The aim of the study was to define the most common causes, symptoms and clinical features of vulvovaginitis in prepubertal girls, and to evaluate treatment success depending on the causative agent involved. The study included 115 girls aged 2-8 (mean 4.8) years, presenting with vulvovaginitis to the Outpatient Clinic for Pediatric and Adolescent Gynecology, Zagreb Children's Hospital, between September 2006 and July 2007. Medical history data were obtained from parents. Vaginal samples were collected for microbiological culture by using cotton-tipped swabs moistened with saline. All samples were referred to microbiology laboratory, where standard microbiological diagnostic procedures were performed. Selective and non-selective media were used. Of 115 study patients, 43 (37.4%) had received antibiotic therapy more than one month prior to their visit to the Clinic, mainly for upper respiratory tract infection. The most common presenting symptom was increased vaginal discharge usually noticed on the pants or diaper, found in 26 of 115 (22.6%) patients, followed by vulvar redness in 16 (13.9%), burning in seven (6.1%), itching in the vulvovaginal area in seven (6.1%), soreness in six (5.2%), odor in three (2.6%) patients, and two or more of these symptoms in another 50 (43.5%) patients. Fifty-nine of 115 children had normal clinical finding on gynecologic examination. Among the remaining 56 children, the most common finding was erythema observed in 19, vaginal discharge in ten, and a combination of discharge and erythema in 13 patients. Of 115 study patients, causative agents were isolated from vaginal culture in 38 (33%) cases. Of these, 21 grew group A beta hemolytic streptococcus, five patients Haemophilus influenzae, three Escherichia coli, two Enterococcus spp., and one each Staphylococcus aureus, Proteus mirabilis, and Streptococcus pneumoniae. Antibiotic therapy was administered in 31 of these 38 patients, except for those cases where intestinal bacteria and Staphylococcus aureus were isolated and topical therapy and hygienic measures were applied alone. Accordingly, vulvovaginitis in girls was most commonly caused by pathogens originating from the patient upper respiratory tract, accompanied by the symptoms of redness and vaginal discharge. In these cases, antibiotic treatment was administered. In the majority of prepubertal girls with either vulvitis or normal genital finding, simple measures to improve hygiene will lead to resolution of all symptoms.