Treatment of enteric fever in children on the basis of current trends of antimicrobial susceptibility of Salmonella enterica serovar typhi and paratyphi A

Purpose: Recent reports indicate decreased susceptibility of S. typhi to fluoroquinolones, especially ciprofloxacin. Chloramphenicol has been suggested as first line therapy of enteric fever in many studies. This is a prospective study that describes the trends of antimicrobial susceptibility of S. typhi and S. paratyphi A causing bacteraemia in children and reports therapeutic failure to ciprofloxacin and evaluates the possible use of chloramphenicol, ampicillin, ciprofloxacin and third generation cephalosporins as first line therapy in the treatment of enteric fever in children. Methods: The present study was conducted from April 2004 to March 2005 in a superspeciality children hospital at New Delhi. A total of 56 S. typhi and five S. paratyphi A isolates were obtained among the 673 blood cultures performed. Antimicrobial testing was done using disk diffusion technique (NCCLS method) for 13 antimicrobials and MICs were calculated for ampicillin, ciprofloxacin, chloramphenicol and cefotaxime. Analysis of data was done using WHONET software. Results: All 56 isolates of S. typhi were sensitive to amoxycillin+clavulanate, gentamicin, cefixime, cefotaxime and ceftazidime. Multidrug resistance (MDR, resistance to three drugs) was seen in 22 cases (39%) and resistance to five drugs was seen in 12 cases (21%). Only two isolates were resistant to chloramphenicol (3%). MIC₉₀ for ampicillin, chloramphenicol, ciprofloxacin and cefotaxime were 1.0 μg/ml, 4.0 μg/ml, 64 μg/ml and 0.125 μg/ml respectively. All S. paratyphi A isolates were sensitive to ampicillin and chloramphenicol and resistant to nalidixic acid.MIC distribution data for chloramphenicol revealed elevated MIC but still in susceptible range. Conclusions: There is an urgent need for further clinical studies to evaluate response to chloramphenicol in such cases. Antimicrobial susceptibility data and MIC distribution favour use of ampicillin as a drug of choice for the treatment of enteric fever. Third generation cephalosporins are also useful but their use should be restricted for complicated cases.     

In-vitro susceptibility testing by agar dilution method to determine the minimum inhibitory concentrations of amphotericin B, fluconazole and ketoconazole against ocular fungal isolates

Purpose: To standardize in-vitro antifungal susceptibility testing by agar dilution method to find out the minimum inhibitory concentration (MIC) of amphotericin B, fluconazole and ketoconazole on ocular fungal isolates. Methods: A total of 180 ocular fungal isolates (130 filamentous fungi and 50 yeasts) were included. The antifungal drugs such as amphotericin B (0.0625-8 μg/mL), fluconazole (0.2-819.6 μg/mL) and ketoconazole (0.025-6.4 μg/mL) were incorporated in doubling dilutions in the yeast nitrogen base medium. The MIC was determined as the lowest concentration of the antifungal drug preventing growth of macroscopically visible colonies on drug containing plates when there was visible growth on the drug - free control plates. Results: All 50 ocular isolates of yeast were susceptible to amphotericin B, while two (4%) and five (10%) strains were resistant to fluconazole and ketoconazole respectively. Of the 130 filamentous fungi tested, six (4.6%) were resistant to amphotericin B, 49 (37.7%) and 10 (7.6%) were resistant to fluconazole and ketoconazole respectively. Percentile 50 (MIC 50) and Percentile 90 (MIC 90) for all the three antifungal agents were calculated. Aspergillus niger, Aspergillus terreus and Candida krusei were found to be resistant to fluconazole and ketoconazole. Conclusion: This technique was found to be reliable, cost effective and easy to perform with consistent results.     

