Mediastinal lymphadenopathies and skin lesions in a 49-year-old Sinhalese man.

Leprosy is a neglected disease sporadically reported in high-income countries. Skin lesion and peripheral nerve involvement represent most common manifestations. Mediastinal lymphadenopathy in the absence of superficial lymph nodes involvement is very rare. Atypical or rare clinical presentations of disease may delay diagnosis and therapy and cause potential life-threatening manifestations and disabilities. We describe the case of a 49-year-old Sinhalese man who was admitted to our hospital with a one-month history of peripheral neurological symptoms and skin lesions on lower limbs. CT scan showed the presence of mediastinal lymphadenopathies without lung parenchymal and superficial lymph nodes involvement. Endobronchial ultrasound-guided transbronchial needle aspiration showed the presence of granulomas while skin biopsy revealed dermo-hypodermic granulomas with perineural lymphohistiocytic inflammatory reaction. Fite-Faraco staining demonstrated the presence of acid-fast bacilli in both lymph nodal and skin biopsy and polymerase chain reaction was positive for Mycobacterium leprae. Multibacillary leprosy was then diagnosed.

Three case reports of West Nile virus neuroinvasive disease: lessons from real-life clinical practice.

BACKGROUND: Despite being an uncommon cause of meningoencephalitis, West Nile virus (WNV) recently provoked significant outbreaks throughout Europe. West Nile neuroinvasive disease (WNND) is associated with significant morbidity and mortality in older and compromised individuals, while its diagnosis may be demanding for the clinician. Here discussed are three cases of WNND with a focus on the diagnostic challenges they presented due to atypical clinical presentation and laboratory findings. CASE PRESENTATION: Between July and September 2020 three patients presented to our attention with signs and symptoms compatible with meningoencephalitis. Among routine procedures, they underwent lumbar puncture and imaging. In the absence of microbiological isolates, biological samples were sent for serology and NAATs for WNV. Following diagnosis, the patients gradually recovered and were discharged either home or to rehabilitation facilities. CONCLUSIONS: The laboratory findings here discussed, in particular CSF parameters, are only partially consistent with those described in the literature, which highlights the need for further research. While serology and NAATs on blood and urine appear the most reliable techniques in the diagnostic work-up of WNND, utility of NAATs on CSF specimens is limited by the kinetics of WNV viremia in biological fluids. This report underlines that WNND should always be included in the differential diagnosis of meningoencephalitis during WNV transmission period.

Impact of daily versus weekly service of infectious diseases consultation on hospital antimicrobial consumption: a retrospective study.

BACKGROUND: To verify whether a daily service of Infectious Diseases consultation (ID-cons) is more effective than a weekly service in reducing antibiotic (ATB) consumption without worsening of clinical outcomes. METHODS: Two-year observational analysis of the ID-cons provided in a hospital setting in Milan, Italy. ID-cons resulted in: start-of-ATB; no-ATB; confirmation; modification-of-ATB. The impact of a weekly (September 1, 2016 - August 31, 2017 versus a daily (September 1, 2017 - September 30, 2018) service of ID-cons was evaluated in terms of: time-from-admission-to-first-ID-cons, type of ATB-intervention and number-of-ID-cons per 100 bed-days (bd). Primary outcomes: reduction of hospital ATB consumption overall and by department and classes expressed as Defined Daily Dose (DDD)/100bd (by Wilcoxon test for paired data). SECONDARY OUTCOMES: overall and sepsis-related in-hospital annual mortality rates (as death/patient's admissions). RESULTS: Overall 2552 ID-cons in 1111 patients (mean, 2.3 ID-cons per patient) were performed (18.6% weekly vs 81.4% daily). No differences in patient characteristics were observed. In the daily-service, compared to the weekly-service, patients were seen by the ID-consultant earlier (time-from-admission-to-ID-cons: 6 days (IQR 2-13) vs 10 days (IQR 6-19), p < 0.001) and ATB was more often started by the ID-consultant (Start-of-ATB: 11.6% vs 8%, p = 0.02), rather than treating physicians. After switching to daily-service, the number-of-ID-cons increased from 0.4/100bd to 1.5/100bd (p = 0.01), with the greatest increase in the emergency department (1.5/100bd vs 6.7/100bd, p < 0.001). Total ATB consumption decreased from 64 to 60 DDD/100bd. As for the number-of-cons, the consumption of ATB decreased mainly in the emergency area. According to ATB classes, glycopeptides consumption was reduced from 3.1 to 2.1 DDD/100bd (p = 0.02) while carbapenem use decreased from 3.7 to 3.1 DDD/100bd (p = 0.07). No changes in overall mortality (5.2% vs 5.2%) and sepsis-related mortality (19.3% vs 20.9%; p = 0.7) were observed among the two time-period. CONCLUSIONS: Daily-ID-cons resulted in a more comprehensive management of the infected patient by the ID-consultant, especially in the emergency area where we also observed the highest rate of reduction of ATB-usage. No change in mortality was observed.

