Search This Blog

CLINICAL BLOOD HAEMATOLOGY - COMPLETE BLOOD COUNT/CBC

 COMPLETE BLOOD COUNT (CBC) or FULL BLOOD COUNT (FBC), also known as HEMOGRAM/HAEMOLEUCOGRAM: red blood cells (RBCs), white blood cells (WBCs) and platelets (PLTs), concentration of haemoglobin and haematocrit (the volume percentage of red blood cells). HEMOGRAM/HEMOLEUCOGRAM/CBC: a basic screening test, often the first step in assessing health and diagnosing various haematological and non-haematological conditions. 

Blood cells: red blood cells, white blood cells, platelets. 

The CBC consists of measuring the following parameters:

- number of leukocytes;

- differential white blood count/leukocyte formula; 

- number of erythrocytes;

- haemoglobin;

- haemoglobin concentration: mean corpuscular haemoglobin (MCH) and mean corpuscular haemoglobin concentration (MCHC); these 2 may also be referred to as erythrocytes indices;

- haematocrit;

- erythrocytes indices: mean corpuscular volume (MCV), mean erythrocyte haemoglobin (MCH), and red cell distribution width (RDW);

- platelet count and platelet indices: average platelet volume (VTM) and platelet distribution width (PDW);

- number of reticulocytes. 



WHITE BLOOD CELLS/LEUKOCYTES

White blood cells (WBC)/leukocytes/leucocytes are the cells of the immune system involved in protecting the body against foreign invaders and infectious diseases. There are 5 categories: basophils, eosinophils, lymphocytes, monocytes and neutrophils, each of them fulfilling a specific function. All white blood cells are produced and derived from multipotent cells in the bone marrow known as hematopoietic stem cells.  

White blood cell count: number of white blood cells in the blood. Differential white blood cell: the percentage of each type of white blood cells present in the blood. 

Granulocytes: a type of white blood cells that have small granules, containing proteins. The specific types of granulocytes: neutrophils, basophils, eosinophils. Granulocytes, specifically neutrophils, help the body fight bacterial infections. 

Mast cell: mastocyte/labrocyte; a resident cell of connective tissue that contains granules rich in histamine and heparin. Mast cell: a type of granulocyte derived from the myeloid stem cell; a part of the immune and neuroimmune systems. 

BASOPHILS

Basophils and mast cells are important factors in allergic inflammation and other immune and inflammatory phenomena. These express on their surface an isoform of the receptor with high IgE affinity (when bound to the sensitising allergen or anti-IgE antibodies, both basophils and mast cell are activated, inducing mediators synthesis and secretion). Hence, basophils and mast cells are important factors in allergic inflammation and other immune and inflammatory phenomena.

 Basophils are the largest granulocytes, much larger than the eosinophils or the neutrophils. Basophils are structurally similar to the mast cells, but generally speaking, basophils have fewer granules than the mast cells and have a more homogeneous morphology than the mast cells. 

Basophils occur in most inflammatory reactions, especially those involving allergies. Heparin, contained in basophils, is an anticoagulant that prevents blood from clotting too quickly. 


EOSINOPHILS

The nucleus is usually bilobed, but 3 or more lobes are often seen. They are in small number in healthy individuals, but become predominant in the blood and tissues in association with various allergic diseases (asthma), parasites or malignancies. Eosinophils contain at least 5 different types of intracytoplasmic granulations. Allergen/parasite-induced eosinophilia: dependent on the T-cells; mediated by cytokines released by the sensitized lymphocytes. 

The "eosinophils" name comes from the fact that these cells have an affinity for acid dyes (eosin), which gives them a specific red-brick coloration. They contain small cytoplasmic granules, which contain many active substances, such as histamine, ribonuclease and eosinophil peroxidase. 


NEUTROPHILS

- play a major role in the body's primary anti-infective defence by phagocytizing and digesting microorganisms. Their improper activation may lead to damage to the body's normal tissues by releasing enzymes and pyogenic agents. 

Upon infection, chemotactic agents are produced, which cause migration of neutrophils to the site of infection. The defensive functions of neutrophils are activated, with phagocytosis of the agent, followed by the release of granules into the phagocytosis vesicle and destruction of the infectious agent. 

Immature forms of neutrophils: bands cells; non-segmented polymorphonuclear neutrophils.

Mature forms of neutrophils: segmented neutrophils; polymorphonuclear neutrophils.


 

LYMPHOCYTES

Although some morphological characteristics (size, granularity, nucleolar-cytoplasmic ratio) differentiate lymphocyte populations from each other, they do not provide indications of their type and function. 

Lymphocytes:

- 65-80%: T cells (maturing in the thymus, where they migrate from the medullary level);

- 8-15%: B cells (maturing in the bone marrow);

- 10%: natural killer cells (some of them are identical with the large granular lymphocytes).

Plasma cells are completely differentiated B cells and are not normally present in the blood. Intermediate cells (lymphoplasmocytes): common in viral infections or in immunological diseases with hypergammaglobulinemia. 

B cells control the humoral immune response mediated by antibodies specific to the offensive antigen. Memory B cells: long lifespan; do not produce antibodies until antigenic restimulation, when they respond to much lower doses of antigen, proliferate clonally and produce 7-10 times more antibodies than unexposed B cells. 

T cells: involved in the cell-mediated immune response; include CD4+ helper T cells, CD8+ suppressor T cells and cytotoxic T cells. T-cells circulate until they encounter specific antigens; a critical part in immunity against foreign substances. 

NK cells are effector lymphocytes of the innate immune system that control several types of tumors and microbial infections.  


