Castleman’s Disease In Parotid Region: A Rare Case From Chhattisgarh.

A thirty five year old patient presented with swelling in parotid region. After surgical removal and histopathological examination unicentric lymphoproliferation was observed. Further, immunohistochemical studies of CD 21 tissue marker confirmed Castleman’s Disease. The disease most commonly involves the mediastinum and neck but the involvement of the parotid gland region is very rare, being reported first time from Chhattisgarh State.

Laryngeal Myxoma: Emergency Management.

A 65-year-old male presented with stridor and dysphonia in emergency clinic of Govt. CIMS Medical College, Bilaspur, Chhattisgarh. Indirect laryngoscopic examination revealed a polypoidal lesion in glottic chink. CT scan evaluation confirmed the findings of clinical examination. Patient was relived of symptoms after emergency tracheotomy followed by surgical removal of polypoidal lesion from right vocal cord by microlaryngeal surgery. Histopathological examination revealed myxoma. Clinical examination after 8 months showed significant improvement in hoarseness of voice with no evidence of recurrence of lesion.

Comparative analysis of protein profiles of wild virulent (E156) and aroA-htrA double deletion mutant vaccine strain (S30) of Salmonella enterica subsp. enterica serovar Abortusequi under in vivo and in vitro growth conditions.

In the present study, cell lysate and cell supernatant of the both strains i.e., virulent wild type (E156) and mutant (S30) vaccine strains of Salmonella enterica subspecies enterica serovar Abortusequi (S. Abortusequi), grown under varied in vivo and in vitro conditions were subjected to SDS PAGE and western blotting (using rabbit hyperimmune serum). Variation in growth conditions did not have any significant effect on expression of different proteins. SDS PAGE of E156 and S30 cell lysate (CL) revealed 26 and 28 bands, respectively with 3 prominent proteins of 71, 46 and 42 kDa in cell lysate of E 156 and 4 prominent proteins 71, 65, 46 and 40 kDa in S30 strain. The cell supernatant (CS) from both the strains, subjected to SDS PAGE, exhibited similarity in protein profile among these strains, however three bands of 65, 53 and 40 kDa were more prominent in CS preparation of S30, whereas a 56 kDa protein was prominent in CS of E156. Western blotting of E156 and S30 revealed 3 unique proteins of 65, 53 and 40 kDa present in CS preparation of S30 strains which could be used for differentiation of mutant and wild strains and also in development of test for differentiating vaccinated animals from naturally infected.

Antigenic competition among different 'O' antigens of Salmonella enterica subspecies enterica serovars during hyperimmunization in pony mares.

The present study on antigenic competition among somatic 'O' antigens of different Salmonella groups (A, B, C1, C2, D and E1) in mares revealed that the immune response to most of the antigens was not (A, B, C2) or little (C1, D) affected by antigenic competition. However, E1 group antigen, which induced high antibody titres (Avg. 12967.3) when given alone, produced almost 3.5 log2 lower antibody titres on giving with other antigens, indicating the antigenic competition among some Salmonella group antigens. The antigenic competition varied for different antigens even of the similar chemical nature. Therefore, antigens belonging to different somatic groups should not be given together for the purpose of raising polyvalent serum or for immunization using multivalent Salmonella vaccines prepared from strains of different 'O' groups revealing antigenic competition.

Novel haemolysins of Salmonella enterica spp. enterica serovar Gallinarum.

Haemolysins of Salmonella are important due to their probable role in pathogenesis of systemic salmonellosis and use in sub-serovar level typing. The present study was undertaken to determine haemolytic potential of Salmonella Gallinarum strains through phenotypic and genotypic methods. Amplification of haemolysin gene (clyA) and cytolysin gene (slyA) was attempted in order to determine their role in haemolysin production. Study on 94 strains of S. Gallinarum revealed the production of two types of haemolysis viz., beneath the colony haemolysis (BCH) or contact haemolysis and clear zone haemolysis (CZH). Haemolysis was observed on blood agar prepared with blood of cattle, buffalo, sheep, goat, horse, rabbit, guinea pig, fowl, and human blood group A, B, AB and O. Although, haemolysis was also observed on blood agar prepared with whole blood, clarity of zone was more evident on blood agar made from washed erythrocytes. Clear zone haemolysis was best observed on blood agar prepared with washed erythrocytes of goat and a total of 12% (11 of 94) S. Gallinarum strains under study produced CZH on it. The clyA gene could not be detected in any of the 94 strains under study, while slyA gene could be amplified uniformly irrespective of haemolytic potential (CZH) and haemolytic pattern (BCH) of the strains. The study suggested that the two types of haemolysis (CZH and BCH) observed among S. Gallinarum strains may not be due to either slyA or clyA gene products and thus there may be some other gene responsible for haemolytic trait in Gallinarum serovar. Different haemolytic patterns of strains under study indicated multiplicity of haemolysins in S. Gallinarum.

