Diagnosis of H. pylori infection

Various tests have been developed for the detection of H. pylori, each with their specific advantages and disadvantages. The available tests are generally divided into invasive tests, based on gastric specimens for histology, culture, or other methods, and noninvasive tests, based on peripheral samples, such as blood, breath samples, stools, urine, or saliva for detection of antibodies, bacterial antigens, or urease activity. The choice of a specific test for an individual patient depends on local experience and the clinical setting. In research protocols, a combination of two methods is often applied. In daily clinical practice, use of a single test is generally adequate, and most tests are sufficiently accurate to be used for this purpose. For routine diagnostic purposes, histology, urea breath testing, and culture are currently most often used, whereas the use of serology is most appropriate for large epidemiological studies. In hospital-based care, many patients undergo endoscopy, which is then combined with an invasive test for H. pylori. Otherwise, breath tests and serology are commonly used. For children, fecal antigen tests offer the opportunity to assess H. pylori status without the need for endoscopy or vena puncture.

Endoscopic Diagnostic Tests (Invasive tests)

In patients who have not been on a PPI (proton pump inhibitor)  within 1–2 wk or an antibiotic or bismuth within 4 wk of endoscopy, the rapid urease test (RUT) provides an accurate, inexpensive means of identifying H. pylori. For patients who have been taking a PPI, antibiotics, or bismuth, endoscopic testing for H. pylori should include biopsies from the gastric body and antrum for histology with or without rapid urease testing. Though culture or polymerase chain reaction (PCR) are the primary means by which antibiotic sensitivities can be determined, neither is widely available for clinical use and therefore, cannot be routinely recommended.

There are presently four biopsy-based diagnostic methods for H. pylori infection. These include the RUT, histology, culture, and PCR.

Rapid Urease Testing-

The RUT identifies active H. pylori infection through the organism’s urease activity. Gastric biopsies are obtained and placed into an agar gel or on a reaction strip containing  urea, a buffer, and a pH-sensitive indicator. In the presence of H. pylori’s urease, urea is metabolized to ammonia and bicarbonate leading to a pH increase in the microenvironment of the organism. A change in color of the pH sensitive indicator signifies the presence of active infection. Commercially available kits yield results in 1–24 h. Medications that reduce the density and/or urease activity of H. pylori, such as bismuth-containing compounds, antibiotics, or PPIs, can decrease the sensitivity of the RUT by up to 25%. Though controversial, acute ulcer bleeding at the time of testing may decrease the sensitivity and negative predictive value of the RUT. As a result of the patchy distribution of H. pylori infection after antibiotics or PPIs, it is recommended that biopsies for the RUT be obtained

from two sites, the body at the gastric anglularis and greater curvature of the antrum. The simplicity, low cost, and relatively rapid results make the RUT a practical and cost effective

means of testing for H. pylori in patients not taking antibiotics, bismuth, or PPIs who require upper endoscopy. Unfortunately, the usefulness of the RUT in routine clinical

practice has been compromised by the widespread use of PPIs as an empiric treatment for upper GI symptoms. As such, the RUT can rarely be used as a sole means of identifying H. pylori infection. More commonly, the RUT is combined with other endoscopic or non  endoscopic modalities to establish the presence or absence of this infection. No studies have been performed to define the duration of a PPI’s deleterious effects on the sensitivity of the RUT. Data with the urea breath test (UBT) suggest that PPI therapy can cause false-negative test results for 1–2 wk (68, 69). As the UBT and RUT rely upon the identification of H. pylori’s urease activity, it is reasonable to suggest that PPIs should be withheld for 1–2 wk before performance of the RUT. In situations where a patient has not taken a PPI for a period of 1–2 wk before their procedure, the sensitivity of the RUT is likely sufficient to justify its use as a single test for H. pylori.

