World of Mycoplasma pneumonia~~: 2008

Friday, April 18, 2008

References:

DC Krause, MF Balish - FEMS Microbiology Letters, 2001 - Blackwell Synergy

http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=93483

Waris, M.E., et al (1998). Diagnosis of Mycoplasma pneumoniae pneumonia in children

http://s99.middlebury.edu/BI330A/projects/Howard/Mpneumoniae.html#anchor723401

http://en.wikipedia.org/wiki/Mycoplasma_pneumoniae
http://jcm.asm.org/cgi/content/full/36/4/1151, Journal of Clinical Microbiology
http://www.roadback.org/index.cfm/fuseaction/education.display.html&display_id=93
Molecular Biology and Pathogenesis. (American Society for Microbiology, Washington, D.C.)
http://nar.oxfordjournals.org/cgi/content/abstract/25/4/701
http://en.wikipedia.org/wiki/Mycoplasma
http://pathmicro.med.sc.edu/mayer/myco.htm
http://www.emedicine.com/EMERG/topic467.htm
http://books.google.com.my/books?id=RVH-CDn-R1kC&pg=PA48&lpg=PA48&dq=vaccine+on+mycoplasma+pneumonia&source=web&ots=T-1IMQfLjX&sig=kIKsx8o0vJ8x-PSuNkPfXj5CV4s&hl=en
http://www.healthscout.com/ency/68/205/main.html
http://www.jstor.org/pss/25184
http://en.wikipedia.org/wiki/Vaccine

Mycoplasma pneumoniae has classically been considered an extracellular (or membrane-associated) organism. Nevertheless, the recently elucidated genomic structure of this pathogen strongly suggest that this organism may have been subjected to the process of reductive genetic evolution which is characteristic of intracellular bacteria.

Besides that, the 5S rRNA sequences of eubacteria and mycoplasmas have been analyzed and a phylogenetic tree constructed. The sequences of 5S rRNA from Clostridium innocuum, Acholeplasma laidlawii, Acholeplasma modicum, Anaeroplasma bactoclasticum, Anaeroplasma abactoclasticum, Ureaplasma urealyticum, Mycoplasma pneumoniae, and Mycoplasma gallisepticum is being analyzed. The analysis of these and published sequences shows that mycoplasmas form a coherent phylogenetic group that, with C. innocuum, arose as a branch of the low G+C Gram-positive tree, near the lactobacilli and streptococci. The initial event in mycoplasma phylogeny was formation of the Acholeplasma branch; hence, loss of cell wall probably occurred at the time of genome reduction to ≈ 1000 MDa. A subsequent branch produced the Spiroplasma. This branch appears to have been the origin of sterol-requiring mycoplasmas. During development of the Spiroplasma branch there were several independent genome reductions, each to ≈ 500 MDa, resulting in Mycoplasma and Ureaplasma species. Mycoplasmas, particularly species with the smallest genomes, have high mutation rates, suggesting that they are in a state of rapid evolution.

Vaccination and Eradication:

We can consider modify the virulent factor and used it as a vaccine. As we know that, when Mycoplasma pneumonia is interacted with the host cells, it will inhibit the host cells’ catalase which can be used to breaks down hydrogen peroxide and toxic form of oxygen which is lethal to the cells. We can introduce anti-toxin as a vaccine which is to be injected to the human for prevention of the disease.

Gene-modified Mycoplasma pneumoniae can be considered to be made into vaccine against itself too. The gene therapy required method eliminates the disease-causing gene and replaced it with a modified gene, which can be inserted with functions that enhance the cell regulatory in host cells. Little amount of the gene-modified M. pneumonia can be injected to human body as a vaccine and trigger the immune response of the human body to produce antibody of immune cells to bind at the surface protein of the pathogens and attract phagocytes to kill them.

New Prevention or Treatment:

Since M. pneumoniae infections are almost exclusively mild, preventative measures are not normally recommended as necessary. But then, for some reason, people still can get vaccinated as recommended by physician.