Six-year susceptibility trends and effect of revised Clinical Laboratory Standards Institute breakpoints on ciprofloxacin susceptibility reporting in typhoidal Salmonellae in a tertiary care paediatric hospital in Northern India

The antimicrobial trends over 6 years were studied, and the effect of revised Clinical Laboratory Standards Institute (CLSI) breakpoints (2012) for ciprofloxacin susceptibility reporting in typhoidal Salmonellae was determined. A total of 874 (95.4%) isolates were nalidixic acid-resistant (NAR). Using the CLSI 2011 guidelines (M100-S21), 585 (66.9%) isolates were ciprofloxacin susceptible. The susceptibility reduced to 11 (1.25%) isolates when interpreted using 2012 guidelines (M100-S22). Among the forty nalidixic acid susceptible (NAS) Salmonellae, susceptibility to ciprofloxacin decreased from 37 isolates (M100-S21) to 12 isolates (M100-S22). The 25 cases which appeared resistant with newer guidelines had a minimum inhibitory concentration (MIC) range between 0.125 and 0.5 μg/ml. MIC50 for the third generation cephalosporins varied between 0.125 and 0.5 μg/ml over 6 years whereas MIC90 varied with a broader range of 0.19–1 μg/ml. The gap between NAR and ciprofloxacin-resistant strains identified using 2011 guidelines has been reduced; however, it remains to be seen whether additional NAS, ciprofloxacin-resistant isolates are truly resistant to ciprofloxacin by other mechanisms of resistance.     

Antimicrobial susceptibility testing of Brucella melitensis isolated from patients with acute brucellosis in a centre of Iran

Structured to Purpose: Human brucellosis is one of the most common zoonotic infections worldwide, which remains one of the major problems for public health. Despite the World Health Organization’s recommendation for human brucellosis treatment, sporadic cases of relapse have been reported. The aim of this study was to assess the susceptibility of Brucella isolates to common antibiotics that are prescribed by the physician for the treatment of brucellosis and also to determine the minimum inhibitory concentration 50% (MIC₅₀) and MIC₉₀ for these antibiotics. Materials and Methods: Forty-eight Brucella strains were collected from patients with acute brucellosis. Species identification was made based on the conventional methods. MIC of rifampin, doxycycline, ciprofloxacin, trimethoprim-sulfamethoxazole, streptomycin, azithromycin and ceftriaxone was determined by E-test. Results: All the 48 Brucella isolates (47 blood samples and one synovial fluid) were identified as Brucella melitensis. No antimicrobial-resistant strains were recognised. Trimethoprim-sulfamethoxazole had the lowest MIC₅₀ (0.016 μg/ml) and MIC₉₀ (0.064 μg/ml), whereas MIC₅₀ and MIC₉₀ of streptomycin and azithromycin had the highest level at 0.625, 1.5 µg/ml and 0.25, 1 µg/ml, respectively. All the isolates were susceptible to rifampin, and only one of the isolates had a reduced sensitivity to rifampin (1 μg/ml). Conclusions: Although all the Brucella isolates were susceptible, antimicrobial susceptibility test should be recommended in patients with recurrent brucellosis or life-threatening organ involvement.     

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 MIC₉₀ 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 MIC₉₀ 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.     

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.     

Determination of minimum inhibitory concentrations of itraconazole,terbinafine and ketoconazole against dermatophyte species by broth microdilution method

Purpose: Various antifungal agents both topical and systemic have been introduced into clinical practice for effectively treating dermatophytic conditions. Dermatophytosis is the infection of keratinised tissues caused by fungal species of genera Trichophyton, Epidermophyton and Microsporum, commonly known as dermatophytes affecting 20–25% of the world’s population. The present study aims at determining the susceptibility patterns of dermatophyte species recovered from superficial mycoses of human patients in Himachal Pradesh to antifungal agents; itraconazole, terbinafine and ketoconazole. The study also aims at determining the minimum inhibitory concentrations (MICs) of these agents following the recommended protocol of Clinical and Laboratory Standards Institute (CLSI) (M38-A2). Methodology: A total of 53 isolates of dermatophytes (T. mentagrophyte-34 in no., T. rubrum-18 and M. gypseum-1) recovered from the superficial mycoses were examined. Broth microdilution method M38-A2 approved protocol of CLSI (2008) for filamentous fungi was followed for determining the susceptibility of dermatophyte species. Results: T. mentagrophyte isolates were found more susceptible to both itraconazole and ketoconazole as compared to terbinafine (MIC₅₀: 0.125 μg/ml for itraconazole, 0.0625 μg/ml for ketoconazole and 0.5 μg/ml for terbinafine). Three isolates of T. mentagrophytes (VBS-5, VBSo-3 and VBSo-73) and one isolate of T. rubrum (VBPo-9) had higher MIC values of itraconazole (1 μg/ml). Similarly, the higher MIC values of ketoconazole were observed in case of only three isolates of T. mentagrophyte (VBSo-30 = 2 μg/ml; VBSo-44, VBM-2 = 1 μg/ml). The comparative analysis of the three antifungal drugs based on t-test revealed that ‘itraconazole and terbinafine’ and ‘terbinafine and ketoconazole’ were found independent based on the P < 0.005 in case of T. mentagrophyte isolates. In case of T. rubrum, the similarity existed between MIC values of ‘itraconazole and ketoconazole’ and ‘terbinafine and ketoconazole’. Conclusion: The MIC values observed in the present study based on standard protocol M38-A2 of CLSI 2008 might serve as reference for further studies covering large number of isolates from different geographic regions of the state. Such studies might reflect on the acquisition of drug resistance among isolates of dermatophyte species based on MIC values.     