 MONOCYTES

- the largest cells in the blood; are part of the mononuclear/reticuloendothelial phagocytic system (composed of: monocytes, macrophages and their medullary precursors).

Monocytes and macrophages produce numerous bioactive factors: enzymes, complement factors, coagulation factors, reactive oxygen and nitrogen species, angiogenetic factors, binding proteins, bioactive lipids, chemotactic factors, cytokines and growth factors. 

Monocytes: a type of phagocytic leukocytes (agranulocytes) that are part of the innate immune system of vertebrates (including humans). The precursors of monocytes: monoblasts, which originate in the bone marrow; they initially evolve into pro-monocytes, then into monocytes. 

Monocytes: part of the monocyte-phagocytic system. Once migrated into tissues, monocytes can be differentiated into: 

- macrophages (tissues - spleen, alveoli etc);

- histiocyte - connective tissues;

- microglia - CNS;

- osteoclasts - bones;

- Kupfer cells - liver;

- dendritic cells - Langerhans cells (skin), digestive tract, lungs etc.

 Macrophages are a type of white blood cells that engulfs and digests anything that does not have, on its surface, proteins that are specific to healthy body cells (phagocytosis).  



DIFFERENTIAL WHITE BLOOD CELL COUNT/LEUKOCYTE FORMULA

The leukocyte formula - a blood test that assesses the number of the 5 types of leukocytes, expressed as a percentage and in absolute numbers. The leukocyte formula is used in diagnosing the specific cause of some diseases. 

Normal leukocyte values, in percentage:

- neutrophils: 60-70%

- basophils: 0-1%

- eosinophils: 1-4%

- monocytes: 4-8%

- lymphocytes: 25-30%

Leukocytes: also divided into 2 main groups, according to the presence/absence of granulations in the cytoplasm:

- granulocytes: neutrophils, eosinophils and basophils; also called polymorphonuclears (multilobed nucleus);

- non-granulocytes: lymphocytes and monocytes; no distinct cytoplasmic granules and have non-lobulate nucleus; also called mononuclear leukocytes. 


RED BLOOD CELLS/ERYTHROCYTES

- the most numerous cells in the blood, anucleate upon maturity and necessary for tissue respiration. Main function: transporting oxygen from the lungs to the tissues and carbon dioxide from the tissues to the lungs. Erythrocytes increase and decrease along with the haemoglobin and the haematocrit. Shape: round with narrow centers resembling a donut without a hole in the middle. 

- red blood cells formation: in the red bone marrow of bones. Stem cells in the red bone marrow: hemocytoblasts and give rise to all of the formed elements in the blood. If a cell commits to becoming a cell called a proerythroblast, it will develop into a new red blood cell. 


HAEMOGLOBIN   

- the protein molecule in red blood cells that carries oxygen from the lungs to the body's tissues and returns carbon dioxide from the tissues back to the lungs;

- 4 protein molecules (globulin chains) connected together, make up the haemoglobin molecule. Normal adult haemoglobin: 2 alpha-globulin chains and 2 beta-globulin chains. Foetuses and infants: 2 alpha chains and 2 gamma chains (gamma-chains gradually replaced by beta-chains upon growth);

- each globulin chain: iron-containing porphyrin compound termed heme. Embedded within the heme: an iron atom (vital for transporting oxygen and carbon dioxide; gives the red colour of the blood). 

- haemoglobin also gives the shape of the red blood cells; abnormal haemoglobin structure can disrupt the shape of the red blood cells and implicitly their function.


HAEMOGLOBIN CONCENTRATION 

- MEAN CORPUSCULAR HAEMOGLOBIN/MCH - average quantity of haemoglobin in a single red blood cell;

MCH (pg) = [ Haemoglobin (g/dL) / RBC (mil/uL) ] x 10

- MEAN CORPUSCULAR HAEMOGLOBIN CONCENTRATION/MCHC - concentration of haemoglobin in a certain amount of blood;

MCHC (g/dL) = [ Haemoglobin (g/dL) / HCT (%) ] x 100%


HAEMATOCRIT  

- HCT: volume of red blood cells relative to the volume of blood, expressed as a percentage; example: HCT 25%: 25 ml of red blood cells in 100 ml of blood. 

HCT value is used to determine erythrocyte indices: mean erythrocyte volume, mean corpuscular haemoglobin concentration, mean corpuscular haemoglobin. All of these together are useful for the differential diagnosis of various types of anaemia. 

- also called Packed Cell Volume/PCT


ERYTHROCYTES INDICES 

- MEAN CORPUSCULAR VOLUME (MCV)/MEAN CELL VOLUME/MEAN ERYTHROCYTE VOLUME: average size of red blood cells in a blood sample. MCV represents the volume occupied by a single erythrocyte.  It is a useful index for classifying anaemias and depends on plasma osmolarity and the number of erythrocyte divisions.  

MCV (fL) = [ Hct (%) / RBC (mil/uL) ] x 10

- MEAN ERYTHROCYTE HAEMOGLOBIN/HEM/MCH - see "Haemoglobin concentration";

- WIDTH OF ERYTHROCYTES DISTRIBUTION/RDW - a measurement of the range in the volume and size of the red blood cells (difference in size between the smallest and largest red blood cells in a sample); differentiates between different types of anaemia. 


PLATELETS

Platelets form blood clots to slow down blood loss, to prevent infection and to promote healing. When an injury occurs, platelets aggregate to plug the wound and send hormone signals through the blood to attract protein clotting factors, which assists in repairing the injury. 

Platelets are small, anucleate cells, produced in the bone marrow from the fragmentation of megakaryocytes (are actually pieces of them). Platelets have a role in haemostasis (participating in thrombi formation), as well as a source of growth factors.