Bacterial coinfections in dengue virus disease: what we know and what is still obscure about an emerging concern.

PURPOSE: Dengue virus is the most frequent arthropod-borne viral infection worldwide. Simultaneously to the growth of its incidence, cases of bacterial coinfection in dengue have been increasingly reported. The clinical course of dual infections may worsen for reciprocal interactions and delays in the diagnosis, so that clinicians should be aware of this eventuality. Therefore, we reviewed literature to provide an overview of the epidemiological, clinical, and physiopathological issues related to bacterial coinfections and bacteremia in dengue. METHODS: Clinical studies and case reports regarding bacteremia and bacterial coinfections in dengue and the interactions between the pathogens published on PubMed were reviewed. RESULTS: We found 26 case reports, only 3 studies on concurrent bacteremia and 12 studies reporting data on bacterial coinfections in dengue. According to the three available studies, the 0.18-7 % of dengue infections are accompanied by concurrent bacteremia, while the 14.3-44.4 % of dengue-related deaths seem associated to bacterial coinfections. Comorbidities, advanced age, and more severe dengue manifestations could be risk factors for dual infections. A longer duration of fever and alterations in laboratory parameters such as procalcitonin, hyponatremia, leukocyte count, and renal function tests can raise the suspicion. CONCLUSIONS: Despite the real burden and consequences of this emerging concern is still not computable accurately due to the lack of a significant number of studies on large cohorts, clinicians need a greater awareness about it to early recognize warning signs, to properly use available diagnostic tools and to readily start antibiotic treatment able to prevent worsening in mortality and morbidity.

Switching to unboosted atazanavir reduces bilirubin and triglycerides without compromising treatment efficacy in UGT1A1*28 polymorphism carriers.

OBJECTIVES: Hyperbilirubinaemia is a frequent complication of atazanavir-containing antiretroviral therapy and its severity is related to UDP-glucuronosyl transferase (UGT) 1A1*28 polymorphism. The aim of this study was to evaluate the safety and outcome of unboosted atazanavir-containing regimens based on the genetic constitution. METHODS: Fifty-one HIV-1-infected patients on boosted atazanavir were prospectively enrolled in the study. Twenty-five patients with a UGT1A1*28 allele switched to 400 mg of unboosted atazanavir. RESULTS: At baseline, UGT1A1 heterozygous and homozygous patients had significantly higher bilirubin levels than wild-type (P = 0.012 and P < 0.001, respectively). After ritonavir removal, a reduction was observed in total bilirubin (from 4.09 to 1.82 mg/dL; P < 0.001), γ-glutamyl transpeptidase (P = 0.015), triglycerides (P = 0.03) and total cholesterol (P = 0.05). No significant changes in CD4 T cell count and no increases in viral load were observed 12 months after unboosting. Plasma drug monitoring after ritonavir removal revealed the presence of therapeutic atazanavir concentrations in all patients except one with poor therapy adherence. CONCLUSIONS: UGT1A1*28 is significantly related to hyperbilirubinaemia in HIV-1 patients receiving atazanavir. Genotyping before the initiation of antiretroviral therapy can reduce the emergence of severe hyperbilirubinaemia. Unboosted atazanavir-containing therapy is safe and efficacious in patients with an undetectable viral load with a UGT1A1*28 polymorphism, allowing the use of atazanavir in patients otherwise likely unable to receive it.

