Antimicrobial susceptibility profiles of gram-negative bacteria causing infections collected across India during 2014–2016: Study for monitoring antimicrobial resistance trend report

Background: The emergence of antibiotic resistance among bacterial pathogens in the hospital and community has increased the concern to the health-care providers due to the limited treatment options. Surveillance of antimicrobial resistance (AMR) in frequently isolated bacterial pathogens causing severe infections is of great importance. The data generated will be useful for the clinicians to decide empiric therapy on the local epidemiological resistance profile of the antimicrobial agents. This study aims to monitor the distribution of bacterial pathogen and their susceptibility pattern to the commonly used antimicrobial agents. Materials and Methods: This study includes Gram-negative bacilli collected from intra-abdominal, urinary tract and respiratory tract infections during 2014–2016. Isolates were collected from seven hospitals across India. All the study isolates were characterised up to species level, and minimum inhibitory concentration was determined for a wide range of antimicrobials included in the study panel. The test results were interpreted as per standard Clinical Laboratory Standards Institute guidelines. Results: A total of 2731 isolates of gram-negative bacteria were tested during study period. The most frequently isolated pathogens were 44% of Escherichia coli (n = 1205) followed by 25% of Klebsiella pneumoniae (n = 676) and 11% of Pseudomonas aeruginosa (n = 308). Among the antimicrobials tested, carbapenems were the most active, followed by amikacin and piperacillin/tazobactam. The rate of extended-spectrum beta-lactamase (ESBL)-positive isolates were ranged from 66%–77% in E. coli to 61%–72% in K. pneumoniae, respectively. Overall, colistin retains its activity in > 90% of the isolates tested and appear promising. Conclusion: Increasing rates of ESBL producers have been noted, which is alarming. Further, carbapenem resistance was also gradually increasing, which needs much attention. Overall, this study data show that carbapenems, amikacin and colistin continue to be the best agents available to treat drug-resistant infections. Thus continuous monitoring of susceptibility profile of the clinically important Gram-negative pathogens is of great importance to guide effective antimicrobial therapy.

Intestinal microsporidiosis in renal transplant recipients: Prevalence, predictors of occurrence and genetic characterization.

Purpose: Intestinal microsporidiosis, which occurs in immunocompromised states such as acquired immunodeficiency syndrome, has rarely been studied in patients with renal transplantation (RT) on immunosuppressive therapy. Materials and Methods: Three hundred and twenty‑four consecutive RT recipients on immunosuppressive treatment and 170 healthy subjects were evaluated for intestinal microsporidiosis and other parasites by modified trichrome staining, wet mount using normal saline, iodine and polymerase chain reaction (PCR). Clinical, demographic and laboratory parameters associated with occurrence of intestinal microsporidiosis were studied using univariate and multivariate analysis. The species of microsporidia were studied using PCR‑restriction fragment length polymorphism (RFLP). Patients were treated with albendazole (400 mg twice daily for 2 weeks). Results: Of 324 RT recipients initially screened, 52 were excluded from final analysis due to incomplete data. Patients with RT [n = 272, age 42 ± 12.54 years, 222 (81.6%) male] more often had microsporidiosis than healthy subjects by modified trichrome stain and PCR [n = 170, age 33.8 ± 6.7 years, 123 (72.3%) male] [16/272 (5.8%) vs. 0/170 (0%), P < 0.001]. Patients with intestinal microsporidiosis were younger (33.9 ± 8.3 years vs. 42.3 ± 12.6 years; P = 0.009), had diarrhoea more often (13/16, 81% vs. 123/256, 48%; P = 0.02), which was longer in duration (60, 32.5-105 days vs. 12, 6.2-18 days; P < 0.001) and had associated giardiasis (2/16, 12.5% vs. 2/256, 0.8%; P = 0.018). Younger age, presence of diarrhoea and associated giardiasis were significant on multivariate analysis. Enterocytozoon bieneusi was detected in 15/16 (93%) patients with intestinal microsporidiosis. Conclusion: Intestinal microsporidiosis occurs frequently in patients with RT on immunosuppressive treatment, particularly among younger patients with longer diarrhoea duration and associated giardiasis. E. bieneusi is the major species identified among these patients.

Use of GenoType® MTBDRplus assay to assess drug resistance and mutation patterns of multidrug-resistant tuberculosis isolates in northern India.

