In vivo acquisition and risk of inter-species spread of blaKPC-3-plasmid from Klebsiella pneumoniae to Serratia marcescens in the lower respiratory tract

In recent years, Serratia marcescens has emerged as an important agent of hospital-acquired infections, such as pneumonia, urinary tract infection, septicaemia and meningitis, particularly in vulnerable patients. Compared to Klebsiella pneumoniae and Escherichia coli, S. marcescens is less commonly associated with blaKPC genes, yet few cases of plasmid transmission at the gastrointestinal level from K. pneumoniae carbapenemase (KPC)-producing Enterobacterales to S. marcescens have been described. Here we report a case of in vivo acquisition, during a 3-month period of hospitalization in the intensive care unit, of a blaKPC-3 gene carried by a pKpQIL-IT plasmid, and its probable transmission at the bronchial level among different species of Enterobacterales, including K. pneumoniae and S. marcescens. By using whole genome sequence analyses we were able provide insight into the dynamics of carbapenem-resistance determinants acquisition in the lower respiratory tract, a novel anatomical region for such plasmid transmission events, that usually involve the gastrointestinal tract. The co-presence at the same time of both wild-type and resistant Enterobacterales could have been the critical factor leading to the spread of plasmids harbouring carbapenem-resistance genes, of particular importance during surveillance screenings. The possibility of such an event may have significant consequences in terms of antimicrobial treatment, with a potential limitation of therapeutic options, thereby further complicating the clinical management of high-risk critically ill patients

Novel human immunodeficiency virus type 1 protease mutations potentially involved in resistance to protease inhibitors

Plasma-derived sequences of human immunodeficiency virus type 1 (HIV-1) protease from 1,162 patients (457 drug-naive patients and 705 patients receiving protease inhibitor [PI]-containing antiretroviral regimens) led to the identification and characterization of 17 novel protease mutations potentially associated with resistance to PIs. Fourteen mutations were positively associated with PIs and significantly correlated in pairs and/or clusters with known PI resistance mutations, suggesting their contribution to PI resistance. In particular, E34Q, K43T, and K55R, which were associated with lopinavir treatment, correlated with mutations associated with lopinavir resistance (E34Q with either L33F or F53L, or K43T with I54A) or clustered with multi-PI resistance mutations (K43T with V82A and I54V or V82A, V32I, and I47V, or K55R with V82A, I54V, and M46I). On the other hand, C95F, which was associated with treatment with saquinavir and indinavir, was highly expressed in clusters with either L90M and I93L or V82A and G48V. K45R and K20T, which were associated with nelfinavir treatment, were specifically associated with D30N and N88D and with L90M, respectively. Structural analysis showed that several correlated positions were within 8 A of each other, confirming the role of the local environment for interactions among mutations. We also identified three protease mutations (T12A, L63Q, and H69N) whose frequencies significantly decreased in PI-treated patients compared with that in drug-naive patients. They never showed positive correlations with PI resistance mutations; if anything, H69N showed a negative correlation with the compensatory mutations M36I and L10I. These mutations may prevent the appearance of PI resistance mutations, thus increasing the genetic barrier to PI resistance. Overall, our study contributes to a better definition of protease mutational patterns that regulate PI resistance and strongly suggests that other (novel) mutations beyond those currently known to confer resistance should be taken into account to better predict resistance to antiretroviral drugs

Consequences of inaccurate hepatitis C virus genotyping on the costs of prescription of direct antiviral agents in an Italian district

Available commercial assays may yield inaccurate hepatitis C virus (HCV) genotype assignment in up to 10% of cases. We investigated the cost-effectiveness of re-evaluating HCV genotype by population sequencing, prior to choosing a direct acting antiviral (DAA) regimen. Between March and September 2015, HCV sequence analysis was performed in order to confirm commercial LiPA-HCV genotype (Versant\uae HCV Genotype 2.0) in patients eligible for treatment with DAAs. Out of 134 consecutive patients enrolled, sequencing yielded 21 (15.7%) cases of discordant results. For three cases of wrong genotype assignment, the putative reduction in efficacy was gauged between 15% and 40%. Among the eight cases for whom G1b was assigned by commercial assays instead of G1a, potentially suboptimal treatments would have been prescribed. Finally, for five patients with G1 and indeterminate subtype, the choice of regimens would have targeted the worst option, with a remarkable increase in costs, as in the case of the four mixed HCV infections for whom pan-genotypic regimens would have been mandatory. Precise assignment of HCV genotype and subtype by sequencing may, therefore, be more beneficial than expected, until more potent pan-genotypic regimens are available for all patients