Rising trend of antimicrobial resistance among Neisseria gonorrhoeae isolates and the emergence of N. gonorrhoeae isolate with decreased susceptibility to ceftriaxone

Context: Gonorrhoea is one of the most common sexually transmitted infections (STI) in developing countries and is a global health problem. Aims: To analyze the trend of antimicrobial susceptibility of Neisseria gonorrhoeae isolates over the years, in a tertiary care hospital of North India. Settings and Design: The study population comprised males with urethritis and females with endocervicitis attending STI clinic of our hospital. Materials and Methods: In our STI laboratory, all gonococcal isolates are subjected to antimicrobial susceptibility testing by disc diffusion method as per CLSI guidelines. ß-lactamase production is determined by chromogenic cephalosporin test. Minimum Inhibitory Concentration (MIC) for ceftriaxone is determined by E-test. Statistical Analysis Used: Data were expressed as percentages. The differences in percentages were tested for statistical significance by using χ² test and P values were determined. Results: The percentage of penicillinase producing N. gonorrhoeae (PPNG) increased from 8% in 1995-96 to 20% in 2004-05 and 88% in 2011-2013. Quinolone-resistant N. gonorrhoeae (QRNG) showed a significant increase from 12% in 1995-96 to 98.3% in 2004-05, while 84% isolates were found to be QRNG by 2011-2013. In January 2013 we detected our first gonococcal isolate with decreased susceptibility to third-generation cephalosporins; Ceftriaxone, Cefixime and Cefpodoxime (MIC for ceftriaxone = 0.19 μg/ml). Conclusions: The results of our study highlighted an alarming increase in the percentage of PPNG and QRNG strains over the years. Emergence of N. gonorrhoeae isolates with decreased susceptibility to third-generation cephalosporins is a cause of concern and thus emphasises the importance of antimicrobial susceptibility testing.     

Changing Trends of Culture-positive Typhoid Fever and Antimicrobial Susceptibility in a Tertiary Care North Indian Hospital over the Last Decade

Purpose: The present study was undertaken to analyse the trend in prevalence of culture-positive typhoid fever during the last decade and to determine antimicrobial susceptibility profile of Salmonella Typhi and Salmonella Paratyphi A isolated from patients of enteric fever presenting to our hospital. Methods: All the culture-positive enteric fever cases during 2005–2016 presenting to our Hospital were included in the study. Antimicrobial susceptibility was done against chloramphenicol, amoxicillin, co-trimoxazole, ciprofloxacin, ofloxacin, levofloxacin, pefloxacin, ceftriaxone and azithromycin as per corresponding CLSI guidelines for each year. We also analysed the proportion of culture positivity during 1993–2016 in light of the antibiotic consumption data from published literature. Results: A total of 1066 strains-S. Typhi (772) and S. Paratyphi A (294) were isolated from the blood cultures during the study. A maximum number of cases were found in July–September. Antimicrobial susceptibility for chloramphenicol, amoxicillin and co-trimoxazole was found to be 87.9%, 75.5%, 87.3% for S. Typhi and 94.2%, 90.1% and 94.2% for S. Paratyphi A, respectively. Ciprofloxacin, ofloxacin and levofloxacin susceptibility were 71.3%, 70.8% and 70.9% for S. Typhi and 58.1%, 57.4% and 57.1% for S. Paratyphi A, respectively. Azithromycin susceptibility was 98.9% in S. Typhi. Although susceptibility to ceftriaxone and cefixime was 100% in our isolates, there is a continuous increase in ceftriaxone minimum inhibitory concentration (MIC)₅₀ and MIC₉₀ values over the time. The proportion of blood culture-positive cases during 1993–2016 ranged from a minimum of 0.0006 in 2014 to a maximum of 0.0087 in 1999. Conclusion: We found that the most common etiological agent of enteric fever is S. Typhi causing the majority of cases from July to October in our region. MIC to ceftriaxone in typhoidal salmonellae is creeping towards resistance and more data are needed to understand the azithromycin susceptibility.     

