Effects of a nutrient-enriched water with and without poultry flavoring on water intake, urine specific gravity, and urine output in healthy domestic cats fed a dry kibble diet

Brian M. Zanghi Nestlé Purina Research, 1 Checkerboard Sq, St Louis, MO 63102.

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Emma Wils-Plotz Nestlé Purina Research, 1 Checkerboard Sq, St Louis, MO 63102.

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Staci DeGeer Nestlé Purina Research, 1 Checkerboard Sq, St Louis, MO 63102.

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Cari L. Gardner Nestlé Purina Research, 1 Checkerboard Sq, St Louis, MO 63102.

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Abstract

OBJECTIVE To investigate water intake and urine measures in healthy cats provided free-choice access to a nutrient-enriched water with (NWP) or without (NW) added poultry flavoring offered at 3 different volumes in addition to tap water (TW).

ANIMALS 36 domestic shorthair cats.

PROCEDURES Control group cats (n = 4) received dry food with TW ad libitum throughout the study. Cats of the NW and NWP groups (n = 16/group) received the same food with TW only (period 1; 7 days) followed by TW and the assigned treatment ad libitum at 1X, 1.5X, and 2X the volume of TW consumed in period 1 during periods 2 (17 days), 3 (10 days), and 4 (10 days), respectively. Liquid consumption, food intake, and total water intake (from all sources) were measured; urine collected over 48 hours in each period was measured, and urine specific gravity (USG) was determined. Data were analyzed with mixed-effects models.

RESULTS TW and food calorie intake were similar among groups in period 1; TW consumption by control cats did not differ during the study. Liquid consumed by drinking increased 18%, 57%, and 96% for the NWP group in periods 2, 3, and 4, respectively, with increases of 25% and 44% for the NW group in periods 3 and 4, respectively, compared with period 1 values for the same groups. Increased urine output and decreased USG were significantly associated with period and treatment.

CONCLUSIONS AND CLINICAL RELEVANCE Increasing the volumes of NW or NWP offered to healthy cats led to increased free liquid consumption and was associated with greater urine output and dilution as measured by USG. Studies are warranted to determine whether these treatments provide health benefits for cats in need of greater water consumption.

Abstract

OBJECTIVE To investigate water intake and urine measures in healthy cats provided free-choice access to a nutrient-enriched water with (NWP) or without (NW) added poultry flavoring offered at 3 different volumes in addition to tap water (TW).

ANIMALS 36 domestic shorthair cats.

PROCEDURES Control group cats (n = 4) received dry food with TW ad libitum throughout the study. Cats of the NW and NWP groups (n = 16/group) received the same food with TW only (period 1; 7 days) followed by TW and the assigned treatment ad libitum at 1X, 1.5X, and 2X the volume of TW consumed in period 1 during periods 2 (17 days), 3 (10 days), and 4 (10 days), respectively. Liquid consumption, food intake, and total water intake (from all sources) were measured; urine collected over 48 hours in each period was measured, and urine specific gravity (USG) was determined. Data were analyzed with mixed-effects models.

RESULTS TW and food calorie intake were similar among groups in period 1; TW consumption by control cats did not differ during the study. Liquid consumed by drinking increased 18%, 57%, and 96% for the NWP group in periods 2, 3, and 4, respectively, with increases of 25% and 44% for the NW group in periods 3 and 4, respectively, compared with period 1 values for the same groups. Increased urine output and decreased USG were significantly associated with period and treatment.

CONCLUSIONS AND CLINICAL RELEVANCE Increasing the volumes of NW or NWP offered to healthy cats led to increased free liquid consumption and was associated with greater urine output and dilution as measured by USG. Studies are warranted to determine whether these treatments provide health benefits for cats in need of greater water consumption.

Estimates of daily water requirements have been reported for cats.1,2 However, daily water needs for healthy cats are only marginally defined, and little is known regarding how incremental changes in water intake may affect urine measures associated with hydration. Daily TW drinking volume is relatively stable in healthy cats ingesting a dry food diet when portion size remains unchanged, resulting in the natural production of highly concentrated urine. Even in this situation, cats may be minimally and chronically in a dehydrated state, whether this is because cats are naturally evolved from arid climates or because they appear to have a generally lower thirst drive. Results of recent studies3–5 suggest cats that are chronically dehydrated or that consume an all-dry-food diet might be predisposed to chronic illnesses such as chronic kidney disease, obesity, LUTD, and diabetes. Therefore, cats eating exclusively dry food might benefit from greater water intake, and it is reasonable to consider that some potential health risks may be decreased if additional water intake and greater hydration could be achieved. However, means of supplementing water intake by encouraging increased drinking have not been fully explored as a strategy to address feline hydration.

Nutrition studies of cats have been conducted to investigate health concerns related to LUTDs with a focus on evaluating increased water ingestion that could be beneficial to such patients. These investigations have provided some evidence that increased water intake can be achieved by modification of dietary moisture content to increase water intake through food6,7,a or provision of food with higher sodium content to stimulate drinking.8,9,b When changing from dry to wet food, the daily water intake measured as water-to-calorie intake ratio (mL/kcal of ME) typically increases from approximately 0.6 to 0.7 with dry food intake1,10,11 to 0.9 with wet food intake.12

More recently, our group performed a study11 in which healthy cats fed dry food with free access to TW were switched to a regimen of free access to an NW, followed by free choice of TW or NW, without changing the diet. Results of that study11 revealed that cats had greater mean free liquid intake (through drinking) and mean total water intake from all sources when only the NW was provided with food, compared with their own baseline values (when only TW was given with food). Consequently, those cats had a mean water-to-calorie intake ratio > 1.0 when drinking the NW, a value that was significantly greater than the baseline ratio for the same cats. Cats with access to NW throughout that study11 also had greater free liquid intake, more dilute urine, and greater urine output than did a cohort of cats that received the same dry diet with TW and no NW option over the 8-week treatment period. These findings revealed that an alternative strategy can encourage cats to increase water intake by greater amounts than are typically observed with wet food ingestion or without added sodium intake delivered through therapeutic dry food.

