Antimicrobial resistance trends in blood culture positive Salmonella Paratyphi A isolates from Pondicherry, India.

Enteric fever is a public health problem with the upsurge in the occurrence of Salmonella isolates that are resistant to ciprofloxacin. In this study, a total of 284 blood culture isolates of S. Paratyphi A were investigated. Of these isolates, 281 (98.9%) were nalidixic acid resistant. A high rate (6.3%) of high-level resistance (≥4 µg/mL) was found to ciprofloxacin. The isolates with ciprofloxacin minimum inhibitory concentrations (MICs) of ≥12 µg/mL had 4 mutations, 2 mutations within the quinolone resistance-determining region of gyrA and 2 mutations also in parC. According to the Clinical Laboratory Standards Institute 2012 MIC breakpoints, 75.0% of isolates were resistant to ciprofloxacin. Finally, 3 major pulsed-field gel electrophoresis patterns were observed among the S. Paratyphi A isolates. The spread of fluoroquinolone resistant S. Paratyphi A necessitates a change toward ‘evidence-based’ treatment for enteric fever. The research provides a perspective on the increasing prevalence of antimicrobial resistant S. Paratyphi A isolates in this region of India.

Plasmid-mediated quinolone resistance in typhoidal Salmonellae: A preliminary report from South India.

Background: Fluoroquinolones are the drugs extensively employed for the treatment of Salmonella infections. Over the couple of decades that have elapsed since the introduction of fl uoroquinolones, resistance to these agents by Enterobacteriaceae family members has become common and widespread. Although fl uoroquinolone resistance is mediated by genomic DNA (deoxyribonucleic acid) as well as plasmid DNA, the plasmid-mediated quinolone resistance (PMQR) facilitates higher level resistance by interacting with genomic mechanism and is capable of horizontal spread. Materials and Methods: During a period of 1-year, 63 typhoidal Salmonellae were isolated from 14,050 blood cultures and one parietal wall abscess. 36 (56.25%) were Salmonella Typhi and 27 (42%) were Salmonella Paratyphi A. They were all screened for resistance by the disc diffusion method and their minimum inhibitory concentrations were determined using agar dilution, broth dilution and E-strip method. Ciprofl oxacin resistant isolates were screened for PMQR determinants by polymerase chain reaction assay. Results: All the 63 isolates were resistant to nalidixic acid. Among the 36 S. Typhi isolates 20 were resistant to ciprofl oxacin, of which 14 carried the plasmid gene qnrB and one carried the aac(6’)-Ib-cr gene. qnrA and qnrS genes were not detected. Ciprofl oxacin resistance was not seen in any of the S. Paratyphi A isolates. Conclusion: The antibiotic sensitivity pattern of typhoidal Salmonellae shows an increasing trend of PMQR. The allele B of qnr gene was found to be the predominant cause of PMQR in this study.

Methicillin resistant Staphylococcus aureus (MRSA) in India Prevalence & susceptibility pattern.

Background & objectives: Methicillin resistant Staphylococcus aureus (MRSA) is endemic in India and is a dangerous pathogen for hospital acquired infections. This study was conducted in 15 Indian tertiary care centres during a two year period from January 2008 to December 2009 to determine the prevalence of MRSA and susceptibility pattern of S. aureus isolates in India. Methods: All S. aureus isolates obtained during the study period in the participating centres were included in the study. Each centre compiled their data in a predefined template which included data of the antimicrobial susceptibility pattern, location of the patient and specimen type. The data in the submitted templates were collated and analysed. Results: A total of 26310 isolates were included in the study. The overall prevalence of methicillin resistance during the study period was 41 per cent. Isolation rates for MRSA from outpatients, ward inpatients and ICU were 28, 42 and 43 per cent, respectively in 2008 and 27, 49 and 47 per cent, respectively in 2009. The majority of S. aureus isolates was obtained from patients with skin and soft tissue infections followed by those suffering from blood stream infections and respiratory infections. Susceptibility to ciprofloxacin was low in both MSSA (53%) and MRSA (21%). MSSA isolates showed a higher susceptibility to gentamicin, co-trimoxazole, erythromycin and clindamycin as compared to MRSA isolates. No isolate was found resistant to vancomycin or linezolid. Interpretation & conclusions: The study showed a high level of MRSA in our country. There is a need to study epidemiology of such infections. Robust antimicrobial stewardship and strengthened infection control measures are required to prevent spread and reduce emergence of resistance.

