these studies variously grouped cows with dystocia, retained placenta, twins, milk fever, ketosis, vaginal discharge, and cystic ovarian condition under the classification of abnormal. This biases the treatment effect toward the null, because these conditions are heterogeneous and may have no direct effect on reproductive performance. PGF2a was equal to (Sheldon and Noakes, 1998) or tended to be more effective (Steffan et al., 1984) than IU penicillin or tetracycline for improvement of reproductive performance in cows with endometritis, although these studies lacked negative controls. Numerous reviewers have concluded that PGF2a appears to be at least as effective for endometritis as any available alternative therapy, and presents minimal risk of harm to the uterus or presence of residues in milk or meat (Paisley et al., 1986; Gilbert and Schwark, 1992; Gilbert, 1992; Olsen, 1996). However, the caveats about the lack of statistical power, and the poor

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quality of the design and analysis of most published reports generally hold for PGF2a as they do for intrauterine antibiotics. As DIM increase and a greater proportion of cows are ovular, the efficacy of PGF2a may be expected to increase. Several studies have reported a benefit of routine treatment of normal cows or all cows with a single injection of PGF2a in the postpartum period (Etherington et al., 1984; Young et al., 1984; Young and Anderson, 1986; Etherington et al., 1988; McClary et al., 1989; Etherington et al., 1994), while others have found no benefit (Stevenson and Call, 1988; Armstrong et al., 1989; White and Dobson 1990; Morton et al., 1992; Gay and Upham, 1994). However, in most studies that showed reduced days open, the cows were undifferentiated with respect to endometritis or related conditions. Such a benefit might be mediated through treatment of (undetected) endometritis, or by increasing the number of estruses prior to breeding, in the absence of any uterine pathology. There is evidence that cows that ovulate early and have at least three estruses prior to first breeding have improved fertility (Thatcher and Wilcox, 1973; Benmrad and Stevenson, 1986). A meta-analysis of studies on postpartum PGF2a revealed a statistically significant but small (2–3 days) reduction in days open in treated cows (Burton and Lean, 1995). Unfortunately, they did not consider the interval postpartum or the luteal status of the cows at treatment, and they used an overly broad definition of abnormal for that sub-group. In summary, there are numerous studies that report improved reproductive performance when cows were routinely given at least one injection of PGF2a between 4 and 6 weeks postpartum, but there are also numerous studies that report no benefit of routine postpartum PGF2a. There is little evidence to support the use of PGF2a before 4 weeks postpartum. On balance, there is reasonable support for routine use of PGF2a at approximately 4 and 6 weeks postpartum in herds with a high prevalence of RP and metritis. There is a lack of specific evidence for improved reproductive performance among cows with, or at risk of, clinical endometritis and treated with PGF2a. Further research is needed on the optimum timing postpartum, relationship with ovular status, and the number of doses of PGF2a that may improve reproductive performance in cows with endometritis. Subclinical endometritis Subclinical endometritis, diagnosed by cytology, affects 35–50% of cows between 35 and 60 DIM and is associated with substantially reduced pregnancy rate, increasing median time to pregnancy by 30–88 days and increasing the proportion of cows that fail to become pregnant by 300 DIM by 20% points (Kasimanickam et al., 2004; Gilbert et al., 2005). The cytological criteria for diagnosis of subclinical endometritis continue to be refined (Kasimanickam et al., 2004; Galva˜o et al., 2007). Treatment with either cephapirin IU or PGF2a improved pregnancy rate in cows

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with subclinical endometritis (Kasimanickam et al., 2005a). Ultrasound examination may be used to identify some, but not all cases (Kasimanickam et al., 2004). The use of cytology to diagnose subclinical endometritis is not practical in field conditions, although use of a cytobrush may be simpler than lavage (Kasimanickam et al., 2005b). The high prevalence and impact of this condition merit consideration. Cow-side diagnostics or surrogate measures of subclinical endometritis would facilitate quantification of the prevalence of the problem and evaluation of treatment. Herd economic considerations in addressing endometritis There are valid criteria to diagnose endometritis, and treatments proven to improve pregnancy rate. Treatment of postpartum endometritis should be reserved for cases diagnosed after 4 weeks postpartum based on criteria that are associated with subsequent pregnancy rate. The value of the effort for individual diagnosis and treatment is herd-specific and depends on the sensitivity and specificity of the diagnostic method, the prevalence of endometritis, the voluntary waiting period, and the cost and efficacy of treatment. The practical question is whether a given herd should invest resources in individual diagnosis and treatment of endometritis, implement systematic treatment of selected or all cows, or neither. Large, multi-herd field studies are needed to address this question. However, some data are available to inform current management decisions. If individual diagnosis and treatment of endometritis is considered, one question is whether to examine all cows, or only those thought to be at greater likelihood of having endometritis. McDougall (2003) calculated that it was economically preferable to treat all cows with RP, dead calf, or owner-observed discharge >13 DIM with one dose of cephapirin IU than to examine those at-risk cows and treat based on vaginoscopy (assuming 20% false-negatives with vaginoscopy). However, data from LeBlanc et al. (reported in Sheldon et al., 2006) indicated that, while cows that had twins, RP, or metritis were very likely to have endometritis, if only these ‘high risk’ animals were examined, over 65% of cases of clinical endometritis would be missed. The interactions of the numerous factors favouring or inhibiting uterine infection and immune response that determine the occurrence of endometritis are not adequately captured in clinical disease history alone. Two field studies (Tenhagen and Heuwieser, 1999; Heuwieser et al., 2000) compared herd-level approaches of individual diagnosis and treatment of endometritis versus systematic treatment of all cows. In both studies, endometritis was diagnosed by palpation between 15 and 21 DIM. In the individually managed group, cows diagnosed with endometritis received an IU antiseptic at 15–21 and 29– 35 DIM, while the other group all received PGF2a every 14 days until inseminated. Both groups were bred based on detected estrus after 50–60 DIM. These studies must be interpreted considering that the diagnostic method and timing for endometritis, as well as the efficacy of the IU

