Serum bactericidal titres for monitoring antimicrobial therapy: current status and potential role in the management of multidrug-resistant Gram-negative infections.

BACKGROUND: Serum bactericidal titres (SBTs) were widely used in the 1970s and 1980s to monitor antimicrobial therapy but are now seldom recommended. It is the only laboratory test that integrates drug pharmacodynamics, host pharmacokinetics and synergistic or antagonistic interactions of antimicrobial combinations into a single index of antimicrobial activity. We hypothesized that SBTs could play a renewed role in monitoring antibiotic treatment of multidrug-resistant Gram-negative infections. However, the last critical appraisal of the test was published over 30 years ago. OBJECTIVES: This narrative review provides an updated assessment of the SBT test and its methodological limitations. We performed a diagnostic meta-analysis to estimate the value of SBTs for predicting clinical failure or death during antibiotic treatment. SOURCES: A comprehensive literature search of PubMed including all English publications was performed in December 2019 using the Medical Subject Headings (MeSH search terms "serum", "bactericidal", "inhibitory", "titre", "monitoring", "anti-infective agents" "antimicrobial therapy" and "therapeutic drug monitoring"). CONTENT: Although standardized methods for performing SBTs were approved in 1999, the test remains labour intensive, and results may not be available until 72 hr. However, the use of non-culture-based endpoints (i.e. spectrophotometric or fluorescent) may shorten test time to 24 hr. Despite considerable heterogeneity in published studies, a meta-analysis of 11 evaluable studies published from 1974 to 2007 indicated a critical SBT result (peak SBT ≤1:8 or trough ≤1:2) is associated with a diagnostic odds ratio for clinical failure during antibiotic treatment of 12.27 (95% confidence interval 5.28-28.54) and a 5.32 (95% 1.32-21.42) odds of death. IMPLICATIONS: SBTs have prognostic value for identifying patients at high risk for antibiotic treatment failure, but the slow turnaround time of the current test limits its clinical utility. Standardization of a more rapid SBT testing method is needed.

Bloodstream infection caused by KPC-producing Klebsiella pneumoniae resistant to ceftazidime/avibactam: epidemiology and genomic characterization.

OBJECTIVES: The aim of this study was to evaluate the incidence of ceftazidime/avibactam resistance among Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae (KPC-Kp) strains isolated from patients with bloodstream infection. METHODS: We collected 120 carbapenemase producing Enterobacteriaceae (CPE) strains from unique patients hospitalized in two Italian hospitals between January 2018 to February 2019. Strains were phenotypically characterized for the type of carbapenemase production and susceptibility to ceftazidime/avibactam. Ceftazidime/avibactam-resistant strains were characterized by whole-genome sequencing. RESULTS: During the study period, we characterized 105 (87.5%) KPC producers among a total of 120 CPE strains. Ceftazidime/avibactam resistance was found in three KPC-Kp strains isolated from patients with no history of previous ceftazidime/avibactam-based treatment. Of note, two out of three ceftazidime-avibactam-resistant KPC-Kp were also resistant to meropenem/vaborbactam. Genomic characterization showed that a ceftazidime/avibactam-resistant KPC-Kp harboured a mixed population with D179Y mutated KPC-2, while the other two ceftazidime-avibactam-resistant KPC-Kp possessed non-functional ompK35-ompK37 and mutated ompK36 porins associated with higher copy number of blaKPC gene. CONCLUSIONS: Our results showed that incidence of ceftazidime/avibactam resistance emerged in KCP-Kp strains independently from previous antimicrobial exposure. Resistance to ceftazidime/avibactam was associated with mutations within the blaKPC gene or porin deficiency associated with higher blaKPC copy number and is also related to the meropenem/vaborbactam resistance.

Colonization and infection due to carbapenemase-producing Enterobacteriaceae in liver and lung transplant recipients and donor-derived transmission: a prospective cohort study conducted in Italy.

