MODULE 6

RIZZA A. CALUAG
MICROBIOLOGY
 Eukaryotes Prokaryotes

Protists, fungi,
Examples Bacteria/archaea
plants, animals

Real nucleus NUCLEUS No real nucleus

Linear DNA Circular

40s + 60s = 80 RIBOSOMES 30s + 50s = 70

3
• A prokaryotic chromosome is usually a circular
molecule (an exception is that of the bacterium Borrelia
burgdorferi, which causes Lyme disease).
• Even without a real nucleus, the DNA is condensed in a
nucleoid.
• Prokaryotes can carry extrachromosomal DNA
elements called plasmids, which are usually
circular. Plasmids can carry additional functions, such as
antibiotic resistance.
Live in extreme environments
Halophiles: HIGH salt concentration
Thermophiles: HIGH temperature (>6OdegC)
Barophiles: HIGH pressure environment
Methanogens: HIGH methane environment

Temperature
Psychrophilic – cold temperature (~ -20degC)
Mesophilic – room temperature
Thermophilic – hot temperature (>60degC) (e.g. C. jejuni)

pH
Basophile (basic)
e.g. Vibrio, Campylobacter, curve MO like Leptospira
Mesophile (neutral)
Acidophile (acidic)
e.g. fungi (loves wet and acidic)
 Obligate • really need O2 • Leptospira
aerobes • TB (upper lobes of lungs:
highly perfused w/ O2)
• Pseudomonas
Facultative • uses O2 when present but
aerobes survives without O2
• can live in anerobic envi coz
can perform anaerobic
glycolysis
Aerotolerant • more anaerobic
• utilize anaerobic glycolysis
but doesn’t die in O2
• bet. aero & anaero
Obligate • die when O2 is present • ABC – Actinomyces,
anaerobes Bacteroides, Clostridium
Capneophile • loves CO2 • Campylobacter (dysentery,
acute gastroenteritis)
Microaerophile • needs only 5% O2 • Campylobacter
 BACTERIAL TOXINS
Endotoxins Exotoxins
G- G+, G-
Lipopolysaccharide More virulent
(w/ Lipid A)
Heat stable Heat-labile
E.g. Meningococcemia E.g. Tetanus pasmin,
Botulinum toxin
Capsulated bacteria–
Some Killers Have Pretty Nice Capsule
• Streptococcus pneumoniae
• Klebsiella pneumoniae
• Haemophilus influenzae
• Pseudomonas aeruginosa
• Neisseria meningitidis
• Cryptococcus neoformans
Gram +
Streptococcus
Staphylococcus
Listeria Mycobacterium
Bacillus Spirochetes
Corynebacterium Mycoplasma
Clostridium

Gram -
Neisseria Vibrio
Campylobacter Haemophilus
Chlamydia Pseudomonads
Rickettsiae Helicobacter
Legionella
Giemsa, Wright and Silver Spirochete
Stain
Welch Capsule Stain
Malachite Green Spores
Fuelgen Nuclei
M. Fadyean B. anthracis

Lowenstein Medium M.Tuberculosis

Modified Thayer Martin Neisseria
Mac Conkey Agar G-
Fletcher’s Media Leptospira interrogans
Chocolate Agar Haemophilus influenzae
COCCI
Novobiocin test to distinguish coagulase +/-
Staphylococci :

• S. epidermidis stays away from the novobiocin

disc like an epidemic.
• Therefore, epidermidis is sensitive to
novobiocin.
• S. saprophyticus, much friendlier, comes up to the
disc and says, "'sap?'” [short for "whassup?”]
• Therefore, saprophyticus is novobiocin
resistant. It’s okay to be exposed.
Streptococci: Quellung reaction

