The Clinical Urine Culture: Enhanced Techniques Improve Detection of Clinically Relevant Microorganisms

Travis K. Price; Tanaka Dune; Evann E. Hilt; Krystal J. Thomas-White; Stephanie Kliethermes; Cynthia Brincat; Linda Brubaker; Alan J. Wolfe; Elizabeth R. Mueller; Paul C. Schreckenberger

doi:10.1128/JCM.00044-16

DOI: 10.1128/JCM.00044-16

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FIG 1
Average CFU per milliliter of uropathogens between the UTI and no-UTI cohorts. Depicted are the average CFU per milliliter with which the various uropathogens were cultured for both cohorts: UTI (blue bars) and no-UTI (red bars). Average CFU of Klebsiella pneumoniae (P = 0.04) and Streptococcus agalactiae (P = 0.02) are significantly higher in the UTI cohort (*). Several uropathogens had substantially lower average CFU-per-milliliter values in the no-UTI cohort than in the UTI cohort: Aerococcus urinae (P = 0.12), Enterococcus faecalis (P = 0.09), Escherichia coli (P = 0.08), Staphylococcus aureus (P = 0.06), and Streptococcus anginosus (P = 0.06). Independent t test (*, P < 0.05). Black bars depict common UTI thresholds (≥10⁵ CFU/ml and ≥10³ CFU/ml).

TABLE 1

Summary of urine cultivation protocols for catheterized urine specimens

Protocol	Vol (μl) of urine	Medium or media	Conditions	Incubation(s) (h) for microbial identification	Patient sample identifier^a
Standard urine culture	1	BAP, MacConkey agar	Aerobic, 35°C	24	1–107
Modified urine culture	1	BAP, MacConkey agar	5% CO₂, 35°C	24, 48	108–150
Expanded-spectrum EQUC	1, 10, and 100	BAP, MacConkey agar	Aerobic, 35°C	24, 48	1–150
		BAP, chocolate agar, CNA agar	5% CO₂, 35°C	24, 48
		CDC anaerobic BAP	Anaerobic, 35°C	48
		CDC anaerobic BAP^b	Microaerophilic gas mixture (5% O₂, 10% CO₂, 85% N), 35°C	48
Streamlined EQUC	100	BAP, MacConkey agar,^c CNA agar	5% CO₂, 35°C	48	1–150^d

↵a Refers to the patient samples on which the corresponding protocol assessed the urinary microbiota. For diagnosis, the standard urine culture protocol was used on patient samples 1 to 107; the modified standard urine culture was used on patient samples 108 to 150. For research, all patient samples were assessed by expanded-spectrum EQUC.
↵b The CDC anaerobic BAP microaerophilic gas mixture condition was used only for samples 10 to 150.
↵c The MacConkey 5% CO₂ condition was not part of the expanded-spectrum EQUC protocol.
↵d The streamlined EQUC protocol was performed using a subgroup of agars/conditions from the expanded-spectrum EQUC protocol; therefore, it was used on all patient samples.

TABLE 2

Demographic characteristics and symptoms

Clinical variable	Entire cohort (n = 150)	No-UTI cohort (n = 75)	UTI cohort (n = 75)	P value^d
Age (yr), mean (SD)	62.3 (14.9)	60.6 (12.3)	64.0 (17.1)	0.16^a
BMI (kg/m²), mean (SD)	29.3 (6.3)	28.8 (5.9)	29.9 (6.6)	0.27^a
Race/ethnicity, no. (%)
White	121 (81)	59 (79)	62 (83)
Hispanic	15 (10)	9 (12)	6 (8)
Black	9 (6)	5 (7)	4 (5)	0.90^c
Asian	4 (3)	2 (3)	2 (3)
Other	1 (1)	0 (0)	1 (1)
No. of vaginal deliveries, median (IQR)	2 (0–11)	2 (0–6)	2 (0–11)	0.80^b
Sexually active, no. (%)	58 (39)	37 (49)	21 (28)	0.01
Previous antibiotic treatment, no. (%)	45 (30)	20 (27)	25 (33)	0.37
Current anticholinergic treatment, no. (%)	26 (17)	9 (12)	17 (23)	0.08
Type of anticholinergic used, no. (%)
Oxybutynin	9 (6)	3 (4)	6 (8)
Solifenacin	8 (5)	3 (4)	5 (7)
Fesoterodine	4 (3)	2 (3)	2 (3)
Tolterodine	2 (1)	1 (1)	1 (1)	0.75^c
Oxybutynin patch	1 (1)	0 (0)	1 (1)
Trospium chloride	1 (1)	0 (0)	1 (1)
Mirabegron (Myrbetriq)	1 (1)	0 (0)	1 (1)
Previous vaginal estrogen use, no. (%)	32 (21)	11 (15)	21 (28)	0.05
Current vaginal estrogen use, no. (%)	29 (19)	10 (13)	19 (25)	0.06
Prior urogynecologic surgery, no. (%)	40 (27)	13 (17)	27 (36)	0.01
Symptoms of incontinence, no. (%)
Stress urinary incontinence	17 (11)	10 (13)	7 (9)	0.44
Urgency urinary incontinence	26 (17)	11 (15)	15 (20)	0.39
Mixed urinary incontinence	42 (28)	27 (36)	15 (20)	0.03
Urgency-frequency syndrome, no. (%)	18 (12)	9 (12)	9 (12)	0.99
Myofascial pain, no. (%)	55 (37)	20 (27)	35 (47)	0.01
Painful bladder syndrome, pelvic pain, and dyspareunia, no. (%)	4 (3)	0 (0)	4 (5)	0.06^c
UTISA score, mean (SD)^e
Urination regularity	6.6 (4.3)	5.4 (4.3)	7.9 (3.9)	<0.001^a
Problems with urination	3.4 (3.6)	2.0 (2.5)	5.4 (3.7)	<0.001^a
Pain associated with UTI	3.2 (3.6)	2.1 (3.1)	4.3 (3.8)	<0.001^a
Blood in the urine	0.2 (0.8)	0.1 (0.4)	0.4 (1.1)	0.08^a

