Infect Dis Obstet Gynecol 2003;11:19–26 Leukocyte esterase activity in vaginal fluid of pregnant and non-pregnant women with vaginitis/vaginosis and in controls Per-Anders Mårdh 1, Natalia Novikova 1, Ola Niklasson 1, Zoltan Bekassy 1 and Lennart Skude 2 1Department of Obstetrics and Gynecology, University Hospital, Lund and 2Department of Clinical Chemistry, County Hospital, Halmstad, Sweden Objectives: Todeterminethe leukocyteesterase (LE) activity invaginal lavage fluid ofwomen with acuteand recurrentvulvovaginal candidosis (VVC and RVVC respectively), bacterial vaginosis(BV), and in pregnant and non-pregnantwomen without evidenceof the threeconditions. Also tocompare the resultof LEtestsin women consultingat differentweeks inthe cycle andtrimesters of pregnancy.The LEactivity was correlatedto vaginal pH, numberof inflammatory cells instained vaginal smears,type of predominatingvaginal bacteria andpresence of yeast morphotypes. Methods: Onehundred and thirteen women with ahistoryof RVVC,i.e. with at least fourattacks of the conditionduring the previousyear and who had consultedwith anassumed new attack ofthe condition,were studied.Furthermore, we studied16 women with VVC,15 women with BV,and 27 women attendingfor control of cytological abnormalities, who all presentedwithout evidenceof either vaginitisor vaginosis. Finally, 73 pregnantwomen wereinvestigated. The LEactivity invaginal fluid duringdifferent weeks inthe cycle of 53 of the women was measured. Results: Inthe non-pregnantwomen, anincreased LE activity was foundin 96, 88, 73 and56% of those with RVVC,VVC and BV andin the non-VVC/BVcases,respectively. In 73% of pregnantwomen inthe second trimester,and 76% of thosein the third,the LEtestwas positive.In all groupsof non-pregnantwomen tested, the LEactivity correlatedwith the numberof leukocytesin vaginal smears,but it didnot in those who were pregnant.There was nocorrelation between LE activity andweek incycle. The vaginal pHshowed nocorrelation to LE activity in any of the groups studied. Conclusions: The useof commercial LEdipstickshas alimited valuein the differential diagnosisof RVVC,VVC and BV.There isno correlation between the LEactivity invaginal secretionon one hand andvaginal pH,week in the menstrualcycle andtrimester in pregnancy on the other.Women with BVoftenhave signsof inflammation as evidenced by a positive LE test and inflammatory cells in genital smears. Key words:L EUKOCYTE ESTERASE; VULVOVAGINAL CANDIDOSIS; RECURRENT VULVOVAGINAL CANDIDOSIS; BACTERIAL VAGINOSIS; PREGNANCY Ifthe office diagnosis ofvaginitis/ vaginosis is genital tract, forexample ofvaginitis, microscopy onlybased on history and clinical examination, ofvaginal fluidfor detection of granulocytes has theaccuracy will beunacceptablylow 1–3. To diag- beenthe standard procedure. Bacterial vaginosis noseinflammatory reactions inthe lower female (BV) has beenconsidered to be an– osis, thatis, a Correspondenceto: Per-AndersMå rdh, Department of Obstetrics and Gynecology, Lund University Hospital, SE22185Lund, Sweden. E-mail: [email protected] ã 2003 The Parthenon Publishing Group 19 Leukocyte esterase activity in vaginal fluid Mårdh et al. non-inflammatorycondition. Amsel’ s criteria 4, TheLE activity was correlatedwith the pre- whichdo not include the presence ofinflamma- dominatingbacterial flora, independentof the torycells invaginal fluid,have generally been presence ofcandida morphotypes. The presence employed to diagnose BV. ofextremely elongatedGram-positive rods(of Leukocyteesterase (LE) tests have been lactobacilli morphotype) was also noted. launchedas oneof several chemical tests ondip- sticks employedin the diagnosis ofurinary tract infections(UTI). Contradictoryresults ofthe Patients with a history of recurrent vulvovaginal diagnosticvalue ofLEtests have beenreported 5,6. candidosis Anumberof studies have reportedon theuse of Oneof thecohorts studied comprised 113 women anLE dipsticktest ofurine to diagnose urthritis witha historyof RVVC, i.e. patients whohad in males7,8,while onlysingle studies have used hadat least fourattacks of the condition during suchtests inthe diagnosis offemale genital theprevious year, andwho had consulted with infections9–11. symptoms suggesting anewattack of RVVC. The TheLE test is basedon detectionof an enzyme diagnosticcriteria appliedhave beendescribed in thathydrolyses indoxyl carboxylic acid ester to detail elsewhere 13,14.Thenumberof these women indoxyl,which in turn reacts witha diazoniumsalt whohad a positive LE test has already been to produce a purple colour 12. reportedin a previous study 13.Inthepresent study, Thepresent study was aimed todetermine the weexpandedour investigation bycomparingthe LEactivity invaginal secretion ofpregnant and LEtest results withfindings