"Quell-lung"
Quell: Capsules swell [+ve test]
Lung: S. pnuemonia
· You get pneumonia in your lung.
 DIARRHEA
EPEC Infant and Children diarrhea
ETEC Traveller’s Diarrhea
EHEC Bloody Diarrhea
Shigella dysenteriae Dysenteric Diarrhea
Vibrio cholerae Rice water diarrhea
Campylobacter jejuni Bloody loose diarrhea
Bacillus Cereus Starchy Diarrhea
Giardia lamblia Non-bloody Diarrhea
Cryptosporidium and Isospora Severe diarrhea among AIDS Px
2 Forms of Fungi
YEAST MOLD
Filamentous (w/ Nonfilamentous
hyphae) (do not have hyphae)
Unicellular Multicellular
Live at body Temp Live at room Temp
(37oC) (25oC) [outside the
body]
 CUTANEOUS MYCOSES
TINEA LOCATION
Tinea pedis (Athlete’s Feet
foot)
Tinea cruris (Jock itch) Groin area

Tinea unguium Nails

(Onychomycosis)
Tinea manuum Hands
Tinea corporis Trunk
Tinea barbae Beard area/chin
Tinea capitis Scalp/head
Cestodes
CAUSATIVE AGENT CLINICAL MANIFESTATION TREATMENT
Taenia solium • Neurocystecorcosis → ✓ DOC: Praziquantel
(pork tapeworm) seizures
• Mild intestinal Sx
Taenia saginata • N/V/D, abdominal pain ✓ DOC: Praziquantel
(beef tapeworm)
Diphyllobothrium latum • Vit B12 deficiency ✓ DOC: Praziquantel
(fish tapeworm) anemia (pernicious
anemia)
Hymenolepsis nana • Abdominal pain, diarrhea ✓ DOC: Praziquantel
(dwarf tapeworm)
Echinococccus ✓ DOC: Albendazole
granulosus
(dog tapeworm)
Trematodes – flukes
DISEASE CAUSATIVE AGENT CLINICAL TREATMENT
MANIFESTATION
Schistosomiasis Schistosoma japonicum Katayama Fever DOC: Praziquantel
(Oriental Blood Fluke) Chronic Liver
Pathology
Lung Fluke Paragonimus westermani Mimics TB ~ lung DOC: Praziquantel
(lung fluke) cavitation
• Hermaphrodite

Liver Fluke • Chlonorchis sinensis • Inflammation of DOC: Praziquantel

(Chinese liver fluke) lymph nodes
• Opisthorchis viverrini • Hepatomegaly
(Southeast Asian liver
fluke)

MOT: Undercooked fish

meat
NEMATODES
DISEASE PARASITE OTHER NAME
Anisakiasis Anisakiasis worm/ Phocanema

Ancylostomiasis Ancylostoma duodenale Old World hookworm

Ascariasis Ascaris lumbricoides Giant Intestinal Roundworm

Capillariasis Capillaria philippinensis
Dracunculiasis Dracuncuculus medinensis Guinea/serpent/dragon worm
Enterobiasis/oxyuriasis Enterobius vermicularis Pinworm/seatworm
Bancroftian filariasis Wuchereria bancrofti Bancroft’s filaria
Malayan Filariasis Brugia malayi Malayan filarial worm
Filariasis, occult Dirofilaria species Heartworm
Loiasis/fugitive swelling/calabar swelling Loa loa Eye worm

Necatoriasis Necator americanus New world Hookworm

American Hookworm
Onchocerciasis Onchocerca volvolus Nodular/Blinding worms
Strongyloidiasis/ Strongyloides stercoralis Threadworm
Cochin-china diarrhea
Trichinosis/Trichinellasis Trichinella spiralis Trichina worm
Trichuriasis/trichocephaliasis Trichuris trichura Whipworm
PROTOZOA TRANSMISSION MORPHOLOGY CLINICAL DIAGNOSIS
Entamoeba Fecal-oral -oocyst -asymptomatic: -fecal exam
histolytica -trophozoite: motile carriage -serology
-not all species bulls-eyed shaped -bloody diarrhea -abdominal CT
of Entamoeba nucleus w/ RBC in -liver abscess scan
are pathogenic cytoplasm -AMEBIASIS