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treatment, were not validated. Nevertheless, herd pregnancy rate was either not different (Tenhagen and Heuwieser, 1999) between groups or better in the systematic PGF2a group (Heuwieser et al., 2000), and in both studies this was the case not only in the herd as a whole, but also among cows diagnosed with endometritis. There is additional indirect evidence to be considered regarding individual-level diagnosis and treatment of endometritis versus routine group or whole-herd management. Kasimanickam et al. (2006) found that when all cows received injections of PGF2a at approximately 35 and 49 DIM followed by the Ovsynch program for timed insemination at approximately 73 DIM, there was no effect of endometritis on the probability of pregnancy at the synchronized first insemination. Tenhagen et al. (2001) also conducted a series of studies that considered the importance of endometritis as a factor in herd reproductive management programs for first insemination. Each of these trials reported a 53% prevalence of endometritis and must be interpreted considering that the diagnostic method and timing for endometritis were not validated. In one experiment, all cows were given PGF2a at 51 and 65 DIM, followed by timed insemination; in a second experiment later in the same herd, all cows were synchronized with the Ovsynch program for timed insemination at approximately 75 DIM (Tenhagen et al., 2001). In both experiments, postpartum endometritis did not affect the probability of pregnancy at first insemination. Tenhagen et al. (2003) diagnosed 24% of 1063 cows with endometritis based on palpation at 24 DIM. All cows were bred between 53 and 102 DIM (most between 73 and 81 DIM) using the Ovsynch program. Accounting for the effect of milk production, postpartum endometritis did not affect the probability of pregnancy at first insemination. Endometritis is important only to the extent that it impairs achievement of pregnancies in the economically optimal time interval. Collectively, these studies (Tenhagen et al., 2001, 2003; Kasimanickam et al., 2006) suggest that one or both of time and two injections of PGF2a may mitigate the effect of endometritis on herd reproductive performance to the point that it may not be profitable to devote resources to identifying individual cows with clinical endometritis. Further research is needed on the optimal timing and the need for repeated administration of PGF2a as a treatment for endometritis. However, a program of two injections of PGF2a at approximately 35 and 49 days postpartum without individual examination, followed by either heat detection or timed insemination, is the standard against which other uterine health management programs should be compared economically. Prevention of uterine disease Retained placenta, metritis, and endometritis are largely diseases of immune function. A lack of modalities to stimulate immunity in transition dairy cows means that there are few drugs or management strategies specific to preven-

