M. Watson

Hardy-Weinberg quality control.

An efficient test of deviation from Hardy–Weinberg frequencies with one degree of freedom was applied to 44 marker loci in a genome scan, and 7 loci had a significant excess of apparent homozygotes (χ2(1) > 6) suggestive of typing error. In this example evidence for linkage did not increase when outliers were censored. Statistical quality control is an essential part of genotyping, and the effect of mistyping and map error should be considered in evaluating any genome scan.

Timed Ablation of Regulatory CD4+ T Cells Can Prevent Murine AIDS Progression

We describe successful immunotherapy of murine AIDS (MAIDS) in C57BL/6J mice based on the elimination of replicating CD4+ regulator T cells. We demonstrate that a single injection of the antimitotic drug vinblastine (Vb) given 14 days postinfection (p.i.) with LP-BM5 can prevent MAIDS progression. Treatment with anti-CD4 mAb at 14 days p.i. is similarly able to prevent MAIDS. Treatment at other time points with Vb or anti-CD4 mAb is ineffective. The effect is based on ablation of a replicating dominantly suppressive CD4+ T cell population, as indicated by adoptive transfer and in vivo depletion experiments using mAbs against CD4 as well as combinations of mAbs against the known regulatory cell surface markers CD25, GITR, and CTLA-4. Cell surface marker analysis shows a population of CD4+CD25+ cells arising shortly before day 14 p.i. Cytokine analyses show a peak in IL-10 production from day 12 to day 16 p.i. MAIDS-infected mice also have CD4+ T cells with significantly higher expression levels of CD38 and particularly CD69, which have been demonstrated to be regulator T cell markers in the Friend retroviral model. The immunotherapy appears to prevent disease progression, although no protection against reinfection with LP-BM5 is generated. These data define a new therapy for murine retroviral infection, which has potential for use in other diseases where T regulator cell-mediated immunosuppression plays a role in the disease process.

Bartonella henselae is a causative agent of cat scratch disease in Australia

We report the first isolation of Bartonella henselae from the blood and fleas of a cat of a patient with cat scratch disease (CSD) in Australia. A 49-year-old man presented with a history that 3 weeks after he had removed fleas from his cat he had developed fever, lethargy and anorexia for 3 days. This was followed by the appearance of axillary lymphadenopathy. There was no history of a bite or scratch and no primary lesion on the skin. Two fine needle aspirates of the axillary lymph node showed granulomatous lymphadenitis with no organisms seen by Warthin-Starry silver staining or electron microscopy. No organism was cultured from the patients lymph node aspirates or blood cultures processed by lysis centrifugation. However, the polymerase chain reaction (PCR) using p24E and p12B primers gave a 280 bp band indistinguishable from Bartonella henselae when using DNA extracted from the lymph node aspirates and the patients blood leucocytes. DNA sequencing of the PCR product from the patients blood showed that the DNA was from Bartonella henselae. The patients serum had a titre of 1024 in an indirect immunofluorescence antibody test for Bartonella henselae. Bartonella henselae was subsequently cultured from fleas and blood taken from the patients cat. This case provides evidence that Bartonella henselae is a causative agent of CSD in Australia and supports a possible role for fleas in transmission of the disease.

Efficacy of low-dose oral use of type I interferon in cytomegalovirus infections in vivo

Oral administration of type I interferons (IFNs; murine IFN-alpha and IFN-beta) reduces early replication of murine cytomegalovirus (MCMV) in both the spleen and liver of MCMV-infected BALB/c mice. Examination of a range of doses of IFN (1 to 1000 IU) showed that 10 IU administered daily for 1 week prior to virus infection was optimal for inhibition of MCMV replication. Furthermore, low-dose orally administered IFN (10 IU/day) was effective in mice challenged with lethal and sublethal virus inocula. The antiviral efficacy of low-dose orally administered IFN was not restricted by either the route of virus inoculation or the mouse genotype. Analysis by immunohistochemistry of IFN-alpha receptor-bearing cells of the gastrointestinal tract revealed predominant staining of perivascular smooth muscle and the lamina propria of the anterior tongue, small intestine and rectum. These tissues, dense in IFN-alpha receptor-bearing cells, are likely to be the sites of interaction of the orally administered IFNs with the mucosal immune system. In conclusion, we propose that low-dose oral use of type I IFN therapy may have broad applications in the treatment of CMV infections.

