A Novel Non-Psychoactive Fatty Acid from a Marine Snail, Conus inscriptus, Signals Cannabinoid Receptor 1 (CB1) to Accumulate Apoptotic C16:0 and C18:0 Ceramides in Teratocarcinoma Cell Line PA1.

The cannabinoid-type I (CB1) receptor functions as a double-edged sword to decide cell fate: apoptosis/survival. Elevated CB1 receptor expression is shown to cause acute ceramide accumulation to meet the energy requirements of fast-growing cancers. However, the flip side of continual CB1 activation is the initiation of a second ceramide peak that leads to cell death. In this study, we used ovarian cancer cells, PA1, which expressed CB1, which increased threefold when treated with a natural compound, bis(palmitoleic acid) ester of a glycerol (C2). This novel compound is isolated from a marine snail, Conus inscriptus, using hexane and the structural details are available in the public domain PubChem database (ID: 14275348). The compound induced two acute ceramide pools to cause G0/G1 arrest and killed cells by apoptosis. The compound increased intracellular ceramides (C:16 to 7 times and C:18 to 10 times), both of which are apoptotic inducers in response to CB1 signaling and thus the compound is a potent CB1 agonist. The compound is not genotoxic because it did not induce micronuclei formation in non-cancerous Chinese hamster ovarian (CHO) cells. Since the compound induced the cannabinoid pathway, we tested if there was a psychotropic effect in zebrafish models, however, it was evident that there were no observable neurobehavioral changes in the treatment groups. With the available data, we propose that this marine compound is safe to be used in non-cancerous cells as well as zebrafish. Thus, this anticancer compound is non-toxic and triggers the CB1 pathway without causing psychotropic effects.

Synthesis, Characterization, and In V ivo Toxicological Evaluation of Copper (II) Oxide Containing Herbometallic Siddha Nanocomplex "Thamira Parpam".

"Thamira parpam" (TP), a copper-based herbometallic oxide (copper (II) oxide) nanodrug has been used in Siddha medicine for centuries because of its anti-ulcerogenic property. However, the physicochemical properties and in vivo toxicity of TP still remain elusive. Rigorous clinical translation requires deciphering these vital properties. We have synthesized TP following a gold standard protocol in the traditional Siddha methodology. We assessed the size, phase, elemental constituents, and thermal stability of TP by SEM and TEM, XRD, EPR, and EDAX analyses, respectively. The results depicted the conversion of metallic copper into copper (II) oxide in the final stages of TP preparation and exhibited nanodimensions ranging between 10 and 50 nm. The XPS spectra revealed the presence of oxygen-deficient state and a carbonaceous coating was found on the surface of TP using TEM analysis. In vivo safety was studied in rat toxicity models by adopting OECD guidelines. Body weight changes, feed, and water intake were unaltered upon TP administration. Hematological, biochemical profiling, and histopathological findings also suggested its nontoxic nature with no abnormalities in major organs and its functions. Interestingly, we found that the metal toxicity could have been subdued because of the carbonaceous coating around the nanoparticle copper (II) oxide, confirming that the drug is safe at a low dose. Overall, our study has enlightened the safety of TP supporting the use of Siddha formulations.

Strategy for COVID-19 vaccination in India: the country with the second highest population and number of cases.

Free vaccination against COVID-19 commenced in India on January 16, 2021, and the government is urging all of its citizens to be immunized, in what is expected to be the largest vaccination program in the world. Out of the eight COVID-19 vaccines that are currently under various stages of clinical trials in India, four were developed in the country. India's drug regulator has approved restricted emergency use of Covishield (the name employed in India for the Oxford-AstraZeneca vaccine) and Covaxin, the home-grown vaccine produced by Bharat Biotech. Indian manufacturers have stated that they have the capacity to meet the country's future needs for COVID-19 vaccines. The manpower and cold-chain infrastructure established before the pandemic are sufficient for the initial vaccination of 30 million healthcare workers. The Indian government has taken urgent measures to expand the country's vaccine manufacturing capacity and has also developed an efficient digital system to address and monitor all the aspects of vaccine administration.

Nanomedicine-based cancer immunotherapy: recent trends and future perspectives.

