Next-generation IgVH sequencing CLL-like monoclonal B-cell lymphocytosis reveals frequent oligoclonality and ongoing hypermutation.

Chronic lymphocytic leukemia (CLL) develops from CLL-like monoclonal B-cell lymphocytosis (MBL) which represents a low-level asymptomatic expansion of cells that phenotypically resemble CLL. Although antigen selection plays a key role during CLL development, it is not known whether this occurs in early MBL or only during progression to CLL. Recent studies suggested that MBL sometimes displays oligoclonality, but these used techniques with limited sensitivity and specificity and were not conclusive. In this study, we combine cell sorting and next-generation sequencing of rearranged immunoglobulin heavy chain variable (IgVH) genes to thoroughly assess the VH repertoire and oligoclonality of purified MBL cells. Clonal functional rearrangements or clonotypes were identified in 29 of 30 sequenced cases, with 7 or 24% having two clonotypes with unrelated CDR3 sequences. In four of the seven cases with unrelated clonotypes, VH segments from the same family were used. In addition, 6 of 29 cases showed clear evidence of ongoing VH gene hypermutation with three of these being among the seven with unrelated clonotypes. This study conclusively shows that MBL cases often contain multiple B-cell clones, the first to report ongoing VH gene mutation in MBL, and that antigen selection appears to occur in early MBL.

Influence of skin colour on diagnostic accuracy of the jaundice meter JM 103 in newborns.

AIM: To assess the diagnostic accuracy of the JM 103 as a screening tool for neonatal jaundice and explore differential effects based on skin colour. METHODS: We prospectively compared the transcutaneous bilirubin (TcB) and serum bilirubin (TSB) measurements of newborns over a 3 month-period. Skin colour was assigned via reference colour swatches. Diagnostic measures of the TcB/TSB comparison were made and clinically relevant TcB cut-off values were determined for each skin colour group. RESULTS: 451 infants (51 light, 326 medium and 74 dark skin colour) were recruited. The association between TcB and TSB was high for all skin colours (rs>0.9). The Bland-Altman analysis showed an absolute mean difference between the two measures of 13.3±26.4 µmol/L with broad limits of agreement (-39.4-66.0 µmol/L), with TcB underestimating TSB in light and medium skin colours and overestimating in dark skin colour. Diagnostic measures were also consistently high across skin colours, with no clinically significant differences observed. CONCLUSIONS: The JM 103 is a useful screening tool to identify infants in need of serum bilirubin, regardless of skin colour. The effect of skin colour on the accuracy of this device at high levels of serum bilirubin could not be assessed fully due to small numbers in the light and dark groups.

Thrombocytopenia: an update.

Thrombocytopenia is a common clinical problem with numerous potential causes including decreased bone marrow platelet production, increased peripheral platelet destruction, increased splenic sequestration, and dilution. Investigation of the etiology of thrombocytopenia requires careful consideration of clinical history and laboratory features. A complete blood count and peripheral smear review are essential components of the diagnostic work-up, and physicians should be knowledgeable about appropriate selection and interpretation of more specialized tests, including bone marrow examination, to assist with diagnosis. This review article aims to summarize and address appropriate work-up of the major and/or life-threatening causes of thrombocytopenia and some of the better-characterized congenital thrombocytopenias.

Evaluation of the impact of fetal fibronectin test implementation on hospital admissions for preterm labour in Ontario: a multiple baseline time-series design.

OBJECTIVE: To determine the impact of a health system-wide fetal fibronectin (fFN) testing programme on the rates of hospital admission for preterm labour (PTL). DESIGN: Multiple baseline time-series design. SETTING: Canadian province of Ontario. POPULATION: A retrospective population-based cohort of antepartum and delivered obstetrical admissions in all Ontario hospitals between 1 April 2002 and 31 March 2010. METHODS: International Classification of Diseases codes in a health system-wide hospital administrative database were used to identify the study population and define the outcome measure. An aggregate time series of monthly rates of hospital admissions for PTL was analysed using segmented regression models after aligning the fFN test implementation date for each institution. MAIN OUTCOME MEASURE: Rate of obstetrical hospital admission for PTL. RESULTS: Estimated rates of hospital admission for PTL following fFN implementation were lower than predicted had pre-implementation trends prevailed. The reduction in the rate was modest, but statistically significant, when estimated at 12 months following fFN implementation (-0.96 hospital admissions for PTL per 100 preterm births; 95% confidence interval [CI], -1.02 to -0.90, P = 0.04). The statistically significant reduction was sustained at 24 and 36 months following implementation. CONCLUSIONS: Using a robust quasi-experimental study design to overcome confounding as a result of underlying secular trends or concurrent interventions, we found evidence of a small but statistically significant reduction in the health system-level rate of hospital admissions for PTL following implementation of fFN testing in a large Canadian province.