Toll-like receptor (TLR) expression on CD4+ and CD8+ T-cells in patients chronically infected with hepatitis C virus

Toll-like receptor (TLR) expression on T-cells and the signalling pathways that lead to the production of cytokines may limit antigen-specific T-cell responses. Here, expression of TLR and retinoic acid inducible gene I (RIG-I) on T-cells were evaluated in patients chronically infected with hepatitis C virus (HCV), before and during pegylated interferon-alpha and ribavirin therapy. Expression of TLR2,3,4,7,9 and retinoic acid inducible gene (RIG)-I on different CD4(+) and CD8(+) T-cell sub-populations (naïve: CD45RA(+)CD57(-); central memory: T(CM) CD45RA(-)CD57(-); effector memory: T(EM) CD45RA(-)CD57(+) and terminally differentiated effector memory: T(EMRA) CD45RA(+)CD57(+)) were measured by flow cytometry. TLR7, TLR9 and RIG-I expression on CD4(+) T-cells and RIG-I expression on CD8(+) T-cells was higher in patients than healthy controls. Therapy increased expression of TLR2, TLR4 and TLR9 and this was observed for all T-cell sub-populations. Evaluation of TLR expression at baseline did not identify patients able to achieve sustained virological response following therapy.

Bartonella henselae Associated with Parinaud's Oculoglandular Syndrome

Bartonella henselae was recovered from the conjunctival scraping of a 38-year-old woman who presented with a 2-week history of tender preauricular lymphadenopathy and a 1-day history of a red left eye. Dry adherent colonies were observed on agar plates at 21 days of incubation, and the isolate was identified through conventional and molecular tests. Polymerase chain reaction (PCR) amplification of a specific region of the 16S rRNA gene and confirmation by a separate PCR reaction with hybridization of the product with a B. henselae-specific probe confirmed the isolate as B. henselae. This is the first reported isolation of the causative agent of cat scratch disease from ocular tissue in a patient with Parinauds oculoglandular syndrome.

Upregulation of protein S by progestins.

Summary.  Background:  Users of progestin‐only contraceptives have raised protein S (PS) levels compared with baseline. This contrasts with the reduction in PS levels observed in users of combined oral contraceptives, which contain both a progestin and an estrogen.

Exclusion from proximal 11q of a common gene with megaphenic effect on atopy

We have typed three markers on proximal 11q in 131 random families with three or more children studied for atopy. A summary map that includes the FCER1B candidate was constructed. Using a 2‐locus disease model, we performed combined segregation and linkage analysis of three models, none of which suggested linkage. Nine marker loci on other chromosomes were also negative. In the regions swept by these 12 markers we cannot rule out a rare gene, perhaps of large effect, nor a common gene of small effect. However, a common gene of large effect is excluded. These results and alternative strategies are discussed in the perspective of inconsistent evidence for a major atopy gene.

Increased Proportion of the CD56 bright NK Cell Subset in Patients Chronically Infected With Hepatitis C Virus (HCV) Receiving Interferon-α and Ribavirin Therapy

Natural killer (NK) cells are implicated in the regulation of a protective immune response in patients chronically infected with hepatitis C virus (HCV), but effects of interferon‐α/ribavirin therapy on NK cell subsets and the consequences of viral clearance during therapy remain unclear. Samples were collected from chronically infected patients (n = 34) at baseline and from a subset after 3–10 months on pegylated interferon‐α and ribavirin therapy (n = 19). NK cells present in cryopreserved PBMC were characterized by flow cytometry. Before therapy, the frequency of CD3−CD56+ NK cells was lower in patients than uninfected controls. Therapy increased proportions of CD56bright NK cells. Frequencies of CD56dim NK cells declined slightly while perforin and CD16 expression on CD56dim NK cells decreased compared to baseline samples. Evaluation of NK cell subsets at baseline did not identify patients able to achieve sustained virological response following therapy. However, therapy may promote the expansion of NK cells able to produce interferon‐γ, while minimizing cytotoxicity to limit liver damage. J. Med. Virol. 82:568–574, 2010.

The CrP operon of Chlamydia psittaci and Chlamydia pneumoniae

One of the critical developmental events during the unique intracellular life cycle of Chlamydiae is their differentiation from a metabolically active, replicative form or reticulate body (RB) to an infectious extracellular form of the organism (elementary body or EB). This process is characterized by the expression of two extraordinarily cysteine-rich envelope proteins of molecular masses 9 kDa and 60 kDa. We describe the molecular cloning and sequence determination of the 9 kDa cysteine-rich proteins (CrPs) of C. pneumoniae and C. psittaci. Comparison of these 9 kDa CrP amino acid sequences with those of C. trachomatis showed regions of structural variation and conservation. Transcription of the 9 kDa CrP genes occurred as both a monocistronic message and as a bicistronic message which included the 60 kDa CrP gene. Transcription of the 9 kDa and 60 kDa CrP genes was tightly linked to the chlamydial growth cycle with synthesis of their mRNAs and consequent translation of the 60 kDa CrPs occurring as RBs differentiated to form EBs. The maximal rate of transcription occurred late in the growth cycle from a single but highly conserved promoter which had close similarity with the Escherichia coli consensus promoter sequences. A stem and loop structure which could be involved in regulating translation of mRNA occurred in all three species between the transcriptional start point and the ribosome binding site. Although transcription is initiated from a single promoter in all three chlamydial species, transcriptional termination points for the monocistronic and bicistronic mRNAs differ in both number and position.