Co-existence of Pseudomonas-derived cephalosporinase among plasmid encoded CMY-2 harbouring isolates of Pseudomonas aeruginosa in north India.

Context: In Pseudomonas aeruginosa, AmpC β-lactamases are often responsible for high-level resistance to β-lactam antibiotics. The co-production of plasmid-mediated AmpC along with chromosomal Pseudomonas-derived cephalosporinases thus remain a serious clinical concern owing to high resistance spectrum towards antibiotics. Aim: The present study was performed to investigate the co-existence of both chromosomally-encoded and plasmid-mediated AmpC β-lactamase among clinical isolates of P. aeruginosa. Setting and Design: It is a cross-sectional study carried out in the Department of Microbiology in a tertiary referral hospital of northern India. Methods and Methods: A total of 329 consecutive, non-duplicate clinical isolates of P. aeruginosa, were selected for the detection of AmpC β-lactamases and confirmed for AmpC production by modified three dimensional (M3D) test. Ceftazidime -imipenem antagonism test was used to detect inducible AmpC producers. Molecular characterisation of chromosomally-encoded blaPDC and plasmid-mediated AmpC gene was studied by performing polymerase chain reaction (PCR). Result: A total of 214 (65%) isolates were confirmed for AmpC production by M3D test. On performing multiplex PCR, 27 isolates were detected posessing blaCMY type of plasmid-mediated AmpC gene. While 48 isolates were found to harbour chromosomally-encoded blaPDC gene co-production of both chromosomal and plasmid-encoded AmpC was reported in eleven isolates. Conclusions: Although these chromosomally-encoded cephalosporinases might spread more slowly than mobilised AmpC, but it is likely that in the present scenario of intense antibiotic pressure, this will become an increasing problem and may further limit our antibiotic choices.

Observation on integron carriage among clinical isolates of Klebsiella pneumoniae producing extended-spectrum β-lactamases.

Purpose: Klebsiella pneumoniae is considered an important pathogen causing nosocomial and community-acquired infections and is often associated with the production of extended-spectrum β-lactamases (ESBL) belonging to SHV and CTX-M families, which are frequently described as a part of complex integrons, facilitate their horizontal transfer to other related as well as unrelated microbes. The present study was undertaken to investigate the occurrence and characterization of integrons among K pneumoniae isolates producing ESBL in a tertiary referral hospital. Materials and Methods: A total of 136 clinical isolates of K pneumoniae were investigated for the presence of ESBL. Their ESBL genes were characterized by multiplex polymerase chain reaction (PCR). Integrase gene PCR was performed to detect the presence of integron. The isolates were further typed by random amplification of polymorphic DNA (RAPD). Result: Out of 136 K pneumoniae isolates, 63 (46%) were confirmed to be ESBL producers. SHV (68%) and CTX-M (67%) ESBL genes were the most common in our study. Of the 63 ESBL-positive isolates, 58 (92%) strains carried integrons; 52 strains (82%) carried only class 1 integron, whereas 6 (9%) isolates harboured both class 2 integrons and the class 1 gene. However, in ESBL negatives, only 29 (40%) strains were positive for class 1 integron and none for class 2 integron. Conclusion: The presence of class 2 integron amongst ESBL-producing K pneumoniae is being described for the first time in this part of the world. The findings of this study strongly suggest that integrons have a role in the dissemination of ESBL-mediated resistance among the nosocomial isolates of K pneumonia.

Increased prevalence of extended spectrum beta lactamase producers in neonatal septicaemic cases at a tertiary referral hospital.

Emergence of extended spectrum beta lactamases (ESBLs) producing strains of gram negative bacteria, as one of the leading cause of septicaemia often complicates the clinical and therapeutic outcome. The present study was undertaken to investigate the prevalence of ESBLs in bacteria isolated from neonatal septicaemic cases along with their antimicrobial sensitivity pattern. Blood samples were collected from 243 suspected cases of neonatal septicaemia. Apart from susceptibility testing, all the gram negative isolates were subjected to phenotypic tests for ESBL production. Amongst the positive test samples (n = 115), 84 were gram negative rods. ESBL was detected in 26 (32%) isolates. Results indicate that routine ESBL detection should be made imperative and empirical use of third generation cephalosporins must be discouraged.

Evaluation of different tests for detection of Staphylococcus aureus using coagulase (coa) gene PCR as the gold standard.