Antimicrobial susceptibility of Neisseria gonorrhoeae isolates and treatment of gonorrhoea patients in Ternopil and Dnipropetrovsk regions of Ukraine, 2013–2018

Antimicrobial resistance (AMR) in Neisseria gonorrhoeae is a major public health concern globally. However, recent gonococcal AMR data from Eastern Europe are extremely limited and no AMR data for strains spreading in Ukraine have ever been internationally published. We investigated the AMR of N. gonorrhoeae isolates in two regions of Ukraine (Ternopil 2013–2018, Dnipropetrovsk 2013–2014), and, where information was available, the treatment administered to the corresponding gonorrhoea patients. Determination of minimum inhibitory concentration (MIC) of eight antimicrobials was performed using Etest and resistance breakpoints from the European Committee on Antimicrobial Susceptibility Testing (EUCAST) were applied. Overall, 9.3% of the examined 150 isolates were resistant to ciprofloxacin, 6.0% to tetracycline, 2.0% to azithromycin, and 0.7% to benzylpenicillin. No isolates were resistant to ceftriaxone, cefixime, spectinomycin, or gentamicin. However, one (0.7%) isolate showed a MIC value of 0.125 mg/L for both ceftriaxone and cefixime, i.e., bordering resistance. Eighty‐eight (67.2%) of 131 patients were administered dual therapy (ceftriaxone 1 g plus doxycycline/clarithromycin/azithromycin/ofloxacin) and 22 (16.8%) ceftriaxone 1 g monotherapy. Worryingly, 21 (16.0%) patients received monotherapy with clarithromycin/doxycycline/azithromycin/ofloxacin/benzylpenicillin. In conclusion, the antimicrobial susceptibility of gonococcal strains spreading in Ternopil and Dnipropetrovsk, Ukraine during 2013–2018 was high. Low levels of resistance to ciprofloxacin, tetracycline, azithromycin, and benzylpenicillin were found, but no resistance to the internationally recommended ceftriaxone, cefixime, or spectinomycin. Ceftriaxone 1 g should remain as empiric first‐line treatment, in dual therapy with azithromycin or doxycycline or in monotherapy. Continued and expanded gonococcal AMR surveillance in Ukraine is essential to monitor the susceptibility to particularly extended‐spectrum cephalosporins, azithromycin and doxycycline.     

Correlation between In vitro susceptibility and treatment outcome with azithromycin in gonorrhoea: a prospective study

PURPOSE: This prospective study was carried out to determine the antimicrobial susceptibility of Neisseria gonorrhoeae isolates by disc diffusion method and minimum inhibitory concentration (MIC) by E -test with special reference to azithromycin. Also, the correlation between in vitro susceptibility and treatment outcome with single 2 g oral dose azithromycin was assessed. METHODS: The study included 75 gonococcal isolates from males with urethritis, females with endocervicitis and their sexual contacts. All isolates were subjected to susceptibility testing for penicillin, ciprofloxacin, tetracycline, ceftriaxone, spectinomycin, cefixime and azithromycin. Males with gonococcal urethritis were randomised to receive a single dose of either azithromycin or ceftriaxone. Forty-two men with urethritis received 2 g single oral dose azithromycin, while all other patients were given 250 mg parentral ceftriaxone. All patients were called for follow-up to assess clinical and microbiological cure rates. RESULTS: While all the isolates were susceptible to ceftriaxone, spectinomycin, cefixime and azithromycin; 74 (98.7%), 24 (32%) and 23 (30.7%) strains were resistant to ciprofloxacin, penicillin and tetracycline respectively, by both disc diffusion method and E -test. The MIC range, MIC50 and MIC90 of N. gonorrhoeae strains, to azithromycin were 0.016-0.25, 0.064 and 0.19 microg/mL, respectively. Follow-up attendance of the patients was 52.4 with 100% clinical and microbiological cure rates. CONCLUSIONS: Results of our study indicate that 2 g single oral dose azithromycin is safe and effective in the treatment of uncomplicated gonorrhoea.     

