Pandemic potential of highly pathogenic avian influenza clade 2.3.4.4 A(H5) viruses.

The panzootic caused by A/goose/Guangdong/1/96-lineage highly pathogenic avian influenza (HPAI) A(H5) viruses has occurred in multiple waves since 1996. From 2013 onwards, clade 2.3.4.4 viruses of subtypes A(H5N2), A(H5N6), and A(H5N8) emerged to cause panzootic waves of unprecedented magnitude among avian species accompanied by severe losses to the poultry industry around the world. Clade 2.3.4.4 A(H5) viruses have expanded in distinct geographical and evolutionary pathways likely via long distance migratory bird dispersal onto several continents and by poultry trade among neighboring countries. Coupled with regional circulation, the viruses have evolved further by reassorting with local viruses. As of February 2019, there have been 23 cases of humans infected with clade 2.3.4.4 H5N6 viruses, 16 (70%) of which had fatal outcomes. To date, no HPAI A(H5) virus has caused sustainable human-to-human transmission. However, due to the lack of population immunity in humans and ongoing evolution of the virus, there is a continuing risk that clade 2.3.4.4 A(H5) viruses could cause an influenza pandemic if the ability to transmit efficiently among humans was gained. Therefore, multisectoral collaborations among the animal, environmental, and public health sectors are essential to conduct risk assessments and develop countermeasures to prevent disease and to control spread. In this article, we describe an assessment of the likelihood of clade 2.3.4.4 A(H5) viruses gaining human-to-human transmissibility and impact on human health should such human-to-human transmission occur. This structured analysis assessed properties of the virus, attributes of the human population, and ecology and epidemiology of these viruses in animal hosts.

Characterization of H7N9 influenza A viruses isolated from humans.

Avian influenza A viruses rarely infect humans; however, when human infection and subsequent human-to-human transmission occurs, worldwide outbreaks (pandemics) can result. The recent sporadic infections of humans in China with a previously unrecognized avian influenza A virus of the H7N9 subtype (A(H7N9)) have caused concern owing to the appreciable case fatality rate associated with these infections (more than 25%), potential instances of human-to-human transmission, and the lack of pre-existing immunity among humans to viruses of this subtype. Here we characterize two early human A(H7N9) isolates, A/Anhui/1/2013 (H7N9) and A/Shanghai/1/2013 (H7N9); hereafter referred to as Anhui/1 and Shanghai/1, respectively. In mice, Anhui/1 and Shanghai/1 were more pathogenic than a control avian H7N9 virus (A/duck/Gunma/466/2011 (H7N9); Dk/GM466) and a representative pandemic 2009 H1N1 virus (A/California/4/2009 (H1N1pdm09); CA04). Anhui/1, Shanghai/1 and Dk/GM466 replicated well in the nasal turbinates of ferrets. In nonhuman primates, Anhui/1 and Dk/GM466 replicated efficiently in the upper and lower respiratory tracts, whereas the replicative ability of conventional human influenza viruses is typically restricted to the upper respiratory tract of infected primates. By contrast, Anhui/1 did not replicate well in miniature pigs after intranasal inoculation. Critically, Anhui/1 transmitted through respiratory droplets in one of three pairs of ferrets. Glycan arrays showed that Anhui/1, Shanghai/1 and A/Hangzhou/1/2013 (H7N9) (a third human A(H7N9) virus tested in this assay) bind to human virus-type receptors, a property that may be critical for virus transmissibility in ferrets. Anhui/1 was found to be less sensitive in mice to neuraminidase inhibitors than a pandemic H1N1 2009 virus, although both viruses were equally susceptible to an experimental antiviral polymerase inhibitor. The robust replicative ability in mice, ferrets and nonhuman primates and the limited transmissibility in ferrets of Anhui/1 suggest that A(H7N9) viruses have pandemic potential.

Persistent Pandemic Lineages of Uropathogenic Escherichia coli in a College Community from 1999 to 2017.