Pathogenic effects of Salmonella enterica subspecies enterica serovar Typhimurium on sprouting and growth of maize.

The study was undertaken to understand effects and survival of S. enterica subspecies enterica serovar Typhimurium (S. Typhimurium), a zoonotic serovar, on maize seed germination and plant growth. All the four strains of S. enterica subspecies enterica serovar Typhimurium significantly reduced germination of maize seeds in sprouting plates as well as in soil. About > or =2.7x10(3) Salmonella cfu ml(-1) of soaking water, while > or =2.7x10(7) Salmonella cfu g(-1) soil were required to significantly inhibit germination of maize. Similar inhibition of germination could be observed using > or = 16 mg of bacteria free Salmonella cell lysate (CL) protein per g of soil or > or =0.5 mg of CL protein per ml of soaking water in sprouting plates. At the constant dose of 3.6x10(7) to 3.8x10(7) Salmonella cfu or 5 mg cell lysate protein ml(-1) of soaking water, four strains of Salmonella significantly reduced germination, however difference between strains was insignificant. After germination too, maize growth was affected both by Salmonella organism and CL with little strain-to-strain variation. All Salmonella persisted in growing plants from 15 to 35 days of plant age and up to 190 days in soil. Maize plants once grown for a week in sterile soil were resistant to invasion of S. enterica subspecies enterica serovar Typhimurium in their leaves even in doses as high as 7.6x10(9) cfu g(-1) of soil. Salmonella persisted better and longer in plants grown from contaminated seed sown in loam soil, but rarely in plants grew in sandy soil. All maize plants had Salmonella in their stumps even after 35 days of sowing irrespective of kind of soil, primary source of infection (soil or seed) and type of S. enterica subspecies enterica serovar Typhimurium strain. The study revealed that Salmonella is not only zoonotic but a phytopathogen also.

Haemolysins of Salmonella, their role in pathogenesis and subtyping of Salmonella serovars.

Haemolysin patterns of 175 strains of different Salmonella enterica subspecies enterica serovars isolated from different animal sources and places were determined using 11 different blood agar media made with either non-washed horse/sheep erythrocytes or with washed erythrocytes of cattle, sheep, horse, goat, rabbit, guinea pig, and human A, O and B blood groups. Study on 47 strains belonging to 10 serovars of Salmonella from buffalo meat (buffen), 42 strains of 11 serovars from goat meat (chevon): 16 strains of Salmonella enterica serovar Paratyphi B and 25 of S. enterica serovar Paratyphi B var Java from fish, meat, meat products and clinical cases; 45 isolates of S. Abortusequi from aborted mares (18), fetal contents (21), aborted donkey mares (2) and 4 reference strains, revealed that all host restricted Salmonella namely, S. enterica serovar Gallinarum, S. enterica serovar Anatum, S. enterica serovar Abortusequi and S. enterica serovar Paratyphi B could be divided into different haemolysin types based on their inability to produce haemolysis on one or more types of blood agar, while strains of all zoonotic Salmonella serovars induced haemolysis on all the 9 types of blood agar made of washed erythrocytes. None of 175 Salmonella could produce hemolytic colonies on blood agar made of non-washed horse/ sheep erythrocytes. Haemolysin type I (lysing all types of washed erythrocytes) was the commonest one among all serovars except S. Abortusequi, none of which lysed horse erythrocytes. Salmonella enterica serovar Abortusequi having hemolytic activity against sheep erythrocytes were more invasive but had lesser ability to survive in sheep mononuclear cells than non-hemolytic strains. Multiplicity of haemolysins appeared significant epidemiological tool.