- PLATELET COUNT measures the total number of platelets in the blood. 

- MEAN PLATELET VOLUME (MPV) - measure of the average size of the platelets/thrombocytes. The MPV indicates the uniformity of platelet population size. 

- PLATELET DISTRIBUTION WIDTH (PDW) - reflects how uniform the platelets are in size. PDW is a measurement of the variability in platelet size distribution in the blood. A normal PDW indicates platelets that are mostly the same size, while a high PDW means that platelet size varies greatly, a clue that there is platelet activation and has been associated with vascular diseases and certain cancers. 


RETICULOCYTES

- newly produced, non-nucleated, relatively immature red blood cells, that contain residual nucleic acids (RNA); a reticulocyte count (number/percentage of reticulocytes in the blood) - a reflection of recent bone marrow function/activity.

Blood-forming (hematopoietic) stem cells differentiate and develop, eventually forming reticulocytes and finally becoming mature red blood cells. Reticulocytes are visually slightly larger than mature red blood cells. Unlike most other cells in the body, red blood cells have no nucleus, but reticulocytes still have some remnant genetic material (RNA). As reticulocytes mature, they lose the last residual RNA and most are fully developed within one day of being released from the bone marrow into the blood. The reticulocyte count shows the bone marrow's ability to produce red blood cells. 


SUMMARY IMAGE OF BLOOD CELLS:



CLINICAL BLOOD HAEMATOLOGY - ERYTHROCYTE SEDIMENTATION RATE (ESR) - VSH

 ESR: the rate of red blood cell/erythrocytes sedimentation/aggregation. The new version of this examination is the "VSH": the Velocity of the Sedimentation of the red blood cells/Haematias. 


Erythrocyte sedimentation is the aggregation of red blood cells in the form of a column. The rate of erythrocyte sedimentation (Erythrocyte sedimentation rate ESR): the measuring of red blood cell aggregation. 




In certain conditions (that cause the growth of acute phase proteins or immunoglobulins), plasma proteins attach to the red blood cell surface and reduce the surface potential, causing red blood cell aggregation and increased sedimentation. 

ESR: the rate at which red blood cells aggregate/settle in an anticoagulated blood sample within an hour. 

Normal ESR range: 0-22mm/h for men and 0-29mm/h for women.

 



CLINICAL BLOOD HAEMOSTASIS - COAGULATION

COAGULATION: FIBRINOGEN, PT, BT, TT, APTT.

FIBRINOGEN

Fibrinogen/Coagulation Factor I is a plasma glycoprotein produced by the liver. Fibrinogen is the action substrate for both thrombin (final coagulation enzyme) and plasmin (fibrinolytic system enzyme). Fibrinogen belongs to the acute phase proteins (appearing at 24-48 hours post-event). 

Defective fibrinogen levels show liver disfunction. Fibrinogen levels help assessing the body's ability to form a blood clot.  

While coagulation approaches its end, soluble fibrinogen is transformed into insoluble fibrinogen fibres. Fibrinogen fibres protect the wound by intercalating; along with the platelets, they form a clotted blood barrier that blocks bleeding until healing. 

Fibrinogen analysis measures the soluble amount of Factor I (fibrinogen dissolved in the blood), before its conversion into insoluble fibrin.

Fibrinogen normal levels: 200-400 mg/dL (according to the laboratory reference values - reactive used etc.).
 
 
PROTHROMBINE TIME (PT; INR): 

Prothrombin time (PT) evaluates the activity of factors involved in the extrinsic and common coagulation pathway. Evaluates both the activity of vitamin K-dependent coagulation factors (except F IX), factor V and fibrinogen, as well as the function of liver protein synthesis, with diagnostic and therapeutic implications. 

In vitro, main route of blood coagulation initiation: the extrinsic system. This includes blood components and vascular elements, the initiation of coagulation occurring when the tissue factor (FT) is bound to F VIIa. 
The F VIIa - FT enzyme complex activates both F IX and F X. 
F Xa interacts with its cofactor F Va, forming the prothrombinase complex; the complex: enough to generate few thrombin amounts next to FT-expressing cells. 

In vitro, PT detects clot formation. This represents fibrin polymerization, resulting from thrombin action.
Being absent in normal plasma, the tissue factor (FT) must be provided from an external source. This is why the cascade of enzymatic reactions triggered in PT is known as the "extrinsic pathway".  

Prolonged PT indicates deficiency of coagulation factors (I, II, V, VII, X) or an inhibitor's presence. 

PT is the most commonly used test monitoring oral anticoagulant therapy. Results may be expressed as follows:
- as clotting time - in seconds; PT - time (s); normal: 11-13.5 s;
- as a percentage (%) of normal prothrombin activity; measurable range: 12.5-120%; AP - volume fraction (%)
- as prothrombin ratio (PR = PT patient in seconds / normal plasma PT in seconds); PR - prothrombin time ratio (1)
- as INR - International Normalized Ratio - (1); normal: 0.8-1.1.
INR = (PT patient / PT normal plasma) x ISI; ISI: international sensitivity index of used thromboplastin, calculated in report to reference thromboplastin for which ISI = 1. 

INR = 2.0-3.0 target of oral anticoagulant treatment for most clinical cases;
INR = 2.5-3.5 target in anticoagulant treatment for: recurrent deep vein thrombosis, recurrent systemic embolism, cardiac stent, mechanical heart valves.


BLEEDING TIME (BT/CT)

A test that investigates primary haemostasis, thus being an indicator of the vascular and platelet phases efficiencies. 

BT depends on the:
- platelets (function and number)
- adhesion plasma proteins
- vascular wall matrix integrity
Critical values:  >15 minutes. 