Purpose: The emergence and spread of multidrug-resistant tuberculosis (MDR-TB) is a major public health problem. The diagnosis of MDR-TB is of paramount importance in establishing appropriate clinical management and infection control measures. The aim of this study was to evaluate drug resistance and mutational patterns in clinical isolates MDR-TB by GenoType® MTBDRplus assay. Material and Methods: A total of 350 non-repeated sputum specimens were collected from highly suspected drug-resistant pulmonary tuberculosis (PTB) cases; which were processed by microscopy, culture, differentiation and first line drug susceptibility testing (DST) using BacT/ALERT 3D system. Results: Among a total of 125 mycobacterium tuberculosis complex (MTBC) strains, readable results were obtained from 120 (96%) strains by GenoType® MTBDRplus assay. Only 45 MDR-TB isolates were analysed for the performance, frequency and mutational patterns by GenoType® MTBDRplus assay. The sensitivity of the GenoType® MDRTBplus assay for detecting individual resistance to rifampicin (RIF), isoniazid (INH) and multidrug resistance was found to be 95.8%, 96.3% and 97.7%, respectively. Mutation in codon S531L of the rpoB gene and codon S315T1 of katG genes were dominated in MDR-TB strains, respectively (P < 0.05). Conclusions: The GenoType® MTBDRplus assay is highly sensitive with short turnaround times and a rapid test for the detection of the most common mutations conferring resistance in MDR-TB strains that can readily be included in a routine laboratory workflow.

Changing patterns and trends of multidrug-resistant tuberculosis at referral centre in Northern India: A 4-year experience.

Purpose: India has a high burden of drug-resistant tuberculosis (TB), although there is little data on multidrug-resistant tuberculosis (MDR-TB). Although MDR-TB has existed for long time in India, very few diagnostic laboratories are well-equipped to test drug sensitivity. The objectives of this study were to determine the prevalence of MDR-TB, first-line drug resistance patterns and its changing trends in northern India in the 4 years. Materials and Methods: This was a prospective study from July 2007 to December 2010. Microscopy, culture by Bactec460 and p-nitro-α-acetylamino-β-hydroxypropiophenone (NAP) test was performed to isolate and identify Mycobacterium tuberculosis (M. tb) complex (MTBC). Drug sensitivity testing (DST) was performed by 1% proportional method (Bactec460) for four drugs: Rifampicin, isoniazid, ethambutol and streptomycin. Various clinical and demographical profiles were evaluated to analyse risk factors for development of drug resistance. Results: We found the overall prevalence rate of MDR-TB to be 38.8%, increasing from 36.4% in 2007 to 40.8% in 2010. we found that the prevalence of MDR-TB in new and previously treated cases was 29.1% and 43.3% ( P < 0.05; CI 95%). The increasing trend of MDR-TB was more likely in pulmonary TB when compared with extra-pulmonary TB ( P < 0.05; CI 95%). Conclusions: we found a high prevalence (38.8%) of MDR-TB both in new cases (29.1%) and previously treated cases (43.3%).This study strongly highlights the need to make strategies for testing, surveillance, monitoring and management of such drug-resistant cases.

Posterior fossa midline cryptococcoma in a patient with idiopathic CD4 lymphocytopenia.

Idiopathic CD4 lymphocytopenia (ICL) is a rare disorder which is often diagnosed as HIV-negative AIDS in the light of poor immunity and AIDS-defining illnesses. We present a case of a 50-year-old male who presented with a midline posterior fossa tumour with ICL diagnosed as cerebellar cryptococcoma.

Detection of 123 bp fragment of insertion element IS6110 Mycobacterium tuberculosis for diagnosis of extrapulmonary tuberculosis.