The combination of cancer immunotherapy with efficient functionalized nanosystems has emerged as a beneficial treatment strategy and its use has increased rapidly. The roles of stimuli-responsive nanosystems and nanomedicine-based cancer immunotherapy, a subsidiary discipline in the field of immunology, are pivotal. The present era is witnessing rapid advancements in the use of nanomedicine as a platform for investigating novel therapeutic applications and modern intelligent healthcare management strategies. The development of cancer nanomedicine has posthaste ratified the outcomes of immunotherapy to the subsequent stage in the current era of medical research. This review focuses on key findings with respect to the effectiveness of nanomedicine-based cancer immunotherapies and their applications, which include i) immune checkpoint inhibitors and nanomedicine, ii) CRISPR-Cas nanoparticles (NPs) in cancer immunotherapy, iii) combination cancer immunotherapy with core-shell nanoparticles, iv) biomimetic NPs for cancer immunotherapy, and v) CAR-T cells and cancer nanoimmunotherapy. By evaluating the state-of-the-art tools and taking the challenges involved into consideration, various aspects of the proposed nano-enabled therapeutic approaches have been discussed in this review.

Towards diaspora-driven research capacity strengthening in low- and middle-income countries: results from India and Nepal.

BACKGROUND: Biomedical research from high-income countries often informs practice and policy in low- and middle-income countries (LMICs) with vastly different socioeconomic and health systems. Engagement of LMIC-based researchers is integral to setting research priorities in the local context. METHODS: A program, comprising a research seminar and workshop, and utilizing diaspora health professionals to understand research needs and build research capacity in LMICs, was created and pilot-tested at two institutions in India (65 participants) and Nepal (30 participants). Pre- and post-program surveys were instituted to assess participants' attitudes towards research. RESULTS: In the pre-program survey, most participants (India: 76%, Nepal: 100%) perceived research as 'very/extremely important' in their careers. However, a majority felt that finding time (India: 75%, Nepal: 81%) and funding (India: 82%, Nepal: 100%) for research was 'difficult/very difficult'. After the program, 86-91% and 86-100% of participants from India and Nepal, respectively, felt that the various courses were very useful/useful for their research careers. CONCLUSIONS: Research is seen as an integral part of educational training and career advancement in LMICs. However, inadequate training, funding and mentorship remain a challenge. Engagement of diaspora health workers may serve as an important avenue for collaborative biomedical research capacity strengthening in LMICs.

Hepatitis B virus and hepatitis C virus dual infection among patients with chronic liver disease.

BACKGROUND AND PURPOSE: Hepatitis B virus (HBV) and hepatitis C virus (HCV) dual infection accounts for a substantial proportion of liver diseases worldwide. Although the exact prevalence is not known, these viral infections are common among patients with chronic liver disease (CLD). This study was performed to determine the prevalence of HBV and HCV dual infection among patients with CLD in Chennai, India. METHODS: 251 patients were tested for the presence of hepatitis B surface antigen (HBsAg), immunoglobulin (Ig)-M/IgG antibody to hepatitis B core antigen (anti-HBc) and anti-HCV antibodies, and HBV-DNA and HCV-RNA by qualitative polymerase chain reaction. RESULTS: Coinfection with HCV and HBV was detected in 15 patients (5.9%), 12 of whom (80.0%) were positive for HCV-RNA and IgG anti-HBc with no evidence of HBV-DNA, while 3 HBsAg-negative patients (20.0%) were positive for HBV-DNA in addition to HCV-RNA. Liver function test profiles were significantly altered for HCV-positive patients compared with HBV-positive and HBV/HCV coinfected patients (p = 0.001). Bilirubin and alanine aminotransferase levels were significantly raised in coinfected patients compared with non-HBV, non-HCV patients (p = 0.001). CONCLUSIONS: The results demonstrated that HBV was predominantly associated with underlying CLD among this group of patients in India and suggest that HBV coinfection in HCV-infected patients should not be excluded by negative HBsAg status alone.

Transmission of hepatitis C virus infection from asymptomatic mother to child in southern India.