Bleeding Time is increased when platelet levels are low or when platelets are qualitatively abnormal. 


THROMBIN TIME (TT)

TT measures the time of fibrin formation under the action of thrombin, and its aggregation to form insoluble clot - the final coagulation step. Assesses the activity of fibrinogen.

Under thrombin action on fibrinogen, fibrinopeptides A and B are released. Upon their cleavage, fibrin monomers form soluble aggregates. 
Due to the action of Factor XIII (activated by thrombin), along with that of Ca ions, a transverse polymerization of fibrin monomers takes place, with the formation of insoluble fibrin. 


Thrombin Time mainly reflects the function and interaction between the exogenous thrombin and the endogenous fibrinogen. 

TT measurable range: 13-240 seconds. Thrombin clotting time is generally <22s, between 14-16s (every lab sets its own reference values, according to the reagent kits used; other labs may have: 7,0-12,0s). Critical values: >60s. 


ACTIVATED PARTIAL THROMBOPLASTINE TIME (APTT)

Also called Partial Thromboplastin Time (PTT), is a functional test evaluating both "intrinsic" and "common" coagulation pathways. 
In vitro, the contact system is involved when blood interacts with a foreign surface, such as the cardiopulmonary bypass. 

Kallikrein cleaves HK, releasing bradykinin and kinin-free kininogen (activated HK). 
In vitro, activation of HK is achieved by adding:
- "partial thromboplastin"; composed only of phospholipids; no protein, no TF;
- silica - a surface activator; provides the negatively charged surface.
Than, calcium chloride is added and time expressed in seconds is measured until the clot's formation. 
The so-called "partial thromboplastin" does not contain the tissue factor (protein) needed to initiate coagulation in PT, as in "complete thromboplastin" stage. The silica provides the negatively-charged surface needed for coagulation contact pathway activation. 

The cascade of reactions triggered in PTT were called as "intrinsic pathway" based on the misconception that coagulation is initiated without the addition of any external factors. The external factor involved in initiating coagulation in PTT: the negatively-charged glass surface of the reaction tube; this can be potentiated by adding silica, kaolin, ellagic acid, in what we now refer to as the "activated" partial thromboplastin test.  

Deficiency/inhibition of HK, prekalikrein or Factors XII, XI, IX and VIII causes prolongation of aPTT with normal PT, while deficiencies of "common pathway" coagulation factors (X, V, II, Fibrinogen) can prolong both aPTT and PT. aPTT is not influenced by FVII or FXIII deficiencies. 

Normal aPTT: 21-35s (therapeutic: 2,0-2,5x normal)



COAGULATION: 


"INTRINSIC PATHWAY": prekalikrein, high-molecular weight kininogen-HK, Factors XII, XI, IX and VIII;
"COMMON PATHWAY": Factors X, V, II and I. 


CLINICAL BLOOD HAEMOSTASIS - BLOOD TYPE IDENTIFICATION

 BLOOD TYPE IDENTIFICATION: Based on characterization of an individual's blood according to the presence/absence of an antigen on the surface of his erythrocytes. Most blood group antigens are glycoprotein in nature and genetically stable. 

The antigen-antibody reaction in which the blood group antigens and their specific antibodies participate is one of agglutination (of red blood cells). Hence, the antigens are also called agglutinogens and the antibodies also agglutinins.  

In current medical practice, the ABO and Rh systems are important. 

The main blood groups: 

A

B

AB

0.

Rh factor (Rhesus) is a group of antigens that may be present or absent on the surface of erythrocytes. Most people have Rh factor present on erythrocytes, so they are called Rh positive. Rh negative: do not have the Rh factor present. 

Incompatibility of Rh positive blood with Rh negative blood is an important cause of transfusion reactions and haemolytic diseases in newborns. 


                                BLOOD TYPE COMPATIBILITY RESUME

                                            +   : Rh present;     -   : Rh absent



MICROORGANISMS IDENTIFICATION - culture growth etc

Microscopy: particularly useful for bacteria identification.

Blood analysis (serology, blood culture etc): identification of microorganisms in blood.

Pharyngeal/nasal exudate, sputum, ocular/otic secretions: collection of biological samples for microbiological examination. 

Urinalysis: first morning urine, from the intermediate jet, after local toilet. Important tool for microorganism presence identification.

Coproparasitological examination, coproculture, from faeces.

Vaginal, urethral, vulvo-vaginal secretion examinations (exudates etc)

Various smears with specific colourings - microscopical examinations of morphology.  

Various molecular genetics techniques for virus identification: RT-PCR, ELISA, Western-Blot, immunological techniques (immunochromatography, immunodifusion, immunofluorescence) etc. 



LABORATORY DIAGNOSIS FOR VIRUSES:


- virus isolation on cell cultures

- viral genome highlighting by gene amplification reaction after reverse transcription (Real-Time Polymerase Chain Reaction - RT-PCR) - frequently used due to high precision

- serological identification - immunochromatography techniques (rapid tests), ELISA, Western Blot etc.


CULTIVATION OF VIRUSES


Viruses have obligatory intracellular parasitism, hence they do not grow on artificial environment/mediums. They can only be cultivated on live substrates:

- laboratory animals - limited use nowadays due to spreading of cell culture usage popularity;

- embryonated chicken egg - offer embryonic tissue for viruses culturing; important in vaccine preparation;

- cell cultures - the mostly used virus-host system in virology research. 

Example: Poliomyelitis

                 - indicated collection of pharyngeal exudate, blood, faeces, cerebrospinal fluid;

                 - isolation of virus on cell cultures; RT-PCR.



