The slowdown of new infections by human retroviruses has reached a plateau in Spain.

The 2022 annual meeting of the HTLV & HIV-2 Spanish Network was held in Madrid on December 14. We summarize here the main information presented and discussed at the workshop and review time trends for human retroviral infections in Spain. As transmissible agents, infections by human retroviruses are of obligatory declaration. Until the end of 2022, the Spanish national registry had recorded 451 cases of HTLV-1, 821 of HTLV-2, and 416 of HIV-2. For HIV-1, estimates are of 150 000 people currently living with HIV-1 and 60 000 cumulative deaths due to AIDS. During year 2022, new diagnoses in Spain were of 22 for HTLV-1, 6 for HTLV-2, and 7 for HIV-2. The last updated figures for HIV-1 are from 2021 and counted 2786 new diagnoses. The slowdown in yearly infections for HIV-1 in Spain points out that new strategies are needed to achieve the United Nations 95-95-95 targets by 2025. For the remaining neglected human retroviral infections, their control might be pushed throughout four interventions: (1) expanding testing; (2) improving education and interventions aimed to reduce risk behaviors; (3) facilitating access to antiretrovirals as treatment and prevention, including further development of long-acting formulations; and (4) increasing vaccine research efforts. Spain is a 47 million population country in South Europe with strong migration flows from HTLV-1 endemic regions in Latin America and Sub-Saharan Africa. At this time universal HTLV screening has been implemented only in the transplantation setting, following the report of 5 cases of HTLV-associated myelopathy shortly after transplantation of organs from HTLV-1 positive donors. There are four target populations for expanding testing and unveiling asymptomatic carriers responsible for silent HTLV-1 transmissions: (1) migrants; (2) individuals with sexually transmitted infections; (3) pregnant women; and (4) blood donors.

Susceptibility to hepatitis B virus infection in adults living in Spain.

BACKGROUND: A protective hepatitis B virus (HBV) vaccine has been available for four decades. Universal HBV vaccination of infants is recommended by the WHO since the 1990s. Furthermore, HBV immunization is advised for all adults with high-risk behaviours and no seroprotection. However, HBV vaccine coverage remains globally suboptimal. The advent of new more efficacious trivalent HBV vaccines has renewed the interest in HBV vaccination. At present, the extent of current HBV susceptibility in adults remains unknown in Spain. METHODS: HBV serological markers were assessed on a large and representative sample of adults in Spain, including blood donors and individuals belonging to high-risk groups. Serum HBsAg, anti-HBc and anti-HBs were tested in specimens collected during the last couple of years. RESULTS: From 13 859 consecutive adults tested at seven cities across the Spanish geography, overall 166 (1.2%) had positive HBsAg. Past HBV infection was recognized in 14% and prior vaccine immunization in 24%. Unexpectedly, 37% of blood donors and 63% of persons belonging to high-risk groups had no serum HBV markers and therefore were potentially HBV susceptible. CONCLUSION: Roughly 60% of adults living in Spain seem to be HBV susceptible. Waning immunity might be more common than expected. Hence, HBV serological testing should be performed at least once in all adults regardless of risk exposures. HBV vaccine full courses or boosters should be administered to all adults lacking serological evidence of HBV protection.

Viral Culture Confirmed SARS-CoV-2 Subgenomic RNA Value as a Good Surrogate Marker of Infectivity.

Determining SARS-CoV-2 viral infectivity is crucial for patient clinical assessment and isolation decisions. We assessed subgenomic RNA (sgRNA) as a surrogate marker of SARS-CoV-2 infectivity in SARS-CoV-2-positive reverse transcription PCR (RT-PCR) respiratory samples (n = 105) in comparison with viral culture as the reference standard for virus replication. sgRNA and viral isolation results were concordant in 99/105 cases (94%), indicating highly significant agreement between the two techniques (Cohen's kappa coefficient 0.88, 95% confidence interval [CI] 0.78 to 0.97, P < 0.001). sgRNA RT-PCR showed a sensitivity of 97% and a positive predictive value of 94% to detect replication-competent virus, further supporting sgRNA as a surrogate marker of SARS-CoV-2 infectivity. sgRNA RT-PCR is an accurate, rapid, and affordable technique that can overcome culture and cycle threshold (CT) value limitations and be routinely implemented in hospital laboratories to detect viral infectivity, which is essential for optimizing patient monitoring, the efficacy of treatments/vaccines, and work reincorporation policies, as well as for safely shortening isolation precautions.

