Monday, September 22, 2014

Staphylococcus Paradox


      "Staphylococcus aureus" or more commonly known as "The Staph", is a gram positive cocci in clusters, famous everywhere in the world for a subtype called as MRSA. It has been a really long time since i have blogged something about this organism. Perhaps this is the right time for me to get back talking about this paper.

    One of the most common infections caused by S aureus is skin infections, with an array of virulence factors involved. There are defense systems mounted on the skin, one of which is free fatty acids. The presence of these bactericidal compounds makes it less hospitable for the organisms such as staphylococcus. Obviously, staphylococcus has evolved counter defense mechanisms. But here is the catch of the question. There is a key enzyme called as SAL-2 which basically breaks down the lipid component of skin, producing high concentrations of fatty acids that inhibited bacterial growth. That seems to defy common sense. Bacteria produces an enzyme to produce a compound that will inhibits its own growth!!! As the authors say , “But in the case of the wildtype, the activity of SAL2 lipase produced high concentrations of fatty acids that inhibited bacterial growth”

      MRSA PFGE strain type USA300 (Or commonly known as USA300 strain) is a CA- MRSA subtype representing hypervirulence first reported in the USA as a cause of skin and soft issue infection in 2000. Genetically it is MLST type 8, clonal complex 8 with SccMec IV. The importance is that this strain is highly adapted to living in the skin. Several genetic factors have been proposed to have a role such as speG which induces tolerance to polyamines in the skin. This is further amplified by the fact that SAL-2 is highly conserved and detectable across S aureus.

       Quote from investigator, "In previous work, SAL-2 was identified as one of just seven secreted proteins that was universally produced by 63 diverse strains of S aureus and, as such, it is unlikely that S. aureus would have evolved to maintain abundant production of SAL2 if it did not confer a substantial benefit to the bacterium". Source

       SAL (Staphylococcus aureus lipase), comes in multiple forms, SAL-1, SAL-2 and SAL-3. SAL-2 is known to be derived into staphylococcus through a bacteriophage (Φ29 like phage, member of podoviridae) called SAP-2. Biochemical studies have shown that in its purified form it has a specific activity against S aureus cell wall with a MIC of 1μg/ml.

     So now you see the contrast. SAL-2 is inhibitory yet it is being carried almost universally and helps in virulence. In the new paper by Heinrichs etal; the paradox was studied by engineering strains of CA-MRSA that could not make SAL2 lipase, and compared the mutant and the wild-type responses to triglycerides. Result, mutants grew well in the presence of triglycerides. The basic findings were that SAL-2 is processed to active form by Aureolysin. The active form hydrolyzes triglycerides of both short and long chain fatty acids. SAL2 hydrolyzes trilinolein to linoleic acid, a fatty acid with known anti-staphylococcal properties.

      The paper has presented with the same challenge that initially was in question. However, it is clear that the SAL-2 is inhibitory, yet consistently carried and helps in better colonization. More studies are needed to elucidate the paradox.
Planet PJ, LaRussa SJ, Dana A, Smith H, Xu A, Ryan C, Uhlemann AC, Boundy S, Goldberg J, Narechania A, Kulkarni R, Ratner AJ, Geoghegan JA, Kolokotronis SO, & Prince A (2013). Emergence of the epidemic methicillin-resistant Staphylococcus aureus strain USA300 coincides with horizontal transfer of the arginine catabolic mobile element and speG-mediated adaptations for survival on skin. mBio, 4 (6) PMID: 24345744

Ziebandt AK, Kusch H, Degner M, Jaglitz S, Sibbald MJ, Arends JP, Chlebowicz MA, Albrecht D, Pantucek R, Doskar J, Ziebuhr W, Bröker BM, Hecker M, van Dijl JM, & Engelmann S (2010). Proteomics uncovers extreme heterogeneity in the Staphylococcus aureus exoproteome due to genomic plasticity and variant gene regulation. Proteomics, 10 (8), 1634-44 PMID: 20186749

