Occurrence Of Mixed Infections Research Articles

The plasma viral communities associate with clinical profiles in a large-scale haematological patients cohort

BackgroundHaematological patients exhibit immune system abnormalities that make them susceptible to viral infections. Understanding the relationship between the virome in the blood plasma of haematological patients and their clinical characteristic is crucial for disease management. We aimed to explore the presence of viral pathogens and identify close associations between viral infections and various clinical features.ResultsA total of 21 DNA viruses and 6 RNA viruses from 12 virus families were identified from 1383 patients. Patients with haematological diseases exhibited significantly higher diversity, prevalence, and co-detection rates of viral pathogens. During fever episodes, pathogen detection was notably higher, with Epstein-Barr virus (EBV) and Mucorales infections being the most probable culprits for fever symptoms in non-haematological patients. The detection rate of torque teno virus (TTV) significantly increases in haematological patients after transplantation and during primary lung infections. Additionally, TTV-positive patients demonstrate significantly higher absolute neutrophil counts, while C-reactive protein and procalcitonin levels are notably lower. Furthermore, TTV, cytomegalovirus, and parvovirus B19 (B19V) were found to be more prevalent in non-neutropenic patients, while non-viral pathogenic infections, such as Gram-negative bacteria and Mucorales, were more common in neutropenic patients. Pegivirus C (HPgV-C) infection often occurred post-transplantation, regardless of neutropenia. Additionally, some viruses such as TTV, B19V, EBV, and HPgV-C showed preferences for age and seasonal infections.ConclusionsAnalysis of the plasma virome revealed the susceptibility of haematological patients to plasma viral infections at specific disease stages, along with the occurrence of mixed infections with non-viral pathogens. Close associations were observed between the plasma virome and various clinical characteristics, as well as clinical detection parameters. Understanding plasma virome aids in auxiliary clinical diagnosis and treatment, enabling early prevention to reduce infection rates in patients and improve their quality of life.EnXnzQJ5tBc4iYM41USpCeVideo

Molecular species determination of cyathostomins from horses in Ireland

Cyathostomins are globally important equine parasites, responsible for both chronic and acute pathogenic effects. The occurrence of mixed infections with numerous cyathostomin species hinders our understanding of parasite epidemiology, host-parasite dynamics, and species pathogenicity. There have been few studies of cyathostomin species occurring in horses in Ireland, where temperate climatic conditions with year-round rainfall provide suitable conditions for infection of grazing animals with bursate nematodes. Here, we amplified and sequenced the ITS-2 region of adult worms harvested at post-mortem from eleven adult horses between August 2018 and June 2020, and recorded species prevalence and abundance of worms recovered from the caecum, right ventral colon and left dorsal colon, using both BLAST and IDTAXA for taxonomic attribution. Phylogenetic relationships and community composition were also recorded and compared with other relevant studies, including a global meta-analysis. Overall, our results agree with previous studies that there does not seem to be a major difference in cyathostomin species occurrence in equids in different geographical regions. We confirmed the results of other workers in relation to the difficulties in discriminating between Cylicostephanus calicatus and Coronocyclus coronatus on the basis of ITS-2 sequences.

Vaginitis Polymerase Chain Reaction (PCR) Testing and Occurrence of Mixed Infections: Results from a Large US Clinical Laboratory

Vaginitis Polymerase Chain Reaction (PCR) Testing and Occurrence of Mixed Infections: Results from a Large US Clinical Laboratory

Quantificação de duas estirpes do Potato vírus Y em infecções simples e mistas por RT-qPCR realça o seu panorama epidemiológico no Brasil

ABSTRACT Potato virus Y (PVY) is recognized as one of the most common and destructive pathogens seriously affecting potato producing areas worldwide. More recently PVYNTN and PVYN:O/N-Wi, have emerged as the main strains present in the PVY infected plants detected in Brazilian potato fields. In this study, samples of potato collected in south part of Minas Gerais - Brazil were first tested by DAS-ELISA and then by RT-PCR multiplex in order to discriminate the PVY strains. Afterward, part of them was tested by RT-qPCR to confirm and quantify the viruses in infected tissues. The sensitivity of the techniques for detecting PVY isolates present in the sampled locations was investigated, as well as the occurrence of mixed infections, aiming to understand the general epidemiological picture of this pathogen in potato producing fields. In the multiplex RT-PCR test, the samples with O and N serotypes were identified as infected with PVYNTN and PVYN:O/N-Wi strains. When tested by RT-PCR for amplification of PVYE, 41 samples (67,2%) were positive, having a characteristic electrophoretic profile for this recombinant strain, and 9 isolates were also observed with atypical patterns for recombinant PVYE. The best technique to detect mixed infection was RT-qPCR, with the concentration of PVYNTN being much higher than that of PVYN:O/N-Wi. These results show the importance of using the most suitable method for the diagnosis and surveying of PVY strains in crop fields and reveal, for the first time, the dissemination of PVYE recombinants to several Brazilian potato fields.