Whole-genome analysis to describe a human adenovirus D8 conjunctivitis outbreak in a tertiary hospital.

Conjunctivitis is a frequent ocular disorder caused by human adenoviruses (HAdVs). Only a few of the 45 HAdV-D species are associated with epidemic keratoconjunctivitis, including HAdV-D8. Nosocomial outbreaks due to HAdV-D8 have been rarely described, because keratoconjunctivitis cases are clinically diagnosed and treated without having to characterize the causative agent. Moreover, molecular typing is tedious when using classical techniques. In this study, a hospital outbreak of conjunctivitis caused by HAdV-D8 was characterized using the recently developed whole-genome sequencing (WGS) method. Of the 363 patients attending the Ophthalmology Department between July 13 and August 13, 2018, 36 may have acquired intrahospital conjunctivitis. Also, 11 of 22 samples sent to the Virology section were selected for WGS analysis. The WGS results revealed that 10 out of 11 HAdV-D8 strains were closely related. The remaining strain (Case 28) was more similar to a strain from an outbreak in Germany obtained from a public sequence database. WGS results showed that outbreak HAdV-D8 strains had a minimum percentage of identity of 94.3%. WGS is useful in a clinical setting, because it avoids carrying out viral culture or specific polymerase chain reaction sequencing. The public availability of sequence reads makes it easier to compare clusters in circulation. In conclusion, WGS can play an important role in standard routines to describe viral outbreaks.

Comparison of Commensal and Clinical Isolates for Diversity of Plasmids in Escherichia coli and Klebsiella pneumoniae.

In this study, the plasmid content of clinical and commensal strains was analyzed and compared. The replicon profile was similar in both populations, except for L, M, A/C, and N (detected only in clinical strains) and HI1 (only in commensal strains). Although I1 and F were the most frequent replicons, only IncI1, sequence type 12 (ST12) was associated with blaCMY-2 in both populations. In contrast, the widespread resistant IncF plasmids were not linked to a single epidemic plasmid.

Faecal phageome of healthy individuals: presence of antibiotic resistance genes and variations caused by ciprofloxacin treatment.

OBJECTIVES: Antimicrobial resistance genes (ARGs) can be transferred by means of mobile genetic elements, which play a critical role in the dissemination of resistance in the bacterial community. ARG transmission within mobile genetic elements has been reported in plasmids and transposons but less frequently in bacteriophages. Here, the bacteriophage fraction of seven human faecal samples was purified and deep-sequenced to detect the presence of ARGs in the phage particles. METHODS: Seven faecal samples (five from healthy individuals and two from a patient before and after receiving ciprofloxacin treatment) were used to extract phage DNA, which was purified and then sequenced in a MiSeq (Illumina). Generated reads were checked for quality and assembled, and then the generated contigs analysed with Kraken, PHASTER, VirSorter and Prokka. Some genes were also validated by quantitative PCR. RESULTS: Analysis of the purified phage DNA by Kraken identified from 4 to 266 viruses in the samples. The viral fraction corresponded mainly to the order Caudovirales, including phages from the Siphoviridae and Myoviridae families. Bacterial genes associated with antimicrobial resistance were detected in the viral DNA, as confirmed by quantitative PCR. Higher densities of ARG-carrying phage particles were observed in the post- versus pre-ciprofloxacin treatment sample. CONCLUSIONS: The finding of ARGs in phage particles supports the description of phages as mobile elements contributing to the dissemination of bacterial antibiotic resistance and suggests ciprofloxacin treatment may play a role in the release of ARG-carrying particles, thereby increasing resistance.

