Reducing the rate of central line-associated bloodstream infections; a quality improvement project.

BACKGROUND: The burden of central line-associated bloodstream infections is significant and has negative implications for healthcare, increasing morbidity and mortality risks, increasing inpatient hospital stays, and increasing the cost of hospitalization. Efforts to reduce the incidence of central line-associated bloodstream infections have utilized quality improvement projects that implement, measure, and monitor outcomes. However, variations in location, healthcare organization, patient risks, and practice gaps are key to the success of interventions and approaches. This study aims to evaluate interventions of a quality improvement project on the reduction of central line-associated bloodstream infection rates at a university teaching hospital. METHODS: This was a retrospective review of a quality improvement project that was implemented using the Plan-Do-Study-Act quality improvement cycle. Active surveillance of processes and outcomes was performed in the critical care areas; compliance to central line care bundles, and central line-associated bloodstream infections. Interrupted time series was used to analyze trends pre and post-intervention and regression modeling to estimate data segments preceding and succeeding the interventions. RESULTS: There were 350 central line insertions, 3912 catheter days, and 20 central line-associated bloodstream infection events during the intervention period. Compliance with central line care bundles was at 94%. There was a trend in the reduction of central line-associated bloodstream infections by 18% that did not reach statistical significance (p = 0.252). CONCLUSIONS: Improvement projects to reduce central line-associated bloodstream infections face challenges and complexities associated with implementing interventions in real-world healthcare settings. There is a great need to continuously monitor, evaluate, readjust, and adapt interventions to achieve desired results, sustain improvements in patient outcomes, and investigate reasons for non-adherence as keys to achieving desired outcomes.

Outpatient management of urinary tract infections by medical officers in Nairobi, Kenya: lack of benefit from audit and feedback on adherence to treatment guidelines.

INTRODUCTION: Acute uncomplicated urinary tract infections are common in outpatient settings but are not treated optimally. Few studies of the outpatient use of antibiotics for specific diagnoses have been done in sub-Saharan Africa, so little is known about the prescribing patterns of medical officers in the region. METHODS: Aga Khan University has 16 outpatient clinics throughout the Nairobi metro area with a medical officer specifically assigned to that clinic. A baseline assessment of evaluation and treatment of suspected UTI was performed from medical records in these clinics. Then the medical officer from each of the 16 clinics was recruited from each clinic was recruited with eight each randomized to control vs. feedback groups. Both groups were given a multimodal educational session including locally adapted UTI guidelines and emphasis on problems identified in the baseline assessment Each record was scored using a scoring system that was developed for the study according to adequacy of history, physical examination, clinical diagnosis matching recorded data, diagnostic workup and treatment. Three audits were done for both groups; baseline (audit 1), post-CME (audit 2), and a final audit, which was after feedback for the feedback group (audit 3). The primary analysis assessed overall guideline adherence in the feedback group versus the CME only group. RESULTS: The overall scores in both groups showed significant improvement after the CME in comparison to baseline and for each group, the scores in most domains also improved. However, audit 3 showed persistence of the gains attained after the CME but no additional benefit from the feedback. Some deficiencies that persisted throughout the study included lack of workup of possible STI and excess use of non-UTI laboratory tests such as CBC, stool culture and H. pylori Ag. After the CME, the use of nitrofurantoin rose from only 4% to 8% and cephalosporin use increased from 49 to 67%, accompanied by a drop in quinolone use. CONCLUSION: The CME led to modest improvements in patient care in the categories of history taking, treatment and investigations, but feedback had no additional effect. Future studies should consider an enforcement element or a more intensive feedback approach.

Vaginal microbiota in women with spontaneous preterm labor versus those with term labor in Kenya: a case control study.