Low-dose chemotherapy and rituximab for posttransplant lymphoproliferative disease (PTLD): a Children's Oncology Group Report.

Optimal therapy for posttransplant lymphoproliferative disease (PTLD) remains problematic. A phase II trial adding rituximab to a low-dose cyclophosphamide and prednisone regimen was conducted for pediatric patients with Epstein-Barr virus (EBV) (+), CD20 (+) PTLD. Fifty-five patients were enrolled. Toxicity was similar for cycles of therapy containing rituximab versus those without. The complete remission (CR) rate was 69% (95% confidence interval (CI); 57%-84%). Of 12 patients with radiographic evidence of persistent disease at the end of therapy, eight were in CR 28 weeks later without further PTLD therapy. There were 10 deaths, 3 due to infections while receiving therapy and 7 from PTLD. The 2-year event-free survival (alive with functioning original allograft and no PTLD) was 71% (95% CI: 57%-82%) and overall survival was 83% (95% CI: 69%-91%) with median follow-up of 4.8 years. Due to small numbers, we were unable to determine significance of tumor histology, stage of disease, allograft type or early response to treatment on outcome. These data suggest rituximab combined with low-dose chemotherapy is safe and effective in treating pediatric with EBV (+) PTLD following solid-organ transplantation.

Do molecules matter more than morphology? Promises and pitfalls in parasites.

Systematics involves resolving both the taxonomy and phylogenetic placement of organisms. We review the advantages and disadvantages of the two kinds of information commonly used for such inferences--morphological and molecular data--as applied to the systematics of metazoan parasites generally, with special attention to the malaria parasites. The problems that potentially confound the use of morphology in parasites include challenges to consistent specimen preservation, plasticity of features depending on hosts or other environmental factors, and morphological convergence. Molecular characters such as DNA sequences present an alternative data source and are particularly useful when not all the parasite's life stages are present or when parasitaemia is low. Nonetheless, molecular data can bring challenges that include troublesome DNA isolation, paralogous gene copies, difficulty in developing molecular markers, and preferential amplification in mixed species infections. Given the differential benefits and shortcomings of both molecular and morphological characters, both should be implemented in parasite taxonomy and phylogenetics.

The de-ubiquitinase UCH-L1 is an oncogene that drives the development of lymphoma in vivo by deregulating PHLPP1 and Akt signaling.

De-ubiquitinating enzymes (DUBs) can reverse the modifications catalyzed by ubiquitin ligases and as such are believed to be important regulators of a variety of cellular processes. Several members of this protein family have been associated with human cancers; however, there is little evidence for a direct link between deregulated de-ubiquitination and neoplastic transformation. Ubiquitin C-terminal hydrolase (UCH)-L1 is a DUB of unknown function that is overexpressed in several human cancers, but whether it has oncogenic properties has not been established. To address this issue, we generated mice that overexpress UCH-L1 under the control of a ubiquitous promoter. Here, we show that UCH-L1 transgenic mice are prone to malignancy, primarily lymphomas and lung tumors. Furthermore, UCH-L1 overexpression strongly accelerated lymphomagenesis in Emu-myc transgenic mice. Aberrantly expressed UCH-L1 boosts signaling through the Akt pathway by downregulating the antagonistic phosphatase PHLPP1, an event that requires its de-ubiquitinase activity. These data provide the first in vivo evidence for DUB-driven oncogenesis and suggest that UCH-L1 hyperactivity deregulates normal Akt signaling.

Heritable T-cell malignancy models established in a zebrafish phenotypic screen.