Purpose: Extrapulmonary tuberculosis (EPTB) is emerging problem in developing and developed countries. The diagnosis of EPTB in its different clinical presentations remains a true challenge. IS6110-based polymerase chain reaction (PCR) is used for rapid identification and positivity rate of the Mycobacterium tuberculosis complex in clinical isolates of different sites of EPTB. The present study was carried out to study the prevalence of M. tuberculosis complex in clinical isolates of EPTB at tertiary care centres in Lucknow. Materials and Methods: Seven hundred fifty-six specimens were collected from the suspected cases of EPTB which were processed for Mycobacteria by Ziehl Neelson (ZN) staining and BACTEC culture. All the specimens were also processed for IS6110-based PCR amplification with primers targeting 123 bp fragment of insertion element IS6110 of the M. tuberculosis complex. Results: Of these 756 specimens, 71(9.3%) were positive for acid fast bacilli (AFB) by ZN staining, 227(30.1%) were positive for mycobacteria by BACTEC culture and IS6110 PCR were positive for M. tuberculosis complex in 165 (20.7%) isolates. We found a significant difference in sensitivities of different tests (P<0.05). Conclusions: This study reveals the positivity of M. tuberculosis complex in clinical isolates of EPTB case in tertiary care hospitals in Northern India. 72.7% of M. tuberculosis complex was confirmed by IS6110-PCR in culture isolates from different sites of EPTB. The high prevalence of the M. tuberculosis complex was seen in lymph node aspirate and synovial fluid. However, utility of PCR may play a potentially significant role in strengthening the diagnosis of EPTB especially targeting IS6110.

Rationale of azithromycin prescribing practices for enteric fever in India.

Purpose: The present study was performed to assess the current susceptibility pattern of blood isolates of Salmonella spp from a super specialty hospital in North India against nalidixic acid, ciprofloxacin and azithromycin and compare the in vitro and in vivo response against azithromycin. Materials and Methods: We evaluated the minimum inhibitory concentration's (MIC's) of 107 blood isolates of Salmonella spp against nalidixic acid, azithromycin and ciprofloxacin and correlated in vitro and in vivo response of azithromycin from the treatment and discharge summaries from the Hospital Information System (HIS) software. Results: Among the 107 isolates evaluated, 94 (87.8%) were nalidixic acid-resistant (NAR) Salmonella and 36 were resistant to azithromycin by MIC testing. The MIC 90 value for azithromycin was 24 μg/mL. Among the 57 treatment histories evaluated using the HIS software, 19 (33%) patients had documented clinical non-response to azithromycin which required change of therapy. Conclusions: The present study observed a higher MIC 90 values for azithromycin compared to Salmonella isolates from Western studies. There was also a documented clinical non-response against azithromycin. The in vitro and in vivo findings in this study suggest a guarded use of azithromycin for cases of enteric fever in India. The study also augments the reversal of resistance pattern in favour of chloramphenicol, ampicillin and trimethoprim - sulfamethoxazole.

Ventilator-associated pneumonia caused by carbapenem-resistant Enterobacteriaceae carrying multiple metallo-beta-lactamase genes.

Context: Ventilator-associated pneumonia (VAP) is a leading nosocomial infection in the intensive care unit (ICU). Members of Enterobacteriaceae are the most common causative agents and carbapenems are the most commonly used antibiotics. Metallo-beta-lactamase (MBL) production leading to treatment failure may go unnoticed by routine disc diffusion susceptibility testing. Moreover, there is not much information on association of MBL-producing Enterobacteriaceae with ICU-acquired VAP. Therefore, a study was undertaken to find out the association of MBL-producing Enterobacteriaceae with VAP. Settings: This study was conducted in a large tertiary care hospital of North India with an eight-bed critical care unit. Materials and Methods: The respiratory samples (bronchoalveolar lavage, protected brush catheter specimens and endotracheal or transtracheal aspirates) obtained from VAP patients (during January 2005-December 2006) were processed, isolated bacteria identified and their antibiotic susceptibilities tested as per standard protocols. The isolates of Enterobacteriaceae resistant to carbapenem were subjected to phenotypic and genotypic tests for the detection of MBLs. Results: Twelve of 64 isolates of Enterobacteriaceae were detected as MBL producers, bla IMP being the most prevalent gene. Additionally, in three strains, simultaneous coexistence of multiple MBL genes was detected. Conclusion: The coexistence of multiple MBL genes in Enterobacteriaceae is an alarming situation. As MBL genes are associated with integrons that can be embedded in transposons, which in turn can be accommodated on plasmids thereby resulting in a highly mobile genetic apparatus, the further spread of these genes in different pathogens is likely to occur.

Progression of chronic pulmonary tuberculosis in mice intravenously infected with ethambutol resistant Mycobacterium tuberculosis.