Efficacy and safety of a dual boosted protease inhibitor-based regimen, atazanavir and fosamprenavir/ritonavir, against HIV: experience in a pediatric population.

BACKGROUND: Although dual-boosted protease inhibitors regimen is not recommended in children with HIV infection, such a strategy could be useful in subjects with a complex resistance profile. This study was aimed at assessing the long term efficacy and safety of a double-boosted protease inhibitor combination, fosamprenavir (fAVP) and atazanavir/ritonavir (ATV/r) in a cohort of HIV-infected children and adolescents who had failed with nucleoside reverse transcriptase inhibitors. METHODS: Seven vertically infected children and adolescents who had previously failed highly active antiretroviral therapy and were resistant to nucleoside reverse transcriptase inhibitors, received a dual protease inhibitor (PI) regimen including fAVP plus ATV/r for 42 months. The patients were assessed at baseline, every month for the first 24 weeks of therapy and every 3 months until month 32. Physical examination, CD4+ cell count, HIV-RNA viral load, lipid profile and hepatic function were assessed throughout the follow up. RESULTS: During the study no serious adverse events were reported. CD4 absolute number increased over-time in all subjects. At baseline the median HIV-RNA was 6562 cp/mL (ranging 1048 -102772 cp/mL) and rapidly decreased below the limit of detection (50 cp/mL) after 2 months of the new treatment and remained undetectable in all cases through the entire study period. At the beginning of the study all cases showed a normal lipid profile. During the study period, 4/7 subjects showed total cholesterol, low density lipoprotein and triglyceride levels >97th cent.le for the males and 94th cent.le for the females. HDL cholesterol showed protective values. Hepatic enzymes remained stable during the entire observation, whereas total bilirubin showed toxicity II/III grade in 6/7 subjects. No change in fat redistribution and insulin resistance was observed. CONCLUSION: Dual-boosted protease inhibitor therapy was virologically and immunologically effective and it could be considered as a possible alternative to a rescue regimen in children and adolescents. However, hypercholesterolemia and hypertriglyceridemia need close follow-up and may limit the use of this therapeutic option.

Real World Estimate of Vaccination Protection in Individuals Hospitalized for COVID-19.

Whether vaccination confers a protective effect against progression after hospital admission for COVID-19 remains to be elucidated. Observational study including all the patients admitted to San Paolo Hospital in Milan for COVID-19 in 2021. Previous vaccination was categorized as: none, one dose, full vaccination (two or three doses >14 days before symptoms onset). Data were collected at hospital admission, including demographic and clinical variables, age-unadjusted Charlson Comorbidity index (CCI). The highest intensity of ventilation during hospitalization was registered. The endpoints were in-hospital death (primary) and mechanical ventilation/death (secondary). Survival analysis was conducted by means of Kaplan-Meier curves and Cox regression models. Effect measure modification by age was formally tested. We included 956 patients: 151 (16%) fully vaccinated (18 also third dose), 62 (7%) one dose vaccinated, 743 (78%) unvaccinated. People fully vaccinated were older and suffering from more comorbidities than unvaccinated. By 28 days, the risk of death was of 35.9% (95%CI: 30.1−41.7) in unvaccinated, 41.5% (24.5−58.5) in one dose and 28.4% (18.2−38.5) in fully vaccinated (p = 0.63). After controlling for age, ethnicity, CCI and month of admission, fully vaccinated participants showed a risk reduction of 50% for both in-hospital death, AHR 0.50 (95%CI: 0.30−0.84) and for mechanical ventilation or death, AHR 0.49 (95%CI: 0.35−0.69) compared to unvaccinated, regardless of age (interaction p > 0.56). Fully vaccinated individuals in whom vaccine failed to keep them out of hospital, appeared to be protected against critical disease or death when compared to non-vaccinated. These data support universal COVID-19 vaccination.