A greater understanding of feline water intake patterns, water balance, and how these factors influence urine indices of hydration status is still needed. The purpose of the study reported here was to investigate water intake and urine measures in healthy cats provided free-choice access to a water supplement (NWP or NW) at 3 incrementally increased volumes in combination with TW ad libitum. Specifically, we sought to determine whether cats consumed more NWP or NW when larger volumes were offered and whether changes in consumption of the supplements would be associated with changes in variables such as total water intake, urine output volume, and USG. Finally, the study further sought to determine the influence of several ingredients in the supplement by evaluating results when poultry flavor, which putatively acts as a palatability enhancer, was included or excluded. This allowed for evaluation of the effects of NW that contained only whey protein concentrate and glycerin as well as gums and potassium for processing.

Materials and Methods

Animals

Adult domestic shorthair cats (n = 36) owned by Nestlé Purina PetCare were included in the study. The mean ± SD age and BW of cats at the start of the study were 7.1 ± 1.7 years and 4.7 ± 0.7 kg, respectively. The median body condition score was 6 (range, 5 to 7) on the basis of a 9-point scale.13 All included cats were required to have good overall general health for pretrial selection and underwent examination, including routine serum biochemical analysis, by a veterinarian at the beginning of the study.

Throughout the study, cats were housed indoors with temperature ranging from 21° to 24°C and 50% humidity, artificial lighting, and exposure to natural light cycles in environmentally controlled individual rooms (1.5 × 1.5 × 2.4 m) with partial glass walls for viewing access to cats in adjacent housing. Each room had 3 shelves on each wall running three-quarters of the length, and cats had access to group housing on a daily basis. There was 1 litter pan/cat and several environmental enrichment items in each room, including hiding places, multiple perches, and toys. In addition, cats had daily direct interaction with caretakers and interaction with other cats for approximately 2 to 2.5 hours daily when in group housing. The trial was conducted in accordance with approved animal care and use committee protocols at the pet-care facilities of Nestlé Purina PetCare.

Experimental design

The study was designed to monitor daily liquid and food intake of individual cats over a 44-day period (a 7-day baseline phase [period 1] followed by a 37-day treatment phase [periods 2 through 4]; Figure 1). Free liquid intake was measured as the free liquid volume consumed by drinking; this differed from total daily water intake, which was calculated from all water intake sources. Calculation methods are described in detail in a subsequent section. Each cat's food and TW consumption in grams was recorded manually once daily by weighing the bowls with the use of a scale,c which was calibrated monthly. Other liquid (ie, study treatment) consumption was measured twice daily by the same method described subsequently in the present report. All cats had access to TW ad libitum throughout the study and were fed amounts of dry kibble calculated to maintain BW on the basis of caloric requirements for adult feline maintenance (57 kcal/kg of BW). Proximate analysisd of the chicken and rice–based dry kibblee indicated that it contained 5,090 kcal/kg with 6.0% moisture and 43.2% crude protein, 18.1% crude fat, 0.4% crude fiber, 5,466 ppm of sodium, 6,641 ppm of potassium, and 8.5% ash on an as-fed basis.

Figure 1—
Figure 1—

Timeline depicting feeding and watering protocols and sample collection times in a study to investigate water intake and urine measures in healthy cats provided free-choice access to an NW (n = 16) or NWP (16) at 3 volumes in addition to TW and in a control group of healthy cats (4) that received only TW for drinking throughout the study. The NW or NWP was offered at volumes 1 (1X), 1.5 (1.5X), or 2 (2X) times the amount of their mean daily period 1 TW consumption during periods 2, 3, and 4, respectively (with half the assigned volume provided in the morning and half in the evening [with access for 2 hours at each time]). All cats were fed the same dry kibble diet.

Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1150

Prior to starting the study, 32 of the 36 cats were randomly assigned to receive NW or NWP (n = 16/group) throughout the 37-day treatment phase. The NW and NWP contents are summarized (Appendix); the water component of both products was from the same source used to supply the TW. The remaining 4 cats were assigned to a control group that continued to receive only TW and dry kibble throughout the study on the basis of previous results11 that indicated variation in TW intake over time is minimal for cats fed the same dry kibble diet. The small control group size was also selected to increase the number of observations for comparisons between the NW and NWP treatment groups and to maximize the number of cats available for examination of potential within-group differences related to changes in the amount of NW or NWP made available.

During period 1 (days -7 through -1), the amount of TW consumed by all cats was measured and recorded daily as described. These baseline data were used to determine the amount of NW or NWP initially offered to cats during the treatment phase. The treatment phase was subdivided into 3 periods to evaluate the potential effects of incremental increases in the volume of NW or NWP offered to the cats (Figure 1). In period 2 (days 1 through 17; no day was designated as day 0) of the treatment phase, each cat in the NW and NWP groups was offered the assigned treatment at a volume equal to its mean daily TW consumption during period 1 (ie, 1X volume). During periods 3 (days 18 through 27) and 4 (days 28 through 37), these cats were offered the assigned treatments at volumes 1.5 times and 2 times their mean daily TW consumption during period 1 (ie, 1.5X and 2X volumes), respectively. The NW or NWP was provided in 1 bowl separate from the TW during the treatment phase so that TW consumption and consumption of the liquid supplement could each be individually measured throughout the study. Half of the daily volume of NW or NWP was provided in the morning with access for 2 hours between 8 am and 10 am, and the remaining half was provided for 2 hours between 3 pm and 5 pm. The volume of NW or NWP consumed was measured (as previously described) and recorded when the bowls containing the supplement were removed. Cats in both the NW and NWP groups continued to have ad libitum access to TW throughout the day during the treatment period. Cats of the control group had ad libitum access to TW only for drinking throughout all 4 periods.

Calculation of total daily water intake

Total water intake, defined as the sum of free water, metabolic water, and food moisture intake, was calculated for each cat daily. Free water (measured in grams) included TW and the water-only component of the NW or NWP (measured weight minus the weight of dry matter content). Metabolic water was calculated with an estimate of 10 mL of water/100 kcal ME as previously described.14 On the basis of proximate analysis data, nutrient substrate oxidation of the protein component ingested through NW or NWP consumption (41 g of water/100 g of protein oxidized1) was also accounted for in metabolic water calculations. Each cat's total daily water intake was calculated as the total daily water-to-calorie intake ratio11 and also as the amount adjusted for BW.