BACKGROUND: Preterm birth is a global problem with about 12% of births in sub-Saharan Africa occurring before 37 weeks of gestation. Several studies have explored a potential association between vaginal microbiota and preterm birth, and some have found an association while others have not. We performed a study designed to determine whether there is an association with vaginal microbiota and/or placental microbiota and preterm birth in an African setting. METHODS: Women presenting to the study hospital in labor with a gestational age of 26 to 36 weeks plus six days were prospectively enrolled in a study of the microbiota in preterm labor along with controls matched for age and parity. A vaginal sample was collected at the time of presentation to the hospital in active labor. In addition, a placental sample was collected when available. Libraries were constructed using PCR primers to amplify the V6/V7/V8 variable regions of the 16S rRNA gene, followed by sequencing with an Illumina MiSeq machine and analysis using QIIME2 2022.2. RESULTS: Forty-nine women presenting with preterm labor and their controls were enrolled in the study of which 23 matched case-control pairs had sufficient sequence data for comparison. Lactobacillus was identified in all subjects, ranging in abundance from < 1% to > 99%, with Lactobacillus iners and Lactobacillus crispatus the most common species. Over half of the vaginal samples contained Gardnerella and/or Prevotella; both species were associated with preterm birth in previous studies. However, we found no significant difference in composition between mothers with preterm and those with full-term deliveries, with both groups showing roughly equal representation of different Lactobacillus species and dysbiosis-associated genera. Placental samples generally had poor DNA recovery, with a mix of probable sequencing artifacts, contamination, and bacteria acquired during passage through the birth canal. However, several placental samples showed strong evidence for the presence of Streptococcus species, which are known to infect the placenta. CONCLUSIONS: The current study showed no association of preterm birth with composition of the vaginal community. It does provide important information on the range of sequence types in African women and supports other data suggesting that women of African ancestry have an increased frequency of non-Lactobacillus types, but without evidence of associated adverse outcomes.

Assessment of Cardiac, Vascular, and Pulmonary Pathobiology In Vivo During Acute COVID-19.

Background Acute COVID-19-related myocardial, pulmonary, and vascular pathology and how these relate to each other remain unclear. To our knowledge, no studies have used complementary imaging techniques, including molecular imaging, to elucidate this. We used multimodality imaging and biochemical sampling in vivo to identify the pathobiology of acute COVID-19. Specifically, we investigated the presence of myocardial inflammation and its association with coronary artery disease, systemic vasculitis, and pneumonitis. Methods and Results Consecutive patients presenting with acute COVID-19 were prospectively recruited during hospital admission in this cross-sectional study. Imaging involved computed tomography coronary angiography (identified coronary disease), cardiac 2-deoxy-2-[fluorine-18]fluoro-D-glucose positron emission tomography/computed tomography (identified vascular, cardiac, and pulmonary inflammatory cell infiltration), and cardiac magnetic resonance (identified myocardial disease) alongside biomarker sampling. Of 33 patients (median age 51 years, 94% men), 24 (73%) had respiratory symptoms, with the remainder having nonspecific viral symptoms. A total of 9 patients (35%, n=9/25) had cardiac magnetic resonance-defined myocarditis. Of these patients, 53% (n=5/8) had myocardial inflammatory cell infiltration. A total of 2 patients (5%) had elevated troponin levels. Cardiac troponin concentrations were not significantly higher in patients with and without myocarditis (8.4 ng/L [interquartile range, IQR: 4.0-55.3] versus 3.5 ng/L [IQR: 2.5-5.5]; P=0.07) or myocardial cell infiltration (4.4 ng/L [IQR: 3.4-8.3] versus 3.5 ng/L [IQR: 2.8-7.2]; P=0.89). No patients had obstructive coronary artery disease or vasculitis. Pulmonary inflammation and consolidation (percentage of total lung volume) was 17% (IQR: 5%-31%) and 11% (IQR: 7%-18%), respectively. Neither were associated with the presence of myocarditis. Conclusions Myocarditis was present in a third patients with acute COVID-19, and the majority had inflammatory cell infiltration. Pneumonitis was ubiquitous, but this inflammation was not associated with myocarditis. The mechanism of cardiac pathology is nonischemic and not attributable to a vasculitic process. Registration URL: https://www.isrctn.com; Unique identifier: ISRCTN12154994.

Giardia duodenalis: Biology and Pathogenesis.