Histology-

Histology has been considered by some to be the gold standard for detection of H. pylori. Unfortunately, histology is an imperfect gold standard as the detection of H. pylori relies upon a number of issues including the site, number, and size of gastric biopsies, method of staining, and the level of experience of the examining pathologist. A significant advantage of histology over other diagnostic methods is the ability to evaluate for pathologic changes associated with H. pylori infection such as inflammation, atrophy, intestinal metaplasia, and malignancy. Certainly the absence of chronic gastritis is a potent negative predictor for the presence of H. pylori infection. As the prevalence and density of H. pylori varies throughout the stomach, particularly in the face of medications that may reduce the density of H. pylori, multiple biopsies are needed for accurate diagnosis. It is therefore recommended that a minimum of three biopsies be obtained, one from the anglularis, one from the greater curvature of the corpus, and one from the greater curvature of the antrum, to maximize the diagnostic yield of histology. A recent study found that the addition of corpus biopsies to antral biopsies increased the detection of H. pylori infection by 10% when compared with antral biopsies alone. Similar to the RUT, the sensitivity of histology is significantly affected by the use of medications such as bismuth, antibiotics, and PPIs. Although widely available and capable of achieving sensitivity and specificity of >95%, the cost and need for properly trained.

Culture

Culture is another highly specific method for identifying active H. pylori infection. Conceptually, culture is attractive because it not only provides a means by which to identify infection, but also allows characterization of antimicrobial sensitivities. Unfortunately, culture is not as sensitive as RUT or histology. Furthermore, culturing techniques for H. pylori are demanding and costly and as a consequence, only available in a limited number of clinical laboratories. Nonculture-based means of determining antibiotic resistance are being developed but have not been adequately standardized and are not widely available.

Polymerase Chain Reaction

PCR is a DNA amplification technique that utilizes the rapid production of multiple copies of a target DNA sequence to identify H. pylori. This testing method is highly specific and may be more sensitive than other biopsy-based diagnostic techniques. A recent study found that PCR was able to detect H. pylori in approximately 20% of gastric biopsies with chronic gastritis but no identifiable organisms by histology PCR also provides a means of identifying mutations associated with antimicrobial resistance (78–80). Although presently restricted to the research arena, this method may some day provide a practical, reproducible method for antibiotic sensitivity testing, organism typing, and organism virulence testing.

Non endoscopic Diagnostic Tests (Non-Invasive tests)

Antibody testing is inexpensive and widely available. The UBTs and fecal antigen tests provide reliable means of identifying active H. pylori infection before antibiotic therapy. The UBT is the most reliable non endoscopic test to document eradication of H. pylori infection. The monclonal fecal antigen test provides another non endoscopic means of establishing H. pylori cure after antibiotic treatment. Testing to prove H. pylori eradication appears to be most accurate if performed at least 4 wk after the completion of antibiotic therapy.

There are currently three non endoscopic diagnostic testing methods for H. pylori infection. Antibody testing identifies an immunological reaction to the infection while the non endoscopic urease tests and fecal antigen test identify the presence of active H. pylori infection.

Antibody Tests

Antibody testing relies upon the detection of IgG antibodies specific to H. pylori in serum, whole blood, or urine. IgG antibodies to H. pylori typically become present approximately 21 days after infection and can remain present long after eradication. Antibodies to H. pylori can be quantitatively assessed using enzyme-linked immunosorbent assay(ELISA) and latex agglutination techniques or qualitatively assessed using office-based kits. The advantages of the antibody tests are their low cost, widespread availability, and rapid results. Unfortunately, several factors limit the usefulness of antibody testing in clinical practice. According to studies, the antibody test kits shows about 85% sensitivity and 79% specificity. One of the limitation is that antibody tests developed using antigens from one region of the world may not perform well when applied to patients in another part of the world suggesting that local validation may be necessary. Finally, antibody tests are of little benefit in documenting eradication as results can remain positive for years following successful cure of the infection.

