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Evaluating the effectiveness of a single application of 7% iodine tincture umbilical dip as a prevention of infection of the external umbilical structures in dairy calves

Open AccessPublished:May 12, 2022DOI:https://doi.org/10.3168/jds.2021-21418

      ABSTRACT

      The objective of this randomized clinical trial was to evaluate the effectiveness of a single application of 7% iodine tincture-based umbilical dip for preventing infection of the external umbilical structures in dairy calves. Five dairy farms in southern Ontario were visited twice weekly from September 2020 through June 2021. Female (n = 244) and male (n = 40) Holstein calves were randomly assigned at birth to receive either a 7% iodine tincture-based umbilical dip (n = 140) or no treatment (n = 144). Calves in the 7% iodine tincture umbilical dip group had the product administered once by the producer shortly after birth. For each newborn calf, the experimental group, calving difficulty, volume and timing of colostrum administration, time of birth, calving pen cleanliness, and the dam ID were recorded. Calf body weight was recorded during the first visit after birth, and a blood sample was collected for measurement of serum IgG concentration. Calves were health scored twice weekly from enrollment until approximately 30 d of age for assessment of external umbilical infection, joint inflammation, respiratory disease, and diarrhea. The primary outcome of the study was external umbilical infection, which was defined as an enlarged umbilicus with pain, heat, or a malodorous discharge. Calves were also weighed at 30 and 60 d to determine average daily gain. Serum IgG concentration and birth weight did not differ significantly between groups. Twenty-nine calves (20%) in the umbilical dip group developed an external umbilical infections, compared with 31 calves (22%) in the control group. A mixed logistic regression model, accounting for farm as a random effect, showed no effect of treatment on the incidence of an external umbilical infection. However, for every additional hour that calves received colostrum after birth, the odds of developing an external umbilical infection increased during the first month of life (odds ratio = 1.15; 95% confidence interval: 1.04–1.26). Additionally, treatment had no effect on respiratory disease, joint inflammation, diarrhea, average daily gain, or mortality, compared with the untreated control. These findings suggest that administering a single application of 7% iodine tincture dip to the umbilicus around the time of birth may not be effective for preventing external umbilical infections. Farm-level management factors, including colostrum management, appear to have more influence on risk of this disease.