OBJECTIVES: A prospective cohort study was conducted in Italy in order to describe the microbiologic aspects of colonization/infection by carbapenemase-producing Enterobacteriaceae (CPE) in donors and recipients of lung and liver transplants and the possible CPE transmission from donors to recipients. METHODS: Between 15 January 2014 and 14 January 2015, all recipients of solid organ transplants (SOT) at ten lung and eight liver transplantation centres and the corresponding donors were enrolled. Screening cultures to detect CPE were performed in donors, and screening and clinical cultures in recipients with a 28-day microbiologic follow-up after receipt of SOT. Detection of carbapenemase genes by PCR, genotyping by multilocus sequence typing, and pulsed-field gel electrophoresis and whole-genome sequencing were performed. RESULTS: Of 588 screened donors, 3.4% were colonized with CPE. Of the liver first transplant recipients (n = 521), 2.5% were colonized before receipt of SOT and 5% acquired CPE during follow-up. CPE colonization was higher in lung first transplant recipients (n = 111, 2.7% before SOT and 14.4% after SOT). CPE infections occurred in 1.9% and 5.3% of liver or lung recipients, respectively. CPE isolates were mostly Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae belonging to CG258. Three events of donor-recipient CPE transmission, confirmed by whole-genome sequencing and/or pulsed-field gel electrophoresis, occurred in lung recipients: two involving K. pneumoniae sequence type 512 and one Verona integron-encoded metallo-ß-lactamase (VIM)-producing Enterobacter aerogenes. CONCLUSIONS: This study showed a low risk of donor-recipient CPE transmission, indicating that donor CPE colonization does not necessarily represent a contraindication for donation unless colonization regards the organ to be transplanted. Donor and recipient screening remains essential to prevent CPE transmission and cross-infection in transplantation centres.

New evidence of cutaneous leishmaniasis in north-eastern Italy.

BACKGROUND: Human leishmaniasis is on increase in the Mediterranean Europe. However, the exact prevalence of cutaneous leishmaniasis (CL) is largely unknown as underdiagnosis and under reporting are common. OBJECTIVE: To evaluate epidemiological, clinicopathological and microbiological aspects of CL cases occurring in the Bologna Province, north-eastern Italy. METHODS: We performed a retrospective, observational study on CL cases diagnosed in the Bologna Province between January 2013 and December 2015. RESULTS: During 2013-2015, 30 cases of CL were identified in the Bologna Province with an average incidence of 1.00/100 000, with an increase of fourfold to 12-fold as compared to previous years. 16 of 30 (53%) CL cases presented as single, typical lesions. CL diagnosis was carried out by histological and molecular techniques, although in 7 of 29 (24%) PCR-positive cases, amastigotes were not visible on histology. CONCLUSIONS: We report new evidence of CL cases in a focal area of north-eastern Italy in 2013-2015. Our study highlights the importance of CL surveillance in the Mediterranean basin and emphasizes the need for the molecular laboratory surveillance of CL in endemic areas.

Characterization of antibody response in neuroinvasive infection caused by Toscana virus.

OBJECTIVES: Among sandfly-borne pathogens, Toscana virus (TOSV) is a prominent cause of summer meningitis in Mediterranean Europe. Here, we assessed the kinetics of anti-TOSV antibodies over time in 41 patients diagnosed with TOSV meningitis or meningoencephalitis in northeastern Italy. METHODS: Acute and follow-up serum samples were collected up to 20 months after diagnosis of TOSV infection and tested for the presence of specific antibody using immunoenzymatic and indirect immunofluorescence assays. In addition, maturation of anti-TOSV IgG over time was evaluated as well as production of neutralizing antibodies. RESULTS: Specific IgM and IgG response was present at diagnosis in 100% of patients; TOSV-specific IgM and IgG were detected in patients' sera up to 6 and 20 months after diagnosis, respectively. The avidity index (AI) increased over the first month after infection in 100% of patients and most cases exceeded 60% by Day 30 post infection. The AI subsequently plateaued then declined at 20 months after diagnosis. Finally, neutralization assay to TOSV was performed in 217 sera collected from 41 patients; 69.6% of tested samples resulted in reactive and moderate levels of neutralizing antibodies observed during all phases of infection despite high titres of total anti-TOSV IgG. CONCLUSIONS: Specific antibody response develops rapidly and is long-lasting for neuroinvasive TOSV infection. Serodiagnosis of neuroinvasive TOSV requires simultaneous detection of specific IgM and IgG. Moderate levels of neutralizing antibodies were maintained over the study period, while the protective role of antibodies lacking neutralizing activity is unclear and requires further evaluation.

Increased number of cases of Chikungunya virus (CHIKV) infection imported from the Caribbean and Central America to northern Italy, 2014.

This report describes an increased number of cases of Chikungunya virus (CHIKV) infection imported in northern Italy (Emilia-Romagna region) during the period May-September 2014, indicating that the recent spread of CHIKV and its establishment in the Caribbean and in central America, resulted in a high number of imported cases in Europe, thus representing a threat to public health. From May to September 2014, 14 imported cases of CHIKV infection were diagnosed; the patients were returning to Italy from Dominican Republic (n = 6), Haiti (n = 3), Guadeloupe (n = 2), Martinique (n = 1), Puerto Rico (n = 1) and Venezuela (n = 1). Phylogenetic analysis performed on the envelope protein (E1) gene sequences, obtained from plasma samples from two patients, indicated that the virus strain belongs to the Caribbean clade of the Asian genotype currently circulating in the Caribbean and Americas. The rise in the number of imported cases of CHIKV infection should increase healthcare professionals' awareness of the epidemiological situation and clinical presentation of CHIKV infection in order to enhance surveillance and early diagnosis in the forthcoming season of vector activity in Europe and North America.