Prevalence of quinolone resistance mechanisms in Enterobacteriaceae producing acquired AmpC ß-lactamases and/or carbapenemases in Spain. / Prevalencia en España de mecanismos de resistencia a quinolonas en enterobacterias productoras de betalactamasas de clase C adquiridas y/o carbapenemasas.

BACKGROUND: Quinolone resistance in Enterobacteriaceae species has increased over the past few years, and is significantly associated to beta-lactam resistance. The aim of this study was to evaluate the prevalence of chromosomal- and plasmid-mediated quinolone resistance in acquired AmpC ß-lactamase and/or carbapenemase-producing Enterobacteriaceae isolates. METHODS: The presence of chromosomal- and plasmid-mediated quinolone resistance mechanisms [mutations in the quinolone resistance determining region (QRDR) of gyrA and parC and qnr, aac(6')-Ib-cr and qepA genes] was evaluated in 289 isolates of acquired AmpC ß-lactamase- and/or carbapenemase-producing Enterobacteriaceae collected between February and July 2009 in 35 Spanish hospitals. RESULTS: Plasmid mediated quinolone resistance (PMQR) genes were detected in 92 isolates (31.8%), qnr genes were detected in 83 isolates (28.7%), and the aac(6')-Ib-cr gene was detected in 20 isolates (7%). qnrB4 gene was the most prevalent qnr gene detected (20%), associated, in most cases, with DHA-1. Only 14.6% of isolates showed no mutations in gyrA or parC with a ciprofloxacin MIC of 0.5mg/L or higher, whereas PMQR genes were detected in 90% of such isolates. CONCLUSION: qnrB4 gene was the most prevalent PMQR gene detected, and was significantly associated with acquired AmpC ß-lactamase DHA-1. PMQR determinants in association with other chromosomal-mediated quinolone resistance mechanisms, different to mutations in gyrA and parC (increased energy-dependent efflux, altered lipopolysaccharide or porin loss), could lead to ciprofloxacin MIC values that exceed breakpoints established by the main international committees to define clinical antimicrobial susceptibility breakpoints.

Prospective multicenter study of carbapenemase-producing Enterobacteriaceae from 83 hospitals in Spain reveals high in vitro susceptibility to colistin and meropenem.

The aim of this study was to determine the impact of carbapenemase-producing Enterobacteriaceae (CPE) in Spain in 2013 by describing the prevalence, dissemination, and geographic distribution of CPE clones, and their population structure and antibiotic susceptibility. From February 2013 to May 2013, 83 hospitals (about 40,000 hospital beds) prospectively collected nonduplicate Enterobacteriaceae using the screening cutoff recommended by EUCAST. Carbapenemase characterization was performed by phenotypic methods and confirmed by PCR and sequencing. Multilocus sequencing types (MLST) were determined for Klebsiella pneumoniae and Escherichia coli. A total of 702 Enterobacteriaceae isolates met the inclusion criteria; 379 (54%) were CPE. OXA-48 (71.5%) and VIM-1 (25.3%) were the most frequent carbapenemases, and K. pneumoniae (74.4%), Enterobacter cloacae (10.3%), and E. coli (8.4%) were the species most affected. Susceptibility to colistin, amikacin, and meropenem was 95.5%, 81.3%, and 74.7%, respectively. The most prevalent sequence types (STs) were ST11 and ST405 for K. pneumoniae and ST131 for E. coli. Forty-five (54.1%) of the hospitals had at least one CPE case. For K. pneumoniae, ST11/OXA-48, ST15/OXA-48, ST405/OXA-48, and ST11/VIM-1 were detected in two or more Spanish provinces. ST11 isolates carried four carbapenemases (VIM-1, OXA-48, KPC-2, and OXA-245), but ST405 isolates carried OXA-48 only. A wide interregional spread of CPE in Spain was observed, mainly due to a few successful clones of OXA-48-producing K. pneumoniae (e.g., ST11 and ST405). The dissemination of OXA-48-producing E. coli is a new finding of public health concern. According to the susceptibilities determined in vitro, most of the CPE (94.5%) had three or more options for antibiotic treatment.