The incidence of drug-resistant community-acquired urinary tract infections (CA-UTI) continues to increase worldwide. In 1999 to 2000, a single lineage of uropathogenic Escherichia coli (UPEC) sequence type 69 (ST69) caused 51% of trimethoprim-sulfamethoxazole-resistant UTI in a Northern California university community. We compared the clonal distributions of UPEC and its impact on antimicrobial resistance prevalence in the same community during two periods separated by 17 years. We analyzed E. coli isolates from urine samples from patients with symptoms of UTI who visited a health service between September 2016 and May 2017 and compared them to UPEC isolates collected similarly between October 1999 and March 2000. Isolates were tested for antimicrobial drug susceptibility and genotyped by multilocus sequence typing. In 1999 to 2000, strains belonging to ST95, ST127, ST73, ST69, ST131, and ST10 caused 125 (56%) of 225 UTI cases, while the same STs caused 148 (64%) of 233 UTI cases in 2016 to 2017. The frequencies of ampicillin resistance and ciprofloxacin resistance rose from 24.4% to 41.6% (P < 0.001) and from 0.9% to 5.1% (P < 0.003), respectively. The six STs accounted for 78.6% and 72.7% of these increases, respectively. Prevalence of drug-resistant UTI in this community appears to be largely influenced by a small set of dominant UPEC STs circulating in the same community 17 years apart. Further research to determine the origin and reasons for persistence of these dominant genotypes is necessary to combat antimicrobial-resistant CA-UTI.

An Enzyme-Mediated Amplification Strategy Enables Detection of ß-Lactamase Activity Directly in Unprocessed Clinical Samples for Phenotypic Detection of ß-Lactam Resistance.

Biochemical assays that can identify ß-lactamase activity directly from patient samples have the potential to significantly improve the treatment of bacterial infections. However, current ß-lactamase probes do not have the sensitivity needed to measure ß-lactam resistance directly from patient samples. Here, we report the development of an instrument-free signal amplification technology, DETECT, that connects the activity of two enzymes in series to effectively amplify the activity of ß-lactamase 40 000-fold, compared to the standard ß-lactamase probe nitrocefin.

Emergency Department Urinary Tract Infections Caused by Extended-Spectrum ß-Lactamase-Producing Enterobacteriaceae: Many Patients Have No Identifiable Risk Factor and Discordant Empiric Therapy Is Common.

STUDY OBJECTIVE: Community-onset urinary tract infections (UTIs) caused by extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae, which are resistant to ceftriaxone and usually coresistant to fluoroquinolones, are increasing worldwide. We investigate and describe in detail UTIs caused by ESBL-producing Enterobacteriaceae in our emergency department (ED), and determine the proportion that occurred in patients without health care-associated risk factors and who received discordant initial antibiotic therapy. METHODS: At an urban public hospital in Northern California, microbiology staff prospectively reviewed ED urine culture results weekly for 1 year and presumptively identified ESBL-producing isolates by ceftriaxone plus ceftazidime resistance. For isolates associated with a clinical UTI, patient demographic and case clinical features were abstracted retrospectively. Health care-associated infections were defined by standard risk factors plus aged 65 years or older, bladder catheter, urologic procedure, functional dependence, or antibiotics in the previous 90 days. Community-associated infections were defined by absence of these. A subset of community-associated ESBL-producing Escherichia coli isolates underwent genotyping. Electronic health record query was used to determine the denominator of ED UTI patients who underwent urine culture during the study period. RESULTS: Between August 2016 and July 2017, there were 1,045 unique ED patients diagnosed with a UTI, whose specimens underwent culture. There were 62 ESBL-producing isolates (5.9%; 95% confidence interval [CI] 4.6% to 7.5%). Selected characteristics of the entire ESBL UTI cohort were median age 50 years, 37 (60%) patients were women, 28 (44%) Hispanic, 11 (18%) had been hospitalized in the previous 3 months, 19 (31%) had pyelonephritis, 49 (79%) of isolates were E coli, 44 (71%) were levofloxacin-resistant, and 24 (23%) nitrofurantoin-resistant. Initial antibiotic choice was discordant with isolate susceptibility in 26 of 56 cases (46%; 95% CI 33% to 60%), and the initial oral antibiotic prescred was discordant in 19 of 41 cases (46%; 95% CI 31% to 63%). Twenty-seven infections (44%; 95% CI 31% to 57%) were categorized as community-associated. Eight patients with community-associated infection were women younger than 50 years, with no comorbidities and no more than 1 UTI in the previous year. Of 12 community-associated E coli isolates tested, all were confirmed to harbor ESBL genes; the CTX-M1 ß-lactamase gene was found in 8 (67%); 4 belong to genotype ST131. CONCLUSION: At this single Northern California ED, greater than 5% of culture-proven UTI were caused by ESBL-producing Enterobacteriaceae, and in nearly half of cases there was no identifiable health care-associated risk factor. Levofloxacin co-resistance and discordant antibiotic therapy were common.