Epidemiological Manifestation of Combined Natural Foci of Tularemia, Leptospirosis and Hemorrhagic Fever with Renal Syndrome: Mixed Infections

Epidemiological Manifestation of Combined Natural Foci of Tularemia, Leptospirosis and Hemorrhagic Fever with Renal Syndrome: Mixed Infections

Enhanced detection of Giardia duodenalis mixed assemblage infections in pre-weaned dairy calves using next generation sequencing

Giardia duodenalis is one of the most common parasitic causes of gastrointestinal illness in humans worldwide with widespread infections in mammalian hosts. It frequently infects cattle, producing a high number of cysts. Cattle can harbor both host-adapted assemblage E and human pathogenic assemblages A and B. Previous studies have demonstrated that conventional molecular methods lack the sensitivity required for detecting mixed infections and that the occurrence of mixed infections in cattle are likely underestimated. To investigate the presence of mixed assemblage infections in cattle, 2539 pre-weaned dairy calves from the United Stated were screened for the presence of G. duodenalis using molecular tools. Next generation amplicon sequencing (NGS) was then performed for a subset of around 30% of positive samples (n = 314) and the ability of NGS and Sanger sequencing to detect mixed assemblage infections was compared. The overall prevalence of G. duodenalis in pre-weaned dairy calves in the sample using PCR was high (1013/2539; 39.9%). Molecular genotyping identified only assemblage A and E, with assemblage E as the predominant assemblage. Out of the 314 samples examined by both Sanger and NGS, 9 samples (2.9%) were identified as mixed A/E infections by Sanger while NGS identified 56 samples (17.8%), which was six-times more mixed infections compared with Sanger sequencing. NGS demonstrated superior sensitivity to Sanger in detecting assemblages present in low abundances. The percentage of mixed A/E infections found in the sampled dairy calves was higher than was hypothesized using values from the literature. This underestimation could be present in the wider cattle population as well, though further exploration would be needed to verify that claim. These findings highlight the advantages of NGS application in molecular epidemiological studies of Giardia. To better understand Giardia epidemiology, establish routes of transmission, and assess the potential role of cattle and other animals as a source of environmental contamination with zoonotic assemblages it is necessary to uncover mixed assemblage infections.

Remarkable diversity, new records and Leishmania detection in the sand fly fauna of an area of high endemicity for cutaneous leishmaniasis in Acre state, Brazilian Amazonian Forest

The species richness of Amazonian phlebotomines is considered to be one of the highest in the world. In the present study, we investigated the richness and diversity of phlebotomine fauna in Xapuri city, Acre state, Western Brazilian Amazonia, which is an area that is highly endemic for cutaneous leishmaniasis. Sand fly collections were performed monthly from August 2013 to July 2015 (288 h total of sampling effort) in intradomiciliary, peridomiciliary, and forested environments of two localities. Collected females were dissected, microscopically examined for flagellates in their guts, and preserved in ethanol. A total of 21,197 specimens comprising 14 genera and 57 species were collected, and the majority of these were Nyssomyia, Psychodopygus, and Trichophoromyia genera. Three new records of phlebotomine species for Acre are presented here, including Brumptomyia brumpti, Psathyromyia pradobarrientosi, and for the first time in Brazil, Th. omagua. In Xapuri, the phlebotomine fauna of different ecotopes was varied in regard to abundance, diversity, and frequency, and they included proven and permissive vectors of Leishmania spp. The fauna discovered in the forested areas (57 species) was richer and more diverse than was that (33 species) identified in the peri‑ and intra-domiciles. The identification of Leishmania subgenera that were present in sand fly guts according to SSU rRNA sequences revealed ten and three species harboring Leishmania of subgenera Viannia and Leishmania (most likely Leishmania amazonensis), respectively. The presence of Leishmania (Leishmania) in sand flies are reported here for the first time in Acre. The presence of L. (Viannia) spp. in Brumptomyia sp. and Lutzomyia sherlocki. and the occurrence of mixed infections with Leishmania of both subgenera in Ps. lainsoni have been reported for the first time in Brazil. Taken together, data from previous studies and from the present study highlight the remarkable complexity of phlebotomine fauna that is possibly due to the well-preserved Xapuri forested areas sustaining vital economic activities of plant extraction and ecological tourism. Our findings also provide new insights into the ongoing adaptation of Trichophoromyia and Psychodopygus species to human habitats.