Sample collection and analysis

The USG and void volume were evaluated to generally assess hydration, as these variables are related to the overall dilution and flow of urine. Voided urine was obtained during 48-hour collection periods throughout the study (days -5 to -3, 12 to 14, 23 to 25, and 33 to 35; Figure 1). Pooled urine was collected by use of a litter box with inert litterf located within the cat's individual housing location. The system used facilitated free flow of the urine into a collection vessel and separation of urine from fecal material. Urine was collected repeatedly into a clean collection vessel containing no preservative and was pooled (for each cat) in a container stored at 4°C. When the 48-hour collection was complete, the total volume was measured, and the pooled urine sample was stored at −80°C until analyzed for USG.g Urine output adjusted for BW of the cats was calculated on a daily basis for each 48-hour measurement period.

Statistical analysis

For BW, liquid and food intake data, and urine measures, a linear mixed-effects model was used to account for nonindependence of the data.15,h Cat identity was a random effect, with the intercept allowed to vary by cat. Treatment (control, NW, or NWP), period (1, 2, 3, or 4), and the interaction between treatment and period were entered as fixed effects. Satterthwaite approximation for degrees of freedom was used to calculate P values. Tukey post hoc tests were then conducted. In addition to examining the linear relationships among urine volume, USG and liquid intake, quadratic polynomial models were also examined. Two models were run—1 without and 1 with the term. The fit of each model was then examined by use of likelihood ratio tests to assess whether adding the quadratic term resulted in improved model fit. Values of P ≤ 0.05 were considered significant.

Results

Calorie intake from food and free liquid intake

Measured amounts of free liquid intake by drinking were summarized as mean ± SE for each group on a daily and study period basis (Figures 2 and 3). The amounts of food consumed were not correctly measured on the first 2 days of period 3 (days 18 [all cats] and 19 [half of the cats]); therefore, caloric and liquid intake data from those 2 days were excluded from the study. With these dates excluded, intake of calories from food was stable throughout the study for all 3 groups (NW, NWP, and control [TW only]), with no differences among periods (P = 0.58) or treatments (P = 0.39; Table 1). Although there was a < 3% change in mean BW at the end of the trial for all cats, compared with the period 1 (baseline data collection period) value, there was a significant (P < 0.01) period × treatment interaction in which cats of the control and NW groups had a slight decrease in weight by period 3. For cats in the NWP group, BW did not differ among periods and overall varied by < 1% over the study.

Figure 2—
Figure 2—

Mean ± SE daily amounts of free liquid consumed (by drinking) for the 36 cats in Figure 1 during periods 1 (days -7 to -1), 2 (days 1 to 17), 3 (days 18 to 27), and 4 (days 28 to 37). Black circles, white circles, and diamonds depict mean results for the NWP, NW, and control groups, respectively.

Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1150

Figure 3—
Figure 3—

Mean ± SE daily amounts of free liquid consumed by the cats in Figure 1 during each of the 4 study periods. a–cWithin a time point, values with different lowercase letters differ significantly among treatment groups. A–DWithin a treatment group, values with different uppercase letters differ significantly among time points. White, black, and gray bars represent data for the NWP, NW, and control groups, respectively.

Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1150

Table 1—

Mean ± SE BW, calorie intake by food consumption, and urine values for 36 healthy cats in a study to assess the effects of an NW or NWP on water intake, USG, and urine output.

 PeriodP value*
Variable and group1234PeriodTreatmentTreatment × period
BW (kg)
 Control4.69 ± 0.13a,A4.64 ± 0.13a,A,B4.58 ± 0.11a,A,B4.56 ± 0.12a,B< 0.0010.98< 0.01
 NWP4.75 ± 0.18a,A4.74 ±0.18a,A4.74 ± 0.18a,A4.79 ± 0.18a,A   
 NW4.73 ± 0.17a,A4.69 ± 0.17a,A,B4.65 ± 0.17a,B4.67 ± 0.17a,B   
Calorie intake from food (kcal ME/d)
 Control176 ± 5177 ± 4182 ± 6178 ± 40.580.390.79
 NWP175 ± 6173 ± 6186 ± 6174 ± 6   
 NW165 ± 5165 ± 5165 ± 5165 ± 5   
USG (g/mL)
 Control1.051 ± 0.004a,A1.049 ± 0.002a,A1.044 ± 0.003a,A1.044 ± 0.006a,A< 0.001< 0.001< 0.001
 NWP1.055 ± 0.001a,A1.033 ± 0.002a,B1.022 ± 0.001b,C1.02I ± 0.002b,C   
 NW1.054 ± 0.002a,A1.043 ± 0.002a,B1.037 ± 0.003a,B1.038 ± 0.004a,B   
Voided urine volume (mL/kg/d)
 Control11.1 ± 0.8a,A9.0 ± 1.2a,A12.0 ± 1.4a,A11.5 ± 3.7a,A< 0.0010.02< 0.001
 NWP7.4 ± 1.0a,A13.5 ± 1.8a,A23.5 ± 2.1a,B27.5 ± 3.2b,B   
 NW7.7 ± 0.7a,A9.0 ± 0.9a,A,B15.4 ± 1.8a,B14.1 ± 2.8a,B   

Cats of the control group (n = 4) were offered a dry kibble diet throughout the study with TW ad libitum. Cats of the NW (n = 16) and NWP (16) groups were offered the same dry diet with TW only (period 1), followed by TW and the assigned treatment (offered in a separate bowl) at IX, 1.5X, and 2X volumes (periods 2, 3, and 4, respectively) ad libitum, where the IX volume for each cat was equal to its mean daily TW consumption during period 1.

P values were generated from a linear mixed model.

Food calorie intake data for the first 2 days of period 3 were excluded owing to a measurement error.

Not all cats had urine samples available for every sample collection period; data represent results for 34, 33, 30, and 33 urine samples in periods 1, 2, 3, and 4, respectively.

Values that differed significantly between treatment groups within a period have different superscript lowercase letters.

Values that differed significantly over time within a treatment group have different superscript capital letters. Pairwise comparisons based on Tukey post hoc tests were considered significant at P < 0.05.