Early diagnosis of typhoid fever by nested PCR for flagellin gene of Salmonella enterica serotype Typhi.

Background & objectives: Typhoid fever caused by Salmonella Typhi continues to be a major health problem in spite of the use of antibiotics and the development of newer antibacterial drugs. Inability to make an early laboratory diagnosis and resort to empirical therapy, often lead to increased morbidity and mortality in cases of typhoid fever. This study was aimed to optimize a nested PCR for early diagnosis of typhoid fever and using it as a diagnostic tool in culture negative cases of suspected typhoid fever. Methods: Eighty patients with clinical diagnosis of typhoid fever and 40 controls were included in the study. The blood samples collected were subjected to culture, Widal and nested PCR targeting the flagellin gene of S. Typhi. Results: The sensitivity of PCR on blood was found to be 100 per cent whereas the specificity was 76.9 per cent. The positive predictive value (PPV) of PCR was calculated to be 76.9 per cent with an accuracy of 86 per cent. None of the 40 control samples gave a positive PCR. Interpretation & conclusions: Due to its high sensitivity and specificity nested PCR can be used as a useful tool to diagnose clinically suspected, culture negative cases of typhoid fever.

Molecular description of plasmid-mediated AmpC β-lactamases among nosocomial isolates of Escherichia coli & Klebsiella pneumoniae from six different hospitals in India.

Background & objectives: Plasmid mediated AmpC β-lactamase (PMABL) resistance in Escherichia coli and Klebsiella spp. is an emerging problem worldwide. Phenotypic methods are commonly used for detection of PMABL production in Gram-negative isolates, but molecular data about the prevalence of plasmid-mediated AmpC-type resistance at the national level are needed. Hence, a prospective study was undertaken to determine the occurrence of PMABL gene and its types among clinical isolates of E. coli and K. pneumoniae obtained from six different hospitals in India. Methods: A total of 241 nosocomial isolates of K. pneumoniae (n=109) and E.coli (n=132) from six geographically distant hospitals in India were included. These were screened for cefoxitin resistance. AmpC disk test and modified three dimensional extraction test were used for phenotypic detection of PMABL production. Molecular types were determined by a multiplex PCR. Results: Among the 241 isolates, 187 (77.5%) were found to be cefoxitin resistant (K. pneumoniae n=83, E. coli n=104). AmpC activity was detectable in 153 (63.4%) isolates, (K. pneumoniae n=69, E. coli n=84). By PCR, the plasmid encoded AmpC genes were found in 92 (38.1%) isolates and the molecular types of the genes detected predominantly were DHA, CIT followed by MOX and ACC types. Interpretation & conclusions: A high percentage of plasmid-encoded AmpC enzymes was noted in E. coli and K. pneumonia isolates obtained from different parts of the country. Phenotypic methods alone may not reflect the true number of PMABL producers. Genotypic methods need to be employed in national surveillance studies.

Antimicrobial resistance in typhoidal salmonellae.

Infections with Salmonella are an important public health problem worldwide. On a global scale, it has been appraised that Salmonella is responsible for an estimated 3 billion human infections each year. The World Health Organization (WHO) has estimated that annually typhoid fever accounts for 21.7 million illnesses (217,000 deaths) and paratyphoid fever accounts for 5.4 million of these cases. Infants, children, and adolescents in south-central and South-eastern Asia experience the greatest burden of illness. In cases of enteric fever, including infections with S. Typhi and S. Paratyphi A and B, it is often necessary to commence treatment before the results of laboratory sensitivity tests are available. Hence, it is important to be aware of options and possible problems before beginning treatment. Ciprofloxacin has become the first-line drug of choice since the widespread emergence and spread of strains resistant to chloramphenicol, ampicillin, and trimethoprim. There is increase in the occurrence of strains resistant to ciprofloxacin. Reports of typhoidal salmonellae with increasing minimum inhibitory concentration (MIC) and resistance to newer quinolones raise the fear of potential treatment failures and necessitate the need for new, alternative antimicrobials. Extended-spectrum cephalosporins and azithromycin are the options available for the treatment of enteric fever. The emergence of broad spectrum β-lactamases in typhoidal salmonellae constitutes a new challenge. Already there are rare reports of azithromycin resistance in typhoidal salmonellae leading to treatment failure. This review is based on published research from our centre and literature from elsewhere in the world. This brief review tries to summarize the history and recent trends in antimicrobial resistance in typhoidal salmonellae.