Get vaccinated. Because pneumonia can be a complication of the flu, getting a yearly flu shot is a good way to prevent viral influenza pneumonia, which can lead to bacterial pneumonia. Your doctor may recommend a pneumonia vaccine if you're a smoker, if you're younger but have a lung or cardiovascular disease, certain types of cancer, diabetes or sickle cell anemia, if your immune system is compromised or if you've had your spleen removed for any reason.

Get flu shot. Yearly flu shot can help to prevent pneumoniae since viral pneumoniae may develop from influenza.

New Diagnostic Strategy:

By using PCR - method for amplifying a selected nucleic acid sequence. To target the amplification to a specific DNA segment, two primers bearing the complementary sequences that are unique to the target gene are used. These two primers hybridize to opposite strands of the target DNA, thus enabling DNA polymerase to extend the sequence between them. Each cycle produces a complementary DNA strand to the target gene. Newer detection methods targeting protein antigens or nucleic acids hold promise for more rapid diagnosis. To diagnosed it, usually the physician use of a nested-PCR assay to detect M. pneumoniae DNA in tissue samples from two patients with severe pneumonia. With this approach, a definitive diagnosis was available within 24 h.

Nowadays, there are many PCR test kit that available that can be used to diagnose the disease and it is easy to handle. Mycoplasma complement fixation or MCF which is the replacement test for cold agglutinin titer, identifies antibodies to a specific mycoplasma and are reported as a titer to the specific mycoplasma(s) resident in the patient. This is more specific than the M. pneumoniae kit.

We can also collect the samples from the patients, screen the samples and find out if there are any presence of DNA or protein sequence of the pathogen by the help of existing software. For this case, we can check the presence of the unique adhesin protein of M. pneumonia which allows close interaction between the pathogen and the host cells. It is a useful method which can be used to identify the pathogen which attack the patient’s system.

Genome Information:

When Mycoplasma species were first cultured, they were thought to have been virus because of their size. However, after discovered the presence of DNA and RNA, they are said to be types of bacteria. They are very tiny and able to fit through 450nm pore diameter membrane filters. One of the members of Mycoplasma is M. pneumomiae. The complete sequence of M. pneumoniae genome indicates that it is a circular duplex DNA of 816,394 bp. The presumptive origin of replication is in an A+T rich region between dnaA and dnaN. A total of 677 open reading frames (ORFs) and 39 genes coding for various RNAs were identified. M. pneumoniae has a degenerate genome because it uses some unique genetic code to make its code more similar to mitochondria than to other bacteria. It lacks the cellular machinery for making new purines and pyrimidines. However, it has tri-carboxylic acid cycle and an incomplete electron transport chain.

The coding regions in M. pneumoniae comprise a total length of 724,174 bp, or 88.7% of the genome. The average size of a gene is 1011 bp, giving an average of one gene every 1140 bp. This is similar to the gene size and density seen with both the smaller M. genitalium genome as well as the H. influenza genome, which is more than twice as large.

Below is the comparison of genome information between M. pneumoniae and M. genitalium:

We compare them with emphasis on genome organization and coding capacity. All the 470 proposed open reading frames (ORFs) of the smaller M.genitalium genome (580 kb) were contained in the larger genome (816 kb) of M.pneumoniae. There were some discrepancies in annotation, but inspection of the DNA sequences showed that the corresponding DNA was always present in M.pneumoniae. The two genomes could be subdivided into six segments. The order of orthologous genes was well conserved within individual segments but the order of these segments in both bacteria was different. We explain the different organization of the segments by translocation via homologous recombination. The translocations did not disturb the continuous bidirectional course of transcription in both genomes, starting at the proposed origin of replication. The additional 236 kb in M.pneumoniae, compared to the M.genitalium genome, were coding for 209 proposed ORFs not identified in M.genitalium. Of this ORFs 110 were specific to M.pneumoniae exhibiting no significant similarity to M.genitalium ORFs, while 76 ORFs were amplifications of ORFs existing mainly as single copies in M.genitalium. In addition, 23 ORFs containing a copy of either one of the three repetitive DNA sequences RepMP2/3, RepMP4 and RepMP5 were annotated in M.pneumoniae but not in M.genitalium, although similar DNA sequences were present. The M.pneumoniae-specific genes included a restriction-modification system, two transport systems for carbohydrates, the complete set of three genes coding for the arginine dihydrolase pathway and 14 copies of the repetitive DNA sequence RepMP1 which were part of several different translated genes with unknown function.