Urea Breath Tests

The UBT, like the RUT, identifies active H. pylori infection by way of the organism’s urease activity. In the presence of H. pylori, the ingestion of urea, labeled with either the nonradioactive

isotope ¹³C or the radioactive isotope ¹⁴C, results in production of  labeled CO₂, which can be quantitated in expired breath. Although the amount of radiation in the ¹⁴C UBT is less than daily background radiation exposure, the ¹³C test is preferred in children and pregnant females. Overall, the performance characteristics of both tests are similar with sensitivity and specificity typically exceeding 95% in most studies. Test reproducibility has been found to be excellent. The UBT also provides an accurate means of post treatment testing. Most tests utilize a citrate test meal (50–75 mg), which is administered before the labeled urea. A urease blood test, which relies upon the detection of labeled bicarbonate in a blood sample, also reliably identifies active H. pylori infection before and after treatment. As the non endoscopic urease tests rely upon the identification of H. pylori’s robust urease activity, test sensitivity is decreased by medications that reduce organism density or urease activity, including bismuth containing compounds, antibiotics and PPIs. It is currently recommended that bismuth and antibiotics be withheld for at least 28 days and a PPI for 7–14 days prior to the UBT. The UBT is more costly than the antibody tests or fecal antigen test. The expense of the UBT is largely driven by equipment costs and the cost of labeled urea. UBTs using lower dose ¹³C, which have recently been found to yield excellent performance characteristics, may in part address this issue.

Fecal Antigen Test

The fecal antigen test (FAT) identifies H. pylori antigen in the stool by enzyme immunoassay with the use of polyclonal anti-H. pylori antibody. Recently, a stool test utilizing a monoclonal

anti-H. pylori antibody has been evaluated. As both tests detect bacterial antigen(s) suggestive of ongoing infection, they can be used to screen for infection and as a means of establishing cure following therapy. Similar to the UBT, the sensitivity of the FAT is affected by the recent use of bismuth compounds, antibiotics, and PPIs. Recent studies also suggest that the specificity of the FAT is reduced in the setting of bleeding peptic ulcer disease and, for this reason, should not be the sole diagnostic test employed in this setting. Although the FAT is simple to administer and perform, issues slowing its widespread use include the unpleasantness of handling and storing stool, limited availability. The development of in-office stool tests is under way and may improve upon some of the practical limitations of the currently available tests. At present,

in-office tests have not been adequately validated in clinical trials. Based upon the available data, it is reasonable to conclude that the FAT can be used interchangeably with the UBT to identify H. pylori before antibiotic therapy. The polyclonal FAT has been less well validated than the UBT in the post treatment setting. Compared with the polyclonal test, the monoclonal FAT appears to provide a more reliable means of proving H. pylori eradication.

Pathogenicity of H. pylori

Helicobacter pylori is the first formally recognized bacterial carcinogen and is one of the most successful human pathogens, as over half of the world's population is colonized with this gram-negative bacterium. Unless treated, colonization usually persists lifelong.After an incubation period of a few days, H. pylori causes, in some persons, a mild gastritis which may last for about two weeks. The infection may be transient in some, but in most, it persists for years or decades. Such colonization is usually asymptomatic, though chronic superficial gastritis may be demonstrable histologically. The bacteria are present only in the overlying mucus and do not invade the mucosa. Gastric antrum is the commonest site of colonization, though any part of the stomach may be involved. The infection is strictly confined to the gastric mucosa, in the stomach, as well as in areas of gastric metaplasia and heteropia in the duodenum.

To colonize the stomach, H. pylori must survive the acidic pH of the lumen and use its flagella to burrow into the mucus to reach its niche, close to the stomach's epithelial cell layer. Many bacteria can be found deep in the mucus, which is continuously secreted by mucus-secreting cells and removed on the luminal side. To avoid being carried into the lumen, H. pylori senses the pH gradient within the mucus layer by chemotaxis and swims away from the acidic contents of the lumen towards the more neutral pH environment of the epithelial cell surface. H. pylori is also found on the inner surface of the stomach epithelial cells and occasionally inside epithelial cells. It produces adhesins which bind to membrane-associated lipids and carbohydrates which helps it adhere to the epithelial cells.

