Vertical and Horizontal Transmission of Unique Candida Species to Premature Newborns

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Vertical and Horizontal Transmission of Unique Candida Species to Premature Newborns Linda A. Waggoner-Fountain, M. Whit Walker,* Richard J. Hollis, Michael A. Pfaller, James E. Ferguson II, Richard P. Wenzel,t and Leigh G. Donowitz

From the Department of Pediatrics and the Department of Obstetrics and Gynecology, University of Virginia, Charlottesville, Virginia; and the Departments of Pathology and Internal Medicine, University of Iowa, Iowa City, Iowa

Greater numbers of extremely low-birth-weight infants now survive in neonatal intensive care units (NICUs) because of the introduction of new medical therapies, such as antenatal steroid administration, postnatal surfactant replacement, the use of intravenous fat emulsions, and improved technological interventional techniques. Unfortunately, the survival of these highrisk infants has been associated with an increase in the incidence of disseminated candidal infections [1-4]. Colonization with Candida in the neonate is believed to be secondary to either vertical transmission from the mother or horizontal transmission (nosocomial acquisition) in the nursery. The risk of vertical transmission is increased because the rate of vaginal colonization with Candida in pregnant women ( ~ 30%) is two to three times the rate of colonization in nonpregnant women [2]. In addition, rates of colonization with Candida in preterm infants are significantly higher than rates of colonization in term infants, presumably because of the relative immunodeficiencies in the preterm infant (such as de-

Received 16 August 1995; revised 30 November 1995. The protocol for this study was obtained from the University of Virginia Health Sciences Center Human Investigation Committee, and verbal consent was obtained from all mothers. Grant support: This work was supported in part by the University of Virginia Children's Medical Center (grant no. 641212). • Current affiliation: Department of Neonatology • Greenville Memorial Hospital, Greenville, South Carolina. t Current affiliation: Department of Internal Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia. Reprints or correspondence: Dr. Linda A. Waggoner-Fountain, Department of Pediatrics, Division of Infectious Diseases, University of Virginia Health Sciences Center, Box 386, Charlottesville. Virginia 22908. Clinical Infectious Diseases 1996;22:803-8 © 1996 by The University of Chicago. All rights reserved. 1058-4838/96/2205-0023$02.00

creased skin and mucosal integrity, decreased function of neutrophils, and the relative quantitative deficiency of protective maternal IgO to Candida) [I, 3]. Candida albicans is frequently identified as a colonizing organism on skin and mucous membranes; however, when clinical infection does occur, species identification does not necessarily determine the infecting strain of C. albicans. It is possible for the same strain to exhibit multiple phenotypic morphologies [5]. Conversely, data from recent studies using DNA-based typing methods have demonstrated that phenotypically identical Candida from multiple sites in women may actually be genetically different [6, 7]. Since strain delineation emanates from the level of the genome, delineation requires the DNAbased techniques of modem molecular biology [8]. Current molecular typing methods using restriction endonuclease analysis of genomic DNA (REAG) and electrophoretic karyotyping (EK) with pulsed-field electrophoresis offer optimal means of discriminating specific Candida strains. These methods appear, at present, to be among the most practical typing methods for both large-scale and small-scale epidemiologic studies [8]. To date, the application of molecular typing for helping to define the relationship between maternal and neonatal colonization has been limited [9]. The goal of this prospective study of mother-infant pairs was to identify unique strains of Candida colonizing selected mothers at delivery and their preterm offspring. An additional goal was to determine if there was any specific strain of Candida that predominated in colonized neonates in our NICU.

Materials and Methods Study Population

Selected mothers admitted to the labor and delivery department at the University of Virginia Health Sciences Center

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The number of nosocomial bloodstreaminfectionsdue to Candida speciesin critically ill newborns is increasing.This pathogen may be verticallytransmitted from the mother or nosocomially acquired in the nursery. The goal of this study was to identify the route of transmission of unique Candida species and strains from mothers to their preterm offspring. Specimens from mothers for fungal cultures were obtained before delivery, and specimens from infants for sequential fungal cultures were obtained at defined intervals. Candida species were identified by standard methods and were typed by electrophoretic karyotyping (EK) and restriction endonuclease analysis of genomic DNA (REAG) with pulsed-field gel electrophoresis. Antifungal susceptibility testing was performed on all isolates. Fungal cultures were positive for Candida speciesin 12 (63%)of 19 mothers' specimens and in seven (33%)of 21 infants' specimens. EK and REAG revealed that both the mother and the infant in three (14%) of 21 mother-infant pairs were colonized with the identical strain of Candida albieans. C. albieans was most commonly transmitted vertically. Candida parapsilosis colonized other infants and could not be accounted for by a maternal reservoir.