Giardia lamblia Fecal-oral -oocyst Foul smelling -fecal exam

-trophozoite: greasy diarrhea and -commercialized
flagellated abdominal gassy immunoassay kit
distention
- GIARDIASIS
Trichomonas Sexually -no cyst stage -painful vaginal -examination of
vaginalis fomites -trophozoite: itching vaginal discharge
flagellated -burning on
urination
-yellow green
malodorous frothy
vaginal discharge
-TRICHOMONIASIS/
VAGINITIS/PINGPO
NG INFECTION
African -Tse tse fly -Trypomastigote: African -visulaize -Suramin
Trypanosome bite motile, Sleeping trypomastigote -Melarsoprol
-Trypanosoma -contaminated extracellular form Sickness in blood, spinal,
rhodesiense BT -Trypomastigote fluid or lymph
-Trypanosoma and epimastigote nodes
gambiense in tse tse fly -serology

American -Kissing Bug: -Trypomastigote: Chaga’s -visualize -Nitrofurtimox

Trypanosome defecates on motile disease trypomastigotes -Benznidazole
-Trypanosoma human skin extracellular form in blood
cruzi while feeding -Amastigote: non -xenodiagnosis
- motile,
Contaminated intracellular
BT -Trypomastigote
and epimastigote:
in kissing bug
 Important Tissue or Blood Protozoans
Malaria Female anopheles -trophozoites Malaria
-Plasmodium falciparum mosquito -schizonts
-Plasmodium vivax -gametocytes
-Plasmodium ovale
-Plasmodium malariae
• CHLOROQUINE: DOC IN ACUTE ATTACK, also used in
PROPHYLAXIS
• Primaquine: eliminates the exoerythrocytic forms in the
liver(relapsing malaria) : radical cure
• Mefloquine- chemoprophylactic drug of choice in chloroquine
resistance
• Proguanil (100 mg) and atovaquone (250 mg)– malarone →
both for prophylaxis and treatment of multi-drug resistant
falciparum malaria
• Pyrimethamine-sulfadoxine → presumptive treatment in
case of malarial breakthrough
• *no drug regimen can ensure prevention of malaria
 Leishmaniasis
• Leishmania tropica, Leishmania chagasi, Leishmania
major, Leishmania braziliensis, Leishmania donovani
• Zoonotic disease
• Sandfly

Leishmania donovani
Visceral leishmaniasis, Kala-azar- Dumdum fever
Leishmania braziliensis
Mucocutaneous leishmaniasis

Tx: antimonial stibogluconate (Stibofen ®)

 The 21 Virus Families
DNA viruses
(HHAPPPPy)
Hepadnaviridae
Herpesviridae
Adenoviridae
Poxviridae
Papovaviridae
Parvoviridae
Polyomaviridae
 The 21 Virus Families
RNA viruses
(PRAyFOR PRC)TB
Picornaviridae
Reoviridae
Astroviridae
Arenaviridae
Flaviviridae
Filoviridae
Orthomyxoviridae
Retroviridae
Paramyxoviridae
Rhabdoviridae
Caliciviridae
Coronaviridae
Togaviridae
Bunyaviridae
 DNA Viruses RNA Viruses
Double EXCEPT Parvovirus Single EXCEPT Reoviridae
stranded stranded and Rotaviridae
Icosahedral EXCEPT Helical/ EXCEPT Rhabdo →
Poxvirus (complex) icosahedral helical bullet

*Naked EXCEPT Enveloped EXCEPT Reoviridae,

Hepadnavirus, Herpes Picornaviridae,
virus, Pox virus Calciviridae (PRC)
(enveloped)
Replicate in EXCEPT Replicate in EXCEPT Influenza
nucleus Pox virus cytoplasm and Retrovirus
(nucleus)
Linear DNA EXCEPT
Hepadnavirus,
Papilloma virus,
Polyoma virus
NAME CLASSIFICATION TRANSMISSION CLINICAL TREATMENT
Hepatitis A *Picornaviridae Fecal-oral Acute viral hepatitis -pooled immune
serum globulin
-supportive care
new HAV vaccine
Hepatitis B *Hepadna viridae -BT -Acute viral hepatitis -Hepa B vaccine
-needle sticks -Fulminant hepatitis -alpha-interferon
-sexual -Chronic hepatitis -Lamivudine
-placental Complications:
-hepatocellular CA
-cirrhosis
Hepatitis C *probably a Flavivirus Acute viral hepatitis -alpha interferon
-Ribavirin

