Antimicrobial resistance pattern of anaerobic bacteria causing lower respiratory tract infections.

BACKGROUND: Anaerobes are normal flora of the human body. However, they can cause serious infections in humans. Anaerobic bacteria are known to cause respiratory infections like pneumonia and acute exacerbation of chronic lower airway infections. These are often missed due to the complexity of their isolation and identification. Hence, this study aimed to study anaerobes causing respiratory tract infections and determine their antibiotic susceptibility. MATERIALS & METHODS: Clinical specimens such as bronchial aspirates and pleural aspirates collected from patients with respiratory diseases attending Vallabhbhai Patel Chest Institute were processed, the anaerobes isolated were identified, and their susceptibilities to various groups of antimicrobials were studied using standard microbiological methods. RESULTS: Three hundred and fourteen patients were included in the study, 154 males and 160 females. Of these 314 patients, 148 (47%) yielded anaerobes in their clinical samples. Seventy patients had more than one type of anaerobic organism. Hence, 235 isolates were recovered belonging to as many as 17 genera. The MIC of seven antibiotics on 154 isolates was tested. The isolates belonged mostly to the genera Bacteroides, Prevotella, Veillonella, and Actinomyces. Variable resistance was observed to most classes of antibiotics by many genera. CONCLUSIONS: Metronidazole is commonly used against anaerobes, but the study showed that the isolates were 20-30% resistant to the antibiotic. Starting this as an empirical therapy might lead to treatment failure.

Comparative analysis of the alveolar microbiome in COPD, ECOPD, Sarcoidosis, and ILD patients to identify respiratory illnesses specific microbial signatures.

Studying respiratory illness-specific microbial signatures and their interaction with other micro-residents could provide a better understanding of lung microbial ecology. Each respiratory illness has a specific disease etiology, however, so far no study has revealed disease-specific microbial markers. The present study was designed to determine disease-specific microbial features and their interactions with other residents in chronic obstructive pulmonary diseases (stable and exacerbated), sarcoidosis, and interstitial lung diseases. Broncho-alveolar lavage samples (n = 43) were analyzed by SSU rRNA gene sequencing to study the alveolar microbiome in these diseases. A predominance of Proteobacteria followed by Firmicutes, Bacteroidetes, Actinobacteria, and Fusobacteria was observed in all the disease subsets. Shannon diversity was significantly higher in stable COPD when compared to exacerbated chronic obstructive pulmonary disease (ECOPD) (p = 0.0061), and ILD patient samples (p = 0.037). The lung microbiome of the patients with stable COPD was more diverse in comparison to ECOPD and ILD patients (p < 0.001). Lefse analysis identified 40 disease-differentiating microbial features (LDA score (log10) > 4). Species network analysis indicated a significant correlation (p < 0.05) of diseases specific microbial signature with other lung microbiome members. The current study strengthens the proposed hypothesis that each respiratory illness has unique microbial signatures. These microbial signatures could be used as diagnostic markers to differentiate among various respiratory illnesses.

Calcium Dynamics Regulate Protective Responses and Growth of Staphylococcus aureus in Macrophages.

Staphylococcus aureus (S. aureus) is a gram-positive bacteria, which causes various fatal respiratory infections including pneumonia. The emergence of Methicillin-Resistance Staphylococcus aureus (MRSA) demands a thorough understanding of host-pathogen interactions. Here we report the role of calcium in regulating defence responses of S. aureus in macrophages. Regulating calcium fluxes in cells by different routes differentially governs the expression of T cell costimulatory molecule CD80 and Th1 promoting IL-12 receptor. Inhibiting calcium influx from extracellular medium increased expression of IFN-γ and IL-10 while blocking calcium release from the intracellular stores inhibited TGF-ß levels. Blocking voltage-gated calcium channels (VGCC) inhibited the expression of multiple cytokines. While VGCC regulated the expression of apoptosis protein Bax, extracellular calcium-regulated the expression of Cytochrome-C. Similarly, VGCC regulated the expression of autophagy initiator Beclin-1. Blocking VGCC or calcium release from intracellular stores promoted phagosome-lysosome fusion, while activating VGCC inhibited phagosomelysosome fusion. Finally, calcium homeostasis regulated intracellular growth of Staphylococcus, although using different mechanisms. While blocking extracellular calcium influx seems to rely on IFN-γ and IL-12Rß receptor mediated reduction in bacterial survival, blocking either intracellular calcium release or via VGCC route seem to rely on enhanced autophagy mediated reduction of intracellular bacterial survival. These results point to fine-tuning of defence responses by routes of calcium homeostasis.