tion of uterine disease. On the other hand, it is clear that uterine disease and immune function are substantially associated with peripartum energy metabolism and nutrition (Overton and Waldron, 2004) and most importantly with feed intake in the transition period (Hammon et al., 2006; Huzzey et al., 2007). Because the cause of RP is multifactorial, no one preventive measure will be universally effective. The principle for prevention is to optimize peripartum immune function, principally through management to encourage feed intake in the transition period (Cook and Nordlund, 2004). In particular, the prepartum diet should include 0.3 ppm selenium (ideally 5 mg/day; Hogan et al., 1993) and 1000–2000 IU/cow/day of vitamin E (Weiss, 1998; Baldi et al., 2000). Injection of vitamin E prepartum may help to prevent RP (Bourne et al., 2006), but the effect is conditional. Among animals with sub-optimum circulating vitamin E in the last week prepartum (serum a-tocopherol:cholesterol ratio <2.5  10 3), injection of 3000 IU a-tocopherol SC one week before expected calving reduced the risk of RP (LeBlanc et al., 2002a). Unfortunately, there is no simple way to identify individuals or herds that may benefit from this treatment. Peripartum oral administration of calcium does not reduce the incidence of RP (Oetzel, 1996; Hernandez et al., 1999). Decisions about preventive interventions should consider the magnitude of the problem as well as the evidence for the expected costs and benefits of proposed measures. Bacterial contamination of the uterus is almost universal in the first two weeks postpartum (Sheldon, 2004). While there are good reasons to have cows calve in a clean environment, it is very unlikely that bacterial infection of the uterus can be prevented altogether. Cleanliness of the calving area may not directly determine uterine contamination postpartum (Noakes et al., 1991). However, healthy cows clear bacteria from the uterus within approximately 3 weeks after calving, and complete involution of the uterus and cervix within 4–6 weeks. Conclusions Retained placenta, metritis, and endometritis are diseases of immune function in the transition period, which begin at least 2 weeks prepartum. The principle for prevention is to optimize peripartum immune function, principally through management to encourage feed intake in the transition period. Fortunately, the management procedures aimed at prevention of postpartum uterine disease are essentially the same as those to favor other aspects of health and production in transition cows. Although more research is needed to develop selection criteria for profitable treatment of early cases of metritis, a reasonable protocol based on available information may be to treat cows with at least two symptoms (RP, rectal temperature >39.5 °C, dullness or inappetance, and fetid uterine discharge) with 3–5 days of systemic ceftiofur or penicillin. Automatic antibiotic treatment of all cows with RP is not beneficial in most circumstances. On

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average, approximately 15–20% of cows have clinical endometritis at 4–6 weeks postpartum, with an additional 30– 35% having subclinical endometritis between 4 and 9 weeks postpartum. Clinical and subclinical endometritis cause substantial impairment of reproductive performance in affected cows, but the economic benefit of efforts to identify and treat these individuals is herd-specific. Conflict of interest statement The author (Stephen LeBlanc) has no financial or personal relationship with other people or organisations that could inappropriately influence or bias the paper entitled Postpartum uterine disease and dairy herd reproductive performance: A review. References Archbald, L.F., Tran, T., Thomas, P.G.A., Lyle, S.K., 1990. Apparent failure of prostaglandin F2a to improve the reproductive efficiency of postpartum dairy cows that had experienced dystocia and/or retained fetal membranes. Theriogenology 34, 1025–1034. Armstrong, J.D., O’Gorman, J., Roche, J.F., 1989. Effects of prostaglandin on the reproductive performance of dairy cows. Veterinary Record 125, 597–600. Baldi, A., Savoini, G., Pinotti, L., Monfardini, E., Cheli, F., Dell’Orto, V., 2000. Effects of vitamin E and different energy sources on vitamin E status, milk quality and reproduction in transition cows. Journal of Veterinary Medicine Series A 47, 599–608. Benmrad, M., Stevenson, J.S., 1986. Gonadotropin-releasing hormone and prostaglandin F2a for postpartum dairy cows: estrous, ovulation, and fertility traits. Journal of Dairy Science 69, 800–811. Bolinder, A., Seguin, B., Kindahl, H., Bouley, D., Otterby, D., 1988. Retained fetal membranes in cows: manual removal versus nonremoval and its effect on reproductive performance. Theriogenology 30, 45– 56. Bondurant, RH., 1999. Inflammation in the bovine female reproductive tract. Journal of Dairy Science 82 (Suppl. 2), 101–110. Bonnett, B.N., Martin, S.W., Meek, A.H., 1993. Associations of clinical findings, bacteriological and histological results of endometrial biopsy with reproductive performance of postpartum dairy cows. Preventive Veterinary Medicine 15, 205–220. Bonnett, B.N., Etherington, W.G., Martin, S.W., Johnson, W.H., 1990. The effect of prostaglandin administration to Holstein–Friesian cows at day 26 postpartum on clinical findings, and histological and bacteriological results of endometrial biopsies at day 40. Theriogenology 33, 877–890. Bonnett, B.N., Martin, S.W., Gannon, V.P., Miller, R.B., Etherington, W.G., 1991. Endometrial biopsy in Holstein–Friesian dairy cows. III. Bacteriological analysis and correlations with histological findings. Canadian Journal of Veterinary Research 55, 168–173. Bourne, N., Laven, R., Wathes, D.C., Martinez, T., McGowan, M., 2006. A meta-analysis of the effects of Vitamin E supplementation on the incidence of retained foetal membranes in dairy cows. Theriogenology. doi:10.1016/j.theriogenology.2006.08.015. Bretzlaff, K.N., 1987. Rationale for treatment of endometritis in the dairy cow. Veterinary Clinics of North America: Food Animal Practice 3, 593–607. Bretzlaff, K.N., Ott, R.S., Koritz, G.D., Bevill, R.F., Gustafsson, B.K., Davis, L.E., 1983. Distribution of oxytetracycline in genital tract tissues of postpartum cows given the drug by intravenous and intrauterine routes. American Journal of Veterinary Research 44, 764–769.

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