Trends in antimicrobial susceptibility of Salmonella Typhi from North India (2001-2012)

Purpose: Enteric fever is endemic in India with Salmonella Typhi being the major causative agent. Antibiotic therapy constitutes the mainstay of management. The present study was undertaken to find the susceptibility profile of Salmonella enterica var Typhi (S. Typhi) blood isolates in a tertiary care hospital between January 2001 and December 2012. Materials and Methods: A retrospective analysis of laboratory records was carried out. Conventional blood culture method was used until 2009; from January 2010 onwards BACTEC 9240 system has been in use. Salmonella were confirmed by serotyping using group and type specific antisera. Antibiotic susceptibility was performed using the disk diffusion method. In addition 116 isolates were subjected to minimum inhibitory concentration testing for chloramphenicol, ciprofloxacin, amoxicillin and nalidixic acid (NA) using agar dilution and for ceftriaxone and azithromycin using E-strips (Biomerieux). Result: A total of 1016 typhoidal salmonellae were obtained. The predominant serotype obtained was S. Typhi (852, 83.8%) followed by Salmonella enterica var Paratyphi A (164, 16.2%). We observed a re-emergence of susceptibility to first line antibiotics and a notable decline in multidrug resistant (MDR) strains. We also found all recent isolates resistant to NA and susceptible to third generation cephalosporins and 84.5% of isolates having decreasing ciprofloxacin susceptibility using revised criteria as per Clinical and Laboratory Standards Institute 2012 guidelines. Conclusion: There has been re-emergence of susceptibility to first line antibiotics and a notable decline in MDR strains of S. Typhi. We have a very high resistance to NA and decreasing susceptibility to ciprofloxacin. Third generation cephalosporins and azithromycin seem to be effective therapeutic options. Judicious use of these antibiotics is mandatory to prevent emergence of resistant strains.     

Alarming Resistance of Neisseria gonorrhoeae in a Tertiary Care Hospital of North India

Background: This study attempted to elucidate the spectrum of sexually transmitted infections in a tertiary care centre in North India and to assess the antimicrobial resistance in Neisseria gonorrhoeae. Materials and Methods: Antimicrobial resistance pattern of N. gonorrhoeae was determined by the standard techniques. Genotypic detection of gyrA, parC and blaTEM genes was also carried out. The results of gyrA gene by polymerase chain reaction were confirmed by DNA sequencing. Results: N. gonorrhoea was identified in 10 (4.98%) patients, and antimicrobial sensitivity was performed in seven patients. All the seven patients tested were quinolone-resistant N. gonorrhoeae (QRNG), 5/7 were penicillinase-producing N. gonorrhoeae, 1/7 was chromosomally mediated penicillin-resistant N. gonorrhoeae and 3/7 were tetracycline-resistant N. gonorrhoeae. Minimal inhibitory concentration (MIC) by E-test was performed in five strains, and we observed that MIC90 for ciprofloxacin was ≥4 μg/ml, for penicillin was ≥6 μg/ml and for tetracycline was 12 μg/ml, which clearly brackets them as resistant isolates. The presence of TEM gene was confirmed genotypically in six out of seven cases. In all seven cases, gyrA and parC were observed, thus confirming their QRNG status. Conclusion: Alarming increase in the resistance to commonly used antimicrobials for gonorrhoea in our study, especially of fluoroquinolones, is a clarion call for the urgent need for prudence in prescribing them. Observing the rampant resistance exhibited by N. gonorrhoeae, it is clear that the day is not far when it will acquire a superbug status and become intractable to treatment by the available antibiotics.     

Epidemiology and Antifungal Susceptibility of Infections Caused by Trichosporon Species: An Emerging Non-Candida and Non-Cryptococcus Yeast Worldwide