Data highlighting miR-155 and GAPDH correlation.

This data represents the effect of miR-155 on the expression of commonly used housekeeping genes, GAPDH, Beta Actin, RPL13A, and U6. The human miR-155 and control RNA were transfected to A549 cells by electroporation. Expression of these genes was compared in both groups by real-time PCR. The significant up-regulation in the expression of GAPDH was observed in the miR-155 transfected samples as compared to control while no major change was observed in the expression of the other three genes.

Protective immunity of recombinant LipL21 and I-LipL21 against Leptospira interrogans serovar Autumnalis N2 infection.

INTRODUCTION: Leptospirosis is a zoonotic disease caused by the spirochete of genus Leptospira with widespread distribution in tropical, subtropical and temperate zones. Leptospirosis is often confused with other febrile illnesses including jaundice, dengue, and malaria. Generally, the disease is often underdiagnosed or misdiagnosed. Though leptospirosis is curable with antibiotic treatment, the laboratory diagnosis of the disease is specialized and open to interpretation with multiple kits available to detect the different serological markers of Leptospira. Moreover, when leptospirosis is misdiagnosed, the disease can lead to multi-organ failure and may have fatal effects. There is a need for strategies to develop vaccines and prevent leptospirosis. In the present study, the immunogenic potential of leptospiral recombinant protein LipL21 (rLipL21) and its truncated form I-LipL21 (rI-LipL21) was evaluated. METHODOLOGY: The recombinant proteins were established in cyclophosphamide treated BALB/c mice model infected with L. interrogans serovar Autumnalis strain N2. RESULTS: The vaccination study showed 66% and 83% survivability among mice immunized with rLipL21 and rI-LipL21 respectively and post-challenge with leptospiral strain N2 compared to control groups that showed 100% lethality. Additionally, a significant increase in antibody levels and cytokine levels (TNF-a, IFN-γ and IL-10) was observed evidencing a marked stimulation of both humoral and cell-mediated immune response in mice immunized with rLipL21/rI-LipL21 compared to whole cell leptospiral lysate (WCL). CONCLUSIONS: This study evidenced protective immunization against leptospirosis with rLipL21 and rI-LipL21 recombinant proteins and are potential candidates for the development of leptospiral vaccine.

Corynebacterium striatum: an emerging respiratory pathogen.

INTRODUCTION: Corynebacterium spp. are primarily considered normal flora and dismissed when isolated from clinical specimens. In recent years, Corynebacterium striatum has emerged as a multi-drug resistant human pathogen which can cause nosocomial outbreaks. The organism has infrequently been noted to cause respiratory infections. A retrospective study was conducted to identify the clinical and microbiological features of respiratory infection by Corynebacterium striatum. METHODOLOGY: C. striatum isolates from clinical and surveillance samples were tested for susceptibility to antimicrobials and typed by Random Amplification of Polymorphic DNA (RAPD). Clinical data was obtained through a retrospective review of records. RESULTS: 15 isolates from clinical and surveillance samples of 11 hospitalised patients were included. The patients suffered from either an exacerbation of COPD (n = 9) or pneumonia (n = 2). The isolates were all multi-drug resistant. RAPD typing found no evidence of an outbreak/ transmission between patients. CONCLUSIONS: Corynebacterium spp. must be considered potential pathogens. Suspicious isolates should be identified to the species level since Corynebacterium striatum is often multi-drug resistant.

Similar virulence properties of infection and colonization associated Pseudomonas aeruginosa.