T-cell neoplasias are common in pediatric oncology, and include acute lymphoblastic leukemia (T-ALL) and lymphoblastic lymphoma (T-LBL). These cancers have worse prognoses than their B-cell counterparts, and their treatments carry significant morbidity. Although many pediatric malignancies have characteristic translocations, most T-lymphocyte-derived diseases lack cytogenetic hallmarks. Lacking these informative lesions, insight into their molecular pathogenesis is less complete. Although dysregulation of the NOTCH1 pathway occurs in a substantial fraction of cases, many other genetic lesions of T-cell malignancy have not yet been determined. To address this deficiency, we pioneered a phenotype-driven forward-genetic screen in zebrafish (Danio rerio). Using transgenic fish with T-lymphocyte-specific expression of enhanced green fluorescent protein (EGFP), we performed chemical mutagenesis, screened animals for GFP(+) tumors, and identified multiple lines with a heritable predisposition to T-cell malignancy. In each line, the patterns of infiltration and morphological appearance resembled human T-ALL and T-LBL. T-cell receptor analyses confirmed their clonality. Malignancies were transplantable and contained leukemia-initiating cells, like their human correlates. In summary, we have identified multiple zebrafish mutants that recapitulate human T-cell neoplasia and show heritable transmission. These vertebrate models provide new genetic platforms for the study of these important human cancers.

Specific cytogenetic abnormalities are associated with a significantly inferior outcome in children and adolescents with mature B-cell non-Hodgkin's lymphoma: results of the FAB/LMB 96 international study.

Clinical studies showed that advanced stage, high LDH, poor response to reduction therapy and combined bone marrow and central nervous system disease are significantly associated with a decreased event-free survival (EFS) in pediatric mature B-cell non-Hodgkin's lymphoma (B-NHL) treated on FAB/LMB96. Although rearranged MYC/8q24 (R8q24) is characteristic of Burkitt lymphoma (BL), little information is available on other cytogenetic abnormalities and their prognostic importance. We performed an international review of 238 abnormal karyotypes in childhood mature B-NHL treated on FAB/LMB96: 76% BL, 8% Burkitt-like lymphoma, 13% diffuse large B-cell lymphoma (DLBCL). The main BL R8q24-associated chromosomal aberrations were +1q (29%), +7q and del(13q) (14% each). The DLBCL appeared heterogeneous and more complex. Incidence of R8q24 (34%) was higher than reported in adult DLBCL. The prognostic value of cytogenetic abnormalities on EFS was studied by Cox model controlling for the known risk factors: R8q24, +7q and del(13q) were independently associated with a significant inferior EFS (hazard ratio: 6.1 (P=0.030), 2.5 (P=0.015) and 4.0 (P=0.0003), respectively). The adverse prognosis of R8q24 was observed only in DLBCL, whereas del(13q) and +7q had a similar effect in DLBCL and BL. These results emphasize the significant biological heterogeneity and the development of cytogenetic risk-adapted therapy in childhood mature B-NHL.

The effect of tobacco exposure on the fetal hypothalamic-pituitary-adrenal axis.

OBJECTIVE: Our objective was to determine if maternal smoking is associated with programming of the fetal hypothalamic-pituitary-adrenal (HPA) axis. Cigarette smoking, which induces a state of hypoxia in the fetus, may promote in utero'programming' of the HPA axis. In utero, adaptations to the HPA axis, which become maladaptive later in life, have been hypothesised to contribute to the development of adult cardiovascular disease and metabolic disorders. DESIGN: This was a prospective cohort study of term infants. POPULATION AND SETTING: The study involved 104 infants born by elective caesarean section, 21 of whom were exposed to in utero tobacco and 83 were nonexposed. METHODS: Healthy women with healthy pregnancies were recruited if they were undergoing elective caesarean section. Maternal blood was drawn for cortisol and cotinine in the morning, and the umbilical blood was drawn immediately after delivery of the baby. MAIN OUTCOME MEASURES: Umbilical arterial cortisol and adrenocorticotropin hormone (ACTH) levels. RESULTS: ACTH levels were significantly elevated in smoke-exposed infants [17 (4-22) pmol/l versus 4 (2-11) pmol/l, respectively, P= 0.005], while cortisol levels were similar [182 (130-240) nmol/l versus 192 (127-265) nmol/l, respectively, P= 0.541]. CONCLUSIONS: For the first time, it was shown that infants exposed to in utero tobacco smoke have significantly elevated ACTH levels compared with nonexposed infants. The results of this study warrant further exploration of the effect of smoking on the neonatal HPA axis as a potential set up for 'programming'.