Giardia duodenalis captured the attention of Leeuwenhoek in 1681 while he was examining his own diarrheal stool, but, ironically, it did not really gain attention as a human pathogen until the 1960s, when outbreaks were reported. Key technological advances, including in vitro cultivation, genomic and proteomic databases, and advances in microscopic and molecular approaches, have led to an understanding that this is a eukaryotic organism with a reduced genome rather than a truly premitochondriate eukaryote. This has included the discovery of mitosomes (vestiges of mitochondria), a transport system with many of the features of the Golgi apparatus, and even evidence for a sexual or parasexual cycle. Cell biology approaches have led to a better understanding of how Giardia survives with two nuclei and how it goes through its life cycle as a noninvasive organism in the hostile environment of the lumen of the host intestine. Studies of its immunology and pathogenesis have moved past the general understanding of the importance of the antibody response in controlling infection to determining the key role of the Th17 response. This work has led to understanding of the requirement for a balanced host immune response that avoids the extremes of an excessive response with collateral damage or one that is unable to clear the organism. This understanding is especially important in view of the remarkable ranges of early manifestations, which range from asymptomatic to persistent diarrhea and weight loss, and longer-term sequelae that include growth stunting in children who had no obvious symptoms and a high frequency of postinfectious irritable bowel syndrome (IBS).

A prospective observational study of community acquired pneumonia in Kenya: the role of viral pathogens.

BACKGROUND: Lower respiratory tract infections continue to contribute significantly to morbidity and mortality across all age groups globally. In sub-Saharan Africa, many studies of community acquired pneumonia in adults have focused on HIV-infected patients and little attention has been given to risk factors and etiologic agents in an urban area with a more moderate HIV prevalence. METHODS: We prospectively enrolled 77 patients admitted to a 280 bed teaching hospital in Kenya with radiographically confirmed community acquired pneumonia from May 2019 to March 2020. The patients were followed for etiology and clinical outcomes. Viral PCR testing was performed using the FTD respiratory pathogen-21 multiplex kit on nasopharyngeal or lower respiratory samples. Additional microbiologic workup was performed as determined by the treating physicians. RESULTS: A potential etiologic agent(s) was identified in 57% including 43% viral, 5% combined viral and bacterial, 5% bacterial and 4% Pneumocystis. The most common etiologic agent was Influenza A which was associated with severe clinical disease. The most common underlying conditions were cardiovascular disease, diabetes and lung disease, while HIV infection was identified in only 13% of patients. Critical care admission was required for 24, and 31% had acute kidney injury, sometimes in combination with acute respiratory distress or sepsis. CONCLUSION: Viruses, especially influenza, were commonly found in patients with CAP. In contrast to other studies from sub-Saharan Africa, the underlying conditions were similar to those reported in high resource areas and point to the growing concern of the double burden of infectious and noncommunicable diseases.

Investigating SOFA, delta-SOFA and MPM-III for mortality prediction among critically ill patients at a private tertiary hospital ICU in Kenya: A retrospective cohort study.

BACKGROUND: Outcomes in well-resourced, intensive care units (ICUs) in Kenya are thought to be comparable to those in high-income countries (HICs) but risk-adjusted mortality data is unavailable. We undertook an evaluation of the Aga Khan University Hospital, Nairobi ICU to analyze patient clinical-demographic characteristics, compare the performance of Sequential Organ Failure Assessment (SOFA), delta-SOFA at 48 hours and Mortality Prediction Model-III (MPM-III) mortality prediction systems, and identify factors associated with increased risk of mortality. METHODS: A retrospective cohort study was conducted of adult patients admitted to the ICU between January 2015 and September 2017. SOFA and MPM-III scores were determined at admission and SOFA repeated at 48 hours. RESULTS: Approximately 33% of patients did not meet ICU admission criteria. Mortality among the population of critically ill patients in the ICU was 31.7%, most of whom were male (61.4%) with a median age of 53.4 years. High adjusted odds of mortality were found among critically ill patients with leukemia (aOR 6.32, p<0.01), tuberculosis (aOR 3.96, p<0.01), post-cardiac arrest (aOR 3.57, p<0.01), admissions from the step-down unit (aOR 3.13, p<0.001), acute kidney injury (aOR 2.97, p<0.001) and metastatic cancer (aOR 2.45, p = 0.04). The area under the receiver-operating characteristic (ROC) curve of admission SOFA was 0.77 (95% CI, 0.73-0.81), MPM-III 0.74 (95% CI, 0.69-0.79), delta-SOFA 0.69 (95% CI, 0.63-0.75) and 48-hour SOFA 0.83 (95% CI, 0.79-0.87). The difference between SOFA at 48 hours and admission SOFA, MPM-III and delta-SOFA was statistically significant (chi2 = 17.1, 24.2 and 26.5 respectively; p<0.001). Admission SOFA, MPM-III and 48-hour SOFA were well calibrated (p >0.05) while delta-SOFA was borderline (p = 0.05). CONCLUSION: Mortality among the critically ill was higher than expected in this well-resourced ICU. 48-hour SOFA performed better than admission SOFA, MPM-III and delta-SOFA in our cohort. While a large proportion of patients did not meet admission criteria but were boarded in the ICU, critically ill patients stepped-up from the step-down unit were unlikely to survive. Patients admitted following a cardiac arrest, and those with advanced disease such as leukemia, stage-4 HIV and metastatic cancer, had particularly poor outcomes. Policies for fair allocation of beds, protocol-driven admission criteria and appropriate case selection could contribute to lowering the risk of mortality among the critically ill to a level on par with HICs.