Evaluation of guinea pig model for experimental Salmonella serovar Abortusequi infection in reference to infertility.

The present study conclusively revealed the role for Salmonella enterica subspecies enterica serovar Abortusequi in conception failure. None of the 12 guinea pigs conceived when orally exposed to sublethal dose of the pathogen during breeding, while 66.67% of animals in control group were found pregnant during same period of observation under similar conditions. Salmonella carrier animals also had drastic reduction in conception rate (16.67%). During mid pregnancy, S. Abortusequi exposure to guinea pigs through intravaginal, intramuscular and subcutaneous routes induced fetal death followed by resorption. While 2 out of 6 orally inoculated and 3 out of 6 intraperitonially inoculated guinea pigs aborted, in rest of the animals fetal death was followed by meceration and resorption. It was interesting to note that S. Abortusequi could not persist longer than a week in males while in pregnant females it could be detected for >10 weeks after inoculation. In late pregnancy, most of the exposed animals aborted and non aborting animals though had normal parturition, survival rate of their babies was nearly zero in comparison to the control group. The study revealed role for S. Abortusequi in impairing conception, abortion, early fetal deaths, fetal meceration and resorption. Further studies are required to identify factors responsible for increased susceptibility of females particularly during pregnancy.

Isolation and characterization of Salmonella Gallinarum cytotoxic factors.

Two distinct cytotoxic factors isolated from a Salmonella Gallinarum strain recovered from a bird died during an outbreak of fowl typhoid were purified to homogeneity through ciprofloxacin extraction, salt precipitation, dialysis, gelfiltration, ionexchange chromatography and chromatofocusing. These were designated as Salmonella Gallinarum cytotoxin I (GCT-I) and II (GCT-II). GCT-I was a glycoprotein having mol.wt and pI of Ca 70 kDa and 8.8, respectively. It was lethal to birds (LD50, 150 micrograms) inducing fowl typhoid like lesions. GCT-II, a protein with Ca 55 kDa mol.wt., was not lethal but caused haemorrhagic diarrhoea on intraperitoneal inoculation in birds. Both the cytotoxins induced cytopathic effects (CPE) in Vero and Madin Darby bovine kidney (MDBK) cells, enterotoxicity in rabbit ileal loop, dermatotoxicity in the rabbit skin and specific neutralizing antibodies in rabbits. These were active only between a narrow pH range of 6 to 8.5 and thermostable at 90 degrees C (1 min) but lost their activities on boiling. Trypsin and chymotrypsin enhanced their cytotoxicity, while pepsin, papain, protease, lipase and urea (5 M) had no appreciable effect on their cytotoxicity. Sodium carbonate (0.05 M) and formaldehyde (0.05%) had no effect on antigenicity of both the cytotoxic factors but rendered them nontoxic. Identification and characterization of cytotoxic moieties of S. Gallinarum not only reveals the important virulence factor but also indicates about the use of toxic factors as a candidate for toxoid vaccine and immunodiagnostics.

Detection, prevalence, purification and characterization of lecithinase of Klebsiella pneumoniae.

Lecithinase activity in Klebsiella is a rare trait as out of 208 strains of Klebsiella belonging to 3 species, viz. K. pneumoniae (168), K. planticola (29) and K. oxytoca (11), only 4 strains of K. pneumoniae produced lecithinase positive colonies on egg-yolk-agar. Although cell lysates of 16 K. pneumoniae yielded positive results for lecithinase assay on egg-yolk-agar, 19 strains were detected positive for lecithinase with ELISA using anti-lecithinase serum. Release of up to 52.12% cell-bound lecithinase could be achieved with polymyxin-B treatment at 100 micrograms/ml concentration. Purified lecithinase was determined to be a high molecular weight (70 kDa), crystalizable, anionic (pI, 3.5) protein. It possessed cytolytic, haemolytic and dermonecrotic activities but did not induce fluid accumulation in rabbit ileal loop or infant mouse guts. It was inactivated by boiling, trypsin and chymotrypsin treatment and alkaline pH. Serologically, it was related to lecithinase of Aeromonas caviae and phospholipase-C of Salmonella.

Purification and characterization of Klebsiella pneumoniae cytotoxins.