Hepatitis D -incomplete RNA virus -coinfection w/ HBV Control of HBV

-superinfection infection
Complication
-fulminant hepatitis
-cirrhosis
Hepatitis E *probably a Calici virus Fecal-oral Hepatitis (like A)
Hepatitis G Flavivirus -transfusion Has not been
-needle sticks conclusively shown to
cause liver disease
Coronaviridae
• Corona virus –
common colds;
SARS
Retroviridae
• Human
immunodefficiency
virus (HIV) - AIDS
NEW VIRUSES
 OTHERS:
ADENOVIRIDAE- childhood URTI

OTHER VECTOR-BORNE VIRAL DISEASE:

REOVIRIDAE- COLORADO TICK FEVER CAUSED
BY COLTIVIRUS
TOGAVIRIDAE- CHIKUNGUNYA
BUNYAVIRIDAE- SANDFLY FEVER CAUSED BY
PHLEBOVIRUS
 Immunotherapy
Active Immunization

Live Virus Killed virus Virion subunit Viral polypeptides DNA Vaccines
Vaccines vaccines vaccines

Measles Polio (Salk) Adenovirus HBV HIV

Mumps Rabies Influenza
Rubella Influenza
Chicken pox HAV
Polio (Sabin)
Yellow fever
Passive Immunization

HAV, HBV
Measles
Rabies
RSV
VZV
Relapsing Fever Body Louse

Leishmaniasis Phlebotomus/sand fly

American Sleeping Reduviid bug/Kissing Bug

sickness
African Sleeping Sickness Tsetse fly

Babesiosis
Ixodes Tick
O. Vulvulus Black flies/simulium fly

Lyme Disease Small Ticks

 FEVER
5- day fever/ Trench Fever Bartonella quintana
Rabbit Fever Fracinsella tularensis
Q-Fever Coxiella burnetti
Cat-scratch Fever Bartonella henselei
Relapsing Fever Borellia recurrentis
Scarlet Fever Streptococcus pyogenes
Typhoid Fever Salmonella typhi
Undulant/Malta Fever Brucella mellitensis
 VIRAL VACCINES
LIVE ATTENUATED KILLED
Stimulate both humoral and Only initiate humoral
cell-mediated immunity
Relatively dangerous to
administer
E.g. small pox, yellow E.g. rabies, influenza, salk
fever, chicken pox, sabin, polio, HAV
polio, MMR (measles,
mumps, rubella)
PUBLIC HEALTH
 SPECIFIC IMMUNITY
• Aka adaptive immunity
• (2) types:
1. Natural
• Naturally acquired active immunity – host produces its
own antibody after recovery from a disease
• Naturally acquired passive immunity – mother to fetus
(maternal)
2. Artificial (by vaccination)
• Artificially acquired passive immunity - an immediate,
but short-term immunization provided by the injection of
antibodies, such as gamma globulin, that are not
produced by the recipient’s cells.
• Artificially acquired active immunity - microbe, or parts
of it, are injected into the person before they are able to
take it in naturally. If whole microbes are used, they are
pre-treated, attenuated vaccines.
QUALITY CONTROL AND
ASSURANCE
SX SIZE
ULTRAMICROANALYSIS <1mg
MICROANALYSIS 1-10mg
SEMI-MICROANALYSIS 10-100mg
MACROANALYSIS 100mg-1000mg

EXTENT OF DETERMINATION

PROXIMATE TOTALITY/GROUP
ULTIMATE SPECIFIC CONSTITUENT/ SINGLE
CHEM.SP.
 PACOP QUESTION
The type of alkaliodal assay where the total alkaloid is
determined.
A. Ultimate
B. Specific
C. Proximate
D. Extraction
ACIDIMETRY
Analyte: Free Base Aqueous Non - aqueous
Titrant: Acid