BACKGROUND: Little information is available on the mother-to-child transmission of hepatitis C virus (HCV) in India, and no interventions to decrease transmission rates have been identified. Hence, we performed a long-term prospective study in infants born to HCV-positive mothers, with the aim of evaluating vertical transmission of HCV and correlated risks factors. METHODS: Three thousand one hundred and fifteen healthy asymptomatic pregnant women were included in the study. We used third-generation (Murex anti-HCV) ELISA and HCV RNA reverse transcription PCR (RT-PCR) for screening, and the commercial line probe assay (Inno-LiPA) and direct sequencing HCV genotyping assays were performed to confirm the transmitted HCV genotypes. RESULTS: Of the total 3115 healthy asymptomatic pregnant women, 18 (0.6%) were positive for anti-HCV. Of the 18 anti-HCV-positive women, eight (44.4%) were positive for HCV RNA RT-PCR. HCV transmission was observed in two of the eight babies born to eight HCV RNA-positive mothers who were followed up for 12 months. HCV genotyping of the mother/child pairs revealed the persistent presence of mixed genotypes 1a and 4 throughout the follow-up period. None of the non-viremic (HCV RNA-negative) mothers transmitted HCV infection to their baby. In our study approximately 25% of vertical/perinatal transmission of HCV was observed among HCV RNA-positive antenatal women. CONCLUSIONS: This study is of importance as it is the first report from India of a successful attempt to analyze the rate of vertical/perinatal transmission of HCV from infected mothers to their children by a prospective longitudinal follow-up study, and to characterize the pattern of genotype(s) of HCV present in the infected mother/baby pairs, so as to confirm the source of HCV acquired by the newborn babies.

Low frequency of precore mutants in anti-hepatitis B e antigen positive subjects with chronic hepatitis B virus infection in Chennai, Southern India.

The natural course of chronic hepatitis B (CH-B) virus infection is reportedly variable, and the long-term outcomes in hepatitis B e antigen (HBeAg)-negative chronic hepatitis B infection are distinct from HBeAg-positive chronic hepatitis. However, the molecular virological factors that contribute to the progression of liver disease in the south Indian setting remain largely unclear. We prospectively studied 679 consecutive patients for HBsAg, HBeAg, anti-HBeAg, and HBV DNA by qualitative PCR. Randomly selected samples were subjected to bidirectional sequencing to reveal core/precore variants. Of the total 679 chronic HBV cases investigated, 23% (154/679) were replicative HBV carriers. Furthermore, amongst the 560 HBV DNA samples analyzed, 26% (146/560) were viremic. Among the 154 HBeAg positive cases, HBV DNA was positive in 118 cases (77%), significantly (p<0.001) higher than the anti-HBe positive (7%) (28/406) cases. Significant increase in liver disease (p<0.01) with ALT enzyme elevation (p<0.001) was observed in both HBe and anti-HBe viremic cases. Interestingly, low frequencies of mutations were seen in the precore region of the HBV strains studied. HBV precore and core promoter variants were less often detected in subjects with "e" negative chronic HBV infection and, therefore, may not have a prognostic role in determining liver disease sequelae in this part of tropical India.

Alpha-fetoprotein as a tumor marker in hepatocellular carcinoma: investigations in south Indian subjects with hepatotropic virus and aflatoxin etiologies.

OBJECTIVES: The prevalence of hepatitis B virus (HBV) is reportedly the main cause of hepatocellular carcinoma (HCC) in India, where hepatitis C virus (HCV)-associated HCC is believed to be relatively less prevalent. We verified the usefulness of alpha-fetoprotein (AFP) as a tumor marker and analyzed the influence of viral etiology on AFP levels in HCC. METHODS: Of a total of 1012 cases with liver disease, 202 were investigated for the presence of AFP (142 HCC cases, 30 cirrhosis cases, and 30 chronic liver disease (CLD) cases). In addition, serum samples from 30 healthy patients, 30 hepatitis B surface antigen (HBsAg) carriers, and 30 acute viral hepatitis cases were included as controls. AFP was quantitatively determined using a commercial ELISA (Quorum Diagnostics, Canada). Out of the 142 HCC cases screened for AFP, aflatoxin B1 (AFB1) detection was carried out in 38 HCC cases using an in-house immunoperoxidase test. RESULTS: In HBV and HCV co-infected HCC cases, the AFP positivity was 85.7%. In HBV alone-associated HCC, the positivity was 62.9%, and 54.5% of AFB1 positive HCC cases showed AFP positivity. In HBV and HCV negative HCC cases, the positivity was 20.5%, and in HCV-associated HCC it was 17.6%. The HBV/HCV co-infected group and HBV alone positive HCC cases had significantly elevated levels of AFP. When AFP positivity was analyzed based on the marker profile of HBV, 89.7% of AFP positive cases were HBV-DNA positive. CONCLUSIONS: The overall positivity pattern of AFP in HCC does indicate that higher levels of AFP are observed with hepatitis virus positivity, especially with HBV. Further studies must be carried out to correlate the serum levels of AFP with the size, number, and degree of differentiation of HCC nodules.