Purpose: Ethambutol (EMB) is an important first line drug, however little information on its molecular mechanism of resistance and pathogenicity of resistant isolates is available. Present work was designed to study virulence of the EMB resistant M. tuberculosis strains and the host responses in-vivo on infection of EMB resistant M. tuberculosis using Balb/c mouse model of infection. Methods: Three groups of Balb/c mice (female, age 4-6 wk; 21 mice in each group) were infected intravenously with 106 CFU of M. tuberculosis H37Rv and two EMB resistant clinical isolates. Age and sex matched control animals were mock inoculated with Middlebrook 7H9 broth alone. At 10, 20, 30, 40, 50, 60, and 70 days post-infection three animals from each group were sacrificed by cervical dislocation and lung tissue was collected for further analysis. Results: Infection with EMB resistant M. tuberculosis led to progressive and chronic disease with significantly high bacillary load (p=0.02). Massive infiltration and exacerbated lung pathology with increased expression of IFN-gamma and TNF-alpha was observed in lungs of mice infected with EMB resistant strains. The present study suggests that infection with EMB resistant M. tuberculosis leads to chronic infection with subsequent loss of lung function, bacterial persistence with elevated expression of TNF-alpha resulting in increased lung pathology. Conclusion: These findings highlight that EMB resistant M. tuberculosis regulates host immune response differentially and its pathogenicity is different from drug sensitive strains of M. tuberculosis.

Seroprevalence and risk factors for hepatitis B virus infection among general population in Northern India / Soroprevalência e fatores de risco para infecção pelo vírus da hepatite B na população geral do nordeste da Índia

BACKGROUND: Infection with hepatitis-B virus has been a significant cause of morbidity claiming more than a million lives every year. Epidemiological data reveals that there are 360 million carriers of hepatitis-B virus throughout the globe and 78 percent of the world populations’ hail from Asia. Though several studies from Indian sub-continent have provided an estimate of the prevalence of this viral infection, there exist only few studies, which reflect the status in the general population. AIM: The present study was designed to investigate the prevalence of hepatitis-B infection in North Indian general population. METHODS: The study population comprised of 20,000 healthy blood donors who were screened for hepatitis-B surface antigen (HBsAg) status using third generation ELISA kit. Seroprevalence rate of seropositive donors was calculated and stratified by age, sex and blood groups. Statistical analysis was performed using tests of proportions, chi-square and confidence interval. RESULTS: The study showed that out of 20,000 donors, 450 (2.25 percent) were HBsAg positive (95 percent confidence interval (CI), 2.0445-2.4554). Higher prevalence of HbsAg was found among males (440/19235) than females (10/765). The age specific prevalence rose from 1.78 percent (108/6058) in donors aged 19-25 years to a maximum of 3.03 percent (96/3161) in donors aged 35-45 years and decreased in older age groups. The peaks were detected in male donors aged 35-45 years and in females aged 25-35 years. Rh-negative blood group donors (21/873) and Rh-positive group donors (429/19127) had almost equivalent prevalence rates of HBsAg. HBsAg was more prevalent in blood group B donors (174/7426) and less prevalent in AB blood group donors (38/2032). CONCLUSION: It was found that variables including gender and age were significantly associated with HBsAg positivity. HBsAg positivity in our population was statistically not associated with ABO blood groups.