Hemophagocytic Syndrome in a Child with Brucellosis.

The authors report case of a two and half years old female child presented with fever for one month with hepatosplenomegaly. Though the child had been symptomatic for a prolonged period, she did not appear very unwell at presentation. However, after admission there was sudden rapid deterioration of her clinical status. Investigations revealed hemophagocytosis with brucella infection.

AmpC beta lactamases among Gram negative clinical isolates from a tertiary hospital, South India

AmpC â-lactamases are cephalosporinases that hydrolyze cephamycins as well as other extended-spectrum cephalosporins and are poorly inhibited by clavulanic acid. Although reported with increasing frequency, the true rate of occurrence of AmpC â-lactamases in different organisms, including members of Enterobacteriaceae, remains unknown. The present study was designed to determine the occurrence of AmpC enzyme-harbouring Gram-negative clinical isolates in a tertiary care hospital in Pondicherry state, South India. A total of 235 Gram negative clinical isolates were tested for resistance to cefoxitin, third generation cephalosporin (3GC) antibiotics, ampicillin, amikacin, co-trimoxazole, gentamicin, meropenem and tetracycline by disc diffusion method. Isolates found resistant to 3GC and cefoxitin were tested for the production of AmpC â -lactamases by three dimensional extraction method and AmpC disc method. Isolates found to sensitive to 3GC were subjected to disc antagonism test for inducible AmpC production. One hundred and thirty four (57 percent) strains were resistant to 3GC, among which 63(47 percent) were positive for plasmid-mediated AmpC beta lactamases production. Among the 101 strains sensitive to 3GC, 23 (22.7 percent) revealed the presence of inducible AmpC beta lactamases by disc approximation test. A total of 80.9 percent (51/63) of screen positive isolates were detected by Amp C disc test and 93.6 percent (59/63) by three dimensional extraction method. Out of the 86 AmpC producers, 67 (77.9 percent) were cefoxitin resistant .Inducible AmpC was not found in Esch.coli and Klebsiella spp. The AmpC producers also concurrently showed multidrug resistance pattern. AmpC producers were found to be prevalent in our hospital and though three dimensional extraction test detects AmpC better, the disk test is easier to perform routinely and is user- friendly.

Comparison of an in-house latex agglutination test with IgM ELISA and MAT in the diagnosis of leptospirosis.

The laboratory diagnosis of leptospirosis is fraught with several problems. Isolation of Leptospira by culture has a low sensitivity and the microscopic agglutination test (MAT) is time consuming To overcome these problems, a rapid latex agglutination test (LAT) has been standardized for the detection of antileptospiral antibodies in serum samples from suspected cases of leptospirosis. We compared the efficiency of the LAT to a commercially available IgM ELISA and MAT. A total of 150 serum samples were tested by LAT, IgM ELISA, and MAT. The positivity was 26.7%, 26% and 24% respectively. The sensitivity and specificity of LAT as compared to MAT was 90.62 and 91.96% respectively. Even though LAT and ELISA showed similar results, its rapidity and simplicity made latex agglutination test more suitable as a rapid screening test.

Neonatal melioidosis: A case report from India.

Melioidosis, caused by Burkholderia pseudomallei , is an infectious disease of major public health importance in Southeast Asia and Australia. We report, for the first time from the Indian subcontinent, a case of melioidosis in a neonate, its clinical presentation, microbiological diagnosis, possible mode of transmission and outcome. A pre-term female baby developed respiratory distress soon after birth. The child was febrile, had tachypnea, grunting, normal heart rate with a low pulse volume and poor peripheral perfusion. Chest X-ray revealed right-sided bronchopneumonia. B. pseudomallei was isolated from the blood culture of the neonate collected aseptically. The neonate was successfully treated with meropenem.