      Key words

      INTRODUCTION

      The umbilical cord connects the placenta and the fetus during gestation to allow for the movement of oxygen, water, essential nutrients, and the removal of waste products (
      • Rings D.M.
      • Anderson D.E.
      Chapter 81—Umbilical surgery in calves.
      ;
      • Robinson A.L.
      • Timms L.L.
      • Stalder K.J.
      • Tyler H.D.
      Short communication: The effect of 4 antiseptic compounds on umbilical cord healing and infection rates in the first 24 hours in dairy calves from a commercial herd.
      ). This cord contains 2 umbilical arteries, an umbilical vein, and an urachus (
      • Trent A.M.
      Surgical management of umbilical masses in calves.
      ;
      • Rings D.M.
      • Anderson D.E.
      Chapter 81—Umbilical surgery in calves.
      ;
      • Ortved K.
      Miscellaneous abnormalities of the calf.
      ). During parturition, the umbilical cord separates from the fetus due to contraction of the smooth muscle around the umbilicus, which exposes the umbilical structures to the external environment (
      • Rings D.M.
      • Anderson D.E.
      Chapter 81—Umbilical surgery in calves.
      ). This exposure can allow pathogens to enter the umbilical cord, causing infection of the urachus, umbilical arteries (omphaloarteritis), or umbilical vein (omphalophlebitis;
      • Trent A.M.
      Surgical management of umbilical masses in calves.
      ;
      • Rings D.M.
      • Anderson D.E.
      Chapter 81—Umbilical surgery in calves.
      ). Infection of any or all the umbilical structures (omphalitis) can produce signs such as inflammation, localized heat, purulent or malodorous discharge, and pain (
      • Steerforth D.D.
      • Van Winden S.
      Development of clinical sign-based scoring system for assessment of omphalitis in neonatal calves.
      ). Furthermore, infection can lead to the development of bacteremia that can cause infection to localize within the joints of the calf and lead to lameness or within the liver and lead to septicemia (
      • House J.K.
      Umbilical enlargement.
      ;
      • Madigan J.E.
      Patent urachus, omphalitis, and other umbilical abnormalities.
      ;
      • Rings D.M.
      • Anderson D.E.
      Chapter 81—Umbilical surgery in calves.
      ).
      According to the
      • USDA
      Dairy 2014: Health and Management Practice on U.S. Dairy Operations, 2014.
      , infection of the umbilicus is the third most common illness in dairy calves, following digestive and respiratory diseases. However, the reported prevalence of umbilical infection has varied in the literature from 1% (
      • Svensson C.
      • Lundborg K.
      • Emanuelson U.
      • Olsson S.-O.
      Morbidity in Swedish dairy calves from birth to 90 days of age and individual calf-level risk factors for infectious diseases.
      ) to 34% (
      • Steerforth D.D.
      • Van Winden S.
      Development of clinical sign-based scoring system for assessment of omphalitis in neonatal calves.
      ). The development of an umbilical infection has been associated with decreased ADG (
      • Virtala A.M.K.
      • Mechor G.D.
      • Grohn Y.T.
      • Erb H.N.
      The effect of calfhood diseases on growth of female dairy calves during the first 3 months of life in New York state.
      ), occurrence of umbilical hernias (
      • Steenholdt C.
      • Hernandez J.
      Risk factors for umbilical hernia in Holstein heifers during the first two months after birth.
      ), and other neonatal morbidities such as joint infections (
      • Virtala A.M.K.
      • Mechor G.D.
      • Grohn Y.T.
      • Erb H.N.
      Morbidity from nonrespiratory diseases and mortality in dairy heifers during the first three months of life.
      ;
      • Miessa L.C.
      • Silva A.A.
      • Botteon R.C.C.M.
      • Botteon P.T.L.
      Morbidity and mortality by umbilical cord inflammation in dairy calves.
      ). Furthermore, umbilical disease has been linked to an increased risk of mortality (Retskii et al., 2007; Renaud et al., 2018), with the
      • USDA
      Dairy 2014: Health and Management Practice on U.S. Dairy Operations, 2014.
      reporting that 2% of neonatal calf mortality is related to an umbilical infection. Additionally, omphalitis has been observed to cause 23% of preslaughter mortality and 54% of postslaughter condemnation in veal calves (
      • Thomas G.W.
      • Jordaan P.
      Pre-slaughter mortality and post-slaughter wastage in bobby veal calves at a slaughter premises in New Zealand.
      ).
      To prevent umbilical infections, a wide variety of management practices are recommended, including using a specific type of bedding (
      • Faradonbeh Y.K.
      • Faradonbeh M.K.
      Evaluate the risk factors umbilical cord bacterial infection in calves in Shahrekord city.
      ), having excellent calving pen cleanliness (
      • Mee J.F.
      Newborn dairy calf management.
      ), applying colostrum management (
      • Mee J.F.
      Newborn dairy calf management.
      ), and using an umbilical antiseptic dip (
      • Grover W.M.
      • Godden S.
      Efficacy of a new navel dip to prevent umbilical infection in dairy calves.
      ;
      • Robinson A.L.
      • Timms L.L.
      • Stalder K.J.
      • Tyler H.D.
      Short communication: The effect of 4 antiseptic compounds on umbilical cord healing and infection rates in the first 24 hours in dairy calves from a commercial herd.
      ;
      • Scott P.
      Navel ill (Omphalitis/Omphaloplebitis).
      ). A high proportion of dairy producers use umbilical antiseptics, including approximately 40% of Canadian producers (
      • Renaud D.L.
      • Duffield T.F.
      • LeBlanc S.J.
      • Haley D.B.
      • Kelton D.F.
      Management practices for male calves on Canadian dairy farms.
      ), 88% of Czech producers (
      • Staněk S.
      • Zink V.
      • Doležal O.
      • Štolc L.
      Survey of preweaning dairy calf-rearing practices in Czech dairy herds.
      ), and 61% of French producers (
      • Lorino T.
      • Daudin J.-J.
      • Robin S.
      • Sanaa M.
      Factors associated with time to neonatal diarrhoea in French beef calves.
      ). Despite such a large proportion of dairy producers using an umbilical dip worldwide, observational studies have yielded conflicting results with regard to their effectiveness (
      • Donovan G.A.
      • Dohoo I.R.
      • Montgomery D.M.
      • Bennett F.L.
      Calf and disease factors affecting growth in female Holstein calves in Florida, USA.
      ;
      • Place N.T.
      • Heinrichs A.J.
      • Erb H.N.
      The effects of disease, management and nutrition on average daily gain of dairy heifers from birth to four months.
      ;
      • Windeyer M.C.
      • Leslie K.E.
      • Godden S.M.
      • Hodgins D.C.
      • Lissemore K.D.
      • LeBlanc S.J.
      Factors associated with morbidity, mortality, and growth of dairy heifer calves up to 3 months of age.
      ). Furthermore, in much of the clinical research, umbilical dips have been compared with each other without the inclusion of a negative control group (
      • Robinson A.L.
      • Timms L.L.
      • Stalder K.J.
      • Tyler H.D.
      Short communication: The effect of 4 antiseptic compounds on umbilical cord healing and infection rates in the first 24 hours in dairy calves from a commercial herd.
      ;
      • Wieland M.
      • Mann S.
      • Guard C.L.
      • Nydam D.V.
      The influence of 3 different navel dips on calf health, growth performance, and umbilical infection assessed by clinical and ultrasonographic examination.
      ;
      • Fordyce A.L.
      • Timms L.L.
      • Stalder K.J.
      • Tyler H.D.
      The effect of novel antiseptic compounds on umbilical cord healing and incidence of infection in dairy calves.
      ).
      • Grover W.M.
      • Godden S.
      Efficacy of a new navel dip to prevent umbilical infection in dairy calves.
      reported the only clinical trial to compare the use of umbilical disinfectants with an untreated control. The results of that study showed that treatment with a novel umbilical disinfectant, Navel Guard (NG; purified water, acidified water, isopropyl alcohol, surfactant, citric acid, FD&C Red #40, FD&C Yellow #5; SCG-Solutions Inc.) was more effective at reducing umbilical infections compared with no treatment. However, no difference was found between the NG group and either of the iodine groups in the study (
      • Grover W.M.
      • Godden S.
      Efficacy of a new navel dip to prevent umbilical infection in dairy calves.
      ). A limitation of this study was that 7% iodine tincture was not compared with an untreated control group to demonstrate its effectiveness (
      • Grover W.M.
      • Godden S.
      Efficacy of a new navel dip to prevent umbilical infection in dairy calves.
      ). Given that the most commonly recommended and widely used umbilical dip is a 7% iodine tincture mixture (
      • Hadley F.B.
      Navel infection.
      ;
      • Grover W.M.
      • Godden S.
      Efficacy of a new navel dip to prevent umbilical infection in dairy calves.
      ;
      • Robinson A.L.
      • Timms L.L.
      • Stalder K.J.
      • Tyler H.D.
      Short communication: The effect of 4 antiseptic compounds on umbilical cord healing and infection rates in the first 24 hours in dairy calves from a commercial herd.
      ), data regarding its effectiveness are needed.
      The primary objective of the current study was to evaluate the effectiveness of a 7% iodine-based umbilical disinfectant relative to no treatment as a preventative method to reduce the incidence of external umbilical infections in neonatal calves. We hypothesized that calves receiving a single dose of 7% iodine tincture to the umbilicus shortly after birth would have a lower incidence of external umbilical infections compared with an untreated control group. A secondary objective was to determine management practices associated with the occurrence of an external umbilical infection.