CULTIVATION OF BACTERIA ON MEDIA CULTURE:


Microscopical examination: first step to bacteria identification. Main elements to establish bacteria identity: 

- cilia

- capsule

- endospores. 


Morphological examination: on microscopic preparations, fixed and coloured: smears

Smears: obtained by spreading a bacterial colony on a clean and degreased blade, which will be dried, fixed and coloured, for examination under a microscope. 


Culture media - provides nutrients and physico-chemical conditions for bacteria growth;

Sowing - contact of pathological product with the culture medium;

Isolation - single colony transition to other culture medium => pure culture.


MEDIUMS: 


Physical classification:

- solid - reflecting colour, diameter, appearance, media adherence etc;

- liquid - turbidity and turbidity type, surface/bottom film/deposit formation.  

Examples: homogeneous turbidity - S. aureus; surface film - Vibrio cholerae, bottom deposit - Streptococcus pyogenes. 


In terms of composition, media can be:

- simple - simple agar;

- composed - besides the basics, also organic substances (serum, blood) required for special growth care; example: blood agar;

- special mediums - isolation, enrichment, differential. 



BACTERIA CLASSIFICATION:


1. Tinctorial affinity to Gram staining (most important staining, dividing bacteria in Gram-positives - lilac and Gram-negatives - red): 

a) Gram-positives: Streptococcus pneumoniae, Staphylococcus aureus;

b) Gram-negatives: Neisseria gonorrhoeae and meningitidis, Escherichia coli, Helicobacter pylori, Haemophilus influenzae. The Enterobacteriacee family - large number of Gram-negative bacillus species living in the intestines; the most important genre it includes are: Escherichia, Shigella, Salmonella, Klebsiella, Proteus. Enterobacteria classification according to the fermentation capacity of lactose: 

- Pathogenic Enterobacteria: lactose-negative: Salmonella, Shigella;

- Conditioned-pathogenic or non-pathogenic enterobacteria: 

                            a) lactose-positive: Escherichia, Klebsiella

                            b) lactose-negative: Proteus.


2. Need for oxygen:

a) aerobic bacteria: Staphylococcus aureus;

b) anaerobic bacteria: Clostridium botulinum.


3. Glucose fermentation capacity:

a) glucose-fermentative bacteria: Vibrio cholerae;

b) glucose-nonfermentative bacteria: Pseudomonas aeruginosa. 


4. Pathogenicity classification: 

a) nonpathogen bacteria: suboptimal conditions inside human host

b) pathogenic bacteria: always causing illness; example: Treponema pallidum. 

c) conditionate pathogenic bacteria: from normal flora, causing illness in various conditions.  


EXAMPLES OF AGARS AND COLORATIONS FREQUENTLY USED IN MICROBIOLOGY FOR BACTERIA IDENTIFICATION:

- MacConkey Agar: selective and differential culture medium for bacteria. Isolates Gram-negative and enteric bacteria. Differentiates them based on lactose fermentation. Lactose fermenters turn red or pink, non-fermenters do not change colour.

- Kligler's Iron Agar: Enterobacteriaceae identification, based on double sugar fermentation and hydrogen sulphide production.

- Ziehl-Neelsen stain: acid-fast stain identifying acid-fast organisms, mainly Mycobacteria (Nocardia as well). Acid-fast bacilli are bright red after staining. 

- Voges-Proskauer test: detects acetoin in a bacterial broth culture. Red: positive, yellow-brown: negative. Principle: digestion of glucose to acetylmethylcarbinol. 

- BD CLED Agar: CLED: Cystine-Lactose-Electrolyte-Deficient; differential culture medium for bacteria in urine; supports pathogen growth but prevents Proteus species accumulation due to the lack of electrolytes. 



DIAGNOSIS OF PARASITES:


- Coproparasitology: - faeces examination for parasites;

- ELISA;

- Direct agglutinations to detect IgM antibodies;

- Blood culture; cerebrospinal fluid examination for parasites;

- Smear and thick drop examination of peripheral blood; May-Grumwald-Giemsa staining;

- Immunological examination: ELISA, hemagglutination; latex agglutinations

- Parasites/parasite eggs identification in faeces and other material through microscopical examination.

- Macroscopical examination of faeces for parasite and/or parasite eggs elimination;



MICOLOGICAL DIAGNOSIS:


- based on the clinical lesion's aspect and microscopic analysis results of hairs and squamous material collected from lesions. 



ANTIBIOGRAM - drug choosing

 An antibiogram is an overall profile of antimicrobial susceptibility testing results of a specific microorganisms to a battery of antimicrobial drugs. 

It shows susceptibility of the microorganism investigated to particular antimicrobial drugs. Obtained results classified as: resistant, intermediate or sensitive to the action of a certain antibiotic. 


White disks: the antimicrobial drugs. Unaffected area around the white disks: growth of the microorganism i.e. the drug does not kill the microbe. Affected area around the disks (radii of inhibited areas) gets bigger (no organism growth) with as much as the drug can affect the microbe's growth. In clinics, this affected area is measured (to be taken into consideration for treatment choosing by the clinican), along with the name of each drug with annotations such as "microbe not resistant to drug X - diameter of inhibition zone measured: MIC (Y mm)".
X: name of the drug/antibiotic
Y: number measured in mm


MICROBIOLOGY - fast kits

VARIOUS RAPID MICROBIOLOGICAL EXAMINATIONS

    - include several rapid test kits with multimicrobial identification;

    - can be performed on blood, serum, urine or faecal material.

Example: 


MOST RECENT HEALTH NEWS!