Prevention and Treatment:

Antibiotics such as erythromycin, clarithromycin (Biaxin) or azithromycin (Zithromax) are effective treatment. However, because mycoplasma infection usually resolves on its own, antibiotic treatment of mild symptoms is not always necessary. The doctor will recommend bedrest, plenty of fluids, therapeutic coughing, breathing exercises, proper diet, cough suppressants, pain relievers and fever reducers such as aspirin or acetaminophen. Home care includes rest and a high-protein diet with adequate fluids. With proper treatment, full recovery is expected.

Since there is no vaccine available, to prevent the occurrence of the disease, we need to apply good and hygiene daily life by cleaning and disinfect common touched surfaces regulary. Do not share cup, toothbrush, eating utensils to prevent exchange of saliva that may have carried the pathogen. Limit eating “community” food that may have been contaminated by someone’s hands. Stay home and always avoid crowding environment if you have a fever. Lastly, do not smoke.

Diagnosis:

To diagnose pneumonia, the doctor begins with a medical history and physical examination. By placing a stethoscope on the chest, the doctor may be able to hear crackling sounds, coarse breathing, wheezing and/or the breathing may be faint in a particular area of the chest. Additionally, the doctor may order a chest x-ray, a sputum gram stain and a blood test. The chest x-ray may show a blotchy-white area, where fluid and pus has accumulated in the lung's air sacs. The sputum grain stain and the blood test may determine the cause and severity of the condition.

A positive blood test for cold-hemagglutinins in 50-70% of patients after 10 days of infection (cold-hemagglutinin-test should be used with caution or not at all since 50% of the tests are false-positive), lack of bacteria in a gram-stained sputum sample, and a lack of growth on blood agar. Mycoplasma atypical pneumonia can be complicated by Stevens-Johnson syndrome, hemolytic anemia, encephalitis or Guillain-Barré syndrome.

Other serology tests include complement fixation which detects both IgG and IgM specific to M.pneumoniae, enzyme-linked immunoassay for M. pneumonia specific IgM, and indirect hemagglutination. All of these have acceptable sensitivity and specificity.

Pathogenesis:

The pathogenesis of the microorganism has two properties: The first is a selective affinity for respiratory epithelial cells, and the second is the ability to produce hydrogen peroxide, which is thought to be responsible for much of the initial cell disruption in the respiratory tract and for damage to erythrocyte membranes. Colonization of the respiratory tract by M. pneumoniae results in the cessation of ciliary movement. The normal clearance mechanisms of the respiratory tract do not function, resulting in contamination of the respiratory tract and the development of a dry cough.

Mycoplasma pneumonia cause infection primarily as extracellular parasites, attaching to the surface of ciliated and nonciliated epithelial cells. This is done by the P1 adhesin protein that localizes themselves at the tips of the bacterial cells and binds to sialic acid residues on host epithelial cells. The attachment site, or receptor, is a complex carbohydrate structurally akin to antigen I of red blood cells. Following attachment, mycoplasmal organisms may cause direct cytotoxic damage to epithelial cells because of hydrogen peroxide generation or cytolysis via an inflammatory response mediated by mononuclear cells or antigen-antibody reactions.

Mycoplasma pneumoniae is also thought to be involved in mediating other diseases and infections in its monopolization of the immune response. Some patients have developed severe bacterial and viral infections just after or during a M. pneumoniae infection. This is thought to occur by the creation of an environment that is anatomically, physiologically, and/or immunologically conducive to other organisms for invasion as well as for cellular damage.

