A study of the reactivity of S(VI)-F containing warheads with nucleophilic amino-acid side chains under physiological conditions.

Sulfonyl fluorides (SFs) have recently emerged as a promising warhead for the targeted covalent modification of proteins. Despite numerous examples of the successful deployment of SFs as covalent probe compounds, a detailed exploration of the factors influencing the stability and reactivity of SFs has not yet appeared. In this work we present an extensive study on the influence of steric and electronic factors on the reactivity and stability of the SF and related SVI-F groups. While SFs react rapidly with N-acetylcysteine, the resulting adducts were found to be unstable, rendering SFs inappropriate for the durable covalent inhibition of cysteine residues. In contrast, SFs afforded stable adducts with both N-acetyltyrosine and N-acetyllysine; furthermore, we show that the reactivity of arylsulfonyl fluorides towards these nucleophilic amino acids can be predictably modulated by adjusting the electronic properties of the warhead. These trends were largely conserved when the covalent reaction occurred within a protein binding pocket. We have also obtained a crystal structure depicting covalent modification of the catalytic lysine of a tyrosine kinase (FGFR1) by the ATP analog 5'-O-3-((fluorosulfonyl)benzoyl)adenosine (m-FSBA). Highly reactive warheads were demonstrated to be unstable with respect to hydrolysis in buffered aqueous solutions, indicating that warhead reactivity must be carefully tuned to provide optimal rates of protein modification. Our results demonstrate that the reactivity of SFs complements that of more commonly studied acrylamides, and we hope that this work spurs the rational design of novel SF-containing covalent probe compounds and inhibitors, particularly in cases where a suitably positioned cysteine residue is not present.

Antibacterial and antiviral evaluation of sulfonoquinovosyldiacylglyceride: a glycolipid isolated from Azadirachta indica leaves.

UNLABELLED: Assessment of antibacterial as well as antiherpes virus activity of sulfonoquinovosyldiacylglyceride (SQDG), a glycolipid, isolated from the leaves of Azadirachta indica has been described. Antimicrobial activity was evaluated against Gram-positive, Gram-negative bacteria and herpes simplex virus. SQDG showed significant inhibitory activity against Salmonella typhi and two isolates of Shigella dysenteriae with MIC values 32 µg ml(-1) , while three isolates of Salm. typhi, Escherichia coli and Vibrio cholerae were inhibited at 64 µg ml(-1) and have shown zone diameter ranging from 6.2 to 12.3 mm. The growth kinetics study of SQDG on Salm. typhi and Sh. dysenteriae revealed that the growths were completely inhibited at their MIC values within 24 h of exposure. Interestingly, SQDG inhibits herpes simplex virus (HSV) type 1 and 2 with the EC50 of 9.1 and 8.5 µg ml(-1) , compared with acyclovir (2.2 and 2.8 µg ml(-1) against HSV-1 and HSV-2). The selectivity index (SI) was found to be 12.4 against HSV-1 and 13.41 with HSV-2. Furthermore, the expression of proinflammatory cytokines of HSV-infected and SQDG-treated macrophages using ELISA kit revealed that SQDG significantly downregulated the production of TNF-α, IL-1ß, IL-12 and IL-6. SIGNIFICANCE AND IMPACT OF THE STUDY: The water-soluble metabolite sulfonoquinovosyldiacylglyceride (SQDG) isolated from Azadirachta indica (Neem) possess significant antibacterial as well as anti-HSV activity. The efficacies as well as the solubility factor of SQDG substantiate a greater attention for its use as phytotherapeutic drug for controlling microbial infections as most consumers have better acceptance of phytomedicines than synthetic drugs.

Evaluation of anti-infective potential of a tribal folklore Odina wodier Roxb against some selected microbes and herpes simplex virus associated with skin infection.

AIM: To evaluate the in vitro antimicrobial activity of aqueous and methanol extracts of Odina wodier bark (OWB), a folk medicine, against representative bacteria, fungi and herpes simplex virus (HSV) associated with skin infections. METHODS AND RESULTS: The OWB extract(s) was found to inhibit the isolates of Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa, Klebsiella pneumonia, Escherichia coli at an MIC of 256-5000 µg ml(-1) and Candida albicans at and above 4000 µg ml(-1) by agar and broth dilution assays. The growth curve of Staph. aureus revealed the highest activity within 2-6 h of methanol extract (ME) exposure. Interestingly, the MTT and plaque reduction assay showed that the extracts can inhibit HSV-1 and HSV-2 at EC50 of 22·4 and 28·8 µg ml(-1) , with Selectivity index of 11·7-15. While the time kinetic and binding assays demonstrated that the ME at 50 µg ml(-1) prevents viral attachment into Vero cells. Phytochemical and HPLC analysis of ME revealed the presence of flavonoids, phytosterols, saponins and tannins including the pseudotannin chlorogenic acid. CONCLUSION: The traditional use of OWB for the management of skin infections has scientific basis. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated the antimicrobial potential of OWB on selected isolates of bacteria, fungi and HSV, associated with skin infections.