Female gender is associated with long COVID syndrome: a prospective cohort study.

OBJECTIVE: We explored the association between female gender and long COVID syndrome, defined as persistence of physical and/or psychological symptoms for more than 4 weeks after recovery from acute COVID-19 disease. The secondary aim was to identify predictors of long COVID syndrome by multivariable logistic regression analysis. METHODS: This was a single-centre prospective cohort study conducted at San Paolo Hospital in Milan, Italy. We enrolled adult patients who were evaluated at the post-COVID outpatient service of our Infectious Diseases Unit between 15 April 2020 and 15 December 2020. Participants were individuals who had clinically recovered from COVID-19 and in whom virological clearance had occurred. Previous infection by SARS-CoV-2 was microbiologically documented by positivity using a reverse-transcriptase polymerase chain reaction (RT-PCR) assay of nasopharyngeal swab. All enrolled patients underwent blood tests and a comprehensive medical examination at follow-up. Individuals were interviewed about resolved and persisting symptoms and were asked to fill in two questionnaires to allow assessment of the Hospital Anxiety and Depression symptoms (HADS) score and of the Impact of Event Scale-Revised (IES-R) score. RESULTS: A total of 377 patients were enrolled in the study. The median time from symtpom onset to virological clerance was 44 (37-53) days. A diagnosis of long COVID syndrome was made in 260/377 (69%) patients. The most common reported symptoms were fatigue (149/377, 39.5%), exertional dyspnoea (109/377, 28.9%), musculoskeletal pain (80/377, 21.2%) and "brain fog" (76/377, 20.2%). Anxiety symptoms were ascertained in 71/377 (18.8%) individuals, whereas 40/377 (10.6%) patients presented symptoms of depression. Post-traumatic stress disorder (defined by a pathological IES-R score) was diagnosed in one-third of patients (85/275, 31%). Female gender was independently associated with long COVID syndrome at multivariable analysis (AOR 3.3 vs. males, 95% CI 1.8-6.2, p < 0.0001). Advanced age (adjusted (A)OR 1.03 for 10 years older, 95% CI 1.01-1.05, p 0.01) and active smoking (AOR 0.19 for former smokers vs. active smokers, 95% CI 0.06-0.62, p 0.002) were also associated with a higher risk of long COVID, while no association was found between severity of disease and long COVID (AOR 0.67 for continuous positive airway pressure (CPAP)/non-invasive mechanical ventilation (NIMV)/orotracheal intubation (OTI) vs. no 02 therapy, 95% CI 0.29-1.55, p 0.85). DISCUSSION: Factors that were found to be associated with a higher risk of developing "long COVID" syndrome were female gender, older age and active smoking, but not severity of the acute disease. Individuals affected by SARS-CoV-2 infection with the aforementioned features should be early identified and involved in follow-up programmes.

Comparative evaluation of seven resistance interpretation algorithms and their derived genotypic inhibitory quotients for the prediction of 48 week virological response to darunavir-based salvage regimens.