of candida morpho- non-pregnantwomen with and without evidence types, i.e. ofblastoconidia and pseudohyphae, in ofvaginitis andvaginosis. Samples collectedat Gram- andmethylene blue-stainedvaginal smears. differentweeks ofthemenstrual cycle andat the Themethodsused to identifythose with a positive secondand third trimesters ofthe gestational candidaculture have beenreported in detail else- periodwere studied.We also studiedany correla- where15.Inbrief, vaginal introitaland posterior tionbetween the LE test results andthe number vaginal fornixsamples were culturedon Sabouraud of inflammatory cells in vaginal smears. andchromogenic agar 16.Speciationof isolated candidastrains was donewithAPI 50C kitsandthe Vitek® automised identificationsystem (bothfrom SUBJECTS AND METHODS BioMerieu, Paris). Thevaginal pHwas deter- mined bypHindicatorstrips (Merck, Darmstadt). Leukocyte esterase tests and pH determinations TheLE activity invaginal flushingfluid samples TheLE activity was measured bya commercial was determined byCombur-4 ® (Roche,Basle) kit (Ecur4-Test, Boehringer–Mannheim, later stick. renamed Combur-4and sold by Roche, Basle). Onedrop (approximately 0.3 ml) ofvaginal lavage Patients attending for control of cytological fluidwas mixed with3 ml of0.1M TRIS HCl abnormalities buffer,pH 7.3 orwith physiological saline. The tests were performedwithin a coupleof minutes Twenty-seven womenin whom a previous cyto- ofsampling. Accordingto the package insert, color logical screening hadshown cell abnormalities changesof grades 1, 2and3 correspondto 10–25, (butno signs ofcervical cancer) were investigated. approximately75, andapproximately 500 granu- Thepatients, all ofwhom were non-pregnant,had locytes/ml, respectively. nosigns orsymptoms ofVVC orBV. Thepatients Another1– 2 dropsof the vaginal lavage were were tested forLE activity, vaginal pHand for placedon anindicator strip forpH determination presence ofinflammatory cells andpredominating (Merck, Darmstadt, Germany). Vaginal samples bacterial flora, i.e. either apredominanceof were tested forLE activity withinsome minutes as Gram-positive rods(of lactobacilli morphotypes) well asafter 6hours’storage 2 at roomtemperature oramixture ofbacterial morphotypes(‘ BVflora’). in TRIS buffer and in physiological saline. Furthermore, Amsel’s criteria were searched for: 20 INFECTIOUSDISEASES IN OBSTETRICS AND GYNECOLOGY Leukocyte esterase activity in vaginal fluid Mårdh et al. Table 1 Relationbetween vaginal pH andleukocyte esterase activityin vaginal lavage fluid of women with ahistory of and an assumed new attack of recurrent vulvovaginal candidiasis Number of women with indicated test result* Number of cases Grades 0–1 Grade 2 Grade 3 Vaginal pH (n = 113) (n = 14) (n = 22) (n = 77) p-value £ 4.4 57 6 13 (12%) 38 (34%) 0.904 4.5–5.4 43 6 7 (6%) 30 (27%) 0.903 ³ 5.5–6.4 13 2 2 9 (69%) 0.927 *Grading according to package insert (Combur-4 Ò, Roche, Basle) increased vaginal pH, acharacteristic gray homo- Statistical analyses geneousvaginal content,presence ofclue cells Data were analysed using c2 and student t-tests. (in methylene blue-and Gram-stained vaginal A p-value <0.05 was considered significant. smears), anda positive amine test obtainedby adding 15% KOH to vaginal fluid. RESULTS Patients with vulvovaginal candidosis and Leukocyte esterase activity in cases with history bacterial vaginosis and non-pregnant healthy of recurrent vulvovaginal candidosis women Inthe 113 cases witha historyof RVVC and Sixteen womenwith the diagnosis ofVVC and withsymptoms suggestive ofanewattack of the 15with BV were tested forLE activity invaginal condition,there was acorrelationbetween the secretion. BVwas diagnosedif three offour of LEactivity andthe result ofthe yeast cultures Amsel’s criteria were fulfilled 4. The VVC and fromthe genital tract( p <0.05). Thus,of the63 BV women were all non-pregnant. candidaculture-positive women,all hadpositive Thediagnostic methods were thesame as LEtests. Ofthe 50 culture-negative women,45 thoseapplied in the cases withcytological had such an increased activity. abnormalities. Anacute attack of VVC was con- Therewas nocorrelationbetween the result of sidered whenthe woman reported symptoms pHdeterminationof vaginal lavage andthe result suchas pruritus,discharge andpain. Signs, suchas oftheLEtests (Table 1). Thus,of 57 womenwith oedema anderythema ofthe introitus and the avaginal pHofless than4.5, 51(89%) hadan LE vaginal mucosawith plaques, and/oranincreased test ofgrades 2and3, while ofthe43 women with vaginal discharge (oftenof cotton-cheese type), apHof 4.5– 5.4, 37(86%) hadsuch a result. were required.The presence ofcandida blasto- Eleven ofthe 113 women with an even higher conidiaand/ orhyphae in stained vaginal smears pH had a positive LE test