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(Charlottesville, VA) from May 1993 through May 1994 who were likely to deliver an infant of 256.00 >256.00

C. albicans C. albicans C. albicans

2 2 2

G G G

1.0 1.0 1.0

0.25 0.12 0.50

>256.00 >256.00 >256.00

C. albicans C. albicans C. albicans C. alb/cans C. albicans C. albicans C. alb/cans C. alb/cans C. alb/cans

2 2 2 2 2 2 2 2 2

G G G G G G G G G

1.0 1.0 1.0

0.25 0.50 0.25

>256.00 >256.00 >256.00

1.0 1.0 0.5 0.5 1.0

0.25 0.50 0.50 0.50 1.00

>256.00 >256.00 >256.00 >256.00 >256.00

C. albicans

14

G

1.0

0.25

C. albicans C. parapsilosis

I I I 2 2 2 4 4 4

REAG

C. krusei C. albicans

C. parapsilosis

I 2 2 4 4

EK

NOTE. Specimens were obtained from mothers only once at time of delivery. AmB = amphotericin B; EK = electrophoretic karyotyping; 5-FC Flu = fluconazole; NT = not typed; REAG = restriction endonuclease analysis of genomic DNA; ... = data not evaluated. , Same organisms cultured during multiple weeks.

1.0

= flucytosine;

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Mother 2 Vaginal Rectal Infant 2b Rectal Crural Mother 4 Oral Vaginal Rectal Infant 4 Crural Oral Rectal Blood Mother 8 Oral Mother 9 Oral Infant 10 Crural Crural Rectal Crural Rectal Tracheal Mother II Oral Mother 12 Oral Infant 12 Rectal Crural Rectal Crural Rectal Mother 13 Vaginal Mother 14 Oral Infant 15 Rectal Mother 17 Vaginal Infant 17 Rectal Mother 18 Crural Oral Vaginal Rectal Mother 19 Crural Vaginal Rectal Infant 19 Crural Oral Rectal Crural Oral Rectal Crural Oral Rectal Mother 20 Crural

Week cultured

MIC (ltg/mL)

1

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2 3 4 5 6 7 8 9 10 S

same strain as their mothers. Despite the fact that the same genotype of albieans (2B) colonized two mother-infant pairs, the isolates from mother-infant pair 2 differed from the isolates from mother-infant pair 4 by resistance to fluconazole and were subsequently shown to be different based on preliminary work done by a PCR-based typing method (M . A. Pfaller, unpublished data). Second, in this study, parapsilosis was found to colonize infants and could not be accounted for by a maternal reservoir. The possibility of an external or environmental reservoir for parapsilosis is suggested by colonization with more than one strain and by colonization with the same strain in two individuals (table 1). parapsilosis is the most common species of Candida found on the hands of health care providers (M . A. Pfaller et aI., unpublished data and [16]) and frequently colonizes infants in NICUs [17]. Introduction of'C, parapsilosis from an exogenous source (e.g., hands) to a microbiologically naive infant could explain the pattern of colonization (multiple strains) and probable cross-infection observed.

e.

e.

e.

e.

Figure 1. Profiles of representative types ofrestriction endonuclease analysis of genomic DNA (REAG) from Candida species that were obtainedwith use of Sft I fol1owed by pulsed-fieldgel electrophoresis. Lanes 1-7, Candida albicans REAG types E, J, B, L, D, K, and H, respectively; lanes 8-10, Candida parapsilosis REAG types C, C, and M, respectively; lane S. phage ~ DNA concatemers as molecular size standards (in kilobascs).

by four different strains, and infant 12 was colonized by two different strains. These two infants were in the NICV during the same period and , despite the diversity of colonizing parapsilosis, were colonized with a common strain, DNA type 10 (figure 3 and table I) . The remaining patients colonized with parapsilosis (mother 11, mother 18, and infant 17) each were colonized with their own distinct DNA type (table I).

e.

e.

Discussion The data obtained from this study illustrate several important features of the epidemiology of colonization with Candida in mother-infant pairs. First, infants colonized or infected with albieans were colonized or infected with the same strain of albieans as their mothers. Although colonization was not demonstrated in most infants in this study, three of the four infants colonized with albieans were colonized with the

e. e.

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Figure 2. Profiles of restriction endonuclease analysis of genomic DNA (REAG) from serial Candida albicans isolates from motherinfant pair 19 that demonstratecolonization by the same strain. Lanes 1-5, C. albicans REAG type G isolated from crural, vaginal, and rectal specimens from mother 19; lanes 6-10, C. albicans REAG

type G isolated from crural, oral, and rectal specimens from infant 19; lane S, phage A DNA concatemers as molecular size standards (in kilobases).