Mean ± SD TW consumption for all 36 cats during period 1 was 118 ± 26 mL/d, with some cats drinking as little as 79 mL/d or as much as 200 mL/d. A significant (P < 0.001) period-by-treatment interaction was detected for the mean amount of free liquid intake by drinking. Mean TW consumption did not differ between groups during period 1 (P = 1.00 for all pairwise comparisons), and free liquid consumption did not differ between groups in period 2 (when NWP or NW treatments were offered at 1X volume; P > 0.33; Figure 3). Within the NWP group, mean daily free liquid consumption during period 2 was significantly (P = 0.04) increased by 21 mL (18.4%) P = 0.04, compared with results during period 1. In contrast, NW group cats had a nonsignificant (P = 0.21) increase of 17 mL (15.4%) and control group cats had a nonsignificant (P = 0.59) decrease of 20 mL (16.7%) for mean daily free liquid intake in period 2, compared with that in period 1. In period 3 (when NWP or NW treatments were offered at 1.5X volume), cats in the NWP and NW groups had significantly (P < 0.01 for both comparisons) increased mean daily free liquid consumption (changes of 57.4% and 25.3%, respectively), compared with period 1 values. Mean daily free liquid intake for the control group during period 3 was nonsignificantly (P = 0.69) decreased by 12.2% from that in period 1 and was not different from that for period 2 (P = 0.99) or period 4 (P = 0.99) for the same group. The NWP group had significantly greater (P < 0.01 for both comparisons) mean daily free liquid intake than the control and NW groups, for which results did not differ significantly (P = 0.38), in period 3. During period 4 (with NWP or NW treatments offered at 2X volume), mean daily free liquid intake increased significantly (P < 0.01 for both comparisons) by 95.6% and 43.8% for the NWP and NW groups, respectively, compared with measurements for the same groups in period 1, whereas results for the control group did not differ significantly (P = 0.64) in period 4 relative to period 1. In this last period of the treatment phase, mean daily free liquid intake differed significantly (P < 0.05 for all comparisons) among the 3 treatment groups.

A mean ± SD of 25 ± 6, 36 ± 10, and 46 ± 15 mL of NWP/kg/d and 20 ± 7, 25 ± 11, and 30 ± 12 mL of NW/kg/d was consumed in periods 2, 3, and 4, respectively, by cats assigned those treatments. Overall, cats of the NWP group drank a mean ± SD of 115 ± 24, 167 ± 43, and 213 ± 70 mL of NWP/d/period, respectively, whereas cats of the NW group drank 94 ± 27, 118 ± 42, and 139 ± 50 mL of NW/d/period.

For cats assigned to receive the NWP treatment, nutrient intake from NWP consumption comprised a mean ± SD of 2.0 ± 0.4, 2.9 ± 0.7, and 3.7 ± 1.2 g of crude protein/d and 15.4 ± 3.3, 22.4 ± 5.7, and 28.5 ± 9.3 mg of sodium/d in periods 2, 3, and 4, respectively. Similarly, but to a lesser degree, cats of the NW group ingested an additional 1.1 ± 0.3, 1.4 ± 0.5, and 1.7 ± 0.6 g of crude protein/d and 4.5 ± 1.3, 5.6 ± 2.0, and 6.7 ± 2.4 mg of sodium/d by consumption of the assigned supplement during periods 2, 3, and 4, respectively.

Relative to the amount of nutrients consumed from dry food, mean daily protein intake for the NWP and NW groups was increased by 18% and 10%, respectively, during period 3 and 25% and 12%, respectively, during period 4 through ingestion of the liquid supplement. Sodium ingested from the dry food during period 1 was a mean ± SD of 230 ± 12, 228 ± 30, and 215 ± 27 mg of sodium/d for cats in the control, NWP, and NW groups, respectively. Because calorie intake from food did not significantly change over time, mean ± SD sodium intake from food was generally similar among groups during period 3 (250 ± 12, 251 ± 36, and 229 ± 28 mg/d for the control, NWP, and NW groups, respectively). The significant increases in amounts of assigned liquid supplement treatments ingested during period 3 equated to (subjectively assessed) mean increases in daily sodium intake of 8.2% for the NWP group and 2.4% for the NW group relative to total sodium intake from food plus liquid supplement during period 3. Interestingly, cats of the control group and the NWP group ingested 8.7% and 10.0% more sodium/d, respectively, in period 3 than in period 1, owing to changes in dry food consumption alone. However, control group cats had a 12.2% decrease in free liquid (TW) consumption that coincided with the 8.7% increase in sodium/d, whereas NWP group cats had a 57.4% increase in free liquid intake during the same interval, which accompanied an 18% increase in total sodium intake (approx 50% owing to NWP and 50% to changes in dry food consumption). For comparison purposes, total sodium ingestion (from NW and changes in dry food consumption) increased 9.1% for the NW group cats during period 3, compared with period 1; this percentage change was similar to that for the control group, but total sodium intake (milligrams) was slightly less. However, cats receiving the NW had a 25.3% increase in free liquid intake, compared with that in period 1.

Total daily TW consumption was also reported for each of the treatment groups (Figure 4). When optional NW or NWP was introduced at the start of period 2, daily TW intake declined substantially for those groups. Following this, TW intake remained relatively unchanged within groups throughout the treatment phase (periods 2, 3, and 4).

Figure 4—
Figure 4—

Mean ± SE daily volumes of TW ingested for the 36 cats in Figure 1. See Figure 2 for key.

Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1150

Total water intake

Significant (P < 0.001) time × treatment interactions were identified for the mean daily water-to-calorie intake ratio and for mean daily total water intake adjusted for BW (both calculated with total water ingestion from all sources as described). The water-to-calorie intake ratio was similar (P = 1.00 for all pairwise comparisons) among groups during period 1 and did not differ significantly (P > 0.78 for all comparisons) for the control group between study periods (Figure 5). In contrast, this ratio significantly (P < 0.001) increased for the NWP group by 17%, 40%, and 82% in periods 2, 3, and 4, respectively, compared with period 1; the ratio was also significantly (P < 0.001) greater for each of these periods than for the preceding period. Similar changes were observed for the NW group at most time points, with significant (P < 0.01) increases of 12%, 15%, and 35% relative to period 1. The ratio did not differ significantly (P = 1.0) between periods 2 and 3, but the increase from period 3 to period 4 was significant (P < 0.01).

Figure 5—
Figure 5—

Mean ± SE measures of daily total water intake (sum of liquid consumed by drinking, metabolic water, food moisture content, and water content of the NW or NWP as applicable) for the 36 cats in Figure 1 during each of the 4 study periods. A—Water-to-calorie intake ratios. B—Total water intake adjusted for BW. See Figure 3 for key.

Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1150

Total water intake adjusted for BW was similar (P = 1.00 for all pairwise comparisons) among groups during period 1 and remained consistent between periods for the control group (with nonsignificant [P > 0.97 for all comparisons] decreases of 13% [period 2], 13% [period 3], and 8% [period 4], relative to period 1; Figure 5). Total water intake for the NWP group had a nonsignificant (P = 0.50) increase of 16% in period 2, compared with period 1. This was followed by significant (P < 0.001) increases of 49% and 80% in periods 3 and 4, compared with period 1. In the NW group, a nonsignificant (P = 0.22) total water intake increase of 13% was observed during period 2, compared with period 1, whereas this variable significantly (P < 0.001) increased from the period 1 value by 24% and 39% during periods 3 and 4, respectively.

USG and voided urine volume

One cat in the NW group urinated outside the litter box when the inert litter was used and had no urine available for analysis. Six other cats (3 each from the NWP and NW groups) did not urinate in the litter box used for collection at 1 time point each during the study, and 1 cat from the NWP group did not urinate in the litter box at 3 time points. Therefore, urine data from 130 samples collected from 35 cats were analyzed (Table 1). A significant (P < 0.001) period × treatment interaction was detected for USG. Mean USG did not differ (P = 1.00) among groups during period 1 and did not differ (P > 0.98) among periods for the control group. The USG measurements varied by < 0.007 g/mL for the control group throughout the study. Urine samples for cats in the NWP group became more dilute over time, with mean USG decreasing by 39% in period 2 (P < 0.001), 60% (P < 0.001) in period 3, and 62% (P < 0.001) in period 4, compared with the period 1 value. Mean USG also decreased (P < 0.001 for all comparisons) from the period 1 value for cats in the NW group, but to a lesser extent with changes of 21%, 31%, and 29% during periods 2, 3, and 4, respectively.

Changes in voided urine volume (adjusted for BW) complemented changes in USG overall (Table 1), with a significant (P < 0.001) period × treatment interaction detected for the output variable. Mean daily urine volume did not differ among treatment groups (P = 1.00) in period 1; for cats in the control group, these measurements also did not differ among periods (P = 1.00), with mean measurements ranging from 9.0 to 12.0 mL/kg/d. Cats of the NWP group had a nonsignificant (P = 0.07) increase in mean daily urine volume for period 2, compared with period 1. This was followed by significant (P < 0.01) increases of 216% and 270% in periods 3 and 4, respectively, compared with period 1. Mean daily urine volume was also significantly (P < 0.01) increased for cats of the NW group in periods 3 and 4, with differences of 100% and 91%, respectively, relative to period 1.

Relationships between urine measures and total daily water intake

For USG and daily voided urine volume adjusted for BW, an inverse quadratic polynomial model (x2 = 38.53; y = 14,412x2 − 30,454x + 16,095; P < 0.001) fit the data better than a linear model (Figure 6). Total daily water intake (from all sources) adjusted for BW was then plotted against USG for all periods to assess the relationship between these variables. An inverse quadratic polynomial model (x2 = 22.17; y = 1.84E − 0.5x2 − 0.0027x + 1.11 [where E represents the exponential function]; P < 0.001) also fit these data better than a linear model and confirmed that increased total daily water intake was associated with decreased USG. A very similar relationship was found between USG and the water-to-calorie intake ratio (x2 = 14.57; y = 0.023x2 − 0.094x + 1.11; P < 0.001 [graphic not shown]). Finally, a significant (P < 0.001) positive linear relationship (β = −14.839; y = 0.819x − 14.839) was observed between total daily water intake and voided urine volume adjusted for BW.

Figure 6—
Figure 6—

Results of mixed-effects regression analysis showing the relationships between USG and voided urine volume (A) and between total water intake and urine variables (B and C) in healthy cats. A—Plot of voided urine volume versus USG. B—Plot of USG versus total water intake. Blue lines indicate the estimated total daily water intake associated with a USG of 1.035 g/mL (37 mL/kg/d). C—Plot of voided urine volume versus total water intake. Blue lines indicate the estimated daily urine output volume associated with a total daily water intake of 37 mL/kg (15.5 mL/kg/d). In each panel, each data point represents paired measurements for an individual cat at sample collection times during each period (up to 4 data points/cat; not all cats had urine samples available for every sample collection period).

Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1150

Discussion

In the present study of healthy cats, increased amounts of NWP or NW were offered free choice daily at 1X (period 2), 1.5X (period 3), and 2X (period 4) volumes relative to mean amounts of TW consumed daily by the same cats during baseline data collection (period 1). The results of the present study showed that the addition of whey protein concentrate and glycerin (with potassium added for processing) to TW can positively influence total water intake in cats, resulting in increased urine output. The results also revealed that incremental increases in mean NWP or NW consumption were significantly associated with increases in mean free liquid consumption by drinking, total water intake, and voided urine volume. Results for the NWP group in the present study confirmed the findings in our previous investigation11 that total water intake can be significantly increased in healthy cats that are offered free access to an NWP supplement in addition to TW and dry food. More specifically, significantly greater total water intake in periods 3 and 4 (attributable to NWP consumption) resulted in significantly lower USG and greater voided urine volume (significant in period 4), compared with results for cats receiving only TW and dry food. Moreover, in the present study, the greatest liquid drinking and urine output occurred for cats with access to NWP, which contained additional crude protein, fat, and minerals such as potassium and sodium, as well as the poultry flavor ingredient. The NWP formula in the present study was slightly different from that of our previous study11; a different poultry flavor ingredient was used that contained lower phosphorus and sodium than in the previous version of NWP. This change in added flavor ingredient resulted in a > 50% reduction in sodium content and a phosphorus concentration that was below the lower limit of detection. Furthermore, when the poultry flavor palatant was excluded, the NW was still associated with significantly increased total water intake that resulted in increased urine output and decreased USG (periods 3 and 4), compared with findings for same cats during baseline data collection (period 1).