Epidemiology and risk factors for infections due to AmpC ß-lactamase-producing Escherichia coli.

OBJECTIVES: To describe the prevalence and risk factors for infection due to AmpC ß-lactamase-producing Escherichia coli (AmpC-EC). METHODS: For the prevalence study, all clinical isolates of E. coli with reduced susceptibility to third-generation cephalosporins were prospectively included from June 2010 to November 2011. For risk factor analysis, a case-control study was conducted. Cases were patients with an infection due to AmpC-EC. Controls were patients infected with cephalosporin-susceptible E. coli, matched 1 : 2. Detection of blaAmpC genes was done with a multiplex AmpC-PCR, and hyperproduction of E. coli chromosomal blaAmpC by quantitative RT-PCR. Alteration of the blaAmpC promoter was studied by PCR and sequencing. RESULTS: We identified 243 (1.1%) AmpC-EC strains out of 21 563 clinical isolates. Three cases with strains carrying ESBLs, 18 strains that were considered due to colonization and 8 cases lost to clinical follow-up were excluded. Finally, 214 cases were included in the analysis. Ninety-one cases (42.5%) and 269 (62.8%) controls were strictly community acquired (P < 0.001). Thirty-five (16.3%) cases and 186 controls (43.5%) did not have any identifiable risk factor (P < 0.001). Among cases, 158 (73.8%) were found to harbour an acquired AmpC (73.4% CMY-2). Previous use of fluoroquinolones [OR 2.6 (95% CI 1.12-3.36); P = 0.008] was independently associated with AmpC-EC in the multivariate analysis. CONCLUSIONS: Prevalence of AmpC in E. coli remains low in our area. Plasmid acquisition (CMY type) represents the main mechanism of AmpC production. A high proportion of community-acquired isolates and patients with no identifiable risk factors were found. Previous use of fluoroquinolones was identified as a risk factor.

Antibiotic resistance genes in the bacteriophage DNA fraction of human fecal samples.

A group of antibiotic resistance genes (ARGs) (blaTEM, blaCTX-M-1, mecA, armA, qnrA, and qnrS) were analyzed by real-time quantitative PCR (qPCR) in bacteriophage DNA isolated from feces from 80 healthy humans. Seventy-seven percent of the samples were positive in phage DNA for one or more ARGs. blaTEM, qnrA, and, blaCTX-M-1 were the most abundant, and armA, qnrS, and mecA were less prevalent. Free bacteriophages carrying ARGs may contribute to the mobilization of ARGs in intra- and extraintestinal environments.

Genetic and kinetic characterization of the novel AmpC ß-lactamases DHA-6 and DHA-7.

During a Spanish surveillance study, two natural variants of DHA ß-lactamases, DHA-6 and DHA-7, were found, with the replacements Ala226Thr and Phe322Ser, respectively, with respect to DHA-1. The DHA-6 and DHA-7 enzymes were isolated from Escherichia coli and Enterobacter cloacae clinical isolates, respectively. The aim of this study was to genetically, microbiologically, and biochemically characterize the DHA-6 and DHA-7 ß-lactamases. The blaDHA-6 and blaDHA-7 genes were located in the I1 and HI2 incompatibility group plasmids of 87.3 and 310.4 kb, respectively. The genetic contexts of blaDHA-6 and blaDHA-7 were similar to that already described for the blaDHA-1 gene and included the qnrB4 and aadA genes. The MICs for cephalothin, aztreonam, cefotaxime, and ceftazidime were 8- to 32-fold lower for DHA-6 than for DHA-1 or DHA-7 expressed in the same isogenic E. coli TG1 strain. Interestingly, the MIC for cefoxitin was higher in the DHA-6-expressing transformant than in DHA-1 or DHA-7. Biochemical studies with pure ß-lactamases revealed slightly lower catalytic efficiencies of DHA-6 against cephalothin, ceftazidime, and cefotaxime than those of DHA-1 and DHA-7. To understand this behavior, stability experiments were carried out and showed that the DHA-6 protein displayed significantly higher stability than the DHA-1 and DHA-7 enzymes. The proximity of Thr226 to the N terminus in the tertiary protein structure in DHA-6 may promote this stabilization and, consequently, may induce a slight reduction in the dynamic of this enzyme that primarily affects the hydrolysis of some of the bulkiest antibiotics.

