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BAGIAN MIKROBIOLOGIFAKULTAS KEDOKTERANUNIVERSITAS HASANUDDIN2015PENJELASAN PRAKTIKUMSISTEM UROGENITALIA

1MICROBIOLOGY LAB ASSISTANTS 2014-2015MUH. RIDHO AKBARMUHAMMAD AZZAMAPRIZAL AZHARANDI MUH OCTAVIAN PRATAMAANOM PARTHA JAYAAGUNG DANAN JAYARADHI ARIFINMUH. FARID FETTY ANDRIANIFADHILAH PUTRI WULANDARINURUL FAJRINAAISYAH NAULI SIHOTANGMUFIDAH NUR QISTHISRI RINIA SARI ISWANTIHANA AMIERAAYU AQILAH BT. KHAZANITATA TERTIBLABORATORIUM MIKROBIOLOGILaki-laki :Memakai KEMEJA, CELANA PANJANG KAIN, dan SEPATU TERTUTUP. Rambut TIDAK BOLEH menyentuh kerah kemeja.Perempuan :Memakai KEMEJA, ROK PANJANG KAIN, dan SEPATU TERTUTUP. Rambut diikat ke atas dan bagi yang berjilbab, jilbabnya dimasukkan ke dalam jas lab.Perlengkapan yang WAJIB dibawa saat praktikum :Jas labName tagPenuntun praktikum + pas foto berwarna 4x6MaskerHandscoenPensil warnaPenggaris 30 cm (per kelompok)ASISTENSI & RESPONSITelah mengumpulkan Tugas PendahuluanPRAKTIKUMTelah mengikuti Asistensi & ResponsiMemenuhi Tata Tertib Laboratorium MikrobiologiMembawa Perlengkapan PraktikumUJIAN PRAKTIKUMTelah mengikuti PraktikumTelah mengumpulkan Laporan PraktikumMemenuhi Tata Tertib Laboratorium MikrobiologiREMEDIAL UJIAN PRAKTIKUMTelah mengikuti Ujian Praktikum namun belum dinyatakan lulus Ujian PraktikumPERSYARATANINTRODUCTIONUrinary Tract InfectionBacterial Agents Causing UTIUrine Sample & CollectionBacteriological Examination5Review : BIOCHEMICAL TESTSlant & ButtAlkali:MerahAsam:KuningNetral:OrangeH2S(+):Hitam (-):Warna dasarGas(+):Medium terangkat/ pecah(-):Medium tidak terangkat

TSI (TRIPLE SUGAR IRON) TESTH2S(+):Hitam (-):Warna dasarIndol(+):Terbentuk cincin merah(-):Tidak terbentuk cincin merahMotilitas(+):Stabline diffuse(-):Stabline tidak berubah

SIM (SULFIDE INDOLE MOTILITY) TESTMethyl Red

(+):Merah(-):Kuning / OrangeVoges Proskauer

(+):Merah(-):Kuning / Colorless

METHYL RED / VOGES PROSKAUER (MR/VP)Citrate Utilization

(+):Biru(-) :Hijau

Urea Hydrolysis

(+):Pink(-) :KuningCITRATE UTILIZATION & UREA HYDROLYSIS TESTLactose FermentationSucrose FermentationGlucose FermentationMannitol(+):Kuning(-):Merah

SUGAR FERMENTATIONURINARY TRACT

12Aninfectionthat affects part of theurinary tract caused by pathogenic bacterial infection or opportunistic infection.Bacteriuria is a condition when the presence of bacteria in urine with the number of 105 cfu in 1 ml of midstream urine or 103 cfu in 1 ml of urine by supra-pubic aspiration.Pyuria is the condition of urine containing pus in urine, or white blood cell with the number of 5 cells/HPF or > 10 cells/mm3.URINARY TRACT INFECTIONcolony forming unit 13Escherichia coli (80%)Coagulase-Negative Staphylococci (7%)Staphylococcus saprophyticus in young womanProteus mirabilis (6%)Other Gram-Negative Bacteria (4%)Other Gram-Positive Bacteria (3%)-Hemolytic Group B Streptococcus in pregnant womanBACTERIAL AGENTS CAUSINGURINARY TRACT INFECTION-Hemolitc Group B Streptococcus > Streptococcus agalactiae14Other organismAnaerobic bacteria (2% - rare)Mycobacterium tuberculosisMycobacterium spp.Campylobacter spp.Chlamydia trachomatisGardnerella vaginalisHaemophilus influenzaeLegionella pneumophilaMycoplasma hominisUreaplasma urealyticumSalmonella spp.Shigella spp.AdenovirusBACTERIAL AGENTS CAUSINGURINARY TRACT INFECTIONMycobacterium spp. > M. bovis & M. africanumCampylobacter spp. > C. jejuni & C. coli

15INTRODUCTIONProvidencia stuartiiProvidencia alcalifaciensAlcaligenes faecalisPseudomonas aeruginosaEscherichia coliKlebsiella pneumoniaeProteus vulgarisProteus mirabilisEnterobacter agglomerans16Family enterobacteriaceaeGram-negative bacteriaFacultative anaerobicRod shapedCommonly found in the lowerintestineHave O, H, and K antigensEscherichia coliEscherichia ColiSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAcid/Acid-++++---++++

O > lipopolisakaridH > flagelK > kapsul17Family enterobacteriaceaeGram-negative bacteriaFacultative anaerobicRod shapedCommonly found in the mouth, skin, and intestineOnly have O and K antigensKlebsiella pneumoniae

Klebsiella pneumoniaeSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAcid/Acid-+--+-++++++