BACKGROUND: the darunavir genotypic inhibitory quotient (gIQ) has been suggested as one of the predictors of virological response to darunavir-containing salvage regimens. Nevertheless, which resistance algorithm should be used to optimize the calculation of gIQ is still debated. The aim of our study was to compare seven different free-access resistance algorithms and their derived gIQs as predictors of 48 week virological response to darunavir-based salvage therapy in the clinical setting. METHODS: patients placed on two nucleoside reverse transcriptase inhibitors + 600/100 mg of darunavir/ritonavir twice daily  ±â€Š enfuvirtide were prospectively evaluated. Virological response was assessed at 48 weeks. Darunavir resistance interpretation was performed according to seven different algorithms, of which two were weighted algorithms. Analysis of other factors potentially associated with virological response at 48 weeks was performed. RESULTS: fifty-six treatment-experienced patients were included. Overall, 35 patients (62.5%) had a virological response at 48 weeks. Receiver operator characteristic curve analysis showed that De Meyer's weighted score (WS) and its derived gIQ (gIQ WS) were the most accurate parameters defining virological response, and related cut-offs showed the best sensitivity/specificity pattern. In univariate logistic regression analysis, baseline log viral load (P = 0.028), optimized background score ≥ 2 (P = 0.048), WS >5 (P = 0.001) and WS gIQ ≥ 600 (P < 0.0001) were independently associated with virological response. In multivariate analysis, only baseline log viral load (P = 0.008) and WS gIQ ≥ 600 (P < 0.0001) remained in the model. CONCLUSIONS: in our study, although different resistance interpretation algorithms and derived gIQs were associated with virological response, gIQ WS was the most accurate predictive model for achieving a successful virological response.

Long-term positive effect of an educational antimicrobial stewardship program implemented in an Internal Medicine Department: a prospective analysis and a point prevalence survey on long-term effect.

The study aims to evaluate antimicrobial consumption and appropriateness one year after the implementation of an antimicrobial stewardship (AMS) program in an Internal Medicine Department in Milan. AMS program structured in two phases: "AMS phase", 5 months AMS-program based on an "audit-and-feedback model"; the "follow-up phase", 5 months long point prevalence survey conducted one year later. Outcomes of the study: antimicrobial consumption and appropriateness of antimicrobial therapy. Secondary outcomes: in-hospital mortality and length of stay (LOS). During the "AMS phase", we obtained a mean decrease of -11.4% of total antibiotic consumption as compared to the previous year (67.9 defined daily dose (DDD)/100 bed-days (bd) vs. 79.4 DDD/100bd, p = 0.07). Antibiotic consumption remained stable during "follow-up phase" (66.3 DDD/100bd, p = 0.9). Rate of appropriateness during the project increased from 48% to 85% (p < 0.01). No difference in in-hospital mortality and in LOS were observed. The study documents a positive long-term effect of AMS program on consumption and appropriate use of antibiotics.

Incidence, Risk Factors and Impact on Clinical Outcomes of Bloodstream Infections in Patients Hospitalised with COVID-19: A Prospective Cohort Study.

With the aim of describing the burden and epidemiology of community-acquired/healthcare-associated and hospital-acquired bloodstream infections (CA/HCA-BSIs and HA-BSIs) in patients hospitalised with COVID-19, and evaluating the risk factors for BSIs and their relative impact on mortality, an observational cohort study was performed on patients hospitalised with COVID-19 at San Paolo Hospital in Milan, Italy from 24 February to 30 November 2020. Among 1351 consecutive patients hospitalised with COVID-19, 18 (1.3%) had CA/HCA-BSI and 51 (3.8%) HA-BSI for a total of 82 episodes of BSI. The overall incidence of HA-BSI was 3.3/1000 patient-days (95% CI 2.4-4.2). Patients with HA-BSI had a longer hospital stay compared to CA/HCA-BSI and no-BSI groups (27 (IQR 21-35) vs. 12 (7-29) vs. 9 (5-17) median-days, p < 0.001) but a similar in-hospital mortality (31% vs. 33% vs. 25%, p = 0.421). BSI was not associated with an increased risk of mortality (CA/HCA-BSI vs. non-BSI aOR 1.27 95% CI 0.41-3.90, p = 0.681; HA-BSI vs. non-BSI aOR 1.29 95% CI 0.65-2.54, p = 0.463). Upon multivariate analysis, NIMV/CPAP (aOR 2.09, 95% CI 1.06-4.12, p = 0.034), IMV (aOR 5.13, 95% CI 2.08-12.65, p < 0.001) and corticosteroid treatment (aOR 2.11, 95% CI 1.06-4.19, p = 0.032) were confirmed as independent factors associated with HA-BSI. Development of HA-BSI did not significantly affect mortality. Patients treated with corticosteroid therapy had double the risk of developing BSI.