Introduction: Over the past four decades, there has been an increase in the number of fatal opportunistic invasive trichosporonosis cases especially in immunocompromised hosts. Objective: The objective of the study is to evaluate the epidemiological, clinical details and antifungal susceptibility pattern of the patients with Trichosporon infections. Materials and Methods: Twenty-four clinical isolates of Trichosporon species isolated from blood, samples, pleural fluid and nail were included in this study, over a period of 12 years (2005–2016) in a tertiary hospital in North India. The isolates were characterised phenotypically and few representative isolates were sequenced also. The minimum inhibitory concentration (MIC) was determined as per Clinical and Laboratory Standards Institute, 2012. Results: Trichosporon spp. from blood culture (57.78%), nail (37.5%) and pleural fluid (4.17%). On phenotypic tests, 79.16% of the isolates were Trichosporon asahii, followed by Trichosporon dermatis (8.33%), Trichosporon japonicum (4.17%), Trichosporon ovoides (4.17%) and Trichosporon mucoides (4.17%). The MIC range of Trichosporon species from invasive infections were fluconazole (0.06–256 μg/ml), amphotericin B (0.125–16 μg/ml), voriconazole (0.0616–8 μg/ml), posaconazole (0.0616–32 μg/ml) and caspofungin (8–32 μg/ml). The isolates from superficial infection were resistant to fluconazole (0.06–256 μg/ml) and itraconazole (0.125–32 μg/ml), all were susceptible to ketoconazole and while only two were resistant to voriconazole (0.25–4 μg/ml). Conclusion: T. asahii was the most common isolate. Disseminated trichosporonosis is being increasingly reported worldwide including India and represents a challenge for both diagnosis and species identification. Prognosis is limited, and antifungal regimens containing triazoles appear to be the best therapeutic approach. In addition, accurate identification, removal of central venous lines and voriconazole-based treatment along with control of underlying conditions were associated with favourable outcomes.     

Changing antimicrobial susceptibility patterns among Streptococcus pneumoniae and Haemophilus influenzae from Brazil: Report from the SENTRY Antimicrobial Surveillance Program (1998-2004)

Although antimicrobial resistance rates among Streptococcus pneumoniae and Haemophilus influenzae have increased significantly in most countries in the last years, most studies from Brazil report relatively low resistance rates among these pathogens. In this study, we analyzed the susceptibility patterns of S. pneumoniae and H. influenzae from Brazil during a 7-year period. A total of 829 S. pneumoniae and 718 H. influenzae consecutively collected from 1998 to 2004, mainly from respiratory tract and bloodstream infections, were susceptibility tested by broth microdilution methods against >30 drugs and the results were analyzed by year. Overall, 77.8% of S. pneumoniae strains were considered susceptible (MIC, < or =0.06 microg/ml) to penicillin. Resistance to penicillin (MIC, > or =2 microg/ml) and ceftriaxone (MIC, > or =4 microg/ml) were detected in 7.5 and 0.5% of strains, respectively. The fluoroquinolones, levofloxacin (MIC90) 1 microg/ml) and gatifloxacin (MIC90, 0.5 microg/ml), were active against 99.8% of the isolates tested. Among the other non-beta-lactam drugs tested, the rank order of susceptibility rates was chloramphenicol (98.9%) > clindamycin (96.4%) > erythromycin (90.6%) > tetracycline (69.8%) > trimethoprim/sulfamethoxazole (36.7%). Resistance to penicillin has increased markedly among S. pneumoniae isolates over 7 years (from 2.9 to 11.0%). Additionally, resistance rates against erythromycin, clindamycin, and tetracycline decreased among pneumococcal strains during the same period. S. pneumoniae recovered from pediatric patients (< or =5 years) showed increased penicillin and trimethoprim/sulfametroxazole resistance rates compared to older populations. The rate of ampicillin resistance among H. influenzae was 14.0%, which also corresponds with the beta -lactamase production rate. All H. influenzae isolates were susceptible to amoxicillin/clavulanate (MIC90, 1 microg/ml), ceftriaxone (MIC90, < or =0.008 microg/ml), cefepime (MIC90, 0.12 microg/ml), ciprofloxacin (MIC90, < or = 0.12microg/ml), levofloxacin (MIC90, < or =0.5 microg/ml), and gatifloxacin (MIC90, < or =0.03 microg/ml). Resistance to the antimicrobials tested remained very stable among H. influenzae isolates during the 7-year study period. The continued emerging antimicrobial resistances found in these pathogens (mainly S. pneumoniae) highlight the need for alternative agents for the treatment of infections caused by these species.     