Simple screening tests for detection of carbapenemases in clinical isolates of nonfermentative Gram-negative bacteria.

Background & objectives: The production of carbapenemases is an important mechanism responsible for the carbapenem resistance. A simple and inexpensive testing method for screening of carbapenemase producers is essential. A prospective study was undertaken to detect metallo-β-lactamases (MBLs) and AmpC β-lactamases in nonfermentative Gram negative bacteria and to evaluate the various methods for detection of carbapenemases and MBLs. Methods: A total of 100 Acinetobacter spp. (78 A. baumannii and 22 A. lwoffi i) and 140 Pseudomonas spp. (103 P. aeruginosa and 37 other Pseudomonas spp.) were screened for meropenem resistance by Kirby- Bauer disc diffusion method. Modifi ed Hodge test, EDTA disk synergy (EDS) test and AmpC disk test were used for the detection of carbapenemases, MBLs and AmpC β-lactamases, respectively. Results: Forty six (59.0%) A. baumannii, 7 (31.8%) A. lwoffi i, 32 (31.1%) P. aeruginosa and 7 (18.9%) Pseudomonas spp. were resistant to meropenem. Among the 32 meropenem resistant P. aeruginosa, 15 (46.9%) were AmpC β-lactamase producers, 16 (50.0%) MBL producers by EDS test, but only 9 (28.1%) found positive for carbapenemases by modifi ed Hodge test. Among the 46 meropenem resistant A. baumannii, 31 (67.4%) were AmpC β-lactamase producers, 3 (6.5%) MBL producers, but only 1 (14.3%) was positive for carbapenemases by modifi ed Hodge test. One P. aeruginosa was positive for carbapenemase by modifi ed Hodge test, but was negative for MBL and AmpC β-lactamase. Interpretation & conclusions: MBL production is an important mechanism of carbapenem resistance among Pseudomonas species but not among Acinetobacter species. EDS is more sensitive for detection of MBLs than modifi ed Hodge test. Both EDTA-meropenem and EDTA-ceftazidime combination must be used to detect all the MBL producers. Carbapenemases other than MBL may also be responsible for carbapenem resistance. AmpC β-lactamase is also a contributory factor for carbapenem resistance among the isolates in the hospital.

Cryptococcal meningoencephalitis diagnosed by blood culture.

Increase in cryptococcal infection has been noticed after acquired immunodeficiency syndrome pandemic. Cryptococcus neoformans can be isolated from blood in the process of dissemination to brain. We report a case of cryptococcal fungaemia in a patient whose cerebrospinal fluid was negative for Cryptococcus neoformans. Retrospective analysis revealed human immunodeficiency virus seropositivity of the patient. He was treated with amphotericin B and fluconazole. Antiretroviral therapy was started, however, the patient succumbed to the infection.

Maternal genital bacteria and surface colonization in early neonatal sepsis.

OBJECTIVE: The study was intended to evaluate the role of maternal genital bacteria and baby's surface colonization in early onset neonatal sepsis. METHODS: Babies (born in the hospital of Jawaharlal Institute of Postgraduate Medical Education and Research) who developed clinical signs of sepsis were studied. Swabs were collected for culture from baby's umbilicus, ear, throat in addition to gastric aspirate and blood culture. The genital tract of the mother was also studied for bacterial colonization. The organisms isolated from the maternal genital tract and baby's surface colonization were correlated with those isolated from blood culture by calculating Phi correlation coefficient. RESULTS: Esch coli was the most common organism isolated from maternal genital tract and surface cultures of babies, but Klebseilla-Spp was the most common organism isolated from blood. There was a significant correlation between surface colonization of babies and maternal genital bacteria, so also was baby's surface culture and blood culture. However, correlation between maternal genital bacteria and baby's blood culture was not significant. CONCLUSION: Surface colonizing bacteria and not maternal genital bacteria are important in early onset neonatal sepsis.