Persistence of anti-chikungunya virus-specific antibodies in a cohort of patients followed from the acute phase of infection after the 2007 outbreak in Italy.

Chikungunya is a mosquito-borne infection of humans, and its diffusion has increased worldwide. In 2007 an outbreak occurred in Italy. In this study, the antibody response of 133 patients followed up starting from the acute phase of infection was investigated. Antibody titres were periodically scored up to 1 year since the infection: 82.7% of the IgM antibody disappeared within 12 months, and the IgG response lasted longer than 12 months. Nevertheless, the IgG mean titre was lower in 95.5% of patients at the end of follow-up, thus suggesting a decrease within a relatively short period.

Human and entomological surveillance of Toscana virus in the Emilia-Romagna region, Italy, 2010 to 2012.

Toscana virus (TOSV), transmitted by phlebotomine sandflies, is recognised as one of the most important causes of viral meningitis in summer in Mediterranean countries. A surveillance plan based on both human and entomological surveys was started in 2010 in the Emilia-Romagna region, Italy. Clinical samples from patients with neurological manifestations were collected during 2010 to 2012. The surveillance protocol was improved during these years, allowing the detection of 65 human infections. Most of these infections were recorded in hilly areas, where sandflies reach the highest density. Entomological sampling around the homes of the patients resulted in a low number of captured sandflies, while later sampling in a hilly area with high number of human cases (n=21) resulted in a larger number of captured sandflies. Using this approach, 25,653 sandflies were sampled, of which there were 21,157 females, which were sorted into 287 pools. TOSV RNA was detected by real-time PCR in 33 of the pools. The results highlighted the role of Phlebotomus perfiliewi as the main vector of TOSV and a potential link between vector density and virus circulation. This integrated system shows that an interdisciplinary approach improves the sensitiveness and effectiveness of health surveillance.

The experience of West Nile virus integrated surveillance system in the Emilia-Romagna region: five years of implementation, Italy, 2009 to 2013.

Predicting West Nile virus (WNV) circulation and the risk of WNV epidemics is difficult due to complex interactions of multiple factors involved. Surveillance systems that timely detect virus activity in targeted areas, and allow evidence-based risk assessments may therefore be necessary. Since 2009, a system integrating environmental (mosquitoes and birds) and human surveillance has been implemented and progressively improved in the Emilia-Romagna region, Italy. The objective is to increase knowledge of WNV circulation and to reduce the probability of virus transmission via blood, tissue and organ donation. As of 2013, the system has shown highly satisfactory results in terms of early detection capacity (the environmental surveillance component allowed detection of WNV circulation 3­4 weeks before human cases of West Nile neuroinvasive disease (WNND) occurred), sensitivity (capacity to detect virus circulation even at the enzootic level) and area specificity (capacity to indicate the spatial distribution of the risk for WNND). Strong correlations were observed between the vector index values and the number of human WNND cases registered at the province level. Taking into consideration two scenarios of surveillance, the first with environmental surveillance and the second without, the total costs for the period from 2009 to 2013 were reduced when environmental surveillance was considered (EUR 2.093 million for the first scenario vs EUR 2.560 million for the second). Environmental surveillance helped to reduce costs by enabling a more targeted blood unit testing strategy. The inclusion of environmental surveillance also increased the efficiency of detecting infected blood units and further allowed evidence-based adoption of preventative public health measures.

A model of laboratory surveillance for neuro-arbovirosis applied during 2012 in the Emilia-Romagna region, Italy.

Arboviruses with neuroinvasive potential are gaining more attention due to the increased number of cases of autochthonous and imported infections in the human host. Diagnosis of infection caused by these viruses in patients with central nervous system (CNS) diseases is still underestimated and these infections represent an emerging threat to public health. We describe a model suitable for the laboratory surveillance of neuro-arbovirosis that was applied in the Emilia-Romagna region, north-eastern Italy, during the 2012 summer season. One hundred and twenty cases of suspected neuroinvasive infection were tested for arboviral agents on the basis of clinical and laboratory signs and epidemiological data. The most common virus detected was Toscana virus (TOSV): anti-TOSV specific antibodies or viral components were detected in 28.3% of the cases; 79.4% of the TOSV cases were in the acute phase of infection. No cases resulted in acute phase for West Nile (WNV), Usutu (USUV), Chikungunya (CHIKV) or Dengue (DENV) virus infection. Conversely, two patients with a history of staying in a tick-borne encephalitis virus (TBEV) endemic area showed a probable TBEV infection. These results emphasize the importance of a complete and 'ready to act' laboratory diagnostic system to be implemented within the larger frame of a regional integrated surveillance system.