Evolution of carbapenemase-producing Enterobacteriaceae at the global and national level: what should be expected in the future?

In recent years, Enterobacteriaceae isolates have increased their potential to become highly drug resistant by acquiring resistance to carbapenems, primarily due to the production of acquired carbapenemases. The carbapenemases detected in Enterobacteriaceae are largely of the KPC, VIM, NDM, IMP and OXA-48 types. Although the epidemiological origin and geographic distribution of carbapenemases are clearly different, they all first appeared in the late 20th Century. Only a decade later, these enzymes have already become established and have expanded globally. An important epidemiological change has occurred in Spain in recent years, characterized by a rapid increase in the number of cases of carbapenemase-producing Enterobacteriaceae (CPE), causing both nosocomial outbreaks and single infections. The impact of CPE in Spain is primarily due to OXA-48-producing and VIM-1-producing Klebsiella pneumoniae isolates, although other species such as Escherichia coli and Enterobacter cloacae are also increasing. The emergence of CPE as a cause of community-onset infections is a matter of great concern. Taking into account recent experience, and considering the fact that increasing numbers of patients are becoming infected by CPE and reservoirs of carbapenemases are growing globally, the trend of the CPE epidemic points toward a rise in its incidence. To prevent a massive CPE pandemic, a well-coordinated response from all health professionals and national and supranational authorities is clearly needed.

Characterization of the new AmpC ß-lactamase FOX-8 reveals a single mutation, Phe313Leu, located in the R2 loop that affects ceftazidime hydrolysis.

A novel class C ß-lactamase (FOX-8) was isolated from a clinical strain of Escherichia coli. The FOX-8 enzyme possessed a unique substitution (Phe313Leu) compared to FOX-3. Isogenic E. coli strains carrying FOX-8 showed an 8-fold reduction in resistance to ceftazidime relative to FOX-3. In a kinetic analysis, FOX-8 displayed a 33-fold reduction in kcat/Km for ceftazidime compared to FOX-3. In the FOX family of ß-lactamases, the Phe313 residue located in the R2 loop affects ceftazidime hydrolysis and alters the phenotype of E. coli strains carrying this variant.

Shiga toxin 2-encoding bacteriophages in human fecal samples from healthy individuals.

Shiga toxin-converting bacteriophages (Stx phages) carry the stx gene and convert nonpathogenic bacterial strains into Shiga toxin-producing bacteria. Previous studies have shown that high densities of free and infectious Stx phages are found in environments polluted with feces and also in food samples. Taken together, these two findings suggest that Stx phages could be excreted through feces, but this has not been tested to date. In this study, we purified Stx phages from 100 fecal samples from 100 healthy individuals showing no enteric symptoms. The phages retrieved from each sample were then quantified by quantitative PCR (qPCR). In total, 62% of the samples carried Stx phages, with an average value of 2.6 × 10(4) Stx phages/g. This result confirms the excretion of free Stx phages by healthy humans. Moreover, the Stx phages from feces were able to propagate in enrichment cultures of stx-negative Escherichia coli (strains C600 and O157:H7) and in Shigella sonnei, indicating that at least a fraction of the Stx phages present were infective. Plaque blot hybridization revealed lysis by Stx phages from feces. Our results confirm the presence of infectious free Stx phages in feces from healthy persons, possibly explaining the environmental prevalence observed in previous studies. It cannot be ruled out, therefore, that some positive stx results obtained during the molecular diagnosis of Shiga toxin-producing Escherichia coli (STEC)-related diseases using stool samples are due to the presence of Stx phages.