Current news about health from all around the world


THE CLINICAL ANALYSIS LABORATORY - BASICS

BASICAL CLINICAL ANALYSIS LABORATORY


CLINICAL SERUM BIOCHEMISTRY


GLYCATED HEMOGLOBIN (BLOOD)


PROTEIN ELECTROFORESIS (SERUM)


URIANALYSIS (1st in the morning/8h URINE):

                     - BIOCHEMISTRY  OF URINE

                     - BASIC MICROSCOPY OF URINE (SEDIMENTATION)


CLINICAL IMMUNOLOGY (BLOOD AND FAECAL SAMPLES)

                    - AUTOMATIC

                    - RAPID MANUAL TESTS


IONOMETRY (SERUM/URINE): 

                            - SODIUM

                            - POTASSIUM

                            - OTHERS


CLINICAL BLOOD HAEMATOLOGY AND HAEMOSTASIS

            - HAEMATOLOGY

                    - COMPLETE BLOOD COUNT (CBC): RBCs, WBCs, PLTs

                    - BLOOD SEDIMENTATION (ESR-ERYTHROCYTE SEDIMENTATION RATE)

            - HAEMOSTASIS:

                    COAGULATION (PLASMA): FIBRINOGEN; PT, INR; CT/BT; TT; APTT                

                    - BLOOD TYPE IDENTIFICATION: A, B, O; Rh Factor


MICROBIOLOGY (SERUM/BLOOD, URINE, FAECAL, CEREBROSPINAL FLUID ETC): 

                     - MICROORGANISMS IDENTIFICATION

                                                             - CULTURE GROWTH

                                                             - ANTIBIOGRAM

                                                             - VARIOUS RAPID MICROBIOLOGICAL EXAMINATIONS



Coronavirus - 30 June 2020

Globally, as of 4:03pm CEST, 30 June 2020, there have been 10,185,374 confirmed cases of COVID-19, including 503,862 deaths, reported to WHO.

A first step for Coronavirus vaccin!

An achievement from outside China came through, which could help combat the global spreading of the disease, according to Reuters. 

Researchers at the Peter Doherty Institute of Infections and Immunity in Melbourne, Australia, who development a potential step towards the vaccine, announced they will make the sample available in the WHO and laboratories worldwide.  

The sample could be used to generate an antibody test, which would allow the virus to be detected in patients without symptoms, as well as the creation of a vaccine, as the institute said. 



Assisted Reproduction Laboratory

MALE:

Semen analysis - Spermogram

Hormonal testing - testosteron

Genetic testing - Y chromosome, SRY

Testicular biopsy - sperm analysis (and collection for IVF)

Imaging investigations - ultrasound, MRI, invasive (venography, vasography)

Other special investigations - sperm quality (DNA analysis from sperm: spermatic FISH)

FEMALE:

Complete gynecological exploracion - mammography, cytology 

Ovulation testing - LH

Permeability of the fallopian tubes and uterine cavity - (contrast sound) / hysterosalpingography

Ovarian reserve testing - available ovulation eggs

Other hormonal testing - ovulatory hormones (FSH), pituitary hormones (prolactin, GnRH, GH etc), estradiol

Imaging investigations - pelvic ultrasound

Others - hysteroscopy, laparoscopy, genetic testing

https://www.reginamaria.ro/articole-medicale/infertilitatea-diagnostic
https://www.cancer.gov/publications/dictionaries/cancer-terms/def/reproductive-system
https://www.carlsonstockart.com/photo/reproductive-system-female-illustration/



Old version of the blog

Clinical Biochemistry
Clinical Immunology
Clinical Hematology
Clinical Genetics
Clinical Microbiology and Parasitology

Bibliography

Leo Poon decoded the coronavirus

Scientist Leo Poon, a virologist at the School of Public Health at the University of Hong Kong who decoded the virus, told CNN that it probably started in an animal and spread to humans. In the early days of the virus, it seemed to only be affecting those who had touched or consumed animals sold at the market.





https://montrealgazette.com/news/local-news/coronavirus-qa-what-you-should-know
https://www.timeout.com/hong-kong/blog/hong-kong-profile-leo-poon-lit-man-020117

Detecting Coronavirus

BGI develops Real-Time Fluorescent RT-PCR kit for detecting the 2019 Novel Coronavirus!!!

Building on BGI's history of developing solutions for rapid pathogen detection, BGI has developed a real-time fluorescent RT-PCR kit for detecting the 2019 Novel Coronavirus. The kits have already been issued to many hospitals and disease control centers.

Founded in 1999, BGI is one of the world's leading life science and genomics organizations. BGI's mission is to use genomics to benefit mankind and to be a leader in the era of life sciences. BGI follows a genomics development model of "research, production and application".

https://www.bgi.com/global/company/news/bgi-develops-real-time-dna-based-kit-for-detecting-the-2019-novel-coronavirus/

CORONAVIRUS!!!


23 January 2020: Wuhan city in China has been quarantined in effort to control the deadly coronavirus outbreak. https://www.youtube.com/watch?v=mM_raeUVCCM
Transportation hubs blocked to stop people from leaving after the virus has already killed at least 17 people in China and infected nearly 600 others in at least 6 other countries. https://www.youtube.com/watch?v=IgBT-tslw3k

24 January 2020: More than 900 people are infected and at least 41 people have died. Travel bans are in effect in more than a dozen cities, impacting some 35 million people. Officials are racing to cope with the worsening outbreak by building a 1000-bed hospital this week. https://www.youtube.com/watch?v=uh6nixCRcXU
A Chicago woman who traveled through O'Hare airport was hospitalized, placed in isolation and is now in stable condition. This is one of two confirmed cases of coronavirus in the U.S. and the CDC is watching some 50 patients for symptoms in 22 states. https://www.youtube.com/watch?v=LB3DlzNSr9g
One confirmed case of coronavirus in Victoria and two probable in NSW, with the number of cases expected to grow. https://www.youtube.com/watch?v=8UVcDSDnr9E

