Physiological Constants of the Entomopathogenic Bacterium Xenorhabdus nematophila Determined by Microbial Growth Kinetics

Xenorhabdus nematophila, an entomopathogenic bacterium that symbiotically associates with the entomoparasitic nematode Steinernema carpocapsae, was studied to determine its physiological parameters of glucose utilization. X. nematophila was cultured in chemically defined media containing various concentrations of glucose under optimal conditions utilizing a two-liter fermentation system. Specific growth rates were obtained from each glucose batch. Specific growth rates and their associated glucose concentrations were used to determine physiological parameters. These parameters include the bacterium’s substrate utilization constant (Ks) and its maximum specific growth rate (μmax). The bacteria exhibited a Ks value of 2.02 mg/L suggesting that X. nematophila has a high affinity for glucose. The μmax of Xenorhabdus was determined to be 1.03 h−1. Further research is needed to determine if microbial affinities to different substrates have any influence on biological relationships (symbiosis, pathogenicity, parasitism, etc.) between prokaryotes and higher organisms.

Antibiotic Resistance Pattern and Evaluation of Metallo-Beta Lactamase Genes Including bla-IMP and bla-VIM Types in Pseudomonas aeruginosa Isolated from Patients in Tehran Hospitals

Beta-lactamase producing strains of Pseudomonas aeruginosa are important etiological agents of hospital infections. Carbapenems are among the most effective antibiotics used against Pseudomonas infections, but they can be rendered infective by group B β-lactamase, commonly called metallo-beta lactamase. In this study, the antimicrobial sensitivity patterns of P. aeruginosa strains isolated from 9 different hospitals in Tehran, Iran, as well as the prevalence of MBLs genes (bla-VIM and bla-IMP) were determined. A total of 212 strains of P. aeruginosa recovered from patients in hospitals in Tehran were confirmed by both biochemical methods and PCR. Their antimicrobial sensitivity patterns were determined by Kirby-Bauer disk diffusion method. Following MIC determination, imipenem resistant strains were selected by DDST method which was followed by PCR tests for determination of MBLs genes: bla-IMP and bla-VIM. The results indicated that, in the DDST phenotypic method, among the 100 imipenem resistant isolates, 75 strains were MBLs positive. The PCR test indicated that 70 strains (33%) carried bla-VIM gene and 20 strains (9%) harbored bla-IMP. The results indicated that the extent of antibiotic resistance among Pseudomonas aeruginosa is on the rise. This may be due to production of MBLs enzymes. Therefore, determination of antibiotic sensitivity patterns and MBLs production by these bacteria, can be important in control of clinical Pseudomonas infection.

Function and Evolution of the Sox Multienzyme Complex in the Marine Gammaproteobacterium Congregibacter litoralis

Core sets of sox genes were detected in several genome sequenced members of the environmental important OM60/NOR5 clade of marine gammaproteobacteria. However, emendation of media with thiosulfate did not result in stimulation of growth in two of these strains and cultures of Congregibacter litoralis DSM 17192T did not oxidize thiosulfate to sulfate in concentrations of one mmol L−1 or above. On the other hand, a significant production of sulfate was detected upon growth with the organic sulfur compounds, cysteine and glutathione. It was found that degradation of glutathione resulted in the formation of submillimolar amounts of thiosulfate in the closely related sox-negative strain Chromatocurvus halotolerans DSM 23344T. It is proposed that the Sox multienzyme complex in Congregibacter litoralis and related members of the OM60/NOR5 clade is adapted to the oxidation of submillimolar amounts of thiosulfate and nonfunctional at higher concentrations of reduced inorganic sulfur compounds. Pelagic bacteria thriving in the oxic zones of marine environments may rarely encounter amounts of thiosulfate, which would allow its utilization as electron donor for lithoautotrophic or mixotrophic growth. Consequently, in evolution the Sox multienzyme complex in some of these bacteria may have been optimized for the effective utilization of trace amounts of thiosulfate generated from the degradation of organic sulfur compounds.