Declining Mortality Rate of Hospitalised Patients in the Second Wave of the COVID-19 Epidemics in Italy: Risk Factors and the Age-Specific Patterns.

BACKGROUND: Mortality rate from COVID-19 in Italy is among the world's highest. We aimed to ascertain whether there was any reduction of in-hospital mortality in patients hospitalised for COVID-19 in the second-wave period (October 2020-January 2021) compared to the first one (February-May 2020); further, we verified whether there were clusters of hospitalised patients who particularly benefitted from reduced mortality rate. METHODS: Data collected related to in-patients' demographics, clinical, laboratory, therapies and outcome. Primary end-point was time to in-hospital death. Factors associated were evaluated by uni- and multivariable analyses. A flow diagram was created to determine the rate of in-hospital death according to individual and disease characteristics. RESULTS: A total of 1561 patients were included. The 14-day cumulative incidence of in-hospital death by competing risk regression was of 24.8% (95% CI: 21.3-28.5) and 15.9% (95% CI: 13.7-18.2) in the first and second wave. We observed that the highest relative reduction of death from first to second wave (more than 47%) occurred mainly in the clusters of patients younger than 70 years. CONCLUSIONS: Progress in care and supporting therapies did affect population over 70 years to a lesser extent. Preventive and vaccination campaigns should focus on individuals whose risk of death from COVID-19 remains high.

Chest pain and a left parasternal soft tissue swelling in an immunocompetent refugee with disseminated tuberculosis.

An immunocompetent migrant with chest pain was admitted to an Italian hospital. Computed tomography showed a left pectoral abscess and osteomyelitis of the sternum. The infection had spread into the anterior mediastinum near to the pericardium and the heart, where an atrial mass was confirmed by echocardiography. Disseminated tuberculosis was diagnosed.

The importance of patients' case-mix for the correct interpretation of the hospital fatality rate in COVID-19 disease.

OBJECTIVE: We aimed to document data on the epidemiology and factors associated with clinical course leading to death of patients hospitalised with COVID-19. METHODS: Prospective observational cohort study on patients hospitalised with COVID-19 disease in February-24th/May-17th 2020 in Milan, Italy. Uni-multivariable Cox regression analyses were performed. Death's percentage by two-weeks' intervals according to age and disease severity was analysed. RESULTS: A total of 174/539 (32.3%) patients died in hospital over 8228 person-day follow-up; the 14-day Kaplan-Meier probability of death was 29.5% (95%CI: 25.5-34.0). Older age, burden of comorbidities, COVID-19 disease severity, inflammatory markers at admission were independent predictors of increased risk, while several drug-combinations were predictors of reduced risk of in-hospital death. The highest fatality rate, 36.5%, occurred during the 2nd-3rd week of March, when 55.4% of patients presented with severe disease, while a second peak, by the end of April, was related to the admission of older patients (55% ≥80 years) with less severe disease, 30% coming from long-term care facilities. CONCLUSIONS: The unusual fatality rate in our setting is likely to be related to age and the clinical conditions of our patients. These findings may be useful to better allocate resources of the national healthcare system, in case of re-intensification of COVID-19 epidemics.

New macrocyclic amines showing activity as HIV entry inhibitors against wild type and multi-drug resistant viruses.

Considering as a lead molecule the chemokine CXCR4 receptor antagonist AMD-3100, which shows significant anti-HIV activity in vitro and in vivo, we investigated a series of structurally related macrocyclic polyamines incorporating o,o'-phenanthroline or 2,2'-bipyridyl scaffolds as potential antiviral agents with lower toxicity and increased activity against both wild type X4-tropic and dual tropic HIV strains. The antiviral activity of these compounds was evaluated by susceptibility assays in PBMC (Peripheral Blood Mononuclear Cells) and compared to that of AMD-3100. The newly investigated compounds showed IC(50)s values in the low micromolar range and significantly inhibited the viral replication of wild type X4-tropic isolate and dual tropic strains. These macrocyclic polyamines constitute a promising class of HIV entry inhibitors.