 H. pylori produces large amounts of the enzyme urease molecules of which are localized inside and outside of the bacterium. Urease breaks down urea (which is normally secreted into the stomach) to carbon dioxide and ammonia. The ammonia is converted to ammonium by accepting a proton (H⁺), which neutralizes gastric acid. The survival of H. pylori in the acidic stomach is dependent on urease. Unlike other bacterial ureases, which are found only in the cytoplasm, H. pylori urease is located and also enzymatically active on the cell surface. Cytoplasmic urease adsorbs onto the surface of intact bacteria after the protein is released from the cytoplasm by spontaneous autolysis of a subpopulation of the bacterium. A recent crystal structure study demonstrated that the H. pylori urease enzyme has a unique supramolecular structure that may be important for survival in acidic conditions. The H. pylori urease has a 3-fold symmetry, spherical assembly of about 12 catalytic sites, containing only α and β subunits in contrast to trimers of α, β, γ subunits in other bacterial ureases. The ammonia produced is toxic to the epithelial cells, and, along with the other products of H. pylori—including proteases, vacuolating cytotoxin A (VacA), and certain phospholipases, damages those cells.

The cagA (cytotoxin-associated gene A) gene is the most extensively studied of the H. pylori genes. Early studies indicated that the CagA protein was a marker for more severe disease because it was more frequently associated with strains isolated from patients with peptic ulcer and gastric cancer. It is almost always associated with increased inflammation and more severe disease. However, it’s almost universal presence in strains in developing countries irrespective of disease presentation indicates that other factors (bacterial, host, and environmental) must also be important for development of peptic ulcer or gastric malignancy. Variability in the 3′ region of the cagA gene may confer greater risk for gastric cancer and an enhanced susceptibility to pH, but direct evidence for this is still needed. Alternatively, it may reflect the geographical clustering of the infection.

The VacA protein influences cellular processes via different routes, thus assisting in chronic colonization of the gastric mucosa by H. pylori. Surface-bound VacA may be directly delivered to the cell membrane whereas, secreted VacA may either bind to a cell membrane receptor and initiate a proinflammatory response, or can be taken up directly by the cell and be trafficked to the mitochondria and induce apoptosis. It can be taken up by pinocytosis and induce vacuolization, or form a membrane channel, resulting in leakage of nutrients to the extracellular space, or pass through the tight junctions and inhibit T-cell activation and proliferation.

Two related mechanisms by which H. pylori could promote cancer are under investigation. One mechanism involves the enhanced production of free radicals near H. pylori and an increased rate of host cell mutation. The other proposed mechanism has been called a "perigenetic pathway", and involves enhancement of the transformed host cell phenotype by means of alterations in cell proteins, such as adhesion proteins. H. pylori has been proposed to induce inflammation and locally high levels of TNF-α and/or interleukin 6 (IL-6). According to the proposed perigenetic mechanism, inflammation-associated signaling molecules, such as TNF-α, can alter gastric epithelial cell adhesion and lead to the dispersion and migration of mutated epithelial cells without the need for additional mutations in tumor suppressor genes, such as genes that code for cell adhesion proteins.  Presence of H. pylori infection triggers a excessive inflammatory and immune response in the host. IL-6 and IL-8 appear early in the cascade of inflammatory molecules. H. pylori products also stimulate IL-10, IL-12 and interferon (IFN)-γ in a Th1 type of gastric immune response.

Tuberculosis Made Easy

Article by IfuM

Tuberculosis is a disease which everyone usually talks about; but they have a very few knowledge about it. Are you one of them? If yes then just read the article and spend a few minutes of your time and know about tuberculosis.