Isolation, purification and characterization of 3 new cytotoxins of a K. pneumoniae strain isolated from ready to eat pork sausage are reported. Purification process involved extraction of cytotoxins with polymyxin B sulphate, salt precipitation, gel filtration and anion exchange chromatography. Klebsiella cytotoxin (KCT) I, a glycoprotein of about 65 kDa was verocytotoxic, enterotoxic and dermonerotic. KCT II was erythemogenic, verocytotoxic and enterotoxic protein of co 55 kDa, while KCT III was about double in MW (110 kDa) hadverocytotoxicity but neither enterotoxicity nor dermatotoxicity. KCT I and II caused granulation, conglomeration, shrinkage, detachment and lysis of MDBK and Vero cells, while KCT III induced enlargement, vacuolation, granulation, multinucleolation and syncytia formation in exposed cells. All the three cytotoxins induced specific neutralizing antibodies and cytotoxins were detectable in nanogram quantities with enzyme-linked immunosorbant assay using homologous antibodies. None of the anticytotoxin cross-reacted with either heterologous Klebsiella cytotoxins or with verocytotoxic preparations of Shigella dysenteriae.

Purification and characterization of phospholipase C of Salmonella gallinarum.

Phospholipase C was isolated from an outbreak strain of Salmonella gallinarum with ciprofloxacin extraction, dialysis, gel filtration, ion exchange chromatography and chromatofocussing. Purified phospholipase C (mol wt. 65 KDa; isoelectric point, pI 3.5) was resistant to pasteurization, stomach enzyme (pepsin), bacterial protease and lipase but lost its activity on trypsin and chymotrypsin treatment. It was sensitive to pH > or = 8.0. It was haemolytic, embryotoxic, enterohaemorrhagic, lethal to birds, cytotoxic to Vero and MDBK cells, dermonecrotoxic in rabbit and antigenically active protein. Antisera raised against purified phospholipase C neutralized its all biological activities and agglutinated the producer Salmonella strains. Serologically it was proved similar to phospholipase C of Klebsiella pneumoniae and S. weltevreden. Fluorescent antibody technique (FAT) was standardized to detect phospholipase producer strains.

Coagglutination test: a simple and rapid immunodiagnostic test for parvovirus infection in dogs.

The coagglutination test (COAT) was developed and standardized to detect canine parvovirus (CPV) antigen in faeces of infected dogs. Anti-parvovirus serum was raised in dogs for coating protein-A containing Staphylococcus aureus Cowan I strain. Agglutination of antibody coated bacteria invariably occurred within 2-3 min when mixed with standard CPV antigen or faecal supernatants of dogs having 8 or more haemagglutination (HA) titre of parvovirus antigen. The test had a perfect correlation with HA test and was found to be slightly more sensitive than agar gel precipitation test (AGPT) in detecting CPV antigens. As COAT is easy and needs no specific equipment or much technical know how to perform, it can be used as a field test for rapid clinical diagnosis of parvovirus infection in dogs.

Toxigenicity of Edwardsiella tarda isolates of fish and pig origin in experimental models.

Indian isolates of E. tarda from fish (7) and pigs (2) were examined for their enterotoxigenicity/cytotoxigenicity in rabbit ligated ileal loop (RLIL), suckling mouse, rabbit skin and Vero cell monolayers. Cell free culture filtrates (CFCF) of isolates from fish and pigs, induced blood tinged fluid accumulation in RLIL, increased vasopermeability in rabbit skin, and caused cytopathic effect in Vero cells but could not induce fluid accumulation in suckling mouse. CFCFs lost their activity on heating at 63 degrees C for 30 min or 72 degrees C for 15 sec, and also at pH < or = 4.5 or > or = 8.5. The toxic factor was released extracellularly and was nondializable.

Survivability of Salmonella paratyphi B var Java on experimentally infected cockroaches.

Cockroaches are inhabitant of sewers and frequent visitors of kitchen and stores in the night to feed on left over. It is liable to disseminate a number of pathogens by contaminating kitchen surface, feeding vessels and food items left open. Salmonella paratyphi B var Java a common pathogen of man and animal was used in the study to evaluate its survival and excretion in cockroaches. The host when fed on semisolid feed containing 1 x 10(7) CFU of S. paratyphi B var Java g-1, it was found that the pathogen was eliminated earlier from the live than euthanized cockroaches.