Staphylococcus aureus bacteremia at a referral medical center in Kenya: A retrospective review of cases from 2010 to 2018.

BACKGROUND: Many studies have shown that Staphylococcus aureus is a leading cause of both community onset and hospital onset bloodstream infections. However, relatively little is known about the occurrence and outcome of S. aureus bacteremia in sub-Saharan Africa. A previous report indicated that S. aureus accounts for 16% of community onset and 6% of hospital onset bloodstream infections at Aga Khan University Hospital Nairobi (AKUHN). Data about the etiology of S. aureus bacteremia in sub-Saharan Africa will help optimize recognition and treatment. This study was performed in order to understand the etiologies and risk factors for S. aureus bacteremia in a sub-Saharan location. MATERIALS AND METHODS: A review of the electronic record of laboratory results from September 2010 through December 2018 identified 201 patients seen at AKUHN with S. aureus bacteremia. The source and/or focus of infection was identified and in-hospital mortality was determined. Cases with bacteremia after three days of hospitalization were considered hospital acquired. Community onset cases were divided into community acquired and health care associated. RESULTS: Most cases (71%; 143/201) were community onset, but only 41% (83/201) of these cases were community acquired. The most commonly identified foci of infection for community acquired bacteremia were musculoskeletal (25%; 21/83) and skin and soft tissue (24%; 20/83). The majority of health care associated (70%; 40/57) and hospital acquired cases (74%; 43/58) were associated with invasive vascular devices, with peripheral IVs being the most common for hospital acquired and dialysis catheters being the most common for health care associated infections. In-hospital mortality rates were 23% (19/83) for community acquired, 19% (11/57) for health care associated and 33% (19/58) for hospital acquired infections. CONCLUSION: Invasive vascular devices were associated with a substantial portion of cases of S. aureus bacteremia and provide an important target for infection control efforts.

Analysis of Candida auris fungemia at a single facility in Kenya.

OBJECTIVES: Candida auris emerged as a human pathogen in 2009 and has subsequently been identified around the world as a cause of invasive candidiasis. We did an analysis from a single institution in order to analyze risk factors and outcomes for C. auris candidemia. METHODS: Patients with candidemia were identified by the electronic medical record and reviewed for risk factors and outcome. Candida isolates were identified by Vitek2 as Candida haemulonii, but species determinations for 21 of the isolates using published molecular and proteomic methods identified all as C. auris. FINDINGS: From September 2010 to December 2016, C. auris accounted for 38% of 201 patients with candidemia, while C. albicans contributed 25%. C. auris patients had been hospitalized longer (mean 32 days vs. 13 days; p<0.001), were more likely to have central lines preceding candidemia than C. albicans patients (84% vs. 54%; p=<0.001) and had more commonly been treated with carbapenems (83% vs 61% for C. albicans [p=0.01]). The crude mortality was 29%, compared to 36% for C. albicans. CONCLUSIONS: These findings suggest an opportunistic pathogen that may be less virulent, but difficult to eradicate and that control efforts should focus on antimicrobial usage.

Seroprevalence data at a private teaching hospital in Kenya: An examination of Toxoplasma gondii, cytomegalovirus, rubella, hepatitis A, and Entamoeba histolytica.