A total of 288 staphylococcal specimens isolated from different clinical specimens were selected for the evaluation of tests used to detect Staphylococcus aureus. The coagulase (coa) gene PCR was preformed, which confirmed 288 specimens as S. aureus and 51 specimens as coagulase negative staphylococci (CONS). All the specimens were subjected to slide coagulase test, Slidex Staph plus test and tube coagulase test. Sensitivity, specificity, positive predictive value and negative predictive values of were calculated using coa gene PCR as gold standard for the detection ofS. aureus. The tube coagulase test showed very good sensitivity (98.7%), specificity (98.1%), PPV (99.5%) and NPV (94.4%) than other methods. Slidex Staph plus test showed fairly good sensitivity and specificity. Slide coagulase test has good specificity but poor sensitivity. Therefore we recommend that tube coagulase test be done routinely for the detection ofS. aureus in microbiology laboratory.

Antimicrobial resistance profile of bacterial isolates from Intensive Care Unit: changing trends.

The incidence of antimicrobial resistance has increased over the years resulting in limitation of therapeutic options. Strategies such as appropriate infection control measures and surveillance of resistance pattern are necessary to address the problem of resistance. Knowledge of the pattern of resistance in Intensive Care Units (ICUs) can help to determine antibiotic prescribing policy. A retrospective study has been carried out to determine the bacterial spectrum and the antibiotic resistance pattern of clinical isolates collected from patients admitted to the ICU. The data was compared with a similar study conducted during 1996-97. Amongst the gram-positive organisms Staphylococcus aureus (23%) was the predominant isolate, while Pseudomonas (23%), Acinetobacter (20.8%), Citrobacter (11.7%) accounted for the majority of the gram-negative organisms. Both gram positive and gram-negative organisms exhibited high resistance to most antimicrobial agents used for testing susceptibility. The frequency of resistance has markedly increased as compared to the previous study.

Prevalence of methicillin resistant staphylococcus aureus in a tertiary referral hospital in eastern Uttar Pradesh.

We report the prevalence of methicillin resistant Staphylococcus aureus (MRSA) infections and their antibiotic susceptibility pattern in our hospital located in eastern Uttar Pradesh. Out of total 549 strains of Staphylococcus aureus isolated from different clinical specimens 301 (54.85%) were found to be methicillin resistant. More than 80% of MRSA were found to be resistant to penicillin, cotrimoxazole, ciprofloxacin, gentamicin, erythromycin, tetracycline, 60.5% to amikacin and 47.5% to netilmicin. However, no strains were resistant to vancomycin. Many MRSA strains (32.0%) were multi-drug resistant. To reduce the prevalence of MRSA, the regular surveillance of hospital associated infection, monitoring of antibiotic sensitivity pattern and formulation of definite antibiotic policy may be helpful.

Antibiotic sensitivity pattern of the bacteria isolated from nosocomial infections in ICU.

The incidence of nosocomial infections in ICU is 4-5 times greater than in general ward. Critically ill patients are always at higher risk of developing nosocomial infections with resistant strains. This study is an attempt to know the antibiotic sensitivity pattern of the common isolates in ICU. Samples comprising urine, blood, endotracheal secretions and throat swabs were collected from 102 ICU patients of them, 56 patients showed evidence of nosocomial infection (54.9%), from whom 120 different organisms were isolated. Antibiotic sensitivity test was done according to Kirby Bauer method. Klebsiella pneumoniae were the most prevalent isolates from respiratory tract infections followed by Proteus spp, Escherichia coli, Staphylococci spp. and Acinetobacter spp. The gram negative enteric bacilli were uniformly resistant to betalactam antibiotics as well as betalactam-betalactamase inhibitors. Resistance to Ciprofloxacin and Ceftriaxone ranged from 50-100% and 25-83.3% respectively. Staphylococci were 100% resistant to penicillin and tetracycline, 80% to cotrimoxazole, 60% to erythromycin and gentamicin and 40% to amikacin. Acinetobacter spp. were highly resistant to most of the antibacterial agents except gentamicin while Pseudomonas spp. showed 75% resistance to it. The increased prevalence of resistant organisms in ICU probably reflects lack of proper antibiotic policy resulting in prolonged and indiscriminate use of antimicrobial agent.

Immunohistopathological reactions for liver-specific membrane lipo-protein in experimental autoimmune hepatitis.

Antibody to the hepatocyte membrane protein, was induced in inbred strain C57BL/6 and C3H mice by immunisation with 100,000 g supernatant of syngeneic liver homogenate in CFA. Three weekly intraperitoneal injection of 200 ul of liver homogenate with CFA for continuous 4 weeks gave the best possible result. Histopathological changes were characterised mainly by perivascular inflammatory infiltrates and hepatocyte necrosis which mimicked human autoimmune hepatitis. In one of the immunological parameters, antibody to hepatocyte membrane protein (LSP) has been demonstrate by ouchterlony method in the test serum of those animals, who had received weekly doses of liver antigen. Thus in experimental autoimmune liver disease, semi-purified syngeneic liver fluid (S-100) leads to hepatic destruction and to an inflammatory process with several features in common with human chronic aggressive hepatitis. The presence of antibody against syngeneic liver antigen (S-100) in the test sera emphasizes that hepatocyte membrane protein does have an important role in liver tissue pathogenesis and disease process in experimental model. In this study we tried to prove that hepatocyte membrane protein may act as a target antigen in developing experimental autoimmune hepatitis.