PURPOSE: Pseudomonas aeruginosa is one of the agents that are commonly implicated in nosocomial infections. However, it is also present as a commensal in various body sites of healthy persons, making the diagnosis of infection by culture difficult. A number of virulence factors expressed by the organism have been implicated in its pathogenicity. We undertook this study to identify the host and organism factors associated with infection. METHODOLOGY: Pathogenic, colonizing and environmental isolates were tested for apr, lasB, the T3SS effector exoenzymes (exoS, exoT, exoU and exoY) and toxA genes, biofilm production and antimicrobial susceptibility. The isolates were further typed by RAPD. RESULTS: Eighty-seven isolates from 61 patients, including 11 environmental isolates, were obtained. None of the virulence factors were found to be significantly associated with infection, and nor was the antimicrobial susceptibility. The presence of the exoU gene and infection by MDR strains correlated significantly with the duration of hospital stay. Positivity for exoS and exoU genes was found to be strongly correlated with multi-drug resistance. exoU positivity correlated strongly with fluoroquinolone resistance. Sinks in the ward and intensive care unit were found to be a niche for XDR P. aeruginosa. Eighty-five isolates were typeable using the ERIC2 primer, showing 71 distinct RAPD patterns with >15 % difference in UPGMA-generated dice coefficients. CONCLUSIONS: exoU positivity is associated with severe disease, as evidenced by the longer duration of hospital stay of these patients. However, the presence of virulence factors or multi-drug resistance in the cultured strain should not prompt the administration of anti-pseudomonal chemotherapy.

Exacerbation of bronchiectasis by Pseudomonas monteilii: a case report.

BACKGROUND: Pseudomonas spp are important opportunistic and nosocomial pathogens. One such species is Pseudomonas monteilii (P. monteilii). It has been described as an environmental contaminant and potential pathogen. We identified this organism as the causative agent of an exacerbation of bronchiectasis and an environmental contaminant in our hospital on two separate occasions. CASE PRESENTATION: P. monteilii was the cause of an exacerbation of bronchiectasis in a 30-year-old HIV negative male. Patient presented with cough with sputum production and exertional dyspnea. The isolate was recovered from a sputum sample in significant counts and definitively identified by Matrix-Assisted Laser Desorption/Ionisation- Time of Flight Mass Spectrometry (MALDI-TOF MS). He was treated with piperacillin-tazobactam and recovered clinically and microbiologically. Another two isolates of the organism were contaminants from the hospital environment. The three isolates were susceptible to all tested antibiotics. Typing by Random amplification of polymorphic DNA (RAPD) found no clonal relationship between them. CONCLUSIONS: Less common species of Pseudomonas need to be identified accurately. This organism is identified by commonly used phenotypic systems as P. putida which may have contributed to a lower reported prevalence. P. monteilii is a known environmental contaminant and must also be considered as a potential pathogen, particularly in patients with chronic lung disease.

Pulmonary Nocardiosis: Review of Cases and an Update.

Nocardia, a branching, filamentous bacteria, is widely distributed in the environment and can cause human infection in immune-compromised hosts. Inhalation of Nocardia leads to pulmonary disease. Microbiology laboratory processed the clinical samples from patients with respiratory infections. Smears were prepared from the samples and were stained and cultured. Five cases were positive for Nocardia. They were treated with the trimethoprim-sulfamethoxazole combination. The disease was cured in three patients, and two died due to other comorbid conditions leading to complications. Nocardiosis is encountered in parts of the world even where it is not endemic due to increased world travel. So physicians and laboratory staff should be aware of this and try to diagnose it. Early detection can lead to the prompt initiation of treatment and reduced mortality in these patients. Patients with disseminated or severe nocardiosis should be treated with combination therapy with two or more active agents.

Phenotypic and molecular characterization of clinical isolates of Acinetobacter baumannii isolated from Delhi, India.

BACKGROUND: Acinetobacter has gained importance as a multi-drug resistant and hence a difficult to treat pathogen. This study was done to characterize our isolates with respect to drug resistance and presence of beta-lactamases which is a major mechanism of resistance and to type using RAPD and MLST so that comparison of our clones can be made with the existing international clones. METHODS: 100 isolates recovered from clinical samples from two hospitals in Delhi were tested for their susceptibility against major groups of antimicrobials. The resistant isolates were screened and confirmed phenotypically for presence of ESBL, MBL and AmpC and MBLs also by PCR. The isolates were typed by RAPD and MLST. RESULTS: Out of the 100 isolates, 91, 78 and 2 % were MDR, XDR and PDR respectively. 97, 100 and 85 were screen positive for ESBL, AmpC and MBL respectively. Of these, 38.1 % were confirmed phenotypically to produce ESBL, 99 % produced AmpC and 29.4 % produced MBL comprising of GIM, VIM, SIM and IMP. MLST showed known STs 110, 188, 146, 69, 103, 108 and 194. Eight new STs were encountered. The RAPD showed a high degree of genetic variability among the isolates. CONCLUSION: Majority of our isolates were MDR, producing one or more types of beta-lactamases. We encountered drug resistant international clones by MLST which are found in other continents there by confirming their spread to Indian sub continent. No data on ST types of other Indian isolates is available in the MLST database and hence comparison is not possible.