      MATERIALS AND METHODS

      This study is reported using guidelines for randomized control trials in livestock and food safety (REFLECT;
      • O'Connor A.
      Reporting guidelines for primary research: Saying what you did.
      ). The randomized controlled trial was conducted between September 2020 and June 2021, at 5 commercial dairy farms in southwestern Ontario, Canada. Animal use and experimental protocols were approved by the University of Guelph Animal Care Committee (AUP#4455).

      Farm and Calf Eligibility for Enrollment

      The participating dairy farms were located in southwestern Ontario and were a convenience sample of farms in the region. They were selected due to their willingness to participate and their proximity to the University of Guelph. None of the enrolled farms had a history of using navel dips at the time of enrollment. Before beginning the study, all producers were taught to fill a disposable cup with approximately 50 mL of 7% iodine tincture and submerge the umbilical stump for 3 to 5 s. Producers also received instruction on how to fill out a calving information form. Four of the farms operated freestall dairies, milking 100 to 300 cows, while one farm operated a tiestall barn milking 80 cows.
      All live female Holstein calves born between September 2020 and April 2021 were eligible to be enrolled into the study at birth and were followed for approximately 60 d. Only male Holstein calves from farm 3 were eligible for inclusion in the trial because they were kept on site for the full 60-d study period. Any calf that died within the 60-d study period was included; however, calves born dead were not included. All calves with missing data were included and censored, and details of the missing data are reported.

      Calf Housing

      At all facilities, calves were moved from a calving pen into an individual pen for the first 2 to 3 wk of life. Calves were then moved into group housing pens, with farms 1, 2, 3, and 5 bedding with straw and farm 4 bedding with wood shavings. Farms 1 and 2 fed approximately 8 L of milk replacer a day using mob feeders, while Farms 4 and 5 fed whole milk ad libitum using automatic milk feeders. Farm 3 differed in management strategies between calves, with heifers being fed milk replacer ad libitum using an automatic milk feeder and males being fed 3 L of milk replacer twice a day using buckets.

      Enrollment and Randomization

      Following calving, producers selected the topmost envelope in a randomized stack that contained a calving form with the specified experimental group: either a 7% iodine tincture disinfectant dip (I; Stronger, Dominion Veterinary Laboratories Ltd.) or no disinfectant control (C). The groups were randomized in blocks of 4 (2 controls, 2 treatments) in Microsoft Excel (Microsoft Corp.) to account for the effect of time. The order was concealed from the producers, and each calving resulted in the selection of the next envelope in the sequence.
      If the calf was allocated to the umbilical dip group, producers used a disposable paper cup to administer the 7% iodine tincture solution directly to the umbilical cord to ensure that the exposed cord and surrounding area were submerged. Each disposable cup was used for a single application of iodine to prevent possible cross-contamination of pathogens between calves. The administration of the umbilical dip was completed by the producer after the birth of each calf, and the amount of time in hours following birth to navel dipping was recorded in the calving form. The approximate amount of time in hours for unobserved births was estimated based on the time that producers were last in the barn. Producers were also asked to record calf ID, dam ID, calf sex, approximate time of birth, estimated time of first colostrum feeding after birth (hours), approximate amount of first colostrum fed (liters), and if the calf was a twin. Producers were also asked to score the calving difficulty on a scale from 1 to 4 (1 = unassisted; 2 = easy pull; 3 = hard pull; 4 = surgical/veterinarian assistance) that was adapted from
      • Bennett G.L.
      • Gregory K.E.
      Genetic (co) variances for calving difficulty score in composite and parental populations of beef cattle: I. Calving difficulty score, birth weight, weaning weight, and postweaning gain.
      . Producers were additionally asked to score calving pen cleanliness on a scale from 1 to 4 (1 = clean, no manure visible; 2 = almost clean, less than 25% of area covered with manure; 3 = somewhat contaminated, between 25 and 75% of area covered with manure; 4 = contaminated, over 75% of the pen covered with manure). At each visit, researchers collected the calving information forms and officially enrolled the calf into the trial.

      Blinding

      Producers were not blinded during the study. However, the researcher evaluating all outcomes was blinded to which group animals were in.

      Data Collection and Outcomes

      Calf Health Scoring.