Mammalian adaptive mutations of the PA protein of highly pathogenic avian H5N1 influenza virus.

UNLABELLED: Highly pathogenic H5N1 influenza A viruses continue to circulate among avian species and cause sporadic cases of human infection. Therefore, the threat of a pandemic persists. However, the human cases of H5N1 infection have been limited mainly to individuals in close contact with infected poultry. These findings suggest that the H5N1 viruses need to acquire adaptive mutations to gain a replicative advantage in mammalian cells to break through the species barrier. Many amino acid mutations of the polymerase complex have been reported to enhance H5N1 virus growth in mammalian cells; however, the mechanism for H5N1 virus of adaptation to humans remains unclear. Here, we propose that the PA of an H5N1 influenza virus isolated from a human in Vietnam (A/Vietnam/UT36285/2010 [36285]) increased the ability of an avian H5N1 virus (A/chicken/Vietnam/TY31/2005 [Ck/TY31]) to grow in human lung epithelial A549 cells. The five PA amino acid substitutions V44I, V127A, C241Y, A343T, and I573V, which are rare in H5N1 viruses from human and avian sources, enhanced the growth capability of this virus in A549 cells. Moreover, these mutations increased the pathogenicity of the virus in mice, suggesting that they contribute to adaptation to mammalian hosts. Intriguingly, PA-241Y, which 36285 encodes, is conserved in more than 90% of human seasonal H1N1 viruses, suggesting that PA-241Y contributes to virus adaptation to human lung cells and mammalian hosts. IMPORTANCE: Many amino acid substitutions in highly pathogenic H5N1 avian influenza viruses have been shown to contribute to adaptation to mammalian hosts. However, no naturally isolated H5N1 virus has caused extensive human-to-human transmission, suggesting that additional, as-yet unidentified amino acid mutations are needed for adaptation to humans. Here, we report that five amino acid substitutions in PA (V44I, V127A, C241Y, A343T, and I573V) contribute to the replicative efficiency of H5N1 viruses in human lung cells and to high virulence in mice. These results are helpful for assessing the pandemic risk of isolates and further our understanding of the mechanism of H5N1 virus adaptation to mammalian hosts.

Identification of PB2 mutations responsible for the efficient replication of H5N1 influenza viruses in human lung epithelial cells.