25-26 January 2020: The Chinese president has described the accelerating spread of coronavirus as a "grave situation", after holding a government meeting on the Lunar New Year holiday. https://www.youtube.com/watch?v=5neX0YTduNQ
Leaders in China say coronavirus is starting to spread faster and well beyond the mainlandhttps://newyork.cbslocal.com/video/4420746-leaders-in-china-say-coronavirus-is-spreading-faster/
The passenger, who arrived in Baltimore from Las Vegas and had recently been to Beijing, was escorted off the plane out of an "abundance of caution" over concerns about the dangerous coronavirus, as Southwest said. There are 2 confirmed cases and more than 60 possible cases in the U.S. . https://www.youtube.com/watch?v=aSSIMHNsluY

27 January 2020: China coronavirus death toll climbs to 80 as government works for containing the outbreak. https://www.nbcnews.com/news/world/china-coronavirus-death-toll-climbs-80-government-scrambles-contain-outbreak-n1123561













https://www.elsevier.com/connect/coronavirus-information-center
https://www.tion.ro/romania/ce-este-temutul-coronavirus-si-cum-ne-putem-proteja-de-el-893897/

Systematic Parasitology

                                                                             Back to Clinical Microbiology and Parasitology page
A) PROTOZOA

1. Intestinal protozoa
I) Amoebae. Entamoeba genus: E. histolytica. Iodamoeba and Blastocystis genera
II) Flagellates. Giardia genus. G. intestinalis
III) Ciliates. Balantidium genus. B. coli
IV) Coccidia and Microsporidia: Isospora genus: I. belli
                                                  Cryptosporidium genus: C. parvum
                                                  Cyclospora genus: C. Cayetanensis
                                                  Sarcocystes genus
                                                                  Microsporidia order

2. Hematic protozoa
- Plasmodium genus: Malaria
- Babesia genus: Babesia spp.
- Leishmania genus: L. donovani, L. tropica, L. braziliensis, L. mexicana
- Trypanosoma genus: T. gambiense, T. rhodesiense, T. cruzi

3. Protozoa of tissue and other localizations
I) Tissue-based: Toxoplasma genus: T. gondii
                           Pneumocystis genus: P. carinii
II) Others: Trichomonas genus: T. vaginalis
                  Naegleria genus
                  Acanthamoeba genus

B) HELMINTHS

1. Platyhelminths: Trematodes
I) Trematodes of the lungs and digestive system. Fasciola genus. F. hepatica
II) Hematic trematodes. Schistosoma genus

2. Platyhelminths: Cestodes
I) Intestinal cestodes (parasitism by adult cestodes): Taenia genus. T. solium and T. saginata Dipylidium caninum, Hymenolepis nana, Diphyllobothrium latum
II) Tissue-based cestodes (parasitism during larval stages). Echinococcus genus. E. granulosus Hydatid cyst

3. Intestinal nematodes:
- with exclusively intestinal habitat: Enterobius vermicularis and Trichuris trichiura                              - with tissue migration of larvae: Ascaris lumbricoides, Strongyloides stercolaris, Ancylostoma duodenale and Necator americanus

4. Tissue-based nematodes
- Trichinella genus. T. spiralis. Trichinosis
- filaria that parasitize humans: Wuchereria, Brugia, Loa, Onchocerca, Mansonella genera
- filariasis: Dracunculus genus. D. medinensis

5. Arthropods of health interest: Arachnida: Acarina (mites, ticks); Araneida and Scorpionida  Insecta: Diptera; Heteroptera; Siphonaptera; Anoplura

Systematic Virology

                                                                             Back to Clinical Microbiology and Parasitology page
1. Herpes viruses
Herpesviridae family.
- Alphaherpesvirinae sub-family: Simplex Virus genus: herpes simplex virus types 1 and 22
- Varicella zoster virus
- Betaherpesvirinae subfamily: Cytomegalovirus genus: cytomegalovirus
- Gammaherpesvirinae sub-family: Lymphocryptovirus genus: Epstein-Barr virus
- Other unclassified herpes viruses: human herpes virus types 6, 7 and 8

2. Respiratory viruses
- F. Orthomyxoviridae
- Influenza virus A and B genus. Influenza virus C genus
- F. Paramyxoviridae. Paramyxovirinae sub-family. Paramyxovirus genus: parainfluenza virus
- Pneumovirinae sub-family. Pneumovirus genus: syncytial respiratory virus
- F. Adenoviridae
- Mastadenovirus genus. Adenovirus
- F. Picornaviridae
- Rhinovirus genus: Rhinovirus
- F. Coronaviridae: Coronavirus genus: Human coronavirus
- F. Reoviridae
- Orthoreovirus genus: Rheovirus

3. Enterovirus
- F. Picornaviridae
- Enterovirus genus. Poliomyelitis virus
- Enterovirus, Coxsackie virus and ECHO virus

4. Viruses which cause gastroenteritis
- F. Reoviridae
- Rotavirus genus. Rotavirus
- F. Caliciviridae. Calicivirus genus. Human calicivirus and Norwalk virus
- F. Adenoviridae: Mastadenovirus genus. Adenovirus: serotypes 40 and 41
- Others: human coronavirus and astrovirus

5. Viral hepatitis
- F. Hepadnaviridae
- Orthohepadnavirus genus. Hepatitis B virus
- F. Flaviviridae. Hepatitis C genus: Hepatitis C virus
- F. Picornaviridae
- Hepatovirus genus: Hepatitis A virus
- Other unclassified viruses that cause hepatitis: hepatitis delta virus and hepatitis E