Feasibility of upfront dose-intensive chemotherapy in children with advanced-stage Hodgkin's lymphoma: preliminary results from the Children's Cancer Group Study CCG-59704.

BACKGROUND: Treatment strategies involving dose intensification have recently demonstrated improvements in cure compared with older trials. However, dose-intensive therapy is associated with increased acute and long-term toxicities, particularly in pediatric patients. The Children's Cancer Group initiated this pilot study to assess the feasibility and toxicity of a moderate dose-intensive regimen, BEACOPP (bleomycin, etoposide, doxorubicin, cyclophosphamide, vincristine, procarbazine and prednisone), in children and adolescents with advanced-stage Hodgkin's lymphoma (HL). PATIENTS AND METHODS: Children with stage IIB or IIIB with bulk disease, or stage IV were eligible. Induction consisted of four cycles of escalated dose BEACOPP. The rapidity of response, defined as >70% reduction in disease burden, was assessed after two and four cycles. Rapid responders then received consolidation therapy as per gender-specific guidelines to reduce the risk of gender-specific long-term toxicities of therapy, i.e. females received four cycles of COPP/ABV (cyclophosphamide, vincristine, procarbazine, prednisone, doxorubicin, bleomycin and vinblastine) without radiation therapy and males received two cycles of ABVD (doxurubicin, bleomycin, vinblastine and dacarbazine) with involved field radiation therapy (IFRT). Slow responders received four cycles of BEACOPP and IFRT. RESULTS: Ninety-nine patients were enrolled. Myelosuppression was frequent. Non-hematological grade 4 toxicities included allergic reaction (two patients), hypotension (one), mucositis (four), infection (three), seizure (one) and elevated transaminases (one). Typhlitis developed in four patients; three recovered and completed dose-modified chemotherapy, while one died of sepsis associated with grade 4 neutropenia. A rapid response was achieved by 45 and 72% of patients after two and four cycles, respectively. There are no disease progressions or secondary malignancies to date. There is only one reported relapse to date. Median follow-up for the cohort is 6 months. CONCLUSIONS: BEACOPP chemotherapy is feasible and generally well tolerated in children with advanced-stage HL. The absence of reported progressive disease and only one relapse to date is encouraging.

Diversity of the apoptotic response to chemotherapy in childhood leukemia.

Apoptosis is the primary mechanism through which most chemotherapeutic agents induce tumor cell death. The purpose of this study was to determine the extent to which blasts from children with leukemia undergo a uniform apoptotic death pathway in vivo. The expression of pro- and anti-apoptotic proteins p53, p21, MDM-2, BCL-2, BCL-X(L), BCL-X(S), and BAX, and caspase-3 activity was determined in circulating blasts collected from the peripheral blood of children with leukemia prior to, and at serial time points following chemotherapy. Culturing blasts ex vivo for 12 h assessed spontaneous apoptosis and the increment induced by chemotherapy. Baseline apoptosis varied between 3% and 29%. Twenty-four hours following chemotherapy the increase in the percentage of cells undergoing apoptosis ranged from <1% to 38%. Eleven of 20 patients who received initial treatment with a p53-dependent drug showed an increase in p53 expression. In these patients, the levels of p53 target genes were also increased. A uniform pattern of BCL-2 family protein expression was not observed and only a minority of samples showed a change that would favor apoptosis. We conclude that that the initial apoptotic response to chemotherapy in children with leukemia is variable involving both p53-dependent and p53-independent pathways.

Are myogenin and myoD1 expression specific for rhabdomyosarcoma? A study of 150 cases, with emphasis on spindle cell mimics.