Antiovarian antibody in premature ovarian failure.

Premature ovarian failure is a syndrome consisting of primary or secondary amenorrhoea, hypergonodotropiremia and hypoestrogenemia in women under the age of 40. An autoimmune mechanism was suggested as possible etiology when Vallolton and Forbes in 1966-67 found antibodies to the cytoplasm of rabbit ova in 29 of 232 tested sera. Immune mechanism in the pathogenesis of premature ovarian failure (POF) is suggested by association of autoimmune phenomenon with POF in some cases and demonstration of circulating antibodies to ovary in serum samples from women with POF. The incidence of presence of antiovarian antibody of POF patients has been reported earlier. Evidence of autoimmunity is present in 18-92% of patients with POF. In the present study we have studied 18 cases of POF without any overt manifestation of autoimmune disorder but the antiovarian antibody was detected, with the idea that this autoantibody might be the cause of ovarian dysfunction which is evident in POF. Presence of antiovarian antibody in 16.67% cases with POF in our study that ovarian antibodies may play a role in or reflect an autoimmune process responsible for the development of POF.

Circulating immune complex in murine autoimmune hepatitis.

High level of circulating immune complexes (CIC) in the serum has been reported in different forms of hepatitis particularly in complicated cases of viral hepatitis due to hepatitis B virus (HBV) infection. In this study CIC level in experimental autoimmune hepatitis were assessed by detection of polyethylene glycol (PEG) index. The sera of mice with established autoimmune hepatitis (EAH) confirmed by histopathological study showed higher PEG index (C57BL/6 mice: 34.56 +/- 6.28 and C3H mice: 31.95 +/- 28.99). The control healthy mice showed lower PEG index (C57BL/6 mice: 19.48 +/- 6.85 and C3H mice: 21.27 +/- 6.1). The high level of PEG index in EAH was found statistically significant. The role of CIC in the development of autoimmune hepatitis is discussed.

Immunoglobulin G and complement C3 levels in pregnancy induced hypertension.

The immunoglobulin G (IgG) and complement C3 (C3) were measured in the maternal as well cord blood sera of 30 cases of pregnancy induced hypertension (PIH) as well as 9 controls with normotensive pregnancy. A depression of IgG as well as C3 level was observed in the maternal as well as cord sera of the mothers with PIH. These findings suggest decreased immunological status of both mother and her offspring in PIH, irrespective of the gestation and intrauterine growth status.

Antitoxoplasma antibody in clinically suspected cases of human toxoplasmosis.

(1) Seventy sera from a variety of patients suffering from different diseases suspected to be caused by Toxoplasma gondii infection and forty from non-toxoplasmic hospital cases and laboratory and hospital staff were collected. (2) Antitoxoplasma antibody was detected in those sera by Indirect Haemagglutination test (IHA test) and Enzyme Linked Immunosorbent Assay (ELISA). (3) Twenty one (30%) sera out of 70 test samples and 2 (5%) out of 40 control samples were positive by ELISA test. With IHA test only 17 (24.3%) of test samples and same 2 (5%) of control samples were positive. (4) Sera collected from Paediatric Department showed the highest positivity (40%) followed by Opthalmological group (35.7%) and obstetrics and Gynaecological group (13.6%). (5) No significant co-relation was found between the seropositivity with sex, diet and history of cat contact of the patients.

Effect of exogenous 5-hydroxytryptamine on pathogenicity of Entamoeba histolytica in experimental animals.

In an effort to find out the mechanism(s) operative in enhancing the pathogenicity of E. histolytica in hosts under heat stress reported earlier, effect of 5-hydroxytryptamine (5-HT) on the virulence of the parasite was examined in just weaned Charles Foster strain of albino rats. Pathogenicity of 10 strains of E. histolytica, from various forms of intestinal amoebiasis, grown in modified Boeck and Drbohlav's medium was assessed by caecal scoring. Administration of 5-HT in infected animals significantly enhanced the pathogenicity of all the seven strains tested. Treatment of the host with the 5-HT precursor L-tryptophan also increased the caecal scores examined with three strains of E. histolytica. Prior blocking of tissue 5-HT receptors by administration of methysergide almost completely abolished the pathogenicity enhancing effect of 5-HT treatment. This suggested that 5-HT itself and not any of its metabolites was responsible for the observed increase in pathogenicity of E. histolytica on 5-HT treatment of the host.