Immunoproteomic analysis of Clostridium botulinum type B secretome for identification of immunogenic proteins against botulism.

OBJECTIVES: To identify immunogenic proteins of C. botulinum type B secretome by immunoproteomic analysis. RESULTS: In the present study, an attempt was made to elucidate the vaccine candidates/diagnostic molecules against botulism using immuno proteomic approach. C. botulinum type B secretome was elucidated when it was grown in TPGY as well as CMM media. Predominant 51 proteins were identified in both the media using 2-DE and mass spectrometry analysis. 2D gels (CMM & TPGY) were probed with respected proteins mice antiserum and obtained 17 and 10 immunogenic proteins in TPGY as well as CMM media respectively. Hypothetical protein CLOSPO_00563, ornithine carbamoyl transferase, FlaA, molecular chaperone GroEL and secreted protease proteins were found as the common immuno dominant proteins in both media. Polyclonal Antibodies raised against C. botulinum types A and E showed cross-reactivity with secretome C. botulinum type B at the lowest dilution (1:1000) but did not show cross reactivity with highest dilution (1:30,000) with C. botulinum type B secretome. Polyclonal antibodies against C. botulinum type F secretome did not show cross reactivity with C. botulinum type B secretome. CONCLUSIONS: Identified immunogenic proteins can be used as vaccine candidates and diagnostic markers for the infant and wound botulism but common immunogenic proteins may be the best vaccine candidate molecule for development of vaccine as well as diagnostic system against the infant and wound botulism.

Multiple-reaction monitoring for multiplex detection of three bacterial toxins using liquid chromatography-tandem mass spectrometry.

Clostridium perfringens epsilon toxin, staphylococcal enterotoxin B and shiga toxin are implicated in a number of diseases and food-borne intoxications and are considered potential agents for bioterrorism and warfare. Artificially generated aerosol is the likely mode of delivery of these for nefarious uses, potentially capable of causing mass destruction to human and animal health by inhalation of toxic bioaerosol. Multiplex and unambiguous detection of these agents is of paramount importance for emergency response in a biothreat scenario and for food safety. Multiple-reaction monitoring (MRM) assay for simultaneous monitoring of the three toxins is reported here using reverse-phase high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry. Three different peptides with two fragment ions each were considered for quantification and confirmation. One of the three MRM transitions from each toxin, which exhibited the best sensitivity, was selected for multiplexing of the assay. Simulating a biothreat scenario wherein the bioaerosol is collected in 10 ml of buffer, the multiplex assay was tested with blind samples with one or more of the three toxins even in the presence of interfering Escherichia coli lysate proteins.

Right Atrial Myxoma: An Uncommon Presentation.

There are two types of tumors found in the cardiac chamber. These are divided into primary intra-cardiac tumors and secondary intra-cardiac tumors. Primary intra cardiac tumors are rare and among them 29% are myxomas. Majority of them are found in the left atrium. Here, we report a case of a myxoma in the right atrium with hepatomegaly and Hepatitis B virus infection. The coexistence of all these conditions is very rare. A 52 years old patient presented with history of shortness of breath on exertion along with fever and generalized weakness for 6 months which aggravated lately for last 2 months. He was then taken for better medical care and hospitalization. On cardiac evaluation he had soft S1 and S2 over the tricuspid region on the right lower parasternal region. He had bilateral mild pitting pedal edema. On further examination, it was revealed that he had mild tender hepatomegaly with jaundice. His blood analysis for HBsAg was positive. Echocardiogram showed right atrial myxoma of 14.3cm² almost completely occupying the right atrium and even protruding into the Inferior Venacava however not fully obstructing it. The inferior vena cava size was mildly dilated (22mm). Abdominal ultrasound report showed hepatomegaly (17.6cm) with coarse hepatic parenchyma. In this report, we emphasize the rarity of myxoma in the Right Atrium, its difficult diagnosis because of the location and the atypical presentation in the echocardiograph.

Cardiofacial Syndrome: A Rare Case Report.

Cardiofacial syndrome is associated with facial abnormality with congenital heart disease. Here, we report a case of cardiofacial syndrome having anotia and facial nerve palsy on the right side in combination with infundibular pulmonary stenosis and patent ductus arteriosus which is a rare presentation of cardiofacial syndrome. A 6 years old girl presented to department of Cardiac surgery of Bangabandhu Sheikh Mujib Medical University, Dhaka, Bangladesh with the complaints of shortness of breath on exertion since 2 years of age. Her physical examination revealed right sided anotia and facial nerve palsy associated harsh ejection systolic murmur in upper left parasternal area. Echocardiography showed critical infundibular pulmonary stenosis with a small patent ductus arteriosus. She underwent ligation of patent ductus arteriosus and excision of infundibular muscular bands and discharged to home without any complication. Any child presented with facial abnormality should be checked for any cardiac abnormality for early intervention and better management of the patient.

Distribution Pattern and Outcome of the Patients in the Intensive Care Unit of Bangabandhu Sheikh Mujib Medical University: A Short Term Analysis.