Hidden in plain sight: a molecular field survey of three wheat leaf blotch fungal diseases in North-Western Europe shows co-infection is widespread

Wheat (Triticum aestivum L.) yields are commonly affected by foliar infection by fungal pathogens. Of these, three wheat leaf blotch fungal diseases, septoria nodorum blotch (SNB), tan spot (TS) and septoria tritici blotch (STB), caused by Parastagonospora nodorum (Pn), Pyrenophora tritici-repentis (Ptr) and Zymoseptoria tritici (Zt), respectively, induce major yield losses. Infection results in necrotic areas on the leaf, and it is often difficult to determine the underlying causative pathogen from visible symptoms alone, especially in mixed infections. Here, a regional survey of 330 wheat samples collected across three seasons (years 2015–2017) from four north-west European countries was undertaken. Using quantitative polymerase chain reaction (qPCR) assays specific for each pathogen, as well as disease assessment of leaf materials, distinct regional differences were identified. Two-thirds (65%) of all samples harbored at least two of the three pathogens. Norway had high SNB abundance, but also showed mixed infections of SNB, TS and STB. In Germany, TS was prevalent, with STB also common. Danish samples commonly possessed all three pathogens, with STB prevalent, followed by TS and SNB. The UK had a major prevalence of STB with minimal occurrence of TS and SNB. Across all samples, qPCR identified Zt, Pn and Ptr in 90%, 54% and 57% of samples, respectively. For each pathogen, average disease levels via visual assessment showed modest positive correlation with fungal DNA concentrations (R2 = 0.13–0.32). Overall, our study highlights that the occurrence of mixed infection is common and widespread, with important implications for wheat disease management and breeding strategies.

Analyses of orthotospovirus populations and dispersion under different environmental conditions in Brazil and in the Dominican Republic

Orthotospoviruses (genus Orthotospovirus, family Tospoviridae) are amongst the most devastating plant viruses worldwide, causing severe damage to many economically important vegetable crops, such as tomato and sweet pepper. Monitoring virus populations is an important step for estimating virus damage and epidemiology, and gaining insights into the adaptation processes undergone by orthotospoviruses. Here, we studied the orthotospovirus populations infecting vegetable crops in Brazil and the Dominican Republic, including species diversity, genome comparison and phylogenetic analyses. Comparisons of virus populations showed that in Brazil, which is considered a center of orthotospovirus diversity, groundnut rinspot virus (GRSV) is prevalent, infecting 41% of the plants, whereas tomato spotted wilt virus (TSWV) and tomato chlorotic spot virus (TCSV) were present in 4% and 9% of the samples, respectively. In the Dominican Republic, which can be considered an environment with low orthotospovirus diversity, 55% of the samples were infected with TSWV, 11% showed TCSV infection and no GRSV was detected. The occurrence of mixed infection was low in Brazil, at only 5%, but no mixed infection was detected in the Dominican Republic. The low rates of mixed infections may prevent the emergence of genomes resulting from reassortment. Indeed, no reassortant viruses were detected in either country, except for TCSV, recently proposed as representing a reassortant orthotospovirus species.

Potato Pathogens in Russia's Regions: An Instrumental Survey with the Use of Real-Time PCR/RT-PCR in Matrix Format.