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1 2 3 4 5 6 7 8 9 10 S

ern

1996; 22 (May)

Moth er-to-I nfant T ransmission of Candida

1 2 3 4 5 6 7 8 9 10 11 12 13 S

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ture high-risk neonates. In contrast, however, either C. parap silos is is not as easily transmitted vertically as C. albicans or the maternal inoculum of C. parapsilosi s may be altered sufficiently in number or viability that subsequent growth is prevented because vaginal colonization in mothers was not observed. Finally, although most isolates in this study were susceptible to commonly used antifungal agents, the detection of strains resistant to azoles and flucytosine is remarkable and should help guide the use of antifungal therapy for infants with invasive fungal disease. It may be that certain strains of Candida species are predisposed to colonizing pregnant women or that certain strains arc more likely to be vertically transmitted to high-risk neonates. If we are able to identify which strains of Candida species and specifically C. albicans are more likely to be vertically transmitted or cause systemic disease in neonates, then it may be reasonable to test the hypothesis that screening for fungal colonization in extremely premature infants could be beneficial in preventing subsequent serious fungal infections.

1. Baley JE, KJiegman RM , Fanaroff AA . Disseminated fungal infections in

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Finally, although most isolates in this study were susceptible to commonly used antifungal agents, the detection of strains resistant to azoles or flucytosine is notable. The combination of amphotericin Band flucytosine has been recommended for the treatment of hematogenous candidiasis in the neonate [3, 4, 18]; however, resistance to flucytosine may preclude its use for these patients. An attack rate of 3%- 4% has been previously described among neonates with invasive candidal infection in the NICU [2, 3]. We noted a similar attack rate for a single premature neonate (infant 4) who had systemic fungal disease due to C. albicans that was confirmed by positive cultures of blood and urine obtained at 8 days of age. The strain of C. albicans isolated from the blood had the same electrophoretic karyotype and restriction fragment digest pattern as the species isolated from the infant's skin and the infant's mother.

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Conclusion In this preliminary study using EK and REAG, we have shown that C. albicans can be vertically transmitted to prema-

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very low-birth-w eight infant s: clinical manifestation s and epidem iology. Pediatric s 1984; 73:144- 52. Miller MJ. Fungal infections. In: Remington JS, Klein JO, eds. Infectious disease of the fetus and newborn. 3rd ed. Philadelphi a: WB Saunders, 1990:475-515. Bale y JE. Neonatal candidiasis : the current challenge . Clin Perinatol 1991; 18:263 - 80. John son DE, Thompso n TR, Green TP, Ferrieri P. Systemi c candidiasis in very low-birth-weight infants (less than 1,500 grams) . Pediatrics 1984; 73: 138-43. Hellstein J, Vawter-Hugart H, Fotos P, Schmid J, Soil DR. Genet ic similarity and phenotypic diversity of commensal and pathogenic strains of Candida albicans isolated from the oral cavity. J Clin Microbiol 1993;3 1:3190-9. Soil DR, Ga lask R, Schmid J, Hanna C, Mac K, Morrow B. Gen etic dissimilarity of commensa l strains of Candida spp. carried in different anatomical loca tions of the same healrhy women. J Clin Microbiol 1991;29: 1702-10. Betremieux P, Chevrier S, Quindos G, Sullivan D, Polonelli L, Guiguen C. Use of DNA fingerprin ting and biotyp ing met hods to study a Candida albicans outbreak in a neonatal intensive care unit Pediatr Infect Dis J 1994; 13:899-905 . Pfaller MA. Epidemiological typing methods for mycos es. Clin Infect Dis 1992; 14(suppJ I ):S4 -1O. Carlson S, Hostetter M, Finkel D, Bendel C. Genotypic and functiona l analysis of Candida albicans strains from mother-infants pairs [abstra ct no 1754]. Pediatr Res 1994;35(4 pt 2):295A . Voss A, Holl is RJ, Pfaller MA, Wenzel RP, Doebbel ing BN. Investigation of the sequence of colo nization and candidemia in nonneutropenic patienrs. J Clin Microbiol 1994; 32:975 - 80. Voss A, Pfaller 11A, Hollis RJ, Rhine-Chalberg J, Doebbeling BN . Investigation of Candida albicuns transmission in a surgical inte nsive care unit cluster by using genomic DNA typing methods. J Clin Microbiol 1995;33:576 -80. Branchini ML, Pfaller MA , Rhine-Chalberg J, Frempong T, Isenberg HD . Genotypic variat ion and slime production among blood and catheter isolates of Candida parapsilosis. J Clin Microbiol 1994;32:452 - 6.

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References

Figure 3. Profiles of restriction endonuclease analysis of genomi c DNA (REAG) from Candida parapsilosis and Candida albieans isolates from infants and mothers . Lanes 1-5, C. parapsilosis REAG type C isolated from crural , rectal , and tracheal specimens from infant 10; lane 6, C. albieans REAG type J isolated from an oral specimen from mother 12; lanes 7-11, C. p arapsilosis REAG type C isolated from crural and rectal specimens from infant 12; lane 12, C. albieans REAG type G isolated from mother 20; lane 13, C. albie ans REAG type K isolated from mother 14; lane S, phage A. DNA concatemers as molecular size standards (in kiloba ses).

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