Son JS, Lee SJ, Jun SY, Yoon SJ, Kang SH, Paik HR, Kang JO, & Choi YJ (2010). Antibacterial and biofilm removal activity of a podoviridae Staphylococcus aureus bacteriophage SAP-2 and a derived recombinant cell-wall-degrading enzyme. Applied microbiology and biotechnology, 86 (5), 1439-49 PMID: 20013118

Cadieux B, Vijayakumaran V, Bernards MA, McGavin MJ, & Heinrichs DE (2014). Role of lipase, from community-associated methicillin-resistant Staphylococcus aureus strain USA300, in hydrolyzing triglycerides into growth inhibitory free fatty acids. Journal of bacteriology PMID: 25225262

Wednesday, September 10, 2014

Zmapp success


       It has been a longtime, since Ebola outbreak has been a concern, for it has been the largest of date in terms of cases, fatalities and geographical spread. I have come across internet and media content designating the current situation as "race wiping", which makes no sense. I have previously talked about the known facts (Link). In this post, I wish to visit the topic again with focus on some new research and findings. As of on 31 August 2014 (WHO report), 3685 cases and 1841 deaths have been reported in the current outbreak of Ebola virus disease by the Ministries of Health of Guinea, Liberia and Sierra Leone. From the data available, the case fatality ratio appears to be about 50%.

       Of all the interventions possible, the most optimistic choice has been Zmapp cocktail. It is a trivalent monoclonal antibody combination which has not been clinically tested. However, considering the grave situation, the product was cleared for use based on compassionate use. The original study used a combination of c13C6, h-13F6, and c6D8 in the cocktail. In a recent study published in Nature, by testing several combinations the most active component was found to be c13C6. The study further tested further combinations and compared it with classic combinations. In the study, The combination of c13C6, c2G4, c4G7 was found to be the best. The most striking feature in the paper is that the therapeutic was significantly effective 3 days to 5 days post infection in a primate model. Moreover the successful use of Zmapp, in treating 2 American patients testifies to the fact.

Photo 1: Workers and authors who died
in EBOV outbreak. Link
   A high coverage genomic sequencing analysis of Ebola outbreak, has predicted that the Sierra Leone outbreak is shot from the introduction of two genetically distinct viruses from Guinea. The sad part of the story is that the workers at Kenema Government Hospital (KGH), 6 authors of the science paper, died by contracting EBOV. The study sequenced 99 Ebola virus genomes collected from 78 Ebola patients in Sierra Leone during the first 24 days of the outbreak. The sequencing had a 2000X coverage, The paper reports 395 mutations, including 50 fixed nonsynonymous changes with 8 at positions with high levels of conservation. As Stephen Gire, a research scientist in the Sabeti lab at the Broad Institute and Harvard puts it “We’ve uncovered more than 300 genetic clues about what sets this outbreak apart from previous outbreaks. Although we don’t know whether these differences are related to the severity of the current outbreak, by sharing these data with the research community, we hope to speed up our understanding of this epidemic and support global efforts to contain it.”. Source

    There has been an observation made in previous outbreaks that the survivors of Ebola infection are resistant to at least the same strain of the virus and their serum can protect fresh patients. In 1995 EBOV outbreak in Democratic Republic of the Congo, the doctors, out of option had used recover's serum, to try and save other sick patients out of desperation. Though it was met with high debate the method had seemed to help. This probably forms the basis for claiming that blood from a survivor should be treated as a priority (See Nature news here).

