4 research outputs found
In vitro exoproteome profile of Stenotrophomonas maltophilia in iron depleted condition
No full textStenotrophomonas maltophilia has been recently identified as the third most common nosocomial infection in the hospital especially among immunocompromised patients. Management of S. maltophilia infection is an inordinate challenge in the hospital due to its intrinsic and acquired resistance to most of the antibiotics. Although S. maltophilia is frequently associated with increased morbidities and mortalities, the pathogenesis mechanisms of S. maltophilia are still not very clear. This is due to S. maltophilia which entered hospital setting from environmental sources heavily colonizes the respiratory tract and other anatomical sites, without showing a clear cut infection in human. Hence, the debate about whether S. maltophilia is a true pathogen or a colonizer is still an ongoing investigation.
Iron is an essential factor for their survival. In the host environment, the level of iron is ~1011 times below than the level required for microorganism and is mostly unavailable as it is bound together with host protein. Microbes develop multiple mechanisms such as siderophores secretion to forage for free iron in an iron limited environment. Under iron starvation, microbes also secrete other virulence potential exoproteins. The present study was aimed at investigating the effect of iron depletion in secretion of proteomes in S. maltophilia.
Briefly, four strains of the S. maltophilia LMG959 (environment), ATCC 13637, CS17 and CS24 (clinical) were grown in normal and iron depleted medium. The siderophores production was screened qualitatively and quantitatively using CAS plate and liquid assay. Followed by, nematocidal assay was performed to test the ability of S. maltophilia supernatant grown in iron depleted medium to kill the nematode Caenorhabditis elegans. Lastly, the putative proteins expressed during the stressed condition were identified by Isobariq Tags for Relative and Absolute Quantification (ITRAQ) mass spectrometry.
Initial screening of siderophores production exhibited largest yellow zone for CS17 and CS24 (10 mm) followed by ATCC13637 (8 mm), LMG959 (6 mm). The siderophores production was further confirmed quantitatively and the highest was detected in both clinical isolates (30.8% [p0.05). Isolates grown under iron-depleted condition (ATCC 13637: 63%; CS17: 96%; CS24: 97%) showed more nematocidal activity than in normal condition (ATCC 13637: 43%; CS17: 76%; CS24: 79%) (p>0.05). None of the worms were killed when infected with LMG959. Based on the above result, only CS17 and LMG959 are subjected to ITRAQ analysis. ITRAQ analysis revealed a total of 122 proteins showed altered expression in response to iron starvation with 96 being up-regulated and 26 were down-regulated for both isolates. ITRAQ analysis identified higher expression of several iron acquisition and pathogenic potential proteins in both isolates grown in iron depleted condition. In another study, clinical and environment isolates grown under normal condition were compared to identify the proteomic profiles. ITRAQ analysis discovered 81 proteins that exhibited differently expressed with 40 proteins up-regulated and 41 proteins down-regulated. In normal condition, several proteins such as Elongation factor G, Endoribonuclease and Fimbrial protein expressed in higher fold in clinical isolate compared to environmental isolate.