BACKGROUND: Relevant seroprevalence data for endemic pathogens in a given region provide insight not only into a population's susceptibility to acute infection or risk for reactivation disease but also into the potential need for policy initiatives aimed at reducing these risks. Data from sub-Saharan Africa are sparse and since Aga Khan University Hospital Nairobi is an internationally accredited hospital equipped with a laboratory electronic medical record system, analysis of pertinent local seroprevalence data has been made possible. METHODS: We have analyzed serology data from laboratory electronic records at a 300 bed tertiary private teaching hospital in Kenya for the dates, 2008 to 2017 for Toxoplasma gondii, cytomegalovirus, and rubella, which were used primarily for antenatal screening. We also analyzed the data from hepatitis A and amebiasis serologies, which were used primarily for diagnostic purposes. RESULTS: For T. gondii, cytomegalovirus, and rubella, we used IgG serology to determine seroprevalence, finding rates of 32%, 86%, and 89%, respectively. There was no significant age-related difference in the 20 to 49 year old age range for any of these three pathogens. Of the Hepatitis A IgM tests that were ordered, 33% were positive with a peak positive rate of 70% in the five to nine year old age range. The seroprevalence of amebiasis was 4% and all cases of seropositivity were accompanied by compatible clinical illness (hepatic abscess). CONCLUSIONS: These data provide insight into seroprevalence rates of selected pathogens that can be used to guide screening and diagnostic laboratory testing as well as private and public immunization practices.

Spectrum of Microbial Diseases and Resistance Patterns at a Private Teaching Hospital in Kenya: Implications for Clinical Practice.

BACKGROUND: Accurate local prevalence of microbial diseases and microbial resistance data are vital for optimal treatment of patients. However, there are few reports of these data from developing countries, especially from sub-Saharan Africa. The status of Aga Khan University Hospital Nairobi as an internationally accredited hospital and a laboratory with an electronic medical record system has made it possible to analyze local prevalence and antimicrobial susceptibility data and compare it with other published data. METHODS: We have analyzed the spectrum of microbial agents and resistance patterns seen at a 300 bed tertiary private teaching hospital in Kenya using microbial identity and susceptibility data captured in hospital and laboratory electronic records between 2010 and 2014. RESULTS: For blood isolates, we used culture collection within the first three days of hospitalization as a surrogate for community onset, and within that group, Escherichia coli was the most common, followed by Staphylococcus aureus. In contrast, Candida spp. and Klebsiella pneumoniae were the most common hospital onset causes of bloodstream infection. Antimicrobial resistance rates for the most commonly isolated Gram negative organisms was higher than many recent reports from Europe and North America. In contrast, Gram positive resistance rates were quite low, with 94% of S. aureus being susceptible to oxacillin and only rare isolates of vancomycin-resistant enterococci. CONCLUSIONS: The current report demonstrates high rates of antimicrobial resistance in Gram negative organisms, even in outpatients with urinary tract infections. On the other hand, rates of resistance in Gram positive organisms, notably S. aureus, are remarkably low. A better understanding of the reasons for these trends may contribute to ongoing efforts to combat antimicrobial resistance globally.

Measurement of improvement achieved by participation in international laboratory accreditation in sub-Saharan Africa: the Aga Khan University Hospital Nairobi experience.

OBJECTIVES: As part of the ISO 15189:2007 accreditation process, the Aga Khan University Hospital Nairobi laboratory became the first internationally accredited hospital laboratory in sub-Saharan Africa outside South Africa in 2011 through the South Africa National Accreditation System. METHODS: Seven preanalytic, 10 analytic, eight postanalytic, and five administrative performance parameters were monitored from 2009 to 2012 to measure the impact of the accreditation process. RESULTS: Most measures in all four categories showed substantial improvement. The seven preanalytic measures all showed major improvement-between a quarter and a half sigma. Real but less dramatic improvement appeared in analytic and postanalytic measures, but greater than one sigma decrease in analytic "procedure violations" and a three-quarter sigma decrease in excessive turnaround time were noted in these categories. Administrative improvements included dramatic decreases in misdirected and missing reports and complaints. CONCLUSIONS: This study demonstrates the correlation of the accreditation process with improvement in quality measures in a low-resource region.

Genome sequencing of Giardia lamblia genotypes A2 and B isolates (DH and GS) and comparative analysis with the genomes of genotypes A1 and E (WB and Pig).