    Given the fact that the antibodies are protective, and with a pilot animal trial results now published, and sequences (of at least a few) in hand it appears that Zmapp is going to be the best available answer for countering the outbreak. Of course at the end the large data that would come from use of the monoclonal cocktail will help us know if this is going to be a viable option for future as well.
Qiu X, Wong G, Audet J, Bello A, Fernando L, Alimonti JB, Fausther-Bovendo H, Wei H, Aviles J, Hiatt E, Johnson A, Morton J, Swope K, Bohorov O, Bohorova N, Goodman C, Kim D, Pauly MH, Velasco J, Pettitt J, Olinger GG, Whaley K, Xu B, Strong JE, Zeitlin L, & Kobinger GP (2014). Reversion of advanced Ebola virus disease in nonhuman primates with ZMapp. Nature PMID: 25171469

Stephen K. Gire etal.(2014). Genomic surveillance elucidates Ebola virus origin and transmission during the 2014 outbreak Science DOI: 10.1126/science.1259657

Tuesday, September 02, 2014

Cytokines and Psychiatry- Few facts


    At a time when the only thing the people are interested in Ebola, it is probably very difficult to impress the fact that more is there on infectious literature. There is no doubt that at this point it is a global priority and hence rightly the interest. Leaving the trend aside, I have many a times in this post talked about the possible role of microbes in psychiatric conditions. For example, there probably is a good possibility that toxoplasma (Link), Endogenous retrovirus (Link) or other microbes (Link) can influence.

   It is been a question for me to think, how can a microbe after all influence a neurochemical process? Of course the answer doesn't look straightforward. But as far as I could gain from current evidence, in some cases at least there looks to be an immunology component influencing. There has been a number of studies more recently concentrated on studying the immunological parameters in psychiatry. The field referred to as PNI (Psychoneuroimmunology) probably will help us uncover more facts. So, I have put together a few important points in this post, that let me argue that immune cytokines and psychiatry have some relation

   In my opinion, the point that Central Nervous System (CNS), is highly protected from immunology is probably because the mediators of immune system, have tremendous cross signalling with important neural functions. It thus also makes sense to me argue that Blood Brain Barrier (BBB), strictly polices the signalling molecules. But the potential of cytokines to be involved with BBB regulation is not well studied. Recently a study published in mBio, studied the importance of PRR (Pattern recognition receptor) in context with West Nile Virus and BBB regulation. The study found that interleukins such as IL-1, can cause loss of BBB integrity and additional lymphocyte trafficking. It occurred to me if interleukin dysregulation alone could explain a lot of psychiatric condition based on following.

1. Infections in prenatal period that induced large quantities of inflammatory cytokines (especially IL-6 and IL 8) in the mother during pregnancy, especially during the 2nd trimester is a known risk factor for schizophrenia.
2. Cytokines have shown to directly influence brain developments in hippocampal regions, which is linked with schizophrenia.
3. Interleukin levels are known to effect and alter neurotransmitters especially serotonin and glutamate.
4. Patients treated with Interferon γ, for HCV infection develop significant mood disorders overtime.
5. Commonly used antidepressant drugs (reboxetine, desipramine, fluoxetine and clomipramine) show anti-inflammatory properties.
6. People administered immunomodulatory drugs with standard anti-psychotic therapy show better response.

      It is very tempting to say that dys-regulated immune function in the CNS can explain a lot of psychiatry conditions. To show how this works, let me try a very simple example. Imbalance in TH1/TH2 response is known to effect IDO (Indoleamine 2, 3-Dioxygenase), effecting the Tryptophan – Kynurenine metabolism. The final product, quinolinic acid directly antagonizes the NMDA receptor, and probably subsequent schizophrenia.

     But yes, the question still remains as to how the immune system comes to be deregulated in the first place. I don't have a good answer for that yet, but thats a good question to begin asking.
Muller, N., & J. Schwarz, M. (2010). The Role of Immune System in Schizophrenia Current Immunology Reviews, 6 (3), 213-220. PMID: 21057585

Gray SM, & Bloch MH (2012). Systematic review of proinflammatory cytokines in obsessive-compulsive disorder. Current psychiatry reports, 14 (3), 220-8 PMID: 22477442

Daniels BP, Holman DW, Cruz-Orengo L, Jujjavarapu H, Durrant DM, & Klein RS (2014). Viral Pathogen-Associated Molecular Patterns Regulate Blood-Brain Barrier Integrity via Competing Innate Cytokine Signals. mBio, 5 (5) PMID: 25161189