Occupational HIV infection in a research laboratory with unknown mode of transmission : a case report

A lab-worker was infected with HIV-1 in a biosafety level-2 of containment, without any apparent breach. Through full-genome sequencing and phylogenetic analyses, we could identify the source of infection in a replication-competent clone, unknowingly contaminating a safe experiment. Mode of transmission remains unclear. Caution is warranted when handling HIV-derived constructs

Optimal cure rate by personalized HCV regimens in real-life : a proof-of-concept study

Background: Pretreatment evaluation of HCV-infected patients is a complex interplay between multiple clinical and viral parameters, leading to a tailored approach that may bring real-life sustained virological response (SVR) rates close to 98%-99%. Objectives: As proof-of-concept, we evaluated the efficacy of all-oral direct-acting antiviral (DAA) regimens in patients whose personalization included pre-therapy evaluation of natural resistance-associated substitutions (RASs), in addition to international guideline recommendations. Methods: One hundred and thirty-one patients who started a first-line all-oral DAA regimen between April 2015 and December 2016 were tested for baseline NS3 and NS5A RASs by Sanger sequencing. SVR12 was defined as HCV-RNA undetectability 12\u2009weeks after treatment discontinuation. Results: Compatibly with a real-life context, 74.0% (97 of 131) of patients presented 652 pretreatment risk factors for failure to achieve SVR12 (infection by GT-1a/GT-3a; cirrhosis; previous treatment experience; HCV-RNA 65800\u202f000\u2009IU/mL) and 33.6% had 653 risk factors. Natural RASs were frequently detected (32.1% prevalence), with substantial prevalence of NS5A RASs (15.3%), mostly represented by Y93H in GT-1b (3 of 36, 8.3%) and GT-3a (3 of 25, 12.0%) and F28C in GT-2c (2 of 11, 18.2%). Overall, personalized treatment led to 100% SVR12, even in those patients for whom treatment strategy was either strengthened (by ribavirin inclusion and/or duration increase) or simplified (by ribavirin exclusion and/or duration reduction), thanks to baseline RAS evaluation. Conclusions: Even with newer DAA regimens, an integrated interpretation of clinical and virological pretreatment parameters, including natural RASs, may play a relevant role in bringing SVR rates close to the highest achievable. Treatment tailoring can be foreseen in 'hard-to-treat' patients, but also in 'easy' patients with favourable indicators, whereby a simplification/shortening of recommended regimens can be indicated

The lowest X4 Geno2Pheno false-positive rate is associated with greater CD4 depletion in HIV-1 infected patients

Through this study we evaluated whether the HIV-1 tropism determined by genotypic analysis correlates with HIV-1 markers, such as CD4 cell count and plasma HIV-RNA. The analysis was performed on 1221 HIV-1 B-subtype infected patients with an available V3 sequence (all maraviroc naive). Of them, 532 were antiretroviral therapy (ART) naive and 689 ART experienced. Tropism determination was performed by using the geno2pheno (co-receptor) algorithm set at a false-positive rate (FPR) of 10% and 2%. Potential associations of FPR with CD4 cell count and viraemia were evaluated. Association of V3 mutations with genotypic-determined tropism was also evaluated according to different FPR ranges. About 26% of patients (either ART naive or ART experienced) were infected by X4-tropic viruses (using the classical 10% FPR cut-off). However, a significantly lower proportion of ART-naive patients had FPR ≤ 2% in comparison with ART-experienced patients (4.9% vs. 12.6%, respectively, p <0.001). The risk of advanced HIV-1 infection (with CD4 cell count ≤ 200 cells/mm(3) ) was significantly greater in X4-infected patients, either ART-naive (OR (95% CI)), 4.2 (1.8-9.2); p 0.0006) or ART-experienced (2.3 (1.4-3.6); p 0.0003), with FPR set at 2% (but not at 10%). This finding was confirmed by multivariable logistic analysis. No relationship was found between viraemia and FPR ≤2%. Some X4-related mutations were significantly associated with FPR ≤2% (ART-naive patients, S11R, Y21V, G24K and G24R, p ≤0.001; ART-experienced patients, Y7K, S11R, H13Y, p ≤0.002). In conclusion, these findings show that within the context of genotypically-assessed CXCR4 tropism, FPR ≤2% defines (far better than 10%-FPR) a viral population associated with low CD4 rank, with potentially greater cytopathic effect, and with more advanced disease