Viral and bacterial diseases of potato cause significant yield loss worldwide. The current data on the occurrence of these diseases in Russia do not provide comprehensive understanding of the phytosanitary situation. Diagnostic systems based on disposable stationary open qPCR micromatrices intended for the detection of eight viral and seven bacterial/oomycetal potato diseases have been used for wide-scale screening of target pathogens to estimate their occurrence in 11 regions of Russia and to assess suitability of the technology for high-throughput diagnostics under conditions of field laboratories. Analysis of 1025 leaf and 725 tuber samples confirmed the earlier reported data on the dominance of potato viruses Y, S, and M in most regions of European Russia, as well as relatively high incidences of Clavibacter michiganensis subsp. sepedonicus, Pectobacterium atrosepticum, and P. carotovorum subsp. carotovorum, and provided detailed information on the phytosanitary status of selected regions and geographical spread of individual pathogens. Information on the occurrence of mixed infections, including their composition, was the first data set of this kind for Russia. The study is the first large-scale screening of a wide range of potato pathogens conducted in network mode using unified methodology and standardized qPCR micromatrices. The data represent valuable information for plant pathologists and potato producers and indicate the high potential of the combined use of matrix PCR technology and network approaches to data collection and analysis with the view to rapidly and accurately assess the prevalence of certain pathogens, as well as the phytosanitary state of large territories.

Prevalence and burden of gastrointestinal helminthes among grey-breasted helmet guinea fowls (Numida meleagris galeata) encountered in Gombe state, Nigeria

This cross sectional survey was conducted from July to December 2015 in order to investigate the burden of gastrointestinal helminthes among guinea fowls in Gombe, Northeastern Nigeria. A total of six hundred fowls (viscera) were purchased from six randomly selected slaughter slabs. Out of the 600 birds examined, 479 (79.83%; 95% CI: 76.4, 82.9) were found harbouring one or more gastrointestinal helminthes. Of this, 238 birds (39.7%; 35.8, 43.6) were infected by nematode species and 392 birds (65.3%; 61.4, 69.0) by cestode species. A total of nine nematodes and seven cestodes species were recovered from these birds. There was no any trematode observed among the studied birds. The prevalences of the nematodes identified in descending order were: Ascaridia galli 56.7% (52.7, 60.6); Ascaridia numidae 38.0% (34.2, 42.0); Heterakis gallinarum 17.2% (14.4, 20.4); Heterakis meleagridis 8.3% (6.4, 10.8); Strongyloides avium 3.5% (2.3, 5.3); Subulura brumpti 3.2% (2.0, 5.0); Gongylonema ingluvicola 2.2% (1.3, 3.7) and both Dispharynx spiralis and Tetrameres numidae had 0.7% (0.3, 1.7). While for cestodes: Raillietina tetragona 72.8% (69.1, 76.2); Raillietina echinobothrida 67.3% (63.5, 71.0); Raillietina cesticillus 50% (46.0, 54.0); Raillietina magninumida 25.7% (22.3, 29.3); Hymenolopsis cantaniana 17.3% (14.5, 20.6); Davainea nana 4.2% (2.8, 6.1) and the lowest was observed in Choanotaenia infundibulum with 2% (1.2, 3.5). Infection rates did not differ significantly based on sex (P > 0.05). However, the occurrence of mixed infection as compared with single infection was statistically significant in both cestodes and nematodes (P < 0.001). The results obtained indicated high prevalence of gastrointestinal helminthes among guinea fowls. These birds may serve as important source of helminthes to other commercial birds in the study area.

Антибиотикорезистентность: эволюционные предпосылки, механизмы, последствия

The article focuses on the problem of studying the antibiotic resistance of microorganisms inhabiting the natural environments of the human and animal organism. It analyzes some data published in Russian and world literature on the importance of this problem in the field of public health, veterinary medicine, the existing mechanisms of the emergence of antibiotic resistance in pathogens and forms of acquired resistance. Widespread use of antibiotics in medicine and veterinary led to the emergence of highly resistant pathogens thus creating both an epizootic and epidemiological problem (A.N. Panin et al., 2017). Extensive use of antimicrobials resulted in the selection of resistant strains among the species of commensal, saprophytic and opportunistic microflora. While resisting the effects of antimicrobial agents, bacteria use different sets of defense mechanisms simultaneously and create new counteraction strategies during the evolution (M.A. Shkurat, I.O. Pokudina, D.V. Battalov, 2014). On the one hand, this is due to the evolution of antibiotic resistance genes determining the formation of new molecular mechanisms of resistance, and, on the other hand, to the formation of new mechanisms of adaptation and maintenance in microecosystems. The latter led to the evolution of microbial communities and the emergence of new nosological units. Microbial communities, interacting with each other, create a barrier (biofilm) as a protective factor against antibacterial agents (D. Hughes, D.I. Andersson, 2017). Currently, the microbial communities of the potentially pathogenic microflora, which are stable by species composition, are widely spread, which is manifested by the occurrence of mixed infections. That raises new problems of ensuring the health of animals and humans.