6. Mumps virus and exanthematic viruses
- F. Paramyxoviridae: Paramyxovirinae sub-family: Paramyxovirus genus. Mumps virus
- Morbillivirus genus. Measles virus
- F. Togaviridae
- Rubivirus genus. Rubella virus
- F. Parvoviridae: Parvovirus genus: Parvovirus B19
- F. Herpesviridae. Type 6 human herpes virus

7. Rabies virus, arbovirus and other viruses which cause zoonosis
- F. Rhabdoviridae. Lyssavirus genus. Rabies virus
- Arbovirus: F. Togaviridae. Alphavirus genus. Western equine encephalitis virus
- F. Flaviviridae. Flavivirus genus. Yellow fever virus and dengue virus
- F. Bunyaviridae. Bunyavirus genus. F. Reoviridae. Orbivirus and Coltivirus genera
- Other viruses which cause zoonosis: F. Arenaviridae. Arenavirus genus. Lymphocytic choriomeningitis virus and lassa fever virus
                                                             F. Filoviridae. Filovirus genus: Marburg and Ebola viruses. F. Bunyaviridae. Hantavirus genus. Hantaan virus

8. Retrovirus
- Oncovirinae sub-family. Group HTLVBLV genus. Human T-cell lymphotropic virus types 1 and 2
- Lentivirinae sub-family. Lentivirus genus. Human immune deficiency virus types 1 and 2

9. Oncogene DNA viruses
- F. Papovaviridae. Papillomavirus genus. Human papilloma virus. Epstein-Barr virus. Hepatitis B and C viruses

10. Non-conventional agents: Viroids and prions





Systematic Bacteriology

                                                                            Back to Clinical Microbiology and Parasitology page
1. Staphylococcus genus
- S. aureus
- S. epidermidis
- S. saprophyticus

2. Streptococcus genus and Enterococcus genus
- Streptococcus pyogens (S. pyogenes and S. agalactiae) and streptococci of the viridans group
- S. pneumoniae
- Enterococcus genus

3. Clostridium genus and Bacillus genus
- C. tetani
- C. botulinum
- C. perfringens
- C. difficile
- Bacillus genus: B. anthracis and B. cereus

4. Corynebacterium genus and Listeria genus
- C. diphteriae
- L. monocytogenes

5. Neisseria genus
- N. gonorrhoeae
- N. meningitidis

6. Enterobacteriaceae family
- Salmonella genus
- Shigella genus
- Escherichia genus: E. coli
- Yersinia genus: Y. enterocolítica and Y. pseudotuberculosis, Y. pestis

7. Vibrio genus, Campylobacter genus and Helicobacter genus
- V. cholerae, V. parahaemolyticus
- C. jejuni
- H. pylori

8. Haemophilus genus and Bordetella genus
- H. influenzae
- B. pertussis

9. Legionella genus
- L. pneumophila

10. Brucella genus and Francisella
- Brucella genus: Brucellosis
- Francisella genus: F. tularensis

11. Strict gram-negative aerobic bacteria and opportunistic anaerobic bacteria
- Pseudomonas genus
- opportunistic enterobacteria
- Moraxella genera
- Acinetobacter
- Aeromonas
- Plesiomonas

12. Strict opportunistic anaerobic bacteria
- Gram-negative bacteria: Veillonella, Bacteroides, Leptotrichia and Fusobacterium genera.
- Gram-positive bacteria: Propionibacterium, Eubacterium, Peptococcus and Peptostreptococcus genera.

13. Spirochaetes
- Treponema genus. T. pallidum
- Borrelia genus. B. burgdorferi
- Leptospira genus. L. interrogans

14. Mycobacterium genus, Nocardia genus and Actinomyces genus
- M. tuberculosis, M. leprae
- Atypical mycobacteria: Nocardia genus, Actinomyces genus

15. Mycoplasma genus and Ureaplasma genus
- M. pneumoniae
- Ureaplasma genus

16. Rickettsia genus and Coxiella genus
- Rickettsia genus: R. conori
- Coxiella genus: C. burnetii

17. Chlamydia genus
- C. trachomatis
- C. psittaci
- C. pneumoniae







Clinical Microbiology and Parasitology

                                                                                              Back to Clinical Analysis Laboratory page
A. GENERAL MICROBIOLOGY:
1. Concept of medical microbiology and parasitology
2. Bacteria
3. Fungi
4. Viruses
5. Parasites: protozoa and helminths
6. Microbial genetics
7. Action of physical and chemical agents on microorganisms

B. PATHOGENY OF INFECTIOUS DISEASES AND MICROBIAL IMMUNOLOGY:
1. Interrelationship between host and microorganism
2. Non-specific defence mechanisms
3. Factors that determine pathogenic action
4. Specific defence mechanisms

C. BASIC PRINCIPLES IN THE DIAGNOSIS, EPIDEMIOLOGY AND PROPHYLAXIS OF INFECTIOUS DISEASES:
1. Laboratory diagnosis of infectious diseases
2. Epidemiology and prophylaxis of infectious diseases
3. Immunity to infection: Vaccines and serums

D. CHEMOTHERAPY:
1. Antibacterial and antifungal agents
2. Antiviral agents: antiparasitic agents

E. SYSTEMATIC BACTERIOLOGY

F. SYSTEMATIC MYCOLOGY
1. Superficial, cutaneous and subcutaneous mycosis
2. Systemic mycosis
3. Opportunistic mycosis

G. SYSTEMATIC VIROLOGY

H. SYSTEMATIC PARASITOLOGY