VS Hydrochloric acid Perchloric acid in

glacial acetic acid

Sulfuric acid Perchloric acid, HBr

Primary Standard Anhyd. Sodium carbonate Potassium biphthalate

Secondary Standard Sodium/Potassium hydroxide --------------------

Examples:
• Direct NaOH, NaHCO3, Na salicylate, Methacholine Cl
Caffeine, NaCO3

• Residual NH4Cl, Methenamine,

ZnO, Potassium sodium
tartrate, tartrate
ALKALIMETRY
Analyte: acid Aqueous Non – aqueous
Titrant: free base
VS Sodium hydroxide Sodium methoxide in
ethanol or Toluene,

Potassium hydroxide Li Methoxide

Primary Standard Potassium biphthalate Benzoic acid

Secondary Standard Hydrochloric acid in Sulfuric acid

Examples:
• Direct Phosphoric acid, Hydrochloric Phenytoin
acid, Tartaric acid

• Residual Aspirin
W/ R agent= RED
W/ O agent= GREEN/BLUE
DIAZOTIZATION
• NOT IODOMETRIC METHOD:
• assay for sulfonamide drugs and other compounds
containing Arylamines
• VS: 0.1M 0.1 M sodium nitrite
• Refernce std: K2Cr2O7
• Endpoint: blue
PACOP Q:
Which of the following volumetric solutions is used in
diazotization analysis of sulfa drugs?
0.1 M sodium nitrite
COMPLEXIOMETRY
PRECIPITATION METHOD
METHOD Manifestation VS Indicators 1 std Example

formation of
VOLHARD insoluble Ferric
Ammonium silver Aminophylline,
(residual) colored Ammonium
complex Thiocyanate nitrate NaCl
Sulfate
(colored ppt)

formation of Potassium
MOHR 2nd Silver Nitrate NaCl SLS
precipitate chromate
PRECIPITATION METHOD
Manifestati
METHOD VS Indicators 1 std Example
on
GAY- cessation of Silver NaCl NaCl
LUSAAC ppt. Nitrate
Sodium
appearance K Organic N
LEIBIG Teraphenyl Bromophenol
of turbidity biphthalate compounds
boron
Adsorption Phenylephrine
Change in Silver indicator HCl
FAJANS NaCl
ppt Nitrate Ex. DCF, Tubocurarine
Eosin Y, TEE HCl
APPROXIMATE TEMPERATURE EQUIVALENTS

Very Dull Red Heat 500C-550C

Dull Red Heat 550C-700C

Bright Red Heat 800C- 1000C

Yellow Red Heat 1000C-1200C

White Heat 1200C- 1600C

 BIOLOGICAL METHODS
DIGITALIS PIGEON
HCG FEMALE RATS
INSULIN RABBIT(HPLC)
HEPARIN SHEEP BLOOD
GLUCAGON CAT PLASMA

OXYTOCIN CHICKEN
PROTAMINE SO4 SHEEP BLOOD
VASOPRESSIN MALE RAT PLASMA
COD LIVER OIL RACHITIC RAT
CORTICOTROPIN RAT
PARATHYROID DOGS
TUBOCURARINE INJ. RABBIT(HPLC) HORMONE

METOCURARINE INJ. RABBIT (HPLC)

 BIOLOGICAL REACTIVITY TESTS
(IN VIVO)
TEST ANIMAL USED NOTES
Systemic Albino mice for elastomeric material
Injection Test esp. plastics
Intracutaneous Albino rabbits for elastomeric material
Test esp. plastics
Eye Irritation Test Albino rabbits for medical devices &
ophthalmic containers

Implantation Test Rabbits for material in direct

contact w/ tissue
 LIMIT TEST
As DEC
Cl AgNO3
SO4 BaCl2
HEAVY H2S
METALS
Fe NH4SCN
Pb AMMONIA CYANIDE,
AMMONIUM CITRATE,
HYDROXYLAMINE, HCl,
DITHIZONE
 ASSAY METHODS
SPECTROPHOTOMETRY Vitamin A
Vitamin K (635 nm)
Vitamin B12 (361nm)
Vitamin B3 (450 nm)
Steroids (525 nm)
HPLC VIT. D, B6, B9, PEN, CEPHA

GC BARBITURATES, VIT.E
FLUOROMETRY VIT. B1,B2
L.plantarum VIT B3, B5, B12
RABBIT BLOOD INSULIN ACTIVITY
SUGAR METHOD