Viruses occurring in watermelon in the tropical lowland of the Tocantins state, Brazil

The cucurbits are subject to various diseases caused by viruses and the identification of species that predominate in one region has great importance to direct the control strategies. The study aimed to diagnose by DAS-ELISA and RT-PCR tests virus isolates collected in commercial plantations of lowland tropical watermelon in the State of Tocantins. Were collected 20 leaf in the municipality of Formoso do Araguaia and 46 leaf samples from Lagoa da Confusao. The samples were packed in plastic bags properly identified and transferred to the Department of Plant Pathology DFP / UFLA and / or stored in a refrigerator at -80 °C. The samples were grown in a greenhouse for plants pumpkin Curcubita pepo cv. Caserta and were performed the DAS-ELISA test serology and molecular evaluation. No sample reacted with the antiserum for the cucumber mosaic virus (CMV), on the other hand reacted to sixteen mosaic virus-of-pumpkin (SqMV). The predominant virus in producing regions of watermelon grown in lowland tropical rainforest in the state of Tocantins were ZYMV and SqMV and the occurrence of mixed infection (ZYMV+SqMV) was achieved in 35% of samples in the municipality of Lagoa da Confusao. In the municipality of Formoso do Araguaia prevailed ZYMV.

Simultaneous Detection of Both RNA and DNA Viruses Infecting Dry Bean and Occurrence of Mixed Infections by BGYMV, BCMV and BCMNV in the Central-West Region of Mexico.

A multiplex reverse transcription polymerase chain reaction (RT-PCR) assay was developed to simultaneously detect bean common mosaic virus (BCMV), bean common mosaic necrotic virus (BCMNV), and bean golden yellow mosaic virus (BGYMV) from common bean leaves dried with silica gel using a single total nucleic acid extraction cetyl trimethyl ammonium bromide (CTAB) method. A mixture of five specific primers was used to amplify three distinct fragments corresponding to 272 bp from the AC1 gene of BGYMV as well as 469 bp and 746 bp from the CP gene of BCMV and BCMNV, respectively. The three viruses were detected in a single plant or in a bulk of five plants. The multiplex RT-PCR was successfully applied to detect these three viruses from 187 field samples collected from 23 municipalities from the states of Guanajuato, Nayarit and Jalisco, Mexico. Rates of single infections were 14/187 (7.5%), 41/187 (21.9%), and 35/187 (18.7%), for BGYMV, BCMV, and BCMNV, respectively; 29/187 (15.5%) samples were co-infected with two of these viruses and 10/187 (5.3%) with the three viruses. This multiplex RT-PCR assay is a simple, rapid, sensitive, and cost-effective method for detecting these viruses in the common bean and can be used for routine molecular diagnosis and epidemiological studies.

Cucurbit-infecting viruses in Brazil

Cucurbit crops are grown throughout many Brazilian states and are affected by several viral diseases that represent one of the main constraints to cucurbit production in the country causing severe economic losses. During 2008-2015, a survey was performed in open-field of watermelon (Citrullus lanatus), melon (Cucumis melo), cucumber (C. sativus), pumpkin (Cucurbita spp.), chayote (Sechium edule), and gherkin (C. anguria) crops to determine distribution and relative incidence of six viruses. More than 1050 leaf samples were collected from plants showing virus-like symptoms in more than 20 counties of eight Brazilian states (Goias, Bahia, Sao Paulo, Alagoas, Pernambuco, Ceara, Rio Grande do Norte, and Maranhao) and the Federal District. Samples were screened by dot-ELISA or DAS-ELISA tests using polyclonal antibodies to investigate the occurrence of Papaya ringspot virus-type watermelon (PRSV-W), Watermelon mosaic virus (WMV), Zucchini yellow mosaic virus (ZYMV), Cucumber mosaic virus (CMV), Zucchini lethal chlorosis virus (ZLCV), and Melon yellowing-associated virus (MYaV). In addition, total DNA obtained from leaf samples was tested for geminivirus infections by PCR using a pair of universal primers. Serological tests results indicated that approximately 60% of the samples were infected with at least one of the viruses tested. PRSV-W (40.89%) and ZYMV (25.46%) were the most prevalent viruses detected in all cucurbit species. WMV and CMV were found at low percentage in the leaf samples, 8.76 and 5.02%, respectively. MYaV (22.46%) occurred mainly in melon, while ZLCV (18.11%) was detected mostly infecting watermelon and pumpkins. The occurrence of mixed infection was observed in 17.41% of the leaf samples, especially in combination with PRSV-W. None of the samples tested positive for geminivirus infection by PCR. These data confirm the economic importance of cucurbit-infecting viruses in different Brazilian regions.