Giardia lamblia (syn G. intestinalis, G. duodenalis) is the most common pathogenic intestinal parasite of humans worldwide and is a frequent cause of endemic and epidemic diarrhea. G. lamblia is divided into eight genotypes (A-H) which infect a wide range of mammals and humans, but human infections are caused by Genotypes A and B. To unambiguously determine the relationship among genotypes, we sequenced GS and DH (Genotypes B and A2) to high depth coverage and compared the assemblies with the nearly completed WB genome and draft sequencing surveys of Genotypes E (P15; pig isolate) and B (GS; human isolate). Our results identified DH as the smallest Giardia genome sequenced to date, while GS is the largest. Our open reading frame analyses and phylogenetic analyses showed that GS was more distant from the other three genomes than any of the other three were from each other. Whole-genome comparisons of DH_A2 and GS_B with the optically mapped WB_A1 demonstrated substantial synteny across all five chromosomes but also included a number of rearrangements, inversions, and chromosomal translocations that were more common toward the chromosome ends. However, the WB_A1/GS_B alignment demonstrated only about 70% sequence identity across the syntenic regions. Our findings add to information presented in previous reports suggesting that GS is a different species of Giardia as supported by the degree of genomic diversity, coding capacity, heterozygosity, phylogenetic distance, and known biological differences from WB_A1 and other G. lamblia genotypes.

Vertebral coccidioidomycosis: presentation and multidisciplinary management.

BACKGROUND: Vertebral involvement is a severe complication of infection caused by Coccidioides species. METHODS: We conducted a retrospective review of patients diagnosed with vertebral coccidioidomycosis at an academic medical center between 1996 and 2009. RESULTS: We identified 39 cases of vertebral coccidioidomycosis. Thirty-four patients (79%) were male, and 23 patients (61%) were black. Black patients were overrepresented in comparison with all other patients by a 50-fold odds ratio (95% confidence interval, 26-95). Only 8 patients (20%) were immunocompromised, including 7 who had received systemic steroids. The number of infected vertebrae ranged from 1 to 24; 8 patients (21%) had epidural involvement. All patients received a triazole as part of medical therapy, and 20 patients also received amphotericin B, typically early in the course. Twenty-six patients (67%) required surgery, 18 of whom also required hardware placement. The most common indication for surgery was pain, but 7 patients had neurologic compromise. No patients developed recurrent or refractory infection at the site of surgical debridement, but 6 patients experienced disease relapse after stopping antifungal therapy. CONCLUSION: Vertebral infection caused by Coccidioides species requires a multispecialty approach that always includes medical therapy and frequently requires surgical intervention for debridement or stabilization. A favorable outcome can usually be achieved, but discontinuation of medical therapy is associated with a high risk of relapse, which can occur years later.

Musculoskeletal coccidioidomycosis.

Coccidioidomycosis (valley fever) is a systemic fungal infection caused by soil fungi, Coccidioides species, and is associated with significant morbidity and mortality. This infection is endemic in northern Mexico, the southwestern United States, and parts of Central and South America. The risk factors include ethnicity (especially African and Pacific Island ancestry), male gender, and immunosuppression. The primary infection occurs in lungs, but fewer than 40% of patients are symptomatic. Fewer than 1% of infections result in disseminated disease, which may involve any organ. Skeletal infection occurs in 10 to 50% of these patients and is frequently multicentric with axial skeleton involvement. The diagnosis can be confirmed by culture of the organism or visual detection from cytological or histological specimens obtained from sites of disease. A presumptive diagnosis can be made in patients with a compatible illness and a positive serologic test. Imaging plays a significant role in diagnosis and follow-up treatment of musculoskeletal coccidioidomycosis and includes radiography, magnetic resonance imaging, computed tomography, and bone scintigraphy. Skeletal coccidioidomycosis is difficult to treat and frequently requires a combination of medical therapy and surgical debridement. Medical therapy must be continued for a prolonged period of time, potentially for a lifetime in some patients.

Optical map of the genotype A1 WB C6 Giardia lamblia genome isolate.

The Giardia lamblia genome consists of 12 Mb divided among 5 chromosomes ranging in size from approximately 1 to 4 Mb. The assembled contigs of the genotype A1 isolate, WB, were previously mapped along the 5 chromosomes on the basis of hybridization of plasmid clones representing the contigs to chromosomes separated by PFGE. In the current report, we have generated an MluI optical map of the WB genome to improve the accuracy of the physical map. This has allowed us to correct several assembly errors and to better define the extent of the subtelomeric regions that are not included in the genome assembly.

Molecular analysis of household transmission of Giardia lamblia in a region of high endemicity in Peru.