      Health scoring of all enrolled calves occurred at the first visit and twice weekly (Monday and Thursday) until the final visit at approximately 30 d of age. Health scoring was completed by a single researcher (MBVC) throughout the trial to remove concerns associated with interrater reliability. In addition, latex gloves were changed, and equipment (e.g., rectal thermometer) used as part of the examination was disinfected using accelerated hydrogen peroxide disinfectant wipes between calves to minimize the transfer of pathogens. During each on-farm visit, researchers used an iPad (Apple Inc.) with Qualtrics software (http://www.qualtrics.com/) forms to record the health data. Specifically, all the following factors were evaluated at every visit. Infection of the external umbilical structures (umbilical infection) was scored using a 4-point ordinal scale (adapted from
      • Fecteau G.
      • Pare J.
      • Van Metre D.
      • Smith B.
      • Holmberg C.
      • Guterbock W.
      • Jang S.
      Use of a clinical sepsis score for predicting bacteremia in neonatal dairy calves on a calf rearing farm.
      ): 0 = normal; 1 = slightly enlarged, not warm or painful; 2 = slightly enlarged with slight pain or moisture; and 3 = enlarged with heat, pain, or malodorous discharge. A score of 3 was diagnostic for the presence of an external umbilical infection (adapted from
      • Fecteau G.
      • Pare J.
      • Van Metre D.
      • Smith B.
      • Holmberg C.
      • Guterbock W.
      • Jang S.
      Use of a clinical sepsis score for predicting bacteremia in neonatal dairy calves on a calf rearing farm.
      ). Diarrhea was scored using a 4-point ordinal scale: 0 = normal (firm but not hard, original form is distorted slightly after dropping to floor and settling); 1 = soft (does not hold form, piles but spreads slightly); 2 = runny (spreads readily); and 3 = watery (liquid consistency, splatters). A score of 2 or 3 indicated a diagnosis of diarrhea (
      • Renaud D.L.
      • Steele M.A.
      • Genore R.
      • Roche S.M.
      • Winder C.B.
      Passive immunity and colostrum management practices on Ontario dairy farms and auction facilities: A cross-sectional study.
      ). Ear, nose, eyes, cough, and rectal temperature were used calculate a respiratory score (
      • McGuirk S.M.
      • Peek S.
      Timely diagnosis of dairy calf respiratory disease using a standardized scoring system.
      ). Calves were considered positive for bovine respiratory disease (BRD) when the cumulative score of the ears, nose, eyes, cough, and rectal temperature was ≥5, as described by
      • McGuirk S.M.
      • Peek S.
      Timely diagnosis of dairy calf respiratory disease using a standardized scoring system.
      . Joint inflammation was determined by palpation of all 4 limbs. It was diagnosed if the joint had swelling or was warm or painful when palpated, or if lameness was noted. An umbilical hernia was diagnosed if a hernial ring was detected or if a protrusion of the abdominal structures was identified by palpation (
      • Wieland M.
      • Mann S.
      • Guard C.L.
      • Nydam D.V.
      The influence of 3 different navel dips on calf health, growth performance, and umbilical infection assessed by clinical and ultrasonographic examination.
      ). Conditions that required antimicrobials were determined and treated by the farm owner using established farm protocols. All treatments given during the first 30 d of life and any mortality were recorded into a Qualtrics form.

      Determining the Concentration of Serum IgG.

      Newly enrolled calves that were over 24 h of age had a blood sample taken during the first farm visit after their birth. Calves that were under 24 h of age at their first visit had blood samples taken at the next semiweekly visit. Each blood sample was collected from the jugular vein of the calf using a 20-gauge 1-inch needle into a 10-mL blood vacuum container without an anticoagulant (BD Vacutainer; Becton, Dickson and Co.). Blood samples were centrifuged at 1,500 × g for 15 min at the University of Guelph. Serum was separated and stored in a −20°C freezer until analysis at the Saskatoon Colostrum Company (Saskatoon, SK, Canada) where serum IgG was evaluated by radial immunodiffusion as described by
      • Chelack B.J.
      • Morley P.S.
      • Haines D.M.
      Evaluation of methods for dehydration of bovine colostrum for total replacement of normal colostrum in calves.
      .

      Average Daily Gain.

      Each calf was weighed at 3 separate point during the trial period using a portable digital scale (S3, Tru-Test). The first weight (W1) was taken upon the first visit after birth, the second weight (W2) was obtained following a calf's final health score at approximately 30 d of age, and the third weight (W3) was conducted at approximately 60 d of age, just before weaning. Average daily gain was calculated from birth to 30 d using (W2 − W1)/number of days between weight measurements and from birth to 60 d using (W3 − W1)/number of days between weight measurements.

      Sample Size Calculation.

      A sample size calculation was performed before the start of the trial using Stata 15 (Stata/IC Version 15.1 for Mac, StataCorp). Based on
      • Wieland M.
      • Mann S.
      • Guard C.L.
      • Nydam D.V.
      The influence of 3 different navel dips on calf health, growth performance, and umbilical infection assessed by clinical and ultrasonographic examination.
      and
      • Renaud D.L.
      • Duffield T.F.
      • LeBlanc S.J.
      • Ferguson S.
      • Haley D.B.
      • Kelton D.F.
      Risk factors associated with mortality at a milk-fed veal calf facility: A prospective cohort study.
      , the incidence of umbilical infections in calves was assumed to be approximately 27%. The anticipated reduction in morbidity for the treatment group was estimated to be 50% (
      • Grover W.M.
      • Godden S.
      Efficacy of a new navel dip to prevent umbilical infection in dairy calves.
      ). Therefore, using a 95% CI and 80% power, a minimum sample size of 276 calves or 138 calves per group was calculated.