RACIONAL: A infecção pelo vírus B da hepatite é considerada uma significante causa de morbidade, responsável por mais de 1 milhão de casos, a cada ano. Dados epidemiológicos revelam que existem 360 milhões de portadores de vírus da hepatite B no mundo e 78 por cento da população natural da Ásia. Embora vários estudos realizados na Ásia sub-continental tenham fornecido uma estimativa de prevalência desta infecção viral, existem poucos estudos que avaliam esta condição na população geral. OBJETIVO: Investigar a prevalência da infecção pelo víirus B da hepatite na população geral do nordeste da Índia. MÉTODOS: A população estudada compunha-se de 20.000 doadores de sangue sadios, selecionados através de positividade do antígeno de superfície da hepatite B (HBsAg), utilizando-se o kit ELISA de 3ª geração. A taxa de soroprevalência dos doadores soropositivos foi calculada e estratificada por idade, sexo e grupos sangüíneos. Análise estatística foi obtida usando-se testes de proporções, do qui ao quadrado e intervalo de confiança (CI). RESULTADOS: O estudo revelou que dos 20.000 doadores, 450 (2,25 por cento) eram HBsAg positivos (95 por cento CI 2.0445-2.4554). Prevalência maior de HBsAg foi encontrada em homens (440/19235) do que em mulheres (10/765). A prevalência por faixa etária aumentou de 1,78 por cento (108/6058) em doadores entre 19 a 25 anos, para máximo de 3,03 por cento (96/3161) naqueles entre 35-45 anos e decresceu nos grupos de maior idade. Os picos foram detectados em doadores masculinos de 35-45 anos e em mulheres de 25-35 anos. O grupo de doadores Rh negativo (21/873) e o grupo Rh positivo (4299/19127) mostraram taxas de prevalência de HBsAg quase equivalentes. HBsAg foi mais prevalente no grupo de doadores de tipo sangüíneo B (174/7426) e menos no grupo de tipo sangüíneo AB (38/2032). CONCLUSÃO: Verificou-se que variáveis incluindo gênero e idade foram significantemente associadas à positividade do HBsAg e que esta não foi...

Molecular characterization of echovirus 11 isolates from India.

BACKGROUND & OBJECTIVES: Echovirus 11 (ECV11) is one of the most frequent non-polio enteroviruses isolated from stool samples of children with acute flaccid paralysis in north India. The present work was undertaken to study the sequence variability in the 440 bp of 5'-non-translated region of ECV11 genome using heteroduplex mobility assay (HMA). METHODS: Twelve ECV11 isolates were studied for sequence variability in the 5'-non-translated region (5'NTR) using the HMA followed by nucleotide sequencing. HMA was used to determine sequence diversity between Indian ECV11 isolates and prototype Gregory strain. HMA results were confirmed by 5'NTR nucleotide sequencing of five Indian ECV11 isolates. RESULTS: HMA results showed high genomic diversity between the prototype Gregory strain and Indian ECV11 isolates. All isolates were grouped into five different types of heteroduplex mobility patterns with respect to Gregory strain. A 440 bp 5'NTR fragment of five ECV11 isolates representing different heteroduplex patterns, was sequenced. The sequence alignment showed that 5'NTR of Indian isolates was different from prototype Gregory strain and identical to the ECV11 isolates of Finland and Hungary. Phylogenetic analysis including ECV11 isolate sequences from different parts of the world showed that Indian ECV11 isolates represented a different subgroup. INTERPRETATION & CONCLUSION: The results of the present study suggested that the HMA could be successfully used as a preliminary screening method for sequence variability determination of enterovirus field isolates. The sequence data generated on ECV11 isolates from India will be useful for future studies of endemic genotypes of echovirus.

Non-polio enteroviruses in acute flaccid paralysis.

OBJECTIVE: Human enteroviruses are the major cause of aseptic meningitis and also cause a wide range of other acute illnesses, including neonatal sepsis like disease, meningitis, acute flaccid paralysis and acute hemorrhagic conjunctivitis. Infection in neonates is particularly life threatening. METHODS: Stool samples of 523 children (age < 4 years) showing symptoms of acute flaccid paralysis (AFP) were studied. National Polio Surveillance Project workers from different parts of Uttar Pradesh and Bihar collected the samples during June to October 1998. Non-polio enteroviruses (NPEV) were isolated in 191 cases only, by cell culture based neutralization assay. These NPEV isolates were further studied to find the frequent enterovirus serotype detected in stool of children having AFP. RESULTS: Data generated will help future studies on NPEV serotypes circulating in this area. CONCLUSION: In addition it may reduce unnecessary hospitalization, allow immune globulin batches of high titres to frequently circulating serotypes, to be reserved for intravenous therapy of neonates and guide the formulation of antigens for rapid and less expensive diagnosis.

Identification of enteric pathogens in HIV-positive patients with diarrhoea in northern India.