Examination of TW consumption in period 1 indicated that cats of the control group (which received TW throughout the study), NW group, and NWP group drank similar amounts at the outset of the study. The mean daily water-to-calorie intake ratio for all groups during period 1, as well as over the entire study for the control group, ranged from approximately 0.7 to 0.8 mL/kcal of ME. When reported on a BW basis, mean total water intake for cats drinking TW ranged from approximately 26 to 30 mL/kg. These findings were similar to those in previous research of cats fed dry food10,11 and slightly higher than in other studies6,16 that identified mean values of 21 to 23 mL/kg. Ultimately, the total water intake for cats that had only TW available for drinking resulted in mean voided urine volumes of approximately 7 to 12 mL/kg and mean USG measurements of 1.044 to 1.055 g/mL, similar to results in other studies6,11,12,a of healthy cats fed dry food.

The use of NW or NWP as supplements to drinking water revealed 2 findings that advance the field of feline nutrition and health. First, because most cats offered NW or NWP demonstrated a strong desire to drink the liquid, we were able to observe the effects of voluntary drinking volumes of free liquid that were much greater than those typically consumed by cats drinking TW only. Second, the incremental increases in total water intake for these cats generated a wide spectrum of total water intake, urine output, and USG values. This facilitated a more complete understanding of liquid intake and urine output dynamics in cats that would not easily be achieved through feeding of combinations of dry and wet food paired with TW only. As a result, a robust relationship between feline water intake and urine concentration (as assessed by urine osmolality)2,11 or voided urine volume11 that has been suggested, but not previously characterized, was revealed.

Because water drinking by healthy cats varies from day to day and water consumption can differ substantially among individual cats, the present study was designed to leverage each cat's typical TW drinking habits to determine the amounts of NW or NWP that would be supplied with each incremental increase. This was somewhat similar to determining a specific animal's metabolic energy requirement and adjusting the calorie ration to achieve a targeted BW or body condition.

When the assigned treatments were provided at the 1X volume (period 2), 14 of 16 cats given NWP drank > 95% of the amount offered and the remaining 2 cats drank 63% of the available volume, whereas 9 of 16 cats receiving NW drank > 95% of the solution and the remaining 7 drank 45% to 87% of the amount offered. With the increase to 2X volume, fewer cats drank nearly all the NWP (10/16) or NW (4/16) supplied. Even though many cats did not drink all the liquid supplement offered, a cat consuming only 50% of the 2X volume would still be drinking the equivalent of its typical TW consumption, and in most cases, cats continued to drink some amount of TW in addition to NWP or NW.

We measured USG and calculated daily voided urine volume from 48-hour urine samples collected throughout the study (typically 4/cat) to assess these values for progressive changes in response to changes in liquid consumption. These measures are routinely used as hallmarks of hydration status in people.17 However, the specific USG values that might be associated with various categories of relative hydration status or whether a combination of variables may be more representative of hydration status in this species has yet to be defined in cats.2 As mentioned, all control cats in the present study maintained USG and voided urine volumes within fairly narrow ranges typical of healthy cats fed dry food. For groups that received NW or NWP, an initial decline in USG was observed when the 1X volume was offered (period 2). The NW group cats had a mean USG decrease from 1.054 to 1.043 g/mL, which corresponded to an apparent, albeit nonsignificant, increase of 17 mL/d in mean free liquid drinking. The NWP group cats increased liquid drinking by 21 mL/d and had an even greater reduction in USG. For the NWP group, this pattern continued with a further decrease in USG as the amount of free liquid drinking increased in period 3, but USG remained unchanged in period 4 even though the cats had greater mean free liquid consumption when offered the 2X volume. By contrast, although the NW group had greater free liquid consumption in period 4 than in previous periods, this did not translate into further dilution of USG, which did not differ significantly from that in periods 2 and 3. Voided urine volume for this group followed a similar response, such that although free liquid consumption increased in period 4, the mean voided urine volume did not increase significantly relative to periods 2 and 3.

Examining the relationship between total daily water intake (from all sources) and USG revealed that the highly palatable NW and NWP may have important roles in beginning to characterize the amounts of water intake needed to effectively dilute the urine to a greater extent than is achieved with normal TW drinking. The significant curvilinear relationship between total daily water intake adjusted for BW and USG that was detected allowed for calculation of a target total water intake needed to exceed the renal water resorption mechanism and promote a more dilute urine in healthy adult cats. This curvilinear model enhanced our initial observations regarding this type of data11 (with urine osmolality used as a measure of dilution) because only a significant linear relationship was previously observed. Presumably, the difference resulted from the greater number of cats (35) included in the evaluation for the present study, compared with the previous evaluation (18 cats). Results of the analysis in the present study indicated that to attain a targeted USG of 1.035 g/mL, a healthy cat would typically need an estimated total daily water intake of 37 mL/kg. By use of the same curvilinear model, this same USG value would predict a water-to-calorie intake ratio estimate of 1.08 mL:1 kcal of ME. At a total daily water intake of 37 mL/kg, the linear regression model generated a predicted voided urine volume of approximately 15.5 mL/kg. However, on the basis of the individual cat variations observed, the urine output volume could range from 8 to 22 mL/kg. Finally, results of our previous study11 identified a significant positive correlation between USG and urine osmolality in cats, with a USG of 1.035 g/mL equating to approximately 1,500 mOsm/kg. Our previous results11 also indicated that a water-to-calorie intake ratio of approximately 1.2 mL:1 kcal of ME would yield a urine osmolality of approximately 1,500 mOsm/kg, generally consistent with the USG data summarized for the present study. Although a USG of 1.035 g/mL or osmolality of 1,500 mOsm/kg represents an arbitrary choice, it does offer targeted values for diluted urine that is concentrated enough to not mimic loss of renal function resulting from nephron dysfunction.

Many situations exist that can result in hypohydration status. The results of the present study built upon previous data to enhance understanding of how daily water intake, largely in the form of free liquid drinking, in healthy cats fed a dry food impacts USG and urine output. This research also provided a basis to help predict specific total water intake requirements that may be needed for healthy cats to achieve a targeted urine output and concentration. Although beyond the scope of the present study, a need still exists to investigate means of improving hydration or increasing water intake in cats at risk for urolith formation and feline patients with renal insufficiency, LUTD, or hypohydration related to age, injury, or surgery. Although a USG persistently < 1.030 g/mL is routinely observed as a sign of advanced renal dysfunction in cats, it may be beneficial to consider USGs in this range as potential targets for generally healthy cats in need of increased liquid intake and corresponding increased urine output. Further investigations are needed to determine whether and to what degree increasing total water intake could benefit feline patients with these types of conditions. The study results suggested that the NW or NWP investigated could provide an alternative and feasible method to explore the effects of increasing free liquid consumption by offering a liquid supplement, compared with dietary modification by high-moisture food provision or sodium enrichment, in cats needing greater total water intake.