      Statistical Analysis

      All data were transferred from Qualtrics into Excel and then was uploaded into Stata 15 for further analysis. Descriptive statistics were generated for each variable, and each variable was reviewed for normality and variation. Only calves with complete information were used in the analyses (complete-case analysis;
      • Pigott T.D.
      A review of methods for missing data.
      ).
      An external umbilical infection was the primary outcome of this study and was analyzed using a mixed logistic regression model (0 = normal, 1 = diseased). The outcomes of BRD and diarrhea were analyzed as a single metric per calf of the proportion of days with a positive health score (0–100%) using a generalized linear model with a logit link and binomial family. Mortality (0 = alive, 1 = dead), antimicrobial treatment (0 = untreated, 1 = treated), and joint inflammation (0 = no inflammation, 1 = inflammation) were assessed using a mixed logistic regression model, whereas ADG was analyzed using a mixed linear regression model. In all models, a random effect for farm was added to account for differences that existed between farms. All collected variables were checked for independence from experimental groups via chi-squared tests for categorical variables and t-tests for continuous variables (Table 1).
      Table 1Descriptive characteristics of variables collected in a randomized clinical trial design to evaluate the effectiveness of a single application of 7% iodine tincture umbilical dip
      Data are stratified and tested for statistical difference between experimental groups (P-value), categorical variables [count (%)], and continuous variables [count; mean ± SD (for normally distributed variables) or median (range) (for nonnormally distributed variables)].
      VariableDescriptionControl
      Control = untreated control group.
      (n = 144)
      Iodine
      Iodine = 7% iodine tincture group.
      (n = 140)
      P-value
      Farm125 (17)21 (15)0.93
      229 (20)30 (21)
      345 (31)42 (30)
      435 (24)39 (28)
      510 (7)8 (6)
      SexFemale123 (85)121 (86)0.81
      Male21 (15)19 (14)
      Season of birthFall (September–November)57 (40)50 (36)0.50
      Winter (December–February)59 (41)67 (48)
      Spring (March–April)28 (19)23 (16)
      IgG, g/dL<19.439 (27)30 (21)0.98
      ≥19.4 to <26.8539 (27)33 (24)
      ≥26.85 to <36.236 (25)33 (24)
      ≥36.229 (20)43 (31)
      Time of birthOvernight: 0001–0600 h39 (27)58 (41)0.08
      Morning: 0601–1200 h45 (31)34 (24)
      Afternoon: 1201–1800 h40 (28)34 (24)
      Evening: 1801–0000 h20 (14)14 (10)
      Approximate time of navel dip after birth, h3.9 (0 to 15)
      Approximate time of colostrum feeding after birth, h4.0 (0 to 14)4.0 (0 to 15)0.23
      Approximate amount of colostrum fed, L2.5 ± 0.862.5 ± 1.000.47
      Calving scoreUnassisted129 (90)137 (98)0.04
      Assisted15 (10)3 (2)
      Calving pen cleanliness scoreClean, no manure visible62 (43)52 (37)0.56
      Almost clean, <25% of area covered with manure58 (40)60 (43)
      Somewhat contaminated, 25–75% of area covered with manure20 (14)20 (14)
      Contaminated, >75% of the pen covered with manure4 (3)8 (6)
      TwinsNo135 (94)136 (97)0.17
      Yes9 (6)4 (3)
      Dam lactation number167 (47)60 (43)0.58
      237 (26)32 (23)
      ≥340 (28)47 (34)
      Number of health scores conducted1,270; 8.8 ± 1.71,271; 9.1 ± 1.20.23
      1 Data are stratified and tested for statistical difference between experimental groups (P-value), categorical variables [count (%)], and continuous variables [count; mean ± SD (for normally distributed variables) or median (range) (for nonnormally distributed variables)].
      2 Control = untreated control group.
      3 Iodine = 7% iodine tincture group.
      For each model, experimental group (I or C), calving difficulty (ordinal variable with 4 categories), calving pen cleanliness (ordinal variable with 4 categories), IgG (g/dL), season of birth (ordinal variable with 3 categories), time of birth (ordinal variable with 4 categories), time to administer first dose of colostrum (hours), and amount of colostrum given (liters) were assessed. Experimental group was forced into each model because it was considered the main variable of interest. For the mortality model, the previously mentioned independent variables were assessed along with morbidity variables, including the presence of an external umbilical infection, umbilical hernia, antimicrobial treatment for BRD, and antimicrobial treatment for diarrhea.
      All independent variables were assessed for co-linearity before analysis. Continuous variables were checked for linearity graphically, and if the variables had a nonlinear relationship with the outcome, they were then categorized. Univariable analysis was conducted on all the independent variables. Only variables that were potentially significant using a liberal P-value (P < 0.20) were offered to the multivariable model. Backward selection was used in building the multivariable models, where variables that were significant (P ≤ 0.05) or tended to be significant (P > 0.05 to P < 0.10) were retained. If the coefficient of a significant variable had a >25% change, the variable was classified as a confounder and retained in the model. Only biologically plausible interaction terms were evaluated, and significant interactions were included in the final models. Final logistic models were assessed using a Hosmer-Lemeshow goodness of fit test (P < 0.05), and Pearson and deviance residuals were assessed graphically to identify outliers, which were then examined to ensure the data were correct. No outliers were removed from any analyses. Mixed linear regression models were assessed for homoscedasticity by visually assessing standardized residuals, normality was assessed using a quantile-normal plot, and best linear unbiased predictions were also assessed graphically for homoscedasticity and normality.

      RESULTS

      In total, 284 female (n = 244) and male (n = 40) Holstein calves were enrolled in the trial, with 140 receiving 7% iodine tincture and 144 serving as negative controls. All enrolled calves were enrolled at birth and were followed until 60 d of age (Figure 1).
      Figure thumbnail gr1
      Figure 1Flow diagram displaying timeline of data collection for a randomized clinical trial design to evaluate the efficacy of a single application of 7% iodine tincture umbilical dip (
      • CONSORT (Consolidated Standards of Reporting Trials)
      CONSORT 2010 Flow Diagram.
      ).

      Baseline Data

      Baseline data are described in Table 1 and are stratified by I and C experimental groups. The level of failed transfer of passive immunity (defined as <10 g/L) was 6.7%. No significant differences were found between I and C groups for baseline variables, except for calving score for which C calves had a higher level of assistance than I calves (Table 1).