Adult T-cell leukemia/lymphoma in HTLV-1 non-endemic regions.

BACKGROUND: HTLV-1 infection is a neglected disease, despite producing neurological and lymphoproliferative severe illnesses and affect over 10 million people worldwide. Roughly 5% of HTLV-1 carriers develop Adult T-cell leukemia/lymphoma (ATLL), one of the most aggressive hematological malignancies. METHODS: A national HTLV-1 register exists since 1989 in Spain, a non-endemic country with a large migrant flow from Latin America and Equatorial Africa, where HTLV-1 is endemic. The main features of all patients diagnosed with ATLL in Spain up to date are reported. RESULTS: A total of 451 cases of HTLV-1 infection had been reported in Spain until the end of year 2022. ATLL had been diagnosed in 35 (7.8%). The current average incidence of ATLL in Spain is of two cases per year. Women represent 57% of ATLL patients. Mean age at diagnosis was 47 years-old. Roughly 57% were Latin Americans and 26% Africans. At diagnosis, the majority presented with acute or lymphoma clinical forms. Survival was shorter than one year in most of them. Mean HTLV-1 proviral load was significantly greater in ATLL patients than in asymptomatic HTLV-1 carriers (2,305 vs 104 copies/104 PBMC). HTLV-1 subtyping in 6 ATLL patients found the 1a transcontinental variant (n = 4) and the Japanese variant (n = 2). All ATLL patients were negative for HIV-1, did not develop HTLV-1-associated myelopathy and were not transplant recipients. CONCLUSION: The rate of ATLL is very low in Spain and mostly associated to migrants from HTLV-1 endemic regions. Given the poor clinical outcome of ATLL, HTLV-1 testing should be performed at least once in all migrants coming from HTLV-1 endemic countries and in natives who have lived in or had sex partners from such regions.

Characterization of a novel IMP-28 metallo-ß-lactamase from a Spanish Klebsiella oxytoca clinical isolate.

An isolate of Klebsiella oxytoca carrying a novel IMP metallo-ß-lactamase was discovered in Madrid, Spain. The bla(IMP-28) gene is part of a chromosomally located class I integron. The IMP-28 k(cat)/K(m) values for ampicillin, ceftazidime, and cefepime and, to a lesser extent, imipenem and meropenem, are clearly lower than those of IMP-1. The His306Gln mutation may induce important modifications of the L3 loop and thus of substrate accessibility and hydrolysis and be the main reason for this behavior.

Plasmid typing and genetic context of AmpC ß-lactamases in Enterobacteriaceae lacking inducible chromosomal ampC genes: findings from a Spanish hospital 1999-2007.