Detection and quantification of ‘Candidatus Phytoplasma asterisʼ and ‘Candidatus Liberibacter asiaticusʼ at early and late stages of Huanglongbing disease development

ʻCandidatus Phytoplasma asterisʼ (CPa) has been identified along with ʻCandidatus Liberibacter asiaticusʼ (CLas) from citrus trees showing Huanglongbing (HLB) symptoms in Mexico. The low titre of CPa and CLas in infected plants is the major constraint to developing an accurate and sensitive method for their detection that would support an effective HLB diagnosis and management programme. This investigation reports the development and validation of a new TaqMan quantitative PCR to detect low titres of CPa in citrus trees with HLB symptoms based on the phytoplasma 16S rRNA gene region. The assay was used to monitor the titre of CPa at early and late stages of disease development in symptomatic and asymptomatic Mexican lime trees in Colima, Mexico. The concentration of CLas was determined by nested-qPCR. Only eight out of 39 sampled branches showed HLB symptoms at the early infection stage. All 39 branches were symptomatic at the late infection stage. The qPCR yielded an increase in the branch incidence and concentration of CLas (49 to 100% and 5 × 103 to 12 × 103 cells/100 ng of total DNA), while CPa showed a decreased incidence and concentration (72to 41% and 199 to 19 cells/100 ng of total DNA) throughout the disease development. Results confirm the occurrence of mixed infection of CLas and CPa in HLB-affected citrus trees, although their epidemiological role in co-infecting remains unknown.

Molecular epidemiology reveals genetic diversity among 363 isolates of the Cryptococcus neoformans and Cryptococcus gattii species complex in 61 Ivorian HIV-positive patients.

Cryptococcal meningitis is a severe opportunistic infection in HIV-infected patients. In Ivory Coast, despite the availability of antiretroviral treatment (ART), this infection is still prevalent. The study investigates the genetic diversity of 363 clinical isolates of Cryptococcus from 61 Ivorian HIV-positive patients, the occurrence of mixed infections and the in vitro antifungal susceptibility of the isolates. Serotyping was performed via LAC1 and CAP64 gene amplification. Genotyping was performed using the phage M13 core (GACA)4 and (GTG)5 primers and restriction fragment length polymorphism analysis of the URA5 gene. By PCR fingerprinting, the presence of the three serotypes were demonstrated among the 363 isolates in the population studied: A (n=318; 87.6%), AD (n=40; 11%) and B (n=4; 1.1%). Using PCR fingerprinting with primers M13 (GACA)4 and (GTG)5 , we grouped the isolates into 56 molecular subtypes. We observed a high frequency (39.3%) of mixed infections, with up to two different genotypes per sample. None of the isolates were resistant to amphotericin B. Only 0.3% and 1.1% of the isolates were resistant to fluconazole and flucytosine respectively. This study revealed the high genetic diversity among Cryptococcus isolates, the occurrence of mixed infections and a high antifungal susceptibility for the majority of Ivorian cryptococcal isolates.

Cryptococcosis Serotypes Impact Outcome and Provide Evidence of Cryptococcus neoformans Speciation.