Susceptibility trends of pseudomonas species from corneal ulcers

Purpose: To assess the changing trends in the antibiotic susceptibility of Pseudomonas spp. isolated from bacterial keratitis over a nine year period with special emphasis on fluoroquinolone susceptibilities. Methods: All corneal scraping cultures positive for Pseudomonas spp. (n=585) isolated from patients with bacterial keratitis at the Aravind Eye Hospital, Madurai from1995-2003 were evaluated. Cultures were performed in liquid and solid media and susceptibility testing was done against amikacin, gentamicin, tobramycin, ciprofloxacin and ofloxacin by Kirby-Bauer disc diffusion method. Results: The susceptibility of Pseudomonas spp. was over 90% from 1995-1998 to ciprofloxacin which decreased to 83% from 1999-2003. The total number of isolates resistant to ciprofloxacin was 51(9.4%). No statistically significant increase in the number of isolates resistant to ciprofloxacin was noted. Ofloxacin showed 54% susceptibility from 1995-1998 but increased to 64% from 1999-2003. Analysis of in vitro activity of amikacin reveals that there was 43% sensitivity from 1995-1998 but later it increased to 76% from 1999-2003. In case of gentamicin, the sensitivity decreased marginally from 80% to 70% through the years. Tobramycin showed 45% sensitivity from 1995-1998 but increased to 75% from 1999-2003. Conclusions: The fluoroquinolones remain a good choice in the treatment of ocular infections, with high susceptibility of Pseudomonas spp. Among the aminoglycosides, gentamicin was found to be highly effective against Pseudomonas corneal ulcers when compared to amikacin and tobramycin. The results show a need for continuous monitoring of bacterial resistance trends.     

Study of the role of efflux pump in ciprofloxacin resistance in Salmonella enterica serotype Typhi

Purpose: There are increasing reports on failure of clinical response to ciprofloxacin in typhoid fever despite the strain being sensitive to drug in in-vitro using standard guidelines and showing mutations in DNA gyrase. But this increased MIC and clinical failures with ciprofloxacin are not always co-related with mutations presently identified in gyrA and parC genes. This shows that there may be other mechanisms such as an active drug efflux pump responsible as has been shown in other Enterobacteriaceae. This study was carried out to determine the role of efflux pump in Salmonella Typhi isolates. Materials and Methods: Total 25 already characterized nalidixic acid sensitive and nalidixic acid resistant S. Typhi strains with different range of ciprofloxacin MIC were included to study the role of efflux pump in the presence of CCCP (efflux pump inhibitor). For genotypic characterization, the entire acrR gene was sequenced to confirm the presence of any mutation in the gene. Results: The MIC of ciprofloxacin remained same in the presence and absence of CCCP in the studied strains and no significant mutations were found in the acrR gene in any of the isolates studied. Conclusions: No role of efflux pump in ciprofloxacin resistance was found in strains studied. There is a need to explore further mechanism of ciprofloxacin resistance in Salmonella Typhi.     

Will Ceftazidime/Avibactam Plus Aztreonam be Effective for NDM and OXA-48-Like Producing Organisms: Lessons Learnt from In vitro Study

Introduction: Carbapenem resistance (CR) in Klebsiella pneumoniae is mainly mediated by blaNDM and blaOXA-48 carbapenemases. Newer Food and Drug Administration-approved antimicrobial ceftazidime/avibactam (C/A) has a potent activity against blaOXA-48-like producers. However, its activity is limited in organisms co-producing blaNDM and blaOXA-48-like. Addition of aztreonam (ATM) to C/A potentially expands the spectrum of coverage for carbapenemase co-producers. With this, we aimed to determine the synergistic activity of combination of C/A plus ATM against blaNDM, blaOXA-48-like and co-producers of blaNDM + blaOXA-48-like producing CR Klebsiella pneumoniae (CRKp). Materials and Methods: A total of 12 isolates of CRKp-harbouring genes encoding blaNDM and blaOXA-48-like were tested. Minimum inhibitory concentrations (MICs) were determined for several antimicrobial agents, including C/A (0.5–8 μg/ml) by broth microdilution method. Checkerboard assay was performed for the combination of C/A plus ATM at varying concentrations. Fold differences in the MIC of C/A with and without addition of ATM were determined to infer synergistic effects. Results: MIC of C/A and ATM ranged from 0.5 to >8 μg/ml and 64 to 2048 μg/ml, respectively. Two isolates were susceptible to C/A with MIC of 0.5 and 1 μg/ml, while others were resistant with MIC of >8 μg/ml. Synergistic effects of >8-fold MIC difference in C/A MIC were noted with addition of ATM at 4 μg/ml. This was observed for all CRKp with profiles of blaNDM, blaOXA-48-like and co-producers of blaNDM + blaOXA-48-like genes, which was a promising effect. Notably, all five of the colistin-resistant CRKp were inhibited with >8-fold MIC difference in the combination of C/A plus ATM at 4 μg/ml. Conclusion: With the increasing burden of CRKp, the use of C/A with ATM combination seems to be very promising, especially for blaNDM, blaOXA-48-like and co-producers of blaNDM + blaOXA-48like carbapenemases.     