Seroprevalence of hepatitis delta virus infection among subjects with underlying hepatic diseases in Chennai, southern India.

Four hundred million people are carriers of hepatitis B virus (HBV) worldwide and approximately 5% of these are reportedly positive for hepatitis delta virus (HDV). Several reports indicate a declining trend in the occurrence of HDV infection in the north of tropical India. To our knowledge, no study has been conducted to evaluate whether a similar epidemiological change is occurring in southern India. Therefore we evaluated the seroprevalence of HDV among 153 individuals with HBV-related liver diseases in Chennai, and assessed any change in epidemiological pattern by comparing the results with seroprevalence figures reported previously. Of the 153 patients screened, nine (5.9%) were reactive to anti-delta antibodies, six (3.9%) presented an evidence of past infection (IgG anti-delta positive) and three (2.0%) showed anti-HDV IgM, suggestive of recent HDV infection. Alanine transaminase elevation was not significant in HDV-associated infection compared with HBV alone-infected acute viral hepatitis (AVH) (P=0.82) and chronic liver disease (P=0.77) patients. The anti-HDV positivity in AVH was considerably low (6.6%), compared with previous Indian reports varying from 10.7% to >30%. HDV infection was relatively low and seems to play a minor determining factor of liver diseases in the tropical south Indian population.

Relationship between T-lymphocyte cytokine levels and sero-response to hepatitis B vaccines.

AIM: To investigate the cellular defects by analyzing the (Th1/Th2) cytokine levels in vaccine responders and non-responders. METHODS: Peripheral blood mononuclear cell (PBMC) from responders and non-responders were stimulated with or with out recombinant HBsAg or PHA. Broad spectrum of cytokines viz (Th1) IFN-gamma, IL-2, TNF-alpha, IL-12 and (Th2) IL-10, IL-4 were measured after in vitro stimulation with recombinant HBsAg and were compared with respective antibody titers. RESULTS: A significant decrease (P = 0.001) in Th1 and Th2 cytokines namely, IL-2, INF-gamma, TNF-alpha and IL-10 in non-responders was observed. The level of IL-4 was not significant between the three groups. Furthermore, despite a strong Th1 and Th2 cytokine response, the level of IL-12 was elevated in high-responders compared to other groups (P = 0.001) and demonstrated a positive correlation with anti-HBs titers and Th1 cytokine response. CONCLUSION: Our findings suggest that unresponsiveness to recombinant hepatitis B vaccines (rHB) is multifactorial, including specific failure of antigen presentation or the lack of both T helper Th1 and Th2 response.

The prevalence of hepatitis B virus and hepatitis C virus infection among patients with chronic liver disease in South India.

OBJECTIVE: Determining the identity of hepatitis C virus (HCV) genotypes in liver disease has key implications for ascertaining the duration of antiviral therapy and disease prognosis. We investigated the presence of various genotypes of HCV among 69 chronic liver diseased (CLD) patients with chronic HCV infection. METHODS: Sixty-nine consecutive subjects with underlying chronic hepatitis (n=28), cirrhosis (n=35), and hepatocellular carcinoma (n=6), diagnosed by clinical, biochemical, and histological means, were studied. Hepatitis B virus (HBV) and HCV diagnostic markers were used. HCV-RNA was extracted from sera of HCV-infected subjects and subsequently the HCV genotypes were determined using a commercial line probe assay (Inno-LiPA HCV II). RESULTS: Of the 69 CLD cases screened for possible markers of HBV and HCV infection, 39 (57%) were positive for HBV and 30 (43%) were HCV infected. The overall HCV-RNA positivity was 77% (23/30). Of these, the majority were genotype 1b (13/23, 57%), followed by 1a (6/23, 26%), mixed genotypes 3 and 4(3/23, 13%), and mixed pattern of 1a, 1b, and 4 (1/23, 4.3%). The genotype 1b infected subjects demonstrated significantly elevated transaminase (ALT) levels (p<0.05) as compared with the other non-1b HCV genotypes. CONCLUSIONS: The predominance of HCV genotype 1b among CLD patients could pose a major challenge for the efficient management of HCV disease and the development of effective therapeutic interventions in peninsular India.