ABSTRACTCryptococcus neoformans is a human opportunistic fungal pathogen causing severe disseminated meningoencephalitis, mostly in patients with cellular immune defects. This species is divided into three serotypes: A, D, and the AD hybrid. Our objectives were to compare population structures of serotype A and D clinical isolates and to assess whether infections with AD hybrids differ from infections with the other serotypes. For this purpose, we analyzed 483 isolates and the corresponding clinical data from 234 patients enrolled during the CryptoA/D study or the nationwide survey on cryptococcosis in France. Isolates were characterized in terms of ploidy, serotype, mating type, and genotype, utilizing flow cytometry, serotype- and mating type-specific PCR amplifications, and multilocus sequence typing (MLST) methods. Our results suggest that C. neoformans serotypes A and D have different routes of multiplication (primarily clonal expansion versus recombination events for serotype A and serotype D, respectively) and important genomic differences. Cryptococcosis includes a high proportion of proven or probable infections (21.5%) due to a mixture of genotypes, serotypes, and/or ploidies. Multivariate analysis showed that parameters independently associated with failure to achieve cerebrospinal fluid (CSF) sterilization by week 2 were a high serum antigen titer, the lack of flucytosine during induction therapy, and the occurrence of mixed infection, while infections caused by AD hybrids were more likely to be associated with CSF sterilization. Our study provides additional evidence for the possible speciation of C. neoformans var. neoformans and grubii and highlights the importance of careful characterization of causative isolates.

Mixed Infections and Rifampin Heteroresistance among Mycobacterium tuberculosis Clinical Isolates.

Mixed infections and heteroresistance of Mycobacterium tuberculosis contribute to the difficulty of diagnosis, treatment, and control of tuberculosis. However, there is still no proper solution for these issues. This study aimed to investigate the potential relationship between mixed infections and heteroresistance and to determine the high-risk groups related to these factors. A total of 499 resistant and susceptible isolates were subjected to spoligotyping and 24-locus variable-number tandem repeat methods to analyze their genotypic lineages and the occurrence of mixed infections. Two hundred ninety-two randomly selected isolates were sequenced on their rpoB gene to examine mutations and heteroresistance. The results showed that 12 patients had mixed infections, and the corresponding isolates belonged to Manu2 (n = 8), Beijing (n = 2), T (n = 1), and unknown (n = 1) lineages. Manu2 was found to be significantly associated with mixed infections (odds ratio, 47.72; confidence interval, 9.68 to 235.23; P < 0.01). Four isolates (1.37%) were confirmed to be heteroresistant, which was caused by mixed infections in three (75%) isolates; these belonged to Manu2. Additionally, 3.8% of the rifampin-resistant isolates showing no mutation in the rpoB gene were significantly associated with mixed infections (χ(2), 56.78; P < 0.01). This study revealed for the first time that Manu2 was the predominant group in the cases of mixed infections, and this might be the main reason for heteroresistance and a possible mechanism for isolates without any mutation in the rpoB gene to become rifampin resistant. Further studies should focus on this lineage to clarify its relevance to mixed infections.

Prevalence of gastrointestinal pathogenic bacteria in patients with diarrhoea attending Groote Schuur Hospital, Cape Town, South Africa.

Diarrhoea due to gastrointestinal infections is a significant problem facing the South African (SA) healthcare system. Infections can be acquired both from the community and from the hospital environment itself, the latter acting as a reservoir for potential pathogenic bacteria. To examine the prevalence of a panel of potential diarrhoea-causing bacteria in patients attending a tertiary healthcare facility in Cape Town, SA. Polymerase chain reaction (PCR) primers specific for Clostridium difficile, Shigella spp., Salmonella spp., Klebsiella oxytoca, enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC/EHEC), Staphylococcus aureus, enterotoxigenic Bacteroides fragilis and Campylobacter spp. were used to screen total bacterial genomic DNA extracted from stool samples provided by 156 patients with diarrhoea attending Groote Schuur Hospital, Cape Town, SA. C. difficile was the most frequently detected pathogen (16% of cases) in the 21-87-year-old patient range, but was not present in samples from the 16-20-year-old range. K. oxytoca (6%), EPEC/EHEC strains (9%) and S. aureus (6%) were also detected. The remaining pathogens were present at low frequencies (0-2.9%), and the occurrence of mixed infections was 5%. The majority of non-C. difficile-related diarrhoeas were community acquired. C. difficile was the main cause of infectious diarrhoea in the sampled patients, while K. oxytoca and EPEC/EHEC strains were present as relatively minor but potentially significant pathogens.