Characterization of Streptococcus pneumoniae isolates from India with special reference to their sequence types.

INTRODUCTION: Streptococcus pneumoniae is a major cause of mortality and morbidity in young children and the elderly. In the present study we evaluated antimicrobial susceptibilities, serotypes, and sequence types of pneumococcal isolates recovered in New Delhi, India. METHODOLOGY: A total of 126 clinical isolates of Streptococcus pneumoniae were investigated. They were subjected to disk diffusion susceptibility testing, broth microdilution testing, serotyping and multilocus sequence typing. RESULTS: Broth microdilution assay showed that 5%, 20% and 23% of the isolates exhibited resistance to penicillin, erythromycin and ciprofloxacin, respectively. Serotypes19, 1 and 6 were more frequently isolated. Thirty per cent of the strains were comprised of serotypes 1, 3, 5, 19A and 7F, which are not included in the seven-valent vaccine. Fifty-nine isolates were typed using multilocus sequence typing. Thirty new sequence types were encountered in this study. Only one clonal complex with 4 isolates was seen; 11 clonal complexes and 96 sequence types (STs) were observed among 115 Indian isolates. Only 18 of the 96 STs were found globally, of which only 4 STs were found in many countries with larger numbers. CONCLUSIONS: This study identifies the non-vaccine serotypes of Streptococcus pneumoniae circulating in India. It is important that an appropriate vaccine which covers all serotypes is used in the region.

Role of Mycoplasma pneumoniae infection in acute exacerbations of chronic obstructive pulmonary disease.

Eighty per cent of the cases of acute exacerbation of chronic obstructive pulmonary disease (AECOPD) have an infective aetiology, atypical bacteria including Mycoplasma pneumoniae accounting for 5-10 % of these. However, the importance of association of M. pneumoniae with episodes of AECOPD still remains doubtful. The present study was therefore undertaken to delineate the extent of involvement of M. pneumoniae in patients with AECOPD at a referral hospital in Delhi, India. Sputum samples and throat swabs from a total of 100 AECOPD patients attending the Clinical Research Center of Vallabhbhai Patel Chest Institute, Delhi, were collected during a 2-year period (January 2004-June 2006). The samples were investigated for the presence of aerobic bacterial pathogens and M. pneumoniae. Diagnosis of infection with M. pneumoniae was based on culture, serology, direct detection of M. pneumoniae specific antigen and PCR. Bacterial aetiology could be established in 16 of the 100 samples studied. Pseudomonas spp. were recovered from eight cases, Streptococcus pneumoniae from four and Klebsiella spp. from two cases. Acinetobacter sp. and Moraxella catarrhalis were isolated from one case each. Serological evidence of M. pneumoniae infection and/or detection of M. pneumoniae specific antigen were seen in 16 % of the cases. One case with definite evidence of M. pneumoniae infection also had coinfection with Pseudomonas spp. However, no direct evidence of M. pneumoniae infection was found in our study population as defined by culture isolation or PCR. In conclusion, although the serological prevalence of M. pneumoniae infection in our study population was significantly higher than in the control group, there was no direct evidence of it playing a role in AECOPD.

Chronic community-acquired Acinetobacter pneumonia that responded slowly to rifampicin in the anti-tuberculous regime.

Acinetobacter baumannii is a well-known cause of hospital-acquired pneumonia. Occasionally, it can present as an acute community-acquired pneumonia with a fulminant course. However, the occurrence of the chronic form of community-acquired Acinetobacter pneumonia is yet to be highlighted. We describe a 62-year-old, HIV negative, non-diabetic male, who was referred for evaluation of consolidation and cavitation in the apicoposterior segment of the left upper lobe for 4 months. For this, he had received anti-tuberculous therapy, which included rifampicin. On investigation, a diagnosis of chronic community-acquired pneumonia due to Acinetobacter baumannii was made. The steady clinico-radiologic improvement observed was attributed to rifampicin in the anti-tuberculous regime. Subsequently, an aspergilloma formed in the cavity.