Chikungunya in west bengal, India.

Fever of unknown origin broke out in several districts of West Bengal, from August 2007 to December 2007. The cases were suffering from high fever, severe joint pain lasting for several weeks after clinical cure and appearance of skin rashes. Patients' sera were collected at least five days after fever and were analyzed to detect specific IgM antibodies. A total of 800 patients were investigated and 321 (40.13%) were found to be reactive for Chikungunya antibodies. Of the patients, 66% were male. Predominant signs and symptoms observed in the sero-positive cases were fever (100%), arthralgia (96%) and diffuse erythematous skin rash (94%). Of the patients, 3% had haemorrhagic manifestations. Re-emerging Chikungunya virus spread in epidemic form in several districts of West Bengal after a gap of four decades.

Death due to chikungunya.

After its first appearance in Kolkata (Calcutta) during 1963-1965, chikungunya occurred in epidemic form in India in 17 states and union territories. There were 13,90,322 cases suspected to be suffering from chikungunya in 2006, 59,535 in 2007 and 11,222 in 2008; in 2006, 2007 and 2008 there were 15,961, 7,837 and 262 samples, respectively, sent to the National Institute of Virology, Pune, and the National Institute of Communicable Disease, Delhi, for serological diagnosis. Of these, 2001, 1826 and 44, respectively, were confirmed as chikungunya. There were no deaths (data from the National Vector Borne Disease Control Programme, Government of India).

A simple medium without blood modified for successful isolation & cultivation of LD bodies.

A simple growth medium for primary isolation and subsequent cultivation of Leishmania donovani promastigotes without using whole blood is described. This medium is modified from Aljeboori's biphasic medium (used originally only for cultivation), containing only beef extract, peptone, sodium chloride, bactoagar and foetal calf serum (FCS). We have modified the medium by adding glucose and ascertaining the pH in the solid phase and by drastically reducing (91%) FCS in the liquid phase. The medium helps in isolation of L. donovani promastigotes from kala-azar patients, in addition to luxurious growth of parasites. The medium is simple, reliable, reproducible and convenient, with minimal interference in using the parasitic cells for immunological, molecular and isoenzyme studies.

Longitudinal study on distribution of Phlebotomus argentipes sandflies at different heights in cattleshed.

Distribution of Ph. argentipes at three levels (level I, 0-0.91 m; level II, 0.91-1.82 m; and level III 1.82-2.74 m) above the ground in 6 cattlesheds was studied during July 1989 to June 1990. Of the 8044 Ph. argentipes caught, 3151 (39.2%), 3936 (48.9%) and 957 (11.9%) were from levels I, II, and III respectively. Both male and fed, half-fed, gravid and unfed female sandflies were found at all the three levels and in all the three seasons viz., rainy, winter and summer, of the year. The maximum height of occurrence of sandflies was 2.51 m (8.3 ft) above the ground level.

Efficacy & safety of glycine fortified oral rehydration solution in the treatment of acute dehydrating diarrhoea in children.

Efficacy and safety of glycine fortified oral rehydration solution (ORS) was compared with a standard ORS (WHO formula) in a randomized clinical trial in children aged between 4 months and 5 yr with moderate degree of dehydration owing to acute watery diarrhoea. No significant differences (P greater than 0.05) were observed in diarrhoeal stool output, duration of diarrhoea and intake of ORS between the study and control groups respectively. Thus glycine fortified ORS does not have any additional advantage over standard ORS. Moreover, two children developed hypernatraemia after receiving glycine fortified ORS in contrast to the control group. It is therefore suggested that glycine supplemented ORS should not be prescribed for the treatment of diarrhoeal dehydration in children.

A retrospective study on scorpion sting in a pediatric age group in a hospital in Calcutta.

During 1985-1989, in Calcutta Medical College Hospitals, of 152 children of 1-6 year age group admitted with the history of scorpion sting 18 (11.8%) died. Maximum numbers of stings were inflicted in the fingers. Important clinical features recorded were circulatory failure, breathlessness, profuse sweating, vomiting, local oedema and convulsion. Incidences of scorpion stings were much more frequent in the summer and rainy seasons than in the winter season.