      External Umbilical Infections and Umbilical Hernias

      During the 30 d that calves were assessed twice weekly, 20% (29/144) and 22% (31/140) of calves in the C and I groups developed an external umbilical infection, respectively. In a multivariable logistic regression model, no differences were found between groups with respect to the development of an external umbilical infection (Table 2). The timing of colostrum feeding influenced the development of an external umbilical infection, with every hour delay in providing colostrum after birth increasing the odds for developing an external umbilical infection by 1.15 times. Additionally, calves born to third-lactation dams tended to have greater odds of developing an external umbilical infection compared with calves born to a first-lactation dam (Table 2).
      Table 2Final mixed multilevel logistic regression model
      Controlling for farm as a random effect.
      evaluating factors associated with the occurrence of an external umbilical infection (0 = normal, 1 = infection) in calves (n = 288) enrolled in a randomized clinical trial design to evaluate the effectiveness of a single application of 7% iodine tincture umbilical dip
      ItemVariable
      Control = untreated control group; iodine = 7% iodine tincture group.
      Odds ratioP-value95% CI
      Experimental groupControlReferent
      Iodine1.070.840.58 to 1.95
      Lactation number1Referent
      20.670.350.30 to 1.54
      ≥31.850.080.94 to 3.65
      Time to first colostrum feedingEvery 1 h increase1.150.011.04 to 1.26
      1 Controlling for farm as a random effect.
      2 Control = untreated control group; iodine = 7% iodine tincture group.
      With respect to umbilical hernias, 6% (8/144) and 9% (13/140) of the calves in the C and I groups developed an umbilical hernia. No differences were found between any of the measured variables and the development of an umbilical hernia, including experimental group, with no statistical differences being observed between groups [I vs. C: odds ratio (OR) = 1.83; 95% CI: 0.72–4.63; P = 0.20].

      Joint Inflammation

      Joint inflammation was identified in 6% (9/144) and 9% (12/140) of calves in the C and I groups; however, none of the calves received antimicrobial therapy for joint inflammation. No differences were found between any of the measured variables and the development of joint inflammation, including experimental group (I vs. C: OR = 1.41; 95% CI: 0.57–3.45; P = 0.46).

      Diarrhea

      In total, 8% (11/144) and 9% (12/140) of calves in the C and I groups were treated for diarrhea at least once with antimicrobials drugs, with no difference between groups based on a logistic regression model (I vs. C: OR = 1.13; 95% CI: 0.48–2.66; P = 0.77). The mean proportion of days that calves had diarrhea in semiweekly assessments in the first 30 d after birth were 21% (SD, ±0.16) and 21% (SD, ±0.16) in the C and I groups, respectively. The final generalized linear model evaluating variables associated with proportion of days with diarrhea is described in Table 3. Calves born in the winter and spring had a lower proportion of days with diarrhea compared with calves born in the fall. Additionally, calves born in the evening had a lower proportion of days with diarrhea compared with calves born overnight (Table 3); however, no differences were identified between experimental groups.
      Table 3Final generalized linear model evaluating factors associated with the proportion of days with diarrhea in calves (n = 288) enrolled in a randomized clinical trial design to evaluate the effectiveness of a single application of 7% iodine tincture umbilical dip
      ItemVariable
      Control = untreated control group; iodine = 7% iodine tincture group; overnight = 0001–0600 h; morning = 0601–1200 h; afternoon = 1201–1800 h; evening = 1801–0000 h; fall = September, October, and November; winter = December, January, and February; spring = March and April.
      Relative proportion ratioP-value95% CI
      Experimental groupControlReferent
      Iodine1.170.660.59 to 2.31
      Season of birthFallReferent
      Winter0.320.010.13 to 0.77
      Spring0.230.010.09 to 0.64
      Time of birthOvernightReferent
      Morning1.070.880.45 to 2.53
      Afternoon1.770.270.64 to 4.90
      Evening0.270.010.11 to 0.70
      1 Control = untreated control group; iodine = 7% iodine tincture group; overnight = 0001–0600 h; morning = 0601–1200 h; afternoon = 1201–1800 h; evening = 1801–0000 h; fall = September, October, and November; winter = December, January, and February; spring = March and April.

      Bovine Respiratory Disease

      Among calves in the C and I groups, 26% (38/144) and 30% (42/140) were treated for BRD at least once with antimicrobial drugs. No variables were associated with antimicrobial drug treatment of BRD, including experimental group in a logistic regression model (I vs. C: OR = 1.16; 95% CI: 0.67–1.99; P = 0.59). In semiweekly assessments in the first 30 d after birth, the mean proportion of days with BRD were 17% (SD, ±0.19) and 17% (SD, ±0.18) of calves in the C and I groups, respectively. No variables were found to be associated with the proportion of days with BRD, including experimental group (I vs. C: relative proportion ratio = 1.18; 95% CI: 0.73–1.92; P = 0.51).

      Mortality

      During the 60-d period that each calf was followed, 10% (15/144) and 9% (12/140) of calves in the C and I groups died, respectively. A mixed multilevel logistic regression found no variables to be associated with the occurrence of mortality, including experimental group (I vs. C: OR = 0.76; 95% CI: 0.34–1.72; P = 0.51).