18Proteus vulgarisFamily enterobacteriaceaeGram-negative bacteriaFacultative anaerobicRod shapedInhabits the intestinal tracts of humans, animals, and raw meat, can also be found in soil, water, and fecal matterProteus vulgarisSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAlkali/Acid+++++-+++-++

Proteus mirabilisFamily enterobacteriaceaeGram-negative bacteriaFacultative anaerobicRod shapedCan lead to the formation ofcrystalsofstruvite and calcium carbonatePart of the normal flora of human gastro-intestinal tract, can also be found free living in water and soilProteus mirabilisSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAlkali/Acid++-++-+++--+

Enterobacter agglomeransFamily enterobacteriaceaeGram-negative bacteriaAlso known as Pantoea agglomeransAerobic bacteriaRod shapedInhabits plants (grows symbiotically with various plants), soil, water, and such species includes bacteria reported as both commensal and pathogen of animals and humansEnterobacter agglomeransSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAlkali/Acid------+++-+-

21Providencia alcalifaciens

Family enterobacteriaceaeGram-negative bacteriaFacultative anaerobicRod shapedCould cause Travelers DiarrheaProvidencia alcalifaciensSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAlkali/Acid---++-+++---Providencia stuartiiProvidencia stuartiiSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAlkali/Acid--+++-+-++--Family enterobacteriaceaeGram-negative bacteriaFacultative anaerobicRod shapedCommonly found in the intestine

Alcaligenes faecalisAlcaligenes faecalisSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAlkali/Neutral-+-+--+-----Family alcaligenaceaeGram-negative bacteriaObligat aerobRod shapedOptimum growth occurs at about 37C

24Pseudomonas aeruginosaPseudomonas aeruginosaSlant & ButtH2SGasIndoleMotilityMRVPCitrateUreaSugar FermentationGlucoseLactoseSucroseMannitolAlkali/Neutral-+-+--++----Family pseudomonadaceaeGram-negative bacteriaObligat aerobCoccobacillus bacteriaHave O, H, and K antigensHave 3 pigments :Pyocyanin (blue - green)Pyoverdine (yellow - green and fluorescent)Pyorubin (red - brown)

TSIAlkaliNeutralEscherichiacoliKlebsiellapneumoniaeIndoleCitrate&UreaProteusProvidenciaIndolMacConkeyBrown + GreenAlcaligenes faecalisPseudomonas aeruginosaH2SKlebsiellapneumoniaeEscherichiacoli-+AlkaliAcid-+-+-+Proteus vulgarisProvidencia stuartiiProteus mirabilisProvidencia alkalifaciensBrownAcidAcid26

The urine sample should be collected in a sterile manner and put in a sterile container.Sterilization cant be done with :Wet heat sterilizationMay decrease the volume of urine sample leading to dilution of urine sample.Chemical sterilizationMay leave the chemical substance at the wall of container and may kill the bacteria.Good urine for bacteriological examination is morning urine, except for Mycobacterium tuberculosis examination, 24 hours urine is usually used.URINE SAMPLE & COLLECTIONURINE COLLECTIONMidstream UrineUrine PocketSupra-Pubic AspirationCatheterizationMost commonly used for urine collection and bacteriological examination of urine.Urine specimen should be taken in the morning (morning urine)High risk of contamination (> 20%) and give a false-positive culture.Doesnt cause discomfort to the patient/client.Easy to do and doesnt require any special skills and tools.Difficult to do in patients with urinary incontinence.Give a false-negative culture if use antiseptics to clean the genital.MIDSTREAM URINEMIDSTREAMURINE

URINE POCKETUsually used for uncooperative patients and collected 24 hours urine.Medium risk of contamination (6-10%).Easy to do and doesnt require any special skills.

URINE POCKET

SUPRA-PUBIC ASPIRATIONThe most sterile method of collecting urine (specificity 99%)The gold standard for bacteriological examination for urine.Requires to be done in the morning (morning urine) and the bladder must be in full condition.Requires any special skills and tools.Cause discomfort to the patient/client (pain).

CATHETERIZATIONShould be avoidedLead to irritation of urogenital tract mucosa and enhance the infectionLow risk of contamination (95%)Discomfort sensation for patients.Must be done by a trained person.

CollectingStoringTransportingLaboratoryMidstream UrineSupra-Pubic AspirationUrine PocketSterilePunction with spoitUrine pocket or bagSterile bottleat 4ocSterile bottleat 4ocSterile bottleat 4ocIce boxCentrifugeGram stainingIsolation & IdentificationBacterial countCOLLECTING, STORING, TRANSPORTING, AND LABORATORIUMGram staining of direct preparationBacterial countPour plate methodSurface drop methodDipslide methodIsolation and identificationAntibiotic sensitivity testingBACTERIAL EXAMINATIONDirect preparation can be made from urine sediments or urine without centrifugation. From this preparation, we can see;- The number of bacteria per field- The number of leucocytes per field (used as a criteria for pyuria)GRAM STAINING OF DIRECT PREPARATION

POUR PLATE METHODPOUR PLATE METHOD

SURFACE DROP METHOD

DIPSLIDE METHOD

DIPSLIDE METHOD

Urine is dripped onto the surface of MacConkey /Nutrient agar/Blood agar/Chocolate agar.Strike with a loop or a swab on the media surfaceCulture is incubated overnight & the appearance of growing colony observed.

*If there are 2 types of bacterial growth; considered as contaminated (repeat)ISOLATION & IDENTIFICATIONANTIBIOTIC SENSITIVITY TESTPengukuran diameter zona jernih (zona inhibisi atau clear zone) di sekitar disk antibiotik lalu hasil pengukuran dibandingkan dengan tabel rujukan untuk menentukan interpretasinya.

GenderObstructionSystemic infectionPregnancyCateterizationDiabetes MellitusTumorIritationPREDISPOSITION FACTORS UTI

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