Current pattern in antimicrobial susceptibility of Salmonella Typhi isolates in Pondicherry.

Typhoid fever continues to remain a health problem as the causative organism Salmonella Typhi has developed resistance to many of the antibiotics used. This study was undertaken to determine the current pattern of resistance to antimicrobial agents and phage types of S.Typhi isolates obtained in a tertiary health care hospital in Pondicherry. Blood culture was done for 1296 suspected cases of enteric fever and 157 strains of S. Typhi were isolated. Sensitivity to ampicillin, chloramphenicol, gentamicin, ciprofloxacin and ceftriaxone was determined by disc diffusion, and the minimum inhibitory concentration (MIC) of ciprofloxacin determined. There were 61 multidrug resistant (MDR) isolates. The MIC of ciprofloxacin for 147 isolates was >0.5 mg/l; of these, 131 were resistant to nalidixic acid. Phage typing was done for 123 isolates and 115 were found to be of phage type E1, biotype 1. A decline in the number of MDR isolates was noted. Concurrently, there has been an increase in the number of isolates sensitive to all antibiotics except nalidixic acid, and all these isolates showed reduced susceptibility to ciprofloxacin. Nalidixic acid susceptibility could be a useful screening test for the detection of decreased susceptibility of S. Typhi to ciprofloxacin. The clinicians should be advised to use ceftriaxone selectively in cases showing non-responsiveness to ciprofloxacin.

Detection of slime production by coagulase-negative Staphylococcus: an assessment of two methods.

The detection of slime can be helpful in determining the diagnostic value of coagulase-negative Staphylococcus (CNS). Here, we have made a comparative study of two phenotypic methods of slime production test from 28 clinically significant blood culture isolates of CNS. A total 16 out of 28 isolates (57.1%) were positive for slime by either of the tests, whereas only 5 out of 28 isolates (17.9%) were positive for slime by both the tests. The tube method detected slime in significantly more number of isolates than the spectrophotometric method (14 vs. 7; p<0.0483279). A weak correlation was found between the tube method and the spectrophotometric method (rs=0.2474). The tube test was found to be superior to the spectrophotometric test in terms of sensitivity, ease of performance and interpretation, and cost effectiveness.

Factors predicting occult bacteremia in young children.

A febrile child without a definite localizing sign of infection may be in initial phase of bacteremia which unless treated would result in systemic complication. These instances are referred to as "Occult bacteremia". The common pathogens isolated in these children are Streptococcus pneumoniae, Hemophilus influenzae and Neisseria meningitidis. A hundred consecutive children in the age group of 3-36 months attending pediatric outpatient department and casualty were clinically evaluated using AIOS (acute illness observation scale) score and were subjected to complete blood counts, smear for malarial parasites, ESR and blood culture. In the 19-month study period, 4 instances of occult bacteremia were identified. Streptococcus pneumoniae was cultured in 3 cases and H. influenzae in one. A febrile and toxic child in the age group of 3-36 months has a high risk of occult bacteremia. High fever of temperature > or = 102 degrees F, ESR > or = 15 mm/hour, and total leukocyte count > or = 15,000/mm3, in a child with AIOS score of > or = 10 may be considered for more detailed investigations and early intervention with antimicrobial therapy.

Utility of gelatin particle agglutination test (MLPA) for rapid serodiagnosis of leprosy in a hyperendemic area.

The anti-PGL M. leprae specific antibodies were estimated by MLPA test in 79 patients of leprosy, 8 contacts of lepromatous cases and 10 healthy controls in a hyperendemic area. The results indicated an over all seropositivity of 50.6% in leprosy patients. Three of the eight contacts and five of the controls also gave positive results. Higher seropositivity rates were noted in multibacillary patients (73% in lepromatous, 53.6% in borderline, 40% each in tuberculoid and indeterminate and 10% in pure neuritic types). The practical application of MLPA test in its present form as a serodiagnostic procedure for screening subclinical or clinical infections in leprosy patients appear to be of limited value in hyperendemic areas. Further studies involving large series of subjects are necessary for reaching definite conclusions.