The diagram above illustrates mycoplasmas (red) infecting epithelial cells (black). The mycoplasmas form a small structure called a "tip" by which they adhere to the epithelial cells. If the mycoplasmas are prevented from adhering to the epithelial cells, they are avirulent.

Virulence Factor:

* You may click on the table to view a bigger size image.

Reservoir and Transmission:

The children are an important M. pneumoniae reservoir. Mycoplasma pneumoniae is transmitted from person-to-person contact through respiratory secretions during coughing and sneezing which is respiratory droplet transmission. Hence, close contact is required for transmission, and the bacteria are more commonly found among members of the same family and in schools and day-care institutions. The contagion requires a continuous contact because of the sensitivity of M. pneumoniae to changes in temperature and humidity levels. There is little point in isolating someone infected with the virus since some people carry the infection without feeling ill.

Once attached to the mucosa of a host organism, M. pneumonia extracts nutrients, grows and reproduces by binary fission. Attachment sites include the upper and lower respiratory tract, causing pharyngitis, bronchitis and pneumonia. The infection caused by this bacterium is called atypical pneumonia or “walking pneumonia” because of its protracted course and lack of sputum production and wealth of extra-pulmonary symptoms.

Mycoplasma pneumoniae has an incubation period of about 1 to 4 weeks, which means it may take up to four weeks after exposure to get Mycoplasma pneumonia. They symptoms of infection are headaches, muscle pain, scratchy aching throat, worsening dry cough, fever, chills, vomiting, slow heartbeat, breathlessness, bluish color to lips and nail beds, diarrhea and rash. The most common clinical syndrome following with M. pneumonia is tracheobronchitis. However, over this time period, symptoms may slowly begin to appear. An epidemic occurs every three to five years.

Its Biological Characteristics:

Mycoplasma pneumoniae is the smallest (range from 0.2-0.8 micrometers) free-living bacteria and it is member of the class Mollicutes. This class of organisms lack of peptidoglycan cell wall. Instead, it has a cell membrane which incorporates sterol compounds, similar to eukaryotic cells. It obtains sterols from host serum, allowing it to retain the cell’s structure. Since it has no cell wall, it is resistance to penicillins and other beta-lactam antibiotic which are designed to disrupt the bacterial cell wall. And so, they can be parasitic and saprophytic.

Mycoplasma pneumonia are extracellular, mucous membrane pathogen that do not invade other tissues. They have a low GC-content (18-40 mol%). The analysis of 16S ribosomal RNA sequences suggest that this kind of bacteria is classified under gram-positive since they are too small to be stained. They may induce cellular changes including changes in metabolism and cell growth and infection.

M. pneumonia has the smallest genome size and as a result, lack of many metabolic pathways and so they need to extract nutrient especially glucose from host’s cells and metabolize it in order to get energy for survival and reproduction. M. pneumonia is strict aerobes. They grow slowly by binary fission and the colonies may take up to 3 weeks to develop and so the physical sign of infection of the patient will present for several days to weeks.

Introduction

After blasting for the nucleotide sequences, we found that the gene that responsible to the sequences is Mycoplasma pneumoniae adhesin protein (P30) gene. It was found in Mycoplasma pneumonia. The gene coded for adhensin protein P30 which is required for cytoadherence and associated with cell development of M.pneumoniae. Loss of P30 caused a mutation and the morphological abnormalities like change in cell shape. It indicates P30 plays a role in mycoplasma cell development and support adhesin P1’s receptor-binding function that enhance the attachment to host cells.

Mycoplasma pneumonia

This is what we found from Neuclotide BLAST

Formating result: Z24WWEVE012

Gene: Mycoplasma pneumoniae adhesin protein (P30) gene

Organism: Mycoplasma pneumoniae

Accesion number: EF614306.1

PubMed: 18032594

Max score: 1524

Totral score: 2739

Query coverage: 100%

E value: 0.0

Max ident: 100%

LOCUS EF614306 825 bp DNA linear BCT 18-MAR-2008

DEFINITION Mycoplasma pneumoniae adhesin protein (P30) gene, complete cds.