Prevalence and Genetic Characterization of Carbapenem- and Polymyxin-Resistant Acinetobacter baumannii Isolated from a Tertiary Hospital in Terengganu, Malaysia

Nosocomial infection caused by Acinetobacter baumannii is of great concern due to its increasing resistance to most antimicrobials. In this study, 54 nonrepeat isolates of A. baumannii from the main tertiary hospital in Terengganu, Malaysia, were analyzed for their antibiograms and genotypes. Out of the 54 isolates, 39 (72.2%) were multidrug resistant (MDR) and resistant to carbapenems whereas 14 (25.9%) were categorized as extensive drug resistant (XDR) with additional resistance to polymyxin B, the drug of “last resort.” Pulsed-field gel electrophoresis analyses showed that the polymyxin-resistant isolates were genetically diverse while the carbapenem-resistant isolates were clonally related. The 14 XDR isolates were further investigated for mutations in genes known to mediate polymyxin resistance, namely, pmrCAB, and the lipopolysaccharide biosynthesis genes, lpxA, lpxC, lpxD, and lpsB. All 14 isolates had a P102H mutation in pmrA with no mutation detected in pmrC and pmrB. No mutation was detected in lpxA but each polymyxin-resistant isolate had 2–4 amino acid substitutions in lpxD and 1-2 substitutions in lpxC. Eight resistant isolates also displayed a unique H181Y mutation in lpsB. The extent of polymyxin resistance is of concern and the novel mutations discovered here warrant further investigations.

Investigation of Bacterial Cellulose Biosynthesis Mechanism in Gluconoacetobacter hansenii

The present study explores the mechanism of cellulose biosynthesis in Gluconoacetobacter hansenii. The cellulose synthase enzyme was purified as membrane fraction and solubilized by treatment with 0.1% digitonin. The enzyme was separated by native-gel electrophoresis and β-D-glucan analysis was carried out using in vitro gel assay. The cellulose synthase has glycoprotein nature and composed two polypeptide subunits of 93 KDa and 85 KDa. The confirmation of β-1,4-glucan (cellulose) was performed in whole and hydrolyzed monomeric sugar form. Tinopal and Congo red were used for cellulose detection on the gel. Thus the in vitro cellulose synthesis assay with cell free enzyme fraction was attempted to improve the understanding of cellulose biosynthesis.

Emergence of blaTEM Type Extended-Spectrum β-Lactamase Producing Salmonella spp. in the Urban Area of Bangladesh

Salmonellosis, an acute invasive enteric infection, is endemic in Bangladesh. We analyzed 128,312 stool samples of diarrheal patients to identify Salmonella spp. during 2005–2013. A total of 2120 (1.7%) Salmonella spp. were isolated and the prevalence of Salmonella spp. decreased significantly over time (2→1%, P<0.001). Among the typhoidal Salmonella (TS) serogroups, S. Typhi was predominant (404, [65.1%]) followed by S. Paratyphi B (139, [22.4%]) and S. Paratyphi A (78, [12.6%]). Of the nontyphoidal Salmonella (NTS) isolates, the serogroup C1 (560, [37%]) was predominant followed by B (379, [25%]), C2 (203, [14%]), E (127, [9%]), and D (94, [6%]). Most of the resistance was found towards nalidixic acid (40%), ampicillin (36%), cotrimoxazole (20%), chloramphenicol (13%), ciprofloxacin (4%), and ceftriaxone (4%). Interestingly, 32% of the isolates showed reduced susceptibility to Cip. Multiantibiotic resistance (MAR, ≥3 drugs) was more common among TS than NTS strains (P<0.001). Among the representative ceftriaxone-resistant isolates, blaTEM gene was detected among 88% (7/8) of the strains, whereas only one strain of S. Typhi was positive for both blaTEM and blaCTX-M genes. The study reflects higher prevalence of MAR Salmonella spp. and is the first to report the blaTEM gene mediated ESBL production among Salmonellae in Bangladesh. Emergence of MAR Salmonella spp. in particular ESBL strains should be considered a public health concern.

Comparative Assessment of Factors Involved in Acetoin Synthesis by Bacillus subtilis 168

Acetoin is widely used as flavor agent and serves as a precursor for chemical synthesis. Here we focused on identifying the best physiological conditions (initial substrate concentrations, pH, temperature, and agitation) for enhanced acetoin accumulation by Bacillus subtilis 168. The optimal physiological conditions support maximum acetoin accumulation by minimizing byproduct (acetate and butanediol) synthesis and a maximum of 75% enhancement in acetoin yield could be achieved. Additionally, the effect of change in ALS (acetolactate synthase) and ALDC (acetolactate decarboxylase) activities was evaluated on acetoin accumulation. Increasing ALS and ALDC enzyme activities led to efficient utilization of pyruvate towards acetoin accumulation and about 80% enhancement in acetoin accumulation was observed.

