Molecular Detection of Microorganisms and Inherited Diseases

Molecular Detection

Methods to identify microorganisms using molecular techniques.

Agarose Gel Electrophoresis

Method for separating DNA based on size.

Sequencing

Identify new sequent variant of a particular organism

Immunoassays

Tests using antibodies to detect microorganisms.

Western Blots

Technique for detecting specific proteins in samples.

Mass Spectrometry

Technique for determining molecular mass and structure.

Mycobacterium tuberculosis

Microorganisms that are difficult and/ or time-consuming to isolate

Histoplasma
Coccidioides

Hazardous to handle in the clinical laboratory

Streptococcus
Neisseria
Chlamydia

Organisms that are received in clinical laboratories in high volumes

Treponema pallidum

Bacterium causing syphilis, detectable via PCR.

Chlamydia trachomatis

Bacterium causing chlamydia, detectable via PCR.

Gonococcal Urethritis

Infection caused by Neisseria gonorrhoeae bacteria.

(UL54) or glycoprotein B (gB) gene

DNA is extracted and amplified by PCR using primers targeting the CMV polymerase _________

Epstein-Barr Virus (EBV)

Virus linked to infectious mononucleosis and cancers.

Varicella-zoster Virus (VZV)

Causes chickenpox and shingles infections.

Cytopathic Effect

Microscopic changes in cells due to viral infection.

Broad-range PCR

It detects Candida, Aspergillus, Rhizopus and other Zygomycetes, and Histoplasma

Direct probe hybridization

It detects Blastomycetes, Coccidioides immitis, and Histoplasma capsulatum

PFGE

used in laboratory-developed methods for molecular typing of yeasts.

MALDI-TOF

application to detection of fungi

Mass spectrometry

methods for identification of Saccharomyces cerevisiae

Single-gene Disorders

affect structural proteins, cell surface receptor proteins, growth regulators, and enzymes.

BK Virus

Polyomavirus associated with kidney transplant complications.

JC Virus

Polyomavirus linked to progressive multifocal leukoencephalopathy.

Candida

Fungal genus commonly causing infections.

Lysosomal Storage Diseases

Disorders caused by enzyme deficiencies in lysosomes.

Leiden Mutation

G to A change in F5 gene causing thrombosis.

Factor V

Coagulation factor associated with hypercoagulable phenotype.

Prothrombin

Factor II gene linked to thrombosis risk.

Epigenetic Alterations

Changes affecting gene expression without DNA sequence change.

DNA Methylation

Modification that typically down regulates gene transcription.

Genomic Imprinting

Selective inactivation of specific chromosomal regions.

Chromatin Remodelling

Modification of chromatin structure affecting gene accessibility.

Germ Cell Mutations

Inherited mutations leading to genetic disorders.

Somatic Cell Mutations

Mutations causing cancer and congenital malformations.

MTHFR

Enzyme converting homocysteine to methionine.

Hyperhomocysteinemia

Condition caused by MTHFR mutations affecting homocysteine levels.

Karyotyping

Technique for detecting chromosomal abnormalities.

Ploidy Analysis

Assessment of chromosome number using flow cytometry.

Fluorescent In Situ Hybridization (FISH)

Technique for visualizing chromosomal abnormalities.

Chromosome Mutations

Structural abnormalities in chromosomes larger than 4 million bp.

Hemochromatosis

Disorder caused by mutations in HFE gene affecting iron metabolism.

C282Y Mutation

Common mutation in hemochromatosis linked to iron overload.

Cystic Fibrosis

Genetic disorder caused by CFTR gene mutations.

PCR-RFLP

used for Point mutations

Allele-Specific PCR

Technique for identifying specific genetic polymorphisms.

P-450 genetic variants

Screened by microarray, bead array, or sequencing.

Penetrance

Frequency of disease phenotype expression in gene carriers.

Variable expressivity

Range of phenotypes from the same gene lesion.

Triplet expansion

Inserts multiple glutamine residues in huntingtin protein.

Huntington Disease

Neurological symptoms from huntingtin protein aggregation.

Idiopathic Congenital Central Hypoventilation Syndrome

CCHS caused by mutations in PHOX2b gene.

Mitochondrial mutations

Affect energy production in muscles and nervous system.

Genomic imprinting

Transcriptional silencing via histone or DNA modification.

Uniparental Disomy

Inheritance of both chromosomes from one parent.

Prader-Willi syndrome

Deletion in paternal chromosome 15, del(15)(q11q13).

Angelman syndrome

Deletion in maternal chromosome 15.

Southern blot

Detects large deletions in mitochondrial genes.

High-resolution karyotyping

Detects smaller deletions in chromosomes.

FISH with labelled probes

can detect over 99% of cases

Nucleotide-Repeat Expansion Disorders

Include diseases like Fragile X and Huntington's.

Fragile X Syndrome

Triplet-repeat (CGG) expansion in FMR-1 gene.

CAG repeat expansion

Huntingtin gene expands from 9-37 to 38-86 repeats.

Next-generation sequencing

Advanced method for comprehensive genomic analysis.

Methylation-specific PCR

Diagnoses disorders through DNA methylation patterns.