Evolution of the HIV-1 protease region in heavily pretreated HIV-1 infected patients receiving Atazanavir.

BACKGROUND: Previous in vitro studies indicated that Atazanavir (ATV) has a distinct resistance profile than other protease inhibitors (PIs). In treatment-experienced patients ATV resistance is characterised by the accumulation of at least four mutations among those that confer cross-resistance to the PIs. OBJECTIVE: We studied the evolution of PIs resistance mutations in 10 HAART-failed patients undergoing ATV enrolled in an early access program. STUDY DESIGN: Virus genotypic resistance was determined from plasma collected at baseline and during treatment. HIV-RNA was extracted and the pol region amplified and sequenced. Genotypic data were used to determine drug susceptibility. Phylogenetic analysis was performed. RESULTS: At baseline, genotypic data showed cross-resistance patterns to approved PIs in 6 patients. In two of these subjects new mutations (I54V and A71V) conferring cross-resistance emerged after 3 months of therapy. The I50L mutation was evidenced in one subject after 12 months of treatment. The "virtual" phenotype analysis mirrored the resistance profiles to ATV and other PIs and evidenced differences with tipranavir and darunavir. CONCLUSION: Genotype evolution within the protease region did not emerge at significant levels during salvage therapy of multidrug-experienced patients. ATV exhibited certain/same virologic effect on the majority of our patients.

Abacavir hypersensitivity reaction after switching from the twice-daily to the once-daily formulation.

A 36-year-old man with HIV infection developed a reaction compatible with an abacavir hypersensitivity reaction after switching from the twice-daily to the once-daily formulation. The switch was determined by a more convenient intake. The patient was treated with abacavir twice-daily plus lamivudine and efavirenz for more than 5 years with no side effects. At the time of this change, his CD4 count was 1069 cell/mm(3) and HIV-RNA undetectable. Our case suggests that patients should be carefully monitored after switching, and warned about the potential effects.

Unusual codon 69 insertions: influence on human immunodeficiency virus type 1 reverse transcriptase drug susceptibility.

INTRODUCTION: Multiple amino acid changes in the reverse transcriptase (RT) enzyme of the human immunodeficiency virus 1 (HIV-1) confer simultaneous resistance to most nucleoside RT inhibitors (NRTI). It may take place through different pathways: one of these is the codon 69 insertion, which can involve several 2-amino acid patterns. MATERIALS AND METHODS: We are reporting the case of three patients treated with various antiretroviral compounds. For these subjects we have conducted both a genotypical and a phenotypical analysis in order to understand what kind of influence these insertions may have on HIV-1 RT drug susceptibility. Plasma samples from these patients have been extracted and the RT region has been amplified, cloned and sequenced; meanwhile their PBMCs have been separated, cultivated and then tested for drug susceptibility. RESULTS: Data obtained from the cloning assay showed that the patients had different mutational patterns but constant multiple resistance to NRTI. In particular, they harbored mutations related to Zidovudine (ZDV), 3TC and various NRTIs. Moreover, all three samples had a T69S substitution followed by three different dual amino acid insertions: SG, TG and VG. Several phenotypic experiments revealed that the viruses were resistant to 3TC as well as to ZDV and ABC. Different results were obtained using d4T and ddI. DISCUSSION: In our three patients, all mutation inserts impaired the use of NRTI, particularly ZDV and 3TC. Patient 001 presented a pattern that should not cause a high phenotypic resistance to 3TC per se, and so we can argue that the concomitant presence of the insertion T69S (SG) makes this isolate moderately resistant to this drug. We observed a similar phenomenon in subject 003. d4T was less involved in the resistance generation caused by the RT insertion (in one out of three cases). Moreover, we identified a new 2aa insertion (TG) that has, to the best of our knowledge, never been reported before. A careful survey of novel RT genotypic insertion is thus warranted.