BACKGROUND: Giardia lamblia is ubiquitous in multiple communities of nonindustrialized nations. Genotypes A1, A2, and B (Nash groups 1, 2, and 3, respectively) are found in humans, whereas genotypes C and D are typically found in dogs. However, genotypes A and B have occasionally been identified in dogs. METHODS: Fecal Giardia isolates from 22 families and their dogs, living in Pampas de San Juan, were collected over 7 weeks in 2002 and 6 weeks in 2003. Samples were genotyped, followed by sequencing and haplotyping of many of these isolates by using loci on chromosomes 3 and 5. RESULTS: Human infections were all caused by isolates of genotypes A2 and B. Human coinfections with genotypes A2 and B were common, and the reassortment pattern of different subtypes of A2 isolates supports prior observations that suggested recombination among genotype A2 isolates. All dogs had genotypes C and/or D, with one exception of a dog with a mixed B/D genotype infection. CONCLUSIONS: In a region of high endemicity where infected dogs and humans constantly commingle, different genotypes of Giardia are almost always found in dogs and humans, suggesting that zoonotic transmission is very uncommon.

Phylogenetic status of Pneumocystis from corticosteroid-treated gerbils.

Pneumocystis spp. infect the lungs of multiple mammalian species and cause disease in immunosuppressed individuals. The Pneumocystis isolates that have been studied to date fall into two major clades, those from primates and those from rodents. Within each of these clades, different species have been described on the basis of host specificity and differences in sequence and morphology. Here, we demonstrate that dexamethasone immunosuppression consistently results in histologically apparent lung infection in gerbils (28/35 animals). Sequence analysis of the 18S, 5.8S and internal transcribed spacer regions of the rDNA and a portion of the mitochondrial large subunit rDNA demonstrated that this gerbil Pneumocystis is grouped with other rodent Pneumocystis spp., but is distinct from them. Our results suggest that gerbil Pneumocystis differs sufficiently from Pneumocystis species found in other rodents to be considered a separate species.

The Giardia lamblia vsp gene repertoire: characteristics, genomic organization, and evolution.

BACKGROUND: Giardia lamblia trophozoites colonize the intestines of susceptible mammals and cause diarrhea, which can be prolonged despite an intestinal immune response. The variable expression of the variant-specific surface protein (VSP) genes may contribute to this prolonged infection. Only one is expressed at a time, and switching expression from one gene to another occurs by an epigenetic mechanism. RESULTS: The WB Giardia isolate has been sequenced at 10x coverage and assembled into 306 contigs as large as 870 kb in size. We have used this assembly to evaluate the genomic organization and evolution of the vsp repertoire. We have identified 228 complete and 75 partial vsp gene sequences for an estimated repertoire of 270 to 303, making up about 4% of the genome. The vsp gene diversity includes 30 genes containing tandem repeats, and 14 vsp pairs of identical genes present in either head to head or tail to tail configurations (designated as inverted pairs), where the two genes are separated by 2 to 4 kb of non-coding DNA. Interestingly, over half the total vsp repertoire is present in the form of linear gene arrays that can contain up to 10 vsp gene members. Lastly, evidence for recombination within and across minor clades of vsp genes is provided. CONCLUSIONS: The data we present here is the first comprehensive analysis of the vsp gene family from the Genotype A1 WB isolate with an emphasis on vsp characterization, function, evolution and contributions to pathogenesis of this important pathogen.

Giardia lamblia: intracellular localization of alpha8-giardin.

Alpha8-giardin (α8-giardin) is a member of the multi-gene α-giardin family in the intestinal parasitic protozoan, Giardia lamblia. This gene family shares an ancestry with the annexin super family, whose common characteristic is calcium dependent binding to membranes that contain acidic phospholipids. In the present study, the antigenicity, hydrophilicity, flexibility, surface probability, and secondary structure of α8-giardin amino acids were predicted by bioinformatics applications. A specific anti-peptide antiserum, anti-P3, was used to determine the intracellular location of α8-giardin with confocal immunofluorescence microscopy and immunoelectron microscopy. The results indicated that α8-giardin was located on the plasma membrane and flagella, but not on the ventral disk. Reduction of α8-giardin transcript levels by ribozyme-mediated cleavage decreased trophozoite motility and growth rate, indicating the functional importance of α8-giardin to Giardia trophozoite biology.