↵a Independent t test.
↵b Wilcoxon rank sum test.
↵c Fisher's exact test.
↵d Chi-square test used unless otherwise indicated. Boldface indicates P values that are significant at ≤0.05.
↵e UTISA scores for urinary regularity, problems with urination, and pain associated with UTI range from 0 to 12. UTISA scores for blood in the urine range from 0 to 6.

TABLE 3
Optimal detection of specific uropathogens by the expanded-spectrum EQUC protocol^a
↵a Listed are the uropathogens and the number of times that each was cultured under each expanded-spectrum EQUC plating condition. The condition(s) that best detected each uropathogen is shaded.

TABLE 4

Detection of uropathogens in UTI cohort without symptom improvement^a

Postenrollment questionnaire response (sample identifier)	Antibiotic prescribed	Uropathogen(s) detected by protocol(s):
Postenrollment questionnaire response (sample identifier)	Antibiotic prescribed	Standard urine culture and expanded-spectrum EQUC	Expanded-spectrum EQUC only^d
Same (145)	SMZ-TMP^b	Escherichia coli	Streptococcus anginosus
Same (048)	Nitrofurantoin	Escherichia coli	Streptococcus anginosus
Same (134)	SMZ-TMP	Escherichia coli	Aerococcus urinae, Corynebacterium riegelii
Same (033)	Ciprofloxacin	Klebsiella pneumoniae
Same (060)	Nitrofurantoin	Escherichia coli
Same (109)	Nitrofurantoin	Escherichia coli
Same (122)	Ciprofloxacin	Escherichia coli
Same (135)	Nitrofurantoin	Escherichia coli
Same (136)	Nitrofurantoin	Escherichia coli
Worse (082)	Nitrofurantoin	Escherichia coli
Same (140)		Staphylococcus lugdunensis
Same (116)		Escherichia coli
Worse (128)		Escherichia coli
Same (126)		Lactobacillus species^c	Staphylococcus lugdunensis, Streptococcus anginosus
Same (121)			Proteus mirabilis
Same (029)			Aerococcus urinae, Klebsiella pneumoniae
Same (139)			Alloscardovia omnicolens, Oligella urethralis, Morganella morganii
Same (052)			Streptococcus anginosus
Same (067)			Streptococcus anginosus
Worse (025)			Candida albicans
Worse (112)			Streptococcus agalactiae, Streptococcus anginosus
Worse (142)			Escherichia coli
Same (084)
Same (108)

↵a Uropathogens detected and missed by the standard urine culture in urine samples obtained by catheter from the UTI cohort patients who reported feeling the same or worse for the postenrollment questionnaire. Antibiotics were prescribed based on the reporting of the standard urine culture results.
↵b SMZ-TMP, sulfamethoxazole-trimethoprim.
↵c Lactobacillus species is not considered a uropathogen, but it was detected at >100,000 CFU/ml by standard urine culture.
↵d All of the uropathogens detected by the expanded-spectrum EQUC protocol would have been detected using the streamlined EQUC protocol.

Additional Files

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Supplemental material

Supplemental file 1 -
Fig. S1 (Species accumulation curves for UTI and no-UTI cohorts), S2 (Frequency of detection of genera in UTI and no-UTI cohorts), S3 (Species significantly different between UTI and no-UTI cohorts), and S4 (Detection of uropathogens in urine volumes by expanded-spectrum EQUC) and Tables S1 (Urinary microbiota characteristics), S2 (Detection of Gram-positive and Gram-negative uropathogens), and S3 (Detection of microorganisms of unknown pathogenicity in UTI cohort patients without symptom improvement)

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