Rhabdomyosarcoma (RMS), the most common soft tissue sarcoma of childhood, displays a variety of histologic patterns. Immunohistochemistry is used extensively to distinguish RMS from its mimics. Myogenin and MyoD1, myogenic transcriptional regulatory proteins expressed early in skeletal muscle differentiation, are considered sensitive and specific markers for RMS and are more specific than desmin and muscle-specific actin and more sensitive than myoglobin. Previous studies have focused on expression of myogenin and MyoD1 in small round cell tumors. This study assesses myogenin and MyoD1 in rhabdomyosarcoma subtypes and spindle cell tumors considered in the differential diagnosis of RMS. Formalin-fixed, paraffin-embedded archival tissue from 32 RMS, 107 non-RMS, and 11 benign skeletal muscle samples was stained for myogenin and MyoD1 with standard immunohistochemical techniques. Nuclear positivity was scored on a three-tiered scale. All RMSs expressed myogenin. Alveolar RMS (ARMS) showed strong nuclear staining, especially in tumor cells lining fibrous septae and perivascular regions. In cases with a subtle alveolar architecture on routinely stained sections, myogenin highlighted and enhanced visualization of the alveolar morphologic pattern. Embryonal RMSs (ERMSs) were more variable in myogenin staining pattern and intensity. No cases of nodular fasciitis, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, inflammatory myofibroblastic tumor, myofibrosarcoma, leiomyoma, leiomyosarcoma, or alveolar soft part sarcoma stained for myogenin. Focal nuclear reactivity was seen in desmoid (2 of 10), infantile myofibromatosis (2 of 10), synovial sarcoma (1 of 10), and infantile fibrosarcoma (2 of 10). Non-neoplastic skeletal muscle fiber nuclei stained positively for myogenin in both tumor-associated samples (25 of 40) and benign skeletal muscle samples (5 of 11). Although all RMSs were immunoreactive for MyoD1, cytoplasmic and nonspecific background staining and reactivity of nonmyoid tissues hindered its practical utility in paraffin-embedded samples in this study. Although myogenin is a highly sensitive and specific marker for RMS, it is rarely seen in other spindle cell soft tissue tumors. As previously reported, ARMS stained more strongly than ERMS. In contrast to previous studies, rare non-RMS (7 of 107) displayed focal nuclear reactivity, and entrapped atrophic or regenerative skeletal muscle fibers also stained positively. Although these are potential pitfalls in the interpretation of myogenin, careful attention to morphology and other features, to the relative paucity of myogenin-positive nuclei in non-RMS. and to the presence of entrapped muscle fibers should prevent incorrect interpretation. Because the extent of myogenin expression in RMS is much greater than in non-RMS, it is a very useful marker when interpreted in the context of other clinicopathologic data.

Phylogeny of nuclear small subunit rRNA genes of hemogregarines amplified with specific primers.

Hemogregarines, apicomplexan intracellular blood parasites, are cosmopolitan in distribution and infect a broad range of vertebrate and invertebrate hosts. Molecular phylogenetic studies have been hampered by lack of hemogregarine-specific polymerase chain reaction primers that would allow amplification of parasite, but not host, DNA. A novel method for separating parasite and host 18S rRNA genes has been developed, and new primers that are specific for hemogregarine rRNA genes have been designed. These primers were used to obtain sequences from 4 isolates of hemogregarines of lizards from California, the Caribbean island of Grenada, eastern Australia, and Israel. Combining these results with already published sequences, a preliminary phylogeny of hemogregarines and several other apicomplexan taxa has been created. The hemogregarines form a monophyletic group and appear to be more closely related to coccidia than to Plasmodium species. The difficulty of using 18S genes that have multiple copies in some apicomplexan parasites was explored for systematic studies.

CD30 expression in follicular lymphoma.

CONTEXT: CD30(+) anaplastic large cell lymphomas were originally described as being of T-cell, null cell, and B-cell origin. CD30, however, is not a specific marker of anaplastic large cell lymphoma and has been found to be expressed in reactive as well as neoplastic populations as a probable activation marker. In addition, CD30(+) cells have also been described in both diffuse large B-cell and follicular lymphomas (FLs), resembling the pattern seen in reactive tonsils and lymph nodes. OBJECTIVE: We report an index case of FL with CD30 expression, which on initial touch preparations and flow cytometric immunophenotyping revealed a prominent population of CD30(+) cells with marked cellular pleomorphism (anaplasia) in a background of typical FL. Immunohistochemistry of the paraffin section for CD30 in our index case confirmed unequivocal CD30(+) pleomorphic cells in the malignant nodules in occasional clusters. This case prompted a study of additional cases of FL for pattern of immunoreactivity with CD30 on paraffin sections. DESIGN: Twenty-two additional cases of FL (grades 1-3) were retrieved for CD30 immunoperoxidase staining as in the index case. RESULTS: This study demonstrated 32% of the additional cases of FL had definitive CD30(+), large, pleomorphic malignant cells by paraffin immunohistochemistry. In 2 cases (9%), the pattern of immunoreactivity with CD30 showed clustering and variable staining of large cells, as our index case. CONCLUSION: This study underscores the morphologic and immunophenotypic spectrum of FL that includes CD30 staining and cellular pleomorphism.