Serological diagnosis of dengue in laboratory practice in Kolkata.

PURPOSE: To find out the most suitable serological investigative procedures to diagnose dengue cases effectively in the laboratory practice identifying primary and secondary cases as well as period of suffering. MATERIALS AND METHODS: Dengue suspected cases sent to the laboratory in 2012 in central Kolkata by the local physicians were categorised into seven panels according to the investigations asked for such as (1) only dengue-specific NS1 antigen (2) only IgM antibodies, (3) NS1+IgM+IgG antibodies, (4) only IgM and IgG, (5) NS1+IgM, (6) NS1+IgG and (7) only IgG. RESULTS: Out of 1892 suspected cases, dengue was diagnosed in 725 (38.3%). Through panels I, II, III, IV, V, VI and VII, it was possible to diagnose dengue in (I) 35.98% (435/1209), (II) 37.5% (24/60), (III) 49% (173/354), (IV) 30.8% (68/221), (V) 60.5% (23/38), (VI) 40% (2/5) and (VII) 0 of cases respectively. Detail information such as confirmed diagnosis, duration of the disease (whether early or prolonged) and classification of primary and secondary dengue in such early or prolonged stages would only be possible in panel III, which information would be helpful for effective monitoring and treatment of dengue patients. In all other panels, merely fragmentary information would be obtained. CONCLUSIONS: Serodiagnostic tests dengue-specific NS1 antigen and IgM and IgG antibodies when conducted simultaneously would be able to diagnose confirmed dengue cases categorising primary and secondary dengue along with the duration of the disease, whether early or prolonged.

Epidemiology of snake bite in the district of Burdwan, West Bengal.

An epidemiological field survey on snake bite was conducted on 26 randomly selected villages with a population of 18,892 in the district of Burdwan, West Bengal to assess the magnitude of the problem in a decade (1980-1989). Total number of snake bite, number of presumably poisonous snake bite and deaths due to snake bite poisoning were 307, 48 and 31 respectively. The death rate among snake bite victims was 10.09%. Males (54.72%) were bitten more than females (45.23%) and highest incidence of snake bite was found in the age group of 21-30 years and during the months of July and August. Majority of the snake bites (53%) were encountered in the lower extremities. Among the snake bite patients 201 (65.47% went to the traditional healers (ozhas) and 68 (22.14%) persons received hospital treatment, while 12 (3.09%) people neither went to the ozhas nor to hospital and 26 (8.46%) persons went to hospital after consulting the ozhas. If the present data are extrapolated for the total population of the district, average number of snake bite and death per year would be 7,857 (0.16%) and 793 (0.016%) respectively. Deaths due to snake bite per 100,000 population varied from 5.28 to 31.75 (average 16.4) over 10 years.

Estimating parasite density in patients suffering from falciparum malaria in an endemic area in Kolkata.

Parasite density of one hundred patients suffering from falciparum malaria in an endemic area in Kolkata was determined using three different methods. In the first of these, parasite density per microlitre of blood in a patient was determined using parasite count adjusted by average WBC count (i.e. 8000/microliter) observed in microscopic fields of the thick film. In the remaining two methods, only raw (i.e. unadjusted) parasite counts in microscopic fields of the same slide were used. A statistical analysis was carried out in detail to compare these methods based on raw and adjusted parasite counts and also to find out a suitable method which can be used in practice. Estimating the density of parasites is of primary importance in determining the severity of infection. Furthermore, parasite density can help in identifying short-treatment and long-treatment failure vis-a-vis detection of development of resistance in P. falciparum against the drug used. This article reports some findings that indicate the existence of a potentially dangerous situation in the study area.

Emerging Plasmodium falciparum in an endemic area in Calcutta.

Calcutta is now intensely malarious. Malarial infections are detected in every month. In an endemic area in central Calcutta a year-long study from August 1996 to July 1997 reveals that 31.47% of all cases are due to Plasmodium falciparum infection. A competition is going on between P vivax and P falciparum to gain upperhand. An equilibrium is there in the month of January (1997), when P vivax and P falciparum cases constitute 51.13% and 49.43% respectively. The highest and lowest percentage of P falciparum infection is in December 1996 (67.98%) and in May 1997 (0.89%) which is just reverse in the case of P vivax in December 1996 (33.60%) and in May 1997 (99.10%). Three imported cases of P malariae have been detected but the patients cannot be traced. Six cases of mixed infections have been found.