Acknowledgments

All authors are employed in the R&D department of Nestlé Purina PetCare and conduct nutrition research for potential use in future commercial applications and products. None of the authors have any conflict of interest or affiliation to disclose related to the equipment used during the study.

The authors thank Mark Miller and Sarah Dionne for management and care of cats and coordination of sample collection and Patricia Turpin and Dakota Marti for performing assays on urine samples.

ABBREVIATIONS

BW

Body weight

LUTD

Lower urinary tract disease

ME

Metabolizable energy

NW

Nutrient-enriched water

NWP

Nutrient-enriched water with poultry flavoring

TW

Tap water

USG

Urine specific gravity

Footnotes

a.

Xu H, Greco DS, Zanghi B, et al. The effect of feeding inversely proportional amounts of dry versus canned food on water consumption, hydration, body composition, and urinary parameters in cats (abstr), in Proceedings. 39th World Small Anim Vet Assoc Cong 2014;852.

b.

Xu H, Laflamme D, Bartges J, et al. Effect of dietary sodium on urine characteristics in healthy adult cats (abstr). J Vet Intern Med 2006;20:738.

c.

BBA422-6PM, Mettler Toledo, Columbus, Ohio.

d.

Proximate analysis performed by NP Analytical Laboratories, St Louis, Mo.

e.

Maintenance diet manufactured by Nestlé Purina PetCare, St Louis, Mo.

f.

NOSORB, Catco Inc, Cape Coral, Fla.

g.

HSK-VET veterinary refractometer, Heska, Loveland, Colo.

h.

Bates D, Maechler M, Bolker B, et al. lme4: Linear Mixed-Effects Models using ‘Eigen’ and S4. R package, version 1.1–14. Available at: CRAN.R-project.org/package=lme4. Accessed Sep 26, 2017.

References

  • 1. National Research Council. Water. In: Nutrient requirements of dogs and cats. Washington, DC; National Academies Press, 2006;246250.

    • Search Google Scholar
    • Export Citation
  • 2. Zanghi BM. Water needs and hydration for cats and dogs, in Proceedings. Nestlé Purina Comp Anim Nutr Summit 2017;1523.

  • 3. Greene JP, Lefebvre SL, Wang M, et al. Risk factors associated with the development of chronic kidney disease in cats evaluated at primary care veterinary hospitals. J Am Vet Med Assoc 2014;244:320327.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 4. Rowe E, Browne W, Casey R, et al. Risk factors identified for owner-reported feline obesity at around one year of age: dry diet and indoor lifestyle. Prev Vet Med 2015;121:273281.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 5. Sallander M, Eliasson J, Hedhammar A. Prevalence and risk factors for the development of diabetes mellitus in Swedish cats. Acta Vet Scand 2012;54:61.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 6. Buckley CM, Hawthorne A, Colyer A, et al. Effect of dietary water intake on urinary output, specific gravity and relative supersaturation for calcium oxalate and struvite in the cat. Br J Nutr 2011;106(suppl 1):S128–S130.

    • Search Google Scholar
    • Export Citation
  • 7. Markwell PJ, Buffington CA, Chew DJ, et al. Clinical evaluation of commercially available urinary acidification diets in the management of idiopathic cystitis in cats. J Am Vet Med Assoc 1999;214:361365.

    • Search Google Scholar
    • Export Citation
  • 8. Hawthorne AJ, Markwell PJ. Dietary sodium promotes increased water intake and urine volume in cats. J Nutr 2004;134(suppl 8):2128S–2129S.

    • Search Google Scholar
    • Export Citation
  • 9. Xu H, Laflamme DP, Long GL. Effects of dietary sodium chloride on health parameters in mature cats (Erratum published in J Feline Med Surg 2009;11:735). J Feline Med Surg 2009;11:435–441.

    • Search Google Scholar
    • Export Citation
  • 10. Seefeldt SL, Chapman TE. Body water content and turnover in cats fed dry and canned rations. Am J Vet Res 1979;40:183185.

  • 11. Zanghi BM, Gerheart LG, Gardner CL. Effects of a nutrient-enriched water on water intake and indices of hydration in healthy domestic cats fed a dry kibble diet. Am J Vet Res 2018;79:733744.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 12. Finco DR, Adams DD, Crowell WA, et al. Food and water intake and urine composition in cats: influence of continuous versus periodic feeding. Am J Vet Res 1986;47:16381642.

    • Search Google Scholar
    • Export Citation
  • 13. Laflamme DP. Development and validation of a body condition score system for cats: a clinical tool. Feline Pract 1997;25(5):1318.

  • 14. National Research Council. Water. In: Nutrient requirements of dogs. Washington, DC; National Academies Press, 1985:39.

  • 15. Bates D, Mächler M, Bolker B, et al. Fitting linear mixed-effects models using lme4. J Stat Softw 2015;67:148.

  • 16. Jackson OF, Tovey JD. Water balance studies in domestic cats. Feline Pract 1977;7(4):3033.

  • 17. Armstrong LE. Hydration biomarkers during daily life: recent advances and future potential. Nutr Today 2012;47:S3S6.

Appendix

Results of proximate analysis of NW and NWP used in a study to evaluate the effects of each supplementary product on measures of water intake, urine output, and USG in healthy domestic cats fed a dry kibble diet ad libitum.

ComponentNWPNW
Ingredients
 Whey protein (%)1.21.2
 Glycerin (%)1.01.0
 Potassium chloride (%)0.1000.100
 Hydrocolloids (%)*0.1100.110
 Poultry flavor (%)1.00
Proximate analysis
 Moisture (%)97.998.5
 Crude protein (%)1.721.20
 Crude fat (%)0.31< 0.1
 Crude fiber (%)< 0.2< 0.2
 Phosphorus(%)< 0.01< 0.01
 Potassium (ppm [%])310 (0.031%)245 (0.0245%)
 Magnesium (ppm [%])29.3 (0.0029%)26.0 (0.0026%)
 Sodium (ppm [%])134 (0.0134%)48.2 (0.0048%)

Sixteen cats received NW and 16 received NWP in addition to TW ad libitum during the treatment period; a control group of 4 cats received only TW ad libitum for drinking throughout the study.