Mantle cell leukemia, prolymphocytoid type: a rarely described form.

Leukemic manifestations of mantle cell lymphoma are seen in a minority of cases, usually associated with extensive tumor. Usually the neoplastic cells in the peripheral blood resemble mantle cells with a mature chromatin pattern and irregular nuclear contours, or less frequently with a more "blastic" chromatin pattern. A prolymphocytic leukemia with t(11;14)(q13;q32) has previously been reported; however, a complete flow cytometric immunophenotypic profile was lacking. Mantle cell leukemia, prolymphocytoid type with complete flow cytometric data has not previously been described and is the purpose of this report. We report such a case in a 90 year-old female who presented with an elevated white blood cell count. The diagnosis was based on flow cytometric immunophenotyping and the cytomorphology of the peripheral blood combined with cyclin D1 immunohistochemical staining of the bone marrow. We describe our findings and her clinical course in order to heighten awareness of this previously rarely described entity.

Large cell variants of CD5+, CD23- B-cell lymphoma/leukemia.

CONTEXT: Mantle cell lymphoma (MCL), and its leukemic phase, constitute a well-studied hematologic malignancy with known overall survival, prognostic indicators, morphologic findings at diagnosis and in bone marrow, and known incidence of the bcl-1 immunoglobulin gene rearrangement. Large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype (CD5+, CD23-), including but not limited to blastic MCL, prolymphocytoid MCL, blastic mantle cell leukemia, and prolymphocytic mantle cell leukemia, are not as well characterized. Although blastic MCL is known to be associated with a shorter overall survival than conventional MCL, the large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype have not been described as fully as conventional MCL. OBJECTIVE: The purpose of the present study was to describe the large cell variants of B-cell lymphoma/leukemia with a mantle cell immunophenotype. DESIGN: Nineteen cases of large cell variants of CD5+, CD23- B-cell lymphoma/leukemia are reviewed and described in regard to morphology, bone marrow morphological findings, Cyclin D1 immunostaining, and bcl-1 analysis. Clinical data were not available owing to the varied clinical sources of the specimens. SETTING: Tertiary-care academic institution. RESULTS: Lymph node involvement in blastic CD5+, CD23- B-cell lymphoma was diffuse (100%) with a nodular component (33%) or focal mantle zone pattern (10%). Bone marrow involvement in blastic CD5+, CD23- B-cell lymphoma was seen in only 27% of cases and was composed predominantly of small, slightly irregular lymphocytes. Cyclin D1 was demonstrated in 60% of the 15 cases analyzed and more sensitive in B5-fixed tissue. Bcl-1 (performed in 5 cases) was not detected in the 4 cases of blastic CD5+, CD23- B-cell lymphoma analyzed and was detected in the case of the prolymphocytoid MCL. Cyclin D1 was demonstrated in all 4 bcl-1 negative cases and was negative in the bcl-1 positive prolymphocytoid MCL. CONCLUSION: Careful analysis of clinical data, morphology, immunophenotype, Cyclin D1 expression, and molecular analysis are required to differentiate the unusual large cell variants of MCL from other processes.

Pharmacokinetics of the ganciclovir implant in the silicone-filled eye.