The aim of the study was to describe the characteristics of patients admitted to intensive care unit and their outcome in Bangabandhu Sheikh Mujib Medical University, Bangladesh. This retrospective, descriptive study was conducted in the intensive care unit of Bangabandhu Sheikh Mujib Medical University (BSMMU) from January 2016 to June 2016. Data was retrieved from hospital records of all admitted patients regarding age, gender, admission source, reason for admission, length of ICU stay, requirement of mechanical ventilation, number of organ failure and their outcome. During this study period, the total number of patients admitted was 225; most of them were males (148, 65.7%). Among the 225 patients, the highest number of admission was comprised of intracranial haemorrhage (22.6%), followed by sepsis (12.4%), acute respiratory distress syndrome (10.2%), acute renal failure (9.3%) malignancy (8.8%) and ischemic stroke (8.0%). Mean age of the patient was 54±18 years and mean length of ICU stay was 6.8±3 days. Out of 225 patients, 87 expired (38.6%). Majority of the patients required mechanical ventilation (69.3%) and had multi organ failure (59.8%). Most of the expiries were due to intracranial haemorrhage (24.1%); followed by acute respiratory distress syndrome (12.6%), malignancy (12.6%) and sepsis (11.4%). Elderly age (>65 years), requirement of mechanical ventilation and multiorgan failure had significant relationship (p<0.05) with overall ICU mortality. Intracranial haemorrhage, sepsis, acute respiratory distress syndrome were the main reasons for admissions in ICU, while mortality was highest for intracranial haemorrhage. Developing a well equipped neurological ICU with adequately trained staff will help to improve the outcome of patients.

Development of an immunodetection test for a botulinum-like neurotoxin produced by Clostridium sp. RKD.

BACKGROUND & OBJECTIVES: Clostridial neurotoxins are among the most toxic substances known and cause severe illnesses in both humans and animals. A neurotoxigenic Clostridium sp. (strain RKD) isolated from intestine of decaying fish produced a novel, botulinum type B like neurotoxin as suggested by mouse bioassay, protection with anti-botulinum antibodies and PCR. The aim of the present investigation was to develop a laboratory based detection assay as an alternative to the mouse bioassay without compromising sensitivity and specificity. METHODS: Growth and toxin production were carried out in trypticase peptone yeast-extract glucose (TPYG) broth. Toxicity was estimated in terms of minimum lethal dose (MLD) by mouse bioassay. The toxin was partially purified by acid precipitation. It was used for toxoid preparation by formaldehyde treatment. This purified IgG was used for detection of neurotoxin using indirect ELISA. The culture supernatant was concentrated using a stirred cell with a 50 kDa cut-off membrane at 4 degrees C. Further purification was carried out using Prep cell. Fractions showing toxicity and sufficient purity were pooled, concentrated and analyzed on sodium dodecyl sulphatepolyacrylamide gel electrophoresis (SDS-PAGE). RESULTS: The toxin was purified with a recovery of 8.56 per cent. Polyclonal antiserum was raised in mice using partially purified toxin with a titre of 1: 80000. A detection assay with sensitivity of approximately 15 and 300 ng/ml for partially purified and crude toxins, respectively were achieved using an indirect ELISA method. INTERPRETATION & CONCLUSION: The Clostridium sp. RKD produced a potent neurotoxin earlier shown to have novelties. A specific detection assay for the neurotoxin has been developed that may be useful both from food safety and clinical point of view.

Psychrophilic Planococcus maitriensis sp.nov. from Antarctica.

An orange pigmented bacterium, S1, was isolated from a cyanobacterial mat sample collected in the vicinity of Schirmacher Oasis, Maitri, the Indian station, in Antarctica. The bacterium is Gram-positive and possesses all the characteristics of the genus Planococcus. It is non-sporulating, motile and has A4alpha type peptidoglycan, MK-7 and MK-8 as the major menaquinones and anteiso-C(15:0) as the major fatty acid. Based on the phylogenetic characteristics, the bacterium S1 is identified as a close relative of Planococcus citreus with which it shares 98.12% similarity at the 16S rRNA gene level but exhibits a low similarity of 52% at the whole genome level. Apart from the above major differences, S1 also exhibits phenotypic differences with Planococcus citreus and other members of the genus Planococcus. Based on these differences, the bacterium S1 is identified as a new species of the genus Planococcus for which the name Planococcus maitriensis is proposed. The type strain of Planococcus maitriensis is S1(T) (= MTCC 4827; DSM 15305).

Glycocalyx positive bacteria isolated from chronic osteomyelitis and septic arthritis.

Bacterial cultures isolated from cases of chronic osteomyelitis and septic arthritis were screened for the production of glycocalyx. The presence of glycocalyx was noted in 76.3% of Staphylococcus aureus, 57.14% of Staphylococcus epidermidis, 50% of Pseudomonas aeruginosa, and 75% of Escherichia coli isolates.

Molecular characterization of 'Bhut Jolokia' the hottest chilli.

The northeast region of India, considered as 'hot spot' of biodiversity, having unique ecological environment with hot and high-humidity conditions, has given rise to the world's hottest chilli, 'Bhut Jolokia', which is at least two times hotter than Red Savina Habanero in terms of Scoville heat units (SHU). This study was undertaken to determine the distinctiveness of 'Bhut Jolokia' from Capsicum frutescens or Capsicum chinense through sequencing of the ribosomal RNA (rRNA) gene-internal transcribed ((ITS) region along with its phylogenetic analysis. Although a compensatory base change (CBC) in the ITS2 region was not observed between the closely related species of C. frutescens and C. chinense when compared with Bhut Jolokia; phylogenetic analysis using ITS1, 5.8S and ITS2 sequences indicated a distinct clade for all the accessions of 'Bhut Joloikia', while C. frutescens and C. chinense occupied discrete lineages. Further, a unique 13-base deletion was observed in all the representative accessions of 'Bhut Jolokia', making it distinct from all other members within the genus and beyond. The degree of genetic variations along with its extreme pungency might be related to ambient environmental factors of northeastern India.

Extracellular protease from Bacillus coagulans, a psychrotrophic, Antarctic bacterium.

Bacillus coagulans, when grown on casein at 20°C, produced an inducible, metalloprotease of 28 kDa at 1.6 U/mg cell protein. (NH4)2SO4 at 2 g/l decreased enzyme production irrespective of carbon source.