Acquired fluconazole resistance and genetic clustering in Diutina (Candida) catenulata from clinical samples

ObjectivesDiutina (Candida) catenulata is an ascomycetous yeast isolated from environmental sources and animals, occasionally infecting humans. The aim of this study is to shed light on the in vitro antifungal susceptibility and genetic diversity of this opportunistic yeast.MethodsForty-five D. catenulata strains isolated from various sources (including human and environmental sources) and originating from nine countries were included. Species identification was performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and confirmed via internal transcribed spacer ribosomal DNA barcoding. In vitro antifungal susceptibility was determined for seven systemic antifungals via the gradient strip method after 48 hours of incubation at 35°C using Etest® (Biomérieux) or Liofilchem® strips. Isolates exhibiting fluconazole minimal inhibitory concentrations (MICs) of ≥8 μg/mL were investigated for mutations in the ERG11 gene. A novel microsatellite genotyping scheme consisting of four markers was developed to assess genetic diversity.ResultsMIC ranges for amphotericin B, caspofungin, micafungin, isavuconazole, and posaconazole were 0.19–1 μg/mL, 0.094–0.5 μg/mL, 0.012–0.064 μg/mL, 0.003–0.047 μg/mL, and 0.006–0.032 μg/mL, respectively. By comparison, a broad range of MICs was noted for fluconazole (0.75 to >256 μg/mL) and voriconazole (0.012–0.38 mg/L), the higher values being observed among clinical strains. The Y132F amino acid substitution, associated with azole resistance in various Candida species (C. albicans, C. tropicalis, C. parapsilosis, and C. orthopsilosis), was the main substitution identified. Although microsatellite typing showed extensive genetic diversity, most strains with high fluconazole MICs clustered together, suggesting human-to-human transmission or a common source of contamination.DiscussionThe high rate of acquired fluconazole resistance among clinical isolates of D. catenulata is of concern. In this study, we highlight a link between the genetic diversity of D. catenulata and its antifungal resistance patterns, suggesting possible clonal transmission of resistant isolates.     

Antifungal drug susceptibility profile of clinically important dermatophytes and determination of point mutations in terbinafine-resistant isolates

With regard to increasing number of antifungal-resistant dermatophytes, antifungal susceptibility testing of dermatophytes serves as a useful tool in managing clinical dermatophytosis. This study aimed to determine antifungal susceptibility profile of clinically important dermatophytes and determination of point mutations in terbinafine-resistant isolates. Based on our results, dermatophytosis was confirmed in 97 cases by direct microscopic examination, culture, and sequencing of ITS region. Antifungal susceptibility of 97 dermatophyte isolates distributed in four species including Trichophyton interdigitale (26 isolates), T. rubrum (19 isolates), T. tonsurans (29 isolates), and Epidermophyton floccosum (21 isolates) was assessed to nine antifungal agents using CLSI M38-A2 guidelines. Minimum inhibitory concentration range (MIC range) for luliconazole and terbinafine was 0.001–0.008 μg/ml and 0.003–> 32 μg/ml, compared to 0.03–64 μg/ml for griseofulvin, 0.01–16 μg/ml for itraconazole and voriconazole, 0.03–8 μg/ml for ketoconazole, 0.03–32 μg/ml for econazole, 0.03–1 μg/ml for lanoconazole, and 0.01–4 μg/ml for butenafine. Trichophyton tonsurans was the most susceptible (MIC = 0.006 μg/ml) and E. floccosum was the most resistant (MIC = 0.02 μg/ml) species to terbinafine. Terbinafine resistance was reported for two species, i.e., T. rubrum and T. tonsurans at the total rate of 2% which was due to Leu393Phe substitution in both species. Taken together, our results assist clinicians and prompt the current knowledge about the necessity of antifungal susceptibility testing to select effective strategies for management of clinical cases of dermatophytosis.