PURPOSE: Ganciclovir (GCV) implants are highly effective in delaying the progression of cytomegalovirus (CMV) retinitis. Rhegmatogenous retinal detachments can occur in untreated eyes with CMV retinitis or in eyes treated with anti-CMV therapy, which may include placement of a GCV implant. The clinical management of CMV retinitis and associated retinal detachment often involves the concurrent use of silicone oil and GCV implants. The authors investigated the effect of silicone oil tamponade on intravitreal drug levels achieved with the GCV implant. METHODS: The authors performed gas compression vitrectomy in the right eyes of 29 New Zealand white rabbits. They then inserted a 5-mg GCV implant into the vitreous cavity through an inferotemporal sclerotomy. Saline (1 cc), silicone oil 0.5 cc, or silicone oil 1.0 cc was then injected into the midvitreous cavity of 9, 8, and 12 rabbits, respectively. On postoperative days 21, 42, and 70, the rabbits were killed and the right eyes were immediately collected and stored at -70 degrees C until all samples were obtained. Vitreous was then isolated and drug levels were determined by high-pressure liquid chromatography. RESULTS: Vitreous GCV levels at days 21 and 42 were similar in both the saline-filled and silicone oil-filled eyes. At day 70, GCV levels in both the saline- and silicone-filled eyes were statistically significantly lower than at day 21 (P < 0.05 for all groups). In addition, at day 70, GCV levels in the saline-filled eyes were significantly lower than in silicone-filled eyes (saline versus 0.5 cc oil, P = 0.01; saline versus 1 cc oil, P = 0.09). CONCLUSIONS: Effective GCV levels are maintained in the aqueous phase of the vitreous cavity of eyes with silicone oil tamponade. Ganciclovir levels may be maintained longer in eyes with silicone oil tamponade than in those without. These results support the use of combined GCV implants and silicone oil tamponade in patients with CMV retinitis and associated retinal detachment.

Transpupillary thermotherapy of occult choroidal neovascularization in age-related macular degeneration.

PURPOSE: To evaluate the efficacy of transpupillary thermotherapy (TTT) in management of occult subfoveal choroidal neovascularization (CNV) in exudative age-related macular degeneration (AMD). METHODS: Retrospective chart review of eyes that were treated with TTT and had at least 12 weeks of follow-up. Base-line and final ETDRS visual acuity and fluorescein angiography (FA) were compared. RESULTS: For the 48 eyes which met inclusion criteria, mean pre-operative visual acuity was 20/128 (range: 20/50-20/500). Average follow-up was 27 weeks (range: 12 weeks-55 weeks). At 3 months after treatment, 12 eyes (25%) improved 2 lines or more, 18 eyes (37.5%) had no change or 1 line of visual improvement, and 18 eyes (37.5%) worsened 1 or more lines. No significant adverse event was noted during treatment. Three eyes developed large submacular hemorrhage within 2 months of treatment. Based on clinical examination and FA, 61% of the eyes appeared to have reduction of subretinal fluid compared to pre-operative evaluations. CONCLUSION: Visual acuity was stable or improved in 62.5% of eyes in our series and the treatment was well tolerated. Longer follow up and larger number of patients would be required to evaluate the ultimate benefit of TTT in management of occult CNV due to AMD.

Phylogeography of Caribbean lizard malaria: tracing the history of vector-borne parasites.

The Anolis lizards of the eastern Caribbean islands are parasitized by several species of malaria parasites (Plasmodium). Here I focus on two species of Plasmodium, using molecular data (mitochondrial cytochrome b sequences) to recover the phylogeography of the parasites throughout the Lesser Antilles and Puerto Rico. The two parasites were originally described as a single species, P. azurophilum, which infects both red and white blood cells. Here the two species are termed P. azurophilum Red and P. azurophilum White based on their host cell type. Six haplotypes were found in 100 infections sequenced of P. azurophilum Red and six in 45 infections of P. azurophilum White. Nested clade analysis revealed a significant association of geographical location and clades as well as a pattern of past fragmentation of parasite populations. This is consistent with the hypothesis that vector-borne parasites such as malaria may be subject to frequent local extinctions and recolonizations. Comparison of the phylogeography of the lizard and parasites shows only weak concordance; that is, the parasites colonized the lizards in the islands, but dispersal events between islands via vectors or failed lizard colonizations were present. The two parasites had different histories, P. azurophilum Red colonized the islands from both the north and south, and P. azurophilum White originated in the central Lesser Antilles, probably from P. azurophilum Red, then moved to both north and south. This is the first study to examine the biogeography of a pair of sibling species of vector-borne parasites within an island archipelago system.