Transmission of "a" determinant variants of hepatitis B virus in immunized babies born to HBsAg carrier mothers.

Hepatitis B virus (HBV) surface antigen mutations may lead to immune escape and eventually cause failure of immunization. In this report, we identified immune escape variants in immunized babies born to hepatitis B surface antigen (HBsAg) carrier mothers. A total of 68 babies were followed up for 2 years after the full course of vaccination; 2.9% (2/68) of babies were found to be infected with the variant HBV in spite of preexisting antibody to surface antigen (anti-HBs) at 24 months post immunization. Both infants were positive for HBV-DNA; sequencing results of the "a" determinant region of the surface gene revealed that both babies had point mutations at a different nucleotide position resulting in various amino acid substitutions. In addition, an intriguing variant having an addition-deletion mutation was observed in one of the babies. This is the first report to show the addition-deletion variant of HBV in India. However, the immunological significance of the above HBV variants needs to be further elucidated.

Comparative efficacy of two dosages of recombinant hepatitis B vaccine in healthy adolescents in India.

BACKGROUND: Inclusion of hepatitis B vaccine in the Universal Programme of Immunization of all Asian and African countries is hampered by the economic burden on the health budget because of the cost of hepatitis B vaccines. Here we evaluated the immunogenicity, safety, efficacy, and the persistence of antibody to hepatitis B surface antigen (anti-HBs) titers of a new and a low cost recombinant hepatitis B vaccine GeneVac B, with 2 different dosages in healthy adolescents in India. METHODS: GeneVac-B, a recombinant hepatitis B vaccine (Serum Institute of India, Pune, India), was administered in 10 or 20 microg dose intramuscularly to 2 groups of 100 healthy school-going adolescents at 0-, 1-, and 6-month intervals, who were followed up for 1 year. Group I received 20 mug doses whereas Group II received 10 mug doses. Blood samples were collected 1 month after each dose and 1 year after the third dose. The anti-HBs titers were assayed using commercially available kits to assess the immunogenicity of the 2 dosage schedules. Safety studies were also carried out. RESULTS: The geometric mean titer value of the anti-HBs titer 1 month after the third dose was 2629 (mlU/mL) in Group I and 1373 mlU/mL for Group II subjects. One year after the third dose, the persistence of anti-HBs in those who had received 20 mug was 2262 mlU/mL whereas it was 1039 mlU/mL in the group receiving 10 microg doses. All the subjects in both the groups were seroprotected at 1 year after vaccination. None of the vaccinees exhibited serious adverse reactions throughout the study period. CONCLUSIONS: The study demonstrated the immunogenicity of the recombinant hepatitis B vaccine, and confirms that the 0.5 mL (10 microg) dose of GeneVac B can be administered with satisfactory safety and immunogenicity to adolescents up to 19 years of age, reducing the cost to less than U.S. $1.00 per dose making it acceptable for the Universal Programme of Immunization of developing and under developed countries.

Coinfection of hepatitis B and hepatitis C virus in HIV-infected patients in south India.

AIM: To screen for the co-infection of hepatitis B (HBV) and hepatitis C virus (HCV) in human immunodeficiency virus (HIV) infected patients in southern India. METHODS: Five hundred consecutive HIV infected patients were screened for Hepatitis B Virus (HBsAg and HBV-DNA) and Hepatitis C virus (anti-HCV and HCV-RNA) using commercially available ELISA kits; HBsAg, HBeAg/anti-HBe (Biorad laboratories, USA) and anti-HCV (Murex Diagnostics, UK). The HBV-DNA PCR was performed to detect the surface antigen region (pre S-S). HCV-RNA was detected by RT-PCR for the detection of the constant 5' putative non-coding region of HCV. RESULTS: HBV co-infection was detected in 45/500 (9%) patients and HCV co-infection in 11/500 (2.2%) subjects. Among the 45 co-infected patients only 40 patients could be studied, where the detection rates of HBe was 55% (22/40), antiHBe was 45% (18/40) and HBV-DNA was 56% (23/40). Among 11 HCV co-infected subjects, 6 (54.5%) were anti-HCV and HCV RNA positive, while 3 (27.2%) were positive for anti-HCV alone and 2 (18%) were positive for HCV RNA alone. CONCLUSION: Since the principal routes for HIV transmission are similar to that followed by the hepatotropic viruses, as a consequence, infections with HBV and HCV are expected in HIV infected patients. Therefore, it would be advisable to screen for these viruses in all the HIV infected individuals and their sexual partners at the earliest.