Enteric pathogens associated with chronic diarrhoea in HIV-positive patients were studied. The study was conducted during January 1995-December 1998. Stool specimens from all diarrhoea patients (n = 26) were examined microscopically for ova and parasites using wet preparations and stained smears. Stool samples from diarrhoea patients were also cultured on appropriate media to isolate enteric bacterial pathogens. Of the 59 patients, 26 (44%) had prolonged diarrhoea for more than 4 weeks. Enteric pathogens were detected in 19 (73%) of the 26 patients: 17 patients harboured a single pathogen, and 2 patients had mixed pathogens. The detection rate of emerging parasites, including Isospora, Cryptosporidium, Blastocystis hominis, and Strongyloides stercoralis as a single agent, was significantly higher than conventional pathogens (50% vs 19.2%; p < 0.05). Only one patient harboured both conventional and emerging pathogens (Entamoeba histolytica and Cryptosporidium). Isospora belli was detected in 8 (31%) of the 26 diarrhoea patients: in 7 (27%) patients as a single agent and in one patient with S. stercoralis. Cryptosporidium was identified in 3 (11%) diarrhoea patients: in 2 (8%) patients as a single agent and in one patient with E. histolytica, followed by B. hominis in 2 (8%) patients. E. histolytica was most commonly isolated (3/26; 11.5%), followed by Giardia lamblia, enteropathogenic Escherichia coli, and Campylobacter jejuni (one patient each). Parasitic pathogens were frequently associated with HIV-positive patients with diarrhoea in northern India. I. belli was the most frequent parasite isolated, followed by Cryptosporidium. Stools of all HIV-positive patients with diarrhoea should thoroughly be investigated to identify aetiologic agents for proper management.

Intratypic differentiation & partial nucleotide sequencing of poliovirus isolates of northern India.

The potential resolving power of molecular epidemiological studies has enhanced the precision and reliability of poliovirus (PV) surveillance. PV has an error prone RNA polymerase responsible for rapid evolution of genome (approximately 10(-2) nt substitution/site/year), during inter and intra-human passages. The present study included a serotyped panel of 60 PV (42 PV type-1, 13 PV type-2 and 5 PV type-3) isolated during 1997. They were differentiated into vaccine (Sabin) and wild strains by two methods viz., genotype specific RNA probe hybridization (Rpro-Hy) based on genotypic variability; and ELISA that uses cross-absorbed antiserum (Pab-E) based on phenotypic variability. For obtaining information on molecular epidemiology, partial nucleotide sequencing (VP1/2A region) of five clinical PV isolates was also done. Three of the 60 isolates (two PV type-1 and one PV type-3) intratyped, could not be differentiated correctly by either method. Genotypic characterization of PV isolates was done for confirmation of intratyping results. All five wild PV1 sequenced belonged to the same genotype (> 85% homology) and sequence divergence among the strains was < or = 4.5 per cent. This indicated circulation of a single genetic lineage in the area.

Role of yeasts as nosocomial pathogens & their susceptibility to fluconazole & amphotericin B.

A total of 213 and 208 yeasts were isolated as nosocomial pathogens from various infected specimens during 1996 and 1997 respectively. Yeasts ranked fifth among uropathogens in both the years and from eighth to eleventh in other specimens. Increasing trend in nosocomial urinary tract yeast infection (11.9 in 1996 to 12.6 in 1997) and decreasing trend in wound and other infections (5.1 in 1996 to 2.9 in 1997) per 1000 patients' discharges were observed; blood stream infection remained unchanged (2/1000 discharges) in both the years. Eighty two (41 from each year) randomly selected yeasts were identified to species level following standard protocol and tested for antifungal susceptibility against fluconazole and amphotericin B by reference broth macrodilution technique and agar dilution (AD) method. The frequency of various yeast species identified was Candida albicans 39 (47.6%), C.tropicalis 29 (35.4%), C. krusei 4 (4.9%), C. glabrata 3 (3.7%), C. zeylanoides 2 (2.4%), C. guilliermondii 2 (2.4%), one strain (1.2%) each of C. kefyr, C. parapsilosis, and Trichosporon beigelii. Resistance to fluconazole (MIC > or = 64 micrograms/ml) as per NCCLS criteria was observed in 2 Candida sp. (2.4%). Significantly higher number of non-albicans Candida sp. (8/43; 18.6%) had MIC > 8 micrograms/ml as compared to C. albicans (2/39; 5.1%) (P < 0.05). Only one strain of C. tropicalis had MIC 8 micrograms/ml to amphotericin B and none had MIC > 8 micrograms/ml. Agreement between the reference and the AD methods for fluconazole was 88 per cent and for amphotericin B was 94 per cent. The present study indicates that Candida sp. are emerging as important nosocomial pathogens and the tendency of yeasts to develop resistance to antifungal agents appears to be a challenge for patient management.