Detection of specific antibodies against West Nile and Usutu viruses in healthy blood donors in northern Italy, 2010-2011.

Neutralizing antibodies against West Nile (WNV) and Usutu (USUV) viruses were measured in 6000 samples collected, between 1 September 2010 and 30 June 2011, from blood donors living in different districts of Emilia-Romagna, northeastern Italy. On the basis of the microneutralization assay (MNTA), 47 (0.78%) subjects were positive for WNV and 14 (0.23%) for USUV. These results were compared with those obtained 2 years ago and suggest an increased circulation of USUV among humans in Emilia-Romagna.

West Nile virus in Europe: emergence, epidemiology, diagnosis, treatment, and prevention.

West Nile virus (WNV), a mosquito-borne flavivirus in the Japanese encephalitis antigenic group, has caused sporadic outbreaks in humans, horses and birds throughout many of the warmer regions of Europe for at least 20 years. Occasional cases of West Nile encephalitis have also been associated with infected blood transfusions and organ donations. Currently, WNV appears to be expanding its geographical range in Europe and causing increasing numbers of epidemics/outbreaks associated with human morbidity and mortality. This brief review reports on the current epidemic situation regarding WNV in Europe, highlighting the clinical, diagnostic and preventive measures available for controlling this apparently emerging human pathogen.

Do mosquito-associated bacteria of the genus Asaia circulate in humans?

Symbiotic bacteria of the genus Asaia have been proposed as tools for control of mosquito-borne diseases, specifically malaria. However, safety issues are a major concern for paratransgenesis strategies. The aim of this study is to investigate, with immunofluorescence assays and quantitative PCR experiments, whether Asaia spp. is circulating among humans. All human sera and whole blood samples analyzed were negative for Asaia spp., thus suggesting that this organism could be utilized, in the future, as a malaria control tool.

Outbreak of NDM-1-producing Enterobacteriaceae in northern Italy, July to August 2011.

Between July 2011 and August 2011, the New Delhi metallo-beta-lactamase 1 (NDM-1) gene was detected in Klebsiella pneumoniae and Escherichia coli isolates obtained from six patients hospitalised in four healthcare facilities in northern Italy. The patient who had been hospitalised in New Delhi, India, from February to May 2011 and subsequently in the Bologna area, Italy, from May to July 2011, may have been the source of the outbreak. Our findings suggest ongoing spread of this carbapenem-resistance gene in Italy and highlight the need for intensive surveillance.

Imported cases of dengue virus infection: Emilia-Romagna, Italy, 2010.

Dengue is a significant mosquito-borne infection in humans, and its worldwide prevalence is rapidly increasing. In 2010, 83 serum samples from febrile travellers returning from dengue-endemic countries to a region in north-eastern Italy, densely infested with Aedes albopictus, were analysed for dengue virus (DENV). DENV RNA was detected in 20.5% of patients. By RT-PCR, DENV serotypes 1 and 3 were the most common. DENV must be identified early in symptomatic travellers returning from high-risk countries, to prevent outbreaks where potential vectors exist.

Rapid increase of carbapenemase-producing Klebsiella pneumoniae strains in a large Italian hospital: surveillance period 1 March - 30 September 2010.

The first case of carbapenemase-producing Enterobacteriaceae in Italy was reported in 2009. We performed a study over a period of seven months in 2010 to survey the circulation of Klebsiella pneumoniae carbapenemases (KPC) ina 1,500-bed university hospital in northern Italy and report the presence and rapid increase of these multidrug-resistant bacteria. The results raise a major concern about these pathogens and demonstrate the urgent need for infection control and antibiotic stewardship programmes.

West Nile virus circulation in Emilia-Romagna, Italy: the integrated surveillance system 2009.

Following a large West Nile virus (WNV) epidemic in northeastern Italy in 2008, human and animal surveillance activities were implemented in Emilia Romagna. Human surveillance was performed by serology or genome detection on blood and cerebrospinal fluid for all suspected cases suffering from acute meningoencephalitis in the regional territory. Animal surveillance consisted of passive and active surveillance of horses and active surveillance of wild birds and mosquitoes. Between 15 June and 31 October 2009, nine of 78 possible cases of West Nile neuroinvasive disease were confirmed (three fatal). From May to October, 26 cases of neurological West Nile disease were confirmed among 46 horses. The overall incidence of seroconversion among horses in 2009 was 13%. In 2009, 44 of 1,218 wild birds yielded positive PCR results for WNV infection. The planned veterinary and entomological surveillance actions detected WNV activity from the end of July 2009, about 2-3 weeks before the onset of the first human neurological case. Passive surveillance of horses seems to be an early and suitable tool for the detection of WNV activity, but it will be less sensitive in the future, because an intensive programme of horse vaccination started in June 2009.