ACCESSION EF614306

VERSION EF614306.1 GI:148729625

KEYWORDS .

SOURCE Mycoplasma pneumoniae

ORGANISM Mycoplasma pneumoniae

Bacteria; Firmicutes; Mollicutes; Mycoplasmataceae; Mycoplasma.

REFERENCE 1 (bases 1 to 825)

AUTHORS Varshney,A.K., Chaudhry,R., Kabra,S.K. and Malhotra,P.

TITLE Cloning, expression, and immunological characterization of the P30

protein of Mycoplasma pneumoniae

JOURNAL Clin. Vaccine Immunol. 15 (2), 215-220 (2008)

PUBMED 18032594

REFERENCE 2 (bases 1 to 825)

AUTHORS Varshney,A.K., Chaudhry,R., Malhotra,P. and Kabra,S.K.

TITLE Direct Submission

JOURNAL Submitted (17-MAY-2007) Microbiology, All India Institute of

Medical Sciences, Ansari Nagar, New Delhi 110029, India

FEATURES Location/Qualifiers

source 1..825

/organism="Mycoplasma pneumoniae"

/mol_type="genomic DNA"

/isolation_source="clinical sample"

/db_xref="taxon:2104"

gene 1..825

/gene="P30"

CDS 1..825

/gene="P30"

/codon_start=1

/transl_table=4

/product="adhesin protein"

/protein_id="ABR09215.1"

/db_xref="GI:148729626"

/translation="MKLPPRRKLKLFLLAWMLVLFSALIVLATLILVQHNNTELTEVK

SELSPLNVVLHAEEDTVQIQGKPITEQAWFIPTVAGCFGFSALAIILGLAIGLPIVKR

KEKRLSEEKERQEQLAEQLQRISAQQEEQQALEQQAAAEAHAEAEVEPAPQPVPVPPQ

PQVQINFGPRTGFPPQPGMAPRPGMPPHPGMAPRSGFPPQPGMAPRPGMPPHPGMAPR

PGFPPQPGMAPRPGMPPHPGMAPRPGFPPQPGMAPRPGMQPPRPGMPPQPGFPPKR"

ORIGIN

1 atgaagttac cacctcgaag aaagcttaaa ctgtttttat tagcctgaat gctagtgctg

61 ttcagcgctt taatagtgct tgcaacctta attttggtac agcacaacaa taccgaactg

121 acagaagtta agagtgaatt gagtcccctt aacgttgttt tacacgcaga agaggataca

181 gtacaaattc agggcaagcc gattactgag caagcatggt ttattcctac agttgctggt

241 tgctttggtt ttagtgccct agccatcatc ttgggccttg ctataggact gccaattgtg

301 aagcgcaagg aaaaacgctt atcggaggaa aaggaacgcc aagaacagtt agcggaacag

361 ctacaacgca tttctgccca acaagaagag caacaagcgt tagaacaaca agcagctgct

421 gaagcccatg ctgaagcgga agttgaacca gcaccacaac cagtaccagt accacctcaa

481 ccccaagtcc aaattaactt cggtccccgt actggtttcc cacctcaacc cggtatggcg

541 cctcgtccag gtatgccgcc acaccccggt atggctccaa gatctggttt cccacctcaa

601 cccggtatgg cgcctcgtcc aggtatgccg ccacaccccg gtatggctcc aagacctggt

661 ttcccacctc aacccggtat ggcgcctcgt ccaggtatgc cgccacaccc cggtatggct

721 ccaagacctg gtttcccacc tcaacccggt atggcgcctc gtcccggaat gcaaccacca

781 cgtcctggca tgccacccca acccggtttt ccaccaaaac gctaa

Which organism that will affect our respiratory system?

Group 1- Respiratory system