UNLABELLED: Highly pathogenic H5N1 avian influenza viruses have caused outbreaks among poultry worldwide, resulting in sporadic infections in humans with approximately 60% mortality. However, efficient transmission of H5N1 viruses among humans has yet to occur, suggesting that further adaptation of H5N1 viruses to humans is required for their efficient transmission among humans. The viral determinants for efficient replication in humans are currently poorly understood. Here, we report that the polymerase PB2 protein of an H5N1 influenza virus isolated from a human in Vietnam (A/Vietnam/UT36285/2010, virus 36285) increased the growth ability of an avian H5N1 virus (A/wild bird/Anhui/82/2005, virus Wb/AH82) in human lung epithelial A549 cells (however, the reassortant virus did not replicate more efficiently than human 36285 virus). Furthermore, we demonstrate that the amino acid residues at positions 249, 309, and 339 of the PB2 protein from this human isolate were responsible for its efficient replication in A549 cells. PB2 residues 249G and 339M, which are found in the human H5N1 virus, are rare in H5N1 viruses from both human and avian sources. Interestingly, PB2-249G is found in over 30% of human seasonal H3N2 viruses, which suggests that H5N1 viruses may replicate well in human cells when they acquire this mutation. Our data are of value to H5N1 virus surveillance. IMPORTANCE: Highly pathogenic H5N1 avian influenza viruses must acquire mutations to overcome the species barrier between avian species and humans. When H5N1 viruses replicate in human respiratory cells, they can acquire amino acid mutations that allow them to adapt to humans through continuous selective pressure. Several amino acid mutations have been shown to be advantageous for virus adaptation to mammalian hosts. Here, we found that amino acid changes at positions 249, 309, and 339 of PB2 contribute to efficient replication of avian H5N1 viruses in human lung cells. These findings are beneficial for evaluating the pandemic risk of circulating avian viruses and for further functional analysis of PB2.

Genetic features of antimicrobial drug-susceptible extraintestinal pathogenic Escherichia coli pandemic sequence type 95.

IMPORTANCE: Despite the increasing prevalence of antibiotic-resistant Escherichia coli strains that cause urinary tract and bloodstream infections, a major pandemic lineage of extraintestinal pathogenic E. coli (ExPEC) ST95 has a comparatively low frequency of drug resistance. We compared the genomes of 1,749 ST95 isolates to identify genetic features that may explain why most strains of ST95 resist becoming drug-resistant. Identification of such genomic features could contribute to the development of novel strategies to prevent the spread of antibiotic-resistant genes and devise new measures to control antibiotic-resistant infections.

A Population-Based Surveillance Study of Shared Genotypes of Escherichia coli Isolates from Retail Meat and Suspected Cases of Urinary Tract Infections.

There is increasing evidence that retail food may serve as a source of Escherichia coli that causes community-acquired urinary tract infections, but the impact of this source in a community is not known. We conducted a prospective, population-based study in one community to examine the frequency of recovery of uropathogenic E. coli genotypes from retail meat samples. We analyzed E. coli isolates from consecutively collected urine samples of patients suspected to have urinary tract infections (UTIs) at a university-affiliated health service and retail meat samples from the same geographic region. We genotyped all E. coli isolates by multilocus sequence typing (MLST) and tested them for antimicrobial susceptibility. From 2016 to 2017, we cultured 233 E. coli isolates from 230 (21%) of 1,087 urine samples and 177 E. coli isolates from 120 (28%) of 427 retail meat samples. Urine samples contained 61 sequence types (STs), and meat samples had 95 STs; 12 STs (ST10, ST38, ST69, ST80, ST88, ST101, ST117, ST131, ST569, ST906, ST1844, and ST2562) were common to both. Thirty-five (81%) of 43 meat isolates among the 12 STs were from poultry. Among 94 isolates in the 12 STs, 26 (60%) of 43 retail meat isolates and 15 (29%) of 51 human isolates were pan-susceptible (P < 0.005). We found that 21% of E. coli isolates from suspected cases of UTIs belonged to STs found in poultry. Poultry may serve as a possible reservoir of uropathogenic E. coli (UPEC). Additional studies are needed to demonstrate transmission pathways of these UPEC genotypes and their food sources.IMPORTANCE Community-acquired urinary tract infection caused by Escherichia coli is one of the most common infectious diseases in the United States, affecting approximately seven million women and costing approximately 11.6 billion dollars annually. In addition, antibiotic resistance among E. coli bacteria causing urinary tract infection continues to increase, which greatly complicates treatment. Identifying sources of uropathogenic E. coli and implementing prevention measures are essential. However, the reservoirs of uropathogenic E. coli have not been well defined. This study demonstrated that poultry sold in retail stores may serve as one possible source of uropathogenic E. coli This finding adds to a growing body of evidence that suggests that urinary tract infection may be a food-borne disease. More research in this area can lead to the development of preventive strategies to control this common and costly infectious disease.