In conclusion, S. maltophilia produced siderophores under iron depleted condition. Based on nematocidal assay, eventhough there is no significant difference in killing rates between iron depleted and normal condition but S. maltophilia did show an increased of ~20% of its killing rates in iron depleted medium except for LMG959. ITRAQ analysis revealed S. maltophilia altered the expression of metabolic, iron acquisition and pathogenic potential proteins under iron starvation. A comparison of clinical and environmental isolates grown in normal medium revealed that clinical isolates expresses more pathogenic potential proteins compared to environmental isolate. The data obtained in the present study, clearly indicates that under iron depleted condition, S. maltophilia are capable of altering the expression of its proteomes to ensure their survival in the host
Putative Iron Acquisition Systems in Stenotrophomonas maltophilia
No full textIron has been shown to regulate biofilm formation, oxidative stress response and several pathogenic mechanisms in Stenotrophomonas maltophilia. Thus, the present study is aimed at identifying various iron acquisition systems and iron sources utilized during iron starvation in S. maltophilia. The annotations of the complete genome of strains K279a, R551-3, D457 and JV3 through Rapid Annotations using Subsystems Technology (RAST) revealed two putative subsystems to be involved in iron acquisition: the iron siderophore sensor and receptor system and the heme, hemin uptake and utilization systems/hemin transport system. Screening for these acquisition systems in S. maltophilia showed the presence of all tested functional genes in clinical isolates, but only a few in environmental isolates. NanoString nCounter Elements technology, applied to determine the expression pattern of the genes under iron-depleted condition, showed significant expression for FeSR (6.15-fold), HmuT (12.21-fold), Hup (5.46-fold), ETFb (2.28-fold), TonB (2.03-fold) and Fur (3.30-fold). The isolates, when further screened for the production and chemical nature of siderophores using CAS agar diffusion (CASAD) and Arnows’s colorimetric assay, revealed S. maltophilia to produce catechol-type siderophore. Siderophore production was also tested through liquid CAS assay and was found to be greater in the clinical isolate (30.8%) compared to environmental isolates (4%). Both clinical and environmental isolates utilized hemoglobin, hemin, transferrin and lactoferrin as iron sources. All data put together indicates that S. maltophilia utilizes siderophore-mediated and heme-mediated systems for iron acquisition during iron starvation. These data need to be further confirmed through several knockout studies
Physiological and proteomic analysis of Stenotrophomonas maltophilia grown under the iron-limited condition
No full textAims: to study physiological and proteomic analysis of Stenotrophomonas maltophilia grown under iron-limited condition. Methods: one clinical and environmental S. maltophilia isolates grown under iron-depleted conditions were studied for siderophore production, ability to kill nematodes and alteration in protein expression using isobaric tags for relative and absolute quantification (ITRAQ). Results & conclusions: siderophore production was observed in both clinical and environmental strains under iron-depleted conditions. Caenorhabditis elegans assay showed higher killing rate under iron-depleted (96%) compared with normal condition (76%). The proteins identified revealed, 96 proteins upregulated and 26 proteins downregulated for the two isolates under iron depletion. The upregulated proteins included several iron acquisition proteins, metabolic proteins and putative virulence proteins
Diagnostic accuracy of fresh drooled saliva for SARS-CoV-2 in travelers
No full textBackground The standard for SARS-CoV-2 diagnosis is RT-PCR from nasopharyngeal or oropharyngeal swabs. Major airports require COVID-19 screening, and saliva has the potential as a substitute specimen for SARS-CoV-2 diagnosis. We investigated the utility of fresh drooled saliva against NPS for COVID-19 screening of travelers.
Methods We recruited 81 travelers and 15 non-travelers (including ten controls) prospectively within a mean of 3⋅22 days of RT-PCR confirmed COVID-19. Each study participant provided 2 mls of early morning fresh drooled whole saliva separately into a sterile plastic container and GeneFiX™ saliva collection kit. The saliva specimens were processed within 4 h and tested for SARS-CoV-2 genes (E, RdRP, and N2) and the results compared to paired NPS RT-PCR for diagnostic accuracy.
Results Majority of travellers were asymptomatic (75⋅0%) with a mean age of 34⋅26 years. 77 travelers were RT-PCR positive at the time of hospitalization whilst three travelers had positive contacts. In this group, the detection rate for SARS-CoV-2 with NPS, whole saliva, and GeneFiX™ were comparable (89⋅3%, 50/56; 87⋅8%, 43/49; 89⋅6%, 43/48). Both saliva collection methods were in good agreement (Kappa = 0⋅69). There was no statistical difference between the detection rates of saliva and NPS (p > 0⋅05). Detection was highest for the N2 gene whilst the E gene provided the highest viral load (mean = 27⋅96 to 30⋅10, SD = 3⋅14 to 3⋅85). Saliva specimens have high sensitivity (80⋅4%) and specificity (90⋅0%) with a high positive predictive value of 91⋅8% for SARS-CoV-2 diagnosis.
Conclusion Saliva for SARS-CoV-2 screening is a simple accurate technique comparable with NPS RT-PCR