Prevalence, Clinical Features, and Outcome of Pseudomonas Bacteremia in Under-Five Diarrheal Children in Bangladesh

We sought to evaluate the prevalence, associated factors, and outcome of under-five diarrheal children with either sex having Pseudomonas bacteremia. A retrospective chart review of under-five diarrheal children admitted to the Dhaka Hospital of the International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b), from January 2011 to December 2011 was performed using an online hospital management system. Children with Pseudomonas bacteremia constituted the cases (n = 31), and the controls (n = 124), without Pseudomonas bacteremia, were randomly selected. The prevalence of Pseudomonas bacteremia was 1% (31/5,179). The Pseudomonas was multidrug resistant but was 84% sensitive to ceftazidime and 100% to imipenem. The case-fatality rate was significantly higher among the cases than the controls (26% versus 5%; P = 0.003). In logistic regression analysis, after adjusting for potential confounders such as severe wasting, severe underweight, severe pneumonia, and young age (11.71 (4.0, 18.0) months), the cases more often presented with absent peripheral pulses in absence of dehydration (95% CI = 2.31–24.45) on admission. This finding underscores the importance of early identification of this simple clinical sign to ensure prompt management including fluid resuscitation and broad spectrum antibiotics to help reduce morbidity and mortality in such children, especially in resource-poor settings.

Antimicrobial Resistance of Salmonella enterica Isolates from Tonsil and Jejunum with Lymph Node Tissues of Slaughtered Swine in Metro Manila, Philippines

Due to frequent antibiotic exposure, swine is now recognized as potential risk in disseminating drug-resistant Salmonella enterica strains. This study thus subjected 20 randomly selected S. enterica isolates from tonsil and jejunum with lymph node (JLN) tissues of swine slaughtered in Metro Manila, Philippines, to VITEK 2 antimicrobial susceptibility testing (AST). The test revealed all 20 isolates had resistance to at least one antimicrobial agent, in which highest occurrence of resistance was to amikacin (100%), cefazolin (100%), cefuroxime (100%), cefuroxime axetil (100%), cefoxitin (100%), and gentamicin (100%), followed by ampicillin (50%), and then by sulfamethoxazole trimethoprim (30%). Three multidrug-resistant (MDR) isolates were detected. The sole S. enterica serotype Enteritidis isolate showed resistance to 12 different antibiotics including ceftazidime, ceftriaxone, amikacin, gentamicin, and tigecycline. This study is the first to report worldwide on the novel resistance to tigecycline of MDR S. enterica serotype Enteritidis isolated from swine tonsil tissues. This finding poses huge therapeutic challenge since MDR S. enterica infections are associated with increased rate of hospitalization or death. Thus, continual regulation of antimicrobial use in food animals and prediction of resistant serotypes are crucial to limit the spread of MDR S. enterica isolates among hogs and humans.

Influence of Stress Connected with Moving to a New Farm on Potentially MAP-Infected Mouflons

There is no European legislation concerning paratuberculosis that requires that imported animals be kept in quarantine and commonly they are directly released into areas with other animals. In this study, detection of latent infection of paratuberculosis in healthy mouflons previously diagnosed as paratuberculosis-free, but originating from a real time quantitative PCR- (qPCR-) positive herd, occurred after their transport to a new farm. During a twelve-day quarantine period, all mouflons irregularly shed Mycobacterium avium subsp. paratuberculosis (MAP) in faeces, and in a small number of cases also in milk. After the animals were released from quarantine, MAP was detected for a further two days, after which, testing was negative, except in one case. Therefore, the stress connected with transport, novel environment, dietary change, or limited area with high density of animals might have contributed to the induction of paratuberculosis and the shedding of MAP from the animals, previously diagnosed as MAP-negative. According to these results, the keeping of imported animals in quarantine and their examination for MAP presence not only before the transport but also afterwards should be recommended. The designation of a particular area of a farm as a quarantine enclosure could help to mitigate the impact of stress caused by a confined space with a high density of animals.