Managing the long surviving HIV patient : a proposal for a multidimensional first-level diagnostic assessment

We propose a multidimensional first-level diagnostic assessment easy to use in routine clinical practice to allow infectious disease specialists to have a general and complete overview of persons living with HIV. Following the Delphi method, articles published from January 1, 2011 on controlled trials, clinical reports and observational studies dealing specifically with HIV and its co-morbidities were selected for review by the authors. Participants in the poll were selected among clinicians and infectious diseases specialists, working in 38 different dedicated HIV centres in Italy. The participants were given access to a website dedicated to the project and received a standardized information package containing a synopsis of the study and a description of the Delphi process and the selected literature. A total of 131 Items were divided into 10 first-level survey areas: anamnesis, objective examination, infectious diseases, osteoporosis diagnosis, metabolic pathologies diagnosis, cardiovascular diagnosis, nephrologic diagnosis, hepatological diagnosis, central nervous system diagnosis, evaluation of quality of life (QoL). This simple and concise first level tool identifies a few areas of multi-organ diagnostic assessment beyond the infectivity area. The identification of these areas will allow us to find shared and validated evaluation procedures with the intent to increase the likelihood of early recognition of patients at risk of comorbidity development, in order to facilitate more effective prevention, thereby reducing the overall impact on the quality of life of patients affected by this chronic illness

Durability of lopinavir/ritonavir monotherapy in individuals with viral load ≤50 copies/ml in an observational setting

BACKGROUND: The main objective is to evaluate the efficacy and durability of lopinavir-ritonavir monotherapy (LPV/r-MT) in virologically controlled HIV-positive individuals switching from combination antiretroviral therapy (cART). METHODS: Criteria to be included in this observational study were to have initiated for the first time LPV/r-MT after ≥2 consecutive HIV RNA≤50 copies/ml achieved on a ≥3-drug-including regimen. The main end points were time to virological rebound (VR; defined in two ways: time of first of two consecutive viral load [VL]>50 and >200 copies/ml), time to discontinuation/intensification and time to experience either a single VL>200 copies/ml or discontinuation/intensification (treatment failure [TF]). Individuals' follow-up accrued from the date of starting LPV/r-MT to event or last available VL. Kaplan-Meier curves and Cox regression analyses were used. RESULTS: A total of 228 individuals were included: median age 46 years (IQR 40-50), 36% females, 36% intravenous drug users and 25% HCV-coinfected. Median CD4(+) T-cell count at nadir was 215 cell/mm(3) (IQR 116-336) and at baseline was 615 cell/mm(3) (IQR 436-768). By 36 months after switching to LPV/r-MT, the proportion of individuals with VR (confirmed VL>200 copies/ml) was 11% and with TF was 35%. In the multivariable Cox model the factors associated with a lower risk of TF was the duration of viral suppression <50 copies/ml prior to baseline (ARH=0.92; 95% CI 0.85, 0.99; P=0.024, per 6 months longer) and having LPV/r as part of last cART (ARH=0.45; 95% CI 0.21, 0.95; P=0.037). CONCLUSIONS: In daily clinical practice, we confirm a relatively safe approach of treatment simplification to LPV-MT in a selected population with long-lasting virological control