      Average Daily Gain

      Average daily gain from birth to 30 d was 0.57 kg (SD, ±0.20) and 0.59 kg (SD, ±0.18) in the C and I groups, respectively, whereas ADG from birth to 60 d was 0.67 kg (SD, ±0.09) and 0.69 kg (SD, ±0.18) in the C and I groups, respectively. Two separate mixed multilevel linear regression models were created to assess ADG from birth to 30 d and from birth to 60 d, both of which are further described in Table 4, Table 5, respectively. Specifically, in the model evaluating ADG from birth to 30 d, calves that had a higher concentration of IgG gained more weight per day, compared with calves with a lower IgG concentration. Furthermore, calves born in the morning gained more weight per day, respectively, compared with calves born overnight. Lastly, calves born from a third or greater lactation dam gained more weight per day more compared with calves born to a first-lactation dam (Table 4).
      Table 4Final mixed multilevel linear regression evaluating factors associated with ADG (kg/d) from birth to 30 d in calves (n = 258) enrolled in a randomized clinical trial design to evaluate the effectiveness of a single application of 7% iodine tincture umbilical dip
      ItemVariable
      Control = untreated control group; iodine = 7% iodine tincture group; overnight = 0001–0600 h; morning = 0601–1200 h; afternoon = 1201–1800 h; evening = 1801–0000 h.
      Coefficient, kg/dP-value95% CI
      Experimental groupControlReferent
      Iodine0.020.45−0.03 to 0.06
      IgG, g/dL<19.4Referent
      ≥19.4 to <26.850.050.10−0.01 to 0.11
      ≥26.85 to <36.20.080.020.01 to 0.14
      ≥36.20.110.010.05 to 0.17
      Time of birthOvernightReferent
      Morning0.060.030.01 to 0.11
      Afternoon0.050.08−0.004 to 0.10
      Evening0.040.24−0.03 to 0.11
      Lactation number1Referent
      20.040.11−0.01 to 0.10
      ≥30.060.010.01 to 0.11
      Intercept0.46<0.010.36 to 0.55
      1 Control = untreated control group; iodine = 7% iodine tincture group; overnight = 0001–0600 h; morning = 0601–1200 h; afternoon = 1201–1800 h; evening = 1801–0000 h.
      Table 5Final mixed multilevel linear regression evaluating factors associated with ADG (kg/d) from birth to 60 d in calves (n = 246) enrolled in a randomized clinical trial design to evaluate the effectiveness of a single application of 7% iodine tincture umbilical dip
      ItemVariable
      Control = untreated control group; iodine = 7% iodine tincture group; fall = September, October, and November; winter = December, January and February; spring = March and April.
      Coefficient, kg/dP-value95% CI
      Experimental groupControlReferent
      Iodine0.020.31−0.02 to 0.06
      Season of birthFallReferent
      Winter0.010.65−0.04 to 0.05
      Spring0.060.040.004 to 0.11
      IgG, g/dL<19.4Referent
      ≥19.4 to <26.850.050.13−0.01 to 0.10
      ≥26.85 to <36.20.060.040.004 to 0.12
      ≥36.20.070.020.01 to 0.12
      Intercept0.53<0.010.44 to 0.62
      1 Control = untreated control group; iodine = 7% iodine tincture group; fall = September, October, and November; winter = December, January and February; spring = March and April.
      In the model assessing ADG from birth to 60 d, calves born in the spring gained 0.06 kg per day more compared with calves born in the fall. In addition, calves that had IgG of ≥26.85 to <36.2 g/L or ≥36.2 g/L gained 0.06 kg and 0.07 kg more per day, respectively, than calves with IgG < 19.4 g/L (Table 5).

      DISCUSSION

      Effectiveness of 7% Iodine Umbilical Dip

      Few studies have compared calves treated with an umbilical dip to a negative control group. This lack of research could be due to the long-standing use of umbilical dips within the dairy industry creating a confirmation bias that they are an effective disease prevention tool. This practice includes the use of 7% iodine, which has traditionally been readily available to producers and has been proven to have antiseptic properties (
      • World Health Organization (WHO)
      Care of the Umbilical Cord: A Review of the Evidence. Reproductive Health (Technical Support).
      ). To our knowledge, the current study is the first randomized control trial to evaluate the effectiveness of a single application of 7% iodine tincture compared with no treatment in neonatal dairy calves. Other studies have evaluated and compared the use of multiple umbilical antiseptic dips, such as
      • Wieland M.
      • Mann S.
      • Guard C.L.
      • Nydam D.V.
      The influence of 3 different navel dips on calf health, growth performance, and umbilical infection assessed by clinical and ultrasonographic examination.
      (NG, 7% iodine tincture vs. 2% chlorhexidine gluconate),
      • Robinson A.L.
      • Timms L.L.
      • Stalder K.J.
      • Tyler H.D.
      Short communication: The effect of 4 antiseptic compounds on umbilical cord healing and infection rates in the first 24 hours in dairy calves from a commercial herd.
      (7% tincture of iodine vs. 0.1% chlorine vs. chlorohexidine gluconate 4.0% wt/vol vs. 10% trisodium citrate),
      • Grover W.M.
      • Godden S.
      Efficacy of a new navel dip to prevent umbilical infection in dairy calves.
      (NG vs. 7% iodine tincture vs. 2% chlorhexidine gluconate), and
      • Fordyce A.L.
      • Timms L.L.
      • Stalder K.J.
      • Tyler H.D.
      The effect of novel antiseptic compounds on umbilical cord healing and incidence of infection in dairy calves.
      [7% iodine vs. a dry dip formulated using an antibacterial peptide (nisin) mixed with talc, liquid nisin vs. 4% chlorhexidine mixed with alcohol in a 50:50 solution]. None of these trials found any treatment differences between the various umbilical antiseptic dips and umbilical infections. Given that each of these trials incorporated a 7% iodine tincture treatment group, the potential exists for many of the umbilical dips examined in these trials to also be ineffective compared with no treatment. However, our results reflect a single clinical trial, and additional clinical research would be needed to determine the true effectiveness of other umbilical dips compared with no treatment.
      Applying a single dose of 7% iodine tincture to the umbilicus of the calf shortly after birth could be another reason for the lack of significant results. Currently, in human medicine, the recommendation is to reapply povidone-iodine multiple times a day to prevent infection (
      • NPS (National Prescribing Service)
      Betadine Antiseptic Topical Ointment.
      ). The World Health Organization also recommends daily umbilical cord care for the first week of life for home births or for births in regions with high neonatal morbidity. Reapplying treatment is important because the antiseptic characteristics of 7% iodine tincture decrease 15 min after its application and only remain active for several hours (
      • World Health Organization (WHO)
      Care of the Umbilical Cord: A Review of the Evidence. Reproductive Health (Technical Support).
      ). Therefore, the effective period of 7% iodine tincture for preventing infection in calves possibly ended before the umbilical cord naturally dried, which has been observed to occur between 1 and 8 d of age (
      • Hides S.J.
      • Hannah M.C.
      Drying times of umbilical cords of dairy calves.
      ). Research by
      • Faradonbeh Y.K.
      • Faradonbeh M.K.
      Evaluate the risk factors umbilical cord bacterial infection in calves in Shahrekord city.
      examined calves (n = 100) with umbilical infections and found that disinfecting the umbilical cord 3 times was associated with the lowest frequency of umbilical infection (20%), followed by dipping twice (35%) and dipping only once (45%). Although that study did not include a comparison with no disinfection of the umbilicus, it is indicative for future randomized control trials to test the effects of multiple doses of umbilical dips with no treatment.