Rapid identification of Campylobacter jejuni strains by polymerase chain reaction & their restriction fragment length polymorphism analysis.

A polymerase chain reaction (PCR) technique was developed for specific identification of C. jejuni. A primer pair of a conserved region of flagellin A (fla A) gene identified all 15 strains of C. jejuni isolated from human faeces. None of the control strains like Helicobacter pylori, Vibrio cholerae Escherichia coli and Salmonella typhimurium except C. coli exhibited any amplified product by PCR. A predicted 450 bp could also be amplified from 4 chicken caecal contents positive for C. jejuni-C coli by culture. The caecal contents remained positive for C. jejuni-C. coli by PCR after preservation at 4 degrees C for one week when no viable organism could be detected. Restriction fragment length polymorphism analysis (RFLP) of fla A amplified product by using Bgl II enzyme classified 15 strains into 5 types. Three C. jejuni strains isolated from the same patient over a period of 3 wk showed the same RFLP pattern. The present study indicates that PCR is specific for C. jejuni-C. coli and it has the potential for rapid diagnosis of infection. RFLP can be a good epidemiological marker for C. jejuni infection.

Nosocomial infection due to enterobacter cloacae in a tertiary care hospital in northern India.

A total of 151 Enterobacter cloacae strains isolated from clinical samples (n = 139) and the hospital environment (n = 12) at a tertiary care hospital in northern India during January to October 1993, were analysed. The maximum isolations were during May (n = 24), June (n = 23) and July (n = 22). Urinary tract infection (n = 56) was the most common complication of E. cloacea infection followed by wound infection (42), respiratory tract infection (23) and bacteraemia/septicaemia (18). The frequency of resistance to different drugs was ampicillin 77.4 per cent, cotrimoxazole 79.5 per cent, gentamicin 57.5 per cent, cefotaxime 47 per cent and ofloxacin 36 per cent. Sixty three (41.7%) strains exhibited resistance to multiple drugs. Environmental isolates from bed, hospital diet, hand swab and water from a leaking drain pipe in a ward showed the same resistance pattern. A single index strain could not be identified using phage and biotyping, indicating that a variety of strains were responsible for the nosocomial infection. Adoption of strict aseptic measures and repair of the pipe brought down the infection rate.

Mannose-resistant haemagglutination by Campylobacter jejuni--a preliminary communication.

Ten strains of C. jejuni each isolated respectively from patients with diarrhoea and from chicken intestine (10 strains from each source) were examined for presumptive colonization factor(s) by measuring their cell surface hydrophobicity and haemagglutination. None of the strains expressed cell surface hydrophobicity. However, 14 strains (7 from either source) showed variable haemagglutination pattern with human, sheep and rabbit erythrocytes in the presence of 0.5 per cent D-mannose. Thus, mannose resistant haemagglutinin(s) may be involved in the colonization of intestinal mucosal surfaces by C. jejuni.

Enterotoxigenic Campylobacter jejuni & C. coli in the etiology of diarrhoea in northern India.

Faecal specimens from subjects with (320) and without (450) diarrhoea were screened for Campylobacter jejuni and C. coli. C. jejuni and C. coli were detected in 5 per cent of subjects with diarrhoea and 0.7 per cent of those without diarrhoea and the difference was significant (P less than or equal to 0.01). The isolation rate was much higher in under five diarrhoeal children (8.3%), in comparison to the older group (3.0%). Eleven diarrhoeal isolates of C. jejuni and C. coli were tested in rat ileal loops for enterotoxigenicity. All the strains caused fluid accumulation in the loop model. However, 6 strains required up to 3 consecutive passage(s) to do so. Therefore, rat ileal loops were found to be sensitive and reproducible animal model for the demonstration of enterotoxin produced by C. jejuni and C. coli. The culture filtrates of 3 strains of C. jejuni and C. coli were subjected to neutralisation with cholera antitoxin. The fluid accumulation was completely neutralised up to 1 in 80 dilution showing immunobiological relationship between C. jejuni and C. coli enterotoxin and cholera toxin.