Tuberculosis is a disease which is cause by the organism Mycobacterium tuberculosis. It is spread by infected people through air when they cough, sneeze, or by transmitting their saliva through air. The organism mostly affects lungs, but can also affect other parts of the body.

Some WHO facts about Tuberculosis: 

• Tuberculosis (TB) is second only to HIV/AIDS as the greatest killer worldwide due to a single infectious agent.
• In 2010, 8.8 million people fell ill with TB and 1.4 million died from TB.
• In 2009, there were about 10 million orphan children as a result of TB deaths among parents.
• TB is a leading killer of people living with HIV causing one quarter of all deaths.
• Multi-drug resistant TB (MDR-TB) is present in virtually all countries surveyed.

Tuberculosis mostly affects young adults. Use of tabacco increases the risk of tuberculosis disease and death. Tuberculosis occurs in almost every part of the world. Common symptoms of active lung TB are cough with sputum and blood at times, chest pains, weakness, weight loss, fever and night sweats.
Tuberculosis is treatable as well a curable.

The treatment is done by a 6 month course by 4 anti-microbial drugs namely isoniazid, rifampicin, pyrazinamide and ethambutol.

Multidrug-resistant tuberculosis:

If the organism is resistant to the drugs ioniazid and rifampicin the disease is know as Multidrug-resistant tuberculosis.Primary cuase of Multidrug-resistant tuberculosis is improper treatment and use of poor quality medicines. The treatment for drug resistant tuberculosis is very expensive and takes around 2 years. 

 

Extra Knowledge:
The bacillus causing tuberculosis, Mycobacterium tuberculosis, was identified and described on 24 March 1882 by Robert Koch. He received the Nobel Prize in physiology or medicine in 1905 for this discovery.

Preventing tuberculosis:
Vaccinations: Infants are vaccinated with bacillus Calmette-Guerin (BCG) vaccine because it can prevent severe tuberculosis.

Staying away from tabacco: Better safe than sorry.

Education & awareness: Last but not the least. How to do it right now?
Simple. Do spread the word by simply recommending/sharing this article.

I AM A BUG .........SUPER-BUG!!!

Bug is a slang word for micro-organisms. Super-bug, it has nothing to do with the  Volkswagen car or superman it is not a bug that saves life but they do have something in common with him that is the power to  survive through difficult situations. How do they get this power? Its genes, they acquire these genes by a mechanism, these genes allow them to resist antibiotics, having one or two of these genes is very common but when they acquire a full set of genes which allow them to resist large number of antibiotics they become multi resistant, which makes it difficult to kill them or at least stop there growth. These genes give resistance to bacteria by producing a enzyme or altering a protein structure in a way that they make the antibiotic inactive and are able to flourish even the antibiotic is present. So u see they are a big problem for us, as of now several bacteria have started showing resistance against first line drugs(preferred because they are relatively cheap and less toxic) so we are forced to use second line drugs(costly and toxic). There is our very old multi-resistant Staphylococcus aureus (MRS) and the latest are the multi-resistant Mycobacteriun tuberculosis, since TB is at large scale in developing countries it makes things worst. We have a NDM-1 gene named after New Delhi which confers high resistance to the bacteria it enters. To make things worst this bacteria has reached to other countries first it was only contained in India but because of medical tourism it has reached UK n USA but India still lives in denial.

 The reason these super-bugs are popping out is, abusive use of antibiotics especially in countries  like India were there is no common board for looking after the use of antibiotics and the lack of information between the doctors and also the patients. The pharmacy also have a role to play since they give out medication without  prescription like they are candies. They have also found out another reason is that, they add drugs in feed of domestic animals which eventually enter the human body, if the bacteria acquires resistance in the animal  they are  transmitted to humans via three pathways, those being through the consumption of meat, from close or direct contact with animals, or through the environment.

The super-bugs are a serious threat  but as of yet we still have second  line drugs to control them but time will come when they will become resistant to the second line drugs.