      Umbilical Infections

      Many risk factors for umbilical infections have been studied, but few have proven to be associated with the occurrence of this disease and many are still debated in the literature. Within this trial, the only association with umbilical infections identified was the timing of the first colostrum feeding, with calves that received colostrum later having higher odds of external umbilical infections. These results agree with expert opinions, such as
      • Mee J.F.
      Newborn dairy calf management.
      , which recommended “adequate early intake of good quality colostrum” as a preventative measure for navel ill.
      Importantly, IgG (g/dL) concentrations did not have a significant impact on umbilical infections. This lack of influence is potentially due to the above-average colostrum feeding regimens that all the study farms exhibited. Generally, calves with an IgG below 10 g/L are considered to have failed transfer of passive immunity (FTPI;
      • Tyler J.W.
      • Hancock D.D.
      • Parish S.M.
      • Rea D.E.
      • Besser T.E.
      • Sanders S.G.
      • Wilson L.K.
      Evaluation of 3 assays for failure of passive transfer in calves.
      ;
      • Weaver D.M.
      • Tyler J.W.
      • VanMetre D.C.
      • Hostetler D.E.
      • Barrington G.M.
      Passive transfer of colostral immunoglobulins in calves.
      ;
      • Beam A.L.
      • Lombard J.E.
      • Kopral C.A.
      • Garber L.P.
      • Winter A.L.
      • Hicks J.A.
      • Schlater J.L.
      Prevalence of failure of passive transfer of immunity in newborn heifer calves and associated management practices on US dairy operations.
      ); however, within this trial, few calves (6.7%) fell within this category, regardless of experimental group, compared with a reported FTPI of 24% in a cross-sectional study of Ontario dairy calves (
      • Renaud D.L.
      • Steele M.A.
      • Genore R.
      • Roche S.M.
      • Winder C.B.
      Passive immunity and colostrum management practices on Ontario dairy farms and auction facilities: A cross-sectional study.
      ). Despite the low level of FTPI, calves with higher levels of IgG were found to have gained more in the first 30 and 60 d of life, highlighting the importance of excellent colostrum management. In the future, conducting similar trials on farms with a greater variation in colostrum management practices may reveal a substantial IgG influence and further emphasize the importance of colostrum in terms of umbilical infections, diarrhea, BRD, and mortality.

      Limitations

      Several limitations need to be considered in interpreting the results of this work. First, this study relied heavily on producers to not only administer the treatments but to also record data via the calving forms. The lack of blinding was a necessary forfeiture to allow application of the treatment at the appropriate time. However, it has the potential to lead to participation bias due to the producer possibly increasing or reducing the treatment dose depending on the calf being born. Additionally, recall of time of birth, colostrum management factors, and the requirement to apply the treatment itself may have influenced the results of this study. However, to minimize potential error and bias, farms were visited twice weekly and all calving information was checked to ensure that the producer properly filled out all the required information. Calves were also randomly assigned to receive either the treatment (I) or control (C) as a means of reducing producer selection bias of providing I to perceived higher value calves. It should also be noted that although the researcher evaluating calves was unaware of whether an animal received treatment, the iodine used in this study stained the navels of the calves. Consequently, a distinguishable visible difference existed between I and C groups. Although these potential drawbacks are important to address, asking producers to administer the treatments has better real-world application and these drawbacks are minimized by having adequately powered studies.
      The inability of this study to detect abnormalities of the different internal umbilical structures is another potential limitation as we could have underestimated the number of infections. However, previous research by
      • Steerforth D.D.
      • Van Winden S.
      Development of clinical sign-based scoring system for assessment of omphalitis in neonatal calves.
      found that the external clinical scoring system had 85.9% sensitivity and 74.8% specificity and it accurately diagnosed 78.6% of the enrolled calves. In addition,
      • Wieland M.
      • Mann S.
      • Guard C.L.
      • Nydam D.V.
      The influence of 3 different navel dips on calf health, growth performance, and umbilical infection assessed by clinical and ultrasonographic examination.
      found a moderate to good level of agreement for detection of omphalitis and very good agreement for detection of omphalophlebitis when they compared clinical scoring and ultrasonographic evaluation. This outcome suggests that this limitation could have been minimized; however, future research is needed to understand the impact of umbilical dips on internal umbilical infections.
      Researchers conducting health scores only twice weekly was another limitation for this study. Given the time between visits, positive umbilical infection, BRD, and diarrhea scores may have been missed. Missed positive disease scores may have decreased the true prevalence of disease used in the final models and resulted in bias toward a null effect. However, the time between visits was kept constant throughout the trial (Monday and Thursday) in an attempt to ensure adequate reporting of disease.

      CONCLUSIONS

      Administering a single dose of a 7% iodine tincture to the umbilicus of calves shortly after birth was not effective in reducing external umbilical infections during the first 30 d of life compared with no treatment. The risk of external umbilical infections was significantly reduced in calves that received colostrum sooner after birth. Future clinical research should compare the use of other recommended dips with no treatment to determine their effectiveness and to analyze the impact of multiple doses at various time points on umbilical health.

      ACKNOWLEDGMENTS

      This project was funded by Veal Farmers of Ontario (Guelph, ON, Canada) and the Ontario Ministry of Agriculture, Food, and Rural Affairs–University of Guelph Ontario Agri-Food Innovation Alliance Research Program (Guelph, ON, Canada). The authors extend special thanks to the producers and research students that made this trial possible. The authors have not stated any conflicts of interest.