Mix of guar gum and xanthan gum.

Values determined on an as-fed basis.

Contributor Notes

Each author contributed equally to the experimental design, data analysis, and manuscript drafting.

Address correspondence to Dr. Zanghi (Brian.Zanghi@rd.nestle.com).
  • Figure 1—

    Timeline depicting feeding and watering protocols and sample collection times in a study to investigate water intake and urine measures in healthy cats provided free-choice access to an NW (n = 16) or NWP (16) at 3 volumes in addition to TW and in a control group of healthy cats (4) that received only TW for drinking throughout the study. The NW or NWP was offered at volumes 1 (1X), 1.5 (1.5X), or 2 (2X) times the amount of their mean daily period 1 TW consumption during periods 2, 3, and 4, respectively (with half the assigned volume provided in the morning and half in the evening [with access for 2 hours at each time]). All cats were fed the same dry kibble diet.

  • Figure 2—

    Mean ± SE daily amounts of free liquid consumed (by drinking) for the 36 cats in Figure 1 during periods 1 (days -7 to -1), 2 (days 1 to 17), 3 (days 18 to 27), and 4 (days 28 to 37). Black circles, white circles, and diamonds depict mean results for the NWP, NW, and control groups, respectively.

  • Figure 3—

    Mean ± SE daily amounts of free liquid consumed by the cats in Figure 1 during each of the 4 study periods. a–cWithin a time point, values with different lowercase letters differ significantly among treatment groups. A–DWithin a treatment group, values with different uppercase letters differ significantly among time points. White, black, and gray bars represent data for the NWP, NW, and control groups, respectively.

  • Figure 4—

    Mean ± SE daily volumes of TW ingested for the 36 cats in Figure 1. See Figure 2 for key.

  • Figure 5—

    Mean ± SE measures of daily total water intake (sum of liquid consumed by drinking, metabolic water, food moisture content, and water content of the NW or NWP as applicable) for the 36 cats in Figure 1 during each of the 4 study periods. A—Water-to-calorie intake ratios. B—Total water intake adjusted for BW. See Figure 3 for key.

  • Figure 6—

    Results of mixed-effects regression analysis showing the relationships between USG and voided urine volume (A) and between total water intake and urine variables (B and C) in healthy cats. A—Plot of voided urine volume versus USG. B—Plot of USG versus total water intake. Blue lines indicate the estimated total daily water intake associated with a USG of 1.035 g/mL (37 mL/kg/d). C—Plot of voided urine volume versus total water intake. Blue lines indicate the estimated daily urine output volume associated with a total daily water intake of 37 mL/kg (15.5 mL/kg/d). In each panel, each data point represents paired measurements for an individual cat at sample collection times during each period (up to 4 data points/cat; not all cats had urine samples available for every sample collection period).

  • 1. National Research Council. Water. In: Nutrient requirements of dogs and cats. Washington, DC; National Academies Press, 2006;246250.

    • Search Google Scholar
    • Export Citation
  • 2. Zanghi BM. Water needs and hydration for cats and dogs, in Proceedings. Nestlé Purina Comp Anim Nutr Summit 2017;1523.

  • 3. Greene JP, Lefebvre SL, Wang M, et al. Risk factors associated with the development of chronic kidney disease in cats evaluated at primary care veterinary hospitals. J Am Vet Med Assoc 2014;244:320327.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 4. Rowe E, Browne W, Casey R, et al. Risk factors identified for owner-reported feline obesity at around one year of age: dry diet and indoor lifestyle. Prev Vet Med 2015;121:273281.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 5. Sallander M, Eliasson J, Hedhammar A. Prevalence and risk factors for the development of diabetes mellitus in Swedish cats. Acta Vet Scand 2012;54:61.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 6. Buckley CM, Hawthorne A, Colyer A, et al. Effect of dietary water intake on urinary output, specific gravity and relative supersaturation for calcium oxalate and struvite in the cat. Br J Nutr 2011;106(suppl 1):S128–S130.

    • Search Google Scholar
    • Export Citation
  • 7. Markwell PJ, Buffington CA, Chew DJ, et al. Clinical evaluation of commercially available urinary acidification diets in the management of idiopathic cystitis in cats. J Am Vet Med Assoc 1999;214:361365.

    • Search Google Scholar
    • Export Citation
  • 8. Hawthorne AJ, Markwell PJ. Dietary sodium promotes increased water intake and urine volume in cats. J Nutr 2004;134(suppl 8):2128S–2129S.

    • Search Google Scholar
    • Export Citation
  • 9. Xu H, Laflamme DP, Long GL. Effects of dietary sodium chloride on health parameters in mature cats (Erratum published in J Feline Med Surg 2009;11:735). J Feline Med Surg 2009;11:435–441.

    • Search Google Scholar
    • Export Citation
  • 10. Seefeldt SL, Chapman TE. Body water content and turnover in cats fed dry and canned rations. Am J Vet Res 1979;40:183185.

  • 11. Zanghi BM, Gerheart LG, Gardner CL. Effects of a nutrient-enriched water on water intake and indices of hydration in healthy domestic cats fed a dry kibble diet. Am J Vet Res 2018;79:733744.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 12. Finco DR, Adams DD, Crowell WA, et al. Food and water intake and urine composition in cats: influence of continuous versus periodic feeding. Am J Vet Res 1986;47:16381642.

    • Search Google Scholar
    • Export Citation
  • 13. Laflamme DP. Development and validation of a body condition score system for cats: a clinical tool. Feline Pract 1997;25(5):1318.

  • 14. National Research Council. Water. In: Nutrient requirements of dogs. Washington, DC; National Academies Press, 1985:39.

  • 15. Bates D, Mächler M, Bolker B, et al. Fitting linear mixed-effects models using lme4. J Stat Softw 2015;67:148.

  • 16. Jackson OF, Tovey JD. Water balance studies in domestic cats. Feline Pract 1977;7(4):3033.

  • 17. Armstrong LE. Hydration biomarkers during daily life: recent advances and future potential. Nutr Today 2012;47:S3S6.

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