Comparison of three different recombinant hepatitis B vaccines: GeneVac-B, Engerix B and Shanvac B in high risk infants born to HBsAg positive mothers in India.

AIM: To evaluate a low cost Indian recombinant hepatitis B vaccine GeneVac-B for its immunogenicity and safety in comparison to Engerix B and Shanvac B vaccine in high risk newborn infants born to hepatitis B surface antigen (HBsAg) positive mothers. METHODS: A total of 158 infants were enrolled in the study. Fifty eight infants were enrolled in the GeneVac-B group while 50 each were included for Engerix B and Shanvac B groups. A three-dose regimen of vaccination; at birth (within 24 h of birth), 1st mo and 6 mo. were adopted with 10 mug dosage administered uniformly in all the three groups. Clinical and immunological parameters were assessed for safety and immunogenicity of the vaccines, in all the enrolled infants. RESULTS: Successful follow up until seven months of age was achieved in 83% (48/58) for GeneVac-B, 76% (38/50) and 64% (32/50) for Engerix B and Shanvac B groups respectively. 100% seroconversion and seroprotection was achieved in all the three groups of infants. The geometric mean titers of anti-HBs one month after the completion of three dose of vaccination were 90.5, 80.9 and 72.5 mIU/mL in GeneVac-B, Engerix B and Shanvac B vaccine group respectively. Furthermore the level of anti-HBs increases with age of babies who were born to HBsAg positive mothers. The GMT values of anti-HBs were 226.7, 193.9 and 173.6 mIU/mL respectively in GeneVac-B, Engerix B and Shanvac B groups one year after the completion of the three doses of vaccine. No systemic reactions were reported in infants during the entire vaccination process of GeneVac-B and the other two vaccines. Clinical safety parameters remained within the normal limits throughout the study period. CONCLUSION: The study concludes that there is no significant difference between the three recombinant hepatitis B vaccines. Administration of these vaccines within 24 h of birth to babies, born to HBsAg positive mothers will reduce the incidence of HBV infection.

Phenotypes of isolates of Pseudomonas aeruginosa in a diabetes care center.

BACKGROUND: Pseudomonas aeruginosa is an invasive organism that frequently causes severe tissue damage in diabetic foot ulcers. A major problem in P. aeruginosa infection may be that this pathogen exhibits a high degree of resistance to a broad spectrum of antibiotics. Some researchers feel that P. aeruginosa is a homogeneous species, whereas others have suggested that they are panmictic. Here we characterized P. aeruginosa populations isolated from diabetic foot ulcer and from hospital environment specimens, both from a tertiary diabetes care center in Chennai, India. METHODS: Phenotypic methods like antibiotic susceptibility determinations using Kirby-Bauer's disc diffusion test and minimum inhibitory concentration (MIC) as well as outer membrane protein SDS-PAGE analysis of P. aeruginosa were performed. RESULTS: Twenty three isolates (29.8%) of P. aeruginosa from 77 diabetic foot ulcers and two environmental isolates (13.3%) from 15 different hospital fomites were detected. Both environmental isolates were sensitive to antibiotics than those isolated from clinical specimens by Kirby-Bauer's disk-diffusion method, which correlated the resistance levels by MIC determination. Outer membrane proteins (OMP) corresponding to 21, 23, 43, 46, 50, and 70 kDa were detected. CONCLUSIONS: The study is captivative as the resistance in P. aeruginosa from diabetic foot ulcers seems very common and because all the isolates were resistant to at least one or more antibiotics tested. Disk-diffusion and MIC results shows that piperacillin, amikacin and imipenem retain high levels of antipseudomonal activities and amikacin two times more active than the aforementioned antibiotics to enable itself as a potent antipseudomonal agent in diabetic foot infections. The OMP profile has revealed that clinical isolates were different from hospital environment isolates, which suggests that the origin of infections by P. aeruginosa is mainly due to growth of bacterial strains acquired by patients prior to hospital admission.