OBJECTIVES: To gain insights into ampC transmission between bacterial strains. METHODS: We examined the genetic context of 117 acquired ampC genes from 27119 Enterobacteriaceae collected between 1999 and 2007. Plasmid analysis was carried out by PCR-based replicon or relaxase typing, S1-PFGE and Southern hybridization. I-CeuI/PFGE was used for isolates not characterized by plasmid analysis. PCR reactions were used to map the genetic organization of the ampC genes. RESULTS: Among the isolates studied, 81.2% of ampC genes were located on plasmids of known Inc/MOB groups, 7.7% were chromosomally located and 11.1% were not determined. A/C, I1 and K were the most commonly found replicons in plasmids carrying bla(CMY-2), while L/M replicons were associated with bla(DHA-1). bla(ACC-1) was linked to I1 and MOB(F11) plasmids; bla(CMY-27) was associated with IncF and MOB(P12) plasmids; the plasmid carrying bla(CMY-25) could not be typed, and bla(CMY-40) was chromosomally located. All 87 isolates carrying bla(CMY-2), bla(CMY-4), bla(CMY-25), bla(CMY-27), bla(CMY-40) or bla(ACC-1) displayed the transposon-like structures ISEcp1/ΔISEcp1-bla(CMY)-blc-sugE or ΔISEcp1-bla(ACC-1)-gdha. The most prevalent structure in bla(DHA-1) (93.3% of cases) was identical to that described in the Klebsiella pneumoniae pTN60013 plasmid. Remarkably, in three isolates containing chromosomal bla(CMY-2), this gene was mobilized by conjugation. CONCLUSIONS: Although plasmids are the main cause of the rapid dissemination of ampC genes among bacteria, we need to be aware that other mobile genetic elements such as integrative and conjugative elements (ICEs) can be involved in the mobilization of these genes.

Viral culture and immunofluorescence for the detection of SARS-CoV-2 infectivity in RT-PCR positive respiratory samples.

BACKGROUND: Knowing how long SARS-CoV-2-positive individuals can remain infective is crucial for the design of infection prevention and control strategies. Viral culture is the gold standard for detecting an active-replicative virus and evaluating its infectious potential. OBJECTIVE: To assess the correlation of SARS-CoV-2 infectivity with the number of days from symptom onset and the Ct value, using culture as a reference method. Also, to describe a detailed protocol for SARS-CoV-2 culture and immunofluorescence confirmation based on our experience with other respiratory viruses. STUDY DESIGN: 100 consecutive respiratory samples positive for SARS-CoV-2 by RT-PCR from different subjects were inoculated into VERO E6 cells. RESULTS: Viral isolation was successful in 58% of samples. The median number of days from symptom onset for culture-positive samples was 2, and 15 for culture-negative samples. Six positive cultures were obtained in patients ≥14 days after symptom onset, all of whom were immunocompromised or with severe COVID-19. The mean Ct value was 12.64 units higher in culture-negative than in culture-positive samples. The probability of successfully isolating SARS-CoV-2 in samples with a Ct value <22 was 100%, decreasing to 3.1% when >27. CONCLUSIONS: Our findings show a significant positive correlation between the probability of isolating SARS-CoV-2 in culture, fewer days of symptoms and a lower RT-PCR Ct value. SARS-CoV-2 infectivity lasts no more than 14 days from symptom onset in immunocompetent individuals. In contrast, in immunocompromised patients or those with severe COVID-19 infectivity may remain after 14 days. Ct value <22 always indicates infectivity.

A New Variant of the aadE-sat4-aphA-3 Gene Cluster Found in a Conjugative Plasmid from a MDR Campylobacter jejuni Isolate.

Campylobacter jejuni is a foodborne pathogen causing bacterial gastroenteritis, with the highest incidence reported in Europe. The prevalence of antibiotic resistance in C. jejuni, as well as in many other bacterial pathogens, has increased over the last few years. In this report, we describe the presence of a plasmid in a multi-drug-resistant C. jejuni strain isolated from a gastroenteritis patient. Mating experiments demonstrated the transference of this genetic element (pCjH01) among C. jejuni by plasmid conjugation. The pCjH01 plasmid was sequenced and assembled, revealing high similarity (97% identity) with pTet, a described tetracycline resistance encoding plasmid. pCjH01 (47.7 kb) is a mosaic plasmid composed of a pTet backbone that has acquired two discrete DNA regions. Remarkably, one of the acquired sequences carried an undescribed variant of the aadE-sat4-aphA-3 gene cluster, providing resistance to at least kanamycin and gentamycin. Aside from the antibiotic resistance genes, the cluster also carries genes coding for putative regulators, such as a sigma factor of the RNA polymerase and an antisigma factor. Homology searches suggest that Campylobacter exchanges genetic material with distant G-positive bacterial genera.